Microorganisms doi: 10.3390/microorganisms14061193

Authors: Zhijing Yang Wentao Xv Yingchun Cai Hailong Gu Yaming Feng

Aquaculture sediments are increasingly recognized as important reservoirs of antibiotic resistance genes (ARGs). Although thermophilic fermentation is widely used to reduce ARGs and pathogens in manure, most biosafety assessments stop at the fertilizer product itself, leaving unresolved whether these benefits persist after application to aquaculture sediments. Here, we compared inorganic fertilizer (IF), raw manure (RM), and fermented fertilizer (FF) to test whether fermentation confers sustained biosafety benefits in aquaculture pond sediments. After a 6-month co-culture period, sediment samples were analyzed using shotgun metagenomic sequencing, ARG and mobile genetic element (MGE) profiling, antibiotic residue determination, and network analyses. Long-term fertilization significantly altered sediment physicochemical properties, microbial community composition, and resistome structure. Among the three groups, the RM exhibited the highest total ARG abundance and the greatest number of unique ARG subtypes, with significant enrichment of multidrug resistance genes as well as pathogen-, disease-, and host-associated mobile genetic elements (MGEs). In contrast, the FF group showed the lowest total ARG abundance and fewest unique ARG subtypes, along with suppression of pathogen-associated MGEs, indicating that FF can effectively reduce the risk of ARG dissemination. However, the potential impact of residual antibiotics still warrants attention. Redundancy analysis showed that TC and TN primarily explained bacteriome and resistome variation under RM, whereas pH, EC, AP, and AK were more strongly associated with FF. Co-occurrence analysis further suggested that fertilizer-driven microbial community shifts may regulate ARG persistence and potential cross-ecosystem dissemination. Overall, fermented fertilizer attenuated, but did not eliminate, manure-derived resistance risks in aquaculture sediments. These findings support fermented fertilizer as a safer management option than raw manure and highlight the need for integrated risk assessment combining ARGs, MGEs, microbial hosts, and antibiotic residues.

Microorganisms doi: 10.3390/microorganisms14061192

Authors: Shu-Chen Chang Jianchi Chen Chung-Chieh Lee Ming-Yao Chiang Hsuan Shentu Hsien-Tzung Shih Adalberto Á. Pérez de León

The two-spotted spider mite, Tetranychus urticae, poses significant agricultural challenges due to its rapid population growth and high capacity for developing chemical resistance. This study evaluated the acaricidal activity of bacterial strain TAM1, isolated from naturally deceased mites in Taiwan. In bioassays, TAM1 caused over 90% adult mite mortality within 48 h. Infected mites showed symptoms of body darkening and deformation of the ventral abdominal crest lines. Enzymatic analysis confirmed significant chitinase and gelatinase activities. Whole genome sequence of TAM1 was acquired with a 5,066,903 bp circular chromosome (CP120954) and a 164,574 bp circular plasmid (CP120955). Refined functional profiling identified a sophisticated enzymatic arsenal including core chitin-active families (GH18, GH20, AA10) and 157 proteases, with a high prevalence of metallopeptidases that correlate with the detected gelatinase activities. Secretome analysis predicted 42 extracytoplasmic proteases primarily utilizing the Sec-dependent pathway, while the presence of multiple CBM50 modules suggests a potential for targeted substrate anchoring. These genomic insights provide a plausible molecular basis for the observed enzymatic potential and the localized ultrastructural disruption of the T. urticae cuticle. The alignment between phenotypic observations, microscopic evidence, enzymatic activities, and genomic data suggests that TAM1 utilizes synergistic, multi-target mechanisms to exert its acaricidal effects. Based on analyses of whole-genome sequence and 16S rRNA gene sequence, TAM1 was tentatively designated as a strain of Kosakonia sacchari. The bacterial strain reported here represents a promising microbial agent for integrated pest management (IPM) programs.

Microorganisms doi: 10.3390/microorganisms14061191

Authors: Wang Liu Mongi Ennasri Christopher A. Hempel Mohammad A. Qurban Carlos M. Duarte Susana Agustí

Benthic prokaryotic communities in deep-sea sediments remain poorly studied. They are constrained by organic matter availability and oxygenation in warm deep-sea ecosystems. Here, we investigated benthic prokaryotic communities and carbon uptake in deep Red Sea sediments (218–2415 m seafloor depth), where persistently warm (~21.5 °C) waters and a strong south–north productivity gradient co-occur. Sediment particulate organic carbon (POC), prokaryotic abundance (PA), and [13C]-D-glucose-based carbon uptake and uptake kinetics were examined in two sediment layers (0–1 and 4–5 cm), while bacterial communities were characterized using 16S rRNA gene sequencing of the 0–1 cm layer. Sediment POC, PA, and carbon uptake declined northward, consistent with reduced organic-carbon supply to the seafloor. Bacterial community composition differed significantly across the ~500 m depth associated with the Red Sea oxygen minimum zone (OMZ). Sediments from the relatively low-oxygen upper OMZ-range (200–500 m) had higher sediment POC and PA, and were enriched in putatively anaerobe-associated taxa, whereas deeper sediments (>500 m) below the OMZ exhibited more fragmented co-occurrence networks. These results suggest that organic-carbon availability defines the basin-scale metabolic backdrop, whereas bacterial community differentiation was more clearly resolved between upper OMZ-range and below-OMZ sediments than along latitude alone.

Microorganisms doi: 10.3390/microorganisms14061189

Authors: Serena Bergamo Giusto Trevisan Giovanna Muffato Diana Sacchi Serena Bonin Alessandro Gatti

Mycobacterium chelonae is a rapidly growing nontuberculous mycobacterium (NTM) that can infect both immunocompetent and immunocompromised hosts. Cutaneous and soft tissue infections are the most common manifestations and occur more frequently in individuals with underlying immune dysfunction. Patients with chronic lymphocytic leukemia (CLL), particularly those receiving targeted therapies such as ibrutinib, may be at increased risk of opportunistic infections. The diagnostic workup, microbiological findings, antimicrobial susceptibility testing, and therapeutic approach adopted for a cutaneous M. chelonae infection arising in a CLL patient four months after the introduction of ibrutinib were described. Clinical course and surgical management are also reported. A 60-year-old beekeeper with B-cell CLL developed a progressive cutaneous lesion on the left lower limb within four months of starting ibrutinib. Culture of a skin biopsy identified M. chelonae. Antimicrobial therapy was initiated based on in vitro susceptibility testing, resulting in partial clinical improvement. Complete resolution required surgical excision of the infected tissue followed by skin grafting. The patient’s underlying hematologic disease, ongoing immunosuppression, and recent exposure to ibrutinib likely contributed to susceptibility and persistence of infection. This case highlights the increasing recognition of nontuberculous mycobacterial infections in immunocompromised individuals and underscores the importance of early diagnosis and susceptibility- guided therapy. Clinical response may be incomplete, and combined medical and surgical approaches may be required in selected cases. NTM infections should be considered in patients receiving Bruton’s tyrosine kinase inhibitors who present with persistent, atypical, or non-healing cutaneous lesions. However, the association between ibrutinib therapy and susceptibility to infection remains uncertain, as multiple predisposing factors may coexist. Increased awareness of this possible association, together with careful clinical evaluation, may facilitate earlier diagnosis and improved management.

Microorganisms doi: 10.3390/microorganisms14061190

Authors: Yixinfei Lin Hongfeng Bian Yanan Liu Pengchen Zhou Xue Wang

Microbial functional traits play a central role in regulating carbon mineralization efficiency (CME) in peatlands, yet how they respond to concurrent warming and atmospheric nitrogen deposition remains unclear. In this study, peat soils from three vegetation types (sedge, reed, and shrub) were subjected to controlled microcosm incubations simulating warming and nitrogen addition gradients. Microbial community composition and functional profiles were characterized using 16S rRNA high-throughput sequencing and Functional Annotation of Prokaryotic Taxa (FAPROTAX) functional prediction, while dissolved organic matter (DOM) composition was analyzed via excitation–emission matrix fluorescence spectroscopy with parallel factor analysis (EEM-PARAFAC) and fluorescence indices. Integrating correlation analysis, Random Forest, and partial least squares path modeling (PLS-PM) modeling, we identified microbial functional traits as key factors linking environmental changes to soil CME, with DOM serving as a substrate-mediated pathway. External nitrogen input primarily drove shifts in microbial functional composition, whereas warming modulated substrate utilization preferences and DOM turnover. The interaction between warming and nitrogen selectively reshaped microbial functional profiles, thereby jointly determining CME. Functional traits explained more variation in CME than taxonomic composition, indicating a “structure–function decoupling” under environmental change. These findings highlight the central role of microbial functional traits in peatland carbon transformation and suggest that the net response of peatland carbon emissions to future environmental change will depend critically on the balance between warming magnitude and nitrogen deposition levels.

Microorganisms doi: 10.3390/microorganisms14061188

Authors: Yazhi Feng Yiyao Liu Guangxin Chen Changxin Wu

Butyrophilin-like 2 (BTNL2) is an immunomodulatory molecule critically involved in regulating the host immune response to infection with the avirulent Mycobacterium tuberculosis strain H37Ra. However, its functional role in modulating pyroptosis and associated inflammatory responses remains incompletely characterized. Here, we demonstrate that BTNL2 deficiency exacerbates pyroptosis and the inflammatory response in H37Ra-infected murine peritoneal macrophages via two distinct pathways. First, the loss of BTNL2 induces excessive mitochondrial damage, which leads to aberrant release of mitochondrial DNA (mtDNA) and accumulation of mitochondrial reactive oxygen species (mtROS), thereby triggering NLRP3 (NOD-like receptor family pyrin domain containing 3) inflammasome activation and gasdermin D (GSDMD)-mediated pyroptosis. Second, cytosolic mtDNA accumulation hyperactivates the cGAS–STING signaling axis, resulting in transcriptional upregulation of NLRP3 and consequent amplification of pro-inflammatory cytokine production. Collectively, these findings demonstrate that BTNL2 acts as a regulator of mitochondrial homeostasis and innate immune balance during H37Ra infection in primary peritoneal macrophages. The results provide mechanistic insights into BTNL2 function in the context of H37Ra-induced pyroptosis.

Microorganisms doi: 10.3390/microorganisms14061187

Authors: Lorenzo Chinellato Lisbeth Truelstrup Hansen Martin L. Kragh Nina Gringer Claus H. Bang-Berthelsen

Underutilized and abundant in the Danish coastal area, Carcinus maenas has become a threat to the environment and to the local fisheries. In this study we investigated the microbiota and presence of microbial hazards of interest for human health in crabs caught over the period of one year, to investigate its potential for human consumption. Between 2023 and 2024, four seasonal samples of live specimens (n = 5) were caught off the Lillebælt (DK) coastal area. To characterize the microbiota of the crabs, a culture-dependent approach was used to determine total aerobic mesophilic count, total aerobic psychrotrophic count, Bacillus spp., Enterobacteriaceae, lactic acid bacteria, fungi, Salmonella spp., Listeria monocytogenes and Bacillus cereus. MALDI-TOF was used to corroborate results and further identify isolated microorganisms. The results were then compared with data obtained from amplicon sequencing of community 16S rRNA genes to compare family-level compositions of the microbiota. Of the pathogens of interest, B. cereus was detected during summer/autumn, reaching a maximum of 2.5 log cfu/g. Salmonella spp., and L. monocytogenes were below the limit of detection (<1 cfu/0.1 g). Spoilage bacteria were detected (e.g., Brochothrix spp., Carnobacterium spp., Photobacterium spp., Pseudomonas spp., Psychrobacter spp. and Shewanella spp.). The study highlighted significant seasonal changes (PERMANOVA, FDR-adjusted p = 0.00003) in the microbial composition. The gathered evidence suggests that with proper handling, the crabs could represent a safe resource.

Microorganisms doi: 10.3390/microorganisms14061186

Authors: Kai Yang Fuchun Huang Wensheng Yang Xupeng Lu Zhengtao Zhu Jianqiang Zhu Qixia Wu Xiaohong Xu

The ecological restoration of degraded sandy land in the Yarlung Zangbo River Valley is constrained by the metabolic functions of soil microorganisms. This study investigates the dynamic mechanisms of microbial elemental use efficiency in walnut plantations, with a focus on seasonal variations in soil chemical stoichiometry, extracellular enzyme activity, and microbial nutrient efficiency in rhizosphere and bulk soils. This paper explores the effects of conventional organic fertilizer (CF) and organic–inorganic compound fertilizer (OIF) on microbial nutrient use strategies and their seasonal dynamics. The results showed significant seasonal fluctuations in soil active nutrients and microbial biomass, while the total nutrient content remained stable. OIF enhanced microbial chemical stoichiometric homeostasis but simultaneously triggered a “carbon–phosphorus metabolic trade-off”, leading to a restraint of microbial carbon use efficiency (CUE) during the growing season. Microbial elemental use efficiency (EUE) exhibited clear seasonal differentiation: CUE was higher in summer, promoting biomass accumulation, whereas NUE and PUE increased in winter and spring, reflecting a nutrient conservation strategy. The EUE pathways were decoupled between rhizosphere and non-rhizosphere microenvironments. The rhizosphere was more directly driven by soil chemical stoichiometry and microbial biomass, while the non-rhizosphere was influenced by nutrient limitation states, represented by vector characteristics. This study provides insights into the seasonal adaptability and microenvironmental heterogeneity of microbial metabolism during the restoration of cold sandy land. It is suggested that future ecological management should focus on N-P balanced fertilization and consider the differential responses between rhizosphere and non-rhizosphere zones to enhance ecosystem productivity and soil carbon, nitrogen, and phosphorus sequestration potential.

Microorganisms doi: 10.3390/microorganisms14061184

Authors: Hongqiao Li Chenyu Liu Yan Tang Zongmei Chen Song Ge

Periodontitis is closely linked to atherosclerosis; however, the role of the keystone periodontal pathogen Porphyromonas gingivalis (P. gingivalis), particularly its virulence factor, outer membrane vesicles (OMVs), in vascular smooth muscle cell (VSMC) dysfunction remains unclear. This study aimed to explore the effects of P. g-OMVs on mouse aortic smooth muscle cells (MOVAS) and the potential involvement of cytoskeleton-associated protein 4 (CKAP4) in this process. OMVs were isolated by ultracentrifugation and characterized by transmission electron microscopy (TEM), nanoparticle tracking analysis (NTA), and Western blotting. MOVAS cells were treated with OMVs; cellular functions were evaluated using CCK-8, colony formation, scratch wound-healing, ELISA, and Western blotting assays. Lentiviral vectors were used to construct CKAP4 overexpression and knockout cell models. Results showed that after P. g-OMVs were internalized by MOVAS cells, the cells showed cytoskeletal disorganization, promoting cell proliferation, wound closure, and contractile-to-synthetic phenotypic switching (decreased α-SMA and increased OPN expression), and enhancing extracellular matrix (ECM) remodeling (upregulated expression of type I collagen, type III collagen, fibronectin, matrix metalloproteinase-2 and -9, and tissue inhibitor of metalloproteinase-1). At the protein level, P. g-OMV treatment was associated with upregulated expression of CKAP4, integrin α5, and integrin β1; CKAP4 overexpression synergized with OMV stimulation to amplify these phenotypic alterations, whereas CKAP4 knockout attenuated these cellular changes. These findings suggest an association between CKAP4 upregulation and P. g-OMV-induced MOVAS dysfunction, indicating that CKAP4 may serve as a potential target in periodontitis-associated atherosclerosis.

Microorganisms doi: 10.3390/microorganisms14061185

Authors: Lina Wu Shuai Lu Fanjin Ye Jinxia Lu Xiaoling Liu Yanfang Tian

This study systematically compared the structural, functional, pathogenic, and assembly-mechanism characteristics of bacterial communities between urban and rural rivers across China, based on integrated water quality data from 421 sampling sites and 16S rRNA gene sequences from 475 sampling sites. The results revealed that urban rivers had significantly higher nutrient concentrations and bacterial α-diversity, along with lower β-diversity. Urban rivers were enriched with organic matter-degrading phyla such as Chloroflexi and Acidobacteriota and might exhibit more complex co-occurrence networks (average degree: 85.41). In contrast, rural rivers were enriched with phyla including Firmicutes and Cyanobacteria, as well as genera such as Exiguobacterium and Limnohabitans, and might display higher network modularity (modularity: 0.59) and greater spatial heterogeneity in community composition. Functional prediction indicated stronger carbon-cycling potential in urban rivers, whereas nitrogen-cycling functions did not differ between the two river types. Regarding pathogen composition, urban rivers contained a higher number of pathogen species than rural rivers. It was suggested that stochastic processes dominated community assembly in both systems; however, heterogeneous selection contributed more strongly in urban rivers (14.7%). Overall, this work elucidated systematic differences in bacterial community structure, function, pathogen profile, and assembly mechanisms between urban and rural rivers, offering a scientific foundation for differentiated watershed management.

Microorganisms doi: 10.3390/microorganisms14061183

Authors: Layla Ahmed Mohammed Abdelhadi Nasir A. Ibrahim Mohammed Osman Abdelrahem Essa Sheham Guma Ibrahim Nosiba S. Basher Hosameldeen Mohamed Husien Guoqiang Zhu

Adherent-invasive Escherichia coli (AIEC) is considered a central pathogen in Crohn’s disease (CD), as it was first identified in 1998 in an ileal biopsy from a CD patient and has received extensive attention worldwide, but there is currently a lack of quantitative evaluation of the literature in this field. This study aims to provide a bibliometric analysis of AIEC in CD research over the past 25 years. Publications on AIEC in CD research from 1999 to 2025 were extracted from the Web of Science Core Collection database. Bibliometric analysis was conducted using VOSviewer (1.6.18), RStudio (version 4.0), and HistCite (version 12.03.17) to visualize by author, country, institution, journal, reference, and keywords. A total of 250 articles were analyzed. Overall, the number of annual publications in this field has increased since 2007, with noticeable fluctuations. Barnich Nicolas published the most articles, 66, and has a high impact in the field of AIEC in CD research. The most cited author within this data set by global citation score was Arlette Darfeuille-Michaud. Among countries and institutions, France and Université Clermont Auvergne (UCA) had the highest number of publications. The journal with the most publications is Inflammatory Bowel Diseases. The most frequent keyword was “adhesion”. Over the past 25 years, France has led the rapid growth in AIEC research. Furthermore, according to trend topic analysis, the prevalence of AIEC in CD continues to be the central research theme. Our study provides valuable insights into AIEC research, supporting international collaboration to advance AIEC research in CD. This study provides a visual overview of the field of AIEC in CD.

Microorganisms doi: 10.3390/microorganisms14061182

Authors: Hosana Dau Ferreira de Souza Thereza Cristina da Costa Vianna Juliana Ferreira Nunes Vinícius Carneiro Assunção Ana Paula Alves do Nascimento Ramon Loureiro Pimenta Alexander Machado Cardoso Maysa Mandetta Clementino Miliane Moreira Soares de Souza Irene da Silva Coelho Kayo Bianco Shana de Mattos de Oliveira Coelho

Wastewater treatment plants (WWTPs) are important interfaces for the persistence and dissemination of antimicrobial resistance genes (ARGs) in the environment. This study investigated colistin resistance and the presence of mobile colistin resistance (mcr) genes in Enterobacterales isolated from a poultry slaughterhouse WWTP in Brazil. Samples were collected from raw sewage, an equalization tank, and treated effluent. A total of 27 Enterobacter spp. isolates were identified, of which 70.4% showed resistance to colistin (MIC range: 2 to ≥512 mg/L). PCR screening detected mcr-1 in two isolates and mcr-10 in three isolates distributed across all treatment stages, including the final effluent. Whole-genome sequencing of a representative isolate from treated effluent identified Enterobacter vonholyi ST3343, carrying a plasmid-borne mcr-10 gene on an ~107 kb IncFII(Yp) plasmid, along with additional resistance determinants. Phylogenetic analysis supported the classification of this gene as a novel allele, mcr-10.6. The persistence of a clonal lineage harboring mcr-10.6 throughout the treatment system indicates that conventional wastewater treatment may not effectively eliminate clinically relevant ARGs. These findings highlight treated effluent as a potential route for environmental dissemination of colistin resistance and reinforce the need for improved monitoring and mitigation strategies within a One Health framework.

Microorganisms doi: 10.3390/microorganisms14061181

Authors: Ke Li Huifang Zhao Di Meng Xinlei Zhang Zemin Fang Juanjuan Liu

Some microorganisms present in the cultivation environment serve as biocontrol agents and contribute to enhanced mushroom production. However, the native bacteria naturally associated with commercial cultivation bags of Pleurotus ostreatus, as well as their growth-promoting roles and underlying mechanisms, remain poorly understood. This study aimed to identify native bacteria that promote Pleurotus ostreatus development and to investigate the underlying mechanisms. Four native bacteria, including Brevibacterium epidermidis (P6), Acinetobacter soli (A7), Pseudomonas parafulva (A8), and Pseudomonas hunanensis (A12), were isolated based on their ability to promote mycelial growth of P. ostreatus. B. epidermidis P6 shortened complete mycelial colonization time from ~30 d to 14 d in dual cultivation bags. All four strains increased fresh mushroom yield, with B. epidermidis P6, A. soli A7, and P. parafulva A8 increasing the number of basidiomata, while P. hunanensis A12 enhanced their size. These strains produced exopolysaccharides that enhanced mycelial growth. Additionally, B. epidermidis P6, A. soli A7, and P. parafulva A8 also secreted extracellular crude proteins that also promoted mycelial growth. Bi-plates and further gas chromatography–mass spectrometry analysis demonstrated that volatile organic compounds from P. hunanensis A12, including acetone, 2-butanone, benzaldehyde, and 1-undecene, enhanced fungal mycelial growth. The mycelial growth rates of Ganoderma lucidum and Pleurotus pulmonarius were also enhanced by these four strains. These results reveal that four native bacterial strains promote mushroom development through complex mechanisms.

Microorganisms doi: 10.3390/microorganisms14061180

Authors: Boyanka Angelova Silvena Boteva Anelia Kenarova

The global methane budget is largely driven by biogenic sources, many of which remain insufficiently characterized. Here, we investigated the community composition and seasonal dynamics of methanogenic and methanotrophic assemblages to elucidate the key contributors to methane cycling and the environmental factors shaping these processes in lake sediments of the Rila Mountains (Bulgaria). Methanogenic communities are primarily composed of Methanothrix, Methanosarcina, Methanobacterium and Methanoregula with summer peaks in Methanothrix and Methanoregula, and cold-season proliferation of Methanobacterium. Methanotrophic communities are dominated by representatives of the Pseudomonadota, including Crenothrix, Methylobacter, and Methylocystis with summer maxima observed for Crenothrix and Methylobacter. Methanosarcina and Methylocystis showed relatively stable abundances throughout the ice-free season. Ordination and correlation analyses revealed that temperature, pH, and carbon (organic and inorganic) concentration and lability emerged as the environmental drivers influencing on microbial communities, with seasonally variable effects on methane-cycling microorganisms. These findings provide a foundation for future research on methane cycling in alpine lake ecosystems of the Rila Mountains and contribute to improving predictions of methane emissions under changing climatic conditions.

Microorganisms doi: 10.3390/microorganisms14061179

Authors: Jianchao Cui Haijiao Xu Liying Fan Yu Wang Limin He Zhaoyuan Wang Jicheng Han Jie Li Qihang Tian Wenshi Zhao Yonghong Li

Peach bacterial shot hole is a major disease limiting the yield and quality in most peach-producing areas worldwide. To clarify its etiology and support the development of targeted management strategies, diseased samples were collected from Changli County peach orchards. The pathogen was isolated, purified and verified by Koch’s postulates. Based on morphological, biochemical and multi-locus phylogenetic analyses, the causal agent was identified as Xanthomonas arboricola pv. pruni (isolate TCK-5). Biological characterization revealed that TCK-5 grew optimally in KB and NB medium at 28 °C, pH 7.0–7.5 and 0.5–1.0% NaCl, efficiently utilized glycerol and organic nitrogen source (proteose peptone, beef extract and yeast extract), with light showing no significant effect on growth. The strain TCK-5 exhibited a lethal temperature of 51 °C, indicating that heat treatment above this threshold effectively disinfects pruning tools and contaminated plant debris. Among 18 bactericides tested in vitro, biological bactericide outperformed chemical ones, with 0.3% Tetramycin AS (EC50 = 0.1051 mg/L) and 3% Zhongshengmycin SL (EC50 = 2.9252 mg/L) exhibiting the strongest inhibitory activity. This study fills a regional knowledge gap in the epidemiological distribution of the pathogen in northern China and advances current understanding of X. arboricola pv. pruni occurrence, providing a scientific basis for subsequent epidemic monitoring and integrated control of peach bacterial shot hole.

Microorganisms doi: 10.3390/microorganisms14061178

Authors: Bei Zhao Liubin Huang Qi Zuo Yanxia Fan Muhammad Yousuf Adnan Sen Wang Fengxia Shao

Xinjiang Ziziphus jujuba ‘Huizao’ wine, a characteristic fruit wine in China, is facing industrial bottlenecks such as flavor homogenization and lack of specialized fermentation yeasts, which limits its high-quality development. To solve these problems, four laboratory-preserved indigenous yeast strains (NZ5, NZ6, BH4, BH2) were compared with four commercial strains (FR, RW, RA, SY) in terms of fermentation dynamics, volatile flavor compound synthesis (HS-SPME-GC-MS/GC-FID), and sensory quality to screen the optimal yeast for Ziziphus jujuba ‘Huizao’ wine fermentation. Molecular identification revealed that NZ5 and NZ6 belong to Saccharomyces cerevisiae, while BH4 and BH2 are closely related to Pichia kudriavzevii, respectively, indicating their non-Saccharomyces characteristics with distinct metabolic potentials. The results showed that indigenous strains exhibited significantly superior performance to commercial strains: (1) Saccharomyces strains NZ5 and NZ6 had higher fermentation efficiency, with 12.5–25% shorter fermentation cycles and 14% higher cumulative CO2 release than commercial strains; (2) Non-Saccharomyces strain BH4 synthesized the most diverse volatile flavors (99 compounds), with ethyl acetate content reaching 314.92 mg/L, which was 13-fold higher than that of commercial yeast FR (24.09 mg/L). Meanwhile, its phenethyl alcohol content reached 3.12 mg/L, 7.2 times that of commercial yeast RW; (3) Sensory evaluation showed that BH4-fermented wine had the highest score (88.59), significantly higher than commercial strains (63.57–67.67). In conclusion, BH4 is the optimal strain for improving the flavor quality of Xinjiang Ziziphus jujuba ‘Huizao’ wine, and NZ5/NZ6 are suitable for efficient industrial fermentation. This study provides valuable microbial resources and technical guidance for the quality improvement and industrial development of Xinjiang characteristic fruit wine.

Microorganisms doi: 10.3390/microorganisms14061177

Authors: Dan Hao Xiaohang Yu Xiangyang Sun Dongdong Cheng Hao Ding Yige Wang Yalin Li Zhewen Geng Guijun Xu

Thermophilic microbial inoculant (CI) has been demonstrated to optimize the green waste composting (GWC) process. The pathways through which it enhances lignocellulose degradation remain unclear. This study evaluated composting performance under four treatments: CI, effective microorganisms (EM), Phanerochaete chrysosporium (WF), and natural composting (CK). To elucidate the biological differences between efficient lignocellulose-degrading systems and CK, metagenomic analyses were conducted on CI and CK based on lignocellulose degradation rates. The results indicated that CI inoculation did not negatively affect the compost heating process and produced a nitrogen-rich, safe, and mature compost product. Compared to other treatments, CI increased the lignocellulose degradation rate by 3.66% to 31.8%. Metagenomic analysis revealed that CI inoculation enriched genes encoding glycoside hydrolases (GHs), glycosyl transferases (GTs), carbohydrate esterases (CEs), and carbohydrate-binding modules (CBMs) across multiple composting phases, positively impacting dominant carbohydrate-active enzyme (CAZyme) families including AA3, CE1, and CE7. CI inoculation also elevated the relative abundance of lignocellulose-degrading microorganisms (0.70~2.73%), simplified microbial network structure, and strengthened microbial cooperation. Within the microbial network, Chryseolinea, Protaetiibacter, and unclassified_f__Burkholderiaceae were identified as core taxa involved in lignocellulose degradation. Redundancy analysis (RDA) identified temperature as the primary factor influencing biological factors, with CI improving composting efficiency by optimizing the microenvironment. Collectively, this work provides a novel strategy for microbial inoculant application in composting and offers new perspectives for identifying core taxa, contributing to advancing composting efficiency.

Microorganisms doi: 10.3390/microorganisms14061176

Authors: Leonardo Miranda dos Santos Rodrigo Covre Vieira Louise de Souza Canto Covre Milena Cristina Martins da Silva Thiago de Matos Bezerra Geraldo Mariano Moraes de Macedo Edna Aoba Yassui Ishikawa Karla Valéria Batista Lima Maísa Silva de Sousa Rodrigo Vellasco Duarte Silvestre

Sexually transmitted infections (STIs) caused by C. trachomatis and HPV are the most prevalent worldwide. College students are characterized by being young women of reproductive age who may have risky sexual behavior. To describe the prevalence and factors associated with endocervical infection by C. trachomatis and HPV in college women in the Brazilian Amazon. Endocervical secretions were collected. The ompA gene of C. trachomatis and the L1 gene of HPV were detected. The Chi-square test, Fisher’s exact test, G test, Odds Ratio, and Multiple Logistic Regression were used with 95% confidence interval and p ≤ 0.05. The overall prevalence of endocervical infection by C. trachomatis was 8.3% (25/302) and by HPV was 28.9% (87/302). Low income was associated with sexually transmitted infection by C. trachomatis (14.8%, p = 0.0336). Those under 25 years old had twice the chance of HPV infection [39.3%, (OR: 2.6989), 95% CI: 1.6054–4.5371, p = 0.0002], as did women without children [31.8%, (OR: 2.333), CI: 1.1235–4.8461, p = 0.0307]. Women who did not study in a public school had 63% reduced risk of acquiring HPV infection [45.8% (OR: 0.3713), CI: 0.1951–0.7064, p = 0.0035]. C. trachomatis and HPV infections were present in low-income, childless young women who attended public schools, requiring the intensification of STI prevention policies in the Amazon region.

Microorganisms doi: 10.3390/microorganisms14061175

Authors: Rosalind T. B. Herrington Zhen Lyu David T. Ellenberger Nathan J. Bivens Zhentian Lei Tanhaul Islam Lloyd W. Sumner R. Michael Roberts Trupti Joshi Cheryl S. Rosenfeld

Probiotic supplements are increasingly being touted to have health benefits for pregnant women consuming such supplements and their unborn offspring. The placenta is in direct communication with maternal blood, and bioactive agents can thus easily be transferred to this organ where they may influence gene expression by the different trophoblast (TB) cell lineages. The underlying hypothesis assessed herein is that maternal probiotic supplementation can influence the placenta and fetal brain. The composition of bacterial short-chain fatty acids (SCFAs) was examined in fecal boli of mouse dams on a maternal probiotic supplement relative to control dams. Further, SCFA and transcriptomic profiles were examined in placenta and fetal brain from conceptuses derived from dams on the probiotic supplement and conceptuses from control dams. While this treatment did not affect bacterial SCFAs, placenta and fetal brain changes were evident in male and female conceptuses carried by dams receiving probiotics relative to controls. For the placenta, females were more sensitive to maternal probiotic supplementation, whereas the opposite was the case for the fetal brain. Slc6a4 showed increased expression in female placenta from probiotic-treated dams, which could enhance uptake of maternal 5-HT. Male placenta from probiotic-treated dams had dramatic reduction in Hsd11b2 that may render them more vulnerable to maternal stress. In the fetal brain, maternal probiotic supplementation was associated with genes linked to forebrain development, suggesting this treatment might impact life-long neurobehavioral responses. Current studies suggest that maternal probiotic supplementation might lead to adverse changes in the placenta and fetal brain of their unborn children.

Microorganisms doi: 10.3390/microorganisms14061174

Authors: Liu Wu Zongyan Li Jinxuan Zhu Zhigang Sun Lujun Yan Mingzhan Luo Huahai Chen Yeshi Yin

The gut microbiome, often termed the human “second genome”, profoundly influences host physiology through metabolic interactions, immune modulation, and gut–brain axis signaling. Dysbiosis is implicated in the pathogenesis of obesity, inflammatory bowel disease (IBD), malignancies, and neuropsychiatric disorders. However, traditional gut microbiota interventions, such as probiotic supplementation and fecal microbiota transplantation (FMT), still exhibit significant limitations in precision therapeutics. Probiotic intervention fails to achieve precise regulation at the strain or genetic level, and although FMT demonstrates definitive efficacy against recurrent Clostridioides difficile infection (rCDI), its therapeutic outcomes and safety profiles show marked interindividual variability in ulcerative colitis (UC), metabolic syndrome, and other diseases, with insufficient treatment specificity to meet the practical demands of clinical precision intervention. Recent advancements in genome editing technologies, particularly Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)–CRISPR-associated (Cas) proteins systems and base editors, have enabled targeted functional manipulation of specific gut commensals and optimization of community architectures. These engineered strategies, combined with sophisticated delivery systems, demonstrate substantial potential in disease treatment, diagnostic monitoring, and immune modulation. This review systematically examines core editing methodologies, innovative delivery platforms, and targeted design strategies, elucidating their applications in metabolic disorders, IBD, cancer immunotherapy, and neuropsychiatric conditions. We critically analyze current technical bottlenecks and biosafety concerns while prospecting future directions, including in situ editing, artificial intelligence (AI)-driven design, and personalized engineering. Collectively, these insights aim to facilitate the clinical translation of gut microbiome engineering from bench to bedside.

Microorganisms doi: 10.3390/microorganisms14061173

Authors: Varvara P. Pavlova Artem G. Chebotarevskii Ecaterina V. Diuvenji Nadezhda A. Loginova Anna M. Mosolova Aleksandra S. Novikova Sergey V. Martyanov Marina V. Sukhacheva Alexander I. Netrusov Andrei V. Gannesen

This pilot study investigated the effects of estradiol and clindamycin on mono- and dual-species biofilms of selected reference and clinical isolates of Lactobacillus gasseri and Cutibacterium acnes, including one vaginal isolate of C. acnes. Our findings demonstrate complex, strain-dependent effects of both compounds and their combinations. Estradiol inhibited biofilm formation in L. gasseri strains but exhibited divergent impacts on C. acnes isolates, stimulating the skin-derived strain while suppressing the vaginal isolate. The observation that pre-adsorbed estradiol tended to enhance its biological activity is consistent with, though does not prove, the hypothesis of a direct hormonal interaction with the bacterial cell envelope. Crucially, estradiol modulated the susceptibility of both species to clindamycin. At the working concentration selected, clindamycin susceptibility varied considerably between strains, with the antibiotic stimulating biofilm growth in skin-derived C. acnes HM514 biofilms. In dual-species communities, an apparent inversion of clindamycin activity was observed, suggesting that estradiol may alter antibiotic efficacy in a manner dependent on community composition and strain identity. Furthermore, while transcriptional changes in bacteriocin genes were evident under hormonal and antibiotic pressure, these shifts did not consistently correlate with observed phenotypic antagonistic activity. These results underscore the limitations of traditional mono-species assays and highlight the importance of considering hormonal background, community context, and the substantial phenotypic variability among individual microbial isolates when evaluating antimicrobial interventions.

Microorganisms doi: 10.3390/microorganisms14061172

Authors: Jianrong Liu Yingying Song Hongruo Ma Chunlan Mao Zuoyan Chen

Anaerobic biodegradation is the most affordable method for the degradation of N,N-dimethylformamide. However, the degradation efficiency depends on the concentration. To elucidate the responses of microbial community to N,N-dimethylformamide load, microbial diversity, composition and functional changes at different concentrations of 100, 2000, and 3500 mg/L were analyzed. Results showed that as the N,N-dimethylformamide influent concentration increased from 100 to 2000 mg/L, the removal rate stabilized at 90%, whereas it decreased to ~75% at concentrations over 2000 mg/L. Microbial community diversity increased, and specialists were enriched at 3500 mg/L. Patescibacteria (42.88% and 42.90%), Bacillota (18.52% and 18.54%), and Pseudomonadota (7.13% and 7.09%) were the dominant phyla at 100 mg/L and 2000 mg/L, respectively, and Patescibacteria (16.88%) and Pseudomonadota (15.34%) were the dominant phyla at 3500 mg/L. Methylotrophic methanogeneic (Methanolobus and Methanomassiliicoccus) and syntrophic electron-donating bacteria (Clostridiumand and Trichococcus) were significantly enriched. DMF-degrading genes (fdh, rfA/nrfH, and ATPase) and methylotrophic methanogenesis genes (mcr, mta, and mtm) were significantly upregulated. Therefore, the degradation of N,N-dimethylformamide was characterized by a parallel carbon flux distribution, “methylamine-driven methanogenesis + further oxidation/integration of single-carbon intermediates”, and the nitrogen flux tended to enter a reductive nitrogen cycle characterized by retention and reuse.

Microorganisms doi: 10.3390/microorganisms14061171

Authors: Ricardo Rodríguez-Martínez Jetsi Mancilla-Rojano Sara A. Ochoa Graciela Castro-Escarpulli Ariadnna Cruz-Córdova Juan Xicohtencatl-Cortes

In this review, the virulence factors involved in enterotoxigenic Escherichia coli (ETEC) colonization and pathogenesis are analyzed, with an emphasis on colonization factors, enterotoxins and antigenic diversity as central challenges in vaccine development. ETEC remains a major cause of diarrhea worldwide, particularly in vulnerable populations. Despite extensive research, no broadly protective licensed vaccines are available largely because of antigenic heterogeneity and the limited understanding of immune correlates of protection. We identified critical knowledge gaps in antigen prioritization and host–pathogen interactions and translational limitations that have hindered vaccine success. We critically evaluated emerging platforms (including mRNA vaccines, nanoparticles, multiepitope strategies, and reverse vaccinology) for their potential to overcome variability and increase immunogenicity. We examined the roles of ecological environmental reservoirs associated with human and animal systems, in addition to antimicrobial pressure, in shaping ETEC evolution and vaccine effectiveness within a One Health framework; moreover, we propose an integrated approach that links genomic surveillance-based vaccine ecology and next-generation vaccine technologies to support adaptive immunogen design. This review provides actionable recommendations for the development of broadly protective and translationally viable ETEC vaccines from the One Health perspective.

Microorganisms doi: 10.3390/microorganisms14061170

Authors: Xinping Zhong Biwei Song Lixin Zhang Tom Hsiang Liming Ouyang Jingyu Zhang

Phosphonate natural products, characterized by a stable carbon–phosphorus (C–P) bond, represent a distinctive class of natural products with broad bioactivities and diverse applications in medicine and agriculture. Despite their potential, the field has lacked a systematic, quantitatively informed synthesis linking discovery trends to biological function and scalable production. In this review, we curate 71 reports of phosphonate natural products between 1959 and the present, primarily from bacterial sources, and analyze them across discovery modalities, bioactivity profiles, and producer taxonomy. We further summarize structure–activity relationships and consolidate engineering strategies—including metabolic engineering, synthetic biology, and biocatalytic cascades—employed to enhance phosphonate yields. Based on these analyses, we propose practical directions to accelerate the discovery of novel phosphonates, and improve their accessibility through advances in production and isolation technologies.

Microorganisms doi: 10.3390/microorganisms14061169

Authors: Kadir Burak Akgün

Pseudomonas fluorescens is a rare, environmental Gram-negative bacterium that has been rarely reported as a cause of respiratory tract infections. This paper presents a case of a 72-year-old male who developed community-acquired pneumonia due to P. fluorescens. The diagnosis was made by sputum culture and he responded to meropenem treatment. A literature search revealed three previously reported cases of P. fluorescens pneumonia. These cases primarily affected elderly male patients. All reported patients demonstrated positive clinical outcomes following appropriate antimicrobial therapy. This case highlights that although P. fluorescens is often considered a colonizer, it may act as a potential pathogen in selected clinical settings.

Microorganisms doi: 10.3390/microorganisms14061168

Authors: Changjun Zhou Yiqing Chen Ying Yu Bing Liu Jidong Yu Yaokun Wu Jianying Li Lan Ma Gang Chen Xu Feng

According to an FAO report, the total area of saline-alkali land worldwide is approximately 954 million hectares, accounting for about 20% of global cultivated land. Saline-alkali stress significantly reduces soybean (Glycine max L.) yield and quality, and saline-alkali-tolerant plant growth-promoting bacteria (PGPB) have shown important application value for soybean planting in such farmlands. In this study, 15 strains of saline-alkali-tolerant bacteria were isolated from saline-alkali soil in Anda City, Heilongjiang Province, China, and identified morphologically, belonging to the genera Enterobacter, Bacillus, Chryseobacterium, Acinetobacter, Enterococcus, and Pseudomonas. Through tests for nitrogen fixation, phosphorus solubilization, potassium solubilization, hydrolase production (including pectinase, amylase, and protease), and germination promotion assays, Bacillus pumilus AD14 was identified as having the best growth-promoting effect on soybean seedlings. Pot experiments in saline-alkali soil showed that AD14 significantly promoted soybean seedling growth, increasing plant height by 5.63–6.37 cm and root length by 3.58–3.99 cm compared to the control. AD14 also enhanced saline-alkali tolerance by improving the activity of antioxidant enzymes including superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT) and increasing soluble sugar and protein contents. Meanwhile, soil pH decreased by 10.94–12.15% and soluble salt content decreased by 9.59–13.39% after planting, and soil enzyme activities (including urease, sucrase, and catalase) increased markedly. These results demonstrate the great potential of AD14 for soybean planting in saline-alkali soil. This study provides a relevant reference for enriching the resources of saline-alkali-tolerant PGPB and developing new biological agents suitable for soybean planting in saline-alkali soils.

Microorganisms doi: 10.3390/microorganisms14061167

Authors: Zeyu Pan Jian Bao Xiangdong Liu Gentu Ge Muqier Zhao

Rhizosphere microorganisms are important components of grassland ecosystems, but the rhizosphere microbiome of the poisonous and medicinal plant Stellera chamaejasme L. remains poorly characterized. In this study, shotgun metagenomic sequencing was used to compare the taxonomic composition, community structure, differentially enriched taxa, and KEGG-based functional potential of rhizosphere microbial communities associated with S. chamaejasme from three typical steppe regions in Inner Mongolia. Acidobacteria, Proteobacteria, and Actinobacteria were the dominant phyla, while Sphingomonas, Bradyrhizobium, and Streptomyces were among the dominant genera. Genus-level profiles and ordination analysis showed region-associated community patterns, and rarefaction curves indicated that sequencing depth was sufficient to capture most detectable taxa. LEfSe analysis identified region-associated differentially enriched taxa, including Sphingomonas-, Bradyrhizobium/Nitrospira-, and Streptomyces/Solirubrobacter-associated taxa. KEGG annotation suggested broadly similar major functional categories across regions, with some differences in the relative abundance of metabolic pathways. These results provide baseline metagenomic information on S. chamaejasme rhizosphere communities. Because of the limited replication and lack of soil physicochemical measurements, ecological mechanisms should be tested in future studies.

Microorganisms doi: 10.3390/microorganisms14051166

Authors: Shiqi Zhang Guo Li Ensheng Zhu Yu Zhao Xiaoying Yang Suzhen Huang Zheng Zheng

Rhizosphere microorganisms play central roles in nutrient cycling and contaminant transformation in sediment-associated freshwater systems, yet their responses to newer pesticides remain insufficiently characterized. In this study, a 28-day Vallisneria natans–rhizosphere sediment microcosm was used to compare the effects of trifluenfuronate and fluopyram at nominal concentrations of 0.01, 0.1, and 1 mg L−1. Bacterial community composition was assessed using 16S rRNA gene sequencing, and shotgun metagenomic data were used to evaluate relative functional annotation patterns. Plant physiological traits and rhizosphere sediment enzyme activities were measured as ecological context for interpreting microorganism-associated responses. Fluopyram, particularly at 1 mg L−1, produced clearer ordination-level shifts in rhizosphere bacterial community composition than trifluenfuronate, although pairwise treatment separation was not statistically resolved after multiple-testing correction. Annotation-based metagenomic profiles also differed between the two pesticides: stronger exposure was associated with reduced relative signals for several xenobiotic-, transport-, and regulation-related annotations, while high-dose fluopyram showed a methane-metabolism-related annotation signal and high-dose trifluenfuronate showed relative enrichment of secondary-metabolism-related annotations. These microbial and annotation-profile responses coincided with stronger inhibition of V. natans growth and greater suppression of rhizosphere sediment enzyme activities under fluopyram exposure. Overall, fluopyram induced more consistent microorganism-associated response patterns than trifluenfuronate in the tested rooted macrophyte–sediment microcosm. The results highlight the sensitivity of rhizosphere microbial communities and metagenomic annotation profiles to pesticide exposure in sediment-associated freshwater systems.

Microorganisms doi: 10.3390/microorganisms14051165

Authors: Luana Granja Jorge Carlos Santamaría José Pintado Camino Gestal Gonzalo Del Olmo

The aquaculture of Octopus vulgaris faces high larval mortality, mainly due to nutritional limitations and susceptibility to pathogens, particularly Vibrio spp. As vaccination is not feasible in cephalopods, host-associated probiotics represent a promising and sustainable alternative to improve survival and reduce infections. This study evaluated bacteria from the Roseobacter clade as probiotic candidates during octopus embryonic and paralarval stages. Characterization of egg-associated microbiota revealed the absence of cultivable bacteria within eggs and a significantly lower bacterial load on egg surfaces under maternal care, highlighting the regulatory role of female cleaning behavior. No bacteria with antagonism against Vibrio lentus, a common pathogen to octopus, were isolated from egg surfaces. Therefore, selected Roseobacter clade strains were screened in vitro against relevant aquaculture pathogens. Phaeobacter strains showed strong inhibitory activity against Vibrio spp., including V. lentus, while Ruegeria strains exhibited higher specificity against Tenacibaculum maritimum. Based on these results, Phaeobacter sp. 4UAC3 was selected for in vivo assays. This strain successfully colonized eggs, water, and paralarvae; however, its application reduced hatching success in eggs by 33%, likely due to surface-associated accumulation of the bacteria linked to the administration method. In contrast, probiotic treatment significantly improved survival at the paralarval stage. Although high variability was observed, probably due to stressful rearing conditions, more than 50% was observed in treated vs. 0% in non-treated cases at day 6. Overall, Phaeobacter sp. 4UAC3 emerges as a promising probiotic candidate to improve O. vulgaris paralarvae survival, potentially contributing to solving this bottleneck in a sustainable way.

Microorganisms doi: 10.3390/microorganisms14051164

Authors: Yaqi Liu Peng He Dongxin Liu Yang Song Chenxi Jia Duochun Wang Qinghua Jin Gang Song Qiang Wei

In the original publication [...]

Microorganisms doi: 10.3390/microorganisms14051163

Authors: Xiaoling Ma Shanshan Nan Li Zhang Yuyang Xue Wenju Zhang

Salvia sclarea L. extract contains various bioactive components such as flavonoids and fatty acids, exhibiting anti-inflammatory, antioxidant, and antibacterial properties. This study aimed to investigate the effects of Salvia sclarea L. extract on the gut microbiota and serum metabolome in lambs. Sixty 2-month-old Chinese Merino female lambs (body weight 20 ± 2 kg) were randomly assigned to five groups. The control (CK) group received the basal diet only, while the treatment groups received the basal diet supplemented with 0.04 mL/kg (CL1), 0.08 mL/kg (CL2), 0.12 mL/kg (CL3), and 0.16 mL/kg (CL4) of Salvia sclarea L. extract, respectively. The results showed that Firmicutes, Bacteroidetes, Spirochaetes, and Proteobacteria were identified as the dominant phyla across all groups (>90%). Compared with the CK group, CL1 and CL2 groups significantly reduced the relative abundance of Tenericutes (decreased by 38.2% and 32.9%, respectively, p < 0.05); the relative abundance of Patescibacteria in the CL1 group was significantly lower (decreased by 55.2%, p < 0.05). At the genus level, Ruminococcaceae constituted a substantial proportion, including Ruminococcaceae UCG-005, UCG-010, UCG-014, and NK4A214 group. STAMP analysis revealed that Klebsiella was significantly enriched in CL2, CL3, and CL4 groups compared to the CK group (p < 0.05). Correlation analysis between microbiota and immune indices showed that Christensenellaceae R-7 group was significantly negatively correlated with TNF-α (p < 0.05); Ruminococcaceae UCG-005 was significantly negatively correlated with IFN-γ (p < 0.05) and showed a negative correlation trend with immunoglobulins (IgA, IgG, IgM). Conversely, Ruminococcaceae UCG-014 was significantly positively correlated with IL-4 (p < 0.05) but showed a negative correlation trend with IgM. Untargeted metabolomics analysis identified 8, 18, 25, and 20 differential metabolites in CL1, CL2, CL3, and CL4 groups, respectively. Notably, 3-hydroxy-7-methoxyflavone and Gamma-Glu-Cys were significantly upregulated across all treatment groups. KEGG pathway enrichment analysis indicated that these differential metabolites were primarily involved in nucleotide metabolism, fatty acid biosynthesis, and oxidative stress-related pathways. Further Spearman correlation analysis revealed significant associations between gut microbiota and differential metabolites. Specifically, g_Klebsiella was significantly positively correlated with 3-Hydroxycapric acid and 3-hydroxy-7-methoxyflavone (p < 0.05). In conclusion, Salvia sclarea L. extract modulates host energy metabolism by regulating nucleotide metabolism and fatty acid biosynthesis, and enhances immune function by alleviating oxidative stress, through the remodeling of gut microbiota and serum metabolome.

Microorganisms doi: 10.3390/microorganisms14051162

Authors: Mengshi Zhao Hongyang Zhao Peimin Li Song Peng Fengqiang Lin Quanwang Wu Phurbu Tashi Zhaolong Li

Age-related changes in the gut significantly impact host health, yet the multi-omics dynamics during the maturation of Tibetan pigs remain unclear. This study aimed to investigate the morphological, microbial, metabolic, and transcriptomic transformations in the intestines of aging Tibetan pigs. We analyzed the ileum and colon of 1-year-old and 3-year-old Tibetan pigs using histological evaluation, 16S rRNA sequencing, metabolomics, and transcriptomics. Aging to 3 years significantly improved ileal architecture, notably increasing the villus height to crypt depth ratio. Older pigs exhibited higher colonic microbial diversity, a decreased Firmicutes to Bacteroidota ratio, and enrichment of homeostasis-associated taxa, including Lactobacillus, Prevotellaceae, and Ruminococcaceae. Metabolomics revealed higher abundance of certain metabolites, including docosahexaenoic and arachidonic acids, enriching lipid metabolism and bile secretion pathways. Transcriptomics identified 2363 differentially expressed genes in the ileum, primarily involved in immune regulation and nutrient digestion. Integrated analysis showed strong positive correlations between enriched microbes (Lactobacillus porci) and up-regulated host genes (UGT2B31, CCL28) governing intestinal homeostasis. The transition from 1 to 3 years of age in Tibetan pigs fosters a synergistic host-microbiome environment, enhancing intestinal barrier function, immune capacity, and metabolic efficiency.

Microorganisms doi: 10.3390/microorganisms14051161

Authors: Bocheng Gao Jiale Yi Linna Gai Jing Liu Xuan Min Ju Yao Mingzhi Li Jiarong Liu Yule Chen Su Wu Yunzi Hu Lingbao Kong

This study aimed to express the canine parvovirus (CPV) VP2 protein prokaryotically and develop an indirect ELISA for detecting CPV-specific antibodies in canine serum. The VP2 gene from a laboratory-isolated CPV strain was amplified and cloned into the pET-28a vector. Following prokaryotic expression optimization, the recombinant protein was purified via Ni-NTA affinity chromatography and validated using Western blotting. An indirect ELISA was established utilizing the purified VP2 as the coating antigen, with optimal parameters determined by checkerboard titration. A 1773 bp VP2 fragment was amplified. Optimal expression of the 64.8 kDa recombinant VP2 was achieved with 2 mmol/L isopropyl β-D-thiogalactoside (IPTG) at 32 °C for 8 h. For the indirect ELISA, the optimal antigen coating concentration was 2 μg/mL, alongside primary (canine serum) and secondary antibody dilutions of 1:320 and 1:4000, respectively. The diagnostic cut-off optical density at 450 nm (OD450) threshold was established at ≥0.2066, and the analytical sensitivity reached a serum dilution of 1:5120. Compared with the hemagglutination inhibition (HI) assay using 192 clinical serum samples, the ELISA showed a diagnostic sensitivity of 85.94%, a diagnostic specificity of 88.28%, and an overall agreement rate of 87.50%. The mean intra-assay and inter-assay coefficients of variation were 4.39% and 3.02%, respectively. These findings indicate that the recombinant VP2-based indirect ELISA showed good analytical sensitivity, reproducibility, and diagnostic agreement with the HI assay for detecting CPV-specific antibodies in canine serum under the tested conditions, although broader cross-reactivity validation is still required.

Microorganisms doi: 10.3390/microorganisms14051160

Authors: John A. Kyndt Kristina O. Petrova Stepan V. Toshchakov Irina A. Bryantseva

Heliobacteraceae are unique endospore-forming photosynthetic bacteria that are known for possessing the simplest photosynthetic apparatus of any known organism. More genomic and physiological analysis is needed to further understand the evolution of photosynthesis and the unique metabolic pathways of nitrogen and sulfur metabolism in this family. Here, we present the genome and phylogenetic analysis of “Heliomicrobium sulfidophilum” strain BR4T, which was isolated previously from an alkaline sulfide-containing hot spring. In addition to the presence of a Type I reaction center, genes for bacteriochlorophyll g synthesis and nitrogenase system, the genomic analysis also explains the need for biotin as a supplied growth factor in Heliomicrobium species. The Heliobacteriaceae genome comparison also revealed a previously unidentified gene cluster of heterodisulfide reductase-like proteins (Hdr genes) and molybdopterin-based enzymes for polysulfide reductase. The whole-genome comparison, including ANI, dDDH, and single-gene phylogenetic analyses, confirms the correct placement of strain BR4 in the Heliomicrobium genus and strengthens the overall phylogenetic distribution of the Heliobacteriaceae.

Microorganisms doi: 10.3390/microorganisms14051159

Authors: Nelly O. Elshafie Rebecca P. Wilkes

Respiratory disease remains one of the mostly costly challenges in the U.S. swine industry and is frequently associated with polymicrobial infections. Routine qPCR assays are highly sensitive but are limited in multiplexing capacity and generally do not provide sequencing information for pathogen characterization. We hypothesized that a target next-generation sequencing (tNGS) panel could provide the broad, simultaneous detection of swine respiratory pathogens while preserving clinically relevant sensitivity. A multiplex Ion Torrent tNGS panel was developed and analytically validated using 20 serially diluted qPCR-positive clinical samples and synthetic gBlock controls, followed by diagnostic validation with 25 qPCR positive and 25 qPCR negative respiratory samples. Most targets were detected across clinically relevant pathogens concentrations. Actinobacillus suis primers showed nonspecific amplification, streptococcus suis serotyping was not consistently achievable in clinical samples, and porcine reproductive and respiratory syndrome virus typing was limited to distinguishing North American and European genotypes. Diagnostic agreement with routine qPCR was high (Cohen’s κ = 0.84), although sensitivity decreased for low-abundance targets. The assay detected mixed infections and additional organisms outside routine qPCR panels. These findings support tNGS as a complementary diagnostic and surveillance tool for swine respiratory disease.

Microorganisms doi: 10.3390/microorganisms14051158

Authors: Andrew C. Schuerger Petra Schwendner Lisa Guan Jerami Mennella Nicholas Heinz Ioannis Mikellides Brian G. Clement

Mars Sample Return Program planning includes a series of spacecraft staged both on the Martian surface and in low-Mars-orbit (LMO). During the transfer of samples into orbit, external spacecraft surfaces might be exposed to Mars dust carried on the sample container exterior and possibly extant microbiota (if present). This study was designed to characterize the synergistic effects of LMO ultraviolet irradiation, vacuum, and solar heating on the survival of two UV-resistant and heat-tolerant bacteria, one yeast, and one fungus. The species tested were Bacillus pumilus SAFR-032 spores, Geobacillus stearothermophilus ATCC 12980 spores, Naganishia onofrii DBVPG 5303 cells, and Aspergillus fumigatus ISSFT-021-30 spores, respectively. Spores of A. fumigatus ISSFT-021-30 and B. pumilus were also exposed to LMO conditions with and without a Mojave Mars Simulant (MMS) dust layer. Based on the data, the time required to reach the desired Sterility Assurance Level (SAL; dose-defined to yield a −12 log reduction) was 2.0 h for A. fumigatus ISSFT-021-30 and 76.6 min for B. pumilus SAFR-032 if exposed directly to the solar UV beam under LMO conditions. With the MMS present, predicted times to reach one SAL were extended to 22 h and 1.72 h, respectively. Analysis of UV transmittance through cell stacks of up to 12 µm thick was performed for A. fumigatus ISSFT-021-30. Results indicated that ~4–5% of UVC photons can penetrate through 12 µm stacked aggregates of spores. These findings indicate that (1) the LMO environment can be used to attain the mandated levels of spacecraft surface bioburden reductions and (2) dust shielding and microbial aggregation attenuate UV irradiation, leading to extended orbital residence times to achieve mandated bioburden reductions.

Microorganisms doi: 10.3390/microorganisms14051157

Authors: Aditi Singh Sumeeta Kumari Gunjan Vasudeva Mohit Kumar Saini Anil Kumar Pinnaka Karel Kopejtka Michal Koblížek Nupur

A novel photoheterotrophic, beige-pigmented, bacteriochlorophyll a-containing strain KR11T was isolated from Kaprový pond in Třeboň, Czechia. KR11T cells were Gram-negative, rod-shaped, and motile. The isolated strain grew under photoheterotrophic conditions between 20 and 40 °C (optimum 22–25 °C), at pH ranges from 6.0 to 9.0 (optimum 7.0). It did not require NaCl for growth but tolerated NaCl concentrations up to 1.5% (w/v). No growth was observed under photoautotrophic conditions. Strain KR11T showed the highest 16S rRNA gene sequence similarity to the type strains of Stagnihabitans tardus CYK-10T (98.84%), Tabrizicola fusiformis SY72T (95.95%), and Rhodobacter sediminis N1T (95.37%). Phylogenetic analyses based on 16S rRNA gene sequences indicated that strain KR11T clusters within the genus Stagnihabitans in the family Paracoccaceae of class Alphaproteobacteria. The whole-genome sequence of strain KR11T comprises 4,085,976 bp with a 65 mol% G+C content. Phylogenomic analysis, including core-genome phylogeny, and the low genomic similarity (<95% ANI and <70% dDDH) to phylogenetically related taxa confirmed the taxonomic separation of strain KR11T at the species level. The distinctive phenotypic traits, chemotaxonomic studies, phylogenetic, and genomic analysis establish strain KR11T as a novel species within the genus Stagnihabitans. Accordingly, we propose the name Stagnihabitans lacustris sp. nov. KR11T (=CCUG 74777T, LMG 31924T), isolated from fresh water.

Microorganisms doi: 10.3390/microorganisms14051154

Authors: Mingzhu Zhu Edmond Massuda Shane Cloney-Clark Urvashi Patel Anand Parekh Andrew Gorinson Andrew Klindworth Ali Ahmadi Miranda R. Cai Chijioke Bennett Raj Kalkeri Joyce S. Plested

Mucosal immunity, including antibodies like immunoglobulin A (IgA), function as the body’s first line of defense in the respiratory tract, particularly against viruses. An anti-rS protein IgA enzyme-linked immunosorbent assay (ELISA) was developed using the Omicron XBB.1.5 subvariant of SARS-CoV-2 and was validated to demonstrate the suitability of the method for testing saliva from SARS-CoV-2 vaccine clinical trials. This assay successfully met acceptance criteria for inter-/intra-assay precision, specificity, selectivity, linearity, lower/upper limits of quantitation, and assay robustness. The IgA in saliva was stable for up to 7 freeze/thaw cycles, for up to 48 h at 24 °C, up to 7 days at 4 °C, up to 3 weeks at −20 °C, and up to 1 year at −80 °C. After validation using Omicron XBB.1.5 rS protein, cross-reactivity was demonstrated with the SARS-CoV-2 variant JN.1. This validated IgA assay can be a valuable tool to assess mucosal IgA levels in SARS-CoV-2 clinical trials.

Microorganisms doi: 10.3390/microorganisms14051155

Authors: Zi-Jun Li Ling-Jiao Yu Ya-Xin Yang Ying Li Emad Beshir Ata Yang Zhou Rong-Rong Zhang Yi-Bing Lian Hong-Liang Chen Chun-Wei Shi Gui-Lian Yang Hai-Bin Huang Yan-Long Jiang Jian-Zhong Wang Xin Cao Nan Wang Yan Zeng Wen-Tao Yang Chun-Feng Wang

Salmonellosis is a global foodborne pathogen with zoonotic importance that seriously threatens livestock breeding and human health. Due to the implementation of an anti-resistance policy, probiotics as an alternative to antibiotics have attracted widespread attention. In this study, the widely used probiotic Escherichia coli Nissle 1917 (EcN) was selected to study its protective effect on mice infected with Salmonella typhimurium. Two mice groups (n = 15) were treated with either EcN and PBS. Flow cytometry showed that the frequency of mature dendritic cells in the Peyer’s patch was significantly increased compared to the PBS group. Previous administration of EcN protected against challenge with Salmonella typhimurium infection as an increased survival rate of the mice, a decreased degree of pathological changes, and the number of live bacteria in the spleen and liver were recorded compared to the control group. The results of 16S rRNA high-throughput sequencing of fecal microbial flora showed that EcN could reduce the abundance of microorganisms in the intestine and reduce the proportion of Lactobacillus, while Ruminococcaceae sp., Rikenella sp. and Bifidobacterium sp. disappeared. In contrast, the abundance of Bacteroides increased, which reduced the effect of Salmonella typhimurium on the distribution of intestinal microorganisms. Our results demonstrated that EcN has a protective effect against Salmonella typhimurium infection and may act as a candidate probiotic bacterium to apply in the future.

Microorganisms doi: 10.3390/microorganisms14051156

Authors: Isabel S. Godinho Gonçalo Queirós Lesya Yefimenko Filomena M. Pereira João Piedade

Angola is one of the countries with the highest HIV-1 genetic diversity, yet the implications of this diversity for antiretroviral therapy remain insufficiently characterised. Following the introduction of dolutegravir (DTG) in Angola in 2021, evaluating transmitted drug resistance prior to its widespread implementation is essential to inform treatment strategies and establish a baseline for future surveillance. In this study, 243 blood samples were collected from treatment-naïve people living with HIV attending the General Hospital of Benguela, Angola. The integrase coding region of proviral DNA was amplified and sequenced using the Sanger method. Phylogenetic relationships were inferred using a maximum likelihood approach, recombinant forms were characterised by bootscanning analysis, and resistance-associated mutations to integrase strand transfer inhibitors were identified using Stanford HIVdb, ANRS-MIE, and IAS-USA algorithms. A total of 92 integrase sequences were successfully obtained, revealing 16 distinct genetic forms, with unique recombinant forms accounting for 50.0%, followed by subtype C (10.9%) and sub-subtype F1 (8.7%). Five accessory mutations (L74I, L74M, Q95K, T97A, and E157Q) and one major mutation (E92G) were detected, corresponding to an overall prevalence of 28.8% (23/80). These findings highlight the extensive HIV-1 genetic complexity in Angola and support the continued use of DTG-based regimens, while underscoring the importance of sustained surveillance of integrase inhibitor resistance.

Microorganisms doi: 10.3390/microorganisms14051153

Authors: Aylin López-Palestino Natali Hernández-Parada Zorba Josué Hernández-Estrada Oscar González-Ríos Olaya Pirene Castellanos-Onorio Rodrigo Alonso-Villegas Aztrid Elena Estrada-Beltrán Mirna Leonor Suárez-Quiroz Claudia Yuritzi Figueroa-Hernández

Yeasts have attracted increasing attention as potential alternatives to traditional bacterial probiotic strains due to their physiological resilience and functional versatility. However, the probiotic potential of yeast strains associated with Mexican cocoa bean fermentation remains largely unexplored. Therefore, this study aimed to conduct a preliminary screening of physiological and surface-related traits associated with probiotic functionality in four autochthonous Saccharomyces cerevisiae strains (YCTA5, YCTA9, YCTA14, and YCTA16), previously isolated from cocoa fermentation, using Saccharomyces boulardii (Jarrow Formulas®) as a reference strain. Evaluated parameters included tolerance to temperature, pH, and bile salts; hemolytic activity; survival in vitro under gastrointestinal (GI) conditions; bile salt hydrolase activity; auto-aggregation; co-aggregation; hydrophobicity; and response to antifungal agents (fluconazole, ciclopirox, nystatin, and clotrimazole). All yeast strains grew at 37 °C and at pH 4–8 and showed no hemolytic activity. All strains exhibited high auto-aggregation (>70%) and hydrophobicity values ranging from 55 to 88%. In the coaggregation assay, strains YCTA9, YCTA14, and YCTA16 showed moderate interactions with Escherichia coli, Bacillus cereus, and Listeria innocua, with some combinations exceeding 50%. Nevertheless, none of the yeast strains exhibited measurable growth at pH 2; bile salt tolerance was limited to 0.1% Oxgall, and viability decreased by approximately 54–56% after simulated gastrointestinal transit. These findings indicate that although some strains exhibited promising surface-related properties, significant physiological constraints restrict their probiotic potential under the tested conditions. Therefore, the studied yeast strains should be regarded as preliminary candidates requiring further validation. This work provides a first-stage evaluation for identifying functional yeast strains from Mexican cocoa bean fermentation, serving as a basis for future in vitro and in vivo studies.

Microorganisms doi: 10.3390/microorganisms14051152

Authors: Ji Sun Park Woo Sik Kim Jaehoon Bae Jinseok Jung Ji-Young Park Hyung Jae Jeong Woo Song Lee Su-Jin Park

Influenza A virus (IAV) remains a major global health threat despite available vaccines and antiviral agents, while current therapies are limited by drug resistance and safety concerns. Curcuminoids exhibit antiviral and anti-inflammatory activities but are constrained by poor water solubility and low bioavailability. To address these limitations, we investigated the antiviral and immunomodulatory properties of a water-solubilized curcuminoid nanoparticle formulation (C–S/M) in both in vitro and in vivo models of IAV infection. To evaluate the potential antiviral and anti-inflammatory effects of C–S/M, we performed a cytopathic effect (CPE) reduction assay in triplicate at 0.001 MOI and quantitative real-time PCR (qRT-PCR) targeting viral NS1 transcripts in MDCK cells. C–S/M suppressed viral NS1 vRNA levels in MDCK cells at lower curcuminoid-equivalent concentrations than native curcuminoids and attenuated IAV-induced TNF-α, IL-6, and IL-8 production. Furthermore, in vivo antiviral efficacy was evaluated in female C57BL/6 mice intranasally infected with IAV and treated orally with C–S/M. Survival, lung viral loads, pulmonary cytokine levels, and splenic immune cell phenotypes were analyzed. In IAV-infected mice, oral administration of C–S/M modestly improved survival and significantly reduced lung viral burden and pulmonary proinflammatory cytokine levels. In addition, in vivo C–S/M treatment was associated with recovery of virus-suppressed T-cell immune responses, including increased Th1 and activated CD8+ T cells, reduced regulatory T-cell expansion, and restoration of multifunctional CD4+ and CD8+ T cells. These findings suggest that C–S/M exerts antiviral and immunomodulatory effects in experimental IAV infection and may serve as a potential adjunctive candidate for further investigation against influenza-associated inflammation.

Microorganisms doi: 10.3390/microorganisms14051151

Authors: Zhen Hu Yifan Rao Lu Liu Zuwen Guo Yuting Wang Weilong Shang Huagang Peng

The emergence of vancomycin-intermediate Staphylococcus aureus (VISA) threatens the efficacy of this last-line antibiotic. The GraSR two-component system is frequently mutated in VISA strains. Here, we demonstrate that the GraS(T136I) point mutation, identified in the clinical VISA isolate XN108, is a key determinant of reduced vancomycin susceptibility. Introducing this mutation into the susceptible strain Newman increased the vancomycin MIC from 1.5 to 4 mg/L, while its reversion in XN108 decreased the MIC from 12 to 8 mg/L. The mutation conferred common phenotypes, including thickened cell wall, decreased autolysis, and reduced cell surface negative charge via upregulation of the dltABCD operon and mprF. Notably, the GraS(T136I) mutation also upregulated virulence genes (efb, hlb, sbi, hld) and enhanced hemolytic activity. Interestingly, despite this hypervirulent profile, the mutant showed impaired long-term survival within macrophages. Our study reveals that a single GraSR mutation can co-regulate vancomycin resistance and virulence, offering new insights into the adaptation of S. aureus to antibiotic pressure.

Microorganisms doi: 10.3390/microorganisms14051150

Authors: Himani Darangwal Bhawna Vyas Munesh Kumari Ojal Bansal Shanmugam Mayilraj Venkata Ramana Vemuluri

Gram-stain-positive, endospore-producing, mesophilic and rod-shaped strain O16T was isolated from a copper mine’s soil and characterized using a polyphasic taxonomic approach. The 16S rRNA gene-sequence analysis revealed that strain O16T belongs to the genus Saccharibacillus. It exhibited the highest sequence similarity to Saccharibacillus endophyticus JM-1350T (97.2%), followed by ‘Saccharibacillus alkalitolerans’ VR-M41T (97.1%), Saccharibacillus sacchari GR21T (96.8%), Saccharibacillus kuerlensis HR1T (96.6%), and Saccharibacillus deserti WLJ055T (95.7%). Genome-based comparisons revealed that the digital DNA–DNA hybridization (dDDH) and average nucleotide identity (ANI) values between strain O16T and its closest relatives, S. endophyticus JM-1350T and ‘S. alkalitolerans’ VR-M41T, were 21.3% and 22.3%, and 76.6% and 77.6%, respectively, which are well below the recommended thresholds for species delineation. The diagnostic diamino acid of the cell wall was meso-diaminopimelic acid. Phosphatidylglycerol and diphosphatidylglycerol were the major polar lipids in strain O16T. The predominant menaquinone was MK-7. The DNA G+C content was 53.4%. The major cellular fatty acids present were anteiso-C15:0 (60.8%), iso-C16:0 (9.5%) and C16:1 ω11c (7.4%). On the basis of phenotypic, chemotaxonomic, and genotypic evidence, strain O16T is considered to represent a novel species within the genus Saccharibacillus. This data strongly supports the classification of the strain O16T as a novel species in the genus Saccharibacillus, for which we propose the name Saccharibacillus soli sp. nov. strain O16T (=CCM 8781T = KCTC 33898T).

Microorganisms doi: 10.3390/microorganisms14051148

Authors: Liting Zhou Yan Yang Li Kang Jiayi You Ye Wang Ailing Xu Guangmin Tu Rui Huang Zhengyu Zhou Minghui Li Shuyan Wu

Salmonella infects a wide range of hosts, causing gastroenteritis or systemic infection in humans and animals, highlighting the urgent need for a deeper understanding of its pathogenesis. SpvC, a critical virulence determinant of salmonella, facilitates bacterial dissemination. Gasdermin D (GSDMD) is the only gasdermin known to protect mice against acute Salmonella enteritis. Our preliminary findings indicated that SpvC counteracts GSDMD-mediated antibacterial effects to enhance bacterial dissemination, although its functional relevance to epithelial-derived GSDMD and the underlying mechanisms remain unclear. To address this, Gsdmd−/− C57BL/6J and wild-type mice were infected with Salmonella Typhimurium (S. Typhimurium) wild-type strain and spvC deletion mutant. Our results demonstrate that SpvC compromises intestinal epithelial barrier integrity, overcoming GSDMD-mediated protection against systemic infection. Specifically, through bioinformatics analysis, LC-MS/MS, and in vivo experiments with Caco-2 cell monolayers and site-directed spvC mutants, we identified SEC23B as a novel target of SpvC. This interaction disrupts the intestinal epithelial barrier through the autophagy–pyroptosis pathway. This study identifies SEC23B as a unique cellular target of SpvC involved in GSDMD activation during S. Typhimurium systemic infection. It also reveals a novel mechanism by which Salmonella evades host defense mechanisms.

Microorganisms doi: 10.3390/microorganisms14051149

Authors: Ajay Badhan Chunli Li Le Luo Guan Tim A. McAllister

Efficient utilization of lignocellulosic biomass by the rumen microbiome is critical for improving feed efficiency in ruminants, yet the development of stable, functionally specialized microbial consortia remains limited. This study aimed to assemble substrate-adapted rumen microbial consortia using an ecology-guided enrichment approach. Rumen fluid collected from cannulated Angus × Hereford heifers was sequentially enriched over 10 generations on four substrates with distinct cell wall characteristics: alfalfa, barley straw, carboxymethyl cellulose (CMC), and xylan. Fermentation parameters, including gas production and volatile fatty acids (VFAs), and bacterial community dynamics were analyzed, and selected consortia (alfalfa and xylan) were evaluated for stability following one month of cryopreservation. Across enrichments, total VFA concentrations declined (e.g., xylan: 109.8 mM (G0) to 56.37 mM (G10)), accompanied by reduced gas production and decreased alpha diversity, indicating substrate-driven selection. Distinct functional profiles emerged, including increased propionate in alfalfa consortia, higher acetate in barley straw, lactate–propionate cross-feeding with CMC, and caproate production (6.3 mM at G10) in xylan enrichments associated with Caproiciproducens and Megasphaera. Cryopreserved consortia retained core community structure and fermentation characteristics upon revival. These results demonstrate that substrate-driven enrichment can generate stable, functionally specialized rumen consortia and provide a framework for developing ecologically compatible microbial communities with potential applications in improving rumen fermentation efficiency.

Microorganisms doi: 10.3390/microorganisms14051147

Authors: Bing Yang Zhihe Zhang Yue Liu Zhidi Wang Yuanlan Sheng Zhisong Yang

Soil nematodes are integral to soil micro-food webs and serve as sensitive bioindicators of soil ecological condition. However, how forest vegetation and soil properties interact to shape nematode community assembly, network structure, and functional stability remains inadequately understood. Using 18S rRNA gene amplicon sequencing coupled with phylogenetic null modeling, we examined soil nematode communities across four forest types along a succession gradient. Although taxonomic diversity (e.g., Shannon and Pielou indices) differed significantly among forest types, network topology and stochastic assembly processes were more closely associated with community restructuring and co-occurrence patterns. This suggests that, while diversity is not irrelevant, network- and assembly-based metrics provide complementary and often more sensitive indicators of nematode community responses to forest type. Co-occurrence network analysis revealed that mixed forests fostered more complex and potentially stable networks, whereas plantations formed dense but potentially vulnerable networks. Assembly processes were not dominated by strong deterministic selection (|βNTI| ≤ 2 for most comparisons), a pattern consistent with undominated processes (e.g., ecological drift, weak environmental filtering). Dispersal limitation played a negligible role in this system. Partial Least Square Path Modeling identified spatial factors and key soil properties (e.g., pH, electrical conductivity, soil water content, and organic carbon) as primary drivers of community structure. Our findings indicate that assessing soil food web health should integrate network analysis and stochasticity metrics rather than rely solely on taxonomic diversity. For sustainable forest management, mixed-species stands are preferable to monoculture plantations, and maintaining soil physicochemical heterogeneity is critical for community stability.

Microorganisms doi: 10.3390/microorganisms14051146

Authors: Xiaolong Cao Xiangrui Zhu Hao Lin Lingyu Guo Hua Shui Zhen Wang Enci Shen Zegao Guo Ruizhe Zhang Xin Li

This study systematically evaluated the modulatory effects of Brassica rapa L. polysaccharides (BRP) on intestinal mucosal injury in a CTX-induced mouse model. The results showed that high-dose BRP (HBRP) significantly alleviated oxidative damage, with CAT activity increased by approximately 2-fold, SOD increased by 1.3-fold, GSH-Px increased by 70%, and MDA levels decreased by 65%. Meanwhile, liver injury was improved, as ALT and AST decreased by 35% and 55%, respectively. CTX markedly suppressed immune function, while BRP intervention significantly restored cytokine levels, with IL-1β increased by up to 2.5-fold, TNF-α by 1.3-fold, and IL-4 by 2.1-fold. In addition, BRP significantly modulated gut microbiota composition. The number of unique ASVs decreased from 316 in the normal control (NC) group to 63 in the model control (MC) group and recovered to 140 in the HBRP group. At the phylum level, Bacillota increased from 55% to 97% (MC) and decreased to 65% after BRP intervention, while Bacteroidota recovered to 25%. At the genus level, Candidatus_Arthromitus decreased from 40% to nearly 0%, whereas beneficial bacteria such as Ligilactobacillus and norank_f__Muribaculaceae were restored. Overall, BRP effectively alleviates CTX-induced intestinal injury in a dose-dependent manner through antioxidant, immunomodulatory, and gut microbiota–regulating mechanisms.

Microorganisms doi: 10.3390/microorganisms14051145

Authors: Abdullah A. Alshehri Jehad A. Aldali Ghina M. Alhuwaymani Farah M. Alanazi Yara K. Alsarhan Shahad A. Almutairi Abrar A. Altayeb

Ventilator-associated pneumonia (VAP) remains a major healthcare-associated infection in intensive care units (ICUs) and is associated with prolonged hospitalization, increased antimicrobial use, and high mortality. In Saudi Arabia, evidence on VAP epidemiology, microbiology, and outcomes is fragmented across settings. This study aimed to systematically review and synthesise the available evidence on VAP in Saudi Arabian ICUs. This study followed PRISMA guidelines and was prospectively registered in PROSPERO (CRD420261332740). A systematic search of PubMed, MEDLINE, Embase, and Web of Science was conducted for studies published between 2015 and 2025. Studies from Saudi Arabia reporting VAP incidence in ICUs were included. A random-effects model was used to pool incidence per 1000 ventilator-days. Risk of bias was assessed using Joanna Briggs Institute tools. Analyses were performed using R. Seven studies involving a total of 15,844 patients, representing multicentre studies, national surveillance data, and single-centre cohorts across diverse ICU settings. The pooled incidence of VAP was 8.50 episodes per 1000 ventilator-days (95% CI: 3.23–13.78), with substantial heterogeneity (I2 = 97%). Subgroup analysis showed higher incidence during baseline phases (12.46 per 1000 ventilator-days) compared with intervention phases (8.06 per 1000 ventilator-days), while surveillance estimates were lower. Gram-negative pathogens predominated, particularly Acinetobacter baumannii, often exhibiting multidrug resistance. VAP was associated with prolonged ICU stay, delayed extubation, and high mortality. Implementation of infection prevention bundles was associated with reductions in VAP incidence. Ventilator-associated pneumonia remains a significant burden in Saudi Arabian ICUs, characterised by substantial variability in incidence and a predominance of multidrug-resistant pathogens. Strengthening infection prevention measures, enhancing antimicrobial stewardship, and improving national surveillance systems are essential to reduce VAP burden and improve patient outcomes.

Microorganisms doi: 10.3390/microorganisms14051143

Authors: Lixu Li Feng Chen Guohua He Xiao Wei Feng Wang Jianmin Tang

Ormosia microphylla is a national first-class protected wild plant in China that faces conservation challenges, including weak natural regeneration and limited environmental adaptability. Plant-associated bacterial communities are important components of host-associated microecosystems, but bacterial community patterns across the whole-plant continuum of O. microphylla remain poorly understood. To provide a descriptive micro-ecological baseline, we characterized bacterial communities across the rhizosphere–root–stem–leaf continuum of O. microphylla in three geographic habitats in Southwest China: karst mountainous area, a plateau-to-plain transitional slope zone, and a hilly area. High-throughput amplicon sequencing was used to analyze bacterial diversity and composition, and co-occurrence network analysis was used to describe statistical associations among bacterial taxa. Three main patterns were observed. First, bacterial alpha diversity generally declined from the rhizosphere to internal tissues (rhizosphere > root > stem > leaf). Second, bacterial composition varied by plant compartment and habitat. Dominant rhizosphere taxa differed among habitats, whereas internal tissues were generally dominated by Proteobacteria. Delftia showed relatively high abundance in several endophytic compartments, suggesting that this genus may be considered a candidate endophytic taxon for future validation. Third, co-occurrence network analysis showed habitat- and compartment-associated differences in network size, complexity, and positive/negative co-occurrence patterns. Overall, these results describe compartment- and habitat-associated bacterial community patterns in O. microphylla and provide a micro-ecological baseline for future culture-dependent and functional studies.

Microorganisms doi: 10.3390/microorganisms14051144

Authors: Carlos J. Jiménez-Sánchez Cristopher Perez Sandra Rivera-Gutiérrez Jorge Soria-Bustos Fernando Chimal-Cázares Roberto Rosales-Reyes Santa Mejía-Ventura Gabriela Hernández-Martínez Miguel A. De la Cruz Jorge A. Yañez-Santos Maria L. Cedillo James G. Fox Miguel A. Ares

Toxigenic Klebsiella oxytoca strains linked to antibiotic-associated hemorrhagic colitis produce the cytotoxins tilimycin and tilivalline, which contribute to intestinal epithelial damage during infection. Tilimycin and tilivalline are synthesized by enzymes encoded within the nonribosomal peptide synthetase (NRPS) operon, yet the regulatory mechanisms controlling operon expression remain poorly understood. SdiA, an orphan LuxR-type quorum-sensing regulator, detects exogenous N-acyl homoserine lactones (AHLs) produced by neighboring bacterial species and modulates gene expression in response to interspecies communication. Although SdiA has been implicated in virulence regulation in several enteric pathogens, its role in K. oxytoca remains unclear. This study demonstrates that SdiA positively regulates npsA, the first gene in the NRPS operon, and that this regulatory effect is enhanced in the presence of exogenous AHL. Electrophoretic mobility shift assays indicate that SdiA directly binds to the upstream regulatory region of npsA, supporting a direct interaction consistent with positive transcriptional regulation. Furthermore, deletion of sdiA significantly reduces cytotoxicity toward HeLa cells under the conditions tested. Collectively, these findings identify SdiA as a quorum-sensing-responsive activator of npsA expression and support its role in modulating cytotoxicity in toxigenic K. oxytoca strains. These results provide new insight into the influence of interspecies quorum-sensing signals on virulence-associated regulatory pathways in K. oxytoca.

Microorganisms doi: 10.3390/microorganisms14051142

Authors: Ayman Alsheikh Raghad Shanabla Ahmad Badawi Hafez Al-Momani Mohammed Nasser-Ali Yaqeen Rjoub Mohammad A. A. Al-Najjar Montasir Al-Mansi Iman Aolymat Lana Al-Shoubaki Nawal Al-Zaa’q

Klebsiella pneumoniae is an opportunistic pathogen associated with both community-acquired and nosocomial infections. Multidrug-resistant (MDR) strains are increasingly implicated in urinary tract infections (UTIs), traveller’s diarrhoea, bacteraemia, and sepsis. β-lactam antibiotics are commonly used for treatment; however, antimicrobial resistance has emerged largely due to the production of extended-spectrum β-lactamases (ESBLs), which confer resistance mainly to penicillins, oxyimino-cephalosporins, and monobactams, while cephamycins and carbapenems usually remain stable to ESBL-mediated hydrolysis and compromise therapeutic efficacy. ESBL-producing strains represent a major cause of severe Gram-negative infections. This study aimed to determine the prevalence of ESBL-producing K. pneumoniae among UTI patients in Jordanian hospitals (Al Mafraq, Ma’an, and Islamic Hospitals), evaluate their antimicrobial susceptibility patterns, and detect antimicrobial resistance genes at the molecular level. A total of 450 urine isolates of K. pneumoniae were collected from UTI patients between November 2023 and May 2024. Isolates were identified in hospital laboratories using standard microbiological methods. Antimicrobial susceptibility testing was performed, and molecular characterisation of ESBL-associated genes was conducted using polymerase chain reaction (PCR). Out of 450 K. pneumoniae isolates collected from UTI patients across three Jordanian regions, 72 (16%) were confirmed as ESBL producers. Among the 72 ESBL-positive K. pneumoniae isolates, 34 (47.2%) were recovered from the Central region, 20 (27.8%) from the North, and 18 (25.0%) from the South. Molecular analysis revealed that 41.7% of ESBL-producing isolates carried the blaCTX-M gene, while 33.3% harboured the blaOXA gene. All ESBL-producing isolates demonstrated antimicrobial resistance to third-generation cephalosporins. A significantly higher proportion of ESBL-producing isolates was identified in female patients (84.7%) compared with males (15.3%). A significant association was observed between blaOXA gene distribution and geographic region (p = 0.016), whereas blaCTX-M gene distribution showed no significant regional association. ESBL-producing K. pneumoniae accounted for a substantial proportion of UTI isolates in Jordan, with blaCTX-M identified as the predominant resistance gene. The higher burden observed in the Central region and among female patients highlights notable distribution patterns in this cohort. These findings emphasise the necessity for sustained molecular surveillance and strengthened antimicrobial stewardship strategies to limit the dissemination of ESBL-producing strains in Jordanian healthcare settings.

Microorganisms doi: 10.3390/microorganisms14051141

Authors: Qiuxia Xiang Guijun Bu Xiaorong Tang Changwu Shi Bing Xiong Lin Wu Jia Xiong

Phenolics in Sphagnum can inhibit its microbial decomposition. Climate warming and drainage have driven vascular plants, such as Ericaceae, to expand into Sphagnum-dominated peatland. However, the impact of fine root invasion by Rhododendron auriculatum Hemsl. on Sphagnum decomposition and changes in phenolic compounds remains unclear. This study compared Sphagnum decomposition in a Sphagnum palustre L.-dominated peatland and an R. auriculatum (Sapling)–S. palustre peatland by examining the microscopic structure of S. palustre and microbial community composition. Decomposition was higher in the R. auriculatum–S. palustre peatland. On this site, bacterial metabolic types such as aerobic chemoheterotrophy and chemoheterotrophy had higher relative abundances, as did fungal trophic modes, including those with combined ectomycorrhizal, ericoid mycorrhizal, and saprotrophic functions. Acid phosphatase, laccase, total nitrogen (TN), C/N ratio (C:N), and pH differed significantly across decomposition stages. Microbial communities are affected by physicochemical factors and enzyme activities. Untargeted metabolomics revealed more downregulated than upregulated phenolics, cinnamic acids, and tannins, indicating loss of phenolic compounds. In summary, R. auriculatum fine root invasion altered enzyme activities and physicochemical properties, driving the restructuring of bacterial and fungal trophic modes and accelerating S. palustre cell wall and hyaline cell decomposition.

Microorganisms doi: 10.3390/microorganisms14051140

Authors: Zhikang Guo Mu Peng Haibo Wang

Heavy metal contamination in soils threatens ecosystem stability, agricultural productivity, and human health due to its persistence, toxicity, and ecological risks. Microbial remediation has emerged as a sustainable and cost-effective strategy, but the knowledge structure and research trends in this field remain insufficiently summarized. This study conducted a bibliometric analysis of publications on microbial remediation of heavy metal-contaminated soils retrieved from the Web of Science Core Collection from 2000 to 2025. VOSviewer (version 1.6.20), CiteSpace (version 7.0.R0), and the bibliometrix package (version 4.5.0) were used to analyze publication trends, major contributors, influential journals, and keyword evolution. The results showed that the number of publications increased continuously, with rapid growth after 2020. China, India, and the United States were the leading contributors, while Poland, Spain, and the United States played important bridging roles in international collaboration. Ravi Naidu was the most cited author, and Journal of Hazardous Materials was the most productive journal. Keyword analysis revealed a shift from pollutant degradation and microbial screening toward plant–microbe synergistic remediation, co-contaminated soil treatment, microbial community responses, and ecological risk assessment. Future research should emphasize multi-omics-based mechanisms, long-term in situ applications, and integrated evaluation frameworks.

Microorganisms doi: 10.3390/microorganisms14051139

Authors: Qi Zhou Zhenyu Huang Han Li Jiaying Xiong Meixia Chen Yan Liu Wei Liu Yanlai Yao Ramon Gonzalez Yu Li Aiqin Shi Fuping Lu

Understanding the interaction between plants and rhizosphere microorganisms is critical for the development of biofertilizers. Fluorescent labeling of rhizosphere microorganisms serves as a key strategy to track their behavior during plant–microbe coculture. However, most newly isolated strains are novel and lack available molecular tools for such studies. In this research, Planococcus dechangensis NEAU-ST10-9T (P. dechangensis NEAU-ST10-9T), a salt-tolerant strain, was obtained from the China General Microbiological Culture Collection Center (CGMCC). It significantly increased maize root length by approximately 1.56-fold. To investigate the underlying mechanism, a donor strain (Ec102) and a shuttle plasmid (pAS104) were engineered to mediate conjugation with P. dechangensis NEAU-ST10-9T and drive GFP overexpression in the bacterium, generating the genetically labeled strain Pd103. The fluorescence intensity (expressed as GFP/OD600, arbitrary units) of Pd103 increased with bacterial growth and was approximately tenfold higher than that of the wild-type strain after 16 h of culture. Following inoculation onto maize seeds, confocal microscopy analysis revealed that Pd103 colonized the epidermis and endodermis of maize roots. These results indicated that P. dechangensis NEAU-ST10-9T could invade maize roots and promote maize seedling growth. In summary, we have successfully established a robust fluorescence labeling and tracking system tailored for P. dechangensis NEAU-ST10-9T, which constitutes a valuable tool for elucidating the cellular and molecular mechanisms governing its plant–microbe interaction.

Microorganisms doi: 10.3390/microorganisms14051138

Authors: Nourhan Fouad Emad M. Elzayat Dina Amr Dina A. El-Khishin Khaled H. Radwan Alaa Youssef Abeer A. Khalaf Hoda A. Ahmed Eman H. Radwan Sawsan Tawkaz Michael Baum

Rhizosphere microbiome engineering is a promising approach that can enhance crop resilience and input use efficiency by redirecting plant–microbe–soil interactions toward predictable functions. Here, we review the mechanistic bases underlying rhizosphere assembly and stability, including root exudate-mediated selection, priority effects, keystone taxa, and metabolite-driven signaling, and connect these principles to proposed design rules for microbial inoculants. We present a generalizable Design–Build–Test–Learn (DBTL) framework for engineering synthetic microbial consortia, covering trait-to-module mapping (nutrient acquisition, phytohormone modulation, ACC deaminase activity, stress-protective metabolites, and biocontrol), compatibility screening, minimal yet robust community architectures, and iterative optimization driven by multi-omics and high-throughput phenotyping. Translation to field settings is framed as an engineering challenge defined by formulation and administration limitations, including carrier type, seed coating and encapsulation methods, shelf life, strain invasiveness, and permanence of colonization amid environmental diversity. We also summarize how integrative measurement pipelines (amplicon and shotgun sequencing, transcriptomics, metabolomics, and network or causal analyses) can advance microbiome studies from correlation to actionability. We describe how precision agriculture (sensors, remote sensing, and variable-rate inputs) and AI/ML (split-sample comparisons, transfer learning, and active learning) approaches can accelerate strain discovery, mixture optimization, and adaptive experimentation, driven by the need for stringent controls, metadata-rich reporting, and cross-site comparability. Use cases focus on stress conditions (drought, salinity, thermal extremes, and biotic stress) to demonstrate how microbial functions translate to agronomic outcomes and to highlight critical bottlenecks for reproducible, scalable microbiome products.

Microorganisms doi: 10.3390/microorganisms14051137

Authors: Yingxi Li Mingfeng He Yao Song Lu Liu Jiling Xiao Jie Wang Bin Yang Shunyi Ouyang Xin Li Di Peng Zheyuan Zhu

Herbicides considered selective to rice are generally evaluated based on their direct crop safety and weed suppression effects, yet it remains unclear whether they may also trigger indirect or context-dependent effects on rice under rice–barnyardgrass co-cultivation. To address this question, we compared rice performance and associated microbial dynamics under six conditions: rice–barnyardgrass co-cultivation and rice monoculture, each treated with a water spray control or sublethal doses of propanil (Pro, 66.7 mg a.i. L−1) or cyhalofop-butyl (Cyh, 5.86 mg a.i. L−1). Barnyardgrass exhibited visible injury and stronger leaf-level oxidative stress responses, whereas rice displayed no discernible phytotoxic symptoms. Nevertheless, under co-cultivation, herbicide treatment significantly suppressed rice growth, with up to 17.8% lower root lengths and 24.8% lower shoot fresh weights, with reductions varying by herbicide and trait. By contrast, comparable suppression was not observed under herbicide exposure or co-cultivation alone, identifying this response as an emergent, context-dependent negative effect. Microbiota reassembly emerged as an early and stage-specific component of the herbicide-associated response under co-cultivation, with the most pronounced changes detected on day 5 and occurring primarily in bacterial communities. Moreover, bacterial community variation was negatively correlated with root length (ρ = −0.664), and urease activity declined under herbicide treatment. Together, these findings indicate that in paddy fields, herbicides act not only on individual plants but also as an external disturbance to the coupled rice–barnyardgrass system, for which microbiota reorganization represents a key component of the ecological response. Our results suggest that herbicide selectivity should be interpreted within a crop–weed–microbiome context, rather than being inferred solely from their direct crop safety and weed suppression effects.

Microorganisms doi: 10.3390/microorganisms14051136

Authors: Jialing Hou Jingyu Miao Yanyan Dai Yu Cui Sheng Yin Yunbo Luo

Mucin is the core component of the mucus layer barrier in the human reproductive tract and the interaction between mucin and commensal bacteria is crucial for the female reproductive health. In this study, we found that the female reproductive tract isolate Lactobacillus gasseri YS1021 could degrade and utilize mucin to obtain the promoted proliferation. After 48 h of cultivation with 1% mucin added, the biomass of strain YS1021 increased fourfold compared to the control group without mucin. Meanwhile, the addition of mucin induced a 5-fold higher expression of the M13 family metalloprotease encoding gene RS7445, revealing that the enzyme was associated with mucin degradation in the strain YS1201. Moreover, the recombinant strain L. gasseri 7445OE with RS7445 over-expression exhibited an enhanced mucin degradation ability and reached a 7-fold higher biomass than the parent strain YS1021 after 48 h of cultivation with addition of 1% mucin. Furthermore, the gene RS7445 was expressed in Escherichia coli BL21, and the 74 kDa recombinant protein of RS7445 was successfully purified. Enzymatic hydrolysis assay in vitro showed that mucin was decomposed into 39 peptide fragments by the recombinant RS7445, which demonstrated that the M13 family metalloprotease contributes to mucin degradation in L. gasseri YS1021. In sum, this study revealed an important mechanism by which the vaginal L. gasseri YS1021 could degrade and utilize mucin to gain a growth advantage.

Microorganisms doi: 10.3390/microorganisms14051135

Authors: Ikeoluwa Adekoya Michelle Maclean Logan Mackie Katharine C. Carter

Cutaneous leishmaniasis is a protozoan disease which is responsible for significant morbidity in humans. Currently, there is no clinically approved vaccine to prevent infections, and, therefore, treatments to cure skin lesions are required. Ideally, a treatment that can be self-administered to affected areas is desirable. In this study, the effect of violet-blue light, of wavelength in the region of 405 nm, on the survival of Leishmania major and Leishmania mexicana was determined using in vitro and in vivo models. Light treatment caused significant killing of both promastigotes and intracellular amastigotes (p < 0.001) of both species in vitro, and L. mexicana intracellular amastigotes were more resistant to light treatment than L. major intracellular amastigotes. Treatment with violet-blue light at a dose of 45 J/cm2 (0.15 W/cm2 for 5 min) per day on days 3–7 post-infection in an in vivo footpad model caused a significant reduction in L. major parasite burdens on day 5 post-infection (p < 0.05) in one of two experiments, though by day 10 post-infection, parasite numbers had recovered to those of controls. The results of this study clearly demonstrate that violet-blue light can kill both L. major and L. mexicana parasites, but application to infected cutaneous tissues requires refinement.

Microorganisms doi: 10.3390/microorganisms14051134

Authors: Bo Kyeung Jung Jeong Su Han Jae Kyung Kim

The coronavirus disease pandemic impacted respiratory virus circulation and seasonality, but its effect on viral co-detection patterns remains unclear. We examined temporal changes in co-detection burden and dominant pairwise patterns across pandemic periods. We analyzed 23,284 respiratory virus multiplex PCR tests performed at a tertiary care center in the Republic of Korea from 2007 to 2024. Co-detection was defined as detection of ≥2 viruses in a single episode. To address temporal panel changes, we used crude full-panel, restricted 12-target, and recent-period sensitivity analyses based on a stable respiratory virus target set. Co-detection burden showed discordant trends across analytical approaches. In the crude analysis, co-detection among positive episodes was the highest during the pandemic. In the restricted analysis, co-detection decreased from 20.5% before to 14.8% during and 12.2% after the pandemic. Pairwise co-detection patterns also shifted: adenovirus–rhinovirus predominated before the pandemic, whereas rhinovirus-containing pairs involving parainfluenza virus type 3 or respiratory syncytial virus B accounted for relatively larger shares during and, to a lesser extent, after the pandemic. These findings suggest that post-pandemic respiratory virus surveillance should consider not only single-virus positivity or overall co-detection frequency, but also the composition of dominant pairwise viral combinations captured by multiplex PCR.

Microorganisms doi: 10.3390/microorganisms14051133

Authors: Ramokone Mothupi Mashilo Matotoka Gabriel Mashabela Peter Masoko

The persistence of Mycobacterium tuberculosis within biofilm-like structures underscores the need for alternative drug discovery strategies aimed at resistance mechanisms. Medicinal plants provide a rich source of chemically diverse compounds with broad biological activities, including potential antimycobacterial properties. This study investigated acetone stem extracts from Buddleja saligna, Combretum hereroense, and Olea europaea subsp. africana for their phytochemical composition, antioxidant capacity, and antimycobacterial activity against planktonic and biofilm forms of Mycobacterium smegmatis. Phytochemical profiles were analyzed using liquid chromatography–mass spectrometry and quantified through colorimetric assays. Antioxidant activity was assessed using DPPH radical scavenging and ferric reducing power assays, while antimycobacterial effects at MIC and sub-MIC levels were determined through microdilution and growth kinetic assays. Phytochemical composition and concentrations varied among extracts, with B. saligna exhibiting the highest levels of tannins (287.18 ± 0.19 mgGAE/g extract) and flavonoids (16.48 ± 0.05 mgQE/g extract) and showing the strongest antioxidant activity (17.66 ± 5.396 and 399.1 ± 3.717 µg/mL). C. hereroense had a notable antimycobacterial activity with an MIC of 0.16 mg/mL followed by B. saligna and O. europaea subsp. afriana with MIC values of 0.31 and 0.63 mg/mL, respectively. All extracts significantly inhibited early biofilm formation by over 80% at sub-MICs. However, the mature biofilms and sliding motility were less susceptible to the extracts. Overall, the results confirm the antioxidant and antimycobacterial potential of the selected plant extracts, while highlighting challenges in targeting mycobacterial biofilms.

Microorganisms doi: 10.3390/microorganisms14051132

Authors: Abdulla Albastaki Judith Smith

Studying soil microbiomes is challenging because soil contains thousands of microbial species at vastly different abundances. The choice of sequencing method has a strong effect on which of these species are detected and how the community is described. Three approaches now dominate soil microbiome research: short-read 16S rRNA amplicon sequencing on Illumina platforms, full-length 16S sequencing on Oxford Nanopore Technologies (ONT) platforms (particularly the R10.4.1 flow cell), and long-read shotgun metagenomics. Each has distinct biases that shape the recovered community, yet researchers routinely select a method based on cost, understanding, or local expertise rather than on a clear knowledge of what each approach methodically over- or under-represents. Here, we review head-to-head benchmarking studies that have applied two or more of these methods to the same soil or directly comparable samples. We show that while long-read and short-read 16S approaches generally converge on dominant taxa and on between-sample differences, they disagree substantially on alpha diversity estimates, rare taxon detection, and the relative abundances of entire phyla. The R10.4.1 flow cell chemistry has narrowed but not eliminated the accuracy gap with Illumina, and shotgun metagenomics reveals systematic biases in both short and long-read assembly that depend on population diversity within the sample. We synthesise this evidence into an evidence-based decision framework tied to specific research questions and recognise the gaps in soil-specific benchmarking that limit current methods. Rather than asking which platform is “best,” we argue that method choice should be framed as an important part of study design, with the biases of the chosen method acknowledged and, where possible, controlled for.

Microorganisms doi: 10.3390/microorganisms14051131

Authors: Guilherme Campos Tavares Sarah Portes Carneiro Angelo Carlo Chaparro Barbanti Angélica Emanuely Costa do Rosário Helena Caldeira Matos Cynthia Rafaela Monteiro da Silva Maia Henrique Lopes Costa Renata Catão Egger Luiz Fagner Ferreira Nogueira Júlio César Câmara Rosa Felipe Luiz Pereira Fabiana Pilarski Silvia Umeda Gallani Esteban Soto Carlos Augusto Gomes Leal Henrique César Pereira Figueiredo

Lactococcosis has emerged as an economically and ecologically significant disease in aquatic animals worldwide. This study employed multilocus sequence typing (MLST) to investigate the genetic diversity of Lactococcus spp. strains from Brazilian fish species and evaluate their phylogenetic relationships with global isolates to elucidate potential epidemiological connections involving multiple host species and distinct geographic regions. A total of 55 isolates from different laboratories had their DNA extracted, followed by the amplification and sequencing of the internal fragments of seven housekeeping genes (als, atpA, tuf, gapC, gyrB, rpoC and galP). Sequence types (STs) and clonal complexes (CCs) were defined. An unrooted neighbor-joining phylogenetic tree was generated using allele profiles from this study and those previously reported from other aquatic animal species. The isolates comprised 29 STs (11 previously reported, 18 novel ones), which were grouped into species-specific CCs: CC5 (L. formosensis); CC4, CC17, CC62 (L. garvieae); CC24, CC29, CC97 (L. petauri). Considerable genetic divergence was observed, with L. formosensis and L. garvieae forming heterogeneous populations, while L. petauri was more homogeneous. These findings describe the MLST structure of the sampled isolates and should be interpreted as hypothesis-generating rather than population-level estimates of genotype prevalence. Phylogenetics confirmed groupings within the CCs and revealed additional phylogenetic clustering patterns. In conclusion, the Brazilian Lactococcus spp. strains analyzed in this study constitute a genetically diverse population based on their STs. MLST and phylogenetic analysis demonstrated genetic relatedness between the L. garvieae and L. formosensis isolates from this study and those from other aquatic animal species. In contrast, all the STs identified for L. petauri in this study were unrelated to the MLST lineages responsible for outbreaks in Brazilian Nile tilapia (Oreochromis niloticus) and North American rainbow trout (Oncorhynchus mykiss). This suggests that piscine L. petauri populations in the Americas evolved from distinct ancestral origins.

Microorganisms doi: 10.3390/microorganisms14051130

Authors: Xuanwei Huang Caner Akıl Peijun Zhang

Respiratory syncytial virus (RSV) remains a leading cause of severe lower respiratory tract disease in infants, older adults, and immunocompromised individuals. Over the past decade, advances in structural biology, particularly cryo-electron microscopy (cryo-EM) and cryo-electron tomography (cryo-ET), have transformed our understanding of RSV architecture, dynamics, and the mechanisms of entry and replication. High-resolution structures of the prefusion F glycoprotein (pre-F) and its complexes with neutralizing antibodies established the rationale for structure-guided antigen stabilization and directly enabled the development of the first licensed RSV vaccines. Complementary structures of the ribonucleoprotein, polymerase complex, and matrix lattice have broadened therapeutic targets beyond F. Here, we summarize these structural advances; review current structure-guided vaccine, antibody, and antiviral development efforts; and highlight priorities for next-generation vaccines and therapeutics.

Microorganisms doi: 10.3390/microorganisms14051129

Authors: Sergiu Dorin Matei Ramona Nicoleta Suciu Tiberia Ilias Grațiela Aneta Avram Corina Suteu Laura Ioana Bondar Cristian Hocopan Carmen Pantis Roland Fazakas Ovidiu Frățilă

Rising antimicrobial resistance has reduced the effectiveness of empirical eradication regimens for Helicobacter pylori (H. pylori) infection, particularly those containing clarithromycin. Local resistance surveillance and identification of clinical predictors of resistance are essential to guide treatment strategies. This study evaluated antimicrobial resistance patterns and clinical determinants of resistance in a real-world tertiary-care cohort. A retrospective observational study was performed, which included 352 adult patients with confirmed H. pylori infection managed between November 2022 and November 2025. Of these, 168 patients underwent culture and antibiotic susceptibility testing, while 184 received empirical therapy. Resistance rates were calculated according to the number of isolates tested for each antimicrobial agent (available-case analysis). Multivariable logistic regression analysis was used to identify independent predictors of resistance. Among susceptibility-tested patients, resistance to at least one antimicrobial agent was detected in 44.6%. Clarithromycin resistance was most frequent (42.5%), followed by metronidazole (36.4%) and levofloxacin (14.0%), whereas amoxicillin resistance remained low (2.4%). Multidrug resistance (MDR) based on available susceptibility data was observed in 12.5% of cases, most commonly involving dual clarithromycin–metronidazole resistance. Prior eradication therapy was independently associated with resistance (adjusted Odds Ratio aOR 2.41; 95% Confidence Interval CI 1.29–4.51; p = 0.006), while demographic factors were not. Clarithromycin resistance substantially exceeds recommended thresholds for empirical triple therapy in this setting. Prior eradication therapy is the principal predictor of resistance, supporting resistance-informed and stewardship-oriented management strategies.

Microorganisms doi: 10.3390/microorganisms14051128

Authors: Sun Hee Kim Dae Gyu Choi Dong Min Han SeongEui Yoo Jin Ju Park Chan-Woo Kim So-Young Kim

Acetobacter cerevisiae KSO5 is an indigenous strain isolated from Korean fruit vinegar and is a potential starter candidate for vinegar fermentation. Here, we report the first complete circular genome of KSO5, comprising a 3.3 Mb chromosome and two plasmids encoding 2898 genes. Core-genome phylogeny clearly placed KSO5 within the A. cerevisiae clade, supported by ANI (97%) and dDDH (71%) values. Comparative analysis with seven draft A. cerevisiae genomes identified strain-specific genomic islands, mobile genetic elements, and plasmid-borne modules potentially related to genetic stability. Comparative COG profiling suggested enhanced potential for carbohydrate utilization, redox balancing, membrane transport, and stress adaptation within a conserved Acetobacter genomic background. The genome encoded a periplasmic oxidative fermentation system, including membrane-bound pyrroloquinoline quinone-dependent alcohol dehydrogenase and molybdopterin-dependent aldehyde dehydrogenase, together with predicted acetate-handling routes that may reduce intracellular acetate accumulation. Consistent with these features, KSO5 maintained growth and titratable acidity production up to 9% ethanol, with the strongest performance at 7–9% ethanol, whereas both traits declined markedly at 10% ethanol. In 5% ethanol medium, KSO5 also showed high ethanol consumption, comparable to that of A. pasteurianus LMG 1262 and higher than that of most reference strains. These findings link the genomic features of KSO5 to efficient ethanol oxidation, sustained acidification, and stable growth, supporting its potential as a starter strain for vinegar fermentation.

Microorganisms doi: 10.3390/microorganisms14051127

Authors: Murat Aral Ayfer Bakır Cemal Çiçek Elif Tuğçe Güner Didem Özkan Muhammed Furkan Kürkçü Gülşah Ceylan Yağız Mehmet Morkoç Ferit Kulalı Ahmet Kürşad Güneş

Background: Cytomegalovirus (CMV) viral load monitoring forms the basis of preemptive treatment strategies in patients undergoing solid organ and hematopoietic stem cell transplantation. This study aimed to evaluate the analytical performance and inter-method agreement of a laboratory-developed CMV real-time PCR (qPCR) test compared to a commercial reference method using plasma samples. Methods: A total of 100 EDTA plasma samples were analyzed in parallel using a laboratory-developed CMV qPCR test and the reference method (Roche Cobas® CMV). Analytical sensitivity was determined us-ing synthetic DNA cloned into the pUC57 plasmid backbone containing the US17 region of the CMV genome, and the limit of detection (LoD95) was calculated using probit regression analysis. The relationship between the quantitative results obtained from clinical samples was evaluated using the Spearman rank correlation coefficient, while inter-method clinical agreement was assessed using the Bland–Altman method. Results: The limit of detection (LoD95) of the laboratory-developed CMV qPCR test, as determined by probit regression analysis, was 63.8 copies/µL. A weak and statistically non-significant correlation was ob-served between the laboratory-developed CMV qPCR test and the reference method in Spearman rank correlation analysis of samples for which numerical quantitative results were available from both methods (ρ = 0.32; p = 0.22; n = 16). Bland–Altman analysis showed a mean difference of −0.48 log10 units, with the vast majority of measurements falling within the 95% limits of agreement. Conclusions: The assay demonstrated measurable analytical performance and inter-method agreement; however, its use for quantitative viral load monitoring, particularly at low CMV DNA levels, should be interpreted with caution.

Microorganisms doi: 10.3390/microorganisms14051126

Authors: Lin Yuan Liting Cheng Guangnian Yuan Chao Liu Zhiwen Song

To address the issues of high water exchange rates and significant negative environmental impacts associated with eel aquaculture, this study explored the use of yellow clay as a carrier for nitrifying bacterial communities. By pre-enhancing the nitrification capacity of the yellow clay, we aimed to improve the control of inorganic nitrogen in the aquaculture water. Three experimental groups were established: NF-YCA (nitrifying-functional yellow clay-added eel aquaculture system); NN-YCA (non-nitrifying yellow clay-added eel aquaculture system); and YC-F (yellow clay-free eel aquaculture system, blank control). The NF-YCA group had zero water exchange, while the YC-F and NN-YCA groups underwent water exchange equivalent to 28.36 times the system volume. Nitrification was most pronounced in the NF-YCA group, where both mean and peak concentrations of total ammonia nitrogen and nitrite nitrogen were lower than in the YC-F and NN-YCA groups, whereas nitrate nitrogen concentrations in the NF-YCA group were significantly higher than in the other two groups. No significant differences were observed in the survival rate and specific growth rate of elvers among the three systems during the experiment. High-throughput sequencing results revealed that Pseudomonadota and Bacteroidota were the most dominant phyla across all systems. However, the bacterial community structure in NF-YCA was more abundant and stable, and nitrification-related genera, such as Nitrosomonas, were detected in high abundance in this system. The preliminary results demonstrate that the eel aquaculture system with enhanced yellow clay nitrification function, can effectively maintain water quality without water exchange, highlighting its potential for practical application.

Microorganisms doi: 10.3390/microorganisms14051124

Authors: Bogdan Adrian Manta Diana-Maria Mateescu Stela Iurciuc Cris Virgiliu Precup Camelia Corina Pescaru Alina Andreea Tischer

Evidence on hospitalised COVID-19 patients during the Omicron BA.5 wave from Eastern European, vaccine-heterogeneous cohorts remains limited. We conducted a retrospective single-centre cohort study of 395 consecutive adults admitted with laboratory-confirmed COVID-19 to a tertiary infectious-diseases unit in western Romania between 1 July and 31 October 2022. Median age was 72 years (IQR 65–81); 33.2% were unvaccinated, 42.8% had documented prior SARS-CoV-2 infection, and 41.3% were obese. Multivariable logistic regression identified independent predictors of in-hospital mortality and post-acute symptom burden. In-hospital mortality was 15.7% (62/395). Vaccination was independently associated with lower mortality (adjusted odds ratio [aOR] 0.55, 95% CI 0.30–0.99; p = 0.048), as was each 1% increase in admission SpO2 (aOR 0.83, 95% CI 0.76–0.92; p < 0.001), whereas COPD independently increased mortality risk (aOR 2.42, 95% CI 1.15–5.10; p = 0.020). Interleukin-6 was the most discriminating admission biomarker for in-hospital mortality (AUROC 0.70). Bloodstream bacterial co-infection, detected in 22.5% of patients tested on clinical suspicion, was dominated by gut-derived organisms with case-fatality ≥30%. At discharge, 90.1% reported persistent symptoms, most commonly cognitive (24.6%). Prior SARS-CoV-2 infection independently predicted post-acute symptom burden (aOR 2.96, 95% CI 1.75–5.01; p < 0.001), with a specific cardiopulmonary signature. In this BA.5 cohort, vaccination remained protective; IL-6 was the most informative admission biomarker; bloodstream infections suggested gut translocation; and prior infection was an independent determinant of early post-acute symptom burden.

Microorganisms doi: 10.3390/microorganisms14051125

Authors: Cosmina Maria Bouari George Cosmin Nadăş Smaranda Crăciun Nicodim Iosif Fiț

Bovine mastitis is a major infectious disease in dairy cattle, causing significant economic losses and compromising animal health and milk quality worldwide. Among its etiological agents, Staphylococcus aureus is a key contagious pathogen due to its ability to establish persistent intramammary infections and evade host immune responses and antimicrobial therapy. This review summarizes current knowledge on the epidemiology, pathogenesis, clinical presentation, diagnosis, and control of S. aureus in bovine mastitis. Particular emphasis is placed on virulence mechanisms, including adhesion, intracellular persistence, biofilm formation, and immune evasion, which contribute to chronic and recurrent infections. The increasing prevalence of antimicrobial resistance, including methicillin-resistant and multidrug-resistant strains, is highlighted as a major challenge limiting treatment efficacy and posing risks within a One Health context. The review also discusses emerging alternative therapies and innovative control strategies, such as anti-biofilm approaches, immunomodulation, and improved diagnostics, aimed at reducing antimicrobial use. Advances in molecular and point-of-care diagnostic tools are considered for their role in early detection and targeted interventions. Overall, effective control of S. aureus mastitis requires integrated strategies combining prudent antimicrobial use, alternative therapies, improved hygiene, and a multidisciplinary One Health approach.

Microorganisms doi: 10.3390/microorganisms14051123

Authors: Branka Bedenić Marina Nađ Vesna Bratić Daniela Bandić Pavlović Mislav Kasalo Mirela Dobrić Rocío Arazo del Pino Tessa Burgwinkel Andrea Grisold Josefa Luxner Gernot Zarfel Paul G. Higgins

During the COVID-19 pandemic carbapenem-resistant Acinetobacter baumannii (CRAB) was found to be the major pathogen associated with ventilator-associated pneumonia in mechanically ventilated patients. This prompted us to analyze the post-pandemic mechanisms of carbapenem resistance, antibiotic resistance trends, and molecular epidemiology of CRAB in Croatia. In total, 94 CRAB isolates from two hospital centers, including outpatient settings, were investigated. Antimicrobial susceptibility testing was performed by broth microdilution. PCR was used to detect genes encoding carbapenemases of group A, B and D and extended-spectrum β-lactamases (ESBL). Randomly selected isolates were subjected to whole resistome analysis by Inter-array CarbaResist Kit and whole-genome sequencing (WGS). Phylogenetic tree and sequence types (STs) were retrieved from WGS. Plasmid incompatibility groups were determined by PCR-based replicon typing (PBRT). All isolates were extensively drug resistant (XDR), showing resistance to ceftazidime, cefepime, piperacillin–tazobactam, imipenem, meropenem, gentamicin, amikacin and ciprofloxacin, and 13% (n = 12) were also resistant to colistin. The Hodge and CIM test exhibited poor sensitivity with only 32 and 30% of isolates being identified as carbapenemase producers, respectively. PCR identified blaOXA-23 as the dominant carbapenemase gene in both hospitals, found in 71% of the isolates (67/94). In an outpatient setting, blaOXA-24/40 was dominant. blaOXA-23 and blaOXA-72 were the only allelic variants. The Inter-array CarbaResist Kit and whole-genome sequencing (WGS) identified a variety of aminoglycoside (armA, ant(3″)-IIa, aph(3″)-Ib, aph(6)-Id) and sulphonamide resistance (sul1 and sul2) genes. The representative blaOXA-23-positive isolates belonged to ST2, while blaOXA-72-positive isolates were allocated to ST492. These data show that there are different populations of XDR A. baumannii between hospital and outpatients.

Microorganisms doi: 10.3390/microorganisms14051121

Authors: Zhendong Li Yang Chen Yajie Li Aidong Ruan

Although sedimentary zones in Lake Taihu differ in external inputs, hydrodynamic conditions, and sedimentary settings, the spatial differentiation of eukaryotic microbial communities and their assembly mechanisms remain insufficiently understood. This study analyzed sediment cores from four typical sedimentary zones of Lake Taihu: Dapu (DP), Gonghu (GH), the central lake area (HX), and Xuhu (XH). By integrating physicochemical measurements, 18S rRNA gene high-throughput sequencing, redundancy analysis, functional annotation, iCAMP, and co-occurrence network analysis, we characterized the composition, environmental associations, and assembly mechanisms of sedimentary eukaryotic microbial communities. The results showed that eukaryotic microbial communities in Lake Taihu sediments exhibited marked spatial heterogeneity, with dominant taxonomic groups including Chlorophyta, Intramacronucleata, and Diatomea. Alpha diversity was higher in the GH zone and lower in the HX zone, whereas beta diversity showed significant separation among lake zones. NH4+-N, NO3−-N, TN, TP, TOC, D50, MWC, and pH were associated with variation in community composition, but the main associated factors differed among zones. FunGuild annotation showed that annotated fungal functional groups exhibited distinct trophic distribution patterns across sedimentary zones. iCAMP analysis indicated that community assembly was generally dominated by stochastic processes, with dispersal limitation prevailing in the GH zone and ecological drift dominating in the DP, HX, and XH zones. Co-occurrence network analysis further revealed marked differentiation in potential biological associations among sedimentary zones. Overall, this study showed that nutrient conditions and sediment physical properties in different sedimentary environments of Lake Taihu jointly shaped the spatial patterns of eukaryotic microbial communities and their ecological associations, providing baseline information for understanding sedimentary ecological processes in eutrophic shallow lakes.

Microorganisms doi: 10.3390/microorganisms14051122

Authors: Ling Qiao Yuqi Wu Jianping Zhao Ye Chen Jianglin Li Qing Hao Yuanming Guo Tiejun Li

Benthic foraminifera are effective indicators of heavy metal contamination in marine ecosystems. Traditional methods for benthic foraminiferal identification and biodiversity assessment rely predominantly on stereomicroscopic analysis. However, this approach is time-consuming, labor-intensive, and cannot effectively identify small or morphologically similar species. In this study, we aimed to enhance the utility of benthic foraminifera as bioindicators. To this end, we investigated the responses of benthic foraminiferal communities to varying concentrations of Cd under controlled laboratory conditions using both morphological assessments and metabarcoding analyses. Cd exposure reduced the abundance of benthic foraminifera. High Cd concentrations led to Cd enrichment in foraminiferal tests and altered the contents of other elements. Quinqueloculina, Ammonia, and Miliammina exhibited tolerance to Cd, whereas Parasorites and Ovammina were more sensitive. This study provides an effective approach for evaluating the short-term effects of heavy metal pollution on benthic foraminiferal communities.

Microorganisms doi: 10.3390/microorganisms14051119

Authors: Xi Du Mingye Sun Shan Cheng Jiang-Shiou Hwang Rulong Liu Jiasong Fang Li Wang

Members of the class Campylobacteria are microaerophilic bacteria widely distributed across diverse environments and are abundant in hydrothermal systems. However, cultivated representatives, particularly from shallow-water vents, remain limited. Here, we investigated the genomic diversity and environmental adaptation of the genus Sulfurospirillum. Phylogenomic analysis revealed a clear separation between terrestrial and marine clades, with relatively few cultured representatives in the marine lineage. Strain 1307, isolated from shallow-water hydrothermal vents, expands the genomic representation of this underexplored clade. Pan-genome analyses based on complete genomes revealed an open pan-genome, indicating ongoing diversification of genus Sulfurospirillum. Further comparison between hydrothermal vent (HTV) and non-HTV lineages identified distinct adaptive features. Vent-associated strains are enriched in genes involved in sulfur metabolism, carbon fixation, the glycine cleavage system (GCS), and the biosynthesis of key cofactors (spermidine, thiamine, lipoate, and heme), reflecting metabolic adaptation to hydrothermal environments. Beyond well-established processes such as sulfur metabolism and autotrophic carbon fixation, the widespread presence of the GCS in vent-associated lineages suggests its potential role as an auxiliary carbon fixation pathway under anaerobic conditions. Overall, this study expands the phylogenetic and genomic diversity of Sulfurospirillum and offers new insights into the mechanisms underlying environmental adaptation and niche differentiation in vent-associated Campylobacteria.

Microorganisms doi: 10.3390/microorganisms14051120

Authors: Kadir Sinan Arslan Meriam Bouri Aissa Bakelli Fikrettin Şahin

Halotolerant plant growth-promoting rhizobacteria (PGPR) represent a promising strategy for enhancing crop productivity in degraded soils. This study evaluated 51 bacterial strains isolated from the rhizosphere of the Saharan halophyte Phragmites communis L. for their capacity to improve the performance of wheat (Triticum aestivum L.) and pepper (Capsicum annuum L.) under nutrient-deficient sandy soil conditions. The selection of halotolerant isolates was based on their potential for cross-tolerance, assuming that their adaptive mechanisms against salinity could also mitigate the osmotic and nutritional constraints inherent to nutrient-poor sandy substrates. Two strains, XE-15 and XR-18, were selected based on in vitro screening and tentatively assigned to the genera Pseudomonas and Bacillus, respectively, using 16S rRNA sequencing and multilocus sequence analysis (MLSA). Greenhouse experiments demonstrated that bacterial inoculation significantly increased plant biomass (up to ~2-fold compared to control) and enhanced pepper fruit yield (0.68 g vs. 0.20 g in control). XR-18 notably increased Fe (up to 198.65 mg kg−1) and P (7.98 mg kg−1) accumulation in wheat, while XE-15 exhibited substantial concentrations of nitrogen (1.08%) and magnesium (4.11 mg kg−1) and zinc (102.3 mg kg−1). Soil properties were also improved, including increased water-holding capacity (~30%) and enhanced micronutrient availability. Zinc showed the most pronounced strain-specific response, increasing by 84% under XE-15 and by more than 160% under XR-18. However, taxonomic resolution remains tentative in the absence of genome-level analyses, and mechanistic insights are primarily inferred from in vitro traits. The simplified greenhouse system further limits ecological interpretation. These findings highlight the potential of halotolerant PGPR in degraded soils while emphasizing the need for genomic validation, mechanistic studies, and field-scale evaluation.

Microorganisms doi: 10.3390/microorganisms14051118

Authors: Zihang Qin Kaili Guo Xin Li Chuanjun Wang Bao Wang Rulong Chen Yunle Cui Kuojun Cai Yuefeng Chu Gang Yao Xuelian Ma Yawei Sun Na Li

To prevent the spread of antibiotic resistance, bacteriophages have gradually become the most promising alternative to antibiotics for treating bacterial infectious diseases. In this study, using E. coli DC1 as the host strain, we isolated a bacteriophage named Escherichia coli phage XJA18 from farm sewage. We conducted morphological identification, host range determination, biological characteristic analysis, genomic feature analysis, and evaluation of in vitro and in vivo antibacterial effects. Electron microscopy revealed that phage XJA18 belongs to the class Caudoviricetes, with an icosahedral head and a non-contractile long tail. Whole-genome sequencing revealed that the phage has dsDNA with a length of 50,572 bp, with a GC content of 45.33%. The genome does not contain any antibiotic resistance genes or virulence genes, indicating good safety. XJA18 showed lytic activity against 24% of clinically isolated E. coli strains. The optimal multiplicity of infection (MOI) was 0.001, with a latent period of 10 min, a burst period of 30 min, and a burst size of 2.22 × 102 PFU/cell. It remained stable at 4–50 °C and pH 4–12. In vitro antibacterial results revealed that XJA18 had the most pronounced initial bacterial growth suppression at MOI = 0.001 during the first 4 h. In vivo experiments demonstrated that both prophylactic and therapeutic administration of XJA18 could protect against E. coli infection, significantly reducing inflammatory cytokine levels and bacterial loads in the livers and spleens of mice (p < 0.001), significantly increasing body weight (p < 0.05), and reducing histopathological damage to the colon, liver, and lungs. In summary, phage XJA18 can effectively inhibit E. coli and is safe and stable. These characteristics indicate that phage XJA18 has great potential as a novel biological agent to replace antibiotics for treating bacterial infectious diarrhea in calves.

Microorganisms doi: 10.3390/microorganisms14051117

Authors: Silvia Ioana Musuroi Adela Voinescu Corina Musuroi Delia Muntean Florin George Horhat Luminita Mirela Baditoiu Oana Izmendi Andrei Cosnita Valentin Ordodi Zorin Crainiceanu Edward Seclaman Monica Licker

Chronic wounds represent a major complication of underlying conditions such as diabetes mellitus, arterial ischemia, surgical wound and burns. This study aimed at the phenotypic and molecular characterization of antimicrobial resistance for a selection of bacterial isolates, originating from wounds harvested from patients hospitalized in the Vascular Surgery and Plastic Surgery wards. The microbiological diagnosis of wound infections was established according to the laboratory’s working protocol. PCR screening of antibiotic resistance genes was performed using a real-time PCR, while the microtiter plate assay was used to determine the biofilm-forming capacity. Testing of biofilm susceptibility to meropenem and amikacin was performed on Calgary biofilm device. Of the 88 bacterial isolates studied, 78.40% were Gram-negative bacilli (GNB)—Klebsiella pneumoniae (K.P), Pseudomonas aeruginosa (P.A), Proteus mirabilis (P.M), Acinetobacter baumannii (A.B), while the remaining 21.60% were Gram-positive cocci (GPC)—Staphylococcus aureus (S.A). All A.B isolates and 92.59% of K.P were carriers of β-lactamase- and carbapenemase-encoding genes, while 57.89% of S. aureus isolates were carriers of mecA (methicillin-resistant). Strong biofilm-forming isolates (B+++) were more frequent in P.A than in K.P (p = 0.002) and P.M (p = 0.02), with a frequency comparable to that of A.B strains (p = 0.212). When analyzing the biofilm reaction to meropenem, a significantly lower susceptibility was detected in the biofilm for K.P isolates, compared to the planktonic ones. Most GNB have been extensively multidrug-resistant, particularly K.P and A.B. Isolates from chronic wounds are major biofilm-formers. A strong and statistically significant association has been identified in the case of K.P and P.M between the presence of resistance genes and the biofilm-forming capacity. These findings highlight the need for a customized therapeutic approach for each chronic wound, considering the mechanisms underlying treatment resistance. These include bacterial virulence factors and the wound microenvironment colonized by the biofilm and the relative contribution of each to the overall resistance profile.

Microorganisms doi: 10.3390/microorganisms14051116

Authors: Matthew S. Tucker Doaa Naguib Celia N. O’Brien Christina Yeager Benjamin M. Rosenthal Mark C. Jenkins Asis Khan

Enteric coccidian parasites harm agriculture and human health. Infectious, sporulated parasites eventually senesce. Here, we examined transcriptional changes in sporulated oocysts of Eimeria acervulina stored for 4–30 months at 4 °C. Precipitous decline in RNA abundance and transcription followed an interval of stability. Sixty constitutively expressed genes each contributed > 1000 transcripts per million (TPM) throughout, including a serine protease inhibitor, surface antigen genes, a cation-transporting ATPase, an oocyst wall protein, a zinc finger DHHC domain-containing protein, and highly expressed hypothetical proteins with no known function. Strikingly, ~82% of 6867 annotated genes underwent differential expression when comparing freshly sporulated parasites to those held for 30 months; nearly one-third of these underwent significant expression change. In freshly sporulated oocysts, 86 significantly DEGs exceeded 1000 TPM; these encoded heat shock proteins, lactate dehydrogenase, glucose-6 isomerase, and various hypothetical proteins. The oldest parasites expressed 66 DEGs, including many ribosomal subunits, a haloacid dehalogenase-like hydrolase domain-containing protein, and various hypothetical proteins. Taken together, these findings helped us to identify markers of mature parasites that remain relatively abundant in the transcript pool as oocysts age and identify other transcripts (e.g., ribosomal RNA) that increase in their relative abundance even as RNA abundance declines in senescent parasites.

Microorganisms doi: 10.3390/microorganisms14051115

Authors: Zhongyan Tang Chen Chen Li Dong Liuyuan Bao Chengcui Yang Xiaodan Wang Xiaoling Chen Xiaokun Li Fajun Xiang Shunqiang Yang

Morchella sextelata a species of high nutritional and economic value, is widely cultivated. To investigate how different cultivation environments affect the soil physicochemical properties and microbial communities associated with common morel, this study established cultivation plots under three distinct settings: apple orchard canopies, dry upland fields, and paddy fields. The objective was to compare the differential impacts of common morel cultivation on soil environmental conditions across these habitats. The results indicate that cultivating common morel effectively enhances soil fertility. Across all environments, soil hydrolyzable nitrogen (HN), available potassium (AK), and organic matter content were higher than in the control. In apple orchard and dryland soils, total phosphorus (TP), total potassium (TK), available phosphorus (AP), and pH values were also elevated compared to the control, with most differences reaching significant levels. Solid Sucrase (S-SC) activity increased in all environments compared to the control, with values of 17.52 mg/d/g in PG, 17.39 mg/d/g in HD, and 21.68 mg/d/g in DT soils. Soil Amylase (S-AL) activity was higher in PG (451.28 μg/h/g) and HD (475.38 μg/h/g) soils. In contrast, Soil-acid phosphatase (S-ACP) activity was significantly elevated in DT soil (2922.08 nmol/h/g). PG soil exhibited significantly higher activities of Solid-Catalase (S-CAT), Solid polyphenol oxidase (S-PPO), and Solid Urease (S-UE), with S-CAT reaching 952.5 μmol/h/g. Following common morel cultivation, bacterial richness and diversity decreased across all conditions, while fungal richness increased but diversity declined. At the phylum level, Proteobacteria remained the dominant bacterial group, accounting for 26.78% in PG, 28.27% in HD, and 20.05% in DT soils. Ascomycota was the predominant fungal phylum, comprising 68.03% in PG, 72.16% in HD, and 68.94% in DT soils. Predicted bacterial functional pathways were primarily associated with metabolism, genetic information processing, environmental information processing, and cellular processes. Key metabolic pathways included carbohydrate metabolism, amino acid metabolism, and metabolism of cofactors and vitamins. fungal functional guilds were mainly classified as pathotrophic, pathotrophic–saprotrophic, pathotrophic–saprotrophic–symbiotrophic, and saprotrophic. Among these, saprotrophic and pathotrophic guilds showed higher abundance compared to the control. This shift is characterized by a reduction in both the diversity and abundance of beneficial microorganisms, alongside an increase in the richness of harmful microbial taxa. The combined effect of these factors disrupts the soil microbial equilibrium. The findings of this study provide a theoretical foundation for the cultivation of common morel and the management of associated soils.

Microorganisms doi: 10.3390/microorganisms14051114

Authors: Akamu Ewunkem Josiah Dixon Jordan Queenie Uchenna Iloghalu Franklin Ezeanowai Sada Boyd

Rising antibiotic resistance necessitates alternatives such as silver nanoparticles (AgNPs), which exhibit bactericidal activity via multi-target mechanisms (e.g., membrane disruption, ROS production). While resistance to chemically synthesized AgNPs exists, the potential for resistance to green-synthesized AgNPs, such as those from reishi mushroom, is unknown. This study compared S. aureus resistance development against both AgNP types using experimental evolution by analyzing genomic and morphological changes. Additionally, this work evaluated potential cross-resistance responses to ionic silver and investigated how adaptation to green-synthesized AgNPs affects sensitivity to chemically synthesized AgNPs (and vice versa). Rapid resistance, along with cross-resistance to silver ions, emerged in bacteria following 14 days of sublethal exposure to silver nanoparticles, regardless of whether they were chemically or biologically synthesized. While green-synthesized AgNPs demonstrated a substantial resistance to chemical variants (p < 0.05), the reverse effect was not as strong, and resistant populations showed distinct morphological adaptations. Genomic analysis highlighted convergent hard selective sweeps, identifying common mutations across both chemical and green AgNP-treated populations, with limited unique mutations found for either. These findings enhance our understanding of bacterial resistance mechanisms to nanomaterials, contributing to the development of safer, eco-friendly, and high-efficacy treatments against multidrug-resistant infection.

Microorganisms doi: 10.3390/microorganisms14051113

Authors: Helena Chacón-Navarrete Marina Barbudo-Lunar Francisco Javier Ruiz-Castilla José Ramos

Increasing consumer demand for natural and safe food products has led to the exploration of biocontrol alternatives to chemical preservatives, especially in the cured meat industry. The yeast Debaryomyces hansenii has emerged as a promising biocontrol candidate due to its antagonistic properties against spoilage fungi. This study assessed the impact of D. hansenii inoculation on the fungal community structure of Iberian cured pork loin using high-throughput sequencing of the ITS1 region. Ion Torrent ITS1 amplicon sequencing, QIIME2/DADA2 pipeline, and ALDEx2 differential abundance analysis were applied to this study. Pork loin samples inoculated with D. hansenii were compared to non-inoculated controls to evaluate changes in the fungal microbiome. Inoculation resulted in a marked decrease in fungal diversity and evenness, indicating strong competition by D. hansenii against native fungal populations. This effect was reflected in a significant reduction in alpha diversity in inoculated samples (Shannon, p = 0.0042; Pielou p = 0.0075; Gini–Simpson, p = 0.0081). Notably, genera associated with spoilage and mycotoxin production, particularly Aspergillus and Penicillium, were significantly reduced in inoculated samples. Simultaneously, D. hansenii became dominant, reducing other yeasts and filamentous fungi. These findings highlight the powerful competitive and biocontrol potential of D. hansenii, demonstrating its ability to improve microbial safety by potentially reducing mycotoxin-associated risks through the suppression of toxigenic genera. This is the first study to characterise the fungal community of Iberian pork loin using metabarcoding under D. hansenii inoculation. The findings confirm that the inoculation of D. hansenii can substantially reduce fungal contamination risks. Overall, the results contribute valuable insights into microbial interactions during meat curing and underscore the practical benefits of targeted starter cultures for enhancing food safety and quality.

Microorganisms doi: 10.3390/microorganisms14051111

Authors: Geril Sekangue Obili Bridy Chelsy Moutombi Ditombi Charlene Manomba Boulingui Roger Hadry Sibi Matotou Joyce Coëlla Mihindou Dimitri Mabicka Moussavou Denise Patricia Mawili Mboumba Marielle Karine Bouyou-Akotet

Digestive candidiasis is a major opportunistic infection among people living with HIV (PLHIV). In Gabon, data on antifungal resistance remain limited. This study aimed to characterise Candida colonisation and antifungal resistance according to anatomical site and species in Libreville. In this cross-sectional study, 108 PLHIV provided paired oral and stool samples. Candida spp. was identified using conventional phenotypic methods. Antifungal susceptibility to azoles and polyenes was assessed by disc diffusion following CLSI guidelines. Resistance burden was classified by drug class and by cumulative number of antifungal agents involved. Digestive colonisation was detected in 97 (89.8%) participants. Oral and intestinal colonisation rates were 78.7% and 66.7%, respectively, with dual-site involvement in 55.6%. Among resistant isolates, Candida albicans accounted for 55.2% (oral) and 48.9% (intestinal), while non-albicans Candida represented 29.8% and 44.4%, respectively. Multidrug resistance was significantly higher in intestinal than oral isolates (36.2% vs. 11.8%; OR = 4.99; 95% CI: 2.04–12.16; p < 0.01). Resistance was predominantly azole-driven, with complex cumulative resistance profiles in intestinal isolates. The intestinal tract showed resistance profiles consistent with a preferential accumulation of MDR Candida populations in PLHIV. Site-specific resistance patterns underscore the importance of targeted sampling and antifungal stewardship strategies in resource-limited settings.

Microorganisms doi: 10.3390/microorganisms14051112

Authors: Marina Ferreira Batista-Zauli Maria Eduarda Carvalho Guimarães Brasil Carlos Roberto de Almeida-Júnior Bárbara Germana Soares de Abreu Nailma Silva Aprigio dos Santos Mayra Fernanda Ricci Santuza Maria Ribeiro Teixeira

Different protozoan parasites are the causative agents of tropical diseases, including malaria, toxoplasmosis, leishmaniasis, and Chagas disease (CD), which, altogether, affect over 300 million people throughout the world. Except for two recently approved malaria vaccines, individuals affected by or at risk of contracting any of these four diseases still experience a lack of effective treatments and vaccines. Many vaccine studies, including those that have reached clinical trials, are based on inactivated parasites, adjuvanted recombinant proteins, or viral vector vaccines. Here, we review the current advances towards the development of vaccines based on genetically modified live-attenuated parasites (GMLAP) as well as RNA formulations encoding parasite antigens. Because these are diseases caused by intracellular pathogens that depend on efficient T-cell responses for parasite control, these two new vaccine platforms have generated great expectations, since they are known to induce a robust cellular immune response. Although preclinical studies aimed at developing new malaria, toxoplasmosis, and leishmaniasis vaccines have led to significant progress that may soon result in clinical trials, advances in next-generation vaccines against CD are lagging behind. Increased collaborative efforts between research groups, governments, and the pharmaceutical industry, particularly in Africa, Asia, and Latin American countries, are urgently needed to accelerate the development of vaccines for all neglected and less-studied diseases.

Microorganisms doi: 10.3390/microorganisms14051108

Authors: Caiyuan Zhou Jing Sun Yihan Luo Fang Wang Luqi Li Tong Wu Peng Xie Chenxi Liu Yibin Hu Leilei Sun Chengbao Wang

The bactericidal mechanisms of fluoroquinolones extend beyond their canonical inhibition of DNA topoisomerases, yet the associated metabolic perturbations remain incompletely understood. In this study, we systematically investigated the metabolic responses of Escherichia coli to three representative FQs—ofloxacin, enrofloxacin, and ciprofloxacin—using untargeted UPLC–Q Exactive Orbitrap–MS-based metabolomics. Bacterial cells were exposed to bactericidal concentrations (2 × MIC) for a single-time point (1 h), followed by comprehensive metabolomic profiling with six biological replicates per group. Our findings demonstrate that FQ-induced metabolic reprogramming serves as a primary driver of oxidative stress and nucleic acid damage, rather than a mere secondary effect. All three FQs induced substantial metabolic reprogramming characterized by disruptions in nucleotide biosynthesis, central carbon metabolism, and redox-related pathways, with notable drug-specific differences. Ciprofloxacin exhibited the most pronounced suppression of energy metabolism and antioxidant systems, whereas ofloxacin and enrofloxacin showed partial compensatory metabolic responses. Consistently, intracellular ROS levels were significantly elevated in all treatment groups, and this effect was attenuated by antioxidant supplementation. Furthermore, increased accumulation of 8-hydroxydeoxyguanosine and 8-hydroxyguanosine confirmed the occurrence of oxidative DNA and RNA damage. Collectively, these findings indicate that FQs induce distinct metabolic perturbations that are closely associated with oxidative stress and nucleic acid damage, providing a metabolic perspective on their bactericidal activity and suggesting potential targets for metabolic adjuvant strategies.

Microorganisms doi: 10.3390/microorganisms14051110

Authors: Bogdan Sendrea Argyrios Periferakis Aristodemos-Theodoros Periferakis Ioannis Xefteris Lamprini Troumpata Konstantinos Periferakis Andreea-Elena Scheau Emi Marinela Preda Dana-Georgiana Nedelea Diana-Elena Vulpe Rares-Mircea Birlutiu Cristian Scheau Romica Cergan

Spinal infections in general, and infectious spondylodiscitis in particular, are increasingly diagnosed in the Western world, in recent decades. This rise in incidence is associated with an ageing population and with an increased availability of accurate diagnostic modalities. Even so, due to the non-specific nature of clinical manifestations, and of the implicated blood and serum markers, there is a risk of underdiagnosis or misdiagnosis of the disease in its initial stages. Ionizing radiation methods, such as plain radiography (X-ray) and computed tomography (CT), are also not reliable in the early stages of the diseases, and the golden standard of imagistic diagnosis, magnetic resonance imaging (MRI), is not always available or requested. Still, MRI remains the most reliable method in most cases where there is a need for differential diagnosis with other pathologies, namely Andersson lesions, destructive spondyloarthropathy, erosive osteochondritis, micro-crystalline spondylitis, Modic 1 lesion, Charcot spinal arthropathy, osteoporotic fractures, SAPHO syndrome with spinal involvement, and Schmorl’s nodes. Infectious spondylodiscitis is caused by bacteria, and, less frequently, by fungi. Rare cases of parasitic causes have also been reported in the literature. Infectious spondylodiscitis of bacterial causes may be pyogenic, more frequently caused by Staphylococcus spp. or Streptococcus spp., or granulomatous, usually caused by Mycobacterium tuberculosis complex (MTBC) or from classical brucellosis. In all these cases, therapy may be conservative, with antibiotics, or surgical, when the former fails or in patients with significant spinal instability or other neurological manifestations. There are various surgical approaches, each with its own drawbacks, and usually used according to the preference of the attending physician. Even in cases of surgical treatment, antibiotic administration is prolonged, and it is important for a proper scheme to be selected based on antimicrobial susceptibility testing. However, given that in many cases, the causative agent cannot be identified, empirical treatment must be initiated. Finally, newer approaches, including the incorporation of antimicrobial substances, may offer better solutions for improving treatment and rehabilitation outcomes.

Microorganisms doi: 10.3390/microorganisms14051109

Authors: Elli-Panagiota Magklara Maria Kkirgia Andreas G. Tsantes Petros Ioannou Alexandra Mpakosi Vasiliki Mougiou Zoi Iliodromiti Theodora Boutsikou Nicoletta Iacovidou Rozeta Sokou

Nipah virus (NiV) is an animal-borne RNA virus of the genus Henipavirus that poses a significant global health threat. This threat is driven by the virus’s high mortality rate, its capacity to cause epidemics, and the lack of licensed therapeutic interventions or vaccines. Since its initial identification during the 1998–1999 outbreak in Malaysia and Singapore, recurrent episodes have occurred primarily in Bangladesh and India, with mortality rates frequently exceeding 70%. Fruit bats of the genus Pteropus serve as the biological host for the virus. Transmission to humans occurs via contact with infected wildlife, consumption of contaminated products, such as freshly harvested date palm sap, or direct person-to-person exposure. Other modes of transmission, such as transplacentally or via breast milk, are still under investigation. The clinical presentation of NiV infection varies widely, from mild flu-like symptoms to life-threatening respiratory disease and acute encephalitis. It frequently attacks the nervous system, which can lead to coma, permanent neurological damage, or relapsing encephalitis. The virus enters host cells via ephrin-B2/B3 receptors, enabling systemic dissemination and infiltration of the central nervous system. Diagnosis relies primarily on RT-PCR and serological assays, and virus isolation requires high-containment laboratories. Management remains largely supportive, as no approved antiviral therapy exists. Experimental agents, such as remdesivir, favipiravir, and monoclonal antibodies such as m102.4, have shown promise in preclinical studies. Multiple vaccine platforms—including subunit, viral vector, mRNA, and nanoparticle-based approaches—are under development, though none is yet licensed for human use. Strengthened surveillance, infection control measures, and continued research are essential to mitigate the threat posed by this emerging pathogen. This review summarizes current knowledge on NiV, including its virology, epidemiology, pathogenesis, transmission, and recent progress in therapeutic and vaccine development.

Microorganisms doi: 10.3390/microorganisms14051105

Authors: Thibault Maumy Claire McCartney Ayodeji Samuel Ajayi Claire Gerkins Gabriela Fragoso Annie Calvé Manuela M. Santos

Host–microbiota interactions during the neonatal window of opportunity have gained significant interest as factors influencing long-term health. Factors such as nutrient availability may shape the microbial community, and iron is no exception to this rule. Although the use of iron supplementation is widespread during infancy, this micronutrient is known to have profound effects on gut microbiota. This study aimed to determine how early-life iron supplementation shapes gut microbiota composition and whether it influences recovery from gut dysbiosis later in life. Three-week-old female C57BL/6 mice were fed an iron-excess diet for five weeks during the critical period of microbiota establishment. After a two-week washout period to normalize luminal iron content, dysbiosis was induced using either dextran sulfate sodium-induced acute colitis or antibiotic treatment. Mice were then allowed an 8-week recovery period. Gut microbiota composition was longitudinally analyzed by 16S rRNA gene sequencing of fecal samples. Early-life iron supplementation induced durable alterations in gut microbiota composition, with differences persisting even after luminal iron normalization (β-diversity, PERMANOVA p < 0.01). At the endpoint, mice exposed to an iron-sufficient diet remained significantly more distant from their baseline compared to the excess iron group in both the dextran sulfate sodium (33% greater distance) and antibiotic (41% greater distance) models (both p < 0.05). Notably, this pattern was not observed when supplementation occurred in adulthood. In the dextran sulfate sodium model, mice that received an iron-sufficient diet during early life showed an expansion of the probiotic Ligilactobacillus murinus, which positively correlated with fecal succinate levels. Conversely, in the antibiotic-induced model, early-life exposure to an iron-sufficient diet was associated with a more pronounced dysbiosis characterized by a nearly two-fold-greater loss in α-diversity compared to 500 ppm mice (∆Shannon: 1.98 ± 0.22 vs. 1.02 ± 0.25, p < 0.01). These findings suggest that early-life iron supplementation influences long-term host–microbiota interactions and recovery from gut dysbiosis.

Microorganisms doi: 10.3390/microorganisms14051107

Authors: Sifan Ai Yu Wang Jiao Pan Gang Hu Ruiting Zhao

Dajue Temple, a representative ancient architectural heritage in North China, houses numerous lacquered wooden components of exceptional historical and artistic value. Prolonged environmental exposure causes severe dark discoloration and black spotting on lacquer surfaces, threatening their structural integrity. This first investigation into the damage identifies the spots as microbial in origin, with Cladosporium spp. as the primary agent driving deterioration and possessing wood-degrading capabilities. Antifungal tests show that thymol, clove essential oil, and nano-silver gel are all effective inhibitors. We proposed targeted, relic-friendly microbial control strategies tailored for ancient lacquered wooden components. These findings provided scientific guidance for the sustainable conservation and restoration of lacquered architectural elements in historic temples and comparable cultural heritage sites. In future work, environmental monitoring and the biocides’ compatibility should be involved, which will help to clarify microbe–environment interactions, enable early warning of biodeterioration risks and explore the wood-friendly biocides.

Microorganisms doi: 10.3390/microorganisms14051106

Authors: Jing Feng Jiancong Huang Shaolu Zhou Xianghai Chen Gang Zhou Xudong Wang Xia Wen Qingshan Shi Pianjuan Guo Qiongfei Li Xiaobao Xie

Seborrheic alopecia (SA) is one of the most common forms of hair loss with a complex pathogenesis involving multiple etiological factors. Although the scalp microbiome has been implicated in various scalp disorders, its specific role in the development and progression of SA remains incompletely understood. To characterize the scalp microbiome in SA, we performed high-throughput sequencing of the 16S rRNA gene and ITS region on scalp samples from 41 Chinese SA participants before and after a 12-week intervention with a shampoo containing herbal extracts (ginger root, Polygonum multiflorum, and Platycladus orientalis leaf) and 29 healthy controls. The untreated SA group exhibited significant microbial dysbiosis compared to the healthy controls, characterized by reduced bacterial and fungal alpha diversity and increased relative abundances of Staphylococcus, Cutibacterium, and Malassezia. LEfSe analysis confirmed the significant enrichment of these three genera. Correlation network analysis revealed a substantial restructuring of microbial interactions in the untreated SA group: Staphylococcus and Malassezia lost all positive correlations with other genera, whereas Cutibacterium displayed relatively stable topological relationships. Following the 12-week intervention, the treated SA group showed significant clinical improvement (reduced hair loss and scalp sebum content), along with a restoration of microbial diversity to levels comparable to the healthy group and a normalization of the abundances of Staphylococcus and Malassezia. Our study confirms the critical role of scalp microecological dysbiosis in SA pathogenesis and identifies Staphylococcus and Malassezia as key taxa strongly associated with this dysbiosis. These findings provide a theoretical foundation for developing microbiome-targeted strategies for SA treatment and support the use of multi-targeted, plant-based interventions to restore microbial homeostasis and promote hair growth.

Microorganisms doi: 10.3390/microorganisms14051104

Authors: Glenda M. Davison Tandi Matsha Shanel Raghubeer Stanton Hector Saarah Davids Yvonne Prince

The oral cavity is home to a diverse community of microbiota comprising bacteria, viruses, protozoa, and fungi. These microorganisms inhabit several oral niches and play a significant role in supporting both oral and systemic health. The fine balance between the microbial communities can be influenced by genetics and environmental factors, potentially leading to dysbiosis. Alterations in the oral microbiota have been implicated in periodontitis, chronic inflammation, and systemic disease. Tobacco has been identified as a major player in altering the oral microenvironment and disturbing the balance between potentially pathogenic and beneficial commensals. The resulting dysbiosis promotes inflammation and assists in the passage of pathogenic microorganisms into the blood system. This narrative review examines current evidence linking the use of tobacco with the dominance of pathogenic oral bacteria and a dysfunctional immune response. We explore how the chemicals and toxins in cigarettes promote a reduction in oxygen and cause changes in the abundance of anaerobic bacteria. After discussing the mechanistic pathways leading to periodontitis and the entry of microorganisms into the circulation, the review will interrogate previous studies and identify opportunities and priorities for future research.

Microorganisms doi: 10.3390/microorganisms14051103

Authors: Jiya Wu Weicheng Yang Xiaona Zhang Xianyu Li Bibo Zhou Tianyu Liang Fen Liu

Alsophila spinulosa is a tree fern designated as a second-class nationally protected species in China and valued for its medicinal and ornamental properties. Its slow growth and susceptibility to environmental stresses pose challenges to its cultivation. Plant-growth-promoting rhizobacteria (PGPR) can enhance plant development by producing phytohormones, such as indole-3-acetic acid (IAA). In this study, 39 IAA-producing strains were isolated from the rhizosphere of A. spinulosa. Morphological and molecular analyses identified the highest IAA-producing strain, R74, as Burkholderia pyrrocinia. Its optimal inoculum age was determined to be 12–20 h, and its optimal culture conditions for IAA production were 24 h of incubation, 32 °C and pH 7.0. Whole-genome sequencing revealed that the genome of strain R74 is 8,347,169 bp in size with a GC content of 67%, comprising 7543 genetic elements. Further genomic analysis showed that IAA biosynthesis in R74 involves the tryptophan side-chain oxidase (TSO) pathway and the tryptophan-independent pathway. Pot experiments revealed that inoculation with R74 increased the height, root length, stem diameter, and biomass of A. spinulosa seedlings. It also increased antioxidant enzyme activities, elevated soluble protein and chlorophyll contents, and reduced malondialdehyde levels. This study provides an empirical basis for the development of Burkholderia-based biofertilizers to promote A. spinulosa growth.

Microorganisms doi: 10.3390/microorganisms14051102

Authors: Ndzalo Mashabela Darian Naidu Ernest Oduro-Kwateng Nompumelelo P. Mkhwanazi

Endophytic fungi are promising sources of novel antiviral compounds, and the crude extract from Alternaria alternata PO4PR2 has previously shown anti-HIV-1 activity. This study evaluated its efficacy against integrase strand-transfer inhibitor (INSTI)-resistant HIV-1 and its mechanism of action. Key resistance mutations (Y143H, G118R, N155H, and R263K) were introduced into the HIV-1 pNL4.3 clone via site-directed mutagenesis and confirmed through Sanger sequencing. Viral infectivity was assessed in TZM-bl cells, while cytotoxicity was measured using an MTT assay. Antiviral activity was determined through a luciferase-based assay, and integration inhibition was evaluated using integrase activity assays and Alu-gag nested PCR. The extract demonstrated potent inhibition of resistant mutants, with low IC50 values (0.02971–0.1652 μg/mL), and showed minimal cytotoxicity (CC50 = 300 μg/mL), maintaining over 80% cell viability. It inhibited integrase activity by 67%, specifically targeting the strand-transfer step, and significantly reduced integrated viral DNA. Molecular docking of 14 compounds identified coumarin derivatives as key bioactive metabolites, exhibiting mutation-tolerant binding within the integrase catalytic pocket. Overall, these findings highlight PO4PR2 as a promising source of compounds for developing new therapies targeting drug-resistant HIV-1 integrase.

Microorganisms doi: 10.3390/microorganisms14051101

Authors: Mina Popovic Boris Rajcic Neveka Rajic

The accumulation of polyethylene (PE) in aquatic ecosystems represents a significant environmental challenge due to the polymer’s high molecular weight and chemical stability. This study investigates the biodegradation potential of Hafnia paralvei UUNT_MP29, a bacterial strain isolated from the gut of common carp (Cyprinus carpio), for low-density polyethylene (LDPE). Initial screening on LDPE-emulsified agar confirmed extracellular enzymatic activity through the formation of distinct clear zones. Quantitative analysis showed a cumulative mass loss of 24.10% by Day 16, with the most intensive degradation occurring between Days 4 and 8, which closely correlated with maximum bacterial count (CFU/mL). Kinetic modeling indicated that the degradation followed a first-order rate law (R2 = 0.9269), with a rate constant (k) of 0.2991 days−1 and a remarkably short half-life (t1/2) of 2.32 days. Structural characterization via FTIR spectroscopy demonstrated oxidative transformation, evidenced by a reduction in sp3 C-H stretching and the emergence of C-O/C-O-C functional groups. SEM micrographs further confirmed extensive bio-deterioration, including surface pitting and macroscale erosion. Thermal analysis (TGA/DTG) supported these findings, showing a significant 10.95 °C decrease in the maximum degradation temperature (Tmax), indicating a reduction in polymer chain length. These results suggest that H. paralvei UUNT_MP29 is a highly efficient agent for the rapid breakdown of polyethylene and highlight the potential of aquatic gut microbiota as reservoirs for plastic-degrading biotechnologies.

Microorganisms doi: 10.3390/microorganisms14051100

Authors: Tania Islas-Flores Estefanía Morales-Ruiz Marco A. Villanueva

Protists arose about 1 [...]

Microorganisms doi: 10.3390/microorganisms14051099

Authors: Zichun Yan Shuichao Fan Wankai Yan Haopeng Ma Tianhao Zhao

To address the problems of short-circuit flow and dead zones, complicated operation and control caused by intermittent influent, and the mismatch between aeration rate and oxygen demand in the Cyclic Activated Sludge System (CASS), a novel Multi-Compartment Fixed-Biofilm Cyclic Activated Sludge System (MCFCASS) was developed. This system operated in continuous-flow mode, and the aeration rate of each compartment was redistributed using a mathematical model. The results show that the plug flow ratio of the MCFCASS reactor increased from 18.75% to 31.25% compared with the CASS reactor. After aeration rate redistribution, the average total nitrogen (TN) removal efficiency of the MCFCASS reactor rose from 83.34% to 86.80%, and the effluent TN concentration consistently met the Grade I-A limit (15 mg/L) specified in the Discharge Standard of Pollutants for Municipal Wastewater Treatment Plant (GB 18918-2002). The average removal efficiencies of chemical oxygen demand (COD) and ammonium nitrogen (NH4+-N) increased from 91.58% and 93.39% to 92.98% and 94.57%, respectively. Microbial community analysis revealed that after aeration rate redistribution, the relative abundances of Pseudomonadota, Bacteroidota, and Bacillota in the pre-reaction zone of MCFCASS were 39.17%, 17.78%, and 10.33%, respectively. In addition, the abundances of some functional bacterial groups in the first and fourth compartments of the main reaction zone shifted adaptively in response to the aeration rate redistribution, consistent with the trends in pollutant removal contributions in these compartments. Hierarchical clustering and principal coordinate analysis (PCoA) further indicated that aeration rate redistribution influenced the microbial community structure. The above laboratory-scale optimization results may provide a preliminary reference for aeration control and improvement of denitrification performance in similar processes.

Microorganisms doi: 10.3390/microorganisms14051098

Authors: Darshana Kadekar Deepak Velayudhan Ester Vinyeta Jianqiang Zhang Ethan Aljets Veeraya Bamrung Panchan Sitthicharoenchai Alyona Michael Keith Frogue Meng Heng Amy Liu Cristina Bongiorni Manasi Bhate David A. Estell Chong Shen Charlotte Poulsen

Porcine reproductive and respiratory syndrome virus (PRRSV) remains a major challenge to swine production worldwide. Current vaccines have limited efficacy against genetically diverse PRRSV strains. Therefore, strategies with alternative modes of action—such as antiviral approaches that target conserved virus–host interactions, including viral attachment and entry, rather than relying solely on adaptive immune responses—are needed. We first evaluated the in vitro effect of griffithsin (GRFT), a high-mannose-binding lectin, in the monkey kidney cell line MARC-145. Cells were pre-treated with GRFT (50–200 µg/mL) prior to PRRSV infection, after which cell morphology and viral RNA replication (measured by RT-qPCR) were assessed. Pre-treatment with 100–200 µg/mL GRFT, followed by PRRSV inoculation at a multiplicity of infection of 1 or 10, reduced viral replication in MARC145 cells in a dose-dependent manner, achieving almost 100% inhibition of ORF5 and ORF7 RNA compared with untreated controls (p < 0.0001). We next investigated the in vivo effects of intranasal GRFT administration (7.5 or 15 mg/day) in pigs (n = 56). Pigs treated with 15 mg/day GRFT exhibited significantly reduced (p < 0.05) viremia 2, 4 and 7 days post-challenge, compared with untreated, challenged, and controls (log10 8.1 ± 0.2 vs. 9.0 ± 0.25, 8.2 ± 0.1 vs. 9.1 ± 0.2, and 8.9 ± 0.2 vs. 9.3 ± 0.2, respectively), along with earlier resolution of fever and a trend toward increased average daily gain over 42 days (p < 0.1). These findings are the first report of GRFT efficacy in pigs and support its potential as an antiviral strategy against PRRSV, alongside existing interventions.

Microorganisms doi: 10.3390/microorganisms14051096

Authors: András P. Bózsik Dömötör M. László Borisz Egri

Research increasingly identifies wild birds, particularly long-distance migratory species, as epidemiologically relevant hosts and vectors for tick-borne Borrelia species that pose risks to both avian and human health. This review contextualizes avian-associated Borrelia research historically and microbiologically, showing the role of avian hosts in the ecology of agents causing relapsing fever and Lyme borreliosis. We identify key publications that trace the evolution of Borrelia research—from early microscopic observations of spirochetes to the modern molecular and serological evidence. The review collects literature on the process by which Borrelia gained early scientific attention due to its characteristic morphology and elevated bloodstream concentrations during septicemic phases, which enabled early etiological links between the microbe and disease. It follows the recognition of avian spirochetosis caused by Borrelia anserina and charts the shift in focus after the discovery of Borrelia burgdorferi sensu lato (Subgen. novum recomm. Borreliella, Lyme-group Borrelia). Publications listed show that birds can transport infected human-parasitic ticks over long distances and, in certain bird species, selectively amplify Lyme-group Borrelia species, especially Borrelia garinii, which has the highest temperature tolerance and is thus potentially viable in avian hosts. The literature supports the role of birds in maintaining and disseminating Borrelia infections and infected ticks across continents.

Microorganisms doi: 10.3390/microorganisms14051097

Authors: Antonia Mataragka Marios Mataragas Nikolaos Tzimotoudis Ioannis Galiatsatos Panagiota Stathopoulou Spiros Paramithiotis John Ikonomopoulos Nikolaos D. Andritsos

Listeria monocytogenes is already a well-known foodborne bacterial pathogen, ubiquitously dispersed not only in the food production environment but also in the primary animal production environment as well. The present study performed whole-genome characterization of the multidrug-resistant (MDR) L. monocytogenes strain BF11, previously isolated from raw pet food and phenotypically described for antimicrobial resistance. To this end, the genomic analysis performed on the isolate confirmed the pathogen’s designation as a serotype 1/2b strain belonging to ST5 and CC5 (Lineage I), carrying multiple MDR genes, stress-related genes, and mobile genetic elements, despite the absence of plasmids. The strain is phylogenetically closely related to Lineage I epidemic strains (e.g., F2365), as it has a full-length inlA and a functional prfA, rendering it capable of invading human cells and marking its high virulence. Overall, this strain may represent a potentially novel genomic profile when core genome multilocus sequence typing (cgMLST) is used, although further data from additional isolates would be required to confirm its classification within a new Complex Type, while displaying a hybrid unique profile. It is an evolved ST5 L. monocytogenes strain that has acquired genetic material conferring a “clinical signature” (Lineage I-like) and an extensive resistance network. Therefore, presence of L. monocytogenes strain BF11 in pet food is alarming, since such hybrid strains often evade surveillance monitoring as they do not fit strictly into classical categories, posing a serious food safety and public health threat in the concept of One Health.

Microorganisms doi: 10.3390/microorganisms14051095

Authors: Edisson Ronaldo Duran Yunga María de Lourdes Rodriguez Coyago

While a conserved core microbiome is shared across healthy individuals, significant interindividual taxonomic variation exists; however, the specific influence of genetic ancestry on supragingival plaque structure in eubiosis remains unclear. This systematic review analyzed evidence regarding taxonomic variations in supragingival plaque associated with ethnicity in systemically healthy populations. A search was conducted in PubMed, Scopus, ScienceDirect, and Scielo following PRISMA 2020 guidelines, covering literature up to October 2025. Cross-sectional studies using genomic sequencing or metagenomics were included, with quality assessed via the GRADE system. Six studies met eligibility criteria. Results identified a universal core microbiome structurally dominated by Corynebacterium spp. and Streptococcus spp. However, distinct ethnic-specific taxonomic signatures emerged, such as the enrichment of Fusobacterium spp. in African Americans and Corynebacterium spp. in Caucasians, alongside the exclusive presence of Sneathia spp. in Burmese individuals. Although a basal microbial architecture necessary for homeostasis exists, ethnicity acts as a biological filter defining distinctive bacterial profiles and differential susceptibilities. These findings suggest that while the core microbiome is conserved, the composition of peripheral species in the dental plaque hedgehog structure varies according to ancestry. This supports a transition from standardized dental care to personalized medicine oriented towards the patient’s biological heritage.

Microorganisms doi: 10.3390/microorganisms14051094

Authors: Dingxiang Lu Lin Liu Zhongwei Yang Tianjiao Chen Dong Yang Danyang Shi Shuqing Zhou Junwen Li Haibei Li Min Jin

Carbapenem-resistant Pseudomonas aeruginosa is increasingly becoming a global health threat. However, although aquatic environments are key resistance reservoirs, data obtained from high-altitude ecosystems are scarce. Whole-genome sequencing of eight carbapenem-resistant P. aeruginosa isolates collected from aquatic environments in the Tibetan Plateau identified three sequence types (STs), with ST1420 predominating (62.5%, 5/8). Phylogenetic analysis revealed a close clustering of isolates with those from distant clinical settings, suggesting potential cross-habitat transmission. All studied strains were multidrug-resistant, exhibiting 100% resistance to imipenem, ceftriaxone, and trimethoprim–sulfamethoxazole. This included the PA6 strain, which showed multiple-antibiotic resistance. Eight strains harbored the intrinsic carbapenemase gene blaOXA-50. The diverse virulence-gene profiles of strains PA2, PA4, and PA6 aligned with their high pathogenicity observed both in vitro and in vivo. However, virulence genotypes sometimes did not correlate with phenotypes, revealing the limitations of relying on static genetic information alone. This study highlights the aquatic environments of the Tibetan Plateau as reservoirs of carbapenem-resistant P. aeruginosa with substantial genetic diversity and divergent pathogenic potential, underscoring their public-health relevance.