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Microorganisms
Volume 14
Issue 3
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Microorganisms, Volume 14, Issue 3 (March 2026) – 213 articles

Cover Story (view full-size image): Herpes simplex virus type 1 (HSV-1) interacts dynamically with host cells within structured tissue microenvironments. Two-dimensional (2D) cultures do not fully capture these spatial features. Here, a three-dimensional (3D) culture platform was developed using the self-assembling peptide RADA16-I to form an extracellular matrix mimetic hydrogel scaffold. This system supported stable Vero cell spheroids viable for over 30 days. Following HSV-1 infection, viral dissemination progressed from the spheroid periphery to the core, with sustained replication for up to 22 days. Unlike 2D cultures, the 3D model exhibited distinct autophagic dynamics: LC3B-II increased while p62 decreased, indicating altered autophagic flux. This platform offers a physiologically relevant system for studying HSV-1 pathogenesis and host–virus interactions. View this paper
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13 pages, 268 KB  
Article
Association of TLR4 Polymorphisms with Increased Susceptibility to Recurrent Vulvovaginal Candidiasis in Greek Women
by Maria Mavrouli, Chrysoula Verra, Athanasios Tsakris and John Routsias
Microorganisms 2026, 14(3), 727; https://doi.org/10.3390/microorganisms14030727 - 23 Mar 2026
Viewed by 429
Abstract
Recurrent vulvovaginal candidiasis (RVVC) affects 5–8% of women of reproductive age. Host genetic factors, particularly single nucleotide polymorphisms (SNPs) in Toll-like receptors (TLRs), may influence RVVC susceptibility by impairing vaginal mucosal antifungal immunity. The aim of this study was to assess the effect [...] Read more.
Recurrent vulvovaginal candidiasis (RVVC) affects 5–8% of women of reproductive age. Host genetic factors, particularly single nucleotide polymorphisms (SNPs) in Toll-like receptors (TLRs), may influence RVVC susceptibility by impairing vaginal mucosal antifungal immunity. The aim of this study was to assess the effect of SNPs in genes encoding TLRs on RVVC susceptibility. Τhe distribution of TLR2 Arg753Gln and TLR4 Asp299Gly/Thr399Ile polymorphisms in Greek women, including RVVC (n = 63), first-episode VVC (n = 37), Gardnerella vaginalis vaginitis (GV, n = 36) patients, and healthy controls (n = 61), was investigated using TaqMan SNP genotyping. Genotype and allele frequencies were analyzed under allelic and dominant models, with odds ratios (ORs), 95% confidence intervals (CIs), and linkage disequilibrium assessed. TLR4 Asp299Gly and Thr399Ile heterozygotes were significantly more frequent in RVVC patients compared with controls and affected RVVC susceptibility (OR: 5.57, 95% CI: 1.17–26.56, p: 0.0172; OR: 4.92, 95% CI: 1.02–23.78, p: 0.0306, respectively). No associations were observed for TLR2 Arg753Gln or for any SNP with GV or first-episode VVC. TLR4 variants co-segregated, indicating a haplotype effect. TLR4 haplotypes, rather than TLR2 polymorphism, confer increased RVVC susceptibility, supporting a genetically distinct, mucosal immunity-driven pathogenesis. Larger, ethnically diverse studies with functional assays are warranted to validate these findings and guide personalized prevention and treatment strategies. Full article
(This article belongs to the Special Issue Laboratory Diagnosis and Epidemiology of Clinical Microbial Infections)
14 pages, 674 KB  
Review
Resistance of Uropathogens to Tebipenem: An Analysis of the Evidence from In Vitro Antimicrobial Susceptibility Studies
by Matthew E. Falagas, Christina-Maria Asimotou, Dimitrios S. Kontogiannis, Laura T. Romanos, Panagiota Poziou and Iva D. Tzvetanova
Microorganisms 2026, 14(3), 726; https://doi.org/10.3390/microorganisms14030726 - 23 Mar 2026
Viewed by 406
Abstract
Tebipenem is a new carbapenem antibiotic that binds to penicillin-binding proteins (PBPs). Given the need for effective antibiotics against multidrug-resistant (MDR) bacteria, this review evaluated the in vitro antimicrobial activity of tebipenem against Gram-negative and Gram-positive bacteria, focusing on uropathogens. Five resources (Google [...] Read more.
Tebipenem is a new carbapenem antibiotic that binds to penicillin-binding proteins (PBPs). Given the need for effective antibiotics against multidrug-resistant (MDR) bacteria, this review evaluated the in vitro antimicrobial activity of tebipenem against Gram-negative and Gram-positive bacteria, focusing on uropathogens. Five resources (Google Scholar, Web of Science, Embase, Scopus, and PubMed) were used to identify relevant articles. Of the 1322 articles identified, 9 relevant studies were included, which evaluated 12,501 Gram-negative and 122 Gram-positive pathogens. All nine studies (100%) assessed the activity of tebipenem against Escherichia coli, with an MIC90 value range of 0.015–>4 mg/L. Seven studies (77.8%) included Klebsiella pneumoniae, with an MIC90 value range of 0.015–0.5 mg/L. Six studies (66.7%) reported data on Proteus mirabilis, with an MIC90 value range of ≤0.125–0.5 mg/L. Two studies (22.2%) evaluated the activity of tebipenem against Enterococcus faecalis, with MIC90 of 1 mg/L among vancomycin-susceptible isolates and 32 mg/L in isolates with not-reported mechanisms of resistance. Two studies (22.2%) evaluated the activity of tebipenem against Enterococcus faecium, with MIC90 of >4 mg/L among both vancomycin-susceptible and vancomycin-resistant isolates and MIC90 of 128 mg/L among isolates with no resistance mechanism reported. Tebipenem demonstrated good activity against Enterobacterales, such as E. coli and K. pneumoniae. The antimicrobial agent exhibited higher MICs and a higher proportion of resistance among P. mirabilis isolates. Tebipenem could be effective for outpatient treatment of infections caused by MDR Gram-negative pathogens. However, given its potential to exert selective pressure for the development of antimicrobial resistance, it should be considered for patients with cUTIs when none of the first-line treatment options demonstrate in vitro antimicrobial activity. Full article
(This article belongs to the Special Issue Laboratory Diagnosis and Epidemiology of Clinical Microbial Infections)
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21 pages, 5259 KB  
Article
Monosodium Glutamate Inhibits Pseudomonas aeruginosa-Induced Acute Lung Injury by Targeting the Type III Secretion Systems and Modulating Host Immunity
by Jing Xu, Weiwei Wang, Yaxin Zhou, Hongxing Zhang, Zixuan Shang, Zhijin Zhang, Bing Li, Yubin Bai and Jiyu Zhang
Microorganisms 2026, 14(3), 725; https://doi.org/10.3390/microorganisms14030725 - 23 Mar 2026
Viewed by 314
Abstract
The opportunistic pathogen Pseudomonas aeruginosa poses a serious threat to immunocompromised patients. Monosodium glutamate (MSG), a widely used flavor enhancer, has been reported to possess anti-inflammatory and antioxidant properties. However, its therapeutic potential and mechanism against Pseudomonas aeruginosa (P. aeruginosa) infection [...] Read more.
The opportunistic pathogen Pseudomonas aeruginosa poses a serious threat to immunocompromised patients. Monosodium glutamate (MSG), a widely used flavor enhancer, has been reported to possess anti-inflammatory and antioxidant properties. However, its therapeutic potential and mechanism against Pseudomonas aeruginosa (P. aeruginosa) infection have remained unexplored. This study systematically elucidated the protective effects and molecular mechanisms of MSG against P. aeruginosa-induced acute lung injury (ALI). In a murine pneumonia model, MSG administration effectively alleviated lung pathological damage, edema, and inflammatory responses. Mechanistically, MSG exerted protection through a multifaceted strategy, including direct suppression of bacterial virulence via binding to PopB of T3SS inhibition of the TLR4/MyD88/MAPK-driven inflammatory cascade and pro-inflammatory cytokine production, enhancement of endogenous antioxidant defense (SOD, CAT), and reshaping of pulmonary macrophages from the M1 to M2 phenotype. Notably, the anti-virulence effect of MSG, achieved by binding to PopB (KD = 3.52 × 10−6 M), presented a distinct advantage over traditional antimicrobials by potentially mitigating resistance development. Collectively, these findings indicated that MSG can alleviate ALI caused by P. aeruginosa infection. Full article
(This article belongs to the Section Molecular Microbiology and Immunology)
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24 pages, 5263 KB  
Article
Post-Transcriptional Regulatory Mechanism Based on CsrA and rpoS in Extremophile Sulfur Oxidizer Acidithiobacillus caldus
by Yiwen Zhu, Panyan Chen, Hailin Yang, Yanjun Tong and Shoushuai Feng
Microorganisms 2026, 14(3), 724; https://doi.org/10.3390/microorganisms14030724 - 23 Mar 2026
Viewed by 302
Abstract
Acidithiobacillus caldus is perpetually exposed to multiple extreme environmental stresses. CsrA, functioning as a post-transcriptional regulator of physiological metabolism, acts as a differential modulator, facilitating more economical and efficient adaptation to extreme environments. The csrA expression recombinant strain was constructed in A. caldus [...] Read more.
Acidithiobacillus caldus is perpetually exposed to multiple extreme environmental stresses. CsrA, functioning as a post-transcriptional regulator of physiological metabolism, acts as a differential modulator, facilitating more economical and efficient adaptation to extreme environments. The csrA expression recombinant strain was constructed in A. caldus MTH-04 by conjugative transfer technology pJD215. Physiological characterization revealed enhanced acid tolerance, significantly elongated flagella, elevated extracellular secretion, and altered biofilm composition. Notably, intracellular concentrations of free glutamate and aspartate increased to 24.18 mg/L and 16.07 mg/L, respectively. The secondary structure of CsrA protein was determined in vitro through circular dichroism spectroscopy and size-exclusion chromatography. Electrophoretic Mobility Shift Assay (EMSA) successfully demonstrated in vitro binding activity of CsrA to the rpoS leader mRNA. CsrA suppresses rpoS mRNA translation by competing with ribosomes for binding sites, thereby negatively regulating rpoS expression. Critical binding sites were further validated through site-directed mutagenesis. Through EMSA, RT-qPCR and the translation reporter system, it was also found that CsrA has a dual regulatory function for nearby flagella- and motility-related gene clusters (flgC, 07035, motD, 15040), which also implies the global regulatory role of CsrA. In summary, a potential overall post-transcriptional regulatory mechanism based on CsrA and rpoS by extremophile A. caldus was proposed. Finally, the efficiency of bioleaching application by csrA overexpression strain was improved by 20.81%. Full article
(This article belongs to the Special Issue Resource Utilization of Microorganisms: Fermentation and Biosynthesis)
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20 pages, 6905 KB  
Article
Comparative Enzymology and Biomass Hydrolysis Reveal Industrial Biorefining Potential of Aspergillus fumigatus Strain VP2T
by Vaniksha Pal, Punam Vishwakarma, Dipayan Samanta, Priya Saxena, Rohit Rai and Rajesh K. Sani
Microorganisms 2026, 14(3), 723; https://doi.org/10.3390/microorganisms14030723 - 23 Mar 2026
Viewed by 360
Abstract
We report on the isolation and comprehensive genomic and biochemical characterization of Aspergillus fumigatus VP2T, a thermophilic filamentous fungus recovered from Himalayan Forest soil with exceptional lignocellulolytic capacity. Whole-genome sequencing revealed a 32.1 Mb genome encoding 12,675 predicted genes, including an extensive repertoire [...] Read more.
We report on the isolation and comprehensive genomic and biochemical characterization of Aspergillus fumigatus VP2T, a thermophilic filamentous fungus recovered from Himalayan Forest soil with exceptional lignocellulolytic capacity. Whole-genome sequencing revealed a 32.1 Mb genome encoding 12,675 predicted genes, including an extensive repertoire of >300 carbohydrate-active enzymes (CAZymes). Notably, the genome harbors multiple auxiliary activity enzymes, including AA9-family lytic polysaccharide monooxygenases and several cellobiose dehydrogenases (CDHs), supporting oxidative–hydrolytic synergism during biomass degradation. Submerged fermentation using a cellulose–wheat bran–rice straw substrate induced high enzyme titers, including 33 U/mL endoglucanase and 131 U/mL CDH, exceeding activities commonly reported for both native and engineered fungal strains. Although exoglucanase (0.02 U/mL) and xylanase (14.22 U/mL) activities were comparatively modest, the strain VP2T demonstrated superior hydrolysis of untreated rice straw, achieving a 1.89-fold increase in saccharification efficiency relative to the commercial enzyme cocktail Cellic® CTec2. Scanning electron microscopy confirmed extensive disruption of lignocellulosic architecture, consistent with enhanced enzyme accessibility and oxidative fiber loosening. Collectively, genomic evidence and functional assays identify A. fumigatus VP2T as a redox-optimized, moderately thermophilic biocatalyst suited for low-pH lignocellulose conversion. This study highlights the value of exploring thermophilic fungal biodiversity to discover native strains with inherent oxidative capacity, offering promising alternatives to pretreatment-intensive biorefinery processes and informing the rational development of tailored enzyme systems. Full article
(This article belongs to the Special Issue Advances in Aspergillus and Aspergillosis)
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15 pages, 4009 KB  
Article
Effects of Microbial Inoculants from Three Nutrient-Poor Environments on Soil Improvement and Plant Growth Promotion in Sandy Soil
by Xin Sun, Xuanran Yu, Xingyu Zhang, Xinxin Yang, Rengui Xue, Aodeng Rong, Xin Liu, Xiongfei Zhang, Chong Li and Jinchi Zhang
Microorganisms 2026, 14(3), 722; https://doi.org/10.3390/microorganisms14030722 - 23 Mar 2026
Viewed by 389
Abstract
Approximately 20% of China’s land area is desertified or highly desertifiable, where loose sandy soil and low nutrient availability restrict plant growth. Microbial inoculants, as an emerging ecological restoration technology, play a key role in plant growth and soil nutrient activation in sandy [...] Read more.
Approximately 20% of China’s land area is desertified or highly desertifiable, where loose sandy soil and low nutrient availability restrict plant growth. Microbial inoculants, as an emerging ecological restoration technology, play a key role in plant growth and soil nutrient activation in sandy regions. However, a systematic understanding of functional differences among microorganisms isolated from different stressed environments remains insufficient. Nine functional microbial strains from three stressed habitats, including sandy land, coastal saline-alkali soil, and heavy metal mining areas, were selected to conduct a three-month pot experiment, investigating their effects on soil nutrient activation, plant growth and microbial communities. Results showed that all inoculants increase plant biomass (by 4.15~25.59%), with KS-33, KS-36, SD-13 and SD-3 significantly promoting biomass in different plant parts (p < 0.05), and with YJ-15 remarkably enhancing root growth (root length increased by 70.83%, p < 0.01). Inoculation reduced bacterial Chao1 by 27.18~53.97%, but increased fungal Chao1 by 12.77~28.38% (except SD-30). Bacterial generalist species proportion increased from 61.12% to 83.78~93.99% after inoculation, higher than the variation degree of the fungal community. Mantel analysis revealed a reverse trend between soil nutrients, water content and plant growth. This may be associated with the increased consumption by plants and microorganisms. In summary, microbial inoculants enhance nutrient cycling processes and plant growth by reshaping soil microbial communities. Performance of microbial inoculants is more likely governed by their inherent ecological functions rather than being entirely determined by their original environments. Despite varying mechanisms, these inoculants can effectively enhance sandy soil microbial communities, providing a theoretical basis for regional ecological restoration. Full article
(This article belongs to the Section Environmental Microbiology)
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19 pages, 905 KB  
Review
Review of Infections in Immunocompromised Travellers: Epidemiology, Infection Prevention and Management
by Anca-Elena Duduveche, Andreea-Lacramioara Mohorea-Neata and Sorina-Nicoleta Badelita
Microorganisms 2026, 14(3), 721; https://doi.org/10.3390/microorganisms14030721 - 23 Mar 2026
Viewed by 541
Abstract
International travel has increased significantly in recent years. Many travellers are immunocompromised, making them more susceptible to infectious diseases due to their weakened immune systems. This narrative review provides an overview of infection epidemiology, prevention strategies, and clinical management for immunocompromised travellers. We [...] Read more.
International travel has increased significantly in recent years. Many travellers are immunocompromised, making them more susceptible to infectious diseases due to their weakened immune systems. This narrative review provides an overview of infection epidemiology, prevention strategies, and clinical management for immunocompromised travellers. We demonstrate the role of pre-travel consultation, emphasizing individualized risk assessment informed by destination-specific epidemiology, immune status, and underlying conditions. Key preventive interventions, including vaccination strategies, antimicrobial prophylaxis, and detailed health counseling, in addition to practical measures that can be taken to minimize infection risk during travel, are addressed. Post-travel surveillance and early recognition of travel-related infections are analyzed, highlighting the importance of prompt symptom evaluation. Effective care for immunocompromised travellers requires a multidisciplinary framework that integrates patients, primary care physicians, infectious disease specialists, and travel medicine experts. Full article
(This article belongs to the Special Issue The Molecular Epidemiology of Infectious Diseases, 2nd Edition)
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18 pages, 1712 KB  
Review
Strain Diversity in the Human Microbiome: Personal Variation, Pathobionts, Therapeutics, and Methodological Challenges
by Hyunjoon Park, Jung Soo Kim, Dong Joon Kim and Ki Tae Suk
Microorganisms 2026, 14(3), 720; https://doi.org/10.3390/microorganisms14030720 - 23 Mar 2026
Viewed by 426
Abstract
Advances in sequencing technologies have transformed human microbiome research, yet most analyses still rely on species-level profiles. However, strains rather than species represent the true ecological and functional units of the microbiome. Individual strains can vary substantially in gene content, metabolic capacity, virulence [...] Read more.
Advances in sequencing technologies have transformed human microbiome research, yet most analyses still rely on species-level profiles. However, strains rather than species represent the true ecological and functional units of the microbiome. Individual strains can vary substantially in gene content, metabolic capacity, virulence factors, antimicrobial resistance, and host-interaction properties. These differences critically influence immune responses, epithelial barrier integrity, disease susceptibility, and therapeutic outcomes. Here, we synthesize recent human microbiome studies that provide robust strain-resolved evidence, focusing on three major themes: (i) the emergence and long-term persistence of personalized strain repertoires, (ii) strain-specific pathobiont traits that drive host pathology, and (iii) the implications of strain-level ecology for the development of next-generation microbiome therapeutics. We also highlight key methodological innovations including high-resolution amplicon profiling, advanced metagenomic and single-cell genomics, and culture-based functional approaches that collectively enable strain-level resolution and are reshaping the field. Full article
(This article belongs to the Section Gut Microbiota)
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25 pages, 2240 KB  
Review
Seeding the Future: How Feeding Mode Shapes the Infant Gut Microbiota
by Felicia Trofin, Aida Corina Badescu, Luminita Smaranda Iancu, Elena Roxana Buzila, Dana-Teodora Anton-Păduraru, Cristina Mihaela Sima, Oana-Raluca Temneanu, Anca Matei, Stefana Catalina Bilha, Ioana Alexandra Benea and Olivia Simona Dorneanu
Microorganisms 2026, 14(3), 719; https://doi.org/10.3390/microorganisms14030719 - 23 Mar 2026
Viewed by 590
Abstract
Early life represents a critical developmental programming window during which nutrition and microbial exposures shape long-term physiological function. Feeding mode is a major determinant of infant gut microbiota assembly and metabolic activity. This narrative review synthesizes current evidence comparing breastfeeding (BF) and formula [...] Read more.
Early life represents a critical developmental programming window during which nutrition and microbial exposures shape long-term physiological function. Feeding mode is a major determinant of infant gut microbiota assembly and metabolic activity. This narrative review synthesizes current evidence comparing breastfeeding (BF) and formula feeding in relation to microbial composition, functional capacity, and immune programming during the preweaning and early postweaning periods. BF may support a relatively stable, bifidobacteria-dominated microbiota enriched in pathways involved in carbohydrate utilization, vitamin biosynthesis, and immune modulation. Human milk oligosaccharides, secretory IgA, lactoferrin, and milk-associated microbes collectively guide microbial succession, enhance barrier integrity, and support immune tolerance. In contrast, formula-fed infants typically exhibit greater microbial diversity, earlier transition toward adult-like profiles, and increased abundance of facultative anaerobes, alongside the enrichment of pathways related to bile acid and amino acid metabolism. Microbiota patterns in formula-fed infants are further influenced by formula composition, including protein load, lipid structure, and supplementation with prebiotics, probiotics, and human milk oligosaccharide analogues. Although advances in formula design have reduced compositional gaps, functional differences in microbial stability and immune programming persist. Recognizing early infancy as a sensitive programming window underscores the need for microbiome-informed nutritional strategies and longitudinal, multi-omics research to clarify causal mechanisms and optimize early-life interventions. Full article
(This article belongs to the Special Issue Milk, Microbes, and Medicine: The Triad Shaping Infant Health)
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13 pages, 3615 KB  
Article
Impact of Transgenic Insect-Resistant Maize LD05 on Rhizosphere Soil Bacterial Communities
by Wenlan Li, Xiaohui Xu, Xinwei Hou and Runqing Yue
Microorganisms 2026, 14(3), 718; https://doi.org/10.3390/microorganisms14030718 - 23 Mar 2026
Viewed by 267
Abstract
The artificially modified Bacillus thuringiensis (Bt) protein can target lepidopteran pests, and planting genetically modified crops with insect-resistant traits is environmentally friendly. However, it is still uncertain whether the exogenous insect-resistant proteins in genetically modified crops will affect the soil rhizosphere microorganisms. This [...] Read more.
The artificially modified Bacillus thuringiensis (Bt) protein can target lepidopteran pests, and planting genetically modified crops with insect-resistant traits is environmentally friendly. However, it is still uncertain whether the exogenous insect-resistant proteins in genetically modified crops will affect the soil rhizosphere microorganisms. This study utilized 16S rDNA sequencing technology to analyze the rhizosphere soil of insect-resistant genetically modified corn LD05 and its control variety Zheng58 at five developmental stages: before sowing, seedling stage, jointing stage, silk emergence stage, and maturity stage. Each sample was taken with six biological replicates, resulting in a total of 60 sequencing samples, with an average of 4368 OTUs obtained per sample. Both alpha and beta analyses showed that LD05 and Zheng58 did not have a significant impact on the soil rhizosphere microbial community. The developmental stage rather than the variety was the main factor causing differences in the bacterial community. Overall, there was no significant difference in the bacterial diversity between the insect-resistant genetically modified corn LD05 and its control variety Zheng58. The results provide useful information for understanding the impact of genetically modified crops on soil microbial communities and also provide a theoretical basis for the safety evaluation of LD05. Full article
(This article belongs to the Section Environmental Microbiology)
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3 pages, 130 KB  
Editorial
Editorial for Special Issue “Animal Viral Infectious Diseases”
by He Zhang
Microorganisms 2026, 14(3), 717; https://doi.org/10.3390/microorganisms14030717 - 23 Mar 2026
Viewed by 342
Abstract
Animal viral infectious diseases continue to pose a persistent threat to the development of global animal husbandry and animal health [...] Full article
(This article belongs to the Special Issue Animal Viral Infectious Diseases)
4 pages, 182 KB  
Editorial
Editorial for “Gut Microbiota, Diet, and Gastrointestinal Cancer”
by Marco Cintoni and Flavio De Maio
Microorganisms 2026, 14(3), 716; https://doi.org/10.3390/microorganisms14030716 - 23 Mar 2026
Viewed by 346
Abstract
Gastrointestinal (GI) cancers remain a leading cause of morbidity and mortality globally, and their etiology is recognized as a multifactorial process driven by the interplay of genetic susceptibility, environmental exposures, and lifestyle-related factors [...] Full article
(This article belongs to the Special Issue Gut Microbiota, Diet, and Gastrointestinal Cancer)
15 pages, 2112 KB  
Article
Enhancing Synthesis Efficiency in Microbial 1,5-Pentanediol Production Through Transcriptomics-Informed Metabolic Engineering of Escherichia coli
by Hongyu Deng, Fei Meng, Yihao Sun, Yang Song, Chunhui Zhao, Xiaonan Wang, Yan Zhang, Ruiming Wang and Ning Chen
Microorganisms 2026, 14(3), 715; https://doi.org/10.3390/microorganisms14030715 - 22 Mar 2026
Viewed by 336
Abstract
The microbial production of 1,5-pentanediol (1,5-PDO), a versatile platform chemical with extensive industrial applications, remains limited by suboptimal fermentation titers and incomplete understanding of metabolic bottlenecks. To address these challenges, this study employed comparative transcriptomics to systematically identify novel genetic targets capable of [...] Read more.
The microbial production of 1,5-pentanediol (1,5-PDO), a versatile platform chemical with extensive industrial applications, remains limited by suboptimal fermentation titers and incomplete understanding of metabolic bottlenecks. To address these challenges, this study employed comparative transcriptomics to systematically identify novel genetic targets capable of enhancing 1,5-PDO biosynthesis in engineered Escherichia coli. Transcriptomic profiling of the 1,5-PDO-producing strain relative to the parental E. coli W3110, conducted at both exponential (24 h) and stationary (96 h) growth phases, revealed 1384 significantly differentially expressed genes, including 851 upregulated and 533 downregulated genes. From these, 20 candidate metabolic genes associated with 1,5-PDO synthesis were selected for functional validation through plasmid-based overexpression or CRISPR interference (CRISPRi)-mediated repression. Reverse engineering confirmed that overexpression of fecA (encoding an iron(III)-citrate transporter) and deletion of gadA (encoding glutamate decarboxylase) significantly enhanced 1,5-PDO production. Subsequent chromosomal integration of fecA at the neutral ilvG locus and deletion of gadA generated the optimized strain S7, which achieved a 1,5-PDO titer of 1.7 g/L in shake flask cultures, representing a 13.3% increase over the parental strain, with a concomitant 50% improvement in glucose yield (0.18 mol/mol). In fed-batch fermentation at the 5 L bioreactor scale, strain S7 attained a titer of 12.45 g/L and a glucose yield of 0.26 mol/mol, marking a 15.6% enhancement in carbon conversion efficiency relative to the parental strain (0.225 mol/mol), while concurrently improving biomass accumulation by 7.6%. These findings demonstrate that transcriptomics-guided reverse engineering constitutes an effective strategy for elucidating nonobvious metabolic determinants and optimizing microbial cell factories for efficient 1,5-PDO production. The identification of fecA and gadA as beneficial targets provides valuable insights into the metabolic rewiring underlying enhanced 1,5-PDO biosynthesis and establishes a foundation for further strain improvement through systems metabolic engineering. Full article
(This article belongs to the Special Issue Resource Utilization of Microorganisms: Fermentation and Biosynthesis)
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13 pages, 2408 KB  
Article
Nematicidal Potential of Purpureocillium takamizusanense PMEPF27 Against Motile Bursaphelenchus rainulfi In Vitro
by Yuh Tzean, Elena Gamboa Chen, Xiao-Yu Wei, I-En Shih, Hui-Yu Hsu, Ya-Zhen Xu, Ying-Hong Lin, Meng-Ling Wu, Tai-Yuan Chen, Yung-Yu Yang and Jen-Chih Chen
Microorganisms 2026, 14(3), 714; https://doi.org/10.3390/microorganisms14030714 - 22 Mar 2026
Viewed by 288
Abstract
Bursaphelenchus rainulfi is a pine-associated, non-pathogenic nematode that serves as a motile comparative species for evaluating nematophagous fungi. We investigated the in vitro biocontrol activity of Purpureocillium takamizusanense strain PMEPF27, originally isolated from insect cadavers in Taiwan, against mixed motile stages of B. [...] Read more.
Bursaphelenchus rainulfi is a pine-associated, non-pathogenic nematode that serves as a motile comparative species for evaluating nematophagous fungi. We investigated the in vitro biocontrol activity of Purpureocillium takamizusanense strain PMEPF27, originally isolated from insect cadavers in Taiwan, against mixed motile stages of B. rainulfi. Identity of the fungus was confirmed by morphology and ITS/EF-1α sequencing. Nematodes were exposed in liquid suspension to PMEPF27 conidia, with sterile water as the negative control and fluopyram as the positive control. Mortality was monitored over 7 days, and scanning electron microscopy was used to observe fungus–nematode interactions. PMEPF27 caused significantly higher mortality than the water control, reaching ~80% by day 7, and showed surface disruption of nematode cuticles, although no direct spore penetration was observed. These findings expand the nematophagous profile of P. takamizusanense beyond egg and sedentary stages, validate B. rainulfi as a motile comparative species in pine-nematode bioassays, and highlight PMEPF27 as a promising candidate for biocontrol development. Full article
(This article belongs to the Special Issue Biological Control of Microbial Pathogens in Plants)
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18 pages, 2202 KB  
Article
The Microbial Network Stability in Cyanobacterial and Moss Biocrusts Respond Differently to Climate Warming
by Chang Tian, Chongfeng Bu, Shufang Wu, Xinhao Li and Kadambot H. M. Siddique
Microorganisms 2026, 14(3), 713; https://doi.org/10.3390/microorganisms14030713 - 22 Mar 2026
Viewed by 399
Abstract
Climate warming—a key driver of global change—significantly affects soil microbial communities and network stability. Biological soil crusts (biocrusts) help mitigate these impacts while maintaining soil ecological functions and biodiversity. However, how microbial networks and community dynamics respond to warming remains poorly understood between [...] Read more.
Climate warming—a key driver of global change—significantly affects soil microbial communities and network stability. Biological soil crusts (biocrusts) help mitigate these impacts while maintaining soil ecological functions and biodiversity. However, how microbial networks and community dynamics respond to warming remains poorly understood between biocrust types, namely cyanobacterial and moss biocrust. In this study, we investigated the effect of warming on microbial communities and network stability in these biocrusts within the Mu Us Sandland, China. Using structural equation modeling (SEM), we found that warming altered microbial network properties: compared to the control, warming increased network vulnerability and decreased robustness specifically in cyanobacterial biocrusts. Warming and decreased soil moisture acted as strong filtering factors, resulting in lower microbial network stability. Although overall network complexity remained unchanged, warming reduced connectivity in cyanobacterial biocrusts, thus undermining network stability. Moreover, under both warming and control conditions, moss biocrusts exhibited lower robustness but higher vulnerability than cyanobacterial biocrusts, indicating cyanobacterial biocrusts displayed greater microbial network stability in comparison. Additionally, warming reduced the number of module hubs and keystone phyla in both biocrust types, decreasing key taxa abundance and weakening direct microbial interactions. We concluded that warming impaired microbial network stability by reducing connectivity in cyanobacterial biocrusts. These findings highlight the superior capacity of cyanobacterial biocrusts to sustain soil microbial network stability under climate warming and identify shifts in network connectivity as a central mechanism driving biocrust responses to environmental stress. Full article
(This article belongs to the Section Environmental Microbiology)
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24 pages, 351 KB  
Article
One Health Investigation of Stage-Dependent Antimicrobial Resistance Patterns Across Intermediate and Ripened Dairy Matrices: The Tyrovolia–Kopanisti Paradigm
by Georgios Rozos, Konstantina Fotou, Vaia Gerokomou, Konstantina Nikolaou, Aikaterini Dadamogia, Lampros Hatzizisis, Ioannis Skoufos, Athina Tzora, Eugenia Bezirtzoglou and Chrysoula (Chrysa) Voidarou
Microorganisms 2026, 14(3), 712; https://doi.org/10.3390/microorganisms14030712 - 22 Mar 2026
Viewed by 380
Abstract
Antimicrobial resistance (AMR) emerges and circulates across interconnected human, animal, food, and environmental reservoirs; however, food fermentation systems remain underexplored as indicators of local AMR pressure, even though artisanal dairy fermentations may function as natural sentinels of AMR. In this study, we used [...] Read more.
Antimicrobial resistance (AMR) emerges and circulates across interconnected human, animal, food, and environmental reservoirs; however, food fermentation systems remain underexplored as indicators of local AMR pressure, even though artisanal dairy fermentations may function as natural sentinels of AMR. In this study, we used an artisanal dairy fermentation chain as a One Health model to investigate whether environmentally exposed lactobacilli can reflect stage-associated shifts in resistance. A total of 1.085 isolates representing 16 Lactobacillus species were recovered from the same artisanal dairy matrix at two fermentation stages: day 5 (“Tyrovolia”; n = 518) and day 30 (“Kopanisti”; n = 567). Susceptibility to 14 antibiotics was evaluated by broth micro-dilution, and L. acidophilus was further screened for selected resistance genes. Overall resistance increased significantly from 69.88% (362/518) at day 5 to 77.25% (438/567) at day 30 (p = 0.0059), while multidrug resistance rose from 37.57% to 60.73% of resistant isolates (p < 0.001). Across the 224 species–antibiotic combinations examined, 129 (57.58%) showed an increased upper MIC limit at day 30, and resistance increased significantly for 9 of the 14 antibiotics tested, with the largest rises observed for metronidazole (RR = 7.67), chloramphenicol (RR = 5.74), quinupristin/dalfopristin (RR = 4.11), vancomycin (RR = 2.78), and trimethoprim (RR = 2.43). In contrast, erythromycin and oxytetracycline resistance declined significantly at the ripened stage. In L. acidophilus, 21 resistance genes were detected in 14/70 day-5 isolates and 19 genes in 13/71 day-30 isolates, but marked genotype–phenotype discordance was observed, including matrix-dependent expression patterns for tetM, ermB, and blaTEM. Collectively, these findings show that environmentally exposed artisanal dairy fermentations can enrich resistance phenotypes and may serve as sensitive sentinels of AMR dynamics at the human–animal–environment interface. Full article
(This article belongs to the Special Issue Microbial Safety and Beneficial Microorganisms in Foods)
10 pages, 1176 KB  
Article
A Proof-of-Concept of a 2-Hours Direct Antimicrobial Susceptibility Test from Inoculated Urine Samples
by Mariana Sousa-Pinheiro, Inês Martins-Oliveira, David Abreu, Rosário Gomes, Ana Silva-Dias, Patrícia Poeta, Cidália Pina-Vaz and António José Soares
Microorganisms 2026, 14(3), 711; https://doi.org/10.3390/microorganisms14030711 - 22 Mar 2026
Viewed by 269
Abstract
Urinary tract infections (UTIs) are the most frequent infections in hospitalized and outpatient settings, where Escherichia coli is the predominant pathogen. Conventional diagnostic and antimicrobial susceptibility testing (AST) methods are time-consuming, often requiring 48 h, leading to empirical antibiotic therapy and contributing to [...] Read more.
Urinary tract infections (UTIs) are the most frequent infections in hospitalized and outpatient settings, where Escherichia coli is the predominant pathogen. Conventional diagnostic and antimicrobial susceptibility testing (AST) methods are time-consuming, often requiring 48 h, leading to empirical antibiotic therapy and contributing to antimicrobial resistance (AMR). FASTinov® developed a rapid phenotypic method that enables AST directly from urine samples within two hours using flow cytometry. In this study, 154 inoculated urine samples were analyzed to evaluate the performance of two diagnostic panels: FASTgramneg for Gram-negative bacteria and FASTgrampos for Gram-positive bacteria. Data analysis was performed using bioFAST® software (version 3.0), providing results in accordance with EUCAST guidelines. The FASTgramneg panel allows detection of resistance mechanisms, including extended-spectrum β-lactamases (ESBLs), and screening of AmpC β-lactamases and carbapenemases; the FASTgrampos panel additionally determines the minimal inhibitory concentration (MIC) of vancomycin for Staphylococcus aureus. Overall agreement with conventional AST methods was 97.5% for Gram-negative bacteria and 95.0% for Gram-positive bacteria. All resistance mechanisms were correctly identified with no false positives. The essential agreement for vancomycin’s MIC was 95.2%, with a BIAS of +14.3%. Reproducibility was 99.5% for FASTgramneg and 95.0% for FASTgrampos. These results demonstrate that the FASTinov® kit significantly reduces turnaround time while maintaining high accuracy, supporting improved UTI management and antimicrobial stewardship. Full article
(This article belongs to the Section Antimicrobial Agents and Resistance)
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3 pages, 153 KB  
Editorial
Editorial for the Special Issue “Water Microorganisms Associated with Human Health, 2nd Edition”
by Ana Machado
Microorganisms 2026, 14(3), 710; https://doi.org/10.3390/microorganisms14030710 - 22 Mar 2026
Viewed by 278
Abstract
Water-associated microorganisms have re-emerged as a central public health concern, reflecting how microbial dynamics in water reflect current social, climatic, and infrastructural pressures [...] Full article
(This article belongs to the Special Issue Water Microorganisms Associated with Human Health, 2nd Edition)
18 pages, 1584 KB  
Article
Development and Validation of a Quantitative RT-qPCR Panel for the Detection and Monitoring of Polioviruses in Wastewater Samples
by Linnet Immaraj, Judy Y. Qiu, Logan A. Brand, Tiejun Gao, Bonita Lee, Michael Parkins, Casey Hubert, Christine O’Grady and Xiaoli Pang
Microorganisms 2026, 14(3), 709; https://doi.org/10.3390/microorganisms14030709 - 21 Mar 2026
Viewed by 370
Abstract
Clusters of acute flaccid paralysis (AFP) caused by oral vaccine-derived poliovirus (VDPV) in 2022 and sporadic outbreaks in New York and Gaza highlight the ongoing risk of polio, alongside the persistent global threat posed by wild-type poliovirus. This study aims to develop and [...] Read more.
Clusters of acute flaccid paralysis (AFP) caused by oral vaccine-derived poliovirus (VDPV) in 2022 and sporadic outbreaks in New York and Gaza highlight the ongoing risk of polio, alongside the persistent global threat posed by wild-type poliovirus. This study aims to develop and validate a quantitative reverse transcription PCR (RT-qPCR) panel that employs different primer–probe sets to simultaneously detect vaccine and wild-type poliovirus (WPV) in wastewater. Using an inactivated poliovirus vaccine (IPV) and engineered DNA fragments (eDNAf), the qPCR master mix (MM) performance, assay specificity, sensitivity (limit of detection, LOD), and recovery from IPV-spiked wastewater were evaluated. Compared with two-step RT-qPCR and qScript MM, one-step RT-qPCR with TaqMan MM improved sensitivity for the following polioviruses (PV): Sabin 1 in IPV and the eDNAf of Sabin 1, 2, and 3; WPV1 and WPV3; and poliovirus type 2 (any serotype 2). The LOD for Sabin 1 in IPV was 2.49 copies/PCR, while LODs for eDNAf of polio targets ranged from 1.06 to 3.12 copies/PCR. Sabin 1 recovery from IPV-spiked wastewater ranged from 10.26% to 57.27%. The RT-qPCR panel for poliovirus exhibited good specificity and sensitivity, with moderate viral recovery, enabling rapid implementation of wastewater monitoring for PV as needed. Full article
(This article belongs to the Special Issue Pathogen Surveillance in Wastewater)
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37 pages, 15545 KB  
Article
Comparative Analysis of the Intestinal Microbiota in Wild and Aquaculture Populations of Sparus aurata
by Maria Lanara, Elias Asimakis, Naima Bel Mokhtar, Pinelopi Koutsodima, Costas Batargias, Kosmas Toskas, Panagiota Stathopoulou and George Tsiamis
Microorganisms 2026, 14(3), 708; https://doi.org/10.3390/microorganisms14030708 - 21 Mar 2026
Viewed by 768
Abstract
Fish host complex intestinal bacterial communities that contribute to a wide range of functions, from nutrient assimilation to modulation of the immune system. Understanding how environmental and host-related factors shape the fish gut microbiota is essential for advancing sustainable aquaculture practices. This study [...] Read more.
Fish host complex intestinal bacterial communities that contribute to a wide range of functions, from nutrient assimilation to modulation of the immune system. Understanding how environmental and host-related factors shape the fish gut microbiota is essential for advancing sustainable aquaculture practices. This study compared the intestinal microbiota of gilthead sea bream (Sparus aurata) between wild and aquaculture populations in western Greece using 16S rRNA gene amplicon sequencing targeting the V3–V4 region, combined with culture-based methods. The analysis was based on a 97% similarity threshold and included 141 gastrointestinal samples of fish collected at two aquaculture facilities and two wild fisheries, representing two different growth phases (150 g and 300 g body weight). High-throughput sequencing data revealed a clear separation of gut microbial communities according to origin (wild vs. aquaculture), geographic location, and body growth phase, with most wild fish groups exhibiting higher microbial diversity than their farmed counterparts, except for group MES_150 which showed similar or lower values. The gut microbiota was dominated by Pseudomonadota (53%), Bacillota (29%), Actinomycetota (7%), Deinococcota (5%), and Bacteroidota (4%). A shared core microbiome, comprising Psychrobacter, Staphylococcus, Geobacillus, Aeromonas, Enterobacter, Pantoea, Bacillus, and Acinetobacter, was detected across all populations. Wild fish were enriched in Psychrobacter, Aeromonas, and Photobacterium, while aquaculture fish displayed higher abundances of Vibrio, Allomeiothermus, and Staphylococcus. Network analysis revealed mostly mutually exclusive interactions in both groups but distinct patterns of co-occurrence, driven mainly by Paenibacillus, Enterobacter, and Staphylococcus in wild samples, and by Vibrio, Aeromonas, and Pseudomonas in farmed fish. Culture-based assays demonstrated greater diversity in wild fish, dominated by Pseudomonas, Staphylococcus, and Vibrio strains, in contrast to the frequent occurrence of Staphylococcus and Psychrobacter in aquaculture samples. The findings suggest that aquaculture practices significantly alter gut microbial community structure and reduce diversity, with potential implications for fish health and disease resistance. The identified core and differentially abundant taxa provide candidates for probiotic development to improve aquaculture sustainability. Full article
(This article belongs to the Special Issue Pathogenomics for Sustainable Aquaculture: From Diagnostics to Disease Resistance)
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21 pages, 4808 KB  
Article
Hungatella hathewayi: A Tumor-Derived Bacterium Enriched in Colorectal Cancer Tissues and a Potential Diagnostic Biomarker
by Wenzhe Zhang, Jin Liu, Shanshan Sha, Qiulong Yan and Yufang Ma
Microorganisms 2026, 14(3), 707; https://doi.org/10.3390/microorganisms14030707 - 21 Mar 2026
Viewed by 391
Abstract
Colorectal cancer (CRC) is the third most common cancer globally and the second leading cause of cancer-related deaths. While intestinal microbiota dysbiosis is linked to CRC, the direct role of intratumoral bacteria in metastasis remains poorly understood. In this study, we isolated pathogenic [...] Read more.
Colorectal cancer (CRC) is the third most common cancer globally and the second leading cause of cancer-related deaths. While intestinal microbiota dysbiosis is linked to CRC, the direct role of intratumoral bacteria in metastasis remains poorly understood. In this study, we isolated pathogenic bacteria from CRC tumor tissues, identified as Hungatella hathewayi (H. hathewayi), through the 16S rRNA gene and whole-genome sequencing. We developed specific primers (P48/P52) and polyclonal antibodies for detecting H. hathewayi in samples. Using quantitative real-time PCR (qPCR), we found significant enrichment of H. hathewayi in fecal samples from CRC patients compared to healthy controls, with mean fold changes of 137-fold and 142-fold for primers P48 and P52, respectively. Analysis of tissue samples revealed that H. hathewayi abundance was higher in CRC tumor tissues compared to normal tissues, with mean fold changes of 2.90 for P48 and 3.97 for P52. Fluorescence in situ hybridization (FISH), immunofluorescence (IF), and immunohistochemistry (IHC) confirmed its spatial distribution within tumor tissues. In vitro assays using CRC cell lines demonstrated that H. hathewayi-derived succinate upregulates HIF-1α and SUCNR1 expression and promotes cell metastasis by inducing epithelial–mesenchymal transition (EMT). Collectively, these findings identify H. hathewayi as a novel pro-metastatic bacterium and a potential non-invasive biomarker for CRC diagnosis, providing direct evidence for the role of intratumoral bacteria in CRC progression. Full article
(This article belongs to the Section Gut Microbiota)
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17 pages, 1109 KB  
Review
African Swine Fever: Vaccine Advancement and Major Gaps
by Lihua Wang and Jishu Shi
Microorganisms 2026, 14(3), 706; https://doi.org/10.3390/microorganisms14030706 - 21 Mar 2026
Viewed by 544
Abstract
African swine fever (ASF), a highly contagious and lethal viral disease caused by the African swine fever virus (ASFV), poses a severe threat to the global swine industry. Recent outbreaks across Asia, Europe, and the Caribbean are exacerbating the challenge. Current control measures [...] Read more.
African swine fever (ASF), a highly contagious and lethal viral disease caused by the African swine fever virus (ASFV), poses a severe threat to the global swine industry. Recent outbreaks across Asia, Europe, and the Caribbean are exacerbating the challenge. Current control measures rely mainly on early detection, culling and strict biosecurity practices, underscoring the urgent need for a safe and effective vaccine. Since the mid-1960s, diverse vaccine strategies, including inactivated, subunit, DNA/mRNA, vectored, and live attenuated virus (LAV) vaccines, have been explored. Inactivated vaccines have consistently failed to confer protection due to insufficient functional antigen presentation and weak cellular immune activation. Subunit vaccines targeting single or multiple ASFV antigens have also shown limited success, often failing to induce sterile or long-lasting immunity. Among these approaches, LAV vaccines have demonstrated the greatest promise in eliciting robust and durable immune responses. However, major knowledge gaps remain regarding ASFV biology, ASFV–host interactions, ASFV immune evasion mechanisms, protective and cross-protective immunity, stable cell lines for LAV production, virulence reversion of LAVs, and the lack of harmonized standards for evaluating vaccine safety and efficacy, all of which impede the development of safe and broadly effective ASF vaccines. This narrative review summarizes recent advances in ASF vaccine research and highlights the critical obstacles that must be overcome to achieve successful ASF vaccine development. Full article
(This article belongs to the Special Issue Virulence-Associated Genes, Pathogenesis and Immunoevasion Mechanisms of African Swine Fever Virus)
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39 pages, 5196 KB  
Article
Harnessing the Enzymatic Potential of Indigenous Yeast Strains: Screening and Evaluation for Biocontrol and Oenological Advancements
by Rowland Adetayo Adesida, Jan Reščič, Lorena Butinar and Melita Sternad Lemut
Microorganisms 2026, 14(3), 705; https://doi.org/10.3390/microorganisms14030705 - 21 Mar 2026
Viewed by 854
Abstract
The growing emphasis on sustainability, regional distinctiveness, and spontaneous fermentation in winemaking necessitates a more comprehensive understanding of local yeast populations and their functional mechanisms. In total, 115 indigenous yeast strains were examined for their enzymatic activities of potential vitivinicultural significance. The yeasts [...] Read more.
The growing emphasis on sustainability, regional distinctiveness, and spontaneous fermentation in winemaking necessitates a more comprehensive understanding of local yeast populations and their functional mechanisms. In total, 115 indigenous yeast strains were examined for their enzymatic activities of potential vitivinicultural significance. The yeasts were screened for chitinase activity (biocontrol potential), glycosidase activity (terpene release), β-lyases (thiol release), and sulfite reductases (off-flavor formation), followed by quantitative analysis of the selected subsets. Yeasts were further evaluated for inhibition of fungal mycelial growth, VOC-mediated inhibition, and tolerance to commonly applied fungicides. Pre-field selection was refined using the niche overlap index and grapevine leaf disc assay. The results confirmed chitinolytic activity in four species; all strains exhibited hydrolase activities, with H. uvarum 116 displaying the highest cell-associated activity (6.32 U/g), while T. delbrueckii Sut94 showed the highest extracellular activity (1.36 U/g). β-glucosidase and β-lyase activities were widespread, whereas hydrogen sulfide production was infrequent. P. guilliermondii ZIM 624 showed the most comprehensive overall enzymatic profile, together with strong inhibition patterns. A field trial on Pinot cultivars (V. vinifera L.) further evaluated P. guilliermondii ZIM 624 within an integrated disease management approach, with responses being more pronounced in ‘Pinot noir’ than in ‘Pinot gris’. Full article
(This article belongs to the Section Food Microbiology)
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14 pages, 514 KB  
Article
High Levels of Helicobacter pylori Antimicrobial Resistance in Ireland—A Multicentre Study
by Thomas J. Butler, Stephen Molloy, Atiyekeogbebe Douglas, Denise Brennan, Rebecca FitzGerald, Conor Costigan, Vikrant Parihar, Kevin Van Der Merwe, Serhiy Semenov, Donal Tighe, Sharon Hough, David Kevans, Colm O’Morain, Deirdre McNamara and Sinéad Marian Smith
Microorganisms 2026, 14(3), 704; https://doi.org/10.3390/microorganisms14030704 - 21 Mar 2026
Viewed by 446
Abstract
Resistance surveillance programmes are essential for choosing the most appropriate eradication therapy for the stomach pathogen Helicobacter pylori. This study aimed to determine H. pylori antimicrobial resistance rates in Ireland. H. pylori was cultured from patients attending four gastroenterology clinics from 2018 [...] Read more.
Resistance surveillance programmes are essential for choosing the most appropriate eradication therapy for the stomach pathogen Helicobacter pylori. This study aimed to determine H. pylori antimicrobial resistance rates in Ireland. H. pylori was cultured from patients attending four gastroenterology clinics from 2018 to 2023. Antimicrobial susceptibility testing (AST) was performed using Etests for metronidazole, clarithromycin, levofloxacin, amoxicillin, tetracycline and rifampicin and resistance classified using EUCAST guidelines. Resistance rates were compared between H. pylori treatment-naïve and previously treated patients (primary and secondary resistance, respectively). Samples from 138 culture-positive patients (mean age 49.4 ± 15.7 years, 47.1% female) were analysed. A total of 28.7% of isolates from treatment-naïve patients were susceptible to all antimicrobials tested. Primary resistance rates to metronidazole, clarithromycin, levofloxacin, amoxicillin, tetracycline and rifampicin were 44.3%, 36.5%, 18.3%, 14.6%, 9.6% and 9.6%, respectively. Primary dual resistance to clarithromycin and metronidazole was 22.6% and primary multidrug resistance was 13.0%. Secondary resistance rates were significantly higher than primary resistance rates for clarithromycin, metronidazole, dual resistance to clarithromycin and either amoxicillin, metronidazole or levofloxacin, and multidrug resistance. Female sex and older age were associated with increased risk of resistance. H. pylori resistance rates were high in our cohort. Clarithromycin-based triple therapy should no longer be used in Ireland in the absence of pre-treatment AST. Resistance to amoxicillin, tetracycline and rifampicin should be monitored closely. Full article
(This article belongs to the Special Issue Gut Microbiota and Human Health: From Mechanisms to Therapeutic Frontiers)
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27 pages, 4087 KB  
Article
Alpinia katsumadai and Wurfbainia vera Extracts Modulate Antioxidant Function and Intestinal Morphology in Danzhou Chickens via Gut Microbiota–Metabolite Interactions Involving Hydroxyoctadecadienoic Acid Metabolism and Bacteroidota Remodeling
by Hongzhi Wu, Haoliang Chai, Xilong Yu, Dexin Zhao, Hanyang Liu, Weiqi Peng, Fengjie Ji, Liangmei Xu and Guanyu Hou
Microorganisms 2026, 14(3), 703; https://doi.org/10.3390/microorganisms14030703 - 20 Mar 2026
Viewed by 323
Abstract
This study evaluated the effects of supplementing Alpinia katsumadai and Wurfbainia vera extracts on the growth performance, antioxidant capacity, intestinal metabolites, and microbiota of Danzhou chickens. Using Danzhou broilers, we examined the individual or combined inclusion of Alpinia katsumadai and Wurfbainia vera extracts [...] Read more.
This study evaluated the effects of supplementing Alpinia katsumadai and Wurfbainia vera extracts on the growth performance, antioxidant capacity, intestinal metabolites, and microbiota of Danzhou chickens. Using Danzhou broilers, we examined the individual or combined inclusion of Alpinia katsumadai and Wurfbainia vera extracts in a 2 × 2 factorial layout. Four hundred and eighty female dual-purpose chickens were randomly assigned to four treatments (six replicates of 20 chicks each): control basal diet (CON), basal + 600 mg kg−1Alpinia katsumadai (T1), basal + 600 mg kg−1 Wurfbainia vera (T2), or basal + 600 mg kg−1 Alpinia katsumadai + 600 mg kg−1 Wurfbainia vera (T3). All treatments differed significantly from CON. For intestinal morphology, T1, T2, and T3 increased jejunal villus height and villus-to-crypt ratio while reducing crypt depth. T1 exceeded CON (p < 0.05), and an interaction was detected. T1 raised the abundances of Bacteroidota, Bifidobacterium, Tidjanibacter, and Phocaeicola relative to CON (p < 0.05). T3 exhibited higher activities of glutathione peroxidase and catalase than CON, T2, and T1 (p < 0.05). Metabolomically, T1, T2, and T3 elevated intestinal Menaquinone-9, lecithin, and L-galactono-1,5-lactone versus CON (p < 0.05). T3 lowered 3-(R)-Hydroxyoctadecadienoic acid and 9-(R)-Hydroxyoctadecadienoic acid versus CON and T1, and increased eugenol versus CON (p < 0.05). Overall, T1 and T2, especially in combination, enhance antioxidant capacity, improve gut morphology, promote beneficial microbiota and activate health-related metabolic pathways in Danzhou broilers. Full article
(This article belongs to the Section Gut Microbiota)
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18 pages, 2508 KB  
Article
Machine Learning-Enhanced MALDI-TOF Mass Spectrometry for Screening HBsAg-Positive Patients
by Tiantian Zhang, Shixuan Huang, Junxun Li, Yuwei Wu, Xinyu Zhao, He Gao, Juan Yang, Lingshuang Yang, Lulu Cao, Xinqiang Xie, Hui Zhao, Jing Cheng, Hongxia Tan, Ying Li and Qingping Wu
Microorganisms 2026, 14(3), 702; https://doi.org/10.3390/microorganisms14030702 - 20 Mar 2026
Viewed by 309
Abstract
Hepatitis B virus (HBV) remains a major global public health challenge, and its early screening is essential for controlling transmission and improving treatment outcomes. We analyzed serum samples from 422 participants via Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) to establish a [...] Read more.
Hepatitis B virus (HBV) remains a major global public health challenge, and its early screening is essential for controlling transmission and improving treatment outcomes. We analyzed serum samples from 422 participants via Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) to establish a screening model for hepatitis B surface antigen (HBsAg)-positive status. Following multi-bin preprocessing and single-sample spectral aggregation, we assessed three machine learning algorithms—random forest, deep neural network, and light gradient boosting machine (LightGBM). Among them, the LightGBM model achieved the best performance, with an optimized F1 score of 0.87 and an area under the receiver operating characteristic curve (AUC) of 0.94. A 100-iteration ensemble feature stabilization strategy identified twelve distinct m/z peaks as stable biomarkers for HBsAg-positive screening. Independent validation yielded sensitivity of 77.7% and specificity of 76.0%—insufficient for individual diagnosis but potentially suitable for population-level surveillance programs combined with confirmatory testing, particularly in resource-limited settings where conventional methods are impractical. Notably, the method offers a detection time of approximately one minute, a per-sample cost of ~$0.14. In conclusion, the combination of MALDI-TOF MS and machine learning enables a rapid, low-cost screening tool for large-scale HBV detection. Full article
(This article belongs to the Special Issue Matrix-Assisted Laser Desorption/Ionization Time of Flight Mass Spectrometry (MALDI-TOF MS) for the Identification of Pathogenic Microorganisms, 3rd Edition)
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18 pages, 4247 KB  
Article
Bacillus coagulans idrc019 Attenuates Irritable Bowel Syndrome by Revealing Multimodal Protective Mechanisms
by Yi-Wei Jin, Feng Chen and Jiang Cao
Microorganisms 2026, 14(3), 701; https://doi.org/10.3390/microorganisms14030701 - 20 Mar 2026
Viewed by 376
Abstract
Bacillus coagulans has attracted widespread attention in the treatment of irritable bowel syndrome due to its multiple probiotic functions, yet its specific molecular mechanisms remain unclear, and the efficacy of probiotics exhibits significant strain specificity, posing a key bottleneck for practical application. To [...] Read more.
Bacillus coagulans has attracted widespread attention in the treatment of irritable bowel syndrome due to its multiple probiotic functions, yet its specific molecular mechanisms remain unclear, and the efficacy of probiotics exhibits significant strain specificity, posing a key bottleneck for practical application. To address this, this study obtained a bile salt-tolerant B. coaguans idrc019 through in vitro screening. This strain demonstrated strong survival and germination in simulated gut conditions, supporting effective intestinal colonization. Further evaluation in an IBS animal model revealed that idrc019 alleviated visceral hypersensitivity and colonic inflammation in a dose-dependent manner. Through enhanced intestinal barrier integrity, microbiota modulation (e.g., Actinobacteria restoration), and elevated metabolites (e.g., kynurenine), the strain exerted IBS-alleviating effects via synchronized immune, microbial, and metabolic regulation. Our findings offer a mechanistically grounded probiotic candidate, underscore functional screening as a critical strategy, and pave the way for clinical application. Full article
(This article belongs to the Section Food Microbiology)
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19 pages, 18921 KB  
Article
Ethanolic Gracilaria fisheri Extract and Purified N-Benzylcinnamamide Inhibit Staphylococcus epidermidis Adhesion and Biofilm Formation on Device-Relevant Surfaces
by Kulwadee Karnjana, Sakun Thala and Kanokpan Wongprasert
Microorganisms 2026, 14(3), 700; https://doi.org/10.3390/microorganisms14030700 - 20 Mar 2026
Viewed by 377
Abstract
Staphylococcus epidermidis is a leading opportunistic pathogen in medical device-associated infections due to its ability to adhere to abiotic materials and develop biofilms that are difficult to eradicate. This study investigated the antibiofilm potential of an ethanolic extract of the red seaweed Gracilaria [...] Read more.
Staphylococcus epidermidis is a leading opportunistic pathogen in medical device-associated infections due to its ability to adhere to abiotic materials and develop biofilms that are difficult to eradicate. This study investigated the antibiofilm potential of an ethanolic extract of the red seaweed Gracilaria fisheri and its purified constituent, N-benzylcinnamamide, against S. epidermidis. Antibacterial activity was determined, and antibiofilm effects were assessed using the crystal violet assay and confocal laser scanning microscopy (CLSM). Early bacterial adhesion on glass and polyurethane (PU) surfaces was measured. The effect on catheter-associated biofilms was evaluated by scanning electron microscopy (SEM). Transcripts of biofilm- and quorum-sensing-associated genes (icaA and luxS) were assessed by semi-quantitative RT-PCR. Cytotoxicity was evaluated by MTT assay. At 200 µg/mL, biofilm biomass decreased to 48.21 ± 5.52% with the extract and to 36.65 ± 6.82% with N-benzylcinnamamide. CLSM time-course imaging showed delayed biofilm maturation and less consolidated, discontinuous structures. Surface exposure to the extract markedly reduced early attachment on both materials. On PU catheter segments, SEM demonstrated that N-benzylcinnamamide markedly reduced surface coverage and disrupted three-dimensional biofilm architecture. At the molecular level, transcription of icaA and luxS was reduced. Both the extract and N-benzylcinnamamide showed minimal cytotoxicity in HeLa cells. These findings support further evaluation of these marine-derived agents as candidates for antibiofilm surface treatments to reduce early medical device colonization. Full article
(This article belongs to the Section Biofilm)
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16 pages, 1689 KB  
Article
Phenotypic and Metabolic Variations in High-Risk Clones of Multidrug-Resistant Pseudomonas aeruginosa
by Sonia J. Gutierrez, Juan David Escobar Prieto, Deninson Alejandro Vargas, Richard Burchmore, Karl Burguess and Adriana Correa
Microorganisms 2026, 14(3), 699; https://doi.org/10.3390/microorganisms14030699 - 20 Mar 2026
Viewed by 334
Abstract
The global spread of high-risk clo1nes (HRCs) of multidrug-resistant (MDR) Pseudomonas aeruginosa has hindered infection control and treatment strategies worldwide. In Colombia, globally relevant HRCs such as ST235 and ST111 have been widely reported. In this study, we evaluated phenotypic and metabolic variations [...] Read more.
The global spread of high-risk clo1nes (HRCs) of multidrug-resistant (MDR) Pseudomonas aeruginosa has hindered infection control and treatment strategies worldwide. In Colombia, globally relevant HRCs such as ST235 and ST111 have been widely reported. In this study, we evaluated phenotypic and metabolic variations associated with intracellular survival and dissemination in P. aeruginosa. A total of 100 clinical isolates were collected from 22 hospitals in Colombia. The isolates had been previously characterized and classified as MDR or susceptible strains (SSs), and their sequence types (STs) had been earlier determined. Based on this prior characterization, isolates were grouped in this study as multidrug-resistant high-risk clones (HRC, n = 50; corresponding to sequence types ST235 and ST111), multidrug-resistant non-high-risk clones (NHRCs, n = 27; non-ST235/ST111), and susceptible strains (SS, n = 23; also, non-ST235/ST111). Phenotypic traits, including motility, spontaneous mutation frequency, biofilm formation, and pigment production, were evaluated. In addition, a subset of 30 isolates was assessed for intracellular survival in vitro and metabolomic profiling using liquid chromatography-mass spectrometry. HRC isolates exhibited significantly reduced motility compared with NHRC and SS isolates (swarming: HRC vs. NHRC, p = 0.0032; HRC vs. SS, p = 0.010; swimming: HRC vs. NHRC and SS, p < 0.0001; twitching: HRC vs. SS, p = 0.0004), as well as lower pigment production (pyocyanin: HRC vs. NHRC and SS, p < 0.0001; pyoverdine: HRC vs. NHRC, p < 0.0001). Metabolomic analysis revealed increased concentrations of metabolites associated with iron acquisition and siderophore-related pathways in HRC isolates. Overall, these findings suggest that P. aeruginosa HRCs display distinct phenotypic and metabolic patterns that may contribute to persistence and dissemination in clinical settings, contributing to their epidemiological success. Full article
(This article belongs to the Topic Healthcare-Associated Infections and Antimicrobial Therapy)
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17 pages, 1908 KB  
Article
Novel Genomes of Sphingomonadales Strains Isolated from Diverse Environments
by Nathan W. Williams, Tahir Ali and Paul D. Boudreau
Microorganisms 2026, 14(3), 698; https://doi.org/10.3390/microorganisms14030698 - 20 Mar 2026
Viewed by 305
Abstract
Glycosphingolipids are amphiphilic compounds that feature sugar or glycan moieties installed onto a ceramide lipid. The synthesis of glycosphingolipids by members of the human gut microbiome, and their known immune stimulating activity, have made them of interest for potential pharmaceutical roles. However, the [...] Read more.
Glycosphingolipids are amphiphilic compounds that feature sugar or glycan moieties installed onto a ceramide lipid. The synthesis of glycosphingolipids by members of the human gut microbiome, and their known immune stimulating activity, have made them of interest for potential pharmaceutical roles. However, the known diversity of glycosphingolipid glycans in bacteria remains limited, highlighting the need to isolate novel glycosphingolipid-producing organisms as a source of these compounds. The order Sphingomonadales, one of the major clades of sphingolipid producing bacteria, conserves a serine palmitoyltransferase (SPT) enzyme needed for the initial biosynthetic step in sphingolipid production which can be targeted as part of isolation efforts. With these bacteria known to live in diverse environments such as soil microbiomes, soap scum biofilms, and cyanobacterial microbiomes, there are many environments to target for the isolation of these bacteria. In this work, we designed a set of polymerase chain reaction (PCR) primers for the isolation of diverse Sphingomonadales strains by targeting the SPT gene (spt), which we used to isolate strains from the genera Sphingomonas and Novosphingobium in soil, soap scum biofilms, and xenic cyanobacterial cultures. In these efforts, streptomycin improved the encounter rate, as represented by the SPT assay true-positive rate. Our isolates represent novel genomic space: with genomes from both genera that have low similarity to known genomes, suggestive of novel species, while several novel plasmids were also missing known marker sequences. Full article
(This article belongs to the Section Environmental Microbiology)
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