Pathogens, Volume 14, Issue 8 (August 2025) – 81 articles

The Asian citrus psyllid (Diaphorina citri) is an economically important pest of citrus as it is a vector of the bacterium (Candidatus Liberibacter asiaticus, CLas) that causes huanglongbing disease (HLB). Understanding the virome of D. citri is important for uncovering factors that influence vector competence, to support biosecurity surveillance, and to identify candidate agents for biological control. Previous studies have identified several D. citri-associated viruses from various geographical populations of this pest. To further investigate virus diversity in this pest, high-throughput sequencing was used to analyse D. citri populations from the Samoan islands of Upolu and Savai’i. Eleven novel viruses from the Yadokariviridae, Botourmiaviridae, Nodaviridae, Mymonaviridae, Partitiviridae, Totiviridae, and Polymycoviridae were identified as well as some that corresponded to unclassified groups. In addition, microbiome analysis revealed the presence of several endosymbiotic microorganisms, including Wolbachia, as well as some plant pathogenic fungi, including Botrytis cinerea. However, the causative agent of HLB disease (CLas) was not detected in the RNA-Seq data. These findings highlight the complex and diverse microbiota associated with D. citri and suggest potential interactions and dynamics between microorganisms and psyllid-associated viruses. Further research is needed to understand the ecological significance of these discoveries, and whether the novel viruses play a role in regulating field populations of the psyllid. Full article

The metastrongyloid nematodes Angiostrongylus vasorum, Aelurostrongylus abstrusus, and Crenosoma vulpis can cause severe cardiopulmonary and respiratory symptoms in domestic dogs and cats and free-ranging canids and felids (e.g., foxes, wolves, wild cats, lynxes). Recent data on the prevalence of A. vasorum infections in dogs and foxes and on the prevalence of Ae. abstrusus and Troglostrongylus brevior infections in free-ranging lynxes and wild cats revealed several endemic and hyperendemic foci in Germany. Nonetheless, long-term investigations on the prevalence of metastrongyloid larvae infecting gastropod intermediate hosts are still scarce for Germany. To fill this gap, we conducted an epidemiological survey on native slugs and snails in a selected meadow close to Obrigheim, previously identified as a hyperendemic focus for canine angiostrongylosis. To re-evaluate this location as a ‘hotspot’ of canine angiostrongylosis, terrestrial slugs and snails (n = 533) were collected in all seasons, artificially digested, and microscopically and molecularly analyzed for the presence of metastrongyloid lungworm larvae. Here, the prevalence ranged greatly between seasons. In summer, 27.46% (59/215) of gastropods were infected with metastrongyloid larvae. In fall, the prevalence dropped to 10.00% (16/160) and lowest infection rates were observed in both winter (5.65%) and spring (1.47%). In total, A. vasorum was detected in 12.01% (64/533), Crenosoma sp. in 0.94% (5/533), and Ae. abstrusus in 0.38% (2/533) of gastropod samples. Even though total A. vasorum infection levels were revealed to be considerably lower than in the prior study, this epidemiological survey in principle reconfirms Obrigheim as a stable hyperendemic focus and thereby as a location with high metastrongyloid infection risk for domestic dogs, cats, and wildlife throughout the year. These results call for continuous epidemiological studies on gastropod populations to better understand metastrongyloid lungworm spread and infection dynamics over the years. Full article

Recurrent vulvovaginal candidiasis (RVVC), predominantly caused by Candida albicans, represents a global health issue, particularly in developing regions. This study explores the antifungal potential of aqueous leaf extract of Schinus weinmanniifolia Mart. ex Engl., a native Latin American plant. The extract was evaluated for phytochemical composition, antifungal efficacy, and safety profile. Phytochemical analyses identified six major compounds, including shikimic acid, gallic acid, and methyl gallate, with antioxidant and antimicrobial properties. The extract showed potent antioxidant activity, with IC₅₀ values between 1.52-5.51 µg/mL. It strongly inhibited C. albicans, with a minimum inhibitory concentration (MIC) of 1.95 µg/mL, and was active against other yeasts (MIC 0.48–62.5 µg/mL). The growth kinetics assay revealed reduced C. albicans viability after 12 h at 2×MIC versus the positive control. Scanning electron microscopy confirmed reduced fungal counts without morphological damage. The extract impaired C. albicans virulence, reducing germ tube formation by 75.49% and hyphal transition by 84.34%, outperforming fluconazole. Biocompatibility assays showed it is non-hemolytic (IC₅₀ >1000 µg/mL), non-mutagenic, and highly selective for fungal cells (SI = 512.82), suggesting minimal human cell toxicity. In conclusion, the extract combines strong antifungal activity and favorable safety, with cost-effective preparation suitable for traditional medicine in resource-limited regions. Full article

Parvovirus B19 (B19V) re-emerged across Europe in 2024, raising concerns about vertical transmission and neonatal morbidity. We undertook a prospective, single-centre cohort study to characterise the early clinical course of congenitally infected neonates born between April and December 2024. Seventy-one pregnancies with serologically or PCR-confirmed maternal infection were enrolled; seven neonates met laboratory criteria for congenital B19V infection and were followed with serial clinical, biochemical and imaging assessments through the first year of life. Troponin I and CK-MB were measured on days 1, 3, 7 and 15; electrocardiogram (ECG) and echocardiography were repeated in parallel, and cranial ultrasound (US), ophthalmologic and audiologic screening were scheduled prospectively. Mean troponin rose from 50.7 ng L⁻¹ on day 1 to a peak of 120.7 ng L⁻¹ on day 7 (p < 0.01), normalising by one month, while echocardiograms remained structurally normal, and only one transient arrhythmia was recorded. CK-MB exceeded the reference range in 29% of infants but showed no clinical sequelae. Multiple periventricular hyperechogenicities were identified in 8/70 neonates (11%), and moderate anaemia (Hb ≤ 9.8 g/dL) occurred in 2 cases. Serum PCR detected high-level viraemia (>10⁸ genome equivalents mL⁻¹) in 40% of those tested; saliva and urine were consistently negative. No instances of myocarditis or hydrops were observed. Our findings indicate that congenital B19V infection during the current outbreak is marked by transient biochemical myocardial stress and subtle neurosonographic changes rather than overt cardiac disease, supporting an outpatient-focused follow-up strategy incorporating serial biomarkers and targeted neuroimaging. Full article

Yersinia pestis, the causative agent of plague, has triggered multiple pandemics throughout human history, yet its long-term evolutionary patterns and reservoir dynamics remain poorly understood. Here, we present a global phylogenomic analysis of ancient and modern Y. pestis strains spanning from the Neolithic and Bronze Age to the present day. We show that pandemic-causing lineages did not arise from a single ancestral strain but instead emerged independently along deep branches of the Y. pestis phylogeny. Pandemic-associated Y. pestis strains were recovered exclusively from human remains and display clear local temporal divergence, indicating evolution driven by human transmission during outbreaks. These findings support the hypothesis that plague emergence is driven by complex, regionally rooted reservoirs, with recurrent spillovers into human populations across millennia. Our work highlights the need to view plague not as a series of isolated outbreaks but as a long-standing zoonotic threat shaped by deep evolutionary history, host ecology, and human societal structures. Full article

Childbearing-aged and pregnant women represent a risk group for Toxoplasma gondii infection due to possible transplacental transmission resulting in congenital toxoplasmosis. We analyzed the seroepidemiological trends of toxoplasmosis in Croatia over ten years (2015–2024). A total of 2791 childbearing-aged and pregnant women were included. Toxoplasma gondii-specific IgM/IgG antibodies were detected using an enzyme-linked fluorescence assay. Samples with positive IgM and IgG antibodies were tested for IgG avidity. IgG antibodies were detected in 695 (24.9%) participants, while acute toxoplasmosis (IgM antibodies and low avidity IgG antibodies) was confirmed in 32 (1.2%) of participants. The IgG seroprevalence showed a declining trend over the years. Residents of suburban/rural areas were more often seropositive than those in urban areas (31.4 vs. 22.3%). Logistic regression analysis revealed that year of testing, age, and settlement were associated with the risk of seropositivity. For each later calendar year, the log odds of being IgG-positive decreased, while for each additional year of age, the log odds increased. Residence in an urban area was associated with lower log odds. The region was not a significant predictor in the logistic regression. The differences in seropositivity observed across regions can be mainly attributed to Pannonian Croatia, which showed significantly higher odds of IgG seropositivity. Data about the toxoplasma serological status is useful for planning prevention campaigns. Full article

Background and Aim: The immunological interactions between soil-transmitted helminths (STHs) and herpes simplex virus type 2 (HSV-2), particularly in the context of co-infection, are poorly understood. Next-generation sequencing (NGS) offers a powerful approach to explore these complex immune responses and uncover potential therapeutic targets. This study leveraged NGS and bioinformatic tools to investigate transcriptional changes and immunological pathways in female genital tract (FGT) tissues of BALB/c mice acutely infected with Nippostrongylus brasiliensis (Nb), HSV-2, or co-infected. Methods: Total RNA was harvested from FGT tissues of BALB/c mice infected with Nb, HSV-2, co-infected with both pathogens, and uninfected controls. Differentially expressed genes (DEGs) were identified by comparing uninfected versus infected FGT tissues in R using edgeR and limma packages. Immune-related genes were identified by intersecting DEGs in each group-wise comparison with immune function gene sets derived from the Mouse Genome Informatics (MGI) database. Functional and pathway enrichment analyses were performed with g: Profiler and protein–protein interaction networks were built using the STRING database and visualized with Cytoscape. Key hub genes and significant gene modules were identified using the Cytoscape plugins CytoHubba and MCODE, followed by further functional analysis of these modules. Results: NGS analysis revealed distinct gene expression profiles in response to single infection with Nb or HSV-2, with both showing significant differences when uninfected controls were compared to infected FGT tissues at a 5% false discovery rate. Notably, there were no significant differences in gene expression profiles between uninfected and co-infected FGT tissues. In the comparison of uninfected versus Nb-infected FGT tissues, 368 DEGs were identified, with 356 genes upregulated and 12 downregulated. Several immune-related genes, such as Ptprc, Ccl11, Ccr2, and Cx3cr1, were significantly altered. Pathway analysis of DEGs, hub genes, and significant modules indicated modulation of immune and defense responses. Notably, Nb infection induced a robust Th2-dominant immune response in the FGT, with downregulation of pro-inflammatory genes. This likely reflects helminth-driven modulation that may impair protective Th1 responses and highlights the systemic impact of Nb on the FGT immunity. In the comparison of uninfected versus HSV-2-infected FGT tissues, 140 DEGs were identified, with 121 upregulated and 19 downregulated. Immune-related genes, including Ldlr, Camk1d, Lrp8 and Epg5, were notably altered. HSV-2 infection led to early and predominant downregulation of immune genes, consistent with viral immune evasion strategies. In addition, functional analysis revealed enrichment in cell cycle and sterol biosynthesis pathways, suggesting that HSV-2 modulates host metabolism to support viral replication while influencing immune responses. In co-infection, no significant transcriptional changes were observed, potentially reflecting immune antagonism where Nb-induced Th2 responses may suppress HSV-2-driven Th1 immune responses. Conclusions: This preliminary study offers insights into the gene expression responses in the FGT to acute single and co-infection with Nb and HSV-2. Together, these findings reveal distinct transcriptomic changes in the FGT following Nb and HSV-2 infection, with co-infection potentially leading to immune antagonism and transcriptional equilibrium. This highlights the complex interplay between helminth- and virus-induced immune modulation in shaping FGT immunity. By leveraging NGS, this study highlights important immune-related pathways and serves as a foundation for further investigations into the mechanistic roles of DEGs in immunity to these pathogens, with potential implications for developing novel therapeutic strategies. Full article

Two key players in the immune system, dendritic cells (DCs) and innate lymphoid cells (ILCs), interact in a crucial way to fight infectious diseases. DCs play a key role in recognizing pathogens, and ILCs respond to cytokines released by DCs. This response triggers the production of specific effector cytokines that help control pathogens and maintain the body’s barrier integrity. DCs have various receptors, including Toll-like receptors (TLRs), that detect microbial components and trigger immune responses. Likewise, ILCs act as essential initial responders in the immune system in viral, bacterial, and parasitic infections. Successfully managing diseases caused by pathogens mainly depends on the combined actions of DCs and ILCs, which work to suppress and eliminate pathogens. DCs also play a crucial role in activating innate and adaptive immune cell subsets, including ILCs. Furthermore, the use of DCs in developing vaccines and immunotherapy for cancers, along with the dedication of many researchers to improve immune responses through DCs, has increased interest in the potential of DC therapies for treating and preventing infectious diseases. This review examines approaches that may enhance DC vaccines and boost anti-infection immune responses by fostering better interactions of DCs with ILCs. Full article

The highly pathogenic avian influenza H5N1 2.3.4.4b clade virus has caused widespread outbreaks across South America, primarily affecting seabirds, poultry, and marine mammals. The virus likely reached the continent through migratory birds from North America, initially spreading along the Pacific coast before advancing into Atlantic-bordering countries such as Argentina, Uruguay, and Brazil. This study investigated the dynamics of H5N1 strains in Uruguay during outbreaks from February and October 2023. We analyzed an updated South American database, including a newly sequenced viral genome from a royal tern (Thalasseus maximus) collected at the end of the outbreaks. Phylogeographic reconstruction revealed two distinct South American phylogroups comprising Uruguayan strains: one mainly driven by wild birds and poultry, with the royal tern strain clustering with Brazilian isolates, and another primarily associated with marine mammals, displaying adaptive residues in the PB2 protein. In Uruguay, these phylogroups delineate two main transmission routes: (i) an avian-derived pathway originating in Argentina and (ii) a pinniped-derived route from Chile. Brazil, initially colonized via the Argentine route, later emerged as a secondary source for Uruguay. This host-pathway interplay underscores the virus’s cross-species potential and highlights the need for coordinated regional surveillance within a One Health framework to mitigate zoonotic risks. Full article

Mosquito blood meals provide a biological sample of host blood which can then be used in downstream applications including host–pathogen detection. We conducted DNA barcoding to identify the host species of blood meals from 4557 blood engorged mosquitoes collected in south central Florida, USA. We identified 314 blood meals from invasive wild pigs, 219 wild turkey blood meals, and 1046 white-tailed deer blood meals. From a subset of these host blood meals, we used molecular assays to detect the nucleic acids of Torque teno sus virus 1 (TTSuV1) in wild pig blood meals, Lymphoproliferative virus (LPDV) in wild turkey blood meals, and bluetongue virus (BTV) in white-tailed deer blood meals. None of these wildlife pathogens are transmitted by mosquitoes, but viral nucleic acids circulate in the peripheral blood of host species during or post infection. Prevalence of TTSuV1 in wild pig blood meals was 34%; in wild turkey blood meals the prevalence of LPDV was 2.7%, and BTV prevalence in blood meals of white-tailed deer was 3.6%. These prevalence values were similar to estimates obtained from peripheral blood collected directly from these hosts in Florida. Our analysis suggests that mosquito blood meals are a valuable sampling tool for the detection of wildlife pathogens. We suggest that this type of exogenous xenosurveillance, using mosquitoes to infer information about the vertebrate host, is distinct from endogenous xenosurveillance in which the goal is to understand the role of the arthropod in vectoring a pathogen. Full article

In this study, astaxanthin, which has previously been shown to have antiviral effects, was examined for its dose-dependent potency to inhibit cellular SARS-CoV-2 infections. Naturally occurring astaxanthin is obtained and orally administered as ASX-oleoresin, a composition of different astaxanthin fatty acid esters. We therefore hypothesized that the compound’s beneficial effects are not only related to astaxanthin. Thus, a “green” algae extract (i.e., poor astaxanthin content < 0.2%; ASX^p) of the microalgae Haematococcus pluvialis, as well as an astaxanthin-rich algae extract (astaxanthin content = 20%; ASX^r), were tested in in vitro cellular viral infection assays. Thereby, it was found that both extracts reduced viral infections significantly. As a potential mode of inhibitory action, the binding of ASX-oleoresin to the viral spike protein was investigated by isothermal fluorescence titration, revealing binding affinities of Kd = 1.05 µM for ASX^r and Kd = 1.42 µM for ASX^p. Based on our data, we conclude that several ASX-oleoresin fractions from H. pluvialis exhibit antiviral activity, which extends beyond the known antioxidant activity of astaxanthin. From a molecular dynamic simulation of ASX-oleoresin, fatty acid domains could be considered as activity-chaperoning factors of ASX. Therefore, microalgae biomass should be considered in the future for further antiviral activities. Full article

Coxiella burnetii, the etiological agent of Q fever, is a globally distributed zoonotic pathogen affecting both animals and humans. Despite its known endemicity in various Mediterranean regions, data on human seroprevalence in Sardinia are still lacking. This study aimed to assess seroprevalence in patients and to analyze the annual positivity rate related to the serum samples collected in Sardinia over a ten-year period (2015–2024). For this purpose, a total of 1792 patients were involved in the survey, and 4310 serum samples were analyzed using indirect immunofluorescence assay (IFI) to detect IgM and IgG antibodies against C. burnetii. The global seroprevalence rates relating to all the patients over a ten-year period were determined along with the annual positivity rate and trends from all sera. An overall seroprevalence of 27.0% and an average of annual positivity rate of 16.0% were determined, with the IFI detecting IgG antibodies in 15.2% of positive samples and IgM antibodies in 0.9%, suggesting significant prior exposure of the population evaluated. Annual positivity rates ranged from 24.8% in 2016 to 8.0% in 2020. These results confirmed the endemic circulation of C. burnetii in Sardinia and the ongoing risk of human exposure. A GIS-based map was built to evidence the spatial distribution of Q fever in Sardinia. Interestingly, areas with higher seroprevalence appear to coincide with the distribution of sheep and goat farms, indicating a link between livestock and human exposure. These findings confirm the circulation of C. burnetii in Sardinia and underscore the importance of epidemiological monitoring, public health interventions, and educational efforts in populations at increased risk. Full article

Background/Objectives: This study investigates the potential of Anisakis sp. as a novel source of α-Gal (Galα1-3Galβ1-4GlcNAc-R) epitopes capable of inducing allergic sensitization in humans. While α-Gal is classically associated with delayed IgE-mediated hypersensitivity following tick bites, emerging evidence suggests that parasitic helminths such as Anisakis sp. may also express α-Gal-containing glycoconjugates, offering an alternative sensitization pathway. Methods: Protein extracts from Anisakis sp. third-stage larvae and mammalian tissues (beef, pork) were analyzed by SDS-PAGE and Western blot using a monoclonal anti-α-Gal antibody (clone M86), and α-Gal epitopes were detected by ELISA. Sera from urticaria patients, stratified by Anisakis sp. sensitization status, were evaluated for anti-α-Gal IgG, IgE, and IgG4 antibodies. Inhibition assays assessed cross-reactivity. Results: Results confirmed the presence of α-Gal epitopes on Anisakis sp. proteins, with prominent bands at ~250 kDa and 65 kDa. Urticaria patients sensitized to Anisakis sp. exhibited significantly elevated anti-α-Gal antibody levels compared to controls. Inhibition ELISA demonstrated substantial reduction in antibody binding with Anisakis sp. extracts, indicating shared antigenic determinants with mammalian α-Gal. Conclusions: These findings establish Anisakis sp. as a source of α-Gal-containing glycoproteins capable of eliciting specific antibody responses in humans, highlighting a potential parasitic route for α-Gal sensitization. Full article

The core perturbome is defined as a central response to multiple disturbances, functioning as a complex molecular network to overcome the disruption of homeostasis under stress conditions, thereby promoting tolerance and survival under stress conditions. Based on the biological and clinical relevance of Escherichia coli and Staphylococcus aureus, we characterized their molecular responses to multiple perturbations. Gene expression data from E. coli (8815 target genes—based on a pangenome—across 132 samples) and S. aureus (3312 target genes across 156 samples) were used. Accordingly, this study aimed to identify and describe the functionality of the core perturbome of these two prokaryotic models using a machine learning approach. For this purpose, feature selection and classification algorithms (KNN, RF and SVM) were implemented to identify a subset of genes as core molecular signatures, distinguishing control and perturbation conditions. After verifying effective dimensional reduction (with median accuracies of 82.6% and 85.1% for E. coli and S. aureus, respectively), a model of molecular interactions and functional enrichment analyses was performed to characterize the selected genes. The core perturbome was composed of 55 genes (including nine hubs) for E. coli and 46 (eight hubs) for S. aureus. Well-defined interactomes were predicted for each model, which are jointly associated with enriched pathways, including energy and macromolecule metabolism, DNA/RNA and protein synthesis and degradation, transcription regulation, virulence factors, and other signaling processes. Taken together, these results may support the identification of potential therapeutic targets and biomarkers of stress responses in future studies. Full article

The cross-species spillover of coronaviruses is considered a serious public health risk. Feline coronavirus (FCoV), canine coronavirus (CCoV), and transmissible gastroenteritis virus (TGEV) are all classified under Alphacoronavirus suis and infect companion animals and livestock. Due to their frequent contact with humans, these viruses pose a potential risk of future cross-species transmission. Molnupiravir, a prodrug of N4-hydroxycytidine, exhibits potent antiviral activity against SARS-CoV-2, a member of the Betacoronavirus genus, and has been approved for the treatment of COVID-19. Molnupiravir was recently shown to be effective against FCoV, suggesting broad-spectrum antiviral activity across coronavirus lineages. Based on these findings, the present study investigated whether molnupiravir is also effective against CCoV and TGEV, which belong to the same Alphacoronavirus suis species as FCoV. We examined the in vitro antiviral effects of molnupiravir using four viral strains: FCoV-1 and -2, CCoV-2, and TGEV. Molnupiravir inhibited plaque formation, viral antigen expression, the production of infectious viral particles, and viral RNA replication in a dose-dependent manner in all strains. IC₅₀ values for CCoV-2 and TGEV, calculated using a feline-derived cell line (fcwf-4), were significantly lower than those for FCoV, suggesting higher sensitivity to molnupiravir. These results demonstrate that molnupiravir exhibited broad antiviral activity against animal coronaviruses classified under Alphacoronavirus suis, providing a foundation for antiviral strategies to mitigate the future risk of cross-species transmission. Full article

White-tailed deer (Odocoileus virginianus) are a cervid species native to the Americas with ecological, social, and economic significance. Managers must consider several factors when working to maintain the health and sustainability of these wild herds, including reproduction, particularly pregnancy and recruitment rates. White-tailed deer have a variable reproductive capacity, with age, health, and habitat influencing this variability. However, it is unknown whether chronic wasting disease (CWD) impacts reproduction and, more specifically, if CWD infection alters a female deer’s probability of pregnancy. Our study addressed this question using data from 9783 female deer culled in northern Illinois between 2003 and 2023 as part of the Illinois Department of Natural Resources’ ongoing CWD management program. Multilevel Bayesian logistic regression was employed to quantify the relationship between pregnancy probability and covariates like maternal age, deer population density, and date of culling. Maternal infection with CWD was found to have no significant effect on pregnancy probability, raising concerns that the equal ability of infected and non-infected females to reproduce could make breeding, which inherently involves close physical contact, an important source of disease transmission between males and females and females and their fawns. The results also identified that female fawns (<1 year old) are sensitive to county-level deer land cover utility (LCU) and deer population density, and that there was no significant difference in how yearlings (1–2 years old) and adult (2+ years old) responded to these variables. Full article

Crimean–Congo hemorrhagic fever (CCHF) is a zoonotic viral disease endemic to parts of Africa, Asia and southeastern Europe. Bulgaria is one of the few European countries with the consistent annual reporting of human CCHF cases. This study provides a descriptive overview of 24 confirmed CCHF cases in Bulgaria between 2015 and 2024. Laboratory confirmation was performed by an enzyme-linked immunosorbent assay (ELISA) and/or real-time reverse transcriptase polymerase chain reaction (RT-qPCR) testing. Common findings included fever, fatigue, gastrointestinal symptoms, thrombocytopenia, leukopenia, liver dysfunction and coagulopathy. Two fatal cases were recorded. Two samples collected in 2016 and 2024 were subjected to whole-genome sequencing. Phylogenetic analysis showed that both strains clustered within the Turkish branch of the Europe 1 genotype and shared high genetic similarity with previous Bulgarian strains, as well as strains from neighboring countries. These findings suggest the long-term persistence of a genetically stable viral lineage in the region. Continuous molecular and clinical surveillance is necessary to monitor the evolution and public health impact of CCHFV in endemic areas. Full article

As direct detection methods of Borrelia burgdorferi are limited, serology plays an important role in diagnosing Lyme disease (LD). There are various types of Lyme serological tests with varying diagnostic accuracy, so it is necessary to compare and rank them. The aim of this study is to compare the accuracy of various serological diagnostic methods for LD using network meta-analysis (NMA). We searched the Cochrane Library and PubMed databases for all serological diagnostic accuracy studies published from the discovery of LD until June 2024. After screening, we assessed the quality of the included studies with QUADAS-C and extracted relevant data. We calculated the Q* index of the receiver operating characteristic curve for each diagnostic test. Meta-disc 2.0 and Stata 15.0 were used to perform traditional meta-analysis and NMA with the gold standard (the comprehensive evaluation) as a reference. We then compared the Q* index values between different methods using two-by-two comparisons and ranked them accordingly. A total of 52 studies with 181,032 participants, including 5318 patients with LD, were included. These studies covered 14 diagnostic methods. The results of the NMA suggest that modified two-tiered testing (MTTT), C6 enzyme immunoassay (EIA), and standard two-tiered testing (STTT) rank in the top three among the 14 methods in terms of Q* index, with MTTT being the highest, followed by C6 EIA and STTT. MTTT and C6 EIA have higher overall diagnostic performance, and their accuracy is not inferior to that of the widely used STTT (PROSPERO CRD42022378326). Full article

Rhabditis (Rhabditella) axei is a predominantly free-living nematode commonly found in sewage systems and decomposing organic matter. While primarily saprophytic, it has been documented as an opportunistic pathogen in human urinary and gastrointestinal tracts. The Chinese red panda (Ailurus styani), a rare and protected species in China, has not previously been reported as a host for Rhabditis (Rhabditella) spp. infections. This study reports the first documented occurrence of R. axei in red panda feces, unambiguously confirmed through integrative taxonomic approaches combining morphological and molecular analyses. The nematodes exhibited key morphological features consistent with R. axei, including a cylindrical rhabditiform esophagus, sexually dimorphic tail structures, and diagnostic spicule morphology. Molecular analysis based on 18S-ITS-28S rDNA sequencing confirmed their identity, showing >99% sequence similarity to R. axei reference strains (GenBank: PP135624.1, PP135622.1). Phylogenetic reconstruction using 18S rDNA and ITS rDNA sequences placed the isolate within a well-supported R. axei clade, clearly distinguishing it from related species such as R. blumi and R. brassicae. The findings demonstrate the ecological plasticity of R. axei as a facultative parasite capable of infecting non-traditional hosts and further highlight potential zoonotic risks associated with environmental exposure in captive wildlife populations. Our results emphasize the indispensable role of molecular diagnostics in accurately distinguishing morphologically similar nematodes within the Rhabditidae family, while providing essential baseline data for health monitoring in both in situ and ex situ conservation programs for this endangered species. Full article

Grapevine (Vitis vinifera L.) cultivation is an important agricultural sector worldwide. Its expansion into new areas, like Kazakhstan, brings significant phytosanitary risks. Viroids, such as grapevine yellow speckle viroid 1 (GYSVd-1) and hop stunt viroid (HSVd), are RNA pathogens that threaten vineyard productivity. They can cause a progressive decline through latent infections. Traditional diagnostic methods are usually targeted and therefore not suitable for thorough surveillance. In contrast, modern high-throughput sequencing (HTS) methods often face challenges due to their high costs and complicated sample preparation, such as ribosomal RNA (rRNA) depletion. This study introduces a simplified diagnostic workflow that overcomes these barriers. We utilized the latest Oxford Nanopore V14 cDNA chemistry, which is designed to prevent internal priming, by substituting a targeted oligo(dT)VN priming strategy to facilitate the sequencing of non-polyadenylated viroids from total RNA extracts, completely bypassing the rRNA depletion step and use of random oligonucleotides for c DNA synthesis. This method effectively detects and identifies both GYSVd-1 and HSVd. This workflow significantly reduces the time, cost, and complexity of HTS-based diagnostics. It provides a powerful and scalable tool for establishing strong genomic surveillance and phytosanitary certification programs, which are essential for supporting the growing viticulture industry in Kazakhstan. Full article

Background: Skin and soft tissue infections (SSTIs) are a major cause of emergency room (ER) visits and hospitalizations. Long-acting lipoglycopeptides (LALs), such as dalbavancin and oritavancin, offer potential for early discharge and outpatient management, especially in patients at risk for methicillin-resistant Staphylococcus aureus (MRSA) or with comorbidities. Methods: We conducted a retrospective observational cohort study from March to December 2024 in an Italian tertiary-care hospital. Adult patients treated in the ER with a single dose of dalbavancin (1500 mg) or oritavancin (1200 mg) for SSTIs were included. Demographic, clinical, and laboratory data were collected. Follow-up evaluations were performed at 14 and 30 days post-treatment to assess outcomes. Results: Nineteen patients were enrolled (median age 59 years; 53% female). Most had lower limb involvement and elevated inflammatory markers. Three patients (16%) were septic. Fourteen patients (74%) were discharged without hospital admission; hospitalization in the remaining cases was due to comorbidities rather than SSTI severity. No adverse drug reactions were observed. At 14 days, 84% of patients had clinical resolution; only 10% had recurrence by day 30, with no mortality nor readmission reported. Conclusions: LALs appear effective and well-tolerated in the ER setting, supporting early discharge and reducing healthcare burden. Broader use may require structured care pathways and multidisciplinary coordination. Full article

Bee products, in particular honey, propolis and bee venom, are of growing scientific interest due to their broad spectrum of antimicrobial activity. In the face of increasing antibiotic resistance and the limitations of conventional therapies, natural bee-derived substances offer a promising alternative or support for the treatment of infections. This paper summarizes the current state of knowledge on the chemical composition, biological properties and antimicrobial activity of key bee products. The main mechanisms of action of honey, propolis and bee venom are presented, and their potential applications in the prevention and treatment of bacterial, viral and fungal infections are discussed. Data on their synergy with conventional drugs and prospects for use in medicine and pharmacology are also included. The available findings suggest that, with appropriate standardization and further preclinical and clinical analyses, bee products could become an effective support for the treatment of infections, especially those caused by pathogens resistant to standard therapies. Full article

Fusarium diseases are among the most dangerous fungal diseases of plants. To date, there are no plant protectants that completely prevent fusariosis. Current breeding trends are therefore focused on increasing genetic resistance. While global modern maize breeding relies on various molecular genetics techniques, they are useless without a precise characterization of genomic regions that determine plant physiological responses to fungi. The aim of this study was thus to characterize the expression of candidate genes that were previously reported by our team as harboring markers linked to fusarium resistance in maize. The plant material included one susceptible and four resistant varieties. Biotic stress was induced in adult plants by inoculation with fungal spores under controlled conditions. qRT-PCR was performed. The analysis focused on four genes that encode for GDSL esterase/lipase (LOC100273960), putrescine hydroxycinnamyltransferase (LOC103649226), peroxidase 72 (LOC100282124), and uncharacterized protein (LOC100501166). Their expression showed differences between analyzed time points and varieties, peaking at 6 hpi. The resistant varieties consistently showed higher levels of expression compared to the susceptible variety, indicating their stronger defense responses. Moreover, to better understand the function of these genes, their expression in various organs and tissues was also evaluated using publicly available transcriptomic data. Our results are consistent with literature reports that clearly indicate the involvement of these genes in the resistance response to fusarium. Thus, they further emphasize the high usefulness of the previously selected markers in breeding programs to select fusarium-resistant maize genotypes. Full article

Ensuring bacterial safety of blood transfusions remains a critical focus in medicine. We investigated a novel pathogen reduction technology utilizing nylon functionalized with synthetic dyes (nylon affinity networks) to capture and remove bacteria from plasma. In the initial screening process, we spiked phosphate buffer solution (PBS) and human plasma (1 mL each) with 10 or 100 colony forming units (cfu) of either Escherichia coli or Staphylococcus epidermidis, exposed the suspensions to affinity networks and assessed the extent of bacterial reduction using agar plate cultures as the assay output. Nineteen synthetic dyes were tested. Among these, Alcian Blue exhibited the best performance with both bacterial strains in both PBS and plasma. Next, bacterial suspensions of approximately 1 and 2 cfu/mL in 10 and 50 mL, respectively, were treated with Alcian Blue affinity networks in three sequential capture steps. This procedure resulted in complete bacterial depletion, as demonstrated by the lack of bacterial growth in the remaining fraction. The viability of the captured bacteria was confirmed by plating the post-treatment affinity networks on agar. Alcian Blue affinity networks captured and sequestered a few plasma proteins identified by liquid chromatography tandem mass spectrometry. These findings support the potential applicability of nylon affinity networks to enhance transfusion safety, although additional investigations are needed. Full article

Cefepime-enmetazobactam is a novel β-lactam/β-lactamase inhibitor combination showing good activity against multidrug-resistant (MDR) Gram-negative bacteria producing a variety of β-lactamases. In this systematic review, we aimed to evaluate the available data on resistance to this drug. We performed a thorough search of four databases (Embase, PubMed, Scopus, and Web of Science), as well as backward citation searching, to identify studies containing data on resistance to cefepime-enmetazobactam. The data were extracted and analyzed according to the breakpoints established by the European Committee on Antimicrobial Susceptibility Testing (EUCAST) and the Food and Drug Administration (FDA), or the specific breakpoints reported by the authors of the respective studies. Analysis based on the type of lactamases produced by the isolates was also performed. Ten studies reported in vitro susceptibility testing and mechanisms of antimicrobial resistance. The total number of isolates was 15,408. The activity of cefepime-enmetazobactam against β-lactamase-producing isolates was variable. The resistance of the studied extended-spectrum β-lactamase (ESBL)-producing and ampicillin C β-lactamase (AmpC)-producing isolates was low (0–2.8% and 0%, respectively). The resistance was higher among oxacillinase-48 β-lactamase (OXA-48)-producing and Klebsiella pneumoniae carbapenemase (KPC)-producing isolates (3.4–13.2% and 36.7–57.8%, respectively). High resistance was noted among metallo-β-^lactamase (MBL)-producing isolates (reaching 87.5% in one study), especially those producing New Delhi metallo-β-lactamase (NDM) and Verona integron-encoded metallo-β-lactamase (VIM), which had the highest rates of resistance. The high activity of cefepime-enmetazobactam against Enterobacterales and selected lactose non-fermenting Gram-negative pathogens, including ESBL-producing and AmpC-producing isolates, makes it a potential carbapenem-sparing agent. The drug should be used after in vitro antimicrobial susceptibility testing in patients with infections caused by OXA-48, KPC, and MBL-producing isolates. Full article

The increasing prevalence of antibiotic resistance and the limited availability of antiviral therapeutics for pathogens such as human respiratory syncytial virus (hRSV) underscore the need for novel, plant-derived antimicrobial substances. In this study, we evaluated the antiproliferative, antibacterial, and antiviral activities of aqueous leaf extracts from two plants commonly found in North America, Osage orange (M. pomifera) and spearmint (M. spicata). Both extracts exhibited no significant cytotoxic or morphologic impact on HEp-2 human cancer cells up to 25 mg/mL. However, both extracts demonstrated strong dose-dependent antibacterial activity, significantly inhibiting replication of E. coli and S. aureus at concentrations ≥ 1 mg/mL. Antiviral assays revealed that both extracts inhibited hRSV infectivity, with spearmint extract showing higher potency (EC₅₀ = 1.01 mg/mL) compared to Osage orange (EC₅₀ = 3.85 mg/mL). Gas chromatography–mass spectrometry (GC-MS) identified three major extract constituents: 3-hydroxybenzyl alcohol, 4-hydroxybenzyl alcohol (Osage orange), and R-(-)-carvone (spearmint). Among these, only carvone significantly inhibited hRSV in vitro, suggesting its key role in spearmint’s antiviral activity. These findings highlight the therapeutic potential of Osage orange and spearmint leaf extracts, particularly as sources of water-soluble compounds with antimicrobial properties, and support further investigation into their mechanisms of action and broader clinical relevance. Full article

Helminth parasites of wild animals represent a major threat to the health of these animals, leading to significant losses in performance, health, and zoonotic implications. In some zoos, anthelmintics have traditionally been used to control these parasites, many of which are also zoonotic. Other actions, such as the removal of organic waste, have also been adopted. Few or no control measures are applied to free-ranging wild animals. Helminthophagous fungi are a promising biological alternative. When animals ingest fungal spores, they are excreted in their feces, where they trap and destroy helminth larvae and eggs, preventing and reducing the parasite load in the environment. Another alternative is to administer fungi by spraying them directly into the environment. This review aims to examine the use of helminthophagous fungi in the control of helminthiases in wild animals, highlighting their potential to minimize dependence on chemical treatments and promote sustainable animal breeding and production. There are many challenges to making this viable, such as environmental variability, stability of formulations, and acceptance of this new technology. These fungi have been shown to reduce parasite burdens in wild animals by up to 75% and can be administered through the animals’ feeding troughs. To date, evidence shows that helminthophagous fungi can reliably curb environmental parasite loads for extended periods, offering a sustainable alternative to repeated anthelmintic dosing. Their use has been linked to tangible gains in body condition, weight, and overall welfare in various captive and free-ranging wildlife species. Full article

Chimeric antigen receptor (CAR)-T immunotherapy has revolutionized the management of patients with relapsed/refractory B-cell hematological malignancies. There is emerging evidence that CAR-engineered cells—not only T cells, but also natural killers and macrophages—might have a crucial role in the treatment of autoimmune disorders and solid tumors. Moreover, given the burden of chronic infectious diseases, the mortality and morbidity of infections in immunocompromised individuals, and the development of multidrug-resistant pathogens, including bacteria, fungi, and mycobacteria, a need for novel and personalized therapeutics in this field is emerging. To this end, the development of CAR cells for the management of chronic infections has been reported. In this literature review, we summarize the ongoing clinical and pre-clinical data about CAR cell products in the field of infectious diseases. Currently, clinical studies on CAR immunotherapy for infections mainly concern human immunodeficiency virus infection treatment, and data regarding other infections largely originate from preclinical in vitro and in vivo models. In the era of personalized medicine, effective and safe therapies for the management of chronic infections and infectious complications in immunocompromised patients are crucial. Full article

Blastocystis spp. is an enteric protist that is present worldwide. Despite being discovered a century ago, there is still much to be learned about its pathogenicity and transmission. Different subtypes (ST) of Blastocystis spp. have been identified in various hosts, including humans, birds, and insects, and there is potential for zoonotic transmission through contact between humans and animals. The prevalence of Blastocystis spp. in humans and macaques in Singapore was understudied, and the findings revealed a significant prevalence of the parasite, with rates of 90% and 100% observed in each respective Macaca fascicularis population 1 and 2, with main subtypes (ST1, ST2, ST3, and ST5). Using metagenomics, the different subtypes of Blastocystis spp. (comprising ST2, ST3, and ST17) were identified in a healthy Singaporean cohort. Additionally, seven incidental findings of Blastocystis spp. were discovered in human patients with other gut parasites, including two ST1, two ST2, two ST3, and one ST8. Several factors such as diet or reverse zoonotic transmission are suggested to play a role in Blastocystis sp. subtype distribution. Full article

Tuberculosis (TB) is an ancient and re-emerging granulomatous infectious disease that continues to challenge public health. Early diagnosis and prompt effective treatment are crucial for preventing disease progression and reducing both morbidity and mortality. These steps play a vital role in infection control and in lowering death rates at both individual and population levels. Although diagnostic methods have improved sufficiently in recent decades, TB can still present with ambiguous laboratory and imaging features. This ambiguity can lead to diagnostic pitfalls and potentially disastrous outcomes due to delayed diagnosis. In this article, we present a case of TB that was difficult to diagnose. The disease had invaded the mediastinum, right atrium, right coronary artery, and inferior vena cava (IVC), resulting in Budd–Chiari syndrome. This rare presentation created clinical, laboratory, and radiological confusion, resulting in a diagnostic dilemma that ultimately led to open cardiac surgery. The patient initially presented with progressive shortness of breath on exertion and fatigue, which suggested possible heart disease. This suspicion was reinforced by computed tomography (CT) imaging, which showed infiltrative mass lesions predominantly in the right side of the heart, invading the right coronary artery and IVC, with imaging features mimicking angiosarcoma. Although laboratory findings revealed an exudative effusion with lymphocyte predominance and elevated adenosine deaminase (ADA), the Gram stain was negative for bacteria, and an acid-fast bacilli (AFB) smear was also negative. These findings contributed to diagnostic uncertainty and delayed the confirmation of TB. Open surgery with excisional biopsy and histopathological analysis ultimately confirmed TB. We conclude that TB should not be ruled out solely based on negative Mycobacterium bacteria in pericardial effusion or AFB smear. TB can mimic aggressive tumors such as angiosarcoma or lymphoma with invasion of the surrounding tissues and blood vessels. Awareness of the clinical presentation, imaging findings, and potential diagnostic pitfalls of TB is essential, especially in endemic regions. Full article