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Meta-analysis of growth and inactivation kinetics of Legionella.

Quantitative risk assessments intended to inform evidence-based water management plans and public health targets for Legionella in engineered water systems are constrained by fragmented and heterogeneous growth and inactivation kinetics. We conducted a meta-analysis of 25 growth and 39 thermal- and chemical-inactivation studies, fitting microbial persistence models to harmonize parameters. Nonlinear models outperformed first-order formulations, indicating that lag phases and resistant or protected subpopulations are central to Legionella persistence. Random forest analysis identified environmental and methodological drivers of variability based on 226 growth rates and reduction times for thermal (209) and chemical (135) inactivation. Growth was primarily governed by temperature, nutrient availability, and compatible Legionella-host pairings; thermal inactivation by quantification method, temperature, and turbidity; and chemical inactivation by inoculum size, disinfectant type, concentration, and host-associations. Accordingly, temperature-dependent growth parameters and exposure metrics for heat, free-chlorine, and monochloramine, expressed as TT (Temperature×time) and CT (Concentration×time), were derived as condition-specific inputs for predictive models. Growth optima around 37-40 °C, together with lag-time estimates, indicate that hot-water temperature setbacks and energy-saving practices may favor Legionella proliferation under repeated or prolonged lukewarm exposure. Culture- and viability-based TT differences highlight the need to consider viable‑but-non-culturable persistence in monitoring programs. CT comparisons suggest monochloramine may be advantageous because of its lower apparent sensitivity to host-associated protection. Although limited by restricted experimental conditions, the findings show that predictive models should account for microbial ecology, water matrix effects, and quantification endpoints. Future kinetic studies should prioritize realistic multi-host systems, strain pre-adaptation, complementary viability measurements, and standardized protocols and reporting to ensure reproducibility and enable robust system-level predictive modeling.

Legionella

Genomic determinants underlying biogenic amine detoxification phenotypes in food-associated lactic acid bacteria: Mechanism, evolutionary origin, and relevance to fermented food safety.

Biogenic amines (BAs) are toxic metabolites that accumulate in fermented foods and pose significant food safety concerns. Although several lactic acid bacteria (LAB) have previously been reported to exhibit strain-specific BA-degrading phenotypes, the genetic determinants underlying these activities have remained largely uncharacterized. Here, we analyzed 8251 LAB genomes to validate BA-degrading phenotypes. We predicted five BA-associated genes, including two direct biogenic amine-degrading genes (BADGs), mco and patA, and three polyamine-modifying genes (PMGs), speG, paiA, and bltD. Among BADGs, mco was broadly distributed across LAB and strongly enriched across food-associated niches. patA, organized within a conserved potD-glnB-potABC-patA cassette, is a putative, functionally distinct BADG in LAB, revealing a nitrogen-responsive polyamine uptake-catabolism module. Phylogenomics, phylogenetic reconciliation, and synteny analysis established that all five genes entered the LAB through episodic horizontal gene transfer followed by lineage-specific fixation. GC compositional bias and mobile genetic element association further corroborated the horizontal origin of the two BADGs. Structural analysis confirmed the conservation of catalytic core residues of BADGs across LAB, indicating strong purifying selection. Phenotype-to-genotype correlation with experimentally reported LAB suggested mco as a reliable genomic predictor of degrading phenotype. Integration of degradation and biosynthetic profiles predicted multiple LAB species capable of both synthesizing and degrading BA, along with 1823 genomes with degradation potential but lacking detectable BA biosynthesis genes. This study provides the first large-scale genome framework linking BA-degrading phenotypes with their genetic determinants in LAB and offers a rational basis for selecting BA-detoxifying strains for fermented food applications.

Biogenic Amines

Molecular characterization and biological characteristics of a highly pathogenic recombinant ALV-J strain (HUE2023) with cross-clade gp85 recombination.

Avian leukosis virus subgroup J (ALV-J) has undergone extensive diversification into phylogenetically distinct clades, yet whether recombination between these clades within the gp85 envelope glycoprotein generates variants with altered pathogenicity has received little direct investigation. A field strain (HUE2023) was recovered from breeding roosters displaying vascular tumors. The viral genome was sequenced and subjected to phylogenetic and recombination analyses. The three-dimensional structure of gp85 was predicted with AlphaFold3; electrostatic surface potentials and surface hydrophobicity were computed using the Adaptive Poisson-Boltzmann Solver and the Eisenberg hydrophobicity scale, respectively. Pathogenicity and immunosuppressive effects were assessed in Hy-Line Brown chickens. Recombination analysis revealed that HUE2023 is an inter-clade recombinant derived from Clade 1.1 (major parent: JS14NT01) and Clade 1.2 (minor parent: JS09GY3). A single-residue deletion at position 61 within receptor-binding domain 1 (RBD-1), unique to the recombinant, induced a localized conformational rearrangement that generated a concentrated electronegative surface patch and a contiguous hydrophobic pocket not observed in either parental gp85. Animal challenge showed that HUE2023 is highly pathogenic: female chickens in the high-dose group reached only 61% survival and displayed significant growth retardation (P&#x202f;<&#x202f;0.05) together with marked immunosuppression. The recombination in the RBD-1 led to local conformational rearrangement, resulting in a concentrated and negatively charged surface area as well as a continuous hydrophobic pocket, which were never present in any of the parental gp85 sequences. These results indicate that gp85 recombination across clades can yield variants with fundamentally altered receptor-binding surfaces and argue for integrating structural surveillance into ALV-J monitoring programmes.

Animals

Genomic epidemiology of clinically critical antibiotic resistance in Salmonella enterica causing bloodstream infections across six Chinese provinces, 1994-2023.

Clinically critical antibiotic-resistant Salmonella enterica (S. enterica) causing bloodstream infections remains a public health challenge. Here, we aim to reveal the emergence and trends of clinically important antibiotic resistance in S. enterica causing bloodstream infections using 833 isolates from six Chinese provincial-level administrative areas during 1994-2023. We identified 48 serovars and 64 sequence types (STs). Overall, 8.52% of 833 isolates were resistant or had decreased susceptibility to ciprofloxacin, 4.32% and 6.84% reported resistance or decreased susceptibility to third- and fourth-generation cephalosporins (3GCs and 4GCs), 1.80% reported resistance to fosfomycin, and 2.16% reported resistance to azithromycin. Across these six regions, azithromycin and fosfomycin resistance is increasing, as is decreased susceptibility or resistance to ciprofloxacin, 3GCs, and 4GCs, especially among younger children and elderly people. Clinically prioritized antibiotic resistance also varies by region, serovar, and age group. S. Paratyphi A genotype 2.3.3 strains are mainly divided into 2 lineages distributed in Guangxi and Shanghai. Within the scope of this passive surveillance dataset, S. Typhi genotype 4.3.1.2.1 was identified as the earliest documented case among the collected isolates. Our retrospective and longitudinal genomic epidemiology study provides critical data for the formulation of treatment guidelines and policies for bloodstream infections and for the monitoring and control of antimicrobial resistance.

Humans

From commensal to pathobiont: The emergence of virulence-enhanced Escherichia coli in China's food-animal systems - insights with future implications.

A fundamental shift in Escherichia coli epidemiology is being driven by convergence of virulence determinants and antimicrobial resistance within linked human-animal-environment systems. In China, the rapid growth of food-animal production, extensive antimicrobial use, and complex food networks are accelerating the emergence and dissemination of virulence-enhanced E. coli pathobionts. This review synthesizes recent epidemiological, genomics, and outbreak data to characterize China's evolving landscape of food-animal-associated E. coli. We highlight a significant shift from classical pathotypes to hybrid lineages that simultaneously carry virulence factors and last-resort antibiotic resistance determinants, including mcr-1, tet(X4), and blaNDM. These traits disseminate rapidly via plasmid-mediated horizontal gene transfer, facilitating rapid adaptation and enabling cross-sectoral One Health transmission. National surveillance, foodborne outbreak investigations, and whole-genome sequencing data show that food-animal reservoirs are active evolutionary niches that drive pathogen diversity and fitness, rather than serving merely as contamination sources. Whole-genome sequencing also pinpoints high-risk clones (e.g., ST394) and plasmid-mediated co-selection of virulence and AMR. The emergence of hybrid pathotypes (e.g., STEC/ETEC) and AMR-virulence co-selection challenges traditional classification and limits the effectiveness of conventional surveillance approaches. The 2017 colistin ban reduced mcr-1, yet ongoing resistance and emerging tet(X4) demand integrated surveillance. Collectively, these findings call for reconceptualizing E. coli as a dynamic genomic entity embedded within a unified ecological network. Addressing this threat requires an integrated One Health strategy including genomic surveillance, agricultural antimicrobial stewardship, and coordinated food-environment-clinical monitoring to prevent high-risk clone emergence and global spread.

Animals

Autodissemination stations suppress Aedes notoscriptus mosquitoes and reduce Buruli ulcer risk in urban Australia: a randomized controlled field trial.

Aedes notoscriptus are mosquito vectors implicated in transmission of Mycobacterium ulcerans. This bacterium causes a destructive infection of skin and soft tissue called Buruli ulcer. Here we ran a randomized controlled trial in an urban Buruli ulcer endemic area in Melbourne, Australia to test whether autodissemination mosquito control stations, containing pyriproxyfen (larvicide) and Beauveria bassiana (entomopathogenic fungus), suppress Ae. notoscriptus populations. Six geographic areas each received 100 autodissemination stations for 8&#x2009;weeks, and six control areas received no stations between 25 January 2024 and 21 March 2024. The primary outcome measure was mosquito population numbers. After the trial, there was a 70% average reduction in mosquito egg counts among the six intervention areas compared to control areas (P&#x2009;=&#x2009;0.0076). In an ad hoc analysis, we then explored human Buruli ulcer notifications in treatment and control areas. After accounting for the 4.8-month mean incubation period, there was an 83% reduction in infection likelihood coinciding with peak intervention effect (intervention zones 1 case, control zones 6 cases, incidence ratio rate 0.167, 95% CI 0.0026-1.054, P&#x2009;=&#x2009;0.047). The effect was not observed during the same time period in the year previous or following 2024, when no interventions were undertaken. A strong correlation (R2&#x2009;=&#x2009;0.85) was observed between decreased disease risk and mosquito suppression. These data show that autodissemination traps can effectively lower urban mosquito populations and reduce the threat of Buruli ulcer in humans.

Animals

Herd-level heterogeneity of antimicrobial resistance in commensal Escherichia coli: A nationwide high-throughput survey of Australian pig herds.

Antimicrobial resistance in commensal Escherichia coli provides a useful indicator for overall antimicrobial resistance burden. We applied this approach to assess antimicrobial resistance within and between commercial pig herds across Australia. A high-throughput robotic workflow was used to isolate 2730 E. coli colonies from rectal contents collected in 2022 from healthy slaughter pigs (n&#x202f;=&#x202f;300) representing 30 herds (&#x223c;70% of national production). Up to 94 isolates per herd underwent antimicrobial susceptibility testing using the Robotic Antimicrobial Susceptibility Platform. Isolate- and herd-level antimicrobial resistance indices were calculated, weighting antimicrobials by their human health importance. Resistance to first-line agents was widespread: ampicillin 77% and tetracycline 79%. By contrast, resistance to critically important antimicrobials was rare (ciprofloxacin 0.11%; extended-spectrum cephalosporins 0.04%), and no clinical resistance to carbapenems or colistin was detected. Overall, 56.9% of isolates were multi-class resistant. Herd-level antimicrobial resistance within indices ranged from 1.51 to 5.76, revealing substantial between-herd heterogeneity. Three herds carried critically important antimicrobials-resistant isolates that would likely have been missed using conventional, lower-density sampling approaches. Whole-genome sequencing identified fluoroquinolone-resistant isolates belonging to ST10 and ST69 (both qnrS1), and ST744 (Quinolone Resistance Determining Region mutations plus blaCTX-M-27). By testing approximately tenfold more isolates than conventional surveys, we uncovered considerable antimicrobial resistance with heterogeneity within and between animals and herds, including farm-specific variability. This expanded sampling also enabled detection of critically important antimicrobial resistance at very low prevalence. In conclusion, high-throughput, high-density testing offers a practical early-warning system and herd-level benchmark to inform surveillance and targeted interventions.

Animals

Diagnostic accuracy of bronchoalveolar lavage fluid-based testing for pulmonary cryptococcosis: A systematic review and meta-analysis.

BACKGROUND: Pulmonary cryptococcosis(PC) presents diagnostic challenges because of its non-specific clinical and radiological manifestations. Bronchoalveolar lavage fluid (BALF)-based testing, which includes latex agglutination (LA) and lateral flow assay (LFA), offers a minimally invasive diagnostic method, yet its pooled diagnostic accuracy remains unclear. METHODS: We systematically searched PubMed, Embase, Cochrane Library, and Scopus from inception to May 2026. Studies evaluating BALF-based testing for PC with extractable 2 &#xd7; 2 data were included. The methodological quality of relevant studies was assessed by the QUADAS-2 tool. Pooled sensitivity, specificity, likelihood ratios, and diagnostic odds ratio (DOR) were estimated using a bivariate random-effects model. Subgroup analyses were performed by testing method and reference standard type. Heterogeneity was evaluated through paired forest plots, HSROC visualization, and exploratory bivariate meta-regression. RESULTS: The pooled sensitivity was 0.87 (95% CI: 0.81-0.91), and the specificity was 0.99 (95% CI: 0.982 - 0.995). The pooled positive likelihood ratio (PLR) was 88.00 (95% CI: 47.39 - 163.42), the negative likelihood ratio (NLR) was 0.13 (95% CI: 0.09 -0.20), and the DOR was 658.50 (95% CI: 285.36-1519.55). No significant threshold effect or publication bias was detected. Exploratory meta-regression suggested a possible assay-method effect in the joint model (P = 0.03), mainly driven by specificity (P = 0.01). CONCLUSIONS: The study demonstrates the high accuracy of CrAg in BALF for the diagnosis of pulmonary cryptococcosis, supporting its role as an important adjunctive diagnostic tool, particularly when tissue biopsy is not feasible or rapid results are needed. Larger prospective studies with standardized protocols are needed to validate these estimates.

Humans

Genome mining of alkaliphilic cyanobacterial consortia: identification of biosynthetic gene clusters in Sodalinema and associated heterotrophs.

Alkaline soda lakes are high-pH environments that host specialized microbial communities with potential for biotechnology and natural product discovery. We characterized three Sodalinema-dominated cyanobacterial consortia enriched from Canadian soda lakes over 510 days. Using hybrid metagenomic sequencing and metatranscriptomics across pH, alkalinity, and temperature gradients, we reconstructed high-quality metagenome-assembled genomes and assessed functional activity. All consortia converged toward cyanobacteria dominance and exhibited temperature optima between 21&#xb0;C and 30&#xb0;C. Phylogenetic analysis placed Sodalinema genomes within a distinct clade affiliated with Candidatus Sodalinema alkaliphilum. Genomic analysis indicated complete biosynthetic pathways for vitamin B5, vitamin B7, and the molybdenum cofactor, but incomplete pathways for vitamins B1, B9, and B12, consistent with patterns observed in Sodalinema yuhuli. Metatranscriptomic profiles showed increased expression of genes involved in phycocyanin and carotenoid biosynthesis at pH 10.2 relative to pH 8.5. Biosynthetic gene cluster analysis revealed that most secondary metabolic potential resided in heterotrophic community members. Roseinatronobacter encoded pathways for N-acyl homoserine lactones, osmoprotectants, betalactones, and prodigiosin, while Alkalimonas, Wenzhouxiangella, and members of the Kiloniellales encoded clusters for lanthipeptides, cyclodipeptides, hydrogen cyanide, and pyrroloquinoline quinone. These findings indicate functional partitioning within the consortia and highlight the contribution of heterotrophs to secondary metabolism.IMPORTANCEAlkaline soda lakes contain microbial communities adapted to high pH that remain underexplored for biotechnology. This study focuses on Sodalinema, a filamentous cyanobacterium that dominates enriched consortia from Canadian soda lakes, and its associated heterotrophic partners. We show that while Sodalinema drives primary productivity, heterotrophic bacteria encode most of the pathways for antimicrobial and signaling compounds. These interactions may support community stability and defense against competing microorganisms. By linking genomic potential with gene expression, this work identifies alkaline cyanobacterial consortia as a source of bioactive compounds and provides a framework for exploring extremophilic microbial communities for natural product discovery.

Sodalinema

Interface-dependent V. parahaemolyticus biofilm under varying temperatures, media, and oxygen conditions: implications for seafood safety.

Vibrio parahaemolyticus biofilms play a critical role in pathogen persistence in marine and seafood-processing environments, where oxygen availability, temperature, and surface interfaces vary widely. This study investigated biofilm development by three strains on partially submerged stainless-steel coupons under gas-liquid-wall (GLW) and fully submerged (SM) interfaces. Viable cell counts (log&#x2081;&#x2080;CFU/cm2) along with normalized protein concentration per viable cell (nProt) and normalized polysaccharide concentration per viable cell (nPol) were measured, under aerobic and anaerobic conditions across a temperature range of 15-30&#xa0;&#xb0;C, using tryptic soy broth with 3% NaCl (TSB) and seawater-based medium (SW). GLW biofilms consistently exhibited higher cell counts (6.4-7.3 log&#x2081;&#x2080;CFU/cm2) compared to SM biofilms (5.9-6.3 log&#x2081;&#x2080;CFU/cm2), suggesting that enhanced oxygen diffusion promotes bacterial proliferation. Conversely, SM biofilms exhibited significantly higher nProt and nPol levels (p&#xa0;<&#xa0;0.001), indicating increased production of the extracellular polymeric substance (EPS) matrix under low-oxygen, high-nutrient conditions. Microscopy and three-dimensional surface plot analyses revealed relatively uniform biofilm layers at the GLW interface, whereas SM biofilms formed heterogeneous, tower-like structures. EPS production was further influenced by medium composition, oxygen, and temperature. SM biofilms grown in SW exhibited significantly higher nProt and nPol than those in TSB under aerobic conditions (p&#xa0;<&#xa0;0.001), indicating enhanced matrix stabilization. Under anaerobic conditions at 15&#xa0;&#xb0;C, nProt and nPol were higher, whereas under aerobic conditions, peak nProt and nPol occurred at elevated temperatures. These findings highlight a trade-off between bacterial growth and matrix production and provide insight into biofilm adaptation and persistence in seafood-processing environments. These insights may help develop improved biofilm control and seafood safety management.

Biofilms

Whole genome sequencing of unusual Hepatitis C virus subtypes and drug resistance analysis during direct-acting antiviral therapy in India.

INTRODUCTION AND OBJECTIVES: Pangenotypic direct-acting antivirals (DAA) are effective against highly prevalent Hepatitis C virus (HCV) subtypes, but have been clinically validated almost exclusively in high-income countries. Unusual HCV subtypes may carry natural polymorphisms, potentially impacting DAA susceptibility. We conducted full-genome characterization and resistance analysis of unusual HCV subtypes in patients receiving DAA treatment. PATIENTS AND METHODS: In this prospective hospital-based study, eligible patients were screened for anti-HCV antibodies and active infection was confirmed by diagnostic 5'NCR-based HCV RNA detection. Genotyping was performed by core region sequencing, and viral load quantified by real-time PCR. For whole genome sequencing, multiplex primers were designed using alignments of global reference sequences. Sequencing was carried out using the Oxford Nanopore Technology platform. Phylogenetic analysis used multiple sequence alignment and the HCV-GLUE resource for resistance-associated substitution (RAS) analysis. RESULTS: Predominant genotype was genotype 3 in 64.3% (n = 45); genotype 6 in 21.4% (n = 15); and genotype 1 in 14.2% (n = 10). Unusual HCV subtype 6xa was detected in two patients and showed no NS5A resistance mutations. One genotype 3b patient relapsed at 24 weeks post-DAA treatment completion and carried NS5A resistance-associated substitutions 30 K and 31 M both at baseline and at relapse, conferring high-level resistance to NS5A inhibitors. CONCLUSION: This is the first report from India of whole genome sequencing of HCV subtype 6xa. The identification of NS5A resistance mutations in the 3b relapse case underscores challenges for global HCV elimination strategies.

Humans

Genomic and One Health insights into Vibrio parahaemolyticus from environmental, seafood and clinical sources.

Vibrio parahaemolyticus is a leading cause of seafood-borne gastroenteritis worldwide, with climate warming facilitating its spread to high-latitude areas. In this study, we analyzed 212 genomes of environmental and seafood-associated isolates collected from seven cities in Zhejiang Province, China (2019-2024), alongside 228 clinical genomes from public databases. The 212 isolates were assigned to 172 sequence types (STs), with ST490 being the most frequent (5/212, 2.36%). Forty-four serotypes were identified, dominated by OL3:KUT (12.68%). High ST and serotype diversity were observed across different sample types and sources, with median pairwise single nucleotide polymorphisms (SNPs) ranging from 57,431 to 58,378, indicating comparable genetic diversity across groups. All isolates carried tlh and T3SS1 but lacked tdh and T3SS2. Resistance rates against ampicillin and cefazolin were 54.72% (116/212) and 44.34% (94/212), respectively, with multidrug resistance (MDR) detected in nine isolates, predominantly from seafood (7/9). A total of 63 distinct antimicrobial resistance genes (ARGs) spanning seven classes were identified. Isolates from aquaculture farms and wet markets exhibited greater resistance category diversity and higher ARG carriage than those from coastal or riverine sites. In contrast, the 228 clinical isolates harbored only 25 ARGs across two classes, with a significantly lower proportion of isolates carrying multiple ARG classes (0.44% vs. 6.13%, P&#xa0;<&#xa0;0.001). Human isolates formed tighter phylogenetic clusters, although a minority were closely related to environmental/foodborne strains. Overall, our findings demonstrate the genetic diversity and resistance potential of V. parahaemolyticus across environmental, seafood, and clinical sources, highlighting the importance of the One Health approach to comprehensive public health risk assessment.

Vibrio parahaemolyticus

Coordinated use of three homocysteine methyltransferases supports l-methionine biosynthesis and environmental adaptation among plant-associated bacteria.

Plant pathogens colonize multiple plant-associated habitats throughout their life cycle, encountering distinct nutrient conditions and microbial communities. l-methionine is required for bacterial growth and environmental adaptation. However, how plant pathogens coordinate l-methionine biosynthetic pathways to adapt to different plant-associated environments remains poorly understood. Here, using the plant pathogen Xanthomonas campestris pv. campestris strain XC1 as a model, we show that three homocysteine methyltransferase pathways allow XC1 to catalyze the final step of l-methionine biosynthesis using different methyl donors and cofactors under different environmental conditions. Bioinformatic and transcriptional analyses identified three homocysteine methyltransferase-associated operons in XC1, mesMXD, mmuPM, and metHRHaHb, corresponding to the MesD-, MmuM-, and MetHaHb-dependent pathways, respectively. MesD uses an endogenously synthesized methyl donor and functions as the dominant homocysteine methyltransferase under l-methionine-limiting conditions, supporting bacterial growth, intracellular l-methionine accumulation, and full virulence. Furthermore, MmuM enables XC1 to use plant-derived S-methylmethionine for l-methionine biosynthesis, whereas MetHaHb enables XC1 to use vitamin B12 supplied by a neighboring bacterium for l-methionine biosynthesis in co-culture. Expression analyses showed that mesMXD was the only homocysteine methyltransferase-associated operon that responded to l-methionine availability, and its expression also decreased when S-methylmethionine- or vitamin B12-dependent pathways supported l-methionine biosynthesis. Comparative genomic analysis further showed that the three-homocysteine methyltransferase configuration is conserved in Xanthomonas and is also present in other plant-associated bacteria. Together, these findings show that a plant pathogen can coordinate endogenous, plant-derived, and microbially supported homocysteine methyltransferase pathways to maintain l-methionine biosynthesis, providing a metabolic strategy for adaptation to plant-associated environments.

Methionine

MIC-based tuberculosis drug susceptibility testing using Sensititre MYCOTB: a diagnostic accuracy meta-analysis.

Accurate drug susceptibility testing (DST) is crucial for designing effective regimens for multidrug-resistant (MDR) and pre-extensively drug-resistant tuberculosis (pre-XDR TB). Sensititre MYCOTB enables simultaneous determination of minimum inhibitory concentrations (MICs) for multiple drugs, but its diagnostic performance varies across studies. This meta-analysis evaluated the diagnostic performance of Sensititre MYCOTB for key MDR and pre-XDR TB drugs. The protocol was registered in PROSPERO (CRD420251230599). PubMed, Cochrane, Google Scholar, Scopus, ONOS, Web of Science, ScienceDirect, and registries were systematically searched for studies published between 2010 and 2025. Studies comparing the Sensititre MYCOTB with reference DST for Mycobacterium tuberculosis complex (MTBC) were included. Bias assessment and pooled diagnostic accuracy estimates were generated. Fourteen studies, including 1,728 isolates, were analyzed. Rifampicin and isoniazid demonstrated high sensitivity (0.976 [95% CI: 0.94-0.99] and 0.977 [95% CI: 0.95-0.99]) and specificity (0.958 [95% CI: 0.84-0.98] and 0.957 [95% CI: 0.83-0.99], respectively) with low heterogeneity. Amikacin, kanamycin, and ofloxacin demonstrate good diagnostic accuracy, with high specificity (>0.98 [95% CI]). Moderate diagnostic accuracy was observed for ethambutol, streptomycin, ethionamide, and rifabutin. Cycloserine, moxifloxacin, and para-aminosalicylic acid showed inconsistent performance despite excellent specificity (>0.97 [95% CI]). Sensitivity analysis partially improved pooled sensitivity for moxifloxacin 0.801 (95% CI: 0.585-0.924) and para-aminosalicylic acid 0.76 (95% CI: 0.518-0.894), whereas cycloserine remained at 0.436 (95% CI: 0.190-0.725), although heterogeneity persisted. Sensititre MYCOTB DST demonstrates high diagnostic accuracy for MDR-TB and pre-XDR-TB drugs, while caution is required with cycloserine, moxifloxacin, and para-aminosalicylic acid. These findings support the integration of MIC-based testing into clinical decision-making.

Microbial Sensitivity Tests

Triazole resistance in clinical Aspergillus fumigatus isolates in India, a multicenter surveillance study.

BACKGROUND: Triazole resistance in Aspergillus fumigatus is a global public health concern associated with treatment failure, notably in invasive aspergillosis. However, population-level data on triazole resistance from India remain limited, with most reports originating from single-center studies. METHODS: We conducted a multicenter surveillance study to assess the prevalence of triazole resistance among clinical A. fumigatus isolates across India. Antifungal susceptibility testing was performed using the CLSI broth microdilution method (M38-Ed3), and molecular characterization was conducted on resistant isolates. A total of 518 isolates were analyzed: 115 prospectively collected from 13 tertiary-care hospitals from 2015-2020, and 403 archived isolates obtained from the National Culture Collection of Pathogenic Fungi (1994-2020). RESULTS: The overall pooled prevalence of non-wildtype isolates was 4.1% for itraconazole (95% CI: 2.54-6.17%), 3.9% for posaconazole (95% CI: 2.39-5.94%), while 1.4% were resistant to voriconazole (95% CI: 0.55-2.77%). One multi-azole-resistant isolate from an immunocompromised, mold-active triazole-na&#xef;ve patient carried the TR34/L98H mutation, suggesting environmental acquisition. Prevalence of resistance did not differ significantly across geographic regions or between public and private sector hospitals. Linear regression analysis revealed a significant temporal increase in median MICs of all three licensed triazoles between 1994 and 2020. Approximately 29% of isolates exhibited amphotericin B MICs exceeding the epidemiological cutoff value; however, the clinical significance of this finding remains uncertain. CONCLUSIONS: Azole resistance among clinical A. fumigatus isolates in India remains uncommon (<5%), supporting the continued use of triazoles as first-line therapy. However, the observed temporal increase in triazole MICs underscores the need for sustained national surveillance to detect emerging resistance trends.

Aspergillus fumigatus

Regional genomic analysis of lineage distribution and transferable multidrug resistance among chicken-associated Salmonella Kentucky isolates in China.

Salmonella enterica serovar Kentucky is an important multidrug-resistant foodborne pathogen in the poultry meat supply chain. Although recent broader genomic studies have elucidated the population structure and epidemiological significance of major lineages in China (e.g., ST198 and ST314), the regional dynamics within local poultry supply chains remain insufficiently characterized. In this study, 31 chicken meat-derived isolates from Shanghai and 39 publicly available genomes from China were analyzed using antimicrobial susceptibility testing, whole-genome sequencing, phylogenetic analysis, conjugation experiments, and complete sequencing of representative plasmids. This enabled a systematic characterization of the molecular epidemiological features of the population and the mechanisms underlying resistance dissemination. Population genomic analysis revealed a lineage composition markedly different from the global epidemiological pattern: ST314 was the predominant sequence type among the Shanghai chicken-derived isolates (74.2%), whereas the internationally recognized high-risk clone ST198 accounted for only 25.8% of the local isolates. However, risk stratification analysis indicated that although ST198 was detected less frequently, it carried a significantly greater burden of acquired resistance genes and therefore represented a higher-risk resistant lineage. Functional and structural validation further elucidated the molecular basis of resistance dissemination within this high-risk lineage. Conjugation experiments confirmed the co-transfer of a multidrug resistance module carrying blaTEM-1 and blaCTX-M-267 to the recipient strain Escherichia coli J53. Complete plasmid analysis revealed that these two &#x3b2;-lactam resistance genes were co-localized on a 242-kb transferable plasmid flanked by Tn1331, Tn3, and multiple transposase-associated elements, thereby providing a structural basis for their horizontal transfer. This study provides important molecular epidemiological evidence for lineage-specific surveillance and risk-stratified control of resistant Salmonella in the poultry meat supply chain and further underscores the need for continuous monitoring of mobile genetic elements within a One Health framework.

Animals

Genomic and food-safety evaluation of Staphylococcus chromogenes in Chinese dairy milk.

Non-aureus staphylococci and mammaliicocci (NASM) cause mastitis and may contaminate milk and dairy products. Milk samples (n&#xa0;=&#xa0;1916) from cows with subclinical or clinical mastitis (SCM and CM, respectively) were collected from 28 large-scale (> 500 lactating cows) Chinese dairy farms. Overall, 999 NASM isolates representing 19 species were identified by MALDI-TOF MS and cpn60 sequencing, with Staphylococcuschromogenes, Mammaliicoccus sciuri and Staphylococcus haemolyticus being most prevalent. Antimicrobial resistance (AMR) was determined with disc diffusion; non-susceptible to penicillin was most common (SCM, 30% and CM, 29%) whereas cefoxitin non-susceptible NASM accounted for 8-10% of isolates; among these, 12.5% carried mecA but none carried mecC. Galleria mellonella was used to assess virulence of 78 strains of S. chromogenes, a dominant species; subsequently, 32 strains, representing higher- and lower-virulence in the Galleria model, were selected for whole-genome sequencing and comparative genomics. S. chromogenes isolates from CM had higher virulence (p&#xa0;<&#xa0;0.05) than those from SCM. The 32 genomes comprised 20 sequence types, indicating high genetic diversity. No robust genomic marker of Galleria virulence phenotype was identified in this selected WGS subset. Acquired resistance genes (n&#xa0;=&#xa0;5) were detected, including a first report of fusC in S. chromogenes; the fusC-positive isolate had an elevated fusidic acid MIC (8&#xa0;mg/L). Although S. chromogenes persisted in milk at 4&#xa0;&#xb0;C, pasteurization (64&#xa0;&#xb0;C for 30&#xa0;min) reduced viable counts to below detection. This study provided new insights into the prevalence, AMR, genomic diversity, and dairy-chain relevance of milk-derived NASM, particularly S. chromogenes. However, the genomic findings were based on an intentionally selected WGS subset and should be interpreted as hypothesis-generating rather than population-representative.

Animals

Genomic diversity and thermal niches of Aspergillus molds disrupting rind formation of surface-ripened cheeses.

Filamentous fungi play important roles in the development of surface-ripened cheese microbial communities and contribute to the aesthetics and flavors of these products. Much is known about the diversity and ecology of desirable cheese fungi, but our understanding of the natural history of cheese spoilage molds is limited. The goal of this work was to characterize the genomic diversity of Aspergillus species contaminating artisan cheeses and to identify how the abiotic environment of cheese (the substrate itself and temperature) may constrain the growth of Aspergillus. Comparative genomics identified two main species of Aspergillus, A. westerdijkiae and A. ostianus, as the spoilage molds across three different facilities in the Northeastern United States that experienced contamination events. Multiple genomic types of A. westerdijkiae were found across the different cheese production facilities, indicating that these contamination events are not caused by a single clonal strain. All A. westerdijkiae isolates produced ochratoxin A, but concentrations varied greatly across strains. RNA-sequencing of A. westerdijkiae on nutrient-rich lab media (malt extract agar) versus cheese curd agar identified a suite of pathways enriched in expression on cheese, including degradation of amino and fatty acids. Experiments measuring growth over a range of temperatures identified that spoilage Aspergillus species have a higher optimal growth temperature compared to desirable fungal species in cheese rinds and are outcompeted by Penicillium species at temperatures lower than 15&#xb0;C. Global fungal metabarcoding databases suggest that A. westerdijkiae is not normally found in natural habitats of the Northeastern United States, and it may be introduced to this region.IMPORTANCEOver the past decade, disruptive contamination events of Aspergillus spoilage molds have occurred at cheese production facilities in Massachusetts, Connecticut, and Vermont in the United States, causing aesthetic, flavor, and potential safety issues. Our work highlights independent introductions of different strains of A. westerdijkiae into multiple cheese facilities and suggests that temperature could be used to control the abundance of Aspergillus spoilage molds. Based on our analysis of the global distribution of A. westerdijkiae, it is not invading cheese facilities from local fungal populations and may be a contaminant in materials used for cheese production.

Aspergillus