Journal of Applied Microbiology最新文献

Aims: This study aimed to characterize a colistin- and carbapenem-resistant Pseudomonas aeruginosa ST1560 strain isolated from Guanabara Bay, Brazil, and to investigate the molecular mechanisms underlying its resistance phenotype.

Methods and results: Six surface water samples from Guanabara Bay were collected, yielding 71 P. aeruginosa subjected to antimicrobial susceptibility testing. Three isolates exhibited elevated minimal inhibitory concentrations (MICs) to colistin (≥512, 64, and 8 mg/L) in the absence of mcr genes (1-10). Among these, only strain CCVSU 5861 demonstrated carbapenemase confirmed by Blue Carba test. This strain was selected for whole-genome sequencing (Illumina). Genomic analysis identified the presence of blaKPC-2 and blaOXA-395, along with additional resistance determinants associated with aminoglycosides and fosfomycin. Genes involved in lipopolysaccharide modification, (arnA, arnT, and basS) were also detected, likely contributing to colistin resistance. The blaKPC-2 gene was located adjacent to the mobile genetic element ISKpn6, suggesting potential horizontal gene transfer.

Conclusions: The P. aeruginosa ST1560 displays a complex multidrug resistance profile, including resistance to both colistin and carbapenems. This phenotype appears to be mediated by a combination of acquired resistance genes and chromosomal mechanisms. The localization of blaKPC-2 within a mobile genetic element underscores the risk of dissemination in aquatic environments.

Aims: This study reports the first documented cases of Castellaniella spp. infection in dogs, describing associated clinical and pathological findings and characterizing a novel species within this genus.

Methods and results: Pleural effusions from two dogs presenting with acute respiratory distress and systemic illness were evaluated via cytology, bacterial culture, and next-generation sequencing. Both cases exhibited neutrophilic-macrophagic inflammation with intracellular gram-negative rods, primarily within macrophages. Bacterial culture failed to identify the organisms. Metagenomic analysis identified organisms belonging to the genus Castellaniella in both cases. In case 2, an unclassified Castellaniella species was detected, suggesting the presence of a previously undescribed species within the genus. Both dogs died shortly after presentation, and necropsy and histopathology findings were described.

Conclusions: Castellaniella spp. warrant consideration as potential emerging pathogens in domestic animals, challenging their previous classification as non-pathogenic environmental bacteria. The identification of a novel species also underscores the genus's genetic diversity and adaptive potential.

Chronic wounds, including diabetic foot ulcers, venous leg ulcers, and pressure ulcers, remain a major global healthcare challenge, associated with substantial morbidity, risk of limb loss, and high healthcare costs. Increasing evidence indicates that the wound microbiome modulates inflammation, tissue repair, and responses to therapy, thereby influencing clinical outcomes. This review summarizes current knowledge on the composition and function of chronic wound microbial communities and discusses their clinical relevance as prognostic biomarkers and therapeutic targets. Microbiome structure is shaped by wound etiology, chronicity, anatomical site, and host comorbidities. Dysbiosis and biofilm formation contribute to persistent inflammation, antimicrobial tolerance, and delayed healing. Advances in sequencing and multi-omics technologies have improved microbial characterization and enabled the identification of candidate microbial signatures associated with healing trajectories. Emerging microbiome-modulating strategies such as probiotics, bacteriophages, topical oxygen approaches and nanotechnology-based interventions show potential to shift wound ecosystems toward a pro-healing state; however, robust clinical validation remains limited. Further clinical studies are needed to validate microbiome-guided diagnostics and interventions and to establish standardized protocols for their application in clinical practice.

Aims: Cyanobacteria produce and release secondary metabolites in waterways, challenging drinking water treatment plants. Benthic Cyanobacteria, a group of species living at the bottom of waterbodies, have been identified as potential Taste and Odour (T&O) compound and toxin producers. Following an increase in customer T&O complaints about water produced from the SA-L1 Reservoir, this study was conducted to better understand benthic cyanobacetria growth patterns and establish whether the source of detrimental metabolites was pelagic or benthic.

Methods and results: A field-survey was performed from December 2014 to December 2015, during which physical samplers were deployed across a transect (1 m, 2 m, 6 m and 14 m depth). Biofilm and water samples were analysed for microbial community composition, chlorophyll-a (Chl-a), nutrients, T&O and toxins and their associated genes. Seasons and sampling depths impacted benthic communities, Chl-a concentrations and biofilm growth. Spring and autumn were established as peak growth periods for benthic Cyanobacteria. Water geosmin concentrations significantly correlated with the abundance of benthic Cyanobacteria. The potential for cylindrospermopsin and saxitoxin production was detected in this reservoir. Mat detachment mid-spring contributed to T&O dispersion.

Conclusions: Benthic Cyanobacteria are major geosmin contributors in this reservoir. The potential cylindrospermopsin-producer was demonstrated to be benthic, while the saxitoxin producer was identified as pelagic Dolichospermum circinale, which is recruited from bentic mats in spring. Utilities should consider regular monitoring of benthic mats, which provides the necessary evidence to better anticipate benthic and pelagic events, to in turn provide safe and palatable drinking water.

Aims: This study was designed to investigate the basis of atypical SAM resistance in Salmonella isolates from raw chicken that lacked extended-spectrum or inhibitor-resistant TEM β-lactamase variants.

Methods and results: The ampicillin (AMP) minimum inhibition concentrations (MICs) of all Salmonella isolates, blaTEM expression, and copy numbers of select isolates were determined. Plasmids from select Salmonella isolates were conjugated into E. coli recipients, and their AMP MIC, SAM resistance phenotypes, and blaTEM expression were quantified. SAM-resistant Salmonella isolates displayed ∼5.4-fold higher average AMP MIC and significantly elevated blaTEM expression levels compared to SAM-susceptible control. Conjugation experiments revealed differences in AMP MICs and SAM resistance phenotypes between Salmonella donors and E. coli transconjugants. An AMP MIC breakpoint of 1024 ppm ostensibly confers resistance to SAM in Salmonella.

Conclusion: Host-dependent blaTEM overexpression and resulting β-lactamase hyperproduction can lead to atypical SAM resistance in Salmonella. This study furthers our understanding of the understudied SAM resistance in Salmonella spp.

Aims: Ready-to-eat dry-cured meats are characterized by combined hurdles such as reduced water activity, elevated salt content, and variable temperatures. Listeria monocytogenes may survive or adapt under these conditions. This study aimed to evaluate the behavior of L. monocytogenes under combined water activity (aw), sodium chloride (NaCl), and temperature stresses using a dry-cured meat-based agar designed to simulate physicochemical conditions encountered on cured meat surfaces and during dry-curing and storage processing conditions. Socol, a traditional Brazilian dry-cured pork loin, was used as the reference product for the development of this model system.

Methods and results: The influence of aw (0.806-0.974), NaCl concentration (1.97-9.53% w/w), and temperature (0.9-31.1°C) on the growth and/or survival of L. monocytogenes was investigated over 12, 24, 48, and 72 h using a Central Composite Rotatable Design (CCRD). Survival after 72 h of incubation was also evaluated in simulated gastric fluid (SGF). The predictive models accurately described the effects of aw, NaCl, and temperature on bacterial behavior, showing excellent validation performance (R² ≥ 0.96; Af ≥ 1.14; Bf ≥ 1.14). Water activity was the dominant factor influencing pathogen behavior, with increased growth observed at aw > 0.92. Temperature had no significant inhibitory effect, indicating that L. monocytogenes can survive under refrigeration conditions. The bacterium was fully inactivated in SGF under all tested conditions.

Conclusions: The results emphasize the importance of water activity control to limit L. monocytogenes survival and growth in ready-to-eat dry-cured meats.

Aims: Pasteurella multocida (Pm) is one of the main pathogens causing bovine respiratory disease in China. The prevention and control measures against Pm are traditionally based on the use of broad-spectrum antibiotics. Previous studies have found that Pm is prone to developing antibiotic resistance and tolerance-related mutations when exposed to low concentrations of antibiotics, ultimately leading to challenges in the prevention and control of Pm. This study aimed to explore the role of the recO gene in Pm in mediating resistance and tolerance to fluoroquinolones.

Methods and results: Highly pathogenic Pm strains (fluoroquinolone-sensitive P3; enrofloxacin-induced resistant P32) were used. RNA-seq screened SOS response-related differentially expressed genes, with recO functionally verified. Its role in Pm's fluoroquinolone resistance/tolerance was clarified via MIC, MBC.The results showed that recO deletion reduced the bacterial tolerance by approximately 10-100-fold after 4 h of exposure to enrofloxacin (ENR) (p < 0.05), decreased the MBC value by 2-fold, and significantly prolonged the time required for resistance development.

Conclusions: In conclusion, inhibiting the expression of the recO gene in Pm not only reduces its resistance to fluoroquinolones but also delays the development of fluoroquinolone resistance. It is hypothesized that the recO gene could serve as a potential target for enhancing the efficacy of fluoroquinolones, thereby improving their antibacterial activity against Pm.

Aims: Targeting bacterial virulence is a promising alternative to traditional antibiotics. In this study, we aimed to identify microbiome-derived factors capable of suppressing virulence traits in pathogenic bacteria.

Methods and results: Eighty-two cell-free extracts (CFEs) applied at 10% (v/v), from human (n = 56) and soil (n = 26) bacterial isolates were screened for effects on planktonic growth, biofilm formation, and virulence using a Galleria mellonella infection model. Growth inhibition was rare, observed only for S. aureus exposed to a Bacillus flexus extract. Anti-biofilm activity was frequent against P. aeruginosa (65/82 CFEs, 79%), less so for S. aureus (20/82, 24%), and absent or reversed for E. coli. Eight CFEs improved larval survival in P. aeruginosa infections; one partly protected against S. aureus, while (47/82, 57%) increased mortality in E. coli-infected larvae. Thirty-eight of 65 anti-biofilm CFEs (58%) for P. aeruginosa tested positive (i.e. observed) in a violacein-based quorum-sensing inhibition assay. One extract (Streptococcus australis) also protected keratinocytes from P. aeruginosa-induced cell loss. Proteomic analysis of P. aeruginosa exposed to S. australis CFE indicated modulation of proteins associated with biofilm regulation and quorum-sensing pathways (e.g. the RsmA/CsrA family), consistent with anti-virulence activity.

Conclusions: Of 82 microbiome-derived CFEs, (65/82, 79%) partially inhibited P. aeruginosa biofilm formation, (20/82, 24%) affected S. aureus, and none inhibited E. coli biofilms. Ten % improved in vivo survival in P. aeruginosa infections, while (47/82, 57%) increased mortality in E. coli-infected larvae. Anti-virulence effects were pathogen-specific and occurred without measurable impacts on planktonic growth under the conditions tested.

Aims: To achieve optimal application of antimicrobials to poultry processing requires an understanding of the potential for resistance by foodborne pathogens such as Salmonella. The objective of this study was to use transposon sequencing (Tn-seq) to identify genetic factors required for Salmonella Typhimurium's tolerance to PAA.

Methods and results: A genome-saturated Tn5 mutant library (Input pool) was inoculated in two replicates into either 6% chicken meat extract (CME) or 11% diluted Luria-Bertani (LB) broth, both supplemented with 15 ppm PAA. Cultures were incubated for 90 minutes at 37°C. Viable Tn5 mutant cells recovered on LB agar plates were combined to form four output pools (two CME and two LB). Genomic DNA extracted from these pools were deep sequenced (Tn5-junction reads). Conditionally essential genes required for fitness in 6% CME and 11% LB were identified and subjected to pathway enrichment analysis (ShinyGO graphical gene-set enrichment tool). We identified two overlapping sets of conditionally essential genes (276 common genes) required for survival in the presence of PAA. In CME, 362 conditionally essential genes were identified, while LB media revealed 536 genes. Pathway enrichment analysis showed that these genes were significantly enriched in pathways such as pyruvate metabolism, the tricarboxylic acid cycle, fumarate reductase/succinate dehydrogenase (transmembrane subunit and 2Fe-2S iron-sulfur cluster binding domain), stress response, and oxidoreductase activity. Notably, genes previously shown to increase sensitivity to PAA upon inactivation (sdhC, zwf, pta, and icdA) were identified as conditionally essential in this study, further validating the Tn-seq data.

Aims: Multidrug-resistant (MDR) foodborne pathogens pose an urgent global threat. We surveyed retail foods in Ismailia Governorate, Egypt, to identify highly resistant strains and develop a phage-based biocontrol strategy.

Methods and results: Staphylococcus aureus was detected in 45% of samples (70% meat) and Escherichia coli in 52.5% (80% in arugula), often exceeding safety standards. From 40 representative isolates, the most resistant strains: Staphylococcus aureus SL4 from lettuce (GenBank OR646818; MAR index 0.44) and Escherichia coli ER2 from arugula (GenBank OR646817; MAR index 0.72), were selected for phage targeting. Two sewage-derived lytic phages, STB (Siphoviridae; host SL4) and ECB (Podoviridae; host ER2), were isolated. Both had ideal biocontrol traits: latent periods ∼10 min, burst sizes ∼2×10¹¹ PFU cell⁻¹, and stability at 28-55 °C and pH 5-9. In simulated decontamination trials, phage soaking on meat (CFU g⁻¹), arugula, and cutting boards (CFU cm⁻²) reduced bacterial counts by 3.4-6.4 log₁₀ after 1 h, 4.7-6.4 log₁₀ after 3 h, and near-complete eradication by 6 h (9.20-8.58 log₁₀ for SL4, 9.41-7.86 log₁₀ for ER2). Soaking, spraying, and the phage cocktail all outperformed 5% vinegar by 0.8-3.8 log₁₀; the cocktail broadened host range but had slightly slower kill kinetics.

Conclusions: Optimized phage application enables rapid, chemical-free eradication of MDR pathogens from foods and food-contact surfaces.