Current Microbiology最新文献

Lactic acid bacteria that persist in the gastrointestinal tract contribute not only to nutrient metabolism but also to the regulation of immune responses. Accordingly, these microorganisms and their components are classified as probiotics when administered in adequate amounts and shown to confer health benefits on the host. However, the mechanisms by which they interact with host cells remain poorly understood. Extracellular vesicles (EVs) have been shown to modulate functions in recipient cells, including both mammalian cells and microorganisms. Surface-layer associated proteins (SLAPs) are preferentially produced by Gram-positive syntrophic bacteria and are secreted to associate with the cell envelope. The dysfunction of SLAPs leads to disruption of the cell envelope, cell division, and cell-cell communication. In this study, we identified SLAP in extracellular vesicle-like nanoparticles (EV-LNPs) derived from Lactobacillus helveticus strain GIF001 (closed to strain VHProbi Y21) and observed enhanced immune responses-including nitric oxide and interleukin-6 production-in RAW264.7 cells when SLAP was added to the culture medium, either in the form of EV-LNPs or as a purified protein. Notably, L. helveticus EV-LNPs were internalized within 3 h and enhanced NF-κB signaling in response to lipopolysaccharide. Intracellular overexpression of SLAP led to increased activity of the inducible nitric oxide synthase promoter about 1.5 times, whereas deletion of the SLAP domain abolished this enhancement. Incubation of RAW264.7 cells with EV-LNPs or SLAP enhanced the uptake of Escherichia coli. These results suggest that EV-LNPs from L. helveticus may upregulate immune responses by activating host cell communication via SLAP.

Salmonella Typhimurium has long been one of the most frequently isolated serovars in animal and human infections. Pork has been involved in the dissemination of S. Typhimurium to humans and plays an important role in the epidemiology of this infection. This study aimed to characterize genotypically and phenotypically S. Typhimurium strains isolated from swine in Brazil. The genomic relatedness among 17 of the 18 S. Typhimurium genomes was ≥ 90% according to Gegenees analysis, while ANI analysis showed ≥ 98.2% similarity across all 18 genomes, with most strains belonging to SNP cluster PDS000201117.2. Virulence factors and stress-related genes were searched at NCBI Pathogen Detection. All strains carried the iroB, iroC, sinH, asr, golS, and golT genes. Under acid stress, all strains survived after 10 min and 1 h. Under oxidative stress, 17 (94.44%) strains survived after 10 min, and 11 (61.11%) strains survived after 1 h. The invasion rates in Caco-2 cells ranged from 37.50% to 100%, and survival in the macrophage assay ranged from 37.50% to 87.50%. In conclusion, the close genetic relationship observed among the S. Typhimurium strains isolated from swine studied may suggest that a predominant subtype may have prevailed in Brazil's swine sources. The high prevalence of some heavy metal tolerance encoding genes is alarming due to their potential to influence in the co-selection of S. Typhimurium strains resistant to antibiotics. Moreover, the presence of some virulence genes and the notable stress survival and cell invasion capacities highlighted the importance of surveillance to prevent the dissemination through food of virulent S. Typhimurium strains present in livestock to humans.

The emergence of antimicrobial resistance (AMR) poses a critical global health threat, underscoring the interconnectedness of human, animal, and environmental health within the One Health framework. This study describes two multidrug-resistant (MDR) extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli strains, classified as critical priority bacteria by the World Health Organization, identified in recreational freshwater in Brazil. A cross-sectional study was conducted to investigate the presence of ESBL-producing Enterobacterales in water samples from a river in a touristic Brazilian city, including the waterfall, stream, and areas outside the city center. Bacteria were cultured and isolated on MacConkey agar supplemented with ceftriaxone, identified by MALDI-TOF, and antimicrobial susceptibility testing was performed by disk diffusion. ESBL-producing strains were then subjected to whole-genome sequencing and bioinformatic analysis. Two ESBL-producing E. coli strains were recovered from the waterfall (USP-MG-W) and steam (USP-MG-B). These strains showed an MDR profile, with resistance to β-lactams, tetracyclines, aminoglycosides, quinolones and phenicols, but remained susceptible to carbapenems. USP-MG-B and USP-MG-W harbor the ESBL genes bla_CTX-M-15 and bla_CTX-M-65, respectively, highlighting their broad-spectrum cephalosporin resistance and clinical relevance. Predicted heavy metal resistance genes suggest AMR co-selection. USP-MG-B belongs to pandemic E. coli sequence type (ST) 10, closely related to Brazilian human isolates. At the same time, USP-MG-W is part of clonal complex 155 (ST683), associated with international livestock and poultry strains. Remarkably, this study is the first report of E. coli ST10 in Brazilian rivers and ST683 with bla_CTX-M-65 in Brazil. The detection of ESBL clones in freshwater underscores the role of the environment as a reservoir of critical priority pathogens. These findings highlight the urgent need for a One Health approach to the AMR crisis and for strengthening the epidemiological surveillance of ESBL-producing Enterobacterales beyond human settings.

Deuterolysin metalloproteases (M35s), a family of zinc-dependent proteolytic enzymes, function as key virulence factors in bacteria and fungi. In plant-pathogenic fungi, these proteins induce host cell death and suppress plant chitinases, thereby protecting fungal cell walls from degradation and preventing the release of chitin oligomers that trigger plant immune responses. Here, we investigated the evolutionary history of the M35 gene family across Dothideomycetes fungi using genomic data from 79 Dothideomycetes species and 61 Corynespora cassiicola isolates. Predicted M35 proteins were classified as putative effectors or putative non-effectors based on bioinformatic criteria, with putative effectors defined as secreted proteins capable of manipulating host immunity. Across Dothideomycetes, we identified 146 M35 genes, of which 107 were putative effectors and 39 putative non-effectors. Family size varied widely (0-7 genes per genome), with the largest repertoires observed in the plant-pathogenic species Botryosphaeria dothidea and Diplodia seriata. In C. cassiicola, most isolates harbored three M35 genes per genome. Phylogenetic analysis revealed four distinct sub-clades within the C. cassiicola M35 family. Sub-clades Cc_M35_1.1 and Cc_M35_2.1 showed evidence of gene duplication followed by retention across most isolates, whereas Cc_M35_1.2 and Cc_M35_2.2 exhibited patterns consistent with gene loss. Notably, putative effector M35s in sub-clade Cc_M35_2.2 were exclusively recovered from isolates associated with cotton, suggesting a specialized role in C. cassiicola-cotton interactions. During soybean infection, all three M35 genes in C. cassiicola were expressed, but CC_29_g9669 (Cc_M35_1.1) displayed the highest relative expression, peaking at 8 days post-inoculation.

Escherichia coli (E. coli) is a leading cause of urinary tract infections (UTIs) and diabetic foot ulcers, especially in individuals with weakened immune systems. Pathogenic strains (PEC) employ virulence genes including fimH, hlyA, usp, traT, and papC to establish infection, evade immune responses, and cause tissue damage. However, despite its clinical significance, there is a lack of integrative studies profiling virulence genes and identifying mutations in PEC, which may contribute to pathogenicity, antimicrobial resistance, and limitations in targeted therapy. In this study, 210 E. coli isolates were collected from diabetic patients with UTIs or foot ulcers at Hayatabad Medical Complex (HMC), Peshawar. Identification was performed through biochemical profiling and confirmed via PCR targeting 16S rRNA. Antibiotic resistance was assessed using the Kirby-Bauer disk diffusion method. The presence of virulence genes was detected by PCR, and sequencing data were analyzed using bioinformatics tools such as Multiple Sequence Alignment, PROVEAN, InterProScan, and SWISS-MODEL. A significantly higher prevalence of all five virulence genes was observed in diabetic isolates (fimH OR = 2.63; hlyA OR = 3.17; usp OR = 3.57; traT OR = 3.31; papC OR = 3.45; p < 0.001). Several mutations were identified, with PROVEAN predicting deleterious mutations including V20G (FimH), S130L (HlyA), G3R (USP), G1V and Q104L (PapC), and E27K (TraT). Structural modeling revealed localized conformational shifts, while InterProScan indicated conserved functional domains. Phylogenetic analysis showed that mutated genes formed distinct clusters, pointing to evolutionary divergence. This study emphasizes the value of molecular monitoring and the development of targeted therapies to control antibiotic-resistant E. coli infections in diabetic patients.

This study reports a green synthesis of zinc oxide nanoparticles (ZnONPs) using Musa paradisiaca peel extract and evaluates their ability to enhance the activity of commercially available antibiotics against multidrug-resistant (MDR) Escherichia coli and Klebsiella pneumoniae. ZnONPs were prepared by mixing 30 mL of peel extract with 30 mL of 1 mM ZnSO₄·7H₂O, with the pH adjusted to 10, followed by centrifugation at 3000 rpm for 5 min, washing, and drying in the dark at 25 °C to obtain ZnONP powder. The synthesized ZnONPs were characterized using UV-Vis spectroscopy, scanning electron microscopy-energy dispersive analysis of X-rays (SEM-EDAX), Fourier transform infrared spectroscopy (FTIR), and X-ray diffraction (XRD). A characteristic UV-Vis absorption peak at 450 nm confirmed ZnONP formation, while XRD indicated a crystalline structure with an average crystallite size of 32 nm. SEM revealed irregular, monodisperse particles, and EDAX confirmed Zn as the dominant element along with signals attributable to organic and mineral components from the peel extract. FTIR suggested that phenolic, hydroxyl, and carboxyl functional groups in the extract likely contributed to nanoparticle reduction and capping. In antibacterial assays, ZnONPs alone produced inhibition zones of 17 ± 1.54 mm against E. coli and 26 ± 1.73 mm against K. pneumoniae. ZnONPs significantly enhanced antibiotic activity compared with non-coated discs (p = 0.01), increasing inhibition by 42.5-55.7% against E. coli and 42.9-52.7% against K. pneumoniae. These findings support the potential of banana peel-derived ZnONPs as low-cost synergistic enhancers of existing antibiotics against MDR pathogens.