Brazilian Journal of Microbiology最新文献

Klebsiella pneumoniae is an important nosocomial pathogen, considered a critical threat to public health, but its genomic data are scarce in Brazil. Therefore, the aim of this study was to identify the mechanisms of virulence and antimicrobial resistance and to evaluate the molecular epidemiology of K. pneumoniae isolates from an outbreak in a university hospital in Brazil. The DNA of the 25 isolates was sequenced, the functional annotation of the genome was performed in Prokka, for virulence and resistance analyses PanViTa was used, and for Multilocus Sequence Typing analyses pyMLST was used. The isolates presented several virulence genes such as those of the fim operon, ecp operon, and T6SS (identified in all isolates), which are related to adhesion, biofilm formation, and toxin delivery. Other predicted virulence genes involve nutritional/metabolic factors, effector delivery systems, and immune modulation, while resistance genes mainly involve efflux (AcrAB-TolC, KpnGH-TolC, OqxAB, and KpnEF), antibiotic target alteration, reduced permeability, and antibiotic inactivation. In addition, the resistance genes identified in K. pneumoniae isolates are related to a total of 28 antimicrobials from different classes. The sequence types (ST) 11, 273, 395, 5209 and 636 were identified, this being the first report of the ST636 in Brazil. Genomic analysis confirmed the presence of complex virulence and antimicrobial resistance mechanisms, highlighting the threat these pathogens pose and raising concerns about the introduction of ST636 into Brazil, emphasizing the complexity of local strains and the need for continuous surveillance.

The widespread use of aromatic herbicides such as 2,4-dichlorophenoxyacetic acid (2,4-D) has led to persistent environmental contamination, requiring efficient and sustainable biodegradation strategies. In this study, we isolated and characterized a novel actinobacterial strain, Streptomyces sp. SCPE-10, from contaminated coastal soil, capable of using 2,4-D as its sole carbon source. Phenotypic assays revealed robust growth on aromatic substrates, while whole-genome sequencing (Submission no. SUB15461668) followed by multilocus sequence analysis (16 S rRNA, recA, rpoB, atpD, gyrB, and trpB) revealed that SCPE-10 is closely related to Streptomyces phaeoluteichromatogenes. Functional genome annotation revealed a high abundance of genes involved in aromatic compound degradation, including cytochrome P450 monooxygenases, ring-cleaving dioxygenases, and dehalogenases. KEGG and antiSMASH analyses identified multiple metabolic pathways and 28 biosynthetic gene clusters (BGCs), including clusters for polyketides, nonribosomal peptides, terpenes, siderophores, and ectoine. Notably, SCPE-10 harbors key genes related to the degradation of benzoate, naphthalene, toluene, xylene, and 2,4-D, indicating broad-spectrum catabolic potential. These findings suggest that Streptomyces sp. SCPE-10 is a promising candidate for the bioremediation of herbicide-contaminated environments and the exploration of novel secondary metabolites.

Background: Bovine mastitis is the most economically significantdisease in dairy farming, with Staphylococcus aureus and Staphylococcus chromogenes asprominent agents, the former posing a major threat. As principal immune sentinels in themammary gland, macrophages orchestrate early pathogen recognition and immune activation,critically influencing the trajectory and outcome of infection. Thus, this study aimed tocharacterize the early macrophage responses to distinct bovine-associated S. aureus and S.chromogenes strains.

Methods: Here, RAW 264.7 cells were challenged with four differentstrains: S. aureus [isolated from nose (SN), and intramammary infection (IMI)] and S.chromogenes [IMI, and teat apex (TA)] were evaluated after 90- and 180-min. Nitric oxide (NO)production was analyzed in the supernatants, and mRNA levels of IL-1β, IL-18, NLRP3, NOS2,Arg1, Bax, and Bcl2 were assessed in the cells.

Results: Macrophages challenged with S. aureusIMI strains showed elevated Nos2 expression but negligible NO production, indicating a potentialimmune evasion mechanism. The commensal S. aureus SN strain uniquely maintained arginaseexpression, suggesting M2-like polarization that may promote immune tolerance and bacterialcolonization. Both S. aureus strains significantly upregulated the anti-apoptotic Bcl2 gene, atranscriptional response that may be associated with host cell survival, which may facilitatebacterial intracellular persistence. In contrast, S. chromogenes strains induced strong NOS2expression, robust NLRP3 inflammasome activation, and increased IL-1β production, indicatingM1 polarization and a pro-inflammatory response. The pro-apoptotic Bax gene showed an earlydecrease followed by a later increase exclusively in S. aureus-infected macrophages, indicating atime-dependent transcriptional modulation of apoptosis-related genes.

Conclusions: Thesegenotype-dependent macrophage responses reveal complex immune modulation shaping mastitis pathogenesis. However, our findings are based solely on transcriptional data on the murine cells and require further validation.

Gliotoxin of Aspergillus fumigatus has been extensively studied for its role in pathogenesis in animals and humans. It triggers pathogenesis by its immunosuppressive and cytotoxic effects. Biosynthetic gene cluster (BGC) consisting of 13 genes regulates its biosynthesis. We targeted gliZ, gliP and gliA genes of this BGC using CRISPR/Cas9 system in a multigene editing approach to check the pathogenesis in broilers. crRNAs were designed using EuPaGDT and 3 single guide RNAs (sgRNA) were commercially synthesized. Each sgRNA was combined with Cas9 to form ribonucleoprotein complexes which were then used for simultaneously transfecting fungal protoplasts. Thin-layer chromatography showed the absence of gliotoxin on silica plate and DNA sequencing showed various indels in target genes. These indels caused amino acid substitutions in all three gene products but, the gliP mutation, since it was synonymous, was likely not functionally relevant. Regenerated protoplasts were matured to form fungal hyphae and spore production was induced. These spores were inoculated intra-air sac in broiler chicks. During one-week infection trial, birds infected with the wild-type spores (group 1) showed morbidity and their mortality rate was 30%. Birds inoculated with RNP-treated spores (group 2) showed mild clinical signs and no mortality. No morbidity or mortality was recorded in birds in negative control group (group 3). Histopathological analysis of lungs showed necrosis and congestion, and presence of mixed population of inflammatory cells in wild-type infected birds, while no such lesions were seen in birds infected with RNP-treated spores. These results show that multigene editing approach was successful in creating indels simultaneously in 3 gliotoxin genes which resulted in amino acid substitution which negatively impacted gliotoxin biosynthesis and export. In vivo experiment results show that RNP-treated fungal spores were unable to cause A. fumigatus pathogenicity in broiler. Targeting gliotoxin biosynthesis could thus be a promising approach to develop antifungal therapy.

The present study aims to evaluate the efficiency of bacterial isolate to produce biosurfactant through nine analytical methods based on surface activity measurements. Bacterial isolate designated 13(1) showed capability to produce surfactant resulting in significant surface tension reduction of minimal salt culture broth medium with 42.84% corresponding to surface activity of 19.48 mN m^-1 and effectively emulsified soybean oil with E24% value reached 41.07%. This bacterium gave positive results in CTAB test indicating presence of extracellular anionic surfactants. Also, it gave highest oil dispersing zone with 2.22 cm diameter corresponding to 3.86 cm² area, and largest flatten droplet with 7.5 mm diameter on hydrophobic surface. Moreover, isolate 13(1) effectively produced lipases and had hydrocarbon degrading and hemolytic activities. Furthermore, the isolated bacterial strain was identified as Brucella anthropi OL469515 based on its morphological, cultural and biochemical characteristics, and analysis of 16S rRNA gene sequence. More interestingly, the biosurfactant production and cell growth were evaluated through five culture media including minimal salt, Hua, modified Hua, Bushnell Haas, and Kim media to investigate the more suitable medium ingredients for the efficiency and magnitude of biosurfactant production. Results have shown that maximum biosurfactant production obtained when B. anthropi grown on Hua medium leading to highest reduction of culture surface tension reached 45.81% with surface activity of 20.30 mN m^-1. Also, largest oil clearing zone was occurred with 2.45 cm diameter with area value of 4.73 cm². Finally, these comparable results concluded that Hua medium is the most suitable production medium to be used for biosurfactant optimization in future studies.