Journal of Applied Microbiology最新文献

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: Hyperosmolarity is widely used for food preservation by inhibiting bacterial survival and growth. Therefore, it is of great significance to reveal bacterial osmotic-response mechanism. Biofilm formation presents a significant challenge for the control and prevention of pathogenic bacteria. Our previous study showed that inactivation of the efflux protein TolC in extraintestinal pathogenic Escherichia coli (ExPEC) decreased biofilm formation by affecting curli production in a medium osmolarity-dependent manner. This study aims to explore the role of the two-component CpxA/R system in mediating TolC regulation of ExPEC biofilm formation in response to osmolarity.

Methods and results: Various mutants derived from the parental ExPEC ΔtolC strain were constructed, and their abilities to form biofilms and produce curli fimbriae in different osmotic media were evaluated using crystal violet staining, scanning electron microscopy, Congo red assay, and real-time quantitative PCR. The results showed that the disruption of CpxA/R system by deleting the gene encoding histidine kinase-CpxA or response regulator-CpxR, or by introducing a point mutation at the phosphorylation site of CpxA, significantly compromised the effect of TolC inactivation on ExPEC biofilm formation and curli biosynthesis under both NaCl- and sucrose-induced osmotic stresses.

Conclusions: Our study firstly demonstrate that the CpxAR system mediated the regulation of TolC inactivation on ExPEC biofilm formation and curli production in response to both NaCl- and sucrose-induced osmotic stresses. These findings expand the regulatory network of bacterial biofilm formation and osmotic-responsiveness, contributing to exploring potential targets for preventing and controlling pathogenic bacteria.

This review provides a comprehensive overview of the current landscape in Bacillus spp. fermentation processes for high-value bioproducts such as proteases, amylases, and surfactin in lab-scale bioreactors, highlighting critical process features. An analysis of key findings over the past five years shows that an efficient fermentation requires customized bioreactor conditions dependent on the substrate, strain type, and the target product. Major challenges include overcoming foaming, ensuring oxygen mass transfer, and precisely balancing the growth and production phases. A combined approach using statistical modeling, such as response surface methodology, along with experimental validation in different bioreactor designs is recommended for the process optimization. This integrated strategy can significantly improve the efficiency, predictability, and overall success of Bacillus-based bioprocesses.

Aims: This study investigated colistin resistance in Gram-negative bacteria isolated from wastewater. The research focuses on understanding the genetic mechanisms of mcr-mediated resistance and the role of wastewater as a reservoir for colistin-resistant bacterial pathogens.

Methods and results: The study utilized 16 sewage effluent samples collected from four discharge points (three hospitals and one municipal wastewater treatment plant), during May 2024. Bacterial isolates were obtained using the membrane filtration method, resulting in the recovery of 50 Gram-negative isolates, including Enterobacteriaceae and Aeromonas species. Identification was conducted using MALDI-TOF mass spectrometry. Whole-genome sequencing (WGS) and comprehensive bioinformatics analysis were performed to characterize resistance genes and phylogenetic relationships. Colistin resistance was found in Escherichia coli (mcr-1), Aeromonas veronii (mcr-3), and Enterobacter kobei (mcr-9). Escherichia coli was the predominant species, accounting for 50% of the isolates. WGS revealed predominant resistance profiles across isolates, with E. coli harboring 95 resistance genes, E. kobei 21, and A. veronii. 14. Genomic analysis identified mobile genetic elements (MGEs) like ISCR1 and tnpA, suggesting the potential for horizontal gene transfer. Comparative genomic analysis identified the genetic context of mcr genes, with the mcr-1 gene found on a plasmid in E. coli, mcr-3 on a conjugative plasmid in A. veronii, and mcr-9 on a plasmid in E. kobei.

Conclusions: These findings highlight the role of wastewater in the spread of colistin-resistant bacteria. The presence of mcr genes on mobile elements underscores the need for robust surveillance strategies to monitor the dissemination of these resistance traits in the environment.

Aims: While drinking water disinfection can significantly reduce bacterial populations it does not eradicate all the bacteria present as many cells enter varying levels of dormancy including the viable but non-culturable (VBNC) state. We posit that the VBNC state represents a continuum, with lower and greater levels of stress leading to Shallow and Deep VBNC, respectively, in exposed cells.

Methods and results: We examined the proteomic differences of Escherichia coli that entered varying depths of the VBNC state upon monochloramine disinfection at different doses. Exposure for 1 h to low (17 μM) and high (50 μM) concentrations of monochloramine led to general VBNC outcomes that showed loss of culturability but retention of viability. However, cells exposed to 17 μM monochloramine exhibited high levels of ATP production, while 50 μM exposure resulted in low levels of ATP productions, confirming that the low and high monochloramine doses resulted in a proposed Shallow and Deep VBNC states, respectively. Differentially expressed proteins analyses showed that proteins related to cell motility (LuxS) and arginine biosynthesis (ArgD) were upregulated, while several oxidative stress response proteins (FumC and Mdh) were downregulated in the Shallow VBNC group. Within the Deep VBNC group, multiple DNA translation-related proteins were downregulated as oligopeptides transport proteins (OppA) and periplasmic proteins (YdeI) were upregulated.

Conclusions: Our results confirmed a gradient in VBNC responses of E. coli in response to different monochloramine doses, accompanied by changes in the proteome.

Aims: We have investigated bacterial colonization and site-specific distribution across hard and soft oral tissues focusing on both intra- and inter-individual patterns of Streptococcus clonality and overall microbial composition.

Methods and results: Thirty samples were collected from five healthy adults, covering six oral sites (left and right buccal mucosa, tongue dorsum, maxillary central incisors, mandibular molar fissure, and saliva). Repetitive sequence-based PCR assessed Streptococcus clonality, and 16S rRNA gene sequencing characterized broader microbial communities. Among 98 Streptococcus isolates, 67.3% were apparently clonal and frequently recovered from multiple sites within individuals. Identical S. vestibularis clones were detected across all sampled sites in one participant and across several sites in others, indicating shared intra-oral reservoirs. Amplicon sequencing revealed distinct microbial compositions across oral sites, with saliva displaying the highest alpha diversity (Shannon index) and significant beta-diversity differences between most sites, except saliva and tongue. Firmicutes predominated overall, and Streptococcus was the most abundant genus. Differential-abundance testing (ANCOM-BC2) identified 26 genera varying significantly across sites, with lower abundance generally observed on soft tissues.

Conclusions: Whilst microbial communities varied by oral site, certain streptococcal clones, including S. vestibularis, occurred across multiple habitats, suggesting potential intra-oral reservoirs that may contribute to microbiome stability.

Aims: This study evaluated the influence of two carrier materials, a nylon pad (NP) and a nylon-silicon carbide pad (NSCP), on fixed-bed reactor (FBR) and microbiomes performances during the anaerobic digestion of tomato liquid fraction.

Methods and results: FBR performance was evaluated during start-up, biofilm formation, and biodegradability tests, applying principal components analysis (PCA) to correlate biological, physicochemical, and operating variables. PCA results showed two positive interactions, the first one between facultatively-syntrophic fermenters and CO2-reducing methanogens in both carriers, increasing chemical oxygen demand (COD) removal and methane yield through interspecies formate/H2 transfer. The second interaction was associated with Geobacter daltonii and Methanosarcina spelaei in NSCP, suggesting a direct interspecies electron transfer (DIET) to acetotrophic methanogens.

Conclusions: Biodegradability tests revealed positive microbial interactions to enhance COD removal and methane yield via sugars→butyrate→acetate→methane pathway in both pads. The use of NP and NSCP carrier materials enhanced syntrophisms, impacting FBR performance. In NSCP syntrophic relationships were associated with the acetotrophic pathway that was increased by a DIET; while in NP a competitive exclusion mechanism where the fermenter Trichococcus alkaliphilus successfully competed against obligately-syntrophic acetogens for sugars and fatty acids was observed.

Aims: This study aimed to elucidate how copper ions (Cu2+) control laccase production in Dichomitus squalens, to identify the Cu2+-responsive laccase gene(s), and to clarify their physiological roles.

Methods and results: The research employed various experimental approaches including enzyme activity assays, transcriptome sequencing, proteomics, RT-qPCR, RNA interference, and ROS detection. Laccase activity and biomass of D. squalens were quantified after exposure to 0-3.0 mM Cu2+, showing that 2.0 mM Cu2+ elevated total laccase activity without inhibiting mycelial growth. Transcriptome sequencing and RT-qPCR revealed that the expression of laccase gene lcc3 was specifically up-regulated by 299.8-fold. Quantitative proteomics revealed that the abundance of LCC3 protein, the predominant extracellular laccase isoform, rose 44.8-fold upon copper supplementation, underscoring its pivotal contribution to overall laccase activity. In addition, RNA interference of lcc3 gene markedly reduced Cu2+-stimulated laccase activity even under copper induction, confirming its indispensable role. Finally, the total reactive oxygen species (ROS) in Cu2+-treated hyphae were lower than in untreated controls, linking lcc3 gene induction to cellular redox homeostasis.

Conclusions: In this study, by integrating transcriptomics, proteomics and functional genetics, we elucidated that Cu2+ specifically induces the expression of specific laccase genes to enhance laccase activity in D. squalens. lcc3 is the sole laccase gene that is massively and specifically induced by physiological Cu2+ concentrations. We further clarified the function of its encoded protein: LCC3 protein not only enhances extracellular laccase activity but also confers Cu2+ tolerance via ROS scavenging and Cu2+ chelation. These findings uncover a unique copper-responsive regulon and establish the lcc3 gene as a promising target for metabolic engineering aimed at high-yield laccase production and robust fungal performance under metal stress.