Microbiology spectrum最新文献

Bacterial membrane vesicles (MVs) are produced by all bacteria and contribute to numerous bacterial functions due to their ability to package and transfer bacterial cargo. In doing so, MVs have been shown to facilitate horizontal gene transfer, mediate antimicrobial activity, and promote biofilm formation. Uropathogenic Escherichia coli is a pathogenic Gram-negative organism that persists in the urinary tract of its host due to its ability to form persistent, antibiotic-resistant biofilms. The formation of these biofilms is dependent upon proteins such as Antigen 43 (Ag43), which belongs to the widespread Autotransporter group of bacterial surface proteins. In E. coli, the autotransporter Ag43 has been shown to contribute to bacterial cell aggregation and biofilm formation via self-association of Ag43 between neighboring Ag43-expressing bacteria. As MVs package bacterial proteins, we investigated whether MVs produced by E. coli contained Ag43, and the ability of Ag43-expressing MVs to facilitate cell aggregation and biofilm formation. We showed that Ag43 expressing E. coli produced MVs that contained Ag43 on their surface and had an enhanced ability to bind to E. coli bacteria. Furthermore, we demonstrated that the addition of Ag43-containing MVs to Ag43-expressing E. coli significantly enhanced biofilm formation. These findings reveal the contribution of MVs harboring autotransporters in promoting bacterial aggregation and enhancing biofilm formation, highlighting the impact of MVs and their specific composition to bacterial adaptation and pathogenesis.IMPORTANCEAutotransporter proteins are the largest family of outer membrane and secreted proteins in Gram-negative bacteria which contribute to pathogenesis by promoting aggregation, biofilm formation, persistence, and cytotoxicity. Although the roles of bacterial autotransporters are well known, the ability of bacterial membrane vesicles (MVs) naturally released from the surface of bacteria to contain autotransporters and their role in promoting virulence remains less investigated. Our findings reveal that MVs produced by E. coli contain the autotransporter protein Ag43. Furthermore, we show that Ag43-containing MVs function to enhance bacterial cell interactions and biofilm formation. By demonstrating the ability of MVs to carry functional autotransporter adhesins, this work highlights the importance of MVs in disseminating autotransporters beyond the bacterial cell membrane to ultimately promote cellular interactions and enhance biofilm development. Overall, these findings have significant implications in furthering our understanding of the numerous ways in which MVs can facilitate bacterial persistence and pathogenesis.

Candida albicans, a major human fungal pathogen, can form biofilms on a variety of inert and biological surfaces. C. albicans biofilms allow for immune evasion, are highly resistant to antifungal therapies, and represent a significant complication for a wide variety of immunocompromised patients in clinical settings. While transcriptional regulators and global transcriptional profiles of C. albicans biofilm formation have been well-characterized, much less is known about translational regulation of this important C. albicans virulence property. Here, using ribosome profiling, we define the first global translational profile of genes that are expressed during early biofilm development in a human fungal pathogen, C. albicans. We show that C. albicans biofilm formation involves altered translational regulation of genes and gene classes associated with protein synthesis, pathogenesis, transport, plasma membrane, polarized growth, cell cycle, secretion, and signal transduction. Interestingly, while similar, but not identical, classes of genes showed transcriptional alterations during early C. albicans biofilm development, we observed very little overlap between specific genes that are upregulated or downregulated at the translational vs transcriptional levels. Our results suggest that distinct translational mechanisms play an important role in regulating early biofilm development of a major human fungal pathogen. These mechanisms, in turn, could serve as potential targets for novel antifungal strategies.IMPORTANCEThe major human fungal pathogen Candida albicans is known to form biofilms or complex aggregated microbial communities encased in an extracellular matrix. These biofilms allow C. albicans to escape detection by the immune system as well as resist a variety of antifungal drugs. In this study, we define the first global profile of genes that show altered translation during C. albicans biofilm formation. These genes are involved in a variety of key cellular processes, including polarized growth, pathogenesis, transport, protein synthesis, cell cycle, plasma membrane, signal transduction, and secretion. Interestingly, while similar classes of genes are induced at both the transcriptional and translational levels during early C. albicans biofilm formation, we observed very little overlap among specific genes with altered transcription and translation. Our results suggest that C. albicans biofilm formation is controlled by distinct translational mechanisms, which could potentially be targeted by novel antifungal drugs.

Bacteria can become resistant to antibiotics in two ways: by acquiring resistance genes through horizontal gene transfer and by de novo development of resistance upon exposure to non-lethal concentrations. The importance of the second process, de novo build-up, has not been investigated systematically over a range of species and may be underestimated as a result. To investigate the DNA mutation patterns accompanying the de novo antibiotic resistance acquisition process, six bacterial species encountered in the food chain were exposed to step-wise increasing sublethal concentrations of six antibiotics to develop high levels of resistance. Phenotypic and mutational landscapes were constructed based on whole-genome sequencing at two time points of the evolutionary trajectory. In this study, we found that (1) all of the six strains can develop high levels of resistance against most antibiotics; (2) increased resistance is accompanied by different mutations for each bacterium-antibiotic combination; (3) the number of mutations varies widely, with Y. enterocolitica having by far the most; (4) in the case of fluoroquinolone resistance, a mutational pattern of gyrA combined with parC is conserved in five of six species; and (5) mutations in genes coding for efflux pumps are widely encountered in gram-negative species. The overall conclusion is that very similar phenotypic outcomes are instigated by very different genetic changes. The outcome of this study may assist policymakers when formulating practical strategies to prevent development of antimicrobial resistance in human and veterinary health care.IMPORTANCEMost studies on de novo development of antimicrobial resistance have been performed on Escherichia coli. To examine whether the conclusions of this research can be applied to more bacterial species, six species of veterinary importance were made resistant to six antibiotics, each of a different class. The rapid build-up of resistance observed in all six species upon exposure to non-lethal concentrations of antimicrobials indicates a similar ability to adjust to the presence of antibiotics. The large differences in the number of DNA mutations accompanying de novo resistance suggest that the mechanisms and pathways involved may differ. Hence, very similar phenotypes can be the result of various genotypes. The implications of the outcome are to be considered by policymakers in the area of veterinary and human healthcare.

Simpler, safer, and faster chemotherapeutic regimens for tuberculosis and other respiratory mycobacterial infections are an urgent need. Many current therapies suffer from suboptimal drug exposure and dose-limiting systemic adverse effects, challenges that could be addressed via controlled delivery of drugs to the primary site of infection. We sought to address this need by designing a flexible formulation platform that targets drugs to the lung macrophages that concentrate at infectious foci. Our approach is based on an encapsulation strategy in which drugs or specifically designed prodrugs are captured in the hydrophobic core of a glucan-lipid particle (GLP). We show chemically diverse antimycobacterial drugs can be efficiently and stably encapsulated within GLP and that these microparticles can be engineered to release drugs upon low pH or reducing conditions that occur upon phagocytosis by macrophages. Encapsulated formulations of clofazimine, isoniazid, and linezolid retain activity against intracellular Mycobacterium tuberculosis (Mtb) in an ex vivo model, demonstrating efficient drug delivery and release. Intranasal administration of GLP-clofazimine to Mtb-infected mice effectively concentrates the drug in the lung and reduces bacterial burden, whereas GLP-delivered linezolid was systemically distributed and failed to inhibit bacterial growth in the lung. This work establishes GLPs as a promising platform for targeted antibiotic delivery to the lung and also illustrates pharmacokinetic parameters that must be considered in future development.

Importance: Tuberculosis (TB) causes an estimated 10.8 million cases each year and remains one of the leading causes of infectious death. Effective treatment is complicated due to the lengthy drug regimen required to prevent relapse and treatment failure. A primary challenge is delivering drugs effectively to lung granulomas, where TB bacteria can persist. Here, we developed yeast-derived glucan lipid microparticles (GLPs) as a novel delivery system to efficiently encapsulate and deliver TB drugs directly to lung tissue via intranasal administration. Of the formulations evaluated, GLP-encapsulated clofazimine achieved increased lung drug levels and reduced bacterial burden in TB-infected mice. The use of GLPs offers a promising approach to improve TB treatment by enabling targeted drug delivery to infection sites within the lungs.

Elizabethkingia species, isolated from clinical and environmental samples, are emerging opportunistic bacterial pathogens with a high mortality rate in clinical settings worldwide. Taxonomically, Elizabethkingia comprises seven species: E. anophelis, E. argenteiflava, E. bruuniana, E. meningoseptica, E. miricola, E. ursingii, and E. occulta. In this study, we identified useful biomarker proteins, including ribosomal L29, L30, S21, and the YtxH domain-containing proteins, for distinguishing Elizabethkingia species using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) profiles. Evaluation of 29 clinical and environmental Elizabethkingia strains revealed that these species could be separated by MALDI-TOF MS profiles into six groups-E. anophelis, E. argenteiflava, E. bruuniana/E. miricola, E. meningoseptica, E. ursingii, and E. occulta-based on the four biomarker protein peaks. This study demonstrates the potential of routine MALDI-TOF MS -based examination methods for the early detection of Elizabethkingia species in clinical laboratories.

Importance: Elizabethkingia species are groups of emerging opportunistic bacterial pathogens with a high mortality rate, causing healthcare-associated outbreaks worldwide. Rapid identification of Elizabethkingia species is important becausethese species show intrinsically carbapenem resistance and there are few data for using appropriate antibiotics. Until now, only whole-genome sequencing could accurately identify the seven Elizabethkingia species. Therefore, establishing rapid and accurate identification methods for Elizabethkingia species in clinical laboratories is vital. In this study, we developed new methods for identifying Elizabethkingia species using four biomarker protein peaks-ribosomal L29, L30, S21, and the YtxH domain-containing proteins by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) proteotyping. This study demonstrates the potential of routine MALDI-TOF MS -based laboratory examination for the early identification of Elizabethkingia species.

Salmonella Enteritidis (S. Enteritidis) stands as a leading cause of human salmonellosis worldwide with a tendency to spread through contaminated foodstuffs and animals. In Hong Kong, a significant proportion of food products are imported, and many cases are often caused by the consumption of contaminated food, hence making the geographical surveillance of drug-resistant S. Enteritidis important for strong public health and food safety measures. We analyzed the whole genomes of 207 S. Enteritidis from Hong Kong, Australia, Canada, mainland China, the United States of America, South Africa, Taiwan, and the United Kingdom to examine associated antimicrobial resistance and the transmission dynamics between continents. Phylogenetic cluster inferences and Bayesian phylogeographical analysis were performed. Overall, sequence type ST11 strains were dominant (92.8%, 192/207). Five phylogenomic clusters A to E were identified, where most isolates from mainland China and Hong Kong were in Cluster E. Among the 22 plasmid types identified, IncX1 was dominant in the Asian isolates. Most of the virulence genes were distributed in Salmonella pathogenicity islands -1 and -2, with two universal virulence operons responsible for the effector delivery system and bacterial cell adhesion. The phylogeographic inference analysis showed a statistically significant link between mainland China and Hong Kong with the highest relative migration rate (relativeGeoRates mean ± standard error = 2.93 ± .07, Bayes Factor [BF] = 1285.5], with some traceable to Canada (0.61 ± 0.03, BF = 6.9) and Australia (1.02 ± 0.04, BF = 4.2). Our analysis suggests hypothetical transmission of S. Enteritidis and its associated antimicrobial resistance across borders.

Importance: Antimicrobial resistance and disease severity in nontyphoidal Salmonella have constituted a serious public health challenge worldwide. Drug-resistant Salmonella Enteritidis is a leading pathogen that causes human infections primarily through the consumption of contaminated food products. Previous research focuses on the whole-genome analysis of antimicrobial resistance and virulence factors in S. Enteritidis; however, details on how this bacterium localized, expanded, and diversified from location to location remain unknown. Our study for the first time addresses this gap by investigating the phylogeographic transmission to estimate the frequency and location of cross-border spread. By evidence-based inferred transmission, we aim to uncover novel insights into the dynamic spread of S. Enteritidis, revealing the route of emergence and migration. This research is crucial for enhancing our understanding of the control strategies to safeguard human health.

Asthma is a chronic respiratory disease with increasing global prevalence, often linked to disrupted airway microbiota. Azithromycin has shown promise in asthma treatment, but whether its effect is owing to its antimicrobial capacity remains largely unknown. A house dust mite (HDM)-induced asthmatic mouse model was used to evaluate the effects of azithromycin on airway inflammation and microbiota. Mice were divided into control, HDM-induced asthma, HDM + azithromycin, and azithromycin-alone groups. Airway microbiota was analyzed using 16S rRNA sequencing, and metabolomic profiles were assessed via liquid chromatography-tandem mass spectrometry. Azithromycin alleviated type 2 airway inflammation in HDM-induced asthma, restoring microbiota diversity by modulating specific genera, including Streptococcus, Staphylococcus, Ruminococcus, Coprococcus, Bifidobacterium, etc. Combination analysis with metabolomics revealed that azithromycin significantly regulated airway microbiota-associated sphingomyelin metabolism. Azithromycin's therapeutic effects in asthma are associated with its ability to regulate airway microbiota and its associated sphingomyelin metabolism, highlighting the potential for microbiota-targeted therapies in asthma.IMPORTANCEAsthma, a prevalent chronic respiratory condition, poses a significant global health challenge due to its increasing prevalence and associated morbidity. The role of airway microbiota in asthma pathogenesis is gaining attention, with evidence suggesting that disruptions in this microbial community contribute to disease severity. Our study investigates the impact of azithromycin, a macrolide antibiotic, on airway inflammation and microbiota in a mouse model of asthma. The findings reveal that azithromycin not only alleviates airway inflammation but also restores microbiota diversity and modulates microbiota-associated sphingomyelin metabolism. This research underscores the potential of microbiota-targeted therapies in asthma management, offering a novel therapeutic strategy that could improve patient outcomes and reduce the healthcare burden associated with asthma.

The characterization of Treponema pallidum proteins is of great significance for the study of the prevention, diagnosis, and pathogenesis of syphilis. The structures and functions of many T. pallidum proteins, including the Tp40 (Tp0134) protein, remain unknown. To explore the expression pattern of the Tp40 protein within T. pallidum, we established an animal model of syphilis infection to compare the variations in serum Tp40 antibody levels between Live and Inactivated Tp groups. The results indicated that the absorbance of Tp40-enzyme-linked immunosorbent assay (ELISA) did not increase in the Inactivated group and the Untreated group, but it increased in the Live Tp infection group, suggesting that the Tp40 protein is an in vivo-induced antigen that is only actively expressed during infection. In addition, the localization of the Tp40 protein was determined by the gel microdrop method. We found that Tp40 may be a transmembrane protein with a signaling peptide present in the intima periplasm of T. pallidum. Finally, 468 patients' sera were collected for diagnostic value evaluation. Tp40-ELISA, LZ-ELISA, and Shanghai Kehua rapid plasma reagin (RPR) reagent kit showed a high degree of consistency in 468 serum samples. This suggests that Tp40 could be a valuable diagnostic antigen. The results of this study provide a new reference for the study of the pathogenesis, protein function, and diagnosis of syphilis.

Importance: In recent years, syphilis, as a chronic infectious disease, has once again attracted much attention. Treponema pallidum exhibits remarkable infectivity, concealment, and aggressiveness, posing considerable challenges to its prevention and control. The underlying pathogenic mechanisms remain elusive, and during the infection process, the roles of numerous proteins are still unclear. Through protein characterization in this study, it was found that the Tp40 protein is highly likely to be a transmembrane protein with a signal peptide and may be located in the periplasm. Besides, based on experiments with animal models and the detection of human serum samples, we believe that the Tp40 protein is a potential in vivo-induced antigen of T. pallidum that can be used for serological diagnosis of syphilis. This study conducted a preliminary exploration of the Tp40 protein and provided a meaningful reference for further exploration of the functional mechanism of the Tp40 protein and its significance in clinical diagnosis.

Enterotoxigenic Escherichia coli (ETEC) is a leading cause of childhood and travelers' diarrhea. The vaccine candidate ETVAX encompasses several ETEC colonization factors (CFs) with a hybrid LT (heat-labile toxin)/cholera toxin B subunit adjuvanted with a double-mutant LT. Stool samples from a Phase 2b ETVAX trial were tested by a PCR-based customized TaqMan Array Card (TAC), including three ETEC toxin genes (LT and heat-stable toxins, STh and STp) and 18 ETEC CFs. Stool samples were also tested with the molecular platform Amplidiag and culture, followed by GM1-enzyme-linked immunosorbent assay (ELISA) and inhibition GM1-ELISA for LT and ST and dot blot for CFs of ETECs identified among six culture isolates (maximum). Compared with Amplidiag, TAC yielded 89.4% sensitivity (320/358) and 96.4% specificity (405/420) for ETEC detection. The two methods demonstrated a good quantitative correlation (quantification cycle R² = 0.827, P < 0.05). Compared with culture, TAC and Amplidiag each exhibited 96.8% (184/190) sensitivity and identified an additional of 151 and 174 PCR positives in 588 culture-negative stools, respectively. The concordance of stool TAC versus ELISA of ETEC colonies for LT and STh/STp was 85.5% (165/193). TAC demonstrated 98% sensitivity and 92% specificity versus the dot blot results of 793 colonies for the ETVAX CFs CFA/I, CS3, CS5, and CS6. Overall ETEC was detected by TAC in 335 (43.1%) and by Amplidiag in 358 (46.0%) of specimens compared to 190 (24.4%) by culture. We conclude that molecular diagnostic approaches of TAC or Amplidiag increase the detection of ETEC compared with culture, and TAC can also provide vaccine-subtype ETEC data.CLINICAL TRIALSThis study was registered with ClinicalTrials.gov as NCT03729219.IMPORTANCEEnterotoxigenic Escherichia coli (ETEC) is an important cause of childhood and travelers' diarrhea. Vaccines in development utilize specific toxins and colonization factors (CFs) as antigens. Therefore, clinical microbiologic diagnostic methods are needed to discriminate specific toxins and CFs, both for vaccine trials and to guide epidemiology. In this work, we assessed the diagnostic performance of several methods for ETEC: a PCR-based customized TaqMan Array Card (TAC) and the molecular platform Amplidiag on stool and E. coli culture, followed by GM1-enzyme-linked immunosorbent assay for toxins and dot blot for CFs. Stool samples from a Phase 2b ETEC vaccine trial were used. Overall, ETEC was detected by TAC in 335 (43.1%) and by Amplidiag in 358 samples (46.0%) compared to 190 (24.4%) by culture. TAC additionally provided CF data with 98% sensitivity and 92% specificity. We conclude that the molecular diagnostic approaches of TAC or Amplidiag increase the detection of ETEC compared with culture.