Gut dysbiosis is associated with acceleration of lupus nephritis

The gut microbiota (GM) exerts a strong influence over the host immune system and dysbiosis of this microbial community can affect the clinical phenotype in chronic inflammatory conditions. To explore the role of the GM in lupus nephritis, we colonized NZM2410 mice with Segmented Filamentous Bacteria (SFB). Gut colonization with SFB was associated with worsening glomerulonephritis, glomerular and tubular immune complex deposition and interstitial inflammation compared to NZM2410 mice free of SFB. With SFB colonization mice experienced an increase in small intestinal lamina propria Th17 cells and group 3 innate lymphoid cells (ILC3s). However, although serum IL-17A expression was elevated in these mice, Th17 cells and ILC3s were not detected in the inflammatory infiltrate in the kidney. In contrast, serum and kidney tissue expression of the macrophage chemoattractants MCP-1 and CXCL1 were significantly elevated in SFB colonized mice. Furthermore, kidney infiltrating F4/80+CD206+M2-like macrophages were significantly increased in these mice. Evidence of increased gut permeability or “leakiness” was also detected in SFB colonized mice. Finally, the intestinal microbiome of SFB colonized mice at 15 and 30 weeks of age exhibited dysbiosis when compared to uncolonized mice at the same time points. Both microbial relative abundance as well as biodiversity of colonized mice was found to be altered. Collectively, SFB gut colonization in the NZM2410 mouse exacerbates kidney disease, promotes kidney M2-like macrophage infiltration and overall intestinal microbiota dysbiosis.

uvrY deletion and acetate reduce gut colonization of Crohn’s disease-associated adherent-invasive Escherichia coli by decreasing expression of type 1 fimbriae

Adherent-invasive Escherichia coli (AIEC) is involved in onset and/or exacerbation of Crohn’s disease. AIEC adapts to the gut environment by altering gene-expression programs, leading to successful gut-lumen colonization. However, the underlying mechanism of gut colonization is still far from clarified. Here, we show the role of UvrY, a response regulator of bacterial two-component signal transduction systems, in AIEC gut colonization. An AIEC mutant lacking the uvrY gene exhibited impairment of competitive colonization in the murine intestinal tract. UvrY contributes to functional expression of type 1 fimbriae by activating expression of small RNA CsrB, which confers adherence and invasion into epithelial cells on AIEC. In contrast, acetate suppresses the UvrY-dependent expression of type 1 fimbriae, resulting in less efficient cell invasion and attenuated gut colonization. Our findings might lead to therapeutic interventions for CD, in which inhibitions of UvrY activation and acetate supplementation reduce the colonization levels of AIEC by decreasing type-1 fimbriae expression.

Metaproteomic Profiling of Fungal Gut Colonization in Gnotobiotic Mice

Background: Eukaryotic microbes can modulate mammalian host health and disease states, yet the molecular contribution of gut fungi remains nascent. We previously showed that mice exclusively colonised with fungi displayed increased sensitivity to allergic airway inflammation and had fecal metabolite profiles similar to germ-free mice. This marginal effect on the host metabolome suggested that fungi do not primarily use metabolites to modulate the host immune system.Methods: To describe functional changes attributed to fungal colonisation, we performed mass spectrometry-based analyses of feces (Label-Free Quantitative; LFQ) and the small intestine (labeling with Tandem Mass Tag; TMT) of gnotobiotic mice colonised with defined consortia of twelve bacterial species, five fungal species, or both. We also evaluated the effect of microbiome perturbances on the metaproteome by analysing feces from mouse pups treated with an antibiotic or antifungal.Results: We detected 6,675 proteins in the mice feces, of which 3,845 had determined LFQ levels. Analysis of variance showed changes in the different gnotobiotic mouse groups; specifically, 46% of 2,860 bacterial, 15% of 580 fungal, and 76% of 405 mouse quantified proteins displayed differential levels. The antimicrobial treatments resulted in lasting changes in the bacterial and fungal proteomes, suggesting that the antimicrobials impacted the entire community. Fungal colonisation resulted in changes in host proteins functional in innate immunity as well as metabolism, predicting specific roles of gut fungi on host systems during early developmental stages. Several of the detected fungal proteins (3% of 1,492) have been previously reported as part of extracellular vesicles and having immunomodulating properties. Using an isobaric labelling TMT approach for profiling low abundant proteins of the jejunal tissue, we confirmed that the five fungal species differentially impacted the host intestinal proteome compared to the bacterial consortium. The detected changes in mouse jejunal proteins (4% of 1,514) were mainly driven by metabolic proteins. Conclusions: We used quantitative proteomic profiling of gnotobiotic conditions to show how colonisation with selected fungal species impacts the host gut proteome. Our results suggest that an increased abundance of certain gut fungal species in early life may affect the developing intracellular attributes of epithelial and immune cells.

Colonization with Multidrug-Resistant Bacteria in the First Week of Life among Hospitalized Preterm Neonates in Serbia: Risk Factors and Outcomes

The aim of this prospective cohort study was to determine the prevalence of gut colonization with multidrug-resistant (MDR) bacteria, risk factors for colonization, infection risk, and outcomes among preterm neonates hospitalized at a tertiary-care center in Serbia. During the period from December 2017 to April 2018, 103 neonates were screened for rectal carriage at admission and on the seventh day of life. Characterization of MDR strains was done by conventional microbiology and molecular methods. Out of 61 (59.2%) colonized neonates, 12 (11.6%) were found colonized at admission, while 49 (47.6%) became colonized at the study site. Among a total of 72 MDR isolates, extended-spectrum beta-lactamase (ESBL)-producing enterobacteria prevailed (56/72, 77%), followed by Acinetobacter baumannii (14/72, 19%). The majority of ESBL-producing strains carried multiple genes (blaTEM/blaCTX-M-15 or blaTEM/blaSHV). Longer previous hospitalization and delivery by cesarean section were associated with MDR colonization, while mechanical ventilation was a risk factor for colonization at the study site. Infections due to MDR bacteria were more frequent among colonized than non-colonized neonates, but not significantly, and mortality was low (1%) in the studied neonates. These results indicate that hospitalized preterm neonates in Serbia are rapidly colonized with a diversity of MDR species and resistance phenotypes/genotypes.

Anticolonization of Carbapenem-Resistant Klebsiella pneumoniae by Lactobacillus plantarum LP1812 Through Accumulated Acetic Acid in Mice Intestinal

Carbapenem-resistant Klebsiella pneumoniae (CRKP) is highly prevalent and poses a significant threat to public health. In critically ill patients, gut colonization is considered to be the reservoir of recurrent CRKP infection. Therefore, eliminating CRKP carriage in the intestine is critical for preventing subsequent CRKP infection. In the present study, Lactobacillus plantarum LP1812, a probiotic that can inhibit CRKP in vitro, was used as a candidate probiotic to investigate its efficacy for CRKP anticolonization. Compared with the control, mice fed with 1×10 8 CFU L. plantarum LP1812 exhibited significant CRKP clearance from 1×10 4 CFU/mg to less than 10 CFU/mg in mice feces. Furthermore, 16S RNA gene sequencing revealed that L. plantarum LP1812 modulated mice microbiota by increasing the relative abundance of the genus Halomanas, Blautia, and Holdemania. Further KEGG pathway enrichment analysis revealed that fatty acid-utilizing bacteria, such as acetate-producing Bacteroidetes and Blautia flourished in mice fed with L. plantarum LP1812. Moreover, we found that the concentration of acetic acid was higher in L. plantarum LP1812, which inhibited the growth of K. pneumoniae strains in vitro. Meanwhile, mice intragastrically administered with acetic acid exhibited significantly increased CRKP elimination in vivo. In conclusion, L. plantarum LP1812 is a potential candidate for intestinal CRKP anticolonization by regulating the intestinal microbiota and inhibiting CRKP via increased acetic acid in the intestinal lumen.

EVALUACIÓN DEL PERFIL DE VIRULENCIA DE Campylobacter TERMOTOLERANTES AISLADOS DE GRANJAS COMERCIALES DE POLLOS PARRILLEROS

The aim of this work was to analyze the presence of genes associated with the virulence of thermotolerant Campylobacter (CT) isolated from different sources in commercial broiler farms. For this, we worked with a collection of CT isolates obtained from three commercial broiler farms during 2015. The presence of 10 genes related to CT virulence was examined with a PCR reaction. The results showed a prevalence of 100% for flaA and flhA; 91% cadF, 51% cdtABC, 48% iam, 46% racR, 32% ciaB and 11% virB11. C. jejuni showed higher prevalence of all genes evaluated than C. coli, except iam and virB11 wich were higher for C. coli. The same gene pattern was presented in different sources of isolates. The high prevalence of flaA and flhA, involved in CT mobility, cadF related to Campylobacter gut colonization of broiler and cdtABC genes involved for the expression of cytotoxin, indicates the importance of these factors in CT virulence. Differences in the prevalence of genes according to the species of CT could demonstrate different mechanisms of pathogenicity, and this is not related to the source of isolates. All the results showed the ability of CT to colonize cells but is not evidence of pathogenesis.

IHF regulates host colonization factors in the bee gut symbiont Frischella perrara

Gut bacteria colonize specific niches in the digestive tract of animals. Yet, the genetic basis of these associations often remains elusive. The gut symbiont Frischella perrara colonizes the anterior hindgut of honey bees, where it causes a characteristic immune response leading to the formation of the scab phenotype. Genetic determinants required for the establishment of this specific association are currently unknown. Here, we independently isolated three point mutations in the two genes encoding the DNA-binding protein integration host factor (IHF). The mutations resulted in the formation of larger colonies on agar plates and the absence of an aryl polyene metabolite conferring the yellow color to colonies of F. perrara. Inoculation of microbiota-free bees with one of these mutants drastically decreased gut colonization of F. perrara and abolished scab development. Using RNAseq we show that IHF affects the expression of potential colonization factors, including a colibactin biosynthetic gene cluster, two Type 6 secretion systems, pili genes, and the aryl polyene biosynthesis pathway. Individual gene deletions of these components revealed different colonization defects indicating that these genetic determinants of F. perrara have distinct roles in the interaction with the host. IHF is conserved across many bacteria and may regulate host colonization also in other animal symbionts.

MgrB dependent colistin resistance in Klebsiella pneumoniae is associated with an increase in host-to-host transmission

Due to its high transmissibility, Klebsiella pneumoniae (Kpn) is one of the leading causes of nosocomial infections. Here, we studied the biological cost of colistin resistance, an antibiotic of last resort, of this opportunistic pathogen using a murine model of gut colonization and transmission. Colistin resistance in Kpn is commonly the result of inactivation of the small regulatory protein MgrB. Without a functional MgrB, the two-component system PhoPQ is constitutively active, leading to increased lipid A modifications and subsequent colistin resistance. Using an engineered MgrB mutant, we observed that MgrB-dependent colistin resistance is not associated with a fitness defect during in vitro growth conditions. However, colistin-resistant Kpn colonizes the murine gut poorly, which may be due to the decreased production of capsular polysaccharide by the mutant. The colistin-resistant mutant of Kpn had increased survival outside the host when compared to the parental colistin-sensitive strain. We attribute this enhanced survivability to dysregulation of the PhoPQ two-component system and accumulation of the master stress regulator RpoS. The enhanced survival of the colistin resistant strain may be a key factor in the observed rapid host-to-host transmission in our model. Together, our data demonstrate that colistin-resistant Kpn experiences a biological cost in gastrointestinal colonization. However, this cost is mitigated by enhanced survival outside the host, increasing the risk of transmission. Additionally, it underscores the importance of considering the entire life cycle of a pathogen to truly determine the biological cost associated with antibiotic resistance.ImportanceThe biological cost associated with colistin resistance in Klebsiella pneumoniae (Kpn) was examined using a murine model of Kpn gut colonization and fecal-oral transmission. A common mutation resulting in colistin resistance in Kpn is a loss-of-function mutation of the small regulatory protein MgrB that regulates the two-component system PhoPQ. Even though colistin resistance in Kpn comes with a fitness defect in gut colonization, it increases bacterial survival outside the host enabling it to more effectively transmit to a new host. The enhanced survival is dependent upon the accumulation of RpoS and dysregulation of the PhoPQ. Hence, our study expands our understanding of the underlying molecular mechanism contributing to the transmission of colistin-resistant Kpn.

Utility of bile esculin azide agar for screening of stool samples for vancomycin resistant enterococci from patients with gut colonization

Background: Due to increased prevalence of vancomycin resistant enterococci (VRE) in hospital settings as an important nosocomial pathogen, microbiology laboratories should be prepared with test protocol for prompt detection and reporting of these resistant organisms. This helps in appropriate treatment of patients without delay and implementation of infection control measures in order to prevent spread of such infections. With this background present study was conducted to demonstrate utility of bile esculin azide agar with vancomycin (BEAV) for screening of enterococci for vancomycin drug resistance.Methods: Over a period of one year 200 stool samples were collected from hospitalized patients in a tertiary care hospital. Samples were inoculated on bile esculin azide agar with vancomycin (6ug/ml) to screen for vancomycin drug resistance in enterococci isolated from stool samples. Vancomycin drug resistance was confirmed by agar dilution method.Results: Out of 200 stool samples collected from hospitalized patients, 13 (6.5%) samples showed growth on bile esculin azide agar with vancomycin (6 µg/ml). Of these 13 isolates, 12 (92.3%) isolates were confirmed as VRE by agar dilution method and demonstrated minimum inhibitory concentration (MIC) of ≥32 µg/ml and all 12 isolates were identified as E. faecium. One (7.7%) isolate grown on BEAV was identified as E. gallinarum and showed MIC value of 8 µg/ml.Conclusions: Present study recommends use of bile esculin azide agar with vancomycin (6 µg/ml) as a screening medium for isolation of VRE from stool samples which usually carries mixed commensal flora of gastrointestinal tract.