Cross-alteration of murine skin and tick microbiome concomitant with pathogen transmission after Ixodes ricinus bite.

BACKGROUND: Ticks are major vectors of diseases affecting humans such as Lyme disease or domestic animals such as anaplasmosis. Cross-alteration of the vertebrate host skin microbiome and the tick microbiome may be essential during the process of tick feeding and for the mechanism of pathogen transmission. However, it has been poorly investigated. METHODS: We used mice bitten by field-collected ticks (nymphs and adult ticks) in different experimental conditions to investigate, by 16S rRNA gene metabarcoding, the impact of blood feeding on both the mouse skin microbiome and the tick microbiome. We also investigated by PCR and 16S rRNA gene metabarcoding, the diversity of microorganisms transmitted to the host during the process of tick bite at the skin interface and the dissemination of the pathogen in host tissues (blood, heart, and spleen). RESULTS: Most of the commensal bacteria present in the skin of control mice were replaced during the blood-feeding process by bacteria originating from the ticks. The microbiome of the ticks was also impacted by the blood feeding. Several pathogens including tick-borne pathogens (Borrelia/Borreliella, Anaplasma, Neoehrlichia, Rickettsia) and opportunistic bacteria (Williamsia) were transmitted to the skin microbiome and some of them disseminated to the blood or spleen of the mice. In the different experiments of this study, skin microbiome alteration and Borrelia/Borreliella transmission were different depending on the tick stages (nymphs or adult female ticks). CONCLUSIONS: Host skin microbiome at the bite site was deeply impacted by the tick bite, to an extent which suggests a role in the tick feeding, in the pathogen transmission, and a potentially important impact on the skin physiopathology. The diversified taxonomic profiles of the tick microbiome were also modified by the blood feeding. Video Abstract.

Blood, Gut, and Oral Microbiome in Kidney Transplant Recipients.

Background and Objective: Recent reports describe the existence of a blood microbiome profile not associated with an infection state. Given the high impact that the dysbiotic human microbiome appears to have in chronic kidney disease and, in particular, in the outcome of kidney transplant recipients (KTRs), we aimed to explore the variations and correlations of the gut, oral, and blood microbiome of recipients, 3 months after kidney transplantation. Materials and Methods: We conducted a cross-sectional study where the microbiome of stool, saliva, and blood collected from recipients 3 months after kidney transplantation (N = 6) was analyzed by polymerase chain reaction (PCR) amplification and sequencing of the V3-V4 hypervariable regions of the 16S rRNA gene using MiSeq Illumina® technology. Results: Blood of KTRs harbors a distinct low-abundance microbiome dominated by Proteobacteria and Firmicutes. Gut and oral microbiome of KTRs also present distinct profiles. The existence of a proportion of shared operational taxonomic units among the different body sites is reported, mainly classified as Proteobacteria and Firmicutes. Conclusions: This study provides evidence of existence a blood microbiome in KTRs, different from the gut and the oral microbiome profiles, with a small number of operational taxonomic units representing a shared microbiome. The clinical relevance of this observation should be further explored in these patients.

The visceral adipose tissue bacterial microbiota provides a signature of obesity based on inferred metagenomic functions.

BACKGROUND: Metabolic inflammation mediated obesity requires bacterial molecules to trigger immune and adipose cells leading to inflammation and adipose depot development. In addition to the well-established gut microbiota dysbiosis, a leaky gut has been identified in patients with obesity and animal models, characterized by the presence of a tissue microbiota in the adipose fat pads. METHODS: To determine its potential role, we sequenced the bacterial 16 S rRNA genes in the visceral adipose depot of patients with obesity. Taking great care (surgical, biochemical, and bioinformatic) to avoid environmental contaminants. We performed statistical discriminant analyses to identify specific signatures and constructed network of interactions between variables. RESULTS: The data showed that a specific 16SrRNA gene signature was composed of numerous bacterial families discriminating between lean versus patients with obesity and people with severe obesity. The main discriminant families were Burkholderiaceae, Yearsiniaceae, and Xanthomonadaceae, all of which were gram-negative. Interestingly, the Morganellaceae were totally absent from people without obesity while preponderant in all in patients with obesity. To generate hypotheses regarding their potential role, we inferred metabolic pathways from the 16SrRNA gene signatures. We identified several pathways associated with adenosyl-cobalamine previously described to be linked with adipose tissue development. We further identified chorismate biosynthesis, which is involved in aromatic amino-acid metabolism and could play a role in fat pad development. This innovative approach generates novel hypotheses regarding the gut to adipose tissue axis. CONCLUSIONS: This innovative approach generates novel hypotheses regarding the gut to adipose tissue axis in obesity and notably the potential role of tissue microbiota.

Low-Diversity Microbiota in Apical Periodontitis and High Blood Pressure Are Signatures of the Severity of Apical Lesions in Humans.

(1) Background: In developed countries, the prevalence of apical periodontitis (AP) varies from 20% to 50% for reasons that could be associated with the apical periodontitis microbiota ecology. (2) Methods: We performed a clinical study in the Odontology department of Toulouse hospital in France, to sequence the 16S rRNA gene of AP microbiota and collect clinical parameters from 94 patients. Forty-four patients were characterized with a PAI (periapical index of AP severity) score lower or equal to 3, while the others had superior scores (n = 50). (3) Results: The low diversity of granuloma microbiota is associated with the highest severity (PAI = 5) of periapical lesions (Odds Ratio 4.592, IC 95% [1.6329; 14.0728]; p = 0.001; notably, a lower relative abundance of Burkholderiaceae and a higher relative abundance of Pseudomonas and Prevotella). We also identified that high blood pressure (HBP) is associated with the increase in PAI scores. (4) Conclusions: Our data show that a low diversity of bacterial ecology of the AP is associated with severe PAI scores, suggesting a causal mechanism. Furthermore, a second risk factor was blood pressure associated with the severity of apical periodontitis.

Human liver microbiota modeling strategy at the early onset of fibrosis.

BACKGROUND: Gut microbiota is involved in the development of liver diseases such as fibrosis. We and others identified that selected sets of gut bacterial DNA and bacteria translocate to tissues, notably the liver, to establish a non-infectious tissue microbiota composed of microbial DNA and a low frequency live bacteria. However, the precise set of bacterial DNA, and thereby the corresponding taxa associated with the early stages of fibrosis need to be identified. Furthermore, to overcome the impact of different group size and patient origins we adapted innovative statistical approaches. Liver samples with low liver fibrosis scores (F0, F1, F2), to study the early stages of the disease, were collected from Romania(n = 36), Austria(n = 10), Italy(n = 19), and Spain(n = 17). The 16S rRNA gene was sequenced. We considered the frequency, sparsity, unbalanced sample size between cohorts to identify taxonomic profiles and statistical differences. RESULTS: Multivariate analyses, including adapted spectral clustering with L1-penalty fair-discriminant strategies, and predicted metagenomics were used to identify that 50% of liver taxa associated with the early stage fibrosis were Enterobacteriaceae, Pseudomonadaceae, Xanthobacteriaceae and Burkholderiaceae. The Flavobacteriaceae and Xanthobacteriaceae discriminated between F0 and F1. Predicted metagenomics analysis identified that the preQ0 biosynthesis and the potential pathways involving glucoryranose and glycogen degradation were negatively associated with liver fibrosis F1-F2 vs F0. CONCLUSIONS: Without demonstrating causality, our results suggest first a role of bacterial translocation to the liver in the progression of fibrosis, notably at the earliest stages. Second, our statistical approach can identify microbial signatures and overcome issues regarding sample size differences, the impact of environment, and sets of analyses. TRIAL REGISTRATION: TirguMECCH ROLIVER Prospective Cohort for the Identification of Liver Microbiota, registration 4065/2014. Registered 01 01 2014.

Gut microbiota dysbiosis of type 2 diabetic mice impairs the intestinal daily rhythms of GLP-1 sensitivity.

The gut-brain-beta cell glucagon-like peptide-1 (GLP-1)-dependent axis and the clock genes both control insulin secretion. Evidence shows that a keystone of this molecular interaction could be the gut microbiota. We analyzed in mice the circadian profile of GLP-1 sensitivity on insulin secretion and the impact of the autonomic neuropathy, antibiotic treated in different diabetic mouse models and in germ-free colonized mice. We show that GLP-1sensitivity is maximal during the dark feeding period, i.e., the postprandial state. Coincidently, the ileum expression of GLP-1 receptor and peripherin is increased and tightly correlated with a subset of clock gene. Since both are markers of enteric neurons, it suggests a role in the gut-brain-beta cell GLP-1-dependent axis. We evaluated the importance of gut microbiota dysbiosis and found that the abundance of ileum bacteria, particularly Ruminococcaceae and Lachnospiraceae, oscillated diurnally, with a maximum during the dark period, along with expression patterns of a subset of clock genes. This diurnal pattern of circadian gene expression and Lachnospiraceae abundance was also observed in two separate mouse models of gut microbiota dysbiosis and of autonomic neuropathy with impaired GLP-1 sensitivity (1.high-fat diet-fed type 2 diabetic, 2.antibiotic-treated/germ-free mice). Our data show that GLP-1 sensitivity relies on specific pattern of intestinal clock gene expression and specific gut bacteria. This new statement opens opportunities to treat diabetic patient with GLP-1-based therapies by using on a possible pre/probiotic co-treatment to improve the time-dependent efficiency of these therapies.

A specific tongue microbiota signature is found in patients displaying an improvement of orosensory lipid perception after a sleeve gastrectomy.

Introduction: A preferential consumption of low-fat foods is reported by most of the patients after a vertical sleeve gastrectomy (VSG). The fact that a recent study shed light on a relationship between oral microbiota and fat taste sensitivity in obese patients prompted us to explore whether such a connection also exists in the context of a VSG. Methods: Thirty-two adult female patients with a severe obesity (BMI = 43.1 ± 0.7 kg/m2) and candidates for a VSG were selected. Oral microbiota composition surrounding the gustatory circumvallate papillae (CVP) and the lipid perception thresholds were explored before and 6 months after surgery. Results: VSG was found to be associated both with a qualitative (compositional changes) and quantitative (lower gene richness) remodeling of the peri-CVP microbiota. Analysis of the lipid perception allowed us to distinguish two subgroups: patients with a post-operative improvement of the fat taste sensitivity (i.e., with a lower threshold, n = 14) and unimproved patients (n = 18). Specific peri-CVP microbiota signatures also discriminated these two subgroups, unimproved patient being characterized by higher levels of Porphyromonas, Fusobacterium, and Haemophilus genera associated with lower levels of Atopobium and Prevotella genera as compared to the lipid-improved patients. Conclusion: Collectively, these data raise the possibility that the microbial environment surrounding gustatory papillae might play a role in the positive changes of fat taste sensitivity observed in some patients after VSG.

Polydextrose with and without Bifidobacterium animalis ssp. lactis 420 drives the prevalence of Akkermansia and improves liver health in a multi-compartmental obesogenic mice study.

The past two decades of research have raised gut microbiota composition as a contributing factor to the development of obesity, and higher abundance of certain bacterial species has been linked to the lean phenotype, such as Akkermansia muciniphila. The ability of pre- and probiotics to affect metabolic health could be via microbial community alterations and subsequently changes in metabolite profiles, modulating for example host energy balance via complex signaling pathways. The aim of this mice study was to determine how administration of a prebiotic fiber, polydextrose (PDX) and a probiotic Bifidobacterium animalis ssp. lactis 420 (B420), during high fat diet (HFD; 60 kcal% fat) affects microbiota composition in the gastrointestinal tract and adipose tissue, and metabolite levels in gut and liver. In this study C57Bl/6J mice (N = 200) were split in five treatments and daily gavaged: 1) Normal control (NC); 2) HFD; 3) HFD + PDX; 4) HFD + B420 or 5) HFD + PDX + B420 (HFD+S). At six weeks of treatment intraperitoneal glucose-tolerance test (IPGTT) was performed, and feces were collected at weeks 0, 3, 6 and 9. At end of the intervention, ileum and colon mucosa, adipose tissue and liver samples were collected. The microbiota composition in fecal, ileum, colon and adipose tissue was analyzed using 16S rDNA sequencing, fecal and liver metabolomics were performed by nuclear magnetic resonance (NMR) spectroscopy. It was found that HFD+PDX intervention reduced body weight gain and hepatic fat compared to HFD. Sequencing the mice adipose tissue (MAT) identified Akkermansia and its prevalence was increased in HFD+S group. Furthermore, by the inclusion of PDX, fecal, lleum and colon levels of Akkermansia were increased and liver health was improved as the detoxification capacity and levels of methyl-donors were increased. These new results demonstrate how PDX and B420 can affect the interactions between gut, liver and adipose tissue.

CX3CR1 regulates gut microbiota and metabolism. A risk factor of type 2 diabetes.

OBJECTIVE: The intestinal microbiota to immune system crosstalk is a major regulator of metabolism and hence metabolic diseases. An impairment of the chemokine receptor CX3CR1, as a key regulator shaping intestinal microbiota under normal chow feeding, could be one of the early events of dysglycemia. METHODS: We studied the gut microbiota ecology by sequencing the gut and tissue microbiota. We studied its role in energy metabolism in CX3CR1-deficent and control mice using various bioassays notably the glycemic regulation during fasting and the respiratory quotient as two highly sensitive physiological features. We used antibiotics and prebiotics treatments, and germ free mouse colonization. RESULTS: We identify that CX3CR1 disruption impairs gut microbiota ecology and identified a specific signature associated to the genotype. The glycemic control during fasting and the respiratory quotient throughout the day are deeply impaired. A selected four-week prebiotic treatment modifies the dysbiotic microbiota and improves the fasting state glycemic control of the CX3CR1-deficent mice and following a glucose tolerance test. A 4 week antibiotic treatment also improves the glycemic control as well. Eventually, germ free mice colonized with the microbiota from CX3CR1-deficent mice developed glucose intolerance. CONCLUSIONS: CX3CR1 is a molecular mechanism in the control of the gut microbiota ecology ensuring the maintenance of a steady glycemia and energy metabolism. Its impairment could be an early mechanism leading to gut microbiota dysbiosis and the onset of metabolic disease.

Liraglutide targets the gut microbiota and the intestinal immune system to regulate insulin secretion.

AIMS: Liraglutide controls type 2 diabetes (T2D) and inflammation. Gut microbiota regulates the immune system and causes at least in part type 2 diabetes. We here evaluated whether liraglutide regulates T2D through both gut microbiota and immunity in dysmetabolic mice. METHODS: Diet-induced dysmetabolic mice were treated for 14 days with intraperitoneal injection of liraglutide (100 µg/kg) or with vehicle or Exendin 4 (10 µg/kg) as controls. Various metabolic parameters, the intestinal immune cells were characterized and the 16SrDNA gene sequenced from the gut. The causal role of gut microbiota was shown using large spectrum antibiotics and by colonization of germ-free mice with the gut microbiota from treated mice. RESULTS: Besides, the expected metabolic impacts liraglutide treatment induced a specific gut microbiota specific signature when compared to vehicle or Ex4-treated mice. However, liraglutide only increased glucose-induced insulin secretion, reduced the frequency of Th1 lymphocytes, and increased that of TReg in the intestine. These effects were abolished by a concomitant antibiotic treatment. Colonization of germ-free mice with gut microbiota from liraglutide-treated diabetic mice improved glucose-induced insulin secretion and regulated the intestinal immune system differently from what observed in germ-free mice colonized with microbiota from non-treated diabetic mice. CONCLUSIONS: Altogether, our result demonstrated first the influence of liraglutide on gut microbiota and the intestinal immune system which could at least in part control glucose-induced insulin secretion.

The Gut and Blood Microbiome in IgA Nephropathy and Healthy Controls.

Background: IgA nephropathy (IgAN) has been associated with gut dysbiosis, intestinal membrane disruption, and translocation of bacteria into blood. Our study aimed to understand the association of gut and blood microbiomes in patients with IgAN in relation to healthy controls. Methods: We conducted a case-control study with 20 patients with progressive IgAN, matched with 20 healthy controls, and analyzed bacterial DNA quantitatively in blood using 16S PCR and qualitatively in blood and stool using 16S metagenomic sequencing. We conducted between-group comparisons as well as comparisons between the blood and gut microbiomes. Results: Higher median 16S bacterial DNA in blood was found in the IgAN group compared with the healthy controls group (7410 versus 6030 16S rDNA copies/µl blood, P=0.04). α- and ß-Diversity in both blood and stool was largely similar between the IgAN and healthy groups. In patients with IgAN, in comparison with healthy controls, we observed higher proportions of the class Coriobacteriia and species of the genera Legionella, Enhydrobacter, and Parabacteroides in blood, and species of the genera Bacteroides, Escherichia-Shigella, and some Ruminococcus in stool. Taxa distribution were markedly different between the blood and stool samples of each subject in both IgAN and healthy groups, without any significant correlation between corresponding gut and blood phyla. Conclusions: Important bacterial taxonomic differences, quantitatively in blood and qualitatively in both blood and stool samples, that were detected between IgAN and healthy groups warrant further investigation into their roles in the pathogenesis of IgAN. Although gut bacterial translocation into blood may be one of the potential sources of the blood microbiome, marked taxonomic differences between gut and blood samples in each subject in both groups confirms that the blood microbiome does not directly reflect the gut microbiome. Further research is needed into other possible sites of origin and internal regulation of the blood microbiome.

Blood Bacterial Profiles Associated With Human Immunodeficiency Virus Infection and Immune Recovery.

Human immunodeficiency virus (HIV) infection impairs mucosal immunity and leads to bacterial translocation, fueling chronic inflammation and disease progression. While this is well established, questions remain about the compositional profile of the translocated bacteria, and to what extent it is influenced by antiretroviral therapy (ART). Using 16S ribosomal DNA targeted sequencing and shotgun proteomics, we showed that HIV increases bacterial translocation from the gut to the blood. HIV increased alpha diversity in the blood, which was dominated by aerobic bacteria belonging to Micrococcaceae (Actinobacteria) and Pseudomonadaceae (Proteobacteria) families, and the number of circulating bacterial proteins was also increased. Forty-eight weeks of ART attenuated this phenomenon. We found that enrichment with Lactobacillales order, and depletion of Actinobacteria class and Moraxellaceae and Corynebacteriacae families, were significantly associated with greater immune recovery and correlated with several inflammatory markers. Our findings suggest that the molecular cross talk between the host and the translocated bacterial products could influence ART-mediated immune recovery.

Type 2 diabetes influences bacterial tissue compartmentalisation in human obesity.

Visceral obesity is a key risk factor for type 2 diabetes (T2D). Whereas gut dysbiosis appears to be instrumental for this relationship, whether gut-associated signatures translocate to extra-intestinal tissues and how this affects host metabolism remain elusive. Here we provide a comparative analysis of the microbial profile found in plasma, liver and in three distinct adipose tissues of individuals with morbid obesity. We explored how these tissue microbial signatures vary between individuals with normoglycaemia and those with T2D that were matched for body mass index. We identified tissue-specific signatures with higher bacterial load in the liver and omental adipose tissue. Gut commensals, but also environmental bacteria, showed tissue- and T2D-specific compartmentalisation. T2D signatures were most evident in mesenteric adipose tissue, in which individuals with diabetes displayed reduced bacterial diversity concomitant with fewer Gram-positive bacteria, such as Faecalibacterium, as opposed to enhanced levels of typically opportunistic Gram-negative Enterobacteriaceae. Plasma samples of individuals with diabetes were similarly enriched in Enterobacteriaceae, including the pathobiont Escherichia-Shigella. Our work provides evidence for the presence of selective plasma and tissue microbial signatures in individuals with severe obesity and identifies new potential microbial targets and biomarkers of T2D.

Blood Microbiota Modification After Myocardial Infarction Depends Upon Low-Density Lipoprotein Cholesterol Levels.

Background The role of bacteria on the onset of cardiovascular disease has been suggested. Reciprocally, increased intestinal bacterial translocation and bloodstream infection are common comorbidities associated with heart failure and myocardial infarction (MI). In this context, the aim of this study was to analyze the blood microbiome in patients shortly after acute myocardial infarction. Methods and Results We carried out a case control study comparing 103 patients at high cardiovascular risk but free of coronary disease and 99 patients who had an MI. The blood microbiome was analyzed both quantitatively by 16S quantitative polymerase chain reaction and qualitatively by 16S targeted metagenomic sequencing specifically optimized for blood samples. A significant increase in blood bacterial 16S rDNA concentration was observed in patients admitted for MI. This increase in blood bacterial DNA concentration was independent of post-MI left ventricular function and was more marked in patients with low-density lipoprotein cholesterol ≥1 g/L. In addition, differences in the proportion of numerous bacterial taxa in blood were significantly modified with the onset of MI, thus defining a blood microbiota signature of MI. Among the bacterial taxa whose proportions are decreased in patients with MI, at least 6 are known to include species able to metabolize cholesterol. Conclusions These results could provide the basis for the identification of blood microbiome-based biomarkers for the stratification of MI patients. Furthermore, these findings should provide insight into the mechanism underlying the negative correlation reported between low-density lipoprotein cholesterol concentration and the prognosis at the acute onset of MI and mortality. Clinical Trial Registration URL: http://www.clinicaltrials.gov. Unique identifier: NCT02405468.

Blood Microbiome Profile in CKD : A Pilot Study.

BACKGROUND AND OBJECTIVES: The association between gut dysbiosis, high intestinal permeability, and endotoxemia-mediated inflammation is well established in CKD. However, changes in the circulating microbiome in patients with CKD have not been studied. In this pilot study, we compare the blood microbiome profile between patients with CKD and healthy controls using 16S ribosomal DNA sequencing. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS: Blood bacterial DNA was studied in buffy coat samples quantitatively by 16S PCR and qualitatively by 16S targeted metagenomic sequencing using a molecular pipeline specifically optimized for blood samples in a cross-sectional study comparing 20 nondiabetic patients with CKD and 20 healthy controls. RESULTS: There were 22 operational taxonomic units significantly different between the two groups. 16S metagenomic sequencing revealed a significant reduction in α diversity (Chao1 index) in the CKD group compared with healthy controls (127±18 versus 145±31; P=0.04). Proteobacteria phylum, Gammaproteobacteria class, and Enterobacteriaceae and Pseudomonadaceae families were more abundant in the CKD group compared with healthy controls. Median 16S ribosomal DNA levels did not significantly differ between CKD and healthy groups (117 versus 122 copies/ng DNA; P=0.38). GFR correlated inversely with the proportion of Proteobacteria (r=-0.54; P≤0.01). CONCLUSIONS: Our pilot study demonstrates qualitative differences in the circulating microbiome profile with lower α diversity and significant taxonomic variations in the blood microbiome in patients with CKD compared with healthy controls.

Author Correction: Obese Subjects With Specific Gustatory Papillae Microbiota and Salivary Cues Display an Impairment to Sense Lipids.

A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has been fixed in the paper.