Shotgun metagenomics and systemic targeted metabolomics highlight indole-3-propionic acid as a protective gut microbial metabolite against influenza infection.

The gut-to-lung axis is critical during respiratory infections, including influenza A virus (IAV) infection. In the present study, we used high-resolution shotgun metagenomics and targeted metabolomic analysis to characterize influenza-associated changes in the composition and metabolism of the mouse gut microbiota. We observed several taxonomic-level changes on day (D)7 post-infection, including a marked reduction in the abundance of members of the Lactobacillaceae and Bifidobacteriaceae families, and an increase in the abundance of Akkermansia muciniphila. On D14, perturbation persisted in some species. Functional scale analysis of metagenomic data revealed transient changes in several metabolic pathways, particularly those leading to the production of short-chain fatty acids (SCFAs), polyamines, and tryptophan metabolites. Quantitative targeted metabolomics analysis of the serum revealed changes in specific classes of gut microbiota metabolites, including SCFAs, trimethylamine, polyamines, and indole-containing tryptophan metabolites. A marked decrease in indole-3-propionic acid (IPA) blood level was observed on D7. Changes in microbiota-associated metabolites correlated with changes in taxon abundance and disease marker levels. In particular, IPA was positively correlated with some Lactobacillaceae and Bifidobacteriaceae species (Limosilactobacillus reuteri, Lactobacillus animalis) and negatively correlated with Bacteroidales bacterium M7, viral load, and inflammation markers. IPA supplementation in diseased animals reduced viral load and lowered local (lung) and systemic inflammation. Treatment of mice with antibiotics targeting IPA-producing bacteria before infection enhanced viral load and lung inflammation, an effect inhibited by IPA supplementation. The results of this integrated metagenomic-metabolomic analysis highlighted IPA as an important contributor to influenza outcomes and a potential biomarker of disease severity.

Type 2-high asthma is associated with a specific indoor mycobiome and microbiome.

BACKGROUND: The links between microbial environmental exposures and asthma are well documented, but no study has combined deep sequencing results from pulmonary and indoor microbiomes of patients with asthma with spirometry, clinical, and endotype parameters. OBJECTIVE: The goal of this study was to investigate the links between indoor microbial exposures and pulmonary microbial communities and to document the role of microbial exposures on inflammatory and clinical outcomes of patients with severe asthma (SA). METHODS: A total of 55 patients with SA from the national Cohort of Bronchial Obstruction and Asthma cohort were enrolled for analyzing their indoor microbial flora through the use of electrostatic dust collectors (EDCs). Among these patients, 22 were able to produce sputum during "stable" or pulmonary "exacerbation" periods and had complete pairs of EDC and sputum samples, both collected and analyzed. We used amplicon targeted metagenomics to compare microbial communities from EDC and sputum samples of patients according to type 2 (T2)-asthma endotypes. RESULTS: Compared with patients with T2-low SA, patients with T2-high SA exhibited an increase in bacterial α-diversity and a decrease in fungal α-diversity of their indoor microbial florae, the latter being significantly correlated with fraction of exhaled nitric oxide levels. The ß-diversity of the EDC mycobiome clustered significantly according to T2 endotypes. Moreover, the proportion of fungal taxa in common between the sputum and EDC samples was significantly higher when patients exhibited acute exacerbation. CONCLUSION: These results illustrated, for the first time, a potential association between the indoor mycobiome and clinical features of patients with SA, which should renew interest in deciphering the interactions between indoor environment, fungi, and host in asthma.

Chlorinated ethene biodegradation and associated bacterial taxa in multi-polluted groundwater: Insights from biomolecular markers and stable isotope analysis.

Chlorinated ethenes (CEs) are most problematic pollutants in groundwater. Dehalogenating bacteria, and in particular organohalide-respiring bacteria (OHRB), can transform PCE to ethene under anaerobic conditions, and thus contribute to bioremediation of contaminated sites. Current approaches to characterize in situ biodegradation of CEs include hydrochemical analyses, quantification of the abundance of key species (e.g. Dehalococcoides mccartyi) and dehalogenase genes (pceA, vcrA, bvcA and tceA) involved in different steps of organohalide respiration (OHR) by qPCR, and compound-specific isotope analysis (CSIA) of CEs. Here we combined these approaches with sequencing of 16S rRNA gene amplicons to consider both OHRB and bacterial taxa involved in CE transformation at a multi-contaminated site. Integrated analysis of hydrogeochemical characteristics, gene abundances and bacterial diversity shows that bacterial diversity and OHRB mainly correlated with hydrogeochemical conditions, suggesting that pollutant exposure acts as a central driver of bacterial diversity. CSIA, abundances of four reductive dehalogenase encoding genes and the prevalence of Dehalococcoides highlighted sustained PCE, DCE and VC degradation in several wells of the polluted plume. These results suggest that bacterial taxa associated with OHR play an essential role in natural attenuation of CEs, and that representatives of taxa including Dehalobacterium and Desulfosporosinus co-occur with Dehalococcoides. Overall, our study emphasizes the benefits of combining several approaches to evaluate the interplay between the dynamics of bacterial diversity in CE-polluted plumes and in situ degradation of CEs, and to contribute to a more robust assessment of natural attenuation at multi-polluted sites.

Fungal Species Diversity in French Bread Sourdoughs Made of Organic Wheat Flour.

Microbial communities are essential for the maintenance and functioning of ecosystems, including fermented food ecosystems. The analysis of food microbial communities is mainly focused on lactic acid bacteria (LAB), while yeast diversity is less understood. Here, we describe the fungal diversity of a typical food fermented product, sourdough bread. The species diversity of 14 sourdoughs collected from bakeries located all over France was analyzed. Bakeries were chosen to represent diverse bakery practices and included bakers and farmer-bakers. Both non-culture-based (pyrosequencing of Internal Transcribed Spacer 1 amplicons) and culture-based methods were used. While both identification methods were in agreement regarding the dominant yeast species of each sourdough, the ITS1 metabarcoding analysis identified an increased number of fungal species in sourdough communities. Two third of the identified sequences obtained from sourdoughs were Saccharomycetales, mostly in the Kazachstania genus. No Saccharomycetales species was shared by all the sourdoughs, whereas five other fungal species, mainly known plant pathogens, were found in all sourdoughs. Interestingly, Saccharomyces cerevisiae, known as "baker's yeast," was identified as the dominant species in only one sourdough. By contrast, five Kazachstania species were identified as the dominant sourdough species, including one recently described Kazachstania species, Kazachstania saulgeensis and an undescribed Kazachstania sp. Sourdoughs from farmer-bakers harbored Kazachstania bulderi, Kazachstania unispora and two newly described Kazachstania species, while sourdough from bakers mostly carried Kazachstania humilis as the dominant species. Such yeast diversity has not been found in sourdoughs before, highlighting the need to maintain different traditional food practices to conserve microbial diversity.

Functional Genes and Bacterial Communities During Organohalide Respiration of Chloroethenes in Microcosms of Multi-Contaminated Groundwater.

Microcosm experiments with CE-contaminated groundwater from a former industrial site were set-up to evaluate the relationships between biological CE dissipation, dehalogenase genes abundance and bacterial genera diversity. Impact of high concentrations of PCE on organohalide respiration was also evaluated. Complete or partial dechlorination of PCE, TCE, cis-DCE and VC was observed independently of the addition of a reducing agent (Na2S) or an electron donor (acetate). The addition of either 10 or 100 µM PCE had no effect on organohalide respiration. qPCR analysis of reductive dehalogenases genes (pceA, tceA, vcrA, and bvcA) indicated that the version of pceA gene found in the genus Dehalococcoides [hereafter named pceA(Dhc)] and vcrA gene increased in abundance by one order of magnitude during the first 10 days of incubation. The version of the pceA gene found, among others, in the genus Dehalobacter, Sulfurospirillum, Desulfuromonas, and Geobacter [hereafter named pceA(Dhb)] and bvcA gene showed very low abundance. The tceA gene was not detected throughout the experiment. The proportion of pceA(Dhc) or vcrA genes relative to the universal 16S ribosomal RNA (16S rRNA) gene increased by up to 6-fold upon completion of cis-DCE dissipation. Sequencing of 16S rRNA amplicons indicated that the abundance of Operational Taxonomic Units (OTUs) affiliated to dehalogenating genera Dehalococcoides, Sulfurospirillum, and Geobacter represented more than 20% sequence abundance in the microcosms. Among organohalide respiration associated genera, only abundance of Dehalococcoides spp. increased up to fourfold upon complete dissipation of PCE and cis-DCE, suggesting a major implication of Dehalococcoides in CEs organohalide respiration. The relative abundance of pceA and vcrA genes correlated with the occurrence of Dehalococcoides and with dissipation extent of PCE, cis-DCE and CV. A new type of dehalogenating Dehalococcoides sp. phylotype affiliated to the Pinellas group, and suggested to contain both pceA(Dhc) and vcrA genes, may be involved in organohalide respiration of CEs in groundwater of the study site. Overall, the results demonstrate in situ dechlorination potential of CE in the plume, and suggest that taxonomic and functional biomarkers in laboratory microcosms of contaminated groundwater following pollutant exposure can help predict bioremediation potential at contaminated industrial sites.

Enabling large-scale feather mite studies: an Illumina DNA metabarcoding pipeline.

Feather mites are among the most common and diverse ectosymbionts of birds, yet basic questions such as the nature of their relationship remain largely unanswered. One reason for feather mites being understudied is that their morphological identification is often virtually impossible when using female or young individuals. Even for adult male specimens this task is tedious and requires advanced taxonomic expertise, thus hampering large-scale studies. In addition, molecular-based methods are challenging because the low DNA amounts usually obtained from these tiny mites do not reach the levels required for high-throughput sequencing. This work aims to overcome these issues by using a DNA metabarcoding approach to accurately identify and quantify the feather mite species present in a sample. DNA metabarcoding is a widely used molecular technique that takes advantage of high-throughput sequencing methodologies to assign the taxonomic identity to all the organisms present in a complex sample (i.e., a sample made up of multiple specimens that are hard or impossible to individualise). We present a high-throughput method for feather mite identification using a fragment of the COI gene as marker and Illumina Miseq technology. We tested this method by performing two experiments plus a field test over a total of 11,861 individual mites (5360 of which were also morphologically identified). In the first experiment, we tested the probability of detecting a single feather mite in a heterogeneous pool of non-conspecific individuals. In the second experiment, we made 2 × 2 combinations of species and studied the relationship between the proportion of individuals of a given species in a sample and the proportion of sequences retrieved to test whether DNA metabarcoding can reliably quantify the relative abundance of mites in a sample. Here we also tested the efficacy of degenerate primers (i.e., a mixture of similar primers that differ in one or several bases that are designed to increase the chance of annealing) and investigated the relationship between the number of mismatches and PCR success. Finally, we applied our DNA metabarcoding pipeline to a total of 6501 unidentified and unsorted feather mite individuals sampled from 380 European passerine birds belonging to 10 bird species (field test). Our results show that this proposed pipeline is suitable for correct identification and quantitative estimation of the relative abundance of feather mite species in complex samples, especially when dealing with a moderate number (> 30) of individuals per sample.

Three novel ascomycetous yeast species of the Kazachstania clade, Kazachstania saulgeensis sp. nov., Kazachstaniaserrabonitensis sp. nov. and Kazachstania australis sp. nov. Reassignment of Candida humilis to Kazachstania humilis f.a. comb. nov. and Candida pseudohumilis to Kazachstania pseudohumilis f.a. comb. nov.

Five ascosporogenous yeast strains related to the genus Kazachstania were isolated. Two strains (CLIB 1764T and CLIB 1780) were isolated from French sourdoughs; three others (UFMG-CM-Y273T, UFMG-CM-Y451 and UFMG-CM-Y452) were from rotting wood in Brazil. The sequences of the French and Brazilian strains differed by one and three substitutions, respectively, in the D1/D2 large subunit (LSU) rRNA gene and the internal transcribed spacer (ITS). The D1/D2 LSU rRNA sequence of these strains differed by 0.5 and 0.7 % from Kazachstania exigua, but their ITS sequences diverged by 8.1 and 8.3 %, respectively, from that of the closest described species Kazachstania barnettii. Analysis of protein coding sequences of RPB1, RPB2 and EF-1α distinguished the French from the Brazilian strains, with respectively 3.3, 6 and 11.7 % substitutions. Two novel species are described to accommodate these newly isolated strains: Kazachstania saulgeensis sp. nov. (type strain CLIB 1764T=CBS 14374T) and Kazachstania serrabonitensis sp. nov. (type strain UFMG-CM-Y273T=CLIB 1783T=CBS 14236T). Further analysis of culture collections revealed a strain previously assigned to the K. exigua species, but having 3.8 % difference (22 substitutions and 2 indels) in its ITS with respect to K. exigua. Hence, we describe a new taxon, Kazachstania australis sp. nov. (type strain CLIB 162T=CBS 2141T), to accommodate this strain. Finally, Candida humilis and Candida pseudohumilis are reassigned to the genus Kazachstania as new combinations. On the basis of sequence analysis, we also propose that Candida milleri and Kazachstania humilis comb. nov. are conspecific.

Sourdough microbial community dynamics: An analysis during French organic bread-making processes.

Natural sourdoughs are commonly used in bread-making processes, especially for organic bread. Despite its role in bread flavor and dough rise, the stability of the sourdough microbial community during and between bread-making processes is debated. We investigated the dynamics of lactic acid bacteria (LAB) and yeast communities in traditional organic sourdoughs of five French bakeries during the bread-making process and several months apart using classical and molecular microbiology techniques. Sourdoughs were sampled at four steps of the bread-making process with repetition. The analysis of microbial density over 68 sourdough/dough samples revealed that both LAB and yeast counts changed along the bread-making process and between bread-making runs. The species composition was less variable. A total of six LAB and nine yeast species was identified from 520 and 1675 isolates, respectively. The dominant LAB species was Lactobacillus sanfranciscensis, found for all bakeries and each bread-making run. The dominant yeast species changed only once between bread-making processes but differed between bakeries. They mostly belonged to the Kazachstania clade. Overall, this study highlights the change of population density within the bread-making process and between bread-making runs and the relative stability of the sourdough species community during bread-making process.