Human Gut Microbial Community Research Articles

Readsynth: short-read simulation for consideration of composition-biases in reduced metagenome sequencing approaches

BackgroundThe application of reduced metagenomic sequencing approaches holds promise as a middle ground between targeted amplicon sequencing and whole metagenome sequencing approaches but has not been widely adopted as a technique. A major barrier to adoption is the lack of read simulation software built to handle characteristic features of these novel approaches. Reduced metagenomic sequencing (RMS) produces unique patterns of fragmentation per genome that are sensitive to restriction enzyme choice, and the non-uniform size selection of these fragments may introduce novel challenges to taxonomic assignment as well as relative abundance estimates.ResultsThrough the development and application of simulation software, readsynth, we compare simulated metagenomic sequencing libraries with existing RMS data to assess the influence of multiple library preparation and sequencing steps on downstream analytical results. Based on read depth per position, readsynth achieved 0.79 Pearson’s correlation and 0.94 Spearman’s correlation to these benchmarks. Application of a novel estimation approach, fixed length taxonomic ratios, improved quantification accuracy of simulated human gut microbial communities when compared to estimates of mean or median coverage.ConclusionsWe investigate the possible strengths and weaknesses of applying the RMS technique to profiling microbial communities via simulations with readsynth. The choice of restriction enzymes and size selection steps in library prep are non-trivial decisions that bias downstream profiling and quantification. The simulations investigated in this study illustrate the possible limits of preparing metagenomic libraries with a reduced representation sequencing approach, but also allow for the development of strategies for producing and handling the sequence data produced by this promising application.

Modeling Dynamics of Human Gut Microbiota Derived from Gluten Metabolism: Obtention, Maintenance and Characterization of Complex Microbial Communities.

Western diets are rich in gluten-containing products, which are frequently poorly digested. The human large intestine harbors microorganisms able to metabolize undigested gluten fragments that have escaped digestion by human enzymatic activities. The aim of this work was obtaining and culturing complex human gut microbial communities derived from gluten metabolism to model the dynamics of healthy human large intestine microbiota associated with different gluten forms. For this purpose, stool samples from six healthy volunteers were inoculated in media containing predigested gluten or predigested gluten plus non-digested gluten. Passages were carried out every 24 h for 15 days in the same medium and community composition along time was studied via V3-V4 16S rDNA sequencing. Diverse microbial communities were successfully obtained. Moreover, communities were shown to be maintained in culture with stable composition for 14 days. Under non-digested gluten presence, communities were enriched in members of Bacillota, such as Lachnospiraceae, Clostridiaceae, Streptococcaceae, Peptoniphilaceae, Selenomonadaceae or Erysipelotrichaceae, and members of Actinomycetota, such as Bifidobacteriaceae and Eggerthellaceae. Contrarily, communities exposed to digested gluten were enriched in Pseudomonadota. Hence, this study shows a method for culture and stable maintenance of gut communities derived from gluten metabolism. This method enables the analysis of microbial metabolism of gluten in the gut from a community perspective.

Massively parallel single-cell sequencing of diverse microbial populations.

Single-cell genetic heterogeneity is ubiquitous in microbial populations and an important aspect of microbial biology; however, we lack a broadly applicable and accessible method to study this heterogeneity in microbial populations. Here, we show a simple, robust and generalizable method for high-throughput single-cell sequencing of target genetic loci in diverse microbes using simple droplet microfluidics devices (droplet targeted amplicon sequencing; DoTA-seq). DoTA-seq serves as a platform to perform diverse assays for single-cell genetic analysis of microbial populations. Using DoTA-seq, we demonstrate the ability to simultaneously track the prevalence and taxonomic associations of >10 antibiotic-resistance genes and plasmids within human and mouse gut microbial communities. This workflow is a powerful and accessible platform for high-throughput single-cell sequencing of diverse microbial populations.

DIVERSITY OF β-GLUCURONIDASE AMONG THE MICROBIOME OF HEALTHY INDIVIDUALS AND PCOS PATIENTS

PCOS is a heterogeneous disorder caused by reproductive and immuno-metabolic abnormalities. It is accompanied by dysbiosis of the human gut microbial community. Bacterial enzyme β-glucuronidase (GUSB) performs deglucuronidation of conjugated estrogen, resulting in abnormal conc. of this hormone in females and PCOS incidence. Present study was initiated to characterize the GUSB enzyme in bacteria associated with the gut of healthy and PCOS individuals. Enzyme sequences from ten bacteria were retrieved from the UniProt database and characterized by CELLO, Protparam, SOPMA, AlphaFold, and HDOCK tools. Analysis revealed the localization of enzymes in periplasm and cytoplasm in most bacteria and, in addition to this, the outer membrane only in B. intestinalis. PCOS-associated enzymes were alkaline, with high thermostability and in-vitro stability compared to healthy gut bacterial enzymes. Secondary (2D) and tertiary (3D) structures were comparable in enzymes of both these groups. The affinity of GUSB was higher for catechin in PCOS-associated bacteria than bacteria found in a healthy gut. Hence, catechin can be an effective ligand for inhibiting the GUSB enzyme in PCOS patients.

Anaerobutyricum hallii promotes the functional depletion of a food carcinogen in diverse healthy fecal microbiota

IntroductionAnaerobutyricum hallii is a human gut commensal that transforms the heterocyclic amine 2-amino-1-methyl-6-phenylimidazo [4,5-b] pyridine (PhIP), a carcinogen from cooked meat. The transformation mechanism involves the microbial production of acrolein from glycerol, and its conjugation with PhIP, thus blocking its mutagenic potential. A potential cancer prevention strategy could therefore involve supplementing complex human microbial communities with metabolically competent bacteria such as A. hallii that can deplete PhIP. However, it has not been established how the proportion of A. hallii in diverse healthy human gut microbial communities relates to functional capacity for PhIP transformation and, moreover, how supplementing microbiomes with A. hallii affects this function.MethodsIn this study, shotgun metagenomics was used to study taxonomic profiling, the abundance of glycerol/diol dehydratase (gdh)-harboring taxa, the proportion of resident A. hallii, and the reconstruction of A. hallii population genomes in the fecal samples of 20 healthy young adult donors. Furthermore, the influence of supplementing 106 cells/mL of A. hallii DSM 3353 with diluted fecal microbiota was characterized.Results and discussionSix microbiota were assigned to Bacteroides, nine to Prevotella, and five to Ruminococcus by enterotype-associated clustering. The total number of gdh copies in the 20 fecal microbiota expressed per 1010 bacterial cells ranged between 1.32 × 108 and 1.15 × 109. Eighteen out of the 20 donors were dominated by A. hallii, representing between 33% and 94% of the total gdh relative abundance of the samples. The microbiota with low A. hallii abundance (i.e., with a relative abundance < 1%) transformed less PhIP than the microbiota with high A. hallii abundance (i.e., with a relative abundance > 1%). Furthermore, supplementing the low-A. hallii-abundant microbiota with glycerol significantly increased the PhIP transformation capacity after 6 h while reducing total short-chain fatty acid (SCFA) levels, which is most likely due to acrolein production. Although acetate decreased in all microbiota with glycerol and with the combination of glycerol and A. hallii, for most of the microbiomes, butyrate production increased over time. Thus, for a significant number of diverse healthy human fecal microbiomes, and especially when they have little of the taxa to start with, supplementing A. hallii increases PhIP transformation. These findings suggest the need to test in vivo whether supplementing microbiomes with A. hallii reduces PhIP exposure.

Genome enrichment of rare and unknown species from complicated microbiome by nanopore selective sequencing.

Rare species are vital members of a microbial community, but retrieving their genomes is difficult due to their low abundance. The ReadUntil (RU) approach allows nanopore devices to sequence specific DNA molecules selectively in real-time, which provides an opportunity for enriching rare species. Despite the robustness of enriching rare species by reducing the sequencing depth of known host sequences, such as the human genome, there is still a gap in RU-based enriching of rare species in environmental samples whose community composition is unclear, and many rare species have poor or incomplete reference genomes in public databases. Therefore, here we present metaRUpore to overcome this challenge. We applied metaRUpore to a thermophilic anaerobic digester (TAD) community and human gut microbial community, it reduced coverage of the high-abundance populations and modestly increased (~2×) the genome coverage of the rare taxa, facilitating successful recovery of near-finished metagenome-assembled genomes (nf-MAGs) of rare species. The simplicity and robustness of the approach make it accessible for labs with moderate computational resources and hold the potential to become the standard practice in future metagenomic sequencing of complicated microbiomes.

Tracing human life trajectory using gut microbial communities by context-aware deep learning.

The gut microbial communities are highly plastic throughout life, and the human gut microbial communities show spatial-temporal dynamic patterns at different life stages. However, the underlying association between gut microbial communities and time-related factors remains unclear. The lack of context-awareness, insufficient data, and the existence of batch effect are the three major issues, making the life trajection of the host based on gut microbial communities problematic. Here, we used a novel computational approach (microDELTA, microbial-based deep life trajectory) to track longitudinal human gut microbial communities' alterations, which employs transfer learning for context-aware mining of gut microbial community dynamics at different life stages. Using an infant cohort, we demonstrated that microDELTA outperformed Neural Network for accurately predicting the age of infant with different delivery mode, especially for newborn infants of vaginal delivery with the area under the receiver operating characteristic curve of microDELTA and Neural Network at 0.811 and 0.436, respectively. In this context, we have discovered the influence of delivery mode on infant gut microbial communities. Along the human lifespan, we also applied microDELTA to a Chinese traveler cohort, a Hadza hunter-gatherer cohort and an elderly cohort. Results revealed the association between long-term dietary shifts during travel and adult gut microbial communities, the seasonal cycling of gut microbial communities for the Hadza hunter-gatherers, and the distinctive microbial pattern of elderly gut microbial communities. In summary, microDELTA can largely solve the issues in tracing the life trajectory of the human microbial communities and generate accurate and flexible models for a broad spectrum of microbial-based longitudinal researches.

Healthy adult gut microbiota sustains its own vitamin B12 requirement in an in vitro batch fermentation model.

Vitamin B12 (cobalamin) is present in the human lower gastrointestinal tract either coming from the unabsorbed dietary fraction or from in situ production of the gut microbiota. However, it is unclear whether the gut microbial communities need exogenous B12 for growth and metabolism, or whether B12 in low and high levels could affect gut community composition and metabolite production. Here, we investigated in vitro B12 production of human fecal microbiota and the effects of different levels of B12 (as cyanocobalamin) on composition and activity. Eight fecal communities from healthy human adults distributed over three enterotypes, dominated by Firmicutes (n = 5), Bacteroides (n = 1) or Prevotella (n = 2) were used to perform batch fermentations in Macfarlane medium supplemented with low B12 medium (Control, 5 ng/ml, within the tested fecal range), no B12 addition (NB12), and high B12 addition (ExtraB12, 2500 ng/ml). The microbiota community composition (qPCR, 16S rRNA metabarcoding), metabolic activity (HPLC-RI), and B12 levels (UHPLC-DAD) were measured after 24 h incubation at 37°C under strict anaerobic conditions. All fecal microbial communities produced B12 in the NB12 condition after 24 h, in the range from 152 ± 4 to 564 ± 25 ng/ml. None of the B12 treatments had an impact on total bacterial growth, community richness, diversity and total metabolite production, compared to the low B12 control. However, a significant increase of propionate was measured in ExtraB12 compared to NB12. Most taxonomic and metabolite changes compared to control incubations were donor-dependent, implying donor-microbiota-specific changes upon B12 treatments. Our in vitro data suggest that healthy human adult gut microbial communities have the capacity to produce B12 at levels fulfilling their own requirements, independently of the initial B12 content tested in the donor's feces. Further, supplementation of exogenous dietary B12 may have limited impact on the healthy human gut microbial community composition and function.

A gut microbial metabolite of dietary polyphenols reverses obesity-driven hepatic steatosis

The molecular mechanisms by which dietary fruits and vegetables confer cardiometabolic benefits remain poorly understood. Historically, these beneficial properties have been attributed to the antioxidant activity of flavonoids. Here, we reveal that the host metabolic benefits associated with flavonoid consumption hinge, in part, on gut microbial metabolism. Specifically, we show that a single gut microbial flavonoid catabolite, 4-hydroxyphenylacetic acid (4-HPAA), is sufficient to reduce diet-induced cardiometabolic disease (CMD) burden in mice. The addition of flavonoids to a high fat diet heightened the levels of 4-HPAA within the portal plasma and attenuated obesity, and continuous delivery of 4-HPAA was sufficient to reverse hepatic steatosis. The antisteatotic effect was shown to be associated with the activation of AMP-activated protein kinase α (AMPKα). In a large survey of healthy human gut metagenomes, just over one percent contained homologs of all four characterized bacterial genes required to catabolize flavonols into 4-HPAA. Our results demonstrate the gut microbial contribution to the metabolic benefits associated with flavonoid consumption and underscore the rarity of this process in human gut microbial communities.

2'-Fucosyllactose inhibits proliferation of Clostridioides difficile ATCC 43599 in the CDi-screen, an in vitro model simulating Clostridioides difficile infection.

Clostridioides difficile is a Gram-positive anaerobic bacterium that can produce the toxins TcdA and/or TcdB and is considered an opportunistic pathogen. C. difficile is mainly transmitted as endospores, which germinate to produce the pathogenic vegetative cells under suitable conditions in the gut. To efficiently screen novel therapeutic- interventions against the proliferation of C. difficile within a complex microbial community, platforms are needed that facilitate parallel experimentation. In order to allow for screening of novel interventions a medium-to-high throughput in vitro system is desirable. To this end, we have developed the 96-well CDi-screen platform that employs an adapted simulated ileal effluent medium (CDi-SIEM) and allows for culturing of pathogenic C. difficile. C. difficile strain ATCC 43599 was inoculated in the form of vegetative cells and spores into the CDi-screen in the presence and absence of a cultured fecal microbiota and incubated for 48h. To demonstrate its utility, we investigated the effect of the human milk oligosaccharide 2'-Fucosyllactose (2'-FL) at 4 and 8 mg/mL on C. difficile outgrowth and toxin production in the CDi-screen. The test conditions were sampled after 24 and 48 hours. C. difficile -specific primers were used to monitor C. difficile growth via qPCR and barcoded 16S rRNA gene amplicon sequencing facilitated the in-depth analysis of gut microbial community dynamics. C. difficile ATCC 43599 proliferated in CDi-SIEM, both when inoculated as spores and as vegetative cells. The strain reached cell numbers expressed as C. difficile genome equivalents of up to 10 8 cells per mL after 24h of incubation. 2'-FL significantly inhibited the outgrowth of the ATTC 43599 strain within a complex human gut microbial community in the CDi-screen. In addition, a dose-dependent modulation of the gut microbial community composition by 2'-FL supplementation was detected, with a significant increase in the relative abundance of the genus Blautia in the presence of 2'-FL. The CDi-screen is suitable for studying C. difficile proliferation in a complex gut ecosystem and for screening for anti-pathogenic interventions that target C. difficile directly and/or indirectly through interactions with the gut microbiota. Different doses of compounds such as in this study the dose of the human milk oligosaccharide 2'-FL can be screened for efficacy in the inhibition of C. difficile proliferation.

MetaLP: An integrative linear programming method for protein inference in metaproteomics.

Metaproteomics based on high-throughput tandem mass spectrometry (MS/MS) plays a crucial role in characterizing microbiome functions. The acquired MS/MS data is searched against a protein sequence database to identify peptides, which are then used to infer a list of proteins present in a metaproteome sample. While the problem of protein inference has been well-studied for proteomics of single organisms, it remains a major challenge for metaproteomics of complex microbial communities because of the large number of degenerate peptides shared among homologous proteins in different organisms. This challenge calls for improved discrimination of true protein identifications from false protein identifications given a set of unique and degenerate peptides identified in metaproteomics. MetaLP was developed here for protein inference in metaproteomics using an integrative linear programming method. Taxonomic abundance information extracted from metagenomics shotgun sequencing or 16s rRNA gene amplicon sequencing, was incorporated as prior information in MetaLP. Benchmarking with mock, human gut, soil, and marine microbial communities demonstrated significantly higher numbers of protein identifications by MetaLP than ProteinLP, PeptideProphet, DeepPep, PIPQ, and Sipros Ensemble. In conclusion, MetaLP could substantially improve protein inference for complex metaproteomes by incorporating taxonomic abundance information in a linear programming model.

Gut microbe metabolism of small molecules supports human development across the early stages of life.

From birth to adulthood, the human gut-associated microbial communities experience profound changes in their structure. However, while the taxonomical composition has been extensively explored, temporal shifts in the microbial metabolic functionalities related to the metabolism of bioactive small molecules are still largely unexplored. Here, we collected a total of 6,617 publicly available human fecal shotgun metagenomes and 42 metatranscriptomes from infants and adults to explore the dynamic changes of the microbial-derived small molecule metabolisms according to the age-related development of the human gut microbiome. Moreover, by selecting metagenomic data from 250 breastfed and 217 formula-fed infants, we also investigated how feeding types can shape the metabolic functionality of the incipient gut microbiome. From the small molecule metabolism perspective, our findings suggested that the human gut microbial communities are genetically equipped and prepared to metabolically evolve toward the adult state as early as 1 month after birth, although at the age of 4 years, it still appeared functionally underdeveloped compared to adults. Furthermore, in respect of formula-fed newborns, breastfed infants showed enrichment in microbial metabolic functions related to specific amino acids present at low concentrations in human milk, highlighting that the infant gut microbiome has specifically evolved to synthesize bioactive molecules that can complement the human breast milk composition contributing to complete nutritional supply of infant.

High throughput genome scale modeling predicts microbial vitamin requirements contribute to gut microbiome community structure

ABSTRACT Human gut microbiome structure and emergent metabolic outputs impact health outcomes. However, what drives such community characteristics remains underexplored. Here, we rely on high throughput genomic reconstruction modeling, to infer the metabolic attributes and nutritional requirements of 816 gut strains, via a framework termed GEMNAST. This has been performed in terms of a group of human vitamins to examine the role vitamin exchanges have at different levels of community organization. We find that only 91 strains can satisfy their vitamin requirements (prototrophs) while the rest show various degrees of auxotrophy/specialization, highlighting their dependence on external sources, such as other members of the microbial community. Further, 79% of the strains in our sample were mapped to 11 distinct vitamin requirement profiles with low phylogenetic consistency. Yet, we find that human gut microbial community enterotype indicators display marked metabolic differences. Prevotella strains display a metabolic profile that can be complemented by strains from other genera often associated with the Prevotella enterotype and agrarian diets, while Bacteroides strains occupy a prototrophic profile. Finally, we identify pre-defined interaction modules (IMs) of gut species from human and mice predicted to be driven by, or highly independent of vitamin exchanges. Our analysis provides mechanistic grounding to gut microbiome stability and to co-abundance-based observations, a fundamental step toward understanding emergent processes that influence health outcomes. Further, our work opens a path to future explorations in the field through applications of GEMNAST to additional nutritional dimensions.

Antidepressants promote the spread of extracellular antibiotic resistance genes via transformation

The development of antibiotic resistance as an unavoidable consequence of the application of antimicrobials is a significant concern for human health. Antidepressants are being increasingly consumed globally. Human gut microbial communities are frequently exposed to antidepressants, yet little is known about the interaction between antidepressants and antibiotic resistance. This study aimed to investigate whether antidepressants can accelerate the dissemination of antibiotic resistance by increasing the rate of the horizontal transfer of antibiotic resistance genes (ARGs). Results demonstrated that some of the commonly-prescribed antidepressants (Duloxetine, Sertraline, Fluoxetine and Bupropion) at clinically relevant concentrations can significantly (n = 9; padj < 0.01) promote the transformation of extracellular ARGs into Acinetobacter baylyi ADP1 for a maximum of 2.3-fold, which is primarily associated with the overproduction of reactive oxygen species. The increased cell membrane permeability and porosity, stimulated transcription and translation of competence, SOS response, universal stress response and ATP synthesis-related genes are also associated with antidepressants-enhanced transformation. This study demonstrated that some antidepressants can speed up the spread of antibiotic resistance by promoting the transformation of ARGs, which emphasizes the necessity to assess the potential risks of antidepressants in spreading antibiotic resistance during clinical antidepressant applications.

The Prebiotic Effect of Australian Seaweeds on Commensal Bacteria and Short Chain Fatty Acid Production in a Simulated Gut Model.

Diet is known to affect the composition and metabolite production of the human gut microbial community, which in turn is linked with the health and immune status of the host. Whole seaweeds (WH) and their extracts contain prebiotic components such as polysaccharides (PS) and polyphenols (PP). In this study, the Australian seaweeds, Phyllospora comosa, Ecklonia radiata, Ulva ohnoi, and their PS and PP extracts were assessed for potential prebiotic activities using an in vitro gut model that included fresh human faecal inoculum. 16S rRNA sequencing post gut simulation treatment revealed that the abundance of several taxa of commensal bacteria within the phylum Firmicutes linked with short chain fatty acid (SCFA) production, and gut and immune function, including the lactic acid producing order Lactobacillales and the chief butyrate-producing genera Faecalibacteria, Roseburia, Blautia, and Butyricicoccus were significantly enhanced by the inclusion of WH, PS and PP extracts. After 24 h fermentation, the abundance of total Firmicutes ranged from 57.35–81.55% in the WH, PS and PP samples, which was significantly greater (p ≤ 0.01) than the inulin (INU) polysaccharide control (32.50%) and the epigallocatechingallate (EGCG) polyphenol control (67.13%); with the exception of P. comosa PP (57.35%), which was significantly greater than INU only. However, all WH, PS and PP samples also increased the abundance of the phylum Proteobacteria; while the abundance of the phylum Actinobacteria was decreased by WH and PS samples. After 24 h incubation, the total and individual SCFAs present, including butyric, acetic and propionic acids produced by bacteria fermented with E. radiata and U. ohnoi, were significantly greater than the SCFAs identified in the INU and EGCG controls. Most notably, total SCFAs in the E. radiata PS and U. ohnoi WH samples were 227.53 and 208.68 µmol/mL, respectively, compared to only 71.05 µmol/mL in INU and 7.76 µmol/mL in the EGCG samples. This study demonstrates that whole seaweeds and their extracts have potential as functional food ingredients to support normal gut and immune function.

Gut microbiota in alopecia areata.

In the past few years, the advancement of 16S rRNA metagenomic analysis sequencing has enabled assessing the impact of gut microbiota on the development of skin disease. Alopecia areata (AA) is a nonscarring hair loss disorder with an unknown etiopathogenesis, however, it is hypothesised that a combination of genetic and environmental factors might be involved. Although numerous studies have shown that the microbiome plays a key role at the beginning of skin diseases, the link between AA and gut dysbiosis remains unclear. To analyse the intestinal microbiome in patients suffering from AA. The study describes the conceivable involvement of gut microbiota in the unclear pathogenesis of AA. We enrolled 25 patients, over 18 years of age with an active state of AA who donated their stool samples. The samples were examined at the human gut microbial community at the species level by metataxonomic analysis of the full-length 16S V3-V4 sequencing. The four major genera that constitute the microbiome's core are Lachnoclostridium, Bifidobacterium, Streptococcus, and Eubacterium, as well as three major phyla: Firmicutes, Proteobacteria, and Actinobacteria. Firmicutes and Proteobacteria are overrepresented in the microflora, which might suggest a disturbed microflora. Furthermore, the composition of bacterial communities suggests a loss of overall richness and a decrease in taxonomic diversity across all samples. This study describes, for the first time, the characteristics of the gut microbiome in AA patients and may provide new insight into the gut microbiome that may play a role in the development of AA.

Human gut Bacteroides uniformis utilizes mixed linked β-glucans via an alternative strategy

Human gut microbial communities have a tremendous impact on well-being of the human health by producing myriad of metabolites. Polysaccharide utilizing capability of these microbial communities is a key driving force in shaping the composition of gut microbiota. Previous studies suggest that genes responsible for β- glucans utilizing in Bacteroides are present in a polysaccharide utilization locus (PUL). However, recent observations show that Bacteroides uniformis lacks such PUL archetypal organization for efficient utilization of pustulan (β1-6) and mixed linkage (β1-4/β1-3) glucans, and its mechanism is yet to be studied. Here, we first used BuGH16 for production of important two mixed linked β- glucan-oligosaccharides, and then demonstrate kinetics of the BuGH3MLG. The B. uniformis JCM13288T coordinates between PUL associated glycoside hydrolase (GH)-16 and distantly localised bespoke GH 3 (BuGH3MLG) for efficient utilization of mixed linkage (β1-4/β1-3) glucans. BuGH3MLG found to be cleaved β1-4, β1-3 and β1-6 glucan-oligosaccharides. Such understanding of glycan utilization mechanism of human gut bacterial communities is crucial for development of nutraceutical therapy and improving health of human being.

Appendectomy Is Associated With Alteration of Human Gut Bacterial and Fungal Communities.

Recent research has revealed the importance of the appendix in regulating the intestinal microbiota and mucosal immunity. However, the changes that occur in human gut microbial communities after appendectomy have never been analyzed. We assessed the alterations in gut bacterial and fungal populations associated with a history of appendectomy. In this cross-sectional study, we investigated the association between appendectomy and the gut microbiome using 16S and ITS2 sequencing on fecal samples from 30 healthy individuals with prior appendectomy (HwA) and 30 healthy individuals without appendectomy (HwoA). Analysis showed that the gut bacterial composition of samples from HwA was less diverse than that of samples from HwoA and had a lower abundance of Roseburia, Barnesiella, Butyricicoccus, Odoribacter, and Butyricimonas species, most of which were short-chain fatty acids-producing microbes. The HwA subgroup analysis indicated a trend toward restoration of the HwoA bacterial microbiome over time after appendectomy. HwA had higher gut fungi composition and diversity than HwoA, even 5 years after appendectomy. Compared with those in samples from HwoA, the abundance correlation networks in samples from HwA displayed more complex fungal–fungal and fungal–bacterial community interactions. This study revealed a marked impact of appendectomy on gut bacteria and fungi, which was particularly durable for fungi.

Breastmilk, Stool, and Meconium: Bacterial Communities in South Africa.

Human milk optimizes gut microbial richness and diversity, and is critical for proper immune development. Research has shown differing microbial composition based on geographic location, providing evidence that diverse biospecimen data is needed when studying human bacterial communities. Yet, limited research describes human milk and infant gut microbial communities in Africa. Our study uses breastmilk, stool, and meconium samples from a South African birth cohort to describe the microbial diversity, identify distinct taxonomic units, and determine correlations between bacterial abundance in breastmilk and stool samples. Mother-infant dyads (N = 20) were identified from a longitudinal birth cohort in the Vhembe district of Limpopo Province, South Africa. Breastmilk, meconium, and stool samples were analyzed using 16S ribosomal RNA sequencing of the V4-V5 gene region using the MiSeq platform for identification and relative quantification of bacterial taxa. A non-metric multidimensional scaling using Bray-Curtis distances of sample Z-scores showed that meconium, stool, and breastmilk microbial communities are distinct with varying genus. Breastmilk was mostly comprised of Streptococcus, Staphylococcus, Veillonella, and Corynebacterium. Stool samples showed the highest levels of Bifidobacterium, Faecalibacterium, Bacteroides, and Streptococcus. Alpha diversity measures found that stool samples have the highest Shannon index score compared to breastmilk and meconium. The abundance of Bifidobacterium (r = 0.57), Blautia (r = 0.59), and Haemophilus (r = 0.69) was correlated (p < 0.1) between breastmilk and stool samples. Despite the importance of breastmilk in seeding the infant gut microbiome, we found evidence of distinct bacterial communities between breastmilk and stool samples from South African mother-infant dyads.

Changes in the Human Gut Microbiota Associated With Colonization by Blastocystis sp. and Entamoeba spp. in Non-Industrialized Populations.

Human gut microbial communities are mainly composed of bacteria, but also include fungi, viruses, archaea, and protozoa, whose role in the gut ecosystem has only recently begun to be recognized. For example, humans colonized by Blastocystis (a gut protozoan with controversial pathogenicity) host a more diverse bacterial microbiota than individuals not carrying it, suggesting that its presence may be beneficial for the host. In parallel, the presence of non-pathogenic Entamoeba spp. has been associated with an increased diversity and compositional shifts in the bacterial microbiota of healthy rural individuals in Cameroon. However, Entamoeba and Blastocystis, the two most prevalent human gut protozoa, have never been studied in the same individuals, preventing the study of their interaction. As Blastocystis is one of the few gut protozoa commonly found in industrialized populations, which are otherwise mostly devoid of gut eukaryotes, we need to focus on rural “traditional” populations, who harbor a higher diversity of gut eukaryotes (whether pathogenic or commensal) in order to study protozoa interactions in the gut ecosystem. To this end, we profiled the gut bacterial microbiota of 134 healthy Cameroonian adults using 16S rRNA gene amplicon sequencing data. Entamoeba and Blastocystis presence and co-occurrence pattern in the same individuals were determined using metagenomic shotgun data. We found that, when taking into account both protozoa jointly, Blastocystis was associated with both a higher richness and a higher evenness of the gut bacterial microbiota, while Entamoeba was associated only with a higher richness. We demonstrated a cumulative influence of these protozoa on bacterial microbiome diversity. Furthermore, while the abundance of several common taxa (for example, Ruminococcaceae, Coprococcus and Butyrivibrio) varied according to Blastocystis colonization, only a single Bacteroides amplicon sequence variant was found to be differentially abundant between Entamoeba-negative and Entamoeba-positive samples. Given the specific signature of each protozoan on the gut microbiota and the seemingly stronger association for Blastocystis, our results suggest that Blastocystis and Entamoeba interact with gut bacteria each in its own way, but experimental studies are needed to explore the precise mechanisms of these interactions.