Isolation of potentially novel species expands the genomic and functional diversity of Lachnospiraceae.

The Lachnospiraceae family holds promise as a source of next-generation probiotics, yet a comprehensive delineation of its diversity is lacking, hampering the identification of suitable strains for future applications. To address this knowledge gap, we conducted an in-depth genomic and functional analysis of 1868 high-quality genomes, combining data from public databases with our new isolates. This data set represented 387 colonization-selective species-level clusters, of which eight genera represented multilineage clusters. Pan-genome analysis, single-nucleotide polymorphism (SNP) identification, and probiotic functional predictions revealed that species taxonomy, habitats, and geography together shape the functional diversity of Lachnospiraceae. Moreover, analyses of associations with atherosclerotic cardiovascular disease (ACVD) and inflammatory bowel disease (IBD) indicated that several strains of potentially novel Lachnospiraceae species possess the capacity to reduce the abundance of opportunistic pathogens, thereby imparting potential health benefits. Our findings shed light on the untapped potential of novel species enabling knowledge-based selection of strains for the development of next-generation probiotics holding promise for improving human health and disease management.

Exploring the Microbiome Landscape of Dental Plaque: A Cross-Sectional Analysis in Periodontal Health and Disease.

OBJECTIVE: This study aimed to comprehensively analyze the microbiome of dental plaque in individuals with varying periodontal statuses, encompassing both periodontal health and disease. The primary objectives were to identify microbial markers associated with different clinical conditions, explore variations in microbial diversity, and investigate potential correlations between the oral microbiome and clinical parameters. METHODS: A cross-sectional design was employed, involving 164 participants aged 18 to 65 years. Inclusion criteria comprised individuals with good oral and systemic health for the periodontal health group and those diagnosed with various stages of periodontal disease for the periodontal disease group. Dental plaque samples were meticulously collected from diverse tooth surfaces, and clinical examinations were conducted to assess periodontal health status. High-throughput sequencing of the 16S ribosomal RNA (rRNA) gene was utilized for microbiome analysis. RESULTS: Demographic characteristics revealed a balanced distribution between the periodontal health and disease groups. Clinical parameters, including probing depth, clinical attachment loss, and bleeding on probing, exhibited significant differences between the two groups (p < 0.001). Microbial diversity indices indicated a higher diversity in the periodontal health group compared to the disease group (p < 0.001). Analysis of relative abundance of bacterial phyla identified significant variations, with Firmicutes, Bacteroidetes, and Actinobacteria showing differential prevalence between health and disease (p < 0.05). Differentially abundant taxa analysis highlighted specific species associated with each clinical condition, including Prevotella intermedia and Porphyromonas gingivalis. Network analysis revealed complex microbial interactions within the oral microbiome. Functional predictions indicated variations in metabolic capabilities between health and disease, with potential implications for virulence and antibiotic resistance. CONCLUSION: This study provides a comprehensive analysis of the oral microbiome in periodontal health and disease, revealing significant associations between microbial composition and clinical parameters. The identification of microbial markers and functional insights enhances our understanding of the complex interplay within the oral ecosystem. These findings hold promise for advancing diagnostic and therapeutic approaches tailored to individual microbial profiles.

Holed up, but thriving: Impact of multitrophic cryoconite communities on glacier elemental cycles.

Cryoconite holes (water and sediment-filled depressions), found on glacier surfaces worldwide, serve as reservoirs of microbes, carbon, trace elements, and nutrients, transferring these components downstream via glacier hydrological networks. Through targeted amplicon sequencing of carbon and nitrogen cycling genes, coupled with functional inference-based methods, we explore the functional diversity of these mini-ecosystems within Antarctica and the Himalayas. These regions showcase distinct environmental gradients and experience varying rates of environmental change influenced by global climatic shifts. Analysis revealed a diverse array of photosynthetic microorganisms, including Stramenopiles, Cyanobacteria, Rhizobiales, Burkholderiales, and photosynthetic purple sulfur Proteobacteria. Functional inference highlighted the high potential for carbohydrate, amino acid, and lipid metabolism in the Himalayan region, where organic carbon concentrations surpassed those in Antarctica by up to 2 orders of magnitude. Nitrogen cycling processes, including fixation, nitrification, and denitrification, are evident, with Antarctic cryoconite exhibiting a pronounced capacity for nitrogen fixation, potentially compensating for the limited nitrate concentrations in this region. Processes associated with the respiration of elemental sulfur and inorganic sulfur compounds such as sulfate, sulfite, thiosulfate, and sulfide suggest the presence of a complete sulfur cycle. The Himalayan region exhibits a higher potential for sulfur cycling, likely due to the abundant sulfate ions and sulfur-bearing minerals in this region. The capability for complete iron cycling through iron oxidation and reduction reactions was also predicted. Methanogenic archaea that produce methane during organic matter decomposition and methanotrophic bacteria that utilize methane as carbon and energy sources co-exist in the cryoconite, suggesting that these niches support the complete cycling of methane. Additionally, the presence of various microfauna suggests the existence of a complex food web. Collectively, these results indicate that cryoconite holes are self-sustaining ecosystems that drive elemental cycles on glaciers and potentially control carbon, nitrogen, sulfur, and iron exports downstream.

Effects of Diet on the Gut Bacterial Community of Aldrichina grahami (Diptera: Calliphoridae) across Developmental Stages.

The blowfly, Aldrichina grahami (Diptera: Calliphoridae), is a well-known forensically important insect. Basic data related to A. grahami have been well documented; but despite the pivotal role of gut microbes in various facets of insect biology, little is known about its gut microbiome. To investigate the gut bacterial community of A. grahami and explore its stability, diet and developmental stage were selected as the two variables. Larvae were reared on bovine liver, swine manure, and chicken manure, and high-throughput sequencing of the 1st, 2nd, and 3rd instar larvae and the newly emerged adults was performed. According to our results, the alpha diversity of the gut bacterial community did not significantly vary among different diets and developmental stages. Principal coordinate analysis revealed that the gut microbiome of A. grahami clustered together among different diets and developmental stages. The main phyla in the gut microbial community of A. grahami were Firmicutes and Proteobacteria, and the dominant genera were Vagococcus, Providencia, Lactobacillus, and Morganella. These findings characterized the gut microbiome of A. grahami and demonstrated that the gut bacterial community is fairly stable. The dominant genera Vagococcus, Providencia, Lactobacillus, and Morganella have the potential to serve as core microbiomes within the gut microbial community of A. grahami.

Exploring the Successions in Microbial Community and Flavor of Daqu during Fermentation Produced by Different Pressing Patterns.

Daqu can be divided into artificially pressed daqu (A-Daqu) and mechanically pressed daqu (M-Daqu) based on pressing patterns. Here, we compared the discrepancies in physicochemical properties, volatile metabolites, and microbiota features between A-Daqu and M-Daqu during fermentation and further investigated the factors causing those differences. A-Daqu microbiota was characterized by six genera (e.g., Bacillus and Thermoactinomyces), while five genera (e.g., Bacillus and Thermomyces) dominated in M-Daqu. The flavor compounds analysis revealed that no obvious difference was observed in the type of esters between the two types of daqu, and M-Daqu was enriched with more alcohols. The factors related to differences between the two types of daqu were five genera (e.g., Hyphopichia). The functional prediction of microbial communities revealed that the functional discrepancies between the two types of daqu were mainly related to ethanol metabolism and 2,3-butanediol metabolism. This study provided a theoretical basis for understanding the heterogeneity of daqu due to the different pressing patterns.

Elevated functional versatility of the soil microbial community associated with the invader Carpobrotus edulis across a broad geographical scale.

Exotic invasive plants may shape their own rhizosphere microbial community during global invasions. Nevertheless, the impacts of such plant invasions on the functional capacities of soil microbial communities remain poorly explored. We used an approach at a broad geographical scale to estimate the composition and abundance of the fungal functional groups, as well as the bacterial metabolic functions, associated with the rhizospheres of Carpobrotus edulis (L.) L. Bolus and the predominant native plants in coastal ecosystems located in different geographical regions. We used the ASV method to infer the potential functions of the soil microbial community with the PICRUSt2 and FUNGuild tools. The predictive functional profiling of the bacterial communities differed between the rhizospheres of the invasive and native plants, regardless of the biogeographic location of the invaded soil. Some predicted pathways related to the biosynthesis of nucleotides such as ppGpp and pppGpp, lipids, carbohydrates and secondary metabolites and the degradation of organic matter were enriched in the C. edulis rhizosphere. Moreover, the invasive microbiota was characterised by a greater richness and diversity of catabolic enzymes involved in nutrients cycling and higher relative abundances of saprotrophs and pathotrophs. Invasion by C. edulis promoted a shift in the potential functional versatility of the soil microbial communities, which can cope with nutrient limitations and biotic stress, and can favour the establishment of the invasive plant, but also alter the functioning and stability of the invaded ecosystems.

Phytase Supplementation Effects on Amino Acid Digestibility in Broiler Chickens are Influenced by Dietary Calcium Concentrations but not by Acid-Binding Capacity.

BACKGROUND: Responses to dietary calcium (Ca) and supplemented phytase on prececal amino acid digestibility (pcAAD) in broiler chickens vary among studies. The variation may arise from the dietary acid-binding capacity (ABC) that influences the activity of enzymes in the digestive tract and from microbial activity. OBJECTIVE: This study aimed to investigate whether the ABC influences phytase effects on pcAAD and whether microbial activity contributes to this. METHODS: Male Ross 308 broiler chickens were provided 1 of 12 diets in 72 pens (15/pen) from day 16 of age until the end of the experiment on days 21 or 22. In a 3 × 2 × 2-factorial arrangement, the ABC was varied by replacing calcium carbonate (CaCO3) with Ca-formate or by adding formic acid to CaCO3-containing diets, and contained 5.6 or 8.2 g Ca/kg and 0 or 1500 phytase units/kg. The ileum content was collected for pcAAD measurement and microbial community composition was used to investigate whether changes in pcAAD are related to the microbiota. RESULTS: Three-factor ANOVA showed that reducing the ABC increased pcAAD (average 1.1 percentage points) and no significant interaction of the ABC with Ca concentration and phytase supplementation including 3-way interactions. Without phytase, increasing dietary Ca concentration decreased pcAAD (average 3.1 percentage points). Phytase supplementation increased pcAAD (average 2.1 and 5.0 percentage points at low and high Ca concentrations, respectively), to reach the same level for both Ca concentrations. Microbial functional predictions pointed towards an influence of the microbiota in the crop and ileum content on amino acid concentrations, as indicated by different relative abundances of predicted genes related to amino acid biosynthesis, degradation, and metabolism. CONCLUSIONS: Dietary Ca concentrations but not the ABC modulates the effect of supplemented phytase on pcAAD in broiler chickens. The microbiota might contribute to differences in pcAAD by changing the amino acid composition of the digesta. The extent of this effect is still unknown.

Colorectal Cancer Stage-Specific Fecal Bacterial Community Fingerprinting of the Taiwanese Population and Underpinning of Potential Taxonomic Biomarkers.

Despite advances in the characterization of colorectal cancer (CRC), it still faces a poor prognosis. There is growing evidence that gut microbiota and their metabolites potentially contribute to the development of CRC. Thus, microbial dysbiosis and their metabolites associated with CRC, based on stool samples, may be used to advantage to provide an excellent opportunity to find possible biomarkers for the screening, early detection, prevention, and treatment of CRC. Using 16S rRNA amplicon sequencing coupled with statistical analysis, this study analyzed the cause-effect shift of the microbial taxa and their metabolites that was associated with the fecal gut microbiota of 17 healthy controls, 21 polyps patients, and 21 cancer patients. The microbial taxonomic shift analysis revealed striking differences among the healthy control, polyps and cancer groups. At the phylum level, Synergistetes was reduced significantly in the polyps group compared to the healthy control and cancer group. Additionally, at the genus level and in association with the cancer group, a total of 12 genera were highly enriched in abundance. In contrast, only Oscillosprira was significantly higher in abundance in the healthy control group. Comparisons of the polyps and cancer groups showed a total of 18 significantly enriched genera. Among them, 78% of the genera associated with the cancer group were in higher abundance, whereas the remaining genera showed a higher abundance in the polyps group. Additionally, the comparison of healthy control and polyp groups showed six significantly abundant genera. More than 66% of these genera showed a reduced abundance in the polyps group than in healthy controls, whereas the remaining genera were highly abundant in the polyps group. Based on tumor presence and absence, the abundance of Olsenella and Lactobacillus at the genus level was significantly reduced in the patient group compared to healthy controls. The significant microbial function prediction revealed an increase in the abundance of metabolites in the polyps and cancer groups compared to healthy controls. A correlation analysis revealed a higher contribution of Dorea in the predicted functions. This study showed dysbiosis of gut microbiota at the taxonomic level and their metabolic functions among healthy subjects and in two stages of colorectal cancer, including adenoma and adenocarcinoma, which might serve as potential biomarkers for the early diagnosis and treatment of CRC.

Bioinformatic analysis and functional predictions of selected regeneration-associated transcripts expressed by zebrafish microglia.

BACKGROUND: Unlike mammals, zebrafish have a remarkable capacity to regenerate a variety of tissues, including central nervous system tissue. The function of macrophages in tissue regeneration is of great interest, as macrophages respond and participate in the landscape of events that occur following tissue injury in all vertebrate species examined. Understanding macrophage populations in regenerating tissue (such as in zebrafish) may inform strategies that aim to regenerate tissue in humans. We recently published an RNA-seq experiment that identified genes enriched in microglia/macrophages in regenerating zebrafish retinas. Interestingly, a small number of transcripts differentially expressed by retinal microglia/macrophages during retinal regeneration did not have predicted orthologs in human or mouse. We reasoned that at least some of these genes could be functionally important for tissue regeneration, but most of these genes have not been studied experimentally and their functions are largely unknown. To reveal their possible functions, we performed a variety of bioinformatic analyses aimed at identifying the presence of functional protein domains as well as orthologous relationships to other species. RESULTS: Our analyses identified putative functional domains in predicted proteins for a number of selected genes. For example, we confidently predict kinase function for one gene, cytokine/chemokine function for another, and carbohydrate enzymatic function for a third. Predicted orthologs were identified for some, but not all, genes in species with described regenerative capacity, and functional domains were consistent with identified orthologs. Comparison to other published gene expression datasets suggest that at least some of these genes could be important in regenerative responses in zebrafish and not necessarily in response to microbial infection. CONCLUSIONS: This work reveals previously undescribed putative function of several genes implicated in regulating tissue regeneration. This will inform future work to experimentally determine the function of these genes in vivo, and how these genes may be involved in microglia/macrophage roles in tissue regeneration.

Agroforestry Management Systems Drive the Composition, Diversity, and Function of Fungal and Bacterial Endophyte Communities in Theobroma Cacao Leaves.

Cacao (Theobroma cacao L.) is one of the most economically important crops worldwide. Despite the important role of endophytes for plant growth and health, very little is known about the effect of agroforestry management systems on the endophyte communities of T. cacao. To close this knowledge gap, we investigated the diversity, community composition, and function of bacterial and fungal endophytes in the leaves of T. cacao trees growing in five major cacao-growing regions in the central region of Cameroon using DNA metabarcoding. Fungal but not bacterial alpha diversity measures differed significantly between the agroforestry management systems. Interestingly, less managed home-garden cacao forests harbored the lowest fungal richness and diversity. Our results suggest that the composition of bacterial and fungal endophyte communities is predominantly affected by agroforestry management systems and, to a lesser extent, by environmental properties. The core microbiome detected comprised important fungal phytopathogens, such as Lasiodiplodia species. Several predicted pathways of bacterial endophytes and functional guilds of fungal endophytes differed between the agroforest systems which might be attributed to bacteria and fungi specifically associated with a single agroforest. Our results provide the basis for future studies on foliar fungal and bacterial endophytes of T. cacao and their responsiveness towards agroforestry management systems.

Tax4Fun2: prediction of habitat-specific functional profiles and functional redundancy based on 16S rRNA gene sequences.

BACKGROUND: Sequencing of 16S rRNA genes has become a powerful technique to study microbial communities and their responses towards changing environmental conditions in various ecosystems. Several tools have been developed for the prediction of functional profiles from 16S rRNA gene sequencing data, because numerous questions in ecosystem ecology require knowledge of community functions in addition to taxonomic composition. However, the accuracy of these tools relies on functional information derived from genomes available in public databases, which are often not representative of the microorganisms present in the studied ecosystem. In addition, there is also a lack of tools to predict functional gene redundancy in microbial communities. RESULTS: To address these challenges, we developed Tax4Fun2, an R package for the prediction of functional profiles and functional gene redundancies of prokaryotic communities from 16S rRNA gene sequences. We demonstrate that functional profiles predicted by Tax4Fun2 are highly correlated to functional profiles derived from metagenomes of the same samples. We further show that Tax4Fun2 has higher accuracies than PICRUSt and Tax4Fun. By incorporating user-defined, habitat-specific genomic information, the accuracy and robustness of predicted functional profiles is substantially enhanced. In addition, functional gene redundancies predicted with Tax4Fun2 are highly correlated to functional gene redundancies determined for simulated microbial communities. CONCLUSIONS: Tax4Fun2 provides researchers with a unique tool to predict and investigate functional profiles of prokaryotic communities based on 16S rRNA gene sequencing data. It is easy-to-use, platform-independent and highly memory-efficient, thus enabling researchers without extensive bioinformatics knowledge or access to high-performance clusters to predict functional profiles. Another unique feature of Tax4Fun2 is that it allows researchers to calculate the redundancy of specific functions, which is a potentially important measure of how resilient a community will be to environmental perturbation. Tax4Fun2 is implemented in R and freely available at https://github.com/bwemheu/Tax4Fun2.

Functional Prediction of Candidate MicroRNAs for CRC Management Using in Silico Approach.

Approximately 30-50% of malignant growths can be prevented by avoiding risk factors and implementing evidence-based strategies. Colorectal cancer (CRC) accounted for the second most common cancer and the third most common cause of cancer death worldwide. This cancer subtype can be reduced by early detection and patients' management. In this study, the functional roles of the identified microRNAs were determined using an in silico pipeline. Five microRNAs identified using an in silico approach alongside their seven target genes from our previous study were used as datasets in this study. Furthermore, the secondary structure and the thermodynamic energies of the microRNAs were revealed by Mfold algorithm. The triplex binding ability of the oligonucleotide with the target promoters were analyzed by Trident. Finally, evolutionary stage-specific somatic events and co-expression analysis of the target genes in CRC were analyzed by SEECancer and GeneMANIA plugin in Cytoscape. Four of the five microRNAs have the potential to form more than one secondary structure. The ranges of the observed/expected ratio of CpG dinucleotides of these genes range from 0.60 to 1.22. Three of the candidate microRNA were capable of forming multiple triplexes along with three of the target mRNAs. Four of the total targets were involved in either early or metastatic stage-specific events while three other genes were either a product of antecedent or subsequent events of the four genes implicated in CRC. The secondary structure of the candidate microRNAs can be used to explain the different degrees of genetic regulation in CRC due to their conformational role to modulate target interaction. Furthermore, due to the regulation of important genes in the CRC pathway and the enrichment of the microRNA with triplex binding sites, they may be a useful diagnostic biomarker for the disease subtype.

Fungiculture in Termites Is Associated with a Mycolytic Gut Bacterial Community.

Termites forage on a range of substrates, and it has been suggested that diet shapes the composition and function of termite gut bacterial communities. Through comparative analyses of gut metagenomes in nine termite species with distinct diets, we characterize bacterial community compositions and use peptide-based functional annotation method to determine biomass-degrading enzymes and the bacterial taxa that encode them. We find that fungus-growing termite guts have relatively more fungal cell wall-degrading enzyme genes, while wood-feeding termite gut communities have relatively more plant cell wall-degrading enzyme genes. Interestingly, wood-feeding termite gut bacterial genes code for abundant chitinolytic enzymes, suggesting that fungal biomass within the decaying wood likely contributes to gut bacterial or termite host nutrition. Across diets, the dominant biomass-degrading enzymes are predominantly coded for by the most abundant bacterial taxa, suggesting tight links between diet and gut community composition, with the most marked difference being the communities coding for the mycolytic capacity of the fungus-growing termite gut.IMPORTANCE Understanding functional capacities of gut microbiomes is important to improve our understanding of symbiotic associations. Here, we use peptide-based functional annotation to show that the gut microbiomes of fungus-farming termites code for a wealth of enzymes that likely target the fungal diet the termites eat. Comparisons to other termites showed that fungus-growing termite guts have relatively more fungal cell wall-degrading enzyme genes, whereas wood-feeding termite gut communities have relatively more plant cell wall-degrading enzyme genes. Across termites with different diets, the dominant biomass-degrading enzymes are predominantly coded for by the most abundant bacterial taxa, suggesting tight links between diet and gut community compositions.

Annotation of Variant Data from High-Throughput DNA Sequencing from Tumor Specimens: Filtering Strategies to Identify Driver Mutations.

The use of next-generation sequencing technologies has enabled the analysis of a wide spectrum of somatic mutations in tumors. This analysis can be carried out using various strategies including the use of small panels of focused, clinically actionable genes, large panels of cancer-related genes, whole exomes, and the entire genome. One of the main goals in these analyses is to identify key mutations in these tumors that drive the oncogenic process. Depending on the gene, mutations can have altering effects, such as loss of function mutations in tumor suppressor genes, to mutations that activate genes such as kinases involved with cell cycle progression or proliferation. Once the sequencing process is complete, and the alignment of the large collection of reads to the reference genome and variant calling has been carried out, one is left with a large collection of variants. The challenge then becomes assigning where the variant resides in the genome with respect to coding regions, splice site regions, regulatory regions, and what potential functional effect these variants may have on the resulting protein. Other helpful information includes determining if the variant has been identified before, and if so, the tumor type associated with the variant. In addition, if the tumor profiling experiment is not conducted with a matched specimen representing the inherited genome, various tools are helpful to determine if the variant is likely to be an inherited polymorphism or a somatic event. In this chapter, we review the various tools available for annotating variants to assist in filtering down and prioritizing the hundreds to thousands of variants down to the key variants likely to be driver mutations and relevant to the tumor being profiled.

Linking Compositional and Functional Predictions to Decipher the Biogeochemical Significance in DFAA Turnover of Abundant Bacterioplankton Lineages in the North Sea.

Deciphering the ecological traits of abundant marine bacteria is a major challenge in marine microbial ecology. In the current study, we linked compositional and functional predictions to elucidate such traits for abundant bacterioplankton lineages in the North Sea. For this purpose, we investigated entire and active bacterioplankton composition along a transect ranging from the German Bight to the northern North Sea by pyrotag sequencing of bacterial 16S rRNA genes and transcripts. Functional profiles were inferred from 16S rRNA data using Tax4Fun. Bacterioplankton communities were dominated by well-known marine lineages including clusters/genera that are affiliated with the Roseobacter group and the Flavobacteria. Variations in community composition and function were significantly explained by measured environmental and microbial properties. Turnover of dissolved free amino acids (DFAA) showed the strongest correlation to community composition and function. We applied multinomial models, which enabled us to identify bacterial lineages involved in DFAA turnover. For instance, the genus Planktomarina was more abundant at higher DFAA turnover rates, suggesting its vital role in amino acid degradation. Functional predictions further indicated that Planktomarina is involved in leucine and isoleucine degradation. Overall, our results provide novel insights into the biogeochemical significance of abundant bacterioplankton lineages in the North Sea.

Composition and Functional Specialists of the Gut Microbiota of Frogs Reflect Habitat Differences and Agricultural Activity.

The physiological impact of agricultural pollution, habitat disturbance, and food source variability on amphibian remains poorly understood. By comparing the composition and predicted functions of gut microbiota of two frog species from forest and farmland, we quantified the effects of the exogenous environment and endogenous filters on gut microbiota and the corresponding functions. However, compositional differences of the gut microbiota between the frog species were not detected, even when removing roughly 80-88% of the confounding effect produced by common and shared bacteria (i.e., generalists) and those taxa deemed too rare. The habitat effect accounted for 14.1% of the compositional difference of gut microbial specialists, but host and host × habitat effects were not significant. Similar trends of a significant habitat effect, at an even higher level (26.0%), for the physiological and metabolic functions of gut microbiota was predicted. A very obvious skewing of the relative abundance of functional groups toward farmland habitats reflects the highly diverse bacterial functions of farmland frogs, in particular related to pathogenic disease and pesticide degradation, which may be indication of poor adaptation or strong selective pressure against disease. These patterns reflect the impacts of agricultural activities on frogs and how such stresses may be applied in an unequal manner for different frog species.