Entire Microbial Community Research Articles

Conversion of natural grassland to cropland alters microbial community assembly across northern China.

To feed the growing human population, natural grasslands are being converted to agricultural use at a massive scale. This conversion may have negative consequences for soil biodiversity, but its impact on the community assembly of differentially microbial groups remains largely unknown. Here, we investigated the diversity and community compositions of bacteria, archaea, fungi and protists, using a paired sampling of grassland and cropland soils across the agro-pastoral ecotone of northern China. Land-use conversion decreased α diversity of bacteria, fungi and protists, and altered the structures of the entire soil microbial community (archaea, bacteria, fungi and protists). The community assembly of archaea and bacteria was dominated by stochastic processes, and that of protists dominated by deterministic processes in both land-use types. By contrast, the fungal community was governed more strongly by stochastic processes in grassland soil, than by deterministic processes in cropland soil. Our findings support the 'size-plasticity' hypothesis that smaller body-sized microorganisms (archaea and bacteria) are more structured by stochastic processes, and larger one (protist) is more influenced by deterministic processes. Our study demonstrates that distinct ecological processes govern microbial community assembly, and land-use change regulates the balance between determinism and stochasticity.

Metagenomics: A Tool for Exploring Key Microbiome With the Potentials for Improving Sustainable Agriculture

Microorganisms are immense in nature and exist in every imaginable ecological niche, performing a wide range of metabolic processes. Unfortunately, using traditional microbiological methods, most microorganisms remain unculturable. The emergence of metagenomics has resolved the challenge of capturing the entire microbial community in an environmental sample by enabling the analysis of whole genomes without requiring culturing. Metagenomics as a non-culture approach encompasses a greater amount of genetic information than traditional approaches. The plant root-associated microbial community is essential for plant growth and development, hence the interactions between microorganisms, soil, and plants is essential to understand and improve crop yields in rural and urban agriculture. Although some of these microorganisms are currently unculturable in the laboratory, metagenomic techniques may nevertheless be used to identify the microorganisms and their functional traits. A detailed understanding of these organisms and their interactions should facilitate an improvement of plant growth and sustainable crop production in soil and soilless agriculture. Therefore, the objective of this review is to provide insights into metagenomic techniques to study plant root-associated microbiota and microbial ecology. In addition, the different DNA-based techniques and their role in elaborating plant microbiomes are discussed. As an understanding of these microorganisms and their biotechnological potentials are unlocked through metagenomics, they can be used to develop new, useful and unique bio-fertilizers and bio-pesticides that are not harmful to the environment.

Milk: A Scientific Model for Diet and Health Research in the 21st Century

The origin of lactation and the composition, structures and functions of milk's biopolymers highlight the Darwinian pressure on lactation as a complete, nourishing and protective diet. Lactation, under the driving pressure to be a sustainable bioreactor, was under selection pressure of its biopolymers with diverse functions acting from the mammary gland through the digestive system of the infant. For example, milk is extensively glycosylated and the glycan structures and their functions are now emerging. Milk contains free oligosaccharides; complex polymers of sugars whose stereospecific linkages are not matched by glycosidic enzymes within the mammalian infant gut. These glycan polymers reach the lower intestine undigested. In this microbe-rich environment, bacteria compete to release and ferment the sugars via different hydrolytic strategies. One specific type of bacteria, Bifidobacterium longum subsp. infantis, (B. infantis) is uniquely equipped with a repertoire of genes encoding enzymes capable of taking up, hydrolyzing and metabolizing the complex glycans of human milk. This combination of a distinct food supply and unique genetic capability shapes the composition and metabolic products of the entire microbial community within the lower intestine of breast fed infants. The intestinal microbiome dominated by B. infantis, shields the infant from the growth of gram negative enteropathogens and their endotoxins as a clear health benefit. The world is facing unprecedented challenges to produce a food supply that is both nourishing, safe and sustainable. Scientists need to guide the future of agriculture and food in response to these 21st century challenges. Lactation provides an inspiring model of what that future research strategy could be.

Identification of lipolytic enzymes using high-throughput single-cell screening and sorting of a metagenomic library

The demand for novel, robust microbial biocatalysts for use in industrial and pharmaceutical applications continues to increase rapidly. As a result, there is a need to develop advanced tools and technologies to exploit the vast metabolic potential of unculturable microorganisms found in various environments. Single-cell and functional metagenomics studies can explore the enzymatic potential of entire microbial communities in a given environment without the need to culture the microorganisms. This approach has contributed substantially to the discovery of unique microbial genes for industrial and medical applications. Functional metagenomics involves the extraction of microbial DNA directly from environmental samples, constructing expression libraries comprising the entire microbial genome, and screening of the libraries for the presence of desired phenotypes. In this study, lipolytic enzymes from the Red Sea were targeted. A high-throughput single-cell microfluidic platform combined with a laser-based fluorescent screening bioassay was employed to discover new genes encoding lipolytic enzymes. Analysis of the metagenomic library led to the identification of three microbial genes encoding lipases based on their functional similarity and sequence homology to known lipases. The results demonstrated that microfluidics is a robust technology that can be used for screening in functional metagenomics. The results also indicate that the Red Sea is a promising, under-investigated source of new genes and gene products.

Dynamic Shifts in the Root Microbiota of Cultivated Paphiopedilum armeniacum during Different Stages of Growth

Paphiopedilum armeniacum S. C. Chen et F. Y. Liu is an endangered lady’s slipper orchid species with high horticultural value. As observed for other orchids, mycorrhizal fungi and endophytic bacteria play important roles in the growth and development of P. armeniacum. In the present study, the community structure dynamics across three growth and development stages of cultivated P. armeniacum were investigated. The potential interactions between Tulasnellaceae fungi and core bacterial genera on one hand and the stability of the presumed mycorrhizal fungi communities on the other were analyzed in three growth stages of P. armeniacum to enhance our understanding of endophytic microbial community structure dynamics in the roots at different development stages. Based on sequencing, 3 and 16 phyla and 59 and 269 genera were identified in the fungal and bacterial communities, respectively. The predominant fungi and bacteria were Basidiomycota (62.90%) and Proteobacteria (43.98%), which exhibited changes in abundance and diversity depending on the growth stage of P. armeniacum. Assessment of the entire microbial communities from different growth stages showed that the seedling stage had the highest richness and diversity. The microbial communities recruited by P. armeniacum at the seedling stage were different from those recruited at the vegetative and reproductive growth stages, and the microbial communities recruited in the latter two stages overlapped. Tulasnellaceae were the only dominant fungal symbionts during P. armeniacum growth. Brevibacillus, Mycobacterium, and Sphingomonas, the three core genera, showed significant interactions with the main OTUs of Tulasnellaceae. Putative mycorrhizal fungi in P. armeniacum were relatively stable across different growth environments, and the core mycorrhizal fungi were uncultured Tulasnellaceae (OTU1). This could facilitate the ex situ conservation and commercial development of the endangered orchid.

Recently Evolved Francisella-Like Endosymbiont Outcompetes an Ancient and Evolutionarily Associated Coxiella-Like Endosymbiont in the Lone Star Tick (Amblyomma americanum) Linked to the Alpha-Gal Syndrome.

BackgroundTicks are hematophagous arthropods that transmit various bacterial, viral, and protozoan pathogens of public health significance. The lone star tick (Amblyomma americanum) is an aggressive human-biting tick that transmits bacterial and viral pathogens, and its bites are suspected of eliciting the alpha-gal syndrome, a newly emerged delayed hypersensitivity following consumption of red meat in the United States. While ongoing studies have attempted to investigate the contribution of different tick-inherent factors to the induction of alpha-gal syndrome, an otherwise understudied aspect is the contribution of the tick microbiome and specifically obligate endosymbionts to the establishment of the alpha-gal syndrome in humans.Materials and MethodsHere we utilized a high-throughput metagenomic sequencing approach to cataloging the entire microbial communities residing within different developmental stages and tissues of unfed and blood-fed ticks from laboratory-maintained ticks and three new geographical locations in the United States. The Quantitative Insights Into Microbial Ecology (QIIME2) pipeline was used to perform data analysis and taxonomic classification. Moreover, using a SparCC (Sparse Correlations for Compositional data) network construction model, we investigated potential interactions between members of the microbial communities from laboratory-maintained and field-collected ticks.ResultsOverall, Francisellaceae was the most dominant bacteria identified in the microbiome of both laboratory-raised and field-collected Am. americanum across all tissues and developmental stages. Likewise, microbial diversity was seen to be significantly higher in field-collected ticks compared with laboratory-maintained ticks as seen with a higher number of both Operational Taxonomic Units and measures of species richness. Several potential positive and negative correlations were identified from our network analysis. We observed a strong positive correlation between Francisellaceae, Rickettsiaceae, and Midichloriaceae in both developmental stages and tissues from laboratory-maintained ticks, whereas ovarian tissues had a strong positive correlation of bacteria in the family Xanthobacteraceae and Rhizobiaceae. A negative interaction was observed between Coxiellaceae and Francisellaceae in Illinois, and all the bacteria detected from ticks from Delaware were negatively correlated.ConclusionThis study is the first to catalog the microbiome of Am. americanum throughout its developmental stages and different tissue niches and report the potential replacement of Coxiellaceae by Francisellaceae across developmental stages and tissues tested except in ovarian tissues. These unique and significant findings advance our knowledge and open a new avenue of research to further understand the role of tick microbiome in tick-borne diseases and develop a holistic strategy to control alpha-gal syndrome.

Interspecies microbiome transplantation recapitulates microbial acquisition in mosquitoes

BackgroundMosquitoes harbor microbial communities that play important roles in their growth, survival, reproduction, and ability to transmit human pathogens. Microbiome transplantation approaches are often used to study host-microbe interactions and identify microbial taxa and assemblages associated with health or disease. However, no such approaches have been developed to manipulate the microbiota of mosquitoes.ResultsHere, we developed an approach to transfer entire microbial communities between mosquito cohorts. We undertook transfers between (Culex quinquefasciatus to Aedes aegypti) and within (Ae. aegypti to Ae. aegypti) species to validate the approach and determine the number of mosquitoes required to prepare donor microbiota. After the transfer, we monitored mosquito development and microbiota dynamics throughout the life cycle. Typical holometabolous lifestyle-related microbiota structures were observed, with higher dynamics of microbial structures in larval stages, including the larval water, and less diversity in adults. Microbiota diversity in recipient adults was also more similar to the microbiota diversity in donor adults.ConclusionsThis study provides the first evidence for successful microbiome transplantation in mosquitoes. Our results highlight the value of such methods for studying mosquito-microbe interactions and lay the foundation for future studies to elucidate the factors underlying microbiota acquisition, assembly, and function in mosquitoes under controlled conditions.1rc3aWT8PxAqCNbXPTfbB8Video

Metagenomic insights to the functional potential of sediment microbial communities in freshwater lakes

Molecular-based techniques offer considerable potential to provide new insights into the impact of anthropogenic stressors on lake ecosystems. Microbial communities are involved in many geochemical cycling processes in lakes and a greater understanding of their functions could assist in guiding more targeted remedial actions. Recent advances in metagenomics now make it possible to determine the functional potential of entire microbial communities. The present study investigated microbial communities and their functional potential in surface sediments collected from three lakes with differing trophic states and characteristics. Surface sediments were analysed for their nutrient and elemental contents and metagenomics and metabarcoding analysis undertaken. The nutrients content of the surface sediments did not show as distinct a gradient as water chemistry monitoring data, likely reflecting effects of other lake characteristics, in particular, depth. Metabarcoding and metagenomics revealed differing bacterial community composition and functional potential amongst lakes. Amongst the differentially abundant metabolic pathways, the most prominent were clusters in the energy and xenobiotics pathways. Differences in the energy metabolism paths of photosynthesis and oxidative phosphorylation were observed. These were most likely related to changes in the community composition and especially the presence of cyanobacteria in two of the three lakes. Xenobiotic pathways, such as those involving polycyclic aromatic hydrocarbons, were highest in the lakes with the greatest agricultural land-use in their catchment. These results highlight how microbial metagenomics can be used to gain insights into the causes of differences in trophic status amongst lakes.

Changes in Microbiome Activity and Sporadic Viral Infection Help Explain Observed Variability in Microcosm Studies.

The environmental conditions experienced by microbial communities are rarely fully simulated in the laboratory. Researchers use experimental containers (“bottles”), where natural samples can be manipulated and evaluated. However, container-based methods are subject to “bottle effects”: changes that occur when enclosing the plankton community that are often times unexplained by standard measures like pigment and nutrient concentrations. We noted variability in a short-term, nutrient amendment experiment during a 2019 Lake Erie, Microcystis spp. bloom. We observed changes in heterotrophic bacteria activity (transcription) on a time-frame consistent with a response to experimental changes in nutrient availability, demonstrating how the often overlooked microbiome of cyanobacterial blooms can be altered. Samples processed at the time of collection (T0) contained abundant transcripts from Bacteroidetes, which reduced in abundance during incubation in all bottles, including controls. Significant biological variability in the expression of Microcystis-infecting phage was observed between replicates, with phosphate-amended treatments showing a 10-fold variation. The expression patterns of Microcystis-infecting phage were significantly correlated with ∼35% of Microcystis-specific functional genes and ∼45% of the cellular-metabolites measured across the entire microbial community, suggesting phage activity not only influenced Microcystis dynamics, but the biochemistry of the microbiome. Our observations demonstrate how natural heterogeneity among replicates can be harnessed to provide further insight on virus and host ecology.

Accurate prediction of metagenome-assembled genome completeness by MAGISTA, a random forest model built on alignment-free intra-bin statistics

BackgroundAlthough the total number of microbial taxa on Earth is under debate, it is clear that only a small fraction of these has been cultivated and validly named. Evidently, the inability to culture most bacteria outside of very specific conditions severely limits their characterization and further studies. In the last decade, a major part of the solution to this problem has been the use of metagenome sequencing, whereby the DNA of an entire microbial community is sequenced, followed by the in silico reconstruction of genomes of its novel component species. The large discrepancy between the number of sequenced type strain genomes (around 12,000) and total microbial diversity (106–1012 species) directs these efforts to de novo assembly and binning. Unfortunately, these steps are error-prone and as such, the results have to be intensely scrutinized to avoid publishing incomplete and low-quality genomes.ResultsWe developed MAGISTA (metagenome-assembled genome intra-bin statistics assessment), a novel approach to assess metagenome-assembled genome quality that tackles some of the often-neglected drawbacks of current reference gene-based methods. MAGISTA is based on alignment-free distance distributions between contig fragments within metagenomic bins, rather than a set of reference genes. For proper training, a highly complex genomic DNA mock community was needed and constructed by pooling genomic DNA of 227 bacterial strains, specifically selected to obtain a wide variety representing the major phylogenetic lineages of cultivable bacteria.ConclusionsMAGISTA achieved a 20% reduction in root-mean-square error in comparison to the marker gene approach when tested on publicly available mock metagenomes. Furthermore, our highly complex genomic DNA mock community is a very valuable tool for benchmarking (new) metagenome analysis methods.

Metagenomics Approaches for Improving Food Safety: A Review

Advancements in next-generation sequencing technology have dramatically reduced the cost and increased the ease of microbial whole genome sequencing. This approach is revolutionizing the identification and analysis of foodborne microbial pathogens, facilitating expedited detection and mitigation of foodborne outbreaks, improving public health outcomes, and limiting costly recalls. However, next-generation sequencing is still anchored in the traditional laboratory practice of the selection and culture of a single isolate. Metagenomic-based approaches, including metabarcoding and shotgun and long-read metagenomics, are part of the next disruptive revolution in food safety diagnostics and offer the potential to directly identify entire microbial communities in a single food, ingredient, or environmental sample. In this review, metagenomic-based approaches are introduced and placed within the context of conventional detection and diagnostic techniques, and essential considerations for undertaking metagenomic assays and data analysis are described. Recent applications of the use of metagenomics for food safety are discussed alongside current limitations and knowledge gaps and new opportunities arising from the use of this technology.

Combining tangential flow filtration and size fractionation of mesocosm water as a method for the investigation of waterborne coral diseases.

The causative agents of most coral diseases today remain unknown, complicating disease response and restoration efforts. Pathogen identifications can be hampered by complex microbial communities naturally associated with corals and seawater, which create complicating "background noise" that can potentially obscure a pathogen's signal. Here, we outline an approach to investigate waterborne coral diseases that use a combination of coral mesocosms, tangential flow filtration, and size fractionation to reduce the impact of this background microbial diversity, compensate for unknown infectious dose, and further narrow the suspect pool of potential pathogens. As proof of concept, we use this method to compare the bacterial communities shed into six Montastraea cavernosa coral mesocosms and demonstrate this method effectively detects differences between diseased and healthy coral colonies. We found several amplicon sequence variants (ASVs) in the diseased mesocosms that represented 100% matches with ASVs identified in prior studies of diseased coral tissue, further illustrating the effectiveness of our approach. Our described method is an effective alternative to using coral tissue or mucus to investigate waterborne coral diseases of unknown etiology and can help more quickly narrow the pool of possible pathogens to better aid in disease response efforts. Additionally, this versatile method can be easily adapted to characterize either the entire microbial community associated with a coral or target-specific microbial groups, making it a beneficial approach regardless of whether a causative agent is suspected or is completely unknown.

Potential Roles of the Free Salivary Microbiome Dysbiosis in Periodontal Diseases.

Saliva is a vital mediator in the oral cavity. The dysbiosis of free bacteria in saliva might be related to the onset, development, prognosis, and recurrence of periodontal diseases, but this potential relationship is still unclear. The objective of this study was to investigate the potential roles of the free salivary microbiome in different periodontal statuses, their reaction to nonsurgical periodontal therapy, and differences between diseased individuals after treatment and healthy persons. We recruited 15 healthy individuals, 15 individuals with gingivitis, and 15 individuals with stage I/II generalized periodontitis. A total of 90 unstimulated whole saliva samples were collected and sequenced using full-length bacterial 16S rRNA gene sequencing. We found that as the severity of disease increased, from healthy to gingivitis and periodontitis, the degree of dysbiosis also increased. A higher abundance of Prevotella intermedia and Catonella morbi and a lower abundance of Porphyromonas pasteri, Prevotella nanceiensis, and Haemophilus parainfluenzae might be biomarkers of periodontitis, with an area under curve (AUC) reaching 0.9733. When patients received supragingival scaling, there were more pathogens related to recolonization in the saliva of periodontitis patients than in healthy persons. Even after effective nonsurgical periodontal therapy, individuals with periodontitis displayed a more dysbiotic and pathogenic microbial community in their saliva than healthy individuals. Therefore, the gradual transition in the entire salivary microbial community from healthy to diseased includes a gradual shift to dysbiosis. Free salivary pathogens might play an important role in the recolonization of bacteria as well as the prognosis and recurrence of periodontal diseases.

Soil microbial community coalescence and fertilization interact to drive the functioning of the legume–rhizobium symbiosis

Abstract Soil microbial community coalescence, where entire microbial communities mix and interact under new conditions, is a widespread phenomenon whose applicability for targeted root microbiome assembly has not been studied. Whether soil mixing can lead to predictable outcomes for community assembly and functioning of specific functional groups, for example, N2‐fixing rhizobia, remains unknown. Using a legume shrub adapted to nutrient‐poor soils, we tested the effects of community coalescence on plant nutrition and growth, and its influence on the rhizobial root nodule symbiosis. We grew seedlings of rooibos [Aspalathus linearis (Burm.f.) Dahlg.] in individual rhizosphere soils collected from cultivated and wild rooibos plants and their mixtures, and included a fertilization treatment. Portions of the taxonomic gyrB and the symbiotic nodA gene makers were sequenced to characterize rhizobial communities present in rooibos root nodules under the different soil conditions. Overall, community coalescence by soil mixing had positive effects on plant nutrition and growth, and interacted with fertilizer addition to concurrently change rhizobial taxonomic evenness and promote higher relative N2 fixation. We identified particular rhizobia preferentially associated with rooibos plants that showed higher N fractions from N2 fixation, raised in mixed fertilized soils. These findings indicate that soil bacterial community coalescence and fertilization can have synergistic effects on plant performance, while promoting the assembly of alternative symbiotic rhizobial communities that provide improved nutritional benefits to host plants. Synthesis and applications. The combination of soil mixing and fertilizer addition may be an important, but hitherto overlooked measure to improve the functioning of rhizobium symbioses in legume crops. Microbial community coalescence should gain recognition as a potentially effective mechanism to improve the functioning of plant microbiomes.

The rice foot rot pathogen Dickeya zeae alters the in-field plant microbiome.

SummaryStudies on bacterial plant diseases have thus far been focused on the single bacterial species causing the disease, with very little attention given to the many other microorganisms present in the microbiome. This study intends to use pathobiome analysis of the rice foot rot disease, caused by Dickeya zeae, as a case study to investigate the effects of this bacterial pathogen to the total resident microbiome and to highlight possible interactions between the pathogen and the members of the community involved in the disease process. The microbiome of asymptomatic and the pathobiome of foot‐rot symptomatic field‐grown rice plants over two growing periods and belonging to two rice cultivars were determined via 16S rRNA gene amplicon sequencing. Results showed that the presence of D. zeae is associated with an alteration of the resident bacterial community in terms of species composition, abundance and richness, leading to the formation of microbial consortia linked to the disease state. Several bacterial species were significantly co‐presented with the pathogen in the two growing periods suggesting that they could be involved in the disease process. Besides, culture‐dependent isolation and in planta inoculation studies of a bacterial member of the pathobiome, identified as positive correlated with the pathogen in our in silico analysis, indicated that it benefits from the presence of D. zeae. A similar microbiome/pathobiome experiment was also performed in a symptomatically different rice disease evidencing that not all plant diseases have the same consequence/relationship with the plant microbiome. This study moves away from a pathogen‐focused stance and goes towards a more ecological perception considering the effect of the entire microbial community which could be involved in the pathogenesis, persistence, transmission and evolution of plant pathogens.

Metagenomic Analysis of Fecal Archaea, Bacteria, Eukaryota, and Virus in Przewalski's Horses Following Anthelmintic Treatment.

Intestinal microbiota is involved in immune response and metabolism of the host. The frequent use of anthelmintic compounds for parasite expulsion causes disturbance to the equine intestinal microbiota. However, most studies were on the effects of such treatment on the intestinal bacterial microbes; none is on the entire microbial community including archaea and eukaryotic and viral community in equine animals. This study is the first to explore the differences of the microbial community composition and structure in Przewalski's horses prior to and following anthelmintic treatment, and to determine the corresponding changes of their functional attributes based on metagenomic sequencing. Results showed that in archaea, the methanogen of Euryarchaeota was the dominant phylum. Under this phylum, anthelmintic treatment increased the Methanobrevibacter genus and decreased the Methanocorpusculum genus and two other dominant archaea species, Methanocorpusculum labreanum and Methanocorpusculum bavaricum. In bacteria, Firmicutes and Bacteroidetes were the dominant phyla. Anthelmintic treatment increased the genera of Clostridium and Eubacterium and decreased those of Bacteroides and Prevotella and dominant bacteria species. These altered genera were associated with immunity and digestion. In eukaryota, anthelmintic treatment also changed the genera related to digestion and substantially decreased the relative abundances of identified species. In virus, anthelmintic treatment increased the genus of unclassified_d__Viruses and decreased those of unclassified_f__Siphoviridae and unclassified_f__Myoviridae. Most of the identified viral species were classified into phage, which were more sensitive to anthelmintic treatment than other viruses. Furthermore, anthelmintic treatment was found to increase the number of pathogens related to some clinical diseases in horses. The COG and KEGG function analysis showed that the intestinal microbiota of Przewalski's horse mainly participated in the carbohydrate and amino acid metabolism. The anthelmintic treatment did not change their overall function; however, it displaced the population of the functional microbes involved in each function or pathway. These results provide a complete view on the changes caused by anthelmintic treatment in the intestinal microbiota of the Przewalski's horses.

Microorganisms: crucial players of smokeless tobacco for several health attributes.

Global consumption of smokeless tobacco (SLT) reached 300 million users worldwide majorly from middle-income countries. More than 4000 chemical compounds represent it as one of the noxious consumable products by humans. Besides toxicants/carcinogens, the heavy microbial load on smokeless tobacco further keeps human health at higher risk. Several of these inhabitant microbes participate in biofilm formation and secrete endotoxin/mycotoxins and proinflammatory-like molecules, leading to several oral diseases. Tobacco-associated bacteria exhibit their role in tobacco-specific nitrosamines (TSNAs) formation and acetaldehyde production; both are well-documented carcinogens. Moreover, tobacco exhibits the potential to alter the oral microbiome and induce dysbiotic conditions that lead to the onset of several oral and systemic diseases. Traditional cultivation approaches of microbiology provide partial information of microbial communities of a habitat; therefore, microbiomics has now been employed to study the metagenomes of entire microbial communities. In the past 5 years, few NGS-based investigations have revealed that SLT harbors four dominant phyla (Firmicutes, Proteobacteria, Actinobacteria, and Bacteroidetes) dominating Bacillus spp. and/or Pseudomonas spp. However, functional characterization of their genetic elements will be a more informative attribute to understand the correlation between inhabitant microbial diversity and their relatedness concerning abundance and diseases. This review provides an update on the microbial diversity of SLT and its associated attributes in human health. KEY POINTS: • Heavy microbial load on smokeless tobacco alarms for poor oral hygiene. • Inhabitant microorganisms of SLT participate in TSNA and biofilm formation. • SLTs alter the oral microbiome and causes oral dysbiosis.

Simple mapping-based quantification of a mock microbial community using total RNA-seq data.

Most microbes in the natural environment are difficult to cultivate. Thus, culture-independent analysis for microbial community structure is important for the understanding of its ecological functions. An immense ribosomal RNA sequence collection is available from phylogenetic research on organisms in all domains. These sequences are available for use in genetic research. However, the amplicon-seq process using PCR requires the construction of a sequence library. Construction can introduce bias into quantitative analyses, and each domain of species needs its own primer set. Total RNA sequencing has the advantage of analyzing an entire microbial community, including bacteria, archea, and eukaryote, at once. Such analysis yields large amounts of ribosomal RNA sequences that can be used for analysis without PCR bias. Evaluation using total RNA-seq for quantitative analysis of microbial communities and comparison with amplicon-seq is still rare. In the present study, we developed a mapping-based total RNA-seq analysis to obtain quantitative information on microbial community structure and compared our results with ordinary amplicon-seq methods. We read total RNA sequences from a commercially available mock community (ATCC MSA-2003) and divided reads into small subunit ribosomal RNA (ssrRNA) origin reads and others, such as mRNA origin reads. We then mapped ssrRNA origin reads on annotated assembled contigs and obtained quantitative results under several analysis strategies. Removal of low complexity sequences, sorting ssrRNA with paired-in mode, and performing homology-based taxonomical assignments (BLAST+ or vsearch) showed superior outcomes to other strategies. Results with this approach showed a median relative abundance among ten mock community members of ~10%; ordinary amplicon-seq showed a much lower percentage. Thus, total RNA-seq can be a powerful tool for analyzing microbial community structure and is not limited to analyzing gene expression profiling of microbiomes.

Joint contributions of the gut microbiota and host genetics to feed efficiency in chickens

BackgroundFeed contributes most to livestock production costs. Improving feed efficiency is crucial to increase profitability and sustainability for animal production. Host genetics and the gut microbiota can both influence the host phenotype. However, the association between the gut microbiota and host genetics and their joint contribution to feed efficiency in chickens is largely unclear.ResultsHere, we examined microbial data from the duodenum, jejunum, ileum, cecum, and feces in 206 chickens and their host genotypes and confirmed that the microbial phenotypes and co-occurrence networks exhibited dramatic spatial heterogeneity along the digestive tract. The correlations between host genetic kinship and gut microbial similarities within different sampling sites were weak, with coefficients ranging from − 0.07 to 0.08. However, microbial genome-wide analysis revealed that genetic markers near or inside the genes MTHFD1L and LARGE1 were associated with the abundances of cecal Megasphaera and Parabacteroides, respectively. The effect of host genetics on residual feed intake (RFI) was 39%. We further identified three independent genetic variations that were related to feed efficiency and had a modest effect on the gut microbiota. The contributions of the gut microbiota from the different parts of the intestinal tract on RFI were distinct. The cecal microbiota accounted for 28% of the RFI variance, a value higher than that explained by the duodenal, jejunal, ileal, and fecal microbiota. Additionally, six bacteria exhibited significant associations with RFI. Specifically, lower abundances of duodenal Akkermansia muciniphila and cecal Parabacteroides and higher abundances of cecal Lactobacillus, Corynebacterium, Coprobacillus, and Slackia were related to better feed efficiency.ConclusionsOur findings solidified the notion that both host genetics and the gut microbiota, especially the cecal microbiota, can drive the variation in feed efficiency. Although host genetics has a limited effect on the entire microbial community, a small fraction of gut microorganisms tends to interact with host genes, jointly contributing to feed efficiency. Therefore, the gut microbiota and host genetic variations can be simultaneously targeted by favoring more-efficient taxa and selective breeding to improve feed efficiency in chickens.EooUS9Np68nMFAp4xE4K1kVideo abstract

Evaluation of acidogenesis products\u2019 effect on biogas production performed with metagenomics and isotopic approaches

BackgroundDuring the acetogenic step of anaerobic digestion, the products of acidogenesis are oxidized to substrates for methanogenesis: hydrogen, carbon dioxide and acetate. Acetogenesis and methanogenesis are highly interconnected processes due to the syntrophic associations between acetogenic bacteria and hydrogenotrophic methanogens, allowing the whole process to become thermodynamically favorable. The aim of this study is to determine the influence of the dominant acidic products on the metabolic pathways of methane formation and to find a core microbiome and substrate-specific species in a mixed biogas-producing system.ResultsFour methane-producing microbial communities were fed with artificial media having one dominant component, respectively, lactate, butyrate, propionate and acetate, for 896 days in 3.5-L Up-flow Anaerobic Sludge Blanket (UASB) bioreactors. All the microbial communities showed moderately different methane production and utilization of the substrates. Analyses of stable carbon isotope composition of the fermentation gas and the substrates showed differences in average values of δ13C(CH4) and δ13C(CO2) revealing that acetate and lactate strongly favored the acetotrophic pathway, while butyrate and propionate favored the hydrogenotrophic pathway of methane formation. Genome-centric metagenomic analysis recovered 234 Metagenome Assembled Genomes (MAGs), including 31 archaeal and 203 bacterial species, mostly unknown and uncultivable. MAGs accounted for 54%–67% of the entire microbial community (depending on the bioreactor) and evidenced that the microbiome is extremely complex in terms of the number of species. The core microbiome was composed of Methanothrix soehngenii (the most abundant), Methanoculleus sp., unknown Bacteroidales and Spirochaetaceae. Relative abundance analysis of all the samples revealed microbes having substrate preferences. Substrate-specific species were mostly unknown and not predominant in the microbial communities.ConclusionsIn this experimental system, the dominant fermentation products subjected to methanogenesis moderately modified the final effect of bioreactor performance. At the molecular level, a different contribution of acetotrophic and hydrogenotrophic pathways for methane production, a very high level of new species recovered, and a moderate variability in microbial composition depending on substrate availability were evidenced. Propionate was not a factor ceasing methane production. All these findings are relevant because lactate, acetate, propionate and butyrate are the universal products of acidogenesis, regardless of feedstock.