Search

The Barents Sea shelf is one of the most economically promising regions in the Arctic in terms of its resources and geographic location. However, benthic microbial communities of the northeastern Barents Sea are still barely studied. Here, we present a detailed systematic description of the structures of microbial communities located in the sediments and bottom water of the northeastern Barents Sea based on 16S rRNA profiling and a qPCR assessment of the total prokaryotic abundance in 177 samples. Beta- and alpha-diversity analyses revealed a clear difference between the microbial communities of diverse sediment layers and bottom-water fractions. We identified 101 microbial taxa whose representatives had statistically reliable distribution patterns between these ecotopes. Analysis of the correlation between microbial community structure and geological data yielded a number of important results—correlations were found between the abundance of individual microbial taxa and bottom relief, thickness of marine sediments, presence of hydrotrolite interlayers, and the values of pH and Eh. We also demonstrated that a relatively high abundance of prokaryotes in sediments can be caused by the proliferation of Deltaproteobacteria representatives, in particular, sulfate and iron reducers. [ABSTRACT FROM AUTHOR]

Metatranscriptomics is a promising technique for the characterization of active members of microbial populations and their functions. Only a subset of the functional potential of the gut microbiota is expressed and many functions are transcribed only by few organisms when broadly present, but corresponding information for many other habitats is lacking. Transcriptional activity may predict bacterial growth and death; undertranscription relative to the DNA abundance indicates loss of the taxa, while overtranscription precedes future growth. Metatranscriptomics has revealed medically relevant expression signatures as exemplified by signatures associated with preterm labor and risk for developing asthma. Metatranscriptomics has in proof-of-concept studies provided a higher diagnostic yield than metagenomics for detecting infectious diseases, but results should be validated. Metatranscriptomics has revolutionized our ability to explore and understand transcriptional programs in microbial communities. Moreover, it has enabled us to gain deeper and more specific insight into the microbial activities in human gut, respiratory, oral, and vaginal communities. Perhaps the most important contribution of metatranscriptomics arises, however, from the analyses of disease-associated communities. We review the advantages and disadvantages of metatranscriptomics analyses in understanding human health and disease. We focus on human tissues low in microbial biomass and conditions associated with dysbiotic microbiota. We conclude that a more widespread use of metatranscriptomics and increased knowledge on microbe activities will uncover critical interactions between microbes and host in human health and provide diagnostic basis for culturing-independent, direct functional pathogen identification. [ABSTRACT FROM AUTHOR]

The Barents Sea shelf is one of the most economically promising regions in the Arctic in terms of its resources and geographic location. However, benthic microbial communities of the northeastern Barents Sea are still barely studied. Here, we present a detailed systematic description of the structures of microbial communities located in the sediments and bottom water of the northeastern Barents Sea based on 16S rRNA profiling and a qPCR assessment of the total prokaryotic abundance in 177 samples. Beta- and alpha-diversity analyses revealed a clear difference between the microbial communities of diverse sediment layers and bottom-water fractions. We identified 101 microbial taxa whose representatives had statistically reliable distribution patterns between these ecotopes. Analysis of the correlation between microbial community structure and geological data yielded a number of important results—correlations were found between the abundance of individual microbial taxa and bottom relief, thickness of marine sediments, presence of hydrotrolite interlayers, and the values of pH and Eh. We also demonstrated that a relatively high abundance of prokaryotes in sediments can be caused by the proliferation of Deltaproteobacteria representatives, in particular, sulfate and iron reducers.

Microbial communities are important for high composting efficiency and good quality composts. This study was conducted to compare the changes of physicochemical and bacterial characteristics in composting from different raw materials, including chicken manure (CM), duck manure (DM), sheep manure (SM), food waste (FW), and vegetable waste (VW). The role and interactions of core bacteria and their contribution to maturity in diverse composts were analyzed by advanced bioinformatics methods combined sequencing with co-occurrence network and structural equation modeling (SEM). Results indicated that there were obviously different bacterial composition and diversity in composting from diverse sources. FW had a low pH and different physiochemical characteristics compared to other composts but they all achieved similar maturity products. Redundancy analysis suggested total organic carbon, phosphorus, and temperature governed the composition of microbial species but key factors were different in diverse composts. Network analysis showed completely different interactions of core bacterial community from diverse composts but Thermobifida was the ubiquitous core bacteria in composting bacterial network. Sphaerobacter and Lactobacillus as core genus were presented in the starting mesophilic and thermophilic phases of composting from manure (CM, DM, SM) and municipal solid waste (FW, VW), respectively. SEM indicated core bacteria had the positive, direct, and the biggest (> 80%) effects on composting maturity. Therefore, this study presents theoretical basis to identify and enhance the core bacteria for improving full-scale composting efficiency facing more and more organic wastes. [ABSTRACT FROM AUTHOR]

Intestinal human microbiota is a dynamic system that is under the pressures of its host organism and external factors. Microbiota disruption caused by these factors can lead to severe diseases including inflammatory and oncological diseases of the gastrointestinal tract. One of the possible approaches in managing the intestinal microbiota is fecal microbiota transplantation (FT) – transfer of the microbiota from the stool of a healthy donor to the intestinal tract of a recipient patient. Currently, this procedure is recognized as an efficacious method to normalize the intestinal microbiota mainly in inflammatory diseases of the gastrointestinal tract. In Russia, pilot studies of the effectiveness of FT in patients with ulcerative colitis have been conducted for several years, and these studies were started in Novosibirsk. The aim of this study was to assess the change of intestinal microbiome in 20 patients with ulcerative colitis after a single FT procedure. The main method is a comparative analysis of 16S ribosomal RNA sequence libraries constructed using fecal samples obtained from patients with ulcerative colitis before and after FT and sequenced on the Illumina MiSeq platform. The obtained results showed that FT led to an increase in average biodiversity in samples after FT compared to samples before FT; however, the difference was not significant. In the samples studied, the proportion of Firmicutes sequences, the major gastrointestinal microbiota of healthy people, was decreased (~32 % vs. >70 %), while the proportion of Proteobacteria sequences was increased (>9 % vs.

Appropriate development of the intestinal microbiota during infancy is known to be important for human health. In fact, aberrant alterations of the microbial composition during childhood may cause short- and/or long-term negative health effects. Many factors influence the initial assembly and subsequent progression of the gut microbiota of a neonate, such as feeding type, delivery mode, gestational age, maternal metabolic status and antibiotic exposure.In the current study, the composition of the infant gut core-microbiota was explored, revealing particular variations of this core-microbiota during the first three years as influenced by delivery mode and feeding type. A multi-population cohort meta-analysis was performed by selecting 15 publicly available datasets pertaining to taxonomic profiles of 1035 fecal samples of healthy infants, as obtained by means of a 16S rRNA gene-based profiling approach. Interestingly, this multi-population cohort meta-analysis revealed great microbial complexity and specific taxonomic shifts in children older than six months, suggesting a major impact by the introduction of solid foods which prompts progression of infant gut microbiota towards that typical of adults. The taxonomic data sets employed in this multi-population cohort meta-analysis possess the statistical robustness to allow the identification of infant community state types (ICSTs). Our analysis therefore reveals the existence of specific taxonomic patterns that correspond to particular nutritional and developmental stages of early life, and that had previously been obscured by the high variability typical of such infant gut microbiota.

Abstract Background Anaerobic digestion, whose final products are methane and carbon dioxide, ensures energy flow and circulation of matter in ecosystems. This naturally occurring process is used for the production of renewable energy from biomass. Lactate, a common product of acidic fermentation, is a key intermediate in anaerobic digestion of biomass in the environment and biogas plants. Effective utilization of lactate has been observed in many experimental approaches used to study anaerobic digestion. Interestingly, anaerobic lactate oxidation and lactate oxidizers as a physiological group in methane-yielding microbial communities have not received enough attention in the context of the acetogenic step of anaerobic digestion. This study focuses on metabolic transformation of lactate during the acetogenic and methanogenic steps of anaerobic digestion in methane-yielding bioreactors. Results Methane-yielding microbial communities instead of pure cultures of acetate producers were used to process artificial lactate-rich media to methane and carbon dioxide in up-flow anaerobic sludge blanket reactors. The media imitated the mixture of acidic products found in anaerobic environments/digesters where lactate fermentation dominates in acidogenesis. Effective utilization of lactate and biogas production was observed. 16S rRNA profiling was used to examine the selected methane-yielding communities. Among Archaea present in the bioreactors, the order Methanosarcinales predominated. The acetoclastic pathway of methane formation was further confirmed by analysis of the stable carbon isotope composition of methane and carbon dioxide. The domain Bacteria was represented by Bacteroidetes, Firmicutes, Proteobacteria, Synergistetes, Actinobacteria, Spirochaetes, Tenericutes, Caldithrix, Verrucomicrobia, Thermotogae, Chloroflexi, Nitrospirae, and Cyanobacteria. Available genome sequences of species and/or genera identified in the microbial communities were searched for genes encoding the lactate-oxidizing metabolic machinery homologous to those of Acetobacterium woodii and Desulfovibrio vulgaris. Furthermore, genes for enzymes of the reductive acetyl-CoA pathway were present in the microbial communities. Conclusions The results indicate that lactate is oxidized mainly to acetate during the acetogenic step of AD and this comprises the acetotrophic pathway of methanogenesis. The genes for lactate utilization under anaerobic conditions are widespread in the domain Bacteria. Lactate oxidation to the substrates for methanogens is the most energetically attractive process in comparison to butyrate, propionate, or ethanol oxidation.

Tropical gar (Atractosteus tropicus) is freshwater and estuarine fish, inhabiting the Earth since the Mesozoic era and undergoing limited physiological variation ever since. Besides its recognized cultural and scientific relevance, the species has seen remarkable growth in its economic impact due to pisciculture. In this study, we present the first report of the whole taxonomic composition of microbial communities in gut contents in juveniles and adults of A. tropicus, by sex and origin (wild and cultivated). For this study, 508 genera were identified, with the most and least abundant being Cetobacterium and Paludibacter, respectively. Fusobacteria, Proteobacteria, Firmicutes, and Bacteroidetes phyla are the core gut microbiome of A. tropicus juvenile and adult by sex and origin. Deinococcus-Thermus phylum sequence was only identified in wild-type males. In the phylogenetic trees reconstruction Lactococcus lactis strains CAU929 and CAU6600, Cp6 and CAU9951, Cetobacterium strain H69, Aeromonas hydrophila strain P5 and WR-5-3-2, Aeromonas sobria strain CP DC28 and Aeromonas hydrophila were identified, some of them with probiotic potential within the three dominant phyla in core gut microbiome in A. tropicus adults, especially in wild-type organisms. Myroides genus was recognized in microbiota gut of the cultivated juvenile A. tropicus. Nevertheless, Alpha diversity indicated that the highest gut microbiota abundance and richness is found in cultivated juvenile and wild-type adult A. tropicus female, rather than adult wild-type males and the least gut microbiota abundance and richness is found in a cultivated adult of A. tropicus for both sexes. [ABSTRACT FROM AUTHOR]

The soil microbiome is inherently complex with high biological diversity, and spatial heterogeneity typically occurring on the submillimetre scale. To study the microbial ecology of soils, and other microbiomes, biomolecules, that is, nucleic acids and proteins, must be efficiently and reliably co‐recovered from the same biological samples. Commercial kits are currently available for the co‐extraction of DNA, RNA and proteins but none has been developed for soil samples. We present a new protocol drawing on existing phenol–chloroform‐based methods for nucleic acids co‐extraction but incorporating targeted precipitation of proteins from the phenol phase. The protocol is cost‐effective and robust, and easily implemented using reagents commonly available in laboratories. The method is estimated to be eight times cheaper than using disparate commercial kits for the isolation of DNA and/or RNA, and proteins, from soil. The method is effective, providing good quality biomolecules from a diverse range of soil types, with clay contents varying from 9.5% to 35.1%, which we successfully used for downstream, high‐throughput gene sequencing and metaproteomics. Additionally, we demonstrate that the protocol can also be easily implemented for biomolecule co‐extraction from other complex microbiome samples, including cattle slurry and microbial communities recovered from anaerobic bioreactors, as well as from Gram‐positive and Gram‐negative pure cultures. [ABSTRACT FROM AUTHOR]

Harvesting valuable bioproducts from various renewable feedstocks is necessary for the critical development of a sustainable bioeconomy. Anaerobic digestion is a well-established technology for the conversion of wastewater and solid feedstocks to energy with the additional potential for production of process intermediates of high market values (e.g., carboxylates). In recent years, first-generation biofuels typically derived from food crops have been widely utilized as a renewable source of energy. The environmental and socioeconomic limitations of such strategy, however, have led to the development of second-generation biofuels utilizing, amongst other feedstocks, lignocellulosic biomass. In this context, the anaerobic digestion of perennial grass holds great promise for the conversion of sustainable renewable feedstock to energy and other process intermediates. The advancement of this technology however, and its implementation for industrial applications, relies on a greater understanding of the microbiome underpinning the process. To this end, microbial communities recovered from replicated anaerobic bioreactors digesting grass were analyzed. The bioreactors leachates were not buffered and acidic pH (between 5.5 and 6.3) prevailed at the time of sampling as a result of microbial activities. Community composition and transcriptionally active taxa were examined using 16S rRNA sequencing and microbial functions were investigated using metaproteomics. Bioreactor fraction, i.e., grass or leachate, was found to be the main discriminator of community analysis across the three molecular level of investigation (DNA, RNA, and proteins). Six taxa, namely Bacteroidia, Betaproteobacteria, Clostridia, Gammaproteobacteria, Methanomicrobia, and Negativicutes accounted for the large majority of the three datasets. The initial stages of grass hydrolysis were carried out by Bacteroidia, Gammaproteobacteria, and Negativicutes in the grass biofilms, in addition to Clostridia in the bioreactor leachates. Numerous glycolytic enzymes and carbohydrate transporters were detected throughout the bioreactors in addition to proteins involved in butanol and lactate production. Finally, evidence of the prevalence of stressful conditions within the bioreactors and particularly impacting Clostridia was observed in the metaproteomes. Taken together, this study highlights the functional importance of Clostridia during the anaerobic digestion of grass and thus research avenues allowing members of this taxon to thrive should be explored.

Harvesting valuable bioproducts from various renewable feedstocks is necessary for the critical development of a sustainable bioeconomy. Anaerobic digestion is a well-established technology for the conversion of wastewater and solid feedstocks to energy with the additional potential for production of process intermediates of high market values (e.g., carboxylates). In recent years, first-generation biofuels typically derived from food crops have been widely utilized as a renewable source of energy. The environmental and socioeconomic limitations of such strategy, however, have led to the development of second-generation biofuels utilizing, amongst other feedstocks, lignocellulosic biomass. In this context, the anaerobic digestion of perennial grass holds great promise for the conversion of sustainable renewable feedstock to energy and other process intermediates. The advancement of this technology however, and its implementation for industrial applications, relies on a greater understanding of the microbiome underpinning the process. To this end, microbial communities recovered from replicated anaerobic bioreactors digesting grass were analyzed. The bioreactors leachates were not buffered and acidic pH (between 5.5 and 6.3) prevailed at the time of sampling as a result of microbial activities. Community composition and transcriptionally active taxa were examined using 16S rRNA sequencing and microbial functions were investigated using metaproteomics. Bioreactor fraction, i.e., grass or leachate, was found to be the main discriminator of community analysis across the three molecular level of investigation (DNA, RNA, and proteins). Six taxa, namely Bacteroidia, Betaproteobacteria, Clostridia, Gammaproteobacteria, Methanomicrobia, and Negativicutes accounted for the large majority of the three datasets. The initial stages of grass hydrolysis were carried out by Bacteroidia, Gammaproteobacteria, and Negativicutes in the grass biofilms, in addition to Clostridia in the bioreactor leachates. Numerous glycolytic enzymes and carbohydrate transporters were detected throughout the bioreactors in addition to proteins involved in butanol and lactate production. Finally, evidence of the prevalence of stressful conditions within the bioreactors and particularly impacting Clostridia was observed in the metaproteomes. Taken together, this study highlights the functional importance of Clostridia during the anaerobic digestion of grass and thus research avenues allowing members of this taxon to thrive should be explored. [ABSTRACT FROM AUTHOR]

Next Generation Sequencing (NGS) technologies have overcome the limitations of cultivation-dependent approaches and allowed detailed study of bacterial populations that inhabit the human body. The consortium of bacteria residing in the human intestinal tract, also known as the gut microbiota, impacts several physiological processes important for preservation of the health status of the host. The most widespread microbiota profiling method is based on amplification and sequencing of a variable portion of the 16S rRNA gene as a universal taxonomic marker among members of the Bacteria domain. Despite its popularity and obvious advantages, this 16S rRNA gene-based approach comes with some important limitations. In particular, the choice of the primer pair for amplification plays a major role in defining the accuracy of the reconstructed bacterial profiles. In the current study, we performed an in silico PCR using all currently described 16S rRNA gene-targeting primer pairs (PP) in order to assess their efficiency. Our results show that V3, V4, V5, and V6 were the optimal regions on which to design 16S rRNA metagenomic primers. In detail, PP39 (Probio_Uni/Probio_Rev), PP41 (341F/534R), and PP72 (970F/1050R) were the most suitable primer pairs with an amplification efficiency of >98.5%. Furthermore, the Bifidobacterium genus was examined as a test case for accurate evaluation of intra-genus performances at subspecies level. Intriguingly, the in silico analysis revealed that primer pair PP55 (527f/1406r) was unable to amplify the targeted region of any member of this bacterial genus, while several other primer pairs seem to rather inefficiently amplify the target region of the main bifidobacterial taxa. These results highlight that selection of a 16S rRNA gene-based PP should be done with utmost care in order to avoid biases in microbiota profiling results.

Worldwide, 10% of babies are born preterm, defined as birth before 37 weeks' gestation. We have had little success in developing strategies to prevent preterm births, the majority of which are due to infection or are idiopathic. An emerging hypothesis is that the maternal microbiome—the bacteria that inhabit the mother's body and play vital functions in normal health—contributes to the etiology of preterm birth. Here, we highlight the latest data revealing correlations between preterm birth and maternal intestinal, vaginal, cervical, and placental microbiomes. Additionally, we describe the most commonly used comparative microbiome analysis methods and highlight important issues to consider when conducting such studies. [ABSTRACT FROM AUTHOR]

D-lactic acidosis (D-LA) is an uncommon complication of short bowel syndrome characterized by elevated plasma D-lactate and encephalopathy. Treatments include rehydration, dietary carbohydrate restriction, and antibiotics to alter the gut microbiota. Fecal microbiota transplantation (FMT) has recently been used in children to successfully treat D-LA. We compared the clinical course and then utilized metagenomic shotgun sequencing to describe changes in the composition and function of the intestinal microbiome following FMT in 2 patients with recurrent D-LA. FMT altered the composition of the fecal microbiota in these 2 patients with recurrent D-LA, though not necessarily in a consistent manner. Importantly, microbial metabolic pathways were also impacted by FMT, which may be critical for achieving desired clinical outcomes. While sample size limits the generalizability of our results, these findings set the stage for further understanding of the role of microbes in the pathogenesis of recurrent D-LA., Competing Interests: The authors report no conflicts of interest., (Copyright © 2023 The Author(s). Published by Wolters Kluwer Health, Inc. on behalf of the European Society for Pediatric Gastroenterology, Hepatology, and Nutrition and the North American Society for Pediatric Gastroenterology, Hepatology, and Nutrition.)

Oil refining waste (ORW) contains complex, hazardous, and refractory components, causing more severe long-term environmental pollution than petroleum. Here, ORW was used to simulate the accelerated domestication of bacteria from oily sludges and polymer-flooding wastewater, and the effects of key factors, oxygen and temperature, on the ORW degradation were evaluated. Bacterial communities acclimated respectively in 30/60 °C, aerobic/anaerobic conditions showed differentiated degradation rates of ORW, ranging from 5% to 34%. High-throughput amplicon sequencing and ORW component analysis revealed significant correlation between bacterial diversity/biomass and degradation efficiency/substrate preference. Under mesophilic and oxygen-rich condition, the high biomass and abundant biodiversity with diverse genes and pathways for petroleum hydrocarbons degradation, effectively promoted the rapid and multi-component degradation of ORW. While under harsh conditions, a few dominant genera still contributed to ORW degradation, although the biodiversity was severely restricted. The typical dominant facultative anaerobes Bacillus (up to 99.8% abundance anaerobically) and Geobacillus (up to 99.9% abundance aerobically and anaerobically) showed oxygen-independent sustainable degradation ability and broad-spectrum of temperature adaptability, making them promising and competitive bioremediation candidates for future application. Our findings provide important strategies for practical bioremediation of varied environments polluted by hazardous ORW. [Display omitted] • Stress-opted biodiversities differentially contribute to hazardous ORW degradation. • Abundant and diverse organic-metabolic genes/pathways aid degradation efficiency. • Bacillus and Geobacillus are competitive, sustainable and broad-adaptable degraders. • Oxygen and suitable temperature maximize remediation effect of in-situ microbiota. • Supplementation of facultative anaerobes helps remove ORW from harsh environments. [ABSTRACT FROM AUTHOR]

Turkish artisanal Tulum cheese is a traditional raw milk cheese and starter cultures are not used for ripening process. In this study, 16S rRNA amplicon-based sequencing approach was used for determining bacterial diversity in Tulum cheeses. The volatile organic compound (VOC) profile which contributes to sensory properties of the cheeses was determined via solid-phase microextraction combined with gas chromatography-mass spectrometry (GC-MS/SPME) and correlated with the microbiota. In Tulum cheese samples, Lactobacillaceae (30,71%) phylum widely varied but Streptococcaceae (59,86%) was dominant. It mainly belongs to the four dominant genera of lactic acid bacteria (LAB) and includes Lactobacillus, Lactococcus, Streptococcus and Pediococcus. The bacterial diversity of TC5 sample was much higher than that of other samples. Moreover, several species of non-starter LAB were frequently detected. A total of 36 volatile compounds were identified including hydrocarbons (3), acids (13), alcohols (5), ketones (3), aldehydes (3), esters (8) and miscellaneous compound (1). Pentadecanoic acid, octadecanoic acid, tetradecanoic acid, octanoic acid, hexanoic acid were determined to be dominant in all cheese samples and to be related to microbial flora. Also, the sensory profiles of the samples were examined. The most popular TC6 did not contain 2-butanone and hexanal from the volatile organic compound (VOC) values, while in the most unpopular TC3, 2-butanone (7.73%) and hexanal (5.95%) were found to be the highest. The outcome of this study will contribute to literature on bacterial diversity and aromatic profile of Tulum cheese which should be investigated in order to improve quality and safety. • The bacterial population diversity of Tulum cheeses was assessed by using amplicon based metagenomic sequencing method. • Lactobacillus, Lactococcus, Streptococcus, Leuconostoc, Enterococcus, Pediococcus prevailed in Tulum cheese samples. • Thirty-six different volatile compounds were identified in Tulum cheese samples. • Pentadecanoic acid, octadecanoic acid, tetradecanoic acid, octanoic acid, hexanoic acid were dominant mainly in all samples. [ABSTRACT FROM AUTHOR]

Background: Bifidobacterium represents an important early life microbiota member. Specific bifidobacterial components, exopolysaccharides (EPS), positively modulate host responses, with purified EPS also suggested to impact microbe–microbe interactions by acting as a nutrient substrate. Thus, we determined the longitudinal effects of bifidobacterial EPS on microbial communities and metabolite profiles using an infant model colon system. Methods: Differential gene expression and growth characteristics were determined for each strain; Bifidobacterium breve UCC2003 and corresponding isogenic EPS-deletion mutant (B. breve UCC2003del). Model colon vessels were inoculated with B. breve and microbiome dynamics monitored using 16S rRNA sequencing and metabolomics (NMR). Results: Transcriptomics of EPS mutant vs. B. breve UCC2003 highlighted discrete differential gene expression (e.g., eps biosynthetic cluster), though overall growth dynamics between strains were unaffected. The EPS-positive vessel had significant shifts in microbiome and metabolite profiles until study end (405 h); with increases of Tyzzerella and Faecalibacterium, and short-chain fatty acids, with further correlations between taxa and metabolites which were not observed within the EPS-negative vessel. Conclusions: These data indicate that B. breve UCC2003 EPS is potentially metabolized by infant microbiota members, leading to differential microbial metabolism and altered metabolite by-products. Overall, these findings may allow development of EPS-specific strategies to promote infant health. [ABSTRACT FROM AUTHOR]

Next Generation Sequencing (NGS) technologies have overcome the limitations of cultivation-dependent approaches and allowed detailed study of bacterial populations that inhabit the human body. The consortium of bacteria residing in the human intestinal tract, also known as the gut microbiota, impacts several physiological processes important for preservation of the health status of the host. The most widespread microbiota profiling method is based on amplification and sequencing of a variable portion of the 16S rRNA gene as a universal taxonomic marker among members of the Bacteria domain. Despite its popularity and obvious advantages, this 16S rRNA gene-based approach comes with some important limitations. In particular, the choice of the primer pair for amplification plays a major role in defining the accuracy of the reconstructed bacterial profiles. In the current study, we performed an in silico PCR using all currently described 16S rRNA gene-targeting primer pairs (PP) in order to assess their efficiency. Our results show that V3, V4, V5, and V6 were the optimal regions on which to design 16S rRNA metagenomic primers. In detail, PP39 (Probio_Uni/Probio_Rev), PP41 (341F/534R), and PP72 (970F/1050R) were the most suitable primer pairs with an amplification efficiency of >98.5%. Furthermore, the Bifidobacterium genus was examined as a test case for accurate evaluation of intra-genus performances at subspecies level. Intriguingly, the in silico analysis revealed that primer pair PP55 (527f/1406r) was unable to amplify the targeted region of any member of this bacterial genus, while several other primer pairs seem to rather inefficiently amplify the target region of the main bifidobacterial taxa. These results highlight that selection of a 16S rRNA gene-based PP should be done with utmost care in order to avoid biases in microbiota profiling results. [ABSTRACT FROM AUTHOR]

Background: Anaerobic digestion, whose final products are methane and carbon dioxide, ensures energy flow and circulation of matter in ecosystems. This naturally occurring process is used for the production of renewable energy from biomass. Lactate, a common product of acidic fermentation, is a key intermediate in anaerobic digestion of biomass in the environment and biogas plants. Effective utilization of lactate has been observed in many experimental approaches used to study anaerobic digestion. Interestingly, anaerobic lactate oxidation and lactate oxidizers as a physiological group in methane-yielding microbial communities have not received enough attention in the context of the acetogenic step of anaerobic digestion. This study focuses on metabolic transformation of lactate during the acetogenic and methanogenic steps of anaerobic digestion in methane-yielding bioreactors. Results: Methane-yielding microbial communities instead of pure cultures of acetate producers were used to process artificial lactate-rich media to methane and carbon dioxide in up-flow anaerobic sludge blanket reactors. The media imitated the mixture of acidic products found in anaerobic environments/digesters where lactate fermentation dominates in acidogenesis. Effective utilization of lactate and biogas production was observed. 16S rRNA profiling was used to examine the selected methane-yielding communities. Among Archaea present in the bioreactors, the order Methanosarcinales predominated. The acetoclastic pathway of methane formation was further confirmed by analysis of the stable carbon isotope composition of methane and carbon dioxide. The domain Bacteria was represented by Bacteroidetes, Firmicutes, Proteobacteria, Synergistetes, Actinobacteria, Spirochaetes, Tenericutes, Caldithrix, Verrucomicrobia, Thermotogae, Chloroflexi, Nitrospirae, and Cyanobacteria. Available genome sequences of species and/or genera identified in the microbial communities were searched for genes encoding the lactate-oxidizing metabolic machinery homologous to those of Acetobacterium woodii and Desulfovibrio vulgaris. Furthermore, genes for enzymes of the reductive acetyl-CoA pathway were present in the microbial communities. Conclusions: The results indicate that lactate is oxidized mainly to acetate during the acetogenic step of AD and this comprises the acetotrophic pathway of methanogenesis. The genes for lactate utilization under anaerobic conditions are widespread in the domain Bacteria. Lactate oxidation to the substrates for methanogens is the most energetically attractive process in comparison to butyrate, propionate, or ethanol oxidation. [ABSTRACT FROM AUTHOR]

Purpose: The aims of this work were: (i) to profile and compare pelvic and cervical microbiota from patients with pelvic inflammatory disease (PID); (ii) to test the ascending infection hypothesis that the microbes in the vagina or cervix spread to the upper genital tract and cause PID., Methodology: Thirty-eight PID patients and 19 control patients were enrolled in this study and received salpingectomy or salpingo-oophorectomy. Both pelvic and cervical samples were collected during surgery, which were subject to culture diagnosis and next-generation sequencing (NGS)-based 16S rRNA profiling., Results: For the PID group, the NGS-based method revealed that half of the pelvic samples were dominated by a single organism while the other half exhibited a polymicrobial infection. The pelvic and cervical microbiota were similar in terms of both taxonomic richness and evenness. Pelvic microbiota was dominated by Acinetobacter, Escherichia, Sneathia and Streptococcus whereas cervical microbiota was dominated by Lactobacillus and Gardnerella. Only six pelvic samples were positive by culture diagnosis, three of which matched the NGS-based results. The PID group was further divided into 23 hydrosalpinx and 15 pyosalpinx samples according to the pelvic mass collected during surgery, the former showing a richer bacterial composition than the latter., Conclusion: The NGS-based method provides a more informative profile of pelvic microbiota than the culture method. The pelvic and cervical microbiota from the same patient were inconsistent with each other, and thus therapeutic decisions based on cervical microbiota may lead to antimicrobial misuse.

Microbiome research is an emerging field in medical sciences. Several studies have made headways in understanding the influence of microbes on our health and disease states. Further progress in mapping microbiome populations across different body sites and understanding the underlying mechanisms of microbiome-host interactions depends critically on study design, collection protocols, analytical genetic techniques, and reference databases. In particular, a shift has appeared going from small sample collections to large-scale population studies (with extensive phenotypic information including disease status) which calls for some adaptions. In this review we will focus on gut microbiome profiling using the 16S ribosomal RNA approach in the setting of large-scale population studies, and discuss some novel developments., (Copyright © 2017. Published by Elsevier Ltd.)