Characterization of Sinus Microbiota by 16S Sequencing from Swabs.
Ontology highlight
ABSTRACT:
SUBMITTER: Vickery TW
PROVIDER: S-EPMC5558891 | biostudies-other | 2017
REPOSITORIES: biostudies-other
Similar Datasets
Project description:Rectal swabs are potentially a valuable method for monitoring the gut microbiome in research and clinical settings, where it is important to adhere to strict timing, or where acute sampling is needed. It is currently unknown whether rectal swabs give comparable results to faecal samples regarding microbiota community composition in neonates and infants. To study how well the two sampling methods correlate in infants, we compared the 16S-rRNA-based sequencing results of 131 paired rectal swabs and faecal samples collected from 116 infants at two timepoints in early life. The paired samples were highly comparable regarding both diversity and overall community composition, and strongly correlated on taxonomical level. We observed no significant nor relevant contribution of sampling method to the variation in overall gut microbiota community composition in a multivariable model. Our study provides evidence supporting the use of rectal swabs as a reliable proxy for faecal samples in infant gut microbiota research.
| S-EPMC6831562 | biostudies-literature
Project description:BACKGROUND:Substantial progress in high-throughput metagenomic sequencing methodologies has enabled the characterisation of bacteria from various origins (for example gut and skin). However, the recently-discovered bacterial microbiota present within animal internal tissues has remained unexplored due to technical difficulties associated with these challenging samples. RESULTS:We have optimized a specific 16S rDNA-targeted metagenomics sequencing (16S metabarcoding) pipeline based on the Illumina MiSeq technology for the analysis of bacterial DNA in human and animal tissues. This was successfully achieved in various mouse tissues despite the high abundance of eukaryotic DNA and PCR inhibitors in these samples. We extensively tested this pipeline on mock communities, negative controls, positive controls and tissues and demonstrated the presence of novel tissue specific bacterial DNA profiles in a variety of organs (including brain, muscle, adipose tissue, liver and heart). CONCLUSION:The high throughput and excellent reproducibility of the method ensured exhaustive and precise coverage of the 16S rDNA bacterial variants present in mouse tissues. This optimized 16S metagenomic sequencing pipeline will allow the scientific community to catalogue the bacterial DNA profiles of different tissues and will provide a database to analyse host/bacterial interactions in relation to homeostasis and disease.
| S-EPMC4636327 | biostudies-literature
Project description:The tissue-resident microbiota is an integral component of multiple tumor types, but it remains challenging to characterize its abundance and composition due to its low biomass. Here, we describe an optimized protocol for quantification and profiling of tissue-resident microbiota. The major optimized steps include DNA extraction, qPCR, 16S library construction, and bioinformatics analysis. This protocol enables robust and accurate characterization of the dynamics of normal and tumor tissue-resident microbiota at its physiological abundance from both mouse and human origins. For complete details on the use and execution of this protocol, please refer to Fu et al. (2022).
| S-EPMC9558047 | biostudies-literature
Project description:As the applications of microbiome science in agriculture expand, laboratory methods should be constantly evaluated to ensure optimization and reliability of downstream results. Most animal microbiome research uses fecal samples or rectal swabs for profiling the gut bacterial community; however, in birds, this is difficult given the unique anatomy of the cloaca where the fecal, urinary, and reproductive tracts converge into one orifice. Therefore, avian gut microbiomes are usually sampled from cloacal swabs, creating a need to evaluate sample preparation methods to optimize 16S sequencing. We compared four different DNA extraction methods from two commercially available kits on cloacal swabs from 10 adult commercial laying hens and included mock communities and negative controls, which were then subjected to 16S rRNA amplicon sequencing. Extracted DNA yield and quality, diversity analyses, and contaminants were assessed. Differences in DNA quality and quantity were observed, and all methods needed further purification for optimal sequencing, suggesting contaminants due to cloacal contents, method reagents, and/or environmental factors. However, no differences were observed in alpha or beta diversity between methods. Importantly, multiple bacterial contaminants were detected in each mock community and negative control, indicating the prevalence of laboratory and handling contamination as well as method-specific reagent contamination. We found that although the extraction methods resulted in different extraction quality and yield, overall sequencing results were not affected, and we did not identify any method that would be an inappropriate choice in extracting DNA from cloacal swabs for 16S rRNA sequencing. Overall, our results highlight the need for careful consideration of positive and negative controls in addition to DNA isolation method and lend guidance to future microbiome research in poultry.
| S-EPMC8151189 | biostudies-literature
Project description:In this paper we compared taxonomic results obtained by metataxonomics (16S rRNA gene sequencing) and metagenomics (whole shotgun metagenomic sequencing) to investigate their reliability for bacteria profiling, studying the chicken gut as a model system. The experimental conditions included two compartments of gastrointestinal tracts and two sampling times. We compared the relative abundance distributions obtained with the two sequencing strategies and then tested their capability to distinguish the experimental conditions. The results showed that 16S rRNA gene sequencing detects only part of the gut microbiota community revealed by shotgun sequencing. Specifically, when a sufficient number of reads is available, Shotgun sequencing has more power to identify less abundant taxa than 16S sequencing. Finally, we showed that the less abundant genera detected only by shotgun sequencing are biologically meaningful, being able to discriminate between the experimental conditions as much as the more abundant genera detected by both sequencing strategies.
| S-EPMC7862389 | biostudies-literature
Project description:BackgroundChildhood obesity constitutes a worldwide health problem, and the gut microbiota play extremely important roles in obesity. Herein, we aimed to characterize the gut microbiota in children of normal weight, overweight, and obesity.MethodsThirty children of normal weight, 35 who were overweight, and 35 with obesity were enrolled from Nanjing, China. We isolated DNA from fecal samples, and employed 16S rRNA gene sequencing to explore the diversity and composition of gut microbiota.ResultsThe operational taxonomic unit (OTU) numbers exhibited a reduction in the gut microbiota abundance with the increase in the body weight. Alpha diversity analysis revealed a sharp decrease in the mean microbial abundance among the three groups (Chao1: F = 5.478, P = 0.006; observed species: F = 7.271, P = 0.001; PD whole tree: F = 8.735, P < 0.001). Beta diversity analysis indicated notable differences in the gut microbial composition between children of normal weight and obesity. However, overweight children had little difference in gut microbiota compared to either children of normal weight or obesity. At the genus level, Oscillospira decreased among the three groups (χ2 = 10.062, P = 0.001), and Sutterella increased (F = 4.052, P = 0.020). There were many remarkably increased species of gut bacteria in the comparison among three groups, 31 in the normal weight group, 32 in the obese group, and only three species of bacteria were identified in the overweight group. These significantly increased species of gut bacteria may have a close relationship with the progression of obesity.ConclusionsThe abundance of species decreased significantly as the BMI increased. Although the gut microbial composition between children of normal weight and obesity was notably different, due to the changing ratio of some microbial communities, gut microbiota in overweight children showed similarities to that of children with normal weight and obesity.
| S-EPMC8194416 | biostudies-literature
Project description:Due to their potential impact on the host's phenotype, organ-specific microbiotas are receiving increasing attention in several animal species, including cattle. Specifically, the vaginal microbiota of ruminants is attracting growing interest, due to its predicted critical role on cows' reproductive functions in livestock contexts. Notably, fertility disorders represent a leading cause for culling, and additional research would help to fill relevant knowledge gaps. In the present study, we aimed to characterize the vaginal microbiota of a large cohort of 1171 female dairy cattle from 19 commercial herds in Northern France. Vaginal samples were collected using a swab and the composition of the microbiota was determined through 16S rRNA sequencing targeting the V3-V4 hypervariable regions. Initial analyses allowed us to define the core bacterial vaginal microbiota, comprising all the taxa observed in more than 90% of the animals. Consequently, four phyla, 16 families, 14 genera and a single amplicon sequence variant (ASV) met the criteria, suggesting a high diversity of bacterial vaginal microbiota within the studied population. This variability was partially attributed to various environmental factors such as the herd, sampling season, parity, and lactation stage. Next, we identified numerous significant associations between the diversity and composition of the vaginal microbiota and several traits related to host's production and reproduction performance, as well as reproductive tract health. Specifically, 169 genera were associated with at least one trait, with 69% of them significantly associated with multiple traits. Among these, the abundances of Negativibacillus and Ruminobacter were positively correlated with the cows' performances (i.e., longevity, production performances). Other genera showed mixed relationships with the phenotypes, such as Leptotrichia being overabundant in cows with improved fertility records and reproductive tract health, but also in cows with lower production levels. Overall, the numerous associations underscored the complex interactions between the vaginal microbiota and its host. Given the large number of samples collected from commercial farms and the diversity of the phenotypes considered, this study marks an initial step towards a better understanding of the intimate relationship between the vaginal microbiota and the dairy cow's phenotypes.
| S-EPMC11336114 | biostudies-literature
Project description:Periodontal abscess is an oral infective disease caused by various kinds of bacteria. We aimed to characterize the microbiota composition of periodontal abscesses by metagenomic methods and compare it to that of the corresponding pocket and healthy gingival crevice to investigate the specific bacteria associated with this disease. Samples from abscess pus (AB), periodontal pocket coronally above the abscess (PO), and the gingival crevice of the periodontal healthy tooth were obtained from 20 periodontal abscess patients. Furthermore, healthy gingival crevice samples were obtained from 25 healthy individuals. Bacterial DNA was extracted and 16S rRNA gene fragments were sequenced to characterize the microbiota and determine taxonomic classification. The beta-diversity analysis results showed that the AB and PO groups had similar compositions. Porphyromonas gingivalis, Prevotella intermedia, and other Prevotella spp. were the predominant bacteria of human periodontal abscesses. The abundances of Filifactor alocis and Atopobium rimae were significantly higher in periodontal abscesses than in the periodontal pocket, suggesting their association with periodontal abscess formation. In conclusion, we characterized the microbiota in periodontal abscess and identified some species that are positively associated with this disease. This provides a better understanding of the components of periodontal abscesses, which will help facilitate the development of antibiotic therapy strategies.
| S-EPMC6682650 | biostudies-literature
Project description:Metagenomics seeks to understand microbial communities and assemblages by DNA sequencing. Technological advances in next generation sequencing technologies are fuelling a rapid growth in the number and scope of projects aiming to analyze complex microbial environments such as marine, soil or the gut. Recent improvements in longer read lengths and paired-sequencing allow better resolution in profiling microbial communities. While both 454 sequencing and Illumina sequencing have been used in numerous metagenomic studies, SOLiD sequencing is not commonly used in this area, as it is believed to be more suitable in the context of reference-guided projects.To investigate the performance of SOLiD sequencing in a metagenomic context, we compared taxonomic profiles of SOLiD mate-pair sequencing reads with Sanger paired reads and 454 single reads. All sequences were obtained from the bacterial 16S rRNA gene, which was amplified from microbial DNA extracted from a human fecal sample. Additionally, from the same fecal sample, complete genomic microbial DNA was extracted and shotgun sequenced using SOLiD sequencing to study the composition of the intestinal microbiota and the existing microbial metabolism. We found that the microbiota composition of 16S rRNA gene sequences obtained using Sanger, 454 and SOLiD sequencing provide results comparable to the result based on shotgun sequencing. Moreover, with SOLiD sequences we obtained more resolution down to the species level. In addition, the shotgun data allowed us to determine a functional profile using the databases SEED and KEGG.This study shows that SOLiD mate-pair sequencing is a viable and cost-efficient option for analyzing a complex microbiome. To the best of our knowledge, this is the first time that SOLiD sequencing has been used in a human sample.
| S-EPMC3852202 | biostudies-literature
Project description:The human bladder contains bacteria, even in the absence of infection. Interest in studying these bacteria and their association with bladder conditions is increasing. However, the chosen experimental method can limit the resolution of the taxonomy that can be assigned to the bacteria found in the bladder. 16S rRNA amplicon sequencing is commonly used to identify bacteria in urinary specimens, but it is typically restricted to genus-level identification. Our primary aim here was to determine if accurate species-level identification of bladder bacteria is possible using 16S rRNA amplicon sequencing. We evaluated the ability of different classification schemes, each consisting of combinations of a reference database, a 16S rRNA gene variable region, and a taxonomic classification algorithm to correctly classify bladder bacteria. We show that species-level identification is possible and that the reference database chosen is the most important component, followed by the 16S variable region sequenced. IMPORTANCE Accurate species-level identification from culture-independent techniques is of importance for microbial niches that are less well characterized, such as that of the bladder. 16S rRNA amplicon sequencing, a common culture-independent way to identify bacteria, is often critiqued for lacking species-level resolution. Here, we extensively evaluate classification schemes for species-level bacterial annotation of 16S amplicon data from bladder bacteria. Our results show that the proper choice of taxonomic database and variable region of the 16S rRNA gene sequence makes species level identification possible. We also show that this improvement can be achieved through the more careful application of existing methods and resources. Species-level information may deepen our understanding of associations between bacteria in the bladder and bladder conditions such as lower urinary tract symptoms and urinary tract infections.
| S-EPMC8547459 | biostudies-literature