scholarly journals Low Maternal Microbiota Sharing across Gut, Breast Milk and Vagina, as Revealed by 16S rRNA Gene and Reduced Metagenomic Sequencing

Genes ◽  
2018 ◽  
Vol 9 (5) ◽  
pp. 231 ◽  
Author(s):  
Ekaterina Avershina ◽  
Inga Angell ◽  
Melanie Simpson ◽  
Ola Storrø ◽  
Torbjørn Øien ◽  
...  
Keyword(s):  
Breast Milk ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Rrna Gene ◽  
Metagenomic Sequencing

scholarly journals Comparison between 16S rRNA and shotgun sequencing data for the taxonomic characterization of the gut microbiota

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Francesco Durazzi ◽  
Claudia Sala ◽  
Gastone Castellani ◽  
Gerardo Manfreda ◽  
Daniel Remondini ◽  
...  
Keyword(s):  
16S Rrna ◽  
Gut Microbiota ◽  
16S Rrna Gene ◽  
Gene Sequencing ◽  
16S Rrna Gene Sequencing ◽  
Shotgun Sequencing ◽  
Rrna Gene ◽  
Metagenomic Sequencing ◽  
Experimental Conditions ◽  
Rrna Gene Sequencing

AbstractIn 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.

scholarly journals Metagenome-validated Parallel Amplicon Sequencing and Text Mining-based Annotations for Simultaneous Profiling of Bacteria and Fungi: Vaginal Microbiome and Mycobiota in Healthy Women

2021 ◽  
Author(s):  
Seppo Virtanen ◽  
Schahzad Saqib ◽  
Tinja Kanerva ◽  
Pekka Nieminen ◽  
Ilkka Kalliala ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Cost Effective ◽  
Amplicon Sequencing ◽  
Rrna Gene ◽  
Extensive Literature ◽  
Metagenomic Sequencing ◽  
Text Extraction ◽  
Healthy Women ◽  
Bacteria And Fungi

Abstract Background: Amplicon sequencing of kingdom-specific tags such as 16S rRNA gene for bacteria and internal transcribed spacer (ITS) region for fungi are widely used for investigating microbial populations. So far most human studies have focused on bacteria while studies on host-associated fungi in health and disease have only recently started to accumulate. To enable cost-effective parallel analysis of bacterial and fungal communities in human and environmental samples, we developed a method where 16S rRNA gene and ITS-1 amplicons were pooled together for a single Illumina MiSeq or HiSeq run and analysed after primer-based segregation. Taxonomic assignments were performed with Blast in combination with an iterative text-extraction based filtration approach, which uses extensive literature records from public databases to select the most probable hits that were further validated by shotgun metagenomic sequencing. Results: Using 50 vaginal samples, we show that the combined run provides comparable results on bacterial composition and diversity to conventional 16S rRNA gene amplicon sequencing. The text-extraction-based taxonomic assignment guided tool provided ecosystem specific annotations that were confirmed by Metagenomic Phylogenetic Analysis (MetaPhlAn). The metagenome analysis revealed distinct functional differences between the bacterial community types while fungi were undetected, despite being identified in all samples based on ITS amplicons. Co-abundance analysis of bacteria and fungi did not show strong between-kingdom correlations within the vaginal ecosystem of healthy women.Conclusion: Combined amplicon sequencing for bacteria and fungi provides a simple and cost-effective method for simultaneous analysis of microbiota and mycobiota within the same samples. Text extraction-based annotation tool facilitates the characterization and interpretation of defined microbial communities from rapidly accumulating sequencing and metadata readily available through public databases.

scholarly journals Alone Yet Not Alone: Frankia Lives Under the Same Roof With Other Bacteria in Actinorhizal Nodules

2021 ◽  
Vol 12 ◽  
Author(s):  
Faten Ghodhbane-Gtari ◽  
Timothy D’Angelo ◽  
Abdellatif Gueddou ◽  
Sabrine Ghazouani ◽  
Maher Gtari ◽  
...  
Keyword(s):  
16S Rrna ◽  
Plant Growth ◽  
16S Rrna Gene ◽  
Root Nodules ◽  
Actinorhizal Plants ◽  
Rrna Gene ◽  
Plant Growth Promoting Bacteria ◽  
Metagenomic Sequencing ◽  
Plant Growth Promoting ◽  
Growth Promoting

Actinorhizal plants host mutualistic symbionts of the nitrogen-fixing actinobacterial genus Frankia within nodule structures formed on their roots. Several plant-growth-promoting bacteria have also been isolated from actinorhizal root nodules, but little is known about them. We were interested investigating the in planta microbial community composition of actinorhizal root nodules using culture-independent techniques. To address this knowledge gap, 16S rRNA gene amplicon and shotgun metagenomic sequencing was performed on DNA from the nodules of Casuarina glauca. DNA was extracted from C. glauca nodules collected in three different sampling sites in Tunisia, along a gradient of aridity ranging from humid to arid. Sequencing libraries were prepared using Illumina NextEra technology and the Illumina HiSeq 2500 platform. Genome bins extracted from the metagenome were taxonomically and functionally profiled. Community structure based off preliminary 16S rRNA gene amplicon data was analyzed via the QIIME pipeline. Reconstructed genomes were comprised of members of Frankia, Micromonospora, Bacillus, Paenibacillus, Phyllobacterium, and Afipia. Frankia dominated the nodule community at the humid sampling site, while the absolute and relative prevalence of Frankia decreased at the semi-arid and arid sampling locations. Actinorhizal plants harbor similar non-Frankia plant-growth-promoting-bacteria as legumes and other plants. The data suggests that the prevalence of Frankia in the nodule community is influenced by environmental factors, with being less abundant under more arid environments.

scholarly journals KAJIAN MOLEKULER PADA PROBIOTIK ASAL AIR SUSU IBU DALAM SINTESIS EKSOPOLISAKARIDA (EPS)

2021 ◽  
Vol 8 (1) ◽  
pp. 114-123
Author(s):  
Hamiyawati Qoimatu Dini Alfaruqi ◽  
Nosa Septiana Anindita ◽  
Arif Bimantara
Keyword(s):  
Breast Milk ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Universal Primers ◽  
Rrna Gene ◽  
Human Breast Milk ◽  
Human Breast ◽  
Adhesion Ability ◽  
The 16S Rrna Gene ◽  
Selection Of

Molecular Studies on Probiotic of Human Breast Milk in the Synthesis of Exopolysaccharide (EPS)  The glucosyltransferase (gtf) gene has an important role in exopolysaccharide (EPS) synthesis in probiotic bacteria. The EPS produced is associated with the adhesion ability of bacteria to the intestinal mucosa. Therefore, the gtf gene can be used as a parameter in the selection of potential probiotic through a molecular approach. This study was conducted to determine the presence of the gtf gene in probiotic from human breast milk using PCR technique. The methods in this study include the following: reculture of probiotic isolates, DNA isolation, amplification of the 16S rRNA gene using universal primers (pA and pB), amplification of specific LAB primers (LABfw and LABrv), specific primary design for the gtf gene, and the amplification of the gtf gene. The results of 16S rRNA gene amplification using universal primers obtained the amplicons of 500-1,000 bp in size. The results of amplification using specific LAB primers obtained an amplicon of about 700 bp in all isolates. The results of amplification of the gtf gene using a specific primer produced an amplicon of 325 bp in all isolates. Based on this study, it was concluded that 16 probiotic isolates from human breast milk were proven to have the gtf gene. Gen glukosiltransferase (gtf) memiliki peran penting dalam sintesis eksopolisakarida (EPS) pada bakteri probiotik. EPS yang diproduksi berhubungan dengan kemampuan adhesi bakteri pada mukosa usus. Oleh karena itu, gen gtf dapat dijadikan sebagai salah satu parameter dalam seleksi probiotik potensial melalui pendekatan molekuler. Penelitian ini dilakukan untuk mengetahui adanya gen gtf pada probiotik asal air susu ibu (ASI) menggunakan teknik PCR. Metode pada penelitian ini meliputi: reculture isolat probiotik, isolasi DNA, amplifikasi gen 16S rRNA menggunakan primer universal (pA dan pB), amplifikasi primer spesifik BAL (LABfw dan LABrv), desain primer spesifik untuk gen gtf dan amplifikasi gen gtf. Hasil amplifikasi gen 16S rRNA menggunakan primer universal diperoleh amplikon berukuran antara 500-1.000 bp. Adapun hasil amplifikasi menggunakan primer spesifik BAL diperoleh amplikon berukuran sekitar 700 bp pada seluruh isolat. Hasil amplifikasi gen gtf menggunakan primer spesifik menghasilkan amplikon berukuran sekitar 325 bp pada seluruh isolat. Berdasarkan penelitian ini dapat disimpulkan bahwa 16 isolat probiotik asal ASI terbukti memiliki gen gtf.

scholarly journals Reduction of Arcobacter at Two Conventional Wastewater Treatment Plants in Southern Arizona, USA

Pathogens ◽  
2019 ◽  
Vol 8 (4) ◽  
pp. 175 ◽  
Author(s):  
Ghaju Shrestha ◽  
Sherchan ◽  
Kitajima ◽  
Tanaka ◽  
Gerba ◽  
...  
Keyword(s):  
Wastewater Treatment ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Quantitative Pcr ◽  
Pathogenic Bacteria ◽  
Wastewater Treatment Plants ◽  
The United States ◽  
Rrna Gene ◽  
Metagenomic Sequencing ◽  
Wwtp Effluent

This study aimed to identify the bacterial community in two wastewater treatment plants (WWTPs) and to determine the occurrence and reduction of Arcobacter, along with virulence genes (ciaB and pldA). A total of 48 samples (24 influent and 24 effluent) were collected at two WWTPs in southern Arizona in the United States, monthly from August 2011 to July 2012. Bacterial DNA extract was utilized for 16S rRNA metagenomic sequencing. Quantification of Arcobacter 16S rRNA gene was conducted using a recently developed SYBR Green-based quantitative PCR assay. Among 847 genera identified, 113 (13%) were identified as potentially pathogenic bacteria. Arcobacter 16S rRNA gene was detected in all influent samples and ten (83%) and nine (75%) effluent samples at each plant, respectively. Log reduction ratios of Arcobacter 16S rRNA gene in Plant A and Plant B were 1.7 ± 0.9 (n = 10) and 2.3 ± 1.5 (n = 9), respectively. The ciaB gene was detected by quantitative PCR in eleven (92%) and twelve (100%) of 12 influent samples from Plant A and Plant B, respectively, while the pldA gene was detected in eight (67%) and six (50%) influent samples from Plant A and Plant B, respectively. The prevalence of potentially pathogenic bacteria in WWTP effluent indicated the need for disinfection before discharge into the environment.

scholarly journals Sensitive Identification of Bacterial DNA in Clinical Specimens by Broad-Range 16S rRNA Gene Enrichment

2020 ◽  
Vol 58 (12) ◽  
Author(s):  
Sara Rassoulian Barrett ◽  
Noah G. Hoffman ◽  
Christopher Rosenthal ◽  
Andrew Bryan ◽  
Desiree A. Marshall ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Nucleic Acids ◽  
Bacterial Species ◽  
False Negative ◽  
Case Series ◽  
Rrna Gene ◽  
Metagenomic Sequencing ◽  
Bacterial Dna ◽  
Clinical Specimens

ABSTRACT The broad-range detection and identification of bacterial DNA from clinical specimens are a foundational approach in the practice of molecular microbiology. However, there are circumstances under which conventional testing may yield false-negative or otherwise uninterpretable results, including the presence of multiple bacterial templates or degraded nucleic acids. Here, we describe an alternative, next-generation sequencing approach for the broad range detection of bacterial DNA using broad-range 16S rRNA gene hybrid capture (“16S Capture”). The method is able to deconvolute multiple bacterial species present in a specimen, is compatible with highly fragmented templates, and can be readily implemented when the overwhelming majority of nucleic acids in a specimen derive from the human host. We find that this approach is sensitive to detecting as few as 17 Staphylococcus aureus genomes from a background of 100 ng of human DNA, providing 19- to 189-fold greater sensitivity for identifying bacterial sequences than standard shotgun metagenomic sequencing, and is able to successfully recover organisms from across the eubacterial tree of life. Application of 16S Capture to a proof-of-principle case series demonstrated its ability to identify bacterial species that were consistent with histological evidence of infection, even when diagnosis could not be established using conventional broad range bacterial detection assays. 16S Capture provides a novel means for the efficient and sensitive detection of bacteria embedded in human tissues and for specimens containing highly fragmented template DNA.

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