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      Impact of the Gut Microbiome on the Progression of Hepatitis B Virus Related Acute-on-Chronic Liver Failure

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          Abstract

          The relationship between the progression of hepatitis B virus-related acute-on-chronic liver failure (HBV-ACLF) and the gut microbiota is poorly understood, and an HBV-ACLF-related microbiome has yet to be identified. In this study alterations in the fecal microbiome of 91 patients with HBV-ACLF (109 stool samples), including a cohort of nine patients at different stages of HBV-ACLF, were determined by high-throughput 16S rDNA sequencing. The operational taxonomic units and Shannon indexes indicated that the diversity and abundance of the gut microbiome significantly decreased with the progression of HBV-ACLF (p <0.05). The relative abundance of the Bacteroidetes phylum in the microbiome was significantly reduced, whereas the abundance of potentially pathogenic bacteria, such as Veilonella, Streptococcus, Enterococcus, and Klebsiella, was highly enriched in the HBV-ACLF group compared with the healthy control group. The abundance of Bacteroidetes was negatively correlated with the level of serum alpha fetoprotein, and the abundance of Veilonella was positively correlated with serum total bilirubin (TBIL). Furthermore, the abundance of Coprococcus was significantly negatively correlated with the level of serum TBIL and the international normalized ratio and positively correlated with prothrombin time activity. Our findings suggest that the gut microbiota plays an important role in the development of HBV-ACLF.

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          Most cited references38

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          UPARSE: highly accurate OTU sequences from microbial amplicon reads.

          Amplified marker-gene sequences can be used to understand microbial community structure, but they suffer from a high level of sequencing and amplification artifacts. The UPARSE pipeline reports operational taxonomic unit (OTU) sequences with ≤1% incorrect bases in artificial microbial community tests, compared with >3% incorrect bases commonly reported by other methods. The improved accuracy results in far fewer OTUs, consistently closer to the expected number of species in a community.
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            Evaluation of general 16S ribosomal RNA gene PCR primers for classical and next-generation sequencing-based diversity studies

            16S ribosomal RNA gene (rDNA) amplicon analysis remains the standard approach for the cultivation-independent investigation of microbial diversity. The accuracy of these analyses depends strongly on the choice of primers. The overall coverage and phylum spectrum of 175 primers and 512 primer pairs were evaluated in silico with respect to the SILVA 16S/18S rDNA non-redundant reference dataset (SSURef 108 NR). Based on this evaluation a selection of ‘best available’ primer pairs for Bacteria and Archaea for three amplicon size classes (100–400, 400–1000, ≥1000 bp) is provided. The most promising bacterial primer pair (S-D-Bact-0341-b-S-17/S-D-Bact-0785-a-A-21), with an amplicon size of 464 bp, was experimentally evaluated by comparing the taxonomic distribution of the 16S rDNA amplicons with 16S rDNA fragments from directly sequenced metagenomes. The results of this study may be used as a guideline for selecting primer pairs with the best overall coverage and phylum spectrum for specific applications, therefore reducing the bias in PCR-based microbial diversity studies.
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              Development of a dual-index sequencing strategy and curation pipeline for analyzing amplicon sequence data on the MiSeq Illumina sequencing platform.

              Rapid advances in sequencing technology have changed the experimental landscape of microbial ecology. In the last 10 years, the field has moved from sequencing hundreds of 16S rRNA gene fragments per study using clone libraries to the sequencing of millions of fragments per study using next-generation sequencing technologies from 454 and Illumina. As these technologies advance, it is critical to assess the strengths, weaknesses, and overall suitability of these platforms for the interrogation of microbial communities. Here, we present an improved method for sequencing variable regions within the 16S rRNA gene using Illumina's MiSeq platform, which is currently capable of producing paired 250-nucleotide reads. We evaluated three overlapping regions of the 16S rRNA gene that vary in length (i.e., V34, V4, and V45) by resequencing a mock community and natural samples from human feces, mouse feces, and soil. By titrating the concentration of 16S rRNA gene amplicons applied to the flow cell and using a quality score-based approach to correct discrepancies between reads used to construct contigs, we were able to reduce error rates by as much as two orders of magnitude. Finally, we reprocessed samples from a previous study to demonstrate that large numbers of samples could be multiplexed and sequenced in parallel with shotgun metagenomes. These analyses demonstrate that our approach can provide data that are at least as good as that generated by the 454 platform while providing considerably higher sequencing coverage for a fraction of the cost.
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                Author and article information

                Contributors
                Journal
                Front Cell Infect Microbiol
                Front Cell Infect Microbiol
                Front. Cell. Infect. Microbiol.
                Frontiers in Cellular and Infection Microbiology
                Frontiers Media S.A.
                2235-2988
                06 April 2021
                2021
                : 11
                : 573923
                Affiliations
                [1] 1 Department of Infectious Diseases, The Second Affiliated Hospital of Nanchang University , Nanchang, China
                [2] 2 Department of Infectious Diseases, Nanchang Centers for Disease Control and Prevention , Nanchang, China
                [3] 3 State Key Laboratory for Infectious Disease Prevention and Control, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention , Beijing, China
                [4] 4 Department of Pharmaceutics, Medical College of Jiaxing University , Jiaxing, China
                Author notes

                Edited by: Yi Xu, Texas A&M Health Science Center, United States

                Reviewed by: Yanfei Chen, Zhejiang University, China; Yousong Peng, Hunan University, China

                *Correspondence: Junrong Liang, liangjunrong@ 123456icdc.cn ; Baogang Xie, xiebaogang49@ 123456zjxu.edu.cn ; Shuilin Sun, sunshuilin2280@ 123456126.com

                This article was submitted to Microbiome in Health and Disease, a section of the journal Frontiers in Cellular and Infection Microbiology

                †These authors have contributed equally to this work

                Article
                10.3389/fcimb.2021.573923
                8056260
                33889550
                53450352-b6de-4049-9534-edfa411cd85e
                Copyright © 2021 Yao, Yu, Fan, Xiang, Long, Xu, Wu, Chen, Xi, Gao, Liu, Gong, Yang, Sun, Yu, Liang, Xie and Sun

                This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

                History
                : 18 June 2020
                : 26 February 2021
                Page count
                Figures: 5, Tables: 1, Equations: 0, References: 39, Pages: 12, Words: 4580
                Categories
                Cellular and Infection Microbiology
                Original Research

                Infectious disease & Microbiology
                fecal microbiome,hepatitis b virus,acute-on-chronic liver failure,biomedical indicators,metabolites

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