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      Microbiota of newborn calves and their mothers reveals possible transfer routes for newborn calves’ gastrointestinal microbiota

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          Abstract

          The intestinal microbiota of newborns plays an important role in the development of immunity and metabolism. In livestock animals, knowledge of the intestinal microbiota is essential not only to prevent diseases but also to optimize weight gain and performance. The aim of our study was to examine faecal samples repeatedly within the first two days of life using 16S rRNA gene High Throughput Sequencing. Additionally, samples from the mouths of the calves and the vaginas, colostrum, and faeces of the dams were included to evaluate possible sources of the calf faecal microbiota. The calf faecal microbiota was highly variable during the first 48 hours post natum ( p. n.). Significant changes were found in species diversity and richness, in copy numbers evaluated by qPCR and in predominant bacteria over time. The most pronounced changes occurred between 6 and 24 hours p. n. All calf faecal samples were dominated by Operational Taxonomic Units (OTUs) belonging to the family Enterobacteriaceae. Cow faecal samples showed significantly higher species richness, diversity, number of observed OTUs, and copy numbers compared to all other samples. OTUs belonging to the family Ruminococcaceae were most abundant in cow faecal and vaginal samples. Colostrum was dominated by Enhydrobacter affiliated OTUs. To identify possible inoculation routes for the calf microbiota, we analysed OTU sharing between samples. The calf microbiota during the first two days of life was clearly distinct from the dam’s faecal microbiota. Furthermore, colostrum microbiota clearly differed from calf and cow faecal microbiota and thus most likely does not play an important role as inoculation source for calf microbiota during the first two days of life. In contrast, the cow vaginal and the calf faecal microbiota were more similar, suggesting that some of the calf faecal microbiota may derive from inoculation from the birth canal during birth.

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          FLASH: fast length adjustment of short reads to improve genome assemblies.

          Next-generation sequencing technologies generate very large numbers of short reads. Even with very deep genome coverage, short read lengths cause problems in de novo assemblies. The use of paired-end libraries with a fragment size shorter than twice the read length provides an opportunity to generate much longer reads by overlapping and merging read pairs before assembling a genome. We present FLASH, a fast computational tool to extend the length of short reads by overlapping paired-end reads from fragment libraries that are sufficiently short. We tested the correctness of the tool on one million simulated read pairs, and we then applied it as a pre-processor for genome assemblies of Illumina reads from the bacterium Staphylococcus aureus and human chromosome 14. FLASH correctly extended and merged reads >99% of the time on simulated reads with an error rate of <1%. With adequately set parameters, FLASH correctly merged reads over 90% of the time even when the reads contained up to 5% errors. When FLASH was used to extend reads prior to assembly, the resulting assemblies had substantially greater N50 lengths for both contigs and scaffolds. The FLASH system is implemented in C and is freely available as open-source code at http://www.cbcb.umd.edu/software/flash. t.magoc@gmail.com.
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            Global patterns of 16S rRNA diversity at a depth of millions of sequences per sample.

            The ongoing revolution in high-throughput sequencing continues to democratize the ability of small groups of investigators to map the microbial component of the biosphere. In particular, the coevolution of new sequencing platforms and new software tools allows data acquisition and analysis on an unprecedented scale. Here we report the next stage in this coevolutionary arms race, using the Illumina GAIIx platform to sequence a diverse array of 25 environmental samples and three known "mock communities" at a depth averaging 3.1 million reads per sample. We demonstrate excellent consistency in taxonomic recovery and recapture diversity patterns that were previously reported on the basis of metaanalysis of many studies from the literature (notably, the saline/nonsaline split in environmental samples and the split between host-associated and free-living communities). We also demonstrate that 2,000 Illumina single-end reads are sufficient to recapture the same relationships among samples that we observe with the full dataset. The results thus open up the possibility of conducting large-scale studies analyzing thousands of samples simultaneously to survey microbial communities at an unprecedented spatial and temporal resolution.
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              Fecal Microbial Diversity in Pre-Weaned Dairy Calves as Described by Pyrosequencing of Metagenomic 16S rDNA. Associations of Faecalibacterium Species with Health and Growth

              In this study, we use barcoded pyrosequencing of the 16S rRNA gene to characterize the fecal microbiota of neonatal calves and identify possible relationships of certain microbiota profiles with health and weight gain. Fecal samples were obtained weekly from 61 calves from birth until weaning (seventh week of the calves' life). Firmicutes was the most prevalent phylum, with a prevalence ranging from 63.84% to 81.90%, followed by Bacteroidetes (8.36% to 23.93%), Proteobacteria (3.72% to 9.75%), Fusobacteria (0.76% to 5.67%), and Actinobacteria (1.02% to 2.35%). Chao1 index gradually increased from the first to the seventh postnatal week. Chao1 index was lower during the third, fourth, and fifth week of life in calves that suffered from pneumonia and were treated with antibiotics. Diarrhea incidence during the first four weeks of the calves' life was also associated with a reduction of microbial diversity during the third week of life. Increased fecal microbial diversity after the second week of life was associated with higher weight gain. Using discriminant analysis we were able to show differences in the microbiota profiles between different weeks of life, between high and low weight gain groups of calves, and between calves affected and not affected with diarrhea during the first four weeks life. The prevalence of Faecalibacterium spp. in the first week of life was associated with weight gain and the incidence of diarrhea, with higher prevalence being associated with higher weight gain and less diarrhea. Representative sequences from Faecalibacterium spp. were closely affiliated to Faecalibacterium prausnitzii. Results presented here provide new information regarding the intestinal microbiota of neonatal calves and its association with health and growth. Fecal microbial diversity was associated with calf age, disease status and growth rates. Results suggesting a possible beneficial effect of Faecalibacterium spp. on health and growth are promising.
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                Author and article information

                Contributors
                Role: ConceptualizationRole: Formal analysisRole: Funding acquisitionRole: InvestigationRole: Project administrationRole: ResourcesRole: SupervisionRole: VisualizationRole: Writing – original draftRole: Writing – review & editing
                Role: Data curationRole: Formal analysisRole: InvestigationRole: MethodologyRole: SoftwareRole: VisualizationRole: Writing – review & editing
                Role: Data curationRole: Formal analysisRole: InvestigationRole: MethodologyRole: ResourcesRole: SupervisionRole: Writing – review & editing
                Role: Data curationRole: InvestigationRole: Methodology
                Role: ResourcesRole: SupervisionRole: Writing – review & editing
                Role: ResourcesRole: SupervisionRole: Writing – review & editing
                Role: ConceptualizationRole: Formal analysisRole: MethodologyRole: Project administrationRole: ResourcesRole: SoftwareRole: SupervisionRole: ValidationRole: VisualizationRole: Writing – original draftRole: Writing – review & editing
                Role: Data curationRole: Formal analysisRole: InvestigationRole: MethodologyRole: Project administrationRole: ResourcesRole: SoftwareRole: SupervisionRole: ValidationRole: VisualizationRole: Writing – original draftRole: Writing – review & editing
                Role: Editor
                Journal
                PLoS One
                PLoS ONE
                plos
                plosone
                PLoS ONE
                Public Library of Science (San Francisco, CA USA )
                1932-6203
                1 August 2019
                2019
                : 14
                : 8
                : e0220554
                Affiliations
                [1 ] Department for Farm Animals and Veterinary Public Health, Clinical Unit for Herd Health Management, University Clinic for Ruminants, University of Veterinary Medicine Vienna, Vienna, Austria
                [2 ] Laboratory of Molecular Biology and Microbiology, Instituto Tecnológico Agrario de Castilla y León, Valladolid, Spain
                [3 ] Department for Farm Animals and Veterinary Public Health, Institute of Milk Hygiene, Milk Technology and Food Science, University of Veterinary Medicine Vienna, Vienna, Austria
                [4 ] Department of Animal Science, Iowa State University, Ames, Iowa, United States of America
                Wageningen University, NETHERLANDS
                Author notes

                Competing Interests: The authors have declared that no competing interests exist.

                Author information
                http://orcid.org/0000-0001-5806-7601
                http://orcid.org/0000-0002-0972-6241
                http://orcid.org/0000-0002-2253-5247
                Article
                PONE-D-18-35697
                10.1371/journal.pone.0220554
                6675284
                31369600
                fe7a0195-b760-4247-8434-bf855af60064
                © 2019 Klein-Jöbstl et al

                This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

                History
                : 14 December 2018
                : 18 July 2019
                Page count
                Figures: 5, Tables: 2, Pages: 18
                Funding
                Funded by: start-up project (Profillinie 2) by the University of Veterinary Medicine Vienna
                Award Recipient :
                Funded by: Ph.D. studentship from the Spanish National Institute for Agriculture and Food Research and Technology (INIA) cofinanced by the European Social Fund
                Award ID: FPI2014-020
                Award Recipient :
                Funded by: COST action
                Award ID: STSM-FA1202
                Award Recipient :
                The authors received funding from: 1. start-up project (Profillinie 2) by the University of Veterinary Medicine Vienna; (no award number available); Recipient Daniela Klein-Jöbstl; 2. Ph.D. studentship from the Spanish National Institute for Agriculture and Food Research and Technology (INIA) cofinanced by the European Social Fund; number: FPI2014-020; Recipient Narciso M. Quijada; and 3. COST action; STSM-FA1202; Recipient Narciso M. Quijada.
                Categories
                Research Article
                Biology and Life Sciences
                Microbiology
                Medical Microbiology
                Microbiome
                Biology and Life Sciences
                Genetics
                Genomics
                Microbial Genomics
                Microbiome
                Biology and Life Sciences
                Microbiology
                Microbial Genomics
                Microbiome
                Biology and Life Sciences
                Anatomy
                Reproductive System
                Genital Anatomy
                Vagina
                Medicine and Health Sciences
                Anatomy
                Reproductive System
                Genital Anatomy
                Vagina
                Biology and Life Sciences
                Ecology
                Ecological Metrics
                Species Diversity
                Ecology and Environmental Sciences
                Ecology
                Ecological Metrics
                Species Diversity
                Biology and Life Sciences
                Anatomy
                Digestive System
                Gastrointestinal Tract
                Medicine and Health Sciences
                Anatomy
                Digestive System
                Gastrointestinal Tract
                Biology and Life Sciences
                Organisms
                Bacteria
                Gut Bacteria
                Ruminococcus
                Biology and Life Sciences
                Developmental Biology
                Neonates
                Biology and Life Sciences
                Organisms
                Bacteria
                Enterobacteriaceae
                Biology and life sciences
                Biochemistry
                Nucleic acids
                RNA
                Non-coding RNA
                Ribosomal RNA
                Biology and life sciences
                Biochemistry
                Ribosomes
                Ribosomal RNA
                Biology and life sciences
                Cell biology
                Cellular structures and organelles
                Ribosomes
                Ribosomal RNA
                Custom metadata
                All relevant data are within the paper and its Supporting Information files. The sequencing data now have been made public with all associated data. Data were submitted to the European Nucleotide Archive (ENA) and can be found under the accession number PRJEB29277 ( http://www.ebi.ac.uk/ena/data/view/PRJEB29277).

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