DOI: 10.5061/dryad.zkh1893m9
title: Fecal microbiota transplants (FMT) of three distinct human communities to germ-free mice exacerbated inflammation and decreased lung function in their offspring

description: Despite explosive rise in allergies, little is known about early life gut
 microbiota effects on postnatal respiratory function. We hypothesized that
 Enterobacteriaceae-dominant gut microbiota from eczemic infants increases
 Type 2 inflammation and decreases lung function in transplanted mice,
 while Bacteroidaceae-dominant gut microbiota from non-eczemic infants is
 protective. FMT from eczemic infants “Infant A” and non-eczemic infants
 “Infant B” were successfully transplanted into germ-free C57BL/6 mice
 passing to offspring unchanged. Infant A and B, Adult C-human-derived
 (positive control), and mouse (negative control) microbiotas all in
 C57BL/6 mice were tested for effects on airway function in non-allergic
 (PBS) and allergic (HDM) conditions. Baseline lung mechanics in mice with
 human microbiotas (HUmicrobiota) were significantly impaired compared to
 mouse microbiota controls (MOmicrobiota) with or without HDM; Respiratory
 system resistance (Rrs) was increased (p&lt; 0.05-p&lt;0.01) and
 Respiratory system compliance (Crs) was decreased
 (p&lt;0.05-p&lt;0.01). HUmicrobiota mice showed statistically
 significant impairment compared to MOmicrobiota mice in lung parameters
 Rrs, Ers, Rn, and G at baseline, and at multiple Mch doses with baseline
 removed. Impairment manifested as increased small airway resistance and
 tissue resistance. HDM significantly elevated IL-4, eosinophils, lung
 inflammation and mucus cell metaplasia and decreased macrophages and lung
 function (p&lt;0.05) in mice of all microbiotas, yet each HUmicrobiota
 produced distinct features. Infant B and Adult C mice had elevated basal
 levels of total IgE compared to MOmicrobiota and Infant A mice
 (p&lt;0.05). In HUmicrobiota mice given HDM, only Adult C had elevated
 IL-5 and IL-13 (p&lt;0.05), only Adult C and Infant B mice had
 elevated neutrophils (p&lt;0.05) and only Infant A had elevated
 lymphocytes (p&lt;0.01).
description: Fecal pellets of C57BL/6 germ-free mice receiving transplant of
 fecal materials collected from 3-month-old infants of the Isle of Wight
 third generation birth cohort were collected at the endpoint of the
 experiment. Bacterial DNA was isolated using the FastDNA SPIN Kit then
 sequeced for the 16S rRNA V4 region using Illumina MiSeq using MiSeq v2
 flow cell and 500 cycle MiSeq v2 reagen kit with paired-end 250 base
 reads. Base calling was performed by Illumina Real Time Analysis (RTA)
 v1.18.54 and output of RTA was de-multiplexed and converted to FastQ
 format files with Illumina Bcl2fastq v1.8.4. 109,085 high-quality reads
 from Illumina sequencing remained - average read number per group: 3896
 (min 3245, max 5053). 
description: # Fecal microbiota transplants (FMT) of three distinct human communities
 to germ-free mice exacerbated inflammation and decreased lung function in
 their offspring
 [https://doi.org/10.5061/dryad.zkh1893m9](https://doi.org/10.5061/dryad.zkh1893m9) ## Description of the data and file structure All sequence files are of fecal pellets of C57BL/6 germ-free mice receiving transplant of fecal materials collected from 3-month-old infants of the Isle of Wight third generation birth cohort were collected at the endpoint of the experiment. Zymo mock community positive control was used as a sequencing positive control. Bacterial DNA was isolated using the FastDNA SPIN Kit then sequenced for the 16S rRNA V4 region using Illumina MiSeq using MiSeq v2 flow cell and 500 cycle MiSeq v2 reagen kit with paired-end 250 base reads. Base calling was performed by Illumina Real Time Analysis (RTA) v1.18.54 and output of RTA was de-multiplexed and converted to FastQ format files with Illumina Bcl2fastq v1.8.4. 109,085 high-quality reads from Illumina sequencing remained - average read number per group: 3896 (min 3245, max 5053).  These raw fasta files were processed using QIIME2 (v. 2019.1) using UCHIME for eukaryota sequence removal, and Silva for taxonomic annotation reference. Sequence clustering of 97% was used and chimera sequences were removed, 128 OTUs remaining for data analysis completed in PAST and R.  ### Files and variables #### File: FMT\_germ\_free\_mice\_fecal\_microbiota.zip **Description:**  178 *.fastq.gz files: gzipped fasta files. First 2-5 digit letters and numbers indicate sample ID's. There are R1 and R2 for each of the samples as they are paired end reads. Perform `gunzip` to unzip .fastq.gz files, then you can open and read the .fastq files using a text editor of your choice.  FastQC: a directory containing QC info ## Code/software These raw fasta files were processed using QIIME2 (v. 2019.1) using UCHIME for eukaryota sequence removal, and Silva for taxonomic annotation reference. Sequence clustering of 97% was used and chimera sequences were removed, 128 OTUs remaining for data analysis completed in PAST and R.