@article {151, title = {High-throughput sequencing of pooled samples to determine community-level microbiome diversity.}, journal = {Ann Epidemiol}, volume = {39}, year = {2019}, month = {2019 11}, pages = {63-68}, abstract = {

PURPOSE: Community-level interventions in cluster randomized controlled trials may alter the gut microbiome of individuals. The current method of estimating community diversities uses microbiome data obtained from multiple individual{\textquoteright}s specimens. Here we propose randomly pooling a number of microbiome samples from the same community into one sample before sequencing to estimate community-level microbiome diversity.

METHODS: We design and analyze an experiment to compare community microbiome diversity (gamma-diversity) estimates derived from 16S rRNA gene sequencing of 1) individually sequenced specimens vs. 2) pooled specimens collected from a community. Pool sizes of 10, 20, and 40 are considered. We then compare the gamma-estimates using Pearson{\textquoteright}s correlation as well as using Bland and Altman agreement analysis for three established diversity indices including richness, Simpson{\textquoteright}s and Shannon{\textquoteright}s.

RESULTS: The gamma-diversity estimates are highly correlated, with most being statistically significant. All correlations between all three diversity estimates are significant in the 10-pooled data. Pools comprising 40 specimens are closest to the line of agreement, but all pooled samples and individual samples fall within the 95\% limits of agreement.

CONCLUSIONS: Pooling microbiome samples before DNA amplification and metagenomics sequencing to estimate community-level diversity is a viable measure to consider in population-level association research studies.

}, keywords = {Administration, Oral, Anti-Bacterial Agents, Azithromycin, Bacteria, Biodiversity, Child, Preschool, Female, Gastrointestinal Microbiome, Gastrointestinal Tract, High-Throughput Nucleotide Sequencing, Humans, Infant, Infant, Newborn, Male, Metagenomics, Microbiota, Niger, Placebos, RNA, Ribosomal, 16S, Sequence Analysis, DNA}, issn = {1873-2585}, doi = {10.1016/j.annepidem.2019.09.002}, author = {Ray, Kathryn J and Cotter, Sun Y and Arzika, Ahmed M and Kim, Jessica and Boubacar, Nameywa and Zhou, Zhaoxia and Zhong, Lina and Porco, Travis C and Keenan, Jeremy D and Lietman, Thomas M and Doan, Thuy} } @article {221, title = {Mass Azithromycin Distribution and Community Microbiome: A Cluster-Randomized Trial.}, journal = {Open Forum Infect Dis}, volume = {5}, year = {2018}, month = {2018 Aug}, pages = {ofy182}, abstract = {

Background: Mass distributions of oral azithromycin have long been used to eliminate trachoma, and they are now being proposed to reduce childhood mortality. The observed benefit appears to be augmented with each additional treatment, suggesting a possible community-level effect. Here, we assess whether 2 biannual mass treatments of preschool children affect the community{\textquoteright}s gut microbiome at 6 months after the last distribution.

Methods: In this cluster-randomized controlled trial, children aged 1-60 months in the Dossa region of Niger were randomized at the village level to receive a single dose of azithromycin or placebo every 6 months. Fecal samples were collected 6 months after the second treatment for metagenomic deep sequencing. The prespecified primary outcome was the Euclidean PERMANOVA of the gut microbiome, or effectively the distance between the genus-level centroid at the community level, with the secondary outcome being the Simpson{\textquoteright}s α diversity.

Results: In the azithromycin arm, the gut microbial structures were significantly different than in the placebo arm (Euclidean PERMANOVA, \< .001). Further, the diversity of the gut microbiome in the azithromycin arm was significantly lower than in the placebo arm (inverse Simpson{\textquoteright}s index, = .005).

Conclusions: Two mass azithromycin administrations, 6 months apart, in preschool children led to long-term alterations of the gut microbiome structure and community diversity. Here, long-term microbial alterations in the community did not imply disease but were associated with an improvement in childhood mortality.

Clinical Trials Registration: NCT02048007.

}, issn = {2328-8957}, doi = {10.1093/ofid/ofy182}, author = {Doan, Thuy and Hinterwirth, Armin and Arzika, Ahmed M and Cotter, Sun Y and Ray, Kathryn J and O{\textquoteright}Brien, Kieran S and Zhong, Lina and Chow, Eric D and Zhou, Zhaoxia and Cummings, Susie L and Fry, Dionna and Oldenburg, Catherine E and Worden, Lee and Porco, Travis C and Keenan, Jeremy D and Lietman, Thomas M} } @article {276, title = {Gut Microbial Diversity in Antibiotic-Naive Children After Systemic Antibiotic Exposure: A Randomized Controlled Trial.}, journal = {Clin Infect Dis}, volume = {64}, year = {2017}, month = {2017 05 01}, pages = {1147-1153}, abstract = {

Background: Antibiotic exposure can alter the gut microbiome. We evaluate the effects of azithromycin on the gut microbiome diversity of children from an antibiotic-naive community in Niger.

Methods: A population-based sample of 80 children aged 1-60 months in the Dosso region of Niger was randomized to receive a single dose of either oral azithromycin or placebo. Fecal samples were collected immediately before treatment and 5 days after treatment for 16S rRNA gene sequencing. The prespecified outcome was α-diversity (inverse Simpson{\textquoteright}s α-diversity index), with secondary outcomes of β and γ Simpson{\textquoteright}s and Shannon{\textquoteright}s diversities.

Results: At 5 days after treatment, 40 children aged 1-60 months were analyzed in the azithromycin-treated group and 40 children in the placebo-treated group. Diversity of the gut microbiome was significantly lower in the treated group (inverse Simpson{\textquoteright}s α-diversity, 5.03; 95\% confidence interval [CI], 4.08-6.14) than in the placebo group (6.91; 95\% CI, 5.82-8.21; P = .03). Similarly, the Shannon{\textquoteright}s α-diversity was lower in the treated group (10.60; 95\% CI, 8.82-12.36) than the placebo group (15.42; 95\% CI, 13.24-17.80; P = .004). Simpson{\textquoteright}s community-level (γ) diversity decreased with azithromycin exposure from 17.72 (95\% CI, 13.80-20.21) to 10.10 (95\% CI, 7.80-11.40; P = .00008), although β-diversity was not significantly reduced (2.56, 95\% CI, 1.88-3.12; to 2.01, 95\% CI, 1.46-2.51; P = .26).

Conclusions: Oral administration of azithromycin definitively decreases the diversity of the gut microbiome of children in an antibiotic-naive community.

Clinical Trials Registration: NCT02048007.

}, keywords = {Administration, Oral, Anti-Bacterial Agents, Azithromycin, Bacteria, Biodiversity, Child, Preschool, Cluster Analysis, DNA, Ribosomal, Female, Gastrointestinal Microbiome, Gastrointestinal Tract, Humans, Infant, Male, Niger, Phylogeny, Placebos, RNA, Ribosomal, 16S, Sequence Analysis, DNA}, issn = {1537-6591}, doi = {10.1093/cid/cix141}, author = {Doan, Thuy and Arzika, Ahmed M and Ray, Kathryn J and Cotter, Sun Y and Kim, Jessica and Maliki, Ramatou and Zhong, Lina and Zhou, Zhaoxia and Porco, Travis C and Vanderschelden, Benjamin and Keenan, Jeremy D and Lietman, Thomas M} }