Early life exercise and age altered beta diversity.

<p>Principle coordinates analysis (PCoA) using unweighted UniFrac distances with an explicit time axis depicts clustering of microbial communities due to age after three days (3d) and six weeks (6 wk) of exercise and 25 days following exercise cessation (25d post). After 6 wk, a clear clustering of juvenile run versus juvenile sed samples is noticeable.</p

Effects of age of exercise onset on body composition across the lifespan.

<p>A) Weekly total running distance across time for the separate cohort used for chemical carcass analyses. Adult runners ran less toward the end of exercise, whereas running distance steadily increased for juvenile runners throughout the duration of exercise. Total distance summed across six weeks did not differ by age. B-D depict body weight, lean mass and fat mass, respectively, for juvenile and adult run and sed chemical carcass rats after six weeks of exercise (6 wk) or 25 days following exercise cessation (25d post). B) Body weight; adult runners weighed less than adult seds at 6 wk, however they returned to sedentary levels 25d post. In contrast, juvenile onset runners consistently weighed more than their sedentary counterparts at both time points. C) Lean mass; juvenile onset runners had sustained increases in lean mass at both time points. D) Fat mass; sustained decreases in fat mass were observed in adult onset runners only. Data are represented as mean ± SEM; *p<0.05.</p

Early life exercise and age altered alpha diversity.

<p>Measures of alpha diversity for adult and juvenile run and sed rats after three days (3d) and six weeks (6 wk) of wheel running, and 25 days following exercise cessation (25d post). A) Shannon entropy, an indicator of an even community structure, was significantly higher in the adults than juveniles. Juvenile runners displayed decreased Shannon entropy overall and at 6 wk. B) Species richness was significantly higher in the adults relative to juveniles. Runners had significantly fewer species overall than their sedentary counterparts, and juvenile runners had significantly fewer species than juvenile sedentary rats 3d following the start of exercise. Data are represented as mean ± SEM; *p<0.05.</p

Exercise Is More Effective at Altering Gut Microbial Composition and Producing Stable Changes in Lean Mass in Juvenile versus Adult Male F344 Rats

<div><p>The mammalian intestine harbors a complex microbial ecosystem that influences many aspects of host physiology. Exposure to specific microbes early in development affects host metabolism, immune function, and behavior across the lifespan. Just as the physiology of the developing organism undergoes a period of plasticity, the developing microbial ecosystem is characterized by instability and may also be more sensitive to change. Early life thus presents a window of opportunity for manipulations that produce adaptive changes in microbial composition. Recent insights have revealed that increasing physical activity can increase the abundance of beneficial microbial species. We therefore investigated whether six weeks of wheel running initiated in the juvenile period (postnatal day 24) would produce more robust and stable changes in microbial communities versus exercise initiated in adulthood (postnatal day 70) in male F344 rats. 16S rRNA gene sequencing was used to characterize the microbial composition of juvenile versus adult runners and their sedentary counterparts across multiple time points during exercise and following exercise cessation. Alpha diversity measures revealed that the microbial communities of young runners were less even and diverse, a community structure that reflects volatility and malleability. Juvenile onset exercise altered several phyla and, notably, increased Bacteroidetes and decreased Firmicutes, a configuration associated with leanness. At the genus level of taxonomy, exercise altered more genera in juveniles than in the adults and produced patterns associated with adaptive metabolic consequences. Given the potential of these changes to contribute to a lean phenotype, we examined body composition in juvenile versus adult runners. Interestingly, exercise produced persistent increases in lean body mass in juvenile but not adult runners. Taken together, these results indicate that the impact of exercise on gut microbiota composition as well as body composition may depend on the developmental stage during which exercise is initiated.</p></div

Running distance and body weight.

<p>A) Weekly total running distance across six weeks of exercise, estimated per rat. Adults ran more in the first half of exercise, whereas juveniles ran more during the second half of exercise, although total distance summed across six weeks did not differ between age groups. B) Body weight across the duration of the experiment; adult runners weighed less than their sedentary counterparts during exercise, then returned to sedentary levels shortly following exercise cessation. Juvenile runners weighed more than their sedentary counterparts toward the end of exercise and continued to weigh more after exercise cessation. Data are represented as mean ± SEM; *p<0.05.</p

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Placing changes in the microbiome in the context of the American Gut. We accumulated samples over sequencing runs to demonstrate the structural consistency in the data. We demonstrate that while the ICU dataset (https://www.ncbi.nlm.nih.gov/pubmed/27602409) falls within the American Gut samples, they do not fall close to most samples at any of the body sites. We then highlight samples from the United Kingdom, Australia, the United States and other countries to show that nationality does not overcome the variation in body site. We then highlight the utility of the American Gut in meta-analysis by reproducing results from (https://www.ncbi.nlm.nih.gov/pubmed/20668239) and (https://www.ncbi.nlm.nih.gov/pubmed/23861384), using the AGP dataset as the context for dynamic microbiome changes instead of the HMP dataset. We show rapid, complete recovery of C. diff patients following fecal material transplantation and also contextualized the change in an infant gut over time until it settles into an adult state. This demonstrates the power of the American Gut dataset, both as a cohesive study and as a context for other investigations

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The SEPP (Mirarab et al Pac Symp Biocomput 2012) fragment insertion tree used for phylogenetic analyses

Unweighted UniFrac distances

The unweighted UniFrac distance (Lozupone and Knight AEM 2005) matrix of the 9511 fecal samples used in the American Gut paper. UniFrac was computed using Striped UniFrac (https://github.com/biocore/unifrac). Prior to execution of UniFrac, Deblur (Amir et al mSystems 2017) was run on the samples, all bloom sOTUs were removed (Amir et al mSystems 2017), and samples were rarefied to a depth of 1250 reads (Weiss et al Microbiome 2017). For the phylogeny, fragments were inserted using SEPP (Mirarab et al Pac Symp Biocomput 2012) into the Greengenes 13_5 99% OTU tree (McDonald et al ISME 2012)

American Gut Project fecal sOTU counts table

The Deblur sOTU counts table for the fecal samples used in the American Gut Project manuscript. The samples were trimmed to a common read length of 125nt, and processed by Deblur (Amir et al mSystems 2017). Blooms were removed (Amir et al mSystems 2017) and any sample with fewer than 1250 sequences was omitted. This table is not rarefied,

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Longitudinal samples from a large bowel resection. We place longitudinal samples collected prior to and following a large bowel resection in the context of samples from the AGP, the Earth Microbiome Project (https://www.ncbi.nlm.nih.gov/pubmed/29088705), intensive care unit patients (https://www.ncbi.nlm.nih.gov/pubmed/27602409), "extreme" diet samples from (https://www.ncbi.nlm.nih.gov/pubmed/24336217), and samples from the Hadza hunter-gatherers (https://www.ncbi.nlm.nih.gov/pubmed/28839072). Unweighted UniFrac was computed on this sample set, and principal coordinates were assessed. Using EMPeror (https://www.ncbi.nlm.nih.gov/pubmed/24280061), we then animate the plot by connect successive data points gut resection time series, while rotating the data frame. We first show the how the extent of change in the microbial community, and how the samples immediately following surgery resemble fecal samples from ICU patients. In the background of the animation, a black line connects a plant rhizosphere sample to a marine sediment sample, which have the same unweighted UniFrac distance (0.78) as the longitudinal sample immediately preceding and immediately following surgery