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Gut microbiota
muscle turnover, creatine, 3-methylhistidine and L-valine, and host metabolism, carnitine, are elevated in the athlete groups. Metabolites derived from vitamins and recovery supplements common in professional sports, including glutamine, lysine, 4- pyridoxic acid and nicotinamide, are also raised in the athlete group. It is notable that PAG, a microbial conversion product of phenylalanine, has been associated with a lean phenotype and is increased in the athletes. 42 Furthermore, PAG positively corre- lates with the genus Erysipelotrichaceae incertae sedis , whichwe have previously noted to be present in relatively higher propor- tions in the athlete group compared with both control groups. PAG is the strongest biomarker postbariatric surgery, where it is associated with an increase in the relative proportions of Proteobacteria as observed here in the athlete group. Within the SCFAs, two distinct clusters were observed; acetic acid, propio- nic acid and butyric acid correlate with dietary contributors ( fi bre and protein), while isobutyric acid, isovaleric acid and valeric acid correlate with microbial diversity. The same clusters are observed when correlating with individual taxa, in support of previously observed links between SCFAs and numerous metabolic bene fi ts and a lean phenotype. 33 – 35 Our ongoing work in this area with non-athletes engaging in a structured exercise regime looks to further explore compo- nents of the exercise and diet – microbiome paradigm, which, along with this study, may inform the design of exercise and fi tness programmes, including diet design in the context of opti- mising microbiota functionality for both athletes and the general population.
It was noted that athletes excreted proportionately higher levels of the metabolite TMAO, an end product metabolite of dietary protein degradation. Elevated TMAO has been observed in patients with cardiovascular disease and atheroscler- osis, highlighting a potential downside to increased protein intake. 15 – 17 22 37 However, TMAO is also found in high levels in the urine of Japanese populations, 38 who do not have high risk for CVD. Similarly to these populations, the athletes ’ diet contained a signi fi cantly greater proportion of fi sh. Our current understanding of the implications of this result remains limited and requires elaboration in future studies. Furthermore, pathway abundance in a metagenome merely re fl ects functional potential and not necessarily increased expression in situ. Variance of metagenomic composition between athletes and controls was exempli fi ed with unique pathway – pathway correla- tions between the two groups. Analysis of categorically arranged pathway abundances within the separate cohorts provided add- itional insight into the previously described dichotomy between the microbiota of athletes and high-BMI controls. The two groups displayed distinct structures of functional capacity, separ- ately oriented to operate under the different physiological milieu of the two groups. Notably, from a functional perspec- tive, the microbiota of the low-BMI group was more similar to the athletes. The low-BMI controls were generally engaged in a modestly active lifestyle, re fl ected by their leanness and increased levels of CK. It is speculative but not implausible that moderate improvements in physical activity for overweight and obese individuals may confer the bene fi cial metabolic functions observed within the athlete microbiome. Dietary contributions to the functional composition of the enteric microbial system are also evident in our study. The rela- tive abundances of pathways related to fundamental metabolic function — AAB, VB and LB — were higher on average within the high-BMI control group when compared with the athlete group. The mechanisms behind these differences are unclear and might re fl ect chronic adaptation of the athlete gut microbiome; pos- sibly due to a reduced reliance on the corresponding biosyn- thetic capacities of their gut microbiota. On the contrary, the athlete microbiome presents a functional capacity that is primed for tissue repair and to harness energy from the diet with increased capacity for carbohydrate, cell structure and nucleo- tide biosynthesis, re fl ecting the signi fi cant energy demands and high cell-turnover evident in elite sport. Remarkably, our examination of pathway correlation to dietary macronutrients and plasma CK, as a biomarker of exer- cise, 39 is suggestive of an impact of physical activity on the use of dietary nutrients by the microbiota of the gut. Comparing athletes to both high-BMI and low-BMI controls, a greater number of pathways correlating to speci fi c macronutrients with the controls suggests a shift in the dynamics of these varied metabolic functions. The impact of the athletes ’ increased protein intake compared with both control groups was evident in the metabolomic phenotyping results. By-products of dietary protein metabolism (mostly by microbes) including TMAO, carnitines, TMA, 3-CMPF and 3-hydroxy-isovaleric acid are all elevated in the athlete cohort. Of particular interest is 3-hydroxy-isovaleric acid (potentially from egg consumption), which has been demonstrated to have ef fi cacy for inhibiting muscle wasting when used in conjunction with physical exercise. 40 41 The compound is also commonly used as a sup- plement by athletes to increase exercise-induced gains in muscle size, muscle strength and lean body mass, reduce exercise-induced muscle damage and speed recovery from high- intensity exercise. 41 Numerous metabolites associated with
MATERIALS AND METHODS Study population
Elite professional male athletes (n=40) and healthy controls (n=46) matched for age and gender were enrolled in 2011 as previously described in the study. 26 Due to the range of physi- ques within a rugby team (player position dictates need for a variety of physical constitutions, ie, forward players tend to have larger BMI values than backs, often in the overweight/ obese range) the recruited control cohort was subdivided into two groups. To more completely include control participants, the BMI parameter for group inclusion was adjusted to BMI ≤ 25.2 and BMI ≥ 26.5 for the low-BMI and high-BMI groups, respectively. Approval for this study was granted by the Cork Clinical Research Ethics Committee. Acquisition of clinical, exercise and dietary data Self-reported dietary intake information was accommodated by a research nutritionist within the parameters of a food frequency questionnaire in conjunction with a photographic food atlas as per the initial investigation. 26 Fasting blood samples were col- lected and analysed at the Mercy University Hospital clinical laboratories, Cork. As the athletes were involved in a rigorous training camp, we needed to assess the physical activity levels of both control groups. To determine this, we used an adapted version of the EPIC-Norfolk questionnaire. 43 Creatine kinase levels were used as a proxy for level of physical activity across all groups. Preparation of metagenomic libraries DNA derived from faecal samples was extracted and puri fi ed using the QIAmp DNA Stool Mini Kit (cat. no 51 504) prior to storage at − 80°C. DNA libraries were prepared with the Nextera XT DNA Library Kit (cat. no FC-131-1096) prior to processing on the Illumina HiSeq 2500 sequencing platform (see online supplementary methods for further detail).
BartonW, et al . Gut 2017; 0 :1 – 9. doi:10.1136/gutjnl-2016-313627
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