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Gut microbiome research has increased dramatically in the last decade, including in renal health and disease. The field is moving from experiments showing mere association to causation using both forward and reverse microbiome approaches, leveraging tools such as germ-free animals, treatment with antibiotics, and fecal microbiota transplantations. However, we are still seeing a gap between discovery and translation that needs to be addressed, so that patients can benefit from microbiome-based therapies. In this guideline paper, we discuss the key considerations that affect the gut microbiome of animals and clinical studies assessing renal function, many of which are often overlooked, resulting in false-positive results. For animal studies, these include suppliers, acclimatization, baseline microbiota and its normalization, littermates and cohort/cage effects, diet, sex differences, age, circadian differences, antibiotics and sweeteners, and models used. Clinical studies have some unique considerations, which include sampling, gut transit time, dietary records, medication, and renal phenotypes. We provide best-practice guidance on sampling, storage, DNA extraction, and methods for microbial DNA sequencing (both 16S rRNA and shotgun metagenome). Finally, we discuss follow-up analyses, including tools available, metrics, and their interpretation, and the key challenges ahead in the microbiome field. By standardizing study designs, methods, and reporting, we will accelerate the findings from discovery to translation and result in new microbiome-based therapies that may improve renal health.

Alterations in gut homeostasis may contribute to the progression of diabetic nephropathy. There has been recent attention on the renoprotective effects of metabolite-sensing receptors in chronic renal injury, including the G protein-coupled receptor (GPR)109a, which ligates the short-chain fatty acid butyrate. However, the role of GPR109a in the development of diabetic nephropathy, a milieu of diminished microbiome-derived metabolites, has not yet been determined. The present study aimed to assess the effects of insufficient GPR109a signaling, via genetic deletion of GPR109a, on the development of renal injury in diabetic nephropathy. Gpr109a -/- mice or their wild-type littermates ( Gpr109a +/+ ) were rendered diabetic with streptozotocin. Mice received a control diet or an isocaloric high-fiber diet (12.5% resistant starch) for 24 wk, and gastrointestinal permeability and renal injury were determined. Diabetes was associated with increased albuminuria, glomerulosclerosis, and inflammation. In comparison, Gpr109a -/- mice with diabetes did not show an altered renal phenotype. Resistant starch supplementation did not afford protection from renal injury in diabetic nephropathy. While diabetes was associated with alterations in intestinal morphology, intestinal permeability assessed in vivo using the FITC-dextran test was unaltered. GPR109a deletion did not worsen gastrointestinal permeability. Furthermore, 12.5% resistant starch supplementation, at physiological concentrations, had no effect on intestinal permeability or morphology. The results of this study indicate that GPR109a does not play a critical role in intestinal homeostasis in a model of type 1 diabetes or in the development of diabetic nephropathy.