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ObjectiveTo delineate the role of the urinary metabolome in the genesis of urinary stone disease (USD).MethodsUntargeted metabolomics was utilized in comparative analyses of calcium-based stones (CBS) and spot urine samples from patients with a history of USD with or without urinary stone activity based on radiologic imaging. Stone and urine metabolomes were stratified by composition and radiographic stone-activity, respectively. Additionally, we quantified highly abundant metabolites that were present in either calcium oxalate (CaOx) or calcium phosphate (CaPhos) stones and also significantly enriched in the urine of active stone formers (SF) compared to non-active SF. These data were used to delineate either a direct involvement of urinary metabolites in lithogenesis or the passive uptake of biomolecules within the stone matrix.ResultsUrinary metabolomes were distinct based on radiographic stone-activity and the 2 types of CBS. Stratification by radiologic stone activity was driven by the enrichment of 14 metabolites in the urine of active SF that were also highly abundant in both CaOx and CaPhos stones, indicative of a potential involvement of these metabolites in lithogenesis. Using the combination of these 14 metabolites in total, we generated a model that correctly classified patients as either active vs non-active SF in a prospectively recruited cohort with 73% success.ConclusionCollectively, our data suggest specific urinary metabolites directly contribute to the formation of urinary stones and that active SF may excrete higher levels of lithogenic metabolites than non-active patients. Future studies are needed to confirm these findings and establish the causative mechanisms associated with these metabolites.

To determine whether functionally relevant questions associated with the urinary or gut microbiome and urinary stone disease (USD) can be answered from metagenome-wide association studies (MWAS), we performed the most comprehensive meta-analysis of published clinical MWAS in USD to date, using publicly available data published prior to April 2021. Six relevant studies met inclusion criteria. For alpha-diversity, significant differences were noted between USD status, stone composition, sample type, study location, age, diet, and sex. For beta-diversity, significant differences were noted by USD status, stone composition, sample type, study location, antibiotic use (30 days and 12 months before sampling), sex, hypertension, water intake, body habitus, and age. Prevotella and Lactobacillus in the gut and urinary tract, respectively, were associated with healthy individuals, while Enterobacteriaceae was associated with USD in the urine and stones. Paradoxically, other Prevotella strains were also strongly associated with USD in the gut microbiome. When data were analyzed together, USD status, stone composition, age group, and study location were the predominant factors associated with microbiome composition. Meta-analysis showed significant microbiome differences based on USD status, stone composition, age group or study location. However, analyses were limited by a lack of public data from published studies, metadata collected, and differing study protocols. Results highlight the need for field-specific standardization of experimental protocols in terms of sample collection procedures and the anatomical niches to assess, as well as in defining clinically relevant metadata and subphenotypes such as stone composition. IMPORTANCE Studies focused on the microbiome broadly support the hypothesis that the microbiome influences the onset of chronic diseases such as urinary stone disease. However, it is unclear what environmental factors shape the microbiome in ways that increase the risk for chronic disease. In addition, it is unclear how differences in study methodology can impact the results of clinical metagenome-wide association studies. In the current meta-analysis, we show that age, stone composition, and study location are the predominant factors that associate with the microbiome and USD status. Furthermore, we reveal differences in results based on specific analytical protocols, which impacts the interpretation of any microbiome study.

Numerous metagenome-wide association studies (MWAS) for urolithiasis have been published, leading to the discovery of potential interactions between the microbiome and urolithiasis. However, questions remain about the reproducibility, applicability and physiological relevance of these data owing to discrepancies in experimental technique and a lack of standardization in the field. One barrier to interpreting MWAS is that experimental biases can be introduced at every step of the experimental pipeline, including sample collection, preservation, storage, processing, sequencing, data analysis and validation. Thus, the introduction of standardized protocols that maintain the flexibility to achieve study-specific objectives is urgently required. To address this need, the first international consortium for microbiome in urinary stone disease - MICROCOSM - was created and consensus panel members were asked to participate in a consensus meeting to develop standardized protocols for microbiome studies if they had published an MWAS on urolithiasis. Study-specific protocols were revised until a consensus was reached. This consensus group generated standardized protocols, which are publicly available via a secure online server, for each step in the typical clinical microbiome-urolithiasis study pipeline. This standardization creates the benchmark for future studies to facilitate consistent interpretation of results and, collectively, to lead to effective interventions to prevent the onset of urolithiasis, and will also be useful for investigators interested in microbiome research in other urological diseases.

The prevalence of many chronic diseases has increased over the last decades. It has been postulated that dysbiosis driven by environmental factors such as antibiotic use is shifting the microbiome in ways that increase inflammation and the onset of chronic disease. Dysbiosis can be defined through the loss or gain of bacteria that either promote health or disease, respectively. Here we use multiple independent datasets to determine the nature of dysbiosis for a cluster of chronic diseases that includes urinary stone disease (USD), obesity, diabetes, cardiovascular disease, and kidney disease, which often exist as co-morbidities. For all disease states, individuals exhibited a statistically significant association with antibiotics in the last year compared to healthy counterparts. There was also a statistically significant association between antibiotic use and gut microbiota composition. Furthermore, each disease state was associated with a loss of microbial diversity in the gut. Three genera, Bacteroides, Prevotella, and Ruminococcus, were the most common dysbiotic taxa in terms of being enriched or depleted in disease populations and was driven in part by the diversity of operational taxonomic units (OTUs) within these genera. Results of the cross-sectional analysis suggest that antibiotic-driven loss of microbial diversity may increase the risk for chronic disease. However, longitudinal studies are needed to confirm the causative effect of diversity loss for chronic disease risk.

The prevalence of urinary stone disease (USD) is rapidly rising. However, the factors driving this increase are unknown. Recent microbiome studies suggest that dysbiosis may in part contribute to the increasing prevalence. The objective of the current study was to determine the nature and location of dysbiosis associated with USD. We conducted microbiome analysis from the gastrointestinal and urinary tracts, along with a metabolomic analysis of the urinary metabolome, from subjects with an active episode of USD or no history of the disease. Higher rates of antibiotic use among USD patients along with integrated microbiome and metabolomic results support the hypothesis that USD is associated with an antibiotic-driven shift in the microbiome from one that protects against USD to one that promotes the disease. Specifically, our study implicates urinary tract Lactobacillus and Enterobacteriaceae in protective and pathogenic roles for USD, respectively, which conventional, culture-based methods of bacterial analysis from urine and kidney stones would not necessarily detect. Results suggest that antibiotics produce a long-term shift in the microbiome that may increase the risk for USD, with the urinary tract microbiome holding more relevance for USD than the gut microbiome.

Research question: Male infertility accounts for almost half of all infertility cases worldwide, with idiopathic male infertility accounting for up to 30% of the cases. Sperm proteomics has revealed critical molecular pathway changes in men with infertility. However, the sperm mitochondrial proteome remains poorly understood. We attempted to answer the following question: Do patients with idiopathic primary male infertility exhibit a proteomic signature associated with mitochondrial dysfunction that could be used as a target for future mechanistic investigations? Design: Patients with idiopathic primary infertility (20–40 years old) referred to the Cleveland Clinic between March 2012 and April 2014 were compared with fertile donor controls. Sperm proteins were analyzed using sodium dodecyl sulphate-polyacrylamide gel electrophoresis page (SDS-PAGE) and liquid chromatography-mass spectrometry (LC-MS), and differentially expressed proteins (DEPs) were identified based on significance test results and fold change thresholds. Protein expression was validated using western blotting. Results: Proteomic analysis of pooled samples from fertile donors (n = 5) and patients with idiopathic primary infertility (n = 5) identified 1,134 proteins, including 344 DEPs. Mitochondrial dysfunction topped the ingenuity toxicity list. Analysis of expression levels of three mitochondrial proteins known to combat oxidative stress revealed that peroxiredoxin-5 (PRDX5) and superoxide dismutase 2 (SOD2), but not glutathione disulphide reductase, were significantly decreased in patient samples compared with those in fertile-donor samples. Conclusions: This study revealed an association of downregulated expression of PRDX5 and SOD2 in sperm samples of patients with idiopathic primary male infertility. Our results support future mechanistic studies and development of advanced diagnostic methods to better identify men with mitochondria-related male infertility. [ABSTRACT FROM AUTHOR]