Urinary Microbiome Shows Distinct Signature in Men with Poor Sperm Motility

Men with asthenozoospermia—the leading form of male-factor infertility in which sperm swim poorly—carry a measurably different community of bacteria in their urine than fertile men, according to a case-control study published in Basic and Clinical Andrology. Using 16S rRNA gene sequencing, researchers profiled the urinary microbiota of 60 men with isolated asthenozoospermia and 60 age-matched fertile controls, finding that while the overall richness of bacterial species was unchanged, the structure and predicted function of the community differed significantly between the two groups.

The study, led by Yang Pan, Shangren Wang, Tingshuai Cao and colleagues, enrolled participants through a hospital health examination center and fertility counseling service. The asthenozoospermia group consisted of men aged 18 to 40 whose progressive sperm motility fell below the World Health Organization reference value while ejaculate volume, sperm concentration, total sperm count and morphology remained within normal ranges. Men with oligozoospermia, teratozoospermia, azoospermia or any combined semen abnormality were excluded, as were those who had used antibiotics, probiotics or immunosuppressive agents within two months of sampling, or who had active urinary or genital tract infection, leukocytospermia, clinically detectable varicocele, recent febrile illness or systemic inflammatory conditions.

Controls were fertile volunteers with normozoospermia, each of whom had fathered at least one child within the preceding three years through natural conception. They were frequency-matched to the patient group by age in two-year strata. The two groups were well balanced: mean age was 28.3 years in both, body mass index differed negligibly, and smoking and alcohol consumption were comparable. Abstinence duration, ejaculate volume, sperm concentration and the proportion of morphologically normal spermatozoa did not differ between groups. As expected from the case definition, median progressive motility was 22.5 percent in the asthenozoospermia group versus 45.5 percent in controls.

Urine was collected as a clean-catch midstream specimen using a standardized sterile technique, with roughly 50 milliliters obtained from each participant and transported on ice. Because urine has low microbial biomass, the microbial fraction was concentrated by centrifugation at 10,000 g for 20 minutes at 4 degrees Celsius before DNA extraction with a modified protocol designed to improve lysis of Gram-positive organisms. Extraction blanks and PCR no-template controls were processed alongside the biological specimens throughout the workflow, and the decontam R package was applied to remove amplicon sequence variants that were more prevalent in negative controls than in samples. Quantitative PCR confirmed that the bacterial load of every urine specimen exceeded that of the controls, and total bacterial load did not differ between the groups.

The central result concerned diversity. Alpha diversity—measured by observed features, Chao1, Shannon and Simpson indices—did not differ between the groups by any metric examined. Beta diversity, however, told a different story. Principal coordinate analysis based on Bray-Curtis dissimilarity showed partial separation of the two groups, and PERMANOVA confirmed a statistically significant difference in overall community structure, with an R2 of 0.13 and p equal to 0.001. PERMDISP verified that this separation was not merely an artifact of unequal within-group dispersion, and the group effect remained significant when body mass index, smoking status, alcohol consumption and abstinence duration were entered as covariates.

The authors are careful to stress the modest size of that effect. Group membership accounted for only about 13 percent of the total variance in community structure, with the remainder presumably attributable to other host-related, behavioral and environmental determinants. Asthenozoospermia-associated microbiota alteration, they write, should be regarded as one modest contributor among many rather than the dominant determinant of the urinary community.

At the taxonomic level, the dominant phyla in both groups were Firmicutes, Proteobacteria, Actinobacteriota and Bacteroidota. Linear discriminant analysis effect size identified 51 discriminative features, among which 18 genera differed significantly in relative abundance. Porphyromonas, Corynebacterium, Streptomyces, Peptoniphilus, Negativicoccus, Agromyces, Paludibacteraceae F0058, Finegoldia, Campylobacter and Gemella were more abundant in the asthenozoospermia group, while Blautia, Bacteroides, Intestinimonas, Sphingomonas, Alistipes, the Christensenellaceae R-7 group, Rhodoplanes and Oscillospiraceae UCG-005 predominated in fertile controls. The five most abundant genera in the patient group were Finegoldia, Streptococcus, Corynebacterium, Staphylococcus and Prevotella, whereas Streptococcus, Staphylococcus, Acinetobacter, Bacillus and Gardnerella predominated in controls.

Several of the patient-enriched taxa have prior connections to urogenital disease. Porphyromonas has been detected in the urine of men with prostate cancer, and increased Porphyromonas abundance has been reported in men with bladder cancer relative to controls. Corynebacterium, often dismissed as a culture contaminant, can cause encrusted cystitis, a chronic urinary tract infection accompanied by pain, bladder necrosis and renal failure; because it is also a commensal of skin, urethra and genital tract, the authors regard it as a conditional pathogen. Streptomyces, a soil-associated genus only occasionally reported as an opportunistic human pathogen, was also more abundant in the patient group, though the researchers suggest its presence in urine more plausibly reflects environmental exposure than pathogenic contribution and requires independent confirmation.

Beyond composition, the team used PICRUSt2 to infer functional potential from the 16S profiles, mapping gene content onto the KEGG hierarchy. Four level 1 categories, 18 level 2 pathways and 89 level 3 pathways differed between the groups. Organismal systems and genetic information processing were enriched in the asthenozoospermia group, while metabolism and cellular processes were relatively enriched in controls. At the finer level, pathways differing between groups included replication and repair modules, base excision repair, peptidoglycan biosynthesis, one-carbon pool by folate, folate biosynthesis, and nicotinate and nicotinamide metabolism. The authors interpret these patterns cautiously: enrichment of DNA repair and translation machinery might indicate taxa equipped for rapid growth, while shifts in folate and nicotinamide metabolism may point to an altered redox and methyl-donor economy of the community. Crucially, these pathways were predicted from taxonomic data rather than measured directly, and PICRUSt2 cannot indicate whether the inferred genes are expressed or resolve strain-level functional variation.

The study’s limitations are laid out explicitly. The cross-sectional, observational design precludes causal inference, and reverse causation—impaired reproductive physiology shaping the microbiota—cannot be excluded. No consensus gold-standard urinary fraction exists for male infertility microbiome research, and the choice of midstream urine characterizes the broader urinary ecosystem rather than serving as a direct proxy for the seminal, prostatic or epididymal microenvironment. Low-biomass specimens such as urine remain vulnerable to residual contamination despite sterile collection and in silico decontamination. Abstinence data were self-reported, idiopathic asthenozoospermia is etiologically heterogeneous, and the single-center design and modest sample size limit generalizability. The authors frame their mechanistic proposals—involving lipopolysaccharide-driven inflammation, oxidative stress and microbial metabolites—as hypotheses derived from associative data that require direct experimental validation, and they call for longitudinal, mechanistic and multi-omic studies, ideally incorporating paired urinary and semen sampling, to establish causality and determine whether urinary signatures could eventually serve as non-invasive biomarkers.

Subject of Research: Urinary microbiota composition and predicted function in men with asthenozoospermia

Article Title: Alterations in urinary microbiota composition and function in men with Asthenozoospermia: a case-control study

Article References: Pan, Y., Wang, S., Cao, T., Li, B., Hao, X., Li, M., Liu, L., & Liu, X. (2026). Alterations in urinary microbiota composition and function in men with Asthenozoospermia: a case-control study. Basic and Clinical Andrology, 36(1), Article 29. https://doi.org/10.1186/s12610-026-00326-0

Image Credits: AI Generated

DOI: 10.1186/s12610-026-00326-0

Keywords: asthenozoospermia, male infertility, urinary microbiota, 16S rRNA sequencing, PICRUSt2, PERMANOVA, dysbiosis, sperm motility, case-control study, Porphyromonas, Corynebacterium, Blautia