Wu Zi Yan Zong Wan enhances sperm quality in boars by regulating spermatogenesis-related pathways and gut microbiota

<i>Wu Zi Yan Zong Wan</i> (<i>WZYZW</i>) is a well-known Traditional Chinese Medicine formula used to treat low sperm quality, although its underlying therapeutic mechanism remains incompletely elucidated. This study aimed to elucidate the therapeutic mechanism of <i>WZYZW</i> for low sperm quality in breeding boars and to evaluate its potential for extending their reproductive lifespan. Boars with low sperm quality were pre-screened using the CASA system and <i>WZYZW</i> was added to the diet as an intervention. Sperm viability, ejaculate volume, sperm density, FSH, LH, testosterone T, Ca²⁺, Zn²⁺, T-AOC, GSH-Px, SOD and Giemsa staining were evaluated after 8 weeks to evaluate the therapeutic effect of <i>WZYZW</i> on boars with low sperm quality. Furthermore, an investigation was conducted into the potential mechanisms of <i>WZYZW</i> for treating low sperm quality in boars. This investigation utilized quantitative polymerase chain reaction (qPCR), 16S RNA sequencing, and thermal correlation analysis. <i>WZYZW</i> significantly restored sperm motility, ejaculate volume, and sperm density in boars with initial poor semen quality. The treatment also effectively reduced sperm deformities and increased the levels of FSH, testosterone (T), fructose, Ca²⁺, Zn²⁺, total antioxidant capacity (T-AOC), glutathione peroxidase (GSH-Px), and superoxide dismutase (SOD). At the molecular level, <i>WZYZW</i> altered chromatin structure and the expression of adhesion protein family genes, potentially via the cAMP-AKT pathway. Furthermore, 16S rRNA sequencing revealed that <i>WZYZW</i> significantly suppressed the relative abundance of the phyla <i>Tenericutes</i> and <i>Proteobacteria</i>. Correlation analysis suggested that the abundance of these gut microbiota taxa may be associated with signaling pathways regulating cell growth and death. Our results demonstrate that <i>WZYZW</i> is a promising therapeutic agent for low sperm quality in boars, which acts by modulating spermatogenesis-related gene expression and reshaping the gut microbiota, thereby extending the reproductive lifespan of breeding boars.