Integrating Near-Infrared Spectroscopy and Proteomics for Semen Quality Biosensing.

Artificial insemination (AI) is a key breeding technique in the swine industry; however, the lack of reliable biomarkers for semen quality limits its effectiveness. Seminal plasma (SP) contains extracellular vesicles (EVs) that present a promising, non-invasive biomarker for semen quality. This study explores the biochemical profiles of boar SP to assess semen quality through near-infrared spectroscopy (NIRS) and proteomics of SP-EVs. Fresh semen from mature Duroc boars was evaluated based on sperm motility, classifying samples as Passed (≥70%) or Failed (<70%). NIRS analysis identified distinct variations in water structures at specific wavelengths (C1, C5, C12 nm), achieving high accuracy (92.2%), sensitivity (94.2%), and specificity (90.3%) through PCA-LDA. Proteomic analysis of SP-EVs revealed 218 proteins in Passed and 238 in Failed samples. Nexin-1 and seminal plasma protein pB1 were upregulated in Passed samples, while LGALS3BP was downregulated. The functional analysis highlighted pathways associated with single fertilization, filament organization, and glutathione metabolism in Passed samples. Integrating NIRS with SP-EV proteomics provides a robust approach to non-invasive assessment of semen quality. These findings suggest that SP-EVs could serve as effective biosensors for rapid semen quality assessment, enabling better boar semen selection and enhancing AI practices in swine breeding.