Male fertility in farm animals is considered as an important economic trait. The phenomenon of spermatogenesis plays adynamic functional role in determining the viability of sperm and thereby can impact on fertility-driven complications. The process ofspermatogenesis is controlled by numerous molecular factors and requires a precisely regulated pattern of gene expression. The role ofsmall noncoding RNAs in altering gene expression has been extensively studied. However, limited information is available apropos theirrole in yak spermatogenesis. The present study aimed to evaluate the assessment of some significant microRNAs and their expressionpattern in the body tissues and sperm of fertile and subfertile yak from Arunachal Pradesh besides identified a novel class of sperm enrichedsmall RNA ‘mature-sperm-enriched small RNA’ (mse-tsRNA) in Yak spermatozoa. The RNAwas extracted from tissue and sperm using 27gauge needles and subsequently reverse transcribed into small RNA cDNAs. The PCR positive sperm-predominant miRNAs were validatedby quantitative reverse transcriptase PCR (qRT-PCR) for their expression in fertile and subfertile yak. Of the 22 microRNAs, themiRNA19a, miRNA142 and miRNA143 showed higher expression in the subfertile yak, whereas expression of miRNA7d, miRNA23a andmiRNA23b were found elevated in the fertile animal. The presence of these small noncoding RNAs in yak sperm and testis indicated thelegitimate involvement of their role in yak bull fertility.