Current evidence suggests the presence of transcripts for several secretory phospholipase A₂ (sPLA₂) enzymes in male genital organs. In this study, we examined by immunohistochemistry the localization of group IIA, IIC, IID, IIE, IIF, V and X sPLA₂s in male genital organs. In sPLA₂-IIA-deficient C57BL/6 mouse testis, sPLA₂-IIC, -IID, -IIE, -IIF, -V and -X were diversely expressed in spermatogenic cells within the seminiferous tubules. Immunoblotting revealed the presence of these sPLA₂s in mouse spermatozoa. In addition, sPLA₂-IIF, -V and -X were localized in the interstitial Leydig cells. The same set of sPLA₂s was detected in a mouse cultured Leydig cell line, and adenovirus-mediated transfer of these sPLA₂s into Leydig cells resulted in increased prostaglandin production. sPLA₂-IIC, -IID, -IIE, -IIF, -V and -X were also detected diversely in the epithelium of the epididymis, vas deferens, seminal vesicles, and prostate. In a sPLA₂-IIA-positive FVB strain, weak expression of sPLA₂-IIA was detected in Leydig cells. Notable differences in the sPLA₂ expression profiles were found in the seminal vesicles and prostate between mice and humans. Taken together, individual sPLA₂s exhibit distinct or partially overlapping localizations in male reproductive organs, suggesting both specific and redundant functions.