XXX/XY chimerism with urogenital malformations in a Japanese black calf.

Abstract

Background: Sex chromosome abnormalities in cattle are rare, and manifestations of genital anomalies due to such abnormalities are even less frequently reported. Among these, XXX/XY chimerism is particularly uncommon. This report presents a Japanese black calf with complex urogenital malformations linked to XXX/XY chimerism, contributing valuable insights into bovine sex determination and reproductive development.

Case presentation: A Japanese black calf of phenotypic indeterminate sex, born co-twin to a phenotypically normal male, presented with hypospadias-like features and ambiguous genitalia. Clinical examination revealed a scrotum-like structure without palpable testes or vulva. An hCG stimulation test indicated a lack of functional testicular tissue. Chromosomal analysis of leukocytes revealed the presence of two distinct cells with 60, XY and 61, XXX, revealing XXX/XY chimerism. The ratio of male to trisomic cells was 63:37 (95% confidence intervals; XY: 54-72%, XXX: 28-46%) in the affected calf. Necropsy revealed both male (testis, epididymis, ductus deferens) and female (uterus-like) reproductive structures, with uterus-like organs embedded within the perineal tissue. Histological and immunohistochemical analyses confirmed the presence of the uterine remnant and revealed Sertoli cell-only testicular tissue, indicating spermatogenic failure. PCR-based sex determination performed on multiple tissues revealed three distinct genotypic patterns, with evidence of tissue-specific variation in the distribution of the X and Y chromosomes. Some tissues lacked detectable Y-linked AMEL-Y, despite the presence of SRY, suggesting a complex chimeric constitution with potential deletion of the AMEL-Y region in some XY cell populations.

Conclusions: This case highlights a rare instance of systemic XXX/XY chimerism associated with ambiguous genitalia and mixed internal reproductive structures, which is distinct from typical freemartinism or isolated X-trisomy syndromes. The differential chromosomal mosaicism across tissues likely influences the phenotypic outcome. These findings emphasize the complexity and plasticity of bovine sex differentiation, particularly in twin pregnancies, and underscore the importance of integrating clinical, cytogenetic, and molecular diagnostics to accurately identify and manage congenital reproductive anomalies in livestock.