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Vaginal cytology evaluation is an economic, non-invasive technique for indirect monitoring of fluctuations in estrogen concentrations, and thus progression of the estrous cycle. This technique is widely used in domestic dogs for determining timing of artificial insemination. There, however, are only a few reports on the vaginal cytology of non-domestic felids, including lions. This study was conducted, therefore, to describe the vaginal epithelial changes throughout the reproductive cycle of African lions, and to investigate the efficacy of vaginal cytology assessments for predicting reproductive stages. During a 12-month period, reproductive behavioral data and vaginal swabs were collected daily from five lionesses. In total, 541 vaginal smears were evaluated for the proportion of mucosal epithelial cells, neutrophils, bacterial cells, and amount of mucous, cellular debris. One single swab with a large proportion of superficial cells, absence of neutrophils, large number of bacteria, without cellular debris was sufficient for detecting lionesses in estrus. Likewise, one cytology sample with a large proportion of parabasal and intermediate cells, few neutrophils, few bacteria, and large amount of mucous, cellular debris enabled detection of females in advanced diestrus or gestation. To distinguish lionesses in early diestrus from those in an inter-estrous period, at least two consecutive swabs were necessary for satisfactory classification. Overall, evaluation of vaginal cytology samples was an effective technique for differentiation among different stages of the reproductive cycle, confirmation of estrus, and pregnancy diagnosis in lionesses. This technique, therefore, has the potential for application in classifying different stages of the reproductive cycle in other feline species.

Despite postulated potential for wildlife conservation, success of assisted reproduction techniques (ART) in ex-situ feline breeding remains25%. The aim of this project was to develop a simplified, non-surgical artificial insemination (AI) protocol for African lions (Panthera leo), using an exogenous GnRH analogue to induce ovulation in females presenting natural oestrus, and minimizing manipulation of the animals. Four protocols were tested in five trained lionesses (3.5-8 years), for a total of 14 inseminations (2-4 per lioness). These protocols differed in the time lapse between GnRH injection and insemination, on days 4, 5, or 6 from onset of natural oestrus, determined by daily behavioural observation and vaginal cytology. Semen was collected from 8 different males by urethral catheterization and electro-ejaculation, during full anaesthesia. Females were immediately immobilized for AI after semen collection. After transrectal ultrasound examination of the reproductive tract, insemination was performed either intravaginal or transcervical using a commercial dog urinary catheter (2.0 × 500 mm, Buster