A non-invasive, improved RIA and overt observation in the study of singleton Apennines’ wolf (Canis lupus) reproductive behavior

The analysis of fecal hormones allows a close but non-invasive monitoring of animals avoiding the stress of restraint/capture, which in turn can upset animalsʹ hormonal profile. Steroid hormone progesterone was analysed in three singleton, female grey wolves of different age, belonging to the endangered species of the Apenninesʹ Canis lupus. The analysis was carried out during the breeding season by using an improved radioimmunoassay on samples collected on the field. To reduce the stress to animals and danger to people, the overt observations were carried out by operators who were already familiar with the animals, saving the money of a camera-monitoring-system. Concurrently, a male and a female gray wolves housed together were monitored as a control. The results indicated the importance of dehydration of fecal samples before the extraction with petroleum ether, which was shown to be more efficient than diethyl ether, and that pre-treatment with methanol greatly enhances extraction (p<0.01). Females of Apenninesʹ grey wolf showed the first sign of oestrus by a vaginal blood loss, that was easily detected on the ground; the analysis carried out on fecal samples revealed a rapidly declining luteal phase, with P4 metabolites reaching the basal values of a non-cyclic female. In the matter of welfare, behavioural observations on Apenninesʹ grey wolf showed that unpaired animals, although familiar with the operators, failed to display a sexual social behavior during the reproductive season, that is the behavioural signs were hidden in overt observational situation.