Non-invasive age estimation based on faecal DNA using methylation-sensitive high-resolution melting for Indo-Pacific bottlenose dolphins.

Age is necessary information for the study of life history of wild animals. A general method to estimate the age of odontocetes is counting dental growth layer groups (GLGs). However, this method is highly invasive as it requires the capture and handling of individuals to collect their teeth. Recently, the development of DNA-based age estimation methods has been actively studied as an alternative to such invasive methods, of which many have relied on used biopsy samples. However, if DNA-based age estimation can be developed from faecal samples, age estimation can be performed entirely non-invasively. We developed an age estimation model using the methylation rate of two gene regions, GRIA2 and CDKN2A, measured through methylation-sensitive high-resolution melting (MS-HRM) from faecal samples of wild Indo-Pacific bottlenose dolphins (Tursiops aduncus). The age of individuals was known through conducting longitudinal individual identification surveys underwater. Methylation rates were quantified from 36 samples collected from 30 individuals. Both gene regions showed a significant correlation between age and methylation rate. The age estimation model was constructed based on the methylation rates of both genes which achieved sufficient accuracy (after LOOCV: MAE = 5.08, R2 = 0.33) for the ecological studies of the Indo-Pacific bottlenose dolphins, with a lifespan of 40-50 years. This is the first study to report the use of non-invasive faecal samples to estimate the age of marine mammals.

A wild Indo-Pacific bottlenose dolphin adopts a socially and genetically distant neonate.

Alloparental behaviour and adoption have been reported in many mammals and birds. Such behaviours are energetically costly, and their causes and functions remain unclear. We observed the adoption behaviour of a wild Indo-Pacific bottlenose dolphin (Tursiops aduncus) near Mikura Island, Japan. A calf was seen with its mother on six observation days. Following the mother's death, the calf was observed with a sub-adult female on all 18 observation days from May to September 2012. On three days, the calf was observed swimming with this female in the suckling position and milk was seen leaking from the female's mammary slit. A five-year dataset revealed no significant social or kin relationships between the biological mother and allomother, indicating that kinship and social relationships did not play an important role in the observed adoption.

Spontaneous ejaculation in a wild Indo-Pacific bottlenose dolphin (Tursiops aduncus).

Spontaneous ejaculation, which is defined as the release of seminal fluids without apparent sexual stimulation, has been documented in boreoeutherian mammals. Here we report spontaneous ejaculation in a wild Indo-Pacific bottlenose dolphin (Tursiops aduncus), and present a video of this rare behavior. This is the first report of spontaneous ejaculation by an aquatic mammal, and the first video of this behavior in animals to be published in a scientific journal.

Fine-scale analysis of synchronous breathing in wild Indo-Pacific bottlenose dolphins (Tursiops aduncus).

We quantitatively analysed synchronous breathing for dyads in Indo-Pacific bottlenose dolphins at Mikura Island, Tokyo, Japan. For most cases, we observed dyads swimming in the same direction (97%), in close proximity (i.e., less than 1.5m) and with their body axes parallel as they breathed synchronously. Moreover, the pairs engaged in identical behaviour before and after the synchronous breathing episodes. These results suggest that the dolphins synchronize their movements, and that synchronous breathing is a component of "pair-swimming", an affiliative social behaviour. Same sex pairs of the same age class frequently engaged in synchronous breathing for adults and subadults, as well as mother-calf and escort-calf pairs. The distance between individuals during synchronous breathing for mother-calf pairs was less than for other pairs. The distance observed between individuals for female pairs was less than for male pairs. The time differences between each exhale for each of the two dolphins involved in synchronous breathing episodes for female pairs were smaller than for male pairs, and time differences for adult pairs were smaller than subadult pairs. These results suggest that age and sex class influenced the characteristics of this behaviour.