T. P. Fletcher

Monitoring reproduction in the critically endangered marsupial, Gilbert’s potoroo (Potorous gilbertii): Preliminary analysis of faecal oestradiol-17β, cortisol and progestagens

Gilberts potoroo (Potorous gilbertii) was rediscovered in 1994 after having been presumed extinct for 120 years. Estimates indicate fewer than 40 individuals remain at Two Peoples Bay Nature Reserve on the south coast of Western Australia although a translocated population of approximately 20 animals has recently been established on nearby Bald Island. A captive breeding facility has been established adjacent to the mainland population but few young have been produced (8 since 1995). Faecal levels of oestradiol-17beta (E(2)) were monitored over a 2-year period in an effort to determine cyclic reproductive activity, and faecal cortisol levels were also monitored to gauge whether chronic stress may be a factor limiting breeding in captivity. Faecal steroids were monitored in six captive females, and four captive male potoroos, and four wild females. The only captive births recorded after 1998 were one in August 1999 and one in February 2001, both to the same female. Peaks in E(2) concentration, up to 10 ng g(-1) of dried faecal mass were measured and results to date suggest the main breeding period to be November-December based on elevated E(2) levels at this time. Clear patterns of reproductive activity in the captive females, however, were not evident. Analysis of epithelial cell counts from urinogenital swabs and faecal E(2) and progestagen (PM) levels from a single female kept at the Perth Zoo, suggest that Gilberts potoroo has an oestrous cycle of approximately 39 days. Faecal cortisol levels in captive females were significantly lower than those in wild-caught individuals and thus there is no indication that elevated cortisol levels per se inhibited reproduction in captive females.

Perturbed growth and development in marsupial young after reciprocal cross-fostering between species.

Cross-fostering of marsupial young between species can potentially facilitate propagation of endangered or rare marsupial species by artificially increasing the number of progeny produced. The present study compares the growth and development of normal and cross-fostered tammar and parma wallabies. Tammars cross-fostered into the pouches of parmas grew at a similar rate to naturally reared tammar young and had developmental milestones at a similar age. However, parma young cross-fostered between the day of birth and 15 days post-partum into tammars that were carrying young of equivalent developmental stages did not grow normally and were lost from the pouch. Parma young cross-fostered at 30 days survived, but had significantly reduced growth rates and their developmental milestones were delayed compared with normally reared parma young. Thus, growth can be affected by cross-fostering, even between species like tammars and parmas that are of similar size and have similar lactation lengths. The results of the present study suggest that maternal milk regulates the timing of development of each species and a mis-match in the time that each young receives critical milk components can have a marked effect on their growth and development.

Early onset of ghrelin production in a marsupial

Ghrelin regulates appetite in mammals and can stimulate growth hormone (GH) release from the pituitary. In rats and humans, ghrelin cells appear in the stomach during late fetal life. Nevertheless, the role of ghrelin in early mammalian development is not well understood. Marsupials deliver highly altricial young that weigh less than 1g so they must feed and digest milk at a comparatively immature stage of development. Since they complete their growth and differentiation while in the pouch, they are accessible models in which to determine the time course of ghrelin production during development. We examined the distribution of gastric ghrelin cells, plasma ghrelin concentrations and pituitary expression of the ghrelin receptor (ghsr-1alpha) and GH in the tammar wallaby, Macropus eugenii. There were ghrelin immunopositive cells in the developing mesenchyme of the stomach from day 10 post partum (pp) to day 150pp. Subsequently ghrelin protein in the fore-stomach declined and was absent by day 250pp but remained in the gastric cells of the hind-stomach. Ghrelin was detected in the developing pancreas from day 10pp but was absent by day 150pp and in the adult. Pituitary ghsr-1alpha expression and plasma concentrations of ghrelin increased significantly up to day 70-120pp while GH expression was also elevated, declining with GH to reach adult levels by day 180pp. These results demonstrate an early onset of gastric ghrelin expression in the tammar in concert with a functional stomach at a relatively earlier stage than that of developmentally more mature eutherian young.

Maturation of the growth axis in marsupials occurs gradually during post-natal life and over an equivalent developmental stage relative to eutherian species

The separation of a nutrition-responsive insulin-like growth factor (IGF) system and a growth hormone (GH) responsive IGF system to control pre- and post-natal growth of developing mammals may originate from the constraints imposed by intra-uterine development. In eutherian species that deliver relatively precocial young, maturation of the GH regulatory system is coincident with the time of birth. We measured the hepatic expression of the four key growth axis genes GH-receptor, IGF-1 and -2, and IGFBBP-3, and plasma protein concentrations of IGF-1 from late fetal life through to adult stages of a marsupial, the tammar wallaby. The data clearly show that maturation of GH-regulated growth in marsupials occurs gradually over the course of post-natal life at an equivalent developmental stage to that of precocial eutherian mammals. This suggests that the timing of GH-regulated growth in marsupials is not related to parturition but instead to the relative developmental stage.

Exon 3 of the growth hormone receptor (GH-R) is specific to eutherian mammals.

Growth hormone receptor (GH-R) plays a critical role in the control of growth and metabolism in all vertebrates. GH-R consists of 9 coding exons (2-10) in all eutherian mammals, while the chicken only has 8 coding exons, and does not have an orthologous region to exon 3 in eutherians. To further understand the evolutionary origins of exon 3 of the GH-R in eutherians we cloned the full-length GH-R sequence in a marsupial, the tammar wallaby to determine whether exon 3 was present or absent in marsupial liver cDNA. There was no evidence for the presence of an exon 3 containing mRNA in sequence of tammar pouch young and adult livers. We next examined the genomes of the platypus (a monotreme mammal) and the grey short-tailed opossum (another marsupial). Like the tammar, the GH-R gene of neither species contained an exon 3. GH receptor can obviously function in the absence of this exon, raising speculation about the function of this domain, if any, in eutherians. A comparison of exon 3 protein sequences within 16 species of eutherian mammals showed that there was approximately 75% homology in the domain but only 3 of the 21 amino acids were identical (Leu12, Gln13 and Pro17). Interestingly, we detected greater evolutionary divergence in exon 3 sequences from species that have variants of GH or prolactin (PRL) in their placentas. These data show that exon 3 was inserted into the GH-R after the divergence of the marsupial and eutherian lineages at least 130 million years ago.

Population ecology of the long-nosed potoroo (Potorous tridactylus) on French Island, Victoria

The elusive nature of the long-nosed potoroo (Potorous tridactylus) has hindered the collection of long-term data for this threatened species. Between June 2005 and May 2009, data on the ecology of a wild population of long-nosed potoroos located on French Island, Victoria, were collected during a series of research projects. Over this period, 33 individual potoroos were trapped a total of 251 times. Up to nine individuals were known to be alive at once on the 15-ha study site of mature remnant native forest. Adult potoroos showed high site fidelity and significant sexual size dimorphism, with males heavier and having longer head and pes lengths than females. Congruent with other studies, we found no evidence of seasonality in breeding. Births occurred in every month of the year and the testis volume of males did not vary throughout the year. In contrast to previous studies, however, we did not observe peaks in breeding activity. Our research and review of existing literature suggests that the ecology of the long-nosed potoroo is strongly influenced by local environmental conditions and emphasises the need to consider long-term and site-specific data when developing management strategies to conserve this ecologically important species.