The koala (Phascolarctos cinereus) is an iconic Australian species that is currently undergoing a number of threatening processes, including disease and habitat loss. A thorough understanding of population genetic structuring and genomic variability of this species is essential to effectively manage populations across the species range. Using a reduced representation genome sequencing method known as double digest restriction-associated sequencing, this study has provided the first genome-wide SNP marker panel in the koala. In this study, 33,019 loci were identified in the koala and a filtered panel of 3060 high-utility SNP markers, including 95 sex-linked markers, were used to provide key insights into population variability and genomic variation in 171 koalas from eight populations across their geographic range. Broad-scale genetic differentiation between geographically separated populations (including sub-species) was assessed and revealed significant differentiation between all populations (F ST range = 0.01-0.28), with the largest divergence observed between the three geographically distant subgroups (QLD, NSW and VIC) along the east coast of Australia (average F ST range = 0.17-0.23). Subgroup divergence appears to be a reflection of an isolation by distance effect and sampling strategy rather than true evidence of sub-speciation. This is further supported by low proportions of AMOVA variation between sub-species groups (11.19 %). Fine-scale analysis using genome-wide SNP loci and the NETVIEW pipeline revealed cryptic genetic sub-structuring within localised geographic regions, which corresponded to the hierarchical mating system of the species. High levels of genome-wide SNP heterozygosity were observed amongst all populations (He = 0.25-0.35), and when evaluating across the species to other vertebrate taxa were amongst the highest values observed. This illustrates that the species as a whole still retains high levels of diversity which is comparable to other outbred vertebrate taxa for genome-wide SNPs. Insights into the potential for adaptive variation in the koala were also gained using 3/43 outlier analysis of genome-wide SNPs. A total of 10 putative outlier SNPs were identified indicating the high likelihood of local adaptations within populations and regions. This is the first use of genome-wide markers to assess population differentiation at a broad-scale in the koala and the first time that sex-linked SNPs have been identified in this species. The application of this novel genomic resource to populations across the species range will provide in-depth information allowing informed conservation priorities and management plans for in situ koalas across Australia and ex situ around the world.

The extent to which sex-biased maternal investment characterizes mammals is controversial, with less information available for evaluating patterns of maternal effort in marsupials than in placentals. Koalas Phascolarctos cinereus are sizedimorphic animals with a lengthy period of dependency and they reside in mating systems that might favour sex-biased maternal investment. We examined 18 years of data recorded from koalas living at the San Diego Zoo in order to examine how joey development and maternal condition might be connected. Koalas are pregnant for only 1 month, but joey emergence from the pouch does not occur until 32 weeks of age. Neither maternal condition nor age affected sex ratio at joey emergence, and both sexes had the same survivorship prospects. Koala dams transport and nurse joeys for close to 1 year, at which time the two sexes are size dimorphic. Given the poor-quality diet of koalas, combined with maternal transport of infants who are at least 25% of maternal mass, we suggest that infant rearing poses high energetic costs on koala females. We suggest that ecological and energetic constraints have moulded koala maternal strategies such that females maximize allocation of resources to offspring, regardless of sex, in order to increase prospects for joey survivorship.

The Australian koala is an iconic marsupial with highly specific dietary requirements distributed across heterogeneous environments, over a large geographic range. The distribution and genetic structure of koala populations has been heavily influenced by human actions, specifically habitat modification, hunting and translocation of koalas. There is currently limited information on population diversity and gene flow at a species-wide scale, or with consideration to the potential impacts of local adaptation. Using species-wide sampling across heterogeneous environments, and high-density genome-wide markers (SNPs and PAVs), we show that most koala populations display levels of diversity comparable to other outbred species, except for those populations impacted by population reductions. Genetic clustering analysis and phylogenetic reconstruction reveals a lack of support for current taxonomic classification of three koala subspecies, with only a single evolutionary significant unit supported. Furthermore, ~70% of genetic variance is accounted for at the individual level. The Sydney Basin region is highlighted as a unique reservoir of genetic diversity, having higher diversity levels (i.e., Blue Mountains region; AvHe=0.20, PL% = 68.6). Broad-scale population differentiation is primarily driven by an isolation by distance genetic structure model (49% of genetic variance), with clinal local adaptation corresponding to habitat bioregions. Signatures of selection were detected between bioregions, with no single region returning evidence of strong selection. The results of this study show that although the koala is widely considered to be a dietary-specialist species, this apparent specialisation has not limited the koala's ability to maintain gene flow and adapt across divergent environments as long as the required food source is available.

Male koalas communicate with conspecifics using scent markings from their sternal scent gland. We examined age and seasonal changes in the profile of organic compounds produced by koala sternal glands. Scent exudates from 12 male koalas were collected over a one-year period and analysed by direct-injection gas chromatography–mass spectrometry (GC–MS) of the methanol extract. Carboxylic acids, aldehydes, ketones, and alcohols were found to dominate the scent mixture. Thirty-seven compounds were identified by comparison of their spectra with those in the Wiley 138K library. The identity, complexity and relative concentrations of these compounds were found to vary both seasonally and by age of the individuals, with the most odorous and complex mixtures occurring during the mating season. We suggest that seasonal and age differences in the complexity of the chemical composition of the koala scent gland exudate provides evidence that olfactory communication probably has important ramifications for male reproductive success in the wild.

We analyzed 18 years of data collected from a breeding colony of captive koalas Phascolarctos cinereus in order to identify factors regulating mating success and to generate expectations about the behavioral ecology of wild koalas. Short-term mate fidelity was associated with reproductive success, but familiarity appeared to diminish reproductive output with time. Male body mass had no effect on reproductive success. Reproductive output was highest when males were slightly older than females. Survivorship of joeys was not dependent on either male or female age. We suggest that sexual selection regulates koala mating tactics primarily in the context of non-agonistic male competition. We hypothesize that field studies will reveal that females in the wild probably capitalize on the link between male vocal and olfactory advertisements and male age as a mechanism to foster positive assortative mating by age.