Non-invasive DNA sampling from feces can provide powerful tools for wildlife research, management, and conservation. However, obtaining high quality and quantity fecal DNA is notoriously problematic, being affected by many different variables. Arguably, the most influential factor within the control of biologists is how samples are collected and stored prior to laboratory analyses. Here, we aimed to compare different fecal DNA preservation methodologies for their performance with a targeted genotyping approach and improve the quality and quantity of DNA extracted to better inform sampling of feces in the field. We assessed the proportion of missing single nucleotide polymorphism (SNP) data resulting from seven different fecal DNA preservation methodologies on fresh koala (Phascolarctos cinereus) scats. DNA was successfully obtained from all preservation methodologies; however, optimal recovery of DNA was obtained via a lysis shaken (i.e., washed) methodology, which provided a yield almost equivalent to that of more invasively sampled, high-quality, tissue samples. Our findings suggest there is a significant advantage of using a lysis buffer and washing technique coupled with targeted genotyping from scats. As a robust sampling method underpins successful data analysis, this optimized fecal sampling technique can further enhance our ability to address critical questions in population ecology, conservation genetics, and population management and help implement improved conservation strategies and decision making.

Recent advances in DNA sequencing and genotyping technologies are rapidly building our capacity to address ecological, evolutionary, and conservation questions for wildlife. However, wildlife genetic research increasingly relies on samples containing low quantities and quality of DNA, such as non-invasive, archival, and environmental DNA. These samples present unique methodological challenges; for samples collected in the wild, it is important to understand the impact of environmental exposure time or sample ’age’ on DNA quality and downstream genetic analyses. Here, we aged koala (Phascolarctos cinereus) scats under natural conditions and quantified DNA degradation. We assessed the effect of age on genetic data quality by measuring the proportion of missing single nucleotide polymorphism (SNP) data using DArTCap, a targeted genotyping methodology hypothesised to tolerate degraded DNA. Contrary to other studies, we found koala scat age was not a significant predictor of genotyping quality (i.e., rate of missing SNP calls) in the first 10 days of environmental exposure. We yielded high quality data from 10-day-old scats, but also low-quality data from fresh scats. This study is the first to investigate the effect of scat age on genotyping success using a targeted approach, and DArTCap specifically. These findings support the use of targeted genotyping (such as DArTCap) from scats and provide insights for future research using DNA from non-invasive samples. Targeted genotyping may extend the timeframe from which accurate data can be obtained from non-invasive samples, increasing potential sample sizes and enhancing our ability to address important questions in population ecology, conservation genetics, and population management.

Linear infrastructure stands as one of the main culprits of anthropogenically caused biodiversity decline. As it fragments landscapes, it ultimately results in a myriad of direct and indirect ecological consequences for wildlife. As transportation networks will continue to grow under increasing human population growth, biodiversity will continue to decline making the need to understand and mitigate their impact on species an urgent need for conservation worldwide. The implementation of mitigation measures to alleviate the barrier effect produced by linear transport infrastructure on local fauna is not new, and research has shown that their effectiveness has been shown to be influenced by their design, their placement and the biology of the impacted species. Our understanding of their effectiveness in preventing the longer-term impacts of linear transport infrastructure on habitat connectivity via gene flow, however, remains poorly understood. Here, we used a pre- and post-habitat fragmentation genetic dataset collected as part of an extensive Koala Management Program to ask questions about the immediate and predicted longer-term genetic consequences of linear transport infrastructure on the impacted species. Importantly, using forward migration simulations, we show that to preserve connectivity would need to result in around 20% of the population mixing to avoid long-term genetic drift. These results have important consequences for the management of species at the forefront of linear infrastructure. In particular, the study shows the importance of considering gene flow in our assessment of the effectiveness of fauna crossings.

The koala, one of the most iconic Australian wildlife species, is facing several concomitant threats that are driving population declines. Some threats are well known and have clear methods of prevention (e.g. habitat loss can be reduced with stronger land-clearing control), whereas others are less easily addressed. One of the major current threats to koalas is chlamydial disease, which can have major impacts on individual survival and reproduction rates, and can translate into population declines. Effective management strategies for the disease in the wild are currently lacking, and to date we know little about the determinants of individual susceptibility to disease. Here we used a rare opportunity to investigate the genetic basis of variation in susceptibility to chlamydia using one of the most intensively studied wild koala populations. We combine data from veterinary examinations, chlamydia testing, genetic sampling and movement monitoring. Out of our sample of 342 wild koalas, 60 were found to have chlamydia. Using genotype information on 8649 SNPs to investigate the role of genetic characteristics in determining disease status, we found no evidence of inbreeding depression, but a heritability of 0.14 (95%CI: 0.06 – 0.23) for the probability that koalas had chlamydia. Heritability of susceptibility to chlamydia could be relevant for future disease management in koalas, as it suggests the potential to select for disease resilience through assisted breeding.