The ghost bat (Macroderma gigas) is endemic to Australia but is under threat, with scarce information available on the genetic health of remaining populations. Here, we develop molecular assays for microsatellite genotyping and molecular sexing of non-invasive samples as a genetic monitoring tool to identify individuals, measure genetic diversity and investigate spatial and temporal patterns of habitat use by ghost bats. We identified novel microsatellites through high-throughput sequencing on the Illumina MiSeq platform. Of 48 loci tested, six markers were added to five previously developed microsatellite loci. We developed three Y-linked (DDX3Y, Zfy and SRY) and one X-linked markers (Zfx) to enable molecular identification of sex. To assess performance, all 11 microsatellite and four sex-linked markers were amplified in three multiplex reactions in 160 M. gigas faecal samples from the Pilbara region, Western Australia. The combined markers offered a high level of individual discrimination (P_IDsibs = 0.00002) and we detected 19 bats in total (11 males, 4 females and 4 sex undetermined). The number of alleles per locus ranged from 5 to 14 and the average observed and expected heterozygosity across loci were H_o = 0.735 (0.58–0.91) and uH_e = 0.785 (0.59–0.89) respectively. Our molecular assays allowed identification of individuals from faecal samples at multiple time points and spatial locations and enabled us to elucidate patterns of habitat usage at the study site. This study highlights the value of our molecular assays as a potential capture-mark-recapture technique for population monitoring for this species.

蝙蝠（Macroderma gigas）是澳大利亚特有的，但受到威胁，缺乏关于剩余种群遗传健康的信息。在这里，我们开发了用于微卫星基因分型和非侵入性样品分子性别鉴定的分子测定法，以作为一种遗传监测工具来识别个体，测量遗传多样性并调查幽灵蝙蝠使用的栖息地的时空格局。我们通过Illumina MiSeq平台上的高通量测序鉴定了新型微卫星。在测试的48个基因座中，将六个标记物添加到五个先前开发的微卫星基因座中。我们开发了三个Y连锁（DDX3Y，Zfy和SRY）和一个X连锁标记（Zfx），以进行性别分子鉴定。为了评估性能，在16 M中的三个多重反应中扩增了所有11个微卫星和四个性别相关标记。来自澳大利亚西部Pilbara地区的gigas粪便样本。组合标记物提供了高水平的个体歧视性（P _IDsibs = 0.00002），我们总共检测到19只蝙蝠（11头雄性，4雌性和4性别未定）。每个基因座的等位基因数量在5至14之间，并且每个基因座的平均观察杂合度和预期杂合度为H _o = 0.735（0.58–0.91）和uH _e分别为0.785（0.59–0.89）。我们的分子分析可以在多个时间点和空间位置上从粪便样本中识别出个体，并使我们能够阐明研究地点的栖息地使用方式。这项研究强调了我们的分子分析作为对该种群进行监测的潜在捕获标记捕获技术的价值。