The short-beaked echidna, Tachyglossus aculeatus, is a cherished and iconic native Australian animal and is 1 of 5 currently existing monotremes. Having diverged from Eutherian/placental mammals roughly 184 million years ago, monotremes have evolved unique reproductive strategies, specifically egg-laying. The composition of the gut microbiota of female mammals is quite important, as it provides an opportunity for vertical transmission of microbes to offspring as well as maintaining foetus health. Very little is known about the composition and role of the echidna microbiome. Hence, this study aims to characterize the gut microbiome of female short-beaked echidnas and gain a better understanding of the changes that may occur in their microbiome as they go through pregnancy. Faecal samples from four female echidnas were obtained from the Currumbin Wildlife Sanctuary in Queensland with samples collected across the gestation cycles for each female. DNA was extracted from each sample, and the V4-V5 region of the bacterial 16S rRNA gene was amplified and sequenced on an Illumina MiSeq (2x150 bp) for metabarcoding. The sample results provided insight into the core taxa and highly abundant bacterial identities across the different stages. A number of these taxa have also been recognised to be abundant in the microbiomes of other native Australian mammals. Analysis of samples revealed that the echidna microbiome remains relatively stable across the different gestation stages and hence microbiomes by gestation were not significantly different from each other. This study is the first to describe the microbiome composition of short-beaked echidnas during gestation phases and hence allows the opportunity for this novel information to be used as a metric of health and during the detection of diseases triggered by microbiota dysbiosis.