Biomonitoring is vital for establishing baseline data that is needed to identify and quantify ecological change and to inform management and conservation activities. However, biomonitoring and biodiversity assessment in arid environments, which are predicted to cover 56% of the Earth's land surface by 2100, can be prohibitively time consuming, expensive, and logistically challenging due to their often remote and inhospitable nature. Sampling of environmental DNA (eDNA) coupled with high‐throughput sequencing is an emerging biodiversity assessment method. Here we explore the application of eDNA metabarcoding and various sampling approaches to estimate vertebrate richness and assemblage at human‐constructed and natural water sources in a semi‐arid region of Western Australia. Three sampling methods: sediment samples, filtering through a membrane with a pump, and membrane sweeping in the water body, were compared using two eDNA metabarcoding assays, 12S‐V5 and 16smam, for 120 eDNA samples collected from four gnammas (gnamma: Australian Indigenous Noongar language term–granite rock pools) and four cattle troughs in the Great Western Woodlands, Western Australia. We detected higher vertebrate richness in samples from cattle troughs and found differences between assemblages detected in gnammas (more birds and amphibians) and cattle troughs (more mammals, including feral taxa). Total vertebrate richness was not different between swept and filtered samples, but all sampling methods yielded different assemblages. Our findings indicate that eDNA surveys in arid lands will benefit from collecting multiple samples at multiple water sources to avoid underestimating vertebrate richness. The high concentration of eDNA in small, isolated water bodies permits the use of sweep sampling that simplifies sample collection, processing, and storage, particularly when assessing vertebrate biodiversity across large spatial scales.