The Development of Environmental DNA (eDNA) Sampling for Arbovirus Vector Surveillance in Southern Nevada

Vector-borne diseases account for 17% of all infectious diseases worldwide and are responsible for more than 700,000 deaths annually. Two-thirds of the global population are at risk of arthropod-borne RNA viruses (arboviruses), spread by mosquitoes, ticks, and sandflies, with these pathogens accounting for 37% of all emerging human diseases. Arboviruses are becoming increasingly important causes of neurological disease in the United States, presenting a significant public health problem. In the absence of available vaccines and efficacious treatment options, accurate, rapid, and cost-effective surveillance of arbovirus vectors is critical for monitoring species distribution and mitigating disease transmission risk. Currently, mosquito vector surveys conducted by the Southern Nevada Health District (SNHD) are limited by infrastructure and logistical requirements. New vector surveillance methods that are simple and unbiased at the sampling stage are needed to enhance the survey capacity of the SNHD. One such methods is to exploit detection of environmental DNA (eDNA), shed by vectors breeding in aquatic environments. In this study, we designed and optimized a novel multiplex TaqMan qPCR assay, using single nucleotide polymorphisms in cytochrome oxidase I, to simultaneously detect environmental DNA (eDNA) shed by the three major local mosquito vector species—Culex (Cx.) quinquefasciatus, Cx. tarsalis, and Aedes (Ae.) aegypti—in aquatic environments. Water samples collected from various locations in Clark County, including water bodies near overnight gravid traps and BG sentinel traps, were analyzed to compare vector species composition. The results demonstrated that eDNA surveillance could effectively identify all three vector species of interest both with and without direct visualization at the breeding site. This approach shows promise for integration into citizen science initiatives, enhancing field-friendly arbovirus vector surveillance and expanding entomological monitoring capabilities in Southern Nevada. The study also highlights the potential of eDNA surveillance to detect changes in vector species dispersal patterns and the emergence of new invasive species.