BackgroundScrub typhus in an emerging vector-borne zoonosis, caused by Orientia spp. and transmitted by larvae of trombiculid mites, called chiggers. It mainly occurs within a certain region of the Asia-Pacific, called tsutsugamushi triangle, where rodents are known as the most relevant hosts for the trombiculid vector. The disease has recently been discovered on Chiloé Island in southern Chile. Still, the reservoir(s) and vector(s) of the scrub typhus outside Asia-Pacific are unknown. The aim of the present work was to study the prevalence of chiggers on different rodent species captured in sites identified as probably hot spots of scrub typhus on Chiloé Island in southern Chile.Methodology/Principal FindingsDuring austral summer 2018, rodents were live-trapped in six sites and examined for chigger infestation. During a total of 4,713 trap-nights, 244 rodents of seven species were captured: the most abundant was Abrothrix olivacea. All study sites were rural areas on Chiloé Island, previously identified as localities of probable human infection with scrub typhus. Chiggers were detected on all seven rodent species with a 55% prevalence rate. Chiggers showed low host specificity and varied according to site specific host abundance. We identified trombiculids of three genera. Colicus was the most abundant chigger (93%), prevalent in five of six sites, followed by Quadraseta (7%) and Paratrombicula (7%), which were in only one site. Infestation rates showed site specific differences, which were statistically different using a GLM model with binomial errors.Conclusions/SignificanceThis study firstly reports the presence of different rodent-associated chigger mites in a region with endemic scrub typhus in southern Chile. Colicus and two other genera of mites were found with high infestation rates in sites previously identified as hot spots of scrub typhus, suggesting their role as vectors and reservoirs of this emerging zoonosis in South America.Author SummaryScrub typhus is a chigger-transmitted zoonotic infection, which is endemic in the tsutsugamushi triangle in Asia-Pacific. Recently, a first focus of scrub typhus in South America has been confirmed on Chiloé Island in southern Chile. Still, the vectors of scrub typhus in this region remain unknown. We undertook a survey to study the prevalence of chiggers on different rodent species in areas identified as probable hot spots of scrub typhus on Chiloé Island. The study showed that 55% of rodents were infested by trombiculids. Three chigger genera were identified, of which Colicus was the most abundant. Chiggers showed low host specificity, but spatial differences. This first demonstration of rodent-associated chigger mites in hot spots of scrub typhus suggests their possible role as vectors of this infection in Chile.