Vector-borne disease prevalence is increasing at a time when surveillance capacity in the United States is decreasing. One way to address this surveillance deficiency is to utilize established infrastructure, such as zoological parks, to investigate animal disease outbreaks and improve our epidemiological understanding of vector-borne pathogens. During fall 2020, an outbreak of epizootic hemorrhagic disease (EHD) at the Minnesota Zoo resulted in morbidity and seroconversion of several collection animals. In response to this outbreak, insect surveillance was conducted, and the collected insects were tested for the presence of epizootic hemorrhagic disease virus (EHDV) by RT-qPCR to better understand the local transmitting vector populations responsible for the outbreak. Six pools of Culicoides biting midges were positive for EHDV, including three pools of Culicoides sonorensis, two pools of Culicoides variipennis, and a pool of degraded C. variipennis complex midges. All three endemic serotypes of EHDV (1, 2, and 6) were detected in both animals and midge pools from the premises. Despite this outbreak, no EHDV cases had been reported in wild animals near the zoo. This highlights the importance and utility of using animal holding facilities, such as zoos, as sentinels to better understand the spatio-temporal dynamics of pathogen transmission.

Vesicular stomatitis (VS) is a viral disease that affects horses, cattle, and swine that is transmitted by direct contact and hematophagous insects. In 2023, a multi-state outbreak of vesicular stomatitis New Jersey virus (VSNJV) occurred in California, Nevada, and Texas, infecting horses, cattle, and rhinoceros. To identify possible insect vectors, we conducted insect surveillance at various locations in San Diego County, CA, including at a wildlife park. CO₂ baited traps set from mid-May to mid-August 2023 collected 2357 Culicoides biting midges and 1215 Simulium black flies, which are insect genera implicated in VSNJV transmission. Insects were pooled by species, location, and date, then tested for viral RNA. Nine RNA-positive pools of Culicoides spp. and sixteen RNA-positive pools of Simulium spp were detected. Infectious virus was detected by cytopathic effect in 96% of the RNA-positive pools. This is the first report of VSNJV in wild-caught C. bergi, C. freeborni, C. occidentalis, S. argus, S. hippovorum, and S. tescorum. The vector competency of these species for VSNJV has yet to be determined but warrants examination. Active vector surveillance and testing during disease outbreaks increases our understanding of the ecology and epidemiology of VS and informs vector control efforts.

Bluetongue virus (BTV) causes internationally reportable hemorrhagic disease in cattle, sheep, and white-tailed deer. The closely related, and often co-circulating, epizootic hemorrhagic disease virus causes a clinically similar devastating disease in white-tailed deer, with increasing levels of disease in cattle in the past 10 years. Transmitted by Culicoides biting midges, together, they constitute constant disease threats to the livelihood of livestock owners. In cattle, serious economic impacts result from decreased animal production, but most significantly from trade regulations. For effective disease surveillance and accurate trade regulation implementation, rapid, sensitive assays that can detect exposure of cattle to BTV and/or EHDV are needed. We describe the development and validation of a duplex fluorescent microsphere immunoassay (FMIA) to simultaneously detect and differentiate antibodies to BTV and EHDV in a single bovine serum sample. Performance of the duplex FMIA for detection and differentiation of BTV and EHDV serogroup antibodies was comparable, with higher sensitivity than commercially available single-plex competitive enzyme-linked immunosorbent assays (cELISA) for detection of each virus antibody separately. The FMIA adds to the currently available diagnostic tools for hemorrhagic orbiviral diseases in cattle as a sensitive, specific assay, with the benefits of serogroup differentiation in a single serum sample, and multiplexing flexibility in a high-throughput platform.

Bluetongue (BT) is a Culicoides midge-borne hemorrhagic disease affecting cervids and ruminant livestock species, resulting in significant economic losses from animal production and trade restrictions. Experimental animal infections using the α/β interferon receptor knockout IFNAR mouse model and susceptible target species are critical for understanding viral pathogenesis, virulence, and evaluating vaccines. However, conducting experimental vector-borne transmission studies with the vector itself are logistically difficult and experimentally problematic. Therefore, experimental infections are induced by hypodermic injection with virus typically derived from baby hamster kidney (BHK) cells. Unfortunately, for many U.S. BTV serotypes, it is difficult to replicate the severity of the disease seen in natural, midge-transmitted infections by injecting BHK-derived virus into target host animals. Using the IFNAR BTV murine model, we compared the virulence of traditional BHK cell-derived BTV-17 with C. sonorensis midge (W8) cell-derived BTV-17 to determine whether using cells of the transmission vector would provide an in vitro virulence aspect of vector-transmitted virus. At both low and high doses, mice inoculated with W8-BTV-17 had an earlier onset of viremia, earlier onset and peak of clinical signs, and significantly higher mortality compared to mice inoculated with BHK-BTV-17. Our results suggest using a Culicoides W8 cell-derived inoculum may provide an in vitro vector-enhanced infection to more closely represent disease levels seen in natural midge-transmitted infections while avoiding the logistical and experimental complexity of working with live midges.

Oropouche virus (OROV) is an emerging zoonotic arthropod-borne virus of public health importance. The host range of OROV is largely unknown, but antibody evidence suggests that wildlife and livestock species could be susceptible hosts. To identify potential North American mammalian reservoir hosts, OROV replication curves were generated using eight cell lines derived from livestock, wildlife, and domestic animal species (cow, sheep, bison, white-tailed deer, elk, pig, horse, and dog). The virus replicated in all cell lines by 48 h post infection, except for the horse cells. OROV replication success was greatest in the bison cells followed by pig and dog cells. Moderate replication was achieved in the deer, elk, sheep, and cow cells. These results indicate that numerous animal species may be susceptible hosts for OROV, including important agricultural and wildlife species, but pathogenesis studies are required to confirm this finding. Identifying the reservoir hosts for OROV will allow livestock producers, veterinarians, and public health officials to prepare appropriate vector and disease control measures should the virus initiate an outbreak in the United States.