The Kunjin strain of West Nile virus (WNV_KUN) is a mosquito-transmitted flavivirus that can infect farmed saltwater crocodiles in Australia and cause skin lesions that devalue the hides of harvested animals. We implemented a surveillance system using honey-baited nucleic acid preservation cards to monitor WNV_KUN and another endemic flavivirus pathogen, Murray Valley encephalitis virus (MVEV), on crocodile farms in northern Australia. The traps were set between February 2018 and July 2020 on three crocodile farms in Darwin (Northern Territory) and one in Cairns (North Queensland) at fortnightly intervals with reduced trapping during the winter months. WNV_KUN RNA was detected on all three crocodile farms near Darwin, predominantly between March and May of each year. Two of the NT crocodile farms also yielded the detection of MVE viral RNA sporadically spread between April and November in 2018 and 2020. In contrast, no viral RNA was detected on crocodile farms in Cairns during the entire trapping period. The detection of WNV_KUN and MVEV transmission by FTA^TM cards on farms in the Northern Territory generally correlated with the detection of their transmission to sentinel chicken flocks in nearby localities around Darwin as part of a separate public health surveillance program. While no isolates of WNV_KUN or MVEV were obtained from mosquitoes collected on Darwin crocodile farms immediately following the FTA^TM card detections, we did isolate another flavivirus, Kokobera virus (KOKV), from Culex annulirostris mosquitoes. Our studies support the use of the FTA^TM card system as a sensitive and accurate method to monitor the transmission of WNV_KUN and other arboviruses on crocodile farms to enable the timely implementation of mosquito control measures. Our detection of MVEV transmission and isolation of KOKV from mosquitoes also warrants further investigation of their potential role in causing diseases in crocodiles and highlights a “One Health” issue concerning arbovirus transmission to crocodile farm workers. In this context, the introduction of FTA^TM cards onto crocodile farms appears to provide an additional surveillance tool to detect arbovirus transmission in the Darwin region, allowing for a more timely intervention of vector control by relevant authorities.

The Northern Territory Top End Health Service, Medical Entomology Section and the City of Darwin council carry out a joint Mosquito Engineering Program targeting the rectification of mosquito breeding sites in the City of Darwin, Northern Territory, Australia. In 2005, an investigation into potential subterranean stormwater breeding sites in the City of Darwin commenced, specifically targeting roadside stormwater side entry pits. There were 79 side entry pits randomly investigated for mosquito breeding in the Darwin suburbs of Nightcliff and Rapid Creek, with 69.6% of the pits containing water holding sumps, and 45.6% of those water holding sumps breeding endemic mosquitoes. Culex quinquefasciatus was the most common mosquito collected, accounting for 73% of all mosquito identifications, with the potential vector mosquito Aedes notoscriptus also recovered from a small number of sumps. The sumps were also considered potential dry season maintenance breeding sites for important exotic Aedes mosquitoes such as Aedes aegypti and Aedes albopictus, which are potential vectors of dengue, chickungunya and Zika virus. Overall, 1229 side entry pits were inspected in ten Darwin suburbs from 2005 to 2008, with 180 water holding sumps identified and rectified by concrete filling.

Mosquitoes (n = 4381 in 198 pools) were collected in March and April 2018 to survey the presence of West Nile virus Kunjin strain in mosquito populations around crocodile farms in the Darwin region of the Northern Territory (NT) of Australia. While no Kunjin virus was detected in these mosquitoes, we applied our viral replicative intermediates screening system termed monoclonal antibodies to viral RNA intermediates in cells or MAVRIC to this set of samples. This resulted in the detection of 28 pools with virus replicating in C6/36 mosquito cells and the identification of three insect viruses from three distinct virus classes. We demonstrate the persistence of the insect-specific flavivirus Palm Creek virus in Coquillettidia xanthogaster mosquitoes from Darwin over almost a decade, with limited genetic drift. We also detected a novel Hubei macula-like virus 3 strain in samples from two mosquito genera, suggesting the virus, for which the sequence was originally detected in spiders and soybean thrips, might be involved in a horizontal transmission cycle between arthropods and plants. Overall, these data demonstrate the strength of the optimized MAVRIC system and contribute to our general knowledge of the mosquito virome and insect viruses.