KFDV primarily replicated in the brain. Other research has expanded on this by described neurological changes that occurred within infected organisms. This experiment was completed by using KFDV-infected mice and discovered that KFDV caused gliosis, inflammation, and cell death in the brain. They posited that KFDV could be primarily a neuropathic disease and other symptoms are due to this pathogenesis. Diagnosis In earlier days suspected case were confirmed in a laboratory by serum inoculation into suckling mice (Swiss Albino mice) and subsequent death of mice was leveled as KFD Positive case. Other methods of diagnosis included hemagglutination inhibition (HI), complement fixation, neutralization tests. However, new research has introduced more efficient molecular based methods to diagnose KFDV. These methods include: RT-PCR, nested RT-PCR, TaqMan-based real-time RT-PCR, Immunoglobin M antibodies and Immunoglobin G detection by ELISA. The two methods involving RT-PCR are able to function by attaching a primer to the NS-5 gene, which is highly conserved among the genus to which KFDV belongs. PCR positivity is limited to 8–10 days from the onset of symptoms. The ELISA based methods allows for the detections of anti-KFDV antibodies in patients typically from 5th day of onset of symptoms up to 3 months. Prevention and treatment Prevention is by vaccination, as well as preventive measures such as protective clothing and tick population control. The vaccine for KFDV consists of formalin-inactivated KFDV. The vaccine has a 62.4% effectiveness rate for individuals who receive two doses. For individuals who receive an additional dose, the effectiveness increases to 82.9%. Specific treatments are not available. Risk factors and risk groups The spill-over of Kyasanur forest disease happens at the crossroads of the animal-human interaction, especially villages adjoining forest areas and inter-state borders. People who frequently visit the forest areas of the Western Ghats region such as forest guards and officials, range forest officer (RFO), forest watchers, shepherds, firewood collectors, dry leaf collectors, hunters, people who handle dead animal carcasses, travelers who camp in the forest areas, tribal communities living inside the forest areas (Jenu kurubas and Betta kurubas), cashew nut workers especially those who engage in cleaning the dry leaves before the harvest season (seen in Pali and Mauxi outbreaks, North Goa), and areca nut farm workers working in infected tick areas will have a high risk of acquiring KFD infection. People who live in the KFD endemic areas and refuse to take KFD vaccination are at risk