Studies on the sequential pathology of Kyasanur Forest Disease (KFD) in Mouse brain: KFD virus induces apoptosis of neurons in cerebrum and hippocampus.

The sequential pathology of Kyasanur forest disease (KFD) in mouse brain was assessed in this study. Kyasanur forest disease virus (KFDV) strain P9605 used in this study was confirmed by real-time reverse transcriptase-polymerase chain reaction targeting the NS5 gene. Mouse Lethal Dose 50 (MLD50) of the virus was determined by in-vivo mice inoculation test. One MLD50 of the KFDV was inoculated intra-cerebrally into 36 mice aged 2-3 weeks. Another group of 36 age-matched mice that served as control group were inoculated with plain media. Six mice each from infected and control groups were euthanized every 24 hrs intervals for six days. Brain tissues were collected in 10% NBF. The collected brain tissues were processed and subjected to histopathological studies by Hematoxylin and Eosin staining. Grossly, the infected mice showed symptoms of dullness, hunched back appearance, weakness, sluggish movements with indication of hind quarter paralysis on day four post-infection. These symptoms got aggravated with complete paralysis of the hind quarters, inability to move and death on 5th and 6th day post-infection. Microscopically, the brain showed apoptosis of neurons, perivascular cuffing, gliosis, congestion, neuropil vacuolation, meningitis, degeneration, and necrotic neurons. The real-time RT-PCR on hippocampus of the KFDV-infected mouse brain showed three-fold higher expression levels of Caspase 3, a crucial mediator of apoptosis. The cerebral cortex, cerebellum and hippocampus that control the motor neuron activities and muscle tone were primarily affected, possibly correlating with the gross symptoms of hind quarter paralysis, ataxia, and other motor neuron dysfunctions noticed. Taken together, these findings reveal that KFDV induces apoptosis of neurons in the cerebrum and hippocampus of KFDV infected mice. Further studies are needed to confirm if the lesions noticed in mice brain simulate the brain lesions in humans since gross motor-neuron symptoms are similar in mice as well as humans.

Cellular Immune Responses Against γ-Inactivated Antigen in the Recovered Cases of Kyasanur Forest Disease.

Kyasanur Forest Disease Virus (KFDV) is a tick-borne flavivirus that causes life-threatening hemorrhagic fever in humans with case fatality rates of 3-5%. Relatively little is known about the mechanism of its pathogenesis or host immune responses to KFDV infection. Here, we investigated KFDV-specific cellular immune responses in the recovered cases of Kyasanur Forest Disease (KFD). Peripheral blood mononuclear cells of the recovered KFD cases and healthy controls were exposed to γ-inactivated KFDV antigen ex vivo. The proliferation index was determined using an enzyme-linked immunosorbent assay-based lymphoproliferative assay. The frequencies of CD4+ and CD8+ T cells expressing intracellular interferon (IFN)-γ in response to stimulation with γ-inactivated KFDV antigen were determined using flow cytometry. A significant increase in lymphoproliferation and a high frequency of CD4+ and CD8+ T cells secreting IFN-γ against γ-inactivated KFDV antigen were found in the recovered KFD group compared to the healthy control group. In conclusion, the study indicated the generation of cellular immune responses in individuals who recovered from KFD and can be used as indicators of cellular immunity in KFD vaccine studies.

Prevalence of ixodid ticks on goats and risk factors for tick-borne diseases in humans in the Kyasanur forest disease endemic area of Kerala, India.

In India, tick-borne diseases are prevalent in many states due to the presence of tick vectors. However, information on disease-causing ticks from domestic animals and the associated risk factors for residents living in the disease-endemic area is lacking. Therefore, we conducted this study to identify ixodid ticks in goats and human risk factors in three villages of the Wayanad district of Kerala. We examined 202 goats and collected 741 ticks, of which 69.8% were ticks belonging to the genus Haemaphysalis. The maximum number of ticks was collected from Thirunelli (81.3%), followed by Noolpuzha (76.27%) and Pulpally (45.6%). Overall, H. bispinosa Neumann, 1897 (54.6%), was the most common species, followed by H. turturis Nuttall and Warburton, 1915 (38.0%), H. spinigera Neumann, 1897 (5.4%), and H. intermedia Warburton and Nuttall, 1909 (1.8%). We included 428 participants (men and women) in this study. The average age of the respondents was 43 years. We found significant associations between accessing the forest for cattle grazing and other activities and tick-borne diseases (χ2 = 9.5, p = 0.002), between workers who were bitten by ticks and tick-borne diseases (χ2 = 3.8, p = 0.05), and between number of tick bites per day > 6 and tick-borne diseases (χ2 = 12.1, p = 0.001). The high frequency of Haemaphysalis spp. found in goats highlighted the risk of tick exposure and tick-borne diseases, such as Kyasanur forest disease in humans, and the need for the development and implementation of effective measures to control ticks.

Knowledge, Attitudes, and Practices Regarding Ticks, Tick-Borne Diseases, and Ethnomedicine Among an at-Risk Population in Kerala.

Tick and tick-borne diseases (TBDs) are increasing annually, and the study of ticks has gained importance after the outbreak of Kyasanur Forest disease (KFD) in the South Western Ghats of India. Wayanad district of Kerala, with the highest tribal population in South India, is a KFD endemic state, owing to the lack of knowledge, attitude, and practice studies on TBDs and ethnomedicines against ticks. This study was carried out to assess their baseline knowledge, attitude, and ethnomedicinal practice against ticks. A structured questionnaire was used to conduct a survey of 499 tribal members living in forest fringe areas. Logistic regression analysis was performed to identify the factors that influence the knowledge, attitude, and practice of tribes on TBDs. More than 70% of the population visit the forests on a regular basis, with 65.7% of the population regularly exposed to tick bites; however, only 47.7% were aware of TBDs. About 47.4% of the respondents took precautions like therapeutics and natural remedies to avoid tick bites. Ten species of medicinal plant belonging to eight different families have been identified from the survey. The tribal population use these plants to repel ticks as well as treat tick bites. From the study, we concluded that the limited in-depth knowledge displayed by the tribes can be strengthened by conducting community programs such as awareness classes on TBD and its control measures. The ethnobotanicals identified can be used to formulate novel tick repellents in the future.

Clinical, epidemiological, and molecular investigation of Kyasanur forest disease from Karnataka state, India during 2018-2019.

BACKGROUND: In this study, we carried out an investigation of Kyasanur Forest Disease (KFD) suspected human cases reported in Karnataka state, India from December 2018 to June 2019. METHODS: The clinical samples of KFD suspected cases (n = 1955) from 14 districts of Karnataka were tested for KFD using real-time RT-PCR and IgM ELISA. Further, the KFD-negative samples were tested for IgM antibodies against dengue and chikungunya viruses. Monkey samples (n = 276) and tick pools (n = 11582) were also screened using real-time RT-PCR. KFD-positive samples were further analysed using next-generation sequencing along with clinico-epidemiological analysis. RESULTS: Of all, 173 (8.8%) cases tested positive for KFD either by real-time RT-PCR (n = 124), IgM ELISA (n = 53) or both tests (n = 4) from seven districts. Among KFD-negative cases, IgM antibody positivity was observed for dengue (2.6%), chikungunya (5.8%), dengue and chikungunya coinfection (3.7%). KFD cases peaked in January 2019 with fever, conjunctivitis, and myalgia as the predominant symptoms and a mortality of 4.6%. Among confirmed cases, 41% received a single dose and 20% received two doses of the KFD vaccine. Of the seven districts with KFDV positivity, Shivamogga and Hassan districts reported KFD viral RNA positivity in humans, monkeys, and ticks. Sequencing analysis of 2019 cases demonstrated a difference of less than 1.5% amino acid compared to prototype KFDV. CONCLUSION: Although the KFD has been endemic in many districts of Karnataka state, our study confirms the presence of KFDV for the first time in two new districts, i.e. Hassan and Mysore. A comparative analysis of KFDV infection among the KFD-vaccinated and non-vaccinated populations demonstrated an insignificant difference.

Single-dose VSV-based vaccine protects against Kyasanur Forest disease in nonhuman primates.

Kyasanur Forest disease virus (KFDV) is an endemic arbovirus in western India mainly transmitted by hard ticks of the genus Haemaphysalis. KFDV causes Kyasanur Forest disease (KFD), a syndrome including fever, gastrointestinal symptoms, and hemorrhages. There are no approved treatments, and the efficacy of the only vaccine licensed in India has recently been questioned. Here, we studied the protective efficacy of a vesicular stomatitis virus (VSV)-based vaccine expressing the KFDV precursor membrane and envelope proteins (VSV-KFDV) in pigtailed macaques. VSV-KFDV vaccination was found to be safe and elicited strong humoral and cellular immune responses. A single-dose vaccination reduced KFDV loads and pathology and protected macaques from KFD-like disease. Furthermore, VSV-KFDV elicited cross-reactive neutralizing immune responses to Alkhurma hemorrhagic fever virus, a KFDV variant found in Saudi Arabia.

Predicting Kyasanur forest disease in resource-limited settings using event-based surveillance and transfer learning.

In recent years, the reports of Kyasanur forest disease (KFD) breaking endemic barriers by spreading to new regions and crossing state boundaries is alarming. Effective disease surveillance and reporting systems are lacking for this emerging zoonosis, hence hindering control and prevention efforts. We compared time-series models using weather data with and without Event-Based Surveillance (EBS) information, i.e., news media reports and internet search trends, to predict monthly KFD cases in humans. We fitted Extreme Gradient Boosting (XGB) and Long Short Term Memory models at the national and regional levels. We utilized the rich epidemiological data from endemic regions by applying Transfer Learning (TL) techniques to predict KFD cases in new outbreak regions where disease surveillance information was scarce. Overall, the inclusion of EBS data, in addition to the weather data, substantially increased the prediction performance across all models. The XGB method produced the best predictions at the national and regional levels. The TL techniques outperformed baseline models in predicting KFD in new outbreak regions. Novel sources of data and advanced machine-learning approaches, e.g., EBS and TL, show great potential towards increasing disease prediction capabilities in data-scarce scenarios and/or resource-limited settings, for better-informed decisions in the face of emerging zoonotic threats.

Simplified visual detection of Kyasanur Forest Disease virus employing Reverse Transcriptase-Polymerase Spiral Reaction (RT-PSR).

Among recently prevalent tick-borne infections in India, Kyasanur Forest Virus Disease (KFD) is an important public health concern. During last decade the emergence of cases apart from endemic zone raised concern about case positivity. Early diagnosis is therefore very important in disease management and primary containment. This study, aimed to develop a simplified viral RNA extraction in combination to dry down format of novel isothermal assay for (Reverse Transcription- Polymerase Spiral reaction) specific and rapid identification of Kyasanur Forest Disease Virus targeting viral envelope gene. The one step method was optimized by magnetic bead based viral RNA extraction followed by isothermal RT-PSR assay in heat bath at 63°C for 60 minutes. Further, visual results interpretation was done by color change of Hydroxy Naphthol Blue dye. The detection limit of the assay was found 10 RNA copies/rxn with comparable to silica column based viral RNA combined real time qPCR. No cross reactivity was observed with other closely related flaviviruses. The assay was evaluated with clinical samples has shown >99% concordance between two methods. This is the first report of sample extraction coupled isothermal detection of KFD in a simplified manner without a need of any hi-end equipment. The assay developed here has potential to use as an alternate for field-based detection in resource limited settings for KFD.

Using mechanistic models to highlight research priorities for tick-borne zoonotic diseases: Improving our understanding of the ecology and maintenance of Kyasanur Forest Disease in India.

The risk of spillover of zoonotic diseases to humans is changing in response to multiple environmental and societal drivers, particularly in tropical regions where the burden of neglected zoonotic diseases is highest and land use change and forest conversion is occurring most rapidly. Neglected zoonotic diseases can have significant impacts on poor and marginalised populations in low-resource settings but ultimately receive less attention and funding for research and interventions. As such, effective control measures and interventions are often hindered by a limited ecological evidence base, which results in a limited understanding of epidemiologically relevant hosts or vectors and the processes that contribute to the maintenance of pathogens and spillover to humans. Here, we develop a generalisable next generation matrix modelling framework to better understand the transmission processes and hosts that have the greatest contribution to the maintenance of tick-borne diseases with the aim of improving the ecological evidence base and framing future research priorities for tick-borne diseases. Using this model we explore the relative contribution of different host groups and transmission routes to the maintenance of a neglected zoonotic tick-borne disease, Kyasanur Forest Disease Virus (KFD), in multiple habitat types. The results highlight the potential importance of transovarial transmission and small mammals and birds in maintaining this disease. This contradicts previous hypotheses that primates play an important role influencing the distribution of infected ticks. There is also a suggestion that risk could vary across different habitat types but currently more research is needed to evaluate this relationship. In light of these results, we outline the key knowledge gaps for this system and future research priorities that could inform effective interventions and control measures.

Dry- down probe free qPCR for detection of KFD in resource limited settings.

Kyasanur Forest Disease is a tick-borne flavivirus is endemic in the Southern India. The recent expansion and resurgence of sporadic outbreaks in southern parts of country is the most important concern. Although only formalin inactivated vaccine is available for treatment with limited efficacy the early detection and timely identification is a only way to prevent spread of cases. If the disease can be identified prior to infection in humans like in forest areas from ticks and vectors the disease spread supposed to be managed quickly. Here we have standardized a single tube ready to use dry-down probe free real time RT-PCR targeted against virus envelope gene for detection of KFDV infection. The assay was standardized in liquid format first, later it was converted into dry-down format with addition of stabilizers with a similar sensitivity and specificity (10RNA Copies/rxn). The sensitivity was comparable to the most widely used and accepted diagnostic platform i.e. TaqMan qRT-PCR. However as the reported assay here omit the need of probes makes it cost effective and dry-down reagents makes more stability to the developed assay in this study if compare to TaqMan qPCR. The assay was evaluated with KFD positive samples and healthy sample panel which revealed high concordance with TaqMan qRT-PCR. Stability was unaffected by temperature fluctuations during transportation even in cold chain free conditions, thus reduce the maintenance of strict cold storage. These findings demonstrated that the reported assay is convenient with 100% sensitivity and specificity to TaqMan qPCR. Thus this assay has the potential usefulness for diagnosis KFDV for routine surveillance in resource limited laboratory settings omitting the use costly and heat sensitive TaqMan qRT-PCR reagents without compromising the sensitivity and specificity of the diagnosis assay.

Sofosbuvir and its tri-phosphate metabolite inhibit the RNA-dependent RNA polymerase activity of non-structural protein 5 from the Kyasanur forest disease virus.

Kyasanur forest disease is a neglected zoonotic disease caused by a single-stranded RNA-based flavivirus, the incidence of which was first recorded in 1957 in the Southern part of India. Kyasanur forest disease virus is transmitted to monkeys and humans through the infected tick bite of Haemophysalis spinigera. Kyasanur forest disease is a febrile illness, which in severe cases, results in neurological complications leading to mortality. The current treatment regimens are symptomatic and supportive, and no targeted therapies are available for this disease. In this study, we evaluated the ability of FDA-approved drugs sofosbuvir (and its active metabolite) and Dasabuvir to inhibit the RNA-dependent RNA polymerase activity of NS5 protein from the Kyasanur forest disease virus. NS5 protein containing the N-terminal methyl transferase domain and C-terminal RNA-dependent RNA polymerase domain was expressed in Escherichia coli, and RNA-dependent RNA polymerase activity was demonstrated with the purified protein. The RNA-dependent RNA polymerase assay conditions were optimized, followed by the determination of apparent Km,ATP to validate the enzyme preparation. Half maximal-inhibitory concentrations against RNA-dependent RNA polymerase activity were determined for Sofosbuvir and its active metabolite. Dasabuvir did not show detectable inhibition with the tested conditions. This is the first demonstration of the inhibition of RNA-dependent RNA polymerase activity of NS5 protein from the Kyasanur forest disease virus with small molecule inhibitors. These initial findings can potentially facilitate the discovery and development of targeted therapies for treating Kyasanur forest disease.

Development and application of a recombinant Envelope Domain III protein based indirect human IgM ELISA for Kyasanur forest disease virus.

Kyasanur forest virus disease (KFD) is a major public health concern in India. Its etiology KFD virus causes haemorrhagic fever with severe sequelae in humans. Due to continuous spatiotemporal expansion of KFD in last decade, the incidences of positive cases have been increasing in both humans and primates. Early diagnosis is of prime importance for disease management and epidemiological containment. In the present study, the highly immunogenic Envelope Domain III (EDIII) antigen was produced using prokaryotic expression system with an yield of 8 mg/L. The protein was purified using affinity chromatography and confirmed for its immuno-reactivity by western blot and UPLCMS/MS analysis. The recombinant EDIII was used as an antigen for the standardization of ELISA to detect anti KFD IgM antibodies in humans. The ROC curve was prepared to set the optimum cut-off OD for the assay. The comparative evaluation of the assay with a reference MAC ELISA revealed 86.96% concordance, 82.22% sensitivity and 91.48% specificity. Inter-rater agreement was performed with kappa index revealing significant agreement between the assays. This is the first study using safe recombinant protein antigen-based detection of anti KFDV antibodies in humans. This simple and scalable ELISA assay will be applicable for large scale screening of samples for combating the emerging threats of KFD in newer territories.

Impact of Plantation Induced Forest Degradation on the Outbreak of Emerging Infectious Diseases-Wayanad District, Kerala, India.

The world has been facing a pandemic owing to COVID-19. We have also seen the geographic expansion and outbreaks of other emerging infectious diseases (EID) in recent years. This paper investigates the direct and indirect effects of land use land cover change (LULCC) on EID outbreaks in the context of Wayanad District of Kerala, India. Wayanad is in the vulnerable tropical forested region, and it is named as one of the four environmental change hotspots. The focus of this project is mainly three EIDs prevalent in this region: Kyasanur forest disease (KFD), Dengue and Leptospirosis. Our results, based on topographical map, remote sensing and extensive field work, show that the natural forest in Wayanad was replaced with agriculture and forest plantation during 1950-2018. This paper further suggests that encroachment of forest by forest plantation causes the human-animal conflict resulting in the outbreak of KFD cases. Our analysis reveals that a high number of Dengue cases is found in the forested regions of the district and over the adjacent human-made agriculture plantation areas. High and medium number of Leptospirosis cases contain a high portion of land area devoted to paddy cultivation and agricultural plantation. In summary, the results clearly show the linkage between the outbreak of above mentioned EIDs and LULCC in the context of Wayanad district, Kerala. We also discuss in detail the causal pathway involving human-environmental dynamics through which plantation leads to the outbreak of KFD. Replacing forests with plantations poses an alarming threat of disease outbreak in the community.

Effectiveness of One Health approach for control of Kyasanur Forest Disease in Wayanad, Kerala, India.

BACKGROUND & OBJECTIVES: Kyasanur Forest Disease (KFD) is a vector borne haemorrhagic fever that is endemic in the Wayanad region located in Northern part of Kerala, India. The region is managing the outbreak well ever since the major epidemic of 2015. This was because of the successful implementation of One Health (OH) initiative concentrating on multisectoral collaboration between regional institutions involved in public, animal and environmental health domains. The article presents how OH was implemented for the first time in the district in the year 2015 and evaluates the degree OH-ness of the Initiative. METHODS: The OH approach involved trans-disciplinary stakeholder meetings and reviews, outbreak management and integrated surveillance targeting ticks, monkeys and humans. The degree of OH-ness used for addressing KFD during the year 2015 was evaluated following the protocol developed by the Network for Evaluation of One Health (NEOH). In detail, we (i) described the OH initiative and its system (Aim, stakeholders, action strategy) and (ii) scored different aspects of this initiative (i.e., OH-thinking, -planning, -working, -sharing, -learning, -organization), with values from 0 (=no OH approach) to 1 (=perfect OH approach). RESULTS: We obtained a median score for each aspect evaluated. We reached high scores for OH systemic organization (1.0), OH thinking (0.83) and OH working (0.83). Lower scores were attributed to OH planning (0.58), OH sharing (0.50) and OH learning (0.33). The OH index was 0.36 and OH ratio was 0.95, indicating a balance between the OH operations and supporting infrastructures. INTERPRETATION & CONCLUSION: With this we could high-light some critical issues related to communication on sharing data as well as learning gaps for consideration to control future outbreaks. The strengths and weaknesses detected may be used to refine the initiative, aiming to provide a basis for the development of shared recommendations in a more OH-oriented perspective. This model of evaluation criteria will serve to create a database of OH success stories in India that will in turn help to institutionalize the approach at ministerial level. Future India is moving towards implementing a One Health, hence, this study data will provide an ideal opportunity for all sectors to control any vector borne diseases.

Kyasanur forest disease and climatic attributes in India.

BACKGROUND & OBJECTIVES: In India, Kyasanur Forest Disease has been reported from the states of Karnataka, Kerala, Goa, and Maharashtra. The relationship between climatic factors and transmission of KFD remains untouched, therefore, the present study was undertaken. METHODS: Based on the occurrence of cases, Shivamogga district (Karnataka) and Wayanad district in Kerala and northern Goa (Goa state) were selected for the study. Data on the incidence of KFD and climate factors were collected from concerned authorities. To determine the relationship between dependent and independent variables, spearman's correlation was calculated for monthly as well as with lag months. RESULTS: KFD cases and temperature (°C) were found significantly correlated up to 1 months' lag period (p<0.05) while with precipitation relationship was found negatively significant for 0-3 months' lag. The range of suitable temperature for KFD in Shivamogga, Goa and Wayanad was found as 20-31°C, 25-29°C and 27-31°C respectively. The cumulative precipitation during transmission months (November-May) ranged from <150-500mm, while in non-transmission months (June-October) from >1100-2400mm. INTERPRETATION & CONCLUSION: The analysis of three sites revealed that with the increase in temperature, the intensity of KFD transmission decreases as corroborated by the seasonal fluctuations in Shivamogga, Goa and Wayanad. High precipitation from June to October rovides suitable ecology to tick vector and sets in transmission season from November to May when cumulative precipitation is <500 mm.

Kinetics of human infection with Kyasanur Forest Disease Virus.

BACKGROUND: Kyasanur Forest disease (KFD) was first reported in 1957 and became an emerging tick-borne viral disease of public health importance in India. However, very little is known about the host-virus interaction and pathogenesis of KFD in humans. This study described the presence, duration, and kinetics of KFDV RNA in body fluids in infected human cases. METHODOLOGY: We enrolled 76 laboratory-confirmed KFD individuals and followed them up in the study. We obtained serial samples of blood, throat swabs in viral transport medium (VTM), urine, stool, and semen during the acute and convalescent phase of KFD illness. In addition, specimens were inactivated, and nucleic acid was extracted and tested for KFDV real-time reverse transcriptase -PCR. Clinical data was also obtained from the subjects. RESULT: The study provides evidence of KFD virus RNA in different biological body fluids of humans. The percentage positivity of KFDV RNA in blood was 100% during the first four days of illness. PCR became negative in most cases by 7-8 days; a subset of cases (14%) had prolonged viremia for up to 15 days post-onset of illness. Relatively low copies of KFDV RNA were also detected in throat swabs and urine in the first week of illness. In addition, we detected KFDV RNA in stool samples of cases of those who had diarrhea at an early stage of infection. CONCLUSION: The study provides evidence of KFDV RNA in different biological body fluids, which will help understand the pathogenesis, transmission pattern and develop diagnostic algorithms of KFDV in humans. In Kyasanur Forest disease infection, the blood has more RNA copies/ml than other body fluids, and viremia may last up to two weeks post-infection.

Neurological manifestations of Kyasanur Forest disease: a retrospective cohort study from South India.

BACKGROUND: Kyasanur Forest disease (KFD) is a viral zoonotic disease where patients present with febrile illness and haemorrhagic manifestations in the first phase. In a small fraction of patients, the fever may be biphasic. This study aimed to describe the neurological manifestations of patients with KFD in the first and second phases of the illness. METHODS: This is a retrospective cohort study of 297 patients admitted with a molecular diagnosis of KFD from December 2018 to December 2020. The case records of these patients were reviewed for evidence of neurological involvement. RESULTS: A total of 34 (11.5%) patients in the first phase and 16 (36.4%) patients in the second phase had neurological involvement. Altered sensorium, seizures and focal infarcts were common in the first phase, while cerebellar signs and leptomeningeal enhancement were common in the second phase. CONCLUSIONS: Neurological involvement is seen in both phases of KFD. While in the first phase it is a result of possible encephalitis/encephalopathy, the second phase involvement is possibly due to postinfectious cerebellitis or meningitis.

Exploring potential inhibitors against Kyasanur forest disease by utilizing molecular dynamics simulations and ensemble docking.

Kyasanur forest disease (KFD) is a tick-borne, neglected tropical disease, caused by KFD virus (KFDV) which belongs to Flavivirus (Flaviviridae family). This emerging viral disease is a major threat to humans. Currently, vaccination is the only controlling method against the KFDV, and its effectiveness is very low. An effective control strategy is required to combat this emerging tropical disease using the existing resources. In this regard, in silico drug repurposing method offers an effective strategy to find suitable antiviral drugs against KFDV proteins. Drug repurposing is an effective strategy to identify new use for approved or investigational drugs that are outside the scope of their initial usage and the repurposed drugs have lower risk and higher safety compared to de novo developed drugs, because their toxicity and safety issues are profoundly investigated during the preclinical trials in human/other models. In the present work, we evaluated the effectiveness of the FDA approved and natural compounds against KFDV proteins using in silico molecular docking and molecular simulations. At present, no experimentally solved 3D structures for the KFD viral proteins are available in Protein Data Bank and hence their homology model was developed and used for the analysis. The present analysis successfully developed the reliable homology model of NS3 of KFDV, in terms of geometry and energy contour. Further, in silico molecular docking and molecular dynamics simulations successfully presented four FDA approved drugs and one natural compound against the NS3 homology model of KFDV. Communicated by Ramaswamy H. Sarma.

Kyasanur forest disease: a state-of-the-art review.

Kyasanur forest disease (KFD) virus is a flavivirus that can be transmitted to humans from monkeys or other mammals through hard ticks (Haemaphysalis spinigera). The disease is endemic to 16 districts in 5 states of Southern India and is reported in the dry season, most commonly in humans travelling to the forests in these areas. The aim of this systematic review is to raise awareness of the clinical and laboratory manifestation of KFD among physicians and travel medicine practitioners. A total of 153 articles were screened of which 16 articles that met the inclusion and exclusion criteria were included for qualitative analysis. KFD is an acute haemorrhagic fever with a biphasic component in some individuals. The second phase is usually marked by neurological symptoms. Leucopoenia, thrombocytopenia and elevated transaminases are the hallmarks of the first phase of KFD. The diagnostic modality of choice in the first few days of illness is polymerase chain reaction assay, whereas serology is used in the late phase. In the absence of a specific antiviral treatment, the clinical management of patients is limited to supportive care. Avoidance of exposure and vaccination is recommended to prevent this infection.

Species distribution and ecological suitability analysis for potential tick vectors of Kyasanur forest disease in the Western Ghats of Kerala.

Kyasanur forest disease (KFD) is a tick-borne zoonotic viral disease with an estimated case fatality rate of 3% to 5% in humans. The KFD virus is transmitted to both humans and animals by infected ticks, primarily the Haemaphysalis species. Our study was undertaken following reports of five confirmed cases and three deaths due to KFD in Malappuram district of Kerala in 2014 to determine the diversity of Ixodidae ticks on vegetation and their possible infection with the KFD virus. Overall, 3502 hard ticks belonging to three genera and eight species were collected from two forest divisions. Haemaphysalis was the predominant tick species (92.72%). Tick positivity for the KFD virus was 4 (5.33%) of 75 pools of ticks tested. Kyasanur forest disease viral RNA was detected from the genera Haemaphysalis and Amblyomma. The KFD virus was detected in 2 of 35 pools (5.71%) of Haemaphysalis spinigera, 1 of 30 pools (3.33%) of Haemaphysalis turturis, and 1 of 3 pools (3.33%) of Amblyomma integrum from the south forest division. The ticks reached their peak density between December and February and then decreased from the end of May. The temperature in the area ranges from 28°C to 30°C, which is suitable for tick survival.