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Kyasanur Forest Disease (KFD), also known as Monkey Fever, is an important tick-borne viral infection in India. It is mainly transmitted by the hard tick vector, Haemaphysalis spinigera . This re-emerging viral disease in humans has a mortality rate of 2–10% and was first reported in the Shimoga district of Karnataka in 1957. In recent years, KFD has been reported from various parts of the country, indicating possible geographical expansion and wider distribution of the tick vector. Assessing the distinct populations of H. spinigera and their local adaptations is important. However, the genetic structure of this vector population remains undocumented due to the lack of well-characterized genetic markers. Although the distribution of H. spinigera has been described, information on its genetic diversity and population structure is still limited. In this study, we characterized and validated a mitochondrial marker, 12S rRNA for assessing the genetic diversity of H. spinigera in Eastern and Western Ghats of India. Analysis of ninety individual ticks revealed a genetically diverse population comprising three haplotypes, which formed three distinct clades. These clades included haplotypes from nine geographical populations. Understanding the genetic structure of tick vectors is crucial for studying the transmission dynamics of tick-borne diseases. Our findings suggest that 12S rRNA is a suitable genetic marker for studying intraspecific variation in H. spinigera . To the best of our knowledge, this is the first to report the population structure of H. spinigera , highlighting its genetic diversity in the Western and Eastern Ghats of India.

The Kyasanur Forest Disease (KFD) has become a major public health problem in the State of Karnataka, India where the disease was first identified and in Tamil Nadu, Maharashtra, Kerala, and Goa covering the Western Ghats region of India. The incidence of positive cases and distribution of the Kyasanur Forest Disease virus (KFDV) in different geographical regions raises the need to understand the evolution and spatiotemporal transmission dynamics. Phylogeography analysis based on 48 whole genomes (46 from this study) and additionally 28 E-gene sequences of KFDV isolated from different regions spanning the period 1957–2017 was thus undertaken. The mean evolutionary rates based the E-gene was marginally higher than that based on the whole genomes. A subgroup of KFDV strains (2006–2017) differing from the early Karnataka strains (1957–1972) by ~2.76% in their whole genomes and representing spread to different geographical areas diverged around 1980. Dispersal from Karnataka to Goa and Maharashtra was indicated. Maharashtra represented a new source for transmission of KFDV since ~2013. Significant evidence of adaptive evolution at site 123 A/T located in the vicinity of the envelope protein dimer interface may have functional implications. The findings indicate the need to curtail the spread of KFDV by surveillance measures and improved vaccination strategies.

In South Asia, tick transmits Kyasanur Forest Disease Virus (KFDV), a flavivirus that causes severe hemorrhagic fever with neurological manifestations such as mental disturbances, severe headache, tremors, and vision deficits in infected human beings with a fatality rate of 3-10%. The disease was first reported in March 1957 from Kyasanur forest of Karnataka (India) from sick and dying monkeys. Since then, between 400 and 500 humans cases per year have been recorded; monkeys and small mammals are common hosts of this virus. KFDV can cause epizootics with high fatality in primates and is a level-4 virus according to the international biosafety rules. The density of tick vectors in a given year correlates with the incidence of human disease. The virus is a positive strand RNA virus and its genome was discovered to code for one polyprotein that is cleaved post-translationally into 3 structural proteins (Capsid protein, Envelope Glycoprotein M and Envelope Glycoprotein E) and 7 non-structural proteins (NS1, NS2A, NS2B, NS3, NS4A, NS4B, and NS5). KFDV has a high degree of sequence homology with most members of the TBEV serocomplex. Alkhurma virus is a KFDV variant sharing a sequence similarity of 97%. KFDV is classified as a NIAID Category C priority pathogen due to its extreme pathogenicity and lack of US FDA approved vaccines and therapeutics; also, the infectious dose is currently unknown for KFD. In India, formalin-inactivated KFDV vaccine produced in chick embryo fibroblast is being used. Nevertheless, further efforts are required to enhance its long-term efficacy. KFDV remains an understudied virus and there remains a lack of insight into its pathogenesis; moreover, specific treatment to the disease is not available to date. Environmental and climatic factors involved in disseminating Kyasanur Forest Disease are required to be fully explored. There should be a mapping of endemic areas and cross-border veterinary surveillance needs to be developed in high-risk regions. The involvement of both animal and health sector is pivotal for circumscribing the spread of this disease to new areas.

Haemaphysalis ticks are significant vectors of multiple pathogens and pose substantial threats to both public and veterinary health. Among them, H. bispinosa and H. turturis are important vectors of Kyasanur Forest Disease Virus (KFDV) in South India. Although both species are well recognized, detailed comparative descriptions of their morphological and morphometric characteristics remain limited. This study aimed to expand the current understanding of these species by describing and quantifying morphological differences across all developmental stages ( n  = 15 per stage) through a combination of morphological, morphometric, and statistical approaches, using specimens collected from two geographically distinct populations in South India. Scanning Electron Microscopy (SEM) provided high-resolution structural comparisons, while Principal Component Analysis (PCA) was used to assess morphometric variation and distinguish key diagnostic features. The results revealed clear interspecific differences, particularly in the scutum and idiosoma. Haemaphysalis bispinosa exhibited a broader basis capituli, while H. turturis displayed a wider and shorter body form, along with a distinct ventral spur morphology on the coxa III segment. These findings confirm that H. bispinosa and H. turturis are morphometrically distinct species and underscore the value of integrative morphological and statistical tools for resolving cryptic species complexes. This work contributes to the biosystematic understanding and accurate identification of medically important Haemaphysalis ticks in endemic regions.

Background Kyasanur forest disease virus (KFDV) is a tick-borne flavivirus causing debilitating and potentially fatal disease in people in the Western Ghats region of India. The transmission cycle is complex, involving multiple vector and host species, but there are significant gaps in ecological knowledge. Empirical data on pathogen-vector-host interactions and incrimination have not been updated since the last century, despite significant local changes in land use and the expansion of KFD to new areas. Mathematical models predict that transovarial transmission, whereby adult female ticks pass KFDV infections to their offspring, plays an important role in the persistence of KFD, but this has not been shown in the wild. Here we set out to establish whether transovarial transmission of KFDV was occurring under natural field conditions by assessing whether host-seeking larvae were positive for KFDV. Methods Ticks were sampled by dragging and flagging across a broad range of habitats within the agro-forest matrix at 49 sites in two districts: Shivamogga, Karnataka and Wayanad, Kerala (September 2018-March 2019), and larvae were tested for KFDV by PCR. Results In total, larval ticks from 7 of the 49 sites sampled tested positive for KFDV, indicating that transovarial transmission is occurring. Of the 13 KFDV-positive larval samples, 3 came from around houses and gardens, 5 from crops (3 from harvested rice paddy and 2 from areca plantation), 1 from teak plantation and 4 (2 from 1 transect) from forests. Five different tick species were found to have KFDV-positive larvae: Haemaphysalis spinigera, H. bispinosa, Rhipicephalus annulatus, R. microplus and an unidentifiable species of Haemaphysalis (no close match in GenBank). Conclusions Our empirical confirmation of transovarial transmission has important implications for understanding and predicting KFD dynamics, suggesting that ticks may act as a reservoir for KFDV. Moreover, small mammals and cattle may play crucial roles in transmission if small mammals are the main hosts for larvae infected via transovarial transmission, and cattle support large numbers of infected female adult ticks. This first report of transovarial transmission of KFDV, and within a hitherto undescribed range of vectors and habitats, will help disease managers improve KFD surveillance and mitigation strategies, ultimately leading to communities becoming more resilient to the risk of this tick-transmitted disease. Graphical Abstract

Kyasanur Forest disease (KFD), a tick-borne viral hemorrhagic fever, is endemic in five districts of Karnataka state, India. Recent reports of the spread of disease to neighboring districts of the Western Ghats, namely Chamarajanagar district in Karnataka, Nilgiri district in Tamil Nadu, Wayanad and Malappuram districts in Kerala, and Pali village in Goa are a cause for concern. Besides vaccination of the affected population, establishing an event-based surveillance system for monkey deaths in the national parks, wildlife sanctuaries and reserve forests of the Western Ghats would help detect the disease early and thereby help implement appropriate control measures.

KFD is more localized in certain geographical areas, namely, five districts of Karnataka, Chamarajanagar, Nilgiri (Tamil Nadu), Wayanad and Malappuram (Kerala), Pali village (Goa) and recently confirmed in Sindhudurg, Maharashtra State [2],[13], where it occurs in outbreak form, whereas only sporadic cases or import cases of CCHF virus (CCHFV) have been reported from Gujarat, Rajasthan and Uttar Pradesh [2]. The humans acquire infection when they come in close contact of this environment. [...]only sporadic cases occur for CCHF though the virus is widely prevalent in various geographical areas. [...]monkeys and forest dwellers acquire KFD infections.

Zoonoses disproportionately affect tropical communities and are associated with human modification and use of ecosystems. Effective management is hampered by poor ecological understanding of disease transmission and often focuses on human vaccination or treatment. Better ecological understanding of multi-vector and multi-host transmission, social and environmental factors altering human exposure, might enable a broader suite of management options. Options may include “ecological interventions” that target vectors or hosts and require good knowledge of underlying transmission processes, which may be more effective, economical, and long lasting than conventional approaches. New frameworks identify the hierarchical series of barriers that a pathogen needs to overcome before human spillover occurs and demonstrate how ecological interventions may strengthen these barriers and complement human-focused disease control. We extend these frameworks for vector-borne zoonoses, focusing on Kyasanur Forest Disease Virus (KFDV), a tick-borne, neglected zoonosis affecting poor forest communities in India, involving complex communities of tick and host species. We identify the hierarchical barriers to pathogen transmission targeted by existing management. We show that existing interventions mainly focus on human barriers (via personal protection and vaccination) or at barriers relating to Kyasanur Forest Disease (KFD) vectors (tick control on cattle and at the sites of host (monkey) deaths). We review the validity of existing management guidance for KFD through literature review and interviews with disease managers. Efficacy of interventions was difficult to quantify due to poor empirical understanding of KFDV–vector–host ecology, particularly the role of cattle and monkeys in the disease transmission cycle. Cattle are hypothesised to amplify tick populations. Monkeys may act as sentinels of human infection or are hypothesised to act as amplifying hosts for KFDV, but the spatial scale of risk arising from ticks infected via monkeys versus small mammal reservoirs is unclear. We identified 19 urgent research priorities for refinement of current management strategies or development of ecological interventions targeting vectors and host barriers to prevent disease spillover in the future.

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 ( χ 2  = 9.5, p  = 0.002), between workers who were bitten by ticks and tick-borne diseases ( χ 2  = 3.8, p  = 0.05), and between number of tick bites per day > 6 and tick-borne diseases ( χ 2  = 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. Graphical abstract

The present manuscript deals with experimental infections of bonnet macaques ( Macaca radiata ) to study disease progression for better insights into the Kyasanur Forest Disease (KFD) pathogenesis and transmission. Experimentally, 10 monkeys were inoculated with KFD virus (KFDV) (high or low dose) and were regularly monitored and sampled for various body fluids and tissues at preset time points. We found that only 2 out of the 10 animals showed marked clinical signs becoming moribund, both in the low dose group, even though viremia, virus shedding in the secretions and excretions were evident in all inoculated monkeys. Anti-KFDV immunoglobulin (Ig)M antibody response was observed around a week after inoculation and anti-KFDV IgG antibody response after two weeks. Anaemia, leucopenia, thrombocytopenia, monocytosis, increase in average clotting time, and reduction in the serum protein levels were evident. The virus could be re-isolated from the skin during the viremic period. The persistence of viral RNA in the gastrointestinal tract and lymph nodes was seen up to 53 and 81 days respectively. Neuro-invasion was observed only in moribund macaques. Re-challenge with the virus after 21 days of initial inoculation in a monkey did not result in virus shedding or immune response boosting.