Location: Zoonotic and Emerging Disease Research
Project Number: 3022-32000-021-006-S
Project Type: Non-Assistance Cooperative Agreement
Start Date: Sep 1, 2022
End Date: Apr 30, 2027
Objective:
The emphasis of the agreement will be on examining Hyalomma species of ticks for pathogens that affect animal and human health. The objectives of this agreement will be to identify tick species diversity of the Ugandan cattle corridor and the Kenyan arid north region, assess tick blood meal sources, delineate temporal and spatial Crimean-Congo Hemorrhagic fever virus (CCHFV) infection rates in tick populations, and characterize the tick virome. CCHFV is an enveloped, negative-sense RNA virus, vectored principally by Hyalomma species of ticks. CCHFV causes a contagious zoonosis resulting in haemorrhagic fever with a case fatality rate between 3% and 80%. CCHFV also possesses a high degree of genetic diversity with a potential of new and highly-pathogenic variants emerging, which can impact pastoralists in contact with animals infested with Hyalomma ticks. Despite the occurrence of multiple outbreaks in East Africa since 2013, CCHFV remains poorly characterized with paucity of knowledge of its epidemiology, population burden, and viral characteristics. The data obtained in this study will establish the prevalence of CCHFV and other tick-borne pathogens, as well as the intra- and interpopulation genetic variability of CCHFV isolated from ticks collected in different seasons and ecological zones, and will identify any novel pathogens transmitted by ticks along the Kenya-Ugandan border. These studies will also help prepare the United States agriculture and public health systems for the potential introduction of CCHFV or other tick-borne emerging pathogens by gaining a greater understanding of the epidemiology of CCHFV transmission in East Africa. Collaborators will collect ticks from livestock, camels, goats and sheep in the homesteads, market, and abattoirs during the wet and dry seasons found in the Ugandan cattle corridor and the Kenyan arid north regions. Taxonomic keys and genetic tools will be used for identification of ticks. Transmission dynamic risks of CCHFV will be assessed by identifying the blood meal sources of each tick species while spill over risks will be determined by comparing the temporal and spatial infection rates associated with different tick species. Further, to identify other viruses transmitted by ticks collected from these regions, collaborators will utilize deep sequencing to characterize the tick virome of a few pools of ticks representing the different tick species collected from the Uganda and Kenya. Additionally, this will assess the variation of the tick virome across seasons. The results of this project will identify the incidence of CCHFV in tick populations and provide detailed information on the species of ticks infected by CCHFV, as well as the identification of novel viruses transmitted by ticks in these regions.
Approach:
1. Identification of tick species diversity in diverse ecosystems of the Ugandan cattle corridor and the Kenyan arid north regions. Tick samples will be collected from animal livestock hosts, pastures/vegetation grazed by cattle, wildlife or mixtures of wildlife and livestock by using the blanket dragging method. Ticks will be collected from predilection sites (e.g. ears, under the tail of cattle) using forceps. All ticks will be flash frozen and stored in liquid nitrogen until speciation. The tick samples will be transported to sites in Uganda or in Kenya. Each tick will be surface sterilized by washing twice with sterile water to remove excess particulate contamination from animal skin, rinsed once with 70% ethanol and then rinsed twice with sterile PBS. Ticks will be identified using morphological features to species and DNA and RNA isolated. Isolated DNA and RNA will be shipped to Cooperator and/or the USDA ARS for further processing and species confirmation using CO1 mitochondrial single-step PCR and sanger sequencing.
2. Assess tick blood meal sources of potential vectors collected from unique spatial regions. Field collected tick spp. will be classified according to levels of engorgement. Genomic DNA of engorged ticks will be extracted and will serve as a template for a standard nested PCR assay to amplify a fragment of a vertebrate cytochrome c oxidase subunit I (COI) mitochondrial gene. PCR amplicons will be cleaned, sequenced using Sanger sequencing, and identified by blasting on the Genbank Blastn database.
3. Delineate temporal and spatial CCHFV infection rates of tick populations. After identification of the ticks collected from the aforementioned sites in Uganda and Kenya, RNA will be isolated using tick specific isolation protocols and used to calculate CCHFV prevalence.
4. Characterize the tick virome of the Ugandan cattle corridor and Kenyan arid north tick populations. To identify other viruses transmitted by ticks and to determine the intra- and interpopulation genetic variability of CCHFV, we will utilize the Illumina sequencing platform to characterize the tick virome of pools of ticks representing the different tick species collected from the Ugandan cattle corridor and Kenyan arid north. In addition, phylogenetic studies to compare Ugandan and Kenyan CCHFV strains and to all publicly available strains in the database will be completed. Bayesian and maximum likelihood methodology will be used to infer phylogenetic relationships and assess temporal and geographic patterns.
