Molecular Survey and Genetic Characterization of Anaplasma marginale in Ticks Collected from Livestock Hosts in Pakistan

Simple Summary Ticks transmit different disease-causing agents to humans and animals. Pakistan is an agricultural country, the rural economy mainly relies on livestock farming, and tick infestation is a severe constraint to its livelihood. The genus Anaplasma comprises obligate Gram-negative intracellular bacteria multiplying within the host cells and can be transmitted to humans and animals through the tick vector. The current study aimed to molecularly characterize the Anaplasma spp. in hard ticks infesting livestock in different districts of Khyber Pakhtunkhwa, Pakistan. The present study reported nine species of hard ticks infesting different hosts. The most prevalent tick life stage was adult females, followed by nymphs and adult males. In the phylogenetic tree, 16S rDNA sequences of Anaplasma spp. clustered with sequences of A. marginale. The hard ticks act as a carrier for the transmission of A. marginale. Further extensive country-wide research is required to explore the diverse tick species and the associated pathogens in Pakistan. Ticks transmit pathogens to animals and humans more often than any other arthropod vector. The rural economy of Pakistan mainly depends on livestock farming, and tick infestations cause severe problems in this sector. The present study aimed to molecularly characterize the Anaplasma spp. in hard ticks collected from six districts of Khyber Pakhtunkhwa, Pakistan. Ticks were collected from various livestock hosts, including cattle breeds (Holstein-Friesian, Jersey, Sahiwal, and Achai), Asian water buffaloes, sheep, and goats from March 2018 to February 2019. Collected ticks were morphologically identified and subjected to molecular screening of Anaplasma spp. by amplifying 16S rDNA sequences. Six hundred seventy-six ticks were collected from infested hosts (224/350, 64%). Among the nine morphologically identified tick species, the highest occurrence was noted for Rhipicephalus microplus (254, 37.6%), followed by Hyalomma anatolicum (136, 20.1%), Rhipicephalus haemaphysaloides (119, 17.6%), Rhipicephalus turanicus (116, 17.1%), Haemaphysalis montgomeryi (14, 2.1%), Hyalomma dromedarii (11, 1.6%), Haemaphysalis bispinosa (10, 1.5%), Hyalomma scupense (8, 1.2%), and Haemaphysalis kashmirensis (8, 1.2%). The occurrence of tick females was highest (260, 38.5%), followed by nymphs (246, 36.4%) and males (170, 25.1%). Overall, the highest occurrence of ticks was recorded in the Peshawar district (239, 35.3%), followed by Mardan (183, 27.1%), Charsadda (110, 16.3%), Swat (52, 7.7%), Shangla (48, 7.1%), and Chitral (44, 6.5%). Among these ticks, Anaplasma marginale was detected in R. microplus, R. turanicus, and R. haemaphysaloides. The 16S rDNA sequences showed high identity (98-100%) with A. marginale reported from Australia, China, Japan, Pakistan, Thailand, Uganda, and the USA. In phylogenetic analysis, the sequence of A. marginale clustered with the same species reported from Australia, China, Pakistan, Thailand, Uruguay, and the USA. Further molecular work regarding the diversity of tick species and associated pathogens is essential across the country.