Multiplex-PCR differentiation of two Hyalomma and two Haemaphysalis species (Acari: Ixodidae)

Hosseini Chegeni, Asadollah; Kayedi, Mohammad Hassan; Hosseini, Reza; Telmadarraiy, Zakkyeh

doi:10.22073/pja.v6i2.24392

Multiplex-PCR differentiation of two Hyalomma and two Haemaphysalis species (Acari: Ixodidae)

Authors

Asadollah Hosseini Chegeni ¹

Mohammad Hassan Kayedi ²

Reza Hosseini ³

Zakkyeh Telmadarraiy ⁴

¹ Razi Herbal Medicines Research Center, Lorestan University of Medical Sciences, Khorramabad, Iran;

² Department of Parasitology and Mycology, School of Medicine, Lorestan University of Medical Sciences, Khorram-abad, Iran;

³ Department of Medical Entomology and Vector Control, School of Public Health, Tehran University of Medical Sciences, Tehran, Iran;

⁴ Department of Plant Protection, Faculty of Agriculture, University of Guilan, Guilan, Iran;

10.22073/pja.v6i2.24392

Abstract

Hyalomma and Haemaphysalis are two most important genera of ticks (Acari: Ixodidae). The rapid and accurate identification of field collected specimens is crucial in faunal works, laboratory assays, anti-tick vaccine experiments, etc. The present study was designed to introduce a rapid and more sensitive method for the differentiation of closely related Hyalomma and Haemaphysalis species namely the pairs Hy. anatolicum-Hy. asiaticum and Ha. punctata-Ha. sulcata, as the main vectors of different animal and human pathogenic agents. Tick specimens were collected from domestic animals in Ardabil, Hormozgan, Khuzestan, Kurdistan, Lorestan, and Mazandaran and identified according to the taxonomic keys. DNA was extracted by the Phenol-Chloroform method. Then, PCR was carried out in a single PCR reaction tube using three pairs designed primers (one forward and two reverse) adapted from the internal transcribe spacer 2 (ITS2) and cytochrome oxidase subunit 1 (COI) genes for Hyalomma and Haemaphysalis, respectively. In the present study, results of Multiplex-PCR revealed that the pairs Hy. anatolicum-H. asiaticum and Ha. punctata-Ha. sulcata could be well differentiated on gel electrophoresis. Morphological misidentification of Hy. anatolicum-Hy. asiaticum and Ha. punctata-Ha. sulcata could be reduced significantly after using Multiplex-PCR.

Keywords

Closely related species

COI

ITS2

Hyalomma

Haemaphysalis

Identification