| Home  | About ScienceAsia  | Publication charge  | Advertise with us  | Subscription for printed version  | Contact us  
Editorial Board
Journal Policy
Instructions for Authors
Online submission
Author Login
Reviewer Login
Volume 50 Number 1
Volume 49 Number 6
Volume 49 Number 5
Volume 49S Number 1
Volume 49 Number 4
Volume 49 Number 3
Earlier issues
Back

Research Article

ScienceAsia 25 (1999) : 085-090 |doi: 10.2306/scienceasia1513-1874.1999.25.085

 

Detection of Bovine Hemoparasite Infection Using Multiplex Polymerase Chain Reaction

Chantra Tananyutthawongesea, Klaokwan Saengsombutb, Wasana Sukhumsirichata, Wanlaya Uthaisanga, Nopporn Sarataphanc and Kosum Chansiria*


ABSTRACT: A rapid and specific multiplex polymerase chain reaction has been developed for the diagnosis of hemoparasitic infection of bovine blood by five of the most common protozoan hemoparasites. This method relied on the detection of the presence of genomic DNA from the five different bovine hemoparasites isolated from red blood cells. Infection was detected by a single multiplex PCR reaction containing five pairs of oligonucleotide primers whereby each primer pair was specific for each parasite species. These primer sets amplify 160, 257, 446, 689 and 1,100 bp fragments for Anaplasma marginale, Trypanosoma evansi, Babesia bovis, Babesia bigemina and Theileria sp, respectively. The PCR products derived from each parasite species were visualized in ethidium bromide-stained agarose gels, thus allowing the rapid identification of any, or all, of the five bovine parasites, if present, in a single amplification reaction. This multiplex PCR was sensitive with the ability to detect the presence of as little as 10-1 pg of parasite DNA. The primers used in this multiplex PCR also showed highly specific amplification of each respective parasite DNA without the presence of non-specific and non-target PCR products. This multiplex PCR system was used to analyze 35 bovine blood samples for the presence of parasitic infection and the results were compared with detection of hemoparasites by light microscopic examination after Giemsa staining of thin film blood smears. A comparison of the two detection methods revealed that 86% of specimens showed concordant diagnoses with both techniques. However, 14% of the samples examined showed single or multiple infection by multiplex PCR analysis, but such infection was not detected by microscopy. The simplicity and rapidity of this specific multiplex PCR method make it suitable for large-scale epidemiological studies and for follow-up of drug treatments.

Download PDF


a Department of Biochemistry, Faculty of Medicine, Srinakharinwirot University, Sukhumvit 23, Bangkok 10110, Thailand.
b Department of Biochemistry, Faculty of Science, Burapa University, Chonburi, Thailand.
c Parasitology Section, National Institute of Animal Health, Department of Livestock Development, Kaset Klang, Bangkhen, Bangkok 10900, Thailand.

* Corresponding author: Tel: 66-2-260 2950 ext 4605 Fax: 66-2-260 0125 E-mail: kosum@psm.swu.ac.th

Received 3 Mar 1999