previous page   Pak Vet J, 2020, 40(1): 81-87   next page
Molecular Detection of Brucellosis, Leptospirosis and Campylobacteriosis by Multiplex PCR and Screening by ELISA Assays in Buffalo Breeding Bulls

Saher Islam1,4, Wasim Shehzad1*, Amna Arshad Bajwa1, Muhammad Imran1, Muhammad Yasir Zahoor1, Muhammad Abdullah2, Muhammad Imran Rashid3, Kamran Ashraf3, Yung-Fu Chang4, Asif Nadeem1Muhammad Younas5, Sayed Aamir Mehmood Bukhari5, Muhammad Muzzammil Hassan6Zafar Iqbal Qureshi6 and Raheela Akhtar7

1Institute of Biochemistry and Biotechnology, University of Veterinary and Animal Sciences, Lahore, 54000, Pakistan;
2Department of Livestock Production, University of Veterinary and Animal Sciences, Pattoki, 553050, Pakistan;
3Department of Parasitology, University of Veterinary and Animal Sciences, Lahore, 54000, Pakistan; 4Department of Population Medicine and Diagnostic Sciences, College of Veterinary Medicine, Cornell University, Ithaca, New York, 14853, USA; 5Semen Production Unit, Qadirabad, District Sahiwal 57000, Pakistan; 6Department of Theriogenology, University of Agriculture, Faisalabad, 38000, Pakistan; 7Department of Pathology, University of Veterinary and Animal Sciences, Lahore, 54000, Pakistan; *Corresponding author:


Abortion in buffalo-breeding industry is one of the key roots of economic losses in the livestock sector. Brucella spp., Leptospira spp. and Campylobacter fetus are main pathogens because of the potential impact on veterinary and human health. This study assessed the relative performance of ELISA and multiplex PCR (mPCR) assay using different primer sets for the detection of venereal pathogens. Variety of samples (blood, urine, preputial washings and semen) were collected from a total of 160 buffalo breeding bulls of Semen Production Unit, Qadirabad and private dairy farms in Punjab Province of Pakistan. Serum and total genomic DNA was extracted for molecular detection by ELISA and PCR respectively. This study comprised 9 sets of primers to optimize multiplex with respect to melting temperature and crosslinking among primers. ELISA assay for Brucella was 2.5%, whereas for Leptospira and Campylobacter fetus it was 1.88% for each. With respect to development of triplex and duplex PCR, results of our assay were completely consistent with monoplex PCR using a combination of newly designed and reported primers. All ELISA suspected samples found negative for Brucella, Leptospira and C. fetus when tested with both monoplex and triplex PCR. This multiplex PCR assay provides a valuable complementary tool in routine, simultaneous and robust detection of these three genital diseases and revealing epidemiological facts about abortion measures in buffalo.

To Cite This Article: Islam S, Shehzad W, Bajwa AA, Imran M, Zahoor MY, Abdullah M, Rashid MI, Ashraf K, Chang YF, Nadeem A, Younas M, Bukhari SAM, Hassan MM, Qureshi ZI and Akhtar R, 2020. Molecular detection of brucellosis, leptospirosis and campylobacteriosis by multiplex PCR and screening by ELISA assays in buffalo breeding bulls. Pak Vet J, 40(1): 81-87.  


ISSN 0253-8318 (Print)
ISSN 2074-7764 (Online)