Rapid Detection of Lawsonia intracellularis by a Quantitative
PCR based on Immunomagnetic Beads
Shunli Yang1†, Muhammad Umar Zafar Khan2†,
Sumin Pan1, Xiaoqing Zhang1, Yupeng Fang1,
Wenyan Gai3, Jie Zhang1* and Yongsheng Liu1*
1Key
Laboratory of Preventive Veterinary Medicine of Hebei, College of
Animal Science and Technology, Hebei Normal University of Science
and Technology
(HNUST),
Qinhuangdao, Hebei, 066004, China; 2Institute of
Microbiology, Faculty of Veterinary Science, University of
Agriculture Faisalabad, Faisalabad, 38040, Pakistan; 3Department
of Pet Technology, Shandong Vocational Animal Science and Veterinary
College, Weifang, Shandong, 261061, China
†Equally
contributed to this work
*Corresponding author:
liuys4031@hevttc.edu.cn
(Yongsheng
Liu);
zhangj4032@hevttc.edu.cn
(Jie
Zhang)
Abstract
Immunomagnetic nanobeads
(IMNBs) have been widely used to isolate and detect microorganisms. In this
study, recombinantly expressed outer membrane proteins LI0902 and LI1024 of
Lawsonia intracellularis (L.
intracellularis) were used to yield polyclonal antibodies.
IMNBs functionalized with anti-LI0902 polyclonal
antibodies display the capture activity to
L. intracellularis that was indicated
by immunofluorescence assay based on anti-LI1024 polyclonal antibody. One-step
quantitative PCR (qPCR) target of the aspartate ammonia-lyase gene based on
IMNBs was developed and evaluated using several pathogens associated with pig
diarrheal diseases and clinical faecal samples. The results showed that IMNBs
had good specificity and sensitivity, no cross-reactivity with other pathogens,
and the detection limit was as low as 3.6×102 copies/mL. Meanwhile,
IMNBs showed good capture efficiency, 4.21×105 copies of
L. intracellularis could be captured
by 1 µg of IMNBs. Moreover, L.
intracellularis in faecal samples were successfully detected by the qPCR
combined with IMNBs. The results demonstrate that this method is promising in
rapidly and effectively detecting L.
intracellularis in clinical faecal samples.
To Cite This Article:
Yang S, Khan MUZ, Pan S, Zhang X, Fang Y, Gai W, Zhang J and Liu Y, 2024.
Rapid detection of Lawsonia
intracellularis by a quantitative PCR based on immunomagnetic beads.
Pak Vet J. http://dx.doi.org/10.29261/pakvetj/2024.209