Original Research

Polymerase chain reaction-based discrimination of viable from non-viable Mycoplasma gallisepticum

Ching Giap Tan, Aini Ideris, Abdul R. Omar, Chen Pei Yii, Stanley H. Kleven
Onderstepoort Journal of Veterinary Research | Vol 81, No 1 | a708 | DOI: https://doi.org/10.4102/ojvr.v81i1.708 | © 2014 Ching Giap Tan, Aini Ideris, Abdul R. Omar, Chen Pei Yii, Stanley H. Kleven | This work is licensed under CC Attribution-NoDerivatives 4.0
Submitted: 08 November 2013 | Published: 02 September 2014

About the author(s)

Ching Giap Tan, Department of Veterinary Clinical studies, Universiti Putra Malaysia, Malaysia
Aini Ideris, Department of Veterinary Clinical studies, Universiti Putra Malaysia, Malaysia; Institute of Bioscience, Universiti Putra Malaysia, Malaysia
Abdul R. Omar, Institute of Bioscience, Universiti Putra Malaysia, Malaysia; Department of Veterinary Pathology and Microbiology, Universiti Putra Malaysia, Malaysia
Chen Pei Yii, Department of Veterinary Clinical studies, Universiti Putra Malaysia, Malaysia
Stanley H. Kleven, Poultry Diagnostic and Research Center, University of Georgia, United States


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Abstract

The present study was based on the reverse transcription polymerase chain reaction (RT-PCR) of the 16S ribosomal nucleic acid (rRNA) of Mycoplasma for detection of viable Mycoplasma gallisepticum. To determine the stability of M. gallisepticum 16S rRNA in vitro, three inactivation methods were used and the suspensions were stored at different temperatures. The 16S rRNA of M. gallisepticum was detected up to approximately 20–25 h at 37 °C, 22–25 h at 16 °C, and 23–27 h at 4 °C. The test, therefore, could detect viable or recently dead M. gallisepticum (< 20 h). The RT-PCR method was applied during an in vivo study of drug efficacy under experimental conditions, where commercial broiler-breeder eggs were inoculated with M. gallisepticum into the yolk. Hatched chicks that had been inoculated in ovo were treated with Macrolide 1. The method was then applied in a flock of day 0 chicks with naturally acquired vertical transmission of M. gallisepticum, treated with Macrolide 2. Swabs of the respiratory tract were obtained for PCR and RT-PCR evaluations to determine the viability of M. gallisepticum. This study proved that the combination of both PCR and RT-PCR enables detection and differentiation of viable from non-viable M. gallisepticum.

Keywords

Viability, Stability, PCR, RT-PCR, 16S rRNA, Mycoplasma gallisepticum

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