Hostname: page-component-cd9895bd7-7cvxr Total loading time: 0 Render date: 2024-12-21T17:16:32.040Z Has data issue: false hasContentIssue false
  • English
  • Français

PCR–ELISA method for Mycoplasma contamination detection in veterinary vaccines

Published online by Cambridge University Press:  13 February 2008

Zhang Ying ,
Jiang Jun-Bing ,
Liu Jin-Ping  and
Jia Li-Yan
Zhang Ying*
Affiliation:
College of Veterinary Medicine and Animal Science, Shanxi Agricultural University, Taigu 030801, China
Jiang Jun-Bing
Affiliation:
College of Veterinary Medicine and Animal Science, Shanxi Agricultural University, Taigu 030801, China
Liu Jin-Ping
Affiliation:
College of Veterinary Medicine and Animal Science, Shanxi Agricultural University, Taigu 030801, China
Jia Li-Yan
Affiliation:
College of Veterinary Medicine and Animal Science, Shanxi Agricultural University, Taigu 030801, China
*
*Corresponding author. E-mail: dkyzy@126.com
Get access Rights & Permissions [Opens in a new window]

Abstract

A polymerase chain reaction–enzyme-linked immunosorbent assay (PCR-ELISA) method was set up to detect Mycoplasma in live vaccines and to establish the optimal experimental conditions. According to the 16S rRNA gene sequences published in GenBank, which include Mycoplasma gallisepticum of chicken and M. hyopneumoniae, M. hyosynoviae and M. flocculare of swine (submitted nos: Mg AY744942, Mhp AE017244, Mhs AY973563 and Mf X63377), PCR primers, labelled with digoxin (Dig), and probe, labelled with biotin, were designed using the software DNAstar and Primer Premier 5.0. The system and conditions of PCR–ELISA were optimized using purified genomic DNA extracted from Mycoplasma as a template. Samples of 30 batches of Newcastle disease live vaccines (I strain), 30 batches of Newcastle disease live vaccines (La Sota strain) and 30 batches of swine fever live vaccine were tested by PCR-ELISA. Results showed that the detection rate of Mycoplasma contamination from different vaccines was higher (36.5%) with this technique than with PCR (24.4%). The PCR–ELISA appeared to be a simple, fast and reliable method for qualitative and quantitative analysis of Mycoplasma. The practical use of PCR–ELISA as a kit to detect Mycoplasma contamination in live vaccine seems very promising.

Keywords

live vaccineMycoplasma detectionPCR–ELISA
Type
Research Article
Information
Chinese Journal of Agricultural Biotechnology , Volume 4 , Issue 3 , December 2007 , pp. 187 - 192
Copyright
Copyright © China Agricultural University and Cambridge University Press 2007

Access options

Get access to the full version of this content by using one of the access options below. (Log in options will check for institutional or personal access. Content may require purchase if you do not have access.)

Footnotes

First published in Journal of Agricultural Biotechnology 2007, 15(1): 102–106

References

Chen, R (2001) Rapid detection of transgene fish by Dig-PCR-ELISA. Journal of Fishery Sciences of China 8(4): 13–15 (in Chinese with English abstract).Google Scholar
Chen, TS (1995) The Preparative and Application of Microorganism Media. Beijing: China Agricultural Publishing Company, pp. 470–491.Google Scholar
Lai, XM, Fang, GY and Li, CX (1999) Detecting Mycoplasma contamination in cell cultivation by comparing PCR with isolate cultivation. Academic Journal of Sun Yat-sen University of Medical Sciences 20(2): 151–154 (in Chinese with English abstract).Google Scholar
Levishon, S, Hyman, H, Perelman, D, et al. (1989) The use of a specific DNA probe for detection of Mycoplasma gallisepticum in field outbreaks. Avian Pathology 18: 535–541.CrossRefGoogle Scholar
Li, YM (2004) Studying the nested-PCR detection technique of Mycoplasma contamination in live vaccine. Masters degree dissertation of Shanxi Agricultural University.Google Scholar
Ning, YB and Ji, XL (1988) Study on Mycoplasma contamination in live vaccine from chick embryo culture. Journal of Veterinary Medicine 3: 15–20 (in Chinese).Google Scholar
Ning, YB and Ji, XL (1989) Study on the Mycoplasma contamination in vaccine of Marek's disease virus from chicken embryo fibroblast cell. Chinese Journal of Veterinary Drug 4: 22–23 (in Chinese).Google Scholar
Ning, YB and Ji, XL (1992) Mycoplasma contamination in live vaccines by culturing cell. Chinese Journal of Veterinary Medicine 18(4): 44–45 (in Chinese).Google Scholar
Ning, YB and Ji, XL (1993) Report on Mycoplasma contamination in live vaccine inland. Chinese Journal of Veterinary Drug 27(1): 34–36 (in Chinese).Google Scholar
Shang, YF and Ji, XL (1991) The application of dot immunobinding assay in the Mycoplasma study. Chinese Journal of Veterinary Medicine 17(8): 3–6 (in Chinese).Google Scholar
Yao, Huochun (2002) Veterinary Microbiology Experiment Manual. Beijing: China Agriculture Press, Vol. 2, pp. 58–66.Google Scholar
Zhang, Y, Jia, GL and Shao, GQ (2003) Mycoplasma hyopneumoniae adhesin factor gene's clone and expression at R1R2 region. Chinese Journal of Veterinary Science 23(4): 338–341 (in Chinese).Google Scholar
Zhu, ZM and Liu, H (2002) Simple Immunology Technology. Beijing: Science Press, Vol. 11, pp. 176–184.Google Scholar