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Appl Environ Microbiol, February 1998, p. 543-548, Vol. 64, No. 2
0099-2240/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.

Detection of Mycoplasma hyopneumoniae by Air Sampling with a Nested PCR Assay

Katharina D. C. Stärk,1,* Jacques Nicolet,2 and Joachim Frey2

Department of Veterinary Clinical Sciences, Massey University, Palmerston North, New Zealand,1 and Institute of Veterinary Bacteriology, University of Berne, CH-3001 Berne, Switzerland2

Received 25 August 1997/Accepted 13 November 1997

This article describes the first successful detection of airborne Mycoplasma hyopneumoniae under experimental and field conditions with a new nested PCR assay. Air was sampled with polyethersulfone membranes (pore size, 0.2 µm) mounted in filter holders. Filters were processed by dissolution and direct extraction of DNA for PCR analysis. For the PCR, two nested pairs of oligonucleotide primers were designed by using an M. hyopneumoniae-specific DNA sequence of a repeated gene segment. A nested PCR assay was developed and used to analyze samples collected in eight pig houses where respiratory problems had been common. Air was also sampled from a mycoplasma-free herd. The nested PCR was highly specific and 104 times as sensitive as a one-step PCR. Under field conditions, the sampling system was able to detect airborne M. hyopneumoniae on 80% of farms where acute respiratory disease was present. No airborne M. hyopneumoniae was detected on infected farms without acute cases. The chance of successful detection was increased if air was sampled at several locations within a room and at a lower air humidity.

* Corresponding author. Mailing address: Department of Veterinary Clinical Sciences, Massey University, Palmerston North, New Zealand. Phone: 64 6 350 61 44. Fax: 64 6 350 56 16. E-mail: K.Staerk{at}massey.ac.nz.




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