Effects of Sample Collection and Handling on Detection of Mycoplasma gallisepticum and M. synoviae by Real-time PCR09 December 2013
Dry tracheal swabs are the best option for routine sampling of Mycoplasma for real-time PCR, based on convenience and stability, according to researchers at the University of Georgia. Pre-moistened tracheal swabs do not store as well as the dry swabs but can be useful in some cases.
The current approach to Mycoplasma gallisepticum (MG) and M. synoviae (MS) control in the US includes continuous surveillance of breeding flocks to ensure early detection of infection, according to Naola Ferguson-Noel from the University of Georgia's Department of Population Health. It is of utmost importance that an early test for mycoplasmosis is as sensitive as possible, as missed infections can lead to more opportunities for horizontal and vertical spread of these economically devastating diseases.
Although advances in real-time polymerase chain reaction (PCR) technology have resulted in extremely sensitive and fast tests that are affordable and amenable to high throughput testing, the added sensitivity of real-time PCR may be severely reduced due to inappropriate handling and processing of swabs.
The main goal of her latest project, sponsored by the US Poultry & Egg Association, was to identify optimum methods for collection and handling of tracheal swabs for real-time PCR detection of MG and MS.
The specific objectives of this study were:
- to evaluate two commercially available liquid media for potential MG and MS real-time PCR inhibition
- to compare the sensitivity of pre-moistened and dry tracheal swabs for MG and MS real-time PCR, and
- to evaluate the effect of storage temperature and time on dry and moist swabs for MG and MS real-time PCR.
The Georgia researchers found that neither of the commercially available media they tested (Liquid Amies or Liquid Stuarts) inhibited the real-time PCR for MG or MS to any great degree.
The Remel BactiSwab® swab transport system and its equivalent dry swab were tested and the results indicated that pre-moistened swabs may collect more Mycoplasma cells and thereby enhance the sensitivity of real-time PCR.
The quantity of DNA detected from tracheal swabs (dry and wet) declined over time regardless of storage at 40°C or room temperature. The decline was greatest in wet samples stored at room temperature and least in dry swabs stored at 40°C.
Dry tracheal swabs are the best option for routine sampling for real-time PCR, based on convenience and stability, especially when swabs may take some time to reach the laboratory, concluded Ferguson-Noel. Although pre-moistened tracheal swabs do not store as well as dry swabs, they may be useful in difficult cases (e.g. chronic infections, antibiotic treatment) where maximum sensitivity is required. It is even more important that pre-moistened swabs are handled properly.
The tracheal swabs should be kept cool and processed as soon as possible for optimum results.
This information allows the poultry industry to maximise the benefits of these costly diagnostic assays and set scientifically based standards in different laboratories for sample submission and handling for MG and MS real-time PCR, added Ferguson-Noel.
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