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Get the most out of M. hyopneumoniae diagnostics use, interpretation

The most important aspects to consider in a diagnostic investigation are to have a comprehensive understanding of the potential assays’ applications and be aware of testing limitations.

By Maria Pieters, DVM, PhD., University of Minnesota College of Veterinary Medicine Department of Veterinary Population Medicine and Veterinary Diagnostic Laboratory
Mycoplasma hyopneumoniae is a bacterial agent that causes enzootic pneumonia, a chronic respiratory condition affecting pigs around the world. The detrimental effect of M. hyopneumoniae infections is expressed in decreased pig performance, increased days to reach market weight, and more importantly, greater susceptibility to other respiratory pathogens of viral and bacterial origin.

Detection of either the pathogen itself, or the host response after infection, are crucial for confirmation of herd status, and for the design and application of strategies that can lead to disease control in the field. However, features inherent to the bacterium and its pathogenesis have largely limited M. hyopneumoniae diagnostics. Such features include an extremely slow replication rate and a delayed humoral response in infected pigs, among others. Thus, diagnostic tools for detection of M. hyopneumoniae infection are recognized to be far from perfect, and at most times frustrating.

From the starting point that clinical signs are not evident during the entire infection period and that they are not unique to disease caused by M. hyopneumoniae, diagnosis in the field and the laboratory are challenging to producers, practitioners and scientists.

Historically, culture and isolation of M. hyopneumoniae had been considered the “goal standard” for bacterial detection, but this continues to be significantly difficult to achieve and is therefore reserved to specific cases in which an isolated is needed, which doesn’t seem to be too frequent.

Other methods allowing direct detection of the bacterium in tissues and respiratory secretions, based on genetic material identification, have gained popularity due to the advantages of their use. But more importantly, our understanding of the potential applications and capabilities of the assays are the main driver of better diagnostic interpretation, leading to appropriate prevention and treatment options.

Here, several examples of increased diagnostic accuracy or direct field applications are described, which allow for improved identification of infected or non-infected pigs and better understanding of the disease processes in the field.

• Increasing the diagnostic sensitivity of a testing protocol by selecting assays that are appropriate for the suspected phase of infection, or put in different words, having the goal of detecting the pathogen instead of an active immune response during the very early days after infection. It has been shown that samples collected deep in the respiratory tract of live animals, such as laryngeal swabs and deep tracheal sampling, can yield detection of M. hyopneumoniae by polymerase chain reaction less than one week after infection, and at least four weeks sooner than detection of M. hyopneumoniae-specific antibodies by ELISA. On the other hand, in many cases, the use of newer diagnostic assays can be accompanied by increased diagnostic costs. Nevertheless, the use of techniques like sample pooling can be used to the advantage of the diagnostic investigation, without a substantial sacrifice in sensitivity, and maintaining costs down.

• Characterization of specific regions of the genome of M. hyopneumoniae has been available for a number of years, but its use and application may have been limited in the past. Recently, the use of sequencing of specific adhesion proteins (P146 sequencing) and molecular characterization of tandem repeats in M. hyopneumoniae (i.e. Multiple Locus Variable number tandem repeat Analysis) have served to identify and track down bacterial variants in the field. Thus, allowing for direct comparison of field circulating Mycoplasmas in the case of outbreak investigations.

• Due to the intrinsic low sensitivity of testing for M. hyopneumoniae and the slow nature of the bacterium, confirming low prevalence or negativity of this pathogen in a herd (post disease elimination or to confirm negative status) has always been a problematic scenario. However, the use of specific pooling testing combined with appropriate sample size calculations for extremely low prevalence, at a high confidence level, has brought light into ways to confirm negative status, while keeping diagnostic efforts and cost realistic.

The examples mentioned above are used in order to obtain the most diagnostic information, while using less than perfect tests. Doubtlessly, a clear need exists for the overall improvement of methods for detection of M. hyopneumoniae infection and host response to it. However, current assays can be better utilized for increased accuracy and specific application. Thus, the most important aspects to consider in a diagnostic investigation are to have a comprehensive understanding of the potential assays’ applications and be aware of testing limitations.

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