How can we do better?
By Albert Canturri and Maria Pieters
Accurate diagnostics are the cornerstone in which disease prevention and control interventions lay on. Every diagnostic investigation should start with two simple questions: "what information do I need to obtain?" and "how will I interpret and use the obtained information?"
For the first question, producers and practitioners in the field most likely seek to determine the presence or absence of a pathogen of interest in a sample, whether it is a clinical specimen from individual pigs, aggregated samples, such as oral fluids, or even environmental samples.
In the case of Mycoplasma hyopneumoniae, the intrinsic difficulty of this pathogen to grow in bacterial culture dampens its use for routine diagnostics. Thus, the most practical tool that diagnosticians have in their hands to investigate the presence of M. hyopneumoniae in a sample is polymerase chain reaction (PCR), which targets and amplifies the DNA present in a sample and gives an approximation of bacterial quantity.
For the second question mentioned above, it is generally understood that techniques based on PCR are widely used in swine disease diagnostics, mainly due to their high sensitivity and specificity, and their cost-benefit advantage over other tests.
However, current PCR testing for M. hyopneumoniae is far from perfect and possesses important features that need to be taken into consideration, as interpretation of the obtained results can be challenging at times.
One example of the latter has been the recent detection of unexpected positive results in non-traditional sample types, such as the processing fluids (PF)1.
Detection of M. hyopneumoniae in PF was an unforeseen result since the pathogen is thought to be restricted to the respiratory system. Researchers and practitioners were therefore puzzled and were set up to figure out this perplexing finding by expanding the sampling in farms with different health status and management conditions for an extended period.
Results from such studies study showed that M. hyopneumoniae was consistently detected in PF in positive farms and that, at least in part, the origin of the detected genetic material could be traced down to vaccine products, which clearly illustrates that environmental contamination of samples is not to be ignored.
Indeed, a study2 presented at the most recent Allen D. Leman Conference (2021) reinforced the importance of this issue, showing that post-vaccination environmental contamination of samples was possible even with tracheal secretion swabs, the preferred sample type for ante mortem detection of M. hyopneumoniae.
These two above-mentioned diagnostic challenges recently experienced emphasize that, for an appropriate interpretation of M. hyopneumoniae PCR results, producers, practitioners and caretakers collecting samples in the farm should center their efforts at minimizing the chances of pre-analytical error, that is, all the procedures that occur before the sample arrives at the laboratory for testing.
A meticulous sampling methodology should be exercised, paying special attention to details. For example, it is good practice to differentiate between a clean and a dirty zone in the cart or the surface in which tubes and catheters or swabs are placed, both before and after sampling. Sampling materials, such as mouth speculums or laryngoscopes should be cleaned and disinfected between sampled pigs, and gloves should be changed often.
These measures should be maximized in high-risk scenarios, such as in the monitoring of gilts for introduction into a naïve herd, especially when gilts have been vaccinated prior to sampling at the gilt development unit (GDU). It is important to clarify that vaccinated pigs will not result positive to M. hyopneumoniae by PCR due to inoculation with bacterins, however, vaccines can be unintentionally sprayed into the surroundings and become a source for environmental contamination.
Additionally, farm management practices should be considered when implementing measures to minimize risks of environmental contamination. For example, there is a hypothetical risk that farm personnel who vaccinate pigs at weaning carry vaccine-derived genetic material on their hands and coveralls, and cross-contamination can eventually occur when they handle piglets at processing or when aid at restraining gilts and sows for tracheal testing in gestation or farrowing.
In summary, nucleic acid detection of M. hyopneumoniae by PCR testing is a great diagnostic technique because of its extremely high sensitivity of detection. Nevertheless, because of the same reason, care should be taken when collecting, handling and transporting samples to the laboratories, always aiming at minimizing both cross- and environmental contamination.
References 1. Vilalta C, Sanhueza JM, Murray D, Johnson L, Pieters M. Detection of Mycoplasma hyopneumoniae in piglet processing fluids. Vet Rec. 2019 Oct 26;185(16):510. doi: 10.1136/vr.105475. Epub 2019 Aug 13.
2. Hensch M, Kellen M, Maschhoff A, Gauger P. Diagnostic problems with unexpected positive PCRs. 2021 Allen D. Leman Swine Conference Proceedings, Saint Paul, Minnesota, USA.
Canturri is a graduate student and Pieters is an associate professor, both in the Department of Veterinary Population Medicine at the University of Minnesota.
