PRRS diagnostics

A significant investment has been made to establish effective prevention, control, and elimination strategies for PRRS. A key part of these strategies has been the development and implementation of reliable, accurate and timely diagnostic procedures. 

Detecting antibodies - ELISA

PRRSv antibody detection is a constantly evolving diagnostic discipline.

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Detecting virus - PCR

Virus detection techniques provide a useful indicator of disease, although some are used primarily as research tools because they are labour-intensive, expensive and slow. One such example is virus isolation, which uses specific cell lines to amplify viruses from tissue ...

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Virus sequencing

Genomic variation is used to understand virus transmission and to identify similarities in virus over time and space. Both virus sequencing and restriction fragment length polymorphism (RFLP) are used.

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Oral fluid sampling

Oral fluids were first reported as a diagnostic specimen for PRRSV in 1997. This review is limited to PRRSV, but it is noteworthy that oral fluids samples have diagnostic value for many diseases. A review by Prickett et al. noted that work has been conducted with oral ...

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As these techniques evolve, it is important that veterinarians and industry stakeholders continue to adopt best practices so that a coordinated response to the disease can be maintained across the industry. In this section we review past and current methods for PRRS diagnosis.

Sampling procedures
An important first step in choosing an appropriate diagnostic approach is to decide the objectives of the tests to be performed. These objectives might include detection of infection in at least one animal in a herd, determining the prevalence of virus within the herd, confirming exposure to virus or vaccine, or assessing the timing of an infection.  These factors will determine which animals to test, the number of samples required, and the appropriate tissues to use. Furthermore, interpreting individual test results may not reflect the status of the entire herd as the timeline of infection will vary for each individual animal, so care must be taken. Once the objective is defined, each practitioner can determine whether to test individual animals, subpopulations or the entire herd.

Getting the numbers right
Many factors need to be considered when choosing an appropriate sample size for PRRS diagnostics. In order to identify at least one positive sample from a herd, considerations include herd size, the expected prevalence of the disease within the herd, the degree of certainty required, and the sensitivity and specificity of the test to be used. Fortunately, tables are available to help with the choice of sample size, which take into consideration many of these factors; and for estimating the sample size required to determine the prevalence of PRRSv within a herd. 

The importance of timing
Timing of tests is critical, as several aspects of infection and immune response, including viraemia, viral shedding, antibody response, and the appearance of lesions, can occur within specific time windows (figure 1).

Figure 1. Appropriate diagnostic tests based on the course of PRRSv infection



Source: Adapted from Cano, 2013; additional information from Langenhorst et al. 2012, and Zimmerman et al. 2012




There are broadly two categories of diagnostic test: viral detection and viral exposure. Viral detection requires collection of virus-laden tissue, bronchoalveolar lavage (BAL) fluid, thoracic fluid, semen, blood, oral fluid, or environmental samples; while detection of viral exposure requires blood, oral fluid collection or muscle transudate in order to test for anti-PRRSv antibodies. Therefore choice of tissue sample will be determined by a variety of factors including the specific test objectives, the choice of diagnostic test, and facilities available.
 
It is also important to bear in mind that it may not always be possible to collect a tissue type, and that some are more complex or difficult to collect, handle and store. For instance oral fluid collection is a very simple and easy technique compared with BAL, but is not suitable for individual diagnosis since samples are taken from a penned population (e.g. from purpose-made ropes that have been chewed by the pigs).

Diagnostic tests
Since its discovery, PRRSv and/or its antibodies have been detected using a variety of techniques, some of which are now seldom used in the field for a variety of reasons: they might be a more research-oriented technique, overly time-consuming, expensive, or require highly trained staff. In the following sections we will briefly describe currently-used techniques, mentioning some of the older ones to illustrate how far diagnostics have evolved over recent years. A summary of key features of the newer and more widely used techniques can be found in Table 1.

Table 1. Summary of characteristics of main diagnostic test
Diagnostic test Range Comment
RT-PCR Useful with peak serum viral levels at 4–7 days DPI; undetectable by 29–35 DPI (persists longer in lymph nodes and tonsils) Highly sensitive, highly specific
IHC Useful up to 28 DPI; not recommended above 90 DPI High specificity; moderate sensitivity. Effective in identifying vertically transmitted virus in piglets; may not be effective for some genetically diverse isolates.
FA staining Can be used in the early stages of infection (1-28 DPI), and directly or immediately after incubation in cell culture of lavage fluid in mid stages (30-70 DPI). Not recommended in the later stages of infection May not detect genetically diverse isolates
ELISA Early detection possible via IgM (by 7 DPI). Detection possible until very late stages of infection (well beyond 100 DPI) Highly sensitive, highly specific; low sensitivity with oral fluids; levels of antibody detected do not necessarily reflect virulence
FMIA In serum, antibodies may be detected as early as 7 DPI and may persist beyond 202 DPI Highly sensitive, highly specific
Indirect IFA Detection of early infections (5 – 9 DPI, IGM and IgG respectively) for 21–28 (IgM) and 90–145 (IgG) DPI High specificity; variable sensitivity
Source: data from Ellingson, 2013


 
The future for PRRSv diagnosis
A combined approach of routine surveillance and disease diagnosis is required if efforts to control and eliminate PRRSv infection from swine herds are to be successful. While a range of techniques has become available to diagnose PRRSv infection, no test is 100% reliable under all conditions and for all animals. For example, routine molecular tests provide a sensitive and specific method for assessing viral infection, but some genetically diverse viruses may not be detected. Similarly, serological tests to detect anti-PRRSv antibodies offer information on previous viral exposure but are less useful for persistent infections. Furthermore, detectable viral and antibody levels change during the course of infection, making the timing of specific testing a crucial factor in successful diagnosis. For all tests, false positive and false negative data can impact significantly on appropriate herd management decisions, so secondary testing of samples using alternative strategies should be conducted where appropriate. Indeed, interpretation of results can often be difficult. As new techniques are added to the armoury, it is increasingly important to make sure the correct tests are carried out at the correct time in the correct animals, and with the correct interpretation to optimise effective management and prevention or control of an outbreak.