Recent decades have seen both rapid growth and extensive consolidation in swine production. As a collateral effect, these changes have exacerbated the circulation of viruses and challenged our ability to prevent, control, and/or eliminate impactful swine diseases. Recent pandemic events in human and animal health, e.g., SARS-CoV-2 and African swine fever virus, highlight the fact that clinical observations are too slow and inaccurate to form the basis for effective health management decisions: systematic processes that provide timely, reliable data are required. Oral fluid-based surveillance reflects the adaptation of conventional testing methods to an alternative diagnostic specimen. The routine use of oral fluids in commercial farms for PRRSV and PCV2 surveillance was first proposed in 2008 as an efficient and practical improvement on individual pig sampling. Subsequent research expanded on this initial report to include the detection of ≥23 swine viral pathogens and the implementation of oral fluid-based surveillance in large swine populations (> 12,000 pigs). Herein we compile the current information regarding oral fluid collection methods, testing, and surveillance applications in swine production.

Chagas disease is an important health problem in Latin America due to its incapacitating effects and associated mortality. Studies on seropositivity for Trypanosoma cruzi in Mexican dogs have demonstrated a direct correlation between seropositivity in humans and dogs, which can act as sentinels for the disease in this region. The objective of this study was to determine the seropositivity for T.cruzi infection in dogs from Sonora, a northern borderstate of Mexico. Responsible pet owners were selected at random from an urban area of Empalme municipality, Sonora, Mexico, and from there, 180 dog samples were collected. Anti-T. cruzi antibodies were determined using the enzyme-linked immunosorbent assay (ELISA) method. Reactive ELISA sera were processed by indirect immunofluorescence to confirm the presence of anti-T. cruzi antibodies. For the statistical analysis, chi-square tests were conducted. Dogs’ sera showed a seropositivity rate of 4.44%. The rate of seropositivity was not associated with the dogs’ age, sex, or socioeconomics pertaining to the geographical area. One sample (1/180, 0.55%) showed the acute state of the disease. The study found a presence of anti-T. cruzi antibodies in dogs in this area, which suggests vector transmission. There is a need for active surveillance programs throughout the state of Sonora and vector control strategies should also be implemented in endemic regions.

The 1890s marked a significant milestone with the introduction of antibody-based agglutination and precipitation assays, revolutionizing the detection of bacterial pathogens in both animals and humans. This era also witnessed pivotal contributions to our understanding of humoral immunity, as researchers elucidated the structure and functions of antibody molecules, laying the groundwork for diagnostic applications. Among antibody isotypes, IgG is of paramount importance in diagnostic investigations given its definitive indication of infection or vaccination, coupled with its widespread presence and detectability across various specimen types, such as serum, colostrum, milk, oral fluids, urine, feces, and tissue exudate. Despite their resilience, immunoglobulins are susceptible to structural alterations induced by physicochemical and enzymatic processes, which can compromise the reliability of their detection. Here we review comprehensively the historical milestones, underlying mechanisms, and influencing factors (e.g., temperature, pH, storage) that shape the structural integrity and stability of IgG antibodies in aqueous solutions and various clinical specimens.

Herein we review broad issues that affect test performance for agents that produce persistent infections. Using PRRSV as an example, the relationship between “disease transition stages” and “diagnostic transition stages” is discussed using meta-analyses of diagnostic data (n = 4307 results) from the refereed literature to highlight the key issues. Although diagnostic technology will continue to improve, it may be concluded from the analysis that there can be no single best diagnostic approach; rather, the choice of specimen and test must be tailored to the specific testing objective. In most cases, meeting the testing objective(s) will require the use of more than one assay and/or specimen type.

Oral fluid (n = 564) samples collected longitudinally from twelve 14-week-old pigs vaccinated with a porcine reproductive and respiratory syndrome virus (PRRSV) modified live vaccine were used to evaluate and compare the diagnostic performance of three commercial PRRSV oral fluid (OF) ELISAs (ELISAs 1, 2, 3). Serum samples (n = 132) tested by a PRRSV serum ELISA (ELISA 'S') provided an antibody response baseline for comparison. The initial analysis comparing the rate of positivity between each OF ELISA versus ELISA 'S' and then pairwise among the three OF ELISAs determined that ELISA 2 (143 false negative results) was significantly different from ELISAs 1 and 3, and from ELISA 'S' (Cochran's Q test, p < 0.05). Receiver operating characteristic (ROC) analyses based on the manufacturers' recommended cutoff were used to estimate the diagnostic sensitivities and specificities of ELISA 1 (100%, 100%), ELISA 2 (62%, 97%), and ELISA 3 (94%, 100%). As an additional aid for interpreting results, the diagnostic sensitivities and specificities of each OF ELISA were also estimated over a range of cutoffs. Area under the curve comparisons found no significant difference between ELISAs 1 and 3, but ELISA 2 differed from both ELISA 1 and 3 (ROC Chi-square, p < 0.05). Based on these analyses, the overall diagnostic performance of the three OF ELISAs ranked ELISA 1 ≥ ELISA 3 > ELISA 2.

The ontogeny of PRRSV antibody in oral fluids has been described using isotype-specific ELISAs. Mirroring the serum response, IgM appears in oral fluid by 7days post inoculation (DPI), IgA after 7 DPI, and IgG by 9 to 10 DPI. Commercial PRRSV ELISAs target the detection of IgG because the higher concentration of IgG relative to other isotypes provides the best diagnostic discrimination. Oral fluids are increasingly used for PRRSV surveillance in commercial herds, but in younger pigs, a positive ELISA result may be due either to maternal antibody or to antibody produced by the pigs in response to infection. To address this issue, a combined IgM-IgA PRRSV oral fluid ELISA was developed and evaluated for its capacity to detect pig-derived PRRSV antibody in the presence of maternal antibody. Two longitudinal studies were conducted. In Study 1 (modified-live PRRS vaccinated pigs), testing of individual pig oral fluid samples by isotype-specific ELISAs demonstrated that the combined IgM-IgA PRRSV ELISA provided better discrimination than individual IgM or IgA ELISAs. In Study 2 (field data), testing of pen-based oral fluid samples confirmed the findings in Study 1 and established that the IgM-IgA ELISA was able to detect antibody produced by pigs in response to wild-type PRRSV infection, despite the presence of maternal IgG. Overall, the combined PRRSV IgM-IgA oral fluid ELISA described in this study is a potential tool for PRRSV surveillance, particularly in populations of growing pigs originating from PRRSV-positive or vaccinated breeding herds.