A cross-sectional assessment of PRRSV nucleic acid detection by RT-qPCR in serum, ear-vein blood swabs, nasal swabs, and oral swabs from weaning-age pigs under field conditions

The porcine reproductive and respiratory syndrome virus (PRRSV) continues to challenge swine production in the US and most parts of the world. Effective PRRSV surveillance in swine herds can be challenging, especially because the virus can persist and sustain a very low prevalence. Although weaning-age pigs are a strategic subpopulation in the surveillance of PRRSV in breeding herds, very few sample types have been validated and characterized for surveillance of this subpopulation. The objectives of this study, therefore, were to compare PRRSV RNA detection rates in serum, oral swabs (OS), nasal swabs (NS), ear-vein blood swabs (ES), and family oral fluids (FOF) obtained from weaning-age pigs and to assess the effect of litter-level pooling on the reverse transcription-quantitative polymerase chain reaction (RT-qPCR) detection of PRRSV RNA.Three eligible PRRSV-positive herds in the Midwestern USA were selected for this study. 666 pigs across 55 litters were sampled for serum, NS, ES, OS, and FOF. RT-qPCR tests were done on these samples individually and on the litter-level pools of the swabs. Litter-level pools of each swab sample type were made by combining equal volumes of each swab taken from the pigs within a litter.Ninety-six piglets distributed across 22 litters were positive by PRRSV RT-qPCR on serum, 80 piglets distributed across 15 litters were positive on ES, 80 piglets distributed across 17 litters were positive on OS, and 72 piglets distributed across 14 litters were positive on NS. Cohen's kappa analyses showed near-perfect agreement between all paired ES, OS, NS, and serum comparisons (). The serum RT-qPCR cycle threshold values (Ct) strongly predicted PRRSV detection in swab samples. There was a ≥ 95% probability of PRRSV detection in ES-, OS-, and NS pools when the proportion of positive swab samples was ≥ 23%, ≥ 27%, and ≥ 26%, respectively.ES, NS, and OS can be used as surveillance samples for detecting PRRSV RNA by RT-qPCR in weaning-age pigs. The minimum number of piglets to be sampled by serum, ES, OS, and NS to be 95% confident of detecting ≥ 1 infected piglet when PRRSV prevalence is ≥ 10% is 30, 36, 36, and 40, respectively.

[1]  D. Linhares,et al.  Evaluating oral swab samples for PRRSV surveillance in weaning-age pigs under field conditions , 2023, Frontiers in Veterinary Science.

[2]  H. Akoğlu User's guide to sample size estimation in diagnostic accuracy studies , 2022, Turkish journal of emergency medicine.

[3]  Arka Ghosh,et al.  Effect of pooling family oral fluids on the probability of PRRSV RNA detection by RT-rtPCR. , 2022, Preventive veterinary medicine.

[4]  D. Holtkamp,et al.  Characterization of changes in productivity parameters as breeding herds transitioned through the 2021 PRRSV breeding herd classification system , 2022, Journal of Swine Health and Production.

[5]  C. Corzo,et al.  Proposed modifications to porcine reproductive and respiratory syndrome virus herd classification , 2021, Journal of Swine Health and Production.

[6]  E. Tchilian,et al.  Simultaneous Infection With Porcine Reproductive and Respiratory Syndrome and Influenza Viruses Abrogates Clinical Protection Induced by Live Attenuated Porcine Reproductive and Respiratory Syndrome Vaccination , 2021, bioRxiv.

[7]  P. Gauger,et al.  Probability of PRRS virus detection in pooled processing fluid samples. , 2021, Veterinary microbiology.

[8]  J. Zimmerman,et al.  Finding PRRSV in sow herds: Family oral fluids vs. serum samples from due-to-wean pigs. , 2021, Preventive veterinary medicine.

[9]  J. Zimmerman,et al.  Longitudinal piglet sampling in commercial sow farms highlights the challenge of PRRSV detection , 2021, Porcine Health Management.

[10]  Alba Frias-De-Diego,et al.  The Local and Systemic Humoral Immune Response Against Homologous and Heterologous Strains of the Type 2 Porcine Reproductive and Respiratory Syndrome Virus , 2021, Frontiers in Immunology.

[11]  M. Stevenson Sample Size Estimation in Veterinary Epidemiologic Research , 2021, Frontiers in Veterinary Science.

[12]  A. Allepuz,et al.  Biosecurity in pig farms: a review , 2021, Porcine Health Management.

[13]  J. Niemi,et al.  Financial Analysis of Herd Status and Vaccination Practices for Porcine Reproductive and Respiratory Syndrome Virus, Swine Influenza Virus, and Mycoplasma hyopneumoniae in Farrow-to-Finish Pig Farms Using a Bio-Economic Simulation Model , 2020, Frontiers in Veterinary Science.

[14]  J. Zimmerman,et al.  Guidelines for oral fluid-based surveillance of viral pathogens in swine , 2020, Porcine Health Management.

[15]  G. Balka,et al.  First report of porcine parainfluenza virus 1 (species Porcine respirovirus 1) in Europe. , 2020, Transboundary and emerging diseases.

[16]  M. Pomorska-Mól,et al.  Noninvasive strategies for surveillance of swine viral diseases: a review , 2020, Journal of veterinary diagnostic investigation : official publication of the American Association of Veterinary Laboratory Diagnosticians, Inc.

[17]  P. Gauger,et al.  Practical aspects of PRRSV RNA detection in processing fluids collected in commercial swine farms. , 2020, Preventive veterinary medicine.

[18]  M. Pieters,et al.  Detection of Mycoplasma hyopneumoniae in nasal and laryngeal swab specimens in endemically infected pig herds , 2019, Veterinary Record.

[19]  R. Main,et al.  Macroepidemiological aspects of porcine reproductive and respiratory syndrome virus detection by major United States veterinary diagnostic laboratories over time, age group, and specimen , 2019, PloS one.

[20]  C. Corzo,et al.  Effect of litter aggregation and pooling on detection of porcine reproductive and respiratory virus in piglet processing fluids , 2019, Journal of veterinary diagnostic investigation : official publication of the American Association of Veterinary Laboratory Diagnosticians, Inc.

[21]  Andres M. Perez,et al.  Detection of Influenza a Virus in Swine Nasal Swab Samples With a Wash-Free Magnetic Bioassay and a Handheld Giant Magnetoresistance Sensing System , 2019, Front. Microbiol..

[22]  T. Opriessnig,et al.  Porcine reproductive and respiratory syndrome virus RNA detection in different matrices under typical storage conditions in the UK , 2019, Veterinary Record.

[23]  V. Normand,et al.  Monitoring PRRSV-1 in suckling piglets in an endemic herd using reverse transcriptase quantitative real time polymerase chain reaction: comparison of the rate of detection in serum and oral fluid samples and evaluation of pooling , 2019, Porcine health management.

[24]  P. Gauger,et al.  Detection, isolation, and in vitro characterization of porcine parainfluenza virus type 1 isolated from respiratory diagnostic specimens in swine. , 2019, Veterinary microbiology.

[25]  A. Bowman,et al.  Evaluation of nonwoven fabrics for nasal wipe sampling for influenza A virus in swine , 2018, Journal of veterinary diagnostic investigation : official publication of the American Association of Veterinary Laboratory Diagnosticians, Inc.

[26]  R. Morrison,et al.  Surveillance of porcine reproductive and respiratory syndrome virus in the United States using risk mapping and species distribution modeling. , 2017, Preventive veterinary medicine.

[27]  T. Hirai,et al.  Nasal swab - A new tool for the detection of porcine respiratory disease complex in natural infected pigs , 2017 .

[28]  Woojoo Lee,et al.  Does McNemar’s test compare the sensitivities and specificities of two diagnostic tests? , 2017, Statistical methods in medical research.

[29]  D. Frei,et al.  A novel quantitative real-time polymerase chain reaction method for detecting toxigenic Pasteurella multocida in nasal swabs from swine , 2016, Acta Veterinaria Scandinavica.

[30]  E. Wodak,et al.  Ability of ELISAs to detect antibodies against porcine respiratory and reproductive syndrome virus in serum of pigs after inactivated vaccination and subsequent challenge , 2016, BMC Veterinary Research.

[31]  Mohamad Adam Bujang,et al.  Requirements for Minimum Sample Size for Sensitivity and Specificity Analysis. , 2016, Journal of clinical and diagnostic research : JCDR.

[32]  Zvonimir Poljak,et al.  Lessons learned and knowledge gaps about the epidemiology and control of porcine reproductive and respiratory syndrome virus in North America. , 2015, Journal of the American Veterinary Medical Association.

[33]  L. Larsen,et al.  Different clinical, virological, serological and tissue tropism outcomes of two new and one old Belgian type 1 subtype 1 porcine reproductive and respiratory virus (PRRSV) isolates , 2015, Veterinary Research.

[34]  R. Morrison,et al.  Comparison of time to PRRSv-stability and production losses between two exposure programs to control PRRSv in sow herds. , 2014, Preventive veterinary medicine.

[35]  C. Szablewski,et al.  Utility of snout wipe samples for influenza A virus surveillance in exhibition swine populations , 2014, Influenza and other respiratory viruses.

[36]  T. Opriessnig,et al.  Comparison of Commercial Real-Time Reverse Transcription-PCR Assays for Reliable, Early, and Rapid Detection of Heterologous Strains of Porcine Reproductive and Respiratory Syndrome Virus in Experimentally Infected or Noninfected Boars by Use of Different Sample Types , 2012, Journal of Clinical Microbiology.

[37]  Catherine Belloc,et al.  Infectiousness of pigs infected by the Porcine Reproductive and Respiratory Syndrome virus (PRRSV) is time-dependent , 2012, Veterinary Research.

[38]  M. McHugh Interrater reliability: the kappa statistic , 2012, Biochemia medica.

[39]  T. Opriessnig,et al.  Shedding and infection dynamics of porcine circovirus type 2 (PCV2) after natural exposure. , 2011, Veterinary microbiology.

[40]  Cheng-Yao Yang,et al.  Shedding pattern and serological profile of porcine circovirus type 2 infection in cesarean-derived, colostrum-deprived and farm-raised pigs. , 2011, The Journal of veterinary medical science.

[41]  S. Dee,et al.  Infection dynamics and clinical manifestations following experimental inoculation of gilts at 90 days of gestation with a low dose of porcine reproductive and respiratory syndrome virus. , 2009, Canadian journal of veterinary research = Revue canadienne de recherche veterinaire.

[42]  Geoffrey T Fosgate,et al.  Practical Sample Size Calculations for Surveillance and Diagnostic Investigations , 2009, Journal of veterinary diagnostic investigation : official publication of the American Association of Veterinary Laboratory Diagnosticians, Inc.

[43]  J. Cano,et al.  Feasibility of pooled-sample testing for the detection of porcine reproductive and respiratory syndrome virus antibodies on serum samples by ELISA. , 2008, Veterinary microbiology.

[44]  J. Zimmerman,et al.  Detection of Porcine reproductive and respiratory syndrome virus infection in porcine oral fluid samples: a longitudinal study under experimental conditions , 2008, Journal of veterinary diagnostic investigation : official publication of the American Association of Veterinary Laboratory Diagnosticians, Inc.

[45]  A. Trajman,et al.  McNemar χ2 test revisited: comparing sensitivity and specificity of diagnostic examinations , 2008, Scandinavian journal of clinical and laboratory investigation.

[46]  M. Engle,et al.  Evaluation of the Sensitivity of Reverse-Transcription Polymerase Chain Reaction to Detect Porcine Reproductive and Respiratory Syndrome Virus on Individual and Pooled Samples from Boars , 2007, Journal of veterinary diagnostic investigation : official publication of the American Association of Veterinary Laboratory Diagnosticians, Inc.

[47]  B. Rossow,et al.  Sampling of adult boars during early infection with porcine reproductive and respiratory syndrome virus for testing by polymerase chain reaction using a new blood collection technique (blood-swab method) , 2006 .

[48]  À. Olvera,et al.  Quantification of porcine circovirus type 2 (PCV2) DNA in serum and tonsillar, nasal, tracheo-bronchial, urinary and faecal swabs of pigs with and without postweaning multisystemic wasting syndrome (PMWS). , 2005, Veterinary microbiology.

[49]  T. Asai,et al.  Detection of Mycoplasma hyopneumoniae in lung and nasal swab samples from pigs by nested PCR and culture methods. , 2005, The Journal of veterinary medical science.

[50]  Carlos García,et al.  Temporal localization of porcine reproductive and respiratory syndrome virus in reproductive tissues of experimentally infected boars. , 2003, Theriogenology.

[51]  Takashi Sasaki,et al.  PCR detection of Porcine circovirus type 2 DNA in whole blood, serum, oropharyngeal swab, nasal swab, and feces from experimentally infected pigs and field cases. , 2003, The Journal of veterinary medical science.

[52]  J. Zimmerman,et al.  Characterization of the carrier state in porcine reproductive and respiratory syndrome virus infection. , 2002, Veterinary microbiology.

[53]  T. de Baère,et al.  Detection of Actinobacillus pleuropneumoniae in cultures from nasal and tonsillar swabs of pigs by a PCR assay based on the nucleotide sequence of a dsbE-like gene. , 2001, Veterinary microbiology.

[54]  C. Chae,et al.  Enhanced detection of toxigenic Pasteurella multocida directly from nasal swabs using a nested polymerase chain reaction. , 2001, Veterinary journal.

[55]  J. Christopher-Hennings,et al.  Detection and Duration of Porcine Reproductive and Respiratory Syndrome Virus in Semen, Serum, Peripheral Blood Mononuclear Cells, and Tissues from Yorkshire, Hampshire, and Landrace Boars , 2001, Journal of veterinary diagnostic investigation : official publication of the American Association of Veterinary Laboratory Diagnosticians, Inc.

[56]  H. Schirrmeier,et al.  Porcine Reproductive and Respiratory Syndrome Virus (PRRSV): Kinetics of Infection in Lymphatic Organs and Lung , 2000, Journal of veterinary medicine. B, Infectious diseases and veterinary public health.

[57]  H. Nauwynck,et al.  Virus quantification and identification of cellular targets in the lungs and lymphoid tissues of pigs at different time intervals after inoculation with porcine reproductive and respiratory syndrome virus (PRRSV). , 1997, Veterinary microbiology.

[58]  J. Zimmerman,et al.  Porcine reproductive and respiratory syndrome virus: a persistent infection. , 1997, Veterinary microbiology.

[59]  J. Zimmerman,et al.  Persistence of Porcine Reproductive and Respiratory Syndrome Virus in Serum and Semen of Adult Boars , 1995, Journal of veterinary diagnostic investigation : official publication of the American Association of Veterinary Laboratory Diagnosticians, Inc.

[60]  J. Zimmerman,et al.  Characterization of the Humoral Immune Response to Porcine Reproductive and Respiratory Syndrome (PRRS) Virus Infection , 1995, Journal of veterinary diagnostic investigation : official publication of the American Association of Veterinary Laboratory Diagnosticians, Inc.

[61]  V. Petersen The development of feeding and investigatory behaviour in free-ranging domestic pigs during their first 18 weeks of life , 1994 .

[62]  W. V. Van Alstine,et al.  Characterization of infection with endemic porcine reproductive and respiratory syndrome virus in a swine herd. , 1994, Journal of the American Veterinary Medical Association.

[63]  R. Connor Sample size for testing differences in proportions for the paired-sample design. , 1987, Biometrics.

[64]  J. R. Landis,et al.  The measurement of observer agreement for categorical data. , 1977, Biometrics.

[65]  Anubrata Das,et al.  Porcine reproductive and respiratory syndrome virus (PRRSV). , 2013 .

[66]  J. Kliebenstein,et al.  Economic Impact of Porcine Reproductive and Respiratory Syndrome Virus on U.S. Pork Producers , 2012 .

[67]  Dale Polson,et al.  Terminology for classifying swine herds by porcine reproductive and respiratory syndrome virus status , 2011, Journal of Swine Health and Production.

[68]  Jacob Benesty,et al.  Pearson Correlation Coefficient , 2009 .

[69]  J. Prickett,et al.  Detection of viral pathogens of swine using oral fluid specimens , 2009 .

[70]  C. Prieto,et al.  Porcine reproductive and respiratory syndrome virus infection in the boar: a review. , 2005, Theriogenology.