Effect of saliva stabilisers on detection of porcine reproductive and respiratory syndrome virus in oral fluid by quantitative reverse transcriptase real-time PCR

Decorte, Inge; Stede, Yves Van der; Nauwynck, Hans; Regge, Nick De; Cay, Ann Brigitte

doi:10.1016/j.tvjl.2013.02.001

Cited by 20 publications

(24 citation statements)

References 16 publications

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“…The analyses of B2M values for blood specimens resulted in a narrow distribution curve as expected for a defined specimen type. The wide range of Ct values observed for swabs reflects the heterogeneity of this specimen type due to collection technique differences resulting in variable number of cells in the specimen, dilution factor based on the device used, and possibly degradation from host enzymes and bacteria in the oral cavity (Figure 1A and 1B)[3]. Specimen quality in general was a concern given the sometimes long distances between collection site and the laboratory with no reliable cold chain in place.…”

Section: Discussionmentioning

confidence: 99%

Ebola Virus Disease Diagnostics, Sierra Leone: Analysis of Real-time Reverse Transcription–Polymerase Chain Reaction Values for Clinical Blood and Oral Swab Specimens

et al. 2016

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During the Ebola virus outbreak of 2013–2016, the Viral Special Pathogens Branch field laboratory in Sierra Leone tested approximately 26,000 specimens between August 2014 and October 2015. Analysis of the B2M endogenous control Ct values showed its utility in monitoring specimen quality, comparing results with different specimen types, and interpretation of results. For live patients, blood is the most sensitive specimen type and oral swabs have little diagnostic utility. However, swabs are highly sensitive for diagnostic testing of corpses.

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Section: Discussionmentioning

confidence: 99%

Ebola Virus Disease Diagnostics, Sierra Leone: Analysis of Real-time Reverse Transcription–Polymerase Chain Reaction Values for Clinical Blood and Oral Swab Specimens

et al. 2016

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“…Briefly, total RNA from serum samples was extracted using the MagMAX Pathogen RNA/DNA kit (Thermofisher Scientific, Gent, Belgium) according to the manufacturer’s protocol. RNA from oral fluid was extracted as previously described [10]. Briefly, 150μl oral fluid was mixed with 225 μl of lysis/binding solution and 1μl carrier RNA, followed by 3 minutes of vortexing.…”

Section: Methodsmentioning

confidence: 99%

“…Oral fluid collected at pen level namely possesses several benefits compared to blood sampling: it is a non-invasive collection technique; samples can be collected at group level, and time and money can be saved due to the ease of collection. Many different aspects related to oral fluid sampling and its use as a diagnostic matrix for PRRSV detection have been studied during the past decade: RNA extraction methods and PCR tests were compared and developed for PRRSV detection [1,9–10]; tests for PRRSV specific antibody detection of different isotypes were developed and compared [11–14]; issues related to collection material, sample collection protocol and sample storage were studied and optimized [10,14–17]; and comparison was made between oral fluid and serum diagnostics for PRRSV in individual pigs [18–23]. All these studies showed that oral fluid could be a promising matrix for PRRSV surveillance.…”

Section: Introductionmentioning

confidence: 99%

Comparison of PRRSV Nucleic Acid and Antibody Detection in Pen-Based Oral Fluid and Individual Serum Samples in Three Different Age Categories of Post-Weaning Pigs from Endemically Infected Farms

Regge

Cay

2016

PLoS ONE

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BackgroundPorcine reproductive and respiratory syndrome virus (PRRSV) is the causative agent of an economically important disease in swine. Since it has been shown that PRRSV and PRRSV specific antibodies can be detected in oral fluid, many different aspects have been studied to show that oral fluid could be a worthy alternative diagnostic sample to serum for monitoring and surveillance of this disease. Thorough field evaluations are however missing to convincingly show its usefulness under representative field conditions.MethodologyPen-based oral fluid samples and serum samples from all individual pigs in the corresponding pens were collected from post-weaning pigs of three different age categories in eight endemically PRRSV infected farms and one PRRSV free farm in Belgium. All samples were tested by quantitative reverse transcription polymerase chain reaction (qRT-PCR) and ELISA to detect PRRSV RNA and PRRSV specific antibodies, respectively.ResultsWhile the relative specificity of PRRSV detection by qRT-PCR in pen-based oral fluid compared to serum collected from individual pigs was high in all age categories (>90%), the relative sensitivity decreased with the age of the pigs (89, 93 and 10% in 8-12w, 16-20w and 24-28w old pigs, respectively). The latter correlated with a lower percentage of PRRSV positive pigs in serum/pen in the different age categories (55, 29 and 6%, respectively). Irrespective of the age category, pen-based oral fluid samples were always found PCR positive when at least 30% of the individual pigs were positive in serum. PRRSV specific antibody detection in oral fluid by ELISA showed a 100% relative sensitivity to detection in serum since oral fluid samples were always positive as soon as one pig in the pen was positive in serum. On the other hand, two false positive oral fluid samples in 11 pens without serum positive pigs were found, resulting in a relative specificity of 82%. Indications are however present that the oral fluid result indicated the correct infection status but the absence of a golden standard test makes it difficult to define definitive test characteristics.ConclusionsOverall it can be concluded that oral fluid seems to be a useful matrix for diagnosis of PRRSV under field conditions and that differences in kinetics of PRRSV and PRRSV specific antibody detection in oral fluid and serum of individual pigs can also be reflected in pen-based oral fluid results.

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“…Blood and oral fluid samples were taken in the morning on days 0, 3,5,7,10,14,21,28,35, and 42 postinfection (dpi) for evaluation by qRT-PCR and ELISA. Oral fluid sampling was conducted before blood sampling to reduce the chance that animals would refuse to bite on the ropes after physical manipulation.…”

Section: Animals Inoculation and Sample Collectionmentioning

confidence: 99%

Diagnosis of the Lelystad strain of Porcine reproductive and respiratory syndrome virus infection in individually housed pigs: comparison between serum and oral fluid samples for viral nucleic acid and antibody detection

et al. 2014

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Abstract. There has been a developing interest in the use of oral fluid for the diagnosis of different pathogens such as Porcine reproductive and respiratory syndrome virus (PRRSV). PRRSV and PRRSV-specific antibodies have been shown to be present in oral fluid samples, but the correlation between diagnostic results in oral fluid and serum samples has been insufficiently addressed. Studies investigating this correlation focused on boars older than 6 months and type 2 strains, but it is known that the outcome of a PRRSV infection is age and strain dependent. To address this gap, the current study reports on the detection of PRRSV and PRRSV-specific antibodies in serum and oral fluid samples collected over a 6-week period after an experimental infection of 8-week-old individually housed pigs with Lelystad virus, the type 1 prototype strain. Quantitative reverse transcription polymerase chain reaction analysis showed that significantly more serum samples were PRRSV RNApositive than oral fluid until 5 days postinfection (dpi). Between 7 and 21 dpi, PRRSV RNA detection was similar in both samples but higher detection rates in oral fluid were found from 28 dpi. Compared with existing literature, this highlights that detection rates at particular time points postinfection might vary in function of strain virulence and animal age and provides useful information for the interpretation of pen-based oral fluid results. An excellent agreement between the oral fluid and serum enzyme-linked immunosorbent assay results was observed at every time point, further supporting the usefulness of oral fluid as a diagnostic sample for antibody detection.

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Effect of saliva stabilisers on detection of porcine reproductive and respiratory syndrome virus in oral fluid by quantitative reverse transcriptase real-time PCR

Cited by 20 publications

References 16 publications

Ebola Virus Disease Diagnostics, Sierra Leone: Analysis of Real-time Reverse Transcription–Polymerase Chain Reaction Values for Clinical Blood and Oral Swab Specimens

Ebola Virus Disease Diagnostics, Sierra Leone: Analysis of Real-time Reverse Transcription–Polymerase Chain Reaction Values for Clinical Blood and Oral Swab Specimens

Comparison of PRRSV Nucleic Acid and Antibody Detection in Pen-Based Oral Fluid and Individual Serum Samples in Three Different Age Categories of Post-Weaning Pigs from Endemically Infected Farms

Diagnosis of the Lelystad strain of Porcine reproductive and respiratory syndrome virus infection in individually housed pigs: comparison between serum and oral fluid samples for viral nucleic acid and antibody detection

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