Detection of viral DNA of myxoma virus using a validated PCR method with an internal amplification control

Kwit, Ewa; Osiński, Zbigniew; Rzeżutka, Artur

doi:10.1016/j.jviromet.2019.113709

Cited by 6 publications

(6 citation statements)

References 34 publications

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“…Analysing ĸ values for the AGID and IAC-PCR, the highest result concordance between methods was obtained for samples containing at least 3,125 PCRU of the virus (ĸ = 1.00) (10). Only IAC-PCR showed 100% sensitivity for samples with low (1 PCRU) virus concentration.…”

Section: Resultsmentioning

confidence: 95%

“…The AGID DSe was 0.65 (0.53-0.76) and DSp 1.00 (0.40-1.00); AC of 0.67 (0.55-0.76) was also attained. In the case of the IAC-PCR method, these parameters were 0.98 (0.91-1.00) as the DSe and 0.95 (0.84-0.99) as the DSp (10). Direct comparison of method parameters showed that AGID gave only half the DSe of the molecular methods for samples containing low virus concentration.…”

Section: Resultsmentioning

confidence: 98%

“…The lowest dilution of viral DNA giving a positive signal on gel was designated as 1 PCRU content. Viral DNA was extracted from 200 µL of tissue homogenates using a DNeasy Blood & Tissue Kit (Qiagen, Hilden, Germany) according to the manufacturer's instructions and subsequently detected using a previously described IAC-PCR method (10). For each set of the tested samples a panel of controls was included to monitor the performance of the MYXV DNA isolation and amplification steps.…”

Section: Methodsmentioning

confidence: 99%

“…AGID DSe and DSp were calculated using the extent of the agreement between the AGID-yielded proportions of positive (PA) and negative (NA) results and the respective total numbers of positive and negative samples with subsequent determination of confidence intervals (CI) by the Clopper-Pearson method. The measures of result agreement for the set of samples when analysed using both the AGID and IAC-PCR methods were expressed as Kappa (ĸ) values, which for AGID were obtained for dichotomous data (presence or absence of the virus) in all tested samples (6, 13) and compared with those obtained for the IAC-PCR method (10). If ĸ is equal to 1 then very good (complete) agreement is observed.…”

Section: Methodsmentioning

confidence: 99%

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Detection of myxoma virus in the classical form of myxomatosis using an AGID assay: statistical assessment of the assay’s diagnostic performance

Kwit

Osiński

Llombart‐Bosch

et al. 2020

Journal of Veterinary Research

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IntroductionThe aim of the study was to estimate the diagnostic sensitivity (DSe) and specificity (DSp) of an agar gel immunodiffusion (AGID) assay for detection of myxoma virus (MYXV) in the classical form of myxomatosis and to compare its diagnostic performance to that of molecular methods (IAC-PCR, OIE PCR, and OIE real-time PCR).Material and MethodsA panel of MYXV-positive samples of tissue homogenates with low (1 PCR unit – PCRU) and high (3,125 PCRU) virus levels and outbreak samples were used for method comparison studies. The validation parameters of the AGID assay were assessed using statistical methods.ResultsThe AGID attained DSe of 0.65 (CI95%: 0.53–0.76), DSp of 1.00 (CI95%: 0.40–1.00), and accuracy of 0.67 (CI95%: 0.55–0.76). The assay confirmed its diagnostic usefulness primarily for testing samples containing ≥3,125 PCRU of MYXV DNA. However, in the assaying of samples containing <3,125 PCRU of the virus there was a higher probability of getting false negative results, and only molecular methods showed a 100% sensitivity for samples with low (1 PCRU) virus concentration. The overall concordance of the results between AGID and IAC-PCR was fair (ĸ = 0.40). Full concordance of the results was observed for OIE PCR and OIE real-time PCR when control reference material was analysed.ConclusionsFindings from this study suggest that AGID can be used with some limitations as a screening tool for detection of MYXV infections.

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Section: Resultsmentioning

confidence: 95%

Section: Resultsmentioning

confidence: 98%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

See 2 more Smart Citations

Detection of myxoma virus in the classical form of myxomatosis using an AGID assay: statistical assessment of the assay’s diagnostic performance

Kwit

Osiński

Llombart‐Bosch

et al. 2020

Journal of Veterinary Research

Self Cite

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show abstract

“…In contrast, molecular methods stand out from other bioanalytical/immunological tests because they help to identify a particular virus quickly. Likewise, they have a high sensitivity and are not so expensive ( Kwit et al, 2019 ). The methods based on Polymerase Chain Reaction (PCR) —as the PCR with reverse transcription (RT-PCR)— are used routinely to diagnose and identify viruses.…”

Section: Introductionmentioning

confidence: 99%