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Inter-laboratory ring trial to evaluate real-time reverse transcription polymerase chain reaction methods used for detection of infectious pancreatic necrosis virus in Chile
Tapia, David; Eissler, Yoanna; Espinoza, Juan Carlos & Kuznar, Juan
Abstract
Background: Infectious Pancreatic Necrosis Virus (IPNV) is the etiological agent of a highly contagious disease that
affects salmonids. In Chile, the second worldwide salmon producer, IPNVcauses great economic loss and is one of
themost frequently detected pathogens. Due to its high level of persistence and the lack of information about the
efficiency of its diagnostic techniques, the National Reference Laboratory (NRL) for IPNV in Chile performed the
first inter-laboratory ring trial, to evaluate the sensitivity, specificity and repeatability of the qRT-PCR detection
methods used in the country.
Results: Results showed 100% in sensitivity and specificity in most of the laboratories. Only three of the twelve
participant laboratories presented problems in sensitivity and one in specificity. Problems in specificity (false
positives) were most likely caused by cross contamination of the samples, while errors in sensitivity (false
negatives) were due to detection problems of the least concentrated viral sample. Regarding repeatability, many
of the laboratories presented great dispersion of the results (Ct values) for replicate samples over the three days
of the trial. Moreover, large differences in the Ct values for each sample were detected among all the laboratories.
Conclusions: Overall, the ring trial showed high values of sensitivity and specificity, with some problems of
repeatability and inter-laboratory variability. This last issue needs to be addressed in order to allow harmonized
diagnostic of IPNV within the country. We recommend the use of the NRL methods as validated and reliable
qRT-PCR protocols for the detection of IPNV.
Keywords
Aquaculture; Diagnosis; Diagnostic techniques; Fish diseases; IPNV; Pathogens; qRT-PCR; Ring test; RNA virus; Salmonids; Validation
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