en |
Chilean IPNV isolates: Robustness analysis of PCR detection
Jorquera, Esteban; Morales, Paz; Tapia, David; Torres, Pamela; Eissler, Yoanna; Espinoza, Juan C.; Conejeros, Pablo & Kuznar, Juan
Abstract
Background: The genomes of several infectious pancreatic necrosis viruses (IPNVs) isolated in Chile were
sequenced with a single amplification approach for both segments A and B. The resulting sequences were then
used to determine the conservation of the primer-binding regions used in polymerase chain reaction
(PCR)-based diagnostic methods proposed in the literature. Thus, the robustness of each technique was
studied, particularly the eventual effect of further mutations within the primer-binding sites.
Results: On analysis, most methods currently used to detect Chilean IPNV varieties were deemed adequate.
However, the primers were designed to be genogroup specific, implying that most detection methods pose
some risk of detecting all strains prevalent in the country, due to the coexistence of genogroups 1 and 5.
Conclusions: Negative resultsmust be interpreted carefully given the high genomic variability of IPNVs. Detection
techniques (quantitative reverse transcription (qRT)-PCR) based on degenerate primers can be used to minimize
the possibilities of false-negative detections.
Keywords
IPNV detection; Mismatch\'s Tm analysis; QPCR
|