Multiplex real‐time PCR assay for the detection of three invasive leafminer species: Liriomyza huidobrensis, L. sativae and L. trifolii (Diptera: Agromyzidae)

Rapid and precise identification of immature stages of leafminers of the genus Liriomyza Mik associated with imported fresh produce is essential to ensure appropriate biosecurity decisions at the border, in quarantine and post border. The leafminers Liriomyza huidobrensis, Liriomyza sativae and Liri...

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Bibliographic Details
Published in:Austral entomology 2017-05, Vol.56 (2), p.153-159
Main Authors: Sooda, Anuradha, Gunawardana, Disna, Li, Dongmei, Kumarasinghe, Lalith
Format: Article
Language:English
Subjects:
Amplification
Assaying
Biosecurity
border detection
Coefficients
Correlation coefficient
Correlation coefficients
Deoxyribonucleic acid
Detection
DNA
DNA probes
Identification
interceptions
Invasive species
Liriomyza huidobrensis
Liriomyza sativae
Liriomyza trifolii
mitochondrial cytochrome oxidase I
Multiplexing
Nucleotide sequence
PCR
Pests
Polymerase chain reaction
Primers
quarantine
Real time
Reliability
Seawater
Species identification
Specificity
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Summary:Rapid and precise identification of immature stages of leafminers of the genus Liriomyza Mik associated with imported fresh produce is essential to ensure appropriate biosecurity decisions at the border, in quarantine and post border. The leafminers Liriomyza huidobrensis, Liriomyza sativae and Liriomyza trifolii are not present in New Zealand and classified as regulated pests when detected at New Zealand's border. To assist rapid species identification of the immature stages of interceptions, a multiplex real‐time TaqMan PCR assay was developed to identify these three species simultaneously in a single test. Species‐specific primers and probes were designed by amplifying the mitochondrial COI gene of each targeting species, respectively. The multiplex real‐time PCR assay demonstrated high specificity for all three target species and the assay detected DNA quantities as low as 0.1 pg for all species. Linear responses and high correlation coefficients between the amount of DNA and Cq values for each species were also achieved. Therefore, the assay demonstrated its sensitivity and reliability for the identification of these three invasive Liriomyza species.
ISSN:2052-174X
2052-1758
DOI:10.1111/aen.12237