Tuesday, November 17, 2015

Two hungry caterpillars

Helicoverpa armigera
The Old World bollworm, Helicoverpa armigera, and the corn earworm, Helicoverpa zea  are considered two of the most important agricultural pests in the world. Both of them are highly polyphagous and the greatest damage is inflicted on cotton, tomatoes, maize, chick peas, alfalfa and tobacco. However, in total Helicoverpa armigera feeds on hosts in 68 plant families and Helicoverpa zea on hosts in 36 plant families. Now these are some really hungry caterpillars.

Identifying these two species is difficult. Adults can only be separated by complex dissection, and larvae cannot be identified to species using morphology. In the past geographic origin was used for identification in most instances but recently several studies showed that both species occur in the same regions. A PCR-RFLP assay using COI, and cytochrome b (Cyt b) was developed to distinguish between the two species and a few other congeners but it turns out that some species such as Chloridea virescens generate RFLP patterns identical to those for Helicoverpa armigera.

A new study published in PLoS ONE now introduces a real-time PCR assay to diagnose and separate the two species. The assay is based on ITS-2 and 18S rRNA but all control specimens were initially identified using some morphology and DNA Barcoding. 

The assay can be completed in 50 minutes when using isolated DNA and is successfully tested on larvae intercepted at ports of entry and adults captured during domestic surveys. We demonstrate that the assay can be run in triplex with no negative effects on sensitivity, can be run using alternative real-time PCR reagents and instruments, and does not cross react with other New World Heliothinae.

This is certainly a significant time saving over e.g. DNA Barcoding, where sequencing of the final PCR product is required. As this is also the most expensive step in the process there is also some significant cost saving involved. Furthermore, the assay is capable of providing a correct diagnosis even in the presence of other species. However, the new method has a few limitations:

Although the assay was tested with a variety of other Helothinae, including other Helicoverpa, caution should be used when applying these protocols outside of the New World. 
Mixed template testing demonstrates that the real-time assay developed here is not applicable for diagnosing bulk samples.

Nothing is perfect but these are rather cautionary remarks than real limitations. After all the test was developed for a very specific task and that's what it does with remarkable precision.

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