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Title: Detection of DNA Polymorphisms in Homalodisca coagulata (Homoptera: Cicadellidae) by Polymerase Chain Reaction-Based DNA Fingerprinting Methods

Author
item De Leon, Jesus
item JONES, WALKER - 0212-05-00

Submitted to: Annals of the Entomological Society of America
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 1/3/2004
Publication Date: 5/1/2004
Citation: De Leon, J.H., Jones, W.A. 2004. Detection of DNA Polymorphisms in Homalodisca coagulata (Homoptera: Cicadellidae) by Polymerase Chain Reaction-Based DNA Fingerprinting Methods. Annuals of the Entomological Society of America. 97(3):574-585.

Interpretive Summary: The glassy-winged sharpshooter (GWSS) Homalodisca coagulata (Say) (Homoptera: Cicadellidae), is a xylem feeding leafhopper that is a serious pest because it vectors a strain of the bacterium Xylella fastidiosa, the causal agent of Pierce's Disease of grapevines. In Texas, the single greatest threat to the production of susceptible grape cultivars is Pierce's Disease. The disease has caused millions of dollars in losses to the state's wine industry since 1990 and the problem has escalated in the past five years. Also within the last 10 years, the GWSS has established itself in southern California where it also poses a potentially serious threat to the entire wine and table grape industry in that region of the country. Pierce's Disease is caused when X. fastidiosa resides, multiplies and interferes with the water-conductive system or xylem of the plant, initially causing dieback of leaves and shoots, and eventually causing the entire plant to collapse and die within a year or two. Strains of this bacterium, which are vectored by several species of sharpshooters, have also been associated with other diseases, such as, plum leaf scald, almond leaf scorch, periwinkle wilt, oleander leaf scorch, ragweed stunt, citrus variegated chlorosis and coffee leaf scorch. The aim of the present study was to develop DNA markers for the GWSS as a foundation for improved biological control. DNA markers or DNA fingerprints allow us to determine the origin of insects and therefore allows us to import the correct natural enemies of pests to naturally keep the numbers of pests in check. In the present study, we developed four DNA fingerprinting methods and identified several polymorphic DNA markers with each technique that can be used to identify GWSS from different geographic regions (so far, Texas and California). In addition, we developed DNA markers that can distinguish three Homalodisca sharpshooter species that vector the bacterium that causes Pierce's Disease. These DNA markers are an important accomplishment since they now will allow us to perform detailed genetic studies of the GWSS and other sharpshooter species.

Technical Abstract: The current work was undertaken to develop molecular genetic markers for the glassy-winged sharpshooter, Homalodisca coagulata (Say) by various PCR-based DNA fingerprinting methods for the purpose of estimating the level of genetic variation within and among populations with the aim of achieving genetic information useful for improving the biological control of this leafhopper. These fingerprinting methods included SSR-PCR (Simple Sequence Repeat-Polymerase Chain Reaction), RAMP (Randomly Amplified Microsatellite Polymorphisms), SAMPL (Selective Amplification of Microsatellite Polymorphic Loci) and RAPD (Random Amplification of Polymorphic DNA). A combined total of about 183 polymorphic bands were detected in sharpshooters from Weslaco, Texas with the four methods and three insects per primer combination (48 total), specifically 54, 58, 34 and 37 polymorphic bands were generated by SSR-PCR, RAMP, SAMPL and RAPD, respectively. We then applied and compared two of these PCR-based DNA fingerprinting methods to geographic populations of glassy-winged sharpshooters from Texas and California. In addition, we compared three Homalodisca sharpshooter species H. coagulata, H. lacerta and H. insolita. Our results demonstrated that the molecular genetic markers that we developed distinguished the three Homalodisca sharpshooter species, but most importantly the markers distinguished geographic populations (from Texas and California) of glassy-winged sharpshooters. This is the first report describing development of molecular genetic markers for the glassy-winged sharpshooter utilizing four different DNA fingerprinting methods.