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Title: The utility of the internal transcribed spacer region 2 (ITS2) in confirming species boundaries in the genus Gonatocerus:comparison to the cytochrome oxidase subunit I(COI) gene and taxonomic data: molecular key based on ITS2

Author
item De Leon, Jesus
item LOGARZO, GUILLERMO - USDA,ARS,SABCL-ARGENTINA
item TRIAPITSYN, SERGUEI - UC-RIVERSIDE, CA

Submitted to: CDFA Pierce's Disease Control Program Research Symposium
Publication Type: Proceedings
Publication Acceptance Date: 9/25/2006
Publication Date: 11/27/2006
Citation: De Leon, J.H., Logarzo, G., Triapitsyn, S.V. 2006. The utility of the internal transcribed spacer region 2 (ITS2) in confirming species boundaries in the genus Gonatocerus:comparison to the cytochrome oxidase subunit I(COI) gene and taxonomic data: molecular key based on ITS2. CDFA Pierce's Disease Control Program Research Symposium, November 27-29, 2006, San Diego, California. p.56.

Interpretive Summary: In the current study, we sequenced a nuclear gene [ribosomal internal transcribed spacer region 2 (ITS2)] to test whether this gene could be used to confirm species boundaries of glassy-winged sharpshooter [Homalodisca vitripennis (=H. coagulata)] egg parasitoids belonging to the genus Gonatocerus, in other words, to see if the known different species could be distinguished. In addition, we wanted to test this gene to see if it could be utilized to test the genetic (phylogenetic) relationships among the known Gonatocerus species. We showed that this gene (ITS2) worked as well as another standard mitochondrial gene [cytochrome oxidase subunit I (COI)], confirming its utility in confirming species boundaries. Furthermore, comparison of our results with taxonomic data demonstrated that the ITS2 gene worked slightly better or more informative at defining phylogenetic relationships among Gonatocerus species than the COI gene. Since we included several Gonatocerus species in the current study, we developed a partial molecular key based on ITS2 sizes to distinguish the various species. The current results are important to the biological control program in California.

Technical Abstract: We sequenced the nuclear ribosomal internal transcribed spacer region 2(ITS2) from several glassy-winged sharpshooter (GWSS) [Homalodisca vitripennis Germar (=H.coagulata Say)] egg parasitoid species (Hymenoptera: Mymaridae) belonging to the genus Gonatocerus Nees to test the utility of this fragment to confirm species boundaries and to define phylogenetic relationships. A total of 35 specimens belonging to 10 named species, one unnamed species, and two specimens from another mymarid genus (Anagrus erythroneurae) (outgroup) were analyzed. A phylogenetic tree generated using the neighbor-joining algorithmic method showed that each named Gonatocerus species formed its own unique taxonomic unit or clade with very strong bootstrap support (100%), confirming species boundaries. The ITS2 fragment confirmed species boundaries as well as cytochrome oxidase subunit I (COI) sequence data. Furthermore, the phlogenetic relationships among species generated by the ITS2 fragment were in excellent agreement with those delineated by taxonomic data. The current results clearly confirm the utility of the ITS2 fragment in confirming species boundaries of egg parasitoids belonging to the genus Gonatocerus. The results showed that the ITS2 fragment appears to be phylogenetically more informative or valuable than that inferred by COI sequence data. Since several important Gonatocerus species were analyzed, a molecular diagnostic key based on ITS2 sizes was developed. In the event two species (G. ashmeadi and G. metanotalis and G. walkerjonesi and G. annulicornis) were found with similarly sized ITS2 fragments, inter-simple sequence repeat-polymerase chain reaction (ISSR-PCR) DNA fingerprinting was performed to distinguish them. ISSR-PCR very clearly distinguished the aformentioned species, demonstrating that it is an excellent molecular diagnostic tool. The current results are important to the biological control program in California.