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Title: Preliminary evidence from reproductive compatibility studies suggests that Gonatocerus tuberculifemur exists as a cryptic species complex or a new species is identified:Development and utility of molecular diagnostic markers.

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.A., Triapitsyn, S.V. 2006. Preliminary evidence from reproductive compatibility studies suggests that Gonatocerus tuberculifemur exists as a cryptic species complex or a new species is identified:Development and utility of molecular diagnostic markers. CDFA Pierce's Disease Control Program Research Symposium, November 27-29, 2006, San Diego, California. p 44.

Interpretive Summary: Recently, we genetically characterized geographic populations of Gonatocerus tuberculifemur from South America and demonstrated that some of these populations were genetically distinct by two molecular methods. The populations clustered into two well-supported clades with the population from San Rafael, Mendoza Province, Argentina clustering into its own unique clade. Gonatocerus tuberculifemur is a prospective natural enemy candidate agent for a neoclassical biological control program in California against the invasive glassy-winged sharpshooter (Homalodisca vitripennis) (=H. coagulata). Identifying the correct natural enemy is critical to a biological control program. After genetically characterizing G. tubeculifemur, we developed molecular diagnostic markers by two methods to distinguish isofemale lines created from freshly field-collected populations [Tunuyán (clade 1) and San Rafael (clade 2)] for reproductive compatibility studies. Analysis of the isofemale lines belonging to the two different clades with our newly developed molecular diagnostic markers correctly genotyped the isofemale lines, confirming the utility of our diagnostic markers. Based on our molecular work, we predicted that individuals belonging to the two separate clades would not hybridize or reproduce. Preliminary reproductive compatibility studies between individuals from the two separate clades indeed confirmed our prediction, that is, the results demonstrated that the two isofemales lines were not reproductively compatible. Taken together, our molecular data and the preliminary reproductive compatibility studies suggest the G. tuberculifemur either exists as a cryptic species complex or a new species is identified. Since G. tuberculifemur is under consideration as a biological control agent against the invasive GWSS in California, understanding cryptic variation of this species is critical.

Technical Abstract: Recent work uncovered divergent clades or distinct lineages in populations of Gonatocerus tuberculifemur from South America. Gonatocerus tuberculifemur is a prospective egg parasitoid candidate agent for a neoclassical biological control program in California against the invasive glassy-winged sharpshooter (GWSS), Homalodisca vitripennis (Germar) [=H. coagulata (SAY]. In the present study, we developed molecular diagnostic markers by two approaches to distinguish field-collected populations of G. tuberculifemur for reproductive compatibility studies. The two diagnostic assays were: polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) of the mitochrondrial cytochrome oxidase subunit I gene (COI) and inter-simple sequence repeat-polymerase chain reaction (ISSR-PCR) DNA fingerprinting. Clade-specific restriction enzymes generated bands of the correct size with high specificity. Analysis of two isofemales lines created from freshly field-collected populations belonging to clade 1 (Tunuyan) and clade 2 (San Rafael) showed that both of our developed molecular diagnostic markers correctly genotyped these isofemale lines, confirming the utility of our diagnostic markers. Based on our molecular work, we predicted that G. tuberculifemur individuals belonging to the two distinct clades would not hybridize. Preliminary mating compatibility studies between these two isofemale lines demonstrated that our prediction was indeed correct. Interspecific crosses produced only male offspring, whereas, the intraspecific control crosses produced both males and females or fertile offspring. Taken together, both our molecular work and the preliminary reproductive compatibility studies strongly suggest that G.tuberulifemur either exists as a cryptic species complex or a new species is identified. Since G. tuberculifemur is under consideration as a biological control agent against the invasive GWSS in California, understanding cryptic variation of this species is critical.