Submitted to: Journal of Virological Methods
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 7/21/2009
Publication Date: 7/29/2009
Citation: Valles, S.M., Varone, L., Ramirez, L., Briano, J. 2009. Multiplex detection of Solenopsis invicta viruses -1, -2, and -3. Journal of Virological Methods. 162:276-279. Interpretive Summary: The red imported fire ant was introduced into the United States in the 1930s and currently infests about 300 million acres. It causes significant economic losses (~6 billion dollars annually) in livestock and agricultural production and poses a serious threat to human health. Recently, USDA-ARS scientists at the Center for Medical, Agricultural and Veterinary Entomology (Gainesville, FL) have discovered three new RNA viruses that may prove to be useful biocontrol agents against the invasive red imported fire ant. A number of questions concerning the new viruses must be investigated in order to proceed with their development as control agents. For example, how do viruses spread from colony to colony in the field and how contagious are the viruses? To address these and related epidemiological questions regarding fire ant virues, USDA-ARS scientists at the Center for Medical, Agricultural and Veterinary Entomology (Gainesville, FL) and the South American Biological Control Laboratory (Hurlingham, Argentina) have developed a multiplex RT-PCR method capable of detecting all three known viruses in the fire ant host simultaneously. This method is expected to facilitate studies concerned with transmission and spread of the Solenopsis invicta viruses.
Technical Abstract: A multiplex polymerase chain reaction (PCR) method was developed to detect simultaneously Solenopsis invicta viruses -1, -2, and -3 in their host, the red imported fire ant, Solenopsis invicta. cDNA synthesis was conducted in a single reaction containing three oligonucleotide primers specific for each virus. Multiplex PCR was subsequently conducted with three oligonucleotide primer sets specific for each virus. The method is specific and sensitive, capable of detecting 500 copies of the virus genomes consistently. Specificity was verified by PCR and amplicon sequencing. The method was evaluated against field-collected samples of ant workers from colonies (n = 307) in Argentina and the United States. The prevalence of each virus in fire ant colonies varied considerably from site to site. A number of colonies exhibited multiple virus infections. However, the multiple SINV infection rate was considerably lower than for single infections. Comparison of viral infection prevalence between S. invicta colonies in Argentina and the U.S. showed no statistical differences, regardless of category. This method is anticipated to facilitate epidemiological and related studies concerning the S. invicta viruses in fire ants.