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ARS Home » Research » Publications at this Location » Publication #47865
Title: IDENTIFICATION OF SCREWWORM SPECIES (DIPTERA: CALLIPHORIDAE) BY POLYMERASE CHAIN REACTION-RESTRICTION FRAGMENT LENGTH POLYMOORPHISM

Author
item TAYLOR D B - 5440-30-00
item SZALANSKI A L - UNIV NEBR
item PETERSON R D II - 5440-30-00

Submitted to: Medical and Veterinary Entomology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/18/1995
Publication Date: N/A
Citation: N/A

Interpretive Summary: New World Screwworm (NWS) is a serious pest of livestock in the American tropics. Eradication programs have eliminated this pest from the United States and Mexico. However, reintroductions of NWS to eradicated regions is a constant problem. This problem is exasperated by the presence of Secondary Screwworm (SS), a non-pest species which is morphologically similar to NWS. Differentiating these two species can be difficult when samples are of poor quality or do not contain mature larvae. It is important to begin control measures of an outbreak as quickly as possible to prevent its spreading, however, such control programs are expensive to mobilize. This research provides a technique for the unambiguous identification of screwworm samples of all ages and common methods of preservation based upon DNA fingerprints. Pieces of DNA are amplified from mscrewworm sample using Polymerase Chain Reaction. These pieces of DNA are then treated with enzymes which cut the DNA at precise locations based upon the DNA sequence. The cut DNA is analyzed by electrophoresis which results in species diagnostic patterns. The technique was used successfully to identify several samples collected by screwworm eradication programs in Central America. The technique provides inexpensive, $2.50 per sample, and rapid, 1 day, identifications of screwworm samples. This provides program managers with the information they need prior to committing program resources to the control of screwworm outbreaks.

Technical Abstract: Restriction fragment length polymorphisms in polymerase chain reaction amplified fragments (PCR-RFLP) of mitochondrial DNA were used to differentiate species of New World Screwworms (Diptera: Calliphoridae). Twenty-seven restriction enzymes were screened on five fragments of mtDNA. Twelve restriction fragment length patterns differentiated New World Screwworm, Cochliomyia hominivorax (Coquerel), from Secondary Screwworm, Cochliomyia macellaria (F.). Four restriction fragment length patterns were variable in C. hominivorax while all fragment patterns were fixed in C. macellaria. Diagnostic restriction fragment length patterns were used for species diagnosis while intraspecific variable patterns were used to characterize field samples and laboratory strains. The PCR-RFLP technique is flexible with regard to developmental stage of the sample and method of preservation. We were able to characterize specimens of all life stages from egg to adult as well as larvae preserved in alcohol and pinned adults. PCR-RFLP is rapid and inexpensive, allowing specimens to be characterized within 24 h for less than $2.50. Because the PCR-RFLP technique uses "universal" primers, it can be applied to other insect groups with minimal developmental investment.