Variabilidade genetica e estrutura de populações de Cochliomyia hominivorax (Diptera: Calliphoridae) : uma nova perspectiva atraves de marcadores microssatelites / Genetic diversity and population structure in the new world screw-worm, Cochliomyia hominivorax (Diptera: Calliphoridae), as revealed by microsatellite markers

Tatiana Teixeira Torres

The cultural and economic importance of animal production in South America dates back to the colonial era. This continent has one of the largest percentages of the world s total cattle population and countries like Brazil, Argentina and Uruguay play a major role as exporters of a variety of livestock products. Diseases affecting livestock can have a significant impact on animal productivity, on trade of live animais, meat and other animal products, which, consequently, affects the overall process of the economic development of South American countries. The New World screw-worm (NWS), Cochliomyia hominivorax (Coquerel), is an important parasitic insect pest in Neotropical regions. NWS myiasis is caused by the larval stage of the fly infesting tissues of warm-blooded vertebrates. This species has been successfully eradicated from North and most of Central America by the sterile insect technique. In South America, however, this pest continues to affect the development of the livestock sector and an international effort is underway to evaluate the feasibility of eradicating the NWS from endemic areas of Central and South America. Therefore, the aim of this study was to provide some insight into the patterns of genetic variation, structure and gene flow of C. hominivorax populations from South America. Those are valuable information required prior to an investment on large-scale efforts aiming at controlling insect pests. Microsatellites stand out as co-dominant markers with a high number of alleles per locus, high polymorphism and high-expected heterozygosities. Due to those features, these markers have been increasingly used to investigate questions regarding population structure, gene flow and mC!ting system even in populations that have low levels of allozyme and mitochondrial variation. The major drawback of microsatellites is that they must often be isolated de novo for each species. In order to investigate patterns of genetic differentiation of C. hominivorax from its current distribution using microsatellite loci, two efforts were made to isolate these markers from enriched genomic libraries. Initially, a set of 10 polymorphic microsatellite loci was isolated from an AC-enriched genomic library. Amplification of the reported loci in 30 screw-worms revealed an average of 6.9 alleles per locus with expected heterozygosities ranging from 0.3831 to 0.8022. In the second effort, seven new pOlymorphic microsatellite markers were developed using a different protocol. An (AG)n probe was used instead of (GT)n in the enrichment step. The number of alleles found in the seven new loci ranged from 3 to 13 per locus, with the expected heterozygosities ranging from 0.4220 to 0.9045. Cross-species amplifications of the 17 microsatellite markers were successful in other Calliphoridae species, suggesting that these loci may be useful in those species after evaluating the extent of polymorphism. These efforts have also motivated the development of a web-based, user-friendly toolkit for microsatellite sequence characterization, the FiRe (Find Repeats). This initiative represents an important contribution to other efforts aimed at isolating and characterizing microsatellite markers. Twelve of the 17 isolated loci were used to characterize genetic variability and population structure across NWS South American populations. In the spatio-temporal analysis of C. hominivorax populations from Uruguay, significant departures from Hardy-Weinberg equilibrium were observed for ali populations. Linkage disequilibrium was also detected for severa I locus pairs. Low, but significant, levels of subdivision were recorded between populations, but no evidence of isolation by distance was observed. The temporal analysis indicated a population differentiation increase over time and ali pairwise comparisons between temporal subpopulations yielded significant estimates of population differentiation. The observed differences between local populations are probably a result of the occurrence of demographic changes that affected NWS populations. Analysis of populations from 21 geographical sites in South America (distances ranging from 15 to 5180 km) revealed a high genetic variability within NWS populations. Deviations from Hardy-Weinberg equilibrium and linkage disequilibrium were frequent. A strikingly low, but significant, genetic differentiation was observed, indicating a broad-scale genetic homogeneity in the continent. This low differentiation was coupled with a lack of isolation by distance. These results are consistent with the patterns expected from a species that has undergone range expansion. The pattern of genetic variation among populations could also be an evidence of colonization-extinction dynamics, where subpopulations of this species might usefully be viewed as a metapopulation. Whichever scenario best describes the population structure of NWS, it will have deep implications in contrai strategies. Further studies should complement these findings and provide support for decision makers in the planning and implementation of new area-wide contrai programs of this important livestock pest

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