Molecular characterization, relatedness of Haematobia irritans (horn fly) populations, by RAPD-PCR

Abstract

Haematobia irritans is a hematophagous parasite of cattle that causes significant economic losses in many parts of the world, including Brazil. In the present work, one American and four Brazilian populations of this species were studied by Random Amplified Polymorpht DNA (RAPD) to assess basically genetic variability within and between populations. Ten different decamer random primers were employed in the genomic DNA amplification, yielding 117 fragments in the five H. irritans populations. In Drosophila prosaltans, used as an outgroup, 81 fragments were produced. Forty-three of these fragments were shared by both species. Among the H. irritans samples, that from Rio Branco (Acre State, Brazil) produced the smallest numbers of fragments and polymorphic bands. This high genetic homogenity may be ascribed to its geographic origin (in the Northwest of Brazil), which causes high isolation and low gene flow, unlike the other Brazilian populations, from the South Central region, in which cattle trade is very intensive. Marker fragments (exclusive bands) detected in every sample enabled the population origin to be characterized, but they are also potentially useful for further approaches such as the putative origin of Brazilian populations from North America. Similarity indices [Nei& Li, 1979, Proc. Natl. Acad. Sci. USA 76: 5269-5273] and phylogenetic trees, rooted by using the outgroup and produced by the Phylogenetic Analysis using Parsimony (PAUP 4.0-Swofford, 2001) program showed the closest relationships between flies from Sao Jose do Rio Preto and Turúba (both from São Paulo State, Brazil) while flies from the geographically distant Rio Branco showed the greatest differentiation relative to the others.

Genetic polymorphism, molecular characterization and relatedness of Macrobrachium species (Palaemonidae) based on RAPD-PCR

“…As already mentioned, only the bands with a frequency of at least 10% were considered, in an attempt to avoid including bands with non-specific amplification (Castro and Madi-Ravazzi, 2000;Castiglioni and de Campos Bicudo, 2005). In this manner, 182 fragments were obtained from the species analyzed, varying from 2.40 to 0.30 kb, with Mb being the most polymorphic (74 fragments) followed by Mj (68 fragments) and Ma (65 fragments).…”

“…As important characteristics, such markers must be exclusive to the species in question and preferentially monomorphic, that is, found with 100% frequency among the individuals analyzed (Castiglioni and de Campos Bicudo, 2005;Rampelotti et al, 2008;Rahman et al, 2009).…”

“…Genomic DNA was prepared as described by Dellaporta et al (1983), with modifications adapted by Castiglioni and de Campos Bicudo (2005). Approximately 1g muscle tissue was homogenized in microtubes with 160 µL cold solution I (10 mM Tris, 60 mM NaCl, 5% sucrose, 10 mM EDTA, pH 7.8).…”

“…Besides, only the bands with frequencies of at least 10% were considered, in order to avoid bands possibly resulting from non-specific amplification (Castro and Madi-Ravazzi, 2000;Castiglioni and de Campos Bicudo, 2005).…”

“…Among the many molecular methods currently available for genetic studies, it appears particularly suitable for analysis of any species, revealing a high degree of polymorphism in many cases (Williams et al, 1990;Garcia et al, 1998;Castiglioni and de Campos Bicudo, 2005;Roratto et al, 2008;Cesniene et al, 2010;Nath et al, 2010). The method uses PCR with a single short oligonucleotide primer that randomly amplifies short fragments of genomic DNA, separated by size, with agarose gel electrophoresis.…”