Taxonomic and evolutionary analysis of Zaprionus indianus and its colonization of Palearctic and Neotropical regions.

Zaprionus indianus is a dipteran (Drosophilidae) with a wide distribution throughout the tropics and temperate Palearctic and Nearctic regions. There have been proposals to reclassify the genus Zaprionus as a subgenus or group of the genus Drosophila because various molecular markers have indicated a close relationship between Zaprionus species and the immigrans-Hirtodrosophila radiation within Drosophila. These markers, together with alloenzymes and quantitative traits, have been used to describe the probable scenario for the expansion of Zaprionus indianus from its center of dispersal (Africa) to regions of Asia (ancient dispersal) and the Americas (recent dispersal). The introduction of Z. indianus into Brazil was first reported in 1999 and the current consensus is that the introduced flies came from high-latitude African populations through the importation of fruit. Once in Brazil, Z. indianus spread rapidly throughout the Southeast and then to the rest of the country, in association with highway-based fruit commerce. These and other aspects of the evolutionary biology of Z. indianus are addressed in this review, including a description of a probable route for this species' dispersal during its recent expansion.

Molecular weight estimation of esterase isoenzymes in closely related Drosophila Species (Diptera: Drosophilidae) in non-denaturing polyacrylamide gel electrophoresis

A method that allows the measure of molecular weight of two well-known and closely related esterases from Drosophila mojavensis and its sibling species, D. arizonae, is here described, using native polyacrylamide gel electrophoresis at several concentrations, applying Fergunson´s principles. These enzymes, namely EST-4 and EST-5, presented molecular weight values between 81 and 91 kDa. In spite of their distinct expression pattern through the insect's life cycle, they showed properties of isoenzymes codified by distinct structural genes, supporting the hypothesis of a rather recent gene duplication event that generated both in D. mojavensis and D. arizonae, as well as in other species of repleta group. The method is simple and adequate to be applied to preliminary molecular weight determination of other enzymes without any previous purification procedure.

Neste trabalho, um método que permite a estimativa do peso molecular de duas esterases conhecidas e intimamente relacionadas, encontradas em Drosophila mojavensis e sua espécie aparentada D. arizonae, é descrito. Este método é realizado utilizando a técnica de eletroforese em diferentes concentrações de gel e aplicando os princípios de Fergunson. As enzimas, denominadas EST-4 e EST-5, apresentaram pesos moleculares entre 81 e 91 kDa. Apesar de seus padrões diferenciados de expressão durante o ciclo de vida do inseto, elas demonstraram propriedades de enzimas codificadas por genes estruturais distintos, corroborando a hipótese de um evento de duplicação gênica recente que gerou ambas em D. mojavensis e D. arizonae, bem como em outras espécies do grupo repleta. O método proposto é simples e adequado para ser utilizado em estimativas preliminares de determinação de pesos moleculares de outras enzimas sem haver a necessidade de um procedimento prévio de purificação.

Esterase enzymes involved in pyrethroid and organophosphate resistance in a Brazilian population of Riphicephallus (Boophilus) microplus (Acari, Ixodidae).

Esterases are a group of enzymes that are reportedly associated with acaricide resistance in Riphicephallus (Boophilus) microplus. A comparative analysis was made of the esterase patterns in malathion and deltamethrin-sensitive, tolerant and resistant tick groups, using non-denaturing polyacrylamide gel electrophoresis. Electrophoretical profiles revealed four bands of esterase activity against alpha-naphthyl acetate; which were dubbed EST-1 to EST-4. The EST-3 and EST-4 were detected in all strains and were classified as carboxylesterases (CaEs). The EST-2, classified as an acetylcholinesterase (AChE), was detected in all groups, but its staining intensity increased from susceptible to resistant groups, indicating an altered production according to the degree of resistance. EST-1, which was also classified as an AChE, was detected exclusively in tolerant and resistant groups to both acaricides, but displayed greater activity in the malathion-resistant group. These data suggest that these AChEs may represent an important detoxification strategy developed to overcome the effects of acaricides.

Differential transcription of ribosomal cistrons denoting nucleolar dominance in hybrids of Drosophila mulleri and Drosophila navojoa (mulleri complex, Repleta group)

The fruit flies Drosophila mulleri and Drosophila navojoa are included in the mulleri complex of the mulleri subgroup and Repleta group. Although there is no demonstration that interspecific crosses between them occur in nature, they intercross in the laboratory in both cross directions. Previous data have shown the occurrence of nucleolar dominance in interspecific hybrids of some species in the mulleri complex. We investigated nucleolar dominance in D. mulleri/D. navojoa hybrids using the transcription profiles of the rDNA internal transcribed spacer (ITS-1) region. The results showed that the ribosomal cistrons present in the X chromosome and in the microchromosome of D. navojoa are exclusively or preferentially transcribed in these hybrids depending on the cross direction, denoting the complete or partial nucleolar dominance of this species over D. mulleri.

Distribution of the Bari-I transposable element in stable hybrid strains between Drosophila melanogaster and Drosophila simulans and in Brazilian populations of these species

We analyzed the distribution of the Bari-I transposable element in Drosophila melanogaster (IN(1)AB), its sibling species Drosophila simulans (C167.4) and in eight hybrid strains derived from initial crosses involving D. simulans females and D. melanogaster males of the above cited strains as well as in Brazilian populations of these species. Polymerase chain reaction (PCR) data showed the presence of the Bari-I element among species populations and hybrid strains. Hybridization with a 703 bp probe homologous to the Bari-I sequence showed that the number of Bari-I copies in D. melanogaster IN(1)AB was higher than in D. simulans C167.4 strains. Hybrid strains presented Bari-I sequences related to both parental species. In addition some strains displayed a Bari-I sequence that came from D. melanogaster, suggesting introgression of D. melanogaster genetic material in the background of D. simulans. In contrast, some hybrids showed deletions of D. simulans Bari-I sequences.

Esterase profile in a pyrethroid-resistant Brazilian strain of the cattle tick Boophilus microplus (Acari, Ixodidae)

The cattle tick Boophilus microplus causes great damage in livestock and is considered one of the most important tropical ectoparasites. The traditional method of control is based on the intensive use of pesticides, however the indiscriminate use of these compounds over the years has led to the selection of resistant ticks. Hydrolases, particularly esterases (EST), have been reported to be associated with acaricide resistance in B. microplus. We compared the esterase profile of susceptible and cypermethrin-resistant strains of adult B. microplus and a pyrethroid susceptible reference strain (the Mozzo strain) using polyacrylamide gel electrophoresis and specific staining. The electrophoretic profiles of protein extracts revealed the presence of four regions with esterase activity in the cypermethrin-resistant strain and three of these regions in the susceptible strains. The bands were numbered EST-1 to EST-4 in sequence (starting from the anode) according to their decrease in negative charge. The EST-1A and EST-1B enzymes were detected only in the resistant strain. The inhibition studies with eserine sulfate, copper sulfate, p- p-chloromercuribenzoate (pCMB), malathion and phenylmethylsulfonyl fluoride (PMSF) indicated that the EST-1A and EST-1B enzymes belong to the acetylcholinesterase class and are probably associated with resistance to acaricides in this Brazilian resistant strain of B. microplus.

Ribosomal DNA ITS-1 intergenic spacer polymorphism in triatomines (Triatominae, Heteroptera).

The length polymorphism of ribosomal DNA ITS-1 intergenic spacer was analyzed in eight species of triatomines belonging to Triatoma, Rhodnius, and Panstrongylus genera. The analyzed species were Rhodnius domesticus, R. neivai, R. robustus, Triatoma brasiliensis, T. infestans, T. vitticeps, Panstrongylus megistus, and P. herreri. These insects are vectors of Chagas' disease, one of the most prominent public health problems among South American countries. This work allowed the differentiation between species of the Triatomini and Rhodniini tribes through the analysis of ITS-1 length polymorphism by PCR and RFLP techniques. The species of the Triatoma and Panstrongylus genera presented an amplified ITS-1 fragment between 600 and 1000 bp, whereas Rhodnius presented a less variable ITS-1 length fragment, around 300 bp, which could reflect the monophyletic origin of the Rhodniini tribe. Species belonging to this genus were further differentiated by RFLP with HaeIII and AluI endonucleases. Our results corroborate the hypothesis of polyphyletic origin in this group of insects and contribute to knowledge about evolutionary relationships in triatomines.

Variation of genetic expression during development, revealed by esterase patterns in Aedes aegypti (Diptera, Culicidae).

Polyacrylamide gel electrophoresis was used to analyze esterase patterns during development of Aedes aegypti from the cities of Marília and São José do Rio Preto (SJRP), Brazil. The zymograms showed a total of 23 esterase bands, 22 of which were in the specimens from Marília and 19 in those from SJRP. These esterase bands were considered to be the product of 23 alleles distributed tentatively in eight genetic loci. Most of the alleles were developmentally regulated. The larval stage expressed the greatest number of them (19 alleles, from the eight loci, in Marília; and 17 alleles, from seven loci, in SJRP). The pupal stage expressed 10 alleles from seven loci, in both populations, and the adult stage expressed 8 alleles from five and six loci in SJRP and Marília, respectively. Some alleles that were active in every stage were developmentally controlled at the level of expression (amount of product). A single allele was constitutively and highly expressed, in larvae, pupae, and adults, in both populations. Differences in esterase synthesis among stages are probably due to regulatory mechanisms acting in agreement with the requirements of a variable number of processes in which esterases are involved. The larval stage is the most active in developmental processes and shows very intense intake of food and very high mobility. These features may demand increased esterase production at that stage. Comparison of the two populations examined showed (besides the existence of alleles that they do not share) that they exhibit differences in the control of expression of other alleles. Such findings may reflect genetic differences between founders in each population, but the possibility of involvement of the intensive use of insecticides in SJRP is also discussed.

Problemática vetorial da doença de chagas / Vectorial problematic of the chagas disease

As doenças de chagas é uma parasitose de natureza endêmica com uma alta prevalência entre as doenças cardíacas na América Latina. Cerca de 11 milhões de indivíduos estão infectados nas áreas endêmicas. O agente etiológico da doença de Chagas é o Trypanosoma cruzi e os vetores são insetos hematófagos peretencentes à ordem Heteroptera e família Reduviidae, mais conhecidos como triatomídeos. estudos entomológicos demonstraram que e´spécies secundárias de triatomíneos vêm aumentando sua densidade nos domicílios nos últimos anos. Existe uma preocupação atualcom estas espécies na transmissão da doença de Chagas é prejudicado pelo progressivo desinteresse público e mesmo em áreas onde a transmissão é controlada, há um contínuo risco de transmissão devido à sobrevivência dos vetores e baixa atividade da vigilância sanitária. É importante que se tenha uma visão crítica quanto ao possível retorno da transmissão vetorial. populações emergentes de tritomíneos em regiões de rsico devem ser analisadas periodicamente

Molecular analysis of the rDNA ITS-1 intergenic spacer in Drosophila mulleri, D. arizonae, and their hybrids.

We analyzed the ITS-1 spacer region of the rDNA in Drosophila mulleri and D. arizonae, two sibling species belonging to the mulleri complex (repleta group) and in hybrids obtained in both cross directions. In spite of several previous studies showing the incompatibility of crosses involving D. arizonae females and D. mulleri males, we were able to obtain hybrids in this direction. Complete ITS-1 region was amplified using primers with homology at the 3'-end of the 18S rDNA and the 5'-end of the 5.8S rDNA genes. Our data demonstrated that D. mulleri and D. arizonae can be differentiated as they present a difference in length for the ITS-1 region. The amplified fragment for this region in D. mulleri has a length of 600 bp, whereas in D. arizonae this fragment is about 500 bp. It was also observed that male and female hybrids obtained in both cross directions present two amplified fragments, confirming the location of the ribosomal cistrons in the X chromosomes and microchromosomes of both parental species.