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Resistência a insecticidas em Anopheles gambiae s.l. na região de Luanda, Angola
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Em Angola, a malária é a principal causa de morbilidade e de mortalidade infantil. O controlo de vectores com recurso aos insecticidas representa uma parte importante da estratégia actual para a prevenção da doença. Em Anopheles gambiae s.s., principal vector de malária em África, estão identificadas duas mutações pontuais no gene que codifica os canais de sódio das membranas das células do sistema nervoso, conferindo resistência knockdown (kdr) aos insecticidas piretróides e ao DDT. Também se encontra descrita para esta espécie uma mutação no gene acetilcolinesterase-1 (ace-1), associada à resistência a carbamatos e organofosfatos. Este trabalho teve como principal objectivo avaliar o nível de resistência aos insecticidas em An. gambiae da província de Luanda, Angola e determinar a frequência destas mutações.
Foram realizadas colheitas entomológicas em 2009 e 2010, através da prospeção de criadouros larvares. Os insectos capturados foram criados até a emergência do adulto e sujeitos a ensaios de susceptibilidade a insecticidas, através de testes da OMS. A identificação de espécies e formas moleculares do complexo An. gambiae, bem como a pesquisa de mutações no gene ace-1 foram feitas por PCR-RFLP. A pesquisa de mutações no gene kdr foi realizada por PIRA-PCR. Amostras selecionadas de mosquitos (incluindo uma amostra proveniente de uma colheita de adultos) foram ainda genotipadas para 11 loci microssatélites.
Os níveis de resistência para a permetrina, DDT e -cialotrina foram elevados, com taxas de mortalidade inferiores a 70% em ambos os anos. Em contraste, as taxas de mortalidade foram sempre acima de 98% para bendiocarb e fenitrotião, indicadoras de susceptibilidade a estes insecticidas. Todas as amostras processadas foram identificadas como An. gambiae s.s., forma molecular M e não se observou a mutação no gene ace-1 associada à resistência. Em ambos os anos, foi detectada apenas a mutação L1014F no locus kdr e a frequência do alelo mutante (TTT) foi bastante elevada. Em 2009, observou-se uma associação entre genótipos homozigóticos para o alelo 1014F e o fenótipo resistente, para os insecticidas piretróides e para o DDT.
O polimorfismo dos loci microssatélites analisados foi elevado, com a riqueza alélica a variar entre 5 (45C1) e 20 (H128) e a heterozigotia esperada entre 0,529 (H577) e 0,862 (H249). A análise genética não revelou um grau de parentesco entre os indivíduos que constituíram as amostras estudadas. Este resultado sugere que os elevados níveis de resistência observados não foram influenciados pelo método de colheita de mosquitos que, em certas condições, poderia contribuir para a amostragem de indivíduos aparentados.
In Angola malaria is still one of the leading causes of child morbidity and mortality. Vector control depends mainly on insecticides which constitutes an important part of the national strategy to prevent and control malaria transmission. In Anopheles gambiae s.s., the main malaria vector in sub-saharan Africa, , two point mutations have been identified in the gene encoding the sodium channels in cells of the nervous system. These mutations confer a knockdown resistance (Kdr) to pyrethroids and DDT. A mutation in the acetylcholinesterase-1 gene (ace-1), associated with resistance to carbamate and organophosphate insecticides, was also described for this species. The aims of this study were to evaluate insecticide resistance levels in A. gambiae from Luanda, Angola and determine the frequency of these mutations. Larval sampling was carried out in breeding sites between in 2009 and 2010. Immature mosquitoes were allowed to reach the adult stage and subsequently used in WHO standardized insecticide susceptibility assays. A. gambiae species identification and mutation detection in the ace-1 gene was carried out by PCR-RFLP. Mutations in the kdr gene were identified by PIRA-PCR. In addition, selected mosquito samples (including a sample of wild caught adults) were genotyped for 11 microsatellites. The WHO insecticide susceptibility assays have shown high levels of resistance to permethrin, DDT and -cialothrin, with mortality rates below 70% in both years. In contrast, the rates of mortality were high for bendiocarb and fenitrothion, indicating susceptibility to these insecticides. All specimens were identified as A. gambiae s.s M-form. The resistance-associated mutation at ace-1 was not detected. In both years, only the L1014F mutation was found at the kdr locus and the frequency of the mutant allele (TTT) was very high. In 2009, an association between homozygotic genotypes for the mutant allele and the resistant phenotype for pyrethroids and DDT was observed. Polymorphism at microsatellite loci was high, with allelic richness varying between 5 (45C1) and 20 (H128) and expected heterozygosity between 0,529 (H577) and 0,862 (H249). Genetic analysis did not reveal a degree of relatedness between the individuals that constituted the samples studied. Taken together, these results suggest that the high levels of resistance observed were not influenced by the sampling method used, which in certain conditions can contribute to the sampling of related individuals.
In Angola malaria is still one of the leading causes of child morbidity and mortality. Vector control depends mainly on insecticides which constitutes an important part of the national strategy to prevent and control malaria transmission. In Anopheles gambiae s.s., the main malaria vector in sub-saharan Africa, , two point mutations have been identified in the gene encoding the sodium channels in cells of the nervous system. These mutations confer a knockdown resistance (Kdr) to pyrethroids and DDT. A mutation in the acetylcholinesterase-1 gene (ace-1), associated with resistance to carbamate and organophosphate insecticides, was also described for this species. The aims of this study were to evaluate insecticide resistance levels in A. gambiae from Luanda, Angola and determine the frequency of these mutations. Larval sampling was carried out in breeding sites between in 2009 and 2010. Immature mosquitoes were allowed to reach the adult stage and subsequently used in WHO standardized insecticide susceptibility assays. A. gambiae species identification and mutation detection in the ace-1 gene was carried out by PCR-RFLP. Mutations in the kdr gene were identified by PIRA-PCR. In addition, selected mosquito samples (including a sample of wild caught adults) were genotyped for 11 microsatellites. The WHO insecticide susceptibility assays have shown high levels of resistance to permethrin, DDT and -cialothrin, with mortality rates below 70% in both years. In contrast, the rates of mortality were high for bendiocarb and fenitrothion, indicating susceptibility to these insecticides. All specimens were identified as A. gambiae s.s M-form. The resistance-associated mutation at ace-1 was not detected. In both years, only the L1014F mutation was found at the kdr locus and the frequency of the mutant allele (TTT) was very high. In 2009, an association between homozygotic genotypes for the mutant allele and the resistant phenotype for pyrethroids and DDT was observed. Polymorphism at microsatellite loci was high, with allelic richness varying between 5 (45C1) and 20 (H128) and expected heterozygosity between 0,529 (H577) and 0,862 (H249). Genetic analysis did not reveal a degree of relatedness between the individuals that constituted the samples studied. Taken together, these results suggest that the high levels of resistance observed were not influenced by the sampling method used, which in certain conditions can contribute to the sampling of related individuals.
Descrição
Palavras-chave
Parasitologia médica Saúde pública Malária Morbilidade Anopheles gambiae Luanda Angola Resistência a insecticidas Genes kdr microssatélites
Contexto Educativo
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Instituto de Higiene e Medicina Tropical