Identification and characterization of microsatellite markers for population genetic studies of Panstrongylus megistus (Burmeister, 1835) (Triatominae: Reduviidae)

Background Panstrongylus megistus is the most important vector of Chagas disease in Brazil. Studies show that the principal factor hindering the control of triatomines is reinfestation of houses previously treated with insecticides. Studies at the microgeographic level are therefore necessary to better understand these events. However, an efficient molecular marker is not yet available for carrying out such analyses in this species. The aim of the present study was to identify and characterize microsatellite loci for future population genetic studies of P. megistus. Methods This study work consisted of five stages: (i) sequencing of genomic DNA; (ii) assembly and selection of contigs containing microsatellites; (iii) validation of amplification and evaluation of polymorphic loci; (iv) standardization of the polymorphic loci; and (v) verification of cross-amplification with other triatomine species. Results Sequencing of males and females generated 7,908,463 contigs with a total length of 2,043,422,613 bp. A total of 2,043,690 regions with microsatellites in 1,441,091 contigs were obtained, with mononucleotide repeats being the most abundant class. From a panel of 96 loci it was possible to visualize polymorphisms in 64.55% of the loci. Of the 20 loci genotyped, the number of alleles varied from two to nine with an average of 4.9. Cross-amplification with other species of triatomines was observed in 13 of the loci. Conclusions Due to the high number of alleles encountered, polymorphism and the capacity to amplify from geographically distant populations, the microsatellites described here show promise for utilization in population genetic studies of P. megistus. Graphic abstract

VIGILÂNCIA ENTOMOLÓGICA DA DOENÇA DE CHAGAS NA REGIÃO LESTE DE MINAS GERAIS, BRASIL

Introdução: Segundo a OMS temos cerca de 6 milhões de indivíduos infectados pelo parasito Trypanosoma cruzi no mundo. O mecanismo de transmissão vetorial continua sendo de grande importância na epidemiologia da doença, sendo, portanto, o alvo dos programas de controle. No Brasil observou-se uma notável diminuição de novos casos da infecção após a implementação das campanhas de controle vetorial. Objetivo: Avaliar a vigilância entomológica da DCh nos municípios da jurisdição da Superintendência Regional de Saúde de Governador Valadares (SRS/GV) nos anos de 2014 a 2019. Métodos: A área estudada corresponde a 51 municípios da mesorregião do Vale do Rio Doce, no Leste de Minas Gerais. Os dados foram disponibilizados pelo laboratório de referência em entomologia da SRS/GV. Os insetos capturados pelos moradores e enviados foram avaliados quanto ao hábito alimentar, estádio evolutivo, local de captura, espécie do triatomíneo e presença ou ausência de infecção pelo T. cruzi. Resultados: Foram encaminhados à SRS/GV 1.704 insetos, sendo 1.491 hematófagos, 107 fitófagos e 99 predadores. Avaliando os insetos hematófagos quanto ao estádio evolutivo constatou-se que 1.425 (95,6%) eram adultos e 66 (4,4%) eram ninfas, sendo a maioria capturados no intradomicílio (1.083/1.491). Foram examinados 1.225 insetos, sendo constatado a infecção pelo T. cruzi em 514 exemplares, revelando assim uma taxa de infecção natural de 42%. Dentre os insetos infectados, 407 (79,5%) exemplares foram capturados no intradomicílio, 96 (18,5%) no peridomicílio e 11 (2%) em locais não identificados. As seguintes espécies de triatomíneos foram identificadas na região: Triatoma vitticeps (936), Panstrongylus megistus (284), Panstrongylus diasi (244), Rhodnius neglectus (11) e Panstrongylus geniculatus (1). Foi constatado a presença de triatomíneos infectados em 46 municípios, com destaque para Conselheiro Pena (64), Tarumirim (48), Sobrália (46), Capitão Andrade (44), Itanhomi (36) e José Raydan (30). Conclusão: Os dados nos permitem afirmar que T. vitticeps é a espécie predominante na região, sendo capturados frequentemente insetos adultos no intradomicílio e com consideráveis taxas de infecção natural pelo T. cruzi. Vale ressaltar ainda o encontro de ninfas no intradomicílio que caracteriza um processo de colonização e reforça a necessidade de aprimoramento das ações de vigilância entomológica na região atendida pela SRS/GV.

Avaliação externa da qualidade da identificação entomológica de triatomíneos realizada na Rede de Laboratórios Públicos em Pernambuco, 2017

Resumo Objetivo: Avaliar a qualidade da identificação de triatomíneos na rede de laboratórios do estado de Pernambuco, Brasil. Métodos: Os nove laboratórios participantes receberam material de apoio com as chaves dicotômicas e um painel composto por sete insetos triatomíneos conhecidos no estado, para identificação da situação do espécime no recebimento (estrutura completa ou danificada), espécie e sexo. Resultados: Nove laboratórios de 12 aderiram ao estudo. A proporção de acerto para identificação do sexo foi de 56/63, e para espécie, 45/63, não apresentando relação direta com a ocorrência de danos nas estruturas morfológicas durante o transporte dos insetos. Para Panstrongylus megistus, houve acerto em todos os espécimes (9/9), enquanto para espécies do gênero Rhodnius a proporção foi menor (3/9). Conclusão: Apesar do bom desempenho na identificação entomológica, as fragilidades observadas poderão orientar ações para melhoria na rede de laboratórios e serão essenciais para os programas de controle vetorial da doença de Chagas.

The Fungal Microbiota of the Digestive Tract of the Chagas Disease Vectors Triatoma infestans Klug, 1834 and Panstrongylus megistus Burmeister, 1835

Chagas is a neglected disease, one of Brazil’s main medical and social problems and a serious public health problem in the Americas, with more recent occurrences in non-endemic countries outside of the Americas. Research into the microbiota of triatomines is relevant because of its potential role in vector competence and as a proposed biological control strategy. Stressing a possible insect-fungal interaction in the development of Trypanosoma cruzi, and considering the lack of studies on the subject, we analyzed the fungal microbiota of the digestive tract of two species considered important vectors of Trypanosoma cruzi: Triatoma infestans and Panstrongylus megistus. Specimens were dissected, digestive tracts macerated and contents serially diluted. Each aliquot was seeded in three culture media. The plates were incubated in type B.O.D. climate chambers for 21 days, after which isolated colonies were morphological characterized and identified. There have been few published studies on the fungal microbiota of the triatomine digestive tract. Comparing the results found here with existing data reveals that the genera Aspergillus and Penicillium are commonly found in the digestive tract of the studied triatomines. Among the several genera identified, the species found in the highest percentages were Aspergillus flavus, Paecilomyces variotii, Penicillium waksmanii, Penicillium raistrickii and Penicillium fellutanum. Quantitative differences in the number of isolated fungal strains were observed according to sex and nymphal stage of the vector. The present findings corroborate those found in the literature, showing that there is a natural fungal microbiota in triatomines. Data revealing quantitative differences in isolated fungal strains found in male, female and nymphs reinforce the idea that their presence is related to physiology and fasting resistance. The secondary metabolite-producing fungi isolated in this work have in their biology great potential to be tested with regard to the establishment of T. cruzi in the digestive tract of its vector.

First report of colonization by Panstrongylus megistus (Burmeister, 1835) (Hemiptera, Reduviidae, Triatominae) in the Metropolitan Region of São Paulo, Brazil

We report the first known occurrence of Panstrongylus megistus (Burmeister, 1835) (Hemiptera, Reduviidae, Triatominae) in the Metropolitan Region of São Paulo, Brazil. In 2018, adult specimens were sent by residents to the competent authorities and, in the inspection of the property, a large focus associated with a marsupial nest was found. This finding reinforces the importance of the species in the state of São Paulo, serves as an alert for epidemiological surveillance and extends the species colonization area in the state of São Paulo.

The Fat Body of the Hematophagous Insect, Panstrongylus megistus (Hemiptera: Reduviidae): Histological Features and Participation of the β-Chain of ATP Synthase in the Lipophorin-Mediated Lipid Transfer

In insects, lipid transfer to the tissues is mediated by lipophorin, the major circulating lipoprotein, mainly through a nonendocytic pathway involving docking receptors. Currently, the role of such receptors in lipid metabolism remains poorly understood. In this work, we performed a histological characterization of the fat body of the Chagas’ disease vector, Panstrongylus megistus (Burmeister), subjected to different nutritional conditions. In addition, we addressed the role of the β-chain of ATP synthase (β-ATPase) in the process of lipid transfer from lipophorin to the fat body. Fifth-instar nymphs in either fasting or fed condition were employed in the assays. Histological examination revealed that the fat body was composed by diverse trophocyte phenotypes. In the fasting condition, the cells were smaller and presented a homogeneous cytoplasmic content. The fat body of fed insects increased in size mainly due to the enlargement of lipid stores. In this condition, trophocytes contained abundant lipid droplets, and the rough endoplasmic reticulum was highly developed and mitochondria appeared elongated. Immunofluorescence assays showed that the β-ATPase, a putative lipophorin receptor, was located on the surface of fat body cells colocalizing partially with lipophorin, which suggests their interaction. No changes in β-ATPase expression were found in fasting and fed insects. Blocking the lipophorin–β-ATPase interaction impaired the lipophorin-mediated lipid transfer to the fat body. The results showed that the nutritional status of the insect influenced the morphohistological features of the tissue. Besides, these findings suggest that β-ATPase functions as a lipophorin docking receptor in the fat body.