title: Characterization of the microbiota in the guts of Triatoma brasiliensis and Triatoma pseudomaculata infected by Trypanosoma cruzi in natural conditions using culture independent methods.
abstract: BACKGROUND
  Chagas disease is caused by Trypanosoma cruzi, which is transmitted by triatomine vectors. The northeastern region of Brazil is endemic for Chagas disease and has the largest diversity of triatomine species. T. cruzi development in its triatomine vector depends on diverse factors, including the composition of bacterial gut microbiota.
  
  
  METHODS
  We characterized the triatomines captured in the municipality of Russas (Ceará) by sequencing the cytochrome c oxidase subunit I (COI) gene. The composition of the bacterial community in the gut of peridomestic Triatoma brasiliensis and Triatoma pseudomaculata was investigated using culture independent methods based on the amplification of the 16S rRNA gene by polymerase chain reaction (PCR), denaturing gradient gel electrophoresis (DGGE), DNA fragment cloning, Sanger sequencing and 454 pyrosequencing. Additionally, we identified TcI and TcII types of T. cruzi by sequencing amplicons from the gut metagenomic DNA with primers for the mini-exon gene.
  
  
  RESULTS
  Triatomines collected in the peridomestic ecotopes were diagnosed as T. pseudomaculata and T. brasiliensis by comparing their COI sequence with GenBank. The rate of infection by T. cruzi in adult triatomines reached 80% for T. pseudomaculata and 90% for T. brasiliensis. According to the DNA sequences from the DGGE bands, the triatomine gut microbiota was primarily composed of Proteobacteria and Actinobacteria. However, Firmicutes and Bacteroidetes were also detected, although in much lower proportions. Serratia was the main genus, as it was encountered in all samples analyzed by DGGE and 454 pyrosequencing. Members of Corynebacterinae, a suborder of the Actinomycetales, formed the next most important group. The cloning and sequencing of full-length 16S rRNA genes confirmed the presence of Serratia marcescens, Dietzia sp., Gordonia terrae, Corynebacterium stationis and Corynebacterium glutamicum.
  
  
  CONCLUSIONS
  The study of the bacterial microbiota in the triatomine gut has gained increased attention because of the possible role it may play in the epidemiology of Chagas disease by competing with T. cruzi. Culture independent methods have shown that the bacterial composition of the microbiota in the guts of peridomestic triatomines is made up by only few bacterial species.
authors.affiliation: Laboratório de Bioquímica e Fisiologia de Insetos, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz (IOC/FIOCRUZ), Rio de Janeiro, RJ, Brazil. mgumiel@ioc.fiocruz.br.
authors.forename: Marcia
authors.initials: M
authors.lastname: Gumiel
authors.affiliation: Laboratório de Biologia Computacional e Sistemas, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz (IOC/FIOCRUZ), Rio de Janeiro, RJ, Brazil. fabio@ioc.fiocruz.br.|Departamento de Entomologia Molecular, Instituto Nacional de Entomologia Molecular (INCT-EM), Rio de Janeiro, RJ, Brazil. fabio@ioc.fiocruz.br.
authors.forename: Fabio Faria
authors.initials: FF
authors.lastname: da Mota
authors.affiliation: Laboratório de Bioquímica e Fisiologia de Insetos, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz (IOC/FIOCRUZ), Rio de Janeiro, RJ, Brazil. vanessa.rizzo7@gmail.com.
authors.forename: Vanessa de Sousa
authors.initials: Vde S
authors.lastname: Rizzo
authors.affiliation: Laboratório de Ecoepidemiologia da Doença de Chagas, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz (IOC/FIOCRUZ), Rio de Janeiro, RJ, Brazil. otiliasarquis@ioc.fiocruz.br.
authors.forename: Otília
authors.initials: O
authors.lastname: Sarquis
authors.affiliation: Laboratório de Bioquímica e Fisiologia de Insetos, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz (IOC/FIOCRUZ), Rio de Janeiro, RJ, Brazil. didan.castro@gmail.com.|Departamento de Entomologia Molecular, Instituto Nacional de Entomologia Molecular (INCT-EM), Rio de Janeiro, RJ, Brazil. didan.castro@gmail.com.
authors.forename: Daniele Pereira
authors.initials: DP
authors.lastname: de Castro
authors.affiliation: Laboratório de Ecoepidemiologia da Doença de Chagas, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz (IOC/FIOCRUZ), Rio de Janeiro, RJ, Brazil. mmlima@ioc.fiocruz.br.
authors.forename: Marli Maria
authors.initials: MM
authors.lastname: Lima
authors.affiliation: Laboratório de Bioquímica e Fisiologia de Insetos, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz (IOC/FIOCRUZ), Rio de Janeiro, RJ, Brazil. garciaeloi@gmail.com.|Departamento de Entomologia Molecular, Instituto Nacional de Entomologia Molecular (INCT-EM), Rio de Janeiro, RJ, Brazil. garciaeloi@gmail.com.
authors.forename: Eloi de Souza
authors.initials: Ede S
authors.lastname: Garcia
authors.affiliation: Laboratório de Modelagem de Sistemas Biológicos, National Institute for Science and Technology on Innovation in Neglected Diseases (INCT-IDN), Centro de Desenvolvimento Tecnológico em Saúde (CDTS), Fundação Oswaldo Cruz (FIOCRUZ), Rio de Janeiro, RJ, Brazil. nicolas.carels@cdts.fiocruz.br.
authors.forename: Nicolas
authors.initials: N
authors.lastname: Carels
authors.affiliation: Laboratório de Bioquímica e Fisiologia de Insetos, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz (IOC/FIOCRUZ), Rio de Janeiro, RJ, Brazil. azambuja.p@gmail.com.|Departamento de Entomologia Molecular, Instituto Nacional de Entomologia Molecular (INCT-EM), Rio de Janeiro, RJ, Brazil. azambuja.p@gmail.com.
authors.forename: Patricia
authors.initials: P
authors.lastname: Azambuja
mesh_terms: D000818:Animals; D001419:Bacteria / Q000145:classification / Q000235:genetics; D004269:DNA, Bacterial; D058645:Denaturing Gradient Gel Electrophoresis; D003576:Electron Transport Complex IV / Q000235:genetics; D005260:Female; D041981:Gastrointestinal Tract / Q000382:microbiology; D015964:Gene Expression Regulation, Bacterial; D009758:Nymph / Q000382:microbiology; D010802:Phylogeny; D016133:Polymerase Chain Reaction; D012329:RNA, Bacterial / Q000235:genetics; D014224:Triatoma / Q000145:classification / Q000235:genetics / Q000382:microbiology* / Q000469:parasitology*; D014349:Trypanosoma cruzi / Q000302:isolation & purification*
chemical_list: D004269:DNA, Bacterial; D012329:RNA, Bacterial; D003576:Electron Transport Complex IV