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Background   Trypanosoma cruzi  infection via oral route results in outbreaks or cases of acute Chagas disease (ACD) in different Brazilian regions and poses a novel epidemiological scenario. In the Espírito Santo state (southeastern Brazil), a fatal case of a patient with ACD led us to investigate the enzootic scenario to avoid the development of new cases. At the studied locality,  Triatoma vitticeps  exhibited high  T. cruzi  infection rates and frequently invaded residences.    Methods  Sylvatic and domestic mammals in the Rio da Prata locality, where the ACD case occurred, and in four surrounding areas (Baia Nova, Buenos Aires, Santa Rita and Todos os Santos) were examined and underwent parasitological and serological tests. Triatomines were collected for a fecal material exam, culturing and mini-exon gene molecular characterization, followed by RFLP-PCR of H3/Alul. Paraffin-embedded cardiac tissue of a patient was washed with xylene to remove paraffin and DNA was extracted using the phenol-chloroform method. For genotype characterization, PCR was performed to amplify the 1f8, GPI and 18S rRNA genes. In the case of V7V8 SSU rRNA, the PCR products were molecularly cloned. PCR products were sequenced and compared to sequences in GenBank. Phylogenetic analysis using maximum likelihood method with 1000 bootstrap replicates was performed.    Results  None of the animals showed positive hemocultures. Three rodents and two dogs showed signs of infection, as inferred from borderline serological titers.  T. vitticeps  was the only triatomine species identified and showed  T. cruzi  infection by DTUs TcI and TcIV. The analysis of cardiac tissue DNA showed mixed infection by  T. cruzi  (DTUs I, II, III and IV) and  Trypanosoma dionisii .    Conclusions  Each case or outbreak of ACD should be analyzed as a particular epidemiological occurrence. The results indicated that mixed infections in humans may play a role in pathogenicity and may be more common than is currently recognized. Direct molecular characterization from biological samples is essential because this procedure avoids parasite selection.  T. dionisii  may under certain and unknown circumstances infect humans. The distribution of  T. cruzi  DTUS TcIII and TcIV in Brazilian biomes is broader than has been assumed to date.    Electronic supplementary material  The online version of this article (doi:10.1186/s13071-016-1754-4) contains supplementary material, which is available to authorized users.

Ecological scenario and Trypanosoma cruzi DTU characterization of a fatal acute Chagas disease case transmitted orally (Espírito Santo state, Brazil)