Caracterização molecular da dieta do morcego hematófago desmodus rotundus (mammalia: chiroptera) na Amazônia brasileira

Abstract

Traditional methods based on the identification of vertebrate´s diet by direct analysis of feces are efficient for studies of feeding ecology but this approach fails for vampire bats. The use of molecular methods based on the isolation and amplification of DNA by PCR from feces has become an important tool for the knowledge of the diet of vertebrate predators such us vampire bats. In the present study a molecular method based on PCR-RFLP and analysis of DNA isolated from feces of Desmodus rotundus was developed for the identification of five prey species (chicken, cattle, pork, human and dog) most commonly attacked by this bat species in Brazilian Amazon communities. An experiment was carried out using individuals of D. rotundus kept on captivity in order to collect feces samples of the bats artificially fed with blood of each prey species. Total genomic DNA was successfully extracted from collected feces as well as from the prey species blood. Universal molecular primers were used for PCR amplification of a mitochondrial cytocrome b gene region (380 bp) using DNA isolated from bat´s feces and prey´s blood. For the RFLP analysis PCR fragments were digested using five restriction enzymes (Hae III, Rsa I, Taq I, Bmg BI, Xho I) selected for discrimination among species. The restricted patterns found were similar between the two treatments (feces and blood). The PCR-RFLP analysis allowed the correct differentiation of the five species of prey used by D. rotundus in the Brazilian Amazon rural communities. This study is pioneer by proposing and developing a method for the precise identification of the prey species used by the vampire bat D. rotundus. Using the method developed here we investigated also the feeding habits of vampire bats in field conditions which allow the precise identification of the prey species of the bats were using for feeding in the Brazilian Amazon rural communities. Eighteen human settlements located in Madeira and Aripuana rivers and in Ayapuá lake, Purus river, were sampled. In these places bats were captured and feces samples were collected. A total of 157 individuals of D. rotundus and six of Diaemus youngi were captured and 88 samples of feces were collected and analysed using PCR-RFLP as described previously. The amplified products not identified by the PCR-RFLP technique were sequenced and compared to Genebank dataset using BLAST. For 58 out of 88 samples we successfully amplified the cyt-b gene region (~380 bp). We were able to unambiguously identify four species of prey (chicken, pork, cattle, and dog) by the analysis of 48 samples (55%) of D. rotundus feces collected in field. The analysis showed that chicken was the most commonly prey consumed by D. rotundus followed by pork (32%), cattle (5%), and dog (2%). The PCR-RFLP analyses of fecal DNA haven´t showed any different restriction patterns from the expected for the prey species analysed here indicating that the vampire bats haven´t consumed blood of wild mammals or avian species around the communities. Also PCR-RFLP analysis of feces samples of D. youngi indicated the ingestion of chicken and pork blood by this species. That is the first record of a mammal species attacked by D. youngi in field. Despite the records of people attacked by D. rotundus in the study communities the molecular analyses carried out here haven´t indicated presence of human DNA in the feces of the bats. The results presented showed that the method developed using PCR-RFLP technique is an useful tool for analysis of the vampire bat diet in the Brazilian Amazon countryside. Future studies about frequency of D. rotundus attacks to people in countryside communities are also suggested to support programs aiming to control the populations of the vampire bats in the region.