The Investigate Of The Bartonella Spp. Infection In Bats And Its Ectoparasite From The Southwestern Yunnan Province

ObjectiveThe objectives of the study were to investigate the epidemiological distribution and carrying situation of Bartonella spp.in bats and its ectoparasite from the southwest Yunnan province,and to explore phylogenetic analysis of its genetic diversity and system.This study will provide scientific basis for the Southwest Yunnan Bartonella host and media animals.MethodsSamples of this investigation were selected from the southwest Yunnan province.After the bats were captured by the net-catching method,the ectoparasite of the bats were collected for morphological classification.The bat spleens and blood samples were collected with dissected aseptically and placed in liquid nitrogen tank back to the laboratory,placed in-80℃ refrigerator frozen.The acquisition of bat ectoparasite and bat spleen samples in strict aseptic conditions and grinded into tissue suspension,according to the TIANamp Genomic DNA Kit operating on the bat ectoparasites、bat spleen and blood clots(serum free)of three kinds of samples were extracted from DNA.First,the species of blood sucking insects were amplified and identified by universal primers of arthropod mitochondrial cytochrome oxidase I(CO I),and then amplified by primers gltA,16S-23 S rRNA ITS,groEL,ftsZ and 16 S rRNA genes from three kinds of samples.The positive samples obtained were sent for sequencing,the sequence fragments were spliced,edited and corrected by SeqMan in the DNAstar software package;base pairing and nucleotide and deduced amino acid sequence homology analysis were performed with Clustal X(1.83)software;MEGA 5.0 software neighbor construction(neighbor-joining)building phylogenetic tree analysis phylogenetic evolution relations.ResultsIn this study,there were three species of bats in 12 counties of 7 cities in southwesternYunnan province,including Rhinolophus luctus,Myotis daubentonii and Rousettus leschenaultii;there are total number of 1052 bat ectoparasites of 6 species in 71 batch,including Nycteribiidae Samouelle、Streblidae Kolenati、Zerconidae、Ixodoidea、Odontopsyllus and Leptidea sinapis eggs.In addition 355 samples of bats’ spleens and 231 bats blood clot were collected.With all primers PCR amplification showed that 50 batches of bat ectoparasites were positive for Bartonella spp.,and the infection rate was 70.42%(50/71);there were 13 bat spleen tissues and 12 bat blood clot samples were positive for Bartonella spp.and the infection rates were 3.66%(13/355)and 5.19%(12/231)respectively.The phylogenetic analysis of the positive samples was similar with Bartonella quintana、Bartonella vinsonii subsp.Berkhoffii、Bartonella australis and Bartonella clarridgeiae,and most of positive samples were belong to undetermined species of Bartonella spp..In addition,Rickettsia and Ehrlichia positive specimens were also detected in three kinds of samples.At last the positive rate varies with the different samples and the primers and there are differences in Bartonella spp.genotypes between different species of bats and their ectoparasites.ConclusionThe study is the first report of detection by PCR amplification of Bartonella spp.infection in bats and their ectoparasites from the southwest of Yunnan province.There are four species of Bartonella spp.genotypes which have three kinds of human pathogenic genotype.The results provided a theoretical basis for the monitoring and prevention of Bartonella disease in Yunnan province.