Investigation On The Animal Hosts Infected With Anaplasma Phagocytophilum In The Northern Of Yun Ling

[Objective]The Northern of Yun Ling in Western Yunnan were considered as the investigated site,dogs、domestic animals and small mammals as the research species. A nested polymerase chain reaction(PCR) method was used to amplify and Sequencing the fragments of Anaplasma 16 Sr RNA. Explora the prevalence of Anaplasma phagocytophilum in dogs、domestic animals and small mammals from the Northern of Yun Ling in Western Yunnan to provide the basis for a reasonable control measures. [Methods]1. Collection sample The Northern of Yun Ling in Western Yunnan were considered as the investigated site,Collectioned 1096 samples. Contains 602 small animals(141 from east of the White Snow Mountain, 71 from west of the White Snow mountain,66 from the village of Xia Ruo,95 from the village of Yang La, 31 from the village of Weng Shui, 25 from the village of Dong Wang,12 from the village of Za Ge, 4 from the village of Ni Xi,3 from the village of Zhong Dian, 2 from the village of Jian Tang, 15 from the village of San Ba, 52 from Rhinopithecus bieti reserves, 20 from the village of Nuoge and 15 from the village of Panquan); 315 dogs(73 from the village of Yun Ling, 73 from the village of Fo Shan,13 from the village of Sheng Ping, 58 from the village of Yan Men, 28 from the village of Yang La, 70 from the village of Xia Ruo); 179 domestic animals(45 oxen blood from the county of De Qin, 110 oxen blood from the county of Shangri-La, 24 sheep blood from the county of Shangri-La).2. Collection DNA The QIANEN bacterial kit has been used to extract Yetsinia.3. Nested Polymearse Chain Reactio The 16 Sr RNA genes of Anaplasma were then amplified by nest PCR.4. Electrophoresis PCR amplification products using 1.5% agarose gel electrophoresis, results analysised by Gel imager analysis.5. Sequencing and Analysis The PCR products were Compared and sequenced in homology by using BLAST program. Phylogenetic analysis for 16 Sr RNA gene sequences published in Gen Bank Was conducted by using Mega 6.0 software. [Results]1. A total number of 602 small animals,which belongs to 4 orders 7 family l 6 genera 30 species were tested,10(1.66%)wore positive in 5 genera 6 species.2. Ten positive small mammals、Canada(HG916766.1)、Siberia and Russia(HM366583.1) at the same banch, The similarities of the nucleotide sequences with them is 99%. The different host individuals such as hedgehogs、ticks and sheeps from Russia(HQ629925.1)、Slovenia(AF481855.1)、Hubei(HQ872463.1)、Jilin(DQ449948.1)、Germany(AY706390.1)、France(EU857671.1)、Spain(AY829457.1) and the positive small mammals at the same banch, But far away.3. 18 positive sanmples find from the 315 dogs, 5.71%(18/315) showed positive in which the positiver rate was 7.14%(12/168) in chinese rural dog, 5.13%(4/78) in tibetan mastiff and 2.99%(2/67) in pug, no positive sample was found in saarloos.4. The similarities of the nucleotide sequences with those deposited in Gen Bank were 99%~100%. Two genetic types of A.phagocytophilum were demonstrated in dogs in these areas and the two groups of epidemiological strains were accounted for 16.67% and 83.33% in tested isolates respectively. The similarities of A.phagocytophilum sequences with Yunnan HGA1 and the ixodes persulcatus on the territory of Ural, Siberia and Far East of Russia(HM366583) were 100%, The similarities of A.phagocytophilum sequences with Yunnan HGA2 and the ixodes scapularis in Canada(HG916766) were 99%.5. 17 positive oxen blood sanmples find from the 315 domestic animals, 9.50%(17/179)showed positive, no positive sample was found in sheep.6. Seventeen positive oxen 、 China(GQ900613.1 、 HM64175.1 、 GQ499995.1 、GQ500045.1)、France(JX841254.1)、Mediterranean(EU436154.1)at the same banch, the similarities of the nucleotide sequences with them is 97-99%, the northern xinjiang(KJ459350.1)、 China(GQ500058.1) and the positive oxen at the same banch, But far away. [Conclusion]It is concluded that the infection of Anaplasma phagocytophilum is existed in small mammals、domestic animals and dogs from the Northern of Yun Ling in Western Yunnan.Further investigation of its vectors and hosts and especially febrile patients who are probable infection with Anaplasma phagocytophilum are indicated.