国内部分省市肉兔及蜱虫携带土拉杆菌的流行病学调查

本调查旨在探明土拉杆菌在国内部分省份的流行情况。利用本实验室建立的土拉杆菌通用及亚种PCR检测研究方法，对国内肉兔饲养密度比较大的省份和牛羊存栏的部分省市开展了肉兔和蜱虫中土拉杆菌的检测，并对阳性样本进行了土拉杆菌亚种的鉴定和基因测序验证。研究结果表明，肉兔肝脏组织和牛羊携带蜱虫中土拉杆菌DNA检测阳性，其中218份肉兔样本中12份阳性（阳性率5.5%），490份蜱虫样本中15份阳性（阳性率3.1%）；从地区分布来看，山东、河南、四川肉兔病料检测阳性，云南、山东蜱虫阳性率高；PCR检测显示其亚种为F.h，为土拉杆菌毒力较强的亚种。本次调查显示土拉杆菌在肉兔、蜱虫中存在，山东、云南地区存在土拉杆菌病公共卫生安全风险，应引起有关科研院校、医疗单位和政府部门的高度重视。     

PCR-based detection of Theileria ovis in Rhipicephalus bursa adult ticks

Tick-borne diseases in ruminants are common in tropical and subtropical regions and lead to meat and milk production losses. In this study, polymerase chain reaction (PCR) was used to assess the presence of Theileria ovis in Rhipicephalus bursa ticks. We have demonstrated that the PCR enabled detection of T. ovis in field isolates of R. bursa collected from naturally infested sheep and goats in eastern Turkey. The sampling was done in spring season (between May and June 2004). A total of 420 R. bursa were collected and randomly selected 192 number of them (97 female and 95 male) were dissected. Primers specific for 520 bp fragments small subunit ribosomal RNA (ssu rRNA) gene of T. ovis amplified products from 37 of the 192 (19.27%) samples. The parasite was detected in 17 (17.52%) female and in 20 (21.05%) male ticks. Two T. ovis amplicons from the tick samples were purified and sequenced. The resulting sequences were identical to the nucleotide sequence of the Turkish sheep strain of T. ovis. These results showed that R. bursa might play an important role in the field as a natural vector of T. ovis.     

我国部分地区牛羊弓形虫血清流行病学调查

为了解我国牛羊弓形虫病流行情况,应用间接血凝试验（IHA）对河南、山东、山西、内蒙古、云南、贵州6省区151份牛血清、50份奶样、490份羊血清进行了弓形虫病血清流行病学调查。结果显示：151份被检牛血清和50份牛奶样品,弓形虫抗体均为阴性。490份羊血清弓形虫抗体总阳性率5.71％,其中母羊、公羊血清阳性率分别为4.03％和9.79％;山羊、绵羊、杂交羊血清阳性率分别为6.58％、4.81％和5.13％;阳性率最高的为公山羊（13.2％）,最低的为母绵羊（2.96％）。28份阳性羊血清中,75％的抗体滴度为1：64,25％的抗体滴度为1：256。1岁后的羊,随年龄增长,血清阳性率升高。     

东南沿海地区水禽源大肠杆菌耐药表型及耐药基因型的调查

为研究水禽中大肠杆菌的耐药表型及耐药基因型,本研究从中国广东、福建、浙江、江苏和山东5个省市的水禽养殖场及周边环境中采集鸭粪便样品、水样以及土样1 505份,采用选择性培养基和基质辅助激光解吸电离飞行时间(MALDI-TOF-MS)质谱分离鉴定大肠杆菌,采用微量肉汤稀释法检测耐药表型,采用PCR方法筛选大肠杆菌中的25种耐药基因。结果显示,1 505份样品中共分离鉴定出449株大肠杆菌,分离率为29.8%,其中335份来源于粪便,52份来源于土壤,62份来源于水,这些样品中浙江(43.3%)和广东(43.2%)的分离率最高,其次是福建(28.1%)和江苏(23.8%),山东(18.3%)的分离率最低。449株大肠杆菌具有多重耐药性,对氨苄西林、环丙沙星、金霉素、氟苯尼考和磺胺类药物的耐药率分别为90.2%、50.3%、96.7%、87.3%和90.6%,而对替加环素和美罗培南则比较敏感,来源不同省份的大肠杆菌对不同抗菌药呈现出不同的耐药率。PCR检测结果发现,449株大肠杆菌中耐药基因tet(A)检出率最高(88.2%),随后是floR(58.13%)、sul2(57.02%)、tet(B)(53.9%)、cmlA(39.2%)和sul1(36.75%),未检测到qnrA、tet(M)和tet(X)耐药基因的大肠杆菌。在这5个省份中,上述耐药基因在山东省样品中分布均较高,而在浙江省和江苏省样品中分布较低。综上所述,本研究调查了中国东南沿海五省水禽及环境中大肠杆菌的耐药现状,为养殖业抗生素的规范使用提供理论依据。     

Identification of the etiologic agent of epizootic bovine abortion in field-collected Ornithodoros coriaceus Koch ticks

Epizootic bovine abortion (EBA), or foothill abortion as it has often been termed, is a tick-borne disease of pregnant cattle recognized in California, Nevada and Oregon. The primary objective of this study was to better define the relationship of a novel deltaproteobacterium, the putative etiological agent of EBA (aoEBA), with the Pajaroello tick (Ornithodoros coriaceus Koch), the recognized vector of EBA. Three developmental stages of O. coriaceus (larva, nymph, and adult) were collected from five locations in California, Nevada and Oregon. A polymerase chain reaction (PCR), developed for detection of aoEBA, was applied to DNA extracted from ticks. Southern blotting of the PCR products increased the number of ticks determined to be carrying the bacteria by seven-fold, suggesting the majority of infected ticks carry relatively low numbers of the pathogen. An effort was made to determine if an artificial blood meal would stimulate replication of the bacterial pathogen, thereby increasing the frequency in which aoEBA could be identified; no statistically significant effect was evident. The number of ticks determined to be carrying aoEBA varied with geographic location and ranged from 5 to 20%. aoEBA was found in both adults (12% of the males and 12% of the females) and nymphs (13%) but not larvae. Comparative analysis of dissected ticks provided strong evidence that the salivary gland was the most common location of aoEBA in field-collected ticks. No significant correlations were identified between the frequency of infection and tick weight, suggesting that increasing tick age and increased number of blood meals did not increase infectivity.     

2012年全国家畜血吸虫病疫情状况

本文通报了2012年全国家畜血吸虫病疫情状况。在2012年,湖南省、湖北省、江西省、四川省、安徽省、云南省、江苏省等7省共有存栏牛1 033 056头,存栏羊2 024 512只,其他家畜存栏数891 301头（匹、只）。2012年共检查了684 899头牛,其中牛血吸虫病阳性3961头,阳性率0.58%,比2011年的阳性数下降了42.57%;检查了71 473只羊,其中羊血吸虫病阳性406只,阳性率0.57%,比2011年的阳性数下降了4.19%,并对142 976头其他家畜进行了检查,其中血吸虫病阳性为21头,阳性率0.01%。湖南省和江西省的病牛数占到了全国病牛数的80.06%。2012年,全国家畜血吸虫病疫情较之2011年继续下降,但是洞庭湖和鄱阳湖地区依然是今后家畜血吸虫病疫情控制的难点和重点。     

蜱携带牛丙型肝炎病毒新亚型巢式PCR检测方法的建立

旨在提高新发现的牛丙型肝炎病毒毒株——广东湛江毒株(bovine hepacivirus-GDZJ,BovHepV-GDZJ)的检测及诊断能力,初步了解广东地区蜱携带BovHepV-GDZJ的情况,本研究结合宏转录组学、高通量测序及PCR技术,针对高通量测序结果中丰度高的基因区域,设计该病毒的巢式PCR特异性引物.通过...     

Serosurvey and molecular detection of Crimean–Congo hemorrhagic fever virus (CCHFV) in northern Turkey

In this study, the hard ticks, whole blood and serum samples collected from small ruminants (sheep and goat) in middle Black Sea region of Turkey where Crimean-Congo hemorrhagic fever (CCHF) human cases were observed in the past years were surveyed for the presence of RNA and specific IgG antibodies from CCFH virus (CCHFV). CCHFV RNA was found in 30 of 255 tick pools (11.76%) and nine of 105 (8.57%) leucocyte samples. No CCHFV genomic RNA was detected from animals in Yildizeli and Vezirkopru. However, CCHFV RNA was found from animals in Gerze and Resadiye. Seventy-eight of 105 goat and sheep blood serum samples tested were antibody-positive for CCHFV by enzyme-linked immunosorbent assay (ELISA) (goat: 42/63; sheep: 36/42). Viral RNA was detected from tick samples in all of four provinces. Positivity rates for the provinces varied and were as follows: Gerze 13.04%, Resadiye 35.41%, Vezirkopru 1.61% and Yildizeli 6.06%. CCHFV genomic RNA was detected in four of seven tick species tested. These results suggest that these hard ticks may act as a reservoir for CCHFV in northern Turkey.     

四川省松潘县牦牛体表蜱、高原鼠兔携带巴尔通体和无形体的PCR检测与进化分析

旨在了解四川省松潘县牦牛体表蜱、高原鼠兔巴尔通体和无形体感染情况。采集牦牛体表的蜱和捕获高原鼠兔，对蜱进行形态学初步鉴定后，提取蜱和高原鼠兔脾总DNA，PCR扩增蜱16S rRNA、巴尔通体rpoB和无形体16S rRNA基因，对PCR产物阳性产物测序、比对及构建系统进化树，从而确定蜱种类及蜱和高原鼠兔感染巴尔通体和无形体的种类及感染率。结果显示：在松潘县进安乡、山巴乡、下八寨乡各采集到蜱102、97和131只，共计330只，经鉴定均为青海血蜱。蜱巴尔通体仅检出1种巴尔通体，与B.melophagi亲缘关系最近，进安乡、山巴乡和下八寨乡检出率分别为16.7%、8.2%和18.3%，其中下八寨乡检出率显著高于进安乡（P<0.05）；蜱源无形体进安乡、山巴乡和下八寨乡检出率分别为9.8%、12.4%和26.7%，下八寨乡检出率显著高于进安乡（P<0.01），检出的无形体均为1种，与牛无形体（A.bovis）亲缘关系最近；下八寨乡检出的鼠兔源巴尔通体与B.queenslandens亲缘关系最近，感染率为6.7%；进安乡、山巴乡和下八寨乡检出的鼠兔源巴尔通体与B.grahamii亲缘关系最近，感染率分别为8.7%、17.9%和13.3%，3个点检出率无显著差异；未定种Bartonella sp.（MN296294）和Bartonella sp.（MN296293）仅分别在进安乡和下巴乡检出；与蜱均检出无形体不同，高原鼠兔均未检出无形体。此外，蜱和高原鼠兔均未发现2种及2种以上病原混合感染。松潘县青海血蜱携带巴尔通体和无形体，高原鼠兔感染巴尔通体，且首次在高原鼠兔体内检测到疑似B.queenslandens的病原体，提示当地居民有感染这两类病原风险。     

多重PCR检测圈养牛、猪和羊源魏氏梭菌

采用多重PCR对圈养源魏氏梭菌的α，β，ε，ι毒素基因进行了检测，结果证实该方法具有很高的特异性。通过对山东德州、枣庄、泰安、蒙阴曾经流行过魏氏梭菌病的猪场、牛场和羊场的162个粪便样品进行检测，检出率为19．1％，均为A型。应用该方法鉴别魏氏梭菌血清型快速、简便，结果对于预防治疗均具有重要的指导作用。     

An antigenic investigation of Crimean-Congo hemorrhagic fever virus (CCHFV) in hard ticks from provinces in northern Turkey

Crimean-Congo hemorrhagic fever virus threatens human health. Exposure of the infected tick is a strong risk factor for human disease. In this study, the hard ticks collected from a variety of mammalian species (cattle, sheep, goat, and buffalo) and a turtle in either coastal or inland Black Sea region of Turkey were surveyed for the presence of antigen from Crimean-Congo hemorrhagic fever virus (CCHFV). CCHFV antigen was found in 46 of 421 tick pools (10.92%). Positivity rates for the provinces varied and were as follows: Samsun 33.87%, Ordu 4.34%, Giresun 8.86%, Sinop 6.09%, Amasya 7.40%, Tokat 5.08%, and Sivas 8.06%. CCHFV antigen was detected in seven of 11 tick species tested. These results suggest that these hard ticks may act as a reservoir for CCHFV in northern Turkey.     

石河子地区宠物犬蜱的种类鉴定及其携带布鲁氏菌的检测

为了解新疆石河子地区宠物犬体表寄生蜱的种类及蜱携带布鲁氏菌情况,本试验在形态学分类的基础上,基于线粒体基因12S rRNA和细胞色素氧化酶Ⅰ（COⅠ）对宠物犬体表采集寄生蜱进行分子生物学检测,使用DNAMAN软件比对分析序列同源性,并应用序列分析软件Mega 7.0邻接法构建蜱种系统发育进化树,分析蜱种的遗传进化关系;基于布鲁氏菌外膜蛋白Omp22基因对宠物犬体表寄生蜱进行布鲁氏菌PCR检测,确定宠物犬蜱布鲁氏菌携带情况。结果显示,宠物犬体表寄生的436只蜱的形态学与线粒体基因（12S rRNA和COⅠ）分子生物学鉴定结果一致,均为新疆优势蜱种之一——图兰扇头蜱（Rhipicephalus turanicus）。基于12S rRNA基因构建的蜱种系统发育树显示,本试验所得宠物犬体表寄生图兰扇头蜱序列与GenBank中已知图兰扇头蜱的序列聚为一大支。基于布鲁氏菌Omp22基因的巢式PCR扩增结果显示,宠物犬体表寄生图兰扇头蜱携带布鲁氏菌的阳性率为4.82%（21/436）,且同源性为100%。BLAST比对显示,本试验所得宠物犬图兰扇头蜱携带的布鲁氏菌与新疆流行株YC31（GenBank登录号：MK201679.1）、QH5（GenBank登录号：MK201678.1）、EM3（GenBank登录号：MK201677.1）和ML9（GenBank登录号：MK201676.1）的同源性均为100%。本研究通过形态学及分子生物学探讨了新疆石河子地区宠物犬体表寄生蜱的种类及携带布鲁氏菌基本情况,为宠物犬体表寄生蜱的种类及蜱传疾病的监测和控制等研究工作提供了基础资料。     

Tick infestations on livestock in the yemen arab republic and their potential as vectors of livestock diseases

A survey of ticks on cattle, camels, sheep, goats and donkeys in four different geographical locations of the Yemen Arab Republic (YAR) was carried out to provide more information on the possible risk of tick-borne diseases to imported exotic cattle included in the YAR's plans for livestock development. The most abundant ticks were Hyalomma spp. particularly on camels. Ticks found on cattle included Hyalomma spp., Amblyomma variegatum, Boophilus annulatus and Rhipicephalus spp. In general with the exception of camels tick burdens on all species of domestic livestock were very low. Two hundred and ninety eight serum samples from miscellaneous adult cattle throughout the country were negative to a test for Anaplasma marginale antibodies. It is speculated that tick burdens in the YAR are too low for significant disease transmission and the implications of the findings are discussed.     

Evaluation of a commercially available enzyme-linked immunosorbent assay for detecting antibodies to Fasciola hepatica and Fasciola gigantica in cattle, sheep and buffaloes in Australia

A commercially available ELISA for detecting antibodies to liver fluke was evaluated for use in Australia. Milk and serum samples from cattle and sheep in which infection with Fasciola hepatica was confirmed by detection of eggs in faeces were used to estimate sensitivity. Similar samples collected from cattle and sheep outside the F. hepatica-endemic area were used to estimate specificity. The ELISA was also evaluated for detecting antibodies to F. hepatica in milk from sheep and antibodies to Fasciola gigantica in sera from cattle and buffaloes, but with small numbers of samples. In cattle, the sensitivity and specificity of the ELISA were 98.2% and 98.3% using serum and 97.7% and 99.3% using milk. In infected herds, 41.4% and 41.5% of animals were positive in the serum and milk ELISAs, respectively, whereas F. hepatica eggs were found in faecal samples from 26.5% of animals. In sheep, the sensitivity of the ELISA was 96.9% and the specificity was 99.4%. In infected flocks, 60.2% of animals were positive in the serum ELISA and F. hepatica eggs were found in faecal samples 52.2% of animals. There was perfect agreement in the ELISA between paired serum and milk samples collected from ewes. The assay detected antibodies in sera from cattle and buffaloes with natural and experimental F. gigantica infections. In the experimentally infected animals, antibodies were detected 2 weeks post-infection. We conclude that the ELISA will be a valuable tool for diagnosing F. hepatica infections in cattle and sheep. The assay may also be useful for diagnosing F. gigantica infections but further studies are required to establish sensitivity and specificity.     

Prevalence and potential risk factors for amitraz resistance in Boophilus microplus ticks in cattle farms in the State of Yucatan, Mexico

Southern cattle tick, Boophilus microplus, control in the Mexican tropics is based on the application by spray, dip or pour-on of acaricides. In Mexico the first report of ticks resistant to amitraz was reported in 2002. Amitraz resistance affecting the cattle industry of Yucatan State, Mexico, has not been investigated. A cross-sectional study was carried out in order to investigate the possible association between management risk factors and the prevalence of B. microplus ticks resistant to amitraz in Yucatan, Mexico. Ninety-eight cattle farms were randomly selected and a sample of 20–30 engorged B. microplus females was collected from at least 10 cattle from each farm. Collected ticks were taken to the laboratory where the larval progeny were obtained. The modified larval immersion test was used to detect ticks resistant to amitraz. A discriminating dose of 0.0002% amitraz was used. A questionnaire was applied on each collaborating farm. For the statistical analysis a 2 × 2 contingency table was used to identify potential risk factors. The prevalence of farms with B. microplus resistant to amitraz was 19.40%. However, even though tick resistance to amitraz is a present problem there was no evidence that it was associated with the risk factors analyzed in this survey.     

Semi-nested PCR detection of Babesia orientalis in its natural hosts Rhipicephalus haemaphysaloides and buffalo

Babesiosis has recently been recognized as an emerging infectious disease of buffalo in China. In order to investigate the epidemiology and enzootic potential of this parasite in Hubei province, we sought to develop a semi-nested PCR to detect Babesia orientalis in buffalo and the potential tick vector-Rhipicephalus haemaphysaloides by amplifying a specific 257bp fragment of B. orientalis 18S rRNA gene. The practical limit of detection showed that it had high sensitivity and an approximate parasitemia of 0.00000012% was detected by the PCR system. The blood samples of 121 asymptomatic buffaloes collected from four babesia endemic counties and that of 71 asymptomatic buffaloes collected from three babesia free counties in Hubei province of China were examined for the presence of B. orientalis using both Wright-Giemsa stained blood smear and semi-nested PCR. Microscopic examination revealed that 5/121 animals were positive, whereas 24/121 animals were positive by the semi-nested PCR assay. Of 378 ticks (R. haemaphysaloides) collected from buffaloes and examined by the semi-nested PCR, 35 were positive. The results showed that the semi-nested PCR was a useful method to investigate the epidemiology of buffalo babesiosis (B. orientalis), which is widely distributed in Hubei province, China.     

The occurrence of Coxiella burnetii in sheep and ticks of the genus Dermacentor in Baden-Wuerttemberg

This study examined the occurrence of Coxiella burnetii (C. burnetii), the infectious agent of Q-fever, in sheep and sheep-ticks in Baden-Wuerttemberg, Germany, as a possible source of infection in Q-fever outbreaks. Using PCR, we examined a total of 1066 Dermacentor ticks from 23 herds and 49 samples of tick excrement from 18 herds for C. burnetii. We found the infectious agent in one non-engorged tick and in one sample of tick excrement from the same herd, in Efringen-Kirchen (district Loerrach). Sequencing the PCR-products confirmed the amplifications as specific for C. burnetii. Further serological tests of random samples of the four districts of Baden-Wuerttemberg showed a seroprevalence from 0 to 1.4% using complement fixation test (CFT), as well as a 0.9 to 10.2% seroprevalence, using ELISA test. Serum samples from a Q-fever-suspicious herd resulted, however, in 6% (CFT) and 53% (ELISA) positive reactions. A comparison between CFT and ELISA showed both a correlation of the two test methods that increased with higher CFT titration levels and positive reactions using ELISA for 9.4% of the serums that had tested negative using CFT. The results of the present study reveal that ticks and their excrements are important vectors of transmission of Q-fever in Baden-Wuerttemberg. Investigations on C. burnetii using PCR as well as serological surveys of sheep are important instruments for diagnosis and disease control of Q-fever.     

Ticks on livestock in Barbados

Cattle, sheep, goats, and horses in most areas of Barbados were examined for ticks. About 68% of the cattle, mostly Holstein dairy cows, and 23% of the sheep, mostly purebred or crossbred black belly sheep, were infested with the southern cattle tick, Boophilus microplus (Canestrini), a vector of bovine babesiosis and anaplasmosis. About 13% of the horses were infested with the tropical horse tick, Anocentor nitens (Neumann), a vector of equine babesiosis. Cattle owners used a variety of acaricides as dusts, sprays, or washes on their cattle for the control of ticks.     

甘肃省牛蜱传性血液原虫生物学特性研究及病原的分离

用病原检查法对甘肃省４２个县市的１３１１头牛进行了牛蜱传性血液原虫病的大面积调查,证实该省牛蜱传染性血液原虫的种类为双芽巴贝斯虫,大巴贝斯虫,瑟氏泰勒虫,环形泰勒虫及国边虫等５种病原,     

Phylogenetic analysis of benign Theileria species based on major piroplasm surface protein (MPSP) genes from ticks of grazing cattle in Korea

Complete major piroplasm surface protein (MPSP) gene sequences of benign Theileria parasites were isolated from ticks of grazing cattle in Korea. A total of 556 tick samples were collected in five provinces: Chungbuk, Jeonbuk, Jeonnam, Gyeongbuk, and Jeju during 2010-2011. Fifteen samples from Chungbuk and Jeonnam were positive for the Theileria MPSP gene by PCR amplification using a specific primer set. A phylogenetic tree was constructed with the amplified gene sequences and 26 additional sequences published in GenBank. The benign Theileria parasites were classified into eight types, those isolated from Korean cattle ticks belonged to Types 1 (Ikeda), 2 (Chitose), 4, and 8. Types 2 and 4 were the most common types, with the rate of 40%, followed by Types 1 and 8 (with the rate of 13% and 7%, respectively). Nucleotide sequence identities of 23 theilerial MPSP sequences (15 MPSP gene sequences amplified and 8 sequences published) ranged from 67.3 to 99.8%. Multiple alignments of the deduced amino acid sequences also showed that each type was characterized by specific amino acids: 7 for Type 1, 9 for Type 2, 4 for Type 4, and 3 for Type 8.