新疆沙雅县图兰扇头蜱的形态学及分子生物学鉴定

为了掌握新疆边境地区沙雅县的蜱种分布状况,从该地绵羊体表采集寄生蜱虫,应用数码体视显微镜对其进行形态学初步鉴定,采用分子生物学技术对采集蜱虫的线粒体16S rDNA进行基因序列扩增并测序,应用序列分析软件MEGA 6.0构建系统发育进化树,分析采集蜱样的分子进化特征。结果表明,采集的蜱样均为图兰扇头蜱,在系统发育上依然和新疆境内石河子地区、乌什地区、皮山地区和图木舒克地区的图兰扇头蜱处于同一分支,具有分子进化的相对保守性。     

新疆南疆部分地区蜱种鉴定及斑点热群立克次体流行病学调查

为了解新疆南疆部分地区不同蜱种携带斑点热群立克次体(spotted fever group Rickettsia,SFGR)情况,采用PCR技术扩增16S rDNA基因和SFGR外膜蛋白A(OmpA)基因,进行蜱种鉴定及SFGR检测。对测序阳性样本进行BLAST序列分析,应用DNAStar和Mega 6.0软件构建遗传进化树。结果显示:从新疆南疆9个县市共采集蜱1 340只,其种类包括3属4种,分别为图兰扇头蜱、边缘革蜱、银盾革蜱、亚洲璃眼蜱;在随机挑选的90份蜱DNA样本中共检测出57份SFGR阳性样本,其中阿拉尔市十一团、七连地区的阳性样本均来自犬体表寄生蜱,其余地区的阳性样本均来自绵羊体表寄生蜱。序列分析及遗传进化树显示,此次检测的亚洲璃眼蜱、边缘革蜱携带的SFGR与GenBank公布的SFGR疆内株Candidatus R.barbariae(登录号为MG668832.1)亲缘关系最近,处于同一分支;图兰扇头蜱携带的SFGR也同疆内株R.massiliae、R.conorii、Candidatus R.barbariae(MG668831.1、KP214024.1、MG668832.1)亲缘关系最近,处于同一分支;而银盾革蜱携带的SFGR与国外株R.raoultii(KT805295.1)亲缘关系最近,处于同一分支,并首次于新疆南疆绵羊寄生银盾革蜱中检出饶氏立克次体。本试验为该地区蜱传立克次体病的研究及防控提供重要依据。     

小亚璃眼蜱、亚洲璃眼蜱和残缘璃眼蜱的分子生物学鉴定

本研究旨在建立小亚璃眼蜱、亚洲璃眼蜱和残缘璃眼蜱的分子生物学鉴定方法,并探讨它们的系统发生关系。在新疆、内蒙古从动物体表采集寄生蜱,形态学鉴定后,PCR扩增测序获得3种璃眼蜱的16S rRNA及线粒体色素氧化酶亚基Ⅰ基因(cytochrome oxidaseⅠ,COⅠ)序列后进行同源性分析。用Mega 5.0和Mrbayes 3.2软件分别构建系统进化树,亚洲璃眼蜱、小亚璃眼蜱样本16S rRNA序列与GenBank中已知相应蜱虫16S rRNA序列聚类,而残缘璃眼蜱COⅠ序列与GenBank中已知残缘璃眼蜱COⅠ序列聚类,与形态学鉴定结果一致。在传统形态学分类的基础上,结合分子生物学鉴定方法能简易、准确鉴定小亚璃眼蜱、亚洲璃眼蜱和残缘璃眼蜱。     

乌鲁木齐市犬体表寄生蜱形态学及分子生物学鉴定

在乌鲁木齐市多家宠物医院、宠物店、犬舍、流浪犬基地、社区随机选取无明显临床症状的犬共1 054只,采集其体表蜱虫889只,通过形态学鉴定及蜱虫的线粒体16S rDNA基因扩增序列进行分子生物学鉴定.鉴定得到假头基呈六角形的蜱虫,所采集的蜱虫初步鉴定为血红扇头蜱和图兰扇头蜱.发现血红扇头蜱和图兰扇头蜱的16S rDNA基...     

草原革蜱和边缘革蜱的分子生物学鉴定

本研究旨在建立草原革蜱和边缘革蜱的分子生物学鉴定方法,并探讨其系统发生关系。在新疆从动物体表采集寄生蜱,形态学鉴定后,PCR扩增获得2种革蜱的16S rRNA及线粒体色素氧化酶亚基Ⅰ(COⅠ)基因序列,测序后进行同源性分析。用Mega 5.0和Mrbayes 3.2软件分别构建系统进化树,草原革蜱16S rRNA序列与GenBank数据库中已知草原革蜱16S rRNA序列聚类,而边缘革蜱COⅠ序列与GenBank数据库中已知边缘革蜱COⅠ序列聚类,与形态学鉴定结果一致。本研究结果表明,在传统形态学分类的基础上,结合分子生物学鉴定方法能简易、准确鉴定草原革蜱和边缘革蜱。     

牛蜱龙岩分离株的分子鉴定

为了解福建省龙岩地区牛场体表蜱的种类,试验采集龙岩地区牛体表寄生蜱样本,基于蜱的核糖体rDNA内转录间隔区(internal transcribed spacer,ITS)序列PCR扩增方法进行分子鉴定,获得ITS1和ITS2基因序列,进行blastn相似性搜索,应用MegAlign和MEGA 7.0软件完成同源性分析及种系发育分析。结果表明,该蜱ITS1序列与中国甘肃微小扇头蜱分离株同源性达99.32%(JQ737124.1、JQ737125.1),与中国湖南微小扇头蜱HAI2分离株同源性达99.16%(MK224531.1);ITS2序列与中国河南、南非豪登微小扇头蜱分离株同源性均为100%(KX450287.1、KY457506.1);种系发育分析结果显示,龙岩地区牛体表分离的牛蜱ITS基因序列与扇头蜱属ITS基因序列聚类,与革蜱属和璃眼蜱属处于两个不同的分支。说明福建龙岩牛蜱分离株为微小扇头蜱。     

我国中东部地区宠物犬蜱种类及其分布调查

为了解我国宠物犬的蜱种类及分布情况,为防治提供基础资料,本研究采集了我国北京、上海、广州等20个城市122家宠物医院犬体表寄生的蜱1 550只,进行犬体寄生蜱的种类鉴定。鉴定结果表明:血红扇头蜱、长角血蜱及镰形扇头蜱是我国宠物犬主要寄生蜱类,不同发育阶段的蜱均可犬体寄生,而成蜱是主要的寄生阶段。调查结果提示,我国宠物犬蜱分布广,种类较多,由于蜱虫同样可感染人,而且携带大量蜱媒病,应引起临床兽医和公共卫生部门的关注。     

伊犁河谷区域银盾革蜱的鉴定及其携带马泰勒虫病原DNA检测

为了解伊犁河谷区域银盾革蜱的分布及其携带马泰勒虫病原情况,通过对银盾革蜱形态学及革蜱属特异性基因(ITS-2)和其携带病原18S rRNA基因PCR扩增及其PCR产物的克隆、测序、同源性比较等,鉴定和检测了采自昭苏种马场马体表银盾革蜱及其携带病原情况。试验结果显示,采回的蜱经形态学鉴定均为银盾革蜱,PCR扩增银盾革蜱ITS-2基因和马泰勒虫18S rRNA基因序列大小分别为825 bp和789 bp。本试验蜱获得的ITS-2序列与已发表的伊朗株(GQ144706.1)同源性为99%,与形态学鉴定结果一致,故鉴定为银盾革蜱。在181只银盾革蜱中携带马泰勒虫的有13只,阳性率为7.1%(13/181),且其携带病原目的基因序列与Gen Bank中已发表的马泰勒虫基因序列相似性为99%。研究表明,结合形态学分类和分子生物学方法对银盾革蜱鉴定具有简易、准确度高等特点,且分离银盾革蜱源性病原更有实际意义,为综合防控奠定了基础。     

新疆和田地区蜱种分布及蜱传立克次体分子流行病学调查

为了解新疆南部和田地区蜱种分布及蜱携带立克次体流行情况，本试验从和田市、和田县、墨玉县、皮山县、洛浦县、策勒县、于田县和民丰县8个县市的养殖绵羊采集体表寄生蜱，运用形态学观察和PCR检测技术对蜱种进行鉴定；运用PCR检测技术对立克次体进行检测；通过系统发育分析方法探究和田地区蜱携带立克次体的种间关系。结果显示，本试验共采集蜱914只，经鉴定包括图兰扇头蜱(Rhipicephalus turanicus)532只、亚洲璃眼蜱(Hyalomma asiaticum)304只、草原革蜱(Dermacentor nuttalli)78只。PCR检测发现，该地区蜱中存在暂定巴布瑞立克次体(Candidatus Rickettsia barbariae)、马赛立克次体(Rickettsia massiliae)、饶氏立克次体(Rickettsia raoultii),蜱传立克次体总体阳性率为59.13%。系统发育分析发现，本试验获得的立克次体均与已知立克次体同源。结果表明，和田地区的优势蜱种为图兰扇头蜱(Rhipicephalus turanicus),该地区的立克次体阳性率较高并且近年来呈上升态...     

上海市闵行区宠物犬体表蜱及蜱传病原体初步调查

为调查上海市闵行区宠物犬媒介蜱及蜱传病原体情况,闵行区宠物医院就诊犬2763只,其中57只犬体表检获183只,根据形态学鉴定蜱种类;采用PCR方法筛查采集的蜱类样本和保留的392份犬血液样本,以发现可能的蜱传病原体。研究结果表明,宠物犬带蜱率为2.06%,带蜱指数为0.066,犬体表寄生的蜱宠物种为血红扇头蜱(Rhipicephalus sanguineus,R.sanguineus);病原学检测发现宠物犬携带犬埃立克体(Ehrlichia canis,E.canis),其在蜱和血液样品检出率分别为0.5%和2.29%,其他病原体如螺旋体(Borrelia.spp)和嗜吞噬细胞无浆体(Anaplasma phagocytophilum,A.phagocytophilum)均未有检出。调查结果提示我们闵行区部分宠物犬体表存在血红扇头蜱和蜱传病原体感染,应加强媒介蜱及蜱传病原体的监测与防控。     

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.     

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。     

Characterization of microbiota diversity of engorged ticks collected from dogs in China

BackgroundTicks are one of the most common external parasites in dogs, and are associated with the transmission of a number of major zoonoses, which result in serious harm to human health and even death. Also, the increasing number of pet dogs and pet owners in China has caused concern regarding human tick-borne illnesses. Accordingly, studies are needed to gain a complete understanding of the bacterial composition and diversity of the ticks that parasitize dogs.ObjectivesTo date, there have been relatively few reports on the analysis of the bacterial community structure and diversity in ticks that parasitize dogs. The objective of this study was to investigate the microbial composition and diversity of parasitic ticks of dogs, and assessed the effect of tick sex and geographical region on the bacterial composition in two tick genera collected from dogs in China.MethodsA total of 178 whole ticks were subjected to a 16S ribosomal RNA (rRNA) next generation sequencing analysis. The Illumina MiSeq platform targeting the V3–V4 region of the 16S rRNA gene was used to characterize the bacterial communities of the collected ticks. Sequence analysis and taxonomic assignment were performed using QIIME 2 and the GreenGene database, respectively. After clustering the sequences into taxonomic units, the sequences were quality-filtered and rarefied.ResultsAfter pooling 24 tick samples, we identified a total of 2,081 operational taxonomic units, which were assigned to 23 phyla and 328 genera, revealing a diverse bacterial community profile. The high, moderate and low prevalent taxa include 46, 101, and 182 genera, respectively. Among them, dominant taxa include environmental bacterial genera, such as Psychrobacter and Burkholderia. Additionally, some known tick-associated endosymbionts were also detected, including Coxiella, Rickettsia, and Ricketssiella. Also, the potentially pathogenic genera Staphylococcus and Pseudomonas were detected in the tick pools. Moreover, our preliminary study found that the differences in microbial communities are more dependent on the sampling location than tick sex in the tick specimens collected from dogs.ConclusionsThe findings of this study support the need for future research on the microbial population present in ticks collected from dogs in China.     

Biology and control of ticks infesting dogs and cats in North America

At least 15 different infectious agents and diseases are transmitted or produced by ticks parasitizing dogs and cats in North America. Those infectious agents and diseases are distributed among eight different tick species that commonly infest dogs and cats. All but one of these species are hard ticks with a three-host development cycle in which each motile stage (larva, nymph, and adult) feeds on a different host after molting. Tick species, disease occurrence, and peak activity of each tick life stage can vary dramatically depending on geographic and climatic conditions. Gaining an understanding of tick distribution, tick ecology, and seasonal occurrence of different tick life stages can help with the management of tick infestations and reduce the incidence of tick-transmitted diseases in dogs and cats. Control should be based on an understanding and management of ecologic factors responsible for tick infestations and selection of appropriate acaricides. Occasionally, topical acaricides will appear not to have prevented tick infestations. This lack of control may be real or perceived based on reinfestation rates and/or pet owner expectations of product performance.     

1株罗曼粉鸡源申氏不动杆菌的分离鉴定及耐药性分析

为了解贵州省某养殖场罗曼粉鸡发病病原的特征及其耐药性,本试验从病死鸡中分离出一株革兰氏阴性菌,命名为GZ2019,并对其进行纯化培养、生化试验、16S rRNA序列分析、药敏试验及动物回归试验。分离菌在普通琼脂培养基上呈圆形凸起、表面光滑、半透明的灰白色小菌落,鲜血营养琼脂培养基上菌落形态与普通琼脂培养基上的一致,但不溶血;革兰氏染色结果镜检显示,分离菌为成双排列的阴性球杆菌,偶见单个存在或链状排列;生化试验结果显示,分离菌枸橼酸盐利用、过氧化氢酶反应为阳性,硝酸还原、氧化酶、葡萄糖发酵等反应为阴性,无运动性;16S rRNA序列分析结果显示,分离株与不动杆菌的核苷酸同源性在72.6%~99.9%之间,与申氏不动杆菌LUH 4760株（GenBank登录号：AJ275041）、SNSK 752株（GenBank登录号：MG584984）、MCDA01株（GenBank登录号：KY385627）及RP1株（GenBank登录号：MG461636）的核苷酸同源性高达99.9%;药敏试验结果显示,分离菌对头孢曲松和痢特灵敏感,对头孢哌酮、头孢呋辛、头孢他啶等5种药物中度敏感,对新霉素、环丙沙星、羧苄西林等17种药物耐药;动物回归试验结果显示,试验组小鼠隔离饲养1周仅死亡1只,死亡率为20%,表明该分离菌致病性较弱。本试验成功分离出1株低致病性申氏不动杆菌GZ2019,可为鸡源申氏不动杆菌病的预防和治疗提供一定的参考依据。     

Efficacy of a combination of imidacloprid 10%/permethrin 50% versus fipronil 10%/(S)-methoprene 12%, against ticks in naturally infected dogs

Preventing tick bites is a fundamental step towards reducing the impact of tick-borne protozoal, bacterial and viral diseases (TBDs) in humans and animals. The aim of this study was to evaluate the efficacy of a combination of imidacloprid 10%/permethrin 50% and of fipronil 10%/S-methoprene 12% against ticks in naturally infected dogs and to assess methodological parameters to calculate drug efficacy on tick immature stages.

From July to August 2004, 45 privately owned dogs of various sexes, ages, breeds, coat length and habits were enrolled in a trial carried out in an area (radius approximately 50 km) in Southern Italy. Three homogeneous groups (both for dog population and tick population) were formed: 15 dogs treated with imidacloprid 10% and permethrin 50% spot-on (group A), 15 dogs treated with fipronil 10% and methoprene 12% spot-on (group B) and 15 untreated dogs (group C). The dogs in each group were then sub-grouped according to their age and weight. Two different treatments were administered (time 0 and +28 days) to groups A and B, and the dogs were checked weekly for tick infestation until day +56 post-treatment (p.t.). Twenty-four areas distributed on the whole body surface were examined for ticks at each follow-up, while only at time 0 and at day +56 p.t., ticks were collected from the dogs and identified.

For the immature stages a semi-quantitative method was adopted and the load of immature stages was evaluated and grouped into four classes up to day +56 p.t. when the mean number of immature ticks (MIT) for each infection class was evaluated.

All the adult ticks collected were identified as brown dog ticks (Rhipicephalus sanguineus). Immature stages were first compared at day +28 p.t.. The efficacy of both products used in groups A and B on adult ticks was high and generally very similar. Conversely, the efficacy of imidacloprid 10% and permethrin 50% against immatures was higher than that of fipronil 10% and methoprene 12% throughout the observation period with statistically significant differences (p < 0.05) at day +28 p.t. (i.e. group A = 98.52%, group B = 72.40%).

On the whole, in analysing the efficacy of both products against adult plus immature ticks, it was found that the combination of imidacloprid 10% and permethrin 50% was more effective than fipronil 10% and methoprene 12%, with the differences being statistically significant at day +28 p.t. (group A = 98.43%, group B = 77.56%).     

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。     

Epidemiological Investigation of Francisella tularensis Transmitted by Rabbits and Ticks in Some Provinces of China

The aim of this investigation was to find out the prevalence of Francesella tularensis in some provinces of China. Using the general and subspecies PCR detection method for Francesella tularensis established in our laboratory, PCR detection of Francesella tularensis in rabbits and ticks was carried out in provinces with high-density rabbits feeding and some provinces and cities with sheep and cattle stock. The results showed that DNA detection of Francesella tularensis from rabbit tissues and ticks carried by cattle and sheep was positive. 12 out of 218 rabbit samples were positive (5.5% positive rate), and 15 tick samples were positive (3.1%) of 490 tick samples. In terms of geographical distribution, most of the positive rabbit samples were from Shandong, Henan and Sichuan province, however, ticks collected from Yunnan and Shandong province showed a higher positive rate. PCR detection showed that Francesella tularensis subspecies in this investigation was F.h, which was a subspecies with strong toxicity. This investigation revealed that Francesella tularensis presented in rabbits and ticks, public health and safety risks of Francesella tularensis existed in Shandong and Yunnan province, which should be paid more attention by the relevant scientific research institutions, medical institutions and government departments.     

Brucella proteomes--a review

The proteomes of selected Brucella spp. have been extensively analyzed by utilizing current proteomic technology involving 2-DE and MALDI-MS. In Brucella melitensis, more than 500 proteins were identified. The rapid and large-scale identification of proteins in this organism was accomplished by using the annotated B. melitensis genome which is now available in the GenBank. Coupled with new and powerful tools for data analysis, differentially expressed proteins were identified and categorized into several classes. A global overview of protein expression patterns emerged, thereby facilitating the simultaneous analysis of different metabolic pathways in B. melitensis. Such a global characterization would not have been possible by using time consuming and traditional biochemical approaches. The era of post-genomic technology offers new and exciting opportunities to understand the complete biology of different Brucella species.