吉林省珲春地区蜱中伯氏疏螺旋体与斑点热群立克次体复合感染研究

目的了解吉林省珲春地区伯氏疏螺旋体与斑点热群立克次体的复合感染情况.方法运用PCR方法对吉林省珲春地区采集的蜱标本,进行伯氏疏螺旋体5S～23S rRNA间隔区基因与斑点热群立克次体外膜蛋白A（ompA）基因的检测.测序并用PHYLIP软件进行序列分析.结果全沟硬蜱中伯氏疏螺旋体感染率为36.0%,在全沟硬蜱中检测到了斑点热群立克次体的感染,其感染率为2 0%.二者的复合感染率为2.0%;森林革蜱中伯氏疏螺旋体感染率30.9%,斑点热群立克次体感染率29.1%,二者的复合感染率16.8%.伯氏疏螺旋体的序列分析显示吉林地区的伯氏疏螺旋体都属于B.garinii基因型,同源性较高.对斑点热阳性片段序列分析表明新测序列与斯洛伐克新发现的IRS3株和IRS4株核苷酸序列同源性为97%.结论吉林省珲春地区全沟硬蜱及森林革蜱中检测到伯氏疏螺旋体与斑点热群立克次体的感染,并检测到2种病原体的复合感染情况.     

黑龙江省旅游区蜱伯氏疏螺旋体和斑点热群立克次体复合感染的动态调查

目的 了解黑龙江省旅游区蜱伯氏疏螺旋体和斑点热群立克次体复合感染的动态变化.方法 运用PCR方法检测2010年捕获蜱中两种病原感染状况并测序进行序列分析.结果 在威虎山和镜泊湖景区采集点捕获蜱标本共849只.两地优势蜱种分别为全沟硬蜱和嗜群血蜱,其中伯氏疏螺旋体和斑点热群立克次体感染率分别为26.15％、10.05％和0.00％、13.33％,提示全沟硬蜱复合感染.两地优势蜱种活动高峰分别出现在7月和6月上旬,蜱携带病原率因时间差异而不同.序列分析显示旅游区伯氏疏螺旋体有3种基因型,除B.garinii型、B.afzelii型外,首次在东北地区发现B.valaisiana-like group型.对斑点热群立克次体序列分析表明,所得两序列分别与Rickettsia sp.HL-93和Rickettsia sp.H820,其同源性为100％.结论 两旅游区蜱中存在伯氏疏螺旋体和斑点热群立克次体复合感染,且携带的基因类型存在多样性;动态调查显示蜱种、蜱密度的分布以及蜱携带病原状况因生境、时间和人群的不同而有差异.     

牡丹江镜泊湖区蜱中伯氏疏螺旋体与嗜吞噬细胞无形体感染的研究

目的了解牡丹江镜泊湖区伯氏疏螺旋体和嗜吞噬细胞无形体的感染情况。方法运用聚合酶链方法(PCR)对牡丹江镜泊湖区采集的蜱标本,进行伯氏疏螺旋体5S~23S基因间隔区基因片段与嗜吞噬细胞无形体柠檬酸盐合成酶基因(glt A)片段的检测。结果 2013年5月,在牡丹江镜泊湖北湖区附近共采集嗜群血蜱和全沟硬蜱643只,其中嗜群血蜱573只,占89.11%;全沟硬蜱70只,占10.89%。嗜群血蜱和全沟硬蜱伯氏疏螺旋体感染率分别为0.17%和15.71%,嗜吞噬细胞无形体感染率分别为0和2.86%;全沟硬蜱中伯氏疏螺旋体和嗜吞噬细胞无形体的复合感染率为1.43%。结论牡丹江镜泊湖区采集的蜱中检测到伯氏疏螺旋体和嗜吞噬细胞无形体的感染,并在全沟硬蜱中检测到复合感染情况,但复合感染的感染率较低。     

巢式PCR法检测鼠肝脾标本伯氏疏螺旋体

目的应用巢式PCR方法检测鼠肝脾组织中伯氏疏螺旋体外膜蛋白A（ospa）基因,了解淳安县鼠中伯氏疏螺旋体感染情况。方法于2012年8月收集淳安县102份鼠肝脾标本,采用巢式PCR法检测伯氏疏螺旋体ospA基因特异片段,统计标本阳性率。结果在38只黑线姬鼠中检测到伯氏疏螺旋体ospA基因特异片段阳性标本3份,在13只黄毛鼠中检测到伯氏疏螺旋体ospA基因特异片段阳性标本3份,鼠伯氏疏螺旋体总阳性率为5．88％（6／102）。结论淳安县鼠中检测到伯氏疏螺旋体ospA基因片段,有引起蜱媒传染病莱姆病发生的潜在可能,应引起公共卫生部门的重视。     

六盘山地区蜱中莱姆病螺旋体检测及分型

目的了解宁夏部分地区蜱中莱姆病螺旋体感染及基因型别情况。方法选择宁夏六盘山地区为调查点,采集当地蜱类,用巢式PCR法进行检测,阳性产物进行限制性片段长度多态性(RFLP)分析,确定莱姆病螺旋体的基因型。结果共检测4个蜱种206只蜱,阳性37只,阳性率17.96%;其中以青海血蜱阳性率最高,达59.37%;RFLP分析表明,蜱中莱姆病螺旋体均为B.garinii基因型,但型内存在一定差别,其中有29份标本属B.garinii基因型Ⅰ群,有8份标本属B.garinii基因型Ⅱ群。结论证实在宁夏六盘山地区蜱中存在莱姆病螺旋体,并发现其属于B.garinii基因型。     

一组长角血蜱中检出莱姆病螺旋体和斑点热群立克次体复合感染

目的 了解浙江省山区野生动物和蜱中莱姆病、斑点热、埃立克体病(无形体病)的感染情况.方法 采用巢式PCR对采集的鼠、蜱标本进行莱姆病伯氏疏螺旋体、斑点热群立克次体、埃立克体(无形体)特异性核酸片段检测分析.结果 从121份鼠标本和105组蜱标本中检出阳性结果 14份.鼠标本中检出伯氏疏螺旋体5S～23S rDNA间隔区片段1份和埃立克体(无形体)16SrDNA 5'端片段2份.蜱标本中检出阳性11份,包括伯氏疏螺旋体5S～23S rDNA间隔区片段3份和斑点热群立克次体外膜蛋白OmpA基因5'端片段8份.其中1组长角血蜱成虫标本为伯氏疏螺旋体和斑点热群立克次体复合感染,5S～23S rRNA基因间隔区和ompA基因片段均阳性,分别与伯氏疏螺旋体法雷氏基因型和马赛立克次体株等关系较近.结论 在同一组长角血蜱成虫中同时检出莱姆病疏螺旋体和斑点热群立克次体复合感染.     

长春地区首次发现莱姆病疫源地

目的确认长春地区是否莱姆病疫源地。方法用酶联免疫吸附试验等方法检测莱姆病人群特异IgG、IgM抗体，全沟硬蜱中肠伯氏疏螺旋体培养。长春地区全沟硬蜱733只，用BSK33℃培养，获得伯氏疏螺旋体培养物278株，用兔抗美国B31特异性单克隆抗体和针对莱姆病螺旋体种属特异M7，间接免疫荧光抗体进行试验。其结果与美国菌株稍有差异，同国内分离株M7相同。血清学调查298人份，15人阳性，平均感染率为5．03％。结果长春地区莱姆病疫源人群、家畜、动物感染莱姆病。结论首次确认长春地区存在莱姆病自然疫源地。     

我国滇西北山区家犬莱姆病螺旋体基因的检测及序列分析

目的了解滇西北山区莱姆病伯氏疏螺旋体的自然感染及基因分型情况。方法以山区农村家犬作为指示动物,使用真空采血管抽取家犬全血,应用巢式PCR扩增犬血中伯氏疏螺旋体5S~23S rRNA间隔区片段,然后对阳性片段进行测序,并将所测序列与GenBank中注册的基因序列进行比较分析。结果共检测贡山、福贡2个县3个乡镇山村63只家犬血液,其中发现9只犬为阳性,阳性率14.29%。基因序列分析结果显示,当地犬感染的伯氏疏螺旋体为Borrelia garinii。结论首次证实云南省西北部山区家犬中存在莱姆病Borrelia garinii型伯氏疏螺旋体感染。     

用聚合酶链方法对北京林区莱姆病疫源地及莱姆病病原体基因型的探索性研究

目的：进一步明确北京林区是否存在莱姆病的自然疫源地及其分布，方法基于莱姆病螺旋体外膜蛋白A基因建立半巢式聚合酶链式反应(polymerase chain reactio,PCR)方法，对从北京6个林区采集的蜱和鼠进行检测和基因分型，选择阳性标准本进行克隆和序列测定，与已知序进行同源性比较，间接免疫荧光法检测抗莱姆病螺旋体IgG抗体，从长角血蜱中分离莱姆病螺旋体。结果：从门头沟区东灵山采集的标本中检测到莱姆病螺旋体DNA片段，3只游离全沟硬蜱1只检测阳性，57只寄全沟硬蜱若蜱中1只检测阳性；119只野鼠中9只检测阳性，其中8只，B.garinii阳性，1只B.afzelii阳性。50份野鼠血清有5份莱姆病螺旋体IgG抗体阳性，采集采的160只长角血蜱（20只／组）。未分离到莱姆病螺旋体菌株。结论：北京门头沟区东灵山可能存在莱姆病的自然疫源地，包括两个基因型，全沟硬蜱可能是莱姆病的传播媒体，野鼠可能是贮存宿主。     

青海省部分地区媒介及宿主动物体内携带伯氏疏螺旋体及基因型别的研究

目的 了解青海省部分地区蜱及野鼠体内莱姆病螺旋体感染及其基因分型情况。方法 采用布旗法采集蜱标本,采用夹夜法捕捉野鼠。经种类鉴定后,提取病原体DNA,应用巢式PCR扩增鼠中莱姆病螺旋体5S-23S rRNA间隔区片段,对阳性产物进行限制性片段长度多态性(RFLP)分析。结果 检测3个区县3个蜱种共计618只蜱标本,其中共有3个蜱种97只标本检测阳性,总阳性率为15.69%。不同蜱种阳性率存在差别(χ~2=9.68,P0.01),其中以青海血蜱阳性率为最高,达到21.66%(47/217)。检测2个鼠种121只野鼠,其中12只检测阳性,总阳性率为9.91%,不同鼠种阳性率存在差别(χ~2=4.64,P0.05),以黄胸鼠的阳性率为最高,达到17.39%(8/46)。基因分型分析结果显示蜱及野鼠标本中莱姆病螺旋体为B.garinii及B.afzelii基因型,其中B.garinii基因型占76.47%。结论 青海省部分地区媒介及宿主动物体内存在莱姆病螺旋体,并至少有B.garinii及B.afzelii两种基因型。     

Tickborne pathogen detection, Western Siberia, Russia

Ixodes persulcatus (n = 125) and Dermacentor reticulatus (n = 84) ticks from Western Siberia, Russia, were tested for infection with Borrelia, Anaplasma/Ehrlichia, Bartonella, and Babesia spp. by using nested polymerase chain reaction assays with subsequent sequencing. I. persulcatus ticks were infected with Borrelia burgdorferi sensu lato (37.6% +/- 4.3% [standard deviation]), Anaplasma phagocytophilum (2.4% +/- 1.4%), Ehrlichia muris (8.8% +/- 2.5%), and Bartonella spp. (37.6% +/- 4.3%). D. reticulatus ticks contained DNA of B. burgdorferi sensu lato (3.6% +/- 2.0%), Bartonella spp. (21.4% +/- 4.5%), and Babesia canis canis (3.6% +/- 2.0%). Borrelia garinii, Borrelia afzelii, and their mixed infections were observed among I. persulcatus, whereas B. garinii NT29 DNA was seen in samples from D. reticulatus. Among the I. persulcatus ticks studied, no Babesia spp. were observed, whereas B. canis canis was the single subspecies found in D. reticulatus.     

Characterization of Borrelia burgdorferi sensu lato from Novosibirsk region (West Siberia, Russia) based on direct PCR

Borrelia burgdorferi sensu lato infecting Ixodes persulcatus ticks near Novosibirsk, Russia were detected using PCR with primers specific to 5S and 23S rRNA genes. Two genospecies, B. afzelii and B. garinii, were identified by the PCR-based restriction fragment length polymorphism analysis with Tru9-I restriction endonuclease. Comparison of the corresponding nucleotide sequences revealed considerable diversity of the 5S-23S intergenic spacer structure among B. garinii.     

陕西省莱姆病自然疫源地调查研究

目的调查了解陕西省莱姆病自然疫源地。方法人群莱姆病螺旋体血清抗体检查,采用间接免疫荧光法(IFA);莱姆病患者,采用个案调查;蜱样,采用拖旗法采集;莱姆病螺旋体检测,采用巢式PCR法;莱姆病螺旋体基因分型,采用RFLP法。结果通过血清流行病学调查,陕西省富县张家湾镇、南郑县碑坝、陇县八渡镇3个农林地区人群莱姆病螺旋体自然感染率分别为8.33%(6/113)、8.88%(3/36)和7.51%(4/45)(P0.05),发现莱姆病患者14例,蜱类以嗜群血蜱和达吉克斯坦革蜱为优势种,检测蜱样1 116只。检测出阳性138只,总阳性率为12.35%。物种的阳性率分别为嗜群血蜱30.06%(107/356),达吉克斯坦革蜱3.88%(29/747),日本血蜱15.38%(2/13);检测出阳性蜱样138只,其中嗜群血蜱107只,84只为伽氏疏螺旋体基因型,23只为阿弗西尼疏螺旋体基因型;达吉克斯坦革蜱29只,24只为伽氏疏螺旋体基因型,5只为阿弗西尼疏螺旋体基因型;日本血蜱2只,均为伽氏疏螺旋体基因型。结论研究提示陕西省富县张家湾镇、南郑县碑坝、陇县八渡镇3个农林地区存在莱姆病自然疫源地。     

东北中朝俄边境地区蜱的生态及防制对策研究

目的调查东北中朝俄边境地区蜱的种类、生态,制定防制措施。方法采用人工布旗小时法和牛诱法采集蜱。结果在东北中朝俄边境地区7县、市采获蜱30 023只,计3属5种。丘陵草地的优势种群为长角血蜱(99.7%);针阔混交林为嗜群血蜱(69.5%);灌丛、林缘和阔叶林均为森林革蜱,分别占96.1%,85.1%和41.2%。不同月份的蜱种组成有所不同。5种蜱的活动时间,嗜群血蜱13旬,日本血蜱12旬,长角血蜱10旬,全沟硬蜱10旬,森林革蜱8旬;高峰期,森林革蜱在5月上旬,全沟硬蜱和日本血蜱均在5月下旬,长角血蜱在6月中旬,嗜群血蜱在6月下旬。结论为东北中朝俄边境地区蜱的生态和防制提供了依据。     

Detection of four Borrelia burgdorferi genospecies and first report of human granulocytic ehrlichiosis agent in Ixodes ricinus ticks collected in central Italy

The presence of Borrelia burgdorferi s.l. and of Ehrlichia phagocytophila group was sought by PCR in Ixodes ricinus collected in a protected area of central Italy. Nymphs (n = 1475, gathered in 295 pools of 5 nymphs each) and adult ticks (n = 28) were examined. B. burgdorferi s.l. was detected in 13.8% of the nymph pools; of these, 63.4% were infected by B. valaisiana, 26.8% by B. afzelii, 7.3% by B. garinii, and 2.5% by B. burgdorferi s.s. Only a single adult male tick proved to host B. afzelii. The agent of human granulocytic ehrlichiosis (HGE) was detected in 2.7% of the nymph pools. Two HGE agent-positive nymph pools were also found to be positive for B. garinii and for B. afzelii, respectively. This is the first report from central Italy of the finding of the HGE agent in ticks.     

中国莱姆病螺旋体主要生物媒介的调查研究

为研究中国莱姆病螺旋体的生物媒介———蜱的种类、带菌状况及在传播莱姆病中的作用 ,采用布旗法和动物诱捕法在山林地区采集蜱 ,进行分类鉴定 ,应用直接荧光抗体染色法查蜱中肠带菌率 ,BSK培养基分离病原体 ,单克隆抗体鉴定菌株。调查蜱叮咬率与人群莱姆病感染率、患病率以及蜱的季节消长与莱姆病慢性游走性红斑 (ECM )病例发生的季节性关系。在全国 2 0个省 (市、区 )共采集到170 0 0多只蜱 ,分属为 2科 8属 2 3种 ,蜱类分布的地区差异较大 ,北方林区以全沟硬蜱为优势种 ,南方则以二棘血蜱和粒形硬蜱为优势种。 9种蜱的中肠携带莱姆病螺旋体 ,其带菌率以全沟硬蜱、二棘血蜱和粒形硬蜱为高 ,分别为 2 0 .0 0 %～ 4 5.0 0 %、16.0 0 %～ 4 0 .0 0 %和 2 4 .0 0 %。从 8种蜱分离出 10 8株莱姆病螺旋体 ,其单克隆抗体反应与美国B31菌株有明显的差异 ,绝大多数中国菌株对H6831呈阴性反应。蜱叮咬率高的人群 ,莱姆病的感染率和患病率也高。全沟硬蜱的季节消长与ECM发生的季节性相一致。证实多种蜱的中肠内携带莱姆病螺旋体 ;我国北方林区莱姆病螺旋体的主要生物媒介是全沟硬蜱 ,南方林区二棘血蜱和粒形硬蜱是重要的生物媒介。     

Prevalence of Borrelia burgdorferi sensu lato genospecies in host-seeking Ixodes ricinus ticks in selected South Bohemian locations (Czech Republic)

In selected localities of Ceské Bud?jovice and Cesky, Krumlov districts, well known by stable high incidence of tick-borne encephalitis (TBE) human cases but with low incidence of Lyme borreliosis, monitoring of Borrelia burgdorferi sensu lato (s.l.) in Ixodes ricinus ticks was performed. Research was also aimed at the spread of I. ricinus to mountain areas of this region (National Park Sumava), as well as at investigating this tick for B. burgdorferi s.l. genospecies and TBE virus infection. Altogether 498 nymphs, 88 females and 11 males of I. ricinus from lower locations and 58 nymphs from mountain locations (760-1080 m above sea level) were tested by polymerase chain reaction. In lower locations total prevalence of Borrelia burgdorferi s.l. in Ixodes ricinus ticks was 35%. Single infection of Borrelia afzelii, B. garinii and B. burgdorferi sensu stricto (s.s.) was found in 59, 50 and 63 ticks, respectively (i.e. in 12.8, 11.2 and 14.1%). Double infection was found in 42 ticks (6.0%) and triple infection in three ticks (0.4%). The high frequency of B. burgdorferi s.s. exceeds the as yet reported occurrence in Central Europe. These circumstances are discussed. In mountain locations B. afzelii was found in five ticks, that including two co-infection with B. garinii, in elevations of 762 m and 1024 m above sea level, respectively. This fact signals a real danger of human infections in a region that was previously deemed to be without risk. Moreover, this region is more and more the target destination of tourist activities. The results also suggest that the penetration of infection can be rapid and formation and establishment of natural focus of Lyme borreliosis might be rather quick.     

The distribution and prevalence of B. burgdorferi genomospecies in Ixodes ricinus ticks in Ireland

Questing Ixodes ricinus ticks were collected from six locations throughout Ireland and 638 nymphs, 111 females and 118 males were investigated for infection with Borrelia burgdorferi sensu lato (s.l.). The total prevalence of B. burgdorferi s.l. in the ticks was determined as 14.9% by polymerase chain reaction (PCR) amplification of the spacer region of 5S-23S rRNA genes. Infection prevalence was significantly higher in adult (20.1%) Ixodes ricinus compared to nymphs (13.1%). The prevalence of infection in adult male and female ticks was similar (19.5% and 20.7% respectively). The genomospecies B. burgdorferi sensu stricto, B. afzelii, B. garinii and group VS116 were identified by reverse line blot (RLB) using genomospecies specific oligonucleotide probes. The most prevalent B. burgdorferi genomospecies identified were VS116 (34.6%), B. garinii (24.3%) and B. burgdorferi sensu stricto (18.4%). B. afzelii was uncommon (6.6%). Multiple infections were observed in 13.2% of the infected ticks. The distribution of the genomospecies showed geographical variation and also seemed to be influenced by the nature of the habitat. A broad range of genomospecies seemed to be associated with the presence of a wide spectrum of potential reservoir hosts in the habitat and also with a high overall prevalence of B. burgdorferi s.l.     

Genetic diversity of Borrelia burgdorferi sensu lato isolates obtained from Ixodes ricinus ticks collected in Slovakia

In Europe, Borrelia burgdorferi sensu lato is diverse, including B.burgdorferi s.s., B.garinii, B.afzelii, B.valaisiana and B.lusitaniae. In this study, we focused on the distribution of the different B.burgdorferi species among Ixodes ricinus adult ticks collected in an endemic area within Slovakia. We compared results of prevalence of B.burgdorferi infection in ticks obtained by immunofluorescence (IF) and by isolation. Isolates were characterized by restriction fragment length polymorphism (RFLP) of the rrf-rrl intergenic spacer genes using MseI. Using immunofluorescence we observed that 56/114 (49%) ticks were infected by B.burgdorferi s.l. Males were found to be more often infected (32/57, 56%) than females (24/57, 42%) but the difference was not significant (p=0.1895). From the same 114 ticks a total of 37 isolates were obtained: 19 from males (33%) and 18 from females (32%). The RFLP identification revealed 25 B.afzelii (68%), 5 B.garinii (14%), 5 B.valaisiana (14%) and 2 B.lusitaniae (5%). The infection in ticks was more often detected by IF than by isolation (p=0.0153) and isolation success was higher when the infection degree in ticks was high (p=0.0397). The infection prevalence observed in this area is among the highest observed in Europe.