新疆博乐和沙湾、阜康市县山地莱姆病螺旋体(Borrelia burgdorferi)的分离及鉴定

1991年5月从新疆博乐市准噶尔阿拉套山山地(E81°18′,N45°13′)、沙湾县北天山西段依连哈比尔尕北坡(E85°40′,N43°55′)和阜康县北天山东段博格达山山地北坡(E88°08′,N43°55′)共采集到全沟硬蜱(Ixodes persulcatus)千余只,经做莱姆病病原体分离,共检出疏螺旋体6株。对其中5株做了单克隆抗体试验,5株疏螺旋体均与单克隆抗体H_(9724)呈阳性反应,而与H_(6831)均呈阴性反应,有3株与H_(5332)呈阳性反应、2株呈阴性反应。从而确认所分离的5株疏螺旋体为Borreliaburgdorferi疏螺旋体。首次从病原学证实上述一市二县山地存在莱姆病。

中国新疆两株Borreliaburgdorferi──XjI_3和XjI_（1

作者用Ｄｉｓｃｏｎｔｉｎｕｏｕｓ ＳＤＳ一ＰＡＧＥ及Ｗｅｓｔｅｒｎ ｂｌｏｔ分析了两株分离自中国新疆的Ｂｏｒｒｅｌｉａ ｂｕｒｇｄｏｒｆｅｒｉ─—ＸｊＩ＿３和ＸｊＩ＿（１２）株的主要蛋白组分及其生物学属性。ＸｊＩ３和ＸｊＩ＿（１２）均具有４１ｋＤ鞭毛蛋白和６０ｋＤＣＡ蛋白，均缺失３４ｋＤ蛋白。ＸｊＩ＿（１２）具有３３ｋＤ的ＯｓｐＡ蛋白，而ＸｊＩ＿３缺失该蛋白，但具有丰富的２３ｋＤ蛋白。

Interpretation Criteria for Standardized Western Blot for the Predominant Species of Borrelia Burgdorferi Sensu Lato in China

Objective Western blotting （WB;immunoblotting） is a widely used tool for the serodiagnosis of Lyme borreliosis （LB）,but so far,no generally accepted criteria for its performance and interpretation have been established in China.The present study was designed to determine the criteria for standardized Western blot for the predominant species of Borrelia burgdorferi sensu lato in China,in which WB was produced with strain PD91 as the representative strain attributed to predominant genospecies Borrelia garinii of Borrelia burgdorferi sensu lato.Methods Approximately 13 bands between 14 and 100 kD were differentiated for strain PD91 by using Gel-Pro analysis software.In a study with 631 serum samples （taken from 127 patients with Lyme borreliosis and 504 controls）,all observed bands were documented.To establish criteria for a positive WB result for strain PD91,receiver operating characteristic （ROC） curves were used.Results The following interpretation criteria were recommended：for IgG,at least one band of P83/100,P58,P39,P30,OspC,P17,P66,and OspA;for IgM,at least one band of P83/100,P58,OspA,P30,OspC,P17 or P41.In addition,syphilis,leptospirosis and other related diseases should be excluded when the positive band is P41 in IgM.For IgG criteria,the sensitivity is 73.2%,the specificity is 99.4% and Youden index is 0.726;for IgM criteria,the sensitivity is 50.6%,the specificity is 93.1% and Youden index is 0.437.Conclusion Standardization of WB assays is necessary for comparison of results from different laboratories.Moreover,the criteria of other genospecies of Borrelia burgdorferi sensu lato should be determined in the future to complete the criteria of WB for the diagnosis of the Lyme disease in China.

BORRELIA BURGDORFERI DNA IN BIOLOGICAL SAMPLES FROM PATIENTS WITH SARCOIDOSIS USING THE POLYMERASE CHAIN REACTION TECHNIQUE

ＢＯＲＲＥＬＩＡＢＵＲＧＤＯＲＦＥＲＩＤＮＡＩＮＢＩＯＬＯＧＩＣＡＬＳＡＭＰＬＥＳＦＲＯＭＰＡＴＩＥＮＴＳＷＩＴＨＳＡＲＣＯＩＤＯＳＩＳＵＳＩＮＧＴＨＥＰＯＬＹＭＥＲＡＳＥ ＣＨＡＩＮ ＲＥＡＣＴＩＯＮ ＴＥＣＨＮＩＱＵＥＬｉａｎＷｅｉ（连伟）ａｎｄ...

Optimization of Pulse-Field Gel Electrophoresis for Borrelia burgdorferi Subtyping

Objective To optimize the performance of Pulsed-Field Gel Electrophoresis （PFGE） for the comparison of inter-laboratory results and information exchange of Borrelia burgdorferi subtypingo Methods A panel of 34 strains of B. burgdorferi were used to optimize PFGE for subtyping. In order to optimize the electrophoretic parameters （EPs）, all 34 strains of B. burgdorferi were analyzed using four EPs, yielding different Simpson diversity index （D） values and the epidemiological concordance was also evaluated. Results The EP of a switch time of l s to 25 s for13 h and l s to10 s for 6 h produced the highest D value and was declared to be optimal for Mlul and 5mal PFGE of B. burgdorferi. Mlul and Smal were selected as the first and second restriction enzymes for PFGE subtyping of B. burgdorferi according to discrimination and consistency with epidemiological data. Conclusion PFGE can be used as a valuable test for routine genospecies identification of B burgdorferi.

Establishment of Multiple Locus Variable-number Tandem Repeat Analysis Assay for Genotyping of Borrelia burgdorferi sensu lato Detected in China

Objective Human Lyme Borreliosis （LB）, which is caused by Borrefia burgdorferi sensu lato （B. burgdorferi）, has been identified as a major arthropod-borne infectious disease in China. We aimed to develop a multiple locus variable-number tandem repeat （VNTR） analysis （MLVA） assay for the genotyping of Borrelia burgdorJ：eri strains detected in China. Methods B. garinii PBi complete 904.246 kb chromosome and two plasmids （cp26 and Ip54） were screened by using Tandem Repeats Finder program for getting potential VNTR loci, the potential VNTR loci were analyzed and identified with PCR and the VNTR loci data were analyzed and MLVA clustering tree were constrcted by using the categorical coefficient and the unweighted pair-group method with arithmetic means （UPGMA）. Results We identified 5 new VNTR loci through analyzing 47 potential VNTR loci. We used the MLVA protocol to analyse 101 B. burgdorferi strains detected in China and finally identified 51 unique genotypes in 4 major clusters including B. burgdorferi sensu stricto （B.b.s.s）, B. garinii, B. a[zelii, and B. valaisiana, consistent with the current MLSA phylogeny studies. The allele numbers of VNTR-1, VNTR-2, VNTR-3, VNTR-4, and VNTR-5 were 7, 3, 9, 7, and 6. The Hunter-Gaston index （HGI） of five VNTR loci were 0.79, 0.22, 0.77, 0.71, and 0.67, respectively. The combined HGI of five VNTR loci was 0.96. Clustering of the strains of Xinjiang, Inner Mongolia and Heilongjiang was confirmed, and this situation was consistent with the close geographical distribution of those provinces. Conclusion The MLVA protocol esytablished in this study is easy and can show strains＇ phylogenetic relationships to distinguish the strains of Borrelia species. It is useful for further phylogenetic and epidemiological analyses of Borrelia strains.

Combination of Loop-Mediated Isothermal Amplification Assay and Nested PCR for Detection of Borrelia burgdorferi sensu lato in Human Serum Samples

A set of universal loop-mediated isothermal amplification （LAMP） primers targeting the flo gene was designed to detect Borrelia burgdorferi sensu lato （B. burgdorferi s.I.） in human samples. The sensitivity of LAMP was 20 copies/reaction, and the assay did not detect false positives among 11 other related bacteria. A positive LAMP result was obtained for 9 of the 24 confirmed cases and for 12 of 94 suspected cases. The positive rate of LAMP was the same as that of nested PCR. The LAMP is a useful diagnostic method that can be developed for rapid detection of B. burgdorferi s.I. in human sera. Combination of the LAMP and nested PCR was more sensitive for detecting B. burgdorferi s.I. in human serum samples.

Infection Prevalence of <i>Borrelia burgdorferi</i>in Ticks Collected from Songbirds in Far-Western Canada

Worldwide, wild birds play a vital role in the dispersal of ticks that harbour tick-borne pathogens, including Borrelia burgdorferi, the Lyme disease bacterium. Using PCR testing, we found 124 (31%) of 405 ticks (4 species), which were collected from 21 species of birds in far-western Canada, to be infected with B. burgdorferi. Transstadial transmission of B. burgdorferi occurred from larva to nymph, plus nymph to adult, in the avian coastal tick, Ixodes auritulus, collected from songbirds in British Columbia (B.C). Collectively, all 3 motile life stages (larva, nymph, adult) of this tick had an infection prevalence of 31% for B. burgdorferi, which suggests vector competency. A Pacific Wren was highly infested with I. auritulus immatures, and 20 (44%) of 45 ticks (2 nymphs, 43 larvae) were infected with B. burgdorferi. This heavy infestation shows the high potential to initiate a new population of ticks and to disseminate Lyme spirochetes. Epidemiologically, B. burgdorferi-infected I. auritulus larvae collected from the Spotted Towhee, Swainson’s Thrush, Pacific Wren, and Fox Sparrow suggest that these avian hosts act as reservoirs for B. burgdorferi. In this study, the western blacklegged tick, Ixodes pacificus, and Ixodes spinipalpis played a limited role in the enzootic transmission cycle of B. burgdorferi along coastal B.C. We document the first record of I. spinipalpis on a bird in Alberta. Because songbirds widely disperse Lyme disease vector ticks, primary health providers and the general public must be vigilant that outdoors people may be bitten by B. burgdorferi-infected ticks throughout far-western Canada.

Chronic Lyme Disease: Persistent Clinical Symptoms Related to Immune Evasion, Antibiotic Resistance and Various Defense Mechanisms of <i>Borrelia burgdorferi</i>

There are several factors involved in the ability of Borrelia burgdorferi to retain a persistent infection within a mammalian host. These factors of immune evasion include regulation of membrane proteins, variable epitopes of surface proteins, protection against the immune system through tick saliva, the ability to migrate to regions where it is not exposed to the immune system or antibiotics, invagination or invasion within various cells, pleomorphic forms, and the potential to produce biofilms. The window of conventional treatment for Lyme disease is short and has the potential to display different symptoms depending on the strain of Borrelia bugdorferi. These symptoms are dependent on the localization of Borrelia burgdorferi which correlates to the significance of diagnosing Lyme disease early to prevent such a spread throughout the body. Such complications of Borrelia burgdorferi may demand new clinical treatment discoveries for patient fighting the chronic form.

<i>Borrelia</i><i>burgdorferi</i>: Cell Biology and Clinical Manifestations in Latent Chronic Lyme

Chronic Lyme disease is predicated by an infection with Borrelia burgdorferi via tick vector. B. burgdorferi has been extensively researched with regard to its genome and cell biology. There are many unique characteristics to the bacteria itself;however, serological diagnostics and diagnosis based on symptoms can be complicated and potentially misleading. Other promising diagnostics were also evaluated in this review. Treatment of the chronic Lyme disease can be complicated and at times ineffective. The purpose of this review is to examine B. burgdorferi from a biological and clinical perspective.

Introduction, Sign, Symptoms, Prevention and Management of Lyme Disease Caused by Borrelia burgdorferi Channeled through Ixodes Ticks as Vector. A Review

God has created many miraculous creature;ticks are one of them. Borrelia burgdorferi is a bacterium that lives into the gut of ticks. When such ticks feed upon the blood of Ticks and other livestock, bacteria are transferred into their blood stream of these hosts. A disease is caused into human beings if such infected tick bites humans that are called as Lyme disease. This review will help to have a keen study on the cause, sign, symptoms, prevention and management of this disease.

Polymorphism of P66 in Borrelia Burgdorferi Strains in China

Objective To study the polymorphism in P66 and its human B-cell epitopes of Borrelia burgdorferi strains in China.Methods Polymerase chain reaction(PCR)and sequencing were used to obtain the P66 sequences of59 Chinese B.burgdorferi.Then the sequences were analyzed by MEGA 5.10 software and compared with the human B-cell epitope sequences from the Immune Epitope Database(IEDB)based on the reference strain of each genotype.Results Results showed that genetic and amino acid diversity presented in the 66 kD protein of all 59 Chinese strains,especially in Borrelia garinii(B.g)and Borrelia afzelii(B.a)strains.B.g strains were divided into three subclusters and two scattered strains JC1-7 and JC2-2 according to the amino acid sequences of P66.The P66 sequences of 15 Xinjiang strains represented by XI91-12 in the B.g subcluster1,changed from CAA to TAA codon at 508 aa position,resulting in early termination.Bases A and C were inserted at sequence position 1523 bp of strains FP1,LB20,LB21,and SZ21 in the B.a genotype,which resulted to early termination at position 511 aa.G base was inserted at 438 bp of LIP94-11 strain,which led to early termination at position 172 aa.Conclusion In P66 of 59 Chinese strains,polymorphisms were widely distributed.More importantly,the P66 amino acid sequences of B.g strains had a certain regional character.One of the characteristics of Xinjiang B.g isolates might be the variation at the 508 aa location in 15 Xinjiang B.g strains,which may be related to the strains’pathogenicity in this area.

Genetic Characterization of Four Strains Borrelia Burgdorferi Isolated in China

To study the genetic characterization of four strains of Borrelia burgdorferi isolated in China. PCR technique was used to amplify the 5S-23S rRNA intergenic spacer DNA from the whole cellular DNA of isolated GXLD-4, 9, 18 and Chang 14, and then the amplified products were cloned into plasmid pGEM-T Easy and sequenced. It was found that the 5S-23S rRNA intergenic spacer DNA of the four isolates was 242?bp, revealing the nucleotide sequence identity of more than 99%. The four isolates had higher sequence identify with Borrelia valaisiana than with other genetic groups. These four isolates most likely belong to Borrelia valaisiana genomic group.

Mucosa-Associated Lymphoid Tissue (MALT) Lymphoma of the Colon in a Patient with <i>Borrelia burgdorferi</i>Infection (Lyme Arthritis)

Mucosa-associated lymphoid tissue (MALT) lymphoma generally occurs in the context of chronic inflammation or autoimmune disorders. The most common infections linked to MALT lymphomas include Helicobacter pylori (Hp)-associated gastritis, Chlamydophila psittacii and hepatitis C infection. Although Borrelia burgdorferi infection has been linked to primary cutaneous B-cell lymphoma (PCBCL), there is no known link between Borrelia burgdorferi infection and MALT lymphomas in the US. We report a patient who developed mucosa-associated lymphoid tissue (MALT) lymphoma of the colon in the context of untreated Lyme arthritis, and no other autoimmune disorders or infections known to be associated with MALT lymphoma. We recommend that Lyme disease due to Borrelia burgdorferi be considered as a possible underlying infection potentially contributing to the emergence of extranodal lymphoma.

莱姆病螺旋体端粒解离酶的表达纯化及其晶体衍射分析

【目的】对莱姆病螺旋体(Borrelia burgdorferi)端粒解离酶(telomere resolvase,ResT)进行原核可溶性表达纯化,并筛选高衍射度的ResT-DNA复合物晶体,为探究ResT-DNA复合物的结构和功能奠定基础。【方法】合成pET28 b-ResT表达质粒,构建原核表达载体pSUMO-ResT,分析ResT可溶性蛋白在BL21(DE3)、BL21(DE3)pLysS、BL21 Gold(DE3)pLysS、BL21 Codon Plus(DE3)、Rosetta(DE3)和Rosetta(DE3)pLysS中的表达量,获得最优的ResT原核表达菌株。基于AlphaFold预测的ResT蛋白模型,利用定点突变及ResT可溶性蛋白表达分析,获得高效可溶性表达ResT突变体。经Ni-NTA亲和层析、HiTrap Heparin HP亲和层析及分子筛层析对ResT突变体ResT(W94H)进行蛋白纯化。利用EMAS检测ResT(W94H)与DNA的结合能力。使用蛋白晶体接种(seeding)和交叉接种(cross-seeding)方法,利用多种商业化结晶试剂盒筛选并优化ResT-DNA复合物的结晶条件,对复合物晶体进行X射线衍射分析。【结果】BL21 Gold(DE3)pLysS菌株是ResT最佳的高效可溶性表达菌株,W94H突变体比野生型具有更强的可溶性表达。获得了高纯度(95%)且高质量浓度(15 mg/mL)的ResT(W94H),其能够与底物DNA形成稳定的ResT-DNA复合物。在18℃下,ResT-DNA最佳结晶条件是:PEG3350150 g/L、二甲苯二酸钠0.1 mol/L(pH 6.6)、NaCl 0.15 mol/L,ResT-DNA晶体分辨率最高为3.52,晶体空间群为P4。【结论】得到了高纯度ResT(W94H)蛋白,获得了ResT-DNA复合物晶体。

云南省西部地区宿主和媒介感染伯氏疏螺旋体的基因多样性调查与分析

目的了解云南省西部地区宿主动物和媒介蜱感染伯氏疏螺旋体的基因多样性,为进一步掌握该地区莱姆病菌株的特性奠定基础。方法对采用伯氏疏螺旋体5S~23S rRNA间隔区基因筛查阳性的云南西部地区采集的家畜、小型兽类、蜱虫样本,进行伯氏疏螺旋体鞭毛蛋白基因(FLA基因)和16S rRNA基因的检测,综合分析病原体的基因信息,了解当地流行株的生物学特性及流行特征。结果共检测301份样本,共检出7种基因型伯氏疏螺旋体,其中宿主动物共检出5种:B.afzelii(阿弗西尼疏螺旋体)、B.garinii(伽氏疏螺旋体)、B.burgdorferi s.s.(狭义伯氏疏螺旋体)、B.japonica(日本疏螺旋体)、B.valaisiana(法雷斯疏螺旋体);媒介蜱检出7种:B.afzelii(阿弗西尼疏螺旋体)、B.garinii(伽氏疏螺旋体)、B.burgdorferi s.s.(狭义伯氏疏螺旋体)、B.japonica(日本疏螺旋体)、B.valaisiana(法雷斯疏螺旋体)、B.sinica(中华基因型伯氏疏螺旋体)及未定种Borrelia sp.。B.afzelii、B.garinii和B.burgdorferi s.s.为优势流行株,占总阳性的85.04%(256/301)。不同地区、不同来源的相同基因型株基因序列存在种内差异,同一地区的宿主和媒介中检出的相同基因型螺旋体具有较高的同源性和较近的遗传进化关系。结论云南西部地区宿主和媒介感染伯氏疏螺旋体基因型具有较高的多样性特征,存在多种致病基因型,给当地人群的健康造成一定危害,需加强防范。

安徽黄山地区中华硬蜱及其宿主感染莱姆病Borrelia afzelii的检测

利用PCR和病原体分离技术对安徽黄山地区中华硬蜱及其宿主感染莱姆病螺旋体的状况进行了检测，结果从40只黄山林区的中华硬蜱中获得了一株莱姆病螺旋体分离株，寄主动物及其他中华硬蜱均未获得阳性分离结果。以16srRNA为靶标进行PCR检测，6．78％雌性和5％雄性中华硬蜱发现有阳性感染，2只草兔（92只）和1只麂子（16只）也发现有阳性感染；若蜱和其余宿主动物未发现阳性感染。阳性分离株和扩增获得的16srRNA序列结果一致，同GenBank中的B．afielii的系统发育关系最近，提示该分离株螺旋体属于B．afzelii。该结果与前期的实验传播结果提示中华硬蜱是我国南方莱姆病的传播媒介。

江西省吉安市莱姆病疑似病例调查研究

目的了解江西省吉安市莱姆病疑似病例中确诊病例情况及其临床表现特征和地区来源分布,为莱姆病及时诊治提供依据。方法2021年12月至2022年8月从吉安市中心人民医院收集了133份莱姆病疑似病例血清,采用国际上通用的“两步法”进行莱姆病抗体检测,并采用巢式PCR方法检测血清中莱姆病螺旋体的基因,阳性标本进一步测序分析。结果133份莱姆病疑似病例血清中有25份伯氏疏螺旋体检测阳性,总阳性率为18.80%。其中,20份样本抗体检测阳性,阳性率为15.04%;6份样本核酸检测阳性,阳性率为4.51%。6例核酸检测阳性的患者的5S-23S rRNA基因间隔区的序列均与扬子疏螺旋体(Borrelia yangtzensis)的相应序列一致,并且其中1份样本抗体检测也为阳性。25例阳性患者来自吉安市的9个县区,其中吉州区的患者最多,占44%。阳性患者出现莱姆病相关症状,包括关节病变、神经疾病、感染性发热、皮炎或胸痛,其中关节病变的病人最多,占72%。6例扬子疏螺旋体感染者来自吉州区,其中4例表现为关节病变,2例表现为感染性发热。结论本次研究首次证实江西省吉安市存在莱姆病患者,并首次在我国南方的病人体内检测出扬子疏螺旋体。研究提示该地区医务人员在接诊具有关节炎、神经系统等症状的病例时,应考虑患者可能罹患莱姆病。研究表明吉安市存在莱姆病自然疫源地,应进一步开展莱姆病的调查和监测。

伯氏疏螺旋体的特征及其啮齿动物感染情况

莱姆病(Lyme disease,LD)是由伯氏疏螺旋体(Borrelia burgdorferi)感染引起的一种新发蜱传人兽共患传染病,啮齿动物是该病重要宿主动物之一。近年来,随着生态旅游的兴起,人与宿主动物接触机会大大增加,莱姆病发生呈现上升趋势。明确莱姆病的病原特征及啮齿动物自然感染伯氏疏螺旋体情况,有助于评估莱姆病在人群中发生的风险和预测疾病发生的可能性,对防控人类莱姆病发生起到关口前移的作用。