准噶尔盆地大沙鼠鼠疫自然疫源地鼠疫耶尔森菌质粒谱的研究

目的 了解和掌握准噶尔瓮地大沙鼠鼠疫自然疫源地鼠疫菌质粒谱类型.方法 提取新疆地区不同鼠疫自然疫源地及青藏高原喜玛拉雅旱獭鼠疫自然疫源地、昆仑山喜马拉雅旱獭鼠疫自然疫源地和内蒙古长爪沙鼠鼠疫自然疫源地的39株鼠疫菌质粒DNA,采用琼脂糖凝胶电泳法对其进行质粒谱图型分析.结果 在检测的26株准噶尔盆地大沙鼠鼠疫自然疫源地鼠疫菌中,有23株质粒谱为6×106、45×106、65×106,3株为6×106、45×106和72×106.结论 准噶尔盆地大沙鼠鼠疫自然疫源地鼠疫菌质粒谱分为两个类型:一为6×106、45×106、65×106质粒谱型,是该疫源地的主要质粒谱型,与邻近的北天山灰旱獭.长尾黄鼠鼠疫自然疫源地及内蒙占长爪沙鼠鼠疫自然疫源地、昆仑山喜马拉雅旱獭鼠疫自然疫源地和青藏高原喜马拉雅旱獭鼠疫自然疫源地的鼠疫菌主要质粒谱型一致;二为6×106、45×106和72×106质粒谱型,为少数型,在我国系首次发现.     

准噶尔盆地鼠疫耶尔森菌的基因组型分析

目的 对我国新发现的准噶尔盆地大沙鼠鼠疫自然疫源地的鼠疫菌株进行基因组差异性区段的比较分析,确定该鼠疫疫源地鼠疫菌株的基因组类型.方法 按常规方法提取鼠疫菌染色体DNA,依据已证实的鼠疫菌22个差异性区段(DFR)设计引物,采用PCR技术检测鼠疫菌基因组差异性基因片段的构成.结果 准噶尔盆地鼠疫菌株的差异性区段结构类型为缺失DFRO1、04、06、07、10、13和DFR15～18,不同于其他鼠疫疫源地鼠疫菌株的结构类型.为我国新的鼠疫菌基因组型.结论 准噶尔盆地大沙鼠鼠疫疫源地鼠疫菌株的基因组结构类型为我国新的基因组类型.     

应用抑制削减杂交技术进行鼠疫耶尔森菌的比较基因组研究

目的确定古典型鼠疫耶尔森菌的特有序列，为建立和完善该菌基因组分型系统提供可靠数据。方法通过抑制削减杂交技术发现差异片段并应用PCR验证。结果发现了5个在不同生物型鼠疫菌问存在差异的DNA片段，其中一个383bp的片段在来自天山山地灰旱獭、长尾黄鼠鼠疫自然疫源地的30株鼠疫菌全部阳性，而来自其它鼠疫自然疫源地的菌株全部阴性，5株假结核耶尔森菌中有2株(生物Ⅰ型和Ⅱ型)阳性。结论该383bp片段为天山山地灰旱獭、长尾黄鼠鼠疫自然疫源地的鼠疫菌所特有，此疫源地的菌株可能是我国较古老的鼠疫菌。     

天山山地灰旱獭-长尾黄鼠鼠疫疫源地鼠疫菌基因分型研究

目的对天山山地灰旱獭．长尾黄鼠鼠疫疫源地鼠疫菌进行基因分型。方法根据已经证实的22个差异区段（DFR）设计引物，对每株鼠疫菌进行PCR扩增。结果61株天山疫源地鼠疫菌株共包括4个基因型，即1、2、3、4型。基因型1、2和3在北天山疫源地西部鼠疫菌株中都有分布，但尼勒克菌株的基因型全部为1型。南天山疫源地阿合奇和阿图什两个县菌株的基因型存在显著差别，阿合奇菌株基因型与北天山疫源地的菌株比较接近，而3株阿图什菌株的基因型均为4型，与帕米尔高原长尾旱獭鼠疫疫源地鼠疫菌株相同。结论基因分型结果与纪树立的生态分型基本一致。天山疫源地和帕米尔高原疫源地在阿图什县有交叉。     

44株中国田鼠型与24株非田鼠型鼠疫菌的DFR检测分析

目的比较我国两个田鼠鼠疫自然疫源地的44株田鼠型鼠疫菌的基因组组成，研究中国田鼠型鼠疫菌的遗传稳定性。方法根据已经证实的22个差异区段(I)FR)设计引物，每株鼠疫菌的每个DFR都采用PCR技术进行验证。结果22个DFR在两种田鼠型鼠疫菌的分布状态完全一致，即均同时缺失了DFR6、DFR11、DFR12、DFR13、DFR18、DFR19和DFR20，与来源于其他鼠疫自然疫源地非田鼠型鼠疫菌的基因组成有显著差别。结论锡林郭勒高原型鼠疫菌和青藏高原青海田鼠型鼠疫菌在基因组组成上高度一致，田鼠型鼠疫菌的基因缺失可能与其对人不致病有关。     

甘宁黄土高原阿拉善黄鼠鼠疫疫源地鼠疫菌的基因分型研究

目的 研究我国甘宁黄土高原阿拉善黄鼠鼠疫疫源地鼠疫菌的基因分型。方法 根据已经证实的22个DFR（差异区段）设计引物，每株鼠疫菌的每个DFR都采用PCR技术进行验证。结果 17株甘宁黄土高原疫源地鼠疫菌株共包括3个基因型，即7、11和13型。7、11和13型所占比例分别为5．9％（1／17）、5．9％（1／17）和88．2％（15／17）。结论 我国甘宁黄土高原阿拉善黄鼠鼠疫疫源地鼠疫菌的基因型主要为13型，而且13型是该疫源地鼠疫菌所特有的基因型。     

西藏地区鼠疫菌生物学特性的分析

目的：通过对西藏地区29个县（市），不同宿主分离的116株鼠疫菌生化，毒力，毒力因子，质粒及外膜蛋白的分析，了解该疫源地内鼠疫病原体间的群落关系及遗传变异的规律性。方法：各项检查均按常规方法进行。结果：目前该地区仅有岗底斯山型和青藏高原型两种生态型菌株，二者分别占据一定的地理区域，以念青唐古拉山脉为天然屏障，从藏北地区分离的菌株为青藏高原型，藏南地区及其西南部所分离的菌株均为岗底斯山型。结论：西藏地区是以喜玛拉雅旱獭为主要宿主的鼠疫自然疫源地，就其鼠疫菌生物学特性分析表明，藏北高原疫源地可能是青藏高原喜玛拉雅旱獭疫源地的延伸，而藏南谷地可能是由于有天然屏障的存在，形成了自己特定的地理环境，从而表现为相对独立的一块鼠疫自然疫源地。     

我国鼠疫菌毒力因子的比较与分析

目的 研究自不同生态型和不同鼠疫自然疫源地分离的鼠疫菌4种毒力因子的性状特征。方法 常璺方法检查鼠疫菌毒力因子性状并比较在不同疫源地之间、不同生态型之间的差异。结果 1326株鼠疫菌中仅1株缺失FI、6株缺失PstI,Pgm和WW阳性分别为92.99%、76.47%。结论 自我国不同疫源地、平淡同生态型和不同年代分离的野生型鼠疫菌绝大多数能产生FI和PstI,且不同生态型和不同疫源地分离的菌株这两种毒力因子无明显差别,性状稳定,VW^-和Pgm^-菌株与生态型及疫源地有一定的关系。经长期人工培养传代后。4种毒力因子相比,VW因子较易丧失,该毒力因子具有更不稳定特点。     

松辽平原达乌尔黄鼠鼠疫自然疫源地鼠疫菌基因分型

为了研究松辽平原达乌尔黄鼠鼠疫自然疫源地鼠疫耶尔森菌(以下简称鼠疫菌)的基因组成,采用PCR技术对分离自该疫源地88株鼠疫菌进行了基因分型,其中黑龙江4株、吉林43株、辽宁4株、内蒙37株,鼠疫菌DNA均由青海省地方病预防控制所鼠疫菌专业实验室提供。引物设计和PCR扩增方法:在周冬生等鉴定的22个差异区段(DFR)中,有3个位于pMT1质粒中,19个位于染色体上。采用ArrayDesigner2.0软件针对每个DFR各设计1对引物,为了证实pMT1的存在,我们还特意针对pMT1质粒设计了1对引物,引物序列为5′AACACTA TCTCATTCCGCAGTAAAG3′。PCR…     

鼠疫耶尔森菌中国分离株菌体脂肪酸成分分析

目的 探讨利用脂肪酸组织对中国鼠疫耶尔森菌菌株进行分型的可能性,获得中国鼠疫耶尔森菌菌株脂肪酸组成的本底资料。方法 选择历年来从我国不同疫源地分离到的58株鼠疫耶尔森菌进行了菌体脂肪酸成分分析。并利用STATISTICA统计软件进行实验菌株的聚类分析。结果 中国鼠疫耶尔森菌菌株的主要脂肪酸分为16：0酸,环式17：0酸,3-羟基14：0酸和ω7c16:1酸以及十八碳单烯酸,与MIDI公司Sherlock系统的菌库中的数据存在一些差异。结论 实验菌株的脂肪酸成分极为接近,聚类图不能充分体现分离株在分离时间,地点以及生物型别上的差异。表明脂肪酸分析目前不适于鼠疫耶尔森菌的分型,根据实验结果,建立了中国鼠疫耶尔森菌菌株的脂肪权组成数据库。     

Genotyping of strains of Yersinia pestis from Marmota baibacina-Spermophilus undulates Plague Focus of Tianshan Mountains in China

Themainreservoirsoftheplaguebaccilusare MarmotabaibacinaandCitellusundulatesin PlagueFocusofTianshanMountainsinChina.BotharepartofRodentiaSciuridae,butdonot belongtothesamegenus(MarmotaandSper mophilusrespectively).Themainvectorsare OropsyllasiantiewiandCallopsylladorablis.Threecounties,Wusu,JingheandNilekein NorthernTianshanMountainsareMarmotabaiba cina SpermophilusundulatesPlagueFocus.Five counties,Changji,HutubiandManasiinEasternTianshanMountainsandAheqiandAtushiin SouthernTia…     

Identification of different regions among strains of Yersinia pestis by suppression subtractive hybridization

Yersinia pestis, the causative agent of bubonic and pneumonic plague, has been classified into four biovars: Antiqua, Mediaevalis, Orientalis and Microtus. Although the entire genome sequences of three Y. pestis strains, CO92, KIM and 91001, of biovar Orientalis, Mediaevalis and Microtus, respectively, have been decoded, the genome sequence of the biovar Antiqua strain is unknown. In an initial effort to find Antiqua-specific sequences, suppression subtractive hybridization (SSH) was performed and four different regions (DFRs) were identified. Among the four DFRs, only DFR4 was specific to the tester (strain 49006, biovar Antiqua). PCR demonstrated that DFR4 was present only in 57 of 60 Antiqua strains from the Marmota baibacina-Spermophilus undulates plague focus in the Tianshan Mountains (focus B) and in three strains of Y. pseudotuberculosis (serotypes I and II), showing that not all Antiqua strains had DFR4. Five DFR profiles were identified based on the presence or absence of these four DFRs in 636 strains of Y. pestis from 10 plague foci in China.     

Human plague in the southwestern United States, 1957-2004: spatial models of elevated risk of human exposure to Yersinia pestis

Plague is a rare but highly virulent flea-borne zoonotic disease caused by the Gram-negative bacterium Yersinia pestis Yersin. Identifying areas at high risk of human exposure to the etiological agent of plague could provide a useful tool for targeting limited public health resources and reduce the likelihood of misdiagnosis by raising awareness of the disease. We created logistic regression models to identify landscape features associated with areas where humans have acquired plague from 1957 to 2004 in the four-corners region of the United States (Arizona, Colorado, New Mexico, and Utah), and we extrapolated those models within a geographical information system to predict where plague cases are likely to occur within the southwestern United States disease focus. The probability of an area being classified as high-risk plague habitat increased with elevation up to approximately 2300 m and declined as elevation increased thereafter, and declined with distance from key habitat types (e.g., southern Rocky Mountain pi?on--juniper [Pinus edulis Engelm. and Juniperus spp.], Colorado plateau pi?on--juniper woodland, Rocky Mountain ponderosa pine (Pinus ponderosa P.& C. Lawson var. scopulorum), and southern Rocky Mountain juniper woodland and savanna). The overall accuracy of the model was >82%. Our most conservative model predicted that 14.4% of the four-corners region represented a high risk of peridomestic exposure to Y. pestis.     

Polymorphisms in the lcrV gene of Yersinia enterocolitica and their effect on plague protective immunity

Current efforts to develop plague vaccines focus on LcrV, a polypeptide that resides at the tip of type III secretion needles. LcrV-specific antibodies block Yersinia pestis type III injection of Yop effectors into host immune cells, thereby enabling phagocytes to kill the invading pathogen. Earlier work reported that antibodies against Y. pestis LcrV cannot block type III injection by Yersinia enterocolitica strains and suggested that lcrV polymorphisms may provide for escape from LcrV-mediated plague immunity. We show here that polyclonal or monoclonal antibodies raised against Y. pestis KIM D27 LcrV (LcrV(D27)) bind LcrV from Y. enterocolitica O:9 strain W22703 (LcrV(W22703)) or O:8 strain WA-314 (LcrV(WA-314)) but are otherwise unable to block type III injection by Y. enterocolitica strains. Replacing the lcrV gene on the pCD1 virulence plasmid of Y. pestis KIM D27 with either lcrV(W22703) or lcrV(WA-314) does not affect the ability of plague bacteria to secrete proteins via the type III pathway, to inject Yops into macrophages, or to cause lethal plague infections in mice. LcrV(D27)-specific antibodies blocked type III injection by Y. pestis expressing lcrV(W22703) or lcrV(WA-314) and protected mice against intravenous lethal plague challenge with these strains. Thus, although antibodies raised against LcrV(D27) are unable to block the type III injection of Y. enterocolitica strains, expression of lcrV(W22703) or lcrV(WA-314) in Y. pestis did not allow these strains to escape LcrV-mediated plague protective immunity in the intravenous challenge model.