张家界市首例C群流脑死亡病例及其密接人群中分离的脑膜炎奈瑟菌实验室分析和PFGE分型

目的了解张家界市首例C群流脑死亡病例及其密切接触人群中分离到的11株脑膜炎奈瑟菌的病原学特征及其流行关系。方法经培养及生化鉴定后,对菌株进行血清学及聚合酶链反应（PCR）鉴定分群,最后采用最低抑菌浓度（MIC）琼脂稀释法进行药敏试验;脉冲场凝胶电泳对菌株进行PFGE分型分析。结果通过生化、血清学和PCR实验共鉴定到9株C群脑膜炎奈瑟菌和2株W135群脑膜炎奈瑟菌,药敏试验中所有菌株对青霉素、氨苄西林、米诺环素、头孢曲松、头孢噻肟、氯霉素、阿奇霉素、美罗培南和利福平全部敏感;对复方磺胺甲口恶唑全部耐药;对环丙沙星和左氧氟沙星部分耐药,PFGE结果显示11株菌株共分为两个带型,其中9株C群脑膜炎奈瑟菌菌株带型完全相同。结论 C群和W135群可能成为新的流脑流行群引起疾病,分离的菌株对大部分抗生素仍较敏感,但要注意耐药趋势,造成该病例死亡的病原菌为C群脑膜炎奈瑟菌,与其密接同学中分离到的C群脑膜炎奈瑟菌的PFGE分型呈现高度一致性,提示为同一克隆群。     

湖南省C群及W135群脑膜炎奈瑟菌的流行病学及病原学特征分型

目的了解湖南省C群和W135群脑膜炎奈瑟菌的流行病学及病原学特征。方法收集2006—2016年湖南省流行性脑脊髓膜炎患者的血液或脑脊液、患者密切接触者以及健康人群的咽拭子标本中分离到的脑膜炎奈瑟菌菌株,进行生化检测、血清学分群。选取其中的C群和W135群部分菌株进行药敏试验,并采用脉冲场凝胶电泳(PFGE)分型以及多位点序列分型(MLST)方法对菌株进行分子分型,分析其流行病学特征。结果经生化和血清学确认后,选取22株C群脑膜炎奈瑟菌和9株W135群脑膜炎奈瑟菌进行药敏试验,结果显示对大部分检测抗菌药物全部敏感,但对复方磺胺甲口恶唑C群菌株全部耐药,而W135群菌株的耐药率为55.56%,两者比较差异有统计学意义(χ~2=7.61,P=0.006)。经PFGE分型后,22株C群脑膜炎奈瑟菌共分为5种PFGE带型,其中5株HNC-01带型和13株HNC-02带型属同一亚型;2006年湖南省第1例C群患者分离菌株的PFGE带型为HNC-02,与2012、2013年患者以及患者密切接触者分离菌株的图谱完全一致,与带型为HNC-01的2008、2010、2013年的患者分离菌株仅有一个带型的差异,均属于优势带型。9株W135群脑膜炎奈瑟菌PFGE分型后共分为2个带型,其中首例患者与2013、2016年患者分离菌株的带型一致,均为HNW-01型。选取其中优势带型菌株经MLST后,结果 C群脑膜炎奈瑟菌为ST4821型,W135群脑膜炎奈瑟菌为ST11型,均属于脑膜炎奈瑟菌的高致病性克隆群。结论湖南省C群流脑和W135群流脑自首例病例出现后,各自都成为了该群病例的优势流行克隆群,C群流脑近年有减少态势,但出现了新的流行型别;W135群自2012年起成为我省新的流脑流行株,其优势菌株与国际上侵袭性的W群分型一致,可能引起新的大流行,应及时制定相应的防控政策。     

合肥市W135群脑膜炎奈瑟菌的分子分型研究

目的利用脉冲场凝胶电泳技术(pulsed-field gel electrophoresis,PFGE)对合肥地区2012年分离到的W135群脑膜炎奈瑟菌进行分型,探讨其同源性。方法 1株疑似流脑病例的血液及3个疑似病例的密切接触者咽拭子进行脑膜炎奈瑟菌的培养分离,对分离的菌株进行生化试验、血清学分群鉴定、PCR扩增和PFGE鉴定试验。结果合肥地区分离出11株W135群脑膜炎奈瑟菌,11株W135群脑膜炎奈瑟菌经PFGE分子分型可以分为合肥1到合肥5型,分别是合肥1型4株,合肥2型1株,合肥3型2株,合肥4型1株和合肥5型3株。结论合肥地区的11株W135群脑膜炎奈瑟菌分为5个基因型。     

江西省2005年脑膜炎奈瑟菌带菌调查及C群脑膜炎奈瑟菌脉冲场凝胶电泳分析

目的了解江西省2005年健康人群脑膜炎奈瑟菌(Nm)带菌状况,为预测疫情和制定防治措施提供依据。方法采集健康人群和流行性脑脊髓膜炎(流脑)病人密切接触者的咽拭子标本进行Nm培养和鉴定,C群Nm菌株进行脉冲场凝胶电泳(PFGE)分子分型。结果健康人群1 441份咽拭子标本中分离到A群Nm2株,B群25株,W135群2株。从37份流脑病人的密切接触者咽拭子标本中分离到C群Nm19株,经PFGE分析18株为相同的脉冲场电泳型(AH1)。结论江西省2005年健康人群带菌以B群为主,C群带菌率上升,PFGE分析为相同的菌株型别。     

2013年湖南省一起B群流脑聚集性疫情菌株分子分型分析

目的了解湖南湘西地区B群流脑聚集性疫情中病例及其密切接触人群中分离到的B群脑膜炎奈瑟菌的分子分型特征及其流行关系。方法对病例的血液标本及其密切接触者的咽拭子标本进行分离、培养及生化鉴定,确认为脑膜炎奈瑟菌后,对菌株进行血清学分群和药敏试验;采用多位点序列分型(multilocus sequence typing,MLST)以及脉冲场凝胶电泳(pulsed-field gel electrophoresis,PFGE)方法对菌株进行分子分型。结果共分离到3株B群脑膜炎奈瑟菌,均对复方磺胺甲恶唑(SXT)耐药,对青霉素(PEN)和氨苄青霉素(AMP)的药敏结果为中介度,对其他9种抗生素米诺环素(MIN)、头孢曲松(CRO)、头孢噻肟(CTX)、利福平(RIF)、阿奇霉素(AZM)、氯霉素(CHL)、环丙沙星(CIP)、左氧氟沙星(LVX)和美洛培南(MEM)均敏感,PFGE结果显示3株菌株为同一带型,MLST结果显示3株菌株均为ST-4821 complex高致病克隆群。结论该起湖南湘西地区B群流脑聚集性疫情中病人与其密切接触者中分离到的B群脑膜炎奈瑟菌在分子分型实验中呈现高度一致性,提示为同一来源。流脑菌在湖南省的菌群已经发生变迁,带致病性克隆群的B群菌株可能成为优势菌株,引起流脑的暴发流行。     

北京地区2005年流行性脑脊髓膜炎病原学监测

目的对2005年北京地区流行性脑脊髓膜炎（流脑）进行病原学监测.方法对临床流脑病例的血液、脑脊液标本进行特异性核酸片段检测,对鉴定阳性的菌株进行脉冲场凝胶电泳（PFGE）和多位点序列（MLST）分析.结果在7份血液和5份脑脊液标本中检测到脑膜炎奈瑟菌的核酸片段.共分离到105株脑膜炎奈瑟菌.A群和C群菌株的群内菌株PFGE图谱无差异条带.MLST分析结果表明北京地区A群脑膜炎奈瑟菌的序列类型主要是ST7,而C群主要是ST4821.结论北京地区目前流脑病例主要由序列类型为ST7的A群和ST4821的C群脑膜炎奈瑟菌引起.     

1例C群流脑病例的密切接触者带菌情况调查

[目的]了解1例由C群脑膜炎奈瑟氏菌所致流行性脑脊髓膜炎病例的密切接触者病原菌携带情况及所携带病原菌的血清分群情况,为制定流脑综合防控措施提供依据。[方法]对采集到的151份咽拭子标本进行菌株分离、鉴定、血清学分群。[结果]共检出脑膜炎奈瑟氏菌49株,其中21株为C群,23株为不可分群,3株为B群,1株为W135群,另外1株为多价Ⅲ中的29E。[结论]经过对密切接触者带菌率的调查,我市可能存在C群流脑流行和暴发的隐患。     

四川省B群脑膜炎奈瑟菌脉冲场凝胶电泳分型分析

[目的]对四川省B群脑膜炎奈瑟菌分离菌株进行脉冲场凝胶电泳(PFGE)分析,了解四川省B群脑膜炎奈瑟菌菌株的分子流行病学特征.[方法]对健康人群和流行性脑脊髓膜炎病例密切接触者分离的55株B群脑膜炎奈瑟菌使用限制性内切酶NheI酶切、脉冲场凝胶电泳,电泳结果用BioNumerics软件进行聚类分析.[结果]55株B群脑膜炎奈瑟菌共分为33种PFGE型,呈现高度的多态性.其中健康带菌者的45株菌分为24个型,带型分布在年份和地区上没有关联,但局部地区的菌株呈现出PFGE型别一致的特征;10株病例的密切接触者分离的菌株分为9个型,无优势型别.[结论]四川省B群脑膜炎奈瑟菌菌株具有高度的遗传多样性,但在局部地区菜一时间段存在优势的PFGE型别的B群脑膜炎奈瑟菌菌株.     

江西省2005-2008年C群脑膜炎奈瑟菌分子分型分析

对2005-2008年江西省流脑暴发、常规监测中分离的79株C群脑膜炎奈瑟菌进行脉冲场凝胶电泳(PFGE)和多位点测序(MLST)分析,描述江西省C群脑膜炎奈瑟菌的菌型分布和变迁,从分子水平了解其流行特征.     

江西省2005-2008年C群脑膜炎奈瑟菌分子分型分析

对2005-2008年江西省流脑暴发、常规监测中分离的79株C群脑膜炎奈瑟菌进行脉冲场凝胶电泳(PFGE)和多位点测序(MLST)分析,描述江西省C群脑膜炎奈瑟菌的菌型分布和变迁,从分子水平了解其流行特征.     

中国流行性脑脊髓膜炎流行菌群变化趋势分析

目的研究中国(未包括香港、澳门特别行政区和台湾地区,下同)流行性脑脊髓膜炎(流脑)流行菌群的变迁趋势。方法对中国1956~2006年分离的1 819株流脑菌株的血清群构成进行分析。结果1956~2002年的922株流脑菌株中,A群占69.20%,B群占27.55%,C群占0.97%,其它群2.28%。2003~2006年的897株流脑菌株中,A群占35.90%,B群占29.21%,C群占23.97%,其它群占10.92%。C群流脑病人来源菌株比例由0.88%上升至48.67%,A群流脑病例菌株比例由80.4%下降至50.44%。健康人群鼻咽部携带C群流脑菌株比例由1.07%上升至20.41%,A群由58.33%下降至33.80%。结论中国流脑病人及健康人群携带菌株中,C群流脑菌株的比例呈上升趋势,流脑流行菌群正在发生从A群到C群的变化。     

可变数目串联重复序列技术用于中国C群脑膜炎奈瑟菌基因分型的研究

目的利用脑膜炎奈瑟菌基因组中可变数目串联重复序列（VNTR）特征,对中国C群脑膜炎奈瑟菌菌株进行基因分型.方法中国C群脑膜炎奈瑟菌菌株109株,选择脑膜炎奈瑟菌DNA中4个VNTR位点,PCR扩增含有串联重复序列的DNA片段,选择每一个VNTR位点有差别的PCR产物进行测序,序列比对,测算串联重复序列的拷贝数.Bio-Rad Gel DocTM XR凝胶成像分析系统计算PCR产物DNA片段的碱基含量,换算成串联重复数;对109株菌株4个位点的串联重复序列拷贝数进行聚类分析,依据聚类分析结果进行基因分型,并将VNTR基因分型结果与脉冲场凝胶电泳基因分型（PFGE）结果进行比较.结果109株C群脑膜炎奈瑟菌菌株分为22个VNTR基因型,同一暴发来源的菌株具有相同VNTR特征;VNTR基因分型方法与PFGE基因分型具有相关关系.结论应用VNTR技术可以对中国C群脑膜炎奈瑟菌进行基因分型和分子流行病学方面的研究,VNTR基因分型可较好地应用于追溯流行性脑脊髓膜炎暴发传染源.     

Molecular surveillance of Neisseria meningitidis capsular switching in Portugal, 2002-2006

Neisseria meningitidis capsular switching has been reported in several countries. In order to establish the genetic relationship within group B and C strains expressing subtypes 2a or 2b, and to evaluate whether C to B capsular switching occurred in Portugal, 64 meningococci (56 serogroup C and 8 serogroup B) isolated from invasive meningococcal disease were typed using molecular methods. The studied phenotypes, 2b:P1.5,2 and 2a:P1.5-1,10-8, were the most frequent among serogroup C, but were uncommon among serogroup B strains. The multi-locus sequence typing (MLST) allelic profile and the pulsed-field gel electrophoresis (PFGE) fingerprints showed that seven serogroup B strains were genotypically identical to C strains, suggesting that capsular switching occurred. Active laboratory surveillance to find evidence of capsule switching is a now priority as MenC was introduced in the Portuguese vaccination schedule in January 2006.     

Antimicrobial susceptibility, serotype and genotype distribution of meningococci in Portugal, 2001-2002

One hundred and eighteen Neisseria meningitidis isolates were recovered from patients with invasive meningococcal disease in Portugal, over one year. Our study was undertaken to evaluate antimicrobial susceptibility, serogroup, serotype and genotype of isolates. One quarter (24.6%) of the isolates showed moderate resistance to penicillin and 47.4% were resistant to sulphadiazine. The two most common serosubtypes were C:2b:P1.5,2 (31.3%) and B:4:P1.15 (3.4%). Half (53.6%) of the isolates with moderate resistance to penicillin were phenotype C:2b:P1.5,2 (n=14), C:2b:P1.2 (n=1) or C:2b:NST (n=1); Pulsed-field gel electrophoresis (PFGE) showed that all these isolates were genetically related. Multilocus sequence typing (MLST) analysis of representative clones from each PFGE pattern showed the predominance of the ST-8 complex/cluster A4 among N. meningitidis with moderate resistance to penicillin. This clonal complex has been principally found in Southern Europe. The apparent emergence and dissemination of the hypervirulent ST-8 complex/cluster A4 among serogroup C strains increases the need for a continued surveillance of antimicrobial susceptibility of meningococci and of genotypic markers in Portugal.     

MLEE and PFGE characterization of Neisseria meningitidis serogroup C and B isolated in the Slovak Republic in 1998

In the Slovak Republic the incidence and mortality of invasive meningococcal disease increased after 1995 when the new meningococcal clone of Neisseria meningitidis C:2a:P1.2,P1.5, ET-1.5/37 emerged. The new clone spread between 1995 and 1998 throughout the whole country. Morbidity of invasive meningococcal disease was 1.6/100,000 of the population and fatality reached the highest level of 23% in the Slovak Republic in 1998. The new clone caused a new emergent epidemiological and clinical situation. The occurrence of invasive meningococcal disease caused by this clone has continually risen since 1995. In 1998 72% of all diseases in Slovakia were caused by serogroup C. The emerging clone C:2a:Pl.2,P1.5 represented 74% of the serogroup C isolates. Clonality and genetic diversity of 15 selected meningococcal strains causing invasive meningococcal disease was compared by multilocus enzyme electrophoresis (MLEE) and DNA macrorestriction analysis by pulsed-field gel electrophoresis (PFGE). The strains of serogroup C and B were isolated in all regions of Slovakia in 1998. The majority of isolates belong to hypervirulent clone ET-15 as determined by MLEE. By PFGE a higher degree of diversity was observed.     

Usefulness of the DNA-fingerprinting pattern and the multilocus enzyme electrophoresis profile in the assessment of outbreaks of meningococcal disease.

The objective of the study was to assess whether genotypic characterization by means of DNA-fingerprinting pattern (DFP) and multilocus enzyme electrophoresis (MEE) profile as compared to phenotypic characterization would improve the differentiation of Neisseria meningitidis strains associated with outbreaks from strains associated with sporadic cases of meningococcal disease. In addition, the differentiation of serogroup C carrier strains from those associated with an outbreak of serogroup C meningococcal disease was investigated. A total of 118 N. meningitidis strains were available for the study: 59 from patients involved in outbreaks of meningococcal disease (2 serogroup B and 2 serogroup C), 37 patients considered to be sporadic cases and 22 serogroup C carrier strains. Among the 59 strains from patients involved in outbreaks the 4 strains isolated from the patient registered as the first in each outbreak were designated the index strains. Among the remaining 55 outbreak strains 52 were either DFP-identical or DFP-indistinguishable when compared with the one relevant out of the 4 index strains. This was only the case for 17 of the 37 strains isolated from sporadic cases caused by the same serogroup of meningococci during the outbreak periods, and 5 of the 22 meningococcal strains isolated from healthy carriers. Among the 56 (52 + 4) DFP-identical or DFP-indistinguishable outbreak strains 5 different electrophoretic types were identified by MEE. Among 59 assumed outbreak strains a total of 4 were identified as genotypically distinct. Among the 37 mainly DFP-indistinguishable or DFP-different strains from sporadic cases 17 different ETs were identified, and among the 22 mainly DFP-different carrier strains 13 different ETs were identified. Two strains among those selected from sporadic cases were identical to the outbreak strain. None of the local serogroup C carrier strains isolated during the outbreak of serogroup C disease were identical to the outbreak strain. Both DNA-fingerprinting and MEE improved the differentiation of meningococci when compared with phenotypic characterization. The results indicate that tracing a virulent strain within a open group of contacts is irrelevant.     

Outbreak of Neisseria meningitidis, Edmonton, Alberta, Canada

From December 1999 to April 2001, the greater Edmonton region had 61 cases of invasive meningococcal infection, two fatal. The outbreak was due to Neisseria meningitidis serogroup C, electrophoretic type 15, serotype 2a. Analysis of the strains showed that 50 of 56 culture-confirmed cases were due to a single clone and close relatives of this clone. This strain had not been previously identified in the province of Alberta dating back to January 1997.