中国百日咳鲍特菌分离株的分子流行病学研究

发布时间：2018-05-22 11:56

本文选题：百日咳 + 抗原漂移　；参考：《河北医科大学》2009年硕士论文

【摘要】： 目的:百日咳是由百日咳鲍特菌感染所引起的一种急性呼吸道传染病,人群普遍易感,尤其以婴幼儿多见。我国全细胞百日咳疫苗(whole-cell pertussis vaccine,WPV )的使用有近50年的历史,而相关百日咳鲍特菌分子流行病学的研究资料较少,本实验对近50年我国收集的百日咳鲍特菌进行了分子水平研究,探讨我国分离百日咳鲍特菌的分子流行病学特点,以便更好的监控和预防百日咳。方法:对我国1953-2005年间分离的96株百日咳鲍特菌,进行了百日咳鲍特菌的血清型和脉冲场凝胶电泳(PFGE)分析以及其主要毒力因子的基因序列测序,包括百日咳毒素S1亚基(ptxS1)、百日咳粘附素(prn)、气管定居因子(tcfA)、凝集素2(fim2)和凝集素3(fim3);同时与我国的疫苗生产菌株(CS,P3S10和18530)和世界卫生组织(WHO)参考菌株18323株进行比较。结合菌株分离时间,分析不同年代流行菌株的基因是否存在抗原漂移现象,血清型分布特点和PFGE型别与基因型的关联。分析百日咳疫苗使用前后流行菌株的基因型和血清型的差异。结果:血清型分析显示96株临床菌株中共有fim2,fim3,fim23三个血清型,fim2和fim23血清型菌株主要分布在90年代以前,在90年代后出现了fim3型。基因序列分析结果显示五个抗原基因均存在多态性现象,prn基因包括6个基因型(prn1, prn2, prn3, prn7, prn10和prn11),ptxS1基因包括4个基因型(ptxS1A, ptxS1B, ptxS1D和ptxS1E),tcfA基因包括4个基因型(tcfA1, tcfA2, tcfA5和tcfA9),fim2基因包括2个基因型(fim2-1和fim2-2)和fim3基因包括3个基因型(fim3A, fim3B和fim3A*)。临床菌株中的prn基因型中prn1基因型最为常见,占81.3%,含prn2和prn3基因型菌株最初分别在2000和2003年分离获得。含ptxS1A基因型的菌株最初在1963年被分离,并且以后逐渐取代了在50年代的流行ptxS1B和ptxS1D基因型的菌株,到90年代以后所占比例已达到100%。研究结果显示5个基因组合(prn/pxS1/tcfA/fim2/fim3),即1/A/2/1/A, 1/B/2/1/A, 7/D/2/2/A, 2/A/2/1/B和3/A/2/1/A占所有基因组合的93.5%,这五个基因组合的菌株分别所占比例为60.4%、18.8%、8.3%、3.1%和2.1%。其中被称为旧型的基因组合1/B/2/1/A和7/D/2/2/A的菌株是50年代最流行的基因组合形式,与疫苗生产菌株基因组合一样。到了60年代以后,1/A/2/1/A基因组合的菌株数量增多,含有这种基因组合的细菌逐渐成为主要的流行菌株,2000年后出现了和3/A/2/1/A基因型的菌株和2/A/2/1/B基因型的菌株,后者是目前国外研究中比较流行百日咳菌株的基因组合形式。通过PFGE分析显示26个PFGE谱型被鉴定,不同的年代流行着不同PFGE谱型的菌株,BPCH2和BPCH3谱型是50年代比较流行菌株主要的谱型所占比例为52%,我国的疫苗生产菌株(CS和P3S10)为BPCH2谱型,疫苗生产菌株18530为BPCH25谱型,在1963-2003年收集的菌株中有49株为BPCH1谱型,占所有研究菌株数量的51%,而此谱型的菌株主要都是prn1/pxS1A基因型的菌株,BPCH6谱型(拥有和BPCH1谱型相同的基因组合形式)的菌株分离于1999年,并且在1999-2003年间所收集的菌株中所占比例达30.8%,聚类分析显示PFGE谱型中ptxS1基因型从ptxA1B向ptxS1A基因型的过渡表现在聚类图中。 PFGE谱型与抗原基因组合prn/pxS1/tcfA/fim2/fim3相关联,6个最常见的PFGE谱型中,除了BPCH1谱型中93.9%的菌株为基因组合形式1/A/2/1/A,其它的谱型100%对应各自不同的基因组合。在1953-1979年间和1980-2005年间,血清型有变化(P=0.0001);菌株ptxS1基因型也有变迁(P=0.001)。结论:我国百日咳鲍特菌处于不断的进化中,血清型和主要抗原基因(ptxS1)发生了漂移,在不同的时期流行着以不同的抗原基因型和PFGE谱型为主的菌株;目前流行百日咳菌株与疫苗生产菌株(CS、18530和P3S10)存在基因水平的差异。
[Abstract]:Objective: pertussis is an acute respiratory infection caused by the infection of Bordetella pertussis. The population is generally susceptible, especially in infants. The use of whole-cell pertussis vaccine (WPV) in our country has a history of nearly 50 years, and the molecular epidemiology of pertussis bauter is less. The molecular level of bautusis pertussis collected in China in the last 50 years was studied. The molecular epidemiological characteristics of isolated Bordetella pertussis in China were discussed in order to better monitor and prevent pertussis.
Methods: 96 strains of Bordetella pertussis isolated in China in the past 1953-2005 years were analyzed by serotype and pulsed field gel electrophoresis (PFGE) and the sequence of its main virulence factors, including pertussis toxin S1 subunit (ptxS1), pertussis adhesion (PRN), trachea settlement factor (tcfA), agglutinin 2 (fim2) and agglutination. Element 3 (fim3) was compared with 18323 strains of vaccine strains (CS, P3S10 and 18530) and WHO (WHO) in China.
According to the isolation time of the strains, it was analyzed whether the genes of the epidemic strains in different ages had the phenomenon of antigen drift, the characteristics of serotype distribution and the association between the PFGE type and genotype. The difference of genotypes and serotypes of the epidemic strains before and after the use of pertussis vaccine was analyzed.
Results: the serotype analysis showed that 96 clinical strains were fim2, fim3, fim23 three serotypes, fim2 and fim23 serotype strains were mainly distributed before 90s, and fim3 type appeared after 90s.
The gene sequence analysis showed that the five antigen genes were polymorphic, and the PRN gene consisted of 6 genotypes (prn1, prn2, prn3, prn7, prn10 and prn11). The ptxS1 gene included 4 genotypes (ptxS1A, ptxS1B, ptxS1D and ptxS1E), and 4 genes including 2 genotypes. 1 and fim2-2) and fim3 genes include 3 genotypes (fim3A, fim3B and fim3A*).
In the PRN genotypes of clinical strains, the prn1 genotype is the most common, accounting for 81.3%. The strains containing prn2 and prn3 genotype were first isolated in 2000 and 2003 respectively. The strains containing the ptxS1A genotype were first isolated in 1963, and then gradually replaced the strains of the prevalent ptxS1B and ptxS1D genotypes in 50s. The example has reached 100%.
The results showed that 5 gene combinations (prn/pxS1/tcfA/fim2/fim3), that is, 1/A/2/1/A, 1/B/2/1/A, 7/D/2/2/A, 2/A/2/1/B and 3/A/2/1/A accounted for 93.5% of all gene combinations, and the proportion of the five gene combinations was 60.4%, 18.8%, 8.3%, 3.1% and 2.1%., which was called the old type of 1/B/2/1/A and 7/D/2/2/A was 50. The most popular form of gene combination in the years was the same as the gene combination of vaccine producing strains. After 60s, the number of 1/A/2/1/A gene combinations increased. Bacteria containing this combination became the main epidemic strains. After 2000, the strains and the 3/A/2/1/A genotypes and 2/A/2/1/B genotypes appeared. The latter was the latter. At present, the genetic composition of pertussis strains is relatively popular in overseas studies.
PFGE analysis showed that 26 PFGE spectrum types were identified, and different PFGE strains were prevalent in different ages. BPCH2 and BPCH3 were the most popular strains in 50s, the proportion of the major strains was 52%. The vaccine producing strains (CS and P3S10) in our country were BPCH2 spectrum, vaccine producing strain 18530 was BPCH25 spectrum, and the bacteria collected in 1963-2003 years were collected. 49 of the strains were BPCH1, accounting for 51% of all the research strains, and the strains were mainly prn1/pxS1A genotypes. The strains of BPCH6 spectrum (with the same genetic combination of BPCH1) were isolated in 1999, and the proportion of the strains collected in 1999-2003 years was 30.8%. Cluster analysis showed that PFGE The transition of ptxS1 genotype from ptxA1B to ptxS1A genotype is shown in the cluster map.
The PFGE spectrum is associated with the antigen gene combination of prn/pxS1/tcfA/fim2/fim3. Among the 6 most common PFGE profiles, 93.9% of the strains in the BPCH1 spectrum are 1/A/2/1/A in the form of gene combination, and the other spectral types 100% correspond to their respective gene combinations.
In 1953-1979 and 1980-2005 years, the serotypes were changed (P=0.0001) and the strain ptxS1 genotypes were also changed (P=0.001). Conclusion: in our country, bauter pertussis is in continuous evolution, the serotype and the main antigen gene (ptxS1) are drifting, and the strains with different antigen genotypes and PFGE profiles are prevalent in different time periods. The prevalence of pertussis strains is different from that of vaccine producing strains (CS, 18530 and P3S10).
【学位授予单位】：河北医科大学
【学位级别】：硕士
【学位授予年份】：2009
【分类号】：R181.3

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