Barkoff Alex-Mikael, Guiso Nicole, Guillot Sophie, Xing Dorothy, Markey Kevin, Berbers Guy, Mertsola Jussi, He Qiushui
Department of Infectious Disease Surveillance and Control, National Institute for Health and Welfare, Turku, Finland.
National Centre of Reference of Whooping Cough and Other Bordetelloses, Pasteur Institute, Paris, France.
J Immunol Methods. 2014 Jun;408:142-8. doi: 10.1016/j.jim.2014.06.001. Epub 2014 Jun 10.
Despite extensive vaccinations, there have been pertussis epidemics in many countries including the Netherlands, the UK, Australia and the USA. During these epidemics Bordetella pertussis strains not producing the vaccine antigen pertactin (Prn) are emerging and increasing in numbers. However, methods for confirming PRN production of B. pertussis isolates are combined PCR or PCR-based sequencing tests and western blotting. Furthermore, data about production of pertussis toxin (PT) and filamentous hemagglutinin (FHA) of these isolates are scarce. Fimbriae (Fim) production is usually determined by agglutination and reported as serotype. In this study we developed an easy, accurate and rapid method for screening PT and FHA production. Methods for Prn and Fim production have been published earlier.
We analyzed altogether 109 B. pertussis strains, including 103 Finnish B. pertussis strains collected during 2006-2013, international strain Tohama I, French strains FR3496 (PT-negative), FR3693 (Prn-negative) and FR4624 (FHA-negative) and Fim-serotype reference strains S1 (producing only Fim2) and S3 (producing only Fim3). An indirect ELISA with whole bacterial cells as coating antigen was developed and used for rapid screening of the B. pertussis strains. Production of different antigens (PT, FHA, Prn, Fim2 and Fim3) was detected with specific monoclonal antibodies (mAbs).
From the 103 Finnish B. pertussis strains tested, all were positive for PT, FHA and Fim. Four were found negative for Prn, and they were isolated during 2011-2013.
The newly developed method proved to be useful and simple for rapid screening of different antigen production of B. pertussis isolates.
尽管进行了广泛的疫苗接种，但包括荷兰、英国、澳大利亚和美国在内的许多国家仍出现了百日咳疫情。在这些疫情期间，不产生疫苗抗原百日咳黏附素（Prn）的百日咳博德特氏菌菌株不断出现且数量增加。然而，确认百日咳博德特氏菌分离株Prn产生情况的方法是联合PCR或基于PCR的测序检测以及蛋白质印迹法。此外，关于这些分离株百日咳毒素（PT）和丝状血凝素（FHA）产生情况的数据很少。菌毛（Fim）的产生通常通过凝集反应确定并报告为血清型。在本研究中，我们开发了一种简便、准确且快速的方法来筛选PT和FHA的产生情况。Prn和Fim产生情况的方法已在之前发表。
我们共分析了109株百日咳博德特氏菌菌株，包括2006 - 2013年期间收集的103株芬兰百日咳博德特氏菌菌株、国际菌株Tohama I、法国菌株FR3496（PT阴性）、FR3693（Prn阴性）和FR4624（FHA阴性）以及Fim血清型参考菌株S1（仅产生Fim2）和S3（仅产生Fim3）。开发了一种以全菌细胞作为包被抗原的间接ELISA法，并用于快速筛选百日咳博德特氏菌菌株。使用特异性单克隆抗体（mAb）检测不同抗原（PT、FHA、Prn、Fim2和Fim3）的产生情况。
在测试的103株芬兰百日咳博德特氏菌菌株中，所有菌株的PT、FHA和Fim均为阳性。发现4株Prn为阴性，它们是在2011 - 2013年期间分离得到的。
新开发的方法被证明对于快速筛选百日咳博德特氏菌分离株的不同抗原产生情况是有用且简便的。
