Weiss A A, Hewlett E L, Myers G A, Falkow S
Infect Immun. 1983 Oct;42(1):33-41. doi: 10.1128/iai.42.1.33-41.1983.
Transposon Tn5 was used to isolate mutants of Bordetella pertussis. Strains with Tn5 insertions were screened for loss of virulence-associated factors, including filamentous hemagglutinin, hemolysin, and pertussis toxin. Several mutants deficient for hemolysin production were obtained. All produced dermonecrotic toxin, pertussis toxin, and filamentous hemagglutinin, but were found to vary with respect to adenylate cyclase production. One hemolysin mutant had no detectable adenylate cyclase activity; others had 0.6% or 16% wild-type activity, whereas a fourth seemed to be unaffected in terms of adenylate cyclase activity. Mutants deficient in the ability to hemagglutinate sheep erythrocytes were also isolated. These mutants either failed to synthesize or produced reduced amounts of three protein species of 200,000, 130,000, and 100,000 daltons, all of which reacted with antiserum to filamentous hemagglutinin. Pertussis toxin mutants were identified by screening culture supernatants for failure to induce a clustered growth pattern in Chinese hamster ovary cells, and identification was confirmed by the standard histamine-sensitizing assay in mice. These mutants will be useful to determine the relative contribution of each virulence factor to pathogenicity as well as to determine the identity of the antigens important in protective immunity.
转座子Tn5被用于分离百日咳博德特氏菌的突变体。筛选带有Tn5插入片段的菌株，以检测其与毒力相关因子的缺失情况，这些因子包括丝状血凝素、溶血素和百日咳毒素。获得了几个溶血素产生缺陷的突变体。所有突变体均产生皮肤坏死毒素、百日咳毒素和丝状血凝素，但发现它们在腺苷酸环化酶产生方面存在差异。一个溶血素突变体没有可检测到的腺苷酸环化酶活性；其他突变体具有0.6%或16%的野生型活性，而第四个突变体在腺苷酸环化酶活性方面似乎未受影响。还分离出了不能凝集绵羊红细胞的突变体。这些突变体要么无法合成，要么产生的三种分子量分别为200,000、130,000和100,000道尔顿的蛋白质数量减少，所有这些蛋白质都能与丝状血凝素抗血清发生反应。通过筛选培养上清液中是否未能在中国仓鼠卵巢细胞中诱导聚集生长模式来鉴定百日咳毒素突变体，并通过小鼠标准组胺致敏试验进行确认。这些突变体将有助于确定每种毒力因子对致病性的相对贡献，以及确定在保护性免疫中重要的抗原的身份。
