Ginns C A, Benham M L, Adams L M, Whithear K G, Bettelheim K A, Crabb B S, Browning G F
Department of Veterinary Science, The University of Melbourne, Parkville, Victoria 3052, Australia.
Infect Immun. 2000 Mar;68(3):1535-41. doi: 10.1128/IAI.68.3.1535-1541.2000.
The E3 strain of E. coli was isolated in an outbreak of respiratory disease in broiler chickens, and experimental aerosol exposure of chickens to this strain induced disease similar to that seen in the field. In order to establish whether the virulent phenotype of this strain was associated with carriage of particular plasmids, four plasmid-cured derivatives, each lacking two or more of the plasmids carried by the wild-type strain, were assessed for virulence. Virulence was found to be associated with one large plasmid, pVM01. Plasmid pVM01 was marked by introduction of the transposon TnphoA, carrying kanamycin resistance, and was then cloned by transformation of E. coli strain DH5alpha. The cloned plasmid was then reintroduced by conjugation into an avirulent plasmid-cured derivative of strain E3 which lacked pVM01. The conjugant was shown to be as virulent as the wild-type strain E3, establishing that this plasmid is required for virulence following aerosol exposure. This virulence plasmid conferred expression of a hydroxamate siderophore, but not colicins, on both strain E3 and strain DH5alpha. Carriage of this plasmid was required for strain E3 to colonize the respiratory tracts of chickens but was not necessary for colonization of the gastrointestinal tract. However, the virulence plasmid did not confer virulence, or the capacity to colonize the respiratory tract, on strain DH5alpha. Thus, these studies have established that infection of chickens with E. coli strain E3 by the respiratory route is dependent on carriage of a conjugative virulence plasmid, which confers the capacity to colonize specifically the respiratory tract and which also carries genes for expression of a hydroxymate siderophore. These findings will facilitate identification of the specific genes required for virulence in these pathogens.
大肠杆菌E3菌株是在肉鸡呼吸道疾病暴发期间分离得到的，通过实验性气溶胶暴露将该菌株接种给鸡，所引发的疾病与在实际发病情况中观察到的相似。为了确定该菌株的毒力表型是否与特定质粒的携带有关，对四种质粒缺失衍生物进行了毒力评估，每种衍生物都缺失野生型菌株携带的两种或更多种质粒。结果发现毒力与一个大质粒pVM01有关。通过导入携带卡那霉素抗性的转座子TnphoA对质粒pVM01进行标记，然后通过转化大肠杆菌DH5α菌株进行克隆。接着通过接合作用将克隆的质粒重新导入缺乏pVM01的E3菌株无毒质粒缺失衍生物中。结果表明该接合子与野生型菌株E3的毒力相同，证实该质粒是气溶胶暴露后产生毒力所必需的。这种毒力质粒在E3菌株和DH5α菌株中都能使异羟肟酸铁载体表达，但不能使大肠杆菌素表达。E3菌株携带该质粒是其在鸡呼吸道中定殖所必需的，但在胃肠道定殖中并非必需。然而，毒力质粒并未赋予DH5α菌株毒力或在呼吸道定殖的能力。因此，这些研究表明，通过呼吸道途径用大肠杆菌E3菌株感染鸡取决于一种接合性毒力质粒的携带，该质粒赋予了特异性定殖呼吸道的能力，并且还携带了异羟肟酸铁载体表达所需的基因。这些发现将有助于鉴定这些病原体中毒力所需的特定基因。
