Clemans D L, Marrs C F, Patel M, Duncan M, Gilsdorf J R
Department of Pediatrics and Communicable Diseases, University of Michigan Medical School, Ann Arbor 48109-2029, USA.
Infect Immun. 1998 Feb;66(2):656-63. doi: 10.1128/IAI.66.2.656-663.1998.
Adherence of Haemophilus influenzae to epithelial cells plays a central role in colonization and is the first step in infection with this organism. Pili, which are large polymorphic surface proteins, have been shown to mediate the binding of H. influenzae to cells of the human respiratory tract. Earlier experiments have demonstrated that the major epitopes of H. influenzae pili are highly conformational and immunologically heterogenous; their subunit pilins are, however, immunologically homogenous. To define the extent of structural variation in pilins, which polymerize to form pili, the pilin genes (hifA) of 26 type a to f and 16 nontypeable strains of H. influenzae were amplified by PCR and subjected to restriction fragment length polymorphism (RFLP) analysis with AluI and RsaI. Six different RFLP patterns were identified. Four further RFLP patterns were identified from published hifA sequences from five nontypeable H. influenzae strains. Two patterns contained only nontypeable isolates; one of these contained H. influenzae biotype aegyptius strains F3031 and F3037. Another pattern contained predominantly H. influenzae type f strains. All other patterns were displayed by a variety of capsular and noncapsular types. Sequence analysis of selected hifA genes confirmed the 10 RFLP patterns and showed strong identity among representatives displaying the same RFLP patterns. In addition, the immunologic reactivity of pili with antipilus antisera correlated with the groupings of strains based on hifA RFLP patterns. Those strains that show greater reactivity with antiserum directed against H. influenzae type b strain M43 pili tend to fall into one RFLP pattern (pattern 3); while those strains that show equal or greater reactivity with antiserum directed against H. influenzae type b strain Eagan pili tend to fall in a different RFLP pattern (pattern 1). Sequence analysis of representative HifA pilins from typeable and nontypeable H. influenzae identified several highly conserved regions that play a role in bacterial pilus assembly and other regions with considerable amino acid heterogeneity. These regions of HifA amino acid sequence heterogeneity may explain the immunologic diversity seen in intact pili.
流感嗜血杆菌对上皮细胞的黏附在定植过程中起核心作用，是该菌感染的第一步。菌毛是一种大型多态性表面蛋白，已被证明可介导流感嗜血杆菌与人呼吸道细胞的结合。早期实验表明，流感嗜血杆菌菌毛的主要表位具有高度构象性且在免疫上具有异质性；然而，其亚基菌毛蛋白在免疫上是同质的。为了确定聚合形成菌毛的菌毛蛋白的结构变异程度，通过聚合酶链反应（PCR）扩增了26株a至f型及16株不可分型流感嗜血杆菌的菌毛蛋白基因（hifA），并用AluI和RsaI进行限制性片段长度多态性（RFLP）分析。鉴定出六种不同的RFLP模式。从已发表的五株不可分型流感嗜血杆菌菌株的hifA序列中又鉴定出四种RFLP模式。两种模式仅包含不可分型菌株；其中一种包含埃及生物型流感嗜血杆菌菌株F3031和F3037。另一种模式主要包含f型流感嗜血杆菌菌株。所有其他模式由多种荚膜型和非荚膜型展示。对选定hifA基因的序列分析证实了这10种RFLP模式，并显示出具有相同RFLP模式的代表菌株之间具有高度一致性。此外，菌毛与抗菌毛抗血清的免疫反应性与基于hifA RFLP模式的菌株分组相关。那些与针对b型流感嗜血杆菌菌株M43菌毛的抗血清反应性更强的菌株往往属于一种RFLP模式（模式3）；而那些与针对b型流感嗜血杆菌菌株伊根菌毛的抗血清反应性相同或更强的菌株往往属于另一种不同的RFLP模式（模式1）。对可分型和不可分型流感嗜血杆菌代表性HifA菌毛蛋白的序列分析确定了几个在细菌菌毛组装中起作用的高度保守区域以及其他具有相当大氨基酸异质性的区域。HifA氨基酸序列异质性的这些区域可能解释了完整菌毛中所见的免疫多样性。
