Pasloske B L, Joffe A M, Sun Q, Volpel K, Paranchych W, Eftekhar F, Speert D P
Department of Biochemistry, University of Alberta, Edmonton, Canada.
Infect Immun. 1988 Mar;56(3):665-72. doi: 10.1128/iai.56.3.665-672.1988.
Five isolates of Pseudomonas aeruginosa (CD2, CD3, CD4, CD5, and CD10) from a patient with cystic fibrosis were examined with regard to several genotypic and phenotypic characteristics to determine whether the patient was colonized with one or several distinct strains. Isolates CD2, CD3, and CD4 were obtained from a single sputum sample, and CD5 and CD10 were obtained 1 and 2 years later, respectively. On the basis of colonial morphology, serotyping, and antibiograms, the five isolates appeared to be different strains. However, Southern blot analysis with a 1.2-kilobase DNA probe containing the P. aeruginosa PAK pilin gene indicated that all five strains were identical at that genetic locus. The pilin genes of the five isolates were cloned and sequenced at the nucleotide level and found to be identical. Southern blot analysis with a probe from a separate region of the P. aeruginosa chromosome, a 741-base-pair PstI-NruI DNA fragment adjacent to the exotoxin A gene, also revealed genetic identity among these five clinical isolates. On this basis, it was concluded that this patient was colonized with a single strain of P. aeruginosa and that the strain had remained genetically stable over a period of 2 years. The predicted pilin sequence of the CD isolates was almost identical to that of strain PA103 (97% homology) and serologically related to PAO pilin, with which it shared 80% homology. No immunological cross-reactivity was detected between the CD and PAK pilins, which shared the least homology (62%) among the four pilins considered in this study. Although all five CD isolates contained identical pilin genes, three had acquired mutations which prevented normal expression of the pilus system. CD3 was a putative regulatory mutant which was unable to produce normal amounts of pilin, and CD4 and CD10 were putative assembly mutants which produced normal amounts of pilin but were unable to assemble the pilin subunit into intact pili.
对一名囊性纤维化患者的五株铜绿假单胞菌（CD2、CD3、CD4、CD5和CD10）进行了多种基因型和表型特征检测，以确定该患者是被一种还是几种不同菌株定植。CD2、CD3和CD4分离株取自同一个痰液样本，CD5和CD10分别在1年和2年后获得。基于菌落形态、血清分型和抗菌谱，这五株分离株似乎是不同的菌株。然而，用一个包含铜绿假单胞菌PAK菌毛蛋白基因的1.2千碱基DNA探针进行的Southern印迹分析表明，所有五株菌株在该基因位点是相同的。对这五株分离株的菌毛蛋白基因进行了克隆并在核苷酸水平测序，发现它们是相同的。用来自铜绿假单胞菌染色体另一个区域的探针，即与外毒素A基因相邻的一个741碱基对的PstI - NruI DNA片段进行Southern印迹分析，也揭示了这五株临床分离株之间的基因一致性。基于此，得出结论：该患者被单一菌株的铜绿假单胞菌定植，并且该菌株在2年时间内保持了基因稳定性。CD分离株的预测菌毛蛋白序列与PA103菌株几乎相同（97%同源性），并且在血清学上与PAO菌毛蛋白相关，与之共享80%的同源性。在本研究中所考虑的四种菌毛蛋白中，CD和PAK菌毛蛋白之间的同源性最低（62%），未检测到它们之间的免疫交叉反应。尽管所有五株CD分离株都含有相同的菌毛蛋白基因，但其中三株发生了突变，阻止了菌毛系统的正常表达。CD3是一个假定的调节突变体，无法产生正常量的菌毛蛋白，而CD4和CD10是假定的组装突变体，能产生正常量的菌毛蛋白，但无法将菌毛蛋白亚基组装成完整的菌毛。
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