Molecular serotyping of Klebsiella species isolates by restriction of the amplified capsular antigen gene cluster
Molecular serotyping of Klebsiella species isolates by restriction of the amplified capsular antigen gene cluster
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DOI：
10.1128/jcm.42.8.3388-3398.2004
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发表时间：
2004-08-01
影响因子：
9.4
通讯作者：
Grimont, PAD
中科院分区：
文献类型：
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作者：
Brisse, S;Issenhuth-Jeanjean, S;Grimont, PAD
目前工作的目的是开发一种分子方法，无需使用抗血清即可确定克雷伯氏菌分离株的荚膜血清型。 PCR 扩增荚膜抗原基因簇 (cps)，然后用限制性内切酶 HincII 消化（cps PCR-限制性片段长度多态性 [RFLP] 分析）。代表 77 种已知 K 血清型的 224 个菌株获得的谱（C 模式）显示有 3 至 13 个片段，大小范围为 0.2 至 4.4 kb。总共获得了97种不同的C模式；两次测试的 61 对样品中 100% 显示出可重现的 C 模式。 C型是K型特异性的；即，任何 K 血清型的 C 模式都与所有其他 K 血清型的 C 模式不同，唯一的例外是血清型 K22 和 K37，已知它们会发生交叉反应。对于至少包含两个菌株的 17 K 型中的 12 种，在具有相同 K 血清型的菌株之间发现了 C 型变异。因此，cps PCR-RFLP 分析比经典 K 血清分型具有更高的区分能力。在具有给定 K 型的菌株中观察到 C 模式同一性，这些菌株是相隔多年且来自不同来源的，表明 C 模式稳定性。由于 PCR 扩增不成功，只有 4.5% 的菌株无法分型（而 8% 至 23% 的菌株通过经典 K 血清分型无法分型）。分析的四种非包膜菌株中的三种显示出可识别的 C 模式。 22 个最近的肺炎克雷伯菌临床分离株中的 18 个（82%）的 K 血清型可以从其 C 模式推断出来。总之，cps PCR-RFLP 分析可以确定 K 血清型，同时比经典血清分型更容易执行且更具区分性。
The objective of the present work was to develop a molecular method that would enable determination of the capsular serotypes of Klebsiella isolates without the use of antiserum. PCR amplification of the capsular antigen gene cluster (cps) was followed by digestion with the restriction enzyme HincII (cps PCR-restriction fragment length polymorphism [RFLP] analysis). The profiles (C patterns) obtained for 224 strains representing the 77 known K serotypes showed 3 to 13 fragments ranging in size from 0.2 to 4.4 kb. A total of 97 distinct C patterns were obtained; 100% of 61 pairs of samples tested twice showed reproducible C patterns. The C patterns were K-type specific; i.e., the C pattern(s) of any K serotype was distinct from the C patterns of all other K serotypes, with the only exceptions being serotypes K22 and K37, which are known to cross-react. For 12 of 17 K types for which at least two strains were included, C-pattern variations were found among strains with the same K serotype. Therefore, cps PCR-RFLP analysis has a higher discriminatory power than classical K serotyping. C-pattern identity was observed among strains with a given K type that were collected many years apart and from distinct sources, indicating C-pattern stability. Only 4.5% of the strains were nontypeable, because of unsuccessful PCR amplification (whereas 8 to 23% are nontypeable by classical K serotyping). Three of four noncapsulated strains analyzed showed recognizable C patterns. The K serotypes of 18 (82%) of 22 recent Klebsiella pneumoniae clinical isolates could be deduced from their C patterns. In conclusion, cps PCR-RFLP analysis allows determination of the K serotype, while it is easier to perform and more discriminatory than classical serotyping.