Department of Microbiology, National Taiwan University College of Medicine, Taipei, Taiwan.
PLoS One. 2013 Aug 2;8(8):e70092. doi: 10.1371/journal.pone.0070092. Print 2013.
Klebsiella pneumoniae is one of the major pathogens causing hospital-acquired multidrug-resistant infections. The capsular polysaccharide (CPS) is an important virulence factor of K. pneumoniae. With 78 capsular types discovered thus far, an association between capsular type and the pathogenicity of K. pneumoniae has been observed.
METHODOLOGY/PRINCIPAL FINDINGS: To investigate an initially non-typeable K. pneumoniae UTI isolate NTUH-K1790N, the cps gene region was sequenced. By NTUH-K1790N cps-PCR genotyping, serotyping and determination using a newly isolated capsular type-specific bacteriophage, we found that NTUH-K1790N and three other isolates Ca0507, Ca0421 and C1975 possessed a new capsular type, which we named KN2. Analysis of a KN2 CPS(-) mutant confirmed the role of capsule as the target recognized by the antiserum and the phage. A newly described lytic phage specific for KN2 K. pneumoniae, named 0507-KN2-1, was isolated and characterized using transmission electron microscopy. Whole-genome sequencing of 0507-KN2-1 revealed a 159 991 bp double-stranded DNA genome with a G+C content of 46.7% and at least 154 open reading frames. Based on its morphological and genomic characteristics, 0507-KN2-1 was classified as a member of the Myoviridae phage family. Further analysis of this phage revealed a 3738-bp gene encoding a putative polysaccharide depolymerase. A recombinant form of this protein was produced and assayed to confirm its enzymatic activity and specificity to KN2 capsular polysaccharides. KN2 K. pneumoniae strains exhibited greater sensitivity to this depolymerase than these did to the cognate phage, as determined by spot analysis.
CONCLUSIONS/SIGNIFICANCE: Here we report that a group of clinical strains possess a novel Klebsiella capsular type. We identified a KN2-specific phage and its polysaccharide depolymerase, which could be used for efficient capsular typing. The lytic phage and depolymerase also have potential as alternative therapeutic agents to antibiotics for treating K. pneumoniae infections, especially against antibiotic-resistant strains.
肺炎克雷伯菌是引起医院获得性多药耐药感染的主要病原体之一。荚膜多糖（CPS）是肺炎克雷伯菌的重要毒力因子。迄今为止，已发现 78 种荚膜类型，观察到荚膜类型与肺炎克雷伯菌的致病性之间存在关联。
方法/主要发现：为了研究最初无法分型的肺炎克雷伯菌 UTI 分离株 NTUH-K1790N，我们对 cps 基因区域进行了测序。通过 NTUH-K1790N cps-PCR 基因分型、血清分型和使用新分离的荚膜型特异性噬菌体的测定，我们发现 NTUH-K1790N 和其他三个分离株 Ca0507、Ca0421 和 C1975 具有一种新的荚膜型，我们将其命名为 KN2。KN2 CPS(-)突变体的分析证实了荚膜是抗血清和噬菌体识别的靶标。分离并表征了一种针对 KN2 肺炎克雷伯菌的新描述的裂解噬菌体，命名为 0507-KN2-1。通过透射电子显微镜。0507-KN2-1 的全基因组测序显示，其基因组为 15991bp 的双链 DNA，G+C 含量为 46.7%，至少有 154 个开放阅读框。根据其形态和基因组特征，0507-KN2-1 被归类为肌病毒科噬菌体家族的成员。进一步分析该噬菌体发现了一个编码多糖解聚酶的 3738bp 基因。该蛋白的重组形式被产生并进行了测定，以确认其对 KN2 荚膜多糖的酶活性和特异性。通过斑点分析，发现与相应噬菌体相比，KN2 肺炎克雷伯菌菌株对这种解聚酶的敏感性更高。
结论/意义：在这里，我们报告了一组临床菌株具有一种新型的肺炎克雷伯菌荚膜类型。我们鉴定了一种针对 KN2 的噬菌体及其多糖解聚酶，可用于有效的荚膜分型。溶菌噬菌体和解聚酶也有可能替代抗生素治疗肺炎克雷伯菌感染，特别是对抗生素耐药菌株。
