Characterization of Streptococcus pneumoniae phage-like element SpnCI reveals an enhanced virulent phenotype in the acute invertebrate infection model Galleria mellonella

Phage-like elements are found in a multitude of streptococcal species, including pneumococcal strain Hungary19A-6 (SpnCI). The aim of our research was to investigate the role of phage-like element SpnCI in enhanced virulence and phenotypic modulation within Streptococcus pneumoniae. SpnCI was found to significantly enhance virulence within the invertebrate infection model Galleria mellonella. Infections with SpnCI led to a lower mean health score (1.6) and survival percentage (20%) compared to SpnCI null TIGR4 infections (3.85 mean health score and 50% survival). SpnCI remained integrated throughout growth, conferring greater sensitivity to UV irradiation. Change in transcriptional patterns occurred, including downregulation of operons involved with cell surface modelling in the SpnCI containing strain of TIGR4. Kanamycin-tagged SpnCI strain in Hungary19A-6 was inducible and isolated from lysate along with both annotated prophages. No phages were identified by PCR nor electron microscopy (EM) following induction of TIGR4 SpnCI∆strA suggesting helper-phage dependence for dissemination. EM of lysate showed typical siphoviridae morphology with an average capsid size of 60 nm. Two of sixty capsids were found to be smaller, suggesting SpnCI disseminates using a similar mechanism described for Staphylococcus aureus phage-like element SaPI. SpnCI from lysate infected capsule null strain T4R but was incapable of infecting the encapsulated TIGR4 strain suggesting that capsule impedes phage infection. Our work demonstrates that SpnCI can modulate virulence, UV susceptibility, alter transcriptional patterns, and furthermore, can disseminate via infection within pneumococcus. Further research is necessary to elucidate how SpnCI modulates virulence and what genes are responsible for the enhanced virulence phenotype. Comparison of two strains of Streptococcus pneumoniae that differ only by the presence or absence of phage-like chromosomal island SpnCI

类噬菌体元件（phage-like element）广泛分布于多种链球菌属（Streptococcus）物种中，包括肺炎链球菌（Streptococcus pneumoniae）菌株Hungary19A-6（SpnCI）。本研究旨在探究类噬菌体元件SpnCI在肺炎链球菌内增强毒力与表型调控的作用。研究发现，SpnCI可在无脊椎动物感染模型大蜡螟（Galleria mellonella）中显著增强菌株毒力。与缺失SpnCI的TIGR4菌株感染组（健康评分均值3.85、存活率50%）相比，携带SpnCI的菌株感染后健康评分均值更低（1.6）、存活率仅为20%。SpnCI在整个生长周期均保持整合状态，可使菌株对紫外线照射的敏感性显著升高。转录谱发生改变，包括携带SpnCI的TIGR4菌株中参与细胞表面重塑的操纵子出现下调。匈牙利19A-6菌株中带有卡那霉素（Kanamycin）标记的SpnCI可被诱导，并与两种注释前噬菌体一同从裂解液中分离得到。对TIGR4 SpnCI∆strA经诱导后，未通过聚合酶链式反应（polymerase chain reaction，PCR）与电子显微镜（electron microscopy，EM）检测到噬菌体，提示其传播依赖辅助噬菌体。裂解液的电子显微镜观察显示其具有典型的长尾噬菌体科（Siphoviridae）形态，衣壳平均直径为60 nm；60个衣壳中有2个尺寸更小，这表明SpnCI的传播机制与金黄色葡萄球菌（Staphylococcus aureus）类噬菌体元件SaPI的已报道机制相似。从裂解液中获取的SpnCI可感染荚膜缺失型菌株T4R，但无法感染有荚膜的TIGR4菌株，提示荚膜会阻碍噬菌体感染。本研究证实，SpnCI可调控菌株毒力、改变紫外线敏感性、重塑转录谱，且可通过在肺炎链球菌内的感染实现传播。后续仍需进一步研究以阐明SpnCI调控毒力的具体机制，以及哪些基因与增强毒力表型相关。本研究针对仅在是否携带类噬菌体染色体岛SpnCI这一差异的两株肺炎链球菌进行了比较分析。