Unbiased homeologous recombination during pneumococcal transformation allows for multiple chromosomal integration events

The rapid spread of antimicrobial resistance and vaccine escape in the opportunistic human pathogen Streptococcus pneumoniae can be largely attributed to competence-induced transformation. To better understand the dynamics of competence-induced transformation, we studied this process at the single-cell level. We show that within isogenic populations, all cells become naturally competent and bind exogenous DNA. In addition, we find that transformation is highly efficient and that the chromosomal location of the integration site or whether the transformed gene is encoded on the leading or lagging strand has limited influence on recombination efficiency. Indeed, we have observed multiple recombination events in single recipients in real-time. However, because of saturation of the DNA uptake and integration machinery and because a single stranded donor DNA replaces the original allele, we find that transformation efficiency has an upper threshold of approximately 50% of the population. Counterintuitively, in the presence of multiple transforming DNAs, the fraction of untransformed cells increases to more than 50%. The fixed mechanism of transformation results in a fail-safe strategy for the population as half of the population generally keeps an intact copy of the original genome. Together, this work advances our understanding of pneumococcal genome plasticity.

机会致病性人类病原体肺炎链球菌（Streptococcus pneumoniae）中抗菌药物耐药性与疫苗逃逸的快速传播，在很大程度上可归因于感受态诱导转化。为更好地解析感受态诱导转化的动力学机制，我们在单细胞水平对该过程展开了研究。研究结果显示，在同基因种群中，所有菌体均可进入自然感受态并结合外源DNA。此外，我们发现转化效率极高，且整合位点的染色体位置，或是待整合基因编码于前导链还是滞后链，对重组效率的影响均极为有限。我们还实时观测到，单个受体菌内可发生多起重组事件。然而，由于DNA摄取与整合系统存在饱和效应，且单链供体DNA会替换原始等位基因，我们发现转化效率存在约50%的种群上限阈值。与直觉相悖的是，当存在多种转化DNA时，未转化细胞的占比会升至50%以上。这种固定的转化机制为种群提供了一种安全保障策略：通常会有半数菌体保留原始基因组的完整拷贝。综上，本研究加深了我们对肺炎链球菌基因组可塑性的认知。