Dissemination of Tn916-Related Integrative and Conjugative Elements in Streptococcus pneumoniae Occurs by Transformation and Homologous Recombination in Nasopharyngeal Biofilms.
Dissemination of Tn916-Related Integrative and Conjugative Elements in Streptococcus pneumoniae Occurs by Transformation and Homologous Recombination in Nasopharyngeal Biofilms.
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DOI：
10.1128/spectrum.03759-22
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发表时间：
2023-03-13
影响因子：
3.7
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中科院分区：
文献类型：
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肺炎链球菌（或肺炎球菌）的多药耐药性仍然是一个全球性的挑战。在沙门氏菌中传播的一类重要的抗生素耐药决定因子。在一些实施方案中，与肺炎链球菌相关的整合和接合元件是>20-kb Tn 916相关的整合和接合元件（ICE），例如Tn 2009、Tn 6002和Tn 2010。尽管在包括S.肺炎球菌Tn 916相关ICE广泛传播的分子基础仍有待充分阐明。我们发现，Tn 2009的获得通过体外转化或与供体GA 16833的接合交配都检测不到，产生的转移频率<10−7。GA 16833 Tn 2009接合基因的表达没有被显著诱导，并且在生物膜中没有检测到ICE环状中间体的形成。因此，Tn 2009在生物膜中的转移效率不受ICE接合基因ftsK缺失的影响。然而，在人鼻咽细胞生物反应器中形成的双菌株生物膜中，GA 16833 Tn 2009转移以10−4的重组频率（rF）有效发生。在D39受体菌株中添加和缺失早期感受态基因comE或转化器基因comEA和comEC阻止了Tn 2009的获得（rF <10−7）。对受体基因型的独立重组体进行基因组测序和单核苷酸多态性分析，鉴定出含有完整Tn 2009的~33-~55-kb供体DNA，支持同源重组。额外的肺炎球菌供体和受体组合被证明可以在生物膜中以10−4的rF有效转移Tn 916相关的ICE。Tn 916相关ICE在人鼻咽S.通过将>30-kb的DNA片段转化和同源重组到肺炎球菌基因组中，从而获得肺炎球菌生物膜。由于肺炎链球菌广泛的多药耐药性，世界卫生组织已将其指定为新药治疗研究和开发的优先病原体。多株S.肺炎克雷伯氏菌在人鼻咽中定殖并形成混合生物膜，这可以使抗生素抗性决定子的交换成为可能。Tn 916相关的整合和接合元件（ICE）在很大程度上负责大环内酯类和四环素耐药的广泛存在的S。肺炎。利用模拟供体和受体S.与人鼻咽中的肺炎杆菌菌株相比，与具有外源DNA的增殖细胞的体外条件相反，Tn 916相关ICE的有效转移发生在人鼻咽生物膜中。这种与Tn 916相关的ICE在S.在生物膜中的pneumoniae菌株是由于转化和同源重组，而不是接合。了解Tn 916相关ICE传播的分子机制可以促进对抗抗生素耐药性的新策略的设计。
Multidrug resistance in Streptococcus pneumoniae (or pneumococcus) continues to be a global challenge. An important class of antibiotic resistance determinants disseminating in S. pneumoniae are >20-kb Tn916-related integrative and conjugative elements (ICEs), such as Tn2009, Tn6002, and Tn2010. Although conjugation has been implicated as the transfer mechanism for ICEs in several bacteria, including S. pneumoniae, the molecular basis for widespread dissemination of pneumococcal Tn916-related ICEs remains to be fully elucidated. We found that Tn2009 acquisition was not detectable via in vitro transformation nor conjugative mating with donor GA16833, yielding a transfer frequency of <10−7. GA16833 Tn2009 conjugative gene expression was not significantly induced, and ICE circular intermediate formation was not detected in biofilms. Consistently, Tn2009 transfer efficiency in biofilms was not affected by deletion of the ICE conjugative gene ftsK. However, GA16833 Tn2009 transfer occurred efficiently at a recombination frequency (rF) of 10−4 in dual-strain biofilms formed in a human nasopharyngeal cell bioreactor. DNase I addition and deletions of the early competence gene comE or transformation apparatus genes comEA and comEC in the D39 recipient strain prevented Tn2009 acquisition (rF of <10−7). Genome sequencing and single nucleotide polymorphism analyses of independent recombinants of recipient genotype identified ~33- to ~55-kb donor DNAs containing intact Tn2009, supporting homologous recombination. Additional pneumococcal donor and recipient combinations were demonstrated to efficiently transfer Tn916-related ICEs at a rF of 10−4 in the biofilms. Tn916-related ICEs horizontally disseminate at high frequency in human nasopharyngeal S. pneumoniae biofilms by transformation and homologous recombination of >30-kb DNA fragments into the pneumococcal genome. IMPORTANCE The World Health Organization has designated Streptococcus pneumoniae as a priority pathogen for research and development of new drug treatments due to extensive multidrug resistance. Multiple strains of S. pneumoniae colonize and form mixed biofilms in the human nasopharynx, which could enable exchange of antibiotic resistance determinants. Tn916-related integrative and conjugative elements (ICEs) are largely responsible for the widespread presence of macrolide and tetracycline resistance in S. pneumoniae. Utilizing a system that simulates colonization of donor and recipient S. pneumoniae strains in the human nasopharynx, efficient transfer of Tn916-related ICEs occurred in human nasopharyngeal biofilms, in contrast to in vitro conditions of planktonic cells with exogenous DNA. This high-frequency Tn916-related ICE transfer between S. pneumoniae strains in biofilms was due to transformation and homologous recombination, not conjugation. Understanding the molecular mechanism for dissemination of Tn916-related ICEs can facilitate the design of new strategies to combat antibiotic resistance.
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