Klebsiella pneumoniae is an opportunistic pathogen that is responsible for community acquired infections and nosocomial infections. Antibiotic-resistant K. pneumoniae and/or hypervirulent K. pneumoniae are emerging as a serious threat to public health. For the sake of alleviating and conquering current dilemma, discovery of effective new drugs against K. pneumoniae is a tough challenge. However, traditional anti-K. pneumoniae drug discovery methods cost considerable amount of time, animals, labor and so on. So an efficient technique for in vitro and in vivo drug screening with the least time duration, animals and labor cost is highly needed for the discovery of new effective compounds. Hence, in this study we constructed a selectable marker-free autoluminescent K. pneumoniae (SfAlKp) harboring luxCDABE by combining Tn7 transposon and Xer-dif system. SfAlKp can be used for discovery of new drugs via detecting luminescence intensity as a surrogate marker. The energy-consuming autoluminescent reaction catalyzed by the LuxAB enzymes which use the substrates produced by LuxCDE using the metabolites of the bacteria. Tn7 can insert exogenous genes into the bacterial genome and the DNA fragment in between dif sequences can be recognized and removed by endogenous XerCD recombinases of K. pneumoniae. The drug susceptibility and growth rate of SfAlKp are identical to its parent strain, meanwhile the luminescence intensity and stability are also significant characteristics of SfAlKp. Compared to conventional techniques, the autoluminescence-based measurement is more applicable to high throughput screening for compounds both in vitro as well as in vivo in animal model.

肺炎克雷伯菌是一种机会性病原体，负责社区获得性感染和医院感染。耐药性肺炎克雷伯菌和/或高毒性肺炎克雷伯菌正在对公共卫生构成严重威胁。为了缓解和克服当前的困境，发现有效的抗肺炎克雷伯菌新药是一项艰巨的挑战。然而，传统的抗肺炎克雷伯菌药物发现方法花费大量时间，动物，劳动等。因此，对于一种有效的技术在体外和体内发现新的有效化合物迫切需要时间，动物和人工成本最少的药物筛选。因此，在这项研究中我们构建了一个小号总统当选标记- ˚F REE一个反对派升uminescent ķ。p neumoniae（SfAlKp）窝藏luxCDABE通过组合Tn7转座子和Xer- DIF系统。通过检测发光强度作为替代标记，SfAlKp可用于发现新药。LuxAB酶催化的耗能自发光反应，该酶使用由LuxCDE利用细菌的代谢产物产生的底物。Tn7可以将外源基因插入细菌基因组，dif序列之间的DNA片段可以被肺炎克雷伯菌的内源XerCD重组酶识别和去除。SfAlKp的药敏性和生长速率与其亲本菌株相同，同时发光强度和稳定性也是SfAlKp的重要特征。与传统技术相比，基于自发光的测量更适用于化合物的高通量筛选在动物模型中的体外和体内。