Department of Biomedical Sciences, College of Medicine, Korea University, Anam-Dong, Seongbuk-Gu, Seoul 136-705, Korea.
School of Systems Biomedical Science and Research Center for Integrative Basic Science, Soongsil University, Seoul 156-743, Korea.
J Antimicrob Chemother. 2018 Feb 1;73(2):332-338. doi: 10.1093/jac/dkx378.
Although meropenem is widely used to treat Burkholderia infections, the response of Burkholderia pathogens to this antibiotic is largely unexplored.
Burkholderia thailandensis, a model for Burkholderia spp., particularly Burkholderia mallei and Burkholderia pseudomallei, was challenged with a lethal level of meropenem and survivors were isolated. The genomes of two of the isolates were analysed to identify mutated genes and these genes were then specifically examined in more isolates to profile mutation diversity. Mutants were characterized to investigate the biological basis underlying survival against meropenem.
One of two genes associated with tRNA metabolism [metG or trmD, encoding methionyl-tRNA synthetase or tRNA (guanine-N1)-methyltransferase, respectively] was found to be mutated in the two survivors. A single nucleotide substitution and a frameshift mutation were found in metG and trmD, respectively. Five different substitution mutations affecting methionine- or tRNA-binding sites were found in metG during further screening. The mutants exhibited slowed growth and increased tolerance not only to meropenem but also various other antibiotics. This tolerance required intact RelA, a key stringent response.
Specific mutations affecting the tRNA pool, particularly those in metG, play a pivotal role in the B. thailandensis response to meropenem challenge. This mechanism of antibiotic tolerance is important because it can reduce the effectiveness of meropenem and thereby facilitate chronic infection by Burkholderia pathogens. In addition, specific mutations found in MetG will prove useful in the effort to develop new drugs to completely inhibit this essential enzyme, while preventing stringent-response-mediated antibiotic tolerance in pathogens.
尽管美罗培南被广泛用于治疗伯克霍尔德氏菌感染，但该抗生素对伯克霍尔德氏菌病原体的作用在很大程度上尚未得到探索。
使用模式生物泰国伯克霍尔德氏菌（尤其包括鼻疽伯克霍尔德氏菌和类鼻疽伯克霍尔德氏菌）对致死剂量的美罗培南进行了挑战，然后分离存活下来的细菌。对其中两个分离株的基因组进行了分析，以确定突变基因，然后在更多的分离株中对这些基因进行了专门检查，以分析突变多样性。对突变体进行了特征分析，以研究对美罗培南产生抗性的生物学基础。
在两个存活下来的细菌中，发现与 tRNA 代谢相关的两个基因之一（metG 或 trmD，分别编码甲硫氨酰-tRNA 合成酶或 tRNA（鸟嘌呤-N1）-甲基转移酶）发生了突变。metG 和 trmD 分别发生了单个核苷酸取代和移码突变。在进一步筛选过程中，在 metG 中发现了 5 个不同的取代突变，影响了甲硫氨酸或 tRNA 的结合位点。突变体的生长速度明显减慢，不仅对美罗培南，而且对各种其他抗生素的耐受性也增加了。这种耐受性需要完整的 RelA，这是一种关键的严格反应。
特别是影响 tRNA 库的特定突变，尤其是 metG 中的突变，在泰国伯克霍尔德氏菌对美罗培南挑战的反应中起着关键作用。这种抗生素耐受机制很重要，因为它可以降低美罗培南的有效性，从而促进伯克霍尔德氏菌病原体的慢性感染。此外，在 MetG 中发现的特定突变将有助于开发完全抑制这种必需酶的新药，同时防止病原体产生严格反应介导的抗生素耐受。
