Oethinger M, Podglajen I, Kern W V, Levy S B
Center for Adaptation Genetics and Drug Resistance, Boston, Massachusetts 02111, USA.
Antimicrob Agents Chemother. 1998 Aug;42(8):2089-94. doi: 10.1128/AAC.42.8.2089.
The contribution of regulatory genes to fluoroquinolone resistance was studied with clinical Escherichia coli strains bearing mutations in gyrA and parC and with different levels of fluoroquinolone resistance. Expression of marA and soxS was evaluated by Northern blot analysis of isolates that demonstrated increased organic solvent tolerance, a phenotype that has been linked to overexpression of marA, soxS, and rob. Among 25 cyclohexane-tolerant strains detected by a screen for increased organic solvent tolerance (M. Oethinger, W. V. Kern, J. D. Goldman, and S. B. Levy, J. Antimicrob. Chemother. 41:111-114, 1998), we found 5 Mar mutants and 4 Sox mutants. A further Mar mutant was detected among 11 fluoroquinolone-resistant, cyclohexane-susceptible E. coli strains used as controls. Comparison of the marOR sequences of clinical Mar mutants with that of E. coli K-12 (GenBank accession no. M96235) revealed point mutations in marR in all mutants which correlated with loss of repressor function as detected in a marO::lacZ transcriptional assay. We found four other amino acid changes in MarR that did not lead to loss of function. Two of these changes, present in 20 of the 35 sequenced marOR fragments, identified a variant genotype of marOR. Isolates with the same gyrA and parC mutations showed increased fluoroquinolone resistance when the mutations were accompanied by overexpression of marA or soxS. These data support the hypothesis that high-level fluoroquinolone resistance involves mutations at several chromosomal loci, comprising structural and regulatory genes.
利用携带gyrA和parC突变且具有不同氟喹诺酮耐药水平的临床大肠杆菌菌株，研究了调控基因对氟喹诺酮耐药性的贡献。通过Northern印迹分析对表现出有机溶剂耐受性增加的分离株进行marA和soxS表达评估，该表型与marA、soxS和rob的过表达有关。在通过有机溶剂耐受性增加筛选检测到的25株耐环己烷菌株中（M. 厄廷格、W. V. 克恩、J. D. 戈德曼和S. B. 利维，《抗菌化疗杂志》41:111 - 114, 1998），我们发现了5株Mar突变体和4株Sox突变体。在用作对照的11株氟喹诺酮耐药、对环己烷敏感的大肠杆菌菌株中又检测到1株Mar突变体。将临床Mar突变体的marOR序列与大肠杆菌K - 12（GenBank登录号M96235）的序列进行比较，发现在所有突变体的marR中均存在点突变，这与在marO::lacZ转录分析中检测到的阻遏物功能丧失相关。我们在MarR中还发现了其他四个未导致功能丧失的氨基酸变化。其中两个变化存在于35个测序的marOR片段中的20个中，确定了marOR的一种变异基因型。当gyrA和parC突变伴有marA或soxS过表达时，具有相同突变的分离株表现出更高的氟喹诺酮耐药性。这些数据支持了高水平氟喹诺酮耐药性涉及多个染色体位点（包括结构基因和调控基因）突变的假说。
