Department of Pathology and Laboratory Medicine, Taipei Veterans General Hospital, Taipei, Taiwan.
Ph.D. Program in Medical Biotechnology, Taipei Medical University, Taipei, Taiwan.
J Antimicrob Chemother. 2020 Aug 1;75(8):2101-2109. doi: 10.1093/jac/dkaa173.
Formaldehyde toxicity is invariably stressful for microbes. Stenotrophomonas maltophilia, a human opportunistic pathogen, is widely distributed in different environments and has evolved an array of systems to alleviate various stresses.
To characterize the role of the formaldehyde detoxification system FadRACB of S. maltophilia in formaldehyde detoxification, oxidative stress alleviation and antibiotic susceptibility.
Presence of the fadRACB operon was verified by RT-PCR. Single or combined deletion mutants of the fadRACB operon were constructed for functional assays. Formaldehyde, menadione and quinolone susceptibilities were assessed by observing cell viability in formaldehyde-, menadione- and quinolone-containing media, respectively. Susceptibility to hydrogen peroxide was evaluated by disc diffusion assay. The agar dilution method was used to assess bacterial antibiotic susceptibilities. Expression of fadRACB was assessed by quantitative RT-PCR.
The fadR, fadA, fadC and fadB genes were arranged in an operon. Mutants of fadA and/or fadB were more susceptible to formaldehyde and oxidative stress than the WT KJ strain of S. maltophilia. No significant difference was observed in the ability of a fadC single mutant to ameliorate formaldehyde and oxidative stress; however, simultaneous inactivation of fadA, fadB and fadC further enhanced susceptibility to formaldehyde and oxidative stress. In addition, compared with WT KJ, the triple mutant KJΔFadACB was more susceptible to quinolones and more resistant to aminoglycosides. FadR functions as a repressor for the fadRACB operon. The FadRACB operon has moderate expression in aerobically grown WT KJ and is further derepressed by formaldehyde challenge or oxidative stress, but not by antibiotics.
The FadACB system contributes to mitigation of formaldehyde toxicity and oxidative stress and cross-protects S. maltophilia from quinolones.
甲醛毒性对微生物来说总是有压力的。嗜麦芽寡养单胞菌是一种人类机会致病菌，广泛分布于不同的环境中，并进化出了一系列系统来缓解各种压力。
描述嗜麦芽寡养单胞菌甲醛解毒系统 FadRACB 在甲醛解毒、缓解氧化应激和抗生素敏感性中的作用。
通过 RT-PCR 验证 fadRACB 操纵子的存在。构建 fadRACB 操纵子的单个或组合缺失突变体，用于功能测定。通过观察含甲醛、甲萘醌和喹诺酮的培养基中的细胞活力来评估甲醛、甲萘醌和喹诺酮的敏感性。通过圆盘扩散试验评估对过氧化氢的敏感性。使用琼脂稀释法评估细菌对抗生素的敏感性。通过定量 RT-PCR 评估 fadRACB 的表达。
fadR、fadA、fadC 和 fadB 基因按操纵子排列。与嗜麦芽寡养单胞菌 WT KJ 菌株相比， fadA 和/或 fadB 突变体对甲醛和氧化应激更敏感。 fadC 单突变体缓解甲醛和氧化应激的能力没有明显差异；然而， fadA、 fadB 和 fadC 的同时失活进一步增强了对甲醛和氧化应激的敏感性。此外，与 WT KJ 相比，三突变体 KJΔFadACB 对喹诺酮更敏感，对氨基糖苷类更耐药。FadR 作为 fadRACB 操纵子的阻遏物发挥作用。FadRACB 操纵子在有氧生长的 WT KJ 中适度表达，并进一步被甲醛挑战或氧化应激去阻遏，但不受抗生素影响。
FadACB 系统有助于缓解甲醛毒性和氧化应激，并使嗜麦芽寡养单胞菌对喹诺酮产生交叉保护。
