碳青霉烯类抗生素是临床治疗多重耐药革兰阴性菌感染的“最后防线”。然而，鲍曼不动杆菌（Acinetobacter baumannii）对碳青霉烯类的耐药性日益严峻，已被世界卫生组织（WHO）列为“亟待研发新药”的耐药菌之一。近日，河北医科大学第二医院赵建宏教授团队在BMC Microbiology杂志发表题为Seasonal meropenem resistance in Acinetobacter baumannii and influence of temperature-driven adaptation的研究成果，在系统监测和实验研究基础上，揭示了鲍曼不动杆菌耐药性的季节性波动及其低温适应的分子机制，为理解细菌适应环境与耐药性演化之间的关联以及进一步寻找临床控制该菌的感染提供了新视角。

一、研究背景

1.  鲍曼不动杆菌耐药率呈现与温度相关的季节性波动

图 1 2020–2022年河北省鲍曼不动杆菌月检出数和抗生素耐药性趋势

Figure 1 Monthly detection number and antibiotic resistance trends of A. baumannii in Hebei Province, 2020–2022

Note: The monthly cumulative total (bars, left axis) and antibiotic resistance (line chart, right axis) of isolates obtained over the study period. MEM: Meropenem; IPM: Imipenem; CTX: Cefotaxime; CAZ: Ceftazidime; FEP: Cefepime; CIP: Ciprofloxacin; LVX: Levofloxacin; POL: Polymyxin B; AMK: Amikacin; SAM: Ampicillin/Sulbactam; MNO: Minocycline; TZP: Piperacillin/Tazobactam; GEN: Gentamicin; TOB: Tobramycin.

图 2 2020–2022鲍曼不动杆菌月检出数量及其美罗培南耐药率

Figure 2 Monthly detection number and meropenem resistance rate of A. baumannii in Hebei Province, 2020–2022

Note: The monthly cumulative total (gray bars, left axis) and meropenem (MEM) resistance rates (line chart, right axis) of isolates obtained over the study period.

图 3 MEM^R和MEM^NR鲍曼不动杆菌的月检出菌株数量及河北省月平均气温

Figure 3 Monthly detection number of MEM^R and MEM^NRA. baumannii and Monthly mean temperature in Hebei Province

Note: The monthly number of MEM^R (dotted line) and MEM^NR (solid line) A. baumannii over the study period (left axis), with the monthly average temperatures in Hebei Province superimposed (grey bars, right axis).

研究团队对2020–2022年间河北省45家医疗机构共计27754株鲍曼不动杆菌进行回顾性分析，发现冬季（低温季节）鲍曼不动杆菌的耐药率显著上升（图1-2），与同期气温呈负相关（r = −0.6862，P < 0.0001）。这一趋势主要由非耐药株在夏季显著扩张所致（图3），提示温度可能对菌群组成产生影响。

2.低温逆转耐药质粒的适应性代价

图 4 温度对鲍曼不动杆菌生长动力学和存活率的影响

Figure 4 Effect of temperature on the growth kinetics and survival rate of A. baumannii

Note: (A) Growth kinetics of A. baumannii were compared between MEM^R (n=13) and MEM^NR (n=7) using an independent two-sample t test. (B) Survival rates of A. baumannii at 4℃ were compared between MEM^R (n=2) and MEM^NR (n=2) using an independent two-sample t test. Asterisks denote statistical significance: *P < 0.05, **P< 0.01, ***P< 0.001.

图 5 菌株AB7434-poxa23和AB7434在不同环境温度下的相对适应性

Figure 5 Relative fitness of A. baumannii AB7434-poxa23 and AB7434 at different temperatures

Competitive index of AB7434-poxa23 at 37℃, 25℃, and 4℃ were compared using one-way ANOVA. Data are presented as the mean ± SEM of n = 3 independent experiments. Asterisks denote statistical significance: ****P< 0.0001.

耐药性的获得往往会给细菌带来适应性代价，常见的表现为细菌的生长传播速度减缓。基于这一假设，当环境中缺乏抗菌药物选择压力时，耐药菌株的生存能力下降，敏感菌株便呈现生长优势，耐药菌株可逐渐被清除。本研究比较美罗培南耐药（MEM^R）及非耐药（MEM^NR）鲍曼不动杆菌在不同培养温度下的生长速率，发现在在常温或37℃下，MEM^NR菌株表现出生长优势，但在低温（4℃）环境下，MEM^R菌株表现出更强的适应力（图4）。竞争实验结果表明，低温能够逆转耐药质粒poxa23（携带blaOXA-23）的适应性代价（图5），这可能促成耐药菌在冬季寒冷环境中获得生存优势。

3.抗生素与低温胁迫触发重叠的应激反应通路

图 6 低温胁迫对鲍曼不动杆菌外排泵、膜蛋白和β-内酰胺酶编码基因表达的影响

Figure 6 Effect of low-temperature stress on the expression of efflux pump, porin, and β-lactamase encoding genes in A. baumannii

Note: Gene expression encoding efflux pumps (adeJ and adeB), porin (ompA), and β-lactamase (blaOXA-51 andblaOXA-23) was analyzed in A. baumannii ATCC19606 and eight clinical isolates. These isolates included four MEM^R isolates carrying blaOXA-23 and four MEM^NR isolates. Nine strains of A. baumannii underwent 16 h at 4℃. The relative levels of expression of genes (versus 37℃) for each tested strain are presented as the mean of n = 3 independent experiments.

图 7 美罗培南使鲍曼不动杆菌adeJ和blaOXA-51基因表达上调

Figure 7 Meropenem upregulates adeJ and blaOXA-51 gene expression in A. baumannii

Note: Gene expression was analyzed in A. baumannii ATCC19606. The independent two-sample t test was used to compare the relative mRNA expression levels of A. baumannii before and after treatment with meropenem. All data are presented as mean ± SEM of n = 3 independent experiments. Asterisks distinguish statistical significance: *P < 0.0001.

图 8 温度适应改变了鲍曼不动杆菌对美罗培南的耐受性

Figure 8 Temperature adaptation alters the tolerance of A. baumannii to meropenem

Note: The difference in survival rates of A. baumannii ATCC19606 in MEM stress at 37°C and 4°C was determined by an independent two-sample t test. All data are presented as the mean ± SEM of n = 3 independent experiments. The asterisk distinguishes statistical significance: **P < 0.01, ****P < 0.0001.

进一步实验发现，在4℃条件下适应培养16小时后，耐药相关基因（如oxa-23、oxa-51、adeJ）均发生不同程度的表达上调（图6）；暴露于美罗培南（10 μg/mL）同样可诱导上述基因表达（图7）。此外，经低温适应后，菌株在美罗培南胁迫下的存活率显著提高（图8）。上述结果提示鲍曼不动杆菌对抗生素和低温胁迫存在交叉适应性机制，耐药相关基因可能赋予细菌耐药性的同时兼具环境胁迫响应功能。