Study of <i>OprD2</i> gene polymorphism in imipenem-resistant <i>Pseucionionas aeruginosa</i>_West China Medical Journal

Authors：

BAI Tingting , WANG Dan , FAN Yingzi , YANG Xianggui , ZHOU Qin ,  XU Ying

Department of Clinical Laboratory, the First Affiliated Hospital of Chengdu Medical College, Chengdu, Sichuan 610500, P. R. China;

Corresponding?author：

XU Ying, Email: yingxu@cmc.edu.cn

Keywords：

Pseudomonas aeruginosa; Imipenem; OprD2 gene; Drug-resistant mechanism

DOI：

10.7507/1002-0179.202006367

Video：

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Objective To explore the relationship between imipenem-resistant Pseudomonas aeruginosa (IRPA) and outer membrane porin protein OprD2 gene mutation.Methods IRPA strains (n=30) and imipenem-sensitive Pseudomonas aeruginosa strains (n=30) isolated from the clinical specimens in the First Affiliated Hospital of Chengdu Medical College from December 2018 to December 2019 were collected. Bacteria identification and drug sensitivity experiments were performed by VITEK-2 Compact combined with Kirby-Bauer method. Quantitative real-time polymerase chain reaction was used to detect the expression levels of OprD2 gene in the imipenem-resistant group and the imipenem-sensitive group, and then the strains with decreased expression were sequenced.Results The expression level of OprD2 gene in the imipenem-resistant group was significantly lower than that in the imipenem-sensitive group (P=0.048). Compared with the X63152 sequence, all the 11 Pseudomonas aeruginosa strains with significantly decreased OprD2 expression carried genetic variation, which occurred in coding regions. The variation sites presented diversity. The missense mutation of c.308C→G, c.344A→C, c.379G→C, c.471G→C, c.508T→C, c.553G→C, c.556-558CCG→GGC and c.565-566TG→AC caused amino acid change in the loop L2 and L3 of OprD2 porin, which affected the binding to imipenem. In addition, the mutations at 127, 169-171, 175, 177, 604, 628-630, 688, 719, 785, 826, 828, 842-843, 886, 901, 928-930, 934, 936, 944-945, 1039, 1041 and 1274 all resulted in the changes of amino acid. We also detected a deletion (c.1114-1115delAT) and other nonsense mutations. Large fragment deletion of OprD2 gene occurred in Strain 12. Conclusions The mutation and deletion of OprD2 gene can reduce the expression lever of OprD2 gene, leading to the resistance to imipenem of Pseudomonas aeruginosa. The variation of OprD2 gene of IRPA from clinical strains is diverse.

Citation： BAI Tingting, WANG Dan, FAN Yingzi, YANG Xianggui, ZHOU Qin, XU Ying. Study of OprD2 gene polymorphism in imipenem-resistant Pseucionionas aeruginosa. West China Medical Journal, 2020, 35(8): 924-929. doi: 10.7507/1002-0179.202006367 Copy

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2.	陳約慧, 應芙蓉, 詹玲玲, 等. 1543 株銅綠假單胞菌感染的臨床分布與耐藥性分析. 中國臨床藥理學雜志, 2018, 34(24): 2814-2817.
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4.	Yoneyama H, Nakae T. Mechanism of efficient elimination of protein-D2 in outer-membrane of imipenem-resistant <italic>Pseudomonas-aeruginosa</italic>. Antimicrob Agents Chemother, 1993, 37(11): 2385-2390.
5.	Fang ZL, Zhang LY, Huang YM, <italic>et al</italic>. <italic>OprD</italic> mutations and inactivation in imipenem-resistant <italic>Pseudomonas aeruginosa</italic> isolates from China. Infect Genet Evol, 2014, 21: 124-128.
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10.	Quinn JP, Dudek EJ, Divincenzo CA, <italic>et al</italic>. Emergence of resistance to imipenem during therapy for <italic>Pseudomonas aeruginosa</italic> infections. J Infect Dis, 1986, 154(2): 289-294.
11.	沈繼錄, 朱德妹, 吳衛紅, 等. 碳青霉烯類抗生素耐藥銅綠假單胞菌外膜孔蛋白 OprD2 的研究. 中國感染與化療雜志, 2011, 11(4): 281-286.
12.	顏英俊, 喻華, 周忠華, 等. 亞胺培南耐藥的銅綠假單胞菌中 <italic>oprD</italic> 基因突變研究. 中華檢驗醫學雜志, 2009, 32(4): 451-454.
13.	Kim CH, Kang HY, Kim BR, <italic>et al</italic>. Mutational inactivation of OprD in carbapenem-resistant <italic>Pseudomonas aeruginosa</italic> isolates from Korean hospitals. J Microbiol, 2016, 54(1): 44-49.
14.	宋詩鐸, 鄭東鈞, 張宇雯, 等. 臨床分離耐亞胺培南銅綠假單胞菌染色體 <italic>oprD</italic> 基因突變的檢測. 中華傳染病雜志, 2003, 21(2): 144-145.
15.	李飛, 李麗娟, 梁德志, 等. 耐亞胺培南銅綠假單胞菌耐藥特征及其耐藥機制的研究. 分子診斷與治療雜志, 2019, 11(2): 101-106, 140.
16.	Epp SF, K?hler T, Plésiat P, <italic>et al</italic>. C-terminal region of <italic>Pseudomonas aeruginosa</italic> outer membrane porin OprD modulates susceptibility to meropenem. Antimicrob Agents Chemother, 2001, 45(6): 1780-1787.
17.	席紅利, 李娟, 楊麗娜, 等. 耐亞胺培南銅綠假單胞菌的耐藥性特征及 <italic>oprD</italic> 基因突變分析. 基因組學與應用生物學, 2019, 38(12): 5684-5688.
18.	李方去, 劉洋, 楊錦紅, 等. 兒科銅綠假單胞菌的 <italic>OprD</italic> 基因突變及表達對碳青霉烯類抗生素耐藥的作用. 中國衛生檢驗雜志, 2011, 21(5): 1063-1066.

1. Karampatakis T, Antachopoulos C, Tsakris A, <italic>et al</italic>. Molecular epidemiology of carbapenem-resistant <italic>Pseudomonas aeruginosa</italic> in an endemic area: comparison with global data. Eur J Clin Microbiol Infect Dis, 2018, 37(7): 1211-1220.
2. 陳約慧, 應芙蓉, 詹玲玲, 等. 1543 株銅綠假單胞菌感染的臨床分布與耐藥性分析. 中國臨床藥理學雜志, 2018, 34(24): 2814-2817.
3. 國家衛生計生委合理用藥專家委員會, 全國細菌耐藥監測網. 2018 年全國細菌耐藥監測報告. 中國合理用藥探索, 2020, 17(1): 1-10.
4. Yoneyama H, Nakae T. Mechanism of efficient elimination of protein-D2 in outer-membrane of imipenem-resistant <italic>Pseudomonas-aeruginosa</italic>. Antimicrob Agents Chemother, 1993, 37(11): 2385-2390.
5. Fang ZL, Zhang LY, Huang YM, <italic>et al</italic>. <italic>OprD</italic> mutations and inactivation in imipenem-resistant <italic>Pseudomonas aeruginosa</italic> isolates from China. Infect Genet Evol, 2014, 21: 124-128.
6. Clinical and Laboratory Standards Institute. Performance standards for antimicrobial susceptibility testing. CLSI supplement M100. 30th ed. Wayne, PA: Clinical and Laboratory Standards Institute, 2020.
7. 梁文, 李博, 董萌萌, 等. 銅綠假單胞菌對碳青霉烯類抗生素耐藥機制研究進展. 中國抗生素雜志, 2013, 38(9): 641-646.
8. Muderris T, Durmaz R, Ozdem B, <italic>et al</italic>. Role of efflux pump and OprD porin expression in carbapenem resistance of <italic>Pseudomonas aeruginosa</italic> clinical isolates. J Infect Dev Ctries, 2018, 12(1): 1-8.
9. Li H, Luo YF, Williams BJ, <italic>et al</italic>. Structure and function of OprD protein in <italic>Pseudomonas aeruginosa</italic>: from antibiotic resistance to novel therapies. Int J Med Microbiol, 2012, 302(2): 63-68.
10. Quinn JP, Dudek EJ, Divincenzo CA, <italic>et al</italic>. Emergence of resistance to imipenem during therapy for <italic>Pseudomonas aeruginosa</italic> infections. J Infect Dis, 1986, 154(2): 289-294.
11. 沈繼錄, 朱德妹, 吳衛紅, 等. 碳青霉烯類抗生素耐藥銅綠假單胞菌外膜孔蛋白 OprD2 的研究. 中國感染與化療雜志, 2011, 11(4): 281-286.
12. 顏英俊, 喻華, 周忠華, 等. 亞胺培南耐藥的銅綠假單胞菌中 <italic>oprD</italic> 基因突變研究. 中華檢驗醫學雜志, 2009, 32(4): 451-454.
13. Kim CH, Kang HY, Kim BR, <italic>et al</italic>. Mutational inactivation of OprD in carbapenem-resistant <italic>Pseudomonas aeruginosa</italic> isolates from Korean hospitals. J Microbiol, 2016, 54(1): 44-49.
14. 宋詩鐸, 鄭東鈞, 張宇雯, 等. 臨床分離耐亞胺培南銅綠假單胞菌染色體 <italic>oprD</italic> 基因突變的檢測. 中華傳染病雜志, 2003, 21(2): 144-145.
15. 李飛, 李麗娟, 梁德志, 等. 耐亞胺培南銅綠假單胞菌耐藥特征及其耐藥機制的研究. 分子診斷與治療雜志, 2019, 11(2): 101-106, 140.
16. Epp SF, K?hler T, Plésiat P, <italic>et al</italic>. C-terminal region of <italic>Pseudomonas aeruginosa</italic> outer membrane porin OprD modulates susceptibility to meropenem. Antimicrob Agents Chemother, 2001, 45(6): 1780-1787.
17. 席紅利, 李娟, 楊麗娜, 等. 耐亞胺培南銅綠假單胞菌的耐藥性特征及 <italic>oprD</italic> 基因突變分析. 基因組學與應用生物學, 2019, 38(12): 5684-5688.
18. 李方去, 劉洋, 楊錦紅, 等. 兒科銅綠假單胞菌的 <italic>OprD</italic> 基因突變及表達對碳青霉烯類抗生素耐藥的作用. 中國衛生檢驗雜志, 2011, 21(5): 1063-1066.

West China Medical Journal

Study of OprD2 gene polymorphism in imipenem-resistant Pseucionionas aeruginosa

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