使用qRT-PCR检测从临床样本中分离出的铜绿假单胞菌菌株外排泵MexAB-OprM和OprD的表达情况。
Expression of efflux pump MexAB-OprM and OprD of Pseudomonas aeruginosa strains isolated from clinical samples using qRT-PCR.
作者信息
Arabestani Mohammad Reza, Rajabpour Mojtaba, Yousefi Mashouf Rasoul, Alikhani Mohammad Yousef, Mousavi Seyed Masoud
机构信息
1)Brucellosis Research Center, Hamadan University of Medical Sciences, Hamadan, Iran. 2)Department of Microbiology, Faculty of Medicine, Hamadan University of Medical Sciences, Hamadan,
Brucellosis Research Center, Hamadan University of Medical Sciences, Hamadan, Iran.
出版信息
Arch Iran Med. 2015 Feb;18(2):102-8.
BACKGROUND
Pseudomonas aeruginosa is one of the most common nosocomial pathogens with high mortality rates. Organisms such as Pseudomonas aeruginosa have the ability to develop high level MDR (Multi drug resistance). The MexAB-OprM system is one of the largest multi-drug resistant efflux pumps with high levels of expression and the first finding of the RND (Resistance-nodulation-division) family in P. aeruginosa.
AIM
For better understanding of the antibiotic resistance mechanism in P. aeruginosa, this study was conducted to determine the expression of the genes encoding these efflux pumps in 100 strains of P. aeruginosa isolated from patients admitted to various hospitals in Hamadan using quantitative Real-Time PCR (qRT-PCR).
METHODS
This study examined 100 strains of P. aeruginosa isolated from patients admitted to various hospitals in Hamadan. Then, 31 samples were selected based on collected specimen type and their antibiotic susceptibility pattern; i.e., the samples with reduced susceptibility to antibiotics, particularly carbapenems. Antibiotic disk diffusion method was performed for aminoglycoside, quinolone and carbapenem. Furthermore, MIC method was performed for ciprofloxacin, gentamicin and imipenem. Finally, qRT-PCR was used for determining the efflux pump genes expression.
RESULTS
Among eight selected antibiotics, the greatest resistance was to levofloxacin (61.2%, n = 19) and the lowest one to imipenem (9.6%, n = 3). All isolates (100%, n = 31) exhibited efflux pump MexAB-OprM genes but different expression was observed in different strains. The result of gene expression indicated that significant differences in expression of MexR (P value = 0.003), OprD (P value < 0.001), and MexB (P value = 0.026) genes. In addition, there was high level of MexR gene expression in bacteria that leads to reduced expression of MexA, MexB, and OprM. The OprD gene was presented in all strains but different expression has been observed.
CONCLUSION
Identifying the bacterial resistance mechanisms is very complicated. Although efflux pump MexAB-OprM plays an important role in antibiotic resistance in P. aeruginosa, because of acting the efflux pumps on antibiotics in a non-specific manner, it is elusive to consider or describe an antibiotic resistance based on the presence or absence of an efflux pump.
背景
铜绿假单胞菌是最常见的医院病原体之一，死亡率很高。像铜绿假单胞菌这样的微生物有能力产生高水平的多重耐药性（MDR）。MexAB - OprM系统是最大的多重耐药性外排泵之一，表达水平高，是在铜绿假单胞菌中首次发现的耐药 - 结瘤 - 分裂（RND）家族成员。
目的
为了更好地理解铜绿假单胞菌的抗生素耐药机制，本研究采用定量实时荧光定量PCR（qRT - PCR）来测定从哈马丹各医院收治患者中分离出的100株铜绿假单胞菌中编码这些外排泵的基因表达情况。
方法
本研究检测了从哈马丹各医院收治患者中分离出的100株铜绿假单胞菌。然后，根据收集的标本类型及其抗生素敏感性模式选择了31个样本；即对抗生素，特别是碳青霉烯类抗生素敏感性降低的样本。对氨基糖苷类、喹诺酮类和碳青霉烯类抗生素采用抗生素纸片扩散法。此外，对环丙沙星、庆大霉素和亚胺培南采用最低抑菌浓度（MIC）法。最后，用qRT - PCR测定外排泵基因表达。
结果
在所选的8种抗生素中，对左氧氟沙星的耐药性最高（61.2%，n = 19），对亚胺培南的耐药性最低（9.6%，n = 3）。所有分离株（100%，n = 31）均表现出外排泵MexAB - OprM基因，但不同菌株中观察到不同的表达情况。基因表达结果表明，MexR（P值 = 0.003）、OprD（P值 < 0.001）和MexB（P值 = 0.026）基因的表达存在显著差异。此外，细菌中MexR基因表达水平高会导致MexA、MexB和OprM表达降低。所有菌株中均存在OprD基因，但观察到不同的表达情况。
结论
确定细菌耐药机制非常复杂。虽然外排泵MexAB - OprM在铜绿假单胞菌的抗生素耐药中起重要作用，但由于外排泵以非特异性方式作用于抗生素，基于外排泵的存在与否来考虑或描述抗生素耐药性是难以捉摸的。
Zotero 插件