School of Pharmacy and Biomolecular Sciences, Liverpool John Moores University,Byrom Street, Liverpool L3 3AF, UK; School of Preclinic, Institute of Science, Suranaree University of Technology, Nakhon Ratchasima 30000, Thailand.
School of Preclinic, Institute of Science, Suranaree University of Technology, Nakhon Ratchasima 30000, Thailand.
J Glob Antimicrob Resist. 2019 Sep;18:22-25. doi: 10.1016/j.jgar.2019.01.012. Epub 2019 Jan 19.
In this study, a rapid and simple chromogenic method for screening of carbapenemase-producing Enterobacteriaceae (CPE), namely the Nitro-Carba test (NCT), was developed.
The NCT was validated using a total of 31 carbapenemase-producing isolates [9 Klebsiella pneumoniae carbapenemase (KPC), 11 metallo-β-lactamase (MBL) and 11 OXA-48] and 56 non-carbapenemase-producing isolates. The assay relies on the hydrolysis of nitrocefin by carbapenemases in the presence of carbapenem antibiotics. Carbapenemases were extracted with lysis buffer prior to addition to wells with and without imipenem (IPM), meropenem (MEM) and ertapenem (ETP). Following addition of nitrocefin, a change in colour from yellow to red, indicating carbapenemase production, was observed within 20min. The susceptibility profiles of each bacterial isolate were also investigated.
The NCT detected all 31 CPE within a timeframe of only 10s to 12min. All carbapenemase-producers hydrolysed nitrocefin in all wells. No colour change in wells with carbapenems was observed in non-carbapenemase-producers. The sensitivity for all three carbapenems was 100%, whilst the specificity of IPM, MEM and ETP was 64.3%, 91.1% and 100%, respectively. IPM, MEM and ETP had minimum inhibitory concentrations (MICs) against all carbapenemase-producing strains ranging from 0.5μg/mL to ≥256μg/mL, 0.25μg/mL to ≥256μg/mL and 1μg/mL to ≥256μg/mL, respectively. OXA-48-producing isolates showed lower MICs compared with MBL- and KPC-producing isolates.
This assay is a promising method for detecting CPE rapidly. The NCT is a simple and reliable method capable of detecting CPE even in carbapenem-susceptible strains.
本研究开发了一种快速、简单的显色法来筛选产碳青霉烯酶肠杆菌科细菌（CPE），即硝基碳青霉烯试验（NCT）。
采用 31 株产碳青霉烯酶分离株（9 株肺炎克雷伯菌碳青霉烯酶（KPC）、11 株金属β-内酰胺酶（MBL）和 11 株 OXA-48）和 56 株非产碳青霉烯酶分离株对 NCT 进行验证。该检测法依赖于碳青霉烯酶在碳青霉烯类抗生素存在的情况下对硝基头孢菌素的水解。在加入有无亚胺培南（IPM）、美罗培南（MEM）和厄他培南（ETP）的孔中，用裂解缓冲液提取碳青霉烯酶。加入硝基头孢菌素后，在 20 分钟内观察到颜色从黄色变为红色，表明产生了碳青霉烯酶。还研究了每个细菌分离株的药敏谱。
NCT 在 10 秒至 12 分钟的时间内检测到所有 31 株 CPE。所有产碳青霉烯酶的细菌均水解了所有孔中的硝基头孢菌素。非产碳青霉烯酶的细菌在含碳青霉烯的孔中没有观察到颜色变化。三种碳青霉烯的灵敏度均为 100%，而 IPM、MEM 和 ETP 的特异性分别为 64.3%、91.1%和 100%。IPM、MEM 和 ETP 对所有产碳青霉烯酶的菌株的最低抑菌浓度（MICs）范围为 0.5μg/mL 至≥256μg/mL、0.25μg/mL 至≥256μg/mL 和 1μg/mL 至≥256μg/mL。OXA-48 产酶株的 MIC 较 MBL 和 KPC 产酶株低。
该检测法是一种快速检测 CPE 的有前途的方法。NCT 是一种简单可靠的方法，即使在碳青霉烯类敏感菌株中也能检测到 CPE。
