Expanding the Repertoire of Carbapenem-Hydrolyzing Metallo-ß-Lactamases by Functional Metagenomic Analysis of Soil Microbiota.
Expanding the Repertoire of Carbapenem-Hydrolyzing Metallo-ß-Lactamases by Functional Metagenomic Analysis of Soil Microbiota.
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DOI：
10.3389/fmicb.2016.01985
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发表时间：
2016
影响因子：
5.2
通讯作者：
Guardabassi L
中科院分区：
文献类型：
--
作者：
Gudeta DD;Bortolaia V;Pollini S;Docquier JD;Rossolini GM;Amos GC;Wellington EM;Guardabassi L
碳青霉烯酶是水解碳青霉烯类的细菌酶，碳青霉烯类是一组用于治疗严重细菌感染的最后手段 β-内酰胺抗生素。它们属于基于氨基酸序列的三个 β-内酰胺酶类别（A、B 和 D）。本研究的目的是通过功能宏基因组学阐明土壤微生物群中碳青霉烯酶编码基因的发生、多样性和功能。通过将农业 (n = 6) 和草原 (n = 4) 土壤的宏基因组 DNA 克隆到大肠杆菌中，生成了 10 个质粒文库。将文库培养在含有阿莫西林的琼脂上，并通过 CarbaNP 测试筛选每个文库多达 100 个菌落以产生碳青霉烯酶。假定的碳青霉烯酶的特征包括 DNA 序列、β-内酰胺的最低抑制浓度 (MIC) 和亚胺培南水解。在六个土壤样品中鉴定出九种不同的 B 类碳青霉烯酶，也称为金属-β-内酰胺酶 (MBL)，包括两个 B1 亚类（GRD23-1 和 SPN79-1）和 7 个 B3 亚类（CRD3-1、PEDO-1、GRD33-1、ESP-2、ALG6-1、ALG11-1 和 DHT2-1）。除了 PEDO-1 和 ESP-2 之外，这些酶与之前描述的任何 MBL 都存在较远的相关性（33% 至 59% 同一性）。 RAIphy 分析表明，六种酶（CRD3-1、GRD23-1、DHT2-1、SPN79-1、ALG6-1 和 ALG11-1）源自变形菌门，两种（PEDO-1 和 ESP-2）来自拟杆菌门，一种（GRD33-1）来自Gemmatimonadetes。在土壤微生物群中检测到的所有 MBL 在大肠杆菌中表达时均具有功能，导致可检测到的亚胺培南水解活性，并显着增加临床相关 β-内酰胺的 MIC。有趣的是，功能宏基因组学产生的 MBL 通常与通过抗生素选择性培养在相同土壤样品中检测到的 MBL 不同，这表明这两种方法针对土壤微生物群中的不同亚群。
Carbapenemases are bacterial enzymes that hydrolyze carbapenems, a group of last-resort β-lactam antibiotics used for treatment of severe bacterial infections. They belong to three β-lactamase classes based amino acid sequence (A, B, and D). The aim of this study was to elucidate occurrence, diversity and functionality of carbapenemase-encoding genes in soil microbiota by functional metagenomics. Ten plasmid libraries were generated by cloning metagenomic DNA from agricultural (n = 6) and grassland (n = 4) soil into Escherichia coli. The libraries were cultured on amoxicillin-containing agar and up to 100 colonies per library were screened for carbapenemase production by CarbaNP test. Presumptive carbapenemases were characterized with regard to DNA sequence, minimum inhibitory concentration (MIC) of β-lactams, and imipenem hydrolysis. Nine distinct class B carbapenemases, also known as metallo-beta-lactamases (MBLs), were identified in six soil samples, including two subclass B1 (GRD23-1 and SPN79-1) and seven subclass B3 (CRD3-1, PEDO-1, GRD33-1, ESP-2, ALG6-1, ALG11-1, and DHT2-1). Except PEDO-1 and ESP-2, these enzymes were distantly related to any previously described MBLs (33 to 59% identity). RAIphy analysis indicated that six enzymes (CRD3-1, GRD23-1, DHT2-1, SPN79-1, ALG6-1, and ALG11-1) originated from Proteobacteria, two (PEDO-1 and ESP-2) from Bacteroidetes and one (GRD33-1) from Gemmatimonadetes. All MBLs detected in soil microbiota were functional when expressed in E. coli, resulting in detectable imipenem-hydrolyzing activity and significantly increased MICs of clinically relevant ß-lactams. Interestingly, the MBLs yielded by functional metagenomics generally differed from those detected in the same soil samples by antibiotic selective culture, showing that the two approaches targeted different subpopulations in soil microbiota.
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影响因子：
4.9
作者：
Bellais, S;Naas, T;Nordmann, P
通讯作者：
Nordmann, P
影响因子：
5.6
作者：
Fonseca, Fatima;Bromley, Elizabeth H. C.;Spencer, James
通讯作者：
Spencer, James
影响因子：
4.9
作者：
Dortet, Laurent;Brechard, Ludivine;Nordmann, Patrice
通讯作者：
Nordmann, Patrice
影响因子：
4.9
作者：
Matsumura, N;Minami, S;Mitsuhashi, S
通讯作者：
Mitsuhashi, S
影响因子：
3
作者：
Nalbantoglu OU;Way SF;Hinrichs SH;Sayood K
通讯作者：
Sayood K