Rapid detection and differentiation of mobile colistin resistance (mcr-1 to mcr-10) genes by real-time PCR and melt-curve analysis

粘菌素 MCR-1型 熔化曲线分析 基因 生物信息学 底漆(化妆品) 多路复用 质粒 多重聚合酶链反应 聚合酶链反应 生物 医学 微生物学 遗传学 抗生素 大肠杆菌 肠杆菌科 化学 有机化学
作者
M. Mentasti,Sophia David,Kirsty Sands,S. Khan,Leanne Davies,Luke Turner,Mandy Wootton
出处
期刊:Journal of Hospital Infection [Elsevier BV]
卷期号:110: 148-155 被引量:15
标识
DOI:10.1016/j.jhin.2021.01.010
摘要

Summary

Background

The emergence of multi-drug-resistant (MDR) micro-organisms prompted new interest in older antibiotics, such as colistin, that had been abandoned previously due to limited efficacy or high toxicity. Over the years, several chromosomal-encoded colistin resistance mechanisms have been described; more recently, 10 plasmid-mediated mobile colistin resistance (mcr) genes have been identified. Spread of these genes among MDR Gram-negative bacteria is a matter of serious concern; therefore, reliable and timely mcr detection is paramount.

Aim

To design and validate a multiplex real-time polymerase chain reaction (PCR) assay for detection and differentiation of mcr genes.

Methods

All available mcr alleles were downloaded from the National Center for Biotechnology Information Reference Gene Catalogue, aligned with Clustal Omega and primers designed using Primer-BLAST. Real-time PCR monoplexes were optimized and validated using a panel of 120 characterized Gram-negative strains carrying a wide range of resistance genes, often in combination. Melt-curve analysis was used to confirm positive results.

Findings

In-silico analysis enabled the design of a 'screening' assay for detection of mcr-1/2/6, mcr-3, mcr-4, mcr-5, mcr-7, mcr-8 and mcr-9/10, paired with an internal control assay to discount inhibition. A 'supplementary' assay was subsequently designed to differentiate mcr-1, mcr-2, mcr-6, mcr-9 and mcr-10. Expected results were obtained for all strains (100% sensitivity and specificity). Melt-curve analysis showed consistent melting temperature results. Inhibition was not observed.

Conclusions

The assay is rapid and easy to perform, enabling unequivocal mcr detection and differentiation even when more than one variant is present. Adoption by clinical and veterinary microbiology laboratories would aid the surveillance of mcr genes amongst Gram-negative bacteria.
最长约 10秒,即可获得该文献文件

科研通智能强力驱动
Strongly Powered by AbleSci AI
更新
PDF的下载单位、IP信息已删除 (2025-6-4)

科研通是完全免费的文献互助平台,具备全网最快的应助速度,最高的求助完成率。 对每一个文献求助,科研通都将尽心尽力,给求助人一个满意的交代。
实时播报
2秒前
cm完成签到,获得积分10
4秒前
酷炫的听枫完成签到 ,获得积分10
5秒前
吱吱吱完成签到 ,获得积分10
6秒前
7秒前
上善若水呦完成签到 ,获得积分10
7秒前
小羡完成签到 ,获得积分10
7秒前
cqwswfl完成签到 ,获得积分20
8秒前
南山无梅落完成签到,获得积分10
9秒前
啵妞完成签到 ,获得积分10
9秒前
上官若男应助qiqi采纳,获得30
11秒前
拼搏的潘子完成签到,获得积分10
12秒前
zsj完成签到,获得积分10
13秒前
dolesy发布了新的文献求助10
14秒前
执着烧鹅完成签到 ,获得积分10
14秒前
哈哈哈完成签到,获得积分10
16秒前
yar应助博修采纳,获得10
18秒前
可爱的函函应助博修采纳,获得10
18秒前
MchemG应助博修采纳,获得10
18秒前
酷波er应助博修采纳,获得10
19秒前
时代更迭完成签到 ,获得积分10
19秒前
20秒前
WGOIST完成签到,获得积分10
21秒前
九九完成签到 ,获得积分10
21秒前
李新宇完成签到 ,获得积分10
22秒前
大橙子发布了新的文献求助10
26秒前
库凯伊完成签到,获得积分10
26秒前
duckspy发布了新的文献求助10
27秒前
CodeCraft应助jenny采纳,获得10
29秒前
lhnsisi完成签到,获得积分10
30秒前
jhlz5879完成签到,获得积分10
31秒前
悦耳曼凝完成签到 ,获得积分10
32秒前
文静的紫萱完成签到,获得积分10
32秒前
拼搏的飞薇完成签到,获得积分10
33秒前
曾建完成签到 ,获得积分10
34秒前
pep完成签到 ,获得积分10
35秒前
mufcyang完成签到,获得积分10
39秒前
了晨完成签到 ,获得积分10
40秒前
yi完成签到 ,获得积分10
43秒前
wxnice完成签到,获得积分10
44秒前
高分求助中
【提示信息,请勿应助】关于scihub 10000
Les Mantodea de Guyane: Insecta, Polyneoptera [The Mantids of French Guiana] 3000
徐淮辽南地区新元古代叠层石及生物地层 3000
The Mother of All Tableaux: Order, Equivalence, and Geometry in the Large-scale Structure of Optimality Theory 3000
Handbook of Industrial Diamonds.Vol2 1100
Global Eyelash Assessment scale (GEA) 1000
Picture Books with Same-sex Parented Families: Unintentional Censorship 550
热门求助领域 (近24小时)
化学 材料科学 医学 生物 工程类 有机化学 生物化学 物理 内科学 纳米技术 计算机科学 化学工程 复合材料 遗传学 基因 物理化学 催化作用 冶金 细胞生物学 免疫学
热门帖子
关注 科研通微信公众号,转发送积分 4038157
求助须知:如何正确求助?哪些是违规求助? 3575869
关于积分的说明 11373842
捐赠科研通 3305650
什么是DOI,文献DOI怎么找? 1819255
邀请新用户注册赠送积分活动 892655
科研通“疑难数据库(出版商)”最低求助积分说明 815022