Impact of various killing methods on EMA/PMA-qPCR efficacy

Kód výsledku v IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F60461373%3A22330%2F18%3A43914945" target="_blank" >RIV/60461373:22330/18:43914945 - isvavai.cz</a>

Nalezeny alternativní kódy

RIV/60461373:22340/18:43914945 RIV/60461373:22810/18:43914945

Výsledek na webu

<a href="https://www.sciencedirect.com/science/article/pii/S0956713517304486" target="_blank" >https://www.sciencedirect.com/science/article/pii/S0956713517304486</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.1016/j.foodcont.2017.09.013" target="_blank" >10.1016/j.foodcont.2017.09.013</a>

Jazyk výsledku

angličtina

Název v původním jazyce

Impact of various killing methods on EMA/PMA-qPCR efficacy

Popis výsledku v původním jazyce

Although qPCR is indisputably very powerful tool for identification and quantification of various microorganisms, one of its main disadvantages is the inability to distinguish between signals originating from different sources (live, dead or compromised cells). Sample pre-treatment with DNA intercalating dyes such as ethidium monoazide (EMA) or propidium monoazide (PMA) is a widely used solution which helps to reduce this issue. Efficacy of this pre-treatment, however, depends on membrane permeability. Therefore in this work, a killing assay was performed, evaluating the impact of several cell inactivation techniques with various modes of action on EMA/PMA-qPCR results. Three main thermotolerant Campylobacter spp. (C. jejuni, C. coli and C. lari) in mixed suspension were inactivated by a panel of eight lethal and one sub-lethal procedures, which bacteria can encounter during their life span (e.g. food processing and storage, surface disinfection or laboratory maintenance in terms of sterilization or aseptic work), and afterwards were quantified using previously designed multiplex EMA/PMA-qPCR. It was shown that not only various inactivation mechanisms affect the efficacy of qPCR, but species-specific differences were apparent as well. C. lari was the most resistant species to EMA/PMA treatment, while C. coli seemed to be the most sensitive. Results of this killing assay might be generally helpful in every research where EMA/PMA-qPCR is used, as they clearly show that this pre-treatment technique cannot be blindly used for every sample without at least considering the way of the bacterial inactivation.

Název v anglickém jazyce

Impact of various killing methods on EMA/PMA-qPCR efficacy

Popis výsledku anglicky

Although qPCR is indisputably very powerful tool for identification and quantification of various microorganisms, one of its main disadvantages is the inability to distinguish between signals originating from different sources (live, dead or compromised cells). Sample pre-treatment with DNA intercalating dyes such as ethidium monoazide (EMA) or propidium monoazide (PMA) is a widely used solution which helps to reduce this issue. Efficacy of this pre-treatment, however, depends on membrane permeability. Therefore in this work, a killing assay was performed, evaluating the impact of several cell inactivation techniques with various modes of action on EMA/PMA-qPCR results. Three main thermotolerant Campylobacter spp. (C. jejuni, C. coli and C. lari) in mixed suspension were inactivated by a panel of eight lethal and one sub-lethal procedures, which bacteria can encounter during their life span (e.g. food processing and storage, surface disinfection or laboratory maintenance in terms of sterilization or aseptic work), and afterwards were quantified using previously designed multiplex EMA/PMA-qPCR. It was shown that not only various inactivation mechanisms affect the efficacy of qPCR, but species-specific differences were apparent as well. C. lari was the most resistant species to EMA/PMA treatment, while C. coli seemed to be the most sensitive. Results of this killing assay might be generally helpful in every research where EMA/PMA-qPCR is used, as they clearly show that this pre-treatment technique cannot be blindly used for every sample without at least considering the way of the bacterial inactivation.

Druh

J_imp - Článek v periodiku v databázi Web of Science

CEP obor

—

OECD FORD obor

10606 - Microbiology

Projekt

<a href="/cs/project/GA17-15936S" target="_blank" >GA17-15936S: Interakce nanočástic modifikovaných přírodními látkami s biofilmy patogenních mikroorganismů</a><br>

Návaznosti

P - Projekt vyzkumu a vyvoje financovany z verejnych zdroju (s odkazem do CEP)

Rok uplatnění

2018

Kód důvěrnosti údajů

S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů

Název periodika

Food control

ISSN

0956-7135

e-ISSN

—

Svazek periodika

85

Číslo periodika v rámci svazku

MAR

Stát vydavatele periodika

BE - Belgické království

Počet stran výsledku

6

Strana od-do

23-28

Kód UT WoS článku

000419409000004

EID výsledku v databázi Scopus

—