A Novel Approach to the Viability Determination of Mycobacterium avium subsp. paratuberculosis Using Platinum Compounds in Combination With Quantitative PCR

Result code in IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F67985904%3A_____%2F21%3A00551995" target="_blank" >RIV/67985904:_____/21:00551995 - isvavai.cz</a>

Alternative codes found

RIV/00027162:_____/21:N0000218 RIV/00216224:14310/21:00123072

Result on the web

<a href="https://www.frontiersin.org/articles/10.3389/fmicb.2021.748337/full" target="_blank" >https://www.frontiersin.org/articles/10.3389/fmicb.2021.748337/full</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.3389/fmicb.2021.748337" target="_blank" >10.3389/fmicb.2021.748337</a>

Result language

angličtina

Original language name

A Novel Approach to the Viability Determination of Mycobacterium avium subsp. paratuberculosis Using Platinum Compounds in Combination With Quantitative PCR

Original language description

Mycobacterium avium subsp. paratuberculosis (MAP) represents a slow-growing bacterium causing paratuberculosis, especially in domestic and wild ruminants. Until recently, the assessment of MAP viability relied mainly on cultivation, which is very time consuming and is unable to detect viable but non-culturable cells. Subsequently, viability PCR, a method combining sample treatment with the DNA-modifying agent ethidium monoazide (EMA) or propidium monoazide (PMA) and quantitative PCR (qPCR), was developed, enabling the selective detection of MAP cells with an intact cell membrane. However, this technology requires a laborious procedure involving the need to work in the dark and on ice. In our study, a method based on a combination of platinum compound treatment and qPCR, which does not require such a demanding procedure, was investigated to determine mycobacterial cell viability. The conditions of platinum compound treatment were optimized for the fast-growing mycobacterium M. smegmatis using live and heat-killed cells. The optimal conditions consisting of a single treatment with 100 mu M cis-dichlorodiammine platinum(II) for 60 min at 5 degrees C resulted in a difference in quantification cycle (Cq) values between live and dead membrane-compromised mycobacterial cells of about 6 Cq corresponding to about 2 log(10) units. This optimized viability assay was eventually applied to MAP cells and demonstrated a better ability to distinguish between live and heat-killed mycobacteria as compared to PMA. The viability assay combining the Pt treatment with qPCR thereby proved to be a promising method for the enumeration of viable MAP cells in foodstuffs, environmental, and clinical samples which could replace the time-consuming cultivation or laborious procedures required when using PMA.

Czech name

—

Czech description

—

Type

J_imp - Article in a specialist periodical, which is included in the Web of Science database

CEP classification

—

OECD FORD branch

10606 - Microbiology

Project

<a href="/en/project/QK1810212" target="_blank" >QK1810212: Rapid, complex and multiplex methods for simultaneous detection of food-borne pathogens in food of animal and plant origin</a><br>

Continuities

I - Institucionalni podpora na dlouhodoby koncepcni rozvoj vyzkumne organizace

Publication year

2021

Confidentiality

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

Name of the periodical

Frontiers in Microbiology

ISSN

1664-302X

e-ISSN

1664-302X

Volume of the periodical

12

Issue of the periodical within the volume

NOV 24

Country of publishing house

CH - SWITZERLAND

Number of pages

11

Pages from-to

748337

UT code for WoS article

000738317500001

EID of the result in the Scopus database

2-s2.0-85120870501