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CRISPR Interference (CRISPRi) Inhibition of luxS Gene Expression in E. coli: An Approach to Inhibit Biofilm

Biofilm is a sessile bacterial accretion embedded in self-producing matrix. It is the root cause of about 80% microbial infections in human. Among them, E. coli biofilms are most prevalent in medical devices associated nosocomial infections. The objective of this study was to inhibit biofilm formati...

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Autores principales: Zuberi, Azna, Misba, Lama, Khan, Asad U.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2017
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5445563/
https://www.ncbi.nlm.nih.gov/pubmed/28603699
http://dx.doi.org/10.3389/fcimb.2017.00214
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author Zuberi, Azna
Misba, Lama
Khan, Asad U.
author_facet Zuberi, Azna
Misba, Lama
Khan, Asad U.
author_sort Zuberi, Azna
collection PubMed
description Biofilm is a sessile bacterial accretion embedded in self-producing matrix. It is the root cause of about 80% microbial infections in human. Among them, E. coli biofilms are most prevalent in medical devices associated nosocomial infections. The objective of this study was to inhibit biofilm formation by targeting gene involved in quorum sensing, one of the main mechanisms of biofilm formation. Hence we have introduced the CRISPRi, first time to target luxS gene. luxS is a synthase, involved in the synthesis of Autoinducer-2(AI-2), which in turn guides the initial stage of biofilm formation. To implement CRISPRi system for luxS gene suppression, we have synthesized complementary sgRNA to target gene sequence and co-expressed with dCas9, a mutated form of an endonuclease. Suppression of luxS expression was confirmed through qRT-PCR. The effect of luxS gene on biofilm inhibition was studied through crystal violet assay, XTT reduction assay and scanning electron microscopy. We conclude that CRISPRi system could be a potential strategy to inhibit bacterial biofilm through mechanism base approach.
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spelling pubmed-54455632017-06-09 CRISPR Interference (CRISPRi) Inhibition of luxS Gene Expression in E. coli: An Approach to Inhibit Biofilm Zuberi, Azna Misba, Lama Khan, Asad U. Front Cell Infect Microbiol Microbiology Biofilm is a sessile bacterial accretion embedded in self-producing matrix. It is the root cause of about 80% microbial infections in human. Among them, E. coli biofilms are most prevalent in medical devices associated nosocomial infections. The objective of this study was to inhibit biofilm formation by targeting gene involved in quorum sensing, one of the main mechanisms of biofilm formation. Hence we have introduced the CRISPRi, first time to target luxS gene. luxS is a synthase, involved in the synthesis of Autoinducer-2(AI-2), which in turn guides the initial stage of biofilm formation. To implement CRISPRi system for luxS gene suppression, we have synthesized complementary sgRNA to target gene sequence and co-expressed with dCas9, a mutated form of an endonuclease. Suppression of luxS expression was confirmed through qRT-PCR. The effect of luxS gene on biofilm inhibition was studied through crystal violet assay, XTT reduction assay and scanning electron microscopy. We conclude that CRISPRi system could be a potential strategy to inhibit bacterial biofilm through mechanism base approach. Frontiers Media S.A. 2017-05-26 /pmc/articles/PMC5445563/ /pubmed/28603699 http://dx.doi.org/10.3389/fcimb.2017.00214 Text en Copyright © 2017 Zuberi, Misba and Khan. http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
spellingShingle Microbiology
Zuberi, Azna
Misba, Lama
Khan, Asad U.
CRISPR Interference (CRISPRi) Inhibition of luxS Gene Expression in E. coli: An Approach to Inhibit Biofilm
title CRISPR Interference (CRISPRi) Inhibition of luxS Gene Expression in E. coli: An Approach to Inhibit Biofilm
title_full CRISPR Interference (CRISPRi) Inhibition of luxS Gene Expression in E. coli: An Approach to Inhibit Biofilm
title_fullStr CRISPR Interference (CRISPRi) Inhibition of luxS Gene Expression in E. coli: An Approach to Inhibit Biofilm
title_full_unstemmed CRISPR Interference (CRISPRi) Inhibition of luxS Gene Expression in E. coli: An Approach to Inhibit Biofilm
title_short CRISPR Interference (CRISPRi) Inhibition of luxS Gene Expression in E. coli: An Approach to Inhibit Biofilm
title_sort crispr interference (crispri) inhibition of luxs gene expression in e. coli: an approach to inhibit biofilm
topic Microbiology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5445563/
https://www.ncbi.nlm.nih.gov/pubmed/28603699
http://dx.doi.org/10.3389/fcimb.2017.00214
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