Specificities and Functions of the recA and pps1 Intein Genes of Mycobacterium tuberculosis and Application for Diagnosis of Tuberculosis

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Published inJournal of Clinical Microbiology Vol. 40; no. 3; pp. 943 - 950
Main Authors SAVES, Isabelle, LEWIS, Lee-Ann, WESTRELIN, Fabrice, WARREN, Robin, DAFFE, Mamadou, MASSON, Jean-Michel
Format Journal Article
LanguageEnglish
Published Washington, DC American Society for Microbiology 01.03.2002
Subjects
Antibacterial agents
Antibiotics. Antiinfectious agents. Antiparasitic agents
Bacterial Proteins - genetics
Bacteriology
Base Sequence
Biological and medical sciences
Escherichia coli
Fundamental and applied biological sciences. Psychology
Genetics
Humans
inteins
Medical sciences
Microbiology
Molecular Sequence Data
MtuPps1 protein
Mycobacteriology and Aerobic Actinomycetes
Mycobacterium bovis
Mycobacterium tuberculosis
Mycobacterium tuberculosis - genetics
Pharmacology. Drug treatments
Polymerase Chain Reaction - methods
pps1 gene
Rec A Recombinases - genetics
recA gene
Sensitivity and Specificity
Tuberculosis - diagnosis
Mycobacterial infection
Infection
Specificity
Tuberculosis
Gene
Mycobacterium tuberculosis
Mycobacteriales
RecA protein
Bacteriosis
Mycobacteriaceae
Bacteria
Actinomycetes
Diagnosis
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Abstract Article Usage Stats Services JCM Citing Articles Google Scholar PubMed Related Content Social Bookmarking CiteULike Delicious Digg Facebook Google+ Mendeley Reddit StumbleUpon Twitter current issue JCM About JCM Subscribers Authors Reviewers Advertisers Inquiries from the Press Permissions & Commercial Reprints ASM Journals Public Access Policy JCM RSS Feeds 1752 N Street N.W. • Washington DC 20036 202.737.3600 • 202.942.9355 fax • journals@asmusa.org Print ISSN: 0095-1137 Online ISSN: 1098-660X Copyright © 2014 by the American Society for Microbiology.   For an alternate route to JCM .asm.org, visit: JCM       
AbstractList The worldwide recrudescence of tuberculosis and the widespread appearance of antibiotic resistance have strengthened the need for rapid and specific diagnostic tools. The prevailing microbiological identification of Mycobacterium tuberculosis , the causative agent of tuberculosis, which implies the use of in vitro cultures and acid-fast staining microscopy, is time-consuming. Detection of M. tuberculosis directly in clinical samples through PCR amplification of mycobacterium-specific genes, designed to shorten diagnostic delay, demonstrated reliability and high sensitivity. However, the quality of the diagnosis depends on the specificity of the target sequence for M. tuberculosis complex strains. In the present study, we demonstrated the specificity of recA and pps1 inteins for this complex and thus the feasibility of using intein-coding sequences as a new target for PCR diagnosis. Indeed, the recA and pps1 genes of 36 clinical isolates of M. tuberculosis and 10 field strains of M. bovis were found to be interrupted by an intein sequence at the RecA-a and Pps1-b sites, respectively, while a large number of nontuberculous mycobacterial species failed to demonstrate these insertions. Besides, the MtuPps1, which was cloned and expressed in Escherichia coli , was shown to possess an endonuclease activity. The intein cleaves the 40-bp sequence spanning the intein insertion site Pps1-b in the inteinless pps1 gene. In addition to the PCR amplification of recA and pps1 intein genes as a tool for diagnosis, the specific endonuclease activity could represent a new molecular approach to identify M. tuberculosis .
The worldwide recrudescence of tuberculosis and the widespread appearance of antibiotic resistance have strengthened the need for rapid and specific diagnostic tools. The prevailing microbiological identification of Mycobacterium tuberculosis, the causative agent of tuberculosis, which implies the use of in vitro cultures and acid-fast staining microscopy, is time-consuming. Detection of M. tuberculosis directly in clinical samples through PCR amplification of mycobacterium-specific genes, designed to shorten diagnostic delay, demonstrated reliability and high sensitivity. However, the quality of the diagnosis depends on the specificity of the target sequence for M. tuberculosis complex strains. In the present study, we demonstrated the specificity of recA and pps1 inteins for this complex and thus the feasibility of using intein-coding sequences as a new target for PCR diagnosis. Indeed, the recA and pps1 genes of 36 clinical isolates of M. tuberculosis and 10 field strains of M. bovis were found to be interrupted by an intein sequence at the RecA-a and Pps1-b sites, respectively, while a large number of nontuberculous mycobacterial species failed to demonstrate these insertions. Besides, the MtuPps1, which was cloned and expressed in Escherichia coli, was shown to possess an endonuclease activity. The intein cleaves the 40-bp sequence spanning the intein insertion site Pps1-b in the inteinless pps1 gene. In addition to the PCR amplification of recA and pps1 intein genes as a tool for diagnosis, the specific endonuclease activity could represent a new molecular approach to identify M. tuberculosis.
Article Usage Stats Services JCM Citing Articles Google Scholar PubMed Related Content Social Bookmarking CiteULike Delicious Digg Facebook Google+ Mendeley Reddit StumbleUpon Twitter current issue JCM About JCM Subscribers Authors Reviewers Advertisers Inquiries from the Press Permissions & Commercial Reprints ASM Journals Public Access Policy JCM RSS Feeds 1752 N Street N.W. • Washington DC 20036 202.737.3600 • 202.942.9355 fax • journals@asmusa.org Print ISSN: 0095-1137 Online ISSN: 1098-660X Copyright © 2014 by the American Society for Microbiology.   For an alternate route to JCM .asm.org, visit: JCM       
ABSTRACT The worldwide recrudescence of tuberculosis and the widespread appearance of antibiotic resistance have strengthened the need for rapid and specific diagnostic tools. The prevailing microbiological identification of Mycobacterium tuberculosis , the causative agent of tuberculosis, which implies the use of in vitro cultures and acid-fast staining microscopy, is time-consuming. Detection of M. tuberculosis directly in clinical samples through PCR amplification of mycobacterium-specific genes, designed to shorten diagnostic delay, demonstrated reliability and high sensitivity. However, the quality of the diagnosis depends on the specificity of the target sequence for M. tuberculosis complex strains. In the present study, we demonstrated the specificity of recA and pps1 inteins for this complex and thus the feasibility of using intein-coding sequences as a new target for PCR diagnosis. Indeed, the recA and pps1 genes of 36 clinical isolates of M. tuberculosis and 10 field strains of M. bovis were found to be interrupted by an intein sequence at the RecA-a and Pps1-b sites, respectively, while a large number of nontuberculous mycobacterial species failed to demonstrate these insertions. Besides, the MtuPps1, which was cloned and expressed in Escherichia coli , was shown to possess an endonuclease activity. The intein cleaves the 40-bp sequence spanning the intein insertion site Pps1-b in the inteinless pps1 gene. In addition to the PCR amplification of recA and pps1 intein genes as a tool for diagnosis, the specific endonuclease activity could represent a new molecular approach to identify M. tuberculosis .
Author Jean-Michel Masson
Lee-Ann Lewis
Robin Warren
Isabelle Saves
Mamadou Daffé
Fabrice Westrelin
AuthorAffiliation Institut de Pharmacologie et Biologie Structurale (UMR5089), CNRS/Université Paul Sabatier Toulouse III, 1 Institut National des Sciences Appliquées, Complexe Scientifique de Rangueil, F-31077 Toulouse Cedex, France, 3 Department of Medical Biochemistry, University of Stellenbosch Medical School, Tygerberg 7505, South Africa 2
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Issue 3
Keywords Mycobacterial infection
Infection
Specificity
Tuberculosis
Gene
Mycobacterium tuberculosis
Mycobacteriales
RecA protein
Bacteriosis
Mycobacteriaceae
Bacteria
Actinomycetes
Diagnosis
Language English
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Corresponding author. Mailing address: Institut de Pharmacologie et Biologie Structurale (UMR5089), CNRS/Université Paul Sabatier Toulouse III, 205 Route de Narbonne, F-31077 Toulouse Cedex, France. Phone: (33) 561 175 471. Fax: (33) 561 175 994. E-mail: saves@ipbs.fr.
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The worldwide recrudescence of tuberculosis and the widespread appearance of antibiotic resistance have strengthened the need for rapid and specific diagnostic...
ABSTRACT The worldwide recrudescence of tuberculosis and the widespread appearance of antibiotic resistance have strengthened the need for rapid and specific...
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StartPage 943
SubjectTerms Antibacterial agents
Antibiotics. Antiinfectious agents. Antiparasitic agents
Bacterial Proteins - genetics
Bacteriology
Base Sequence
Biological and medical sciences
Escherichia coli
Fundamental and applied biological sciences. Psychology
Genetics
Humans
inteins
Medical sciences
Microbiology
Molecular Sequence Data
MtuPps1 protein
Mycobacteriology and Aerobic Actinomycetes
Mycobacterium bovis
Mycobacterium tuberculosis
Mycobacterium tuberculosis - genetics
Pharmacology. Drug treatments
Polymerase Chain Reaction - methods
pps1 gene
Rec A Recombinases - genetics
recA gene
Sensitivity and Specificity
Tuberculosis - diagnosis
Title Specificities and Functions of the recA and pps1 Intein Genes of Mycobacterium tuberculosis and Application for Diagnosis of Tuberculosis
URI http://jcm.asm.org/content/40/3/943.abstract
https://www.ncbi.nlm.nih.gov/pubmed/11880421
https://search.proquest.com/docview/18280915
https://search.proquest.com/docview/71499194
https://pubmed.ncbi.nlm.nih.gov/PMC120251
Volume 40
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