{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,10,20]],"date-time":"2025-10-20T10:13:19Z","timestamp":1760955199673},"reference-count":46,"publisher":"Microbiology Society","issue":"4","content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"published-print":{"date-parts":[[2010,4,1]]},"abstract":"<jats:p>Continuous subculture has been observed to produce changes in the virulenceof micro-organisms, e.g. rabies virus, poliovirus and <jats:italic>Mycobacterium bovis<\/jats:italic> BCG. The latter has been used as a vaccine for tuberculosis for thelast 100 years; however, in some instances its efficacy has been observedto be very low. In order to determine whether similar changes can be producedin <jats:italic>Mycobacterium tuberculosis<\/jats:italic>, we selected four isolates, <jats:italic>M.tuberculosis<\/jats:italic> H37Rv, a Beijing strain (DR-689), and two moreisolates with deletion of the phospholipase C locus (<jats:italic>plcA-plcB-plcC<\/jats:italic>), and subjected them to serial culturing on Middlebrook 7H9 medium,with or without ox bile. After 100 passages, we performed RFLP-IS<jats:italic>6110<\/jats:italic> analysis to determine whether genomic changes were produced. We alsochecked their genomic composition by microarray analysis. Changes in virulencewere studied by measuring the cytotoxic effect of parental and subculturedisolates on a THP-1 macrophage monolayer. The most visible change was thechange of position of an IS<jats:italic>6110<\/jats:italic> band of \u223c1400\u2005bp to \u223c1600\u2005bpin the Beijing isolate subcultured in the ox bile medium. Analysis by microarrayand PCR confirmation did not reveal any genomic changes. Cytotoxic activitywas decreased in the isolates at levels close to that of BCG, and more consistentlyin those subcultured in the presence of ox bile.<\/jats:p>","DOI":"10.1099\/jmm.0.015966-0","type":"journal-article","created":{"date-parts":[[2010,1,8]],"date-time":"2010-01-08T03:52:17Z","timestamp":1262922737000},"page":"384-391","source":"Crossref","is-referenced-by-count":11,"title":["Effect of serial subculturing on the genetic composition and cytotoxicactivity of Mycobacterium tuberculosis"],"prefix":"10.1099","volume":"59","author":[{"given":"C. A.","family":"Molina-Torres","sequence":"first","affiliation":[{"name":"Servicio de Dermatolog\u00eda, Hospital Universitario \u2018Jos\u00e9E. Gonz\u00e1lez\u2019, Monterrey NL, Mexico"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"J.","family":"Castro-Garza","sequence":"additional","affiliation":[{"name":"Divisi\u00f3n de Biolog\u00eda Celular y Molecular,Centro de Investigaci\u00f3n Biom\u00e9dica del Noreste, IMSS, MonterreyNL, Mexico"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"J.","family":"Ocampo-Candiani","sequence":"additional","affiliation":[{"name":"Servicio de Dermatolog\u00eda, Hospital Universitario \u2018Jos\u00e9E. Gonz\u00e1lez\u2019, Monterrey NL, Mexico"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"M.","family":"Monot","sequence":"additional","affiliation":[{"name":"Global Health Institute, Ecole Polytechnique F\u00e9d\u00e9ralede Lausanne, EPFL SV\/GHI\/UPCOL, Station no. 15, CH-1015 Lausanne,Switzerland"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"S. T.","family":"Cole","sequence":"additional","affiliation":[{"name":"Global Health Institute, Ecole Polytechnique F\u00e9d\u00e9ralede Lausanne, EPFL SV\/GHI\/UPCOL, Station no. 15, CH-1015 Lausanne,Switzerland"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"L.","family":"Vera-Cabrera","sequence":"additional","affiliation":[{"name":"Servicio de Dermatolog\u00eda, Hospital Universitario \u2018Jos\u00e9E. Gonz\u00e1lez\u2019, Monterrey NL, Mexico"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"345","reference":[{"key":"R1","doi-asserted-by":"crossref","first-page":"151","DOI":"10.1186\/1471-2180-8-151","article-title":"Alteredcellular infiltration and cytokine levels during early Mycobacterium tuberculosis sigC mutant infection are associated with late-stage diseaseattenuation and milder immunopathology in mice","volume":"8","author":"Abdul-Majid","year":"2008","journal-title":"BMC Microbiol"},{"key":"R2","doi-asserted-by":"crossref","first-page":"553","DOI":"10.1016\/j.tube.2008.03.005","article-title":"population heterogeneity in Mycobacterium tuberculosis H37Rv","volume":"88","author":"Andreu","year":"2008).Cell","journal-title":"Tuberculosis (Edinb"},{"key":"R3","doi-asserted-by":"crossref","first-page":"5388","DOI":"10.1128\/JCM.42.11.5388-5391.2004","article-title":"profile change caused by deletion of a direct variable repeatin a Mycobacterium tuberculosis isogenic laboratory strain","volume":"42","author":"Aranaz","year":"2004).Spoligotyping","journal-title":"J Clin Microbiol"},{"key":"R4","doi-asserted-by":"crossref","first-page":"175","DOI":"10.1590\/S0074-02762000000700029","article-title":"Studies on the virulence and attenuationof Trypanosoma cruzi using immunodeficient animals","volume":"95","author":"Basombrio","year":"2000","journal-title":"MemInst Oswaldo Cruz"},{"key":"R5","first-page":"37","article-title":"Activation of thepromoters of genes associated with DNA damage, oxidative stress, ER stressand protein malfolding by the bile salt, deoxycholate","volume":"108","author":"Bernstein","year":"1999","journal-title":"ToxicolLett"},{"key":"R6","doi-asserted-by":"crossref","first-page":"47","DOI":"10.1016\/j.mrrev.2004.08.001","article-title":"Bile acids as carcinogens in humangastrointestinal cancers","volume":"589","author":"Bernstein","year":"2005","journal-title":"Mutat Res"},{"key":"R7","doi-asserted-by":"crossref","first-page":"5596","DOI":"10.1073\/pnas.0700869104","article-title":"Genome plasticity of BCG and impact on vaccineefficacy","volume":"104","author":"Brosch","year":"2007","journal-title":"Proc Natl Acad Sci U S A"},{"key":"R8","first-page":"716","article-title":"Sur quelques propri\u00e9t\u00e9s du bacille tuberculeux d'origine,cultiv\u00e9 sur la bile de boeuf glyc\u00e9rin\u00e9e","volume":"149","author":"Calmette","year":"1909","journal-title":"C R Acad Sci"},{"key":"R9","doi-asserted-by":"crossref","first-page":"183","DOI":"10.1016\/j.micinf.2005.06.011","article-title":"Neutral-red reaction isrelated to virulence and cell wall methyl-branched lipids in Mycobacteriumtuberculosis","volume":"8","author":"Cardona","year":"2006","journal-title":"Microbes Infect"},{"key":"R10","doi-asserted-by":"crossref","first-page":"145","DOI":"10.1111\/j.1574-6968.2002.tb11258.x","article-title":"Demonstration of spread by Mycobacteriumtuberculosis bacilli in A549 epithelial cell monolayers","volume":"212","author":"Castro-Garza","year":"2002","journal-title":"FEMS Microbiol Lett"},{"key":"R11","doi-asserted-by":"crossref","first-page":"733","DOI":"10.1099\/jmm.0.46915-0","article-title":"Use of a colorimetric assayto measure differences in cytotoxicity of Mycobacterium tuberculosis strains","volume":"56","author":"Castro-Garza","year":"2007","journal-title":"J Med Microbiol"},{"key":"R12","first-page":"301","article-title":"Emergence of Mycobacteriumtuberculosis with extensive resistance to second-line drugs \u2013 worldwide,2000\u20132004","volume":"55","year":"2006","journal-title":"MMWR Morb Mortal Wkly Rep"},{"key":"R13","doi-asserted-by":"crossref","first-page":"1329","DOI":"10.1128\/JB.01465-07","article-title":"A point mutation in thetwo-component regulator PhoP-PhoR accounts for the absence of polyketide-derivedacyltrehaloses but not that of phthiocerol dimycocerosates in Mycobacteriumtuberculosis H37Ra","volume":"190","author":"Chesne-Seck","year":"2008","journal-title":"J Bacteriol"},{"key":"R14","doi-asserted-by":"crossref","first-page":"2362","DOI":"10.1001\/jama.1983.03330410048027","article-title":"The BCG controversy: a methodological and statisticalreappraisal","volume":"249","author":"Clemens","year":"1983","journal-title":"JAMA"},{"key":"R15","doi-asserted-by":"crossref","first-page":"537","DOI":"10.1038\/31159","article-title":"Deciphering the biology of Mycobacteriumtuberculosis from the complete genome sequence","volume":"393","author":"Cole","year":"1998","journal-title":"Nature"},{"key":"R16","doi-asserted-by":"crossref","first-page":"649","DOI":"10.1046\/j.1462-5822.2003.00312.x","article-title":"Mycobacterium tuberculosis infectioncauses different levels of apoptosis and necrosis in human macrophages andalveolar epithelial cells","volume":"5","author":"Danelishvili","year":"2003","journal-title":"Cell Microbiol"},{"key":"R17","doi-asserted-by":"crossref","first-page":"6300","DOI":"10.1128\/IAI.68.11.6300-6310.2000","article-title":"Necrosis of lung epithelial cells duringinfection with Mycobacterium tuberculosis is preceded by cell permeation","volume":"68","author":"Dobos","year":"2000","journal-title":"Infect Immun"},{"key":"R18","doi-asserted-by":"crossref","first-page":"3532","DOI":"10.1099\/mic.0.029199-0","article-title":"and spontaneous loss of phthiocerol dimycocerosate (PDIM)from Mycobacterium tuberculosis grown in vitro : implicationsfor virulence studies","volume":"155","author":"Domenech","year":"2009).Rapid","journal-title":"Microbiology"},{"key":"R19","doi-asserted-by":"crossref","first-page":"116","DOI":"10.1006\/expr.2000.4554","article-title":"Entamoebahistolytica : diminution of erythrophagocytosis, phospholipase A(2),and hemolytic activities is related to virulence impairment in long-term axeniccultures","volume":"96","author":"Gonz\u00e1lez-Garza","year":"2000","journal-title":"Exp Parasitol"},{"key":"R20","doi-asserted-by":"crossref","first-page":"359","DOI":"10.1046\/j.1365-2958.2003.03844.x","article-title":"Individual RD1-region genes are required for export of ESAT-6\/CFP-10and for virulence of Mycobacterium tuberculosis","volume":"51","author":"Guinn","year":"2004","journal-title":"Mol Microbiol"},{"key":"R21","doi-asserted-by":"crossref","first-page":"3742","DOI":"10.1016\/j.vaccine.2007.01.112","article-title":"Current status of TB vaccines","volume":"25","author":"Gupta","year":"2007","journal-title":"Vaccine"},{"key":"R22","doi-asserted-by":"crossref","first-page":"164","DOI":"10.1016\/S0140-6736(08)61036-3","article-title":"Tuberculosis vaccinedevelopment: goals, immunological design, and evaluation","volume":"372","author":"Hoft","year":"2008","journal-title":"Lancet"},{"key":"R23","doi-asserted-by":"crossref","first-page":"12420","DOI":"10.1073\/pnas.1635213100","article-title":"The primary mechanism of attenuation of bacillusCalmette\u2013Gu\u00e9rin is a loss of secreted lytic function requiredfor invasion of lung interstitial tissue","volume":"100","author":"Hsu","year":"2003","journal-title":"Proc Natl Acad Sci US A"},{"key":"R24","first-page":"156","article-title":"In Virus Culture: a Practical Approach","author":"Jennings","year":"1999).Virus vaccines"},{"key":"R25","doi-asserted-by":"crossref","first-page":"907","DOI":"10.1128\/JCM.35.4.907-914.1997","article-title":"&other authors","volume":"35","author":"Kamerbeek","year":"1997","journal-title":"J Clin Microbiol"},{"key":"R26","doi-asserted-by":"crossref","first-page":"547","DOI":"10.1101\/gr.166401","article-title":"Comparinggenomes within the species Mycobacterium tuberculosis","volume":"11","author":"Kato-Maeda","year":"2001","journal-title":"Genome Res"},{"key":"R27","doi-asserted-by":"crossref","first-page":"117","DOI":"10.1086\/345862","article-title":"Deletionof RD1 from M. tuberculosis mimics BCG attenuation","volume":"187","author":"Lewis","year":"2003","journal-title":"JInfect Dis"},{"key":"R28","doi-asserted-by":"crossref","first-page":"635","DOI":"10.1586\/14760584.7.5.635","article-title":"Tuberculosis: vaccines in the pipeline","volume":"7","author":"Ly","year":"2008","journal-title":"Expert Rev Vaccines"},{"key":"R29","doi-asserted-by":"crossref","first-page":"1274","DOI":"10.1128\/JB.178.5.1274-1282.1996","article-title":"Molecular analysis of geneticdifferences between Mycobacterium bovis BCG and virulent M. bovis","volume":"178","author":"Mahairas","year":"1996","journal-title":"J Bacteriol"},{"key":"R30","first-page":"341","article-title":"New vaccines against tuberculosis. In Tuberculosis 2007From Basic Science to Patient Care","author":"Mart\u00edn","year":"2007"},{"key":"R31","doi-asserted-by":"crossref","first-page":"4802","DOI":"10.1128\/IAI.63.12.4802-4811.1995","article-title":"Cytotoxicity for lung epithelial cells is a virulence-associated phenotypeof Mycobacterium tuberculosis","volume":"63","author":"McDonough","year":"1995","journal-title":"Infect Immun"},{"key":"R32","doi-asserted-by":"crossref","first-page":"393","DOI":"10.1016\/j.tube.2007.05.010","article-title":"role of IS 6110 in the evolution of Mycobacterium tuberculosis","volume":"87","author":"McEvoy","year":"2007).The","journal-title":"Tuberculosis (Edinb"},{"key":"R33","doi-asserted-by":"crossref","first-page":"1787","DOI":"10.1534\/genetics.104.031062","article-title":"Bile-induced DNA damage in Salmonellaenterica","volume":"168","author":"Prieto","year":"2004","journal-title":"Genetics"},{"key":"R34","doi-asserted-by":"crossref","first-page":"15918","DOI":"10.1073\/pnas.2433882100","article-title":"Hypervirulentmutant of Mycobacterium tuberculosis resulting from disruption ofthe mce1 operon","volume":"100","author":"Shimono","year":"2003","journal-title":"Proc Natl Acad Sci U S A"},{"key":"R35","doi-asserted-by":"crossref","first-page":"1137","DOI":"10.1056\/NEJM199304223281601","article-title":"Exogenous reinfection with multidrug-resistant Mycobacteriumtuberculosis in patients with advanced HIV infection","volume":"328","author":"Small","year":"1993","journal-title":"N EnglJ Med"},{"key":"R36","doi-asserted-by":"crossref","first-page":"3021","DOI":"10.1128\/JCM.40.8.3021-3024.2002","article-title":"Simple and rapid differentiation of Mycobacteriumtuberculosis H37Ra from Mycobacterium tuberculosis clinicalisolates through two cytochemical tests using neutral red and nile blue stains","volume":"40","author":"Soto","year":"2002","journal-title":"J Clin Microbiol"},{"key":"R37","doi-asserted-by":"crossref","first-page":"13001","DOI":"10.1073\/pnas.2235593100","article-title":"Acute infection and macrophage subversionby Mycobacterium tuberculosis require a specialized secretion system","volume":"100","author":"Stanley","year":"2003","journal-title":"Proc Natl Acad Sci U S A"},{"key":"R38","first-page":"22","article-title":"Dissociationof the tubercle bacillus: a review","volume":"62","author":"Steenken","year":"1950","journal-title":"Am Rev Tuberc"},{"key":"R39","doi-asserted-by":"crossref","first-page":"406","DOI":"10.1128\/JCM.31.2.406-409.1993","article-title":"&other authors","volume":"31","author":"van Embden","year":"1993","journal-title":"J Clin Microbiol"},{"key":"R40","doi-asserted-by":"crossref","first-page":"1174","DOI":"10.1056\/NEJM199910143411602","article-title":"Exogenous reinfection as a cause of recurrent tuberculosis aftercurative treatment","volume":"341","author":"van Rie","year":"1999","journal-title":"N Engl J Med"},{"key":"R41","doi-asserted-by":"crossref","first-page":"2578","DOI":"10.1128\/JCM.29.11.2578-2586.1991","article-title":"Occurrence andstability of insertion sequences in Mycobacterium tuberculosis complexstrains: evaluation of an insertion sequence-dependent DNA polymorphism asa tool in the epidemiology of tuberculosis","volume":"29","author":"van Soolingen","year":"1991","journal-title":"J Clin Microbiol"},{"key":"R42","doi-asserted-by":"crossref","first-page":"21","DOI":"10.1016\/j.tube.2006.01.023","article-title":"Geneticcharacterization of Mycobacterium tuberculosis clinical isolateswith deletions in the plcA-plcB-plcC locus","volume":"87","author":"Vera-Cabrera","year":"2007","journal-title":"Tuberculosis (Edinb"},{"key":"R43","doi-asserted-by":"crossref","first-page":"3169","DOI":"10.1099\/00221287-145-11-3169","article-title":"Context-sensitive transposition of IS 6110 in mycobacteria","volume":"145","author":"Wall","year":"1999","journal-title":"Microbiology"},{"key":"R44","first-page":"2.4.1","article-title":"Preparation of genomicDNA from bacteria","volume-title":"In Current Protocols in Molecular Biology","author":"Wilson","year":"1987"},{"key":"R45","volume-title":"Global TuberculosisControl: Epidemiology, Strategy, Financing","year":"2009"},{"key":"R46","doi-asserted-by":"crossref","first-page":"e2375","DOI":"10.1371\/journal.pone.0002375","article-title":"Genetic basis of virulence attenuation revealed by comparative genomic analysisof Mycobacterium tuberculosis strain H37Ra versus H37Rv","volume":"3","author":"Zheng","year":"2008","journal-title":"PLoS One"}],"container-title":["Journal of Medical Microbiology"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.microbiologyresearch.org\/content\/journal\/jmm\/10.1099\/jmm.0.015966-0?crawler=true","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2020,9,30]],"date-time":"2020-09-30T15:57:02Z","timestamp":1601481422000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.microbiologyresearch.org\/content\/journal\/jmm\/10.1099\/jmm.0.015966-0"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2010,4,1]]},"references-count":46,"journal-issue":{"issue":"4","published-print":{"date-parts":[[2010,4,1]]}},"URL":"https:\/\/doi.org\/10.1099\/jmm.0.015966-0","relation":{},"ISSN":["0022-2615","1473-5644"],"issn-type":[{"value":"0022-2615","type":"print"},{"value":"1473-5644","type":"electronic"}],"subject":[],"published":{"date-parts":[[2010,4,1]]}}}