title: Alternative use of polymerase chain reaction instead of rho-nitro-alpha-acetylamino-beta-hydroxypropiophenone test for the early detection of Mycobacterium tuberculosis in BACTEC 12B cultures.
abstract: Compared with conventional culture media, the TB BACTEC system has demonstrated improved isolation rates as well as an earlier detection time for mycobacterial species. However, the identification of M. tuberculosis by the rho-nitro-alpha-acetylamino-beta-hydroxypropiophenone (NAP) test in the TB BACTEC 460 system may require 6 days for interpretable results. We evaluated the usefulness of a polymerase chain reaction (PCR) assay for earlier identification of M. tuberculosis in positive BACTEC 12B cultures. A total of 262 TB BACTEC culture specimens with GIs > or = 10 were assayed by PCR, and the results were compared with those of the NAP test. The aliquot from BACTEC 12B vials was boiled for 10 min, and 2 microliters of the boiled suspension was used for the PCR assay. One set of primers based on the IS 6110 sequence of M. tuberculosis was used to amplify a 457 bp fragment of DNA. Of the 173 TB BACTEC culture specimens which were identified as M. tuberculosis by the NAP test. 171 were PCR positive. Of the 21 TB BACTEC cultures identified as MOTT by the NAP test. 19 were PCR negative, but 2 were PCR positive: these two cultures were shown to grow both M. tuberculosis and MOTT in BACTEC 12B vials. Of the remaining 68 cultures which were contaminated with AFB-negative bacteria, the PCR identified M. tuberculosis in 13, in agreement with the NAP results in the reprocessed specimens. Overall, the PCR results in the 262 BACTEC culture specimens with GIs > or = 10 were sensitive in 99.5% (186/187) and specific in 100% (68/68). The mean time for the identification of M. tuberculosis in TB BACTEC cultures with GIs > or = 10 was 7 h by the PCR compared to 5.9 days by the NAP test. These results suggest that the PCR could be used as an alternative to the NAP test for the rapid identification of M. tuberculosis in BACTEC 12B cultures, particularly in those which contained both M. tuberculosis and MOTT or M. tuberculosis and AFB-negative bacteria.
authors.affiliation: Department of Clinical Pathology, Chonnam University Medical School, Dong-ku, Kwangju, Republic of Korea.
authors.forename: D W
authors.initials: DW
authors.lastname: Ryang
authors.forename: D H
authors.initials: DH
authors.lastname: Ryang
authors.forename: M G
authors.initials: MG
authors.lastname: Shin
authors.forename: J H
authors.initials: JH
authors.lastname: Shin
authors.forename: S J
authors.initials: SJ
authors.lastname: Kee
authors.forename: S P
authors.initials: SP
authors.lastname: Suh
mesh_terms: D001431:Bacteriological Techniques; D006801:Humans; D006910:Hydroxypropiophenone / Q000031:analogs & derivatives*; D009169:Mycobacterium tuberculosis / Q000235:genetics / Q000254:growth & development / Q000302:isolation & purification*; D016133:Polymerase Chain Reaction; D012680:Sensitivity and Specificity; D014376:Tuberculosis / Q000175:diagnosis*
chemical_list: D006910:Hydroxypropiophenone; C017561:4-nitro-alpha-acetylamino-beta-hydroxypropiophenone