TY - JOUR
T1 - Culture-independent prediction of isoniazid resistance in Mycobacterium tuberculosis by katG gene analysis directly from sputum samples
AU - Rinder, Heinz
AU - Feldmann, Knut
AU - Tortoli, Enrico
AU - Grosset, Jacques
AU - Casal, Manuel
AU - Richter, Elvira
AU - Rifai, Mohammed
AU - Jarlier, Vincent
AU - Vaquero, Manuel
AU - Rüsch-Gerdes, Sabine
AU - Cambau, Emmanuelle
AU - Gutierrez, Juan
AU - Löscher, Thomas
N1 - Funding Information:
Supported by a grant from the European Commission,D G 12 (contract BMH4-CT97-2339).
PY - 1999
Y1 - 1999
N2 - Background: The molecular prediction of isoniazid (INH) resistance in Mycobacterium tuberculosis is hampered by the need for specialized equipment, expertise, high costs, a limited range of detectable mutations, or several of these factors. The rationale for the study was to find a practical alternative and to demonstrate generally valid problems. Methods and Results: DNA extracted from decontaminated sputum pellets was used to amplify a 0.26 kb target sequence within the katG gene. Mutations of codon 315, frequently found in isoniazid-resistant isolates, could be discriminated in a simple agarose minigel format following an AciI digest of the nested polymerase chain reaction (PCR) product. Within a panel of 22 sputum samples, INH resistance could be predicted in 5 of 10 samples containing isoniazid- resistant M. tuberculosis. The protocol is robust, requires little expertise and no specialized equipment, and provides the test results within 2 days. Conclusion: The results show the feasibility to rapidly and easily detect mutations highly predictive of isoniazid resistance. Nevertheless, this, like any other molecular resistance prediction test, is affected by often neglected factors, including mutation prevalences, the phenomenon of heteroresistance, and a possible bias toward one's own method.
AB - Background: The molecular prediction of isoniazid (INH) resistance in Mycobacterium tuberculosis is hampered by the need for specialized equipment, expertise, high costs, a limited range of detectable mutations, or several of these factors. The rationale for the study was to find a practical alternative and to demonstrate generally valid problems. Methods and Results: DNA extracted from decontaminated sputum pellets was used to amplify a 0.26 kb target sequence within the katG gene. Mutations of codon 315, frequently found in isoniazid-resistant isolates, could be discriminated in a simple agarose minigel format following an AciI digest of the nested polymerase chain reaction (PCR) product. Within a panel of 22 sputum samples, INH resistance could be predicted in 5 of 10 samples containing isoniazid- resistant M. tuberculosis. The protocol is robust, requires little expertise and no specialized equipment, and provides the test results within 2 days. Conclusion: The results show the feasibility to rapidly and easily detect mutations highly predictive of isoniazid resistance. Nevertheless, this, like any other molecular resistance prediction test, is affected by often neglected factors, including mutation prevalences, the phenomenon of heteroresistance, and a possible bias toward one's own method.
KW - Genotyping
KW - Heteroresistance
KW - PCR-RFLP
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U2 - 10.1016/S1084-8592(99)80038-1
DO - 10.1016/S1084-8592(99)80038-1
M3 - Article
C2 - 10462629
AN - SCOPUS:0032812660
SN - 1084-8592
VL - 4
SP - 145
EP - 152
JO - Molecular Diagnosis
JF - Molecular Diagnosis
IS - 2
ER -