[Phenotypic and genotypic characteristics of Mycobacterium kansasii strains isolated in Spain (2000-2003)]. / Caracterización fenotípica y genotípica de cepas de Mycobacterium kansasii aisladas en España (2000-2003).

INTRODUCTION: Mycobacterium kansasii is an opportunistic pathogen that mainly causes pulmonary infections. This species accounted for 9.7% of Mycobacteria other than tuberculosis complex identified in the reference laboratory of the Spanish Centro Nacional de Microbiologia during the period of 2000-2003. METHODS: In this study we analyzed the phenotypic and genotypic characteristics of 298 M. kansasii strains isolated over this 4-year period. The phenotypic characteristics were determined by conventional methods: biochemical testing, culture and morphological study. Genotypic characteristics were studied using PCR restriction fragment analysis of a fragment of the hsp65 gene and digestion with BstEII and HaeIII, according to the method of Telenti. RESULTS: Among the total of tested strains, 57.4% had the typical phenotypic characteristics described for M. kansasii. The rest had atypical patterns that we grouped into 17 biotypes. Strains belonging to six of the seven described genotypes were identified, with 86.6% of the strains falling into genotype I. CONCLUSION: Analysis of the phenotypic characteristics of M. kansasii showed a higher discrimination index for intraspecific differentiation than genotypic methods. Nevertheless, the high variability of phenotypic characteristics, some of which were very specific for the species (e.g., photochromogenicity), could complicate their identification. Hence both conventional and molecular methods should be used to accurately identify the atypical isolates.

Caracterización fenotípica y genotípica de cepas de Mycobacterium kansasii aisladas en España (2000-2003) / Phenotypic and genotypic characteristics of Mycobacterium kansasii strains isolated in Spain (2000-2003)

Introducción. Mycobacterium kansasii se define como un patógeno oportunista que causa mayoritariamente enfermedad pulmonar. Esta especie representa el 9,7% de las micobacterias no tuberculosas identificadas en el Centro Nacional de Microbiología durante los años 2000-2003. Métodos. En el presente estudio se analizan las características fenotípicas y genotípicas de 298 cepas identificadas como M. kansasii durante este período de tiempo. La caracterización fenotípica se realizó mediante métodos convencionales: características morfológicas, de crecimiento y pruebas bioquímicas. La caracterización genotípica se realizó mediante amplificación de un fragmento del gen hsp65 y posterior digestión con las enzimas BstEII y HaeIII según el método descrito por Telenti. Resultados. El 57,4% de las cepas estudiadas poseían las características fenotípicas propias de M. kansasii, pero en el resto se obtuvieron otros resultados que nos permitieron definir 17 biotipos. Se identificaron cepas pertenecientes a 6 de los 7 genotipos descritos, el 86,6% de las cepas se agrupaban en el genotipo I. Conclusión. Al analizar las características fenotípicas, se obtiene una mayor discriminación para la diferenciación intraespecífica de M. kansasii que al aplicar métodos genotípicos, pero debido al gran número de cepas con características fenotípicas atípicas, algunas de ellas muy específicas para la especie como la fotocromogenicidad, se aconseja la utilización de métodos convencionales y la aplicación de métodos moleculares para la correcta identificación de la especie (AU)

Introduction. Mycobacterium kansasii is an opportunistic pathogen that mainly causes pulmonary infections. This species accounted for 9.7% of Mycobacteria other than tuberculosis complex identified in the reference laboratory of the Spanish Centro Nacional de Microbiología during the period of 2000-2003. Methods. In this study we analyzed the phenotypic and genotypic characteristics of 298 M. kansasii strains isolated over this 4-year period. The phenotypic characteristics were determined by conventional methods: biochemical testing, culture and morphological study. Genotypic characteristics were studied using PCR restriction fragment analysis of a fragment of the hsp65 gene and digestion with BstEII and HaeIII, according to the method of Telenti. Results. Among the total of tested strains, 57.4% had the typical phenotypic characteristics described for M. kansasii. The rest had atypical patterns that we grouped into 17 biotypes. Strains belonging to six of the seven described genotypes were identified, with 86.6% of the strains falling into genotype I. Conclusion. Analysis of the phenotypic characteristics of M. kansasii showed a higher discrimination index for intraspecific differentiation than genotypic methods. Nevertheless, the high variability of phenotypic characteristics, some of which were very specific for the species (e.g., photochromogenicity), could complicate their identification. Hence both conventional and molecular methods should be used to accurately identify the atypical isolates (AU)

Molecular characterization of isoniazid-resistant Mycobacterium tuberculosis clinical strains isolated in the Philippines.

The prevalence of mutations in the katG, inhA and oxyR-ahpC genes of isoniazid (INH)-resistant Mycobacterium tuberculosis isolates in the Philippines were determined. Of 306 M. tuberculosis isolates studied, 81 (26.5%) exhibited INH-resistance. Forty-four strains (54.3%) had mutations in the katG gene, eighteen strains (22.2%) had mutations in the putative inhA locus region, seven had mutations in both regions and five strains had mutations in the oxyR-ahpC operon. Only seven strains had no mutations. A total of 71 of the 81 (87.6%) resistant strains and 65 of the 72 (90.3%) INH sensitive randomly selected strains showed amino acid substitution in codon 463 (Arg to Leu) (88.9%). This fact supports the hypothesis that mutations at codon 463 are independent of INH-resistance and are linked to the geographical origins of the strains.

Molecular analysis of rifampicin-resistant Mycobacterium tuberculosis isolated in Spain (1996-2001). Description of new mutations in the rpoB gene and review of the literature.

Mutations of the 81-bp-core region of rpoB gene associated with rifampicin resistance were studied in 169 clinical strains of Mycobacterium tuberculosis isolated from Spain between 1996 and 2001. The analysis identified 16 single base changes, one deletion, two insertions and five multiple mutations (with two or three codons implicated). Eight strains had no mutations, although they were resistant by the proportion method. The five single nucleotide changes with two or three codons implicated and one insertion have not been previously reported. The analysis of the RFLP-IS6110 pattern showed a great heterogeneity (159 individual patterns and four patterns with two or three strains) that suggests no geographical or temporal relations between the studied strains. The concordance rate of InnoLipa Rif-Tb probe with sequencing and phenotypic methods was 94.6%. The results indicate that the InnoLipa probe assay may be useful for rapid screening of the most frequent mutations found in rifampicin-resistant M. tuberculosis strains.