RpoB gene mutations in rifampin-resistant Mycobacterium tuberculosis strains isolated in the Aegean region of Turkey.

Although DNA sequencing method has not been used routinely for detecting resistance of Mycobacterium tuberculosis to antituberculosis drugs, it is suggested for the investigation of gene mutations causing resistance. Using DNA sequencing (Automated Applied Biosystems), we attempted to determine mutations in the 81-bp cor region (rifampin resistance determining region) of the rpoB gene in 48 Mycobacterium tuberculosis strains found to be rifampin resistant by classical phenotypic methods. Of the 48 strains, 46 (95.8%) were found to have rpo gene mutations with 13 different types while in two (4.2%) of the 48 strains, no mutations were detected. None of the strains had mutations at more than one codon. Point mutations at the 531st (52.1%) and 526th (18.9%) codons were frequent. The most frequent point mutation was Ser531Leu, and it was found in 21 (43,8 %) of 48 strains. This is the first study from Turkey, reporting Ser522Leu point mutation in one isolate and deletion of 515th codon (ATG - Met) in two isolates.

Distinguishing hepatitis B virus (HBV) genotype D from non-D by a simple PCR.

Different HBV genotypes have characteristic geographical distribution, which is important epidemiologically. HBV strains have been classified into eight different genotypes (A-H) on the basis of >8% differences in the entire genomic sequence. Genotypes A and D are predominant in Europe, Africa, and the USA, genotypes B and C are restricted to East Asia, genotype E is found in Africa, and genotype F is found in indigenous populations in Central and South America. Genotype D is prevalent in the Turkish population. HBV genotype D shows a 33-bp deletion in the pre-S1 region that accounts for their smaller genomic size (3182 bp). This deletion can be used to facilitate the identification of genotype D. A primer in the pre-S1 region was designed to discriminate genotype D from non-D by PCR. Sixty genotype D (40 acute and 20 chronic) and 4 genotype A sera identified by restriction fragment length polymorphism (RFLP) were included in the study. Using this simple PCR method, all genotype D sera were identified correctly and the test was able to detect HBV DNA at 1000 genomes per ml. An advantage of this method is that it can differentiate in a mixture of genotypes (genotype D from non-D) provided that one isn't present below 1 x 10(4) copies/ml. In conclusion this method is rapid (approximately 5h) and it will contribute to the epidemiological study of HBV in high prevalence areas of genotype D. It can also differentiate between genotype D from non-D in cases of co-infection.

Determination of minumum inhibitory concentrations of Candida species isolated from vaginal swab specimens by using broth macrodilution and E-test.

The purpose of the present study was to evaluate the utility of the E-test in determining the antifungal susceptibility of Candida species. A total of 50 Candida strains, including 34 Candida albicans and 16 non-albicans were isolated from vaginal swab specimens from women suffering from vaginitis. The minimum inhibitory concentrations (MICs) of amphotericin B, fluconazole and ketoconazole were detected by using broth macrodilution and the E-test. When the results of the two tests were compared, the MIC values were considered acceptable if the difference between the two assays was no more than two-fold (+/-1dilution). The acceptable rates were: 84% for amphotericin B, 97% for fluconazole and 78% for ketoconazole. Finally, MICs of C. albicans against the tested antifungal agents were generally lower than for non-albicans strains. These results suggest that the E-test can be used for the determination of MIC values for Candida species isolates.

A comparison of the E-test and proportion methods for susceptibility testing of Mycobacterium tuberculosis.

The aim of this study was to investigate the correlation between three antibiotic susceptibility methods, the proportion method on Löwenstein Jensen medium (LJ medium), the proportion method on Middlebrook 7H11 agar (7H11 agar), and the E-test for Mycobacterium tuberculosis. Fifty M. tuberculosis isolates were tested in vitro against isoniazid, rifampicin, streptomycin, and ethambutol according to the E-test, the proportion methods on 7H11 agar and LJ medium and then compared with a reference test which was the proportion method on 7H11 agar. The correlations between proportion method on 7H11 agar and proportion method on LJ medium for isoniazid, rifampicin, streptomycin, and ethambutol were 93.9%, 85.1%, 85.4% and 78.7% respectively. The correlations between the proportion method on 7H11 agar and the E-test were 83.1%, 78.8%, 84.7% and 80.5% respectively. There were no significant differences observed between the E-test and LJ medium compared to 7H11 agar. The average times to obtain susceptibility test results were 7 and 21 days for the E-test and agar proportion methods, respectively. The E-test may be suitable for replacing the proportion methods (7H11 agar and LJ medium) in routine practice due to its fast and easy application.