Gen-probe test should not be considered final in Mycobacterium tuberculosis identification.
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Biomedical subjects
Publications and source records attributed to L Heifets.
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MICs of isoniazid, rifampin, ethionamide, streptomycin, amikacin, kanamycin, and capreomycin were determined for Mycobacterium avium complex strains by two methods: broth dilution in 7H12 medium radiometrically and agar dilution on 7H10 agar plates. The broth-determined MICs of all drugs with the exception of isoniazid were two to eight times lower than the agar-determined MICs for most of the tested M. avium strains, which is probably due to the higher absorption and degradation of the drugs in solid media. The MICs, especially those determined in broth, are suggested as quantitative measurements of the degree of susceptibility of M. avium complex strains. For a certain percentage of the M. avium strains the broth-determined MICs were within the limits of MICs found for wild susceptible Mycobacterium tuberculosis strains. These M. avium strains were classified as presumably susceptible. In contrast to M. tuberculosis, the MICs for M. avium strains had a wide range. When the MICs for M. avium strains were only one dilution higher than those for M. tuberculosis, they were tentatively classified as moderately susceptible. The designation moderately resistant or resistant, respectively, is suggested for those M. avium strains for which the MICs were at or above the concentrations achievable in blood. The quantitation of the degree of susceptibility by the MICs and the tentative interpretation of the MICs are suggested for future use in clinical trials as a means of evaluating the patients' responses to chemotherapy compared with the degree of susceptibility of the initial strain isolated before treatment.
Three methods were used for the recovery of mycobacteria from blood specimens obtained from acquired immunodeficiency syndrome patients: (i) inoculation of 7H11 agar plates with a concentrated specimen, (ii) inoculation of 7H12 BACTEC vials with a concentrated specimen, and (iii) inoculation of 7H13 BACTEC vials with a nonconcentrated specimen. In this study, we examined 255 specimens and obtained positive mycobacterial growth in 47 of them. Among these 47 cultures, 40 were found to be positive by all three methods, and the total recovery rates in relation to these culture-positive specimens were 94% for method 1, 89% for method 2, and 96% for method 3. The advantages and disadvantages of these three methods are discussed.
Of 92 drug-resistant Mycobacterium tuberculosis strains isolated from sputum specimens, 86 showed growth in two types of 7H12 broth, one with egg yolk and the other without egg yolk. In addition, two strains grew only in plain 7H12 broth without yolk, and four others were recovered only in the medium supplemented with egg yolk. The radiometrically detected growth was higher in the presence of egg yolk, corresponding to a higher number of CFU per milliliter in these cultures. The improvement of growth in 7H12 broth supplemented with egg yolk was most noticeable in cultures isolated from sputum specimens having a low number of acid-fast bacilli in the smear and producing only a few colonies on solid media.
Qualitative methods of susceptibility testing have been employed widely around the world during the last 37 years and have proved to be a generally reliable tool to guide the chemotherapy of tuberculosis with conventional antituberculosis drugs. However, we believe there is a need now for application of modern quantitative methods to mycobacteriology: (1) for susceptibility testing of Mycobacterium avium complex, (2) for evaluation of the bactericidal activity of the antimycobacterial drugs, (3) for evaluation of the inhibitory and bactericidal activity of drug combinations, (4) for search and evaluation of new drugs against both Mycobacterium tuberculosis and Mycobacterium avium complex. Newly developed technology (radiometric automated bacterial growth recording in the BACTEC system) has opened new opportunities for development of quantitative methods in mycobacteriology using liquid culture medium. This paper analyzes the potential applications to mycobacteriology of 4 quantitative criteria: Minimal Inhibitory Concentration (MIC), Minimal Bactericidal Concentration (MBC), Fractional Inhibitory Concentration (FIC), and Fractional Bactericidal Concentration (FBC).
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A total of 463 respiratory specimens, all smear positive for acid-fast bacteria, were inoculated onto routine solid media and into BACTEC 7H12 Middlebrook medium for detection of mycobacterial growth. Conventional drug susceptibility testing (1% proportion method) was performed on Middlebrook 7H10/7H11 medium, and radiometric susceptibility testing was performed on BACTEC 7H12 medium. The average detection times for BACTEC-positive cultures were 8.3 days for Mycobacterium tuberculosis and 5.2 days for mycobacteria other than tuberculosis; by conventional methods, they were 19.4 and 17.8 days, respectively. These detection times do not include time required for identification, which was done by the conventional method only. There was an excellent correlation in the recovery rates of mycobacteria by the two methods. Drug susceptibility test results of M. tuberculosis isolates by the two methods showed 95.1 to 100% overall agreement. The average reporting time for drug susceptibility results ranged from 4.2 to 6.9 days for the BACTEC method and 13.7 to 21 days for the conventional methods. An average of 18 days was required by the BACTEC method for complete recovery and drug susceptibility testing of M. tuberculosis, as compared with 38.5 days for the conventional methods.
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Phagocytosis was discovered by Elie Metchnikoff (Ilia Mechnikov) in 1882. Although the phenomenon of endocytosis by the leukocytes (more related to the pinocytosis) had been described 30 years earlier [8], it was Metchnikoff who explained its function, and alerted the scientific community to the importance of phagocytosis in immunity. He devoted most of his life to studying different aspects of phagocytosis and related immunological phenomena. It required 25 years of intense effort to achieve recognition of the phagocytosis theory, the first experimentally based theory in immunology. This struggle culminated in 1908 with the awarding of the Nobel Prize to Metchnikoff and Ehrlich for their two theories of immunity, "cellular" and "humoral," respectively. That was an official recognition of the existence and importance of immunology, but modern immunology began at the moment of Metchnikoff's discovery, which is why 1982 may be considered the 100th year anniversary of immunology.
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Clarithromycin is known to accumulate in polymorphonuclear leukocytes, but no accumulation studies with macrophages have been reported. We exposed J774 macrophages, grown for 4-6 days, to clarithromycin 3.0 micrograms/ml for 2 hours. The cells were separated from the extracellular fluid, and the concentration of clarithromycin was determined in an agar diffusion bioassay. The accumulation of clarithromycin was 15.8-fold greater in the cells than it was in the extracellular fluid when the test was performed with noninfected cells, and 17.3-fold greater for cells infected with Mycobacterium avium. However, the ratio was substantially lower, only 3.7 for dead macrophages, suggesting that intracellular accumulation is probably an active process. These data may clarify the nature of the activity of clarithromycin against M. avium in macrophages.