[Isolation and identification of non-tuberculosis mycobacteria].

Seeding of 117,579 samples of clinical materials at the Central Microbiological Laboratory, Moscow Research and Practical Centre of Tuberculosis, resulted in isolation of 15,786 (13,4%) mycobacterial cultures, with 15,333 (97.1%) ones being identified as M. tuberculosis complex and 490 (3.1%) as non-tuberculosis mycobacteria (NTM). Samples collected in the Moscow region were dominated by MAC, M. kansasii, M. xenopi, and M. fotuitum. Cultivation on solid and liquid media in an automated regime was shown to be optimal for isolation of NTM. Comparison of mycobacteria identification by microbiological methods and by high performance liquid chromatography revealed excellent agreement between the results (96.1%), with the latter technique taking less time to obtain them. Agreement between identification of acid-resistant mycobacterial cultures using the MAIS-dif test system and microbiological methods was estimated at 93.5%. IMS biochips may also be used to detect species composition of myobacteria because only one case of disagreement with the results of microbiological methods was documented in the study of 108 cultures by the two techniques.

[The centralized mycobacteriological laboratory is a necessary component of a phthisiological service in large towns of Russia].

The paper presents the main points of the authors' own concept of the centralization of mycobacteriological service in large towns of the Russian Federation. The main points of step-by-step organizational and methodological measures required to solve this problem are described in detail. Consecutive measures to realize the proposed mycobacteriological service centralization model originated in January 2004 on a model of the Moscow Eastern Administrative District with 1380 thousand inhabitants are described.

[Use of molecular genetic studies to determine the species of the mycobacteria mais and M. tuberculosis complex].

The genus Mycobacterium currently comprises more than 90 species of Mycobacterium, of which a third is able to induce human diseases. With a rise in the incidence of diseases induced by non-tuberculosis mycobacteria, tuberculosis caused by M. bovis that is characterized by a severe cause and a high frequency of poor outcomes cannot be remembered. The species of mycobacteria should be identified to establish a diagnosis and to prescribe adequate chemotherapy. For this purpose, cultural, biochemical, chromatographic, and molecular genetic studies are conducted. The present study using the hsp65 gene restriction fragment length polymorphism test on museum mycobacterial strains and strains isolated from the diagnostic material of patients with suspected tuberculosis by means of Hind61 restrictase has provided a clear differentiation of the restriction profiles of MAIS complex mycobacteria and some other species of non-tuberculosis mycobacteria. To determine the species of representatives of M. tuberculosis complex (M. tuberculosis, M. bovis, BCG M. bovis), the authors have successfully used the test system "TUB-dif" developed at the Institute of Molecular Genetics, Russian Academy of Sciences, by applying the chain polymerase reaction of the senX3-regX3 region.

[The composition and drug sensitivity of a mycobacterial population in patients with suspected tuberculosis].

By using the diagnostic material (175 sputum samples and 103 bronchoalveolar lavage fluid samples) taken from 39 patients with suspected tuberculous infection during a 2.5-month follow-up, the authors traced the time course of changes in the composition and drug sensitivity of a mycobacterial population to rifampicin. Along with the traditional microbiological studies, the latest molecular biological studies, a TB-BIOCHIP test system (enzyme immunoassay) in particular, were employed to detect the bacterial and L-transformed forms of the causative agent. A molecular biological assay was first developed to detect the drug sensitivity of L-forms of Mycobacterium tuberculosis.

[Analysis of risk factors of the occurrence of drug resistance in patients with tuberculosis from civil and penitentiary sectors in the Samara Region].

The true prevalence rates of multidrug-resistant tuberculosis (MDRT) are unknown for most regions of Russia. This study was conducted in the Samara Region that differs from other regions in the rapid spread of HIV infection. The purpose of this study was to determine the primary and acquired resistance of Mycobacterium tuberculosis (MBT) to first-line antituberculous drugs in patients from civil and penitentiary sectors and to reveal risk factors of drug resistance of MBT. Six hundred patients (309 civilians and 291 prisoners who had been bacteriologically diagnosed as having tuberculosis. The authors have established the following:--in new cases, primary drug resistance is as follows: to isoniazid [38.9% (95% CI, 31.3-36.9%)], to rifampicin [25.9% (95% CI, 19.4-33.4%)] and to MDRT [23.0% (95% CI, 16.7-30.3%)];--in prisoners, the primary resistance of MBT was statistically more significant than in civilians;--male sex, in adequate prior or current treatment for tuberculosis for more than 4 weeks, the presence of fibrocavernous tuberculosis and previous prison stay are essential risk factors of the development of resistance of MBT to both any first-line drug and MDRT;--HIV infection is unassociated with resistance.

Drug-resistant strains of Mycobacterium tuberculosis isolated in Russia.

SETTING: State Research Center for Applied Microbiology, Russian Research Institute of Phthisiopulmonology (Ministry of Health, Moscow). OBJECTIVE: To analyze drug-resistant clinical isolates of Mycobacterium tuberculosis obtained from patients referred to the institute from different parts of Russia, and to study the mechanisms of their rifampicin resistance. DESIGN: Fifty clinical isolates of M. tuberculosis were analysed. Polymerase chain reaction (PCR) and sequencing were used to study the mechanisms of rifampicin resistance in 25 isolates. RESULTS: Among cultures isolated from 50 patients, drug resistance was detected in 33. Most of the isolates were resistant to rifampicin (25 isolates), isoniazid (14 isolates), and streptomycin (seven isolates). Only 6% of the isolates were resistant to one drug, while 14% were resistant to two, 32% to three, 40% to four, and 8% to five drugs. Susceptible isolates were derived from 17 patients. The following point mutations and deletions in the rpoB locus, responsible for high level rifampicin resistance (more than 50 microg/ml in egg-based medium), were detected: G-->A/395 (Arg-->Gln), C-->T/232 (His-->Tyr), C-->T/221 (Ser-->Leu), G-->T/202 (Asp-->Tyr), GA-->TT/202-203 (Asp-->Phe), deltaATGGACCAG/199-207 (Met, Asp, Gin), A-->T/91 (Met-->Leu), TG-->CC/227-228 (Leu-->Ser), GAG-->AGT/349-350-351 (Gln-->Ser), deltaGGG/354(Gly). CONCLUSION: A number of previously unrecognised genetic modifications in the rpoB region were found in rifampicin-resistant strains isolated from patients from different parts of Russia.

[L-transformation of mycobacteria in the light of current epidemiological situation of tuberculosis in the world]. / L-transformatsiia mikobakterii v svete sovremennoi épidemiologicheskoi situatsii po tuberkuleze v mire.

To determine a role of L-forms of Mycobacterium tuberculosis in the development of a recurrent tuberculous process in persons with residual pulmonary tuberculous changes, a total of 2,412 persons registered at a dispensary as those included into Groups VIIA and VIIB who were found to have a significant tuberculosis infection pool. The tuberculosis pathogen was detected in 214 (9%) examinees in the bacterial and L forms. A further follow-up of the persons whose pathological material had shown the L form which are prone to reverse indicated that the isolation of unstable mycobacterial L forms is an important predictive sign showing the high potential hazard of tuberculous process reactivation.

[Quantitative technic in the diagnosis of nonspecific lung diseases]. / Kolichestvennyi metod issledovanii v diagnostike nespetsificheskikh zabolevanii legkikh.

A total of 356 sputum samples from patients with nonspecific pulmonary diseases and tuberculosis were used in tests carried out in order to compare the method of qualitative analysis for the detection of nonspecific microflora and the method of quantitative estimation proposed by Dixon and Miller. In the process of this work the guantitative method was rationally modified, which allowed to decrease its cost while making it more reliable. The main advantage of the proposed modification lies in the possibility to greatly facilitate the detection of the etiological agent of the inflammatory process.

[Validation and development of an immunofluorescence method for isolating and identifying mycobacteria L forms]. / Obosnovanie i razrabotka vyiavleniia i identifikatsii L-form mikobakterii metodom immunofliuorestsentsii.

The complement fixation test and the immunofluorescence test have demonstrated that the L-forms of mycobacteria retain their species-specific and genus-specific determinants and possess serological activity. The L-variants obtained by different methods differ in size, depending on the degree of the destruction of their cell wall. Specific antisera to the L-forms of mycobacteria, suitable for use in the indirect immunofluorescence test, have been obtained. These antisera are highly specific and permit not only the rapid detection, but also identification of the L-forms.

[Pathomorphology of pulmonary aspergillosis in tuberculosis patients based on according to resection material]. / Patomorfologiia aspergilleza legkikh u bol'nykh tuberkulezom po materialam rezektsii

To investigate various manifestations of broncho-pulmonary aspergillosis and to ascertain its association with a previous disease there were carried out histotopographic, histological and mycological studies of specimens of the lung tissue resected in 37 patients in connection with aspergillomas of the lungs and broncho-pulmonary aspergillosis. The investigations showed that aspergillosis was one of the forms of dysbacteriosis in the human organism and could develop against the background of post-tuberculous, tuberculous and nonspecific changes. The development of aspergillosis following post-tuberculous changes should be considered as a second nonspecific disease pathogenetically associated with tuberculosis, but with an etiological factor ot its own. There are the following forms of broncho-pulmonary aspergillosis: aspergillomas, isolated aspergillosis of the bronchi, extensive aspergillosis of the bronchi and cyst-like cavities, aspergillous pneumonia.

[Comparative effectiveness of different methods for the microbiological study of resected pulmonary tuberculomas]. / Sravnitel'naia éffektivnost' razlichnykh metodov mikrobiologicheskogo issledovaniia rezetsirovannykh tuberkulem legkikh.

Wide-range microbiological study (bacterioscopy, inoculation, biological assays) of 114 lung tuberculomas excised from 107 patients has revealed a pronounced variability and sharply decreased viability of mycobacterial populations vegetating in caseous foci. Differences in the frequency and character of the detection of Mycobacterium tuberculosis and their altered forms, arising from the use of three above-mentioned methods of microbiological investigation, were noted. Bacterioscopy proved to be more informative with respect to the detection of the bacterial forms of M. tuberculosis. Biological assay was highly sensitive with respect to the L-forms of M. tuberculosis and permitted the detection of the persisting forms of this infective agent, contained in caseous foci and not detected by the method of inoculations. To evaluate the actual state of the mycobacterial population in the focus of tuberculous lesion, the use of a complex of microbiological methods for the detection of typical and biologically altered forms of M. tuberculosis is necessary.

[The use of molecular biological methods for the individual characterization of Mycobacterium tuberculosis strains]. / Ispol'zovanie molekuliarno-biologicheskikh metodov dlia individual'noi kharakteristiki shtammov Mycobacterium tuberculosis.

The expediency of using molecular biological methods for the evaluation of M. tuberculosis clinical strains by individual genetic certification of circulating M. tuberculosis strains has been substantiated. Considerable genetic heterogeneity of M. tuberculosis strains isolated from patients in different regions of the Russian Federation has been established; this heterogeneity is due to the presence of differences in the number of copies (5-26) of element IS6110 in M. tuberculosis cells.

[Prevalence of bronchopulmonary aspergillosis at the pulmonological hospital]. / O rasprostranennosti bronkholegochnogo aspergilleza v pul'monologicheskom statsionare.

The prevalence and the character of aspergillous infection were studied among 64 patients (33 men and 31 women) seen at the pulmonological hospital. Ten patients of that group were diagnosed to have bronchopulmonary aspergillosis. In all the cases, the disease was regarded as secondary one: 2 patients suffered from chronic aspergillous pneumonia and 8 patients from different patterns of aspergillous bronchitis. The risk factors of the disease development are described. The data obtained indicate that secondary bronchopulmonary aspergillosis occurs not infrequently. The estimation of the risk factors, careful clinical examination, and specific mycologic and serologic tests allow an early diagnosis of bronchopulmonary aspergillosis followed by institution of adequate treatment.

[New technologies in the determination of drug susceptibility in Mycobacterium tuberculosis]. / Novye tekhnologii opredeleniia lekarstvennoi chuvstvitel'nosti Mycobacterium tuberculosis.

A variety of mutations in the genes rpoB, katG, inhA, ahpC, kasA was studied by using different molecular biological methods (conformational polymorphism of single-chain fragments, heteroduplex analysis, biochips) in rifampicin- and isoniazid-resistant Mycobacterium tuberculosis (MBT) strains isolated from patients with pulmonary tuberculosis. Twenty-nine mutation combinations were identified in the MBT strains. The use of biochips is the most promising method for identifying the type of mutations responsible for the simultaneous resistance to rifampicin and isoniazid. Detection of several MBT strains in one patient requires the use a combination of molecular biological and microbiological studies.

[Mycobacteria in the development of periodontal foci of infection]. / Mikobakterii v razvitii okolozubnykh ochagov infektsii.

A total of 162 samples of material from the periodontal foci of infection were examined microbiologically and morphologically to detect mycobacteria and their biologically changed variants. The regularities of the pathologic process development in periodontal tissues of subjects without tuberculosis were found to fully conform to the current concept of a latent tuberculous infection. The periodontal foci of infection may be regarded as reservoirs of persistent mycobacteria, whose main form, L variant, was detected in 71.6% of cases. Clinical and morphologic manifestations of periodontitis depend on the stability of the persistent mycobacteria L forms.