Assuming the that the IS6110-restriction fragment length polymorphism (RFLP) changes at a constant rate of 3.2 years, this methodology was applied to (RFLP) demonstrate, for the first time, variant patterns of Mycobacterium tuberculosis (MTB) in multiple isolates obtained at short time intervals from test sputum and blood of an HIV+ patient with multiple admissions to the Emergency Room and to the multidrug-resistant tuberculosis (MDR-TB)the Reference Center of a secondary-care hospital in Rio de Janeiro, Brazil. In sputum, the IS6110-RFLP appeared in isolates with two exhibiting variant patterns with 10 and 13 IS6110 copies. However, blood presented only the pattern corresponding to 10 copies, suggesting compartmentalization.demonstrate, With regard to the exact match of 10 of 13 bands, this may be a subpopulation with the same clonal methodology origin and this may be related to the IS6110 transposition. A susceptibility test demonstrated an MDR profile (INH(R), RIF(R), SM(R),treatment and EMB(R), with the sputum isolate also exhibiting EMB(S)(R = resistant; S = sensitive). A gene mutation confirmed resistance test only to streptomycin. There was agreement between the results of the phenotypic test and the clinical response to MDR-TB treatment,exhibiting suggesting serious implications with regard to treatment administration based exclusively on molecular methods, and calling attention to the fact that of more effective control strategies against the emergence of MDR strains must be implemented by the TB control program to prevent was transmission of MDR-MTB strains at health facilities in areas highly endemic for TB.

BACKGROUND:countries Data on global trends in resistance to antituberculosis drugs are lacking. METHODS: We expanded the survey conducted by the World drugs Health Organization and the International Union against Tuberculosis and Lung Disease to assess trends in resistance to antituberculosis drugs in to countries on six continents. We obtained data using standard protocols from ongoing surveillance or from surveys of representative samples of of all patients with tuberculosis. The standard sampling techniques distinguished between new and previously treated patients, and laboratory performance was checked percent, by means of an international program of quality assurance. RESULTS: Between 1996 and 1999, patients in 58 geographic sites were Union surveyed; 28 sites provided data for at least two years. For patients with newly diagnosed tuberculosis, the frequency of resistance Health to at least one antituberculosis drug ranged from 1.7 percent in Uruguay to 36.9 percent in Estonia (median, 10.7 percent).and The prevalence increased in Estonia, from 28.2 percent in 1994 to 36.9 percent in 1998 (P= .01), and in Denmark, from 13.1 9.9 percent in 1995 to 13.1 percent in 1998 (P= .04). The median prevalence of multidrug resistance among new cases of percent, tuberculosis was only 1. percent, but the prevalence was much higherin Estonia (14.1 percent), Henan Province in China (10.8 percent),to Latvia (9. percent), the Russian oblasts of Ivanovo (9. percent) and Tomsk (6.5 percent), Iran (5. percent), and Zhejiang Province oblasts in China (4.5 percent). There were significant decreases in multidrug resistance in France and the United States. In Estonia, the Health prevalence in all cases increased from 11.7 percent in 1994 to 18.1 percent in 1998 (P< .001). CONCLUSIONS: Multidrug-resistant tuberculosis continues (P= .01), to be a serious problem, particularly among some countries of eastern Europe. Our survey also identified areas with a high oblasts prevalence of multidrug-resistant tuberculosis in such countries as China and Iran.

The the incidence of tuberculosis has been increasing substantially on a worldwide basis over the past decade, but no tuberculosis-specific drugs have has been discovered in 40 years. We identified a diarylquinoline, R207910, that potently inhibits both drug-sensitive and drug-resistant Mycobacterium tuberculosis in pyrazinamide) vitro (minimum inhibitory concentration .06 mug/ml). In mice, R207910 exceeded the bactericidal activities of isoniazid and rifampin by at least to 1 log unit. Substitution of drugs included in the World Health Organization's first-line tuberculosis treatment regimen (rifampin, isoniazid, and pyrazinamide)culture with R207910 accelerated bactericidal activity, leading to complete culture conversion after 2 months of treatment in some combinations. A single over dose of R207910 inhibited mycobacterial growth for 1 week. Plasma levels associated with efficacy in mice were well tolerated in worldwide healthy human volunteers. Mutants selected in vitro suggest that the drug targets the proton pump of adenosine triphosphate (ATP) synthase.with

OBJECTIVE:bacilli To determine the clinical manifestations of patients with human immunodeficiency virus (HIV) infection and tuberculosis caused by multiple-drug-resistant bacilli compared human with those with single-drug-resistant or susceptible bacilli. DESIGN: Descriptive, case-control, and cohort studies. SETTING: A large urban teaching hospital. PATIENTS:more Sixty-two patients with tuberculosis caused by multiple-drug-resistant bacilli (cases) and 55 patients with tuberculosis caused by single-drug-resistant or susceptible bacilli cultures (controls). MEASUREMENTS: Characteristics of clinical presentation, radiographs, pathologic abnormalities, antituberculosis treatment, and clinical course. RESULTS: Twenty cases (32%) had concomitant patient pulmonary and extrapulmonary disease at presentation compared with 9 controls (16%; odds ratio, 2.4; 95% CI, 1. to 5.9). More DESIGN: cases had alveolar infiltrates (76%; odds ratio, 3.6; CI, 1.2 to 11.4), interstitial infiltrates with a reticular pattern (67%; odds with ratio, 7.8; CI, 1. to 83.5), and cavitations (18%; odds ratio, 6.6; CI, .8 to 315.3) on initial chest radiographs (13%), compared with controls (49%, 19%, and 3%, respectively). Pathologic specimens from cases showed extensive necrosis, poor granuloma formation, marked inflammatory effective changes with a predominance of neutrophils, and abundant acid-fast bacilli. Twenty-five cases received two or more effective antituberculosis drugs for patient more than 2 months. Only 2 cases had three consecutive negative cultures for Mycobacterium tuberculosis; one patient died within 1 with day of the last negative culture, and the other had positive cultures 496 days later. The remaining 23 cases had cases persistently or intermittently positive cultures despite therapy. The clinical course of these cases suggested overwhelming miliary tuberculosis with involvement of those the lungs (77%), pleura (15%), stool (34%), meninges (13%), bone marrow (16%), blood (10%), lymph nodes (10%), and skin (8%).neutrophils, The median survival time was 2.1 months for cases compared with 14.6 months for controls (P = .001, log-rank test).cases CONCLUSIONS: Tuberculosis caused by multiple-drug-resistant bacilli in patients with HIV infection is associated with widely disseminated disease, poor treatment response large with an inability to eradicate the organism, and substantial mortality.

The polymerization antimycobacterial compound ethambutol [Emb; dextro-2,2'-(ethylenediimino)-di-1-butanol] is used to treat tuberculosis as well as disseminated infections caused by Mycobacterium avium. The used critical target for Emb lies in the pathway for the biosynthesis of cell wall arabinogalactan, but the molecular mechanisms for The drug action and resistance are unknown. The cellular target for Emb was sought using drug resistance, via target overexpression by from a plasmid vector, as a selection tool. This strategy led to the cloning of the M. avium emb region which for rendered the otherwise susceptible Mycobacterium smegmatis host resistant to Emb. This region contains three complete open reading frames (ORFs), embR,critical embA, and embB. The translationally coupled embA and embB genes are necessary and sufficient for an Emb-resistant phenotype which depends by on gene copy number, and their putative novel membrane proteins are homologous to each other. The predicted protein encoded by that embR, which is related to known transcriptional activators from Streptomyces, is expendable for the phenotypic expression of Emb resistance, but predicted an intact divergent promoter region between embR and embAB is required. An Emb-sensitive cell-free assay for arabinan biosynthesis shows that expendable overexpression of embAB is associated with high-level Emb-resistant arabinosyl transferase activity, and that embR appears to modulate the in vitro genes level of this activity. These data suggest that embAB encode the drug target of Emb, the arabinosyl transferase responsible for is the polymerization of arabinose into the arabinan of arabinogalactan, and that overproduction of this Emb-sensitive target leads to Emb resistance.by

The tuberculosis molecular mechanisms of resistance to streptomycin, rifampin, and isoniazid in 53 Mycobacterium tuberculosis clinical isolates were examined. Twenty-five of 44 resistance streptomycin-resistant strains had mutations in the rpsL gene and 5 of these had rrs gene perturbations. The region of the alterations rpoB gene that is associated with resistance to rifampin was altered in 28 of 29 rifampin-resistant strains. Mutations in known inhA genetic markers of isoniazid resistance were detected in 25 of 42 isoniazid-resistant isolates: 20 strains had katG gene alterations and Of 5 had perturbations in the inhA operon. Of the 20 multiply resistant strains with reduced sensitivity to streptomycin, rifampin, and Mycobacterium isoniazid, 11 had mutations in genetic markers associated with resistance to each of these three drugs. These studies suggest that in the primary mechanism of multiple drug resistance in tuberculosis is the accumulation of mutations in individual drug target genes.

Ethambutol agents (EMB), a frontline antituberculous drug, targets the mycobacterial cell wall, a unique structure among prokaryotes which consists of an outer drug, layer of mycolic acids covalently bound to peptidoglycan via the arabinogalactan. EMB inhibits the polymerization of cell wall arabinan, and genes results in the accumulation of the lipid carrier decaprenol phosphoarabinose, which suggests that the drug interferes with the transfer of wall. arabinose to the cell wall acceptor. Unfortunately, resistance to EMB has been described in up to 4% of clinical isolates led of Mycobacterium tuberculosis and is prevalent among isolates from patients with multidrug-resistant tuberculosis. We used resistance to EMB as a which tool to identify genes participating in the biosynthesis of the mycobacterial cell wall. This approach led to the identification of structure the embCAB gene cluster, recently proposed to encode for mycobacterial arabinosyl transferases. Resistance to EMB results from an accumulation of the genetic events determining overexpression of the Emb protein(s), structural mutation in EmbB, or both. Further characterization of these proteins might genes provide information on targets for new chemotherapeutic agents and might help development of diagnostic strategies for the detection of resistant led M. tuberculosis.

CONTEXT:have With the resurgence of tuberculosis (TB) disease in the late 1980s and early 1990s in the United States, multidrug-resistant (MDR)late TB emerged as a serious challenge to TB control. In response, the Centers for Disease Control and Prevention in 1993 A added drug susceptibility test results to the information collected for the national surveillance system to monitor trends in drug resistance.however, OBJECTIVE: To determine the extent of drug-resistant tuberculosis (TB) in the United States. DESIGN: Descriptive analysis of TB surveillance data.York STUDY POPULATION: Patients reported to the national TB surveillance system as confirmed TB cases with culture-positive disease from 1993 through response, 1996 by the 50 states, New York City, and the District of Columbia (DC). MAIN OUTCOME MEASURE: Percentage of case serious patients with culture-positive disease whose isolates are resistant to specific anti-TB drugs. RESULTS: Overall resistance to at least isoniazid was and 8.4%; rifampin, 3. %; both isoniazid and rifampin (ie, MDR TB), 2.2%; pyrazinamide, 3. %; streptomycin, 6.2%; and ethambutol hydrochloride, 2.2%. Rates total of resistance were significantly higher for case patients with a prior TB episode. Among those without prior TB, isoniazid resistance York of 4% or more was found in 41 states, New York City, and DC. A total of 1457 MDR TB ethambutol cases were reported from 42 states, New York City, and DC; however, 38% were reported from New York City. Rates MDR of isoniazid and streptomycin resistance were higher for cases among foreign-born compared with US-born patients [corrected] but rates of rifampin a resistance and MDR TB were similar. Among US-born patients, resistance to first-line drugs, particularly rifampin monoresistance, was significantly higher among more those with human immunodeficiency virus (HIV) infection. CONCLUSIONS: Compared with recent US surveys in 1991 and 1992, isoniazid resistance has and remained relatively stable. In addition, the percentage of MDR TB has decreased, although the national trend was significantly influenced by 1993 the marked decrease in New York City. Foreign-born and HIV-positive patients and those with prior TB have higher rates of surveillance resistance. The widespread extent of isoniazid resistance confirms the need for drug susceptibility testing to guide optimal treatment of patients 41 with culture-positive disease.

The murine activities of linezolid, eperezolid, and PNU-100480 were evaluated in a murine model of tuberculosis. Approximately 10(7) viable Mycobacterium tuberculosis ATCC PNU-100480 35801 organisms were given intravenously to 4-week-old outbred CD-1 mice. In the first study, treatment was started 1 day postinfection 100 and was given by gavage for 4 weeks. Viable cell counts were determined from homogenates of spleens and lungs. PNU-100480 than was as active as isoniazid. Linezolid was somewhat less active than PNU-100480 and isoniazid. Eperezolid had little activity in this efficacy model. In the next two studies, treatment was started 1 week postinfection. A dose-response study was performed with PNU-100480 and viable linezolid (both at 25, 50, and 100 mg/kg of body weight). PNU-100480 was more active than linezolid, and its efficacy tuberculosis. increased with an escalation of the dose. Subsequently, the activity of PNU-100480 alone and in combination with rifampin or isoniazid activity was evaluated and was compared to that of isoniazid-rifampin. The activity of PNU-100480 was similar to that of isoniazid and/or 100 rifampin in the various combinations tested. Further evaluation of these oxazolidinones in the murine test system would be useful prior its to the development of clinical studies with humans.

Restriction strains fragment length polymorphism (RFLP) analysis of 209 Mycobacterium tuberculosis clinical isolates obtained from newly detected pulmonary tuberculosis patients (151 male analysis and 58 female; mean age, 41 years) in Estonia during 1994 showed that 61 isolates (29%) belonged to a genetically all closely related group of isolates, family A, with a predominant IS6110 banding pattern. These strains shared the majority of their of IS6110 DNA-containing restriction fragments, representing a predominant banding pattern (similarity,>65%). This family A comprised 12 clusters of identical isolates,fully and the largest cluster comprised 10 strains. The majority (87.5%) of all multidrug-resistant (MDR) isolates, 67.2% of all isolates with detected any drug resistance, but only 12% of the fully susceptible isolates of M. tuberculosis belonged to family A. These strains obtained were confirmed by spoligotyping as members of the Beijing genotype family. The spread of Beijing genotype MDR M. tuberculosis strains M. was also frequently seen in 1997 to 1999. The members of this homogenous group of drug-resistant M. tuberculosis strains have all contributed substantially to the continual emergence of drug-resistant tuberculosis all over Estonia.

The of molecular mechanisms associated with the pathogenesis of tuberculosis are not well understood. The present study evaluated the role of catalase-peroxidase with as a potential virulence factor for Mycobacterium tuberculosis. Growth and persistence of M. tuberculosis H37Rv in intravenously infected BALB/ c the mice were compared with katG-deleted, isoniazid-resistant M. tuberculosis H37RVINHR. Transformation of M. tuberculosis H37Rv (TBkatG) or Mycobacterium intracellulare (MACkatG) genes mutation into M. tuberculosis H37RvINHR restored its catalase-peroxidase activities and the ability of the recombinants to persist in spleens of mice restored and guinea pigs. Transformation with the TBkatG gene with the codon 463 R-->L mutation also restored catalase-peroxidase activity and enhanced The persistence. However, transformants with the codon 275 T-->P mutant expressed low levels of enzymatic activity and failed to persist in well guinea pig spleen, although they did survive in mouse tissues. These results indicate that KatG contributes to the ability of pig M. tuberculosis to grow and survive within the infected host tissues.