Julian González-Martín

Epidemiology of uveitis in a Western urban multiethnic population. The challenge of globalization

To report the anatomical pattern and etiological spectrum of uveitis in an urban multi‐ethnic population from Barcelona, Spain. General and specific epidemiological data for the most prevalent aetiologies are also calculated.

Tuberculosis transmission patterns among Spanish‐born and foreign‐born populations in the city of Barcelona

During a 2-year period (2003-2004), tuberculosis (TB) transmission in Barcelona and the factors related to transmission among the Spanish- and foreign-born populations were studied by molecular epidemiology. Data were obtained from TB cases and Conventional Contact Tracing registries and genotyping was performed using restriction fragment length polymorphism (RFLP)-IS6110 and MIRU12 as a secondary typing method. Of the 892 TB cases reported, 583 (65.3%) corresponded to Spanish-born and 309 (34.6%) to foreign-born. Six hundred and eighty-seven cases (77%) were confirmed by culture. RFLP typing of 463/687 (67.4%) isolates was performed, revealing 280 (60.5%) unique and 183 (39.5%) shared patterns, which were grouped into 65 clusters. Spanish-born individuals were significantly more clustered than foreign-born individuals (44.6% vs. 28.8%; p 0.016). Clustering in foreign-born individuals was associated with HIV (p 0.051, odds ratio = 3.1, 95% confidence interval 1-10.9) and alcohol abuse (p 0.022), whereas, in the Spanish-born individuals, clustering was associated with age in the range 21-50 years, (p 0.024). Of the total clusters, 36/65 (55.3%) included only Spanish-born patients, whereas 22/65 (33.8%) included individuals from both populations. In mixed clusters, the index case was Spanish-born in 53% and foreign-born in 47%. Among the foreign-born, 2.8% were ill on arrival, 30% developed TB within the first year and 50.3% developed TB within the first 2 years; 58.3% were from South America. In conclusion, half of the foreign-born TB patients developed the disease during the first 2 years after arrival, which, in most cases, was the result of endogenous reactivation. Recent TB transmission among Spanish-born and foreign-born populations, as well as bidirectional transmission between communities, contributed significantly to the burden of TB in Barcelona, suggesting the need to improve Public Health interventions in both populations.

Pulmonary Infiltrates in Immunosuppressed Patients: Analysis of a Diagnostic Protocol

ABSTRACT A diagnostic protocol was started to study the etiology of pulmonary infiltrates in immunosuppressed patients. The diagnostic yields of the different techniques were analyzed, with special emphasis on the importance of the sample quality and the role of rapid techniques in the diagnostic strategy. In total, 241 patients with newly developed pulmonary infiltrates within a period of 19 months were included. Noninvasive or invasive evaluation was performed according to the characteristics of the infiltrates. Diagnosis was achieved in 202 patients (84%); 173 patients (72%) had pneumonia, and specific etiologic agents were found in 114 (66%). Bronchoaspirate and bronchoalveolar lavage showed the highest yields, either on global analysis (23 of 35 specimens [66%] and 70 of 134 specimens [52%], respectively) or on analysis of each type of pneumonia. A tendency toward better results with optimal-quality samples was observed, and a statistically significant difference was found in sputum bacterial culture. Rapid diagnostic tests yielded results in 71 of 114 (62.2%) diagnoses of etiological pneumonia.

Multicenter Laboratory Evaluation of the MB/BacT Mycobacterium Detection System and the BACTEC MGIT 960 System in Comparison with the BACTEC 460TB System for Susceptibility Testing of Mycobacterium tuberculosis

ABSTRACT In this multicenter study, the reliability of two nonradiometric, fully automated systems, the MB/BacT and BACTEC MGIT 960 systems, for testing the susceptibilities of 82 Mycobacterium tuberculosis strains to isoniazid, rifampin, ethambutol, and streptomycin was evaluated in comparison with the radiometric BACTEC 460TB system. The arbitration of discrepant results was done by the reanalysis of the strain, the determination of the MIC, and the molecular characterization of some resistance determinants. The overall level of agreement with BACTEC 460TB results was 96% with the MB/BacT test and 97.2% with the BACTEC MGIT 960 system. With both methods, the level of agreement with BACTEC 460TB results was 96.3% for isoniazid, 98.8% for rifampin, and 98.8% for ethambutol. The level of agreement for streptomycin was 90.2% with MB/BacT and 97.5% with BACTEC MGIT 960. Overall, there were 11 very major errors and 2 major errors with the MB/BacT method and 5 very major errors and 2 major errors with the BACTEC MGIT 960 system. In general, the MB/BacT and BACTEC MGIT 960 systems showed good performance for susceptibility testing with first-line antituberculosis drugs.

Simultaneous Identification of Mycobacterium Genus and Mycobacterium tuberculosis Complex in Clinical Samples by 5′-Exonuclease Fluorogenic PCR

ABSTRACT Early diagnosis of tuberculosis and screening of other mycobacteria is required for the appropriate management of patients. We have therefore developed a 5′-exonuclease fluorogenic PCR assay in a single-tube balanced heminested format that simultaneously detects Mycobacterium tuberculosis complex (MTC) and members of the Mycobacterium genus (MYC) using the 16S ribosomal DNA target directly on clinical samples. One hundred twenty-seven clinical samples (65 smear negative and 62 smear positive) with a positive culture result from 127 patients were tested, including 40 negative control specimens. The finding of both a positive MTC and probe value and a positive MYC probe value confirmed the presence of MTC or mycobacteria with a 100% positive predictive value. However, a negative value for MTC or MYC did not discount the presence of mycobacteria in the specimen. Interestingly, the addition of the MYC probe allowed the diagnosis of an additional 7% of patients with tuberculosis and rapid screening of nontuberculous mycobacteria (NTM). Thus, over 75% of the patients were diagnosed with mycobacterial disease by PCR. The sensitivity was much higher on smear-positive samples (90.3%) than smear-negative samples (49.2%) and was slightly higher for MTC than NTM samples. With regard to the origin of the sample, MTC pulmonary samples gave better results than others. In conclusion, we believe this test may be useful for the rapid detection of mycobacteria in clinical samples and may be a valuable tool when used together with conventional methods and the clinical data available.

Factors associated with differences between conventional contact tracing and molecular epidemiology in study of tuberculosis transmission and analysis in the city of Barcelona, Spain.

ABSTRACT The aim of this study was to analyze the factors associated with conventional contact tracing (CCT) and molecular epidemiology (ME) methods in assessing tuberculosis (TB) transmission, comparing the populations studied and the epidemiological links established by both methods. Data were obtained from TB case and CCT registries, and ME was performed using IS6110-based restriction fragment length polymorphism (RFLP) analysis and mycobacterial interspersed repetitive unit 12 (MIRU12) typing as a secondary typing method. During two years (2003 and 2004), 892 cases of TB were reported, of which 687 (77%) were confirmed by culture. RFLP analysis was performed with 463 (67.4%) of the 687 isolated strains, and MIRU12 types in 75 strains were evaluated; 280 strains (60.5%) had a unique RFLP pattern, and 183 (39.5%) shared patterns, grouping into 65 clusters. CCT of 613 (68.7%) of 892 cases detected 44 clusters involving 101 patients. The results of both CCT and ME methods yielded 96 clusters involving 255 patients. The household link was the one most frequently identified by CCT (corresponding to 80.7% of the cases clustered by this method), whereas nonhousehold and unknown links were associated with 94.1% of the strains clustered by ME. When both methods were used in 351 cases (39.3%), they showed the same results in 214 cases (61%). Of the remainder, 106 (30.2%) were clustered only by ME, 19 (5.5%) were clustered only by CCT, and 12 (3.4%) were clustered by both methods but into different clusters. Patients with factors potentially associated with social problems were less frequently studied by CCT (P = 0.002), whereas patients of <15 years of age, most with negative cultures, were less frequently studied by ME (P = 0.005). Significant differences in the populations studied by ME versus CCT were observed, possibly explaining the scarce correlation found between the results of these methods. Moreover, ME allowed the detection of nonhousehold contact relationships, whereas CCT was more useful for tracing transmission chains involving patients of <15 years of age. In conclusion, the two methods are complementary, suggesting the need to improve the methodology of contact study protocols.

In vitro effect of three-drug combinations of antituberculous agents against multidrug-resistant Mycobacterium tuberculosis isolates

Multidrug resistance has become a problem in the management of tuberculosis, leading to an urgent need for research related to new regimens including the currently available drugs. The objectives of this study were: (i) to study the effect of the following second-choice three-drug combinations against multidrug-resistant (MDR) and drug-susceptible clinical isolates (levofloxacin, linezolid and ethambutol; levofloxacin, amikacin and ethambutol; and levofloxacin, linezolid and amikacin); and (ii) to compare the effect of these combinations with an isoniazid, rifampicin and ethambutol combination against drug-susceptible clinical isolates. A total of 9 MDR clinical and 12 drug-susceptible isolates (11 clinical isolates and the H37Rv reference strain) were studied using an adaptation of the chequerboard assay. The fractional inhibitory concentration index (FICI) was calculated as follows: FICI=FIC(A)+FIC(B)+FIC(C)=A/MIC(A)+B/MIC(B)+C/MIC(C), where A, B and C are the minimum inhibitory concentrations (MICs) of each antibiotic in combination and MIC(A), MIC(B) and MIC(C) are the individual MICs. The FICI was interpreted as synergism when the value was <0.75. The FICI of all the combinations ranged from 1.5 to 3, showing indifferent activity. No differences were found between MDR and drug-susceptible isolates, or between the second-choice combinations and the fourth combination against drug-susceptible isolates. In conclusion, the second-choice drugs are equally effective as the combination of isoniazid, rifampicin and ethambutol.

Activity and interactions of levofloxacin, linezolid, ethambutol and amikacin in three-drug combinations against Mycobacterium tuberculosis isolates in a human macrophage model

Multidrug resistance is a problem in the management of tuberculosis, creating an urgent need for new regimens including currently available drugs. Macrophage models allow an evaluation of the effect of drugs against intracellular bacilli. The effect of the following different drug combinations against six multidrug-resistant and six drug-susceptible clinical isolates of Mycobacterium tuberculosis multiplying inside the human macrophage THP-1 cell line was studied: levofloxacin/linezolid/ethambutol; levofloxacin/amikacin/ethambutol; and levofloxacin/linezolid/amikacin. Macrophages were lysed, seeded onto Middlebrook 7H11 plates and CFU were counted after 21 days of incubation. The interaction of the drugs in combination was interpreted by the effect of the combination compared with the most active single drug alone. The antimicrobial activity of the drugs was evaluated comparing the log(10)CFU/well of the isolate with and without the drug. Drug concentrations within infected macrophages and in extracellular medium were simultaneously determined by chromatography. The levofloxacin/linezolid/amikacin and levofloxacin/linezolid/ethambutol combinations showed antagonism against most of the isolates (91.7%) after a 4-day protocol, whereas levofloxacin/amikacin/ethambutol displayed indifference. Levofloxacin alone and levofloxacin/amikacin/ethambutol were the most potent antimicrobials, presenting reductions up to 5.49 log(10) and 5.86 log(10), respectively. The drug penetration percentages ranged from 5.46% to 11.10%. Intracellular concentrations for the drug alone compared with those for the drugs in combination were not significantly different. All of the combinations tested against M. tuberculosis-infected macrophages showed antimicrobial activity, with combinations including linezolid and levofloxacin showing an antagonistic effect that may be explained by efflux transporters or changes in the macrophage environment.

Indirect supportive evidence for diagnosis of tuberculosis-related uveitis: from the tuberculin skin test to the new interferon gamma release assays.

Purpose:  To evaluate clinical and paraclinical parameters for the indirect diagnosis of tuberculosis‐related uveitis (TRU).

Evaluation of the VersaTREK System Compared to the Bactec MGIT 960 System for First-Line Drug Susceptibility Testing of Mycobacterium tuberculosis

ABSTRACT The aim of this study was to evaluate the reliability of the VersaTREK system for Mycobacterium tuberculosis drug susceptibility testing compared with results obtained with the Bactec MGIT 960 system. A total of 67 strains were evaluated. Overall agreement was at 98.5%. Kappa indexes were 1.0 for isoniazid, rifampin, and ethambutol, 0.937 for pyrazinamide, and 0.907 for streptomycin. The VersaTREK system is validated for M. tuberculosis drug susceptibility testing.