GenoType MTBDRsl for detection of second-line drugs and ethambutol resistance in multidrug-resistant Mycobacterium tuberculosis isolates at a high-throughput laboratory

We assessed the performance of MTBDRsl for detection of resistance to fluoroquinolones, aminoglycosides/cyclic peptides, and ethambutol compared to BACTEC MGIT 960 by subjecting simultaneously to both tests 385 phenotypically multidrug-resistant-Mycobacterium tuberculosis isolates from Sao Paulo, Brazil. Discordances were resolved by Sanger sequencing. MTBDRsl correctly detected 99.7% of the multidrug-resistant isolates, 87.8% of the pre-XDR, and 73.9% of the XDR. The assay showed sensitivity of 86.4%, 100%, 85.2% and 76.4% for fluoroquinolones, amikacin/kanamycin, capreomycin and ethambutol, respectively. Specificity was 100% for fluoroquinolones and aminoglycosides/cyclic peptides, and 93.6% for ethambutol. Most fluoroquinolone-discordances were due to mutations in genome regions not targeted by the MTBDRsl v. 1.0: gyrA_H70R and gyrB_R446C, D461N, D449V, and N488D. Capreomycin-resistant isolates with wild-type rrs results on MTBDRsl presented tlyA mutations. MTBDRsl presented good performance for detecting resistance to second-line drugs and ethambutol in clinical isolates. In our setting, multidrug-resistant. isolates presented mutations not targeted by the molecular assay.

Transmission and prevalence of drug-resistant tuberculosis in a Brazilian setting under a directly observed therapy short-course strategy

Abstract INTRODUCTION: We aimed to estimate the prevalence and transmission of drug-resistant tuberculosis in a high-burden Brazilian setting under directly observed therapy short-course strategy. METHODS: Isolates of culture-confirmed pulmonary tuberculosis patients from Guarulhos, Brazil, diagnosed in October 2007-2011 were subjected to drug susceptibility and IS6110-restriction fragment length polymorphism testing. RESULTS: The overall resistance prevalence was 11.5% and the multi-drug resistance rate was 4.2%. Twenty-six (43.3%) of 60 drug-resistant isolates were clustered. Epidemiological relationships were identified in 11 (42.3%) patients; 30.8% of the cases were transmitted in households. CONCLUSIONS: Drug-resistant tuberculosis was relatively low and transmitted in households and the community.

Speeding up the diagnosis of multidrug-resistant tuberculosis in a high-burden region with the use of a commercial line probe assay / Agilizando o diagnóstico da tuberculose multirresistente em uma região endêmica com o uso de um teste comercial de sondas em linha

ABSTRACT Objective: To evaluate the rapid diagnosis of multidrug-resistant tuberculosis, by using a commercial line probe assay for rifampicin and isoniazid detection (LPA-plus), in the routine workflow of a tuberculosis reference laboratory. Methods: The LPA-plus was prospectively evaluated on 341 isolates concurrently submitted to the automated liquid drug susceptibility testing system. Results: Among 303 phenotypically valid results, none was genotypically rifampicin false-susceptible (13/13; 100% sensitivity). Two rifampicin-susceptible isolates harboured rpoB mutations (288/290; 99.3% specificity) which, however, were non-resistance-conferring mutations. LPA-plus missed three isoniazid-resistant isolates (23/26; 88.5% sensitivity) and detected all isoniazid-susceptible isolates (277/277; 100% specificity). Among the 38 (11%) invalid phenotypic results, LPA-plus identified 31 rifampicin- and isoniazid-susceptible isolates, one isoniazid-resistant and six as non-Mycobacterium tuberculosis complex. Conclusions: LPA-plus showed excellent agreement (≥91%) and accuracy (≥99%). Implementing LPA-plus in our setting can speed up the diagnosis of multidrug-resistant tuberculosis, yield a significantly higher number of valid results than phenotypic drug susceptibility testing and provide further information on the drug-resistance level.

RESUMO Objetivo: Avaliar o diagnóstico rápido de tuberculose multirresistente, utilizando um teste comercial de sondas em linha (LPA-plus), na rotina de um laboratório de referência de tuberculose. Métodos: O teste LPA-plus foi avaliado prospectivamente em 341 isolados simultaneamente submetidos ao teste de suscetibilidade aos antimicrobianos em meio líquido, pelo sistema automatizado. Resultados: Entre os 303 resultados fenotipicamente válidos, nenhum foi genotipicamente falso suscetível à rifampicina (13/13; 100% de sensibilidade). Dois isolados sensíveis à rifampicina apresentavam mutações no gene rpoB (288/290; especificidade de 99,3%), as quais, no entanto, não são associadas à resistência a rifampicina. O LPA-plus não identificou resistência à isoniazida em três isolados fenotipicamente resistentes (23/26; 88,5% de sensibilidade) e detectou todos os isolados sensíveis à isoniazida (277/277; especificidade de 100%). Entre os 38 (11%) resultados fenotípicos inválidos, o LPA-plus identificou 31 isolados sensíveis à rifampicina e à isoniazida, um resistente à isoniazida e seis como micobactérias não tuberculosas. Conclusões: O LPA-plus mostrou excelente concordância (≥91%) e acurácia (≥99%). Sua implementação pode acelerar o diagnóstico da tuberculose multirresistente, produzir número significativamente maior de resultados válidos do que o teste fenotípico de suscetibilidade aos antimicrobianos e fornecer informações adicionais sobre o nível de resistência aos fármacos.

Evaluation of Haemophilus influenzae type b carrier status among children 10 years after the introduction of Hib vaccine in Brazil

The aim of the present study was to assess the prevalence of Haemophilus influenzaetype b (Hib) nasopharyngeal (NP) colonisation among healthy children where Hib vaccination using a 3p+0 dosing schedule has been routinely administered for 10 years with sustained coverage (> 90%). NP swabs were collected from 2,558 children who had received the Hib vaccine, of whom 1,379 were 12-< 24 months (m) old and 1,179 were 48-< 60 m old. Hi strains were identified by molecular methods. Hi carriage prevalence was 45.1% (1,153/2,558) and the prevalence in the 12-< 24 m and 48-< 60 m age groups were 37.5% (517/1,379) and 53.9% (636/1,179), respectively. Hib was identified in 0.6% (16/2,558) of all children in the study, being 0.8% (11/1,379) and 0.4% (5/1,179) among the 12-< 24 m and 48-< 60 m age groups, respectively. The nonencapsulate Hi colonisation was 43% (n = 1,099) and was significantly more frequent at 48-< 60 m of age (51.6%, n = 608) compared with that at 12-< 24 m of age (35.6%, n = 491). The overall resistance rates to ampicillin and chloramphenicol were 16.5% and 3.7%, respectively; the co-resistance was detected in 2.6%. Our findings showed that the Hib carrier rate in healthy children under five years was very low after 10 years of the introduction of the Hib vaccine.

Tuberculose, HIV e coinfecção por TB/HIV no Sistema Prisional de Itirapina, São Paulo, Brasil / Tuberculosis, HIV and TB/HIV co-infection in the Prison System of Itirapina, São Paulo, Brazil

As prevalências do HIV, da tuberculose (TB) e da coinfecção TB/HIV foram avaliadas no Sistema Prisional de Itirapina/SP (Penitenciárias I e II). Foi efetuado estudo retrospectivo dos registros dos exames realizados no IAL-Rio Claro no período de janeiro/2003 a dezembro/2010. A baciloscopia/cultura para TB foi realizada em 24,3 % (1.375 amostras) da PI e em 25, 9 % (3.332 amostras) da PII, e a sorologia para HIV, em 1.810 amostras da PI (32,0 %) e em 2.634 (20,4 %) da PII. Foram detectadas 157 culturas positivas para micobactérias (3,3 %), 177 amostras soropositivas para HIV (4,0 %), e 16 positivas para TB/HIV (11,5 %). A prevalência de TB foi de 2,8 % e 3,6 % (p=0,160), de HIV foi de 2,1 % e 5,3 %(p<0,001), respectivamente, para PI e PII; e a de coinfecção TB/HIV foi 0,4 % (p=0,744) em PI e PII. A soroprevalência de HIV nos indivíduos com TB foi de 13,2 % em PI e 9,2 % (p=0,487) em PII; a prevalência maior em PII sugere que o comportamento da doença difere com a característica populacional. Elevada prevalência de TB, HIV e coinfecção TB/HIV mostra a importância do diagnóstico destas infecções em todos os indivíduos no momento da admissão para definir as medidas de prevenção e de tratamentos dos internos e seus contatos.

HIV, tuberculosis (TB) and TB/HIV co-infection prevalences were assessed in prisons PI and PII inItirapina, SP, Brazil. This retrospective study was performed by consulting the records of diagnosticassays carried out at the Instituto Adolfo Lutz (IAL) Rio Claro Laboratory from January 2003 toDecember 2010. Culture for TB was performed on 24.3 % (1,375 samples) of PI and 25.8 % (3,332samples) of PII, and HIV serology in 32.0 % (1,810 samples) of PI and 20.4 % (2,634 samples) ofPII. There were 177 (4.0 %) HIV positive samples, 157 (3.3 %) positive culture for mycobacteria,and 16 (11.5 %) for TB/HIV. The prevalences of TB were 2.8 % and 3.6 % (p=0.160) for PI and PIIrespectively; of HIV, 2.1 % and 5.3 % (p<0.001) for PI and PII; and of TB/HIV co-infection, 0.4 %(p=0.744) for both PI and PII. Among TB patients, HIV prevalence was 13.2 % in PI and 9.2 %(p=0.487) in PII. Higher HIV prevalence in PII suggests that it depends on the population characteristics.The high prevalence rates of TB, HIV and TB/HIV show that it is crucial to perform the diagnostic testingin all of imprisoned individuals at admission to define the disease treatment and prevention measuresamong inmates and their contacts.

Resistencia a antibióticos no betalactámicos de aislamientos invasores de Streptococcus pneumoniae en niños latinoamericanos: SIREVA II, 2000-2005 / Resistance to non-beta-lactam antibiotics in the clinical isolates of Streptococcus pneumoniae of children in Latin America: SIREVA II, 2000-2005

OBJETIVO. Determinar la evolución de la resistencia a la eritromicina, el cloranfenicol, el trimetoprim-sulfametozaxol (SXT) y la vancomicina de aislamientos invasores de Streptococcus pneumoniae obtenidos de niños de 10 países de América Latina y del Caribe en seis años de vigilancia. MÉTODOS. Se analizaron 8 993 aislamientos de S. pneumoniae recuperados entre 2000 y 2005 de niños menores de 6 años con infecciones invasoras, procedentes de Argentina, Brasil, Chile, Colombia, Cuba, México, Paraguay, República Dominicana, Uruguay y Venezuela. La sensibilidad a los antibióticos se determinó mediante los métodos establecidos y estandarizados en el proyecto SIREVA. La resistencia a múltiples antibióticos se definió como la resistencia a tres o más familias de antibióticos, de los no betalactámicos analizados en este estudio o de los betalactámicos evaluados en un estudio previo en el que 37,8% de estos aislamientos presentaron sensibilidad disminuida a la penicilina. RESULTADOS. Se encontró algún grado de resistencia al SXT y la eritromicina (56,4% y 15,4% de los aislamientos estudiados, respectivamente) y 4,6% presentó alta resistencia al cloranfenicol. Todos los aislamientos fueron sensibles a la vancomicina. Se observó la mayor frecuencia de resistencia al SXT en los aislamientos de neumonía y a la eritromicina en los casos de sepsis (61,6% y 25,5%, respectivamente; P < 0,01). La mayor frecuencia de resistencia al SXT se observó en Brasil (71,9%) y a la eritromicina en México (38,2%) y Venezuela (32,9%). Los serotipos 14, 6B, 19F y 23F fueron los que más frecuentemente se asociaron con la resistencia a los antibióticos estudiados. CONCLUSIONES. Se observó una elevada y creciente frecuencia de aislamientos resistentes al SXT y la eritromicina, y una disminución en la proporción de aislamientos resistentes al cloranfenicol. Estas tendencias mostraron diferencias entre los países estudiados.

OBJECTIVE. To examine the development of resistance to erythromycin, chloramphenicol, trimethoprim-sulfamethoxazole (TMP-SMZ), and vancomycin of the invasive isolates of Streptococcus pneumoniae obtained from children in 10 Latin American/Caribbean countries during six years of surveillance. METHODS. Analysis of 8 993 isolates of S. pneumoniae recovered in 2000­2005 from children with invasive infections, who were less than 6 years of age, and from Argentina, Brazil, Chile, Colombia, Cuba, Dominican Republic, Mexico, Paraguay, Uruguay, or Venezuela. Antibiotic susceptibility was determined through the methods established and standardized by the SIREVA project. Multidrug resistance was defined as: resistance to three or more antibiotics of the same class; to the non-beta-lactams analyzed by this study; or, to the beta-lactams evaluated by a previous study, in which 37.8% of these isolates showed decreased susceptibility to penicillin. RESULTS. Some degree of resistance was found to TMP-SMZ and erythromycin (56.4% and 15.4% of the isolates studied, respectively), with 4.6% highly resistant to chloramphenicol. All isolates were susceptible to vancomycin. The highest prevalence of TMP-SMZ resistance was observed in the pneumonia isolates; and that of erythromycin, in cases of sepsis (61.6% and 25.5%, respectively; P < 0.01). The highest prevalence of TMP-SMZ resistance was found in Brazil (71.9%), and that of erythromycin, in Mexico (38.2%) and Venezuela (32.9%). The 14, 6B, 19F, and 23F serotypes were most often associated with resistance to the antibiotics in the study. CONCLUSIONS. High and increasing rates of isolates resistant to TMP-SMZ and erythromycin were observed, as well as a decreasing percentage of isolates resistant to chloramphenicol. These trends highlight differences among the countries studied

DOT-ELISA-IgM in saliva for the diagnosis of human leptospirosis using polyester fabric-resin as support: preliminary report

Com a finalidade de melhorar o diagnostico da leptospirose humana, padronizou-se o teste dot-ELISA para a pesquisa de anticorpos especificos da classe IgM na saliva. Empregaram-se amostras de saliva e soro coletadas simultaneamente de 20 pacientes com a forma icterohemorragica da doenca, de 10 pacientes com outras patologias e 5 controles negativos. Culturas de Leptospira em meio EMJH, dos sorovares: icterohaemorrhagiae, canicola, hebdomadis, brasiliensis e cynopteri, foram utilizadas como antigeno, na concentracao proteica individual de 0,2 ug/ul, misturadas em volumes iguais. Na fase solida do teste empregou-se tecido de poliester impregnado com resina de N-metilol-acrilamida....

Contribuiçäo do conhecimento da epidemiologia da leptospirose humana, com especial referência ao Grande Rio, Brasil, no período de 1970 a 1982 / Contribution to the know ledge of the epidemiology of human leptospirosis with special reference to Greater Rio de Janeiro, in the period 1970 to 1982

Com o intuito de fornecer dados ao estudo da leptospirose humana, foram analisados 884 pacientes diagnosticados sorologicamente no Laboratório de Leptospirose da Fundaçäo Oswaldo Cruz - FIOCRUZ, no período de 1970 a 1982. Deste total, 775 estiveram internados em hospitais da cidade do Rio de Janeiro e 109 eram oriundos de outras cidades do Brasil. Sorologicamente, Icterohaemorrhagiae foi o sorogrupo mais freqüente, tanto para os pacientes do Grande Rio (76,4%), como para os demais (41,3%). No decorrer dos 13 anos, foram isoladas duas cepas de Icterohaemorrhagiae. Com relaçäo ao Grande Rio, observou-se uma freqüência maior da doença entre janeiro e abril (53,7%); o sexo masculino concorreu para 89,9% dos casos; e a faixa etária mais atingida compôs-se de indivíduos entre 16 e 45 anos (74,9%)