Phenotypic and genotypic variant of MDR-Mycobacterium tuberculosis multiple isolates in the same tuberculosis episode, Rio de Janeiro, Brazil

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

Estudo da transmissão horizontal de Mycobacterium avium-intracellulare em suínos

Experimental infections were performed to study the horizontal transmission of Mycobacterium avium-intracellulare (MAI) in swine. Twelve 35 to 42 day-old pigs were divided into three groups of four pigs each. Each group was housed in separate rooms, and inoculated with three MAI strains: VPS1, SC1 and ATCC-13950. After inoculation, four non-inoculated swines were added to each group and maintained in the same pen up to 150 days of age, when were slaughtered. Other four non-inoculated swines were maintained as a control group. The infection was diagnosed by the tuberculin test at 140 days of age, by detection of macroscopic lesions at slaughter and histopathologic and bacteriologic examination of tissues collected at necropsy. Culture of fecal samples were made during the experiment indicating active excretion of MAI between 14 and 42 days post-inoculation. Transmission of MAI among pigs kept in the same pen was observed with all the three strains used. The most evident transmission was with strain SC1, in which four "in contact" pigs were infected, followed by ATCC-13950 (two pigs) and VPS1 (one pig). These results indicate that infected swine can be an important source of infection, favoring the dissemination of this disease.

O trabalho teve como objetivo estudar a transmissão horizontal de Mycobacterium avium-intracellulare (MAI) entre suínos. Doze suínos com 35 a 42 dias de idade, divididos em três grupos de quatro e alojados em salas independentes, foram inoculados por via oral com as cepas VPS1, SC1 e ATCC-13950 de Mycobacterium avium-intracellulare. A seguir, quatro suínos não inoculados foram colocados em cada grupo e mantidos na mesma baia até o abate aos 150 dias da idade. Outro grupo de quatro suínos não inoculados foi utilizado como controle. Pelo exame de fezes verificou-se que houve excreção ativa de MAI entre 14 e 42 dias pós-inoculação. A transmissão da infecção foi avaliada pelo teste de tuberculina aos 140 dias de idade e pelos exames de patologia macro e microscópica e de isolamento do MAI de órgãos. Os quatro suínos em contato com aqueles inoculados com a cepa SC1 foram contaminados, enquanto que com a cepa VPS1 apenas um e com a ATCC-13950 somente dois. Concluiu-se que houve transmissão de MAI entre os suínos mantidos na mesma baia, com as três cepas utilizadas, mas foi mais evidente com a cepa SC1, indicando que o suíno pode ser importante fonte de infecção na disseminação da doença.

Imunidade celular e humoral na hanseníase bordeline tuberculoid (BT) / Humoral and cellular immunity in bordeline tuberculoid (BT) leprosy

Foram estudados 20 pacientes com diagnóstico de hanseníase bordeline tuberculoid (BT), classificados segundo os critérios de Ridley & Jopling, bem como dois pacientes com forma Tuberculoid tuberculoid (TT), todos com baciloscopia negativa, exceto um, que apresentou índice baciloscópico 1+. Todos os pacientes foram avaliados quanto a sua capacidade de resposta imune humoral ao DBSA (antígeno sintético semelhante ao glicolipídeo fenólico I, específico do M, leprae) e 18 pacientes foram submetidos a testes de avaliaçåo da resposta imunocelular in vivo (teste Mitsuda) e in vitro (linfoproliferaçåo e produçåo de interferon-gama) frente ao Mycobacterium leprae. Observamos que 90 por cento dos pacientess apresentaram resultados negativos quanto à pesquisa de IgM anti-DBSA pelo método imunoenzimático ELISA (densidade óptica , 0,27), o que demonstra ser este teste inadequado para a detecçåo de pacientes paucibacilares. Quanto aos testes de imunidade celular, oito pacientes (44,4 por cento) apresentaram teste de Mitsuda positivo (>= 5mm), sendo os demais considerados negativos. Cerca de 89 por cento dos pacientes tiveram teste de Mitsuda maior ou igual a 3mm. Doze pacientes (66,7 por cento) tiveram resposta linfoproliferativa positiva (índice estimulatório >= 3,0) para o M. leprae. Vinte e dois por cento dos pacientes apresentaram níveis de interferon-gama acima do limite de positividade (40 U/ml). Houve 66,7 por cento de correlaçåo entre os testes de Mitsuda e interferon-gama; 55,6 por cento de correlaçåo entre os testes in vitro (linfoproliferaçåo e interferon-gama). Quando estes três testes foram considerados em conjunto, uma correlaçåo de 38,9 por cento foi observada. Este estudo demonstra a heterogeneidade do comportamento imunológico mediado por células e anticorpos em pacientes com hanseníase BT, apesar de todos histologicamente serem capazes de conter a multiplicaçåo bacilar e de formar granulomas epitelióides

Relationship mbetween immune response and clinical manifestations in patients with tuberculosis

Cutaneous reactivityy to purified protein derivative (PPD) and antibody levels were investigated in 122 adults and 28 children with tuberculosis. IgG anti-PPD levelks (measured by ELISA and reported as absorbance at 405 nm) were higher in adult patients with the disease for more than one year (0.533 ñ 0.391, N=38 vs 0.224 ñ 0.256, n+50 in patients with the disease for less than one year) as well as inn children with disseminated tuberculosis (0.138 ñ 0.137, N=11, vs 0.072 ñ 0.55, N=15 in children with localized disease). The cut-off values (mean + 2SD) for healthy children and adults were 0.09 and 0.22 absorbance at 405 nm. In both adult and pediatric patients, cutaneous reactivity to PPD was inversely correlated with antibody levels. The present data provide additional evidence for the existence of an unstable spectrum of immune response in tuberculosis patients in whom changes in clinical situation are dynamic

The dot-ELISA test for the detection of anti-PGL-1 IgM in leprosy patients and their contacts

Since dot-ELISA has recently been reported to be a sensitive, simple and method, we have compared it with the conventional microplate ELISA method. Sera of 124 leprosy patients, 136 household and professional contacts, and 92 controls were tested for a antibodies against a Mycobacterium leprae antigen using dot-ELISA on nitrocellulose membrane filters and microplate ELISA. The sensitive of the techniques was similar for multibacillary patients, but dot-ELISA was less sensitive for paucibacillary patients although it was more specific (100%) than ELISA (93,4%). Of 21 household contacts that gave a response by ELISA, 3 were also positive by dot-ELISA; one of these 3 developed indeterminate leprosy 12 months later and the other was diagnosed as borderline lepromatous after 28 months. These data indicate that dot-ELISA has a high spedificity and can be a useful tool in field evaluation

Use of the anti-PGL-I antibody ELISA and the Mitsuda reaction in early diagnosis of leprosy

Cutaneous reaction to the Mitsuda antigen and anti-PGL-I IgM antibodies (ELISA) were determined in 134 leprosy patients, 290 household contacts, 52 healthy controls and 43 tuberculous individuals. The multibacillary patients did nort developed cell-mediated immunity (CMI), although they presented high levels of IgM (absorbance at 492 m > 0.5). The paucibacillatry patients presented CMI, although in varying degrees, and IgM levels did not exceed 0.5 absorbance units. Most of the contacts (107) showed a Mitsuda-postive test, and 25 of then were anti-PGL-I IgM seropositive (absorbance < 0.5 but > 0.22), although none became ill during the twoyear follow-up. Of the 17 Mitsuda-negative contacts, two wxhibited an immunological status of lepromatous leprosy (negative Mitsuda test and positive serology; absorvance > 0.05) and became ill (one borderline lepromatous and one indeterminate leprosy). These results show that the immunological status of lepromatous leprosy can appear prior to clinical symptoms, and thus serology for anti-PGL-I IgM together with the Mitsuda test can be useful in an activbe search for new preclinical cases among high risk populations

IgM immunoglobulins reacting with the phenolic glycolipid-1 antigen from Mycobacterium leprae in sera of leprosy patients and their contacts

For the first time in Brazil it was investigated the occurrence of IgM anti-PGL-1 in the sera of household contacts of leprozy patients using the ELISA methodology. The sera of the multipatients. It was observed a high subclinical infection incidence among household contacts (19.4%). The percentage of leprosy development was 5% (1/21) among the seropositive contact group. This finding suggests that serology could be useful as prognostic test, but for better definition is necessary to tet a population from endemic area for long period time