Cohabitation with a sick cage mate: Effects on ascitic form of Ehrlich tumor growth and macrophage activity

The present study was designed to evaluate the effects of mice cohabitation with a sick conspecific cage mate on peritoneal macrophage activity and on resistance to Ehrlich tumor growth. Female mice housed in pairs were divided into control and experimental groups. One mouse of each control pair was inoculated with NaCl (0.1 ml/10 g) intraperitoneally and the other, called `companion of healthy partner` (CHP), was kept undisturbed. One animal of each experimental pair of mice was inoculated with 5.0 x 10(6) Ehrlich tumor cells intraperitoneally and the other, the subject of this study, was called `companion of sick partner` (CSP). Peritoneal macrophages were removed from CSP and CHP mice to analyze resident macrophage activity (experiment 1), macrophage activity after Mycobacterium bovis (experiment 2) or Ehrlich tumor cells (experiment 3) in vivo inoculations. The resistance of CSP and CHP mice to Ehrlich tumor growth was also analyzed (experiment 4). Differences between groups were not found on resident macrophage activity. However, Onco-BCG- and Ehrlich tumor-activated macrophages from CSP mice presented a decreased intensity and percentage of phagocytosis and an increased respiratory burst in the presence of Staphylococcus aureus stimulation in vitro. CSP animals at the same time displayed a decreased resistance to Ehrlich tumor growth. These data were discussed in light of a possible psychological stress effect imposed by the housing condition on mice`s peritoneal macrophage activity and, as a consequence, on their resistance to Ehrlich tumor growth. Copyright (c) 2008 S. Karger AG, Basel.

Utilização do teste de eritroimunoadsorção por captura no imunodiagnóstico da neurocisticercose

Foi padronizado o teste de eritroimunoadsorção por captura (EIAC) para detecção de anticorpos específicos anti-cisticercos de Taenia solium, classe IgG, no líquido cefalorraquidiano (LCR) de pacientes com neurocisticercose. O reagente empregado para detecção de anticorpos específicos foi preparado com hemácias de carneiro em uma concentração de 0,25%, sensibilizadas com antígeno extrato salino bruto (ESB) obtido do Cysticercus cellulosae. A concentração ótima de ESB para sensibilização das hemácias de carneiro foi de 40ug/ml. O rendimento do ESB foi de 0,lug proteína/cavidade. A sensibilidade do teste foi de 84,5% (limite de confiança 95% de 75% a 94%), quando aplicado a 58 amostras de LCR de pacientes com neurocisticercose; e a especificidade foi de 95,3% (limite de confiança 95% de 90,7% a 99,9%) quando 85 amostras de LCR do grupo controle foram analisadas. O teste EIAC foi eficiente para o diagnóstico da neurocisticercose, e é importante para os laboratórios de saúde pública, tendo em vista a fácil execução, alto rendimento e baixo custo.

Comparação entre os testes de eritroimunoadsorção por captura, imunoenzimático e hemaglutinação passiva utilizados no diagnóstico da neurocisticercose

Foi realizada a comparação entre os testes de eritroimunoadsorção por captura (EIAC), imunoenzimático (ELISA) e de hemaglutinação passiva (HAP) utilizados no diagnóstico da neurocisticercose. Foram comparados dois testes já anteriormente utilizados na rotina diagnóstica da neurocisticercose (ELISA e HAP) e um recentemente padronizado (EIAC) para a detecção de anticorpos anti-Cysticercus cellulosae. O antígeno empregado nos três testes foi o extrato salino bruto (ESB), com um rendimento de 0,1; 1 e 1µg proteína/cavidade para os testes EIAC, ELISA e HAP, respectivamente. Quando se analisou um grupo de 58 pacientes com neurocisticercose, a sensibilidade observada foi de 98,2%, 84,5% e 77,2% nos testes ELISA, EIAC e HAP, respectivamente, para um grupo controle de 85 indivíduos, saudáveis ou com outras encefalotipatias, mas sem neurocisticercose, a especificidade foi de 94,1%, 95,3%, 91,8% , respectivamente, nos testes. Esta ordem de escolha poderia ser obedecida na medida dos recursos disponíveis.

Cellular uptake of BCG-loaded chitosan microparticles and in vivo evaluation of immune response following intranasal immunisation

Attenuated Mycobacterium bovis bacillus Calmette-Guérin (BCG) is the only currently available vaccine against tuberculosis. It is highly effective in pre-exposure immunisation against TB in children when administered by subcutaneous route to newborns. However, it does not provide permanent protection in adults. In this work, polymeric chitosan-alginate microparticles have been evaluated as potential nasal delivery systems and mucosal adjuvants for live attenuated BCG. Chitosan (CS) has been employed as adjuvant and mucosal permeation-enhancer, and, together with alginate (ALG), as additive to enhance BCG-loaded microparticles (MPs) cellular uptake in a human monocyte cell line, by particle surface modification. The most suitable particles were used for vaccine formulation and evaluation of immune response following intranasal immunisation of BALB/c mice.

Diagnóstico molecular da tuberculose bovina

Dissertação apresentada para a obtenção do Grau de Mestre em Genética Molecular e Biomedicina, pela Universidade Nova de Lisboa, Faculdade de Ciências e Tecnologia

Tratamento da cisticercose subcutânea co praziquantel: um novo agente cestoidicida

Twenty adult patients presenting dermal cysticercosis without cerebral or ocular involvement were treated with praziquantel. The first eleven cases received 60 mg/kg/day and the last nine cases 30 mg/kg/day. In both groups the daily dose was split into three oral intakes 4 to 6 hours apart and the drug administration lasted for 6 consecutive days. The latter group of patients also got dexamethasone, 3 mg daily, from one day before until four days after the treatment period with praziquantel. The drug proved to be 100% efficacious as demonstrated histopathologically by the death of the cysticerci of Taenia solium (Cysticercus cellulosae) in serial biopsies taken from the 2nd week on after the end of treatment, as well as clinically by the steady disappearence of the dermal nodules during the 6 months following the therapy. Tolerance of praziquantel was good as the incidence and severity of side-effects were not relevant. The drug safety was confirmed through laboratory tests which failed to detect any abnormal findings related to the hematopoietic, liver and kidney functions.

Radiometric studies on the oxidation of (1-14c) fatty acids by drug-susceptible and drug-resistant mycobacteria

A radiometric assay system has been used to study oxidation patterns of (1-14C) fatty acids by drug-susceptible and drug-resistant organisms of the genus Mycobacterium. Two strains of M. tuberculosis susceptible to all drugs, H37Rv and Erdman, were used. Drug-resistant organisms included in this investigation were M. tuberculosis H37Rv resistant to 5 ug/ml isoniazid, M. bovis, M. avium, M. intracellular, M. kansasii and M. chelonei. The organisms were inoculated in sterile reaction vials containing liquid 7H9 medium, 10% ADC enrichment and 1.0 uCi of one of the (1-14C) fatty acids (butyric, hexánoic, octanoic, decanoic, lauric, myristic, palmitic, stearic, oleic, linoleic, linolenic). Vials were incubated at 37°C and the 14CO2 envolved was measured daily for 3 days with a Bactec R-301 instrument. Although each individual organism displayed a different pattern of fatty oxidation, these patterns were not distinctive enough for identification of the organism. No combination of fatty acids nor preferential oxidation of long chain or of short chain fatty acids were able to separate susceptible from resistant organisms. Further investigation with a larger number of drug susceptible mycobacteria including assimilation studies and oxidation of other substrates may be required to achieve a distinction between drug-susceptible and drug-resistant mycobacteria.

Radiometric studies of mycobacterium tuberculosis

An in vitro assay system that included automated radiometric quantification of 14CO2 released as a result of oxidation of 14C- substrates was applied for studying the metabolic activity of M. tuberculosis under various experimental conditions. These experiments included the study of a) mtabolic pathways, b) detection times for various inoculum sizes, c) effect of filtration on reproducibility of results, d) influence of stress environment e) minimal inhibitory concentrations for isoniazid, streptomycin, ethambutol and rifampin, and f) generation times of M. tuberculosis and M. bovis. These organisms were found to metabolize 14C-for-mate, (U-14C) acetate, (U-14C) glycerol, (1-14C) palmitic acid, 1-14C) lauric acid, (U-14C) L-malic acid, (U-14C) D-glucose, and (U-14C) D-glucose, but not (1-14C) L-glucose, (U-14C) glycine, or (U-14C) pyruvate to 14CO2. By using either 14C-for-mate, (1-14C) palmitic acid, or (1-14C) lauric acid, 10(7) organisms/vial could be detected within 24 48 hours and as few as 10 organisms/vial within 16-20 days. Reproducible results could be obtained without filtering the bacterial suspension, provided that the organisms were grown in liquid 7H9 medium with 0.05% polysorbate 80 and homogenized prior to the study. Drugs that block protein synthesis were found to have lower minimal inhibitory concentrations with the radiometric method when compared to the conventional agar dilution method. The mean generation time obtained for M. bovis and different strains of M. tuberculosis with various substrates was 9 ± 1 hours.

Radiometric studies on the oxidation of (U-14C) L-amino acids by drug-susceptible and drug-resistant mycobacteria

A radiometric assay system has been used to study oxidation patterns of (U-14C) L-amino acids by drug-susceptible and drug-resistant mycobacteria. Drug-susceptible M. tuberculosis (H37Rv TMC 102 and Erdman) along with the drug-resistant organism M. tuberculosis (H37 Rv TMC 303), M. bovis, M. avium, M. intracellulare, M. kansasii and M. chelonei were used. The organisms were inoculated into a sterile reaction system with liquid 7H9 medium and one of the (U-14C) L-amino acids. Each organism displayed a different pattern of amino acid oxidation, but these patterns were not distinctive enough for identification of the organism. Complex amino acids such as proline, phenylalanine and tyrosine were of no use in identification of mycobacteria, since virtually all organisms failed to oxidize them. There was no combination of substrates able to separate susceptible from resistant organisms.

Imunodiagnóstico da neurocisticercose: teste imunoenzimático com antígenos quimicamente ligados a suportes para pesquisa de anticorpos em soro e líquido cefalorraquiano

Foi padronizado o teste imunoenzimático, ELISA, utilizando-se componentes antigênicos de Cysticercus cellulosae quimicamente ligados a suportes sólidos constituídos de discos de tecido-resina (ELISA-d), para pesquisa de anticorpos em soro líquido cefalorraquiano (LCR), ensaiando-se uma única diluição do espécime clínico. O suporte tecido-resina foi composto de tecido de poliéster impregnado com resina polimerizada de N-metilol-acrilamida, apresentando grupos N-metilol livres, capazes de reagir covalentemente com grupos funcionais de proteínas e polissacarídeos presentes no extrato antigênico salino total obtido de cisticercos. Foram ensaiados 38 soros e 74 LCR de pacientes com neurocisticercose comprovada e 50 soros e 107 LCR do grupo controle (pacientes com quadros clínicos neurológicos diversos e indivíduos supostamente normais). Obtivemos os seguintes índices de sensibilidade e especificidade: 94,7% e 92,0% para o teste realizado no soro e 98,6% e 100% para o teste realizado no LCR. O teste ELISA-d mostrou-se eficiente para o diagnóstico da neurocisticercose, principalmente quando realizado no LCR, com vantagens de estabilidade, facilidade de execução e baixo custo.

Cisticercose músculo-cutânea e visceral - doença rara?

O presente trabalho teve como objetivos estimar a freqüência das formas músculo-cutánea e visceral da cisticercose em exames anátomo-patológicos e necrópsias realizados em Brasilia, Distrito Federal (estudo retrospectivo) e diagnosticar a cisticercose músculo-cutânea em pacientes residentes na mesma região geográfica (estudo prospectivo). Em 64.911 protocolos de exames anátomo-patológicos, o diagnóstico de cisticercose foi observado em 30 (0,05%), sendo que em 27 (90,0%) os cistos estavam nos tecidos músculo-cutâneo-mucoso, em 1 (3,3%) em gânglio e em 2 (6,7%) no sistema nervoso central. Entre aqueles com cistos nos tecidos músculo-cutâneo-mucoso 2 (7,4%) tinham cisticercos em língua. Em 1520 protocolos de necrópsia, encontraram-se 25 (1,6%) com diagnóstico de cisticercose, sendo: 24(96,0%) com neurocisticercose, seja isolada ou associada a outras formas da doença; e 2 (8,0%) com cisticercos em coração, 2 (8,0%) em músculo esquelético e 1 (4,0%) em fígado, seja isolados ou associados a outras localizações do parasito. Foram também examinados 1122 indivíduos, realizando-se em todos eles as reações sorológicas de imunofluorescência indireta e ELISA para cisticercose e a investigação radiológica de partes moles e crânio. Encontraram-se 59 (5,3%) com ambas reações sorológicas reagentes (10 entre eles com o diagnóstico de cisticercose confirmado por biópsias); e 32 (2,8%) com calcificações nas radiografias de partes moles e/ou crânio, mas apresentando ambas reações sorológicas não-reagentes. Entre os pacientes com os testes imunológicos reagentes, a neurocisticercose foi diagnosticada em 39 (66,1%), a cisticercose muscular em 25 (42,4%); a cutânea em 12 (20,3%); e a visceral em 2 (3,4%), sendo em 1 (1,7%) ovariana e em 1 (1,7%) miocárdica, pleural e renal. Os resultados permitem concluir que a forma músculo-cutânea é observada freqüentemente entre pacientes com cisticercose residentes em Brasília. A forma visceral também foi encontrada, com os cisticercos localizados em diferentes órgãos, sendo que os pacientes afetados não apresentavam as manifestações clínicas.

Un caso de infeccion humana por cisticerco racemoso cerebral de localizacion parenquimatosa en Valdivia, Chile

Se presenta un caso clínico de infección por cisticerco racemoso cerebral de localización parenquimatosa en un paciente de la ciudad de Valdivia (Chile) cuyo diagnóstico definitivo se efectuó a través del estudio morfológico del parásito. Se discute brevemente la escasa frecuencia de la localización parenquimatosa del cisticerco racemoso, así como su diagnóstico diferencial con otros estados larvarios de cestodos que desarrollan en el sistema nervioso.

The diagnostic importance of species specific and cross-reactive components of Taenia solium, Echinococcus granulosus, and Hymenolepis nana

Sera from patients infected with Taenia solium, Hymenolepis nana and Echinococcus granulosus were tested against homologous and heterologous parasite antigens using an ELISA assay, and a high degree of cross-reactivity was verified. To identify polypeptides responsible for this cross reactivity, the Enzyme Linked Immunoelectro Transfer Blot (EITB) was used. Sera from infected patients with T.solium, H.nana, and E.granulosus were assessed against crude, ammonium sulphate precipitated (TSASP), and lentil-lectin purified antigens of T.solium and crude antigens of.H.nana and E.granulosus. Several bands, recognized by sera from patients with T.solium, H.nana, and E.granulosus infections, were common to either two or all three cestodes. Unique reactive bands in H.nana were noted at 49 and 66 K-Da and in E.granulosus at 17-21 K-Da and at 27-32 K-Da. In the crude cysticercosis extract, a specific non glycoprotein band was present at 61-67 K-Da in addiction to specific glycoprotein bands of 50, 42, 24, 21, 18, 14, and 13 K-Da. None of the sera from patients with H.nana or E.granulosus infection cross reacted with these seven glycoprotein bands considered specific for T.solium infection.

HEMAGGLUTINATION TEST FOR THE DIAGNOSIS OF HUMAN NEUROCYSTICERCOSIS: DEVELOPMENT OF A STABLE REAGENT USING HOMOLOGOUS AND HETEROLOGOUS ANTIGENS

A hemagglutination (HA) test was standardized using formalin- and tannin-treated gander red blood cells sensitized with a total salt extract of C. cellulosae (HA-Cc) and an antigenic extract of Cysticercus longicollis (HA-Cl) vesicular fluid. A total of 61 cerebrospinal fluid (CSF) samples were assayed, 41 from patients with neurocysticercosis and 20 from a control group, which were, respectively, reactive and non-reactive to ELISA using C. cellulosae. The CSF samples from the control group did not react and 35 (85.4%) and 34 (82.9%) CSF samples from patients were reactive to the HA-Cc and HA-Cl tests, respectively. The reagents ready for use were stable up to 6 months when stored at 4°C in 50% glycerol. The present results confirm that the reagent using Cysticercus longicollis stabilized with glycerol can be used as an alternative in the immunological diagnosis of neurocysticercosis