Predictors and outcomes of disseminated tuberculosis in an intermediate burden setting.

SETTING: University-affiliated hospital located in Porto, North Portugal, an area with a low to intermediate incidence of tuberculosis (TB). OBJECTIVE: To identify predictors and outcomes of disseminated TB (dTB). DESIGN: A cohort of patients diagnosed with TB between 2007 and 2013 was retrospectively analysed. Patients with dTB criteria were characterized and compared to single organ TB cases. Factors independently associated with dTB were determined by multivariate logistic regression analysis. RESULTS: A total of 744 patients were analysed, including 145 with dTB. Independent risk factors for dTB were pharmacological immunosuppression (OR 5.6, 95% CI 2.8-11.3), HIV infection (OR 5.1, 95% CI 3.1-8.3), chronic liver failure or cirrhosis (OR 2.3, 95% CI 1.4-4.1) and duration of symptoms (OR 2.3, 95% CI 1.4-3.8). Compared to single organ TB, the clinical presentation of dTB patients differed by the absence of haemoptysis (OR 3.2, 95% CI 1.3-8.4) and of dyspnoea (OR 1.9, 95% CI 1.2-3.1), presence of weight loss (OR 1.8, 95% CI 1.1-2.9), night sweats (OR 1.7, 95% CI 1.1-2.7) and bilateral lung involvement (OR 4.4, 95% CI 2.8-7.1). Mortality and time until culture conversion were higher for dTB patients, although not reaching statistical significance. CONCLUSION: Immunosuppressive conditions and chronic liver failure or cirrhosis were associated with increased risk of dTB. The haematogenous spread may be dependent on longer symptomatic disease and usually progresses with bilateral lung involvement.

Short communication: Linear discriminant analysis and type of oil added to dairy goat diets.

Gas chromatography fatty acid (FA) analysis of 112 milk fat samples from dairy goats fed a basal diet with no added oil or the same diet with 1 of 3 vegetable oils added [high oleic sunflower oil (HOSFO), regular sunflower oil (RSFO), or linseed oil (LO)] was used to identify the type of diet consumed through linear discriminant analysis. Twenty variables (19 FA and 1 FA ratio) were selected as valid predictors out of 84 variables tested. The Mahalanobis squared distance was minimal between HOSFO and RSFO groups and maximal between control and LO groups. Cross-validation showed that only one observation from RSFO group was misclassified into the HOSFO group. We concluded that linear discriminant analysis is a useful method to classify milk fat samples from dairy goats according to the particular vegetable oil (of the 3 oils tested here) added to the basal diet.

The rs5743836 polymorphism in TLR9 confers a population-based increased risk of non-Hodgkin lymphoma.

Non-Hodgkin lymphoma (NHL) has been associated with immunological defects, chronic inflammatory and autoimmune conditions. Given the link between immune dysfunction and NHL, genetic variants in toll-like receptors (TLRs) have been regarded as potential predictive factors of susceptibility to NHL. Adequate anti-tumoral responses are known to depend on TLR9 function, such that the use of its synthetic ligand is being targeted as a therapeutic strategy. We investigated the association between the functional rs5743836 polymorphism in the TLR9 promoter and risk for B-cell NHL and its major subtypes in three independent case-control association studies from Portugal (1160 controls, 797 patients), Italy (468 controls, 494 patients) and the US (972 controls, 868 patients). We found that the rs5743836 polymorphism was significantly overtransmitted in both Portuguese (odds ratio (OR), 1.85; P=7.3E-9) and Italian (OR, 1.84; P=6.0E-5) and not in the US cohort of NHL patients. Moreover, the increased transcriptional activity of TLR9 in mononuclear cells from patients harboring rs5743836 further supports a functional effect of this polymorphism on NHL susceptibility in a population-dependent manner.

Cdc42p controls yeast-cell shape and virulence of Paracoccidioides brasiliensis.

Paracoccidioides brasiliensis is characterized by a multiple budding phenotype and a polymorphic cell growth, leading to the formation of cells with extreme variations in shape and size. Since Cdc42 is a pivotal molecule in establishing and maintaining polarized growth for diverse cell types, as well as during pathogenesis of certain fungi, we evaluated its role during cell growth and virulence of the yeast-form of P. brasiliensis. We used antisense technology to knock-down PbCDC42's expression in P. brasiliensis yeast cells, promoting a decrease in cell size and more homogenous cell growth, altering the typical polymorphism of wild-type cells. Reduced expression levels also lead to increased phagocytosis and decreased virulence in a mouse model of infection. We provide genetic evidences underlying Pbcdc42p as an important protein during host-pathogen interaction and the relevance of the polymorphic nature and cell size in the pathogenesis of P. brasiliensis.

Prevalência de listeria spp em amostras de carne de frango obtidas em abatedouros do Estado de São Paulo / Prevalence of listeria spp in samples of chicken meat obtained in s slaughtherhouses of the State of São Paulo

Listeria monocytogenes está envolvida em surtos humanos relacionados a alimentos, embora, no Brasil a doença em humanos seja pouco relatada. Enfoque renovado foi dado a esta bactéria após surtos de doenças de origem alimentar (DOA) ocorridos na América do Norte e Europa durante os anos de 1980 e 1990. Listeria spp é freqüentemente isolada de carnes cruas, incluindo as de frango, como resultado de ampla contaminação cruzada em plantas industriais. A carne de frango é parte integrante da dieta dos brasileiros como fonte de proteína animal, assim, é importante que se conheça a prevalência deste agente neste tipo de alimento. Para tanto, foram examinadas 74 (setenta e quatro) amostras de carne de frango (coxa, sobrecoxa, peito, frango à passarinho e inteiro) utilizando-se métodos de isolamento de Listeria spp com meios de enriquecimento e seletivos, e reação em cadeia de polimerase (PCR), para confirmação dos testes bioquímicos. Em apenas uma das amostras foi detectada a presença de L. monocytogenes. Aventa-se, para o baixo índice de listérias encontrado, a ação antimicrobiana determinada pelo uso de descontaminantes nos tanques de resfriamento dos abatedouros.

Prevalência de listeria spp em amostras de carne de frango obtidas em abatedouros do Estado de São Paulo / Prevalence of listeria spp in samples of chicken meat obtained in s slaughtherhouses of the State of São Paulo

Listeria monocytogenes está envolvida em surtos humanos relacionados a alimentos, embora, no Brasil a doença em humanos seja pouco relatada. Enfoque renovado foi dado a esta bactéria após surtos de doenças de origem alimentar (DOA) ocorridos na América do Norte e Europa durante os anos de 1980 e 1990. Listeria spp é freqüentemente isolada de carnes cruas, incluindo as de frango, como resultado de ampla contaminação cruzada em plantas industriais. A carne de frango é parte integrante da dieta dos brasileiros como fonte de proteína animal, assim, é importante que se conheça a prevalência deste agente neste tipo de alimento. Para tanto, foram examinadas 74 (setenta e quatro) amostras de carne de frango (coxa, sobrecoxa, peito, frango à passarinho e inteiro) utilizando-se métodos de isolamento de Listeria spp com meios de enriquecimento e seletivos, e reação em cadeia de polimerase (PCR), para confirmação dos testes bioquímicos. Em apenas uma das amostras foi detectada a presença de L. monocytogenes. Aventa-se, para o baixo índice de listérias encontrado, a ação antimicrobiana determinada pelo uso de descontaminantes nos tanques de resfriamento dos abatedouros.

Listeria monocytogenes is responsible for food borne disease, although in Brazil there is little data about this agent. Renewed emphasis has been given to this bacterium after the North American and European outbreaks during the 1980s and 1990s. Listeria spp is usually isolated from raw meat, including chicken, due to cross contamination in industrial plants. Chicken meat plays an important role in the diet of Brazilian people as an animal protein source. This work was aimed at investigating this bacterium' s prevalence in different cuts of chicken sampled at slaughterhouses. To achieve this, 74 chicken samples (drumsticks, thighs, breasts, entire chickens cut-up and whole) were cultivated in Listeria spp enrichment and selective culture media, and submitted to polymerase chain reaction to confirm the biochemical tests. Just one sample was positive for L. monocytogenes. As one possible explanation for the low level of listeria found, the authors point to the antiimicrobial action of disinfectant products used in the chilled tanks of slaughther houses.

Anti-interleukin 10 receptor monoclonal antibody is an adjuvant for T helper cell type 1 responses to soluble antigen only in the presence of lipopolysaccharide.

Soluble foreign antigen usually leads to a transient clonal expansion of antigen-specific T cells followed by the deletion and/or functional inactivation of the cells. As interleukin (IL)-10 is a key immunoregulatory cytokine, we questioned whether neutralization of IL-10 during priming with soluble antigen could prime for a subsequent T helper cell type 1 (Th1) effector recall response. By using an adoptive transfer model to track the fate of antigen-specific T cell receptor (TCR)-transgenic CD4(+) T cells, we show that administration of soluble ovalbumin (OVA) protein, but not OVA(323-339) peptide antigen, together with an anti-IL-10 receptor (R) mAb led to the enhancement of a Th1 response upon rechallenge. Lipopolysaccharide (LPS) present in the protein was necessary for priming for Th1 recall responses in the presence of anti-IL-10R mAb, as removal of LPS abrogated this effect. Moreover, addition of LPS to the peptide did not itself allow priming for recall Th1 effector responses unless endogenous levels of IL-10 were neutralized with an anti-IL-10R mAb. A significant increase in OVA-specific IgG1 and IgG2a isotypes was observed when the protein antigen was administered with anti-IL-10R mAb; however, this was not the case with peptide antigen administered together with anti-IL-10R and LPS. Our data, showing that LPS receptor signaling and neutralization of endogenous immunosuppressive cytokines is essential for Th1 priming, has important implications for the design of relevant vaccines for effective in vivo immunotherapy.

Molecular and functional characterization of mouse signaling lymphocytic activation molecule (SLAM): differential expression and responsiveness in Th1 and Th2 cells.

Optimal T cell activation and expansion require engagement of the TCR plus costimulatory signals delivered through accessory molecules. SLAM (signaling lymphocytic activation molecule), a 70-kDa costimulatory molecule belonging to the Ig superfamily, was defined as a human cell surface molecule that mediated CD28-independent proliferation of human T cells and IFN-gamma production by human Th1 and Th2 clones. In this study, we describe the cloning of mouse SLAM and the production of mAb against it which reveal its expression on primary mouse T and B cells. Mouse SLAM is expressed on highly polarized Th1 and Th2 populations, and is maintained on Th1, but not on Th2 clones. Anti-mouse SLAM mAb augmented IFN-gamma production by Th1 cells and Th1 clones stimulated through the TCR, but did not induce IFN-gamma production by Th2 cells, nor their production of IL-4 or their proliferation. Mouse SLAM is a 75-kDa glycoprotein that upon tyrosine phosphorylation associates with the src homology 2-domain-containing protein tyrosine phosphatase SHP-2, but not SHP-1. Mouse SLAM also associates with the recently described human SLAM-associated protein. These studies may provide new insights into the regulation of Th1 responses.

Susceptibility of beige mice to Mycobacterium avium: role of neutrophils.

The beige mutation in C57BL/6 mice has been shown to increase the susceptibility to infection by Mycobacterium avium. In this study, we confirmed those results and showed that the effect of the beige mutation was most obvious after infection with a strain of lower virulence than with a highly virulent isolate of M. avium. The dissemination of M. avium from the gut was observed with both C57BL/6 and beige mice but was faster in the latter. The expression of gamma interferon (IFN-gamma) and the priming for tumor necrosis factor production during an in vivo infection were similar between beige and immunocompetent C57BL/6 mice. IFN-gamma produced during the infection of beige mice was protective in the spleen, and the administration of recombinant IFN-gamma restored the resistance in the spleen to levels similar to those found in control mice. There were no histological differences between wild-type and beige mice with respect to granuloma formation in the liver. The increased susceptibility of beige mice to M. avium as manifested in the liver was reduced by transfusing neutrophils from wild-type C57BL/6 mice. Likewise, depletion of neutrophils from C57BL/6 mice rendered them as susceptible to M. avium infection of the liver as beige mice. Our results point to the participation of neutrophils in the defect of beige mice in addition to other defects. Furthermore, these results show that neutrophils play a significant role in the defense mechanisms against mycobacterial infections and that beige animals may be a useful model for study of the role of neutrophils in mycobacteriosis.

Tumour necrosis factor-alpha (TNF-alpha) in the host resistance to mycobacteria of distinct virulence.

The relative virulence of different isolates of Mycobacterium avium has been linked to their capacity to trigger the secretion of TNF from the macrophages they infect. Smooth opaque (SmOp) variants of Myco. avium have been shown to trigger higher expression of TNF-alpha by macrophages in vitro than the smooth transparent (SmTr) variants. To analyse the role of TNF in resistance to infection by Myco. avium, we studied the infection by two different morphotypes of strain 2.151 of Myco. avium both in vitro and in vivo in the presence or absence of neutralizing antibodies to TNF. No effects were found in vitro regarding the growth of either isolate of Myco. avium. In vivo, only the virulent SmTr morphotype showed enhanced growth in the presence of the neutralizing antibodies. This enhancement occurred relatively late when priming for TNF secretion in vivo was evident. Among four isolates of Myco. avium, three virulent ones induced a marked priming for TNF release and one avirulent strain did not. Mycobacterium tuberculosis H37Ra, which is very active in inducing TNF release due to its lipoarabinomannan moiety, was used to compare with the previous results. The growth of H37Ra in macrophages was increased in vitro by the neutralization of TNF and neutralization of either TNF and/or interferon-gamma (IFN-gamma) enhanced the in vivo proliferation of this microbe in the spleen and liver of infected animals, whereas only the combination of both anti-TNF and anti-IFN-gamma enhanced bacterial proliferation in the lung. We conclude that resistance to the avirulent strains of Myco. avium did not involve TNF, but rather antimicrobial mechanisms expressed constitutively in the mononuclear phagocytes. In contrast, TNF plays an important role in the control of Myco. tuberculosis H37Ra infection.

Associaçäo entre salinomicina e tiamulin em raçöes de aves para corte / Effect of salinomycin and tiamulin association in broiler chicken diets

A pesquisa foi conduzida no Aviário Experimental do Departamento de Produçäo Animal da Faculdade de Medicina Veterinária e Zootecnia da Universidade de Säo Paulo, em Pirassununga, Säo Paulo, e teve a finalidade de verificar a possível toxicidade do anticoccidiano salinomicina pela administraçäo simultânea do antibiótico tiamulin na raçäo. Foram utilizados 1.296 pintos de 01 dia, distribuídos em 36 boxes experimentais e submetidos a 6 tratamentos, com 6 repetiçöes de 36 aves cada. O delineamento experimental foi o de blocos casualizados. Para os tratamentos foram empregados 60 ppm de salinomicina e 0; 10; 15; 20; 30 e 40 ppm de tiamulin. Os dados obtidos indicaram näo haver depressäo no crescimento das aves, nem tampouco no consumo de raçäo e conversäo alimentar. Näo ocorreram lesöes nos sacos aéreos e os testes de soroaglutinaçäo rápida para Mycoplasma gallissepticum apresentaram resultados negativos. Tendo em vista os resultados obtidos, pode-se dizer que a adiçäo simultânea de tiamulin e salinomicina em raçöes de aves para corte, nos níveis utilizados, näo acarreta toxicidade

Characterization of the virulence of Mycobacterium avium complex (MAC) isolates in mice.

The virulence of different isolates of MAC was studied in naturally susceptible BALB/c mice. In preliminary experiments, MAC bacteria forming smooth transparent colonies on solid media (SmT variants) were found to be virulent for BALB/c mice, causing progressive infection; smooth opaque (SmOp) were generally avirulent, being slowly eliminated from the infected organs; and rough (Rg) variants were either avirulent or as virulent as SmT variants. We chose to compare the virulence of different isolates of MAC of different origins, studying only the SmT morphotype. Strains of MAC isolated from naturally infected animals were those that most consistently caused progressive infections. AIDS patients-derived isolates were of intermediate virulence or devoid of virulence in mice. The environmental strains were eliminated from mice or did not proliferate. Strains of MAC isolated from individuals who were not infected by HIV varied in virulence from completely avirulent to highly virulent. There was no close correlation between virulence and restriction fragment length polymorphism (RFLP) type, although all highly virulent strains were of the A/I type. There was also no correlation between virulence analysed in vivo and the ability to grow in cultured macrophages.

Role of gamma interferon and tumor necrosis factor alpha during T-cell-independent and -dependent phases of Mycobacterium avium infection.

To design an effective immunotherapy for Mycobacterium avium infections, the protective host response to the infection must be known. Here we analyzed the role of gamma interferon (IFN-gamma) and tumor necrosis factor alpha (TNF-alpha) in the innate and acquired responses to M. avium infections in mice. T-cell depletion studies showed that CD4+ T cells were required for control of the infection. CD(4+)-depleted mice showed enhanced bacterial proliferation and at the same time showed a reduction in the level of expression of both IFN-gamma and TNF-alpha mRNAs in spleen cells. In contrast, M. bovis BCG immunization restricted M. avium proliferation and at the same time promoted expression of the mRNAs for the two cytokines. In vivo depletion studies using specific monoclonal antibodies showed that both IFN-gamma and TNF-alpha are involved in an early protection possibly involving NK cells, and furthermore, IFN-gamma is involved in the later T-cell-protective response to infection. In vivo neutralization of IFN-gamma during M. avium infection also blocked the priming for enhanced TNF-alpha secretion triggered by endotoxin. Both cytokines were found to be involved in the resistance expressed in BCG-immunized animals and exhibited additive bacteriostatic effects in vitro on bone marrow-derived macrophages infected with different strains of M. avium. These data suggest that both cytokines act in an additive or synergistic fashion in the induction of bacteriostasis and that IFN-gamma is also involved in priming TNF-alpha secretion.

Role of interleukin-6 in the induction of protective T cells during mycobacterial infections in mice.

Interleukin-6 (IL-6) has been shown to regulate numerous functions of the immune system including the differentiation of T-cell subpopulations. Here we examined the involvement of this cytokine in the in vivo generation of a population of T cells able to protect mice against mycobacterial infections. BALB/c mice were infected intravenously with Mycobacterium avium 2447 and anti-IL-6 monoclonal antibodies were administered intraperitoneally throughout the course of the infection. Control mice were able to control the mycobacterial proliferation 1 month after inoculation, whereas mice whose IL-6 had been blocked showed progressive bacterial growth. To distinguish a role for IL-6 associated to the induction or expression of immunity mediated by T cells, we immunized mice with M. bovis bacillus Calmette-Guérin (BCG) Pasteur and challenged them 2 months later with M. avium. One group of mice received anti-IL-6 during the BCG vaccination and another during the M. avium challenge. When M. avium proliferation was assessed at day 30 of the challenge, it was found that the administration of anti-IL-6 during vaccination reduced the protection afforded by BCG compared to administration of the isotype control antibody. No difference in bacterial proliferation was observed at day 30 of challenge when antibodies were administered during M. avium challenge. Our results show that protective T cells arise during M. avium infections in mice after differentiating in the presence of IL-6.

[Acute appendicitis in children. Experience at a general hospital]. / Apendicitis aguda en el niño. Experiencia en un hospital general.

In order to establish a guide for the diagnosis of acute appendicitis, we reviewed the charts of patients with appendectomy. In a 7-year period, 385 patients were studied. The age range was 3 to 15 years. In 53% there was an administration of medications prior to surgery. Perforated appendicitis was found in the majority (53%) of the cases. We could not find any association between age and perforation. Only localized, persistent abdominal pain, peritoneal irritation, anorexia, and vomiting were useful for differential diagnosis. In patients with acute appendicitis (p greater than 0.05), leukocytosis (greater than 10,000/mm3), neutrophilia (greater than 70%) and bands (greater than 3%) were observed in 80% of the cases. The frequency of complications was elevated (39.5%), and the mortality was five times higher than referred in other studies. We propose an algorithm for both opportune diagnosis and treatment of the disease.