Processing of Mycobacterium tuberculosis bacilli by human monocytes for CD4+ alphabeta and gammadelta T cells: role of particulate antigen.

Mycobacterium tuberculosis readily activates both CD4+ and Vdelta2+ gammadelta T cells. Despite similarity in function, these T-cell subsets differ in the antigens they recognize and the manners in which these antigens are presented by M. tuberculosis-infected monocytes. We investigated mechanisms of antigen processing of M. tuberculosis antigens to human CD4 and gammadelta T cells by monocytes. Initial uptake of M. tuberculosis bacilli and subsequent processing were required for efficient presentation not only to CD4 T cells but also to Vdelta2+ gammadelta T cells. For gammadelta T cells, recognition of M. tuberculosis-infected monocytes was dependent on Vdelta2+ T-cell-receptor expression. Recognition of M. tuberculosis antigens by CD4+ T cells was restricted by the class II major histocompatibility complex molecule HLA-DR. Processing of M. tuberculosis bacilli for Vdelta2+ gammadelta T cells was inhibitable by Brefeldin A, whereas processing of soluble mycobacterial antigens for gammadelta T cells was not sensitive to Brefeldin A. Processing of M. tuberculosis bacilli for CD4+ T cells was unaffected by Brefeldin A. Lysosomotropic agents such as chloroquine and ammonium chloride did not affect the processing of M. tuberculosis bacilli for CD4+ and gammadelta T cells. In contrast, both inhibitors blocked processing of soluble mycobacterial antigens for CD4+ T cells. Chloroquine and ammonium chloride insensitivity of processing of M. tuberculosis bacilli was not dependent on the viability of the bacteria, since processing of both formaldehyde-fixed dead bacteria and mycobacterial antigens covalently coupled to latex beads was chloroquine insensitive. Thus, the manner in which mycobacterial antigens were taken up by monocytes (particulate versus soluble) influenced the antigen processing pathway for CD4+ and gammadelta T cells.

Regulation of human CD4(+) alphabeta T-cell-receptor-positive (TCR(+)) and gammadelta TCR(+) T-cell responses to Mycobacterium tuberculosis by interleukin-10 and transforming growth factor beta.

Mycobacterium tuberculosis is the etiologic agent of human tuberculosis and is estimated to infect one-third of the world's population. Control of M. tuberculosis requires T cells and macrophages. T-cell function is modulated by the cytokine environment, which in mycobacterial infection is a balance of proinflammatory (interleukin-1 [IL-1], IL-6, IL-8, IL-12, and tumor necrosis factor alpha) and inhibitory (IL-10 and transforming growth factor beta [TGF-beta]) cytokines. IL-10 and TGF-beta are produced by M. tuberculosis-infected macrophages. The effect of IL-10 and TGF-beta on M. tuberculosis-reactive human CD4(+) and gammadelta T cells, the two major human T-cell subsets activated by M. tuberculosis, was investigated. Both IL-10 and TGF-beta inhibited proliferation and gamma interferon production by CD4(+) and gammadelta T cells. IL-10 was a more potent inhibitor than TGF-beta for both T-cell subsets. Combinations of IL-10 and TGF-beta did not result in additive or synergistic inhibition. IL-10 inhibited gammadelta and CD4(+) T cells directly and inhibited monocyte antigen-presenting cell (APC) function for CD4(+) T cells and, to a lesser extent, for gammadelta T cells. TGF-beta inhibited both CD4(+) and gammadelta T cells directly and had little effect on APC function for gammadelta and CD4(+) T cells. IL-10 down-regulated major histocompatibility complex (MHC) class I, MHC class II, CD40, B7-1, and B7-2 expression on M. tuberculosis-infected monocytes to a greater extent than TGF-beta. Neither cytokine affected the uptake of M. tuberculosis by monocytes. Thus, IL-10 and TGF-beta both inhibited CD4(+) and gammadelta T cells but differed in the mechanism used to inhibit T-cell responses to M. tuberculosis.

Alveolar macrophages as accessory cells for human gamma delta T cells activated by Mycobacterium tuberculosis.

Alveolar macrophages form the first line of defense against inhaled droplets containing Mycobacterium tuberculosis by controlling mycobacterial growth and regulating T cell responses. CD4+ and gamma delta T cells, two major T cell subsets activated by M. tuberculosis, require accessory cells for activation. However, the ability of alveolar macrophages to function as accessory cells for T cell activation remains controversial. We sought to determine the ability of alveolar macrophages to serve as accessory cells for resting (HLA-DR-, IL-2R-) and activated (HLA-DR+, IL-2R+) gamma delta T cells in response to M. tuberculosis and its Ag, and to compare accessory cell function for gamma delta T cells of alveolar macrophages and blood monocytes obtained from the same donor. Alveolar macrophages were found to serve as accessory cells for both resting and activated gamma delta T cells in response to M. tuberculosis Ag. At high alveolar macrophage to T cell ratios (> 3:1), however, expansion of resting gamma delta T cells was inhibited by alveolar macrophages. The inhibition of resting gamma delta T cells by alveolar macrophages was dose-dependent, required their presence during the first 24 h, and was partially overcome by IL-2. Alveolar macrophages did not inhibit activated gamma delta T cells even at high accessory cell to T cell ratios, and alveolar macrophages functioned as well as monocytes as accessory cells. Monocytes were not inhibitory for either resting or activated gamma delta T cells. These findings support the following model. In the normal alveolus the alveolar macrophage to T cell ratio is > or = 9:1, and therefore the threshold for resting gamma delta T cell activation is likely to be high. Once a nonspecific inflammatory response occurs, such as after invasion by M. tuberculosis, this ratio is altered, favoring gamma delta T cell activation by alveolar macrophages.

CD4+ alpha beta T cell and gamma delta T cell responses to Mycobacterium tuberculosis. Similarities and differences in Ag recognition, cytotoxic effector function, and cytokine production.

CD4+ and gamma delta T cells are activated readily by Mycobacterium tuberculosis. To examine their role in the human immune response to M. tuberculosis, CD4+ and gamma delta T cells from healthy tuberculin-positive donor were studied for patterns of Ag recognition, cytotoxicity, and cytokine production in response to M. tuberculosis-infected mononuclear phagocytes. Both T cell subsets responded to intact M. tuberculosis and its cytosolic Ags. However, CD4+ and gamma delta T cells differed in the range of cytosolic Ags recognized: reactivity to a wide m.w. range of Ags for CD4+ T cells, and a restricted pattern for gamma delta T cells, with dominance of Ags of 10 to 15 kDa. Both T cell subsets were equally cytotoxic for M. tuberculosis-infected monocytes. Furthermore, both CD4+ and gamma delta T cells produced large amounts of IFN-gamma: mean pg/ml of IFN-gamma in supernatants was 2458 +/- 213 for CD4+ and 2349 +/- 245 for gamma delta T cells. By filter-spot ELISA (ELISPOT), the frequency of IFN-gamma-secreting gamma delta T cells was one-half of that of CD4+ T cells in response to M. tuberculosis, suggesting that gamma delta T cells on a per cell basis were more efficient producers of IFN-gamma than CD4+ T cells. In contrast, CD4+ T cells produced more IL-2 than gamma delta T cells, which correlated with diminished T cell proliferation of gamma delta T cells compared with CD4+ T cells. These results indicate that CD4+ and gamma delta T cell subsets have similar effector functions (cytotoxicity, IFN-gamma production) in response to M. tuberculosis-infected macrophages, despite differences in the Ags recognized, IL-2 production, and efficiency of IFN-gamma production.

On the duality between rate and power optimizations

Sequence design problems are considered in this paper. The problem of sum power minimization in a spread spectrum system can be reduced to the problem of sum capacity maximization, and vice versa. A solution to one of the problems yields a solution to the other. Subsequently, conceptually simple sequence design algorithms known to hold for the white-noise case are extended to the colored noise case. The algorithms yield an upper bound of 2N - L on the number of sequences where N is the processing gain and L the number of non-interfering subsets of users. If some users (at most N - 1) are allowed to signal along a limited number of multiple dimensions, then N orthogonal sequences suffice.

Assessment of the mutagenic potential of a fungicide Bavistin using multiple assays

The fungicide Bavistin was assessed for mutagenic potential by various assays. Bavistin was found to be unable to induce gene mutation in Salmonella typhimurium, but it was able to induce transfection inhibition in Mycobacterium smegmatis. Bavistin was able to induce immediate genotoxic effects in plants but these were not carried through in development as in the long term no genotoxic effects were observed by the progeny test. Bavistin did induce micronuclei formation and did cause an increase in the ratio of normochromatic to polychromatic erythrocytes in mice. It was able to induce a very low frequency of sister-chromatid exchange in human lymphocytes and in addition, it was observed that the chemical affected the mitotic index but did not affect the cell cycle duration. Present studies indicate that the pesticide shows a positive response in 4 out of 5 different test systems (Table 8) and most of the observations support that Bavistin is genotoxic.

Design of crosspoint-irredundant PLAs using minimal number of control inputs

The problem of determining a minimal number of control inputs for converting a programmable logic array (PLA) with undetectable faults to crosspoint-irredundant PLA for testing has been formulated as a nonstandard set covering problem. By representing subsets of sets as cubes, this problem has been reformulated as familiar problems. It is noted that this result has significance because a crosspoint-irredundant PLA can be converted to a completely testable PLA in a straightforward fashion, thus achieving very good fault coverage and easy testability.

An algorithm to detect linearly separable clusters of binary patterns

The clusters of binary patterns can be considered as Boolean functions of the (binary) features. Such a relationship between the linearly separable (LS) Boolean functions and LS clusters of binary patterns is examined. An algorithm is presented to answer the questions of the type: “Is the cluster formed by the subsets of the (binary) data set having certain features AND/NOT having certain other features, LS from the remaining set?” The algorithm uses the sequences of Numbered Binary Form (NBF) notation and some elementary (NPN) transformations of the binary data.

An algorithm to detect linearly separable clusters of binary patterns

The clusters of binary patterns can be considered as Boolean functions of the (binary) features. Such a relationship between the linearly separable (LS) Boolean functions and LS clusters of binary patterns is examined. An algorithm is presented to answer the questions of the type: “Is the cluster formed by the subsets of the (binary) data set having certain features AND/NOT having certain other features, LS from the remaining set?” The algorithm uses the sequences of Numbered Binary Form (NBF) notation and some elementary (NPN) transformations of the binary data.

Immediate hypersensitivity to Parthenium hysterophorus. II. Clinical studies on the prevalence of parthenium rhinitis

The airborne pollen of the South American weed, Parthenium hysterophorus (American feverfew), accidentally introduced into India was found to be responsible for severe allergic rhinitis. A random clinical survey conducted on 2035 residents of Bangalore city with the aid of questionnaires and skin tests revealed that 7.1% of the study population was suffering from allergic rhinitis due to exposure to Parthenium pollen. Skin-prick tests performed on 1294 clinic patients suffering from nasobronchial allergy during the past 4 years have also shown that 42.5% were sensitive to Parthenium pollen. IgE and IgG antibodies specific for Parthenium pollen allergens were demonstrable in the sera of Parthenium-sensitive rhinitis patients. The specificity of these antibodies to Parthenium allergens was established by ELISA. A 7- to 11-fold higher stimulation was observed when lymphocytes from two Parthenium-sensitive rhinitis patients were treated in vitro with Parthenium pollen extract. To our knowledge, nowhere in the world has such a high incidence of allergic rhinitis due to a single pollen ever been reported.

Immediate hypersensitivity to Parthenium hysterophorus. I. Association of HLA antigens and Parthenium rhinitis

Lymphocytes collected from rhinitis subjects with strong positive skin reactions to the pollen allergens of Parthenium hysterophorus (American feverfew) having moderate to high titres of Parthenium-specific serum IgE were analysed for association of HLA-antigens covering 13 specificities of HLA-A, 17 specificities of HLA-B and eight specificities of HLA-DR loci by the NIH two-stage microlymphocytotoxicity assay. Comparison of the phenotypic frequencies of HLA-A and B antigens between Parthenium rhinitis subjects (n= 22) and control subjects (n= 137) did not suggest any significant association when tested for these antigen specificities. A significant correlation in the association of HLA-DR3 antigen with a relative risk of 11·33, however, was observed in Parthenium rhinitis subjects (n= 30) when compared to controls (n= 50).

Allergenic Cross-Reactivity between Parthenium and Ragweed Pollen Allergens

Cross-reactivity of allergens from the pollen of the Compositae weeds, Parthenium hysterophorus (American feverfew) and Ambrosia (ragweed), in 2 groups of patients with different geographic distributions was studied. Parthenium-sensitive Indian patients, who were never exposed to ragweed, elicited positive skin reactions with ragweed pollen extracts. A significant correlation in the RAST scores of Parthenium and ragweed-specific IgE was observed with the sera of Parthenium and ragweed-sensitive Indian and US patients, respectively. RAST inhibition experiments demonstrated that the binding of IgE antibodies in the sera of ragweed-sensitive patients to short (Wl) and giant (W3) ragweed allergen discs could be inhibited by up to 94% by Parthenium pollen extracts. Similar inhibition (up to 82%) was obtained when the sera of Parthenium rhinitis patients were incubated with ragweed allergen extracts. A dose-dependent proliferation of lymphocytes from a Parthenium-sensitive rhinitis patient with elevated levels of both Parthenium and ragweed-specific IgE was observed when incubated with Parthenium and ragweed pollen extracts. A 1.6-fold higher proliferation, however, was observed with Parthenium pollen extract at a concentration of 100 µg/ml. These results suggest that shared epitopes present on Parthenium and ragweed pollen allergens are recognized by both Indian and US patients sensitized by exposure to Parthenium and ragweed pollen, respectively. The high degree of cross-reactivity between Parthenium and ragweed pollen allergens suggests that individuals sensitized to Parthenium may develop type-I hypersensitivity reactions to ragweed and vice versa when they travel to regions infested with the weed to which they had not been previously exposed.

Salmonella enterica Serovar Typhimurium Lacking hfq Gene Confers Protective Immunity against Murine Typhoid

Salmonella enterica is an important enteric pathogen and its various serovars are involved in causing both systemic and intestinal diseases in humans and domestic animals. The emergence of multidrug-resistant strains of Salmonella leading to increased morbidity and mortality has further complicated its management. Live attenuated vaccines have been proven superior over killed or subunit vaccines due to their ability to induce protective immunity. Of the various strategies used for the generation of live attenuated vaccine strains, focus has gradually shifted towards manipulation of virulence regulator genes. Hfq is a RNA chaperon which mediates the binding of small RNAs to the mRNA and assists in post-transcriptional gene regulation in bacteria. In this study, we evaluated the efficacy of the Salmonella Typhimurium Dhfq strain as a candidate for live oral vaccine in murine model of typhoid fever. Salmonella hfq deletion mutant is highly attenuated in cell culture and animal model implying a significant role of Hfq in bacterial virulence. Oral immunization with the Salmonella hfq deletion mutant efficiently protects mice against subsequent oral challenge with virulent strain of Salmonella Typhimurium. Moreover, protection was induced upon both multiple as well as single dose of immunizations. The vaccine strain appears to be safe for use in pregnant mice and the protection is mediated by the increase in the number of CD4(+) T lymphocytes upon vaccination. The levels of serum IgG and secretory-IgA in intestinal washes specific to lipopolysaccharide and outer membrane protein were significantly increased upon vaccination. Furthermore, hfq deletion mutant showed enhanced antigen presentation by dendritic cells compared to the wild type strain. Taken together, the studies in murine immunization model suggest that the Salmonella hfq deletion mutant can be a novel live oral vaccine candidate.

Characterization of a variant glucose-6-phosphate dehydrogenase from the south Indian population.

Glucose-6-phosphate dehydrogenase (G6PD) is coded by a gene on the X-chromosome. Earlier studies have shown that the South Indian population has a high incidence of this enzyme deficiency. The electrophoretic mobility, pH optimum and the K-m values for G6PD from normal and variant individuals were identical. However, the specific activity of the variant enzyme was 8 times less compared to the value of the normal enzyme. Western blot analysis of partially purified G6PD from normal and variant individuals performed using equal amounts of total protein showed that the variant protein was 3 times less in concentration. Similar analysis performed using protein corresponding to equal enzyme activity units in the normal and variant samples showed that the variant enzyme was 2.25 times less efficient compared to the normal enzyme. RNA dot blot analysis using full length G6PD cDNA probe (PGDT5B, a kind gift from Prof. L Luzzatto) revealed that lymphocytes from normal and variant individuals had equal amounts of G6PD specific mRNA.

Optimal feature extraction for bilingual OCR

Feature extraction in bilingual OCR is handicapped by the increase in the number of classes or characters to be handled. This is evident in the case of Indian languages whose alphabet set is large. It is expected that the complexity of the feature extraction process increases with the number of classes. Though the determination of the best set of features that could be used cannot be ascertained through any quantitative measures, the characteristics of the scripts can help decide on the feature extraction procedure. This paper describes a hierarchical feature extraction scheme for recognition of printed bilingual (Tamil and Roman) text. The scheme divides the combined alphabet set of both the scripts into subsets by the extraction of certain spatial and structural features. Three features viz geometric moments, DCT based features and Wavelet transform based features are extracted from the grouped symbols and a linear transformation is performed on them for the purpose of efficient representation in the feature space. The transformation is obtained by the maximization of certain criterion functions. Three techniques : Principal component analysis, maximization of Fisher's ratio and maximization of divergence measure have been employed to estimate the transformation matrix. It has been observed that the proposed hierarchical scheme allows for easier handling of the alphabets and there is an appreciable rise in the recognition accuracy as a result of the transformations.