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J Ivanyi

Publications and source records attributed to J Ivanyi.

At least 55 records · Page 3Linked to original sources
PubMed1996

Genetic and serovar typing of clinical isolates of the Mycobacterium avium-intracellulare complex.

SETTING: One hundred and thirty-four Mycobacterium avium-intracellulare complex (MAC) isolates were obtained from 121 patients in the UK. OBJECTIVE: To compare serotyping and genetic analysis for species identification of MAC isolates from patients with and without the acquired immunodeficiency syndrome (AIDS). DESIGN: Clinical MAC isolates were cultured and analyzed by serotyping, the commercially available Accuprobe kit, hybridization with genes coding for the 19 kDa and 38 kDa antigens of M. tuberculosis and fingerprinting with the pMB22 probe derived from M. paratuberculosis. RESULTS: Species classification on the basis of genetic analysis was similar to serovar typing, with only exceptional discrepancies. Serovar prevalence was different in the two groups of patients, and different from those reported in other countries. MAC isolates from AIDS patients were exclusively M. avium, whereas patients without AIDS had MAC infections with M. avium and M. intracellulare in about equal proportion. M. intracellulare clinical isolates were genetically more heterogeneous than M. avium. Only M. intracellulare hybridized with the 38 kDa gene probe. CONCLUSIONS: Serovars are strongly linked with species in clinical MAC isolates, confirming results previously obtained with reference strains. M. intracellulare can be easily identified by the presence of a 38 kDa gene.

AIDS-Related Opportunistic Infections↗
PubMed1996

T cell responses to a mixture of Mycobacterium tuberculosis peptides with complementary HLA-DR binding profiles.

The T cell response to a mixture of eight peptides derived from sequences of the Mycobacterium tuberculosis 16-, 19- and 38-kD antigens (MTBmix-8) has been studied. The peptides were selected on the basis of complementary binding to nine HLA-DR molecules (HLA-DR1 to DR9). MTBmix-8 at 6.25 and 50 micrograms/ml gave rise to significant stimulation (P < 0.05) of peripheral blood mononuclear cells (PBMC) from healthy tuberculin-positive and both untreated and treated diseased subjects, but not in any of a control group of healthy tuberculin-negative subjects. MTB-mix-8 stimulated proliferation of PBMC from healthy tuberculin-positive individuals at lower concentrations than the individual component peptides. However, the maximal stimulation achieved was only slightly higher than that achieved with individual peptides. MTBmix-8 also stimulated the production of interferon-gamma (IFN-gamma) in vitro. Using the mean +/- 2 s.d. of the values for IFN-gamma production in the tuberculin-negative population as a cut-off, MTBmix-8 at 6.25 micrograms/ml was able to detect infection with a sensitivity of 100% in untreated patients, 87% in treated patients, and 82% in tuberculin-positive controls. The corresponding figures for the most potent single peptide (16p91-110) were: 66% in untreated patients, 71% in treated patients and only 42% in controls. Thus, using the IFN-gamma-based assay, which has the additional advantages of speed and does not require radioactivity, the mixture of peptides is more sensitive than single peptides in diagnosing infection.

Adult↗
PubMed1996

Increase of tuberculous infection in the organs of B cell-deficient mice.

Protective immunity against infection with Mycobacterium tuberculosis is imparted by T cells rather than antibodies, but B cells can play a role as antigen-presenting cells and in granuloma formation. We re-evaluated the role of B cells in the course of tuberculous infection in mu-chain knock-out (Ig-) mice. Surprisingly, the organs of M. tuberculosis-infected Ig- mice were found to have three- to eight-fold elevated counts of viable bacilli compared with normal littermates at 3-6 weeks post-infection. Splenic interferon-gamma responses to whole antigen were unimpaired, whilst proliferation to certain mycobacterial peptides was found to be diminished. However, bacille Calmette-Guérin (BCG) vaccination significantly reduced the infection in Ig- mice. The mechanisms by which B cells can influence primary tuberculous infection need further study.

Animals↗
PubMed1996

Immunogenicity of peptides for B cells is not impaired by overlapping T-cell epitope topology.

The epitope specificity of T-cell help to B cells and of surface immunoglobulin-mediated B-cell-binding of antigens usually involves topographically distinct antigenic determinants. The possibility of cross-recognition of the same peptide sequence by both T cells and antibodies has been a matter of conflicting opinions. We investigated this subject by detailed mapping of T- and B-cell epitopes within four immunogenic mycobacterial peptides. The identified core sequences of T- and B-cell epitopes showed different topology within each peptide: they were partially overlapping or adjacent in two P38-derived peptides, but entirely overlapping in two P19-derived peptides. The critically important result using the two truncated peptides (P19/67-78 and P19/146-155) containing only the fully overlapping epitope cores was, that they retained full potency for inducing antibody responses. However, despite this desirable overlap of determinants, antipeptide sera failed to block the proliferation of corresponding T-cell hybridomas. We conclude, that our study, in contrast to previous findings, suggests that overlapping topology of T- and B-cell epitopes within synthetic peptides does not necessarily impair B-cell immunogenicity.

Amino Acid Sequence↗
PubMed1996

Distinct conformations of a peptide bound to HLA-DR1 or DRB5*0101 suggested by molecular modelling.

The conformation of peptides when bound to different HLA class II molecules is of interest in the study of specificity and function of responding T cells. Here, we report the investigation of the HLA-DR binding profiles of an immunodominant and HLA-promiscuous mycobacterial peptide, p38G. Its binding affinities were found to be high for DR1, moderate for DR2, DR7 and DR8, low for DR4, DR5, DR6 and DR9, and below detection for DR3. The minimum peptide length required for binding was, in the majority of cases, nine residues and 11 in two instances (DR2 and DR4). Peptide binding to DR2 was attributed to the DRB5*0101 and not to the DRB1*1501 gene product. Substitution analysis of the amino acid residues involved in binding to DR1 and DRB5*0101 identified F-354 as the common primary contact residue (P1), while allele-specific differences were found in positions P4, P6 and in the C-terminal anchor residue (valine at P9 for DR1 or lysine at P10 for DRB5*0101). Computer-assisted evaluation of these empirical data produced a molecular model, suggesting that the peptide binds to DR1 in an elongated conformation, similar to that of other peptide MHC class II complexes. In contrast, the DRB5*0101 bound peptide is likely to be kinked, which so far was considered characteristic only for peptides within MHC class I complexes. The different conformations imposed on the same peptide by distinct HLA alleles may represent an important mechanism for the control of T cell responses.

Alleles↗
PubMed1996

Multiple sclerosis: a protective or a pathogenic role for heat shock protein 60 in the central nervous system?

The stress proteins belonging to the heat shock protein 60 (hsp6O) family of molecular chaperones with known immunogenic properties are expressed at increased levels in a number of autoimmune conditions. Because previous studies from this laboratory suggested that hsp6O may be involved in the pathogenesis of the chronic multiple sclerosis (MS) plaque, we have examined autopsied central nervous system tissue from 10 cases of MS, ranging in clinical history from acute to chronic inactive. MS lesions ranged from acute, actively demyelinating and edematous, to fibrous astrogliotic and chronically demyelinated. As controls, central nervous system tissue from other neurologic diseases and nonneurologic conditions was used. Frozen, paraffin, and epoxy-embedded sections were studied immunocytochemically with the ML30 mAb to hsp6O. Acute MS lesions displayed the greatest reactivity, with particularly prominent staining of hypertrophic astrocytes, reactive macrophages, and hyperplastic oligodendrocytes. In all these cells, elevated expression occurred in the constitutive site for hsp6O (mitochondria) and within the cytosol, which is suggestive of a shift in expression. The hsp6O-reactive oligodendrocytes were structurally intact. Chronic active MS lesions also revealed the highest levels of hsp6O in hypertrophic astrocytes and oligodendrocytes. Chronic silent MS lesions displayed elevated hsp6O in hypertrophic astrocytes only while constitutive expression occurred elsewhere in the central nervous system at levels slightly higher than normal. Other neurologic disease tissue displayed expression elevated above that found in nonneurologic cases, but this was considerably less than that seen in acute MS. Of the other neurologic diseases, AIDS encephalitis revealed the greatest activity for hsp6O, with both mitochondrial and cytosolic staining of astrocytes. It is proposed that the high levels of hsp6O in hyperplastic, structurally intact oligodendrocytes in acute MS lesions may bespeak a protective mechanism, whereas hsp6O in chronic active lesions may serve a pathogenic role in the later depletion of these cells.

Acute Disease↗
PubMed1995

Abundance of H-2 promiscuous T cells specific for mycobacterial determinants in H-2b/d F1 hybrid mice.

A majority of immunodominant epitopes of mycobacterial antigens are known to be recognized by murine T cells in the context of several H-2 haplotypes. In this study, we established the frequency of T cells able to recognize these peptides promiscuously, i.e. in the context of allogeneic antigen-presenting cells, using hybridomas from peptide-immunized H-2 homologous and heterologous mice. The degree of promiscuity in homozygous mice varied between 4-27% between different specificities and genetic backgrounds. In particular, the results showed that promiscuity between Ab and Ad in respect to a peptide from the Mycobacterium tuberculosis 38-kDa protein (residues 350-369) was displayed by 22% of BALB/c and 4% of C57BL/10-derived hybrids, but by 42% of [BALB/c x C57BL/10] F1-derived clones. This represents a significant increase (p < 0.001) of T cell promiscuity compared to the parental haplotypes. It is noteworthy that considerably lower peptide concentrations were able to stimulate the promiscuous hybridomas compared to the H-2-restricted hybrids. This finding suggests a functional advantage of promiscuous T cells which enables them to expand preferentially in the initial stages of infections with M. tuberculosis and thus enables the host to mount a rapid protective immune response.

Animals↗
PubMed1995

Cross-recognition by T cells of an epitope shared by two unrelated mycobacterial antigens.

The molecular mimicry represented by cross-recognition of determinants shared by unrelated antigens by antibodies or T cells is of broad immunological interest. In this study, we analyzed the cross-recognition by CD4+ T cells of a peptide epitope shared by two mycobacterial proteins of diverse sequence, represented by the 19-kDa antigen of Mycobacterium tuberculosis and the 28-kDa antigen of Mycobacterium leprae. This epitope was immunodominant with respect to the 19-kDa antigen, but cryptic in relation to the 28-kDa antigen. The cross-reactive epitope cores were identified by Pepscan window analysis and found to be eight residues long in both antigens (residues 69-76 and 127-134). Alignment of these octameric sequences revealed two identical and five conservatively related amino acids. Within the epitope core, two residues (73Asn and 76Ile) were identified as critical for recognition on the basis of inhibition of the cross-reactive T cell proliferative response using singly substituted analog peptides. These results suggest that T cell cross-reactive epitopes can exist in proteins with apparently not more than random levels of sequence homology. Their potential for unsuspected cross-sensitization may play a role in the maintenance of T cell memory, in the pathogenesis of autoimmune diseases and possibly in a wide range of host immune responses to infectious pathogens.

Amino Acid Sequence↗
PubMed1995

The nature of the immunogen determines the specificity of antibodies and T cells to selected peptides of the 38 kDa mycobacterial antigen.

We report in this study that the nature of the injected immunogen or infection of mice can change the specificity of antibodies against different epitopes of the mycobacterial 38 kDa antigen. Following immunization with a selection of 14 peptides, only p65-83 and p285-304 resulted in antibodies binding to the whole antigen, whereas the recombinant antigen induced antibodies to p201-220 and p230-249, which are non-immunogenic following peptide immunization. Killed tubercle bacilli stimulated antibodies exclusively against the N-terminal 1-20 peptide, whilst infection with live tubercle bacilli led to antibodies recognizing non-continuous epitopes, demonstrable by binding to the 38 kDa protein, but to none of the tested peptides. The most striking finding is that protein immunization failed to induce antibodies to p65-83, although this epitope is immunodominant for T cells, irrespective of the mode of sensitization. Since this peptide contains non-overlapping B (69-75) and T (75-81) cell stimulatory cores, it appears that T help dedicated to the adjacent B epitope following peptide immunization may have shifted in favour of B cells reacting either with distantly located linear epitopes p1-20 and p201-220 following immunization with the whole protein, or to conformational epitopes following tuberculous infection of mice. The mechanisms which can deviate T help from one to another B cell specificity could involve regulation of antigen processing, liable to the nature of the immunogen. The described results suggesting distinct functional pairing between T and B epitopes, but with limited fidelity, may have general implications for the design of subunit vaccines.

Amino Acid Sequence↗
PubMed1995

Ribosomal internal transcribed spacer sequences are identical among Mycobacterium avium-intracellulare complex isolates from AIDS patients, but vary among isolates from elderly pulmonary disease patients.

Sequencing 280 bp of the internal transcribed spacer (ITS) between the 16S and 23S rRNA genes in a collection of 46 clinical isolates of the Mycobacterium avium-intracellulare complex (MAI complex) identified nine different sequences, grouping these isolates in nine 'ITS sequevars'. This analysis extends the subdivision within the MAI complex to 18 ITS sequevars and also improves discrimination from other mycobacterial species. Evaluation of the sequevar grouping among different clinical sources revealed strong association of the M. avium sequevar Mav-B with AIDS and with lymphadenitis in children (18 out of 20 and 3 out of 3 respectively). Isolates from elderly patients with pulmonary disease and not suspected of being HIV infected belonged predominantly to M. intracellulare ITS sequevars and sequevars not assigned to either M. avium or M. intracellulare. On the other hand, animal isolates were of both the Mav-A and Mav-B sequevars. We conclude that ITS sequevar typing is an accurate way of identifying distinct MAI complex strains. The observed differences between clinical sources suggest that ITS sequevars reflect possibly important, biologically and clinically relevant polymorphisms between MAI complex organisms.

Acquired Immunodeficiency Syndrome↗
PubMed1995

Human T cell responses to peptide epitopes of the 16-kD antigen in tuberculosis.

The 16-kD protein constituent of the Mycobacterium tuberculosis complex has been known mainly for its prominent serological immunogenicity and species specificity in tuberculous infection. In this study, we evaluated the T cell immune repertoire in 27 sensitized healthy subjects and 46 patients with active tuberculosis using 14 overlapping 20mer peptides spanning the entire sequence of this protein. Four of the tested peptides individually stimulated proliferation of blood mononuclear cells from more than 50% of healthy controls. Tuberculosis patients reacted to a narrower peptide range and with a 17-27% lower rate of responses to the four most immunogenic peptides, but these differences do not distinguish in any simple way between the T cell repertoire of patients and sensitized healthy subjects. The most immunogenic peptide (91-110) was recognized by 67% of healthy subjects and by 50% of tuberculosis patients. Importantly, several non-responders to this peptide were stimulated with the other three most permissive peptides with sequences of 111-130, 71-91 and 21-40, resulting in an overall response rate to at least one of these four peptides of 93% in healthy controls and 74% in tuberculosis patients. In view of this additive effect between the most immunogenic peptides, their combined use may achieve sufficient sensitivity in a test aimed at the specific discrimination between infected and non-infected healthy subjects. The major interest in testing with these peptides rests in their species specificity, which is not achieved using purified protein derivative (PPD).

Amino Acid Sequence↗
PubMed1995

Delayed hypersensitivity and HLA in smear-positive pulmonary tuberculosis.

Tuberculin responses were quantified by induration of the skin, velocity of blood flow in dermal microcirculation and composition of the cellular infiltrate in 125 patients with tuberculosis and 39 healty controls. The diameters of the tuberculin responses were greater in HLA-DR15-positive patients than in DR15-negative patients. The density of infiltrating CD(4+)-positive cells showed a positive correlation with induration in DR15-negative subjects (r = 0.54). A fraction of DR15-positive patients gave large tuberculin responses (> or = 15 mm) but with few CD4+ cells in the test site (< 500/mm2); these patients had a greater percentage of cells in the diffuse dermal infiltrate than in the perivascular region, greater blood flow velocities in the tuberculin response but more frequently with central relative slowing of blood flow and had higher total IgG and specific antimycobacterial antibody levels compared to other DR15-positive patients. If the inflammatory infiltrate in the lungs parallels that in the tuberculin skin test, the lack of immunocompetent cells and tissue hypoxia could permit pulmonary cavitation and explain the association of HLA-DR15 with smear-positive pulmonary tuberculosis.

Blood Flow Velocity↗
PubMed1995

H-2-associated effects of flanking residues on the recognition of a permissive mycobacterial T-cell epitope.

Previously we have identified an immunodominant, eight-residue, epitope core sequence (TAAGNVNI) from the 19,000 MW protein of Mycobacterium tuberculosis, which is recognized in the context of multiple H-2 I-A molecules. In this study, the role of residues flanking this T-cell epitope core was examined, using a series of 20 mer analogue peptides in which the native flanking residues were progressively replaced with L-alanine. Analogue peptides were tested for their capacity to stimulate a CD4+ 19,000 MW protein-specific T-cell line, revealing that all but one N-terminal flanking residue could be replaced collectively by alanine without significant loss of stimulatory activity. However, clear H-2-associated differences in the requirement for flanking residues were demonstrated with peptide-specific T-cell hybridomas. In particular, H-2d-derived hybridomas were much more stringent in their requirement for flanking residues than were H-2b hybridomas. All polyalanine-substituted peptides bound I-Ab molecules, with affinities similar to the native unsubstituted peptide. In contrast, significantly reduced binding to I-Ad was observed with several analogue peptides, although without a clear relationship to the degree of substitution. Furthermore, in H-2b mice, neither immunogenicity nor cross-reactivity with the native peptide showed a clear inverse relationship with respect to the degree of alanine substitution. The results presented in this paper indicate that flanking residues can influence T-cell specificity and that these effects may be controlled by major histocompatibility complex (MHC) haplotype.

Alanine↗
PubMed1995

Sustained T-cell reactivity to Mycobacterium tuberculosis specific antigens in 'split-anergic' leprosy.

Split anergy represented by delayed-type hypersensitivity skin reaction to tuberculin, but not to leprosin, is known to occur in a distinct proportion of leprosy patients. The mechanism was originally attributed to Mycobacterium leprae-specific suppression of T cells toward common mycobacterial antigens. This study ascertained an alternative explanation, attributing the phenomenon to selective responsiveness to M. tuberculosis-specific epitopes. Indeed, the results of blood T-cell proliferative responses in 11 split-anergic patients showed normal responsiveness to the M. tuberculosis-specific 38 kDa lipoprotein and peptide 71-91 of the 16 kDa antigen but diminished responsiveness to 2 common mycobacterial antigens, represented by the 65 kDa heat shock protein and the fibronectin-binding Ag85 complex, as compared with leprosin responsive patients and healthy contacts. These findings support the hypothesis that split anergy is due to selective recognition of M. tuberculosis-specific epitopes and deletion of T cells reacting to shared mycobacterial antigens.

Adolescent↗
PubMed1995

Human CD4-CD8- alpha beta + T-cell receptor T cells recognize different mycobacteria strains in the context of CD1b.

Double-negative alpha beta+ T-cell receptor (TCR) human T cells have been reported to recognize antigen in the context of the HLA class I-like (Ib) CD1 complex. In particular, the CD1b molecule has been shown to act as the element of genetic restriction for antigens derived from Mycobacterium tuberculosis. The stenotopic nature of these major histocompatibility complex (MHC) class Ib molecules raised the question of whether the antigenic moiety recognized by CD4-CD8- alpha beta+ TCR T cells was shared by different mycobacteria. We demonstrate here that a CD4-CD8- alpha beta+ TCR T-cell line and three clones raised against M. tuberculosis proliferated following stimulation with soluble extracts from organisms of the M. tuberculosis complex, M. leprae and 10 out of 16 tested isolates of M. avium complex; however, four species of weakly or non-pathogenic mycobacteria were not stimulatory. Furthermore, the M. tuberculosis soluble extract (MTSE)-derived, recognized antigenic moiety proved to be proteinase K resistant and to have a molecular weight greater than 5000 MW, thus it differed from the reported antigenic moiety, recognized by CD4-CD8- gamma delta+ TCR cells. Our results suggest that a common antigenic moiety, presented by CD1b molecules to CD4-CD8- alpha beta+ TCR T cells, is shared by many mycobacterial species. Therefore they raise interest in the question of whether CD4-CD8- alpha beta+ TCR T cells, elicited by M. tuberculosis, may play a role in the natural history of other mycobacterial infections.

Antigens, Bacterial↗
PubMed1995

Permissive recognition of a mycobacterial T-cell epitope: localization of overlapping epitope core sequences recognized in association with multiple major histocompatibility complex class II I-A molecules.

Most T-cell epitopes are recognized in the context of a single or limited number of major histocompatibility complex (MHC) class II molecules. We have shown previously, however, that the immunodominant p61-80 epitope from the Mycobacterium tuberculosis 19,000 MW protein is recognized in a genetically permissive manner. In this study, permissive recognition of p61-80 was analysed in three murine MHC haplotypes (H-2b,d and k) with respect to: (i) T-cell-epitope core structure; (ii) I-A/I-E class II MHC restriction; and (iii) the identification of critical amino acid residues within the core region. Overlapping epitope core sequences composed of 6 to 8 amino acids were identified for each of the three H-2 haplotypes by T-cell epitope scanning (PEPSCAN) using peptide-specific T-cell lines. The epitope core sequences recognized by peptide and 19,000 MW protein-specific T cells were similar. In all three haplotypes, responses to p61-80 were restricted by class II MHC I-A molecules. To identify residues within the epitope core critically required for recognition, single substitution (alanine or leucine) analogue peptides were tested for their capacity to stimulate p61-80-specific T-cell hybridomas. A heterogeneous pattern of reactivity was observed, even among individual hybridomas derived from the same H-2 haplotype. Although every core residue could be defined as critical for at least one hybridoma, only one critical substitution (74Val-->Ala) was common to all hybridomas. The identification and structural analysis of genetically permissive epitopes of mycobacteria may be a useful strategy for the rational design of peptide-based vaccines for tuberculosis.

Amino Acid Sequence↗