Adult human buccal epithelial stem cells: identification, ex-vivo expansion, and transplantation for corneal surface reconstruction.

PURPOSE: To identify adult human buccal epithelial stem cells (SCs) on the basis of two parameters (high p63 expression and greater nucleus/cytoplasmic (N/C) ratio) and to evaluate clinical efficacy of ex-vivo expanded autologous epithelium in bilateral limbal SC-deficient (LSCD) patients. METHODS: The epithelial cells were isolated from buccal biopsy and cultured on human amnion in culture inserts with 3T3 feeder layer. The SCs were identified on the basis of two-parameter analysis using confocal microscopy, surface markers, and colony-forming efficiency (CFE). The cultured epithelium was transplanted in 10 LSCD patients followed by penetrating keratoplasty in 4 patients. The clinical outcome was followed up to 3 years. RESULTS: A distinct population (3.0±1.7%) of small cells expressing high levels of p63 with greater N/C ratio was observed in buccal epithelium. The N/C ratio was found to be more appropriate than cell diameter for two-parameter analysis. These cells located in the basal layer were negative for connexin-43 and positive for melanoma-associated chondroitin sulfate proteoglycan, containing holoclones with 0.2% CFE, thus representing the SC population. After transplantation of cultured epithelium with increased (sixfold) SC content, anatomical and visual improvement was observed at 13-34 months in 3/10 LSCD patients. CONCLUSIONS: The two-parameter SC marker is useful to identify and quantify buccal epithelial SCs. The transplantation of bioengineered SC-rich buccal epithelium is a strategy for corneal surface reconstruction in bilateral LSCD. However, further studies are required to optimize the culture conditions and to look for other sources of adult SCs for better visual outcome.

LruA and LruB antibodies in sera of humans with leptospiral uveitis.

Uveitis can be a serious complication of leptospirosis. Previous studies indicated that the leptospiral lipoproteins LruA and LruB are expressed in the eyes of uveitic horses and that antibodies directed against those proteins show in vitro cross-reactivity with components of equine lens, ciliary body, and/or retina. We now demonstrate that sera from a significant proportion of humans who have leptospiral uveitis also contain antibodies against LruA and LruB. Different categories of nonleptospiral uveitis and autoimmune uveitis were also screened; patients diagnosed with Fuchs uveitis or Behçet's syndrome produced antibodies that cross-reacted with LruA and LruB, suggesting similarities of the autoimmune responses in those diseases with those of leptospiral uveitis.

Field rats form a major infection source of leptospirosis in and around Madurai, India.

AIMS: To determine the seroprevalence of leptospires and to isolate Leptospira spp. from field rats and bandicoots in and around Madurai. MATERIALS AND METHODS: Thirteen rats and five bandicoots were trapped alive from fields in and around Madurai. Blood samples were tested for anti-leptospiral antibodies by microscopic agglutination test while the urine and kidney samples were used for isolation of leptospires. The isolated leptospires were tested for pathogenic status (13 degrees C test and PCR) followed by serological and genetic characterization. RESULTS: Serology revealed the presence of anti-leptospiral antibodies in 58% (7/12) of field rats and leptospires were isolated from two urine and six kidney samples. The bandicoots were negative in both serology and culture. Analysis of the isolates from field rats revealed that all the isolates were pathogenic except for one, which was further confirmed by serological and genetic characterization. Six of the seven pathogenic isolates were identified as L. interrogans serogroup Autumnalis serovar Akiyami A and one as L. borgpetersenii serogroup Javanica serovar Veldrat Batavia 46. CONCLUSIONS: Serology and isolation reveals that field rats are major natural carriers and shedders of leptospires in and around Madurai.

Prevalence of eye signs in congenital rubella syndrome in South India: a role for population screening.

PURPOSE: Congenital rubella syndrome (CRS) resulting from maternal rubella infection, especially in the first trimester, affects an estimated 100 000 infants each year worldwide. Immunisation has reduced its occurrence in the developed world, though it remains a problem in countries with poor immunisation coverage. This population-based study was aimed at screening children below 5 years of age for ocular signs suspicious of CRS. METHODS: Suspected CRS cases were recruited from hospital and outreach services of the Aravind Eye Care System over a 24-month period. Clinical confirmation was based on the fulfilment of the World Health Organization (WHO) definition, and laboratory confirmation was based on a positive test for IgM antibody. RESULTS: Children under 5 years of age (n = 51 548) with ocular complaints were screened for eye signs suspicious of CRS; CRS compatible signs were detected in 1.92% (1090) children. Of these suspects (299), 27.42% were subsequently confirmed clinically according to WHO definition, and (46) 4.2% were serologically (Laboratory) confirmed. Of all the eye signs evaluated for screening, cataracts were the most sensitive (80.43%). CONCLUSIONS: Cataracts among children have a high sensitivity for detecting CRS in India. It is the only clinical eye finding that has a high enough sensitivity and specificity to be useful as a screening tool for CRS.

Evaluation of a commercial rubella IgM assay for use on oral fluid samples for diagnosis and surveillance of congenital rubella syndrome and postnatal rubella.

BACKGROUND: Clinical diagnosis (surveillance) of rubella is unreliable and laboratory confirmation is essential. Detection of virus specific IgM in serum is the most commonly used method. However, the use of serum necessitates the drawing of blood, either through venipuncture or finger/heel prick, which can be difficult in young babies. Oral fluid samples have proved useful as an alternative, less invasive sample for virus specific IgM detection however until recently no commercial rubella IgM tests were available, restricting the usefulness of this approach. OBJECTIVES: To evaluate the performance of the Microimmune Rubella IgM capture EIA using oral fluid samples from outbreaks as well as in cases of suspected congenital rubella syndrome (CRS). STUDY DESIGN: Paired serum and oral fluids were collected from cases during a rubella outbreak in three provinces in Turkey. Matched serum and oral fluid samples were collected from children with suspected CRS in an active surveillance programme at the Aravind Eye Hospital in South India. Serum samples were collected as part of the measles surveillance programme in Ethiopia. RESULTS: On serum samples the sensitivity and specificity of the Microimmune Rubella IgM capture EIA compared to Behring Enzygnost rubella IgM test was 96.9% (62/64; 95% CI 94.2-100%) and 100% (53/53; 95% CI 93.2-100%). On oral fluids compared to matched Behring results on serum the sensitivity was 95.5% (42/44; 95% CI 84.5-99.4%). The sensitivity and specificity of Microimmune Rubella IgM capture EIA on oral fluids from suspected CRS cases compared to serum results using Behring Enzygnost IgM assay was 100% (95% CI 84.5-100%) and 100% (95% CI 95.8-100.0%) respectively. CONCLUSION: Microimmune Rubella IgM capture EIA has adequate performance for diagnosis and surveillance of rubella in outbreak using either serum or oral fluid specimens.

Identification and evaluation of LPS antigen for serodiagnosis of uveitis associated with leptospirosis.

Leptospirosis is a widespread zoonotic disease that affects all mammals in different parts of the world. Though there are many commercial kits available for the diagnosis of systemic leptospirosis, the nature of the antigen has not been described. Therefore, identification of a specific antigen is important. Since ocular involvement in leptospirosis has been reported, there is a need to identify and characterize the leptospiral antigen for diagnosis of uveitis associated with past leptospiral infection (leptospiral uveitis) and for confirming the clinical diagnosis. Seven-day-old culture of Leptospira biflexa serovar Patoc was used for preparing the antigen. The present study included serum samples from 81 patients with clinical criteria for leptospiral uveitis, 15 cataract controls and 15 non-leptospiral uveitis controls. Serum samples were assayed by ELISA using our antigenic preparation and by a microscopic agglutination test (MAT) using 19 serovars. The antigen prepared had 280 micro g LPS ml(-1) and no detectable amount of protein. Silver-staining of SDS-PAGE for protein and LPS, dot blot and Western blot analysis and proteinase K and periodate treatment showed that LPS (13-21 kDa and 28 kDa) in our preparation was the relevant antigen for serodiagnosis. IgG antibodies showed reactivity in both leptospiral uveitis patients and controls. However, on the basis of IgM response to LPS, 48 % of the leptospiral uveitis patients were significantly positive compared with controls; 58 % of leptospiral uveitis patients and none of the controls were positive for MAT. When MAT and IgM ELISA results were considered together, 77 % were significantly positive. LPS is identified as a candidate antigen for serodiagnosis of leptospiral uveitis and has sensitivity and specificity of 48 and 90 %, respectively, in ELISA for IgM antibodies. Confirmation of clinical diagnosis with a specific laboratory test would help to initiate the most appropriate treatment for leptospiral uveitis.

Influence of immunopotentiators on the antiporin immunoglobulin G subclass: distribution and protective immunity against murine salmonellosis.

To improve the immune potential of porin (a pore-forming protein of Salmonella sp.), different immunopotentiators such as Freund's complete adjuvant (FCA), lipopolysaccharide (LPS) and polyoxydonium (PO) were evaluated by studying the nature of the protective immune response induced against murine Salmonellosis. The nontoxic, synthetic heteropolymer polyoxydonium was as good as LPS at inducing antiporin immunoglobulin G (IgG) antibodies and protective immunity. Analysis of the antiporin IgG subclass pattern revealed a preferential increase in a particular subclass based on the immunopotentiator used. Porin, alone or emulsified in FCA, elicited predominantly antiporin IgG1 antibodies, whereas LPS preferentially evoked antiporin IgG2a, IgG2b and IgG3 antibodies. Polyoxydonium induced a clear shift towards antiporin IgG2b antibodies. The significance of these antiporin IgG subclass antibodies in protection against murine Salmonellosis was studied by passive immunization and by analysing the infected mouse sera.

IgG subclass antibodies to mycobacterial sonicate and recombinant antigens in leprosy.

In this study the IgG subclass antibodies to sonicated preparations of Mycobacterium leprae (leprosin A) and BCG (BCG-S) as well as to purified recombinant 65 kDa protein of M. leprae (rML65) were analysed in sera from leprosy patients and healthy household contacts (HFC) and noncontacts (HNC) in a leprosy endemic population. In LBI+ (lepromatous bacterial index positive) patients, IgG3 was predominant in the responses to sonicated antigens of M. leprae. Following chemotherapy, IgG3 responses were reduced while IgG2 levels were increased. On the other hand, IgG response to rML65 was dominated by IgG1 in all the patient and control groups. Interestingly, the level of antileprosin A IgG antibody in erythema nodosum leprosum (ENL) was similar to that of lepromatous groups, while the level of anti-rML65 IgG antibody was significantly reduced in ENL. IgG4 antibodies to the antigens studied were only at low levels in all groups, including ENL. Significant differences were observed between HNC and HFC in the pattern of IgG subclass antibodies to sonicated antigens, even though their antigen specific IgG levels were similar. While HNC showed equivalent proportion of IgG1 and IgG2 in their responses to leprosin A and BCG-S, HFC showed a specific increase in IgG1 levels, suggesting that both groups are distinctly different. Further studies are required to elucidate the functional significance of IgG subclass pattern in pathogenesis and the mechanism of immunoregulation resulting in the high levels of IgG1 and IgG3 antibodies to M. leprae protein antigens in lepromatous leprosy.

Antibody response to recombinant 65-kDa, 70-kDa and 18-kDa mycobacterial antigens in leprosy patients and healthy contacts in a leprosy-endemic population.

Antibody responses to recombinant Mycobacterium leprae 65-kDa (rML65) and 18-kDa (rML18), M. bovis BCG 65-kDa (rMB65) and M. tuberculosis 70-kDa (rMT70) antigens were measured by indirect ELISA in sera from leprosy patients and healthy contacts in a leprosy-endemic area in southern India. Antibody responses to M. leprae-specific epitopes on phenolic glycolipid-I (PGL-I) and a 35-kDa protein antigens also were measured simultaneously by PGL-I ELISA and the serum antibody competition test (SACT), respectively. Significantly higher levels of antibodies of the IgG isotype to rML65 and rMB65 were observed in bacterial index (BI)-positive, lepromatous (LBI+) patients but not in other groups of leprosy patients and endemic controls [healthy family contacts (HFC), healthy hospital contacts (HHC), and healthy non-contacts (HNC)]. LBI+ patients could be distinguished from LBI- patients on the basis of their higher levels of IgG antibodies to rML65, rMB65 and rMT70; lower levels of IgM antibodies to these antigens and higher levels of anti-PGL-I IgM levels. In the former group, 84% were SACT positive in contrast to 39% in the latter groups. Among lepromatous patients good positive correlations were observed between IgG antibody responses to rML65 and rMB65 and anti-PGL-I IgM levels, SACT ID50 titers as well as BIs. Among healthy controls, HFC had higher levels of IgG antibodies to rML65, but lower levels to rMB65 than did HNC. Thirty-nine percent of the HFC were seropositive to anti-PGL-I IgM antibodies in contrast to 4% in the HNC. On the basis of these criteria, the immune profile of the HFC appears to be distinctly different from that of the HNC, even though both groups are from the same endemic area. It is therefore possible that antibody response to defined protein antigens of mycobacteria is influenced by the lesional bacterial load in leprosy patients and by exposure to homologous proteins of M. leprae and/or related environmental mycobacteria in the case of healthy contacts and noncontacts. The above results are discussed in relation to T- and B-cell activity toward M. leprae antigens and the immunoregulatory mechanisms of antibody production in leprosy.

Cellular and humoral immune responses to recombinant 65-kD antigen of Mycobacterium leprae in leprosy patients and healthy controls.

Cellular and humoral immune responses to recombinant 65-kD antigen of Mycobacterium leprae (rML65) were studied in leprosy patients and healthy contacts from a leprosy-endemic population. Peripheral blood mononuclear cells from a considerable proportion of tuberculoid leprosy patients, healthy contacts and non-contacts showed proliferative response to rML65 in vitro. A strong positive correlation was observed between the responses to rML65 and bacille Calmette-Guérin (BCG) or leprosin A. Addition of recombinant IL-2 (rIL-2) enhanced the proportion of responders to rML65 considerably in all groups of leprosy patients, healthy contacts and non-contacts. Among lepromatous patients this enhancement was more pronounced in the bacterial index (BI)-negative group. These results indicate that the 65-kD antigen of Myco. leprae is a dominant T cell immunogen in our study population. Though lepromatous patients showed poor lymphoproliferative response to rML65, their IgG antibody levels to the same antigen were markedly high. Most of the BI-positive lepromatous patients with elevated anti-rML65 IgG levels did not show T cell reactivity even with the addition of rIL-2. On the other hand, tuberculoid leprosy patients, healthy contacts and non-contacts showed good T cell reactivity but low levels of IgG antibodies to rML65, thus indicating the presence of an inverse relationship between cell-mediated and humoral immune responses to a defined protein antigen of Myco. leprae in humans. A significant proportion of individuals among tuberculoid leprosy patients, healthy contacts and non-contacts showed neither T cell reactivity nor elevated levels of IgG antibody to rML65. However, in most of these subjects, a T cell response to rML65 was demonstrable with the addition of rIL-2. These results are discussed with reference to the immunoregulatory mechanisms occurring during Myco. leprae infection on the basis of differential activation of Th1 and Th2 subsets.

T lymphocyte reactivity of leprosy patients and healthy contacts from a leprosy-endemic population to delipidified cell components of Mycobacterium leprae.

In this study, we measured in vitro proliferative responses of peripheral blood mononuclear cells from both leprosy patients across the clinical spectrum and also healthy contacts from a leprosy-endemic population to delipidified cell components of Mycobacterium leprae (DCC) and Dharmendra lepromin. Dharmendra lepromin was poor in inducing in vitro T cell proliferation in all the study groups, even though it elicited marked in vivo skin test reaction in tuberculoid leprosy patients and healthy contacts. In contrast, Dharmendra preparation of BCG induced marked T-cell response in tuberculoid as well as bacterial index negative lepromatous patients. DCC induced a significantly higher lymphoproliferative response than Dharmendra lepromin in all study groups. A significant positive correlation was observed between the lymphoproliferative responses to DCC and BCG. The present study, based on a large number of leprosy patients and healthy contacts, clearly demonstrates that DCC, depleted of glycolipids and lipopolysaccharides, is a good antigenic preparation for evaluating T-cell reactivity to M. leprae.

Class-specific immunoglobulins and antibodies to mycobacterial sonicates and autoantigens in leprosy patients and contacts.

A comprehensive analysis of the humoral immune response in leprosy patients and contacts was undertaken. Class-specific antibodies to four mycobacterial sonicates, three autoantigens and three haptens were estimated by ELISA. It was found that IgG levels varied more extensively than IgM or IgA and that total serum IgG was significantly higher in lepromatous bacterial index positive (LL+ve) and negative LL-ve) leprosy patients than in tuberculoid (TT/BT) patients and controls. The high levels of anti-mycobacterial antibodies found in untreated LL+ve patients were significantly reduced in LL-ve patients after effective chemotherapy. Considerable amount of anti-mycobacterial IgG was also detected in TT/BT patients. Each serum when assayed against sonicates of Mycobacterium leprae, Mycobacterium tuberculosis, ICRC bacilli and BCG gave a similar antibody profile suggesting that these antibodies were directed predominantly against cross-reactive antigens. Up to 75% of LL patients and 35% of TT/BT patients were found to be positive for antibodies to histone, collagen and fibronectin. However, antibodies to several haptens were not detected in any of the patients and controls studied. Taken together, these results suggested that the amount of IgG antibodies is directly correlated with the antigenic load in the system, and that there is no evidence for polyclonal activation. It may be speculated that the regulatory mechanism of antibody production is severely deranged in lepromatous leprosy patients.

Diagnosis of typhoid fever: detection of Salmonella typhi porins-specific antibodies by inhibition ELISA.

Porins are highly immunogenic outer membrane proteins of Salmonella. Sera from typhoid patients contained a high level of IgG antibodies directed to porins of Salm. typhi. Since porins are highly conserved proteins, anti-porins antibodies both from typhoid patients and healthy normals reacted with porins from several Gram-negative bacteria. Therefore, in order to improve the specificity of detecting Salm. typhi porins-specific antibodies, an inhibition ELISA was developed using enzyme-conjugated MoAbs (MP1 and MPN4) specific to Salm. typhi porins. Sera from typhoid patients with positive haemoculture (16 out of 17) inhibited the binding of MP1 to porins, thus showing a positive test for typhoid, whereas sera from patients with other Gram-negative bacterial infections (n = 7) and from healthy volunteers (66 out of 67) were found to be negative. The sensitivity, specificity, accuracy, positive predictive value and negative predictive value of this assay were 94.1, 98.7, 97.8, 94.1 and 98.7% respectively. The validity of our inhibition ELISA for typhoid was higher than that of the Widal test. The diagnosis of typhoid fever as early as 3 days after the onset of fever, using a single specimen is possible.

A ribonuclease inhibitor expresses anti-angiogenic properties and leads to reduced tumor growth in mice.

Our experiments were designed to determine whether recombinant ribonuclease inhibitor (RNasin) could inhibit angiogenesis and reduce tumor growth in adult mice. We used the Fajardo disc angiogenesis assay as the primary means of measuring new blood vessel growth. This assay measures the penetration of cells into a polyvinyl alcohol sponge with a central core of ELVAX-coated sponge containing test substances. Cell penetration was reduced to 29.3% of control (phosphate-buffered saline; heat-inactivated RNasin) values. Endothelial cell influx was measured by lectin staining and confirmed by culturing cells isolated from sponges by collagenase treatment. RNasin also reduced the augmented reaction evoked by either basic fibroblast growth factor (bFGF) or sodium orthovanadate. To confirm the anti-angiogenic activity of RNasin, Hydron-coated polyvinyl sponges containing bFGF or bFGF plus RNasin were implanted into adult mouse corneas. bFGF induced a strong angiogenic response that was almost completely inhibited by RNasin. RNasin-containing ELVAX-coated sponges implanted subcutaneously underneath an intradermal inoculum of C755 mammary tumor cells caused significant reduction in tumor growth (P < 0.005). The antitumor effect of RNasin correlated with its effect on tumor-induced neovascularization, suggesting that the ability of RNasin to affect tumor growth was due to its ability to inhibit angiogenesis.

Mechanism of protective immunity induced by porin-lipopolysaccharide against murine salmonellosis.

Investigations were undertaken to characterize the protective immunity induced by porin-lipopolysaccharide (LPS) against Salmonella typhimurium infection in mice. Mice immunized with porin-LPS showed higher levels of antiporin immunoglobulin G than mice which received porin alone. Further, T cells from porin-LPS-immunized mice showed an augmented proliferative response to porin in vitro compared with the response of T cells from porin-injected animals. The passive transfer of anti-LPS antibodies conferred significant protection (17%), while antiporin serum failed to protect mice against lethal challenge, indicating the protective ability of anti-LPS antibodies. However, the transfer of serum obtained from porin-LPS-immunized mice resulted in better protection (30%) than did anti-LPS or antiporin antibodies alone. In contrast to LPS, monophosphoryl lipid A completely failed to induce protection against lethal infection. However, comparable to the effect of LPS, injection of porin with monophosphoryl lipid A enhanced antibody response and the protective ability of porin (81.25%). The transfer of T cells from porin-LPS-immunized mice provided higher levels of protection (47%) against lethal challenge than did T cells from porin-immunized mice (23%). The combination of T cells and serum from porin-immunized mice transferred 36% protection. However, a combination of T cells and serum from porin-LPS-immunized mice conferred the highest level of protection (92%), which was reflected by the number of survivors (100%) in the porin-LPS-immunized group. These results demonstrate that besides the protective effect of anti-LPS antibodies, the ability of LPS to augment humoral and cell-mediated immune responses to porin confers effective protection against Salmonella infection.

Monoclonal antibodies against Salmonella porins: generation and characterization.

Monoclonal antibodies (mAbs) were generated against porins, one of the major outer membrane proteins of Salmonella typhi. Six clones, designated MP1, MP2, MP3 (IgG2ak), MPN4, MPN6 (IgG1k) and MPN5 (IgG2bk) were characterized by enzyme immunoassay (ELISA) for their reactivity to porins from S. typhi, Salmonella paratyphi A, S. paratyphi B, S. paratyphi C, Salmonella choleraesuis, Salmonella enteritidis, Salmonella krefeld, Salmonella panama, Salmonella typhimurium, Escherichia coli B, Shigella flexneri 1b and Pseudomonas aeruginosa. All the clones positive for S. typhi porins showed varying reactivity towards several Salmonella species. However, none of them was positive for porins from other Gram-negative bacteria or for lipopolysaccharide (LPS). The affinity constant of these mAbs, except MPN4, was found to be in the higher range. Dot ELISA revealed that the mAbs recognized porins only in their native form. The results of inhibition ELISA using horseradish peroxidase (HRP)-conjugated MP1 suggest that the clones MP1, MP2, MP3, MPN5 and MPN6 secreted antibodies to identical epitope(s) of a 36-kDa peptide and MPN4 to a different epitope of a 35-kDa peptide. The possible applications of these mAbs were discussed.

Detection of porin antigen in serum for early diagnosis of mouse infections with Salmonella typhimurium.

The monoclonal antibodies to porin, an outer membrane protein isolated from Salmonella typhimurium and sandwich enzyme linked immunosorbent assay (ELISA) has made possible the detection of porin from sera of S. typhimurium-infected mice. The specificity of the monoclonal antibodies was ascertained based on their cross-reactivity with porins isolated from S. typhi, Shigella flexneri and Escherichia coli and lipopolysaccharide (LPS) of S. typhimurium and E. coli. Serum samples were found to be positive for porin as early as 3 days after intravenous and 5 days after oral infection. In addition, a positive correlation was observed between the bacterial load and the concentration of porin detected in the sera. On the other hand, analysis of sera for anti-porin antibody showed diametrically opposite time kinetics with antigenaemia. These results indicate that porin accumulates in the serum of infected mice much earlier than the appearance of antibodies to porin. Thus detection of porin holds promise for early diagnosis of typhoid.

Flow cytometric analysis of CD2 modulation on human peripheral blood T lymphocytes by Dharmendra preparation of Mycobacterium leprae.

It has been reported previously that Mycobacterium leprae modulated CD2 on human peripheral blood T lymphocytes and that this modulation was accompanied by a marked reduction in the proliferative response of these cells to mitogens and antigens. In this study, we report that treatment of peripheral blood mononuclear cells from healthy individuals with Dharmendra preparation of M. leprae inhibited their ability to form rosettes with sheep red blood cells. Flow cytometric analysis of Dharmendra lepromin-treated cells showed that, in addition to CD2, CD4 and CD8 were modulated while the surface expression of CD3 was not affected. The specificity of CD2 modulation was confirmed by similar effects of Dharmendra lepromin on thymocytes and lymph node cells from human CD2 transgenic mice. The modulatory effect of Dharmendra lepromin was not observed at lower temperatures. Dharmendra lepromin treatment of activated T cells resulted in reduced binding of monoclonal antibodies to IL-2R and D66 epitope of CD2. The modulatory effects were not observed with Dharmendra preparation of BCG or other preparations of M. leprae. Our results indicate that certain M. leprae factor(s) specifically modulate(s) CD2, CD4, CD8 and IL-2R but not CD3 on T lymphocytes. The suppressive effect of Dharmendra lepromin on the T-cell proliferative response reported earlier may be explained by its modulatory effect on a number of T-cell surface molecules.