Harpreet Vohra

Group A streptococcal sore throat in a periurban population of northern India: a one-year prospective study

OBJECTIVE To estimate the incidence and risk factors of group A streptococcus (GAS) sore throat among school-aged children living in a periurban slum area of Chandigarh, North India. METHODS A total of 536 children aged 5-15 years from 261 families identified by a systematic random selection method were enrolled in the study. Episodes of sore throat were recorded through fortnightly home visits over a one-year period. The local vernacular (Hindi) terms gala kharab (bad throat) and khansi jukam (cough and cold) were used to identify symptoms of sore throat, and throat swab specimens were collected from children who had these symptoms on the day of the home visit. Bacterial culture was carried out and the isolation of GAS was confirmed using group-A-specific antiserum. FINDINGS The incidences of sore throat and GAS sore throat were, respectively, 7.05 and 0.95 episodes per child-year. The incidence was higher in the following situations: among 11-year-olds, during the winter (November to January) and rainy (August) months (a bimodal peak), among children living in houses where there was no separate room for the kitchen, and in homes that included a tobacco smoker. CONCLUSION The results show that the incidence of GAS sore throat was related to age, season, and indoor air pollution.

Protective Efficacy and Immunogenicity of Vi-Porin Conjugate against Salmonella typhi

A conjugate vaccine against Salmonella typhi was prepared by covalently binding capsular polysaccharide (Vi) with porin, both isolated from S. typhi. First, Vi and porins were extracted. The Vi was purified from S. typhi Ty2. The purified Vi conformed to the requirements of the World Health Organization. Porins were purified from S. typhi 0901. The Vi was bound to the porins by a heterobifunctional cross‐linking reagent, N‐succinimidyl‐3‐(2‐pyridyl dithio)‐propionate (SPDP). After preparing the Vi‐porin conjugate, its protective ability and immunogenicity were studied in mice following systemic immunization. The results showed that the conjugate is 6.5‐fold more protective than Vi alone against S. typhi. The mice immunized with conjugate elicited higher anti‐Vi antibody (IgG) levels (P<0.01) than the mice immunized with Vi alone. Anti‐porin antibodies were also induced by the conjugate. To study the mucosal immune responses, secretory IgA (sIgA) in the intestinal fluid was measured. Conjugate‐immunized mice showed the induction of sIgA as compared to Vi alone. The results showed that when Vi is bound to porins, both isolated from same organism, the resultant conjugate induced both systemic and mucosal immune responses and provided better protection against S. typhi than Vi alone.

Leishmania donovani Infection of a Susceptible Host Results in Apoptosis of Th1‐Like Cells: Rescue of Anti‐Leishmanial CMI by Providing Th1‐Specific Bystander Costimulation

A protective immune response against Leishmania donovani infection is mediated by T‐helper type 1 (Th1) cells. Th1 induced cell‐mediated immunity (CMI), as assessed by anti‐leishmanial DTH response, is lost in a susceptible host such as BALB/c mice. Although the impaired Th1 function eventuates in unhindered parasite growth and in manifestation of the susceptible phenotype, the mechanism of down‐regulation of the Th1 function is yet to be elucidated. Here, we provide evidence that the parasite downregulates the expression of a Th1‐specific costimulatory molecule, M150, on the surface of infected BALB/c mice‐derived macrophages. Th cells are rendered unresponsive to anti‐CD3 Ab‐mediated stimulation after interaction with infected macrophages. The anergized T cells produce much less IL‐2, IL‐4 and IFN‐γ compared to those T cells which were costimulated using normal macrophages. The defect in proliferation, anti‐CD3 Ab induced unresponsiveness and IFN‐γ but not IL‐4 production can be restored by providing bystander costimulation through M150. These results not only unfold a novel immune evasion strategy used by the parasite but also clarify the mechanism of Th1 cell debilitation during the disease. Recovery of Th1 cytokine production by bystander costimulation through M150 may help in formulating a new strategy for the elimination of intracellular parasites.

Effective human defense against E. histolytica: high amoebicidal activity of lymphocytes and monocytes in amoebic liver abscess patients until 3 months follow-up.

Adherence of pathogenic Entamoeba histolytica trophozoites mediated by Gal/GalNAc lectin is a prerequisite for killing naïve T cells and monocytes but the activated T cells and monocyte derived macrophages (MDMs) not only resist the attack but can kill the parasite. In the present study, we have analysed the adherence and cytotoxicity of the immunecompetent cells from patients of amoebic liver abscess at the time of their diagnosis and after 3 months to elucidate the development of cell mediated cytotoxicity, a major mechanism of resistance to amoebic infection. The results show that CD3+ cells from amoebic liver abscess cases, when stimulated, in vitro, bound E. histolytica trophozoites with increased intensity and their viability was also increased. The activated lymphocytes (taken at 3 months post treatment) were also able to kill amoebae. MDMs bound amoebae with greater intensity than lymphocytes, until 3 months post infection. These MDMs were effective in killing approximately 40% amoebae which was significantly less than at the time of diagnosis but was very significant as compared to the controls. The data suggest that cell mediated cytotoxic responses are maximum until 1 month post treatment and are significantly reduced thereafter.

Delivery of antigen in allogeneic cells preferentially generates CD4+ Th1 cells

We have examined the possibility of evoking antigen‐specific T cell immune response by using allogeneic cells as a source of adjuvant and also as a vehicle to deliver antigen. The mice were immunized with different preparations of antigen‐pulsed allogeneic and syngeneic splenocytes. It was observed during the study that the animals immunized with antigen‐pulsed mitomycin C treated allogeneic cells elicited antigen specific CD4+ Th1 cell response. Predominant release of IL‐2, interferon (IFN)‐γ and IgG2a‐isotype also occurred. In contrast, mice immunized with antigen‐pulsed syngeneic cells chiefly enhanced the production of interleukin (IL)‐4 and IgG1‐isotype. Further, allogeneic macrophages induced better T cell response than B cells or splenocytes and prominently induced the expression of B7‐1 and B7‐2. Immunization with antigen‐pulsed macrophages provided better recall responses compared to B cells. This was manifested by the high LFA‐1α and low CD45RB expression on T cells. Because it is already known that mitomycin C‐treated cells undergo apoptosis and dendritic cells engulf apoptotic cells, we therefore propose that generation of T cell response using antigen‐pulsed allogeneic cells may be due to the engulfment of these cells by dendritic cells, which may then process and present antigen entrapped in allogeneic cells to activate naive CD4+ T cells and differentiate them to Th1 cells. This study therefore provides a rational basis for manipulating antigen‐specific responses by immunizing with antigen‐pulsed allogeneic cells.

Spectrum of Gut Immunologic Reactions:Selective Induction of Distinct Responses to Vibrio cholerae WO7 and Its Toxin

Past studies with Vibrio cholerae have shown that cholera toxin (CT) is mainly responsible for inducing T helper type 2 (Th2) responses with systemic IgG1, IgE and mucosal secretory IgA (sIgA) antibodies. In this study, V. cholerae WO7, which produces novel toxin unrelated to CT, was given orally to mice in order to determine whether the strain V. cholerae WO7 differs from V. cholerae 569B, which produces CT, in the nature of responses generated at the gut and splenic level. The analysis of immune responses evoked by V. cholerae WO7 in the gut of mice revealed striking differences as compared to those elicited by V. cholerae 569B infection. To assess the T helper cell type responses, lymphocytes from Peyers patches and the spleen were stimulated in vitro for studying the cytokine patterns. PP and SP lymphoid cells from V. cholerae WO7 infected animals elaborated significant amounts of IL‐2, IFN‐γ and IL‐12 by 7 days p.i., suggesting a Th1 type of response. However by 15 days p.i., the PP and SP lymphoid cells secreted only IL‐6 and IL‐10 with traces of IFN‐γ. On the other hand, infection with V. cholerae 569B yielded mainly Th2 type responses at Peyers patches as well as the splenic level. Infection with both V. cholerae WO7 and 569B induced toxin‐specific IgA secreting cells at the gut and splenic level along with IgG1 secreting cells, indicating that both V. cholerae WO7 and 569B evoke an antigen‐specific Th2 type of response in the gut as well as spleen. The persistence of IgA along with Th1‐type cytokines indicates an alternate induction mechanism since mucosal IgA responses are usually associated with Th2‐type responses. These observations are suggestive of a common mechanism employed by the host to clear different strains of V. cholerae infection (569B and WO7 in this case), while the nature of toxins elaborated failed to modulate the net outcome of the infection caused by V. cholerae.

Modulation of the expression of M150 on macrophages by Th1/Th2 cytokines and co-stimulatory molecules CD40, B7-1, B7-2 and ICAM-1

M150 is an 150‐kDa protein associated with the surface of macrophages and is responsible chiefly for the activation of Th1 cells. It is a unique subset of the lysosome‐associated membrane protein‐1 glycoprotein and its co‐stimulatory activity depends on its post‐translational modification, which has a distinct glycosylation pattern restricted to macrophages. In the present study, we have observed that M150 is expressed constitutively on peritoneal but not splenic macrophages isolated from mice of different genetic backgrounds: Balb/c, C57BL/6 and C3He. However, M150 was expressed not only on peritoneal but also on splenic macrophages of non‐obese diabetic (NOD) mice. Expression on splenic macrophages was induced by culture with lipopolysaccharide (LPS). Expression could also be significantly up‐regulated by interferon (IFN)‐γ and granulocyte‐macrophage colony stimulating factor (GM‐CSF) but was inhibited by interleukin (IL)‐10; IL‐4 exhibited no effect. Further, cross‐linking of B7‐2, CD40, ICAM‐1 but not B7‐1 enhanced the level of M150 significantly. IFN‐γ and GM‐CSF acted synergistically with CD40. The significance of these findings is that cytokines IFN‐γ, GM‐CSF and IL‐10 and the co‐stimulatory molecules B7‐2, CD40 and ICAM‐1 can regulate the expression of M150 on macrophages.