The Department of Respiratory Allergy and Applied Immunology was started by (late) Dr. D.N. Shivpuri in early 1960's with the objective of undertaking research on diagnostic and therapeutic aspects of patients suffering with respiratory allergic disorders. At that time, identification of the offending allergens in patients of broncho-nasal allergy and their treatment by immunotherapy was carried out 'with imported stocks of allergens. the latter were considered unsatisfactory and the need to identify allergens in the actual environment of patients was urgently felt. Systematic studies were undertaken on the identification of clinically important local inhalant allergens: in Delhi atmosphere. Consequently, due to the pioneering work of the Department a large number of allergens of different groups; pollen, fungal, insect, algae, dust and dander were identified and were used for the diagnosis and immunotherapy of the patients with respiratory allergy. The results of immunotherapy with specific local allergens yielded encouraging results. Other faculty members associated with the Departmental activities include; Prof. Kunal Saha, Prof. M.P .S. Menon, Prof. M.K. Agarwal, (Late) Dr. Dharam Parkash and (Late) Dr. Kamal Arora.

Major Activities and Achievements

Research

1. Studies on pollen allergy in Delhi metropolitan area

Systematic and comprehensive studies on the identification of clinically important pollen aeroallergens were undertaken. These studies involved, (i) Field surveys of Delhi and adjoining areas. (ii) Aerobiological surveys to identify and enumerate common airborne pollen types to study their seasonal prevalence. (iii) Collection and processing of common pollen types. (iv) Preparation and storage of pollen extracts, and (v) Clinico- immunologic studies to evaluate their allergenic significance. A large number of pollen types were identified 'and quantified in the air and their seasonal patterns were recorded. More than 150 different pollen extracts were prepared and were evaluated for their allergenic significance in patients with allergic respiratory disorders. About 50 of them were. found to be clinically significant. Besides these common anemophillous pollen allergens, allergy to pollen of an entomophillous plant (papaya tree) was also reported.

2. Studies on the allergenic significance of plants parts Parthenium and its relationship to pollen allergens

Allergenic -potential of pollen and various parts of Parthenium hysterophorous was also studied: Pollen extract was found to be allergenic in 28% patients followed by flower (8%) and leaf extracts (6%) in that order. These studies suggested close relationship among pollen, leaf and flower extracts.

3. Atopic relationship amongst some species of chenopodiaceae and amarant- haceae

Antigens from pollen of the four species of chenopodiaceae (viz. Chenopodium album, C. murale, C. ambrosioides and a tetraploid of C. album) and two species of Amaranthaceae (viz. Amaranthus spinosus and A. hybridus) were skin tested on about 100 patients of nasobronchial allergy. The results indicated that positivity proneness of the patients varied from species to species; probably due to species in chenopodiaceae specific allergenic factors in this family.

4. Studies on allergenic fungi of Delhi metropolitan area

A comprehensive, systematic study of aerial fungal spores in Delhi area was undertaken. In early sixties the fungal spores were enumerated using gravimetric methods by simultaneous exposure of slides and petri-plates. Two peak periods of fungal spore concentration were reported, one from September to November and the other from February to April. A comparison of results of the two gravinetric techniques with those obtained with a volumetric sampler was made in late 1980's. Of the 150 fungal types encountered in Delhi air, 20 were show:1 to be allergenic and aetiologically related to respiratory allergic disorders. Several new fungal allergens including 12 species of Aspergillus have also been identified.

5. Identification of new fungal allergens

Volumetric aeromycological survey of Delhi was undertaken to identify and quantify yet unknown airborne allergenic fungi of Delhi area. With a view to evaluating their significance as inhalant allergens, intradermal tests with a number of new fungal extracts were performed on the patients with respiratory diseases. Results showed that Penicillium nigricans, P. chrysogenum, P. funiculosam, Aspergillus clavatus, Fusarium chamydosporium, Nigrospora sp., Syncephalastrum sp., Gliocladium sp., and Trichoderma viridae were allergenically significant. Inclusion of these yet unknown fungal allergens in the skin test kit will increase the scope of identification of causative allergens in the patients of allergic respiratory disorders.

6. Presence of shared as well as unique allergenic and antigenic components in different fungal species

Systematic studies were undertaken on the presence of shared or specific allergens or antigenic components among various genera, species and varieties of airborne fungi. .The results revealed that the allergenic and antigenic components among different fungal genera were largely specific and only a few of them were cross-reacting. Among various species of Aspergillus, a number of both shared as well as species specific components were observed. However, most of the allergenic and antigenic components among different varieties of P. funiculosum were found to be cross-reacting and only a few of them were variety specific.

7. Studies on insects

Aetiologic significance of about 35 insects, prevalent in Delhi atmosphere was evaluated and all of them were found to be important inhalant allergens. Elevated levels of insect allergen specific IgE antibodies in the sera of respiratory allergy patients by radioimmunoassay were found.

8. lmmunochemical quantification of airborne insects and other allergens

The etiologic significance of insects as producers of inhalant allergens causing type I respiratory allergic diseases was demonstrated by immunochemical quantification of airborne insect allergens for the first time in India, The insects studied were butterfly, moth, mosquito and housefly. These results have established conclusively that insects play an important role in type I hypersensitivity disorders of respiratory system.

On immunochemical quantification, seasonal varieties were noted in the amounts of airborne horse and dog dander allergens. Horse dander allergen varied from 0-16.5 ug/m3 of air, its peak periods being January to April and September to October. Airborne dog dander allergen ranged from 0-4.7 ug/m3 of air and was maximum during the months of January to March and September to December.

9. Demonstration of shared as well as unique allergenic and antigenic components in different insects genera and species and male/female cockroaches

Clinicoimmunologic and immunochemical analysis of the allergen extracts of male and female cockroaches (Periplaneta americana), different genera of cockroaches (P. americana and Blatella germanica) and mosquitoes (Anopheles stephensi, Culex quinquefasciatus and Aedes aegypti) revealed the presence of shared as well as specific allergenic and antigenic components in different sexes and genera of insects, These observations will have significant bearing on the immunotherapy of patients with respiratory allergy.

10. Etiologic significance of various dusts in respiratory allergy

Of the four dusts investigated, house dust and rice dust were found to be important as inhalant allergens, while the other two (cotton and paper dusts) were almost of no allergenic significance. All the dust related arthropods (housefly, rice weevil, male and female cockroaches and house dust mite) were also found to be important sources of inhalant allergens. However, these arthropods were not found to contribute significantly to the allergenicity of house dust and rice dust allergens.

11. Purification and characterization of significant allergenic components of clinically important inhalant allergens

Allergen extracts used for diagnosis and immunotherapy of patients with respiratory allergic disorders are crude, complex mixtures, each comprising a number of biochemical components of different shapes, sizes and properties. While some of them may be allergenic others are non-allergenic. A few of the allergenic components may be major while some others are minor contributors in the allergic activity of the crude extract. For efficient diagnosis and efficacious immunotherapy in patients, it is important that extracts of appropriate biological activity are used. However, techniques/ immunological reagents are not available for the quality control of various commercially available allergen extracts. The Department has been working on the purification, isolation and characterization of major allergenic components of different groups of allergens. Partial purification of the whole body extract of cockroach (male and female) suggested the presence of shared low molecular weight significant allergens and cockroach (male) specific high molecular weight minor allergens. In housefly, Fr. III (~66 ,KD) has been identified as one of the significant allergens.

12. Study on the time course of retention of allergenic activity of some important allergen extracts

Usually an allergen extract, when stored at 4^oC in aqueous form, is considered to retain its allergenic activity for a period of one year. However, loss of biological activity was observed in some extract within 9 months, while some other extracts showed retention of activity for 15 or more months, even when stored at room temperature. While extracts with reduced allergenic activity should not be used for diagnosis and therapy of patients, discarding the batches of latter extracts after one year storage, while they are still biologically active, will lead to financial loss. Thus, there is an urgent need of studying the shelf life of other clinically important extracts.

13. Comparison of biological activity in some clinically important, allergen extracts supplied by different manufacturers

The diagnostic efficiency and therapeutic efficacy of various allergen extracts used for skin tests and immunotherapy in patients with IgE mediated bronchial asthma and/or allergic rhinitis depend on the biological activity in the allergens extracts used. No quality control methods/reagents are available for standardization of allergen extracts in India. For this purpose, the present study has been undertaken to compare the biological activity in commercially available allergen extracts prepared by different manufacturers in India. Biological activity varied from allergen to allergen. The results emphasize the need for developing immunologic reagents and procedures for quality control of clinically important allergen extracts in our country.

14. New methods for enumeration of airborne (i) total (viable + non-viable), (ii) only viable, (iii) immuno-cheniically quantified fungal allergens

A new technique to quantify (i) total (viable + non viable) and (ii) (only viable) fungal species using the Burkard's Air Sampler was developed. Subsequently a new approach for simultaneous counting of air borne (i) total, (ii) only viable fungal spores and (iii) Immuno-chemically quantified fungal aeroallergens using membrane filtration technique was developed. The amount of allergen quantitated had a good correlation with the spore colony counts. House dust mites and their significance as aeroallergens. Mite fauna of house dust samples was studied. Various species of house dust mites (HDM) were identified and counted in the house dust samples of respiratory allergy patients in Delhi area. The results established that HDM is an important inhalant allergen in India.

15. Clinico-immunologic study on comparative evaluation of the sensitivity and specificity of common methods of diagnostic tests used in patients suffering with IgE mediated type I in allergic respiratory diseases

The sensitivity and specificity of various diagnostic methods to identify the offending allergens in patients of respiratory allergy was compared employing skin tests, bronchial provocation tests (BPT), conjunctival tests and radio- allergosorbent tests (RAST). The results showed that the skin tests performed and graded as per the method and criteria developed in the Department give reliable information and the technique is very sensitive and specific. Results of skin tests showed a good co-relation with the BPT and RAST.

16. Development of a rapid method for preparation of allergen extracts

A rapid method of preparation of allergen extracts was developed in which a mixture of chloroform and methanol was found to be a better defatting agent than conventionally used diethyl ether, and two hours period of extraction in the buffered saline was sufficient for optimal allergen recovery.

17. Studies on the causative allergens in patients

Several studies were undertaken to evaluate the etiologic significance of offending allergens in patients of bronchial asthma and allergic rhinitis using a number of allergens of different groups i.e. pollen, fungal, dust, animal dander and insect extracts. Results showed that all these groups played an important role in the etiology of allergic disorders in patients. The causative allergens varied from patient to patient and in different permutation and combination to various allergens of different groups.

18. Investigation on the role of immunotherapy in patients suffering with nasobronchial allergy

Several studies on the therapeutic efficacy of immunotherapy in patients with nasobronchial allergy were undertaken. The best results were obtained in the patients suffering with seasonal nasobronchial allergy. It was observed that immunotherapy administered in properly selected patients gives encouraging results.

19. Studies on bronchial tolerance to histamine, methacholine chloride and specific antigen inhalation

Increase in bronchial tolerance to histamine, methacholine chloride and specific allergens inhalation challenge was observed after treatment with Tylophora indica leaves, its alcoholic extract or its alkaloids. Similar therapeutic response was observed in patients after ingestion of whole plant powder of Solanum xaenthocarpum whole plant.

20. Effect of prior medication on intradermal test response in patients with respiratory allergies

Our studies revealed that certain drugs used for symptomatic treatment of asthma, such as salbutamol, theophyline, adrenaline, etc., were found not to interfere with the results of intradermal tests in patients with type I respiratory allergy. Thus, skin testing of patients need not be postponed due to their inability to discontinue these drugs. However, use of long acting antihistamine-Astemizole resulted in abrogation of interdermal test response to allergens up to four weeks. Thus, it is recommended that prior to performance of skin tests such anti- histamines should be discontinued for a suitable period.

21. Studies on the role of animal danders in the etiology of respiratory allergic disorders

Evaluation of seven animal danders as inhalant allergens was undertaken and relative allergenic significance was studied. The presence of shared and specific allergenic components in different animal dander was found.

22. Influenza virus infection and levels of total allergen specific and virus specific IgE in patients with respiratory allergic disorders

The role of influenza virus in the etiology of type I respiratory allergic disorders was studied. The results suggested that respiratory viruses may precipitate symptoms of asthma either by inducing an increase in the total, or allergen-specific IgE (non-virus) and/or virus-specific IgE antibodies.

23. Role of food allergens in the etiology of nasobronchial allergy

The role of 25 food allergen in etiology of respiratory allergy was studied. A sizable percentage of patients showed marked skin positivity to one or more food allergens suggesting their etiologic significance in the causation of nasobronchial allergy. Our results suggested that a markedly positive skin reaction to food allergens should not be ignored which may be confirmed by performing bronchial provocation test and/or RAST.

24. Heterogeneity of IgE mediated immune response to insects in patients with naso-bronchial allergic disorders

A total number of 346 patients of bronchial asthma and/or allergic rhinitis were tested with 17 insect extracts. Sera of 200 of these patients were studied in various insect RASTs for the presence of allergen-specific IgE antibodies. The patients showed skin reactivity and RAST positivity to more than one of the 17 insect extract in various permutations and combinations. These results crossed- RAST -inhibition and absorption studies showed heterogeneity of immune response of patients to these insect allergens.

25. Comparative efficacy of 2 RHZE/4RH (4D) regime Vs 2 RHZ /4 RH (3D) regimen in Category I sputum positive pulmonary tuberculosis(R: Rifampicin; H: Isoniazid; Z: Pyrazinamide; E: Ethambutol).

26. Long term effects of beclomethasone inhalation (MDI) in bronchial asthma: Two year evaluation involving 100 patients was done.

27. Initial drug resistance in pulmonary tuberculosis and- efficacy of 6 months short course chemotherapy in such patients.

28. The incidence of sexually transmitted diseases (STD) as well as acquired immunodeficiency syndrome (AIDS) in patients of leprosy was studied. In spite of high prevalence of STD in leprosy patients, no case of AIDS could be detected.

29. Studies involving T cell homeostasis in tuberculosis patients at the start and following three months of chemotherapy revealed that there was a drastic alteration in T cell subsets in these patients which reverted to normal after three months of successful chemotherapy.

30. The T4/T8 homeostasis was also altered in patients with various stages of breast carcinoma and benign breast diseases.

31. Lymphocytes with HLA-DR II and IL-2 receptor markers were also reduced in patients with breast carcinoma. The blood levels of IL-2 of carcinoma pa1ients were comparable to that of normal controls although their blood levels of sIL-2R were increased.

32. The nutritional deficiency in leprosy patients was found to be poverty related and not disease related. The serum Zn, vitamin A and E levels were associated with disease.

33. Dapsone and Clofazimine, the important constituents of multi-drug therapy (MDT) were found to alter the early components of the complement pathway, in- vitro, only at a very high concentration.

34. Clofazimine alone enhanced O₂ production of rat resident peritoneal macrophages in a dose-dependent manner. The drug lost. this property after exposure to light. Detailed physico-chemical properties (i.e. UV , IR, NMR and mass spectra) of light reacted Clofazimine were also studied.

35. Rats treated with MDT showed altered T cell homeostasis, while there was no effect on CH-50, C3, total T cells (W3/13), B cells (OX-4) or immunoglobulin levels.

36. The haemagglutinins isolated from Indian fresh water catfish were shown to be specific towards L-galactopyranose. This activity was radioresistant suggesting that the haemagglutinin could be a lectin.

37. The purified haemagglutinin from Indian fresh water catfish had native molecules of molecular weights of 187 KD and 55 KD. The ultrastructure of immune apparatus of this catfish was also studied.

Achievements

1. For the first time in India, systematic comprehensive studies on the etiologic significance of pollen, insects, fungi, algae and animal dander, etc., in respiratory allergic disorders have been undertaken which involved the development of various modern in vitro techniques to study their allergenic and antigenic properties. These techniques include Radio-allergosorbent test (RAST), Rocket immunoelectrophoresis, Crossed immunoelectrophoresis (CIE), Crossed-line immunoelectrophoresis (CLIE), Sodium dodecyl sulphate -Polyacrylamide gel electrophoresis (SDS-PAGE), Thin layer isoelectric-focussing (TLIEF), etc. In addition, the immunochemical quantitation of airborne insect-derived allergen particles has been carried out for the first time in our country.

2. Development of an easy method of inhalation provocation test to detect the cause of allergic asthma which has been referred to, in standard text book of Respiratory Medicine by Crofton.

3. Discovery of three radiological signs which made the diagnosis of allergic-bronchopulmonary aspergillosis simple. These signs are well accepted in India and abroad.

4. Establishment of monoclonal antibody staining techniques for the study of immunology of tuberculosis and leprosy.

5. Some of. the studies have helped in understanding the alterations of immune system (i) in diseases like tuberculosis, leprosy and breast cancer and (ii) on treatment with drugs.

6. Study on phylogeny of immune response revealed that as one goes higher in the evolutionary tree, the classical complement pathway becomes more active as compared to alternative complement pathway.

Patients Care

The Department has been the pioneer centre for research and teaching in the identification of local allergens and management of respiratory allergic disorders by administering immunotherapy. The skin test facility provided by the Department for a large number of patients has been helpful in proper diagnosis and efficient management of respiratory allergy diseases.

Transfer of Technology

Identification of local allergens lead to proper diagnosis and treatment of the patients with respiratory allergy. The results of immunotherapy with specific local allergens were encouraging. This created an exceedingly heavy demand on the Department for supply of these allergens from all over the country. Until 1969, the Department tried to cope with the ever increasing demand but the patients had to wait for several months for investigations and treatment. The Department at that time transferred the technology of allergen preparation to the CSIR, Centre for Biochemicals, a Govt. of India Undertaking to make them available on commercial scale for the use on patients.

Teaching/Training Programmes

Workshops on allergy, allergens and immune response

In 1975, the Department started conducting biannual Workshops on Allergy, Allergens and Immunotherapy in order to impart training, in clinical and laboratory aspects of diagnosis and management of allergic respiratory diseases to medical practitioners and faculty members of hospitals/research institutions working in various parts of India. From 1981 onwards, the Institute 'has been conducting these workshops in collaboration with the Centre for Biochemicals Technology (CSIR). Faculty members and scientists from the V .P .Chest Institute, the Centre for Biochemicals Technology and other medical institutions from Delhi and other parts of India participate in the teaching programme. About 50 medical graduates are trained annually in these workshops. The latest workshop in this series on "Respiratory Allergy : Diagnosis and Management" was held on February 3-8, 1999.