Polymorphism in ADAM33 gene associated with asthmatics in West Bengal, India - An investigation by in-silico analysis.

Introduction: Asthma is one of the common chronic polygenic inflammatory diseases. Genome wide association studies have identified ADAM33 as an asthma candidate gene. The present study investigated possible association of rs2280090 (T1), rs2280091 (T2) and rs3918396 (S1) single nucleotide polymorphisms (SNPs) of ADAM33 with aeroallergen induced asthma in West Bengal population, India. In addition, in-silico analysis was performed to find out changes in protein function. Methods: Forced expiratory volume in 1 second (FEV1)/Forced vital capacity (FVC), peak expiratory flow rate (PEFR) were assessed using spirometry in 1039 participants. Allergic sensitivity of 619 spirometry positive asthma patients was assessed by skin prick test (SPT) against 22 aeroallergens. For genotyping of T1, T2, and S1 SNPs in 540 allergic asthma patient and 420 control subjects, polymerase chain reaction-based restriction fragment length polymorphism was performed. Total Immunoglobulin-E (IgE) level was measured in both patients and controls. ADAM333 haplotype blocks were constructed using Haploview software v.4.2. Structural model of transmembrane and cytoplasmic domains of ADAM33 was generated using RaptorX. Protein-protein interaction was analysed using the STRING server. Results: Highest number of patient sensitivity was observed towards Cocos nusifera (n = 215) and Dermatophagoides farinae (n = 229). Significant difference in sensitivity was observed between child and late adult (P = 0.03), child and early adult (P = 0.02), adolescent and late adult (P = 0.02) and adolescent and early adult (P = 0.01). Genotypic frequencies differed significantly between patients and controls (P < 0.05). rs2280090 GG, rs2280091GG and AG genotype, and rs3918396 AA carried significant risk for asthma (P = 0.02, P = 0.008, P = 0.04, P = 0.01 respectively). ADAM33 T1, T2, and S1 polymorphisms were in high Linkage Disequilibrium (D = 0.98). Haplotype consisting of rs2280090G, rs2280091G and rs3918396A alleles were found significantly higher in patient population in comparison with controls (OR = 2.03). IgE level differed significantly among different genotypes for T1, T2, and S1 SNPs analysed in pair (P < 0.0001). FEV1/FVC ratio differed significantly among different genotypes for T1, T2 and S1 SNPs analysed in pair (P < 0.0001). Significant difference of FEV1/FVC was also found between GGA and AAG haplotype (P < 0.0001). In-silico analysis revealed T1 and T2 polymorphisms are located in cytoplasmic domain of ADAM33 may cause bronchial smooth muscle cell mobility and cellular hyperplasia as well as cytoskeletal remodelling by altered interaction with different cytoplasmic proteins found by string analysis. Conclusion: Present study showed significant association of T1, T2, and S1 polymorphisms of ADAM33 with aeroallergen-induced asthma in West Bengal, India. These polymorphisms may be used as prognostic markers and possible targets for therapeutics in future.

In silico analysis of single nucleotide polymorphism (rs34377097) of TBXA2R gene and pollen induced bronchial asthma susceptibility in West Bengal population, India.

Introduction: Prevalence of asthma is increasing steadily among general population in developing countries over past two decades. One of the causative agents of broncho-constriction in asthma is thromboxane A2 receptor (TBXA2R). However few studies of TBXA2R polymorphism were performed so far. The present study aimed to assess potential association of TBXA2R rs34377097 polymorphism causing missense substitution of Arginine to Leucine (R60L) among 482 patients diagnosed with pollen-induced asthma and 122 control participants from West Bengal, India. Also we performed in-silico analysis of mutated TBXA2R protein (R60L) using homology modeling. Methods: Clinical parameters like Forced expiratory volume in 1 second (FEV1), FEV1/Forced vital capacity (FVC) and Peak expiratory flow rate (PEFR) were assessed using spirometry. Patients' sensitivity was measured by skin prick test (SPT) against 16 pollen allergens. Polymerase chain reaction-based Restriction fragment length polymorphism was done for genotyping. Structural model of wild type and homology model of polymorphic TBXA2R was generated using AlphaFold2 and MODELLER respectively. Electrostatic surface potential was calculated using APBS plugin in PyMol. Results: Genotype frequencies differed significantly between the study groups (P=0.03). There was no significant deviation from Hardy-Weinberg equilibrium in control population (χ2=1.56). Asthmatic patients have significantly higher frequency of rs34377097TT genotype than control subjects (P=0.03). SPT of patients showed maximum sensitivity in A. indica (87.68%) followed by C. nusifera (83.29%) and C. pulcherima (74.94%). Significant difference existed for pollen sensitivity in adolescent and young adult (P=0.01) and between young and old adult (P=0.0003). Significant negative correlation was found between FEV1/FVC ratio and intensity of SPT reactions (P<0.0001). Significant association of FEV1, FEV1/FVC and PEFR was observed with pollen-induced asthma. Furthermore, risk allele T was found to be clinically correlated with lower FEV1/FVC ratio (P=0.015) in patients. Our data showed R60L polymorphism, which was conserved across mammals, significantly reduced positive electrostatic charge of polymorphic protein in cytoplasmic domain thus altered downstream pathway and induced asthma response. Discussion: The present in-silico study is the first one to report association of TBXA2R rs34377097 polymorphism in an Indian population. It may be used as prognostic marker of clinical response to asthma in West Bengal and possible target of therapeutics in future.

A review on aero-allergen induced allergy in India.

Allergic diseases such as bronchial asthma, rhinitis and eczema are an important public health issue globally in 21st century. Their prevalence and impact in India is high, in terms of total numbers of people affected. Due to diverse climate and lifestyle, the Indian population is exposed to a rich and varied range of aerobiological particles. In the last two decades in India, 83 pollens, 34 fungi, six dust mite and 19 insect species were reported as allergenic, often contributing to heavy allergenic exposure in indoor and outdoor environments. There are currently few practitioners who offer allergy diagnosis and treatment in India, but there is active basic and molecular research aiming to improve the health of allergic patients. This review explores the field of aero-allergy in India during the last two decades, with special emphasis on prevalence across various centres and relevant scientific advances such as genetic associations and new therapeutics. This review will be helpful for clinicians to select aero-allergens for sensitisation testing in patients suffering from symptoms in different regions of India, and to support patients undertaking prophylaxis of allergic symptoms in India according to local environmental conditions. This study also helps to define patient selection for allergy testing and the role of test results in guiding allergic disease management in the Indian context. Finally, we summarize genetic findings relevant to aero-allergy in India and discuss the development of novel therapeutics for investigation and management of aero-allergy in India.

Assessment of Co-Sensitization between Pollen and Food Allergen Sources among Bengali Population, West Bengal, India.

INTRODUCTION: Secondary food allergies due to cross-reactivity between pollen and food allergens are a significant health burden in India. Prevalence of cross-reactivity varies by region, and particularly in West Bengal, it remains unexplored. The present study investigated sensitivities of pollen-related food allergy (PFA) patients of West Bengal by empirically determining co-sensitization patterns. METHODS: The present study retrospectively analysed 1,310 pollen-allergic (PA) patients from the Allergy and Asthma Research Centre, West Bengal. A questionnaire survey was performed to record patients' demographic and clinical features and categorized into seasonal and perennial ones. All patients were subjected to the skin prick test and specific IgE for evaluation of sensitivity against 16 pollen and 16 food allergens. Co-relation and principal component analysis (PCA) was performed to determine co-sensitivity. RESULTS: Seasonal allergic patients were comparatively higher than perennial ones. PFA patients were mostly sensitive to eggplant (n = 387), while Azadirachta indica (n = 341) was predominant sensitizer among pollen allergen sources. Cynodon dactylon showed maximum significant positive correlation with banana, orange, peanut, pineapple, cucumber, eggplant, potato, tomato, and wheat. In the seasonal group, Cynodon dactylon-specific IgE in PFA patients (median = 4.60 kU/L) was significantly higher from that of PA ones (median = 3.44 kU/L). Among 32 allergens tested, 27 were placed in first two dimensions of the PCA biplot. CONCLUSION: The present study established that co-sensitivity between pollen and food allergen sources were predominant in allergic patients of West Bengal. The co-sensitization patterns among unrelated allergen sources may be due to possible expression of biologic cross-reactivity to similar allergens.

Assessment of egg and milk allergies among Indians by revalidating a food allergy predictive model.

Background: The recent upsurge in food allergy indicates the need for accurate medical diagnostics. The application of predictive diagnostic models can envisage the outcome of oral food challenge (OFC), reducing cost and time. A logistic regression model was developed by DunnGalvin for children predicting OFC outcome using six predictors viz: sex, age, history, specific IgE, total IgE minus specific IgE, and skin prick test. This model was later updated by Klemans, reducing the number of predictors enhancing the calibration and discrimination of outcome. Objective: Our aim was to revalidate both the models for assessment of egg and milk allergies among Indians in the age group 0-19 years and to determine regression coefficients for our study population. Methods: Revalidation was done at the allergy clinic using OFC outcomes of egg and milk allergic patients. Precise values of the predictors were set up for which calibration (predicted against observed outcome) and discrimination (area under curve [AUC] of receiver operator characteristic curve [ROC]) would be better. Results: The Klemans model with reduced number of predictors showed better accuracy, calibration and discrimination than the DunnGalvin. Best calibration for egg allergy was achieved in the Klemans model with correlation coefficient (r2) of 0.90 and accuracy of 97%. The AUC of ROC was 0.90. For milk allergy, the coefficient was 0.94 with accuracy of 98%. The AUC was 0.91. Conclusion: The present study showed that mathematical models are non-invasive and can be successfully used as appropriate alternative to OFC in Indian population after proper validation.

Association of HLA-DQ and IL13 gene variants with challenge-proven shrimp allergy in West Bengal, India.

Little is known about genetic factors and mechanisms underlying shrimp allergy. Genome-wide association studies identified HLA class-II and IL13 genes as highly plausible candidates for shrimp allergy. The present study was designed to investigate potential associations of HLA-DQ rs9275596, IL13 rs20541, and IL13 rs1800925 polymorphisms with challenge-proven shrimp allergy using the data from 532 people of West Bengal, India; selected on basis of positive skin prick test, elevated specific IgE and medical history. Risk genotypes, i.e., HLA-DQ rs9275596 CC, IL13 rs20541 AA, and IL13 rs1800925 TT, were found to be significantly associated with challenge positive shrimp allergy (P = 0.04, 0.01, and 0.03, respectively). Distribution of genotypes for HLA-DQ and IL13 polymorphisms in allergic and control subjects showed significant difference between younger (20-40 years) and older (> 40 years) age group (P = 0.006). Risk genotypes significantly associated with elevated shrimp-specific IgE. IL13 TA haplotype significantly associated with shrimp allergy and elevated specific IgE (P = 0.02). Synergistic effect of IL13 TA haplotype-HLA-DQ rs9275596 CC genotype interaction significantly elevated specific IgE (P = 0.03). The present study suggests that HLA-DQ and IL13 polymorphisms pose major risk for shrimp allergic patients in West Bengal, India and thus could be helpful for early target-specific therapeutic intervention in near future.

Biocompatible implant mimicking cartilage: A new horizon for reconstructive facial field.

Cartilage is avascular with limited to no regenerative capacity, so its loss could be a challenge for reconstructive surgery. Current treatment options for damaged cartilage are also limited. In this aspect there is a tremendous need to develop an ideal cartilage-mimicking biomaterial that could repair maxillofacial defects. Considering this fact in this study we have prepared twelve silicone-based materials (using Silicone 40, 60, and 80) reinforced with hydroxyapatite, tri-calcium phosphate, and titanium dioxide which itself has proven their efficacy in several studies and able to complement the shortcomings of using silicones. Among the mechanical properties (Young's modulus, tensile strength, percent elongation, and hardness), hardness of Silicone-40 showed similarities with goat ear (P > .05). Silicone peaks have been detected in FTIR. Both AFM morphology and SEM images of the samples confirmed more roughed surfaces. All the materials were nonhemolytic in hemocompatibility tests, but among the twelve materials S2, S3, S5, and S6 showed the least hemolysis. For all tested bacterial strains, adherence was lower on each material than that grown on the plain industrial silicone material which was used as a positive control. S2, S3, S5, and S6 samples were selected as the best based on mechanical characterizations, surface characterizations, in vitro hemocompatibility tests and bacterial adherence activity. So, outcomes of this present study would be promising when developing ideal cartilage-mimicking biocomposites and their emerging applications to treat maxillofacial defects due to cartilage damage.

Assessment of Hymenoptera and Non-Hymenoptera Insect Bite and Sting Allergy Among Patients of Tropical Region of West Bengal, India.

West Bengal, India, is inhabited by abundance and variety of insects that triggers sensitization in some humans to inhalant allergens and/or insect stings/venoms. Lack of research on this topic prevented accurate diagnosis and proper follow-up treatments to patients suffering from insect-induced allergies. The aim of our study was to identify the allergy-causing insects and evaluate resulting sensitization among a study population in West Bengal, India. The skin prick test (SPT) evaluated sensitivity of 450 patients who sought treatment at the Allergy and Asthma Research Center from July 2017 to June 2018. Eight insect allergens were tested: Common Black Ant (Lasius niger, Linnaeus 1758), Fire Ant (Solenopsis invicta, Buren 1972), Honey Bee (Apis cerana indica, Fabricius 1798), Common Wasp (Vespula vulgaris, Linnaeus 1758), Mosquito (Aedes aegypti, Linnaeus in Hasselquist 1762), American Cockroach (Periplaneta americana, Linnaeus 1758), House Fly (Musca domestica, Linnaeus 1758), and Grasshopper (Gesonula punctifrons, Stal 1861). From a total of 450 patients evaluated, 370 patients had positive SPT reaction from at least one of the 8 insect allergens tested. Sensitivity to some Hymenoptera insects (common black ant, 87.62%; fire ant, 84.59%; and honey bee, 67.02%) was found in higher proportion than non-Hymenoptera group (mosquito, 66.67%; American cockroach, 33.33%; house fly, 10.41%; and grasshopper, 5.14%). There was significant difference in sensitivity among child, adolescent, and adult (P < 0.001). While female patients showed more sensitivity than males to SPT, the difference was statistically insignificant. In regards to occupation, farmers and bee keepers were most sensitive of field workers sensitive to Hymenoptera-derived allergens.

Phytochemical analysis of mite-infested tea leaves of Darjeeling Hills, India.

INTRODUCTION: Darjeeling tea of India is one of the most famous beverages globally. However, a large amount of tea crop is damaged every year by the attack of mites. OBJECTIVES: The study aimed to determine the changes in different pigments and biochemical components of tea leaves due to mite infestation. MATERIALS AND METHODS: We used UV-visible and Fourier-transform infrared (FTIR) spectroscopy simultaneously to understand the damages in pigment contents of the leaves of tea (Camellia sinensis (L.) Kuntze) due to attack of phytophagus mite, Oligonychus coffeae Nietner. Furthermore, chemical analysis of infested tea leaves was also performed to compare the nutrients of the plants, namely total phenol, protein and soluble sugar. RESULTS: The UV-visible study reveals severe reduction of the pigments such as chlorophyll (Chl), carotenoids and xanthophylls in the tea leaf due to mite infestation. The findings of the FTIR study, also shows variation in different physiochemical components in the leaf Chl. The sugar and protein content of the infested leaves have been reduced compared to uninfested ones. Results in the case of tea leaves plucked during first (March) and third (November) flushes show similar trends. CONCLUSION: The increase in phenolic and alcoholic components and decrease in Chl contents may affect the quality of Darjeeling tea. The extent of damages done by the pests measured here could aid the pest management in tea gardens.

Association of the STAT6 rs3024974 (C/T) Polymorphism with IgE-Mediated Food Sensitization among West Bengal Population in India.

INTRODUCTION: Genetic polymorphisms associated with IgE-mediated food sensitization have been a robust area of research for decades. A genome-wide search for susceptible loci regulating the IgE response (atopy) identified the candidate gene STAT6, which is important in the context of food allergic manifestations. OBJECTIVE: The present study was designed to investigate the sensitization of West Bengal population against some common allergenic food items and to study the role of the STAT6 gene polymorphism in elevating food-specific IgE levels among sensitized individuals. METHODS: Skin prick test was performed for 6 food items among 501 patients (126 children, 85 adolescents, and 290 adults)from West Bengal, India. Among them, 165 patients were selected for measurement of total IgE and food-specific IgE levels along with 165 controls. Finally, the STAT6 (rs3024974 (C/T) polymorphism was genotyped in 139 cases and control subjects. RESULTS: Shrimp was identified as a dominant food allergen in adolescents and adults, whereas milk sensitization was highest in children. Food-sensitized patients with onset during childhood had significantly higher total IgE levels compared to patients with onset during adulthood (p < 0.00001). The frequency of the rs3024974 CC genotype in both cases and control subjects (55.40 and 46.76%, respectively) was higher than that of CT or TT. Patients with childhood onset bearing the CC genotype had significantly higher specific IgE levels in comparison to those with adult onset (p = 0.001). CONCLUSION: Food sensitization has a genetic background and the rs3024974 polymorphism is associated with susceptibility and reaction severity in food-sensitized patients in West Bengal population in India.

Evaluation of Sensitivity Toward Storage Mites and House Dust Mites Among Nasobronchial Allergic Patients of Kolkata, India.

House dust mites (HDMs) are the major constituents of house dust (HD). HD and HDM sensitization is well documented worldwide. Storage mite (SM) sensitization is presently lacking from India. The present study evaluated the sensitization of both HDM and SM among 372 allergic rhinitis patients reported to the Allergy and Asthma Research Center of Kolkata metropolitan, India. HD samples were collected from the patients' home and analyzed for the major constituent mites. HD and six constituent mites Dermatophagoides pteronyssinus (DP), Dermatophagoides farina Hughes (Acari: Pyroglyphidae) (DF), Blomia tropicalis (BT), Acarus siro Linnaeus (Acari: Acaridae) (AS), Lepidoglyphus destructor (LD), and Tyrophagus putrescentiae (Schrank) (Acari: Acaridae) (TP) are tested for the allergenic potential through Skin Prick Test (SPT). Three SMs, namely AS, LD, and TP, were newly included in the mite SPT extract for the first time in Kolkata. In total, 330 patients showed significant positive SPT toward any one allergen tested. HD was the major elicitor exhibiting 92.42% response. Individuals of age group 15-40 were the worst sufferers. DF showed the highest sensitization (87.87%) among the dust mites. The SMs also contributed significantly to prove their sensitizing potential. SPT rates for AS, LD, and TP were 33, 25, and 18%, respectively. SPT grades and total Immunoglobulin E (IgE) were positively correlated for each of the allergens. Most of the patients were multi-sensitized (95%) and represented markedly high total IgE levels (>500 IU/ml). Three SMs proved to be significant allergens for the studied population. The sensitization toward these SMs is first time reported from India and can be recommended for inclusion of routine SPT for better outcome in the future.

Sensitization to Common Aeroallergens in the Atopic Population of West Bengal, India: An Investigation by Skin Prick Test.

BACKGROUND: Aeroallergen sensitization is increasing worldwide. The optimal management of allergic disease requires the identification of the allergic sensitivities of the patient. Complete data of offending aeroallergens are scant from West Bengal, India. Hence, we employed a retrospective analysis of an atopic population of West Bengal to investigate the rate of sensitivity toward common aeroallergens including pollens, molds and house dust mites. OBJECTIVES: The aim of the present study was to evaluate the profile of skin prick test (SPT) reactivity of an atopic population of eastern India. METHODS: The prevalence aeroallergen sensitization was investigated by SPT along with a questionnaire survey of patients who visited the Allergy and Asthma Research Center of West Bengal, India, from June 2016 to May 2018. Patients were categorized according to the demographic (age, gender) and clinical aspects. Statistical tests were applied to differentiate between age and gender of the sufferers. RESULTS: 524 out of 605 patients exhibited a positive SPT response toward one or more aeroallergens tested. One-way ANOVA following Tukey's post hoc test reveals the age group of 15-40 years was more sensitized than that of < 15 years and no male-female discrimination was found using the paired t test. House dust mites, namely Dermatophagoides pteronyssinus and Dermatophagoides farinae, elicited most significant SPT results (80.34 and 84.92%, respectively) followed by Cocos (73.28%) and Azaridacta (57.25%) pollen. The most sensitive mold was Alternaria alternata (26.9%). Allergic rhinitis, bronchial asthma and atopic dermatitis were recorded as the common symptoms. Other influencing factors were family history, nature and season/time of onset and house conditions. CONCLUSION: Following the increasing trend, house dust mite sensitization still held the dominance like in any other Indian population reported earlier. This study would pave the way to construct a standard and minimal SPT panel for the atopic patients of eastern India.

Sensitivity to House Dust Mites Allergens with Atopic Asthma and Its Relationship with CD14 C(-159T) Polymorphism in Patients of West Bengal, India.

India is the home to around 15-20 million asthmatics, and asthma prevalence is increasing in Indian metropolitan area, including Kolkata, West Bengal. Complex interactions of genetic and environmental factors are involved in asthma. Genome-wide search for susceptible loci regulating IgE response (atopy) have identified a candidate gene CD14 which is most important in the context of allergic responses of respiratory system. This study was aimed to investigate the role of house dust and house dust mites in development of bronchial asthma and to explore the possible association of candidate gene CD14 with disease manifestation among Kolkata patient population. Skin-prick test was done among 950 asthmatic patients against 8 aeroallergens, including house dust and house dust mites and total serum IgE and allergen-specific IgE were measured. Polymerase chain reaction-restriction fragment length polymorphism was done in patients and nonasthmatic control (n = 255 in each) to characterize a functional polymorphism, C(-159)T, of CD14, a positional candidate gene for allergy. We identified house dust as the most common aeroallergen sensitizer among atopic patients in Kolkata followed by Dermatophagoides pteronyssinus and Dermatophagoides farinae Hughes (Acari: Pyroglyphidae) mites. Patient's sera contain significantly higher IgE level than that of control. Allergen-specific IgE antibody test revealed that 76.36% patients had specific IgE antibody against D. pteronyssinus mite. There was a significant difference in the distribution of alleles and genotypes for CD14 polymorphism with an increase in disease severity. So, in Kolkata, house dust mite is a common aeroallergen and D. pteronyssinus is predominant among mites. The present study revealed that bronchial asthma has a genetic background.