Comprehensive characterization and molecular insights into the salt tolerance of a Cu, Zn-superoxide dismutase from an Indian Mangrove, Avicennia marina.

Superoxide dismutases are important group of antioxidant metallozyme and play important role in ROS homeostasis in salinity stress. The present study reports the biochemical properties of a salt-tolerant Cu, Zn-superoxide from Avicennia marina (Am_SOD). Am_SOD was purified from the leaf and identified by mass-spectrometry. Recombinant Am_SOD cDNA was bacterially expressed as a homodimeric protein. Enzyme kinetics revealed a high substrate affinity and specific activity of Am_SOD as compared to many earlier reported SODs. An electronic transition in 360-400 nm spectra of Am_SOD is indicative of Cu2+-binding. Am_SOD activity was potentially inhibited by diethyldithiocarbamate and H2O2, a characteristic of Cu, Zn-SOD. Am_SOD exhibited conformational and functional stability at high NaCl concentration as well in alkaline pH. Introgression of Am_SOD in E. coli conferred tolerance to oxidative stress under highly saline condition. Am_SOD was moderately thermostable and retained functional activity at ~ 60 °C. In-silico analyses revealed 5 solvent-accessible N-terminal residues of Am_SOD that were less hydrophobic than those at similar positions of non-halophilic SODs. Substituting these 5 residues with non-halophilic counterparts resulted in > 50% reduction in salt-tolerance of Am_SOD. This indicates a cumulative role of these residues in maintaining low surface hydrophobicity of Am_SOD and consequently high salt tolerance. The molecular information on antioxidant activity and salt-tolerance of Am_SOD may have potential application in biotechnology research. To our knowledge, this is the first report on salt-tolerant SOD from mangrove.

Differentiating Wild and Apiary Honey by Elemental Profiling: a Case Study from Mangroves of Indian Sundarban.

Honey is a natural substance produced by honeybees from the nectar or secretion of flowering plants. Along with the botanical and geographical origin, several environmental factors also play a major role in determining the characteristics of honey. The aim of this study is to determine and compare the elemental concentration of various macro and trace elements in apiary and wild honeys collected from different parts of Indian Sundarbans. The elemental analysis was performed in inductively coupled plasma optical emission spectroscopy preceded by microwave digestion method. The concentrations of 19 elements (Ag, Al, As, B, Ca, Cd, Co, Cr, Cu, Fe, K, Li, Mg, Mn, Na, Ni, Pb, Se and Zn) were investigated from thirteen locations of Indian Sundarbans. This comparative study shows in wild honey samples, the concentration of K was highest followed by Ca, Mg and Na and Zn was lowest among all. In contrast, in apiary honey samples, Ca had maximum concentration followed by K, Mg and Na and Ag had minimum among all. The elemental concentration in honey from apiary was either equal or higher than their wild counterpart. The results of the factor analysis of PCA algorithm for wild and apiary honey samples were highly variable which implies that the elements are not coming from the same origin. The concentration of element was found to be highly variable across sites and across sources of honey samples.

Molecular characterization of a fungal cyclophilin allergen Rhi o 2 and elucidation of antigenic determinants responsible for IgE-cross-reactivity.

Cyclophilins are structurally conserved pan-allergens showing extensive cross-reactivity. So far, no precise information on cross-reactive IgE-epitopes of cyclophilins is available. Here, an 18-kDa IgE-reactive cyclophilin (Rhi o 2) was purified from Rhizopus oryzae, an indoor mold causing allergic sensitization. Based on LC-MS/MS-derived sequences of natural Rhi o 2, the full-length cDNA was cloned, and expressed as recombinant (r) allergen. Purified rRhi o 2 displayed IgE-reactivity and basophil degranulation with sera from all cyclophilin-positive patients. The melting curve of properly folded rRhi o 2 showed partial refolding after heat denaturation. The allergen displayed monomeric functional peptidyl-prolyl cis-trans isomerase (PPIase) activity. In IgE-inhibition assays, rRhi o 2 exhibited extensive cross-reactivity with various other cyclophilins reported as allergens from diverse sources including its homologous human autoantigen. By generating a series of deletion mutants, a conserved 69-residue (Asn81-Asn149) fragment at C terminus of Rhi o 2 was identified as crucial for IgE-recognition and cross-reactivity. Grafting of the Asn81-Asn149 fragment within the primary structure of yeast cyclophilin CPR1 by replacing its homologous sequence resulted in a hybrid molecule with structural folds similar to Rhi o 2. The IgE-reactivity and allergenic activity of the hybrid cyclophilin were greater than that of CPR1. Therefore, the Asn81-Asn149 fragment can be considered as the site of IgE recognition of Rhi o 2. Hence, Rhi o 2 serves as a candidate antigen for the molecular diagnosis of mold allergy, and determination of a major cross-reactive IgE-epitope has clinical potential for the design of next-generation immunotherapeutics against cyclophilin-induced allergies.

Identification of airborne pollen allergens from two avenue trees of India.

An attempt has been made to detect airborne pollen of Lagerstroemia speciosa (LS) and Spathodea campanulata (SC) - two common avenue trees of India as potential sources of aeroallergens and also to identify the major IgE-reactive components present in them. The airborne pollen concentration was assessed using a Burkard sampler. A detailed questionnaire on clinical data of 1490 patients was recorded based on hospital data. We assessed the allergenicity of pollen by in vivo and in vitro tests. The correlation among meteorological factors, pollen seasons and allergenic potency of patients was assessed by multiple regression analysis. The sensitivity of patients to pollen antigens was highly correlated with pollen seasons. In SDS-PAGE, 15 protein bands were detected from LS pollen, while 14 bands from SC. The IgE-specific immunoblotting with patients' sera allergic to LS displayed five major allergens, while four major allergens were detected from SC. This would be the first report from India to prove the allergenic potentiality of airborne pollen of these two common avenue trees of India.

Spectrum of Allergens and Allergen Biology in India.

The growing prevalence of allergy and asthma in India has become a major health concern with symptoms ranging from mild rhinitis to severe asthma and even life-threatening anaphylaxis. The "allergen repertoire" of this subcontinent is highly diverse due to the varied climate, flora, and food habits. The proper identification, purification, and molecular characterization of allergy-eliciting molecules are essential in order to facilitate an accurate diagnosis and to design immunotherapeutic vaccines. Although several reports on prevalent allergens are available, most of these studies were based on preliminary detection and identification of the allergens. Only a few of these allergen molecules have been characterized by recombinant technology and structural biology. The present review first describes the composition, distribution pattern, and natural sources of the predominant allergens in India along with the prevalence of sensitization to these allergens across the country. We go on to present a comprehensive report on the biochemical, immunological, and molecular information on the allergens reported so far from India. The review also covers the studies on allergy- related biosafety assessment of transgenic plants. Finally, we discuss the allergen-specific immunotherapy trials performed in India.

Monitoring of airborne fungal spore load in relation to meteorological factors, air pollutants and allergic symptoms in Farakka, an unexplored biozone of eastern India.

Fungi are important components of atmosphere which play a major role in causing respiratory allergy upon inhalation. An airborne fungal spore survey was carried out in two outdoor environments in Farakka-an unexplored township covering the National Thermal Power Station, West Bengal, India for a period of 2 years (October 2013 to September 2015). A Burkard personal volumetric air sampler was used at 15 days interval to collect the total fungal spore load. A fungal spore calendar has been prepared depicting monthly average spore concentration in the air. The relationships between fungal spore concentration and different climatic factors were analysed statistically. Higher concentration levels of aerospora and pollutants were recorded during winter season. A detailed questionnaire was used to obtain medical history data of 523 local patients visiting the outpatients department of Farakka NTPC (National Thermal Power Station) hospital. A significant positive correlation was found between fungal spore concentration, atmospheric pollutants and allergic manifestation. The dominant fungal spores were isolated, sub-cultured and tested for allergenic potential by skin prick tests (SPTs) among subjects with clinical history of respiratory allergy, which evoked ˃45.0% skin reactivity upon individuals.

Partial purification and characterization of an antimicrobial activity from the wood extract of mangrove plant Ceriops decandra.

The development of resistance towards the antibiotics in use today has been a source of growing concern in the modern healthcare system around the world. To counter this major threat, there is an urgent need for discovery of new antimicrobials. Many plants, like mangroves, possess highly diversified list of natural phytochemicals which are known to have wide range of bioactivities. These phytochemicals can be good sources for the discovery of new drugs. In this study, we report the partial phytochemical characterization and antimicrobial activities of a semi-purified fraction isolated from the wood tissue of Ceriops decandra, a mangrove plant. This fraction named CD-3PM was chromatographically separated from C. decandra wood extract and was subjected to different spectral analyses to determine its partial chemical nature. The structural investigation indicates the presence of two diterpenoids, i) 3ß, 13ß-Dihydroxy-8-abietaen-7-one and ii) 3ß-Hydroxy-8,13-abietadien-7-one in the CD-3PM fraction. The antimicrobial potential of this fraction was evaluated by microdilution-MTT assay against several organisms. Among the nine microorganisms found to be sensitive to the CD-3PM fraction, six organisms are reported to be pathogenic in nature. The CD-3PM fraction with broad spectrum antimicrobial efficacy revealed the presence of two diterpenoids and possesses potential applications in drug discovery process and food processing industries.

Aerobiological, clinical, and immunobiochemical studies on Alstonia scholaris pollen from Eastern India.

Alstonia scholaris or Indian devil tree is a common, evergreen, tropical tree of the Apocynaceae family. The objectives of this study were (a) to observe the seasonal variation of A. scholaris pollen in the atmosphere of an industrial and rural area of West Bengal, India by conducting a 2-year aerobiological survey with a Burkard personal volumetric sampler, (b) to study its allergenicity in the local population by in vivo (skin-prick test) and in vitro tests (enzyme-linked immunosorbent assay and dot blotting), (c) to identify the immunoglobulin E (IgE)-binding proteins present in the pollen extract (sodium dodecyl sulphate-polyacrylamide gel electrophoresis and immunoblotting), (d) to study its chemical composition. A. scholaris pollen were present in the air from September until November. They contained 14.3 % carbohydrate, 9.2 % lipid, and 4.3 % protein. Among 140 respiratory allergic local patients, 28.57 % showed positive skin reaction to A. scholaris pollen extract. Twelve protein bands in the range of 94.4-13.3 kDa were observed in the pollen extract. Seven IgE-binding proteins were found. Among them, one component of 29.9 kDa was the most important in A. scholaris pollen extract. This component could be purified and would be helpful in the diagnosis and therapy of A. scholaris pollen-susceptible patients.

Airborne load of Cassia pollen in West Bengal, eastern India: its atmospheric variation and health impact.

A Burkard personal volumetric sampler was used at Sriniketan, a town about 150 km northwest of Calcutta, in the state of West Bengal, in eastern India to record the frequency of three common airborne Cassia pollen types, Cassia tora, Cassia occidentalis, and Cassia fistula for two consecutive years (2004-2006). Correlation was made between the meteorological factors and the pollen concentration in the atmosphere. The study reports Cassia pollinosis by in vivo skin prick test in respiratory allergic patients. The highest positive reactions were exhibited by C. tora (34.7 %), C. fistula (33.3 %), and C. occidentalis (28.5 %). The allergic potential of these was investigated by in vitro enzyme linked immunosorbent assay test. Their protein components were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, in the range of 15.8-81.5 kDa. In C. occidentalis and C. fistula, 11 bands were found, while it was 10 in C. tora. The results show that the Cassia pollen occur significantly in the atmosphere with the potential to elicit an allergic response in susceptible patients.

The occurrence and allergising potential of airborne pollen in West Bengal, India.

A continuous 2-year volumetric aerobiological survey was conducted in Berhampore town, a centrally located and representative part of West Bengal, India. The aim of the study was to assess the allergising potential of airborne pollen grains of West Bengal. A total of 31 pollen types were identified of which Poaceae (grasses) pollen showed maximum frequency, followed by Cyperaceae, Cassia sp., Acacia auriculiformis, etc. The seasonal periodicities of the pollen types and their relationship to meteorological conditions were investigated. It was found that the pollen concentration is positively correlated with temperature and negatively correlated with rainfall and relative humidity. Clinical investigations by skin prick test were carried out to detect allergenicity of pollen types. Eighteen common airborne pollen types induced positive responses of which pollen extracts of Saccharum officinarum (grass), Azadirachta indica, Cocos nucifera, Phoenix sylvestris, Cyperus rotundus and Eucalyptus citriodora showed strongest sensitising potential. This result is consistent with previous investigations in different parts of West Bengal.

Deteriorative changes of maize, groundnut and soybean seeds by fungi in storage.

This paper deals with investigations on fungal infection, moisture content, germinability and deterioration of three seeds, viz., maize (starchy), groundnut (oily) and soybean (proteinaceous) in storage at the locality of Santiniketan, West Bengal, India, under natural condition for 1 year. The airspora of storage environment was trapped using culture plate method. Different species of Aspergillus (A. candidus, A. flavus, A. niger, A. terreus, and A. ruber) were dominant followed by Rhizopus, Penicillium, Curvularia, Fusarium, Alternaria, etc. Seed moisture was maximum in the rainy season followed by a gradual decrease during longer storage. A gradual decrease in field fungi with simultaneous increase in storage fungi accompanied by a reduction in germinability occurred in all seeds as storage proceeded. A gradual loss of carbohydrate (both soluble and insoluble) content in all the seeds were recorded. A loss of protein content was recorded followed by a small increase. Oil content decreased in prolonged storage with simultaneous increase in fatty acid.