Lasiosan, a new exopolysaccharide from Lasiodiplodia sp. strain B2 (MTCC 6000): Structural characterization and biological evaluation
Graphical abstract
Introduction
Exopolysaccharides (EPS) produced by some fungal species have recently garnered renewed interest since they exhibited biological response modifying (BRM) activities and possibly function as leads for application in medical therapy [1]. There is growing evidence that bioactivities exhibited by certain EPS are dependent on the chemical structural features of the carbohydrate biopolymer, including monosaccharide composition, molecular weight, the type and configuration of the glycosidic linkages, the degree and size of the branch points, the conformation of the EPS itself, solubility and charge on the polymer [2]. Furthermore, the physiological roles of EPS are mainly related to pathogenicity, quorum sensing, biofilm formation, etc. [3]. The knowledge on biological properties, the structures and fermentation process of different microbial-derived EPS is very important for understanding their physiological activities and industrial applicability. These macromolecular EPS are mainly composed of sugars such as glucose, mannose and uronic acids [4]. EPS acts as a physical barrier to many pathogenic bacteria [5], apart from their anti-tumor [6] and immunostimulatory activities [7]. The fungal diversity is estimated to have 1.5 million species [8] and among them a few fungal genera produce EPS exhibiting diverse biological activities and chemical properties can be explored from an application perspective. Basidiomycete fungi including Ganoderma lucidum, Agaricus blazei, Lentinus edodes and Grifola frondosa were reported to produce a repertoire of diverse bioactive polysaccharides having anti-tumor and immunomodulating properties [9].
In continuation to our earlier efforts on microbial EPS from a biotechnological perspective [[10], [11], [12]], different fungal strains isolated from diverse biosphere zones of India and maintained as in-house culture collection were screened for EPS production. Among them, Lasiodiplodia sp. strain B2 (GenBank accession number of 18S rDNA sequence: KX878985) identified as a EPS producer was isolated earlier from a spoiled banana sample and maintained in our laboratory and also deposited in the Microbial Type Culture Collection, CSIR-Institute of Microbial Technology, Chandigarh, India, bearing accession number MTCC 6000. Lasiodiplodia sp. strain B2, a ascomyceteous fungus belongs to the family Botryosphaeriaceae. The culture on potato dextrose agar (PDA) medium grows rapidly at 28–30 °C, with a light to dark gray aerial mycelium and later produces abundant black pigments, which can be easily noticed from the reverse side of the PDA culture. Based on lactophenol cotton blue staining, the conidia were ellipsoidal, one-celled, hyaline when immature, becoming dark brown and formed septa with age, whilst enclosed within the pycnidia. Normally, the pigmentation and septation happen after they get discharged. Furthermore, the conidia invariably took on a striate appearance. Externally the walls were smooth, but melanin deposited on the inner surface of the walls often gave the conidia a verruculose appearance. The main characteristics were the presence of aseptate paraphyses within the pycnidial conidiomata, but these paraphyses developed septa as they matured. In the present study, the EPS produced by Lasiodiplodia sp. strain B2 was purified, structurally characterized and further evaluated for various biological activities.
Section snippets
Fungal species and fermentation conditions
EPS production by Lasiodiplodia sp. strain B2 was carried out in a minimal salts medium (pH 8) containing (per litre): 20 g maltodextrin, 2 g NaNO3, 5 g MgSO4 and 1 g KH2PO4 and incubated at 30 °C with agitation at 150 rev min−1 in an Innova 43R (New Brunswick Scientific Co. Inc., Edison, New Jersey, USA) rotary shaker for 120 h.
Purification and structural characterization of EPS
After fermentation, the medium was filtered through a muslin cheese cloth to remove the fungal biomass and then the infiltrate was centrifuged at 2000 × g to obtain a
Purification and structural elucidation of EPS
Lasiodiplodia theobromae is an endophyte to many different fruits and vegetables, while its teleomorphic form, Botryosphaeria rhodina is a plant pathogen causing spoilage of fruits and vegetables worldwide. In the present study, the purified EPS from Lasiodiplodia sp. strain B2 was a white fibrous solid. The final pure EPS of 1.405 g L−1 was obtained from 3 g L−1 of the original isopropanol precipitated EPS. The average molecular weight (Mw) recorded was 29.3 kDa based on HPLC analysis using
Conclusions
Lasiodiplodia sp. strain B2 isolated from spoiled banana sample produced a bioactive exopolysaccharide. After isolation, purification and structural elucidation based on biochemical and spectral analysis, the EPS was identified as a glucomannan and it was named as Lasiosan. Results demonstrated that the exploration for novel fungal polysaccharides seems to be very interesting from a medicinal application perspective, since Lasiosan purified from Lasiodiplodia sp. strain B2 exhibited
Conflicts of interest
The authors declare that there is no conflict of interest with any researcher or funding agency.
Acknowledgement
The authors acknowledge the financial assistance provided in the form of a Senior Research Fellowships by the Council of Scientific and Industrial Research (CSIR), New Delhi, Government of India to Ms. Poornima Mongolla and Ms. Sujitha Pombala.
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