Quantifying the ultrastructure changes of air-dried and irradiated human amniotic membrane using atomic force microscopy: a preliminary study

Air-dried and sterilized amnion has been widely used as a dressing to treat burn and partial thickness wounds. Sterilisation at the standard dose of 25 kGy was reported to cause changes in the morphological structure as observed under the scanning electron microscope. This study aimed to quantify the changes in the ultrastructure of the air-dried amnion after gamma-irradiated at several doses by using atomic force microscope. Human placentae were retrieved from mothers who had undergone cesarean elective surgery. Amnion separated from chorion was processed and air-dried for 16 h. It was cut into 10?×?10 mm, individually packed and exposed to gamma irradiation at 5, 15, 25 and 35 kGy. Changes in the ultrastructural images of the amnion were quantified in term of diameter of the epithelial cells, size of the intercellular gap and membrane surface roughness. The longest diameter of the amnion cells reduced significantly after radiation (p?<?0.01) however the effect was not dose dependent. No significant changes in the shortest diameter after radiation, except at 35 kGy which decreased significantly when compared to 5 kGy (p?<?0.01). The size of the irradiated air-dried amnion cells reduced in the same direction without affecting the gross ultrastructure. At 15 kGy the intercellular gap decreased significantly (p?<?0.01) with Ra and Rq, values reflecting surface roughness, were significantly the highest (p?<?0.01). Changes in the ultrastructure quantified by using atomic force microscope could complement results from other microscopic techniques.     

Goat mammary epithelial cells provide a better expression system for production of recombinant human bone morphogenetic protein 2 compared to Chinese hamster ovarian cells

Bone Morphogenetic Protein 2 (BMP2), a member of the Transforming Growth Factor-β (TGF-β) super family of proteins and is instrumental in the repair of fractures. The synthesis of BMP2 involves extensive post-translational processing and several studies have demonstrated the abysmally low production of rhBMP2 in eukaryotic systems, which may be due to the short half-life of the bioactive protein. Consequently, production costs of rhBMP2 are quite high, limiting its availability to the general populace. Therefore, there is an urgent need to identify better in-vitro systems for large scale production of rhBMP2. In the present study, we have carried out a comparative analysis of rhBMP2 production by the conventionally used Chinese Hamster ovarian cells (CHO) and goat mammary epithelial cells (GMEC), upon transfection with appropriate construct. Udder gland cells are highly secretory, and we reasoned that such cells may serve as a better in-vitro model for large scale production of rhBMP2. Our results indicated that the synthesis and secretion of bioactive rhBMP2 by goat mammary epithelial cells was significantly higher as compared to that by CHO-K1 cells. Our results provide strong evidence that GMECs may serve as a better alternative to other mammalian cells used for therapeutic protein production.     

Antimicrobial potential and taxonomic investigation of piezotolerant <Emphasis Type="Italic">Streptomyces</Emphasis> sp. NIOT-Ch-40 isolated from deep-sea sediment

Microbial-derived natural products from extreme niches such as deepsea are known to possess structural and functional novelty. With this background, the present study was designed to investigate the bioprospecting potential and systematics of a deep-sea derived piezotolerant bacterial strain NIOT-Ch-40, showing affiliation to the genus Streptomyces based on 16S RNA gene similarity. Preliminary screening for the presence of biosynthetic genes like polyketide synthase I, polyketide synthase II, non ribosomal peptide synthase, 3-amino-5-hydroxybenzoic acid synthase and spiroindimicin followed by antibacterial activity testing confirmed the presence of potent bioactivity. The secondary metabolites produced during fermentation in Streptomyces broth at 28?°C for 7 days were extracted with ethyl acetate. The extract exhibited a specific inhibitory activity against Gram-positive bacteria and was significantly effective (p?<?0.0001) against methicillin-resistant Staphylococcus aureus (MRSA). The minimum inhibitory concentration and minimum bactericidal concentration against MRSA was 1.5 µg/mL, which was statistically significant in comparison with erythromycin. A multifaceted analysis of the Streptomyces spp. was carried out to delineate the strain NIOT-Ch-40 at a higher resolution which includes morphological, biochemical and molecular studies. Piezotolerance studies and comparison of fatty acid profiles at high pressures revealed that it could be considered as one of the taxonomic markers, especially for the strains isolated from the deep sea environments. In conclusion, the observation of comparative studies with reference strains indicated towards the strain NIOT-Ch-40 as an indigenous marine piezotolerant Streptomyces sp. with a higher probability of obtaining novel bioactive metabolites.     

Microbial lipids from renewable resources: production and characterization

A number of microorganisms belonging to the genera of algae, yeast, bacteria, and fungi have ability to accumulate neutral lipids under specific cultivation conditions. The microbial lipids contain high fractions of polyunsaturated fatty acids and have the potential to serve as a source of significant quantities of transportation fuels. This paper reviews the current state of the art of this field. It summarizes the various microorganism used, feed stocks available, environmental factors that influence growth of cells and accumulation of lipids, major fatty acid composition of lipids, and the technology.     

Phosphate Solubilizing Bacillus megaterium mj1212 Regulates Endogenous Plant Carbohydrates and Amino Acids Contents to Promote Mustard Plant Growth

The current study was conducted to explore the potential of a phosphate solubilizing soil bacterium, Bacillus megaterium mj1212 for enhancing the growth of mustard plants. The newly isolated bacterial strain mj1212 was identified as B. megaterium using phylogenetic analysis and, its phosphate solubilization ability was shown by the clear zone formation on National Botanical Research Institute’s Phosphate medium. Moreover, the phosphate solubilization ability of B. megaterium mj1212 was enhanced by optimal culture conditions at pH 7.0 and 35 °C which might be due to the presence of malic and quinic acid in the culture medium. The beneficial effect of B. megaterium mj1212 in mustard plants was determined by an increasing shoot length, root length and fresh weight of plants. In the biochemical analysis revealed that chlorophyll, sucrose, glucose, fructose and amino acids (Asp, Thr, Ser, Glu, Gly, Ala, Cys, Val, Met, Ilu, Leu, Tyr, Phe, Lys, His, Arg and Pro) were higher in B. megaterium mj1212 treated plants, when compared to their control. The result of present study suggests that B. megaterium mj1212 treatment could be act as phosphate biofertilizer to improve the plant growth.     

Phylogenetic diversity of rhizobia associated with horsegram [Macrotyloma uniflorum (Lam.) Verdc.] grown in South India based on glnII, recA and 16S-23S intergenic sequence analyses

Horsegram [Macrotyloma uniflorum (Lam.) Verdc.) is an important grain legume and fodder crop in India. Information on root nodule endosymbionts of this legume in India is limited. In the present study, 69 isolates from naturally occurring root nodules of horsegram collected from two agro-eco-climatic regions of South India was analyzed by generation rate, acid/alkali reaction on YMA medium, restriction fragment length polymorphism analysis of 16S-23S rDNA intergenic spacer region (IGS), and sequence analyses of IGS and housekeeping genes glnII and recA. Based on the rDNA IGS RFLP by means of three restriction enzymes rhizobia were grouped in five clusters (I–V). By sequence analysis of 16S-23S rDNA IGS identified genotypes of horsegram rhizobia were distributed into five divergent lineages of Bradyrhizobium genus which comprised (I) the IGS type IV rhizobia and valid species B. yuanmingense, (II) the strains of IGS type I and Bradyrhizobium sp. ORS 3257 isolated from Vigna sp., (III) the strains of the IGS type II and Bradyrhizobium sp. CIRADAc12 from Acacia sp., (IV) the IGS type V strains and Bradyrhizobium sp. genospecies IV, and (V) comprising genetically distinct IGS type III strains which probably represent an uncharacterized new genomic species. Nearly, 87% of indigenous horsegram isolates (IGS types I, II, III, and V) could not be related to any other species within the genus Bradyrhizobium. Phylogeny based on housekeeping glnII and recA genes confirmed those results found by the analysis of the IGS sequence. All the isolated rhizobia nodulated Macrotyloma sp. and Vigna spp., and only some of them formed nodules on Arachis hypogeae. The isolates within each IGS type varied in their ability to fix nitrogen. Selection for high symbiotic effective strains could reward horsegram production in poor soils of South India where this legume is largely cultivated.