Showing papers by "Joydeep Mukherjee published in 2009"

Purification and characterization of a collagenolytic enzyme from a pathogen of the great barrier reef sponge, Rhopaloeides odorabile.

Abstract: Background In recent years there has been a global increase in reports of disease affecting marine sponges. While disease outbreaks have the potential to seriously impact on the survival of sponge populations, the ecology of the marine environment and the health of associated invertebrates, our understanding of sponge disease is extremely limited. Methodology/principal findings A collagenolytic enzyme suspected to enhance pathogenicity of bacterial strain NW4327 against the sponge Rhopaloeides odorabile was purified using combinations of size exclusion and anion exchange chromatography. After achieving a 77-fold increase in specific activity, continued purification decreased the yield to 21-fold with 7.2% recovery (specific activity 2575 collagen degrading units mg(-1)protein) possibly due to removal of co-factors. SDS-PAGE of the partially pure enzyme showed two proteins weighing approximately 116 and 45 kDa with the heavier band being similar to reported molecular weights of collagenases from Clostridium and marine Vibrios. The enzyme degraded tissue fibres of several sponge genera suggesting that NW4327 could be deleterious to other sponge species. Activity towards casein and bird feather keratin indicates that the partially purified collagenase is either a non-selective protease able to digest collagen or is contaminated with non-specific proteases. Enzyme activity was highest at pH 5 (the internal pH of R. odorabile) and 30 degrees C (the average ambient seawater temperature). Activity under partially anaerobic conditions also supports the role of this enzyme in the degradation of the spongin tissue. Cultivation of NW4327 in the presence of collagen increased production of collagenase by 30%. Enhanced enzyme activity when NW4327 was cultivated in media formulated in sterile natural seawater indicates the presence of other factors that influence enzyme synthesis. Conclusions/significance Several aspects of the sponge disease etiology were revealed, particularly the strong correlation with the internal tissue chemistry and environmental temperature. This research provides a platform for further investigations into the virulence mechanisms of sponge pathogens.

Antibiotic production by a marine isolate (MS 310) in an ultra-low-speed rotating disk bioreactor

Abstract: Actinomycin-D production by a biofilm-forming estuarine isolate viz Streptomyces sp. MS 310 is studied in small-scale shaken cultures, as well as in a 25 L rotating disk bioreactor, (RDBR) which, when operated at a disk rotational speed of 1 revolution/day with 50% disk submergence, mimics the intertidal conditions of the microbe’s niche estuarine habitat-alternating 12 h periods of inundation and exposure. The ideal pH and temperature for antibiotic production are determined (pH 10, 30°C) through a designed experimental study using shaken flasks. Subsequently, operating conditions in the RDBR are investigated employing a 3n experimental design wherein each of two (n = 2) parameters viz. aeration and disk submergence are considered at three levels viz high, medium, and low: 9.0, 6.0, and 3.0 L/min for aeration rate; and 75, 50, and 25% for disk submergence, (while maintaining the rotational speed at 1.0 rev/day). The niche-mimic condition along with maximum permissible aeration is found to be most favorable for antibiotic production — peak antibiotic activity (PAA) and peak activity attainment rate (PAAR) simultaneously attaining their highest values: 40 mm and 2.13 mm/h, respectively. Both PAA and PAAR increase with increasing aeration at all operating conditions examined — particularly, at the niche-mimic condition, a threefold increase in aeration rate (3∼9 L/min) causes PAA to increase by 33%, whereas PAAR increases by 2.5 times, thus pointing to the strong aeration dependence of this actinomycin-D producer. Again, compared to the best values obtained in the 500 mL shaken flask experiments, corresponding RDBR values are 16% higher for PAA and more than 5 times higher for PAAR — strong evidence for employing these novel bioreactors for cultivation of antibiotic-producer marine microbes.

Antibacterial activity and anticorrosive efficiency of aqueous methanolic extract of Artemisia pallens (Asteraceae) and its major constituent.

Abstract: An aqueous-methanolic [methanol:water (1:1)1 extract of the whole plant of Artemisia pallens (Family: Asteraceae) has shown good antibacterial activity against Pseudomonas aeruginosa and Shigella flexneri at the concentration of 100 mg/mL. The activity-guided fractionation has led to isolation of compound 4-hydroxyphenyl-/3-D-glucopyranoside (which is known as arbutin), as a major constituent and exhibited the same antibacterial activity at a concen-tration of 17 mg/mL. Both the crude extract and arbutin also showed 94% anticorrosive efficiency against mild steel in 1M HC1 at concentration of 200 mg/L.