ABSTRACT
BACKGROUND: Serological techniques like enzyme linked immunosorbent assay (ELISA) and immunoblotting are useful for detection of mycobacterial antigens of diagnostic importance in tuberculosis. AIM: To isolate and identify circulating tuberculous antigens reactive with sputum positive and sputum negative pulmonary tuberculosis (PTB) sera. METHODS: Circulating tuberculous antigen was isolated by ammonium sulphate fractionation from the sera of sputum positive and sputum negative (clinically and radiologically diagnosed) PTB cases. The circulating antigen fractions and individual patients' serum samples were resolved by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting was performed using anti M.tb sonicate IgG as a probe to detect antigens. RESULTS: Anti M.tb sonicate IgG was found to be reactive with mycobacterial proteins 170 kDa, 140 kDa, 85 kDa, 55 kDa, 43 kDa, 20 kDa and 16 kDa in the antigen fraction isolated from sputum positive tuberculosis sera by immunoblotting. However only 85 kDa, 55kDa, 43 kDa and 20 kDa antigenic proteins were found to be recognized by anti sonicate IgG in the antigen isolated from sputum negative sera. These observations were further confirmed by analysis of individual S+ and S- PTB serum by immunoblotting. CONCLUSION: Seroreactive studies of circulating tuberculous antigens showed the presence of 170 kDa, 140 kDa, 85 kDa, 55 kDa, 43 kDa, 20 kDa and 16 kDa protein antigens in sputum positive sera, while 85 kDa, 55 kDa, 43 kDa and 20 kDa antigens were found to be present in sputum negative PTB which need further evaluation for their use in serological diagnosis of tuberculosis.
Subject(s)
Ammonium Sulfate/chemistry , Animals , Antigens, Bacterial/chemistry , Electrophoresis, Polyacrylamide Gel , Enzyme-Linked Immunosorbent Assay/methods , Goats , Humans , Immunoblotting , Immunoglobulin G/chemistry , Mycobacterium tuberculosis/immunology , Penicillinase/chemistry , Serologic Tests/methods , Sputum/metabolism , Tuberculosis, Pulmonary/immunologyABSTRACT
Optimization of the fermentation medium for maximum alkaline protease production was carried out with a new strain of Pseudomonas aeruginosa (B-2). Replacing the protein source/inducer (albumin in place of casein) brought about significant increase in yield after 48 hr of inoculation. Three most effective medium constituents identified by initial screening method of Plackett-Burman were albumin, (NH4)2SO4 and glucose. Central Composite Design (CCD) and Response Surface Methodology (RSM) were used in the design of the experiment and in the analysis of the results. Optimum levels of the effective medium constituents were albumin (6.586%); (NH4)2SO4, 0.164%; and glucose, 6.72%. The alkaline protease production increased from 533460 to 793492 Ul(-1).
Subject(s)
Ammonium Sulfate/chemistry , Bacterial Proteins/biosynthesis , Bacteriological Techniques , Caseins/chemistry , Cell Culture Techniques , Culture Media/chemistry , Endopeptidases/biosynthesis , Fermentation , Glucose/chemistry , Models, Statistical , Pseudomonas aeruginosa/drug effects , Serum Albumin/chemistry , Time FactorsABSTRACT
Two types of buffered media, strictly defined-Ammonium sulphate-basal salts and complex Peptone-basal salts, were used for the cultivation of Dimorphomyces pleomorphis, one of two dimorphic fungi isolated from fermenting juice of soursop fruit, Annona muricata L. The growth count was taken every twenty-four hours. Transient morphologies were observed to change from sporangiospores through enlarged globose cells, to granular particles and eventually, polar budding yeast cells in the strictly defined medium at 15 degrees, 20 degrees, or 37 degrees C, but the complex medium casually terminally induced polar budding yeast cells and multipolar budding yeast like cells in between the growth phases, at 15 degrees and 20 degrees C, while mainly multipolar budding yeastlike morphology was observed at elevated temperature. There was obvious influence of nutritional factor or morphological expression (p < 0.01). After analysis of variance, the growth data could not fit into predictive quadratic polynomial model because the organism's response curves were incongruent with basic assumptions of the model. Furthermore, a stepwise regression analysis gave very low coefficients of determination, r2, for the interactive combinations. They were therefore, considered unfit for the data. Construction of the pII-profiles led to inference being drawn from the chemiosmotic theory, polyelectrolyte theory to account for the behaviour in the buffered multiionic media. It was also thought that inherent cellular mitotic division and glycolytic activity led to a prelogarithmic growth response.
Subject(s)
Ammonium Sulfate/chemistry , Culture Media , Fermentation , Fruit , Fungi/genetics , Nigeria , Peptones/chemistry , Regression Analysis , Spores, Fungal/genetics , TemperatureABSTRACT
A culture medium for batch production of d-endotoxin by Bacillus thuringiensis (B., t.) has been modified. Through batch and continuous cultivation studies, the original medium was diagnosed to be limited in organic nitrogen. Corn steep liquor was found to be an excellent source for the organic nitrogen and its addition resulted in a carbon limited medium and in a significant increase in the amount of spore-toxin complex formed in shake flasks. Results of bioassay, conducted on Trichoplusia ni, suggest enhancement of larvicidal efficacy under carbon-limited growth conditions.