An Objective Method to Assess Bioluminescent Properties of Selected Bacterial Strains (original) (raw)
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Isolation and Characterization of Bioluminescent Bacteria
ICSESD-2017, 2017
Bacteria are the most abundant bioluminescent organisms. These luminescent bacteria are recognized to emit a blue/green light for numerous purposes and are provoked by the enzyme luciferase, regulated by the lux gene operon. Emission of light as a result of biochemical activities of living bacteria makes it possible to monitor environmental changes in ecosystems. The bioluminescent test offers a rapid, simple and sensitive method to test a wide spectrum of chemical substances and environmental samples including water, wastewater, sludge extracts, radioactive waste, hazardous radiations etc. Present study aimed at isolation and characterization of luminescent bacterial strain isolated from beaches in India. The bacterial isolates were grown at different temperatures to standardize the optimum temperature which gave maximum growth and hence luminescence. These strains were subjected to biochemical and molecular identification. Also, the bioluminescent activity was measured by calculating the luminescence using a laboratory luminometer and was expressed in RLU (Relative Luminescent Units). The isolated strain was confirmed as Vibrio harveyi and modified Nutrient Agar medium were indicated as the finest demonstrating proper bioluminescent activity most suitable for further applications.
3 Bacterial bioluminescence and its application
Bioluminescent analyses are one of the most promising express methods for biologically monitoring the environment because the luminescent system is highly sensitive to even micro quantities of toxicants. Bioassays based on luminous bacteria give an integral estimation of toxicity and frequently surpass other known bioassays in speed, accuracy, sensitivity and simplicity. The enzymes of bacterial luminescent system are used in developing highly sensitive analytical methods for practical purposes. This paper considers questions about the development and use of bioluminescent bioassays, and the influence of analyzed substances on bacterial luminescence.
Isolation and Characterisation of Bioluminescent Bacteria
2016
AbstractBioluminescent bacteria are widespread in natural environment. Over the years many researcher have been studying the physiology, biochemistry & genetic control of bacterial bioluminescence. So far many problems have been experience in cultivating these bacteria in labs & maintaining their luminescence with the use of simple laboratory level microbiology practices. This project related to isolation of bioluminescent bacteria from various sample sources such as Shrimp, Squid, Octopus & fish like Raja-Rani (vernacular name) & their characterization. Bioluminescent bacteria are symbiotically associated with these organisms. These bacteria can be grown on specially designed media such as BOSS medium, NaCl complete medium. These bacteria can be used in area of environmental Microbiology using bioluminescent genes as biosensors for environmental studies. These bacteria glow in dark so they are easily identified by observing in dark..
Oxygen Affinities of the Bioluminescence Systems of Various Species of Luminous Bacteria
Microbiology, 1985
O2 affinities of the bioluminescence systems have been measured in five species of luminous bacteria. Photobacterium phosphoreum He-1 a, recently isolated from a fish light-organ, showed half maximal bioluminescence at 0.01 5 ~M-O , , whereas another symbiotic strain (PJ-la) gave a 33-fold higher value. Vibrio fischeri MJ-1 gave a value of 0.60 yM-O2, and the symbiotic Photobacterium leiognathi LN-la gave half maximal emission at 0.7 pM-0,. These results indicate the importance of strain selection for sensitive O2 measurements.
Bioluminescent System of Luminous Bacteria for Detection of Microbial Contamination
Journal of Siberian Federal University. Biology, 2018
Microbial contamination is usually analyzed using luciferin-luciferase system of fireflies by the detection of adenosine-5'-triphosphate (ATP). There is an opportunity to assess the bacterial contamination of various objects based on a quantitative analysis of other nucleotides. In the present study, a bioluminescent enzyme system of luminous bacteria NADH:FMN-oxidoreductase (Red) and luciferase (BLuc) was investigated to understand if it can be used for quantitative measurements of bacterial cells by nicotinamide adenine dinucleotide (NADH) and flavin mononucleotide (FMN) detection. To increase the sensitivity of bioluminescent system to FMN and NADH, optimization of assay conditions was performed by varying enzymes and substrates concentrations. The lowest limits of detection were 1.2 nM FMN and 0.1 pM NADH. Escherichia coli cells were used as a model bacterial sample. FMN and NADH extraction was made by destructing cell membrane by ultrasonication. Cell suspension was added into the reaction mixture instead of FMN and NADH, and light intensity depended on number of bacterial cells in the reaction mixture. Centrifugation of sonicated sample as an additional step of sample preparation did not improve the sensitivity of method. The experimental results showed that Red and BLuc system could detect at least 800 thousand bacterial cells mL-1 by determining concentration of NADH extracted from lysed cells, while 3.9 million cells mL-1 can be detected by determining concentration of FMN.
Isolation of bioluminescent bacteria from marine organisms
2020
Bioluminescence is an emission of cold light by enzyme driven reaction within certain living organisms. The most abundant and widely distributed light emitting organisms are luminescent bacteria. Such organisms are either found as freeliving in the ocean or in symbiotic relationship with the marine host. To employ bioluminescence in environmental monitoring, isolation of bioluminescent bacteria from the two different marine samples (sea water sample and various organs of squid and fish) were collected from different sites of Veraval seashore and fish markets located nearby seashore respectively. The marine organisms used in the study were 20-25 days old. Cultivation media that were used for isolation were sea water agar (SWA), luminous agar (LA) and nutrient agar (NA); out of which SWA proved to be the most suitable medium for their growth and luminescence. No bioluminescent bacterium was found in water samples and total five bioluminescent bacteria were isolated from five different organs of fish and squid each. Out of these five isolates, two were selected based on their maximum light intensity. These two isolates, PBS1 and PBF1, were further characterized biochemically. PBS1 was able to utilize glucose, galactose, maltose and were tested positive for catalase and oxidase tests. Similar results were obtained in case of PBF1 except it was tested positive for urease urea but was unable to utilize glucose. Both isolates thrived at neutral pH and showed maximum bioluminescence. Effect of NaCl concentration on luminescence revealed that the two isolates were not able to grow in media devoid of NaCl and the luminescence was found to be maximum at 3 % (w/v) NaCl supplementation.
Luminescence, 2011
Bioluminescence is a biochemical process occurring in many organisms. Bacterial bioluminescence has been investigated extensively that lead to many application of such knowledge. Quorum sensing in the bioluminescent bacteria is a chemical signal process to recognize the strength of its own population to start luminescence in harmony. There is a mechanism in these bacteria to recognize inter-species strength also. When there is higher number of these bacteria, the possibility and frequency of cell-cell physical contact will be high. In this study, the physical proximity was artificially enhanced between cells and the effect on luminescence in the concentrated cells in the normal culture medium and in presence of other non-bacterial cell free supernatant were investigated. The role of such physical contact in the quorum sensing in the bioluminescence is not known. Increase in the luminescence of V. fischeri when concentrated shows the presence of physical proximity facilitates the quorum sensing for their bioluminescence.
Screening and Characterization of Luminescent Bacterial Strain
Journal of Basic & Applied Sciences, 2012
Several samples of sea water and sea organisms were collected for the isolation of luminescent bacterial strain. The samples were collected from Clifton, Hawksbay, fisheri at Landhi and Jeti at Korangi, Karachi, Pakistan, the sea organisms such as Crab, Prawn, Fish, Eel, Fish Gut, Electric Fish were collected. Three different media were used for enrichment of samples that is BOSS, Luminescent Agar (LA) and nutrient broth. A total of twenty one different bacterial strains were selected for further studies on the basis of different colonial morphologies. They were designated as DGU101 to DGU116, DGU221-DGU224 and DGU227. These strains were checked for luminescent and tolerance against antibiotics and heavy metal salts. Only one strain isolated from sea water (Clifton beach) designated as DGU227 showed luminescent on Luminescent Agar (LA) medium. It also exhibited multiple metal and antibiotic resistances. The two large plasmids were isolated from DGU227 which were more than 10 kb in s...
Evaluation of the Ecotoxicity of Pollutants with Bioluminescent Microorganisms
Advances in Biochemical Engineering/Biotechnology, 2014
This chapter deals with the use of bioluminescent microorganisms in environmental monitoring, particularly in the assessment of the ecotoxicity of pollutants. Toxicity bioassays based on bioluminescent microorganisms are an interesting complement to classical toxicity assays, providing easiness of use, rapid response, mass production, and cost effectiveness. A description of the characteristics and main environmental applications in ecotoxicity testing of naturally bioluminescent microorganisms, covering bacteria and eukaryotes such as fungi and dinoglagellates, is reported in this chapter. The main features and applications of a wide variety of recombinant bioluminescent microorganisms, both prokaryotic and eukaryotic, are also summarized and critically considered. Quantitative structure-activity relationship models and hormesis are two important concepts in ecotoxicology; bioluminescent microorganisms have played a pivotal role in their development. As pollutants usually occur in complex mixtures in the environment, the use of both natural and recombinant bioluminescent microorganisms to assess mixture toxicity has been discussed. The main information has been summarized in tables, allowing quick consultation of the variety of luminescent organisms, bioluminescence gene systems, commercially available bioluminescent tests, environmental applications, and relevant references.