Date post: | 11-Nov-2014 |
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Technology |
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Bacteriophage Based Bioluminescence Biosensor For Pathogen Detection In Food
N.A.G. Sandaruwan
Why detection of micro organisms in food is important?
“2.2 million people die annually due to food and water borne illnesses, and 1.9 millions among them are children” -WHO
Conventional methods used for this are microbiological and biochemical analysis, ELISA, PCR etc.
They are overly time consuming, not amendable to on site detection and need a extensive training.
Biosensors have been identified as attractive alternatives to the existing conventional pathogen detection methods.
What is a Biosensor?
A biosensor is an analytical device, used for the detection of an analyte, that combines a biological component with a physicochemical detector.
Classification of Biosensors
Bioluminescence
History of Bioluminescence Technique
In 1667 Robert Boyle, English chemist discovered bioluminescence.
He observed few spot of bluish-green light on a chicken carcasses which was purchased the week prior.
Dubois confirmed oxygen dependency and demonstrated the necessity of an enzyme (luciferase) and a luciferase substrate for light emission.
BioluminescenceNeed luciferase enzyme.Substrate is catalytic oxidated by
the enzyme to produce light in the presence of molecular oxygen.
FMNH2 + RCHO + O2 FMN + RCOOH + H2O
+ Light
Bioluminescence in a microorganism
Membrane synthesis
Myristol ACP
Myristic acid
Myristic Aldehyde
Light (490nm)
FMNH2
Luciferase
Luciferase
Genetic Background
Luciferase can be found in bacteria ( lux ) and eukaryotes ( luc ).
It’s encoded by lux A and B genes.
In addition light generation requires lux C,D and E genes.
How to use a Bacteriophage?
Select a specific bacteriophage
Change the DNA component
Introduce to the medium
Multification
Reporter phage with lux gene
Introducing to the
sample
Contamination with
targeted MO
Reporter phage attack
to MO
Inject lux gene
Luciferase enzyme is produced
Light emmission
Industrial applications
A511::lux AB recombinant phage is used to identify Listeria bacterium in Spikked ricotta cheese, chocolate pudding etc.
A511::lux AB is used to check the quality of minced meat and soft cheese.
P22::lux AB is used to identify Salmonella Spp.
( ex:- in eggs )
Advantages of bacteriophage based biosensers
Very sensitive.Less time consumingMinimal sample preparation.Specificity of detection can distinguish between live
and dead MO s.
Limitations
Presence of specific phage inhibition.
Antiviral bacterial immunity system.
Thank you!