الفهرس 
History of Bacillus thuriengiensis
Field application problems of B. thuringiensis productsOnly 14 pages are availabe for public view
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Abstract
The genomic era (study of genes and their functions) started 15
years ago and several innovated technology have developed in
association with it. These technologies could be adopted to deep study
B.thuringiensis, to discover the information wealth hidden inside its
genome. These could lead to discovery of novel proteins with different
applications, not only for pest control but also for medical or
pharmaceutical importance.
B. thuringiensis-based biopesticides (Bt) are great environmental
friendly alternative to the hazardous chemical pesticides. However, the
major drawback of these pesticides is the inactivation by solar irradiation,
especially from the ultraviolet componenet. The UV causes damage to
the Bt-based biopesticide and hence the persistence of the compound in
the fields is compromised and this leads to the great economic loss and
raises the total application costs to the farmers. This also, hinders the
wide spread use of this environmental friendly biopesticide.
Industry did not stand still, they innovated various techniques to
achieve protection of Bt-based biopesticides from solar inactivation
which included encapsulation and/or addition of variety of chemical
sunscreens. The drawbacks of chemical additives are the negative
ecological impact and the hazardous effect on human health; they are
reported to cause allergy, eye irritation to the applicants and have
potencial mutangenic and/or carcinogenic effects, as well.
The approach adopted in this thesis is to enhance the local stains
of Bt, which are used to produce the first Egyptian bioinsecticide
AGERIN®, to produce their own natural UV-protecting agent so that the
AGERIN® will be totally biological product. Moreover,the study aimed
to optimize the fermentation conditions at the bench- and pilot-scale to
produce not only the UV-protecting agent but also the insecticidal crystal
protein (ICP) as well.
In order to attain these goals, experiments were designed to:
1. Characterize the Bt mutants able to produce UV-protecting agent
(melanin) using classical bacteriological and molecular biology
techniques; especially their ability to produce ICP by endospore
staining.
2. Chemical characterization of the produced pigment using UV and
FTIR spectra in comparison to standard melanin.
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3. PCR detection and cloning of the gene or gene encoding for the
production of the UV protecting agent, melanin. The cloned gene
was sequenced, as well.
4. Optimizing the production of melanin and ICP either at bench or
pilot-scale fermentation several parameters were studied, they
listed in the following:
- Determination of the best carbon source using from
amongst the locally available raw materials such as corn
starch, soy meal, lactose, sucrose and the best source was
corn starch.
- Determination of the best nitrogen source from amongst
soy meal, urea, and corn gluten and the best source was
soy meal.
- Determination of the best C/N ratio for the production of
melanin pigment and ICP.
- Determination of the best inculum size, incubation
temperature, pH value and aeration. The physical factors
were then standardized as follows: inoculum size one
tenth of the fermenter volume, run fermentation at 30ºC,
adjust the pH value between 7-7.5, aeration rate was best
when the fermenter volume ranged from one third to two
thirds of the vessel.
- Several antifoam agent were studied (maize oil, strucktol
670, sigma, rodersiil silicon) and the best foam breaker
was strucktol 670.
5. Qulaity control measures were applied such as the estimation of
melanin production and ICP formation and ability of melanin to
ptotect ICP under the harshest sun exposure time.
- Melanin was determined spectrophotometrically at 550nm
and its value was estimated from a standard curve.
- Bioassay experiments to determine the potecy and
efficacy of ICP with and without sunscreeners and/or
melanin. Melanin at its lowest concentration 0.01 mg /ml
was able to protect ICP from sun damage. This is much
more superior compared to the chemical sunscreeners
which are normally added at a ranged from 25-35%.