Scribd
Upload
49 views36 pages

Bacillus Subtilis

Bacillus subtilis is a Gram-positive, rod-shaped soil bacterium. It is used as a model organism and can produce endospores. It is mass produced industrially using fermentation and formulated as a biopesticide or biofertilizer to control plant pathogens.

Uploaded by

Gomathi Pichandi
Copyright
© © All Rights Reserved
We take content rights seriously. If you suspect this is your content, claim it here.
Available Formats
Download as PPTX, PDF, TXT or read online on Scribd
49 views36 pages

Bacillus Subtilis

Bacillus subtilis is a Gram-positive, rod-shaped soil bacterium. It is used as a model organism and can produce endospores. It is mass produced industrially using fermentation and formulated as a biopesticide or biofertilizer to control plant pathogens.

Uploaded by

Gomathi Pichandi
Copyright
© © All Rights Reserved
We take content rights seriously. If you suspect this is your content, claim it here.
Available Formats
Download as PPTX, PDF, TXT or read online on Scribd
You are on page 1/ 36

Bacillus subtilis

➢ Bacillus subtilis, known also as the hay


bacillus or grass bacillus.

➢ It is a Gram-positive rod shaped bacteria.

➢ Found in soil and the gastrointestinal


tract of ruminants, humans and marine
sponges.
➢ Historically been classified as an obligate aerobe
though evidence exists that it is facultative
aerobe.
➢ Used as a MODEL ORGANISM to study bacterial
chromosome replication and cell differentiation.
➢ Produces endospores.
➢ It is one of the bacterial champions in secreted
enzyme production and used on an industrial
scale by biotechnology.
CLASSIFICATION
Domain: Bacteria
Phylum. Firmicutes
Class: Bacilli
Order: Bacillales
Family: Bacillaceae
Order: Bacillales
Genus: Bacillus
Species: B. subtilis
Binomial name : Bacillus subtilis
MORPHOLOGY

● It is Rod-shaped bacteria - 4-
10 micrometers (um) long
0.25-1.0 µm in diameter.
● It is Gram-positive (Perez
2000).
● The cell wall - rigid structure
outside the cell.
● It is heavily flagellated.
● It is composed of peptidoglycan, which is a
polymer of sugars and amino acids.
● The peptidoglycan that is found in bacteria
is known as murein.
● They produce colonies which are dry, flat
and irregular with lobate margins.
PHYSIOLOGY

● It produces Endospores.
● In order for a recipient bacterium to bind, take up
exogenous DNA from another bacterium of the same
species and recombine it into its chromosome, it
must enter a special physiological state called
competence.
● Bacillus subtilis bacteria secrete enzymes, such as
amylase, protease, pullulanase, chitinase, xylanase,
lipase, among others.
● Optimal temperature range is 25-35°c.
ANATOMY

Most bacteria share


similarities in their cellular
structure. A typical bacterial
cell contains key structural
components that include:

Cell wall, cell membrane,


cytoplasm, circular DNA
(Nucleoid), ribosomes,
flagella, and pilli.
A bacterial cell wall is a rigid additional covering external to
the cell membrane that provides extra strength to the cell.
The composition of the bacterial cell wall is complex, with
carbohydrates as the chief component.
MASS
PRODUCTION
Collection of soil sample

The soil samples were taken


from 5 cm depth.

A zigzag format of sampling


was used.
Isolation by-serial dilution technique
Isolated in Nutrient Agar (NA) medium by serial
dilution technique.
The procedure adopted was as follows
● 10 g soil sample diluted in 90 ml of
sterile distilled water in 250 ml conical
flask and kept it a orbital shaker at 150
rpm to get a homogenized soil
suspension.
● Serial dilution was made.

● Dilution of 10-7 was inoculated into NA plates

● Incubated at 37°C for 24 h.

● Isolated colonies growing on each diluted plates were


transferred into freshly prepared nutrient agar slants.

● The bacterial strains isolated on NA slants were kept at


in refrigerator for further study.
Serial dilution
Identification and characterization
of the isolate
• The isolate was characterized by Gram staining
technique.
● Based on Gram's staining, the strain was found to be
Gram-positive.
● Microscopic observation-Rod shaped bacterium
arranged in chain
● .•(NOTE - Gram positive bacteria: Stain dark purple
due to retaining the primary dye called Crystal Violet
in the cell wall.
Gram staining :
During a Gram staining test, Bacillus subtilis
retain the crystal violet dye, appearing purplish-
blue under a microscope. This is due to the
presence of a thick peptidoglycan layer.
Mass multiplication of B. subtilis
Preparation of mother culture:
The nutrient broth medium is prepared as detailed
below:
Glucose 5.0 g

Peptone 5.0 g

Beef extract 3.0 g

Sodium chloride 3.0 g

Distilled water 1000ml


● The above medium is take in conical flasks.

● Autoclaved at 15 lb pressure for 15 min.

● A loop of B.subtilis is inoculated into the


medium and incubated for 2 days.

● This serve as the mother culture.


MASS MULTIPLICATION
● Sterilized nutrient broth medium
● A loopful of bacillus subtilis culture is
inoculated to the medium, incubated for 24hrs.
- Mother culture.
● Medium is prepared in a fermentor and
sterilized.
● Mother culture is added to the fermentor @ 1
liter/100 liters of medium .
● Incubate for 72 hrs
● Medium is mixed with carrier substrates
viz., flyash, peatsoil, talk powder at 1:2 ratio
● Carrier substances pH adjusted to 7 by
adding CaCo3 @150g/kg.
● Carrier substances sterilized at 20 p.s.i for
30 minutes for two sussessive days.
● Mixture is air dried & mixed with Carboxyl
Methyl Cellulose (CMC) @ 50g/kg of the
product for sticking.
● Packed in white polythene bags & used for
treating the seeds
QUALITY CONTROL OF BIOCONTROL AGENTS:

● Fresh produce should contain 10-8 cfu/g


Bacillus subtilis.
● Maximum storage period is 90days Bacillus
subtilis.
● After storage period at room temperature the
population should be 10-7cfu/g Bacillus subtilis.
● Size of the carrier substrate should be 50µ .
● Moisture content of the final product should not
be more than 20%
MODE OF ACTION
THE DIRECT MECHANISM :-
It includes the synthesis of many secondary
metabolites, hormones, cell-wall-degrading
enzymes, and antioxidants that assist the plant
in its defense against pathogen attack.

THE INDIRECT MECHANISM :-


It includes the stimulation of plant growth
and the induction of acquired systemic
resistance.
Formulation of B.subtilis
Product Formulation type Primary target
Serenade WP, aqueous Fungi, bacteria on
suspension multiple vegetables,
fruits

Codes WP (Conc.), flowable Fungi on cotton, large-


seeded legumes,
soybeans

Subtilex WP (Conc.) Fungi on cotton, large-


seeded legumes,
soybeans

Hi Stick L + Flowable Fungi on soybeans,


Subtilex peanut
METHOD OF APPLICATION
USES
● Feed additives in agriculture and animal husbandry to
enhance animal fiber digestion and intestinal health.

● Bio control agent

● Biofertilizer
Marketing
THANK YOU

You might also like