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Practical of microbiology
General characteristics of bacteria
1-it exists in all habitats (air-water-soil)
2-it is unicellular
3-prokaryotes have no organelles,DNA exist in cytoplasm
Shapes of bacteria
(1)cocci
-mono cocci -diplo coccus -streptococcus
-tetrads -staphylococcus -sacrina
(2)Bacilli (have two types)
a)straight
-bacillus -diplobacilli -streptobacilli
B)curved
-vibrio -spirillum
اﻋﺮف اﻻﺷﲀل *ٮﺘﺎﻋﺘﻬﻢ
-Sterilization All microbes endospore condition
-Disinfection vegetative bacteria Normal condition
Sterilization : the process of making something free from bacteria or other living microorganisms
There are three methods of sterilization
(1)physical methods (two types)
A)By heat (by two methods)
I)moist
-autoclave,used in operation tools-media and sterilized at 121 ^o and pressure 1.5 and 20 min
-boiling water ,effect vegetative bacteria only
-Fraction sterilization, in media sensitive to temperature above 100 ^o
-pasterilization (milk-milk product) make rapid cooling at 71 ^o
II)Dry
-by hot air oven
B)By radiation (by two method)
I)ionizing
-X-ray,Gamma ray and cathode ray
II)Non ionizing
-operation yooms and isolation units
(2)chemical methods
-(I2,Cl2,ethanol)
(3)mechanical
-By filterization
Types of media
1)function
-Selective
-Differential
-Maintenance (example nutrient broth)
2)composition
-Natural
-synthetic (example nutrient agar as beef extract, peptone,Nacl,agar,dil H2O)
-semi synthetic (potato dextrose agar)
Colony description
1)Shape
-oval -spherical -circular
-(Rossete -shaped ) -irregular
2)Surface
-shiny -Dull
3)edge
-entire -labbled -waved -Ciliated
4)Colour
-white -Buff -Yellow
-orange
5)Size
-normal -small -large
6)elevation
-flat -raised(unblicate-umbinatic)
7)internal structure
-smooth -Rough
Isolation bacteria from different sources
Tools
1-media
2-flame
3-petri-dishes
4-hair
5-alcohol
6-gloves
7-Isolation needle
Procedure
1-sterile media
2-sterile petri-dishes
3-pour media into plates,near flame
4-soldify
5-inoculate plates with (hair,air,water ,breath)
6-lable plates with (name ,source and date)
7-Incubate plates 30^o ,7 days
Observation
-after 7 days,different bacteria appear on petri-dishes
ڡوق/ ال اﺗكﻠﻤﻨﺎ ﻋﻠيﻬﺎcolony description ڡروض ﻫﻨﻌﻤﻞ/"ٮﻌﺪ اﻟﺘ"ﺤر"ٮﻪ دي اﻟﻤ
Staining of bacteria (two types)
-simple -complex
Simple staining
-Positive (basic )
-Negative (acidic indicate as Nigrocine ,india ink)
Positive simple staining
Tools
1-crystal violet,safranin,methylene blue(M.b)
2-flame
3-slides
4-microscope
5-bacterial suspension
6-Isolation needle
7-tube,gloves,glass dropper and alcohol
Procedure
1-prepare bacterial suspension (by mixing part of bacterial colony and dis H2O)
2-fill this bacterial suspension with glass dropper on slides and distribute the bacterial
suspension with isolation needle on slide
3-then dry it behind the flame
4-then drop it with (crystal violet,safranin,M.b)
5-wait 5 minutes then wash it and dry behind the flame
6-then put the slide under microscope and adjust it 40X
Observation
-Positive staining colors the body of the cell.
Negative simple staining
Tools
1-Nigrocine
2-bacterial suspension
2-bacterial suspension
3-microscope
4-Isolation needle
5-tube,slides,gloves,glass dropper and alcohol
Procedure
1-prepare bacterial suspension (by mixing part of bacterial colony and dis H2O)
2-fill this bacterial suspension with glass dropper on slides with drops of Nigrocine and
distribute the bacterial suspension with isolation needle on slide
3-then dry in the air
4-then put the slide under microscope and adjust it 40X
Observation
-Negative staining colors the background surrounding the cell but not the cell itself. The result is
a dark colored halo around a clear cell.
Positive gram staining of bacteria
Tools
1-crystal Violet
2-iodine
3-microscope
4-safranine
5-Isolation needle
6-tube,slides,gloves,glass dropper and alcohol
Procedure
1-prepare bacterial suspension (by mixing part of bacterial colony and dis H2O)
2-fill this bacterial suspension with glass dropper on slides the dry it behind the flame
3-add crystal violet stain for 1 minute then wash it
4-then add grams of iodine 1 minute then wash it and add alcohol for 30 s then wash it
5-add safranin for 1 minute then wash it then dry it inthe flame
6-then put the slide under microscope and adjust it 40X
Observation
-Purple cell wall appear therefore it is positive staining
Effect of antibiotics on bacterial culture
Tools
1-Antibiotic
2-petri-dishes
3-cotton swap
4-autoclave
5-onion extract
6-Nanoparticles as zno
7-flame
7-flame
Procedure
1-sterile nutrient agar compounds medium into plate sterile on bacteria bacterial growth
2-make it more thicker and solidify it away from the flame
3-turn the dishes 3 times of turnes with small angle and culture bacteria with cotton swap near
to the flame
4-devide the dish to four quarters and make four holes,add anti biotics ,Nano particles and onion
extract
5-then test their sensitivity
6-Inocubate at 37c for 24 h in auto clave
Observation
-Anti-biotics or onion extract or Nano particles make inhibition zone ,calculate it
