00 Sush 2yr Microbio v1.0

2nd year Microbio spot notes
Fb@ <sushrutingawale@gmail.com>
Re-Edited by and Special Thanks to the following people for their

devotion of time to make this pdf happen:
1. KEYUR DESAI
2. AJITH VARRIER
3. YASH CHANDAK
There’s a whole New world to Discover under those two tiny lenses of a Microscope!
Agatha Cristie: “Nothing, I believe, is so full of life under the
microscope as a drop of water from a stagnant pool”
Hence…
4. CHAxxxxxxxxxxxxxxxx
NOTE:
1. This is a crude version of notes….will be re-edited later. A lot of changes are

pending which will be made as and when possible.
2. Purpose of this pdf: The handwritten notes get degraded each year as every
senior writes on them before they are passed on to make a xerox. Hence you
can directly use this eco-friendly online version or even may a Xerox of these
digital notes.
3. DO NOT forward this document to anyone outside Seth GSMC….There are
certain clearance issues that need to be resolved first with the department.
4. FEEDBACK: Requesting your feedback or any mistake
rectification/addition/deletion etc. in the form of WORD DOCUMENT sent to
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INDEX
Sr. No. Sr. No. Sr. No.

1 Culture Media 3
2 Microbiological Tests 12
3 Mycology 15
4 Virology 18
5 Disinfectants, Sterilisation & Lab intruments 20
6 Filters 27
7 Serological & Immunological tests 28
8 Parasitology 31
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1. CULTURE MEDIA
Points to be mentioned about culture media (un-inoculated) in viva:

i. Name
ii. Type of medium
iii. Contents
iv. Method of sterilization (same for most – autoclaving at 121 ° C for 15 mins)
v. Uses
1. NUTRIENT AGAR
Un-inoculated:
a) Nutrient broth Peptone + meat extract + NaCl + water.
Nutrient agar prepared by adding 2% agar to nutrient broth
b) Simple/basal media
c) Uses:
 Growth of non-fastidious bacteria
 To perform antibiotic sensitivity test when Mueller-Hinton agar is not
available
 As a base for preparing blood agar, chocolate agar, etc.
 To demonstrate pigment production. Eg. In Staphylococcus species
Inoculated NA
Organism S. aureus S. albus P. Aeruginosa
Description Golden yellow colonies, oil Whitish, opaque Light whitish colonies with diffuse bluish
paint appearance colonies green pigment (pyocyanin)
Infections Abscesses, sinusitis, Same as S. aureus Nosocomial infections: - Urinary infections
endocarditis, osteomyelitis, following catheterization, eye inf., inf. in
pneumonia, TSS, SSSS, etc. burns, iatrogenic meningitis following
lumbar puncture, etc.
Community acquired: Suppurative otitis
Virulence  Cell wall polysaccharide  Exotoxin A
factors  Teichoic acid  Extracellular enzymes – proteases,
 Capsular polysaccharide elastases, hemolysins
 Protein A  Slime layer
_  Pili
 Clumping factor
 Enzymes – coagulase,
lipase, hyaluronidase,
nuclease
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ANTIBIOTIC SUSCEPTIBILITY TESTS

Kirby Bauer Disc Diffusion Method:
 Agar – Mueller Hinton Agar
 Principle – Antibiotic impregnated in discs spreads radially causing clearing d/t
inhibition of susceptible bacteria.
“Zone of inhibition” formed – proportional to effectiveness of antibiotic
 As per standard zone size recommended by CLSI (Clinical & Laboratory Standards
Institute)
Other methods:
 Stroke’s method – diffusion
 Tahe dilution method
 Plate dilution method (green pigment on ABS test for Pseudomonas)
 Epsilometer test
2. 5 % SHEEP BLOOD AGAR

Un-inoculated:
a) Contents – 5% sheep blood + peptone + agar (+ NaCl + tryptic soya broth)
After autoclaving, cool agar base to 50 °C, then add sterile blood gradually onto the
plate
Viva Q. Why is the temperature adjusted to around 50 °C?
A. Because agar solidifies around 40-45 °C
b) Enriched media and Differential media (but not selective)
c) Uses:
 Isolation of fastidious bacteria
 Growth of most aerobic and anaerobic bacteria and fungi
 To differentiate Gram +ve cocci depending on degree of hemolysis caused
Characteristics Organisms
Blood agar with α – hemolytic colonies Greenish discolouration, zone S. pneumoniae,
of hemolysis 1-2 mm wide with S. viridans,
indefinite margins (partial
hemolysis)
Blood gar with β – hemolytic colonies Colourless zone of hemolysis 2- Most pathogenic
4 mm wide, clear, sharply streptococci and
defined margins (complete staphylococci
hemolysis)
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Note:
 Bacillus, Clostridium and Corynebacterium can also be hemolytic

 Hemolysis should always be checked for in transmitted light (Hold the culture plate
against the light and comment. This may impress the examiner)
 Staphylococci - large, convex, disc-like opaque colonies
Streptococci – small, low convex, semi-transparent colonies, clear zone of hemolysis
 Staph β-hemolysis starts at 37°C and is evident only after chilling conditions, hence
may appear non-hemolytic (hot-cold variety of hemolysis by β-hemolysin)
Proteus on blood agar

 Swarming with concentric rings of growth
 Also seen with C. tetani on blood agar in anaerobic conditions
 Significance: For identification and typing (If two swarming patterns cross,
strains are related, if they only meet a point, strains are partially related)
3. CHOCOLATE AGAR
a) Prepared by heating sheep blood agar to 56 °C to:

 Inactivate certain inhibitors that may be present in SBA for
Neisseria/Hemophilus
 To release essential growth factors – X factor (hemin) and V factor (NAD)
b) Enriched media
c) Use: Growth of fastidious organisms like N. gonorrhoeae, N. meningitides, H.
influenzae, Moraxella sp.
Note:
 α – hemolysis is better appreciated on chocolate agar compared to SBA
 Neisseria may also be grown on:
 New York City agar (also for Mycoplasma and Ureaplasma)
 Thayer Martin medium
 Chaco-Nair egg media
4. MAC-CONKEY AGAR
Un-inoculated:
a) Peptone + lactose (differential component) + agar + sod. Taurochlorate (bile salt -
selective) + neutral red (indicator)
b) Selective medium – inhibits Gram +ve bacteria (d/t presence of bile salts)
Differential medium – to differentiate lactose fermenting and NLF Gram –ve bacilli
Mechanism: Fermentation of lactose by bacilli acid production decrease
in pH of medium below 6.8 colour change to pink
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c) Uses:
 Isolation of organisms like Enterobacteriaceae, Klebsiella
 As an indicator medium (neutral red serves as indicator)
MacConkey Agar with lactose fermenting colonies
 Early fermenters – Klebsiella; forms mucoid colonies d/t capsule formation

 E.coli – non-mucoid colonies
 Late fermenters – Shigella sonnei, Vibrio cholerae, paracolons
MacConkey Agar with NLF:
 Straw coloured colonies (Mention mucoid/non-mucoid)

 Eg. Salmonella, Shigella, Proteus, Pseudomonas
5. ROBERTSON’S COOKED MEAT BROTH
a) Cooked minced meat pieces of ox/beef heart + nutrient broth

b) Enrichment media for anaerobes
Indicator media………(1)
Transport media
Differential media……(2)
Mechanism:
 Unsaturated fatty acids in meat take up oxygen, reaction being catalyzed by
hematin in meat
 Sulphydryl compounds cause reduction in oxidation reduction (OR) potential
 Glutathione and cysteine in meat are reducing agents favouring growth of
anaerobes
(1) Saccharolytic species (eg. C. welchi) turn the medium pink; proteolytics (eg. C.
tetani) turn it black
(2) Differentiate putrefactive and saccharolytic anaerobes
Note:
 Direct inoculation of wound swabs/aspirates is done in RCM at the bottom of tube as

anaerobes grow better in medium depth
 Other reducing substances in RCM:
 Dextrose
 Iron
 Ascorbic acid
Points from the internet:
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 After inoculation, the medium is covered with a layer of sterile paraffin oil to prevent
oxygen entry
 Meat particles are cooked because reducing substances are available more in
denatured proteins
6. TCBS AGAR
a) Thiosulfate citrate bile salt sucrose agar
Contents:
 Yeast extract
 Sodium thiosulfate and sod. Citrate – to inhibit growth of Enterobacteriaceae
 Ox bile - inhibits Gram +ve bacteria
 Sucrose
 Ferric citrate
 Peptone
 Agar
pH: 8.6 – to enhance growth of V. cholera and inhibit other intestinal flora
b) Selective medium – inhibitory to Gram +ve bacteria and most coliforms
Differential medium – differentiates sucrose fermenting (yellow colonies) and non-SF
bacteria (green colonies)
Indicator medium – Bromothymol blue serves as indicator (alkaline – green; neutral – blue;
acidic – yellow)
b) Uses:
 To isolate Vibrio cholera (yellow colonies) and V. parahemolyticus (green
colonies)
 Bacteria producing hydrogen sulfide can be detected…Na thiosulfate acts as a
source of sulfur, which in combination with Fe from Ferric citrate produce black
ferric sulfide
7. LOWENSTEIN - JENSEN MEDIUM

a) Contents:
 Coagulated hens eggs – solidifying agent
 Mineral salt solution
 Malachite green – imparts green colour to medium; inhibits most other bacteria
 Glycerol – favors the growth of MTB; For cultivating M. bovis, pyruvate is used
instead of glycerol
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 Asparagine
Note:
 LJ Medium is contained in Mc Cartney’s culture bottle (30 ml capacity)
 Culture bottle is preferred over petri dish because of long incubation time; reduces
risk of contamination and drying up of medium. Also to prevent infection by
inhalation
b) Selective and enriched media
c) Method of sterilization here is inspissation at 85° C for 30 min for 3 consecutive
days
MTB appears as dry, rough, raised, buff colonies (“buff, rough and tough”) on LJ
Medium in 2-8 weeks when incubated at 37°C
M. bovis appears as flat, smooth, moist white colonies
d) Use:
 Diagnosis of MTB, M. bovis and NTM infections

 To differentiate different species of Mycobacteria (M. leprae does not grow
in LJ medium)
7. CASTANEDA’S CULTURE MEDIUM

Bile broth with an agar slant
Biphasic medium – solid + liquid phase
Advantages: - Refer card shown in practical
Used for isolation of Salmonella and Brucella
8. BLOOD CULTURE BOTTLE WITH HARTLEY’S BROTH

(Brain heart infusion broth)
Blood is added through hole provided above
a) Contents – peptone water, calf brain, cow heart infusion, dextrose
b) Enrichment and transport media
c) Used in lab. Diagnosis of bacteremia/septicemia, brucellosis, typhoid, PUO
Pediatric 2-3 mL blood in 20-30 mL of medium
Adult 5-10 mL blood in 50-100 mL medium
Advantages:
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 Dilution (1:10) of sample occurs so that antibiotics or other inhibitors in blood are
less likely to suppress growth
 Enhanced growth of bacteria present in small numbers and may be difficult to
isolate if solid media are used alone
D.Adv: - Quantification of amount of growth can’t be done
10. THIOGLYCOLLATE BROTH

a) Contents:
 Yeast extract
 Casein
 L-cysteine
 Na thioglycollate (Thioglycollic acid) – reducing agent, maintains low O2 tension
 NaCl – for osmotic equilibrium
 Resazurin – indicator: Turns pink in the presence of O2
 Agar (0.075%) – favors growth of anaerobes by impeding diffusion of oxygen in
medium
 Methylene blue – may also be used as indicator: turns blue in presence of O2
b) Enrichment medium
Differential medium – To differentiate aerobes, facultative anaerobes, obligatory anaerobes,
microaerophilic bacteria
Indicator medium
c) Use:
 Isolation of anaerobes from blood when anaerobic infection is suspected
 To study growth characteristics of various bacteria based on their O2 requirements
Note:
 Medium is boiled before use to eliminate O2 since it’s less soluble at high temp.
 Distribution of types of bacteria:
Obligate anaerobes – bottom
Obligate aerobes – top
Facultative anaerobes – Scattered throughout, more at top
Microaerophilic- just below the top
 If more than 1/3 rd of the medium is pink, it is unsuitable for growth
11. TELLURITE BLOOD AGAR

a) Potassium tellurite selectively inhibits growth of most Gram +ve and Gram –ve
bacteria C. diphtheria and Staphylococcus aureus reduce K-tellurite to metallic
tellurium producing black colonies
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b) Selective and indicator medium
12. XLD AGAR

(Xylose, lysine, deoxycholate agar)
a) Contents (apart from XLD; D – Na deoxycholate)

 Sucrose
 Lactose
 Na thiosulfate
 NaCl
 Phenol red - indicator
 Agar
b) Selective medium – Grows Salmonella and Shigella selectively

Differential medium – Can be used to diff. the two.
 Salmonella produces red colonies with a black center d/t H2S formation;
no black center in Shigella
 Salmonella can ferment xylose, thereby turning the medium yellow
initially; not observed with Shigella
Indicator medium
13. LOEFFLER’S SERUM SLOPE

a) Glucose, horse/beef serum, eggs, heart muscle infusion
b) Enriched media
c) Sterilization by inspissation at 85° C for 30 mins on 3 consecutive days
Growth of C. diphtheriae and mycobacteria is produced
14. MISCELLANEOUS CULTURE MEDIA

A. Mannitol Salt Agar – Growth of S. aureus turns phenol red indicator yellow
B. Alkaline peptone water:
 Enrichment and transport medium
 Isolation of V. cholerae from stool sample
 pH=8.6
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 Incubation period: 6-8 hrs; sample must reach lab within 8 hrs
C. Tetrathione broth – enrichment medium for Salmonella - pinkish ppt.
D. Bile broth – enrichment medium for Salmonella, especially clot culture
E. Selenite F broth (tomato-red colour) :
 F stands for “feces”
 Enrichment medium for Salmonella from stool specimen
 Incubated for 24-48 hrs before being transferred to selective media
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2. MICROBIOLOGICAL TESTS
I. Urease Test
 Medium may be broth/agar slant
 Original colour: yellow; +ve test: colour change to pink
 Differs from TSI in having no butt
 Phenol red indicator
 Principle: Organisms break down urea to release NH3 and CO2. NH3
combines with CO2 and water to form ammonium carbonate which turns the
medium alkaline causing a colour change
 Christensen’s Urease Medium:

-ve test: E.coli, Providencia sp, Salmonella, Shigella
+ve test: Proteus, H. pylori, Ureaplasma, Brucella, Klebsiella (less active)
 Urea breath test for detection of H. pylori infection – radioactive urea

administered, radio-labelled CO2 measured
II. Citrate Test

 Simmon’s citrate medium:
 Na citrate is the sole source of carbon
 Ammonium dihydrogen phosphate is source of nitrogen
 Bromothymol blue – indicator; changes colour from green to blue if pH
>7.5
 Principle: Organisms which can utilize citrate as carbon source cause change
in colour of medium from green to blue…Also, the ammonium dihydrogen
phosphate is converted to NH3 and ammonium Hydroxide, creating an
alkaline environment in the medium
 Citrate +ve (blue): Klebsiella, Enterobacter, Citrobacter, Salmonella (except S.
typhi), Pseudomonas, Acinetobacter
Citrate –ve (green): E. coli, S. typhi
III. Indole Test
 Tests ability of bacteria to convert tryptophan to indole.
Tryptophanase catalyzes reductive deamination of tryptophan to produce
indole and pyruvic acid
 Kovac’s reagent (isoamyl alcohol + para-dimethylaminobenzaldehyde
reacts with indole to form rosindole dye; isoamyl alcohol forms a complex
with dye, causing it to precipitate) is used as indicator
+ve test: Red colour
-ve test: Yellow
 Variation: Ehrlich’s reagent (ethyl alcohol instead of Kovac’s reagent) may be
used when performing the test on non-fermenters and anaerobes
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 +ve: E.coli, V. cholera, P. vulgaris

-ve: Klebsiella, Enterobacter, P. mirabilis, Salmonella, S. aureus
Plate method
IV. Oxidase Test
Kovac’s method
 Tests ability of bacteria to utilize O2 for energy production
 Reaction is d/t cytochrome c oxidase catalyzes oxidation of reduced
cytochrome by oxygen
 Indicator – Tetra methylene p-Phenyl diamine hydrochloride
 +ve test: Maroon/dark purple colour
-ve test: Colourless
 +ve organisms: Pseudomonas, Neisseria, V. cholerae, Moraxella, H.pylori
-ve organisms: Enterobacteriaceae
V. Triple Sugar Iron (TSI) agar – Composite medium

 Components – slant and butt
 Tests ability of bacterium to:
 Ferment sugars (glucose, lactose and sucrose)
 Produce gas (bubbles in the butt)
 Produce H2S
 Contents:
 Lactose - 1%
 Sucrose – 1%
 Glucose – 0.1%
 Sodium thiosulfate
 Ferrous ammonium sulfate
 Phenol red – indicator
 Agar – as solidifying agent
 Peptone
Working:
 If only glucose is fermented, enough acid is produced to turn only the butt yellow;
slant remains red
 If either sucrose or lactose or both are fermented, large amounts of acid are
produced turning both the butt and slant yellow
 If none of the sugars is fermented, both butt and slant remain red. The slant can
become a deeper red-purple (more alkaline) d/t production of ammonia from the
oxidative deamination of amino acids
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 Bacteria producing H2S produce a blackish discolouration d/t ferrous sulfide

formation
[At the slant, there is auto-oxidation of acidic substance, so when acid production is
less as when only glucose is utilized, the small acid production is overcome and the
slant is again neutralized and is red]
Results:
 Alkaline slant/no change in butt (K/NC) i.e. Red/Red = glucose, lactose and sucrose
non-fermenter
 Alkaline slant/Alkaline butt (K/K) i.e. Red/Red = glucose, lactose and sucrose non-
fermenter
 Alkaline slant/acidic butt (K/A); Red/Yellow = glucose fermentation only
 Acidic slant/acidic butt (A/A); Yellow/Yellow = glucose, lactose and/or sucrose
fermenter
Kligler’s iron agar is similar to TSI, only sucrose is absent
Name of the
organisms Slant Butt Gas H2S
Escherichia,
Klebsiella,
Enterobacter Acid (A) Acid (A) Pos (+) Neg (-)
Shigella,
Serratia Alkaline (K) Acid (A) Neg (-) Neg (- )
Salmonella,
Proteus Alkaline (K) Acid (A) Pos (+) Pos (+)
Pseudomonas Alkaline (K) Alkaline (K) Neg (-) Neg (-)
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3. MYCOLOGY
CANDIDA ALBICANS / CRYPTOCOCCUS ON SABOURAUDS AGAR SLANT
CRYPTOCOCCUS (White tennis ball )

Smooth,mucoid,creamy oil paint like colonies on Sabourauds agar slant with yeasty odour.
Diseses : Meningitis in HIV patients
Pulmonary cryptococcosis (mild pneumonia)
Virulent factor : Capsule (Confirmation -capsule seen in India Ink preparation able to grow at
37 degree celsius and hydrolyse urea.
CANDIDA ALBICANS (White convex football)

Non mucoid creamy ,white ,smooth with yeasty odour .
Test - germ tube/Reynold Braude phenomenon.
Germ tube- Candida + human sera >>>> 37 degree celsius for 2 hour
Confirmation - gram stained smear ,Reynolds Braude phenomenon, forms chlamydospores
on corn meal agar art 20 degree celsius.
SABOURAUDS DEXTROSE AGAR WITH GROWTH OF ASPERIGILLUS NIGER (black cottony

growth)
Commonly laboratory contaminant and opportunistic fungus that grows rapidly on SDA
giving black coloured growth due to dark conidiospores.
Diseases : otomycosis
Pulmonary aspergillosis - invasive aspergillosis
Capable of growing within cavities formed within body due to pathogenic
processes to form asperigillus ball (Aspergilloma)
Allergic bronchopulmonary aspergillosis
ASPERGILLUS FUMIGATUS - dark green

ASPERIGILLUS FLAVUS -yellow green
ASPERIGILLUS NIGER -black
SABOURAUDS SLANT WITH MUCOR SPP.(bread fungi)
White cottony growth

Mucormycosis /zygomycosis
Opportunistic sytemic mycosis -
URT / lung infection in DM ....thrombosis due to invasion of vessels
Page | 15
Wooly growth till 3/4th of TT

may be filling the entire tube
White gray like spider net
SABOURAUDS MEDIUM (UNINOCULATED)
Gram/ plane yellow coloured

Selective medium for growing fungi because increased glucose concentration and acidic pH
Fungi grow on this in 2-30 days either at room temperature or at 37 degrees celsius or both
depending on the type kept for at least one month .
Content : 4 % dextrose
Agar
Antibiotics (Cycloheximide - to prevent saprophytic fungi eg. Aspergillus
Method os sterilisation : autoclaving at 121 degrees celsius for 15 minutes
Use : Observe slide - colony morphology
Reverse slide - pigmentation
SDA is slant because
-media may dry off in plate
- prevent infection by spore inhalation
-slow growing (1 month )
SLIDES OF MYCOLOGY :
CANDIDA ALBICANS
Gram positive yeast like candida
Diseases : Cutaneous - Intertrigo (papular erythematpus lesions )
Mucosal- Vaginal candidiasis,oral thrush
Intestinal candidiasis
Others - Systemic ;Bronchopulmonary septicemia endocarditis meningitis
Predisposing factors : diabetes prolonged , immunosuppression ,oral antibiotic therapy
Virulence test :germ tube test
Virulence factor : Formation of pseudohyphae to colonise and invade tissues
RHINOSPORIDIUM SEEBERI
Culture -only fungus that cannot be cultivated
Diseases : Rhinosporidiosis -chronic granulomatous disease
Nasal polyps ( common in swimmers ) shows fungal spherules with thousand
Page | 16
endospores .
Mode of infection : From stagnant water /aquatic life
L.P.C.B. MOUNT OF MUCOR / RHIZOPUS

Aseptate mucor hyphae and sac like sporangia bearing sporangiospores .
Sporangia are borne on sporangiophores with ends in columella near sporangia.
L.P.C.B. MOUNT OF ASPERIGILLUS

MUCOR ASPERIGILLUS
Aseptate Septate hyphae
3/4th or 1/2 of the fruiting body Fruiting body is complete sphere
Showing the conidiophores which enlarges into vesicle . The vesicles bear short sterigmata (
phialides) which in turn bear chains of conidia .
L.P.C.B. MOUNT OF MACROCONIDIA OF MICROSPORUM /TRICHOPHYTON

Infections caused : tinea ( ringworm ) dermatophytosis
Eg. Tinea corporis , capitis , pedis , etc.
Skin is mostly affected .
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4. VIROLOGY
1) LEIGHTANT TUBE :
To see cytopathic effect produced by viruses
1/2 potion is flat .
Primary isolation of virus from the sample .
Incubation : a) in a sloped horizontal position ( stationary culture )
b) rolled in special roller drum to provide better aeration
Media kept in it : Hanks medium
Eagles medium
Indicator : Phenol red
2) POCKS ON CHORIOALLANTOIC MEMBRANE

1 virion = 1 pock ( lesion produced by virus)
One of the best method of cultivation of viruses using embryonated hens egg
Pocks are formed by - Variola and Vaccinia
Variola Vaccinia
Small ,shiny,white,pearly,convex Larger, irregular ,flat ,greyish
Non necrotic , non hemorrhagic lesion. Necrotic , hemorrhagic lesion.
Cooling temperature - 35 degrees celsius. 41 degree celsius .
Pock counting can be used for assay of pock forming viruses such as variola and vaccinia .
Allantoic cavity : influenza some paramyxovirus
Amniotic sac : influenza
Yolk sac : Chlamydia and ricketssiae
Other uses : Yellow fever vaccine
Rabies ( flury ) vaccine
3) TISSUE CULTURE BOTTLE

The cells adhere to the glass surface from cell suspension and on inoculation , divide to form
confluent monolayer sheet of cells
Media : Hanks medium , Eagles medium , RPMI medium
MAJOR INGREDIENTS - amino acids , vitamins , salts ,sugars ,bicarbonate buffering system, 5
%carbondioxide , 5 % calf serum , antibiotics balanced salt solution , protein supplement .
METHODS OF STERILISATION - autoclave @ 121 degrees celsius for 15 minutes
USES : To see opacities / plaques production by plaques assay
Harvesting viruses
Large quantities for antigen production : vaccine preparation
4) NEGRI BODIES ( RABIES )
Imp - Laboratory diagnosis of viral infection
Intracytoplasmic inclusion bodies
Stained by Seller's technique ( basic fuschin + methylene blue)
Staining - Giemsa , eosin , methylene blue
Page | 18
Acidophilic - Rabies
Basophilic - Adenovirus , CMV ( intranuclear)
5) EGG
It contain live embryo ( 5 - 7 days old ) and is very handy in cultivating viruses , chlamydia ,
rickettsia .
Candling is used to know where embryo in egg is living or not
Uses : preparation of vaccine , culture of viruses , counting of virus.
Page | 19
5. DISINFECTION & STERILIZATION & LAB INSTRUMENTS

1) Antiseptics
 Spirit, chlorhexidine, betadine, tincture iodine, liquid soap
 Definition : Antiseptic is one used for prevention of infection usually by
inhibiting the growth of bacteria in wounds and tissues
 Chlorhexidine : Thermometer and cheatte forceps
 Tincture iodine : iodine(2.5%) + potassium iodide(2.5%) in alcohol(90%)
Skin disinfectant
Application in microbiology
Use of purpose
Indication for use
2) Disinfectants
 Lysol(2-5%), glutaraldehyde(2%), formalin(4%), Na hypochlorite(7-5%)
 Lysol : for sharp instruments
 Glutaraldehyde : Bronchoscope, endoscope, cystoscope, corrugated rubber
anaesthetic tube
Face mask, plastic endotracheal tubes, catheter, instruments, polythene
tubing
 Formalin : preserve anatomical specimen, destroy anthrax spores, sterilise
clean metal instrument(10% formalin + 0.5% Na hypochlorite)
Heat sensitive catheter
 Na hypochlorite : 1% handwash, 2% for disinfecting workplace, liquid waste,
sharps, blood spill, 5% for disinfecting soiled linen and clothes
Mode of action : Bactericidal
3) Needle cutter device

Uses : 2% Na hypochlorite for remaining part of needle
Importance
4) Leak proof, puncture proof container(Discard can)

 White coloured
 Identify
 Uses
 Chemical kept in container is Na hypochlorite(2%)
Page | 20
5) Red bag / yellow bag / black bag

Government of India, Ministry of Environment, forest and climate change ( modified
in March 2016…BMW guidelines)
 Red bag : infectious waste, solids like I.V sets, blood, catheter, microbiological
and biochemical waste
Gloves
Autoclave / chemical treatment
 Yellow bag : Human anatomical waste
Soiled waste, blood bags, mask, cotton (according to new guidelines)
Incineration / deep burial
Chlorine containing compounds should not be incinerated
By 2018, blood bags should be chlorine free
 Black bag : non infectious waste
Paper waste disposed in secure land fills
 Blue bag : for plastic waste, empty vials / ampoules
6) Autoclave
 Like pressure cooker
 121oC at 15 lbs for 15 min
 Sterilisation by moist heat
 For culture media, gowns, masks, dressings, suture material, pharmaceutical
products
 126oC for 10 min, 134oC for 3 min
 Sterilising control : Indicator(chemical / biological)
a)Chemical : Browne’s tube
Bowie dick tapes to test the presence of vacuum
Chemical dyes, chemicals with known melting point
b)Biological : Spores of Bacillus steaerothermophilus
 Principle : latent heat, vacuum, penetrating power
 Precautions :
a) Discharge tap is opened when atmospheric pressure is equal to pressure
inside autoclave, otherwise explosion of boiling liquid media.
b) All air present initially should be removed through discharge tap
7) Hot air oven

 Principle : Protein denaturation, oxidative damage, toxic effect from
electrolytic imbalance
 Uses : glassware, forceps, scissors, scalpels, all glass syringes, swabs, some
pharmaceutical products like liquid paraffin, dusting powder, fats and grease
Page | 21
 160oC for 2 hours

 Indicator : Non toxigenic strain of Clostridium tetani / Bacillus subtilis var
niger
Physical + chemical ( Browne’s tube – Green spot)
Q) Classification of methods of sterilisation and disinfection
8) Water bath
 Sterilisation by moist heat at temp <100oC
 Uses : Serum or body fluids containing coagulable proteins
 56oC for 1 hour on several successive days(open from above)
 Vaccine bath for inactivation of bacterial vaccines : 60oC for 1 hour
 ? Uses in serology tests
9) Inspissator
 80-85oC for half an hour on 3 successive days
 Uses : Media like Lowenstein Jensen, Loeffler’s serum(egg and serum
containing media)
 Slant is because these media are required as sloped(VIVA)
 Glass lid covered
 Principle : Similar to tyndallisation
 LJ medium : Asparagine, malachite green, mineral salts, hen’s egg, glycerol
10) Mac Intosh Filde’s Jar

 Application : For creating anaerobic atmosphere
 Made of steel
 Catalyst : Alumina pellets coated with palladium
 Container is evacuated and then filled with N2-CO2 mixture
2H2 + O2 pallidium 2H2O
 Methylene blue used as an indicator (P. aeruginosa is a strict aerobe, hence
does not grow in oxygen absence)
 Risk of explosion : Asbestos carries risk of explosion so replaced by alumina
pellets
 Methylene blue turns blue in oxygen presence(colourless in oxygen absence)
 Anaerobes : Peptococcus, peptostreptococcus
B. fragilis, clostridia
Fusobacterium
Treponema, Borrelia
Actinomycetes
Page | 22
11) Oil used for microscopy

 Cedar wood oil(costly; used in KEM)
 Alternative : liquid paraffin, glycerol
 To reduce error of refraction, to increase resolving power
12) Candle jar

 Used for culture of capnophilic organisms
 Provide 5-10% carbon dioxide atmosphere
 Examples : Neisseria, Hemophilus influenza, Brucella abortus, Staph aureus,
Strep pyogenes, Pneumococcus
13) Vaccine vials

 Duration of protection ?
 OPV(10years) : Live attenuated, oral, 2-4 drops
5 doses(0,6,10,14,18 weeks)
Local + CMI
 TT(6years) : Toxoid vaccine, 0.5 ml
Deep IM on lateral aspect of thigh
Artificially acquired immunity
 DPT(3 years) : combined vaccine, 0.5 ml
Toxoid + killed
Deep IM on lateral aspect of thigh
Artificially acquired immunity
 BCG : live attenuated
<1 month : 0.05 ml
>1 month : 0.1 ml
Intradermal just above insertion of deltoid muscle
Duration of protection : 15-20 years
 HBV : 1ml at interval of 0,1,6 month for adults
0.5 ml at interval of 0,1,6 month for children
Plasma derived
Killed recombinant subunit vaccine
IM on lateral aspect of thigh
 MMR : Live attenuated vaccine given subcutaneously
14) Cotton swab in a test tube

 Sterile test tube with cotton swab
 Sterilisation by autoclave, hot air oven(test tube glass)
Page | 23
 Cotton fibres of swab are detrimental for some organism(e.g. Neisseria

gonorrhoea), for which chemically inert material swabs(charcoal coated,
alginate coated, Dacron, Rayon) are used
 Used in ABS test : to do lawn culture, collection of variety of clinical material
for microbiological processing named after use according to material
collected or site(e.g. nasal swab, throat swab)
15) Sterile petri dish

 9cm in diameter
 Sterilisation by autoclave(culture), hot air oven(glass)
 Use : For holding culture, media, to collect sample(especially fungal sample)
16) Sterile test tube

 For collection of urine to carry out biochemical tests
 Autoclave, hot air oven
17) Cavity slide / slide with depression

 For hanging drop preparation to see motility of organism like V. Cholera
 V. cholera will show daring motility
 Seen at periphery(where oxygen is available) VIVA
18) West’s postnasal swab

 Used when there is unproductive cough
e.g. Pertussis
 For detecting meningococcal carriers
19) NIH swab(National Institute of Health) VIVA

 Has a label, cellophane swab, glass rod and rubber band
 For collection of eggs of Enterobius vermicularis form perianal skin
20) Tuberculin syringe

Uses :
a) Tuberculin test for Mycobacteria(Montoux)
b) Casoni test for E. Granulosus
c) Histoplasmin test
d) Trichophytin test
e) Test dose of penicillin(intradermal)
f) Small volume of injection of culture into experimental animals
26 no. Needle
Calibre : 0-1 ml
Page | 24
Sterilisation by autoclave, hot air oven
21) Sputum collection bottle / urine bottle
Uses : Collection of sputum, urine, other body fluids in bacterial pneumonia, TB

Glass - raised plastic – disposable
Wide mouth, screw capped container
Sterilisation : Ethylene oxide / gamma radiation
22) Nichrome wire loop / Bacteriological loop

 Uses :
a) Plating and striking on agar
b) Isolation of bacteria, isolating poor culture from specimen
c) Can be used in semiquantitation of colony count especially in UTI
d) To carry out biochemical tests(e.g. String test for cholera)
 Sterilisation by Bunsen burner flame
 Nichrome heats and cools rapidly
 Platinum can also be used but expensive
 Diameter :
4 mm – 0.01 ml of urine
2 mm – 0.001 ml of urine
23) Syringe, needle and rubber cork

 To collect sample in anaerobic infection
 From deep seated abscess, lesion, gas gangrene
 Method of anaerobic sample collection so that organisms are not exposed to
oxygen
 Collection of pus for isolation of anaerobic organisms
24) Sterile syringe with needle

For blood collection, sending anaerobic specimens, etc
25) Vial / Plain bulb
For all the “serum” studies
26) Gas pack system(transparent jar)

 Chemical present in sachet
 H2O + chemical
Page | 25
 2H2 + O2 cold catalysis 2H2O

 Anaerobiosis
 Chemical : Na borohydrate(NaBH4) + NaHCO3 + cobalt chloride + citric acid
 Water and carbon dioxide are released on addition of water
Use : To create anaerobic environment for organism like Neisseria
Page | 26
7. FILTERS
1) Seitz filter (Asbestos filter)
 Made up of asbestos(magnesium silicate)
 To sterilise heat labile substances like toxin, antibiotic solution, blood
products, hydatid fluid, etc
 Sterilised by autoclave
 Single use / disposable
 High absorbing capacity / tends to alkalanise liquids
 Has carcinogenic potential
 Examples of asbestos filters : Seitz and Sterimat and Carlson filter
2) Sintered glass filter

 Isolation of bacteria from sera and body fluids for culture
 Composed of fused ground glass particles(finely powdered glass particles)
 Sterilised by autoclave
3) Candle filter / Berkefeld filter

 Use : Water purification(large scale)
 Unglazed ceramic : Chamberland, Doulton
Porcelain filter
Kaolin(hydrous aluminium silicate)
 Diatomaceous earth : Berkefeld, Mandler
Keiselguhr, asbestos, P.O.P
Uses of filters :
a) Removing bacteria from heat labile liquids – sera, sugar solution,
antibiotic solution
b) Obtain bacteria free filtrates for virus isolation
c) Concentrate bacteria e.g. testing water sample for cholera vibrio or
typhoid bacilli
d) Obtain bacterial toxins by passing cultures through filters
Page | 27
7. SEROLOGICAL & IMMUNOLOGICAL TESTS
1) Latex agglutination tile

 For latex agglutination test
 For detection of hepatitis B antigen(HBsAg), rota virus antigen,
rheumatoid arthritis (RA) factor, Antistreptolysin O(ASO) test, C-reactive
protein test(CRP) test
 Passive agglutination test
2) Radial Immuno Diffusion Assay(RIDA) slide

 Single diffusion in double dimension occurs
 Uses :
a) Estimation of IgG, IgM, IgA and complements in sera
b )For screening antibodies to influenza
c) Detection of morphine, phenobarbitone
 Disadvantages : Carcinogenic
3) VDRL tile(Venereal Disease Research Laboratory)

 Slide flocculation test for diagnosis of syphilis
 Has 12 depression (3x4)
 Transparent because microagglutination has to be checked under microscope
(VIVA)
 Qualitative test, may be semiquantified to rule out false positive
 Antigen used is purified lipid extract of beef heart(cardiolipin - diphosphatidyl
glycerol) or alcoholic extract of ox heart tissue to which lecithin and
cholesterol are added
 Serum is initially inactivated by heat at 56oC for 2 hours
 Tests for reaginAb :
 Drop of serum + drop of Ag Rotated (4 min)
 Clumps under low power Positive
 Biological false positive :
Acute : (<6 months) acute infection, inflammation, injury
Chronic : (>6 months) SLE, collagen disease, leprosy, malaria, relapsing fever,
infection mononucleosis, hepatitis
 Other STS are Kahn test, RPR card test and Wassermann test
4) Microtitre plate
 For ELISA / IHA / HA / antibiotic sensitivity testing(AMST) for measuring
MIC(by micro broth dilution method) amongst antibiotics
 For detecting antigens of HBsAg, HBeAg, P24Ag, rota virus Ag, dengue
Page | 28
 For detecting antibodies of anti HIV, anti HBc, toxoplasma, CMV, herpes,
leptospira, E. Histolytica, neurocysticercosis(NCC)
 For detecting food toxins (e.g. aflatoxin), food adulterants
 12x8 = 96 depressions
 Made up of polypropylene or polystyrene
5) Widal test rack

 Antibody is detected
 Tube agglutination test
 Significant titre : O – 1:100(IgM); H – 1:200(IgG)
 Widal test – overnight incubation at 37oC in water bath
 Test positive at end of first week infection
 Also positive in case of prior infection or immunisation
 7x4 test tube stand with 28 test tubes
 Used for diagnosis of enteric fever using Salmonella typhi O and H antigen
and S. paratyphi AH and BH antigen
 Dreyer’s tubes(conical) for H agglutination – Pink colour
 Felix tubes (round bottom) for O agglutination – Blue colour
 Ag H – S typhi, S paratyphi A & B(species specific)
 Ag O – S typhi (genus specific)
6) Tridot test HIV

 Principle : Immuno-filtration
 Immunoconcentration vertical flow
 Cylinder / cassette ELISA
 On nitrocellulose membrane
7) Lepto tek dri dot

 For leptospirosis
 Rapid IgM dipstick?
 Results in 30 sec
 If positive – agglutination
 If negative – no agglutination
8) Rapid dengue IgM / IgG test
Page | 29
 Principle : Antibody in serum reacts with coated antigen to produce visible

reaction.
 Based on immunochromatography
 Results are available in 20 min. They appear in the form of coloured bands
 Test comes positive after 7 days of infection or after 4 days of onset of illness
 Antibody is detected
Q) Which antigens are coated on filter paper strips for diagnosis of HIV infection?
P24 Ag
gag – p55 – p15, p18, p24
env – gp160 – gp120, gp41
pol – p31, p51, p66
Page | 30
8. PARASITOLOGY
SPECIMENS
1. Ascaris female worm in intestine . (Read textbook)

2. Ascaris female worm in bottle.
3.Ascaris male worm in appendix.
-female worms don't have curved ends .
Infective stage - embryonated egg containing rhabditiform larvae
Route of entry - ingestion
Complications - Intusussception ,ectopic , ascariasis / appendicitis, jaundice, pancreatitis ,
intestinal obstruction
Disease caused : 1) PEM
2) Loefflers syndrome
4. Calcified Guinea worm specimen -
Dracunculus medinensis
Diseases - guinea worm disease
Sex - female
Mode of entry - by ingestion of water containing cyclope
Incubation period : 6months - 1 year
Intermediate host : cyclopes
5. Diphyllobothrium Latum Full worm ( Fish tapeworm ,longest tapeworm ,

multisegmented )
Sex: female ,central genital pore with hermaphrodite rosette uterus .
Breadth of each segment / strobila > length
Mode of entry- undercooked raw fish flesh
Definitive host - Man
First intermediate - Cyclops ( copepods)
Second intermediate- freshwater fishes
Infective stage: plerocercoid larvae
Infections - usually asymptomatic , megaloblastic anaemia
6. Multiple nematode specimen

Approximate sizes of all
A) guinea worm 50 - 80 mm
B) whipworm 30-50 mm
C) Roundworm 15 - 30 mm
D) hookworm 10 mm
E) pinworm 10mm
Page | 31
SLIDES PARASITOLOGY
BINOCULAR MICROSCOPE
1) Gravid segment of taenia saginata ( beef tapeworm)

Infections caused : Abdominal discomfort , diarrhoea , constipation , obstruction
Definitive host : man
Intermediate host : cattle
Infective stage : cysticercus bovis in muscle ( beef)
Size of adult worm : 4-6 metres
Diagram of egg- same as for granulosus
Length > breadth
2) Fasciola hepatica (liver fluke)

Intermediate host 1st - snail
2nd - aquatic vegetation
Route of entry -ingestion of aquatic plantsv
Infective stage - Metacercaria
3)Tse-Tse fly
( 2-4) are seen
Disease : Sleeping sickness ,trypansomiosis ( west and east African)
Infective agent : Trypanosoma brucei
Mode of transmission - bite of tse tse fly .
4) Sand fly ( Phlebotonius argeutipes)

Diseases : kala azar ( Visceral leishmaniasis,dum dum fever)
Infective agent : Leishmania donovani
Page | 32
LIGHT MICROSCOPE
1) Plasmodium falciparum ring form

(Note: 1 RBC contain more than 1 chromatin dot )
Diseases caused : Malaria
DH - female anopheles mosquitoes
Intermediate host - man
Complication cerbral malaria,algid malaria , septicaemia malaria
2) Gametocyte of Plasmodium falciparum

IH - Man
DH -female anopheles mosquito
Infective stage - sporozoite
3)Plasmodium vivax - schizonts

Each RBC contain single ring form
4)Trypanosomes- form of T. Brucei

Disease caused - Trypanosomiasis
Sleeping sickness
Mode of transmission- bite of tse tse fly
Various morphological stages -metacyclic form ,
Epimastogote (short stumpy form , long slender form ,intermediate form)
Site affected : blood ,lymphatic system , lymph nodes, reticuloendothelial system .
5) Microfilaria in peripheral smear

IH - Culex mosquito ,anopheles ,aedes
DH - man
System affected -lymphatic
Complications - adult filariasis,topical pulmonary eosinophilia,elephantiasis,scrotal edema .
Specimen: Blood at night due to nocturnal periodicity
6) Cyclops
Drucunculus medenesis causing guinea worm disease
7) Rat flea ( Xenopsylla cheopsis )

Wingless ,bilaterally compressed ,conical head without neck
Thorax has 3pairs of strong legs ,abdomen has 10 segments .
Diseases - endemic typhus ,plague,
Page | 33
8) Head louse (Mostly not kept for spot)
9) Body louse ( peduculis corporis)

Wingless ectoparasite with a pair of 5 jointed antennae .Three pairs of leg with claws . Body
flattened dorsoventrally
Abdomen has 9 segments
Epidemic typhus fever (trench fever)
R.prowazekki epidemic
Relapsing fever : Borellia recurreutis
10) Toxoplasma culture smear

Infection : toxoplasmosis
Route of infection: oral
Sites affected: CNS
Epidermic typhus : R.prowazekii - body louse

Endemic typhus R.typhi - Rat flea
Epidemic relapsing fever - B. Recurreutics - body loue b
Endemic relapsing fever -B . Recurreutics - ticks
11) Head part of T. Saginata

4 suckers without hooklet
Unarmed beef tapeworm
Infection caused: GIT(as noted earlier)
IH :cattle
DH : man
Size of adult worm :4 - 6 metres
Infective form : cysticercus bovis in beef
Entry : undercooked beef
Diagram of egg : same as E. granulosus
12) Head part of T. Solium (pork tapeworm)

Infections caused : neurocysticercosis , cysticercosis ( subcutaneous
muscles, eye , git symptoms)
4 suckers with hooklets
Double row of hooklets (armed tapeworm )
Infective stage: cysticercus cellulosae in muscles
Entry : undercooked pork , egg with embryo for cysticercosis
Size of adult worm : 2-4 metres
Diagram of egg : Same as for E. granulosus
Page | 34
13) T. Spiralis coiled larvae

DH : man
IH : NO
Diseases produced : trichenellosis
GIT ( mucositis ) ( stage of invasion )
Fever, periorbital edema, hemorrhages in nail beds
Sclera , mucous membranes ( stage of migration ) .
Also transient myocarditis , CNS infection
Size : male : 1.4-1.6 metres
Female : 3-4 metres
14) D.latum gravid segment ( fish tapeworm )

Width > Length
Infections caused rarely Megaloblastic anemia ( worm that is close to stomach uses vitamin
B12 ) , fatigue , weakness , diarrhoea , obstruction
DH: man
IH : Cyclops , fish
Infective stage : plerocercoid larvae
Size of adult worm : 10 metres ( longest tapeworm )
15) T. Trichura ( male worm)

DH : man
IH : no(NO INTERMEDIATE HOST FOR INTESTINAL WORM )
Diseases - Whipworm dysentery
Complication : Iron deficiency anemia
Rectal prolapse
Male size : 30 -45 mm
Female size : 35 -45 mm
Mode of entry : ingestion of embryonated egg( infective stage)
16) E. Vermicularis ( female Pinworm / threadworm / seat worm)

Showing bilobed oesophagus ( double bulb oesophagus )
Long tapering end
Diseases caused : pinworm enterobiasis
Male size : 2-4 mm
Female size : 8-12 mm
Mode of entry : Ingestion of eggs
Eggs : nonbile stained
Floats in salt solution, asymmetrical
17)Ancylostoma duodenale ( hookworm )

Male worm with copulatory bursa having 13 chitinous rays .
Infection caused : Ancylostoma dermatitis / ground itch
Page | 35
Pulmonary lesion ( bronchitis / bronchopneumonia)

Hookworm disease
Infective stage : Filariform larvae
Male size : 5-11 mm
Female size : 9-13mm
Complications: anaemia ,hypoproteinemia , digestive disturbances and
Retarded development .
Mode of entry: through skin
18) Filariform larvae of A. Duodenale

This is the infective stage
19)Egg of Schistosoma japonicum in intestine

Oval shape and lateral knob
Route of entry : skin penetration by bifid tailed cercaria larva ( from water)
Infections : dermatitis at the site of entry
Lungs cough and mild fever
Acute schistosomiasis- katayama fever
Granulomas or pseudotubercles at the site of eggs
Hepatoslenomegaly
Female genital schistosomiasis
Cerebral involvement- seizures ,paraesthesia , poor vision
20) Egg of S.hematobium in urine deposits

Route of entry : skin penetration
Part affected: bladder
Class : fluke , trematode
IH : freewater snail
DH : human beings
Infections caused : same as described above.
Page | 36

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2nd year Microbio spot notes

Re-Edited by and Special Thanks to the following people for their

1. This is a crude version of notes….will be re-edited later. A lot of changes are

Sr. No. Sr. No. Sr. No.

Points to be mentioned about culture media (un-inoculated) in viva:

Organism S. aureus S. albus P. Aeruginosa

ANTIBIOTIC SUSCEPTIBILITY TESTS

2. 5 % SHEEP BLOOD AGAR

 Bacillus, Clostridium and Corynebacterium can also be hemolytic

Proteus on blood agar

a) Prepared by heating sheep blood agar to 56 °C to:

 Early fermenters – Klebsiella; forms mucoid colonies d/t capsule formation

 Straw coloured colonies (Mention mucoid/non-mucoid)

5. ROBERTSON’S COOKED MEAT BROTH

a) Cooked minced meat pieces of ox/beef heart + nutrient broth

 Direct inoculation of wound swabs/aspirates is done in RCM at the bottom of tube as

a) Thiosulfate citrate bile salt sucrose agar

7. LOWENSTEIN - JENSEN MEDIUM

 Diagnosis of MTB, M. bovis and NTM infections

7. CASTANEDA’S CULTURE MEDIUM

8. BLOOD CULTURE BOTTLE WITH HARTLEY’S BROTH

10. THIOGLYCOLLATE BROTH

11. TELLURITE BLOOD AGAR

b) Selective and indicator medium

12. XLD AGAR

a) Contents (apart from XLD; D – Na deoxycholate)

b) Selective medium – Grows Salmonella and Shigella selectively

13. LOEFFLER’S SERUM SLOPE

14. MISCELLANEOUS CULTURE MEDIA

 Christensen’s Urease Medium:

 Urea breath test for detection of H. pylori infection – radioactive urea

II. Citrate Test

 +ve: E.coli, V. cholera, P. vulgaris

V. Triple Sugar Iron (TSI) agar – Composite medium

 Bacteria producing H2S produce a blackish discolouration d/t ferrous sulfide

Kligler’s iron agar is similar to TSI, only sucrose is absent

Pseudomonas Alkaline (K) Alkaline (K) Neg (-) Neg (-)

CANDIDA ALBICANS / CRYPTOCOCCUS ON SABOURAUDS AGAR SLANT

CRYPTOCOCCUS (White tennis ball )

CANDIDA ALBICANS (White convex football)

SABOURAUDS DEXTROSE AGAR WITH GROWTH OF ASPERIGILLUS NIGER (black cottony

ASPERGILLUS FUMIGATUS - dark green

SABOURAUDS SLANT WITH MUCOR SPP.(bread fungi)

White cottony growth

Wooly growth till 3/4th of TT

SABOURAUDS MEDIUM (UNINOCULATED)

Gram/ plane yellow coloured

L.P.C.B. MOUNT OF MUCOR / RHIZOPUS

L.P.C.B. MOUNT OF ASPERIGILLUS

L.P.C.B. MOUNT OF MACROCONIDIA OF MICROSPORUM /TRICHOPHYTON

2) POCKS ON CHORIOALLANTOIC MEMBRANE

3) TISSUE CULTURE BOTTLE

5. DISINFECTION & STERILIZATION & LAB INSTRUMENTS

3) Needle cutter device

4) Leak proof, puncture proof container(Discard can)

5) Red bag / yellow bag / black bag

7) Hot air oven

 160oC for 2 hours

Q) Classification of methods of sterilisation and disinfection

10) Mac Intosh Filde’s Jar

11) Oil used for microscopy

12) Candle jar

13) Vaccine vials