Pathology OSPE

M. Faraz Khalid, M. Ahsan Asif

Batch 2k22

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 Dedicated to the Teacher of Mankind,
Hazrat Muhammad (Peace Be Upon Him)

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  Urease test .............................. 15
Table of Contents  Specific Bacteria.............. 15
(Click on the content to directly access the
respective page) PATHOLOGY SPECIMENS
MICRO APPARATUS........ 5 ...................................... 19
Culture plates (agars) ..... 7
 Nutrient agar ..........................7
 Blood agar ...................................7
 Chocolate agar.....................8
 MacConkey's agar ..........8
 CLED agar......................................9
 SS agar ...............................................9
 LJ agar...............................................10
Biochemical Tests ......... 12
 Urease test...............................12
 Citrate test................................12
 TSI agar...........................................13
 Oxidase test............................14
 Indole test .................................14

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MICROBIOLOGY APPARATUS

Blood culture bottles already contain a liquid, called a

“transport media” (a transport medium can be powder
or liquid), which allows the microbes in the blood to
Uses: for containing samples of *urine *stool *sputum remain viable if there's a delay in transporting the
*semen * vaginal discharge *CSF *fluids (pleural, sample to the lab.
pericardial, peritoneal, synovial)  For urineBoric acid (powder) transport
medium
 For stoolCarry-Blaire transport medium
(except for the stool from a patient of cholera;
in that case Alkaline peptone medium is used)
 For swabAmis transport medium
 For bloodThioglycolate broth (word “broth”
indicates liquid medium) transport medium

Uses: *throat swab *nasal swab *buccal swab

*pharyngeal swab *vaginal swab *conjunctival swab
*ear swab *wound swab *skin swab

Capnic bacteria(CO2 loving):

Neisseria, H. influenzae, Strep pneumoniae

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Uses: a very fine pipette for measuring, transferring, or
injecting very small quantities of liquids, chemicals or
samples.

Uses: this screening device is usually used when a large

population has to be screened for a particular disease
e.g. Hap B, Hap C, Dengue, Malaria. Another important
use is pregnancy screening.

Principle: in ELISA (Enzyme Linked Immunosorbent

assay) test Ag-Ab reaction is observed.
Uses: diagnosing Hepatitis B & C, HIV, Dengue etc.

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Culture plates (agars) 2) Blood agar:

1) Nutrient agar:

 Off-white color
 Every bacteria can grow in it but characters of
bacteria can't be# identified

 Blood agar=nutrient agar+ fresh whole blood (5-

10%)
 No special indicator present. Blood itself is
indicator
 Used to differentiate between hemolytic and
non-hemolytic bacteria
 Alpha hemolysis: partial RBC lysis by
peroxidases (greenish discoloration around
colony bcz of oxidation of Hb to Biliverdin)
 Uses: when a bacteria has been identified,
(Strep.pneumonae, Strep.viridans)
antibiotic sensitivity test can be performed on
this plate  Beta hemolysis: complete lysis of RBCs by
hemolyins, causing clearing of medium around
 The test is called Antibiotic sensitivity test
the colonies (Strep.pyogenes, Strep.agalactiae,
 Antibiogram= agar+ bacterial growth+
Staph.aureus)
antibiotic disc+ result
 Gamma hemolysis: means no hemolysis &
therefore no change around the colonies

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 pg:146)
colonies of other organism” (Levinson
blood agar plate, obscuring the
characteristically swarm over the
“Proteus sp. are very motile and
 This swarming/ wave like/ rippling pattern on
blood agar confirms that the bacteria belongs to Chocolate agar=Nutrient agar+ blood+ heat (80⁰C to
the Proteus species convert Hb to hematin & to inactivate the growth
 This swarming pattern is produced due to: inhibitors)
motility, very rapid multiplication, very short
doubling time  Used for growth of Neisseria (gonorrhea,
 This can be prevented by adding a small amount meningitidis), Strep.pneumonoiae,
of antibiotic to limit the rapid bacterial growth H.influenzae (factors 5 & 10 are added to
 This can be inhibited by: enhance growth)
1)Bile salts 2)Cystine 3)Inc. amount of agar  These 3 organisms also have the distinction of
4)Small amount of specific antibiotic being capneic, encapsulated and causing
meningitis (thus isolated from spinal fluid)

3) Chocolate agar: 4) MacConkey's agar:

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 MacConkey agar= nutrient agar+ lactose+ 6) Salmonella-Shigella (SS) agar:
indicator
 The indicator used is Neutral red indicator
(acidic ph pink; basic phyellow)
 Used to differentiate btw Lactose fermenters
and non lactose fermenter i.e. for Gram-ve rods
 Lactose fermenters (E.coli, Klebsiella,
Enterobacter)acidic environmentpink
colonies
 Non lactose fermenter (Salmonella,Shigella,
Proteus, Pseudomonas)no acidic
environmentagar remains yellow

5) CLED agar:

 “CLED"= Cystine Lactose Electrolyte Deficient

 Cystine & Lactose present; electrolytes absent
 Uses: differentiate between lactose fermenters
and non fermenters in urine sample  Neutral red indicator is present in this medium
 This medium is specific for urine samples which turns red in presence of acidic pH and
thus shows red color in presence of a lactose
 Indicator present in it color Bromothymol blue
fermenter bacteria
(this agar is originally light green to bluish in
color due to this indicator)  H2S producers: Salmonella, Proteus
 Colored pus: Pseudomonas (produces pigments
Pyocyaninblue & Pyoverdinyellow-green)
due to which pus becomes colored

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 7) Löwenstein Jensen agar:

 Use: for culturing Mycobacterium tuberculosis

 The indicator present is Malachite green dye
 Buff colored colonies are seen when
M.tuberculosis is present in the sample

MEDIUM INDICATOR

1 Nutrient agar No indicator

2 Blood agar Blood itself is indicator

3 Chocolate agar Blood itself is indicator

4 MacConkey agar Neutral red

5 CLED agar Bromothymol blue

6 Shigella-Salmonella Neutral red

(SS) agar
7 Löwenstein-Jensen Malachite green
(LJ) agar (Also an inhibitor of
normal
flora present in sputum
sample
8 TSI agar Phenol red

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 CULTURE MEDIA PROPERTIES ORGANISMS IDENTIFIED

1 Blood agar Identification of hemolytic (alpha, beta) and non- Various bacteria
hemolytic (gamma) bacteria
2 Bordet-Gengou Inc. conc. of blood allows growth Bordetella pertussis
medium
3 Chocolate agar Heating the blood inactivated the growth inhibitors N.meningitidis & N.gonorrhoae
from sterile sites
4 Chocolate agar (plus Factor V & X are required for growth Haemophilus influenzae
factor V & X)
5 Chocolate- yeast Inc. conc. of iron and cysteine allows growth Legionella pneumophila
extract
6 CLED agar Specific for urine samples (differentiates b/w lactose E.coli, Klebsiella sp, Proteus sp,
fermenters and non-fermenters) Salmonella sp, P.aeruginosa,
E,fecalis, Lactobacilli
7 Egg yolk agar Lecithinase produced by the organism degrades egg yolk Clostridium perfringens
to produce insoluble precipitates
8 Eosin Methylene Blue Selects against gram +ve bacteria & differentiates b/w Various enteric gram –ve rods
(EMB) agar lactose fermenters and non-fermenters
9 Loeffler medium Primary value of Loeffler medium is in the growth and Corynebacterium sp
morphological characterization of members of the genus
Corynebacterium. This formulation enhances the
formation of metachromatic granules within the cells of
the organisms.
10 Löwenstein-Jensen Selects against gram +ve bacteria in respiratory tract flora M.tuberculosis
agar and contains lipids required for growth
11 MacConkey agar Selects against gram +ve bacteria & differentiates b/w Various enteric gram –ve rods
lactose fermenters and non-fermenters
12 Nutrient agar Growth of all kinds of bacteria No specific bacteria identified

13 Sabouraud’s agar Sabouraud’s Dextrose Agar is primarily used for the Various fungal species
selective cultivation of yeasts, molds
14 Shigella Salmonella Used as a selective and differential medium for the Shigella, some Salmonella sp
(SS) agar isolation of Salmonella and some Shigella species from
clinical and non-clinical specimens
15 Tellurite plate Causes tellurite to become tellurium, which has black Corynebacterium diphtheria
color
16 Thayer-Martin Chocolate agar with antibiotics to inhibit growth of N.gonorrhoeae from non-sterile
medium normal flora sites
17 TSI agar Distinguishes lactose fermenters from non-fermenters Various enteric gram –ve rods
and H2S producers from non-producers

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Biochemical Tests: 2) Citrate test:

1) Urease test:

 Test: Urease test  Test: Citrate test

 Agar: Urease agar  Agar: Citrate agar
 Urease agar= Nutrient agar+ UREA+ Phenol red  Citrate agar= Nutrient agar+ CITRATE+
 Contains Phenol red indicator (acidic pH Bromothymol blue
yellow; basic pHpink)  Indicator used is Bromothymol blue
 Use: for differentiating urease producing (Original colorGreen
(urease enzyme breaks down urea) bacteria If citrate used by bacteria Citrate +ve
from non urease producers bacteria Alkaline mediumcolor turns Blue)
 Urease test +ve organisms (i.e. urease  Another agar containing Bromothymol blue
producers): H.pylori, Proteus indicator CLED agar
 Diseases caused by H.pylori: 1) Duodenal ulcer
Citrate +ve bacteria Citrate -ve bacteria
2) Acid peptic disease 3) Gastritis 4) Tumors Blue Green
[Adenocarcinoma of stomach & MALT Klebsiella E.coli
lymphoma of stomach] Campylobacter Shigella
Tumor associated Associated tumor: Citrobacter Proteus vulgaris
bacteria Salmonella (other than
1 Helicobacter Adenocarcinoma and MALT typhi)
pylori lymphoma of stomach Proteus mirabilis
2 Salmonella typhi Gallbladder carcinoma

3 Streptococcus Colorectal carcinoma

bovis
4 Chlamydia Lung cancer
pneumoniae

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 3) TSI agar (Triple Sugar Iron):
 Helps in differentiation b/w various gram –ve
rods
 Components: 1)Phenol red Indicator
2) Ferrous sulfate
3) 3 sugars: glucose (1/10 conc.
than other sugars), lactose, sucrose

 Parts of agar: *butt (poorly oxygenated)

*slant (well oxygenated)
 Result interpretation:
Result: Possible Possible genera:
visual:
1 No fermentation at all Pseudomonas
 both the slant & butt
remain red

2 Only glucose Shigella, Serratia

fermented small
amount of acid (since
glucose is only 1/10th
conc.) phenol red
turns yellow (in butt)
the acid produced
on/near to slant is
oxidized to CO2 h
H2Oslant will remain
red

3 Lactose/sucrose E.coli, Klebsiella

fermentedlarge
amount of acid
phenol red turns yellow
(both in butt & slant)

(In this picture note gas

bubbles at bottom)

4 If H2S produced Salmonella (tube

ferrous sulfate of the becomes black
agar changes to ferrous but not
sulfide black color of completely),
ferrous sulfide Proteus (lot of
H2S; tube
completely
becomes black)

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4) Oxidase test:

5) Indole test:

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 6) Urease test: Specific Tests for Bacteria:

Chlamydia
Rickettsia
Obligate Intracellular
Ehrlichia
Bacteria
Anaplasma
Mycobacterium leprae

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 E.coli
H.influenzae
Indole +ve Bacteria
Klebsiella
Proteus sp.

(Following picture is just for understanding)

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PATHOLOGY SPECIMENS

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In above picture, dry gangrene (microscopically In above picture, wet gangrene (microscopically
coagulative necrosis) is visible. Black shrunken liquefactive necrosis) can be seen. Edematous wrinkled
appearance with a clear demarcation b/w healthy & skin with no clear demarcation b/w healthy & necrosed
necrosed tissue tissue
Causes of (pure) dry gangrene: Causes of wet gangrene:
*Electric shock *Diabetes mellitus
* Frost bite * Post op
*Puerperal gangrene

(Wet gangrene may have started off as dry gangrene &

later on becomes wet due to super added bacterial
infection)

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Difference between:
Hemosiderin pigmentusually accumulates at site
where the excess iron, released from the breakdown of
RBCs, is stored as hemosiderin in local macrophages
(e.g. hemosiderin pigmentation of bruised skin)
Hemosiderosisoccurs when there is systemic iron
overload usually bcz of excessive hemolysis, IV iron
therapy or multiple blood transfusions (e.g. in
thalassemia pts). In hemosiderosis, usually hepatocytes
are affected first, and the extra iron is stored in them as
hemosiderin
Hemochromatosisa hereditary disorder in which
excessive iron is absorbed from intestine. Multiple
organs are affected by iron accumulation e.g. liver
(hepatomegaly), pancreas (bronze diabetes), gonads
(testicular dystrophy)

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 Types of fibrillar collagen
Types Sites Deficiency
(disease states)
I Hard & soft tissues Osteogenesis
imperfecta (OI),
Ehlers-Danlos
syndrome
II Cartilage, intervertebral Achodrogenesis
discs type II
III Hollow organs, soft Vascular Ehlers-
tissues Danlos syndrome
V Soft tissues, blood Classical Ehlers-
vessels Danlos syndrome
XI Stickler
syndrome

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