General
characters
1-Members of this family are found mostly in the intestinal tract of
man and animals as normal flora or as pathogens
2-They are Gram –ve, relatively small size and non-spore forming
rods
3-Capsule production (Klebsiella pneumoniae, and enterobacter
aerogenes) give rise a large mucoid colonies
4-Members of this family are primarily motile with peritrichous
flagella although some strains are non-motile
5-Several strains possess fimbriae or pili
6-Grows readily onto ordinary media under aerobic and facultative
anaerobic
7-The genera within the family on the basis of biochemical
characteristics as:
•Capacity to ferment specific carbohydrates
•Utilize of citrate
•Indole and urea tests
•Hydrogen sulfide test
8-The genus distinguished only by serological procedures into
numerous serotypes (serovars) using Kauffman-White Scheme
9-All species ferment glucose, nitrate are reduce to nitrite and
oxidase is not produced.
On MacConkey`s media
*Lactose fermenter (Coliform) as E.coli, Citrobacter, Klebsiella
*Intermediate (Late fermentation) (Paracolon):
As Arizona, Serratia, Hafnia
*Lactose not ferment:
As Proteus, Salmonella, Providence, Shigella, Morganella
G.: Escherichia
(E.coli(
Morphology
• Gr-ve bacilli, arranged in single, pairs or groups
• Size: 2-3x0.4-0.6 µ
• Motile by peritrichous flagella
• Capsulated by polysaccharide capsule
• Non sporulated
Gram –ve bacilli
Peritrichous flagella of E.coli
Culture characters
 Aerobic and facultative anaerobic
 Grow on ordinary media, optimum temperature at
37°C (for man and animals), it can grow at wide range
of temperature (20-44)
 Colonies are circular outline, smooth, glistening,
transparent, unpigmented.
 MaCconkey agar: Pink colonies (lactose fermenter)
Selective media for E.coli:
Endomedia , Jaster media Eosin Methylene Blue
Leiven media
MaCconkey agar
Kauffman media
Brillient green media
EMB
E.Coli on MaCconkey agar
E.Coli on EMB media
Pink colonies
Yellow green colonies
Lactose fermenter
Biochemical reactions
Sugar fermentation: glucose and lactose (acid and gas)
Citrate –ve
H2S: -ve
I
Urea hydrolysis: -ve
+
Gelatin liquefaction: +ve
Litmus milk test: +ve (acid and clot)
Indole: +ve
V.P.: -ve
M.R.: +ve
M V C
+
-
-
Indole +ve: Pink ring
Citrate +ve: blue colour
Indole –ve: Yellow ring
Citrate –ve: green colour
M.R. +ve: Red colour
Voges-proskauer +ve: Pink ring
M.R. –ve: Yellow colour
Voges-proskauer –ve: No ring
E.Coli on Triple Sugar
Iron(TSI): Y⁄ Y
Urease test +ve: purple colour
Gas production
Urease test –ve: yellow colour
Antigenic structure
O Ag
Somatic Ag
171 types
It composed of polysaccharide phospholipid- protein complex
Heat stable (not destroyed at 100-120°C)
Not destroyed by alcohol
K Ag
Envelop or capsular Ag
80 types
It composed of polysaccharide
L, B Ags: inactivated by heating at 100°C for an hour
A Ag : inactivated by heating at 121°C for 2.5 hours
H Ag
Flagellar Ag
56 types
It composed of protein called flagellin
F Ag
Fimbrial Ag
K88: F4
K99: F5
It composed of glycoprotein
Heat labile
Acts as adhesion or colonization factors, indicate
virulence
Toxins
Exotoxin
Enterotoxin that composed of heat labile and heat stable
fractions
Endotoxin
Vomition, diarrhea, fever
Colicin
Antibiotic like substance that produced from virulent
strains of E.coli
It has bactericidal effect causing death for M.O. but not
used for treatment as toxic for tissues
Resistanc
e
M.O.can survive for weeks or months in water, faecal
matter especially if away from sun light
Destroyed at 55°C for an hour, 60°C for 20 minutes
(vegetative M.O.)
Relatively susceptible to physical and chemical
agents
Susceptible to lethal action of phenol, cresol and lysol
Pathogenicity
Man
Food poisoning
Factors affecting pathogenicity
Polysaccharide somatic Ag
Fimbrial Ag
Capsular Ag
Enterotoxins, shiga like toxin
Sidrophore (combete with iron of host cell)
Diagnosi
s
Clinical symptom
Microscopical examination
Culture characters
Biochemical reaction
Serological tests
Slide agglutination
Tube agglutination
Coli titer
Minimum amount of fluid or Gram of solid sample that
contain one E.coli cell
Coli index
Amount of E.coli bacterial cells that present in one liter
(fluid) or one Kg (solid) sample
Klebsiella
Morphology
Gr-ve bacilli, arranged in single, pairs or short chains
Size: 1-3x0.5 µ
Non motile
Capsulated by polysaccharide capsule give rise to
mucoid colonies
Non sporulated
Culture characters
 Aerobic and facultative
temperature 37°C
anaerobic,
optimum
 Colonies are circular outline, smooth, glistening,
transparent, unpigmented, 1-4 mm in diameter,
mucoid due to present of capsule but after repeated
subculture, it lost mucoid capsule and colonies
become smaller in size
 On MaCconkey
fermenter)
agar:
Pink
 On blood agar: No haemolysis
colonies
(lactose
Klebsiella mucoid colonies
Klebsiella on EMB
Large mucoid pink to purple
colonies with no metallic
green sheen
Biochemical reactions
 Sugar fermentation: glucose, lactose, sucrose and
salicin (acid and gas)
 Citrate: +ve
 Urea hydrolysis: +ve (slowly after 4 days)
 H2S: -ve
I
M V C
 Indole: -ve
-
-
 Gelatin liquefaction: -ve
 Litmus milk test: -ve
 V.P.: +ve
 M.R.: -ve
+ +
Indole –ve
Citrate +ve
V.P. +ve
M.R. –ve Urease +ve
Species
K.pneumoniae K.rhinoscleromatis
K.ozaenae
K.arerogenous:
Causing:
Some members are present normally as normal flora in intestine
and respiratory tract.
K.Pneumoniae: sever Pneumonea, Urinary tract infection
K.rhinoscleromatis:
rhinoscleroma,
chronic
granulomatous
condition affecting mucous membrane of nose , throat extended to
larynx
K.Ozaenae : atrophic rhinitis, nasal bad odour
K. aerogenous: Urinary tract infection
Shigella
Morphology
Gr -ve small rods
Size: 2-3 x 0.5 µ
Non motile
Non sporulated
Non capsulated
Culture characters
0n MaCconkey media:pale colonies (non lactose fermenter)
Biochemical reactions
Sugar fermentation:
Sucrose non lactose fermenter: subgroups A, B, C
Lactose non sucrose fermenter: subgroup D
Citrate: -ve
Urea: -ve
I
M V C
H2S: -ve
-
+
Indole: -ve
M.R.: +ve
V.P.: -ve
Gelatin liquefaction: -ve
- -
Species
Sh.dysentery
(A)
Sh.flexneri
(B)
Sh.boydii
(C)
Sh. Sonnei
(D)
Causing:
Human bacillary dysentery especially in children
Salmonella
Morphology
Gram-ve short rods, arranged singly, in pairs or groups
Size: 2-4 x 0.5µ
Motile by peritrichous flagella except S.pullorum and
S.gallinarum
Sometimes capsulated (polysaccharide capsule)
Non sporulated
Culture characters
* Aerobic and facultative anaerobic
* Colonies are round, smooth and transperant 2-3mm
For isolation it require
Enrichment media as:
Brillient green agar
Selenite-F- broth
Desoxycholate citrate agar
Tetrathionate broth
Followed by
Solid media
Xylose lysin desoxycholate
(Salmonella Shigella agar
MaCconkey agar
Incubation at 37°C
Incubation at 37°C
18-20 hours
24-48 hours
Non lactose fermenter
Salmonella on XLD media
On MaCconkey agar
Red colonies with black center
Pale colonies
Salmonella 0n Brillient green agar
Yellow to green colonies
Salmonella on SS agar
Biochemical reactions
Sugar fermentation: It ferment sugars with acid and gas
except lactose, sucrose and salicin
Citrate +ve
H2S: +ve
Urea hydrolysis: -ve
Gelatin liquefaction: -ve
Nitrate: +ve
Indole: -ve
V.P.: -ve
M.R.: +ve
I
M V C
-
+
-
+
Citrate +ve
Indole –ve
Urease-ve
M.R. +ve
V.P. -ve
TSI media
**Alkaline (Red colour)
**Black colour
(H2S production)
**Acid (Yellow colour)
Glucose fermenter
**Gases
(due to fermentation)
Antigenic structure
O Ag:
Thermostable
Designated by arabic numbers
H Ag:
Phase I: More specific
Designated by small letters of alphabetic
Phase II: Less specific
Designated by arabic numbers
K or envelope Ag
Vi Ag
M or mucoid Ag
It was found that:
Addition of alcohol destroy H Ag to prepare O Ag
Addition of formalin destroy O Ag to prepare H Ag
Boiling for 15-20 minutes destroy Vi Ag
Kauffman-White scheme:
Classify Salmonella into different O- groups each of
which contain No. of serotypes have common O Ag not
found in the other O- group.
It designated by capital letters from A to Z
Groups A to E nearly contain all Salmonella species that
are important pathogens for man and animal
Identification of serotypes by tube agglutination test
Kauffman-White scheme:
Serotype
Group O Ag
H Ag
Phase I
Phase II
S.paratyphi A
A
1,2,12
a
-
S.paratyphi B
B
1,4,5,12
b
1,2
S.typhimurium
B
1,4,5,12
i
1,2
S.cholera-suis
C
6,7
c
1,5
S.newport
C
6,8
e, h
1,2
S.abortus ovis
B
4,12
c
1,6
S. abortus equi
B
4,12
-
enx
S.enteritidis
D
1,9,12
c ,m
-
S.Pullorum and
S.gallinarum
D
1,9,12
-
-
Pathogenicity
 Classification:
A. Salmonella causing enteric fever
1.
S.typhi
2.
S.paratyphi A
3.
S.paratyphi B
4.
S.paratyphi C
B. Salmonealla causing food poisoning
1.
S.Typhi murium
2.
S.Enteritidis
C. Salmonella causing septicemia
1.
S.cholerae-suis
2.
S.virchow
Enteric fever

Pathogenicity:
Source of infection: Human case and Human
carrier (Urine and stool)
Mode of transmission: contamination food and
water.
• 1. M.O enter body via mouth
payer’s patches of small intestine
into mesenteric Ln, spleen, liver
multiplies in
via lymphatic
this called
incubation period which still (14 days)
• 2. M.O pass to blood and produced bacteremia
with first appearance of symptoms and this called
(First week)
• 3. M.O stimulate immune system to produced Ab
and this called (2nd week).
• 3. M.O disappear from blood to internal
organ, liver, gall bladder, spleen, kidney to be
extracted in intestine again and M.O excreted
in stool and urine this called (3rd week).
• 4. M.O are localized in peyer’s patches with
typical pathology which include Necrosis,
ulceration and proliferation of intestine in
sever case.
Laboratory
Human case:
A. First week
1. sample: (stool)
2. Direct film stain by Gram stain:
G-ve bacilli, motile. Non-spore, non-capsulated
3. culture characters:
Blood culture
5-10ml of venous blood are added to 50 ml of
broth containing 0.5% Na+ taurocholate and
incubation at 37C/24h. And the subculture on:

a.
MacConky’s medium
pale yellow colonies
b.
DCA (desoxy cholate citrate agar)
c.
S-S
d.
XLD (xyline lysine dextrose agar)
pale yellow
pale yellow
pink
colonies
Cloteculture:
5ml of venous blood are left to clot and then clote is
added to 15 ml of bile salt broth the incubation
37C/24h. And subculture

2nd week:
By Widal test
3rd week: sample (stool or urine)
Stool sample riched by different type of M.O so
to detected salmonella must be
5-10 ml of stool is added on (selneite broth)
where kill all organismis except salmonella and
shigella. Incubation 37C/24h and subculture
Human carrier
Sample: stool or urine
 Direct film
 Culture character:
At first: M.O present in gall bladder (cholamate)
Patient give laxative
diarrhea
M.O in
large amount in stool.
Media: 5-10ml of stool+ salenite broth 37C/24h,
subclture.
Urine centerfugation then deposite is taken and
examined

Diagnosi
s
Clinical symptom
Microscopical examination
Culture characters
Biochemical reaction
Serological tests
Slide agglutination
Tube agglutination
Rapid agglutination
Whole blood agglutination