ENTEROBACTERIACEAE

ENTEROBACTERIACEAE
Gram negative, non sporeforming, facultative anaerobe
bacilli
Most are normal flora of GIT except Salmonella,
Shigella, Yersinia
Catalase (+) except Shigella dysenteriae
Motile with peritrichous flagella except Klebsiella,
Shigella and Yersinia
Non encapsulated except Klebsiella and Enterobacter
All members ferment glucose and reduce nitrate to
nitrite
ANTIGEN DETERMINANTS
Used for serological identifications

1. SOMATIC O ANTIGEN
- heat stable
- located in the cell wall
- for E. coli and Shigella serotyping
2. FLAGELLAR H ANTIGEN
- heat labile
- found in the flagellum
- for Salmonella serotyping
3. CAPSULAR K ANTIGEN
- heat labile polysaccharide
- covers the O antigen
-found as K
1
antigen of E. coli and Vi antigen of S.
enterica subsp. Enterica serotype Typhi


ESCHERICHIA COLI
Part of the normal bowel flora of humans and may also
inhabit female genital tract
It is a primary marker of fecal contamination in water
purification
Leading cause of nosocomial infection UTI
Exhibits green metallic sheen on EMB
Virulence factors: endotoxin, common pili, K1 antigen
TSI: a/a + -
IMViC: ++--
E. coli strains:
a. Enteropathogenic E. coli (EPEC)
b. Enterotoxigenic E. coli (ETEC)
c. Enteroinvasive E. coli (EIEC)
d. Enterohemorrhagic E. coli (EHEC)
STRAIN DESCRIPTION INFECTION
EPEC - Loss of microville; no toxin production
- Attach to the brush border of the
intestinal cells causing cell damage
(adhesive property)
- Infantile diarrhea (stool
without blood)
ETEC - Colonization occurs on the proximal
small intestine
- Virulence factors: heat stable and heat
labile enterotoxin
Traveller's diarrhea/
montezumas revenge
EIEC - Penetrates and multiplies within the
intestinal epithelial cells
- Virulence factor: invasin
Dysentery-like/shigella-like
infection
Watery diarrhea (stool with
wbcs)
EHEC
(serotype
0157:H7)
- Toxins destroy vascular endothelial cells
- Virulence factors: verotoxin (shiga-like
toxin)
Hemolytic Uremic
Syndrome
Hemorrhagic colitis
Bloody diarrhea
KLEBSIELLA
KLEBSIELLA PNEUMONIAE
Common name: friedlanders bacillus
Most common isolates of klebsiella
Causative agent of community acquired pneumonia
(currant jelly-like sputum)
Virulence factor: polysaccharide capsule
String test (+)
Neufeld Quellang (+)
TSI: a/a + -
IMViC: --++ (except for K. oxytoca: +-++)
ENTEROBACTER
Oppurtunistic infections: UTI, RT and wount infections
TSI: a/a + -
IMViC: --++
Exhibits fish eye colonies on EMB


Biochemical test E. aerogenes E. cloacae
Lysine decarboxylase + -
Arginine decarboxylase - +
Ornithine decarboxylase + +
Urease - -
Gelatin (22C) - +
ENTEROBACTER
CRONOBACTER SAKAZAKII
Formerly known as Enterobacter sakazakii
A pathogen of neonates causing meningitis and
bacteremia, often coming from powdered infant
formula
Produces a yellow pigmentation that intensifies at
25C

PANTOEA
PANTOEA AGGLOMERANS
Formerly known as Enterobacter agglomerans
Causes nosocomial outbreak of septicemia due to
contaminated IV fluids
It shows triple decarboxylase negative reaction


SERRATIA
Opportunistic pathogen
DNAse, lipase and gelatinase (+)
ONPG (+)
TSI: k/a + -
IMViC: --++

SERRATIA ODORIFERA
Produces musty-pungent odor or potato-like odor

SERRATIA MARCESCENS
Most clinically significant species
Produces a pink to red pigment (prodigiosin) at
25C


PROTEUS
Isolated in urine, wound and ear infections
Rapid urease producer
Human pathogens: P. mirabilis and P. vulgaris
Exhibits swarming phenomenon and burnt-gun
powder odor
PAD test (+)
IMViC: P. mirabilis : -+--; P. vulgaris: ++--
TSI: k/a ++


PROVIDENCIA
One of the causes of nosocomial outbreaks involving
burn units
Species: P. rettggeri (only urease +), P. stuartii, P.
alcalifaciens
PAD test (+)
IMViC: ++-+
TSI: k/a - -


MORGANELLA
Same biochemical reaction with P. vulgaris except
citrate (-)
Species: M. morganii
PAD test (+)
IMViC: ++--
TSI: k/a + -


EDWARDSIELLA
It has been isolated from cold-blooded and warm
blooded animals
Species: E. tarda (human pathogen)
LDC (+)
IMViC: ++--
TSI: k/a + +


ERWINIA
A plant pathogen and are not significant in human
infections

CITROBACTER
CITROBACTER FREUNDII
Can be isolated in diarrheal stool cultures
Associated with endocarditis in IV drug users
IMViC: -+-+
TSI: k/a + +
CITROBACTER KOSERI
Formely known as C. diversus
Causes nursery outbreaks of neonatal meningitis
and brain abscesses
IMViC: ++-+
TSI: k/a + -
SALMONELLA
Most serious pathogenic enterobacteria for humans,
causing enteric fever (typhoid fever) and acute
gastroenteritis (food poisining)
Acquired by ingestion of contaminated animal food
products or improperly cooked poultry, milk, eggs and
dairy products
May also be transmitted by human carriers
Virulence factors: fimbriae and enterotoxin
LDC (+)
IMViC: -+-+ except S. typhi (-+--)
TSI: k/a ++ except S. typhi (k/a -+)
SALMONELLA
3 general categories of Salmonella Infection
1. Gastroenteritis
- One of the most common forms of food poisoning
- Salmonella strain: S. enterica subsp. Enterica
- S. typhimurium: associated with peanut butter
outbreak
- Infective dose: 10
6
bacteria

- symptoms: nausea, vomiting, fever and chills,
watery diarrhea and abdominal pain
2. Bacteremia
- occurs with of without extraintestinal foci of
infection caused by nontyphoidal Salmonella (S.
typhimurium, S. paratyphi, S. cholerasuis)
- characterized by prolonged fever
SALMONELLA
3 general categories of Salmonella Infection

3. Enteric fever
- aka typhoid fever
- causative agent: S. typhi
- febrile disease that results from the ingestion of
contaminated food from infected individuals or carriers
- symptoms: malaise, anorexia, lethargy, myalgia,
and continous frontal dull headache
-characteristic red spots appears during the 2
nd

week of fever
- complications: necrosis in the gall bladder
(necrotizing cholecystitis) and peyers patches
SALMONELLA
Specimens for Salmonella identification

1. blood- 1
st
week of infection

2. Stool- 2
nd
week of infection

3. Urine- 3
rd
week of infection
SHIGELLA
Most inert enterobacteriaceae
Not a member of the normal GIT flora
An intracellular organism they multiply within the
cells of the colon epithelium
Transmitted through fecal-oral routes
No animal reservoir
Virulence factor: shiga-toxin
Antigenic structure: O antigen
Specimen: rectal swab, stool
Non-motile
TSI: k/a - -
IMViC: -+--
SHIGELLA
Bacillary dysentery
Caused by S. dysenteriae type 1
Characterized by acute inflammatory colitis and
bloody diarrhea
Infective dose: <200bacilli (highly communicable)
In young children, rectal prolapse occurs due to
excessive straining
Symptoms: fever, chills, abdominal cramps, painful
bowel movement and tenesmus
Complications: ileus (obstruction of the intestine),
seizures and HUS
SHIGELLA
PRESENT
DESIGNATION
GROUP TYPE CATALASE ONPG MANNITOL
S. dysenteriae A - - -
S. flexneri B + - +
S. boydii C + - +
S. sonnei D + + +
YERSINIA
Yersinia pestis
Aka plague bacillus
Considered class A bioterrorism agent
Only member of Enterobacteriaceae that is
transmitted through bite of an infected flea
(Xenopsylla cheopsis)
Non-motile
Causative agent of bubonic plague- black death or
6
th
century pandemic
Inclusions: bipolar bodies
Appears closed safety pin appearance after
waysons stain or methylene blue stain
Exhibits stalactite pattern in broth cultures
TSI: k/a - -
YERSINIA
Yersinia enterocolitica
Most commonly isolated species of Yersinia
Causative agent of enterocolitis (waterborne
gastroenteritis)
Motile at 22C but non-motile at 35C
Has the ability to survive in cold environment
Exhibits bulls eye colonies in CIN agar
Ferments sucrose and mannitol
Grows best at 25-30C
IMViC: -+--
TSI: k/a - -
LABORATORY DIAGNOSIS
1. Culture
a. EMB (Eosin-Methylene Blue Agar)
-CHO: lactose
b. MAC (MacConkey Agar)
- contains crystal violet and bile salts
- CHO: lactose
- pH indicator: neutral red
c. HEA (Hektoen-Enteric Agar)
- inhibitor: bile salts
- CHO: lactose, sucrose and salicin
- pH indicator: bromthymol blue
- H
2
S indicator: ferric ammonium citrate
LABORATORY DIAGNOSIS
1. Culture
d. SSA (Salmonella-Shigella Agar)
- CHO: lactose
- pH indicator: neutral red
- H
2
S indicator: ferric citrate
- Salmonella: non-LF, H
2
S (+) colorless
colonies with black center
- Shigella: non-LF, H
2
S (-) colorless colonies
without black center



LABORATORY DIAGNOSIS
2. LIA (Lysine Iron Agar)
- detects lysine deamination and decarboxylation
- pH indicator: bromcresol purple
- H
2
S indicator: ferric ammonium citrate
a. alkaline slant/alkaline butt (K/K)
(-) lysine deamination
(+) lysine decarboxylation
b. alkaline slant/ acid butt (K/A)
(-) lysine deamination
(-) lysine decarboxylation
c. red slant/acid butt (R/A)
(+) lysine deamination
(-) lysine decarboxylation
LABORATORY DIAGNOSIS
LYSINE IRON AGAR
K/K K/A R/A
E. Coli
Enterobacter spp
Serratia
Salmonaella
Edwardsiella
Shigella (some strains)
Shigella
Citrobacter
Yersinia
Enterobacter
Serratia
Proteus
Providencia
Morganella
LABORATORY DIAGNOSIS
3. TSI (Triple Sugar Iron)
- sugar ratio: 10:10:1 (lactose:sucrose:glucose)
- pH indicator: phenol red
- H
2
S indicator: ferrous sulfate and sodium
thiosulfate
TRIPLE SUGAR IRON
A/A + - A/A - + K/A + + K/A - + K/A + - K/A - -
E. Coli
K. Pnuemoniae
K. Oxytoca
E. Aerogenes
E. cloaca
P. agglomerans E. Tarda
Salmonella spp
C. Freundii
P. Vulgaris
P. mirabilis
S. Typhi

C. Diversus
H. Alvei
M. morganii
Shigella spp
LABORATORY DIAGNOSIS
4. SIM (Sulfide-Indole-Motility)
- (+) sulfide: black color formation
- (+) indole: pink to wine colored ring after the
addition of kovacs reagent
- (+) motolity: spread out/movement away from
the stab line

5. Methyl Red
- organisms that produces enough acid form
glucose fermentation will overcome the neutralizing effect
of the buffer
-mixed acid production: lactic acid, acetic acid,
formic acid, succinic acid


LABORATORY DIAGNOSIS
6. Voges Proskauer
- detects acetoin
- reagent: 40% KOH and -naphthol
- positive result: red complex

7. Citrate Utilization test
- determines if an organism can utilize citrate as
sole source of carbon
- pH indicator: bromthymol blue
- (+) result: intense blue

8. Malonate utilization test
- determines if an organisms is capable of utilizing
malonate as sole source of carbon
- (+) result: blue color


LABORATORY DIAGNOSIS
9. Orthonitrophenyl -galactosidase (ONPG)
- determines whether an organism is a slow or late
lactose fermenter (LLF) or true non-lactose fermenter
(NLF)
- (+) result: yellow color

10. Phenylalanine Deaminase (PAD)
- used to determine the ability of an organism to
oxidatively deaminate phenylalanine to phenyl pyruvic
acid
- (+) result: green colored complex

LABORATORY DIAGNOSIS
11. Urea Hydrolysis Test (Christensens Method)
- used to determine the ability of an organism to
produce the enzyme urease , which hydrolyzes urea
- (+) result: change in the color of slant from
orange to magenta
- (+) organism: Proteus and Morganella

12. Gelatin liquefaction
- determines the ability of a baterium proteases
that hydrolyze gelatin and liquefy solid gelatin medium
- (+) organism: Serratia


Organism TSI gas H2S Indole MR VP Citrate PAD Urease
Escherichia coli A/A + - + + - - - +
Shigella sonnei K/A - - - + - - - +
Edwardsiella tarda K/A + - + + - - - +
Salmonella K/A + + - + - + - +
Citrobacter freundii K/A + + - + - + - +/-
Citrobacter koseri K/A + + + + - + - +/-
Klebsiella pneumoniae A/A + - - - + + - +
Klebsiella oxytoca A/A + - + - + + - +
Enterobacter aerogenes A/A + - - - + + - -
Enterbacter cloacae A/A + - - - + + - +/-
Hafnia alvei K/A + - - +/- + - - -
Serratia marcescens K/A + - - +/- + + - -
Proteus vulgaris K/A +/- + + + - +/- + +
Proteus mirabilis K/A + + - + +/- +/- + +
Providencia rettgeri K/A - - + + - + + -
Providencia stuartii K/A - - + + - + + -
Morganella morganii K/A + - + + - - + +
Yersinia enterocolitica K/A - - +/- + - - - +/-