1. Chapter 16 Enterobacteriaceae
MLAB 2434 – Clinical Microbiology
Cecile Sanders & Keri Brophy-Martinez

2. Chapter 16 - Enterics
Family Enterobacteriaceae often
referred to as “enterics”
 Four major features:

 All
ferment glucose (dextrose)
 All reduce nitrates to nitrites
 All are oxidase negative
 All except Klebsiella, Shigella and
Yersinia are motile

3. Microscopic and Colony
Morphology
Gram negative bacilli or coccobacilli
 Non-spore forming
 Colony morphology on BAP or CA of
little value, as they look the same,
except for Klebsiella
 Selective and differential media are
used for initial colony evaluation
(ex. MacConkey, HE, XLD agars)


4. Classification of Enterics
Due to the very large number of
organisms in the Family
Enterobacteriaceae (see Table 1611), species are grouped into Tribes,
which have similar characteristics
(Table 16-1, page 466)
 Within each Tribe, species are
further subgrouped under genera


5. Virulence and Antigenic
Factors of Enterics



Ability to colonize, adhere, produce
various toxins and invade tissues
Some possess plasmids that may mediate
resistance to antibiotics
Many enterics possess antigens that can
be used to identify groups
O
antigen – somatic, heat-stable antigen
located in the cell wall
 H antigen – flagellar, heat labile antigen
 K antigen – capsular, heat-labile antigen

6. Clinical Significance of
Enterics
Enterics are ubiquitous in nature
 Except for few, most are present in
the intestinal tract of animals and
humans as commensal flora;
therefore, they are sometimes call
“fecal coliforms”
 Some live in water, soil and sewage


7. Clinical Significance of
Enterics (cont’d)

Based on clinical infections
produced, enterics are divided into
two categories:
 Opportunistic
pathogens – normally
part of the usual intestinal flora
that may produce infection outside
the intestine
 Primary intestinal pathogens –
Salmonella, Shigella, and Yersinia sp.

8. Escherichia coli
Most significant species in the
genus
 Important potential pathogen in
humans
 Common isolate from colon flora


9. Escherichia coli (cont’d)

Characteristics
 Dry,
pink (lactose positive) colony
with surrounding pink area on
MacConkey

10. Escherichia coli (cont’d)
 Ferments
glucose, lactose,
trehalose, & xylose
 Positive indole and methyl red tests
 Does NOT produce H2S or
phenylalanine deaminase
 Simmons citrate negative
 Usually motile
 Voges-Proskauer test negative

11. Escherichia coli (cont’d)

Infections
 Wide
range including meningitis,
gastrointestinal, urinary tract,
wound, and bacteremia
 Gastrointestinal Infections
• Enteropathogenic (EPEC) – primarily in
infants and children; outbreaks in
hospital nurseries and day care
centers; stool has mucous but not
blood; identified by serotyping

12. Escherichia coli (cont’d)
• Enterotoxigenic (ETEC) – “traveler’s
diarrhea”; watery diarrhea without
blood; self-limiting; usually not
identified, other than patient history
and lactose-positive organisms cultured
on differential media
• Enteroinvasive (EIEC) – produce
dysentery with bowel penetration,
invasion and destruction of intestinal
mucosa; watery diarrhea with blood; do
NOT ferment lactose; identified via
DNA probes

13. Escherichia coli (cont’d)
• Enterohemorrhagic (EHEC serotype
0157:H7) – associated with
hemorrhagic diarrhea and hemolyticuremic syndrome (HUS), which includes
low platelet count, hemolytic anemia,
and kidney failure; potentially fatal,
especially in young children;
undercooked hamburger, unpasteurized
milk and apple cider have spread the
infection; does NOT ferment sucrose;
identified by serotyping

14. Escherichia coli (cont’d)
• Enteroaggregative (EaggEC) – cause
diarrhea by adhering to the mucosal
surface of the intestine; watery
diarrhea; symptoms may persist for
over two weeks
 Urinary
Tract Infections
• E. coli is most common cause of UTI
and kidney infection in humans
• Usually originate in the large instestine
• Able to adhere to epithelial cells in the
urinary tract

15. Escherichia coli (cont’d)
 Septicemia
& Meningitis
• E. coli is one of the most common causes of
septicemia and meningitis among neonates;
acquired in the birth canal before or during
delivery
• E. coli also causes bacteremia in adults,
primarily from a genitourinary tract infection
or a gastrointestinal source


Escherichia hermannii – yellow pigmented;
isolated from CSF, wounds and blood
Escherichia vulneris - wounds

16. Klebsiella, Enterobacter,
Serratia & Hafnia sp.



Usually found in intestinal tract
Wide variety of infections, primarily
pneumonia, wound, and UTI
General characteristics:
 Some
species are non-motile
 Simmons citrate positive
 H2S negative
 Phenylalanine deaminase negative
 Some weakly urease positive
 MR negative; VP positive

17. Klebsiella species



Usually found in GI tract
Four major species
K. pneumoniae is mostly commonly isolated
species
 Possesses
a polysaccharide capsule, which
protects against phagocytosis and
antibiotics AND makes the colonies moist
and mucoid
 Has a distinctive “yeasty” odor
 Frequent cause of nosocomial pneumonia

18. Klebsiella species (cont’d)
 Significant
biochemical reactions
• Lactose positive
• Most are urease positive
• Non-motile

19. Enterobacter species
Comprised of 12 species; E. cloacae
and E. aerogenes are most common
 Isolated from wounds, urine, blood
and CSF
 Major characteristics
 Colonies resemble Klebsiella

 Motile
 MR
negative; VP positive

20. Enterobacter species
(cont’d)

21. Serratia species
Seven species, but S. marcescens is
the only one clinically important
 Frequently found in nosocomial
infections of urinary or respiratory
tracts
 Implicated in bacteremic outbreaks
in nurseries, cardiac surgery, and
burn units
 Fairly resistant to antibiotics


22. Serratia species (cont’d)

Major characteristics
 Ferments
lactose slowly
 Produce characteristic pink pigment,
especially when cultures are left at
room temperature
S. marscens on
nutrient agar →

23. Hafnia species
Hafnia alvei is only species
 Has been isolated from many
anatomical sites in humans and the
environment
 Occasionally isolated from stools
 Delayed citrate reaction is major
characteristic


24. Proteus, Morganella &
Providencia species
All are normal intestinal flora
 Opportunistic pathogens
 Deaminate phenylalanine
 All are lactose negative


25. Proteus species





P. mirabilis and P. vulgaris are widely
recognized human pathogens
Isolated from urine, wounds, and ear and
bacteremic infections
Both produce swarming colonies on nonselective media and have a distinctive
“burned chocolate” odor
Both are strongly urease positive
Both are phenylalanine deaminase positive

26. Proteus species (cont’d)
A exhibits characteristic “swarming”
 B shows urease positive on right


27. Morganella species
Morganella morganii is only species
 Documented cause of UTI
 Isolated from other anatomical
sites
 Urease positive
 Phenylalanine deaminase positive


28. Providencia species
Providencia rettgeri is pathogen of
urinary tract and has caused
nosocomial outbreaks
 Providenicia stuartii can cause
nosocomial outbreaks in burn units
and has been isolated from urine
 Both are phenylalanine deaminase
positive


29. Citrobacter species
Citrobacter freundii associated
with nosocomial infections (UTI,
pneumonias, and intraabdominal
abscesses)
 Ferments lactose and hydrolyzes
urea slowly
 Resembles Salmonella sp.


30. Salmonella
Produce significant infections in
humans and certain animals
 On differential selective agar,
produces clear, colorless, nonlactose fermenting colonies with
black centers (if media contains
indicator for hydrogen sulfide)


31. Salmonella (cont’d)

Salmonella on MacConkey

32. Salmonella (cont’d)
Lactose negative
 Negative for indole, VP,
phenylalanine deaminase, and urease
 Most produce H S
2
 Do not grow in potassium cyanide
 Large and complex group of
organisms; grouped by O, H, and Vi
(for virulence) antigens


33. Salmonella (cont’d)

Clinical Infections
 Acute
gastroenteritis or food poisoning
• Source = handling pets, insufficiently cooked
eggs and chicken, and contaminated cooking
utensils
• Occurs 8 to 36 hours after ingestion
• Requires a high microbial load for infection
• Self-limiting in health individuals (antibiotics
and antidiarrheal agents may prolong
symptoms)

34. Salmonella (cont’d)
 Typhoid
and Other Enteric Fevers
• Prolonged fever
• Bacteremia
• Involvement of the RE system,
particularly liver, spleen, intestines, and
mesentery
• Dissemination to multiple organs
• Occurs more often in tropical and
subtropical countries

35. Salmonella (cont’d)
 Salmonella
Bacteremia
 Carrier State
• Organisms shed in feces
• Gallbladder is the site of organisms
(removal of gallbladder may be the only
solution to carrier state)

36. Shigella species
Closely related to the Escherichia
 All species cause bacillary
dysentery
 S. dysenteriae (Group A)
 S. flexneri (Group B)
 S. boydii (Group C)
 S. sonnei (Group D)


37. Shigella (cont’d)

Characteristics
 Non-motile
 Do
not produce gas from glucose
 Do not hydrolyze urea
 Do not produce H2S on TSI
 Lysine
decarboxylase negative
 ONPG positive (delayed lactose +)
 Fragile organisms
 Possess O and some have K antigens

38. Shigella (cont’d)

Clinical Infections
 Cause
dysentery (bloody stools, mucous,
and numerous WBC)
 S. sonnei is most common, followed by S.
flexneri (“gay bowel syndrome”)
 Humans are only known reservoir
 Oral-fecal transmission
 Fewer than 200 bacilli are needed for
infection in health individuals

39. Shigella (cont’d)

40. Yersinia species


Consists of 11 named species
Yersinia pestis
 Causes
plague, which is a disease primarily
of rodents; transmitted by fleas
 Two forms of plague, bubonic and
pneumonic
 Gram-negative, short, plump bacillus,
exhibiting “safety-pin” or “bipolar”
staining

41. Yersinia species

Yersinia enterocolitica







Most common form of Yersinia
Found worldwide
Found in pigs, cats and dogs
Human also infected by ingestion of contaminated
food or water
Some infections result from eating contaminated
market meat and vacuum-packed beef
Is able to survive refrigerator temperatures (can
use “cold enrichment” to isolate)
Mainly causes acute gastroenteritis with fever

42. Yersinia species

Yersinia pseudotuberculosis
 Pathogen
of rodents, particularly
guinea pigs
 Septicemia with mesenteric
lymphadenitis, similar to
appendicitis
 Motile at 18 to 22 degrees C

43. Laboratory Diagnosis of
Enterics

Collection and Handling
 If
not processed quickly, should be
collected and transported in CaryBlair, Amies, or Stuart media

Isolation and Identification
 Site
of origin must be considered
 Enterics from sterile body sites are
highly significant
 Routinely cultured from stool

44. Laboratory Diagnosis of
Enterics (cont’d)

Media for Isolation and
Identification of Enterics
 Most
labs use BAP, CA and a
selective/differential medium such
as MacConkey
 On MacConkey, lactose positive are
pink; lactose negative are clear and
colorless

45. Laboratory Diagnosis of
Enterics (cont’d)
 For
stools, highly selective media,
such as Hektoen Enteric (HE), XLD,
or SS is used along with MacConkey
agar

Identification
 Most
labs use a miniaturized or
automated commercial identification
system, rather than multiple tubes
inoculated manually

46. Laboratory Diagnosis of
Enterics (cont’d)

Identification (cont’d)
 All
enterics are
• Oxidase negative
• Ferment glucose
• Reduce nitrates to nitrites

47. Laboratory Diagnosis of
Enterics (cont’d)

Common Biochemical Tests
 Lactose
fermentation and utilization
of carbohydrates
 Triple Sugar Iron (TSI)
 ONPG
 Glucose metabolism
• Methyl red
• Voges-Proskauer

48. Laboratory Diagnosis of
Enterics (cont’d)

Common Biochemical Tests (cont’d)
 Miscellaneous
•
•
•
•
•
•
Reactions
Indole
Citrate utilization
Urease production
Motility
Phenylalanine deaminase
Decarboxylase tests

49. Screening Stools for
Pathogens
Because stools have numerous
microbial flora, efficient screening
methods must be used to recover
any pathogens
 Enteric pathogens include
Salmonella, Shigella, Aeromonas,
Campylobacter, Yersinia, Vibrio, and
E. coli 0157:H7


50. Screening Stools for
Pathogens (cont’d)
Most labs screen for Salmonella,
Shigella, and Campylobacter; many
screen for E. coli 0157:H7
 Fecal pathogens are generally
lactose-negative (although Proteus,
Providencia, Serratia, Citrobacter
and Pseudomonas are also lactosenegative)
