Chapter 16 - Enterobacteriaceae

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Chapter 16 - Enterobacteriaceae

• MLAB 2434 Clinical Microbiology
• Cecile Sanders Keri Brophy-Martinez

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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

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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)

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Classification of Enterics

• Due to the very large number of organisms in the
  Family Enterobacteriaceae (see Table 16-11),
  species are grouped into Tribes, which have
  similar characteristics (Table 16-1, page 466)
• Within each Tribe, species are further subgrouped
  under genera

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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

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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

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Clinical Significance of Enterics (contd)

• 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.

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Escherichia coli

• Most significant species in the genus
• Important potential pathogen in humans
• Common isolate from colon flora

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Escherichia coli (contd)

• Characteristics
• Dry, pink (lactose positive) colony with
  surrounding pink area on MacConkey

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Escherichia coli (contd)

• 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

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Escherichia coli (contd)

• 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

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Escherichia coli (contd)

• Enterotoxigenic (ETEC) travelers 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

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Escherichia coli (contd)

• Enterohemorrhagic (EHEC serotype 0157H7)
  associated with hemorrhagic diarrhea and
  hemolytic-uremic 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

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Escherichia coli (contd)

• 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

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Escherichia coli (contd)

• 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

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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

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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

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Klebsiella species (contd)

• Significant biochemical reactions
• Lactose positive
• Most are urease positive
• Non-motile

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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

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Enterobacter species (contd)
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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

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Serratia species (contd)

• Major characteristics
• Ferments lactose slowly
• Produce characteristic pink pigment, especially
  when cultures are left at room temperature
• S. marscens on
• nutrient agar ?

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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

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Proteus, Morganella Providencia species

• All are normal intestinal flora
• Opportunistic pathogens
• Deaminate phenylalanine
• All are lactose negative

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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 non-selective
  media and have a distinctive burned chocolate
  odor
• Both are strongly urease positive
• Both are phenylalanine deaminase positive

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Proteus species (contd)

• A exhibits characteristic swarming
• B shows urease positive on right

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Morganella species

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

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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

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Citrobacter species

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

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Salmonella

• Produce significant infections in humans and
  certain animals
• On differential selective agar, produces clear,
  colorless, non-lactose fermenting colonies with
  black centers (if media contains indicator for
  hydrogen sulfide)

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Salmonella (contd)

• Salmonella on MacConkey

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Salmonella (contd)

• Lactose negative
• Negative for indole, VP, phenylalanine deaminase,
  and urease
• Most produce H2S
• Do not grow in potassium cyanide
• Large and complex group of organisms grouped by
  O, H, and Vi (for virulence) antigens

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Salmonella (contd)

• 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)

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Salmonella (contd)

• 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

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Salmonella (contd)

• 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)

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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)

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Shigella (contd)

• 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

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Shigella (contd)

• 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

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Shigella (contd)
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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

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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

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Yersinia species

• Yersinia pseudotuberculosis
• Pathogen of rodents, particularly guinea pigs
• Septicemia with mesenteric lymphadenitis, similar
  to appendicitis
• Motile at 18 to 22 degrees C

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Laboratory Diagnosis of Enterics

• Collection and Handling
• If not processed quickly, should be collected and
  transported in Cary-Blair, 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

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Laboratory Diagnosis of Enterics (contd)

• 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

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Laboratory Diagnosis of Enterics (contd)

• 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

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Laboratory Diagnosis of Enterics (contd)

• Identification (contd)
• All enterics are
• Oxidase negative
• Ferment glucose
• Reduce nitrates to nitrites

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Laboratory Diagnosis of Enterics (contd)

• Common Biochemical Tests
• Lactose fermentation and utilization of
  carbohydrates
• Triple Sugar Iron (TSI)
• ONPG
• Glucose metabolism
• Methyl red
• Voges-Proskauer

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Laboratory Diagnosis of Enterics (contd)

• Common Biochemical Tests (contd)
• Miscellaneous Reactions
• Indole
• Citrate utilization
• Urease production
• Motility
• Phenylalanine deaminase
• Decarboxylase tests

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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 0157H7

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Screening Stools for Pathogens (contd)

• Most labs screen for Salmonella, Shigella, and
  Campylobacter many screen for E. coli 0157H7
• Fecal pathogens are generally lactose-negative
  (although Proteus, Providencia, Serratia,
  Citrobacter and Pseudomonas are also
  lactose-negative)