Most Common Pathogens

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Most Common Pathogens

• Gastrointestinal Tract

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Gastrointestinal Tract

• Nontyphoidal Salmonella species
• Disease Vomiting and diarrhea
• Epidemiology the most common cause of food
  poisoning in industrialized countries often
  associated with meat and poultry

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Gastrointestinal Tract

• Nontyphoidal Salmonella species
• Isolation
• Colonies like E. coli and other enteric bacteria
  on non-selective media
• Colonies are clear and colorless on MacConkey and
  EMB due to the fact that Salmonella fails to
  ferment lactose

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Gastrointestinal Tract

• Nontyphoidal Salmonella species
• Isolation
• Colonies have clear centers with black centers on
  Salmonella-Shigella (SS), Xylose-Lysine-Deoxychola
  te (XLD), Hektoen Enteric (HE), and other
  enteric agar this is an indication of
  hydrogen sulfide production

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Xylose Lysine Deoxycholate Agar
Salmonella species
Escherichia coli
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Salmonella on Hektoen and SS
Hektoen
SS (Salmonella Shigella)
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Gastrointestinal Tract

• Nontyphoidal Salmonella species
• Isolation
• Enrichment Broths A portion of the fecal
  specimen is suspended in Selenite F and/or
  Gram-negative (GN) broths Salmonella grows
  faster in these media allowing it to attain the
  log phase of growth quicker than indigenous
  bacteria a subculture after short incubation
  onto an enteric agar is more likely to yield
  isolated Salmonella colonies than direct plated
  specimens

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Gastrointestinal Tract

• Nontyphoidal Salmonella species
• Incubation Same as for E. coli and other enteric
  bacteria
• Preliminary identification typical enteric
  bacterium on gram stain oxidase negative
• Preliminary identification Alkaline slant, acid
  butt, volatile gas, and hydrogen sulfide produced
  in Kliglers Iron Agar (KIA) Alkaline slant,
  alkaline butt, and hydrogen sulfide produced in
  Lysine Iron Agar (LIA)

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Salmonella in Kliglers and Lysine Iron Agar
LIA
KIA
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Gastrointestinal Tract

• Nontyphoidal Salmonella species
• Genus identification Typical biochemical profile
  in API, Vitek, and other commercially available
  enteric panels of biochemical substrates
• Species identification Direct slide
  agglutination with Salmonella grouping antisera
  preliminary identification in house definitive
  identification sent to reference lab
  (salmonellosis is a reportable disease)

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Gastrointestinal Tract

• Salmonella typhi
• Disease typhoid fever (bacterium enters the body
  via macrophages and affects many body systems)
• Epidemiology Strictly a human pathogen passed
  by direct or indirect contact or vehicle
  transmission, most often by contaminated food and
  water (can you say Typhoid Mary?)

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Gastrointestinal Tract

• Salmonella typhi
• Isolation and identification is the same for S.
  typhi as it is for non-typhoidal Salmonella
  species S. typhi can be ruled out if isolate
  fails to agglutinate with Salmonella somatic
  group D antisera (S. typhi possesses somatic
  group D antigens)

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Gastrointestinal Tract

• Shigella species
• Disease diarrhea and dysentery (shigellosis)
• Epidemiology strictly a human pathogen, usually
  from contaminated drinking water
• Isolation
• Colonies are like E. coli and other enteric
  bacteria on non-selective media

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Gastrointestinal Tract

• Shigella species
• Isolation
• Colonies are clear and colorless on MacConkey,
  EMB, XLD, HE, and SS due to the fact that
  Shigella fails to ferment lactose

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Gastrointestinal Tract

• Shigella species
• Isolation
• Colonies on Salmonella-Shigella (SS),
  Xylose-Lysine-Deoxycholate (XLD), Hektoen Enteric
  (HE), do not have black centers due to the lack
  of hydrogen sulfide production

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Shigella species
MacConkey
XLD
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Gastrointestinal Tract

• Shigella species
• Incubation Same as for E. coli and other enteric
  bacteria
• Preliminary identification typical enteric
  bacterium on gram stain oxidase negative
• Preliminary identification alkaline slant, acid
  butt, no volatile gas, no hydrogen sulfide in
  Kliglers medium Alkaline slant, acid butt, no
  hydrogen sulfide in Lysine Iron Agar Non-motile
  in semisolid agar deep

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Gastrointestinal Tract

• Shigella species
• Genus identification typical biochemical profile
  in API, Vitek, and other commercially available
  enteric panels of biochemical substrates
• Species identification Direct slide
  agglutination with Shigella grouping antisera
  preliminary identification in house definitive
  identification sent to reference lab
  (shigellosis is a reportable disease)

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Gastrointestinal Tract

• Shiga toxin producing Escherichia coli (STEC)
• Synonyms Enterohemorrhagic (EHEC) Verotoxin
  producing (VTEC) (colloquial name
  Jack-In-The-Box E. coli)
• Disease bloody diarrhea, hemolytic uremic
  syndrome (HUS)
• Epidemiology Naturally found in farm animals,
  especially calves and cattle, contaminant of food
  (undercooked hamburgers, apple juice, etc) and
  water borne

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Gastrointestinal Tract

• Shiga toxin producing Escherichia coli (STEC)
• Incubation same as for other enteric bacteria
• Isolation identical to garden variety
  non-toxigenic E. coli on SBA and EMB or regular
  MAC produces clear colorless colonies on
  sorbitol MacConkey (SMAC) because, unlike other
  E. coli isolates, it fails to ferment sorbitol)

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E. coli on MacConkey Sorbitol
STEC
Non STEC
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Gastrointestinal Tract

• Shiga toxin producing Escherichia coli (STEC)
• Preliminary identification typical enteric
  bacterium on gram stain oxidase negative
• Identified as E. coli on commercial biochemical
  panels sorbitol negative E. coli must be tested
  for shiga-toxin commercial ELISA tests are
  available, cases of STEC diseases are reportable
  complete identification and toxigenicity tests
  performed in reference lab

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Gastrointestinal Tract

• Campylobacter jejuni
• Disease explosive bloody diarrhea
• Epidemiology naturally found in animals,
  especially birds contaminates human food and
  water
• Isolation limited growth on regular sheep blood
  agar, no growth on enteric agars requires
  specially enriched blood agar with contains
  antibiotics to inhibit indigenous fecal
  microbiota (a selective blood agar)

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Gastrointestinal Tract

• Campylobacter jejuni
• Incubation C. jejuni is microaerophilic and
  thermophilic the selective blood agar is usually
  put in a Campy pouch (a sealable pouch which
  can be made microaerophilic with gas generating
  ampoules) and incubated at 42oC for 48 hours

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Gastrointestinal Tract

• Campylobacter jejuni
• Colony morphology 1-3mm, moist and shiny may be
  round or very irregular due to the tendency for
  colonies to run together
• Preliminary identification very pale
  gram-negative, curved to loosley spiraled,
  sometimes described as gull-wing-like, basic
  fuchsin is a better counterstain than safranin

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Campylobacter Morphology
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Gastrointestinal Tract

• Campylobacter jejuni
• Species identification oxidase positive, rapidly
  darting motility in wet mount further testing
  not required if typical colonies grow on
  selective blood agar C. jejuni is the only
  Campylobacter species to hydrolyze sodium
  hippurate

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Gastrointestinal Tract

• Yersinia entercolitica
• Disease diarrhea and mesenteric lymphadenitis
• Epidemiology food and water borne is mostly
  isolated in colder climates (e.g. Northern USA,
  Canada, Scandinavia)
• Isolation grows slower than other enteric
  bacteria on routine primary isolation enteric
  agar and blood agar in ambient air at 35oC

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Gastrointestinal Tract

• Yersinia entercolitica
• Incubation Y. enterocolitica is the only
  frequently isolated enteric pathogen that grows
  optimally at room temperature
• Colony morphology blood agar lt1mm, round,
  smooth, and gray after 24h at 35oC typical
  enteric type colonies develop if incubated an
  additional day at room temperature

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Gastrointestinal Tract

• Yersinia entercolitica
• Colony morphology MAC agar lt1mm, round, smooth,
  bulls-eye colony (pink center clear periphery)
  after 24 hours that grow to typical enteric
  colonies 24h more at room temperature
• Preliminary identification typical enteric
  bacterium on gram stain oxidase negative
• Species identification commercial biochemical
  panel for enteric bacteria usually suffices

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Gastrointestinal Tract

• Helicobacter pylori
• Disease chronic active gastritis and peptic
  ulcers
• Epidemiology high percentage of humans are
  carriers, person-to-person transmission probable,
  reinfection is common
• Isolation poor growth on any routine isolation
  media culture rarely performed

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Gastrointestinal Tract

• Helicobacter pylori
• Incubation if cultured, rich blood or chocolate
  agar incubated several days in CO2 rich
  environment at 35oC
• Preliminary identification H. pylori is
  morphologically similar to Campylobacter on Gram
  stain Warthin-Starry silver stain on gastric
  biopsy is the staining method of choice

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Helicobacter pylori
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Gastrointestinal Tract

• Helicobacter pylori
• Presumptive identification H. pylori produces a
  very potent urease that can be detected when a
  small piece of gastric biopsy is placed on a
  special urea agar (the pH indicator turns deep
  purple within 2 hours due to ammonia
  accumulation)
• Presumptive identification the patient swallows
  a pill containing radioactive urea radioactive
  CO2 is detected in the patients breath

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Gastrointestinal Tract

• Helicobacter pylori
• Definitive diagnosis IgA, IgM, and IgG can be
  detected in patients serum, the presence of IgM
  usually indicates active disease
• The radioactive CO2 breath test is used ascertain
  effectiveness of antibiotic therapy

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Gastrointestinal Tract

• Clostridium difficile
• Disease antibiotic associated diarrhea and
  pseudomembrane colitis most common cause of
  diarrhea in hospitalized patients
• Epidemiology normal indigenous bacteria of the
  human gut normally present in small numbers the
  numbers increase rapidly when other normal gut
  bacteria are eliminated when patient is treated
  with antibiotics

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Gastrointestinal Tract

• Clostridium difficile
• Culture usually not recommended because C.
  difficile can be isolated from feces collected
  from healthy people
• Cycloserine cefoxitin fructose agar (CCFA) is a
  selective and differential medium if culture is
  performed
• Incubation C. difficile is an obligate anaerobe

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Gastrointestinal Tract

• Clostridium difficile
• Preliminary identification (if cultured) typical
  colonies on CCFA gram-positive endospore forming
  rod
• Species identification biochemical
  identification using commercial panel designed to
  identify obligate anaerobes
• Definitive diagnosis serological detection of
  specific exotoxin (antigen) in feces using ELISA
  is the most common method used (performed daily
  at LGH)

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Gastrointestinal Tract

• Rotavirus
• Disease gastroenteritis, (very common,
  especially in children)
• Epidemiology human are carriers,
  person-to-person contact probable
• Detection and identification like other viruses,
  rotavirus requires equipment, supplies and
  trained personnel not available in most clinical
  laboratories

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Gastrointestinal Tract

• Rotavirus
• Definitive diagnosis simple, accurate,
  efficient, and cost effective in house serology
  tests available ELISA tests for rotavirus
  antigens in feces are performed regularly at LGH