Vibrio, Aeromonas, Campylobacter

Vibrionaceae

 Gram negative
 Oxidase positive
 Fermenters
 Straight to Curved rods
 Motile by polar monotrichous flagella
 Facultative anaerobic
 Halophilic

 Gram negative, Straight to Curved rods
 Oxidase positive
 Not H2S Producing
 Indole Positive
 Urea Negative
 VP Variable
 Fermenters: Lactose/Sucrose
 String Test Positive
 Motile by Polar Monotrichous Flagella
 Facultative anaerobe
 Halophilic (thrives in High Salt Concentrations)

Vibrio gram stain

routes of transmission for vibriosis

 Raw or undercooked shellfish, especially oysters
 Drinking contaminated water
 Exposing open wounds to salt water or brackish water
(including surgery, tattoo, piercing)
 Handling raw seafood or seafood juices
 Fishing in marine coastal waters and estuaries

specimens for vibrio

 Stool Samples in Transport Media/Preservative (Cary-Blair)
o Collected during the Acute Stage of the Disease
o Tested at Guthrie
o BioFire Instrument - PCR
o Test TAT = 1hr 15 min
 Vomit or Rectal Swabs in Transport Media (Amies Swabs)
o Acceptable if stool is unable to be collected
o Tested at Quest – Reference Lab - Culture
o Test TAT = 3 days

 Blood Cultures
o Recommended if the patient is febrile, has hemorrhagic bullae, or
signs of sepsis

 Other Samples include Wound swabs and Tissue samples

 Vibrio is susceptible to drying and sunlight
o Making correct sample collection imperative for detection

culture media for vibrio

 Vibrio is non-fastidious
o Do not have any special growth requirements
 Standard Stool Culture Plating
o Blood, MacConkey, Hektoen, Brilliant Green, Campy
o TCBS Added if Vibrio is Suspected
 Indicated by physician's order

thiosulfate-citrated-bile salts-sucrose agar (TCBS)

 Selective and Differential media
 Used for the Isolation of Vibrio, exclusively
o Increased pH enhances the growth of V. cholerae and inhibits the growth of
normal GI flora; Vibrio = alkaline loving

 Bromothymol blue indicator
o AGAR turns yellow in presence of acid
o pH Indicator

 H2S indicator
o H2S producing colonies have a black center

 Sucrose Fermentation
o Fermenters: Colonies are YELLOW
(V. Cholerae and V. alginolyticus)
o Non-fermenters: Colonies are Blue/green
(V. parahaemolyticus and V. vulnificus)

growth characteristics on TCBS

H2S production/sucrose fermentation

vibrio NaCl tolerance

vibrio cholerae

 All strains share a common flagella H antigen
 Further subdivided into serogroups by their somatic O
antigens
o At least 150 serogroups have been recognized
 Two main groups:
o Toxigenic V. cholerae O1
 Associated with epidemic cholera
o Non-O1 V. cholerae
 Do not agglutinate with O1 antisera

V. cholera serotypes and biotypes

 V. cholerae O1 biotypes
o Classic: Asiatic or epidemic cholera
o El Tor variant seen in recent cases
 Non-O1
o V. cholerae 3 serotypes (Classic/El Tor)
 Differentiated by LPS somatic antigens
 Inaba (A & C antigens)
 Ogawa (A & B )
 Hikojima (A, B & C)

o V. cholerae O139
 A.k.a. V. cholerae Bengal, 1992
 Associated with epidemic outbreaks
 Produces same cholera toxins as O1 biotypes

ferment glucose and other carbohydrates?

vibrio, aeromonas, plesiomonas

microaerophilic and do not ferment carbohydrates

Campylobacter spp

cholera: classic VS El Tor

 Classic O1 Strain
o Non-hemolytic on Blood Agar
o Sensitive to Polymyxin B
o Non-agglutinating Chick RBCs
o VP -
 El Tor O1 Strain
o Beta hemolytic on Blood Agar
o Resistant to Polymyxin B
o Agglutinates Chick RBC
o VP +

clinical disease spectrum

 Epidemic cholera
o Mild to fatal illness
 Extraintestinal infections
o Wounds and bacteremia
 Gastroenteritis
o Cholera
 Endemic or epidemic in areas with poor sanitation; it occurs
sporadically
 Potentially life-threatening secretory diarrhea characterized by
numerous, voluminous watery stools, often accompanied by
vomiting, and resulting in hypovolemic shock and acidosis
 Transmitted by the fecal-oral route

epidemic cholera outbreaks

o 1816-1826 - First Cholera pandemic – Bengal, India.
o 1829-1851 - Second Cholera pandemic
o 1849 - Second major outbreak in Paris, London, N.A., Mississippi River, New
Orleans, California & Oregon Trail.
o 1852-1860 - Third Cholera pandemic
o 1863-1875 - Fourth Cholera pandemic
o 1881-1896 - Fifth Cholera pandemic
o 1899-1923 - Sixth Cholera pandemic
o 1961-1970s -Seventh Cholera pandemic
o 1992 a new strain appeared in Asia, a non-O1, (NAG) named O139 Bengal.
o 2008-2009 Outbreak in Zimbabwe
o 2010-2011 Outbreak in Haiti; after earthquake
WHO Task Force Efforts:
o Reduce deaths & spread of Cholera by 2030
o Vaccinate in high-risk population

extraintestinal infections

 Often caused by non-O1 strains
 Usually in immunocompromised hosts
 Bacteremia/Sepsis
 Wound infections
 Ear infections

gastroenteritis

 Watery diarrhea
 Caused by Choleragen (cholera toxin) CTX
 CTX consist of two A subunits and 5 B subunit
 B subunit binds to GM1 receptor on jejunal cells
 Delivery of A2 subunit to cytosolic target
 This facilitates entrance of A1 subunit into the cell
 Activated A1 subunit stimulates the production of Adenylate cyclase
through inactivation of the G protein
 This results in an accumulation of intracellular cyclic adenosine
monophosphate (cAMP)

 Stimulates the Hypersecretion of electrolytes (Na+, K+, CL-)
out of the cell into the lumen of the intestine
 Water passively diffuses out of the cell to maintain osmolality
 Massive accumulation of water in the GI lumen
 When the volume of fluid exceeds the capacity of the GI tract
to reabsorb it, results in massive outpouring of watery diarrhea

 Fluid loss can be severe,
o up to 20 L per day
 Stools may have “rice water” appearance
o colorless w/mucus flecks
 Death can be caused by dehydration and low electrolyte levels
 Cholera occurs as epidemics, pandemics, or sporadically
 Cholera epidemics are usually caused by V. cholerae O1
 Treatment:
o Administration of IV/oral fluids
o Antibiotics

mechanism of action of cholera toxin

vibrio cholerae O1 biochemicals/test results

 Does not require salt
o BUT will grow in media up to 3% NaCl
 Sucrose Fermentation (Yellow on TCBS)
 Non-Lactose Fermentation (Colorless on MacConkey)
 Oxidase +
 Motile
 Nitrate +
 Susceptible to O/129
 Lysine +, ornithine +, arginine -
 String test +
 O1 agglutination +

V. parahaemolyticus biochemicals

 Non-Sucrose Fermentation (Green on TCBS)
 Lactose Fermentation (Pink on MacConkey)
 H2S Producer (Black/Green on TCBS)
 Requires 1-7% NaCl for growth
 Oxidase +
 Motile
 Ornithine +
 Sensitive to O/129

V. vulnificus biochemicals

 Non-Sucrose Fermentation (Green on TCBS)
 Lactose Fermentation (Pink on MacConkey)
 Oxidase +
 Requires 1-6% NaCl for growth
 Motile
 Underlying liver disease- Predisposition
 Hepatitis, cirrhosis

V. alignolyticus biochemicals

 Sucrose Fermentation (Yellow on TCBS)
 Non-Lactose Fermentation (Colorless on MacConkey)
 Strict Halophile
 Growth 3-10% NaCl
 Oxidase +
 Motile
 Sensitive to O/129
 Ornithine +/-

differentiation of vibrio sp

aeromonadaceae

• Straight, gram negative bacilli
• Motile by polar flagella
• Glucose fermentation
• (Several) Sucrose and Lactose fermenation
• Oxidase Positive
• Catalase Positive
• H2S Producer
• Indole Positive
• Facultative Anerobe
• String Test Negative
• Urea Negative
• VP Positive

routes of transmission for aeromoniasis

 Fecal/Oral transmission from contaminated food/water
 Ubiquitous in terrestrial environments
 Close association with humans and animals
 Originally known to cause infections in fish and other cold-blooded
animals
 Widely targets immunocompromised animals and human hosts,
resulting in wound infections, cellulitis, septicemia, and urinary tract
infections.

disease and specimens/culture media

 Gastroenteritis
 Wound infections
 Bacteremia
 UTI
 Meningitis
 Ear infections

Specimens/Culture Media
 Same as Vibrio

complexes

Aeromonas sp are grouped into complexes:
o Aeromonas hydrophila complex
 A. hydrophila, A.bestiarum, and A. dhakensis
o Aeromonas veronii complex
 A. veronii, A. jandaei, A.trota, A.schubertii, A. diversa, and
A.encheleia
o Aeromonas caviae complex
 A. caviae, A.media, A. riviopollensis, and A. eucrenophila

aeromonas hydrophila

 Most strains are β-hemolytic
 Oxidase +
 Nitrate +
 Resistant to O/129
 No growth in 6.5 % NaCl
 Indole +
 Esculin +
 TSI = K/A w/gas
 Gel +
 DNase +
 Resistant to Amoxicillin and Ampicillin

aeromonas hydrophila growth on blood and Mac agar resembles pseudomonas aeruginosa

o Major Differences
 Aeromonas = Indole Positive
 Pseudomonas smells like Grapes

plesiomonaceae

 Straight-to-rounded, short, Gram-negative rod
 Motile
 Have lophotrichous flagella
 nonhemolytic on sheep blood agar
 Glucose fermenter
 Non-Lactose and Non-Sucrose fermenter
 Oxidase Positive
 Indole Positive
 DNase Negative
 VP Negative
 Faculatative Anaerobe

routes of transmission for plesiomoniasis

 Transmitted through the fecal-oral route
 Consumption of infected seafood (especially oysters)
 Contaminated water and contaminated vegetables
 Exposure to amphibians and reptiles
Routes of Transmission for Plesiomoniasis

vibrio vs aeromonas vs plesiomonas

O/129 susceptibility testing for vibrio, aeromonas, and plesiomonas

 O/129 disk is the vibriostatic agent
o 2,4-diamino-6,7-diisopropylpteridine phosphate
 O/129 disks containing 10 and 150 μg are used in a method resembling disk susceptibility testing to
differentiate Vibrio species from Aeromonas species. Aeromonas species are resistant, with no zone
of inhibition at 24 hours, with both disks. Vibrio and Plesiomonas species will show susceptibility, with
a distinct zone of inhibition with the 150 μg disk; results for the 10 μg disk will vary among
the Vibrio and Plesiomonas species. Some Vibrio spp. require salt for growth. Therefore, the test is
run in duplicate on Mueller-Hinton agar (MHA) with low salt (0.5%) and with added NaCl (4%) to
ensure growth on at least one of the plates

string test

 Distinguishes Vibrio from Aeromonas
 Mix a colony of test organism with 0.5 % deoxycholate
 If a string forms as loop is lifted from the slide, the test is
positive
 The string forms because bile lyses the organism and
releases DNA
 Vibrio is string test positive (>60 seconds)
 Aeromonas does not string

campylobacteraceae

 Curved, gram negative rod
 Beard hairs in the sink
 Catalase Positive
 Non-H2S Producer
 Non-hemolytic
 Motile
 Oxidase Positive
 Urea Negative
 DNase Negative
 Hippurate Positive
 Microaerophilic
 Stain best with Safranin instead of Carbol Fuchsin

campylobacter gram stain

route of transmission for campylobacter

Route of Transmission for Campylobacter
 Widely distributed in most warm-blooded animals
 Prevalent in food animals such as poultry, cattle, pigs, sheep and
ostriches; and in pets, including cats and dogs
 Has also been found in shellfish.
 Main route of transmission is generally believed to be foodborne, via
undercooked meat and meat products, as well as raw or contaminated
milk
 Contaminated water or ice is also a source of infection.
o Contact with contaminated water during recreational activities.
 Campylobacteriosis is a zoonosis, a disease transmitted to humans
from animals or animal products.
o Most often, carcasses or meat are contaminated by Campylobacter from faeces during
slaughtering
o In animals, Campylobacter seldom causes disease

specimen and culture media for campylobacter

 Same as Vibrio, Aeromonas, and Plesiomonas

campy agar

 Developed for isolating C. jejuni and C. coli
 Nutrient rich base
 Contain several antimicrobial agents to inhibit normal GI
flora
 Brucella agar base
 with 10% sheep blood and antibiotics
 Incubated in Microaerophilic Environment at 42o C

Guillain-Barre syndrome (GBS)

o Low incidence potential Sequelae
o Reactive, self-limited, autoimmune disease
o Campylobacter jejuni most frequent antecedent pathogen
o Immune response to specific O-antigens cross-reacts with
ganglioside surface components of peripheral nerves
(molecular or antigenic mimicry)
 Acute inflammatory demyelinating neuropathy (85% of
cases) from cross reaction with Schwann-cells or myelin
 Acute axonal forms of GBS (15% of cases) from molecular
mimicry of axonal membrane

helicobacter pylori

 Spiral, helical, curved or seagull shaped
 Gram negative bacilli
 Colonizes the epithelial cells of the stomach
 Associated with type B gastritis
 Most common cause of peptic (gastric) ulcers
 Results from the excessive production of urease
 With its flagella, the bacterium moves through the stomach
lumen and drills into the mucus gel layer of the stomach
 Produces adhesins which bind to membrane-associated lipids
and carbohydrates and help its adhesion to epithelial cells
 An example of this is the Lewis b antigen

additional helicobacter species

 H. cinaedi
Hampsters primary host

 H. fennelliae & CLO-3
Human primary host

helicobacter biochemicals

 Microaerophilic
 Oxidase +
 Catalase +
 Rapid urease Positive:
 Place in Christensen’s media for 2 hours
 A positive (pink color) within 15 minutes is
indicative of Helicobacter
 Motile

specimen for helicobacter

 Gastric biopsy specimen
 Minced tissue: use touch preparation for
gram stain

helicobacter additional testing

 CO2 Breath Test:
 patients drinks 14C labeled urea
 Urea is metabolized by urease
 Measure liberated 14CO2 in breath
 Other tests:
 Enzyme immunoassays (EIA)-Anti H pylori IgG
 Nucleic acid probes
 Stool antigen tests

helicobacter treatment

 Metronidazole
 Amoxicillin
 Clarithromycin
 Proton pump inhibitors
 Irreversibly blocking H+/K+ ATPase
 Prilosec, Prevacid, Nexium

tests and colony reactions

campy vs helicobactor