Gram (-) Rods

Enterobacteriaceae (Citrobacter, Enterobacter, Escherichia, Klebsiella, Morganella, Proteus, Providencia, Salmonella, Serratia, Shigella, Plesiomonas)

• Peritrichous flagella when motile

• Ferment glucose

• Reduce NO₃ to NO₂

• Most oxidase negative

• Antigens used in typing:

  • Flagella = H Ag

  • Envelope = K ag

  • Cell wall LPS = O Ag

  • All possess LPS endotoxin; some produce exotoxins

General characteristics of Enterbacteriaceae

• Indole and lactose (+)

• IMViC = + + - -

Key characteristics of Escherichia coli

What organism is the most common cause of UTI in females?

Escherichia coli

Intestinal infections of Escherichia coli:

• Enterotoxigenic E. coli (ETEC)

• Enteroinvasive E. coil

• Enterohemorrhagic E. coli (EHEC)

Which intestinal infection of E. coli has an LT toxin (heat labile) and ST toxin (heat stable)?

ETEC (Enterotoxigenic E. coli)

Which intestinal infection of E.coli penetrate epithelial cells in large intestine?

Enteroinvasive E. coli

Which intestinal infection of E. coil has:

• Shigella-like toxin

• Food poisoning associated with undercooked meat (hamburger)

• Hemolytic uremic syndrome (HUS)

EHEC (Enterohemorrhagic E. coli)

KI strains in Escherichia coli can cause…

neonatal meningitis

These are key characteristics of…

• Lactose (-)

• Non-motile

• Anaerogenic

Shigella

In Shigella, bacillary dysentery penetrates…

epithelial cells in small intestine

Types of Shigella

• S. dysenteriae (Group A)

• S. flexneri (Group B)

• S. boydii (Group C)

• S. sonnei (Group D)

Which Shigella is most severe?

S. dysenteriae (Group A)

Which Shigella is most common?

S. sonnei (Group D)

How to differentiate Enterobacter and Serratia?

Both Ornithine (+) and VP (+), but Enterobacter is DNAase (-) while Serratia is DNAase (+)

Which gram (-) organism is DNAase (+)?

Serratia

Which organism is negative for lysine, motility, and H₂S?

Shigella

Which organism(s) are Deaminase positive?

• Morganella

• Proteus

• Providencia

Opportunist; UTI, pneumonia; ampicillin-resistant

Klebsiellae

• Non-motile

• Has capsule

• Urea variable

• Ornithine negative

• VP positive

• Can cause lobar pneumoniA

Klebsiella

• Motile

• Ornithine positive

Enterobacter

• May produce red pigment

• DNase, gelatinase positive

• VP positive

Serratia

A large number of this organism is needed for infection (>100,000)

Salmonella

• H2S positive

• Lysine positive

• Indole negative

• Urea negative

Salmonella

Which Salmonella may cause septicemia?

S. cholerasuis

When is S. typhi blood positive and stool positive?

• Blood positive early - 1st week

• Stool positive in 2nd to 3rd week

Which Salmonella is ONPG pos (others neg)?

S. arizona

• Opportunist

• Lysine negative

• Similar to Salmonella biochemically

Citrobacter

• Urea positive

• Deaminase positive

Proteus

Which Proteus is most sensitive to penicillins and is indole negative?

P. mirabilis

Which Proteus is indole positive and H₂S positive?

P. vulgaris

Which Yersinia has an optimal growth of RT (cold enrichment), and is invasive and toxigenic?

Y. enterocolitica

Which Yersinia causes acute mesenteric lymphadenitis and “pseudotubercules”?

Y. pseudotuberculosis

Which Yersinia causes bubonic plague?

Y. pestis

• Oxidase positive

• Reclassified into Enterobacteriaceae

• Lophotrichous flagella

• Associated with diarrheal disease

Plesiomonas

What biochemical test uses reagent tetramethyl p-phenylenediamine dihyrochloride and turns purple when positive?

Oxidase test

What biochemical test uses reagents α-naphthylamine and sulfanilic acid and turns pink when positive?

Nitrate test

What biochemical test detects B-d-galactosidase, uses reagent O-nitrophenyl-B-d-galactopyranoside, and turns yellow when positive?

ONPG test

Yellow butt in TSI means:

Glucose fermented

Yellow slant in TSI means:

Lactose or sucrose fermented

Red slant in TSI means:

Neither lactose nor sucrose fermented

Black butt in TSI means:

H₂S produced

TSI contains:

0.1% glucose, 1% sucrose, and 1% lactose

KIA contains:

Same as TSI but only with glucose and lactose, no sucrose

Which biochemical test uses media green and turns blue when positive?

Citrate test

Which biochemical test measures ability to decarboxylate amino acids and has bromcresol purple as indicator dye?

Decarboxylase test

• Has H2S indicator

• 0.1% glucose and 1% lysine

• Positive = purple butt

• Slant turns red for lysine deaminase

LIA (Lysine iron agar)

• 0.1% glucose and 1% ornithine

• Positive = purple butt

• Also tests for motility and indole (Kovac’s)

MIO (Motility-indole-ornithine)

Which biochemical test uses reagent p-dimethyl-aminobenzaldehyde and turns pink when positive?

Indole test

Which biochemical test hydrolyzes to ammonia and CO₂ and the phenol red indicator turns pink if positive?

Urease test

Which biochemical test detects acetylmethylcarbinol (acetoin), uses reagents KOH and α-naphthol, and turns pink when positive?

VP (Voges-Proskauer)

Which biochemical test uses reagent ferric chloride and turns green when positive?

Phenylalanine deaminase

A/A (Sun over Desert)
SEEK shelter in the hot desert

• Serratia

• Escherchia

• Enterobacter

• Klebsiella

A/A, H₂S + (Sun over Desert)

Wear a CAP to protect yourself from the geyser (H₂S+)

• Citrobacter

• Arizona

• Proteus

K/A, H₂S + (Moon over Desert)
A CASE of firecrackers going off smells like sulfur (H₂S +)

• Citrobacter

• Arizona

• Salmonella

• Edwardsiella

K/A (Moon over Desert)

Sk( c ) ipppy Coyote howls at the moon

• Shigella

• Citrobacter

• Providencia

• Plesiomonas

• Proteus

• Yersinia

K/K (Moon over Mountain)

Pseudomonas (Psuedo “moon” as)

How to differentiate the Gram negative rods that are A/A?

• Serratia (sugars)

• Escherichia (indole +)

• Enterobacter (ODC +, sugars)

• Klebsiella (ODC -)

How to differentiate the Gram negative rods that are A/A, H₂S+?

• Citrobacter (LDC -)

• Arizona (LDC +)

• Proteus (urea +, deam +)

How to differentiate the Gram negative rods that are K/A, H₂S+?

• Salmonella (mal -, ONPG -)

• Citrobacter (LDC -)

• Edwardsiella (indole +, LDC +)

How to differentiate the Gram negative rods that are K/A?

• Shigella (citrate -, non-motile)

• Citrobacter (citrate +)

• Providencia (deam +)

• Plesiomonas (oxidase +)

• Proteus ( urea +, deam +)

• Yersinia (small colonies, urea +, deam -)

How to differentiate the Gram negative rods that are K/K?

• Pseudomonas (ox +, blue-green pigment, growth at 42^oC, growth in centrimide)

Possible Enterobacteriacae that is A/A, H₂S+

• Proteus vulgaris

• Proteus mirabilis

• Arizona

• Citrobacter

Possible Enterobacteriacae that is A/A, H₂S-

• Citrobacter

• E. coli

• Enterobacter cloacae

• Enterobacter agglomerans

• Klebsiella

• Serratia

• Enterobacter aerogenes

• Enterobacter hafniae

Possible Enterobacteriacae that is A/A, H₂S+, Urea+, Deaminase+

• Proteus vulgaris (Indole +)

• Proteus mirabilis (Indole -)

Possible Enterobacteriacae that is A/A, H₂S+, Urea -, Deaminase -

• Arizona (LDC +)

• Citrobacter (LDC -)

Possible Enterobacteriacae that is A/A, H₂S-, Indole +

• Citrobacter (Citrate +)

• E. coli (Citrate -)

Possible Enterobacteriacae that is A/A, H₂S+, Indole -, LDC -

• Enterobacter cloacae (ODC+)

• Enterobacter agglomerans (ODC-)

Possible Enterobacteriacae that is A/A, H₂S+, Indole -, LDC+, ODC-

Klebsiella

Possible Enterobacteriacae that is A/A, H₂S+, Indole -, LDC+, ODC+, Gelatin+

Serratia

Possible Enterobacteriacae that is A/A, H₂S+, Indole -, LDC+, ODC+, Gelatin -

• Enterobacter aerogenes (Lactose, Citrate+)

• Enterobacter hafniae (Lactose, Citrate -)

Possible Enterobacteriacae that is K/A, H₂S+, Urea or Phe Deam+

• Proteus vulgaris (Indole +)

• Proteus mirabilis (Indole -)

Possible Enterobacteriacae that is K/A, H₂S+, Urea or Phe Deam -, Indole +

Edwardsiella

Possible Enterobacteriacae that is K/A, H₂S+, Indole -

• Citrobacter freundii (LDC -)

• Salmonella sp. (LDC+, ODC+)

• Salmonella typhii (LDC+, ODC-)

• Oxidase positive (some exceptions: Stenotrophomonas

• May not grow on MacConkey’s agar

• Glucose NOT fermented

Gram Negative Non-Fermenters

• Semi-solid

• Low protein

• Bromthymol blue indicator

• Incubate tubes in pairs

  • Open - oxygen

  • Closed - oxygen-restricted

  • Pair of tubes yellow = fermenter

  • Pair of tubes green = asaccharolytic (non-utilizer)

  • Open tube yellow and closed tube green = oxidizer

Oxidative Fermentative (OF) media

• Most common

• Oxidase positive

• Growth on MAC (lactose negative)

• Oxidizer (OF)

• Polymixin B susceptible

• Opportunistic pathogen

• Drug resistant

• Infections: Burns, pneumonia, swimmer’s ear, eye infections, UTI

Pseudomonas aeruginosa

• Oxidase weakly positive

• Growth on MAC

• Oxidizer (OF)

• Polymixin B resistant

• Opportunistic pathogen

  • Serious infection in CF patients

  • Yellow-brown insoluble pigment

Burkholderia cepacia

Gram Negative, Oxidase Positive Fermenters

• Vibrio cholera

• Vibrio parahemolyticus

• Vibrio vulnificus

• Aeromonas

• Campylobacter jejuni

• Helicobacter pylori

• Curved rods with polar flagella

• TCBS - Selective for this organism

• BAP - Hemolytic colonies

• Lactose negative

• NO₃ positive (NO₃ to NO₂)

Vibrio

• Gastroenteritis (rapid onset 3-10 hrs; profuse diarrhea)

• Need high concentration of organism unless hypochlorohydric

• Stools contain mucus flecks (described as “rice" water” stools)

• Yellow on TCBS

V. cholera

• Green on TCBS

• Enteritis

V. parahemolytics

• Green on TCBS

• Septicemia - can kill immunocompromised or diabetics

V. vulnificus

• Motile by polar flagella

• Most are indole positive

• Growth on MAC

• Cellulitis, wound infections and diarrhea

Aeromonas

• Small curved rods, “seagull appearance” (light staining)

• Miccroaerophilic, 2-4 days for growth

• Growth on C agar

• Found in raw poultry and contaminated water (like Salmonella)

• Erythromycin or tetracycline for treatment

• Biochemical rxns

  • Catalase positive

  • Oxidase positive

  • Hippurate positive

Campylobacter jejuni

• Associated with gastric and duodenal ulcers

• Produces large amounts of urease

Heliobacter pylori