Gram Negative Bacilli (Enterobacteriaceae)

Front 1

Enterobacteriaceae (Enterics) are facultative (aerobes/anaerobes)

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anaerobes

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Fecal coliforms

Back 2

another name for the enterics found commonly as part of intestinal flora

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Pirmary intestinal pathogens

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Salmonella, shigella, yersinia

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Opportunistic pathogens

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Normally part of intestinal flora that may produce infection outside the intestine when the immune system is compromised

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Enterics produce lethal (endotoxin/exotoxin)

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Endotoxin, called LPS composed of 3 distinct parts

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What are the 3 parts of an LPS?

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O polysaccharide
Polysaccharide core
Lipid A

Front 7

The flagella is also called the _ Antigen

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H antigen (these are heat liable)

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The capsule is also called the _ antigen

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K antigen (overlay the surface O antigen and may block agglutination by O specific antisera)

Front 9

In salmonella, the envelope or capsule is called the

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Vi antigen

Front 10

What are the modes of infection for enterics?

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Contaminated food and water (Salmonella, Shigella, Yersinia enterocolitica, Escherichida coli)
Endogenous infection (UTI - esp female, bacterial peritonitis, abdominal abscess)
Abnormal host colonization (nosocomial pneumonia)
Transfer between debilitated patients
Insect (flea) vector) - unique for Yersinia

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Enterobacterieae opportunistic infections include

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Gram neg sepsis, UTI, pneumonia (older or kids), abdominal sepsis, meningitis, spontaneous bacterial peritonitis, endocarditis

Front 12

Why is selective media useful for identifying Gram neg bacteria

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Bile salts and dyes suppress growth of G+ org's, and differentiate species based on lactose fermentation or H2S production

Front 13

What are the charatacteristics for all enterobacterieae?

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All ferment glucose
All reduce nitrates to nitrites
All are oxidase negative
All (except Klebsiella and Shigella) are motile

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Enterobacterieae opportunistic infections include

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Gram neg sepsis, UTI, pneumonia (older or kids), abdominal sepsis, meningitis, spontaneous bacterial peritonitis, endocarditis

Front 15

What is serotyping?

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Identification based on antigenic features

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How are enterics identified?

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Based on growth charactaristics (using SELECTIVE media)
Biochemical testing (based on matabolism and production of certain acids)
Serotyping (antigenic testing)

Front 17

What type of media should one begin with when dealing with enterics?

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Selective media (MacConkey, HE, XLD agars)

Front 18

Why are non-selective agars an issue for growing enterobacterieae?

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Colonies all look virtually identical on BAP or CBA and may or may not be hemolytic

Front 19

Why is selective media useful for identifying Gram neg bacteria

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Bile salts and dyes suppress growth of G+ org's, and differentiate species based on lactose fermentation or H2S production

Front 20

Why are non-selective agars an issue for growing enterobacterieae?

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Colonies all look virtually identical on BAP or CBA and may or may not be hemolytic

Front 21

Why is selective media useful for identifying Gram neg bacteria

Back 21

Bile salts and dyes suppress growth of G+ org's, and differentiate species based on lactose fermentation or H2S production

Front 22

What are three ways bacteria can utilize carbohydrates?

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oxidation (aerobic)
fermentation (anaerobic)
both (facultative anaerobes)

Front 23

What are the most common carbohydrates used by enterics?

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glucose, lactose, maltose, sucrose, arabinose and rhamnose

Front 24

What are the most common carbohydrates used by enterics?

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glucose, lactose, maltose, sucrose, arabinose and rhamnose

Front 25

What are three ways bacteria can utilize carbohydrates?

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oxidation (aerobic)
fermentation (anaerobic)
both (facultative anaerobes)

Front 26

Asacchrolytic bacteria

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Do not use carbohydrates for metabolism

Front 27

Asacchrolytic bacteria

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Do not use carbohydrates for metabolism

Front 28

What are the most common carbohydrates used by enterics?

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glucose, lactose, maltose, sucrose, arabinose and rhamnose

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Asacchrolytic bacteria

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Do not use carbohydrates for metabolism

Front 30

Oxidation-fermentation tests

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determine the ability of a bacteria to utilize a specific carbohydrate incorporated into a basal medium aerobically or anaerobically

Front 31

What are the two different pathways pyruvic acid enters after glycolysis?

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Mixed acid fermentation pathway
Butylene glycol pathway

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Mixed acid fermentation pathway

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Pyruvic acid (and H+ + NADH + ATP) made after glucose fermentation is converted into lactose and NAD+ (that is fed back into glycolytic pathway)

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Butylene glycol pathway

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Pyruvate is converted into CO2, acetaldehyde, NAD+. Acetaldehyde is converted to ethanol.

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How would one test for mixed acid fermentation bacteria?

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Methyl red test (MR test):
positive: red (acidic)
negative: yellow (neutral)

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What test is used to test for butylene glycol pathways?

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alpha-naphthol and 40% KOH (VP test):
positive: red (presence of alcholol)
negative: no color change or pink (little to no alcohol)

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Durham tubes

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detect small amounts of gas production

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TSI or KIA slants

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detect only large amounts of gas production

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KIA agar is differentiated from TSI by the

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absence of sucrose (only lactose and glucose)

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MacConkey agar

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selects for gram negative organisms. pH<6.8 causes the dye to turn red.

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HE (Hektoen enteric) agar

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High bile salt inhibits nonenteric organisms - differentiates pathogenic salmonella and shigella

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XLD (Xylose Lysine Deoxycholate) agar

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Contains xylose, lactose, sucrose and lysine. Salmonella fermentates sugar, will have purple colonies

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On XLD agar, proteus vulgaris will show

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black on a yellow agar

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What two enzymes are necessary for lactose fermentation?

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beta-galactosidase and beta-galactose permease

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ONPG test

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Quickly detects lactose fermentation (Galactose-Glucose replaced by Galactose-Nitrophenyl)
negative: cloudy white
positive: yellow

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OPNG test detects beta-galactose permease activity (T/F)

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False, only detects beta-galactosidase activity

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Cytochrome oxidase

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tests for oxidase presence, enterics will all be negative

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Nitrate reducion test

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Addition of alpha-naphthylamine and sulfanilic acid
red: positive (nitrate is reduced)
not red: negative (nitrate is not reduced)

Negative tests must then be treated with zinc dust to test for a false negative (zinc reduces any residual nitrates to confirm a negative test)

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IMViC test

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Detects presence of tryptophanase
Methyl red test looks for strong acids.
Voges Proskauer test: detects acetoin and butylene glycol from glucose fermentation
Citrate utilization: determines organisms ability to utilize sodium citrate

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IMViC test results for E. coli?

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I: +
M: +
V: -
C: -

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Urease test

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determines an organisms's ability to metabolize urea and release ammonia

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Phenylalanine deaminase test

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detects oxidative deamination

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IMViC results for E. coli?

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pos/pos/neg/neg

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What two tribes of pathogenic enterobacterieae are very similar?

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Eschirichiae and Shigella

Front 54

What antigens are used to test for E. coli?

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O antigen (LPS)
H antigen (flagella)
K antigen (capsule)

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TSI is A/A + gas
LIA is K/K
IMViC: pos/pos/neg/neg
Urea: neg
Motility: pos
Phenylalanine: neg

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E. Coli

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What is a very common site of infection for E. coli?

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Urinary tract

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What is an ETEC

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Enterotoxigenic: traveler's diarrhea, watery stool, abdominal cramps, (NO FEVER)

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EPEC

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Enteropathogenic E. Coli - seen commonly in babies, fever, vomiting, diarrhea, NO BLOOD IN STOOL

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EIEC

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Enteroinvasive E. coli - FEVER, blood in stool, mucus, white cells in stoll

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EaggEC

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Enteroaggregative e. coli - watery diarrhea, vomiting

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EHEC

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enterohemorrhagic e.coli - commonly 0157:H7

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O157:H7 produces what results on MAC?

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pink colonies

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What is the strain of E. coli currently spreading through Europe?

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O104:H4: shigella like toxins

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All species of Shigella possess which antigen?

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O antigen. Shigella never has an H antigen and sometimes has K antigen.

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How are Shigella species divided?

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Four serologic groups based on the kind of O antigen.

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What are the four clinically significant species of Shigella?

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S. dysenteriae (serogroup A), S. flexneri (B), S. boydii (C), S. sonnei (D)

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Characterize Shigella sonnei based on lactose fermentation, motility, gas production, ureaase, H2S, lysine, arginine and ornithine decarboxylation.

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Lactose negative
Nonmotile
No gas production
No urease activity
No H2S production
Do not decarboxylate lysine or arginine
DO decarboxylate ornithine (slowly ferment lactose)

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What are some symptoms of genums Shigella?

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dysentery, seizures, HUS (hemolytic uremic syndrome)

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Dysentery

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High fever, chills, abdominal cramps, pain and TENESMUS (unique to shigella)

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Tenesmus

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Feels like constipation but there is no actual stool is in intestine

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Shigella sonnei

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Organism most commonly associated with diarrheal disease in the US. Asymptomatic or mild symptom.

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S. dysenteriae

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Rare in developed nations, but very virulent and common in 3rd world. Cannot ferment mannitol.

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S. flexneri

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Mild to asymptomatic, difficult to distinguish

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How would a person commonly contract Edwardsiella tarda?

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Usually contracted from fresh water fish or reptiles.

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Pathogenesis and symptoms of E. tarda

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Bacteremia, wound infection (usually water related) Septicemia seems to be more severe when iron is abundant

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What are the biochemical charactaristics of Edwardsielleae?

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No lactose fermentation
H2S producer
**Lysine and ornithine decarboxylation**

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How do Edwardsielleae look on HE and MAC agar?

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MAC: Black
HE: colorless

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What are the two species of Salmonelaa?

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S. enteritica and S. bongori

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What happened to the many species of Salmonella?

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Many were rolled into S. enterica and are now subspecies strains of that species. (ie. S. typhi is now S. enterica ss serotype typhi)

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Does Salmonella ferment lactose?

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No

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Is Salmonella motile?

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Yes

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Does Salmonella decarboxylate lysine or ornithine?

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Lysine for both
Salmonella: Non-typhi is pos for ornithine, typhi is neg

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Is Salmonellla Citrate positive or negative?

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Positive

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Virulence factors of Salmonellae?

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Endotoxins - play a role in intracellular survival
Capsule - anti-phagocytosis
Adhesions - attach to epithelial cells
Type III secretion systems

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What type of disease is caused by Salmonella?

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Enterics fevers (typhoid fever) and gastroenteritis (food poisoning) - also non-typhoidal septicemia. Salmonella can be a carrier.

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What is the most common cause of food poisoning in the US?

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Salmonella enterica ss enterica (formerly S. enteritidis)

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What is commonly contaminated by S. enterica ss enterica (S. enteritidis)

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Contaminated food, water and milk. Common: uncooked eggs

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Typhoid fever

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Caused by Salmonella enterica ss enterica serotype typhi and serotype paratyphi. First 2 weeks: fever, constipation, bacteria in blood but not in stool.
After 2 weeks: diarrhea begins, blood culture negative and stool positive

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What's the difference between gastroenteritis and enteric fevers?

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Enteric fever (typhoid) involve the spread of the bacteria into the blood, causing bacteremia.

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Non-typhoidal bacteremia or septicemia

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Occurs without GI symptoms; high fever and positive blood culture

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Carrier state (salmonella)

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Organisms centralized in gall bladder (removal necessary) and patient will have organism in stool for up to a year after having typhoid fever

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Salmonella:
H2S:
Citrate:
Ornithine:
Gas:

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H2S: trace
Citrate: neg
Ornithine: neg
Gas: none

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What media are best for Salmonella isolation?

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MAC, HE, XLD

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Biochemical results of S. enterica ss arizonae

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Malonate positive
Dulcitol neg
OPNG pos
Lactose fermentation (some)

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Citrobactereae

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grow on citrate medium
methyl red positive
lactose fermenter
urease pos
looks similar to E. Coli on MAC

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Lactose fermenters show up which color on MAC agar?

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pink

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What diseases are associated with C. freundii?

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Nosocomial infection
UTI
Pneumonias

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C. koseri?

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Associated with infant meningitis and brain abscess. Most infants will not survive infection and those that do will have neurological damage.

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What tests can be used to differentiate C. freundii?

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H2S positive
Urease activity
Lysine decarboxylase negative

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What are the most common species of Klebsiella?

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K. pneumonieae and K oxytoca

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Klebsiella is motile (T/F)

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False

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Some characteristics of Klebsiella?

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H2S negative
MR negative
VP positive
Growth on citrate and KCN
Urease activity positive
Most do not produce indole (K. oxytoca does)

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K. pneumonieae ss pneumonieae

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frequently recovered lower resp. tract infection.
Classic pneumoni: necrosis, hemorrhage, thick brick red sputum

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K. oxytoca

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similar symptoms to K. pneumonieae

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What is special about Klebsiella?

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Produces large mucoid colonies on primary isolation media (because of polysaccharide capsule)

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How does one differentiate K. oxytoca and K. pneumonieae?

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K. oxytoca: Indole positive
K. pneumonieae: Indole negative

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What is ESBL?

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Extended spectrum beta lactamase - antibiotic resistance encoding plasmid present in non-K. pneumonieae species of Klebsiella

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What is an important test to perform when dealing with Klebsiella for patient treatment?

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Identification of antibiotic susceptability and resistance

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Two common species of Enterobacter?

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E. aerogenes and E. cloacae

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Is Enterobacter ornithine decarboxylase positive or negative? Is it motile?

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Positive; Motile

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How are E. aerogenes and E.cloacae differentiated?

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E. aerogenes
Lys: +
arg: -

E. cloaceae:
lys -
arg: +

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Cronobacter sakazakii

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Related to infant meningitis. Bright yellow on TSA.

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Most common species of Pantoea?

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P. agglomerans

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Where is P. agglomerans commonly isolated from?

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P. agglomerans is commonly isolated from plant matter and animal and human waste.

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What is the pathogenesis of P. agglomerans?

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Opportunist; causes septicemia, UTI, wound infection

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How does P. agglomerans utilize lysine arginine or ornithine?

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Does not produce any of these.

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How does P. agglomerans frow on MAC and SBA?

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MAC: pimply red
SBA: yellow

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what are the two Hafnia (Klebsielleae) species?

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one species but two biotype Hafnia alvei and Hafnia alvei biotype 1

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Common species of Serratia?

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S. marcescens

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What enzymes are charactarized by Serratia

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Lipase, gelatinase and DNAase

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Color of S. marcescens on MAC?

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pink to red pigmented colonies on mac

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Proteus are (normal/abnormal) in intestinal flora

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normal flora

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Proteus is a lactose fermenter (T/F)

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False; proteus does not ferment lactose.

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Proteus is phenylalanine deamninase (positive/negative)

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positive

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Which species of Proteus are commonly pathogenic?

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P. mirabilis and P. vulgaris

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Proteus are commonly found in nosocomial cases (T/F)

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True. Proteus are responsible for ~3% of nosocomial infections in the US.

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What colony morphology is charactaristic of Proteus on SBA?

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Swarming colonies on non-selective media with a distinct odor.

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Proteus are H2S (positive/negative)

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Positive

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Proteus are urease (pos/neg)

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pos

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What test is used to diff P. mirabilis and P. vulgaris?

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Indole and ornithine
P. mirabilis is indole neg and orn pos
P. vulgaris is indole pos and orn neg

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What are the two subspecies of Morganella morganii?

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M. morganii ss morganii
M. morganii ss sibonii

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What are the common charactaristics of Providencia?

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Lactose negative
PDA positive
Indole positive
Citrate positive
H2S negative

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P. stuartii is commonly isolated from?

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From burn unit patients

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P rettgeri

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Common in nosocomial, traveler's diarrhea and

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How does one diff P. stuartii and P. rettgeri?

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Urease and arabitol fermentation
P. stuartii: arabitol neg and urease neg
P . rettgeri: arabitol pos and urease pos

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Yersinia pestis is caused by..

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Caused by fleas carried by rats (bubonic plague)

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What are the two forms of Yersinia pestis infection in humans?

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Bubonic (glandular) - 2 - 5 days after infection, high fever, swollen lymph nodes, black spots on skin

Pneumonic - secondary to bubonic: bacteremia, sepsis, respiratory infection

Front 138

What is the optimal temperature for Y. pestis growth?

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25 - 30 C

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Y. pestis is ornithine (neg/pos)

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Negative (also urease negative)

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How is Y. entercolitica transmitted?

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Via pets, contaminated food (esp. meat, chocolate milk, water), infected infusion

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Why is Y. entercolitica so dangerous?

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Can survive in colder temperatures

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What four conditions are associated with Y. entercolitica?

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Acute enteritis: *fever*, abd. pain, nausea, diarrhea, bloody stool (TODDLERS AND CHILDREN)

Mimic acute appendicitis (ADULTS)

Arthritis - most common extraintestinal form (usually followed by GI)

Erythema nodosum - tender red itchy nodules

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What are some special ID charactaristics of Y. entercolitis?

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Growth at 25 - 30 C, loses motility at 35+ C. Better growth with cold enrichment (4 C)

Front 144

What selective media is used for Y. entercolitica?

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CIN: produces red "bull's eye" colonies

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Why is Y. pseudotuberculosis so dangerous?

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Causes mesenteric lymphadenitis, very high mortality rate in children.

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HACEK

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Haemophilus aphrophilus/paraphrophilus
Actinobacillus actinomycetemcomitans
Cardiobacterium hominis
Eikenella corrodens
Kingella spp

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General char's of HACEK

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gram neg bacilli, oropharyngeal and urogenital flora
Pathogenicity: endocarditis, bacteremia, mixed-flora wound infections

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How long do HACEK organisms take to grow?

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at least 2-3 days sometimes 2 weeks

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What medium is used to grow HACEK?

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SBA or chocolate agar; MAC, HEK, etc do not work

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What are causes and symptoms of H. aphrophilus/paraphrophilus infection?

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endocarditis, brain abscess, meningitis, osteomyelitis, soft tissue infection - caued by dental disease, animal bites and skin piercing

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ONPG, catalase and acid for H. aphrophilus

Back 151

Positive; negative; positive

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A. actinomycetemcomitans symptoms

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Abscesses, bacterial endocarditis, periodontitis, Papillion-Lefevre syndrome

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Papillon-Lefevre syndromeassociated with A. actinomcetemcomitans

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Hyperkeratosis of palms and soles, and periodontal destruction

Front 154

Portals of entry for A. actinomcytemcomitans

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oral lesions, skin abrasions, pulmonary infection, thoracotomy, UT instruments (ie. catheter, cystoscope)

Front 155

A. actinomcytemcomitans virulence factors

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Leukotoxin - kill neutrophils, monocytes and T-lymphocytes by damaging chromosomal DNA

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What is special about A. actinomcyetomcomitans colonies?

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Star-shaped area after 1 wk on SBA/chocolate

Front 157

A. ureae colonies show

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distinct filaments

Front 158

Cardiobacterium hominis is mainly associated with which condition?

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endocarditis primarily,followed by dental disease

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How do you get a good gram stain of C. hominis? good culture?

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Must be spun down first; requires subcultures since initial culture growth is usually weak

Front 160

Eikenella corrodens route of infection?

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human bite, head and neck infection, frequent in the immunosuppressed and IV drug users.

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Eikenella virulence factors?

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Pili, slime layer (prevents phagocytosis) proteins for adherence

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Eikenella cultures do what to agar?

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Pitting

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Eikenella smells like...

Back 163

bleach

Front 164

Kingella denitrificans can be confused with N. gonorrheae. What test is used to distinguish the two?

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K. dentrificans ans catalase neg and superoxol neg

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K. dentrificans colonies are

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smooth and opaque

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Kingella kingae

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Associated with child-to-child transmission

Front 167

K. kingae morphology on SBA

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Plump GN rods, Beta hemolytics, pitting

Front 168

Suttonella indologenes

Back 168

similar to K. kingae except for indol positive

Front 169

S. indologenes is resistant to which antibiotics

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clinidamcyin, erythromycin, lincomycin, vancomycin

Front 170

HACEK stands for?

Back 170

H = Haemophilus aphrophilus/paraphrophilus
A = Actinobacillus actinomycetomcomitans
C = Cardiobacterium hominis
E = Eikenella corrodens
K = Kingella spp

Front 171

HACEK organisms generally cause which disorders?

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Endocarditis, bacteremia, mixed-flora wound infections

Front 172

HACEK is a normal flora of what part of the body?

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Oropharyngeal and urogenital tracts

Front 173

What are the growth requirements for HACEK?

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Enriched media (ie. SBA, chocolate) capnophilic (5-7% CO2) and time (2-3 day minimum, sometimes weeks)

Front 174

What is the normal habitat of H. aphrophilus/paraphrophius in humans?

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Oral cavity (dental plaque) and upper resp. tract

Front 175

What conditions are assocaited with H. aphrophilus/paraphrophilus?

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Endocarditis (esp. with valvular heart disease) brain abscess, meningitis, osteomyelitis, soft tissue infection

Front 176

What is mode of transmission for H. aphrophilus/paraphrophilus?

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Dental disease, dog bites, piercings

Front 177

What distinguishes H. aphrophilus/paraphrophilus from other HACEK?

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ONPG positive
catalase negative
acid producer

Front 178

How does one distiguish H. aphrophilus from paraphrophilus?

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H. aphrophilus does NOT require X or V factor

H. paraphrophilus requires V factor

Neither require X factor

Front 179

What are H. aphrophilus/paraphrophilus susceptible to/resistant to?

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Tetracycline, chloramphenical, streptomycin, 2nd generation cephalosporins, quinolones

Resistant to PCN and ampicillin.

Front 180

Actinobacillus actinomycetmcomitans is involved in what diseases/conditions?

Back 180

Subactute bacterial endocarditis (with hx of heart disease)

Juvenile periodontitis

Front 181

What organism is associated with Papillon-Lefevre syndrome?

Back 181

Actinobacillus actinomycetmcomitans

Front 182

What are the portals of entry for Actinobacillus actinomycetmcomitans?

Back 182

Oral lesions, skin abrasions, pulomary infections, thoracotomy, catheters

Front 183

Virulence factors of Actinobacillus actinomycetmcomitans

Back 183

Leukotoxins that inhibit immune response by killing WBC's

Front 184

Growth requirements of Actinobacillus actinomycetmcomitans?

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Enriched media, at least 48 hrs, capnophilic env.

No X or V factor required.

Front 185

Which bacteria is commonly found canine oral cavities?

Back 185

C. canimorsus

Front 186

What are the symptoms of Capnocytophaga?

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Fulminant septicemia and peripheal gangrene

Front 187

How can Capnocytophaga cause juvenile periodontitis?

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When it's with an actinomycetemcomitans infection

Front 188

Salidases/aminopepsidases

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Capnocytophaga virulence proteins

Front 189

What mode of xmission is associated with Streptobacillus moniliformis

Back 189

Rodent

Front 190

Is Streptobacillus motile?

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No

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What special measures need to be taken to culture Streptobacillus moniliformis?

Back 191

Need to be cultured in live host

Front 192

Haverhill fever

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Caused by S. moniliformis. Fever, GI symptoms, rash. MIMICS LYME'S DISEASE

Front 193

The selective agar used for Bordatella is..

Back 193

Bordet-Gengot (high starch)
Regen-Lowe agar (mother of pearl colonies on this)

Front 194

The slective agar for Francisellais...

Back 194

BCYE