Chapter 16 : Aerobic Gram + Bacilli - can produce the toxin when infected with tox-carrying

β-phage
Diphtheria Toxin
- secreted by the bacterial cell and is nontoxic until exposed to
trypsin
● A fragment
- responsible for the cytotoxicity
● B fragment
- binds to receptors on the eukaryootic cells
- mediates th entry of fragment A into the cytoplasm
Toxin production:
● alkaline pH (7.8-8.0)
● Oxygen
● iron concentration
Clinical Infections:
● Respiratory diphtheria
Non–Spore-Forming, Nonbranching Catalase-Positive Bacilli
- Transmission: droplet, hand-to-mouth
- Incubation: 2-5 days
➢ CORYNEBACTERIUM
- Most common site: tonsil or pharynx
● lipophilic
● Cutaneous diphtheria
- often considered fastidious Ø grow slowly on standard
culture media
● nonlipophilic
Gram stain​:
- slightly curved, gram + rods with unparallel sides and
slightly wider ends
- “club shape” or coryneform

❖ C. diphtheriae
Virulence Factors​:
Diphtheria Toxin ​Lab Diagnosis
- major virulence factor - Pleomorphic
- produced by C. diphtheriae strains infected with - gram + bacilli
lysogenic β-phage - appears in palisades or forming “V”/ “L”angles
❖ C. ulcerans - stains irregularly with methylene blue (beaded appearance)
❖ C. pseudotuberculosis - Babes-Ernst granules
- metachromatic areas
 - accumulation of polymerized polyphosphates Lab Diagnosis: Toxi- genecity Test
Lab Diagnosis: Culture Characteristics ELEK TEST
- facultative anaerobe - immunodiffusion test for in vitro diphtheria toxin detection
- Loeffler medium - organisms (ctrl and unknowns) are streaked (single straight
- well demonstrated morphology line parallel to each other, 100 mm apart) on medium of low
- SBA iron content
- small zone of B-hemolysis - incubated at 35°C
Cystine-tellurite blood agar (CTBA) - examined after 18,24, and 48 hours
- Tinsdale medium modification
- selective and differential
- potassium tellurite
- inhibits non-coryneform bacteria
- brown halo
● C. diphtheriae
● C. ulcerans
● C. pseudotuberculosis
Lab Diagnosis: ID
- Catalase-positive
● C. amycolatum
- nonmotile
- most frequently recovered sp. from human clinical
● C. diphtheriae
material
- lack of urease production
- associated with prosthetic joint infection
- glucose and maltose fermenter
- reported to cause bloodstream infection and
- acid producing
endocarditis, typically in immunocompromised patients
- reduces nitrate to nitrite
or in patients in a health care setting
- Nonlipophilic
- Colonies : flat, dry. matte/waxy appearance

● C. jeikeium
- most common cause of corynebacterium- associated
prosthetic valve endocarditis in adults
- causes septicemia, meningitis,
- prosthetic joint infections, and skin complications such
as rash and subcutaneous nodules.
- Lipophilic
- strict aerobe
- Nonhemolytic
 - urease production - negative - urease production - positive
- nitrate reduction - negative
- vancomycin - drug of choice ● C. urealyticum
- most commonly associated with UTIs
● C. pseudodiphtheriticum - Lipophilic
- most frequently associated with respiratory tract - strict aerobe
infections in immunocompromised individuals or - nitrate - negative
patients with other underlying diseases - catalase - positive
- reported to cause endocarditis, urinary tract infections - urease - positive (w/n minutes on Christensen urea
(UTIs), and cutaneous wound infections slant)
- does not show the characteristic pleomorphic - Presumptive ID: pinpoint, nonhemolytic, white colonies
morphology
- urease production - positive ➢ ROTHIA
- nitrate reduction - positive ● R. mucilagonisa
- inked to bacteremia, endocarditis, pneumonia, and
● C. pseudotuberculosis other infections
- causes a granulomatous lymphadenitis ● R. dentocariosa
- produces a dermonecrotic toxin - found in saliva and supragigival plaque
- urease production - positive - isolated from patients with endocarditis
- yellowish-white colonies - SBA
Listeria monocytogenes
● C. striatum Virulence factors​:
- most commonly associated with device-related - hemolysin (listeriolysin O)
infection and has been reported in cases of - damages phagosome membrane
endocarditis, septic arthritis, meningitis, and - Catalase
pneumonia - superoxide dismutase
- nonlipophilic - phospholipase C
- pleomorphic - surface protein (p60)
- Colonies (after 24 hrs) : small, shiny, convex - induces phagocytosis
Clinical Infection​:
● C. ulcerans - infection commonly seen during third trimester on pregnant
- veterinary pathogen women
- human infxns acquired thru contact or by ingestion of - spontaneuos abortion and stillborn neonates
unpasteurized dairy products - food contamination (cheese, coleslaw, and chicken)
- grows well on Loeffler agar and SBA (narrow zone of Lab Diagnosis:
b-hemolysis) - gram positive coccobacillus→ coccoidal (subculture)
- nitrate reduction - negative - older cultures- gram variable
 - singly, short chains, pallisades

Non–Spore-Forming, Nonbranching Catalase-Negative Bacilli

➢ Erysipelothrix rhusiopathiae
- Gram-positive
- catalase negative
- Pleomorphic
Cold Enrichment - tendency to form lng
- broth inoculation filaments
- incubation at 4°C for several weeks - decolorizes easily (gram
- subcultures - weekly intervals variable)
Lab Diagnosis: ID - workers handling fish and
- Catalase-positive animal products are at risk
- motile (RT) Clinical infection:
- tumbling motility in wet mount prep ● Erysipeloid
- “umbrella” pattern - most common infection (skin) in humans
- B-hemolytic - esembles streptococcal
- hydrolyzes esculin erysipelas
- hydrolysis hippurate ● Septicemia
● cutaneous infection
- Rare
- exacerbation of the
erysipeloid lesion
- lasts longer
Culture:
CAMP test - nutrient broth w/ 1% glucose
- Rhodococcus equi - incubated at 5% CO2 @ 35°C
- “block” type hemolysis SBA colonies (24 hrs):
- distinguishes L. monocytogenes from other Listeria spp. - nonhemolytic
- pinpoint

➢ Arcanobacterium
● A. haemolyticum
- recovered from patients with pharyngitis (10-20 y.o)
- associated with soft tissue infections, sepsis,
endocarditis, and other infections
● A. pyogenes
- best known for causing infections in cattle
- causes sepsis and wound infections
● A. bernardiae
- associated with bacteremia, wound infections, UTIs
and septic arthritis
A. haemolyticum
- smal colonies with narrow zone of b- hemolysis (24-48 hrs)
- black opaque dot
- pitting of the agar beneath the colony
Reverse CAMP test (inhibition reaction)
- β-lysin–producing S. aureus is inhibited by a phospholipase D
- S. agalactiae (positive)
Gardnerella vaginalis
- gram positive (cell wall) rod/coccobacillus
- stains gram neg or gram-variable
- primarily known for bacterial vaginosis (BV) in humans
- results fom a reduction in the Lactobacillus population
in the vagina→ increase in pH
- UTIs in men and women
Lab Diagnosis:
- can be visualized in wet mounts of vaginal fluid
- “clue cells”
- large squamous epith cells w/ gram + and
gram-variable bacilli on edges
 Virulence Factor:
- superoxide disutase & catalase
- Nocobactin
- iron-chelating compound
Clinical Infection:
● Pulmonary
- inhalation of organism present in dust or soil
- N. cyriacigeorgica and N. farcinica
- progresses more rapidly than TB
- no sulfur granules
● Cutaneous
- N. brasiliensis
- actinomycotic mycetomas lesion
- with sulfur granules

Culture:
- grows best at 5%-7% CO2 (35-37°C)
- human blood bilayer Tween (HBT) agar - medium of choice
● SBA colonies:
- Pinpoint
- nonhemolytic
Culture:
● HBT agar:
- grows on buffered charcoal-yest extract (BCYE) agar
- b-hemolytic
- nonselective
- small, gray, opaque
Colonies:
- chalky, matte, velvety/powdery
Non–Spore-Forming, Branching
- dry, crumbly( similar to breadcrumbs)
Aerobic Actinomycetes
➢ Nocardia
- beaded gram-positive
- weakly acid-fast
- referred to as
modified acid-fast
positive
- weak acid
decolorizer
 - increases the conc of cAMP in host cells
● lethal factor (LF)
- Protease
- kills host cells
❏ PA + EF = edema
❏ PA + LF = death

Clinical infection:
● anthrax
○ cutaneous
○ inhalation/pulmonary
○ gastrointestinal Ø injectional
- recent form
- direct injection during the administration of
drugs of abuse Clinical Infection:
Clinical Infection: Cutaneous Anthrax
- wound contamination
➢ Tropheryma whipplei - small pimple for papule→ ring of
- agent of Whipple's dss vesicles→ erythematous ring
- more common in middle-aged men - Eschar
- identified from duodenal biopsy in 1991 - small dark area at the center of
thering
Spore-Forming, Nonbranching Catalase Positive Bacilli - sometimes referred to as
➢ Bacillus malignant pustule
- gram + or gram-variable - Painless
- divided into two groups: - no pus
● B. cereus ● Inhalation anthrax
● B. mycoides - also known as woolsorter's dss
- most relevant group ● Gastrointestinal anthrax
- ingestion of spores
B. cereus grp: B. anthracis - bloody diarrhea
- glutamic acid capsule - abdominal pain, nausea, anorexia, vomiting
- exotoxin ● Injectional anthrax
● protective antigen (PA) - soft tissue infection assoc with “skin popping”
- binding molecule for EF & LF
● edema factor (EF)
- adenyl cyclase
Lab Diagnosis: Mx - incubate for 3-6 hrs at 37°C
- large, square-ended
- gram-pos (young cultures)
- gram-variable (old or under B. cereus grp: B. cereus
nutritional stress) - common cause of food poisoning
- Singly ● diarrheal
- in chains - associated with ingestion of meat/poultry, veggies and
- bamboo rods pasta
appearance (unstained - incubation period: 8-16 hrs
central spore) ● Emetic
- capsule production - culture in - associated with ingestion of fried rice
CO2 incubator ● eye infection
Culture: - most common type of nonGI infxn
- nonhemolytic, large, gray, flat with
irregular margin
- “medusa head”
- beaten egg white characteristic
CDC lvl A testing protocol recommendation:
- PEA agar for stools in addition to
SBA and other commonly used
media.

“String of pearls” phenomenon

- noculate the isolate onto penicillin-containing (0.05 to 0.5
U/mL) agar