9/9/2010

1

Corynebacterium

Bacillus

General

● Gram-positive

● Aerobic or facultative anaerobic

● Catalase positive, oxidase negative, nonspore forming

● Nonmotile except C. aquaticum

- Pathogenic species are nonmotile

Gram

+

O2

Catalase + Oxidase -

9/9/2010

2

Morphology

● Pleomorphic rod shaped

- Individual cells may be swollen, clubbed,

Palisades of parallel cells

Angular clusters resembling “Chinese letters”

八eight

人people

Memory Game

9/9/2010

3

Presumptive Identification

Gram stain

Positive ?Negative ?

Morphology

Cocci ? Bacilli (rod)?

Spore formation

Positive ? Negative ?Bacillus

Catalase

Positive ? Negative ?

Motility

Positive ? Negative ?Listeria

C. renale

● Three species (renale, cystitidis and pilosum) were formerly

classified as a single species: “renale is the major”

● Morphology: short rods, thicker at one end

● Growth: favored by blood or serum, grows well in sterile urine

● “Urease” positive: (NH2)2CO + H2O → CO2 + 2NH3

● Habitat: Widely distributed in the urethra and prepuce in normal

cattle and sheep (considered as normal flora)

● Transmission: May be venereal and indirect

● Disease

- Cattle: Infections are almost always in cows

Ascending infection of the urinary tract, cystitis, pyelonephritis

Enlarged kidney, bloody urine

- Sheep: ↑Protein (diet) Urea (urine) Ammonia (by urease)

Ammonia irritates to the mucosa of prepuce of male sheep

(mild irritation to severe inflammation and blockage of the prepuce)

● Treatment: Penicillin (effective if given early in the disease)

9/9/2010

4

C. renale: Story

Habitat: Normal flora in

the lower urogenital tract

(Urethra and Prepuce)

Bladder

Kidney

Pennis (pizzle rot)

Venereally

Well surviving in soil

Indirectly ???

Penicillin

C. pseudotuberculosis

● Morphology: Pleomorphic, short rod, coccoid in some isolates

● Biochemical: Urease pos.

● Pathogenesis

- A facultative intracellular parasite

- Phospholipase D: an exotoxin associated with spread

within the host

- Surface lipid: resistance to killing by macrophages

● Transmission

- Discharges from pus containing bacteria

- The bacteria residing in soil may enter via abraded mucosa or

inhalation

9/9/2010

5

C. pseudotuberculosis:

In sheep and goats

● Caseous lymphadenitis (inflammation of a lymph node)

- Also called “Cheesy gland”: dryish pus

- Two forms

1) Superficial (abscesses of lymph node)

:Visible under the skin near the lymph glands

2) Visceral (abscesses of

internal organs)

- Metastatic lesions

- Inhalation

lung infection

● Ulcerative lymphangitis (inflammation of a lymphatic vessel)

- Infection occurs in lower limbs

- Usually, only one leg is involved

● Pectoral abscesses

- Develop abscesses in their pectoral muscles

Swell and resemble a pigeon's chest “Pigeon fever” (horses)

Bovine ulcerative lymphangitis

Hi guy!, are we siblings?

C. pseudotuberculosis:

In horses and cattle

9/9/2010

6

C. pseudotuberculosis

● Diagnosis

- Isolation of C. pseudotuberculosis

- ELISA

- PCR

● Treatment

- Drainage of the abscesses

- Antibiotic control is not easy:

because bacteria stay protected inside abscesses

- Penicillin may prolong the disease by delaying abscess maturation

● Vaccination

- Vaccination is recommended

- Live attenuated vaccine from Brazil

with good (83%) protection

C. pseudotuberculosis

Direct contact

Bacterial release to soil Contact or inhalation

Ulcerative lymphangitisPigeon fever

Caseous lymphadenitis

Antibioticslittle effect

Vaccination is advised

Bang!

9/9/2010

7

C. diphtheriae

● Human pathogen

-Diphtheria: Upper respiratory infection especially

in kids under 4 yrs

● Diphtheria toxin

- Prevents protein synthesis cause cell death

- Associated with bacteriophage encoding

the toxin (tox+)

● Vaccination: DPT (Diphtheria-Pertussis-Tetanus)

- routine vaccination program in developed countries

- only 10% of children in developing countries

(1 million deaths per year)

● Treatment: Administration of antitoxin is the only successful method

Diphtheria toxin

Lysogenic β-pahge

9/9/2010

8

C. ulcerans

● Disease

- Pharyngitis in humans

- Often isolated from bovine mastitis cases

● Able to produce diphtheria toxin

- When lysogenizied by tox-carrying phage

serious illness to unvaccinated individuals

Pathogenic corynebacteria

C. renale group

C. renale (type I) Cattle Bladder, kidney infections

Sheep Penis infections (pizzle rot)

C. pilosum (type II) Cattle Bladder and kidney infections

C. cystitidis (type III) Cattle Severe bladder infection (cystitis)

Rare kidney infection

C. pseudotuberculosis Sheep, Goats Caseous lymphadenitis

Horses, Cattle Ulcerative lymphangitis, ventral abscesses

C. ulcerans Cattle

Human

Mastitis

Pharyngitis

C. diphtheriae Human Diphtheria

9/9/2010

9

Do you know?

Q: What Corynbacterium is involved in ovine caseous lymphadenitis?

1. C. renale2. C. diphtheriae

3. C. pseudotuberculosis

4. C. ulcerans

Corynebacterium

Bacillus

9/9/2010

10

Bacillus

● Gram-positive, large rods

● A large number (>50) of species

and widespread in nature,

but majority are non-pathogenic

● Endospore formation

● Aerobes or facultative anaerobes, catalase positive

● Majority are motile (except for B. anthracis and B. mycoides)

Flagellar staining (by Leifson's Method) of various Bacillus species (CDC)

Identification

Gram stain

Positive Negative

Morphology

Cocci Bacilli (rod)

Spore formation

Positive Negative Bacillus

Catalase

Positive Negative

Motility

Positive Negative Listeria Corynebacterium

9/9/2010

11

B. anthracis: general

● Cause “anthrax” - A severe disease affecting almost all

mammalian species

● Non-motile, non-hemolytic

● Capsules will form in cultures

grown on media containing

bicarbonate

or 50% serum agar in CO2

Medusa headBicarbonate plate

B. anthracis: Anthrax

● Cattle and sheep

- Herbivores are highly susceptible

- Often developing a rapidly

fatal septicemic disease

Sudden death

● Pigs and horses: Moderately susceptible

● Carnivores: Comparatively resistant

● Birds: Almost totally resistant

What about me?

9/9/2010

12

B. anthracis: Anthrax

● Humans

1. Malignant carbuncle: cutaneous anthrax

- organisms is introduced through a wound

develop dark painless lesions

if untreated bacteremia and death

2. Gastroenteritis

- Rare, but from eating infected carcasses

3. Respiratory form (woolsorter’s disease)

- Inhalation of spores by those who

handles hides, wool, and hair

from infected animals

- Preferred route for biowarfare

B. anthracis: Endospore

● Endospore formation

- Important in persistence and spread of anthrax

- Spores are resistant to heat, acid, etc.

:Spores of B. anthracis can

survive for more than 50 years!

Spores in soil

Germination

Septicemia

Toxemia Host death

Spread of spores

9/9/2010

13

B. anthracis: Virulence Factors

● Two large plasmids (pXO1 and pXO2) carry the genes for

anthrax toxins and capsule

pXO1pXO2Capsule Anthrax toxins

B. anthracis: Anthrax toxins

● Protective antigen (PA)

- Transports toxin components to the host cell cytosol

● Edema factor (EF)

- Adenylate cyclase (ATP cAMP)

- cAMP ↑ accumulation of fluids

edema

● Lethal factor (LF)

- LF is a protease

- Proteolytically cleaves MAPKKs,

intracellular signaling proteins

causing cell death

Inhibit the regulation of

immune response

(Adenylate cyclase) (protease)

acidification

9/9/2010

14

B. anthracis: Capsule

● Virulence factors

- “Polypeptide” capsule (poly-D-glutamic acid)

cf. capsule is usually “polysaccarides”

● Resistance to phagocytosis

- Unencapulated bacilli are more susceptible to phagocytosis

Journal of Bacteriology (1998) 180, 52-58

Peptidoglycan

S-layer (Surface-layer):

consists of two surface proteins (EA1 and Sap)

Capsule

FA test of capsule

B. anthracis: Diagnosis

● Performing necropsies on animals dead of anthrax is

“undesirable” since exposing the tissues to the atmosphere

will result in the formation of large numbers of spores

may contaminate the environment for many years

● Aspirate the aqueous/blood discharge for culture

- B. anthracis grows readily on blood agar (non-hemolytic)

- Produces capsules on BHI agar containing 0.5%

sodium bicarbonate incubated in 10-30% CO2

● PCR

- To detect genes encoding the toxins and the capsule

9/9/2010

15

B. anthracis:

Prevention and Treatment

● Vaccniation

- Spore vaccines and avirulent nonencapsulated strain were

used successfully

- Use of the vaccine in livestock has declined markedly in the US

due to the decreased incidence

● Antibiotics

- Penicillin: effective

Tetracycline

- Ciprofloxacin is used in humans

B. cereus

● B. cereus

-Human: Food poisoning often associated with

cooked rice held at improper temperatures

-Cattle: Mastitis occasionally

-Dogs: Food poisoning when fed highly

contaminated canned food

● B. licheniformis

- Associated with abortions in cattle and sheep

9/9/2010

16

Bacillus & Diseases

B. anthracis Cattle, sheep Fatal peracute or acute septicemic anthrax

Humans Skin, pulmonary and intestinal forms of anthrax

Pigs, dogs, cats Subacute anthrax with edematous swelling in

pharyngeal region

Horses Subacute anthrax with localized edematous

swellings of the head, neck, shoulders, throat

B. cereus Cattle Mastitis (rare)

Humans Food poisoning

B. licheniformis Cattle, sheep Sporadic abortion

Do you know?

Q: What is not the virulence factor of B. anthracis?

1. Toxin production

3. Spore formation

2. Bacterial motility

4. Capsule