Lab 2 - Bacillus, Clostridium

Question 1

Bacillus habitat

Answer 1

• diseased animals (vegetative)

- environment (spore)

Question 2

Bacillus morphology/resistance

Answer 2

• 4-5 µm rod, central spore
• capsule
• no flagella
• R: vegetative bacterium: several days in a carcass
• 56°C: 15 min
• spore
• boiling: 5-10 min, formalin: 12-24 h

Question 3

Bacillus staining

Answer 3

• Gram-positive

- toluidine-blue

Question 4

Bacillus culture

Answer 4

-simple (nutrient broth or agar, blood agar, air,
37°C)
-R-S colonies
-capsule: poly-D-glutamic acid
-plasmid encoded spore – for spore formation: at least 15°C, water, oxygen, cations are needed
-37°C: finished during 16 hours
-18°C: it will start after 50 hours

Question 5

Bacillus biochemistry/antigens

Answer 5

• active metabolism, catalase +, oxidase -
• Ag: capsule - saprophytes, polysaccharide hapten: heat stabile
• Ascoli test, B. cereus – cross reaction!
• oedema factor
• lethal factor
• protective Ag
• Ascoli thermoprecipitation test: for detection of B.anthracis heat stable cell wall Ag.

Question 6

Bacillus pathogenicity

Answer 6

• mammals (mainly herbivorous animals)
• Cattle, sheep: fatal peracut or acut septicaemic anthrax
• Pigs: subacute anthrax with oedematous swelling in pharyngeal region, an intestinal form with higher mortality is less common
• Horses: subacute anthrax with localised oedema, septicaemia with colic and enteritis sometimes occur
• Humans: skin, pulmonary and intestinal forms of anthrax
• Carnivores are comparatively resistant, birds are totally resistant

Question 7

Bacillus species

Answer 7

-B.cereus
*Cattle: mastitis (rare)
*Humans: food poisoning, eye infections
-B.licheniformis: cattle, sheep: sporadic abortion
-Paenibacillus larvae: honey bees: American foulbrood
Saprophyte Bacilli:
-B.subtilis
-B.megaterium
-B.cereus
-B.licheniformis: bacitracin
-B.thuringiensis: insect pathogen, pest-control (moth)
-Paenibacillus (B.)polymyxa: polymyxins
-Geobacillus (B.)stearothermophilus: heat resistant test organism

Question 8

B.anthracis

1. Flagella
2. Capsule prod. in air
3. Haemolysis
4. Pathogenecity

Answer 8

1. Flagella: no
2. Capsule prod. in air: no
3. Haemolysis: no
4. Pathogenecity: yes

Question 9

Clostridium habitat

Answer 9

• soil
• mud
• water
• gut

Question 10

Clostridium morphology/resistance

Answer 10

• 7-10 µm rod
• Flagella (exc. C.perfr.)
• Spore (terminal, subterminal, central)
• no capsule (exc. C.perfr.)
• R: Vegetative: (like Gr+)
• Spore: (in dry specimens: for years)

Question 11

Clostridium staining

Answer 11

Gram-positive

Question 12

Clostridium culture

Answer 12

• anaerobic
• oxygen tolerance different:
  (c. tetani, c. novyi b: strict)
  (c. histolyticum, c. botulinum: can tolerate o2)
• nutrient agar
• blood agar
• optimal temperature: 37oc
• 40-45oc - c.perfringens
• 15-22oc - c.putrefaciens

Question 13

Clostridium biochemistry/antigens

Answer 13

• catalase –, oxidase -
• OF: fermentative
• active metabolism, carbohydrates are fermented
  (exc. C. tetani, C. histolyticum)
• proteolytic: C. tetani, C. histolyticum, C. botulinum
• exotoxins (toxin, toxoid or anatoxin, antitoxin)
• Ag: complex, close relationship

Question 14

Clostridium pathogenicity

Answer 14

• gas gangrenic diseases:
  1. malignant oedema (cattle, pigs, sheep): C.septicum, C.novyi, C.haemolyticum, C.histolyticum, C.sordellii
  2. Blackleg (cattle, sheep): C. chauvoei
  3. human gas gangrene: C.perfringens A, C.novyi A
• enterotoxaemic diseases:
  1. necrotic enteritis of chicken: C.perfringens A/C
  2. lamb dysentery: C.perfringens B
  3. necrotic enteritis of pigs, struck of sheep: C.perfringens C
  4. pulpy kidney disease of sheep: C.perfringens D
  5. ulcerative enteritis of chicken: C.colinum
• intoxications
  1. tetanus: C.tetani
  2. botulism: C.botulinum
• C.septicum

Question 15

C.septicum

Answer 15

• culture: nutrient agar
• toxins: alpha (haemolytic, necrotizing, lethal), beta (DN-ase), gamma (hyaluronidase), delta (haemolysin)
• cattle, pig, sheep: malignant oedema
• sheep and calves: braxy (abomasitis)

Question 16

The vegetative bacteria of the B. anthracis are widely distributed in the normal intestinal flora and on the mucosal surfaces of domestic animals, especially ruminants.

Answer 16

False

Question 17

Bacillus anthracis is an obligate aerobic bacterium, but for the capsule forming CO2 is also required.

Answer 17

True

Question 18

There are cross reactions with Bacillus cereus by using of Ascoli-test to detect Bacillus anthracis.

Answer 18

True

Question 19

Horses are resistant to Bacillus anthracis

Answer 19

False

Question 20

B.anthracis and B.cereus can be differentiated by their haemolysis

Answer 20

True ?

Question 21

The pulpy kidney disease (sheep enterotoxaemia) and the necrotic enteritis of chickens are caused by the same bacterium species

Answer 21

True

Question 22

Bacillus thüringiensis is the causative agent of American foulbrood of hoenybees.

Answer 22

False (Paenibacillus (Bacillus) larvae)

Question 23

One causative agent of malignant edema is C.histiolyticum

Answer 23

True

Question 24

C.perfringens belongs to the group of the neurotoxic Clostridia

Answer 24

False (enterotoxaemic disease)

Question 25

The toxin of the C.botulinum is quite resistant to heat treatment.

Answer 25

True

Question 26

There are no cross reactions by using Ascoli-test to detect B.anthracis

Answer 26

False (w. B.cereus)

Question 27

The pulpy kidney disease and the necrotic enteritis of piglets are caused by the same bacterium species.

Answer 27

True

Question 28

Paenobacillus polymixa is the causative agent of American foulbrood of honeybees.

Answer 28

False (Paenibacillus (Bacillus) larvae)

Question 29

One causative agent of blackleg is C.haemolyticum.

Answer 29

False (C. chauvoei)

Question 30

C.perfringens belongs to the group of the histotoxic Clostridia.

Answer 30

True

Question 31

The toxin of the C.botulinum causes rigid paralysis especially to water birds, like ducks.

Answer 31

True

Question 32

The spores of B.anthracis are widely distributed in the soil and natural water of endemic areas.

Answer 32

?

Question 33

A B.anthracis is an obligate anaerobic bacterium, O2 is needed only for the spore prod.

Answer 33

False (aerobic)

Question 34

By using toluidine blue staining in a smear of B.anthracis, we can see pink rods in chains surrounded by blue capsular material.

Answer 34

True ?

Question 35

C.difficile belongs to the group of the histotoxic Clostridia.

Answer 35

False

Question 36

The necrotic enteritis in chicken and the calf enterotoxaemia are caused by the same bacterium species

Answer 36

False (not calf)

Question 37

The causative agent of the lamb dysentery is a flagellated bacterium.

Answer 37

True

Question 38

The causative agent of malignant edema is C.difficile.

Answer 38

False

Question 39

The toxin of the C.tetani causes flaccid paralysis.

Answer 39

False (C. botulinum)

Question 40

C.piliforme cannot be cultured on artificial media.

Answer 40

True