Anaerobic Bacteria

Question 1

What are the toxic by-products produced by biochemical reactions when anaerobic bacteria are exposed to oxygen?

Answer 1

(a) superoxide radical (O₂⁻)
(b) hydrogen peroxide (H₂O₂)

Further notes:
These substances are also produced in aerobic organisms, but anaerobic bacteria do not have mechanisms to neutralize their toxic effects.

Question 2

List some enzymes that are present in aerobic bacteria but which are absent in anaerobic bacteria.

Answer 2

(1) cytochrome system
(2) catalase
(3) peroxidase
(4) superoxide dismutase

Question 3

Where are strict anaerobes generally found?

Answer 3

◾ Gastrointestinal tracts of animals
◾ Deep soil layers
◾ Sediments: bottom layers of lakes, rivers, oceans
◾ Sewage

Question 4

Where are strict anaerobes found in humans?

Answer 4

◾ Mouth
◾ Deeper areas of the skin
◾ Genitourinary tract
◾ Large intestine

Question 5

Key features of Clostridium

Answer 5

◾ Gram-positive bacilli
◾ Most species are flagellated and motile
◾ Anaerobic
◾ Form spores

Question 6

Comment on spores of Clostridium spp.

Answer 6

◾ They are centrally or sub-terminally or terminally placed.
◾ They are resistant to adverse physical conditions.
◾ They enable prolonged survival of the organism in the environment.
◾ [Image]

Question 7

Name five species of Clostridium associated with disease in humans.

Answer 7

◾ C. perfringens [aka. C. welchii]
◾ C. tetani
◾ C. botulinum
◾ C. difficile
◾ C. histolyticum

Question 8

Discuss the main pathogenicity properties of Clostridium spp.

Answer 8

(1) Spores: increase the chances of survival and enhance transmission.
(2) Ability to retain viability and multiply in tissues without oxygen or with reduced blood supply.
(3) Ability to produce exotoxins which are responsible for major clinical manifestations of associated diseases.
(4) Some species produce aggressins and other enzymes also responsible for pathogenicity.

Question 9

How does Clostridium perfringens spread in the environment?

Answer 9

It spreads via feces from humans and other animals.

Question 10

Morphology of spores of C. perfringens

Answer 10

The spores are centrally placed on the rod.

Question 11

Is Clostridium perfringens motile or non-motile?

Answer 11

It is non-motile.

Question 12

What are the significant biochemical properties of Clostridium perfringens?

Answer 12

◾ saccharolytic activity as the predominant biochemical activity
◾ mildly proteolytic
◾ nitrate reduction test positive

Further notes:
🔎 Saccharolytic activity refers to the ability of an organism to break down sugars (saccharides) into simpler compounds, such as acids or alcohols, through fermentation or other metabolic processes.
🔎 The nitrate reduction test is used to determine an organism’s ability to reduce nitrate (NO₃^-) to nitrite (NO₂^-) or other nitrogenous compounds through anaerobic respiration. For C. perfringens, a positive nitrate reduction test means that it can utilize nitrate as an electron acceptor in the absence of oxygen, which is important for its survival in anaerobic environments.

Question 13

What are the serotypes of C. perfringens?

Answer 13

C. perfringens is classified into five serotypes based on the production of major toxins. These serotypes are designated A, B, C, D and E.

Question 14

Briefly discuss the exotoxins of C. perfringens.

Answer 14

(1) Alpha (α) toxin
◾ Released by all serotypes.
◾ Major toxin of C. perfringens serotype A.
◾ Relatively heat stable.
◾ Acts as an enzyme phospholipase or lecithinase, splitting lecithin, a constituent of cell membranes, leading to cell lysis and tissue damage.

(2) Beta (β) toxin
◾ Serotypes B and C.
◾ Causes necrotizing enteritis by damaging intestinal mucosa.

(3) Epsilon (ε) toxin
◾ Serotypes B and D.
◾ Increases vascular permeability, leading to edema and damage to various organs, including the brain.

(4) Iota (ι) toxin
◾ Serotype E.
◾ Has ADP-ribosylating activity that disrupts the cytoskeleton of host cells, leading to cell death.

(5) Enterotoxin
◾ Released by specific strains.
◾ Causes food poisoning by disrupting the intestinal epithelium, leading to diarrhea and abdominal cramps.

Further notes:
Lecithin is a phospholipid. It is a complex mixture of phospholipids, primarily phosphatidylcholine, along with other components such as phosphatidylethanolamine and phosphatidylinositol. Lecithin is commonly found in cell membranes and plays a crucial role in maintaining their structure and function.

Question 15

List clinical conditions associated with C. perfringens.

Answer 15

(a) gas gangrene
(b) wound and other soft tissue infections
(c) food related gastroenteritis or food poisoning
(d) infections involving pelvic tissues in adult females
(e) intra-abdominal infections

Question 16

Discuss clinical features of gas gangrene [aka. clostridial myonecrosis].

Answer 16

🩺 Rapidly progressing infection that causes severe pain and swelling.
🩺 Skin discoloration, often with a bronze or purple hue.
🩺 Blisters or bullae filled with gas or fluid.
🩺 Foul smelling discharge from the wound.
🩺 Rapid heart rate and low blood pressure.
🩺 Crepitus [crackling sensation in affected tissues]

Question 17

predisposing conditions to gas gangrene

Answer 17

(1) Traumatic injury with skeletal muscle involvement [responsible for most cases]
(2) Injury involving the abdomen together with the large intestine. Infection may result from leakage of intestinal contents from the lumen to adjacent tissues.

Question 18

(a) Which serotype of C. perfringens is most commonly associated with gas gangrene?
(b) List other species of Clostridium that are associated with gas gangrene (3).

Answer 18

(a) Serotype A
(b) C. histolyticum, C. novyi, C. septicum

Question 19

Discuss facilitating factors in the development of gas gangrene.

Answer 19

(1) Injury
✔ Open wounds and damaged skeletal muscle are exposed to contamination with dirty material containing bacterial spores.
✔ Impairment of normal blood flow leads to reduction in oxygen tension in the injured part. This creates a suitable condition for viability and multiplication of C. perfringens.

(2) Spores germinate into bacilli which multiply and release exotoxins. Released toxins cause tissue destruction and gas production
✔ Alpha toxin is the major cause of the tissue damage and severity of clinical manifestations.

Question 20

Explain how certain strains of C. perfringens may cause food borne-gastroenteritis.

Answer 20

✔ The strains form spores which are able to survive boiling for several hours.
✔ When conditions are favorable, the spores germinate into bacilli in the food.
✔ Bacilli are ingested as the food is consumed.
✔ Bacilli produce enterotoxin in the intestinal tract

Question 21

manifestations of food-borne gastroenteritis caused by C. perfringens

Answer 21

✔ mainly mild abdominal cramps and diarrhoea
✔ experienced 8 to 12 hours after ingestion
✔ fever and vomiting are not common
✔ illness subsides within 24 to 48 hours

Question 22

What is clostridial endometritis?

Answer 22

This refers to infection of the uterine lining by C. perfringens, often associated with postpartum or post-abortion complications.

Question 23

Briefly discuss culturing of C. perfringens on BA medium.

Answer 23

🔬 Colonies of C. perfringens are relatively large, transulcent and have a smooth surface.
🔬 C. perfringens also exhibits a characteristic double-zone of haemolysis on blood agar:
◾ Inner zone: beta-haemolysis resulting in a clear zone around the colony.
◾ Outer zone: incomplete haemolysis where red blood cells are partially lysed, resulting in a larger hazy zone around the colony.

[Image 1] [Image 2]

Question 24

Besides blood agar, name one other medium that is particularly useful for isolating and maintaining cultures of Clostridium perfringens.

Answer 24

Robertson’s Cooked Meat medium

Question 25

What test can be used to demonstrate alpha toxin production in C. perfringens?

Answer 25

Nagler reaction/Nagler test

Question 26

(a) Name two effective agents that can be used against C. perfringens.
(b) Briefly explain how the named agents work.

Answer 26

penicillin: interferes with the synthesis of bacterial cell wall

metronidazole: disrupts DNA

Question 27

Explain how gas gangrene can be prevented.

Answer 27

(1) Proper care of wounds by cleaning and removal of foreign materials and dead tissue.
(2) Proper cleansing of skin before invasive procedures and use of sterilized instruments and materials.
(3) Antibiotic prophylaxis in management of fresh or trauma associated wounds.

Question 28

one key differnce between C. perfringens and C. tetani

Answer 28

C. tetani is flagellated and hence motile; C. perfringens is immotile

Question 29

Where is C. tetani found?

Answer 29

It is naturally found as normal flora in the large intestinal tract in humans and other animals including horses, cattle and donkeys.

Question 30

What is one significant biochemical property of C. tetani?

Answer 30

It is predominantly proteolytic.

Question 30

microscopic morphology of spores of C. tetani

Answer 30

The spores are observable microscopically as a distended part at one end of the bacillus, described as terminal spores with a “drumstick” appearance. [Image]

Question 31

What are the significant physical properties of Clostridium tetani spores in terms of resistance to adverse physical conditions?

Answer 31

✔ Some strains form spores that are readily inactivated by commonly used physical methods of treatment of items.
✔ Others form spores that can survive commonly used methods of disinfection, adverse environmental conditions for years, boiling in water for less than 3 hours, and dry heat at 150 degrees Celsius for 1 hour.

Question 32

Briefly discuss the two main extracellular products of C. tetani.

Answer 32

(1) Tetanolysin
✔ causes lysis of red blood cells
✔ effects observable in BA cultures

(2) Tetanospasmin
✔ the exotoxin of C. tetani
✔ neurotoxin that blocks the release of inhibitory neurotransmitters, leading to unopposed muscle contractions and severe spasms.
✔ It can be inactivated to produce tetanus toxoid, which is used in vaccines to provide immunity against tetanus.

Question 33

Explain how C. tetani infection may be acquired.

Answer 33

Infection is acquired through open wounds which get contaminated through contact with foreign material containing spores e.g. soil, dirty clothes, dust, and instruments items used when working on wounds.
Spores in the wound germinate into bacilli, facilitated by reduced oxygen tension and devitalized tissues. Bacilli multiply in the wound but do not spread beyond the wound.

Question 34

Explain how neonatal tetanus may be acquired.

Answer 34

Neonatal tetanus occurs when C. tetani infects the umbilical stump of a newborn infant. This can happen due to non-sterile delivery practices or poor postnatal care.

Question 34

Describe the appearance of C. tetani colonies on blood agar.

Answer 34

✔ The colonies of C. tetani are not distinct.
✔ This is because the organism spreads and grows as a layer on the surface of the BA.

Question 34

Briefly discuss the overall effects and manifestations of tetanospasmin in the CNS.

Answer 34

◾ Tetanospasmin blocks the release of inhibitory neurotransmitters, such as gamma-aminobutyric acid (GABA) and glycine.
◾ The inhibition of inhibitory neurotransmitters leads to unopposed muscle contractions, resulting in severe muscle stiffness and spasms.
◾ One of the earliest and most characteristic symptoms is the stiffness of the jaw muscles, making it difficult to open the mouth. This is known as lockjaw/trismus.
◾ Spasms of the respiratory muscles may lead to breathing difficulties and potentially life-threatening complications.

Question 35

Outline management methods of tetanus.

Answer 35

✔ Administration of tetanus antitoxin
✔ Antimicrobial agents: C. tetani is susceptible to penicillin and its derivatives
✔ General supportive care
✔ Administration of muscle relaxants
✔ Surgical management of wounds where required

Question 36

When is immunization with tetanus toxoid performed/recommended?

Answer 36

◾ As part of management of traumatic wounds
◾ During antenatal care for expectant mothers
◾ In patients undergoing recovery from tetanus
◾ As part of routine immunization of children

Question 37

Significant morphological features of C. botulinum bacilli and spres

Answer 37

◾ Bacilli are flagellated and hence motile
◾ Spores are placed centrally or sub-terminally in the bacillus

Question 38

How may serotypes of C. botulinum are there?

Answer 38

There are 7 serotypes designated A to G according to various antigens also designated A to G.

Question 38

Discuss C. botulinum exotoxin.

Answer 38

◾ The exotoxin produced by C. botulinum is referred to as botulinum toxin.
◾ It is a potent neurotoxin that affects the nervous system. It blocks the release of acetylcholine at the neuromuscular junction, leading to flaccid paralysis. This can result in muscle weakness, respiratory failure, and potentially death if not treated promptly.
◾ The toxin is heat labile, meaning it can be destroyed by heat [80°C for 30 - 40 min, thorough cooking of food].

Question 38

significant physical properties of C. botulinum spores

Answer 38

◾ withstand moist heat at 100°C for several hours
◾ destroyed at 120°C for 5 minutes

Question 38

Name two main types of botulism.

Answer 38

Food-borne botulism, wound botulism

Question 39

What specimens are examined in the laboratory investigation of botulism?

Answer 39

stool and other GI tract contents;
suspected food

Question 40

management of botulism

Answer 40

◾ administration of antitoxin specific to the C. botulinum serotype associated wth the illness
◾ wound management by surgical methods
◾ administration of antimicrobial agents e.g. penicillin and metronidazole
◾ proper handling and processing of foods
◾ proper processing of food for canning
◾ sufficient heating of food before consumption
◾ use of food preservatives

Question 41

clinical relevance of C. difficile

Answer 41

◾ It is more resistant to commonly used antibiotics than other bacterial flora in the large intestines.
◾ Broad spectrum antibiotics administered for treatment of infection in other parts of the GI tract inhibit the susceptible large intestinal bacterial flora.
◾ C. difficile is not inhibited and is able to multiply.
◾ It produces two toxins, Toxin A (enterotoxin) and Toxin B (cytotoxin) which cause diarrhoea and inflammation.
◾ Inflammation may progress to manifest as pseudomembranous colitis.

Question 42

Name three clinically relevant genera of gram-negative non-spore forming anaerobic bacilli.

Answer 42

✔ Bacteroides
✔ Porphyromonas
✔ Prevotella

Question 42

key features of the genus Bacteroides

Answer 42

◾ Non-motile
◾ Gram-negative capsulated bacilli
◾ Obligate anaerobe; though they have the enzymes catalase and superoxide dismutase that allow them to survive transient exposure to oxygen

Question 42

clinical significance of B. fragilis

Answer 42

B. fragilis is commonly associated with intra-abdominal infections, such as abscesses, appendicitis and peritonitis.

Question 43

Fusobacterium are a group of gram-negative, anaerobic, non-spore-forming bacteria that constitute the normal flora of the mouth. Name the two most well known members of this genus.

Answer 43

F. nucleatum
F. nerophorum