Mycobacterium

Question 1

Characteristics

Answer 1

Gram positive
Rod-shapedd
Acid-fast bacili
Slow growing (colonies only visible after 3-4 weeks of incubation)
Intracellular bacteria

Question 2

Grows on which media?

Answer 2

Lowenstein Jensen and middlebrook’s media

Question 3

Causes ?

Answer 3

Chronic, granulomatous infection

Question 4

Important species?

Answer 4

M. tuberculosis, M. bovis, M. avium, M. paratuberculosis

Question 5

Causes what important diseases?

Answer 5

Bovine tuberculosis, Johne’s disease, Avian tuberculosis

Question 6

Virulence Factors (1)

Sulfolipids and Phosphatidyl inositol mannoside (PIM)

Answer 6

Prevent respiratoty burst and phagolysosomal fusion, interfere with function of reactive oxygen intermediates following ingestion by macrophage

Question 7

Virulent factor (2)

Surface mycosides

Answer 7

Aid in ensuring bacterial survival within macrophages

Question 8

Virulent Factor (3)

Waxes

Answer 8

Contained within cell wall, have adjuvant activuty, activate macrophages leading to granuloma formation

Question 9

Virulent factors (4)

Alkyl hyroperoxidase reductase

Answer 9

Resistance to superoxides and reactive nitrogen, intermediates found within macrophage phagolysosomes.

Question 10

Virulent factor (5)

Cord factor

Answer 10

Immobilize neutrohils, act as adjuvant, evokes granulomtous responses, cause mitochondrial disruption > disturbances in cellular respiration

Question 11

Virulent factor (6)

Mycobactins and exochelins

Answer 11

Exochelins remove ferric iron from ferritin.
Mycobactin resides in cell membrane, responsible for transfer of iron from iron-exochelin complexes to the bacterium

Question 12

Bovine Tuberculosis

Answer 12

Agent: M. bovis
Host: deer, pigs, people, domestic & wild animals
Case presentation: progressive emaciation and other non-specific signs
- Progressive development of granulomatous lesions or tubercles in multiple organs
- Tuberculous lesion, caseous in nature on organs
- Lung: chronic cough due to bronchopneumonia, dyspnoea in latter stager
- Common involves retropharyngeal, mediastinal

Question 13

BT in deer

Answer 13

Subacute or acute
Often undetectable clinically
Emaciation (severly infected animal), small or large abscess in lymph node of head, thorax and abdomen, enlargement of superficial nodes may be apparent, more liquefactive than those in cattle, resemble a pyogenic iinfection

Question 14

Transmission of BT

Answer 14

Inhalation of droplets
Ingesting raw milk from infected cows
Through movement of undetected infected domestic animals and contact with infected wild animals

Question 15

Diagnosis of BT

Answer 15

Immunidiagnostics
- Tuberculin skin test (CFT or CCT)
- Serology (IFN GAMMA TEST)
Isolation and identification of bacteria
Polymerase chain reaction (pcr) - molecular detection

Question 16

Mechanism of Tuberculin skin test

Answer 16

Cell-mediated hypersensitivity, acquired through infection.
Demonstrated by fever when given subcutaneous
Demonstrated by conjunctivitis when given conjuctival
Demonstrated by local swelling of dermis when given intradermal
CFT Technique - Positive cases in cattle:characterised by thickening and oedema >5mm swelling
False positive following sensitization of animals with other Mycobacterium spp

Question 17

CCT

Answer 17

Injection site: skin of neck region
Positive: increase in skin thickness
M. bovis site is 4 mm or greater than thickness of m, Avium site

Question 18

Immunodiagnostic of BT

Answer 18

Serology (interferon gamma testing/BOVIGAM)
- to detect immune response to bovine TB
- blood in heparinised tube transport at room temperature
- use freshly collected blood sample
- measure level of cytokine called IFN-gamma
- released by WBC infected with TB when stimulates with bovine and avian tuberculins
- Greater responses of IFN-gamma to bovine TB to avian Tb

Question 19

JOHNE’S DISEASE

Answer 19

Agent: Mycobacterium paratuberculosis
Effects: chronic, infectious, granulomatous enteritis of domestic and wild ruminants
Clinical signs: chronic diarrhoea, progressive emaciation

Question 20

Effects on cattle

@3 stages

Answer 20

1. subclinical
   excretory phase , without cs
2. Clinical phase
   chronic diarrhoea, wasting, decrease milk production, oedema and infertility
3. Death as result

Question 21

Effects on sheeps and goats

Answer 21

Emaciation, diarrhoea uncommon
Intermandibular eodema or bottle jaw resulting from Hypoproeinaemia especially in pregnant ewes, lethargy, depression and decrease parasite resistance in terminal stages

Question 22

Avian Tuberculosis

Answer 22

Widest host range
Difficult to eradicate in cattle and pigs as infected animals with M.avium are sensitive to mammalian tuberculin and gives false positive
In birds; usually chronic, but in highly susceptible population is acute
Lesions at: liver and spleen
In cattle; Lesion usually confined in mesenteric lymph node. Majority infections are asymptomatic

Question 23

Sheeps and goats

Answer 23

Resistant to M. avium although mesenteric node infections can be found (rarely)

Question 24

Type of samples and sample preparation

Answer 24

Serology: plasma sample (lithium/heparin tube)
Isolation and identification: milk, tracheobronchial lavage, faeces, biopsy samples, organ samples especially retropharynheal lymph node, mediastinal lymph node, lung with nodules (tubercle), liver, spleen, intestinal lymph node, intestine with lesion
Dont freeze blood sample
Keep organ or fluid samples chill during long transportation (4-15c)
Organ or fluid samples need to be treated with 4% NaOH or 3% HCl or 5% oxalic acid for 15 to 30 minutes to decontaminate the sample. Tubercle bacili can survive the exposure

Question 25

Lab diagnosis

Answer 25

Lowenstein-Jensen with glycerol – for M. tuberculosis, M.avium
Lowenstein-Jensen without glycerol for M. bovis
- malachite green as selective agent, inhibits contaminants

Middlebrook’s 7H10 agar (oleic acid-albumin media)

Incubation time: atleast 8 weeks, generation timesof tubercle bacill - 12 hours upwards at 37c, aerobic cond.

Acid fast or ziehl-neelsen staining technique
1. make smear
2. flood with concentrated carbol fuchsin
3. Heat until steam appears
4. Decolorize the smear with acid-alcohol
5. Counter stain with methhylene blue for one minute
6. Wash the slide with tap water, blot dry and observe under microscope

Question 26

Expected findings

Answer 26

Colony morphology
- mammalian type: dry and crumbly
- avian type- dome shaped colony

Ziehl-Neelsen staining
- mycobacterium stains red, rod shape while others stain blue

Question 27

Control and prevention

Answer 27

Identification and elimination of infected animals; test and slaughter
Post-mortem meat inspection (tubercles in lungs and lymph node, to prevent unsafe meat from entering the food chain, allows vet. service to trace back to the herd of origin of the infected animal for testing and eliminate if required), intensive surveillance (on-farm visits, systematic individual testing of cattle and removal of infected and in-contact animals, movement controls)
Pastereurization of milk for consumption
Vaccination for human