Bacteriology

Question 1

Define bacteria

Answer 1

Single cell microbe with a simple cell structure.

Question 2

Most bacteria are vital for…

Answer 2

human life (commensal) with very few being parasites or pathogens that cause disease

Question 3

Describe bacteria cells

Answer 3

The cell contains no nucleus, allowing the cell to reproduce, very rapidly, by binary fission. Cytokinesis

Require a suitable environment to live and reproduce

Question 4

Some bacteria can form…

Answer 4

endospores that are dormant and very resistant to hostile environmental conditions e.g. heat, cold, radiation, disinfectants.

Question 5

What can endospore producing bacteria cause?

Answer 5

seriously debilitating infectious disease e.g. bacillus anthracis (anthrax)

Question 6

What are the three categories of bacteria?

Answer 6

Obligate aerobes

Obligate anaerobes

Facultative anaerobe

Question 7

Name four classifications of bacteria

Answer 7

Size

Shape

Arrangement

Structure

Question 8

Why might a vet choose to conduct a bacteria culture?

Answer 8

Joint pain/chronic lameness

Dermatitis

Otitis

Non healing wound

Chronic or aggressive gastroenteritis

Surgical wound breakdown

Pyoderma

Cystitis

Question 9

Give examples of fluid samples

Answer 9

Blood

Urine

CSF

Synovial fluid

Exudates

Semen

Sputum

Question 10

Give examples of swab samples

Answer 10

Eye

Ear

Skin

Wounds

Abscesses

Penis

Vagina

Question 11

Give examples of tissue samples

Answer 11

Liver

Lung

Kidney

Spleen

Cardiac

Lymph node

Skin

Question 12

Describe patient prep for bacteria samples

Answer 12

Obtain sample prior to administration of antimicrobial medication.

Dermatology samples should not be clipped or aseptically prepared prior to sampling

Question 13

Describe facultative anaerobes

Answer 13

Grow aerobically when oxygen is present but can also function in the absence of oxygen e.g.

Question 14

Describe preservation and storage of bacterial samples

Answer 14

Clean, sterile and leak proof container.

Correct use of transport media. This will depend upon the sample obtained.

Aimies media with charcoal added is commonly used for aerobic bacteria. – useful for fastitious bacteria

Aimies without charcoal

Virus transport medium

Anaerobic transport medium

Can be refrigerated in not testing immediately

Do not freeze samples

Question 15

Describe solid media

Answer 15

Agar alone provides no nutrition for bacterial growth.

Specific nutrients and growth factors are impregnated in to the agar base. These are designed to encourage growth of particular organisms.

Isolation of individual species can only be achieved on solid media.

Un-innoculated Media plates should be stored with in the refrigerator and NOT frozen

Question 16

Name 4 types of media types

Answer 16

Simple media.

Enrichment media.

Selective / differential media.

Transport media.

Question 17

Describe simple (basal) media

Answer 17

Provides basic nutrition for growth of nutritionally undemanding species e.g. E Coli

Nutrient broth, nutrient agar

Question 18

Describe enrichment media

Answer 18

Specific nutrients are added to basal media for growth of fastidious bacteria (complex nutritional requirements) e.g.

Blood agar - used to detect haemolytic bacteria strains.

chocolate agar – contains haemoglobin, for detection of respiratory bacteria

Question 19

Describe selective/differential media

Answer 19

Designed to inhibit growth of certain bacteria while not affecting growth of others.

Deoxycholatecitrate used for growth of salmonella spp.

McConkey’s bile lactose agar contains bile salts which encourage the growth of enteric species e.g. E coli

Sabouraud’s media used for the growth of ringworm.

Question 20

Describe transport media

Answer 20

Used to transport or store samples e.g. Swabs

Not used to support or encourage growth but to ensure survival of organisms until analysis can be carried out.

Question 21

Describe how to grow bacteria

Answer 21

Once inoculated replace the lid and put in an incubator with agar side upper most, to avoid condensation droplets spoiling the bacteria growth.

Incubate at 37 ̊C for 18-24hrs.

If no growth after 18-24hrs incubate for further 24hrs.

Question 22

What results will be seen when growing bacteria?

Answer 22

Colonies of bacteria will appear as round raised lumps along the streak lines.

Colonies of different species may show different characteristics helpful for identification e.g. colour.

Question 23

What techniques may be used to help identify bacterial colonies?

Answer 23

Microscopic evaluation

Grams staining

Zeihl Neelson – acid fast

Question 24

How can you identify bacteria?

Answer 24

Staining with a suitable stain e.g. Gram stain allows cell morphology to be observed.

Bacteria can be classified by the thickness or composition of the cell wall structure.

Gram’s positive

Gram’s negative

Acid fast

Question 25

Describe Simple stains (methylene blue)

Answer 25

Colours cells allowing shape, size and arrangement to be observed.

Question 26

Describe Differential stains (Gram’s stain, Ziehl-Neelsen)

Answer 26

This is a combination of 2 dyes, a primary and a counter stain, that allows differentiation of cell types.

Question 27

Describe Structural stains

Answer 27

This stains certain parts of the cell e.g. Flagella, microspore and capsule also aiding in identification of the bacterium.

Question 28

Describe gram stains

Answer 28

This is the most frequently used stain in microbiology that provides the first step in identifying bacteria.

Bacteria are categorised by the colour that they stain

Gram’s positive – blue / purple

Gram’s negative – red / pink

Question 29

Describe gram positive bacteria

Answer 29

Gram positive have a thick, porous petidoglycan layer (outer layer of the cell) which absorbs the purple crystal violet stain.

e.g. a number of staphylococcal and streptococcal spp.

Absorbent layer also will absorb toxic substances so much easier to destroy.

Question 30

Describe gram negative bacteria

Answer 30

Gram’s negative bacteria have a thin peptidoglycan layer which is protected by a further outer membrane.

Small holes ‘porins’ are contained within the outer membrane

The thin peptidoglycan layer does not retain the crystal violet stain but absorbs the pink safranin stain.

e.g. pseudomonus aeruginosa

Question 31

Describe acid fast bacteria

Answer 31

Rarely diagnosed in vet practice.

Have a waxy (mycolic acid) outer layer.

Highly resistant to staining and treatment.

Gram positive bacteria.

Ziehl neelsen stain required ( red on blue background.)

All other organisms stain blue.

e.g. mycoplasma, mycobacterium tuberculosis, m leprae, nocardia

Can be zoonotic

Question 32

What is sensitivity training?

Answer 32

Indicates which antimicrobial is most appropriate to eliminate infection

Allows the patient to be treated quickly and effectively

Reduces the risks associated with antimicrobial resistances

Question 33

Describe disc agar infusion

Answer 33

Most commonly used method to determine the susceptibility of a microorganism to a specific antimicrobial agent (efficiency, cost and convenience)

Antimicrobial agent diffuses into the solid medium from the filter paper resulting in the inhibition of reproduction of the microorganism on its surface.

A zone of inhibition forms around the disc, beyond this zone an unaffected area of normal microbial growth is present

Question 34

What is the method for disc agar infusion?

Answer 34

A bacterial colony from loop or swab is spread over an agar plate

A manufactured disc containing a range of antibiotics is applied to the plate surface.

Incubate for 18-24hrs at 37 ̊C

The resistance zone cut off is specific to each antibiotic.

Accurate measurement of the zone diameter and comparison to an interpretation chart is necessary to properly interpret this test.

Depending on these results the isolate will be classed as being susceptible, intermediate or resistant to that specific antimicrobial agent

Question 35

How should you dispose of bacteriological sample equipment?

Answer 35

Place ALL equipment in suitable disinfectant, following use, during the procedure.

Seal all equipment in autoclave bag and autoclave prior to disposal or re-use.

Once autoclaved, equipment to be disposed of should be placed in infectious waste (orange bag).