3.4 Microbiology

Name of rod shaped bacteria

bacillus

Name of spherical bacteria

coccus

Name of corkscrew shaped bacteria

spirillum

What are the functions of the bacterial cell wall

• Providing overall strength to the cell

• Maintains cell shape

• Protects the cell from osmotic lysis

What will gram-positive bacteria stain under the Gram stain

purple

What will Gram-negative bacteria stain under the Gram stain

pink

What do both bacteria contain in their cell walls

peptidoglycan

Describe the structure of peptidoglycan

a polysaccharide made of long chains of sugars

The chains are cross linked to one another by short peptides

Why do Gram positive bacteria stain purple?

have a thicker outer wall made of peptidoglycan with no outer lipopolysaccharide layer.

This allows the crystal violet/iodine complex to be retained within the cell - staining the cells purple.

Why do Gram negative bacteria stain pink

Gram negative bacteria have a thinner peptidoglycan cell wall and an outer lipopolysaccharide membrane.

On treatment with alcohol, the Gram negative cell walls lose the outer lipopolysaccharide membrane, and the thin peptidoglycan wall allows the purple stain complex to be washed away.

Gram negative cells are not stained by the Gram stain (they remain colourless), but stain pink after counterstaining with safranin.

State the 4 steps of Gram staining

1. Applying a primary stain (crystal violet)

2. Adding a mordant (Gram's iodine)

3. Rapid decolourisation with ethanol

4. Counterstaining with safranin

State the conditions required for microorganisms to grow

• Nutrients

• Temperature

• pH

• Oxygen

Function of carbon as a nutrient

a source of energy

Function of nitrogen as a nutrient

Amino acid synthesis

Function of temperature as a condition

bacterial metabolism is regulated by enymes

Obligate aerobes

microorganisms that only grow in the presence of oxygen

faculative anaerobes

microorganisms that can grow with or without oxygen

obligate anaerobes

Microorganisms that only grow in the absence of oxygen

What are growth factors, vitamins and minerals used for by microorganisms

May act as cofactors to help enzymes function; purines and pyrimidines used to make nucleotides

What are bacteria cultured on or in

on agar jelly or in nutrient broth

What is phosphorous used for by mircoorganisms

Used to make phospholipids / ATP/ DNA/ RNA

What is sulphur used for by microorganisms

As a component of some amino acids and coenzymes

Define aseptic technique

Laboratory practice that maintains sterility in apparatus and prevents contamination of the equipment and the environment.

Describe how to prevent the contamination of pure cultures and apparatus by bacteria from the environment

- sterilise the work surface using a disinfectant-carry out process of inoculation

-Use an autoclave

- After use, disposable materials can be sealed inside autoclavable plastic bags, autoclaved and then placed in a dustbin

Describe how to prevent the contamination to the environment by the bacteria being used in experiments

• sterilise the work surface using a disinfectant-carry out process of inoculation

-Use an autoclave

• After use

Describe the process of inoculation

-flame the mouth of the bottle for 2 or 3 seconds.

• pass the inoculating loop through a flame until red hot.

-lift the lid of the petri dish just enough to allow entry of inoculating loop.

• secure the petri dish with adhesive tape

• incubate at 25 degrees

Descrie how to achieve a dilution of 10-4

1) Transfer 0.1 cm3 of the original culture to 9.9 cm3 sterilised water using a sterile syringe. Mix contents well.

2) Transfer 0.1 cm3 of this dilution (10-2) to a different tube containing 9.9 cm3 sterile water using a (different) sterile syringe. Mix well. This makes 10-4 dilution.

3) Aseptic technique should be used e.g flame the mouth of the tubes by passing through a blue bunsen burner flame quickly (before and after removing a sample).

How are disposable materials such as petri dishes autoclaved

by being sealed in autoclavable plastic bags

State the assumption that is being made in this method determining bacterial cell number

each colony derived from one bacterial cell

Why is the lid of the petri dish lifted only slightly

to avoid contamination of microbes from the air

why do you not seal the petri dish completely

this could create anaerobic conditions and encourage the growth of possible pathogenic microorganisms

why do we not incubate cultures at 37 degrees

as this is an ideal temperature for the growth for many pathogenic species

Describe an autoclave

a sealed container in which glass and metal equipment is heated at 121 degrees in steam under pressure for 15 minutes

Define total count

include both living and dead cells

Define viable count

living cells only

Define a pathogen

an organism that causes disease in its host

Define a colony

A cluster of cells which arises from a single bacterium