Disease

1Getting sick

2Feeling sick

3Fighting back

Killing microbes with chemicals

Attacking bacteria experiment

H Disease

Antibiotic resistant bacteria

Example: MRSA – Methicillin Resistant Staphylococcus Aureus

A disease is any condition that impairs (makes worse) the normal functioning of an organism. We are concerned with those diseases that are caused by micro-organisms (ie pathogens).

When microbes invade and reproduce in the organism we say it has an infection.

Disease

1 Getting sick...

While a few microbes (such as the athlete’s foot fungus) live on the body, the majority must find a way inside before they can reproduce. The majority come in through either the nose or the mouth:

A few microbes get into our blood via insect bites, or through cuts (or dirty needles).

Each pathogen infects specific tissues. Cold and flu viruses infect the cells of the respiratory system.

in airborne dust or droplets

in our food or drink

off our hands.

2 Feeling sick...

Human pathogens grow best at normal blood heat, 37 °C, so the body tries to kill the invader by raising its temperature. You might feel shivery as your temperature starts to rise.

Since your body needs to devote energy to fighting the invader, it diverts energy from other functions. You feel weak and have no appetite.

You may produce more mucus from your respiratory cells, then sneeze or cough to get rid of that mucus. This helps rid your body of the pathogen, but it can also spread the infection to others. Similarly, vomiting or diarrhoea are other ways in which your body rids itself of invaders.

In addition, the waste products of some bacteria include toxins that have damaging effects on the human body.

3 Fighting back...

Your body’s ‘soldiers’ are the white blood cells that attack and destroy any foreign particle, and make antibodies designed to lock onto and neutralise specific pathogens.

When the pathogens are destroyed your fever drops but it takes a while to get rid of the excess mucus and recover strength and appetite.

The antibodies remain on patrol in your blood in case that particular invader should return. We say you are now immune to that particular illness.

Disease

Disease is something that impairs the normal functioning of an organism. Many diseases are caused by microbes.

Microbes usually get into our bodies through the nose or mouth – in particles carried in the air, on food or drink, or via our hands. Some microbes get in through breaks in the skin.

The body tries to destroy the microbes with fever (high temperature), white blood cells and antibodies. Coughing, sneezing, vomiting and diarrhoea eject the microbes from the body.

Sometimes bacterial toxins cause more damage.

After the illness, antibodies remain to prevent future infection by that particular pathogen.6H 1 There’s a cold going round

Killing microbes with chemicals

Chlorine gas and certain chlorine compounds (such as in bleach) kill microbes in water or on surfaces. They are called disinfectants. Hydrogen peroxide (in ‘oxy’ laundry soakers) is another disinfectant.

Most swimming pools and public water supplies in New Zealand are treated with chlorine.

To kill microbes on skin we use milder agents called antiseptics. Dettol and Savlon are two well-known brands of antiseptic solution.

Alcohol also kills microbes on skin.

This man is having his ear sprayed with alcohol

before it is pierced.

Antibiotics are compounds that will kill bacteria inside the body, without harming the patient’s own cells. The first antibiotics were made by fungi, to prevent the growth of bacteria that would otherwise compete with them for nutrients.This agar plate has four circular discs soaked in different antibiotics (C, T, G and P), while the W square was soaked in water. The clear zone around the discs show how well each antibiotic inhibits the growth of this strain of bacteria. Only C and G are effective.

Antibiotics can only be used to treat bacterial infections, but many illnesses are caused by viruses, not bacteria. There are a few antiviral compounds that interfere with the replication of specific viruses inside the human body. Tamiflu® is an antiviral compound that is effective against some strains of the influenza virus. Zovirax® cream contains an antiviral substance that stops replication of the cold sore virus.

Killing microbes with chemicals

Chlorine gas, some chlorine compounds and hydrogen peroxide are disinfectants. They kill microbes on surfaces, in drinking water and in swimming pool water.

Dettol and Savlon are antiseptic solutions which can be safely used to kill microbes on skin.

Antibiotics such as penicillin kill bacteria (but not viruses) inside the body without harming body cells.

A few antiviral compounds, such as Tamiflu, slow the replication of specific viruses inside the body.

Attacking bacteria

In this experiment we want to investigate the effectiveness of different types of antiseptic solution and antibiotics on a particular kind of skin bacteria.

If the antibacterial solutions are effective then bacteria will not be able to grow close to the paper squares.

1. Multiply the skin bacteria in nutrient broth.

2. Inoculate sterile agar plates with this bacteria.

3. Place paper squares soaked in the various test solutions on the inoculated plates and incubate them.

1 Multiply the bacteria in nutrient broth

Prepare some nutrient broth suitable for the bacteria you will culture.

Nutrient broth contains the same nutrients as nutrient agar, but lacks the agar jelly (so it remains liquid).

Different kinds of bacteria require different nutrients, so if you’re getting the bacteria from a university or polytech, make sure you get their recipe for the nutrient broth and agar.

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Pure strain of M Luteus (a common skin bacterium) obtained from Christchurch Polytech.

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Soak the wire inoculating loop in pure ethanol for 70 seconds.

Sterilise the wire loop.

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Heat the loop in a flame until it glows red hot for at least 10 seconds.

Rest the handle of the loop on something that allows it to cool without the sterile loop touching anything.

Keeping the jelly upside down, open the plate with the pure culture on it and use the sterile loop to scoop up a little of the bacteria.

Add bacteria to broth.

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Keep the loop in your hand. Use your last two fingers to open the bottle containing the nutrient broth. Keep holding the lid and the loop.

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Flame the mouth of the bottle to sterilise it.

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Put the loop containing bacteria into the broth...

... and stir it around.

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Flame the mouth of the bottle again.

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Put the lid of the bottle back on.

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To kill the bacteria remaining on the loop, sit it in ethanol for 70 seconds...

Resterilise the loop.

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...then hold the loop in a flame until it is red hot and allow it to cool.

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Label the bottle with the culture and date.

Multiply the bacteria.

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Place the bottle in an incubator at the recommended temperature (30 °C for skin bacteria) for about 24 hours.

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Check that the broth has turned cloudy.

If it is not cloudy, the numbers of bacteria in the broth will be low. Either leave it for another day or start again with a fresh solution.

Cloudy

Clear

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2 Inoculate the agar plates

For this step it is important that the bacteria in the broth be actively growing — that is, fresh out of a warm incubator, not a solution kept in the fridge for a week.

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Open a packet of sterile cotton swabs.

(Wooden swabs with cotton tips should be wrapped in foil and heated in the oven at 200 °C for an hour, then allowed to cool. Alternatively, use a new packet of cotton buds.)

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Open the bottle of nutrient broth and flame the neck as before.

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Dip the first cotton tip into the broth.

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Open the first sterile plate of nutrient agar, holding the lid over it as shown.

Gently wipe the surface of the agar with the wet cotton, rolling it to transfer as much of the bacteria as possible.

Use a zig zag motion to cover the whole surface.

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Warning: Do not dip the swab back into the broth — you might contaminate the broth with stray bacteria.

Rotate the plate 60° and zig zag again to spread out the bacteria.

Rotate again, with a final zig zag.

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Put the lid back on the agar plate, discard the swab, and repeat for the next plate.

All used swabs should be sterilised before disposal.

Flame the neck of the bottle of broth before dipping in each new swab.

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3 Test the antibacterial compounds

Prepare antiseptic solutions according to the manufacturer’s directions.

If you wish to investigate dilution effects, try using the solution at the stated concentration, then half, quarter and eighth strength.

Your teacher will give you a set of sterile filter paper squares. (Cut filter paper into 1 cm squares, or use a cork borer to stamp out paper discs. Wrap these in foil and sterilise at 200 °C in the oven as with the cotton swabs.)

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Dip metal tweezers in alcohol, flame them and allow to cool (as for the wire loop).

Use the tweezers to dip a sterile square of filter paper into the prepared solution, allow it to drip for a second...

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... and place the square on an inoculated agar plate.

The squares should be wet, but not dripping. Make sure they make good contact with the agar and that you know where each square went.

Reflame the tweezers between each solution.

Each plate should also have a square of paper dipped in sterile water (as a control).

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Once the plate is completed, seal it with sellotape.

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If you have access to antibiotic discs, use sterile tweezers to place them on another inoculated agar plate.

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Place the prepared and sealed plates upside down in an incubator at the recommended temperature for 2–3 days.

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The white parts of this plate are the bacteria. Clear zones around some filter paper squares show regions where the bacteria could not grow.

Cetrimide and Savlon are the antiseptics most effective against this particular strain of bacteria.

W = sterile water

S = Savlon

D = Dettol

L = Listerine

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6H 2 Attacking bacteria

6H 3 Disinfectant trials

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Antibiotic-resistant bacteria

Soon after the discovery of antibiotics, doctors started to find strains of bacteria that had become resistant to antibiotics that had worked only a few months earlier.

Although bacterial reproduction through binary fission is asexual – producing identical copies of the parent cell – bacteria have developed different ways to change their genes.

Small errors in the reproduction process may cause a ‘spelling mistake’ in the DNA. Just as some spelling mistakes are real words, so sometimes these mutations create new genes that might be useful.

Sometimes a bacterium acquires DNA by exchanging part of its chromosome with another bacterium – including those of quite different types.

Many bacteria contain plasmids, which are small loops of DNA containing only a few genes each (a bit like applications on a computer). The plasmids are copied each time the cells reproduce, but any time two bacteria touch, they can transfer plasmids. Antibiotic resistance genes are frequently found in plasmids.

A defence that works on a particular antibiotic is often equally effective against other compounds – antibiotics or antiseptics – that kill in the same way.

Every time we use an antimicrobial agent, we assist in the development of resistant strains. Thus, we need to be sensible in our use of these compounds. Don’t take antibiotics to combat colds, flu or

other viral diseases: antibiotics only kill bacteria.

Always finish the course (take all the pills) of antibiotics prescribed, even if you feel better after a day or two.

Only take antibiotics prescribed by a doctor – they know which one is best for each kind of bacteria.

Only use disinfectants around the home when they are needed to fight disease, and not for routine household cleaning.

Example: MRSA – Methicillin Resistant Staphylococcus Aureus

Soon after the invention of the antibiotic methicillin, strains of resistant staphylococci were found and became known as MRSA. The bacteria quickly became resistant to other antibiotics similar to methicillin, and have since also become resistant to several other types of antibiotics as well. Some people call it a superbug.

MRSA is particularly dangerous in hospitals where it may infect surgery wounds.

Antibiotic resistant bacteria

Antibiotic resistant strains of bacteria are not killed by an antibiotic that would normally kill that kind of bacteria.

Resistant strains sometimes occur because of mutations (mistakes) that occur during reproduction. More commonly, bacteria acquire resistant genes by exchanging DNA or plasmids (small loops of DNA) with other bacteria.

To reduce the development of resistant strains of bacteria: don’t use antibiotics to fight viral diseases; take all the antibiotic pills prescribed; only take antibiotics prescribed by a doctor; and only use antiseptics at home when they are needed.6H 5 Friendly fungi 6H 6 Resisting resistance

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