BUG & SUPERBUGBUG & SUPERBUG(with apologies to George (with apologies to George

Bernard Shaw)Bernard Shaw)

DR PETER VICKERS

UNIVERSITY OF HERTFORDSHIRE

THE SCALE OF THE PROBLEMTHE SCALE OF THE PROBLEM

The 1990s brought a world-wide resurgence of bacterial and viral diseases - an important factor of this being antibiotic resistance genes in virtually all major bacterial pathogens

US National Institute of Health estimated in 2000 that 50 000 tons of antibiotics will be used every year throughout the world on humans, animals and plants.

It makes the possibility of a mass mutation of microbes (bugs) and thus new diseases, inevitable.

ENTEROCOCCIENTEROCOCCI

In the late 1980s, certain species and strains of enterococci had evolved to become immune to all known antibiotics, including vancomycin.

By 1994, some mutant enterococci were not just immune to vancomycin, but actually fed off it.

One strain of Enterococcus faecalis had actually become dependent on vancomycin for growth.

ANTIBIOTICS ARE ABUSEDANTIBIOTICS ARE ABUSEDAs far back as 1983, a WHO report

estimated that doctors used antibiotics irrationally or inappropriately on anything between 40% and 66% of all occasions.

By 1994, between 60 - 70 000 patients died from nosocomial infections, 50% of which are caused by drug-resistant micro-organisms

Things have not improved - think of MRSA, resistant TB, E coli 0147 - the list goes on.

Are we running out of viable antibiotics?

BACTERIA & ANTIBIOTICS (1)BACTERIA & ANTIBIOTICS (1)

When you take antibiotics, the drug kills vast numbers of bacteria in the gut and elsewhere on and in the body - but countless millions remain.

The bacteria that survive include species unaffected by the drug, or else drug-resistant variants of a vulnerable species.

After a number of courses of antibiotics, these surviving bacteria are liable to multiply and to fill the space created by the drug.

BACTERIA & ANTIBIOTICS (2)BACTERIA & ANTIBIOTICS (2)What antibiotics do is to exert ‘selective

pressure’ on bacteria. This distorts and accelerates their evolution,

so that previously vulnerable species become drug resistant.

In a population of many millions of bacteria, there will be some chance mutants - maybe one in a million - that just happen to be invulnerable to a drug that kills all the others.

These once insignificant mutants survive, multiply and colonise the space left by the destruction of other bacteria in that species that were previously vulnerable to the drug.

BACTERIA & ANTIBIOTICS (3)BACTERIA & ANTIBIOTICS (3)However, the creation of drug-resistant

bacteria is not just a matter of chance mutation.

Although microscopic, bacteria are very complex and adaptable living organisms.

Their genetic organisation includes structures that probably evolved in order to resist naturally occurring chemicals contained in rival living things (think of fungi and penicillin), just as plants, insects, birds, animals and humans have evolved ways to resist predators.

BACTERIA & ANTIBIOTICS (4)BACTERIA & ANTIBIOTICS (4)These structures, which are rings of genetic

material contained within the bacterial cell wall, but not the nucleus, are PLASMIDS.

The codes for bacterial resistance to one or more antibiotics are contained within the plasmids.

Under selective pressure from drugs, these plasmids can be transferred within bacteria from the same or different species.

Therefore, bacteria are able to transfer multiple drug resistance within and between species.

GENETIC RECOMBINATIONGENETIC RECOMBINATIONIn order for a genetic mutation - for example,

drug resistance - to be passed from one bacterium to another, there needs to be some form of genetic recombination.

This is not something exclusive to bacteria - we do the same thing at the beginning of meiosis.

There are 3 ways in which bacteria are able to transfer genetic information:

1. Transformation2. Conjugation3. Transduction

TRANSFORMATION (1)TRANSFORMATION (1)During this process, genes are transferred

from one bacterium to another as ‘naked’ DNA in solution

Some bacteria, perhaps after death, release their DNA into the environment (our bodies).

Other bacteria can then encounter the DNA and, depending on the particular species and growth conditions, take up fragments of the DNA into their cytoplasm and then combine that DNA into their own DNA

A cell having this new combination of genes is a kind of hybrid - or recombinant cell.

TRANSFORMATION (2)TRANSFORMATION (2)All descendants of this recombinant cell will

be identical to it.Transformation occurs naturally among very

few types of bacteria, including bacilli, haemophilus, neisseria and certain strains of streptococcus and staphylococcus, because even though only a small portion of DNA is transferred, it is still a very large molecule to pass through the cell wall.

So, this works best when the donor and recipient cells are very closely related.

CONJUGATION (1)CONJUGATION (1)Conjugation is mediated by plasmids.These are sub-cellular organisms (rings of

nucleic acid) that live and multiply only within the bacterial cell wall.

They are always beneficial to the bacterium and can be vital to bacterial health and survival.

They are like mini-chromosomes, but are not usually essential for cell growth.

Plasmids carry information that is not contained in bacterial DNA.

CONJUGATION (2)CONJUGATION (2)This information includes genes that protect

the bacterial host against poisons in the environment - including antibiotics.

Conjugation requires that there be direct cell-to-cell contact

The conjugating cells must generally be of opposite ‘mating types’ - donor cells must carry the plasmid and recipient cells usually do not.

The bacterial chromosomes themselves do not cross from cell to cell, only the plasmid.

CONJUGATION (3)CONJUGATION (3)

Since most plasmids occur in more than one copy in any cell, the original carrier retains its plasmid, and the recipient gets a copy.

The plasmid carries genes that code for the synthesis of sex pilli - projections from the donor’s cell surface that contact the recipient and help to bring 2 cells into direct contact.

During conjugation, the plasmid is replicated through transfer of a single-stranded DNA copy to the recipient, where the complementary strand is replicated.

CONJUGATION (4)CONJUGATION (4)In E. coli, the F factor is the first plasmid

observed to transfer between cells.Donors carrying F factor (F+ cells) transfer the

plasmid to recipients (F- cells), which become F+ cells as a result.

bacterial chromosome

plasmid

F+ cell F- cell

Replication & transfer of F factor

F+ cell F+ cell

CONJUGATION (5)CONJUGATION (5)

Once within the recipient cell, donor DNA can combine with the recipient’s DNA, as occurs in transformation.

By the process of conjugation between an these cells, a cell can acquire new versions of chromosomal genes (just as in transformation).

TRANSDUCTION (1)TRANSDUCTION (1)In this process, bacterial DNA is transferred

from the donor cell to the recipient cell inside a virus that infects bacteria - a bacteriophage (phage).

During the process of infection, the phage attaches to the bacterial cell wall and injects its DNA into the bacterium.

Infecting phage

Phage DNA

Bacterial DNA

TRANSDUCTION (2)TRANSDUCTION (2)

The phage DNA acts as a template for the production of new phage DNA, and also directs the production of phage protein coats.

During phage development inside the infected bacterium, the bacterial chromosome breaks apart, and at least some fragments of the bacterial chromosome happen to be packaged inside phage protein coats.

TRANSDUCTION (3)TRANSDUCTION (3)These resulting phage particles thus carry

bacterial DNA instead of phage DNA.

Phage DNA

Bacterial DNA

lysis

TRANSDUCTION (4)TRANSDUCTION (4)When the released phage particles later

infect a new population of bacteria, bacterial genes will be transferred to the newly infected cells.

Bacterial DNA

TRANSDUCTION (5)TRANSDUCTION (5)

So, transduction of cell DNA by a phage virus can lead to recombination between the DNA of the first host bacterium and the DNA of the second host bacterium.

This method of generalised transduction is typical of phages such as P1 of E. coli and phage P2 of Salmonella.

All genes contained within a bacterium infected by a generalised transducing phage are equally likely to be packaged in a phage coat and transferred.

CAUSES OF DRUG CAUSES OF DRUG RESISTANCE (1)RESISTANCE (1)

OveruseNot completing courses of antibioticsOver-the-counter sales in developing

countriesFactory farming - used on animals: found in

meat and milkAgricultural waste (which carries antibiotics

faeces, etc.) polluting earth, rivers

CAUSES OF DRUG CAUSES OF DRUG RESISTANCE (2)RESISTANCE (2)

Antibiotics getting into the food chainOver-crowded environments, e.g.

hospitals, factory farms, developing world

Poor social and physical infrastructuresIrresponsible attitudes to antibioticsHigh costs of developing antibiotics

WHAT TO DO? (1)WHAT TO DO? (1)

Need for a balance between benefit and risk?

Publish an annual list of essential antibiotics?

Keep a reserve list of antibiotics - only to be used in an emergency (such as vancomycin at the moment)?

Make invasive drug-resistant bacterial infection notifiable?

Ensure that antibiotics are only available on prescription?

WHAT TO DO? (2)WHAT TO DO? (2)

Encourage drug companies to market drugs consistently world-wide?

Create an independent drug-regulating agency?

Establish national and/or international groups to audit antibiotics and antibiotic use?

Publicise information about drug-resistant infections?

Give microbiology/immunology a higher profile in medical and nursing education?

WHAT TO DO? (3)WHAT TO DO? (3)

Protect yourself against infection (good nutrition, etc.) - make the individual responsible for self?

Patients to keep a note of drugs taken, what for, and for how long - a drug diary?

Educate the general population to work with doctors and not demand antibiotics for everything?

Cut down on travel?Develop new antibiotics - gene therapy?

Finally – something to Finally – something to think aboutthink about