Download Mechanisms of Antibiotic Resistance

Mechanisms of Antibiotic Resistance
Antibiotics kill or inhibit the growth of bacteria. Antibiotics are derived from three
sources: moulds or fungi; bacteria; or synthetic or semi-synthetic compounds.
1. Identify, with examples, the mechanisms by which micro-organisms resist the
actions of antibiotics
Bacteria demonstrate 4 basic mechanisms of antibiotic resistance:

Decreased uptake by:
o decreased penetration
 e.g. penicillin and gram negative bacteria and aminoglycosides and
streptococci
o increased efflux


Enzymatic inactivation or modification of the antibiotic


e.g. via energy requiring efflux pump is a well known mechanism of
resistance to tetracyclines
e.g. β–lactamases, which are widespread among many kinds of bacteria,
cleave the 4-membered β-lactam ring found in antibiotics such as
penicillin and cephalosporin
Altered target site; modification of the antibiotic receptor site
 e.g. alteration of PBP (penicillin binding protein) in penicillin resistant
bacteria

Resistant or ‘bypass’ metabolic pathways
 e.g. some sulfonamide-resistant bacteria do not require paraaminobenzoic acid (PABA, an important precursor for the synthesis of
folic acid and nucleic acids in bacteria) which is inhibited by
sulfonamides. Instead, like mammalian cells, they turn to utilizing
preformed folic acid.
2. Give examples of mechanisms of acquisition of bacterial resistance to antibiotics
Bacterial antibiotic resistance can be:
 intrinsic – naturally occurring trait arising from the biology of the
organism
 acquired - the organism was sensitive to the antibiotic but has
developed resistance either by:
o mutations

spontaneous and random, and occur regardless of presence of antibiotic,
but resistant bacteria have advantage in presence of antibiotic and
therefore survive
o acquisition of new DNA

plasmids – self-replicating pieces of circular DNA smaller than bacterial
genome. Transfer can occur across different bacterial species and they
can carry multiple antibiotic resistance genes. Transfer can occur via:
o conjugation - transfer of DNA between bacteria via direct cell to
cell contact,
o transformation - when bacteria die, they release their DNA which
can be taken up by other competent bacteria and
o transduction - transfer of DNA from one bacterium to another by
a virus

transposons carried on plasmids and inserted into chromosomes
3. Identify and briefly describe culture techniques for pathogens and the methods of
assessing antibiotic resistance in vitro
Pathogens can be cultured in a liquid broth or on solid agar plates.
Susceptibility to antibiotics can be tested via:
 broth dilutions for MIC (minimum inhibitory concentration) via manual or
complex automated methods
o serial dilution of antibiotic in broth and addition of a standard inoculum to
determine the MIC (99.9% kill)

Agar Diffusion – Disc methods (Kirby-Bauer Method).
o
o



A known quantity of bacteria is grown on agar plates in the presence of thin
wafers containing relevant antibiotics. If the bacteria are susceptible to a
particular antibiotic, an area of clearing surrounds the wafer where bacteria are
not capable of growing (called a zone of inhibition).
Etest system: comprises a predefined and continuous concentration gradient of
different antimicrobial agents, which when applied to inoculated agar plates and
incubated, create ellipses of microbial inhibition. The MIC is determined where
the ellipse of inhibition intersects the strip, and is easily read off the MIC reading
scale on the strip.
Agar Dilution Method – antibiotic in plates, uses breakpoints
(discriminatory antimicrobial concentrations used in the interpretation of
results of susceptibility testing to define isolates as susceptible,
intermediate or resistant.)
Detection of resistance mechanisms e.g β-lactamases
Oxacillin Resistance Screening Agar – for rapid detection of MRSA
(Methicillin-resistant Staphylococcus aureus), only MRSA can grow
To classify MICs into susceptible or resistant categories, i.e. to assess whether it
is possible to treat an infection with a given antibiotic, reference is made to the
critical values recommended by national committees e.g, in Australia, the
Calibrated dichotomous sensitivity (CDS). Others are used in other countries.