Antimicrobial Medications

Antibiotics• Antimicrobial drugs naturally

produced by microorganisms– Penicillium species: Penicillins– Cephalosporium specis:

cephalosporins– Streptomyces species:• lincosamides, aminoglycosides,

tetracyclines, chloramphenicol

Features of antimicrobial drugs• Selective toxicity

– Therapeutic index• Antimicrobial action

– Bactericidal– Bacteristatic

• Spectrum of activity– Broad spectrum– Narrow spectrum

• Combination effects– Antogonistic– Synergistic– Additive

Pharmacokinetics: what happens to the drug in the body?

• Absorption• Tissue distribution• Metabolism• Route of excretion• Rate of elimination

Adverse effects• Adverse drug reaction• Toxic effects• Suppression of normal

microbiota

Mechanisms of antimicrobial drugs

• Inhibition of cell wall synthesis• Inhibition of protein synthesis• Inhibition of nucleic acid synthesis• Inhibition of biosynthetic pathways • Disruption of cell membrane integrity

Targets of Cell Wall Synthesis

Penicillin

• Inhibits formation of tetrapeptide side chains

How organisms degrade penicillins

Family of Penicillins

• Natural penicillins• Penicillinase-resistant penicillins• Broad-spectrum penicillins• Extended-spectrum penicillins• Penicillins plus beta-lactamase inhibitors

Family tree of penicillins

β-lactams

• Penicillins• Cephalosporins• Carbapenems• Vancomycin• Bacitracin

Cephalosporins

• Derived from fungus, Acremonium cephalosporium

• Chemical structure makes them resistant to beta-lactamase: low affinity for penicillin binding proteins

• Grouped into first, second, third, and fourth generation cephalosporins

Vancomycin

• Binds to the terminal amino acids of the peptide chain of NAM molecules, blocks peptidoglycan formation

Antibiotics that inhibit protein synthesis

Oxazolidinones

• Reversibly bind to the 50S subunit, interfere with initiation of protein synthesis

• Used for treating gram positive infections resistant to Beta-lactam drugs and Vancomycin

• Ex: Linezolid

Aminoglycosides

• Bactericidal • Irreversibly bind to 30S ribosome, cause

misreading of the mRNA• Transported into cells that actively respire (not

effective against ananerobes, streptococci, enterococci)

• Ex: streptomycin, gentamicin, tobramycin

Tetracyclines & Glycylcyclines

• Bind reversibly to 30S, block attachment of the tRNA to ribosome

• Actively transported into bacterial cells• Effective against gram positive and gram

negative• Resistance: due to decrease in uptake or

increase in excretion• Ex: Doxycycline

Macrolides

• Reversibly bind to the 50S, prevent continuation of protein synthesis

• Drug of choice for patients allergic to penicillins

• Not good for Enterobacteriaceae• Ex: Erythromycin, Azithromycin• Resistance: enzymes that alter drug,

decreased uptake

Inhibition of protein synthesis: Chloramphenicol

• Rare side effect = irreversible bone marrow suppression

• Banned in food animals• Making a come-back in companion animal

medicine due to effectiveness against multi-drug resistant staphylococci

Antibiotics that inhibit nucleic acid synthesis

• Fluoroquinolones– Interferes with function of topoisomerase

• Rifamycins– Blocks prokaryotic RNA polymerase from initiating

transcription

Antibiotics that inhibit biosynthetic pathways

• Sulfonamides• Trimethoprims

Sulfonamides (sulfa drugs)

• First synthetic drugs to treat microbial infections

• Used to treat urinary tract infections (UTIs)• Combination of trimethoprim and

sulfamethoxazole (TMP-SMZ) example of synergism

Drugs used together inhibit folic acid synthesis

Tests for microbial susceptibility

• Kirby-Bauer (disk diffusion method)

Tests for microbial susceptibility

• Minimum Inhibitory Concentration: MIC– Grow bacteria in a serial dilution of the

antimicrobial being tested– Fixed concentration of bacterial cells– Observation of turbidity after 16 hrs-24 hrs of

growth– Lowest concentration of drug that inhibits growth

= MIC

Minimum Inhibitory Concentration

• Manual broth dilution method• Automated broth dilution method• E-test

Determining the Minimum Inhibitory Concentration (MIC)

Automated MIC

E-test for MIC

Zone size & MIC values

• Raw data• Meaningless without interpretation• Correlation of in vitro results with achievable

levels of drug concentration in a live patient• Correlation with actual clinical outcome

What resistance looks like…

Mutant Prevention Concentration?

Mechanisms of acquired drug resistance

• Destruction or inactivation of the drug: drug inactivation enzymes

• Alteration of target molecule (mutation)

• Decreased uptake: alteration of porins

• Increased elimination: efflux pumps

Acquiring resistance

• Spontaneous mutation• Gene transfer– R plasmids

Emerging antimicrobial resistance

Streptococcus pneumoniae

• Altered penicillin binding proteins– DNA-mediated transformation

Enterococcus species

• Gram positive enteric cocci; facultative anaerobes; formerly classified as Group D streptococcus

• Common cause of nosocomial infections– Enterococcus faecalis,

Enterococcus faecium• Intrinsic resistance:• Acquired resistance

Mycobacterium tuberculosis

• Multidrug-resistant M. tuberculosis– Resistance to isoniazid

& rifampin• Extensively drug-

resistant M. tuberculosis– Resistance to isoniazid

& rifampin + 3 or more of the 2nd line drugs

Enterococcus species: intrinsic resistance

• Low affinity of penicillin binding proteins for many β-lactam antibiotics, esp. cephalosporins

• Resistance to potentiated sulfonamides (i.e., trimethoprim-sulfa): able to utilize external sources of folate

• Low permeability for aminoglycosides– Treatment with a cell-wall active drug such as

ampicillin is synergistic (allows the drug to get into the cell) UNLESS high-level gentamicin resistance is present

Enterococcus species: acquired resistance

• High-level gentamicin-resistance: plasmid-encoded inactivating enzymes

• Tetracycine resistance: efflux pumps, ribosomal protection

• Macrolide resistance: efflux pumps• Vancomycin resistance: altered drug binding

site on cell wall

Enterobacteriaceae• Gram negative enteric

rods• Intrinsic resistance to

many drugs due to outer membrane

• β-lactamases: enzymatic inactivation of the lactam ring

• Extended spectrum β-lactamases (ESBL+)

• Carbapenem-resistance: enzymatic inactivation

Staphylococcus species

• Staph aureus• Staph pseudintermedius• Staph schleiferi• Methicillin-resistant staph: penicillinase +

altered penicillin-binding proteins with low affinity for β-lactam drugs (mecA gene on R plasmid)

• Vancomycin-resistant staph

Methicillin-resistant staphylococci• MRSA: methicillin-resistant Staph aureus– drug resistance + increased pathgenicity

• MRSP: methicillin-resistant Staph pseudintermedius– Acquisition of drug resistance is not associated

with acquisition of new virulence factors• MRSS: methicillin-resistant Staph scheiferi– Drug resistance/no new virulence factors

• Coagulase-negative MRS

Coagulase test

• Tests for coagulase enzyme = virulence factor produced by Staphylococcus aureus, Staph. pseudintermedius and Staph. schleiferi subspecies coagulans

• Important in differentiating potentially pathogenic from non-pathogenic species of staphylococci

Coagulase enzymes

• Bound coagulase (“clumping factor”) – attached to bacterial cell wall– Coagulase enzyme + fibrinogen in plasma → fibrin

clot surrounding bacteria: prevents antibody and complement binding, prevents phagocytosis, protects from NETs

• Free coagulase – secreted enzyme– Coagulase enzyme + CRF → conversion of

prothrombin to thrombin and fibrinogen to fibrin

Coagulase slide test

• Rabbit plasma + bacteria: agglutination within 1-2 minutes = positive result– Detects only bound coagulase– False negatives or equivocal results are common– Negative or equivocal tests have to be confirmed

with tube test

Coagulase tube test

• Rabbit plasma + bacteria: coagulation of the plasma = thickening OR formation of fibrin clumps or threads– Standard practice = read at 4 hrs, if negative

recheck at 24 hrs• Tests not read at 4 hrs that are negative at > 4 hrs will

have to be repeated because early positive results may revert to a negative result

Coagulase negative staph

• Common isolates from skin cultures• Non-pathogenic commensuals• Rarely part of mixed population in deep skin/

wound infections (furuncles)• Rarely cause bacteremia or other systemic

infections in immune-compromised individuals

• Commonly carry plasmids with mecA gene

MRS: colonization vs. infection

• Sharing of plasmids + high antimicrobial use• Selecting for MRS population• Increasing % of staphylococcal isolates from

non-lesional skin and nasal mucosa are methicillin resistant

Responsible drug use

• Use vs misuse of antimicrobial drugs• Responsibilities of health care professionals?• Responsibilities of patients?• Responsibilities of pet owners?• Public education• Over the counter antimicrobial drugs– Developing countries– US feed stores

Antimicrobial Stewardship

• Increasing drug resistance• Fewer drugs in development• Drugs being developed don’t have novel

targets• Drugs being developed are broad spectrum• “Bad Bugs, No Drugs” task force 10x20

initiative

Antimicrobial Stewardship

• 4 D’s of antimicrobial therapy– Right Drug, – Right Dose, – De-escelation to pathogen directed therapy – Right Duration of therapy

• Prevent overuse, misuse and abuse• Minimize the development of resistance