Mechanism of antimicrobial resistance I and II Flashcards
- Describe the difference between intrinsic and acquired antibiotic resistance and list ways a bacterium may acquire antibiotic resistance.
What is antibiotic tolerance?
A. Categories of antibiotic resistance.
1) Intrinsic resistance occurs “just because” of the natural properties of the bacteria in question. An example would be vancomycin resistance of Gram-negative organisms due to the inability of vancomycin to penetrate the outer membrane of Gram-negative organisms.
2) Acquired resistance develops by genetic mutation or by acquisition of new genes. New genetic material mediating antibiotic resistance is usually spread from cell to cell by way of mobile genetic elements such as plasmids, transposons, or bacteriophages.
- Recognize the role of porins and efflux pumps in mediating antibiotic resistance and for which types of bacteria they can be important.
Where are porins found?
What are they?
A. Porins
1) Found in the *outer membrane of gram-negative bacteria (remember there is NO outer membrane in gram-positive bacteria).
2) Porins form hydrophilic channels to allow the selective uptake of essential nutrients and other compounds, including some hydrophilic antibiotics.
3) Changes in configuration or number of porin channels may adversely affect uptake of antibiotics.
- Recognize the role of porins and efflux pumps in mediating antibiotic resistance and for which types of bacteria they can be important.
What is the function of efflux pumps?
Present in what type of bacteria?
Efflux pumps
1) Structures located in bacterial cell membranes that function to eliminate certain substrates from the bacterial cytoplasm. They originated as mechanisms to get rid of toxic substances from inside the bacterium, including antibiotics.
2) Can be present in both gram-positive and gram-negative organisms.
3) Can adversely affect uptake of antibiotics.
4) Efflux pumps can be specific for a certain drug or class of drugs, or may be more general and result in multi-drug class resistance.
Bottom line is to understand that porin channels and efflux pumps can prevent drug/target interaction that can contribute to antibiotic resistance
Describe structure of peptodyglycans:
1) Structure: backbone of two alternating sugars, N-acetylglucosamine (GlcNAc) and N-acetylmuramic acid (MurNAc) cross-linked via peptide bridge.
2) Formation: Peptidoglycan is formed by the addition of subunits called precursors (a GlcNAc-MurNAc disaccharide with its five attached amino acids on the MurNAc) that are assembled in the cytoplasm and transported through the cytoplasmic membrane to the cell surface.
3) Cross-linking (transpeptidation): is driven by cleavage of the terminal stem-peptide amino acid. **The enzymes that perform cross-linking of peptidoglycan are called Penicillin Binding Proteins (PBPs)
- Distinguish between types of beta-lactamases (e.g., narrow spectrum, ESBLs, carbapenemases) based on their spectrum of activity and sensitivity to inhibition.
How do b-lactam antibiotics work??
Biosynthesis of peptidoglycan is accomplished by membrane-bound enzymes known as penicillin-binding proteins (PBPs). PBPs perform transpeptidase and/or transglycosylase reactions. These PBPs are involved in peptidoglycan synthesis during cell growth, cell septation and, in some species, sporulation. b-lactam antibiotics act by irreversibly binding to and inactivating the transpeptidase reaction of penicillin-binding proteins, thereby inhibiting peptide cross-linking and peptidoglycan synthesis.
How do bacteria become resistant to b-lactams??
1) Modifying the drug—through enzymatic destruction of b-lactams by b-lactamases
2) Modifying the target—through production of altered penicillin-binding proteins (PBPs) with low affinity for b-lactam binding.
3) Preventing drug-target interaction—both porin channel mutations and drug efflux mechanisms can increase resistance to b-lactams by preventing b-lactam interaction with the PBPs.
- Distinguish between types of beta-lactamases (e.g., narrow spectrum lactamases, ESBLs, carbapenemases) based on their spectrum of activity and sensitivity to inhibition.
What type of antibiotics do they hydrolyze?
What antibiotics do they don’t hydrolize?
Found on what type of bacteria?
Why are they special for gram-negative bacteria?
a) These beta-lactamases generally hydrolyze penicillin-type antibiotics (penicillin, ampicillin, amoxicillin and possibly piperacillin) but don’t have much activity against cephalosporins or carbapenems.
They can be found in both gram-positive and gram-negative organisms AND they can occur frequently (e.g. more than 95% of S. aureus isolates contain a narrow spectrum beta-lactamase) or less commonly like TEM-1 in E. coli.
b) **For gram-negative bacteria, these initial “emerging” resistant strains that elaborated narrow-spectrum b-lactamases, such as the TEM-1 of E. coli and SHV-1 of Klebsiella pneumoniae prompted the ***development of newer b-lactams (the extended spectrum b-lactams like 2nd and especially the 3rd generation cephalosporins).
Beta-lactamamses Narrow spectrum:
- Distinguish between types of beta-lactamases (e.g., narrow spectrum, ESBLs, carbapenemases) based on their spectrum of activity and sensitivity to inhibition.
What is the characteristic that distinguishes from them parental types?
Where are they usually found?
They are founds mostly on what organisms?
a) Most “extended-spectrum b-lactamases” (ESBLs) began to arise in the 1980s and are mutants of TEM-1, TEM-2 and SHV-1 (“narrow” spectrum b-lactamases) with 1-to-4 amino acid sequence substitutions. In addition, the capture and spread through horizontal transfer of new genes that encode enzymes with ESBL activity occurred in the 1990s (e.g., CTX-M enzymes)
b) The most notable feature of these enzymes, distinguishing them from their parent types, is their **ability to attack most cephalosporins (like the 3rd generation cephalosporins—ceftriaxone and ceftazidime)
c) **ESBLs are usually found on plasmids, which means they are highly mobile and can disseminate, and are often present with additional genes encoding resistance to other classes of antibiotics.
d) **Found almost exclusively in Gram-negative aerobic rods, especially E. coli and Klebsiella pneumoniae, but prevalence is still pretty low (most isolates do NOT have them).
- Distinguish between types of beta-lactamases (e.g., narrow spectrum, ESBLs, carbapenemases) based on their spectrum of activity and sensitivity to inhibition.
Carbapenemases
- Describe the enzymatic activity and gene regulation of the beta-lactamase, AmpC and distinguish between inducible and constitutive expression based on antibiotic susceptibility data.
Where is ampC found?
Is ampC always expressed? What is the difference between inducible and constitutive?
What two B-lactams can induce ampC?
What can lead to constitutive expression of ampC?
a) ampC was first characterized as a chromosomally located gene in a number of gram-negative organisms. It encodes for a b-lactamase (technically a cephalosporinase) that is capable of hydrolyzing penicillins, and all 1st, 2nd, and 3rd-generation cephalosporins, and its activity is not inhibited by beta-lactamase inhibitors (ESBLs can be inhibited by some b-lactamase inhibitors but some question whether they should be used clinically).
b) Found in the chromosome of certain Gram-negative rods: Enterobacter, Pseudomonas, and a few others.
Is the chromosomal ampC beta-lactamase always expressed??
The tricky part of understanding chromosomal ampC is that its expression is either inducible (can be turned on) or constitutive (is on all the time).
a) Under normal conditions, only trace amounts of AmpC are expressed, but this expression can be induced with some (not all!!) b-lactams, conferring inducible resistance to b-lactams like ampicillin and cefazolin (1st generation cephalosporin).
b) Acquisition of some mutations can lead to constitutive expression of AmpC. When that occurs, ampC is expressed all the time and mediates resistance to all b-lactams except carbapenems
See image:
What does this mean clinically??
The mutational events that lead to permanent high-level expression of ampC can occur during therapy! Thus, if you treated the first Enterobacter strain with ceftriaxone, you might select for a constitutive mutant during therapy.
- Describe the enzymatic activity and gene regulation of the beta-lactamase, AmpC and distinguish between inducible and constitutive expression based on antibiotic susceptibility data.
Difference between ESBL and ampC
- Compare and contrast the mechanisms that result in modified penicillin-binding proteins (PBPs) and their impact on resistance to beta-lactam antibiotics.
A second mechanism of bacterial resistance to b-lactam antibiotics is production of a PBP that has a markedly reduced affinity for the drug (beta-lactams). While this alteration may occur by mutation of existing genes, the acquisition of new PBP genes or new pieces of PBP genes is more important.
mecA of staphylococcus Aureus
- Describe the mechanism of vancomycin resistance mediated by VanA/VanB, and recall which organisms might house these genetic elements.
Vancomycin
Resistance:
- Altered PG precursor: change in terminal peptide
- **Plasmid mediated (transposon encoded)
- VanA, VanB (and others), encode enzyme that changes terminal peptide (d-ala-d-ala -> d-ala-d-lac)
- Seen almost exclusively in Enterococci (VRE)
**Much more commonly seen with ****E. faecium than with E. faecalis.**
**Large worry that plasmid would transfer to S. aureus, but it has been rare (VRSA–see Dr. French handouts)
- Describe the process of target modification that leads to fluoroquinolone resistance and recall in which types of organisms this may occur.
How do quinolones work?
• Target DNA gyrase and DNA topoisomerase IV.
Quinolones trap the enzymes in a drug-enzyme-DNA complex leading to lethal DNA strand breaks
- Recall the aminoglycoside-modifying enzymatic reactions associated with antibiotic resistance.
