Maybe They're Born With It: Intrinsic Antibiotic Resistance

March 30, 2017

Some microbes are naturally able to resist the antimicrobial activities of antibiotics due to their structure or functional processes. In one example, Enterococci species are intrinsically resistant to cephalosporins because this drug binds to a peptidoglycan binding protein PBP) that these bacteria don’t have. In another example, Klebsiella species are intrinsically resistant to ampicillin based on their production of beta-lactamases that destroy the drug before it can reach its PBP targets.

The first example, in which the bacteria lack the drug target, is hard to circumvent (it would be hard to force Enterococci to express the targeted PBP gene), but the second can be counteracted – by exposing the bacteria to both ampicillin and a beta-lactamase inhibitor, the bacterium can be rendered susceptible. Several recently published mBio studies describe new mechanisms of intrinsic antibiotic resistance, and these mechanisms may themselves become therapeutic targets to broaden the application of currently available drugs.

intrinsic resistance 1A simplified schematic of peptidoglycan cell wall synthesis and recyclingSource.

In the presence of good nutrient availability, bacteria will grow, and to do so they must be able to rearrange the cell wall. Cell wall rearrangements are necessary to accommodate cell expansion and division, but the cleaved peptidoglycan fragments are valuable, energy-expensive molecules that many bacteria internalize and recycle into new cell wall material. While breaking down parts of the peptidoglycan cell wall, there are two major pathways that recycle N-aceytlmuramic acid (MurNAc), one of the major cell wall components along with N-acetylglucose (GlcNAc). Some bacteria, like Escherichia coli, can convert MurNAC into GlcNAc-6-P, which then enters the peptidoglycan synthesis pathway; this pathway can be inhibited by fosfomycin, which binds to MurA (see figure, right). Other bacteria, particularly gram-negative bacteria, can recycle MurNAC directly into UDP-MurNAc through a salvage pathway, bypassing the steps inhibited by fosfomycin. Two recent mBio studies identify a Pseudomonas gene important for this salvage pathway and its intrinsic fosfomycin resistance. 

mBio: The N-Acetylmuramic Acid 6-Phosphate Phosphatase MupP Completes the Pseudomonas Peptidoglycan Recycling Pathway Leading to Intrinsic Fosfomycin Resistance.

mBio: Identification of MupP as a New Peptidoglycan Recycling Factor and Antibiotic Resistance Determinant in Pseudomonas aeruginosa.

Scientists Coralie Fumeaux and Thomas Bernhardt used a genetic approach to identify P. aeruginosa transposon-insertion mutants resistant to cell wall stress. Their screen uncovered several genes, including regulators of beta-lactamase activity, but also identified PA3172, annotated as encoding a phosphoglycolate phosphatase that the researchers called mupP. A mupP mutant was more resistant to ceftazidime and cefotaxime, a phenotype that depends on ampR and ampC, regulators of the cell-wall stress pathway, but this mutant was also was more sensitive to fosfomycin. These changes in drug susceptibility are due to the disruption of the salvage pathway and reliance of the mutant strain on its peptidoglycan synthesis pathway.

intrinsic resistance 2Fosfomycin resistance of WT (L), mupP mutant (C), and reconstituted (R) P. putidaSource.

Scientists Marina Borisova, Jonathan Gisin, and Christoph Mayer used a complementary approach to identify mupP. They first demonstrated the biochemical activity of a MurNAc-6-P phosphatase from P. putida cell extracts and then used bioinformatics to identify candidate genes responsible for this activity. Demonstration that the candidate mutant mupP strain has increased fosfomycin susceptibility (see figure, right), as well as aberrant levels of the peptidoglycan precursors UDP-GlcNAc and UDP-MurNAc, helped confirm a role for this gene in the Pseudomonas peptidoglycan salvage pathway.

These two studies suggest that a MupP inhibitor co-administered with fosfomycin may enable anti-Pseudomonas fosfomycin activity. Administering drug combination cocktails isn’t a new concept; co-therapy of the beta-lactamase inhibitor avibactam with certain antibiotics shows promise, especially in treating multidrug-resistant gram-negative bacteria. An inhibitor that aids fosfomycin activity would be a big help in fighting Pseudomonas infections, which are especially difficult to eliminate; the intrinsic and acquired antibiotic resistance of this bacterium, along with its ability to form recalcitrant biofilms, has meant increasingly fewer therapeutic options for infected patients. Identifying its mechanisms of intrinsic antibiotic resistance is the first step toward discovering new, effective drugs.

mBio: Antibiotic Capture by Bacterial Lipocalins Uncovers an Extracellular Mechanism of Intrinsic Antibiotic Resistance

intrinsic resistance 3Antibiotic concentration is reduced after incubation with purified B. cenocepacia lipocalinSource.

A second mechanism of intrinsic resistance was recently described in a third mBio article. Here, first author Omar El-Halfawy and senior scientist Miguel Valvano describe a role for the Burkholderia cenocepacia YceI lipocalin. Lipocalins are widely conserved bacterial small proteins with poorly defined functions. Here, the researchers show that B. cenocepacia, P. aeruginosa, Mycobacterium tuberculosis, and Staphylococcal aureus lipocalin gene products are able to bind and prevent the antimicrobial activity of a variety of hydrophobic antibiotics, such as norfloxacin, ceftazidime, and rifampicin (see figure, right).

Just as MupP presents a promising target for cotherapy, so do the lipocalins. By preventing their interaction with antibiotics, the drugs may reach their targets to enact their antibiotic activity. The El-Halfawy report offers promise toward reversing lipocalin activity: vitamin E, a liposoluble vitamin, may counteract the ability of lipocalins to bind and sequester the drugs in the extracellular space. Counteracting intrinsic antibiotic resistance through co-therapy may increase the utility of currently-available antibiotics and allow us to apply drugs to previously nonsusceptible organisms. Neutralizing natural resistance offers one way to fight the growing threat of drug-resistant infections.

Author: Julie Wolf

Julie Wolf
Dr. Julie Wolf is in science communications at Indie Bio, and was a former ASM employee. Follow Julie on Twitter for more ASM and microbiology highlights at @JulieMarieWolf.