Pseudomonas aeruginosa is a Gram-negative pathogen that forms
biofilms, which increase tolerance to antibiotics. Biofilms are dense, surfaceassociated
communities of bacteria that grow in a self-produced matrix of
polysaccharides, proteins, and extracellular DNA (eDNA). Sub-minimal inhibitory
concentration (sub-MIC) levels of antibiotics induce the formation of biofilms,
indicating a potential role in response to antibiotic stress. However, the
mechanisms behind sub-MIC antibiotic-induced biofilm formation are unknown.
We show that treatment with sub-MIC levels of cefixime (cephalosporin),
carbenicillin (β-lactam), tobramycin (aminoglycoside), chloramphenicol
(chloramphenicol), thiostrepton (thiopeptide), novobiocin (aminocoumarin),
ciprofloxacin (fluoroquinolone), or trimethoprim (antifolate) induces biofilm
formation, with maximal induction at ~ ¼ to ½ MIC. We demonstrate that
addition of exogenous eDNA or cell lysate does not stimulate biofilm formation
to the same extent as antibiotics, suggesting that the release of common goods
by antibiotic action does not solely drive the biofilm response. We show that
increased biofilm formation upon antibiotic exposure requires the outer
membrane porin OprF and the extracytoplasmic function sigma factor SigX.
Through transposon mutant screening and deletion studies, we found that OprF
is important for biofilm induction, as mutants lacking this protein did not form
increased biofilm when exposed to sub-MIC antibiotics. OprF expression is
v
controlled by SigX, and its loss increases SigX activity. Loss of SigX also prevents
biofilm induction by sub-MIC antibiotics. Together, these results show that
antibiotic-induced biofilm formation may constitute a type of stress response.
This response may be useful to screen for new antibiotics due to its ability to
reveal antibiotic activity at concentrations below the MIC. Further study of this
response may also provide targets for adjuvant therapies that reduce biofilm
formation in P. aeruginosa infections and increase the efficacy of current
antibiotics. / Thesis / Master of Science (MSc) / Pseudomonas aeruginosa is a bacterium that causes illness in patients
with compromised immune systems, like patients with cystic fibrosis. This
bacterium forms biofilms, which are dense groups that stick to surfaces within a
protective slime that contains proteins, sugars, and DNA. Biofilms make the
bacteria harder to treat with antibiotics. If the bacteria are treated with low
levels of antibiotics, they respond by forming more biofilm but how this happens
is unknown. We showed that adding DNA does not increase biofilm formation,
while adding dead cell debris only causes a small increase. By testing a library of
mutant bacteria, we found that they need two genes, oprF and sigX, to form
more biofilm when they are treated with low levels of antibiotic. By studying
how bacteria respond to low levels of antibiotics, we can find ways to identify
new antibiotics and to make our current antibiotics work better.
| Identifer | oai:union.ndltd.org:mcmaster.ca/oai:macsphere.mcmaster.ca:11375/28186 |
| Date | 11 1900 |
| Creators | Ranieri, Michael |
| Contributors | Burrows, Lori, Biochemistry and Biomedical Sciences |
| Source Sets | McMaster University |
| Language | en_US |
| Detected Language | English |
| Type | Thesis |
Page generated in 0.0023 seconds