Proteomic analysis of ampicillin-resistant oral Fusobacterium nucleatum.
BACKGROUND
Fusobacterium nucleatum represents one of the predominant anaerobic species in the oral microbiota. Penicillin-resistant F. nucleatum have been isolated from intra- and extraoral infections. This study aimed to assess ampicillin resistance in F. nucleatum by investigating the synthesis of resistance-associated proteins.


METHODS
Ampicillin-resistant and ampicillin-susceptible F. nucleatum isolates were obtained from 22 dental plaque samples. Two-dimensional gel electrophoresis and mass spectrometry were used to investigate bacterial protein synthesis. Proteins exhibiting statistically significant quantitative changes between sensitive and resistant isolates were identified using peptide mass mapping and matrix-assisted laser desorption/ionization - time of flight/time of flight (MALDI-TOF/TOF) mass spectrometry.


RESULTS
Twenty-three F. nucleatum isolates were recovered from plaque samples and their ampicillin minimum inhibitory concentrations ranged between 0.125 microg/ml and 256 microg/ml. Analysis of the bacterial cellular proteins by two-dimensional gel electrophoresis resolved 154-246 distinct protein spots (mean 212, n = 9). Between 32% and 83% of the protein spots were common for the F. nucleatum isolates. Comparisons of the protein profiles of sensitive and resistant isolates revealed the presence of a 29 kDa protein and significant increases in the synthesis of two proteins at 37 and 46 kDa in the ampicillin-resistant F. nucleatum isolates. These proteins were identified as a class D beta-lactamase, ATP-binding cassette (ABC) transporter ATP-binding protein and enolase, respectively.


CONCLUSIONS
Synthesis of a class D beta-lactamase by ampicillin-resistant F. nucleatum isolates could complicate antimicrobial treatment because these enzymes might confer resistance to many classes of beta-lactam antibiotics. The differences observed in protein synthesis between ampicillin-resistant and ampicillin-susceptible F. nucleatum may contribute to the antibiotic resistance and virulence of these bacteria.
Al-Haroni
M
M
Department of Oral Sciences - Oral Microbiology, Faculty of Dentistry, and Centre of International Health, University of Bergen, Bergen, Norway. mohammed.al-haroni@student.uib
Skaug
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Bakken
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Cash
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D018528:ATP-Binding Cassette Transporters / Q000032:analysis; D000667:Ampicillin / Q000494:pharmacology; D000668:Ampicillin Resistance; D001426:Bacterial Proteins / Q000032:analysis*; D002352:Carrier Proteins / Q000032:analysis; D003773:Dental Plaque / Q000382:microbiology; D015180:Electrophoresis, Gel, Two-Dimensional; D016967:Fusobacterium nucleatum / Q000145:classification / Q000187:drug effects / Q000378:metabolism*; D006801:Humans; D008826:Microbial Sensitivity Tests; D008970:Molecular Weight; D010449:Peptide Mapping; D010751:Phosphopyruvate Hydratase / Q000032:analysis; D020543:Proteome / Q000032:analysis*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D001618:beta-Lactamases / Q000032:analysis
D018528:ATP-Binding Cassette Transporters; C090796:ATP-binding protein, bacteria; D001426:Bacterial Proteins; D002352:Carrier Proteins; D020543:Proteome; D000667:Ampicillin; D001618:beta-Lactamases; D010751:Phosphopyruvate Hydratase