<i>Campylobacter</i> is the most common bacterial agent causing human gastroenteritis and is mainly transmitted through foods. <i>Campylobacter</i> is a zoonotic agent, and commonly colonizes poultry and other meat animals. Whereas erythromycin resistance in <i>Campylobacter jejuni</i> rarely occurs, erythromycin resistance in <i>Campylobacter coli</i> from meat animals is frequently encountered, and could represent a substantial barrier to antibiotic treatment of human infections. Erythromycin resistance in <i>C. coli</i> has been associated with a point mutation (A2075G) in the 23S rRNA gene, acting synergistically with the CmeABC efflux pump. However, the mechanisms responsible for possible dissemination of erythromycin resistance in <i>C. coli</i> remain poorly understood. In this study we investigated transformation-mediated acquisition of erythromycin resistance by genotypically diverse <i>C. coli</i> strains from turkeys and swine, with total genomic DNA from erythromycin-resistant <i>C. coli</i> used as donor. In addition, we studied the effects of environmental factors and species (<i>C. coli</i> vs. <i>C. jejuni</i>), and fitness costs of erythromycin resistance in transformants. Overall, transformation to erythromycin resistance was significantly more frequent in <i>C. coli</i> from turkeys than in swine-derived strains (P<0.01) with frequency of transformation 10<sup>-4</sup> to 10<sup>-6</sup> in turkey-derived strains, but 10<sup>-7</sup> or less in <i>C. coli</i> from swine. Transformants harbored the point mutation, A2075G in the 23S rRNA gene. Erythromycin resistance was stable in transformants following serial transfers, and most transformants had high MIC values (>256µg/ml), as did the <i>C. coli</i> donor strains. In contrast to results obtained with transformation, spontaneous mutants had relatively low erythromycin MIC (32-64µg/ml) and lacked the A2075G mutation. Temperature profoundly affected frequency of transformation to erythromycin resistance in <i>C. coli</i> and transformation frequency at 42°C was significantly higher than at 25°C, 32°C and even 37°C. However, transformation to nalidixic acid resistance was not significantly affected by temperature. No significant difference in transformation frequency was detected between microaerobic (5-10% CO<sub>2</sub>) and aerobic conditions. Starvation conditions did not affect transformation frequency to nalidixic acid resistance. Increasing incubation time from 3-4h to 15-17h significantly increased transformation frequency to erythromycin resistance (P<0.05). Transformation of <i>C. jejuni</i> using genomic DNA from erythromycin resistant <i>C. coli</i> revealed that transformation frequency of <i>C. jejuni</i> to erythromycin resistance was lower than <i>C. coli</i>, suggesting that erythromycin resistance in <i>C. coli</i> may not be disseminated via transformation in <i>C. jejuni</i> as frequently as in <i>C. coli</i>. Transformants derived from <i>C. jejuni</i>, however, had high erythromycin MIC values (>256µg/ml) and harbored the A2075G transition, similarly to <i>C. coli</i> transformants. When grown separately at 42°C, an erythromycin-resistant transformant derived from <i>C. coli</i> strain 961 had a similar growth rate as its erythromycin-sensitive parental strain, whereas an erythromycin-resistant transformant derived from <i>C. jejuni</i> strain SC49 had a significantly longer generation time compared to its parental strain. In competitive growth studies, however, the <i>C. coli</i> transformant was at competitive disadvantage in relation to its parental strain in stationary phase, whereas the <i>C. jejuni</i> transformant was at a slight fitness advantage after 14days. Furthermore, in the mixed culture the generation time of the <i>C. jejuni</i> transformant was not significantly different from that of the parental strain. In conclusion, natural transformation has the potential to contribute to dissemination of high-level resistance to erythromycin among <i>C. coli</i> strains colonizing meat animals and temperature can greatly affect transformation to erythromycin resistance, but not to nalidixic acid resistance. These findings suggest that ecological attributes may play an important role and exert differential impact on the potential of the organism to acquire antimicrobial resistance determinants via natural transformation. However, further study is necessary to characterize the fitness of erythromycin resistant transformants in <i>Campylobacter</i> and identify possible mechanisms underlying the relatively low frequency of erythromycin resistance in <i>C. jejuni</i>.
| Identifer | oai:union.ndltd.org:NCSU/oai:NCSU:etd-10272005-170447 |
| Date | 09 November 2005 |
| Creators | Kim, Joo-Sung |
| Contributors | Sophia Kathariou, Fred Breidt, Donna K. Carver, Paul Orndorff |
| Publisher | NCSU |
| Source Sets | North Carolina State University |
| Language | English |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | http://www.lib.ncsu.edu/theses/available/etd-10272005-170447/ |
| Rights | unrestricted, I hereby certify that, if appropriate, I have obtained and attached hereto a written permission statement from the owner(s) of each third party copyrighted matter to be included in my thesis, dissertation, or project report, allowing distribution as specified below. I certify that the version I submitted is the same as that approved by my advisory committee. I hereby grant to NC State University or its agents the non-exclusive license to archive and make accessible, under the conditions specified below, my thesis, dissertation, or project report in whole or in part in all forms of media, now or hereafter known. I retain all other ownership rights to the copyright of the thesis, dissertation or project report. I also retain the right to use in future works (such as articles or books) all or part of this thesis, dissertation, or project report. |
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