Macrolides are regarded as drugs of choice for treatment of human campylobacteriosis. The use of antimicrobials for this purpose as well as in food animal production could result in macrolide resistance in Campylobacter species. Campylobacter isolates exhibit two different phenotypes with regard to erythromycin resistance: high-level resistance (HLR) and low-level resistance (LLR). Thirty-six food/animal and human isolates of Campylobacter jejuni and C. coli were examined for their mechanisms of resistance to erythromycin. The data presented here confirm the previous findings that the A2075G mutation in the 23S rRNA gene is the most frequently reported mechanism of high-level erythromycin resistance in Campylobacter isolates. The efflux pump inhibitor PAbetaN increased susceptibility to erythromycin for at least 16-32-fold in all examined HLR isolates, suggesting that the efflux mechanism acts in synergy with the 23S rRNA mutation to confer high-level erythromycin resistance. This was also confirmed in the isolates with sequence variation in the efflux pump cmeB gene. Additionally, the PAbetaN restored the susceptibility of LLR strains to the level of minimal inhibitory concentrations (MICs) of the susceptible strains and also reduced the MICs of the susceptible C. jejuni and C. coli isolates. The data suggest that active efflux contributes to the intrinsic resistance to erythromycin in Campylobacter and also contribute to high-level resistance.