Davenport Emily K, Call Douglas R, Beyenal Haluk
Gene and Linda Voiland School of Chemical Engineering and Bioengineering, Washington State University, Pullman, Washington, USA.
Paul G. Allen School for Global Animal Health, Washington State University, Pullman, Washington, USA.
Antimicrob Agents Chemother. 2014 Aug;58(8):4755-61. doi: 10.1128/AAC.03071-14. Epub 2014 Jun 9.
We investigated biofilms of two pathogens, Acinetobacter baumannii and Staphylococcus aureus, to characterize mechanisms by which the extracellular polymeric substance (EPS) found in biofilms can protect bacteria against tobramycin exposure. To do so, it is critical to study EPS-antibiotic interactions in a homogeneous environment without mass transfer limitations. Consequently, we developed a method to grow biofilms, harvest EPS, and then augment planktonic cultures with isolated EPS and tobramycin. We demonstrated that planktonic cultures respond differently to being treated with different types of EPS (A. baumannii versus S. aureus) in the presence of tobramycin. By harvesting EPS from the biofilms, we found that A. baumannii EPS acts as a "universal protector" by inhibiting tobramycin activity against bacterial cells regardless of species; S. aureus EPS did not show any protective ability in cell cultures. Adding Mg(2+) or Ca(2+) reduced the protective effect of A. baumannii EPS. Finally, when we selectively digested the proteins or DNA of the EPS, we found that the protective ability did not change, suggesting that neither has a significant role in protection. To the best of our knowledge, this is the first study that demonstrates how EPS protects pathogens against antibiotics in a homogeneous system without mass transfer limitations. Our results suggest that EPS protects biofilm communities, in part, by adsorbing antibiotics near the surface. This may limit antibiotic diffusion to the bottom of the biofilms but is not likely to be the only mechanism of protection.
我们研究了两种病原体——鲍曼不动杆菌和金黄色葡萄球菌的生物膜，以确定生物膜中发现的细胞外聚合物（EPS）能够保护细菌免受妥布霉素作用的机制。为此，在没有传质限制的均匀环境中研究EPS与抗生素的相互作用至关重要。因此，我们开发了一种方法来培养生物膜、收获EPS，然后用分离出的EPS和妥布霉素增强浮游培养物。我们证明，在妥布霉素存在的情况下，浮游培养物对用不同类型的EPS（鲍曼不动杆菌与金黄色葡萄球菌）处理的反应不同。通过从生物膜中收获EPS，我们发现鲍曼不动杆菌EPS通过抑制妥布霉素对细菌细胞的活性而起到“通用保护剂”的作用，无论细菌种类如何；金黄色葡萄球菌EPS在细胞培养中未显示出任何保护能力。添加Mg(2+)或Ca(2+)会降低鲍曼不动杆菌EPS的保护作用。最后，当我们选择性地消化EPS中的蛋白质或DNA时，发现保护能力没有变化，这表明两者在保护作用中都没有显著作用。据我们所知，这是第一项证明EPS如何在没有传质限制的均匀系统中保护病原体免受抗生素作用的研究。我们的结果表明，EPS部分地通过在表面附近吸附抗生素来保护生物膜群落。这可能会限制抗生素扩散到生物膜底部，但不太可能是唯一的保护机制。
