The strains still resistant to metronidazole even after treatment with polysorbate 80 could also have undergone a mutation of the reduction systems, i.e. it had a double mechanism of resistance. The increased susceptibility to clarithromycin used in combination with polysorbate 80 could also be due to an augmented permeability of membranes exerted by the detergent. The main constituent of the outer membrane in Gram-negative bacteria is lipopolysaccharide

(LPS); it coats the cell surface and works to exclude OSI-906 datasheet large hydrophobic compounds, such as antibiotics, from invading the cell. LPS has a significant role in membrane transport: the lipid compositions of LPS and the associated proteins have a strong impact on the sensitivity of bacteria to many types of antibiotics [34]. Unlike small hydrophilic antibiotics, large lipophilic agents, such FK228 manufacturer as macrolides, have difficulty in diffusing through the LPS. Previous studies indicate that membrane permeabilizers, such as Tris/EDTA, polymyxin B

etc., have the ability to increase the levels of antibiotic inflow [34] and consequently the sensitivity of Gram-negative bacteria to hydrophobic antibiotics, including macrolides [35, 36]. In this study, two strains were highly resistant to clarithromycin, with MBCs of 320 see more μg/mL and 2500 μg/mL. In the presence of polysorbate 80, clarithromycin’s MBCs decreased by 16 times and 1000 times, respectively, i.e. to 20 μg/mL and 2.5 μg/mL, which still are in the range of resistant values (threshold = 1 μg/mL). In these

cases, we hypothesize the concomitance of two mechanisms of resistance. In a large number of bacterial species, in fact, the existence of drug-resistant strains is due to modifications in the lipid or protein composition of the outer membrane, which work in synergy with other resistance mechanisms [34]. Point mutations in 23S rRNA normally account for the development of resistance to clarithromycin in H. pylori and reduce the chances of eradication when the classical triple therapy is employed [37]. It is likely that in our strains the presence of an efflux apparatus cooperates with putative 23S rRNA mutations to make these two strains highly resistant to clarithromycin [38]. Polysorbate 80 conceivably increased their sensitivity by destroying the outer membrane; strains, however, were still resistant because of the existence of another putative mechanism, such as ribosome mutation. A plausible explanation for the observation that the association of polysorbate 80 with amoxicillin, levofloxacin and tetracycline was not synergistic may consist in the sizes and hydrophilic nature of antimicrobials.