دانلود رایگان مقاله تغییرات در ساختار پوشش سلول استافیلوکوکوس اورئوس و گسترش مقاومت کیتوزان

ABSTRACT

The bacterial cell envelope is believed to be a principal target for initiating the staphylocidal pathway of chitosan. The present study was therefore designed to investigate possible changes in cell surface phenotypes related to the in vitro chitosan resistance development in the laboratory strain S. aureus SG511-Berlin. Following a serial passage experiment, a stable chitosan-resistant variant (CRV) was identified, exhibiting >50-fold reduction in its sensitivity towards chitosan. Our analyses of the CRV identified phenotypic and genotypic features that readily distinguished it from its chitosan-susceptible parental strain, including: (i) a lower overall negative cell surface charge; (ii) cross-resistance to a number of antimicrobial agents; (iii) major alterations in cell envelope structure, cellular bioenergetics and metabolism (based on transcriptional profiling); and (iv) a repaired sensor histidine kinase GraS. Our data therefore suggest a close nexus between changes in cell envelope properties with the in vitro chitosan-resistant phenotype in S. aureus SG511-Berlin.

4. Conclusions

Although the precise mechanisms by which S. aureus SG511- Berlin develops resistance against the antimicrobial activities of chitosan remain incompletely defined, yet the current study provided valuable insights into the nature of the chitosan resistance displayed by CRV (summarized in Fig. 5). Both biochemical/phenotypic and genotypic analyses were complementary and confirmed the key association of changes in cell surface properties with the in vitro development of chitosan resistance in S. aureus SG511-Berlin. This involves a reduced overall negative charge of both the cell wall and cell membrane (resulting in reduced chitosan binding), as well as changes in the expression of genes involved in sugar metabolism. This is in fair agreement with several previous studies, which have shown an association between the activity of certain antimicrobials and cell envelope structures (Friedman et al., 2006;Jones et al., 2008; Mukhopadhyay et al., 2007; Muthaiyan et al., 2008; Peschel et al., 1999; Utaida et al., 2003;Xiong et al., 2005). Furthermore,the phenotype ofthe current CRV (increased L-PG production, increased relative surface positive charge, and resistance to cationic AMPs) suggests a “gain in mprF function”. However, the transcriptional analysis revealed no direct implication of the regulation of mprF in the reduced susceptibility of CRV to chitosan, indicating that the regulation of MprF occurs at a level other than the transcriptional one. Interestingly, most of the observed phenotypes are interrelated, and suggest a broad model of resistance that is closely associated with both cell wall and cell membrane structures. These proposed resistance mechanisms are not mutually exclusive; indeed we believe that combinations of these general types of resistance work in concert, ultimately contributing to the patterns of in vitro resistance that we observed. More importantly, the relatively quick development of stable resistance to chitosan, and the cross-resistance of the emerged isolate to other antimicrobials would warrant more caution in the indiscriminate use of chitosan as an additive to a number of pharmaceutical and biomedical formulations.