RESUMO
A novel nano-formulation (NF) that sensitizes Acinetobacter baumannii (AB) to otherwise ineffective colistin is described in the present study. Infections due to multidrug resistant (MDR) AB represent a major therapeutic challenge, especially in situations of pre-existing colistin resistance (colR). Subsequently, boosting the effectiveness of colistin would be a better alternative tactic to treat AB infections rather than discovering a new class of antibiotics. We have previously demonstrated an NF comprising self-assembled guanidinium and ionic silver nanoparticles [AD-L@Ag(0)] to have anti-biofilm and bactericidal activity. We report NF AD-L@Ag(0) for the very first time for the potentiation of colistin in Gram-negative colistin-resistant bacteria. Our results implied that a combination of clinically relevant concentrations of colistin and AD-L@Ag(0) significantly decreased colistin-resistant AB bacterial growth and viability, which otherwise was elevated in the presence of only colistin. In this study, we have described various combinations of minimum inhibitory concentration (MIC) of colistin (MICcol, 1/2 MICcol, and 1/4 MICcol) and that of AD-L@Ag(0) [MICAD-L@Ag(0), 1/2 MICAD-L@Ag(0), and 1/4 MICAD-L@Ag(0)] and tested them against MDR AB culture. The results (in broth as well as in solid media) signified that AD-L@Ag(0) was able to potentiate the anti-microbial activity of colistin at sub-MIC concentrations. Furthermore, the viability and metabolic activity of bacterial cells were also measured by CTC fluorescence assay and ATP bioluminescence assay. The results of these assays were in perfect concordance with the scores of cultures (colony forming unit and culture turbidity). In addition, quantitative real-time PCR (qRT-PCR) was performed to unveil the expression of selected genes, DNAgyrA, DNAgyrB, and dac. These genes introduce negative supercoiling in the DNA, and hence are important for basic cellular processes. These genes, due to mutation, modified the Lipid A of bacteria, further resisting the uptake of colistin. Therefore, the expression of these genes was upregulated when AB was treated with only colistin, substantiating that AB is resistant to colistin, whereas the combinations of MICcol + MICAD-L@Ag(0) downregulated the expression of these genes, implying that the developed formulation can potentiate the efficiency of colistin. In conclusion, AD-L@Ag(0) can potentiate the proficiency of colistin, further enhancing colistin-mediated death of AB by putatively disrupting the outer membrane (OM) and facilitating bacterial death.
RESUMO
With recent advances in the field of diagnostics and theranostics, liposomal technology has secured a fortified position as a potential nanocarrier. Specifically, radiation/photo-sensitive liposomes containing photo-polymerizable cross-linking lipids are intriguing as they can impart the vesicles with highly interesting properties such as response to stimulus and improved shell stability. In this work, 1,2-bis(10,12-tricosadiynoyl)-sn-glycero-3-phosphoethanolamine (DTPE) is used as a photo-polymerizable lipid to form functional hybrid-liposomes as it can form intermolecular cross-linking through the diacetylenic groups. Hybrid-liposomes were formulated using mixtures of DTPE and saturated lipids of different chain lengths (dipalmitoylphosphatidylcholine (DPPC) and dimirystoilphosphatidylcholine (DMPC)) at different molar ratios. The physico-chemical characteristics of the liposomes has been studied before and after UV irradiation using a combination of techniques: DSC, QCM-D and solid-state NMR. The results signify the importance of a subtle modification in alkyl chain length on the phase behavior of the hybrid-liposomes and on the degree of crosslinking in the shell.