The self-assembly of a cyclic lipopeptides mixture secreted by a B. megaterium strain and its implications on activity against a sensitive Bacillus species.

Cyclic lipopeptides are produced by a soil Bacillus megaterium strain and several other Bacillus species. In this work, they are detected both in the Bacillus intact cells and the cells culture medium by MALDI-TOF mass spectrometry. The cyclic lipopeptides self-assemble in water media producing negatively charged and large aggregates (300-800 nm of mean hydrodynamic radius) as evaluated by dynamic light scattering and zeta-potential analysis. The aggregate size depends on pH and ionic strength. However, it is not affected by changes in the osmolarity of the outer medium suggesting the absence of an internal aqueous compartment despite the occurrence of low molecular weight phospholipids in their composition as determined from inorganic phosphorus analysis. The activity against a sensitive Bacillus cereus strain was evaluated from inhibition halos and B. cereus lysis. Essential features determining the antibiotic activity on susceptible Bacillus cereus cells are the preserved cyclic moiety conferring cyclic lipopeptides resistance to proteases and the medium pH. The aggregates are inactive per se at the pH of the culture medium which is around 6 or below. The knock out of the sensitive cells only takes place when the aggregates are disassembled due to a high negative charge at pH above 6.

Silica-based cationic bilayers as immunoadjuvants.

BACKGROUND: Silica particles cationized by dioctadecyldimethylammonium bromide (DODAB) bilayer were previously described. This work shows the efficiency of these particulates for antigen adsorption and presentation to the immune system and proves the concept that silica-based cationic bilayers exhibit better performance than alum regarding colloid stability and cellular immune responses for vaccine design. RESULTS: Firstly, the silica/DODAB assembly was characterized at 1 mM NaCl, pH 6.3 or 5 mM Tris.HCl, pH 7.4 and 0.1 mg/ml silica over a range of DODAB concentrations (0.001-1 mM) by means of dynamic light scattering for particle sizing and zeta-potential analysis. 0.05 mM DODAB is enough to produce cationic bilayer-covered particles with good colloid stability. Secondly, conditions for maximal adsorption of bovine serum albumin (BSA) or a recombinant, heat-shock protein from Mycobacterium leprae (18 kDa-hsp) onto DODAB-covered or onto bare silica were determined. At maximal antigen adsorption, cellular immune responses in vivo from delayed-type hypersensitivity reactions determined by foot-pad swelling tests (DTH) and cytokines analysis evidenced the superior performance of the silica/DODAB adjuvant as compared to alum or antigens alone whereas humoral response from IgG in serum was equal to the one elicited by alum as adjuvant. CONCLUSION: Cationized silica is a biocompatible, inexpensive, easily prepared and possibly general immunoadjuvant for antigen presentation which displays higher colloid stability than alum, better performance regarding cellular immune responses and employs very low, micromolar doses of cationic and toxic synthetic lipid.

In vivo activity of a novel amphotericin B formulation with synthetic cationic bilayer fragments.

Solubilization of amphotericin B (AMB) by dioctadecyldimethylammonium bromide (DODAB) bilayer fragments inspired this evaluation of its in vivo activity from survival and tissue burden experiments against systemic candidiasis in a mouse model. AMB (< or =0.1 g/L) was simply added to a DODAB powder dispersion in water (10 g/L) previously prepared by sonication in the absence of organic solvents. The AMB aggregation state was evaluated from UV-visible light absorption and dynamic light scattering for aggregate sizing. AMB was stabilized by the DODAB bilayer fragments in its monomeric form, mixing of AMB and DODAB dispersion in pure water causing disappearance of large water-insoluble drug aggregates. From survival experiments, both the bilayer, DODAB/AMB, and the traditional deoxycholate/AMB formulation (DOC/AMB) had identical effect when given by the same route at the same dose of 0.4 mg/kg/day given intraperitoneally for 10 days. From spleen and kidneys tissue burden experiments, similar efficacy of both preparations in reducing tissue cfu counts was obtained. In summary, DODAB/AMB was as effective as DOC/AMB for treating systemic candidiasis in a mouse model.