Activity of dalbavancin, alone and in combination with rifampicin, against meticillin-resistant Staphylococcus aureus in a foreign-body infection model.

The activity of dalbavancin, a representative of the lipoglycopeptide antibiotics, alone and in combination with rifampicin, was investigated against meticillin-resistant Staphylococcus aureus (MRSA) in a foreign-body infection model in guinea pigs. The MIC, MBC and time-kill profile of dalbavancin were determined for MRSA ATCC 43300 in the logarithmic (MBC(log)) and stationary (MBC(stat)) growth phases. The pharmacokinetic profile of dalbavancin was determined in sterile cage fluid in guinea pigs. The activity of intraperitoneal dalbavancin (40, 60 or 80 mg/kg as a single dose), rifampicin (12.5 mg/kg/12 h for 4 days) and their combination was assessed against planktonic and biofilm MRSA. The MIC of dalbavancin was 0.078 mg/L; MBC(log) and MBC(stat) were both >128× MIC. In time-kill studies, bacterial reduction of 3log(10)CFU/mL was achieved after 48 h at ≥32× MIC (logarithmic growth) and at ≥1× MIC (stationary growth). Dalbavancin was neither synergistic nor antagonistic with rifampicin, and prevented the emergence of rifampicin resistance in vitro. The half-life of dalbavancin in cage fluid was 35.8-45.4 h and the concentration remained above the MIC of MRSA during 7 days after a single dose. Dalbavancin reduced planktonic MRSA in cage fluid at high dose (60 mg/kg and 80 mg/kg) but failed to eradicate biofilm MRSA from cages. In combination with rifampicin, dalbavancin at 80 mg/kg cured 36% of infected cages, and emergence of rifampicin resistance was completely prevented. Dalbavancin at 80 mg/kg and in combination with rifampicin eradicated approximately one-third of cage-associated MRSA infections and prevented emergence of rifampicin resistance.

Size fractionation of microscopic protein aggregates using a preparative fluorescence-activated cell sorter.

Protein aggregation, which takes place both in vivo and in vitro, is an important degradative pathway for all proteins. Protein aggregates have distinct physicochemical and biological properties that are important to study and characterize from the perspective of both fundamental and applied sciences. The size of protein aggregates varies across a huge range, spanning several orders of magnitude. Currently, protein aggregates larger than hundreds of nanometers in diameter are impossible to physically fractionate. Here, we present a new method to fractionate microscopic proteinaceous particles using preparative fluorescence-activated cell sorting technology.

A novel secondary acyl chain in the lipopolysaccharide of Bordetella pertussis required for efficient infection of human macrophages.

Lipopolysaccharide is one of the major constituents of the Gram-negative bacterial outer membrane and is a potent stimulator of the host innate immune response. The biosynthesis of the lipid A moiety of lipopolysaccharide is a complex process in which multiple gene products are involved. Two late lipid A acyl transferases, LpxL and LpxM, were first identified in Escherichia coli and shown to be responsible for the addition of secondary acyl chains to the 2' and 3' positions of lipid A, respectively. Here, we describe the identification of two lpxL homologues in the genome of Bordetella pertussis. We show that one of them, LpxL2, is responsible for the addition of the secondary myristate group that is normally present at the 2' position of B. pertussis lipid A, whereas the other one, LpxL1, mediates the addition of a previously unrecognized secondary 2-hydroxy laurate at the 2 position. Increased expression of lpxL1 results in the appearance of a hexa-acylated lipopolysaccharide form with strongly increased endotoxic activity. In addition, we show that an lpxL1-deficient mutant of B. pertussis displays a defect in the infection of human macrophages.