Influence of Miscibility of Protein-Sugar Lyophilizates on Their Storage Stability

For sugars to act as successful stabilizers of proteins during lyophilization and subsequent storage, they need to have several characteristics. One of them is that they need to be able to form interactions with the protein and for that miscibility is essential. To evaluate the influence of protein-sugar miscibility on protein storage stability, model protein IgG was lyophilized in the presence of various sugars of different molecular weight. By comparing solid-state nuclear magnetic resonance spectroscopy relaxation times of both protein and sugar on two different timescales, i.e., H-1 T-1 and H-1 T-1 rho, miscibility of the two components was established on a 2-5- and a 20-50-nm length scale, respectively, and related to protein storage stability. Smaller sugars showed better miscibility with IgG, and the tendency of IgG to aggregate during storage was lower for smaller sugars. The largest sugar performed worst and was phase separated on both length scales. Additionally, shorter protein H-1 T-1 relaxation times correlated with higher aggregation rates during storage. The enzyme-linked immunosorbent assay (ELISA) assay showed overlapping effects of aggregation and chemical degradation and did not correspond as well with the miscibility. Because of the small scale at which miscibility was determined (2-5 nm) and the size of the protein domains (similar to 2.5 x 2.5 x 5 nm), the miscibility data give an indirect measure of interaction between protein and sugar. This reduced interaction could be the result of steric hindrance, providing a possible explanation as to why smaller sugars show better miscibility and storage stability with the protein

Genetic and Molecular Predictors of High Vancomycin MIC in Staphylococcus aureus Bacteremia Isolates

An elevated vancomycin MIC is associated with poor outcomes in Staphylococcus aureus bacteremia (SAB) and is reported in patients with methicillin-susceptible S. aureus (MSSA) bacteremia in the absence of vancomycin treatment. Here, using DNA microarray and phenotype analysis, we investigated the genetic predictors and accessory gene regulator (agr) function and their relationship with elevated vancomycin MIC using blood culture isolates from a multicenter binational cohort of patients with SAB. Specific clonal complexes were associated with elevated (clonal complex 8 [CC8] [P < 0.001]) or low (CC22 [P < 0.001], CC88 [P < 0.001], and CC188 [P = 0.002]) vancomycin MIC. agr dysfunction (P = 0.014) or agr genotype II (P = 0.043) were also associated with an elevated vancomycin MIC. Specific resistance and virulence genes were also linked to an elevated vancomycin MIC, including blaZ (P = 0.002), sea (P < 0.001), clfA (P < 0.001), splA (P = 0.001), and the arginine catabolic mobile element (ACME) locus (P = 0.02). These data suggest that inherent organism characteristics may explain the link between elevated vancomycin MICs and poor outcomes in patients with SAB, regardless of the antibiotic treatment received. A consideration of clonal specificity should be included in future research when attempting to ascertain treatment effects or clinical outcomes