Robust estimation of bacterial cell count from optical density

Optical density (OD) is widely used to estimate the density of cells in liquid culture, but cannot be compared between instruments without a standardized calibration protocol and is challenging to relate to actual cell count. We address this with an interlaboratory study comparing three simple, low-cost, and highly accessible OD calibration protocols across 244 laboratories, applied to eight strains of constitutive GFP-expressing E. coli. Based on our results, we recommend calibrating OD to estimated cell count using serial dilution of silica microspheres, which produces highly precise calibration (95.5% of residuals <1.2-fold), is easily assessed for quality control, also assesses instrument effective linear range, and can be combined with fluorescence calibration to obtain units of Molecules of Equivalent Fluorescein (MEFL) per cell, allowing direct comparison and data fusion with flow cytometry measurements: in our study, fluorescence per cell measurements showed only a 1.07-fold mean difference between plate reader and flow cytometry data

Safety and Immunogenicity of a Novel Recombinant Subunit Respiratory Syncytial Virus Vaccine (BBG2Na) in Healthy Young Adults

A novel recombinant respiratory syncytial virus (RSV) subunit vaccine, designated BBG2Na, was administered to 108 healthy adults randomly assigned to receive 10, 100, or 300 μg of BBG2Na in aluminum phosphate or saline placebo. Each subject received 1, 2, or 3 intramuscular injections of the assigned dose at monthly intervals. Local and systemic reactions were mild, and no evidence of harmful properties of BBG2Na was reported. The highest ELISA and virus-neutralizing (VN) antibody responses were evident in the 100- and 300-μg groups; second or third injections provided no significant boosts against RSV-derived antigens. BBG2Na induced ≥2-fold and ≥4-fold increases in G2Na-specific ELISA units in up to 100% and 57% of subjects, respectively; corresponding RSV-A-specific responses were 89% and 67%. Furthermore, up to 71% of subjects had ≥2-fold VN titer increases. Antibody responses to 2 murine lung protective epitopes were also highly boosted after vaccination. Therefore, BBG2Na is safe, well tolerated, and highly immunogenic in RSV-seropositive adults

Effect-directed analysis and chemical identification of agonists of peroxisome proliferator-activated receptors in white button mushroom

Obesity has a serious effect on human health. It relates to metabolic syndrome, including the associated disorders such as type 2 diabetes, heart disease, stroke and hyperemia. The peroxisome proliferator-activated receptors (PPARs) are important receptors to control fat metabolism in the human body. Because of the safety concerns of synthetic drugs targeting PPARs, ligands from natural sources have drawn interest. Earlier, we have found high PPAR activities in extracts from Agaricus bisporus (white button mushroom, WBM). WBM contains a wide range of candidate compounds which could be agonists of PPARs. To identify which compounds are responsible for PPAR activation by WBM extracts, we used fractionation coupled to effect-directed analysis with reporter gene assays specific for all three PPARs for purification and LC/MS-TOF and NMR for compound identification in purified active fractions. Surprisingly, we identified the relatively common dietary fatty acid, linoleic acid, as the main ligand of PPARs in WBM. Possibly, the relatively low levels of linoleic acid in WBM are sufficient and instrumental in inducing its anti-obesogenic effects, avoiding high energy intake and negative health effects associated with high levels of linoleic acid consumption. However, it could not be excluded that a minor relatively potent compound contributes towards PPAR activation, while the anti-obesity effects of WBM may be further enhanced by receptor expression modulating compounds or compounds with completely PPAR unrelated modes of action