Development and Validation of the Vending Evaluation for Nutrient-Density (VEND)ing Audit.

Background: This paper describes the development and validation of the Vending Evaluation for Nutrient-Density (VEND)ing audit to comprehensively evaluate vended products based upon healthfulness, price and promotion, and machine accessibility. Methods: A novel vending nutrient-density score was created to determine the healthfulness of vended snack/beverage products. Field tested in United States colleges, VENDing audit (∑nutrient-density + 10 × % healthy products) and Support sub-scores (price + promotion + accessibility) were calculated for snack/beverage machines. Higher scores indicate more healthful vending options and supports for choosing healthfully. Nutrition Environment Measures Survey-Vending (NEMS-V) was used to validate the nutrient-density score for a sub-sample of machines. Sensitivity and specificity were computed by comparing the number of healthy snacks/beverages determined by NEMS-V and the VENDing nutrient-density scores. Results: Researchers conducted the VENDing audit on 228 snack/beverage vending machines at 9 universities within the United States and used both VENDing and NEMS-V on 33 snack and 52 beverage vending machines. Mean VENDing audit scores were 4.5 ± 2.0 (2.6, 3.4) and 2.6 ± 2.0 (0, 12) for snack/beverage machines, respectively. The number of products considered healthy assessed with both the VENDing nutrient-density scores and the NEMS-V were positively correlated for beverages (r = 0.687, p < 0.001) and snacks (r = 0.366, p < 0.05). The sensitivity was excellent for beverages (0.83) and moderate for snacks (0.69); while the specificity was moderate for both beverages (0.66) and snacks (0.50). Conclusions: The VENDing audit uses unique, valid, and reliable nutrient-density scoring to evaluate snacks/beverages along a continuum of healthful criteria and comprehensively evaluates the full vending environment.

Chloroplast-derived vaccine antigens confer dual immunity against cholera and malaria by oral or injectable delivery.

Cholera and malaria are major diseases causing high mortality. The only licensed cholera vaccine is expensive; immunity is lost in children within 3 years and adults are not fully protected. No vaccine is yet available for malaria. Therefore, in this study, the cholera toxin-B subunit (CTB) of Vibrio cholerae fused to malarial vaccine antigens apical membrane antigen-1 (AMA1) and merozoite surface protein-1 (MSP1) was expressed in lettuce and tobacco chloroplasts. Southern blot analysis confirmed homoplasmy and stable integration of transgenes. CTB-AMA1 and CTB-MSP1 fusion proteins accumulated up to 13.17% and 10.11% (total soluble protein, TSP) in tobacco and up to 7.3% and 6.1% (TSP) in lettuce, respectively. Nine groups of mice (n = 10/group) were immunized subcutaneously (SQV) or orally (ORV) with purified antigens or transplastomic tobacco leaves. Significant levels of antigen-specific antibody titres of immunized mice completely inhibited proliferation of the malarial parasite and cross-reacted with the native parasite proteins in immunoblots and immunofluorescence studies. Protection against cholera toxin challenge in both ORV (100%) and SQV (89%) mice correlated with CTB-specific titres of intestinal, serum IgA and IgG1 in ORV and only IgG1 in SQV mice, but no other immunoglobulin. Increasing numbers of interleukin-10(+) T cell but not Foxp3(+) regulatory T cells, suppression of interferon-gamma and absence of interleukin-17 were observed in protected mice, suggesting that immunity is conferred via the Tr1/Th2 immune response. Dual immunity against two major infectious diseases provided by chloroplast-derived vaccine antigens for long-term (>300 days, 50% of mouse life span) offers a realistic platform for low cost vaccines and insight into mucosal and systemic immunity.

Characterization of a PRL protein tyrosine phosphatase from Plasmodium falciparum.

Isoprenylated proteins have important functions in cell growth and differentiation of eukaryotic cells. Inhibitors of protein prenylation in malaria have recently shown strong promise as effective antimalarials. In studying protein prenylation in the malaria protozoan parasite Plasmodium falciparum, we have shown earlier that the incubation of P. falciparum cells with (3)H-prenol precursors resulted in various size classes of labeled proteins. To understand the physiological function of prenylated proteins of malaria parasites, that are targets of prenyltransferase inhibitors, we searched the PlasmoDB database for proteins containing the C-terminus prenylation motif. We have identified, among other potentially prenylated proteins, an orthologue of a PRL (protein of regenerating liver) subgroup protein tyrosine phosphatases, termed PfPRL. Here, we show that PfPRL is expressed in the parasite's intraerythrocytic stages, where it partially associates with endoplasmic reticulum and within a subcompartment of the food vacuole. Additionally, PfPRL targeting parallels that of apical membrane antigen-1 in developing merozoites. Recombinant PfPRL shows phosphatase activity that is preferentially inhibited by a tyrosine phosphatase inhibitor suggesting that PfPRL functions as a tyrosine phosphatase. Recombinant PfPRL can also be farnesylated in vitro. Inhibition of malarial farnesyltransferase activity can be achieved with the heptapetide RKCHFM, which corresponds to the C-terminus of PfPRL. This study provides the first evidence for expression of enzymatically active PRL-related protein tyrosine phosphatases in malarial parasites, and demonstrates the potential of peptides derived from Plasmodium prenylated proteins as malarial farnesyltransferase inhibitors.