Use of a Web-Based Dietary Assessment Tool (RiksmatenFlex) in Swedish Adolescents: Comparison and Validation Study.

BACKGROUND: A Web-based dietary assessment tool-RiksmatenFlex-was developed for the national dietary survey of adolescents in Sweden. OBJECTIVE: This study aimed to describe the Web-based method RiksmatenFlex and to test the validity of the reported dietary intake by comparing dietary intake with 24-hour dietary recalls (recall interviews), estimated energy expenditure, and biomarkers. METHODS: Adolescents aged 11-12, 14-15, and 17-18 years were recruited through schools. In total, 78 students had complete dietary information and were included in the study. Diet was reported a few weeks apart with either RiksmatenFlexDiet (the day before and a random later day) or recall interviews (face-to-face, a random day later by phone) in a cross-over, randomized design. At a school visit, weight and height were measured and blood samples were drawn for biomarker analyses. Students wore an accelerometer for 7 days for physical activity measurements. Dietary intake captured by both dietary methods was compared, and energy intake captured by both methods was compared with the accelerometer-estimated energy expenditure (EEest). Intake of whole grain wheat and rye and fruit and vegetables by both methods was compared with alkylresorcinol and carotenoid concentrations in plasma, respectively. RESULTS: The mean of the reported energy intake was 8.92 (SD 2.77) MJ by RiksmatenFlexDiet and 8.04 (SD 2.67) MJ by the recall interviews (P=.01). Intake of fruit and vegetables was 224 (169) g and 227 (150) g, and whole grain wheat and rye intake was 12.4 (SD 13.2) g and 12.0 (SD 13.1) g, respectively; the intakes of fruit and vegetables as well as whole grain wheat and rye did not differ between methods. Intraclass correlation coefficients ranged from 0.57 for protein and carbohydrates to 0.23 for vegetables. Energy intake by RiksmatenFlexDiet was overreported by 8% (P=.03) but not by the recall interviews (P=.53) compared with EEest. The Spearman correlation coefficient between reported energy intake and EEest was 0.34 (P=.008) for RiksmatenFlexDiet and 0.16 (P=.21) for the recall interviews. Spearman correlation coefficient between whole grain wheat and rye and plasma total alkylresorcinol homologs was 0.36 (P=.002) for RiksmatenFlexDiet and 0.29 (P=.02) for the recall interviews. Spearman correlations between intake of fruit and vegetables and plasma carotenoids were weak for both dietary tools. The strongest correlations were observed between fruit and vegetable intake and lutein/zeaxanthin for RiksmatenFlexDiet (0.46; P<.001) and for recall interviews (0.28; P=.02). CONCLUSIONS: RiksmatenFlexDiet provides information on energy, fruit, vegetables, and whole grain wheat and rye intake, which is comparable with intake obtained from recall interviews in Swedish adolescents. The results are promising for cost-effective dietary data collection in upcoming national dietary surveys and other studies in Sweden. Future research should focus on how, and if, new technological solutions could reduce dietary reporting biases.

Riksmaten Adolescents 2016-17: A national dietary survey in Sweden - design, methods, and participation.

BACKGROUND: Nationally representative information on food consumption data is essential to evaluate dietary habits, inform policy-making and nutritional guidelines, as well as forming a basis for risk assessment and identification of risk groups. OBJECTIVE: To describe the methods used in the Swedish national dietary survey of adolescents, Riksmaten Adolescents 2016-2017. DESIGN: Students in grades 5, 8, and 11 (mean ages 12, 15, and 18 years) were recruited in this school-based cross-sectional survey. A new, validated, web-based method was used to assess dietary intake. Information on physical activity, health, and socioeconomic background was collected through web questionnaires. Physical activity was also evaluated by accelerometers. Weight and height were measured in all participants, while blood and urine samples were collected in a subsample of 40% of the participants. RESULTS: A total of 3,477 (68%) respondents participated and 3,099 (60%) had complete dietary information. In the subsample, 1,305 (55%) respondents participated and 1,105 (46%) had complete dietary information. The participants were overall representative for the population with regard to socioeconomic background and school organization (public or independent). All types of municipalities were represented in the survey and overall, the geographic distribution corresponded to the underlying population. Some differences by school grade were observed. Sample weights were calculated for the total sample and the subsample. CONCLUSION: The Riksmaten Adolescents 2016-2017 provides valuable national data on diet, physical activity, and markers of exposure in age groups where data have been lacking. The data will provide a valuable basis for risk assessment, public health policy, and in-depth analyses.

Effects of dietary n-3 fatty acids on Toll-like receptor activation in primary leucocytes from Atlantic salmon (Salmo salar).

The shortage of the n-3 fatty acids eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) on the international markets has led to increasing substitution of fish oil by plant oils in Atlantic salmon (Salmo salar) feed and thereby reducing the EPA and DHA content in salmon. However, the minimum required levels of these fatty acids in fish diets for securing fish health are unknown. Fish were fed with 0, 1 or 2% EPA or DHA alone or in combination of both over a period, growing from 50 to 400 g. Primary head kidney leucocytes were isolated and stimulated with Toll-like receptor (TLR) ligands to determine if EPA and DHA deficiency can affect expression of important immune genes and eicosanoid production. Several genes related to viral immune response did not vary between groups. However, there was a tendency that the high-level EPA and DHA groups expressed lower levels of IL-1ß in non-stimulated leucocytes. These leucocytes were also more responsive to the TLR ligands, inducing higher expression levels of IL-1ß and Mx1 after stimulation. The levels of prostaglandin E2 and leukotriene B4 in serum and media from stimulated leucocytes were lower in both low and high EPA and DHA groups. In conclusion, leucocytes from low EPA and DHA groups seemed to be less responsive towards immunostimulants, like TLR ligands, indicating that low levels or absence of dietary EPA and DHA may have immunosuppressive effects.

Immune responses of a meningococcal A + W outer membrane vesicle (OMV) vaccine with and without aluminium hydroxide adjuvant in two different mouse strains.

Meningococci (Neisseria meningiditis) of serogroups A and W have caused large epidemics of meningitis in sub-Saharan Africa for decades, and affordable and multivalent vaccines, effective in all age groups, are needed. A bivalent serogroup A and W (A + W) meningococcal vaccine candidate consisting of deoxycholate-extracted outer membrane vesicles (OMV) from representative African disease isolates was previously found to be highly immunogenic in outbred mice when formulated with the adjuvant aluminium hydroxide (AH). OMV has been shown to have inherent adjuvant properties. In order to study the importance of AH and genetical differences between mice strains on immune responses, we compared the immunogenicity of the A + W OMV vaccine when formulated with or without AH in inbred C57BL/6J and BALB/cJ mice (Th1 and Th2 dominant strains, respectively). The immunogenicity of the vaccine was found to be comparable in the two mice strains despite their different immune profiles. Adsorption to AH increased anti-OMV IgG levels and serum bactericidal activity (SBA). The immune responses were increased by each dose for the adsorbed vaccine, but the third dose did not significantly improve the immunogenicity further. Thus, a vaccine formulation with the A and W OMV will likely benefit from including AH as adjuvant.

Protective effect of a recombinant VHSV-G vaccine using poly(I:C) loaded nanoparticles as an adjuvant in zebrafish (Danio rerio) infection model.

There is a constant need to increase the efficiency of vaccines in the aquaculture industry. Although several nano-based vaccine formulations have been reported, to the best of our knowledge so far only one of them have been implemented in the industry. Here we report on chitosan-poly(I:C) nanoparticles (NPs) that could be used as a non-specific adjuvant in antiviral vaccines in aquaculture. We have characterized the physical parameters of the NPs, studied the in vivo and in vitro bio-distribution of fluorescent NPs and verified NP uptake by zebrafish leucocytes. We used the zebrafish model to test the protective efficiency of the recombinant glycoprotein G (rgpG) of VHSV compared to inactivated whole virus (iV) against VHSV using NPs as an adjuvant in both formulations. In parallel we tested free poly(I:C) and rgpG (pICrgpG), and free chitosan and rgpG (CSrgpG) vaccine formulations. While the iV group (with NP adjuvant) provided the highest overall survival, all vaccine formulations with poly(I:C) provided a significant protection against VHSV; possibly through an early induction of an anti-viral state. Our results suggest that chitosan-poly(I:C) NPs are a promising adjuvant candidate for future vaccine formulations.

Stability of a Vesicular Stomatitis Virus-Vectored Ebola Vaccine.

The live attenuated vesicular stomatitis virus-vectored Ebola vaccine rVSV-ZEBOV is currently undergoing clinical trials in West Africa. The vaccine is to be stored at -70°C or less. Since maintaining the cold chain is challenging in rural areas, the rVSV-ZEBOV vaccine's short-term and long-term stability at different temperatures was examined. Different dilutions were tested since the optimal vaccine dosage had not yet been determined at the start of this experiment. The results demonstrate that the original vaccine formulation was stable for 1 week at 4°C and for 24 hours at 25°C. The stability of the vaccine was compromised by both high temperatures and dilution.

Activation of unfolded protein response pathway during infectious salmon anemia virus (ISAV) infection in vitro an in vivo.

Infectious salmon anemia virus (ISAV) is a salmon pathogen causing serious outbreaks in fish farms world-wide. There is currently no effective commercially available vaccine and there is a need for better understanding of host pathogen interactions with this virus. Various strains can cause both acute and persistent infections and therefore establish a balance with the host immune responses. We have studied host responses to this infection by analyzing the main branches of the unfolded protein response (UPR) in salmon cells in vitro and in tissues from infected fish to gain a better understanding of virus-host interactions. ISAV induce the main symptoms and signaling pathways of UPR (ATF6, PERK and IRE1) without inducing translational attenuation. This may be due to concomitant induction of an important negative feedback loop via the phosphatase regulator GADD34. The host cells can therefore respond with translation of cytokine and antiviral proteins to curb or control infection.

Structurally diverse genes encode Tlr2 in rainbow trout: The conserved receptor cannot be stimulated by classical ligands to activate NF-κB in vitro.

The mammalian toll-like receptor 2 (TLR2) is a dominant receptor for the recognition of Gram-positive bacteria. Its structure and functional properties were unknown in salmonid fish. In RT-PCR and RACE experiments, we obtained the full-length cDNA sequence encoding Tlr2 from rainbow trout (Oncorhynchus mykiss) as well as a copy of an unspliced nonsense message from a highly segmented gene. The primary structure of the encoded receptor complies with the domain structure and ligand-binding sites known from mammals and other fish species and sorts well into the evolutionary tree of teleostean Tlr2s. We retrieved a gene version encoding the receptor on a single exon (tlr2a) and also a partial sequence of a second gene variant being segmented into multiple exons (tlr2b). Surprisingly, the abundances of both transcript variants accounted only for ∼10% of all Tlr2-encoding transcripts in various tissues and cell types of healthy fish. This suggests the expression of several distinct tlr2 gene variants in rainbow trout. We expressed tlr2a in HEK-293 cells, but were unable to demonstrate its functionality through NF-κB activation. Neither synthetic lipopeptides known to stimulate mammalian TLR2 nor different bacterial challenges induced OmTLR2-mediated NF-κB activation, not in HEK-293 or in salmon CHSE-214 cells. Positive demonstration of TLR2-MYD88 interaction excluded that its functional impairment caused the failure of NF-κB activation. We discuss impaired heterodimerization with a necessary Tlr partner as one from among several alternatives to explain the dysfunction of Tlr2a in the interspecies reconstitution system of TLR signaling.

Use of Poly(I:C) Stabilized with Chitosan As a Vaccine-Adjuvant Against Viral Hemorrhagic Septicemia Virus Infection in Zebrafish.

There is an urgent need for more efficient viral vaccines in finfish aquaculture worldwide. Here, we report the use of poly(I:C) stabilized with chitosan as an adjuvant for development of better finfish vaccines. The adjuvant was co-injected with inactivated viral hemorrhagic septicemia virus (VHSV) (CSpIC+iV vaccine) in adult zebrafish and its efficiency in protection against VHSV infection was compared to a live, attenuated VHS virus vaccine (aV). Both free and stabilized poly(I:C) were strong inducers of an antiviral state, measured by transcriptional activation of the genes of viral sensors: toll-like receptors, interferons, and interferon-stimulated genes, such as MXa within 48 h after injection. Both the CSpIC+iV and the aV formulations provided a significant protection against VHSV-induced mortality. However, when plasma from survivors was tested for neutralizing antibodies in an in vitro protection assay, we could not demonstrate any protective effect. On the contrary, plasma from aV vaccinated fish enhanced cytopathic effects, indicating that antibody-dependent entry may play a role in this system. Our results show that poly(I:C) is a promising candidate as an adjuvant for fish vaccination against viral pathogens, and that the zebrafish is a promising model for aquaculture-relevant vaccination studies.

Effects of TLR agonists and viral infection on cytokine and TLR expression in Atlantic salmon (Salmo salar).

The development of efficient and cheap vaccines against several aquatic viruses is necessary for a sustainable fish farming industry. Toll-like receptor (TLR) ligands have already been used as good adjuvants in human vaccines. With more understanding of TLR expression, function, and ligand specificity in fish, more efficient adjuvants for fish viral vaccines can be developed. In this paper, we examine all known TLRs in Atlantic salmon (Salmo salar) and demonstrate that head kidney and spleen are the main organs expressing TLRs in salmon. We also show that adherent head kidney leucocytes from salmon are able to respond to many of the known agonists for human TLRs, and that viral infection can induce up-regulation of several TLRs. These findings substantiate these receptors' role in immune responses to pathogens in salmonids making their ligands attractive as vaccine adjuvant candidates.