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Improved understanding of lipoproteins, particles that transport lipids throughout the circulation, is vital to developing new treatments for the dyslipidemias associated with metabolic syndrome. Apolipoproteins are a key component of lipoproteins. Apolipoproteins are proteins that structure lipoproteins and regulate lipid metabolism through control of cellular lipid exchange. Constraints of cell culture and mouse models mean that there is a need for a complementary model that can replicate the complex in vivo milieu that regulates apolipoprotein and lipoprotein biology. Here, we further establish the utility of the genetically tractable and optically clear larval zebrafish as a model of apolipoprotein biology. Gene ancestry analyses were implemented to determine the closest human orthologs of the zebrafish apolipoprotein A-I (apoA-I), apoB, apoE and apoA-IV genes and therefore ensure that they have been correctly named. Their expression patterns throughout development were also analyzed, by whole-mount mRNA in situ hybridization (ISH). The ISH results emphasized the importance of apolipoproteins in transporting yolk and dietary lipids: mRNA expression of all apolipoproteins was observed in the yolk syncytial layer, and intestinal and liver expression was observed from 4-6 days post-fertilization (dpf). Furthermore, real-time PCR confirmed that transcription of three of the four zebrafish apoA-IV genes was increased 4 hours after the onset of a 1-hour high-fat feed. Therefore, we tested the hypothesis that zebrafish ApoA-IV performs a conserved role to that in rat in the regulation of food intake by transiently overexpressing ApoA-IVb.1 in transgenic larvae and quantifying ingestion of co-fed fluorescently labeled fatty acid during a high-fat meal as an indicator of food intake. Indeed, ApoA-IVb.1 overexpression decreased food intake by approximately one-third. This study comprehensively describes the expression and function of eleven zebrafish apolipoproteins and serves as a springboard for future investigations to elucidate their roles in development and disease in the larval zebrafish model.

OBJECTIVE To determine the effect of mandatory and nonmandatory influenza vaccination policies on vaccination rates and symptomatic absenteeism among healthcare personnel (HCP). DESIGN Retrospective observational cohort study. SETTING This study took place at 3 university medical centers with mandatory influenza vaccination policies and 4 Veterans Affairs (VA) healthcare systems with nonmandatory influenza vaccination policies. PARTICIPANTS The study included 2,304 outpatient HCP at mandatory vaccination sites and 1,759 outpatient HCP at nonmandatory vaccination sites. METHODS To determine the incidence and duration of absenteeism in outpatient settings, HCP participating in the Respiratory Protection Effectiveness Clinical Trial at both mandatory and nonmandatory vaccination sites over 3 viral respiratory illness (VRI) seasons (2012-2015) reported their influenza vaccination status and symptomatic days absent from work weekly throughout a 12-week period during the peak VRI season each year. The adjusted effects of vaccination and other modulating factors on absenteeism rates were estimated using multivariable regression models. RESULTS The proportion of participants who received influenza vaccination was lower each year at nonmandatory than at mandatory vaccination sites (odds ratio [OR], 0.09; 95% confidence interval [CI], 0.07-0.11). Among HCP who reported at least 1 sick day, vaccinated HCP had lower symptomatic days absent compared to unvaccinated HCP (OR for 2012-2013 and 2013-2014, 0.82; 95% CI, 0.72-0.93; OR for 2014-2015, 0.81; 95% CI, 0.69-0.95). CONCLUSIONS These data suggest that mandatory HCP influenza vaccination policies increase influenza vaccination rates and that HCP symptomatic absenteeism diminishes as rates of influenza vaccination increase. These findings should be considered in formulating HCP influenza vaccination policies. Infect Control Hosp Epidemiol 2018;39:452-461.

Many animals use carotenoid pigments derived from their diet for coloration and immunity. The carotenoid trade‐off hypothesis predicts that, under conditions of carotenoid scarcity, individuals may be forced to allocate limited carotenoids to either coloration or immunity. In polychromatic species, the pattern of allocation may differ among individuals. We tested the carotenoid trade‐off hypothesis in the Midas cichlid,Amphilophus citrinellus, a species with two ontogenetic color morphs, barred and gold, the latter of which is the result of carotenoid expression. We performed a diet‐supplementation experiment in which cichlids of both color morphs were assigned to one of two diet treatments that differed only in carotenoid content (β‐carotene, lutein, and zeaxanthin). We measured integument color using spectrometry, quantified carotenoid concentrations in tissue and plasma, and assessed innate immunity using lysozyme activity and alternative complement pathway assays. In both color morphs, dietary carotenoid supplementation elevated plasma carotenoid circulation but failed to affect skin coloration. Consistent with observable differences in integument coloration, we found that gold fish sequestered more carotenoids in skin tissue than barred fish, but barred fish had higher concentrations of carotenoids in plasma than gold fish. Neither measure of innate immunity differed between gold and barred fish, or as a function of dietary carotenoid supplementation. Lysozyme activity, but not complement activity, was strongly affected by body condition. Our data show that a diet low in carotenoids is sufficient to maintain both coloration and innate immunity in Midas cichlids. Our data also suggest that the developmental transition from the barred to gold morph is not accompanied by a decrease in innate immunity in this species.

Carotenoid pigments have myriad functions in fish, including coloration and immunity. The “carotenoid trade-off hypothesis” posits that dietary limitation of carotenoids imposes constraints on animals to allocate to one function at the expense of another. This hypothesis rarely has been tested in fish. We quantified tissue carotenoids in breeding and non-breeding female convict cichlids in Lake Xiloá, Nicaragua. This species is reverse sexually dichromatic such that females possess carotenoid-based coloration that males lack. We also collected algae samples near nest sites to assess carotenoid availability, recorded water depth, and examined cichlids’ behavioral interactions with pair mates, conspecifics, heterospecific competitors, and predators. Each of these, we predicted, would mediate potential carotenoid trade-offs. We found that non-breeding females had significantly higher levels of carotenoids in their integument, liver, and gonads compared to breeding fish. We found that algae and total carotenoids declined with depth across our study transects at 9, 11, 13, and 15 m, but the concentration of carotenoids (ng carotenoid g−1 algae, or algal quality) did not vary with depth. Furthermore, relationships among carotenoid concentrations of the three tissue types did not vary with depth, and female color status (orange or not) was not affected by behavioral interactions with other community members, reproductive status, or water depth. Our results support previous studies showing that carotenoid pigmentation may serve a signal function that facilitates the establishment of non-breeding females within the breeding population. Our study also uncovered no evidence indicating that carotenoids are limiting in the diet of breeding female convict cichlids.

Carotenoid‐ornamented animals may transfer dietary carotenoid pigments to offspring as well as to their ornament and other competing physiological demands. If carotenoid supplementation increases offspring growth or survival, females should preferentially allocate supplemental carotenoids to offspring rather than ornamentation. We tested whether ornamented convict cichlid (Amatitlania siquia) mothers increase carotenoid allocation to eggs with access to dietary carotenoid supplementation, and whether beneficial maternal effects of carotenoid supplementation were transferred to offspring. Maternal effects of carotenoid supplementation were found; offspring from mothers that ate carotenoids benefited in terms of growth and survival parameters compared with offspring from non‐supplemented mothers. Contrary to predictions, maternal coloration was not sacrificed for fecundity: mothers maintained their coloration and integument carotenoids even though increased carotenoids in the maternal diet benefited the offspring. Furthermore, carotenoid supplementation did not increase the carotenoids deposited in eggs. This work is the first to show that indirect maternal effects of carotenoid supplementation increased the survival and growth of offspring. Indirect benefits may reduce the cost of carotenoid allocation to offspring, but the mechanism for how carotenoids support offspring development is still unknown.

Honest signaling of carotenoid-based ornaments may be reinforced by dietary limitation and by competing physiological demands for carotenoids. This study measured dietary carotenoids in a natural population of the convict cichlid Amatitlania siquia , a species in which females possess carotenoid-based ventral coloration. The zoonotic pathogen Edwardsiella spp. was detected in wild A. siquia , but carotenoids in the stomachs of wild fish did not vary significantly with parasite infection. We followed this with a laboratory experiment on domestic convict cichlids to test whether increased dietary carotenoid (ß-carotene) would decrease oxidative stress and facilitate clearance of experimental infections of Edwardsiella tarda . Fish maintained on a medium carotenoid diet (similar to the diets of wild fish) recovered from E. tarda infections more rapidly than fish on control diets, though fish on high carotenoid diets did not experience a similar benefit. There was an interaction between carotenoid diet and E. tarda such that uninfected fish on the medium carotenoid diet were significantly more colorful along their ventral surface compared to fish on the control and high carotenoid diets. Neither diet nor E. tarda infection affected oxidative damage, antioxidant capacity, or carotenoid content of the skin. From our field and laboratory data we conclude that carotenoid intake by wild convict cichlids occurs at a rate sufficient to affect bacterial infection and ventral coloration, but more data are needed from wild populations to verify the functional significance of these interactions.

Carotenoids may act as antioxidants under many circumstances. We examined the importance of carotenoids as antioxidants in the gonads of male convict cichlids (Amatitlania nigrofasciata), a species in which males lack the carotenoid-based breeding coloration that characterizes females. Male fish were fed one of four diets that included different combinations of xanthophyll and carotene carotenoids, and then we measured carotenoid concentration of the gonads, gonadosomatic index (GSI), sperm motility, and the antioxidant capacity of the gonads. Significant differences were found in gonadal carotenoid content among treatment groups, suggesting that dietary carotenoids were indeed sequestered in the gonads. There were no differences among diet groups, however, in GSI, sperm motility, or gonadal antioxidant capacity. These findings suggest that carotenoids are required only in small amounts in the testes of male convict cichlids or that they play a limited role in protecting sperm from oxidative damage.

The carotenoid trade-off hypothesis states that diet-derived carotenoids are traded off among competing physiological demands, but this statement is rarely tested in ornamented females. In this study, reverse sexually dimorphic convict cichlids (Amantitlania nigrofasciata) were fed diets containing carotenoid supplementation at three biologically relevant levels for 12 wk. This treatment was followed by spectral, microscopic, and chemical analysis to determine how females allocated the pigments to tissues and how those decisions affected their ventral patch coloration. Yellow coloration of the integument increased with carotenoids in the diet, as did carotenoids deposited in ovaries, but diet did not change carotenoid allocation to skin. The results of this study suggest that females have the ability to modulate their expression of yellow coloration via an alternative coloration strategy. Gonadosomatic index and tank environment were also related to ventral patch color, supporting previous behavioral work highlighting the importance of social selection in reinforcing signal honesty.

Phytoestrogens are plant compounds that can act as endocrine disruptors in vertebrates. Biologically active levels of phytoestrogens have been found in aquatic habitats near wood pulp and paper mills, biofuel manufacturing plants, sewage-treatment plants, and agricultural fields. Phytoestrogens are known to cause hormonal and gonadal changes in male fish, but few studies have connected these effects to outcomes relevant to reproductive success. In one experiment, we exposed sexually mature male fighting fish Betta splendens to environmentally relevant (1 μg L −1 ) and pharmacological concentrations (1000 μg L −1 ) of the phytoestrogen genistein as well as to a positive control of waterborne 17β-estradiol (E2; 1 μg L −1 ), and a negative control of untreated water. In a second experiment, we exposed male B. splendens to environmentally relevant concentrations (1 μg L −1 ) of genistein and β-sitosterol singly and in combination as well as to the positive and negative controls. All exposures were 21 days in duration. We measured sex-steroid hormone levels, gonadosomatic index (GSI), sperm concentration and motility, and fertilization success in these fish. We found that exposure to genistein did not affect circulating levels of the androgen 11-ketotestosterone or the estrogen E2 relative to negative-control fish. We also found that neither of the compounds nor their mixture affected GSI, sperm concentration or motility, or fertilization success in exposed fish relative to negative-control fish. However, fish exposed to phytoestrogens showed some evidence of fewer but more motile sperm than fish exposed to the positive control E2. We conclude that sexually mature male B. splendens are relatively immune to reproductive impairments from short-term exposure to waterborne phytoestrogens.

A wide range of research has promised new tools for forecasting infectious disease dynamics, but little of that research is currently being applied in practice, because tools do not address key public health needs, do not produce probabilistic forecasts, have not been evaluated on external data, or do not provide sufficient forecast skill to be useful. We developed an open collaborative forecasting challenge to assess probabilistic forecasts for seasonal epidemics of dengue, a major global public health problem. Sixteen teams used a variety of methods and data to generate forecasts for 3 epidemiological targets (peak incidence, the week of the peak, and total incidence) over 8 dengue seasons in Iquitos, Peru and San Juan, Puerto Rico. Forecast skill was highly variable across teams and targets. While numerous forecasts showed high skill for midseason situational awareness, early season skill was low, and skill was generally lowest for high incidence seasons, those for which forecasts would be most valuable. A comparison of modeling approaches revealed that average forecast skill was lower for models including biologically meaningful data and mechanisms and that both multimodel and multiteam ensemble forecasts consistently outperformed individual model forecasts. Leveraging these insights, data, and the forecasting framework will be critical to improve forecast skill and the application of forecasts in real time for epidemic preparedness and response. Moreover, key components of this project—integration with public health needs, a common forecasting framework, shared and standardized data, and open participation—can help advance infectious disease forecasting beyond dengue.