Obesogens and Obesity: State-of-the-Science and Future Directions Summary from a Healthy Environment and Endocrine Disruptors Strategies Workshop.

On September 7 and 8, 2022, Healthy Environment and Endocrine Disruptors Strategies, an Environmental Health Sciences program, convened a scientific workshop of relevant stakeholders involved in obesity, toxicology, or obesogen research to review the state of the science regarding the role of obesogenic chemicals that might be contributing to the obesity pandemic. The workshop's objectives were to examine the evidence supporting the hypothesis that obesogens contribute to the etiology of human obesity; to discuss opportunities for improved understanding, acceptance, and dissemination of obesogens as contributors to the obesity pandemic; and to consider the need for future research and potential mitigation strategies. This report details the discussions, key areas of agreement, and future opportunities to prevent obesity. The attendees agreed that environmental obesogens are real, significant, and a contributor at some degree to weight gain at the individual level and to the global obesity and metabolic disease pandemic at a societal level; moreover, it is at least, in theory, remediable.

APOBEC3 selects V179I in HIV-1 reverse transcriptase to provide selective advantage for non-nucleoside reverse transcriptase inhibitor-resistant mutants.

The V179I substitution in human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) is selected in humans or mouse models treated with certain nonnucleoside reverse transcriptase inhibitors (NNRTIs). While it is often observed together with other NNRTI resistance mutations, V179I does not confer drug resistance. To understand how V179I arises during NNRTI treatment, we characterized it in HIV-1 molecular clones with or without the NNRTI resistance mutations Y181C or Y181V. While V179I alone did not confer resistance to any NNRTIs tested, when present with Y181C/V it enhanced drug resistance to some NNRTIs by 3- to 8-fold. In replication competition experiments in the presence of the NNRTI rilpivirine (RPV), V179I modestly enhanced Y181C HIV-1 or Y181V HIV-1 replication compared to viruses without V179I. As V179I arises from a G to A mutation, we evaluated whether it could arise due to host APOBEC3 deaminase activity and be maintained in the presence of a NNRTI to provide a selective advantage for the virus. V179I was detected in some humanized mice treated with RPV and was associated with G to A mutations characteristic of APOBEC3 activity. In RPV selection experiments, the frequency of V179I in HIV-1 was accelerated in CD4+ T cells expressing higher APOBEC3F and APOBEC3G levels. Our results provide evidence that V179I in HIV-1 RT can arise due to APOBEC-mediated G to A hypermutation and can confer a selective advantage to drug-resistant HIV-1 isolates in the presence of some NNRTIs.

Construction and characterization of two SARS-CoV-2 minigenome replicon systems.

The ongoing COVID-19 pandemic severely impacts global public health and economies. To facilitate research on severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virology and antiviral discovery, a noninfectious viral replicon system operating under biosafety level 2 containment is warranted. We report herein the construction and characterization of two SARS-CoV-2 minigenome replicon systems. First, we constructed the IVT-CoV2-Rep complementary DNA template to generate a replicon messenger RNA (mRNA) with nanoluciferase (NLuc) reporter via in vitro transcription (IVT). The replicon mRNA transfection assay demonstrated a rapid and transient replication of IVT-CoV2-Rep in a variety of cell lines, which could be completely abolished by known SARS-CoV-2 replication inhibitors. Our data also suggest that the transient phenotype of IVT-CoV2-Rep is not due to host innate antiviral responses. In addition, we have developed a DNA-launched replicon BAC-CoV2-Rep, which supports the in-cell transcription of a replicon mRNA as initial replication template. The BAC-CoV2-Rep transient transfection system exhibited a much stronger and longer replicon signal compared to the IVT-CoV2-Rep version. We also found that a portion of the NLuc reporter signal was derived from the spliced BAC-CoV2-Rep mRNA and was resistant to antiviral treatment, especially during the early phase after transfection. In summary, the established SARS-CoV-2 transient replicon systems are suitable for basic and antiviral research, and hold promise for stable replicon cell line development with further optimization.

Disassembling the Nature of Capsid: Biochemical, Genetic, and Imaging Approaches to Assess HIV-1 Capsid Functions.

The human immunodeficiency virus type 1 (HIV-1) capsid and its disassembly, or capsid uncoating, has remained an active area of study over the past several decades. Our understanding of the HIV-1 capsid as solely a protective shell has since shifted with discoveries linking it to other complex replication events. The interplay of the HIV-1 capsid with reverse transcription, nuclear import, and integration has led to an expansion of knowledge of capsid functionality. Coincident with advances in microscopy, cell, and biochemistry assays, several models of capsid disassembly have been proposed, in which it occurs in either the cytoplasmic, nuclear envelope, or nuclear regions of the cell. Here, we discuss how the understanding of the HIV-1 capsid has evolved and the key methods that made these discoveries possible.

Permeability of the HIV-1 capsid to metabolites modulates viral DNA synthesis.

Reverse transcription, an essential event in the HIV-1 life cycle, requires deoxynucleotide triphosphates (dNTPs) to fuel DNA synthesis, thus requiring penetration of dNTPs into the viral capsid. The central cavity of the capsid protein (CA) hexamer reveals itself as a plausible channel that allows the passage of dNTPs into assembled capsids. Nevertheless, the molecular mechanism of nucleotide import into the capsid remains unknown. Employing all-atom molecular dynamics (MD) simulations, we established that cooperative binding between nucleotides inside a CA hexamer cavity results in energetically favorable conditions for passive translocation of dNTPs into the HIV-1 capsid. Furthermore, binding of the host cell metabolite inositol hexakisphosphate (IP6) enhances dNTP import, while binding of synthesized molecules like benzenehexacarboxylic acid (BHC) inhibits it. The enhancing effect on reverse transcription by IP6 and the consequences of interactions between CA and nucleotides were corroborated using atomic force microscopy, transmission electron microscopy, and virological assays. Collectively, our results provide an atomistic description of the permeability of the HIV-1 capsid to small molecules and reveal a novel mechanism for the involvement of metabolites in HIV-1 capsid stabilization, nucleotide import, and reverse transcription.

A Novel Phenotype Links HIV-1 Capsid Stability to cGAS-Mediated DNA Sensing.

The HIV-1 capsid executes essential functions that are regulated by capsid stability and host factors. In contrast to increasing knowledge on functional roles of capsid-interacting host proteins during postentry steps, less is known about capsid stability and its impact on intracellular events. Here, using the antiviral compound PF-3450074 (PF74) as a probe for capsid function, we uncovered a novel phenotype of capsid stability that has a profound effect on innate sensing of viral DNA by the DNA sensor cGAS. A single mutation, R143A, in the capsid protein conferred resistance to high concentrations of PF74, without affecting capsid binding to PF74. A cell-free assay showed that the R143A mutant partially counteracted the capsid-destabilizing activity of PF74, pointing to capsid stabilization as a resistance mechanism for the R143A mutant. In monocytic THP-1 cells, the R143A virus, but not the wild-type virus, suppressed cGAS-dependent innate immune activation. These results suggest that capsid stabilization improves the shielding of viral DNA from innate sensing. We found that a naturally occurring transmitted founder (T/F) variant shares the same properties as the R143A mutant with respect to PF74 resistance and DNA sensing. Imaging assays revealed delayed uncoating kinetics of this T/F variant and the R143A mutant. All these phenotypes of this T/F variant were controlled by a genetic polymorphism located at the trimeric interface between capsid hexamers, thus linking these capsid-dependent properties. Overall, this work functionally connects capsid stability to innate sensing of viral DNA and reveals naturally occurring phenotypic variation in HIV-1 capsid stability.IMPORTANCE The HIV-1 capsid, which is made from individual viral capsid proteins (CA), is a target for a number of antiviral compounds, including the small-molecule inhibitor PF74. In the present study, we utilized PF74 to identify a transmitted/founder (T/F) strain that shows increased capsid stability. Interestingly, PF74-resistant variants prevented cGAS-dependent innate immune activation under a condition where the other T/F strains induced type I interferon. These observations thus reveal a new CA-specific phenotype that couples capsid stability to viral DNA recognition by cytosolic DNA sensors.

CA Mutation N57A Has Distinct Strain-Specific HIV-1 Capsid Uncoating and Infectivity Phenotypes.

The ability of human immunodeficiency virus type 1 (HIV-1) to transduce nondividing cells is key to infecting terminally differentiated macrophages, which can serve as a long-term reservoir of HIV-1 infection. The mutation N57A in the viral CA protein renders HIV-1 cell cycle dependent, allowing examination of HIV-1 infection of nondividing cells. Here, we show that the N57A mutation confers a postentry infectivity defect that significantly differs in magnitude between the common lab-adapted molecular clones HIV-1NL4-3 (>10-fold) and HIV-1LAI (2- to 5-fold) in multiple human cell lines and primary CD4+ T cells. Capsid permeabilization and reverse transcription are altered when N57A is incorporated into HIV-1NL4-3 but not HIV-1LAI The N57A infectivity defect is significantly exacerbated in both virus strains in the presence of cyclosporine (CsA), indicating that N57A infectivity is dependent upon CA interacting with host factor cyclophilin A (CypA). Adaptation of N57A HIV-1LAI selected for a second CA mutation, G94D, which rescued the N57A infectivity defect in HIV-1LAI but not HIV-1NL4-3 The rescue of N57A by G94D in HIV-1LAI is abrogated by CsA treatment in some cell types, demonstrating that this rescue is CypA dependent. An examination of over 40,000 HIV-1 CA sequences revealed that the four amino acids that differ between HIV-1NL4-3 and HIV-1LAI CA are polymorphic, and the residues at these positions in the two strains are widely prevalent in clinical isolates. Overall, a few polymorphic amino acid differences between two closely related HIV-1 molecular clones affect the phenotype of capsid mutants in different cell types.IMPORTANCE The specific mechanisms by which HIV-1 infects nondividing cells are unclear. A mutation in the HIV-1 capsid protein abolishes the ability of the virus to infect nondividing cells, serving as a tool to examine cell cycle dependence of HIV-1 infection. We have shown that two widely used HIV-1 molecular clones exhibit significantly different N57A infectivity phenotypes due to fewer than a handful of CA amino acid differences and that these clones are both represented in HIV-infected individuals. As such minor differences in closely related HIV-1 strains may impart significant infectivity differences, careful consideration should be given to drawing conclusions from one particular HIV-1 clone. This study highlights the potential for significant variation in results with the use of multiple strains and possible unanticipated effects of natural polymorphisms.

Two Coselected Distal Mutations in HIV-1 Reverse Transcriptase (RT) Alter Susceptibility to Nonnucleoside RT Inhibitors and Nucleoside Analogs.

Two mutations, G112D and M230I, were selected in the reverse transcriptase (RT) of human immunodeficiency virus type 1 (HIV-1) by a novel nonnucleoside reverse transcriptase inhibitor (NNRTI). G112D is located near the HIV-1 polymerase active site; M230I is located near the hydrophobic region where NNRTIs bind. Thus, M230I could directly interfere with NNRTI binding but G112D could not. Biochemical and virological assays were performed to analyze the effects of these mutations individually and in combination. M230I alone caused a reduction in susceptibility to NNRTIs, while G112D alone did not. The G112D/M230I double mutant was less susceptible to NNRTIs than was M230I alone. In contrast, both mutations affected the ability of RT to incorporate nucleoside analogs. We suggest that the mutations interact with each other via the bound nucleic acid substrate; the nucleic acid forms part of the polymerase active site, which is near G112D. The positioning of the nucleic acid is influenced by its interactions with the "primer grip" region and could be influenced by the M230I mutation.IMPORTANCE Although antiretroviral therapy (ART) is highly successful, drug-resistant variants can arise that blunt the efficacy of ART. New inhibitors that are broadly effective against known drug-resistant variants are needed, although such compounds might select for novel resistance mutations that affect the sensitivity of the virus to other compounds. Compound 13 selects for resistance mutations that differ from traditional NNRTI resistance mutations. These mutations cause increased sensitivity to NRTIs, such as AZT.

Truncated CPSF6 Forms Higher-Order Complexes That Bind and Disrupt HIV-1 Capsid.

Cleavage and polyadenylation specificity factor 6 (CPSF6) is a human protein that binds HIV-1 capsid and mediates nuclear transport and integration targeting of HIV-1 preintegration complexes. Truncation of the protein at its C-terminal nuclear-targeting arginine/serine-rich (RS) domain produces a protein, CPSF6-358, that potently inhibits HIV-1 infection by targeting the capsid and inhibiting nuclear entry. To understand the molecular mechanism behind this restriction, the interaction between CPSF6-358 and HIV-1 capsid was characterized using in vitro and in vivo assays. Purified CPSF6-358 protein formed oligomers and bound in vitro-assembled wild-type (WT) capsid protein (CA) tubes, but not CA tubes containing a mutation in the putative binding site of CPSF6. Intriguingly, binding of CPSF6-358 oligomers to WT CA tubes physically disrupted the tubular assemblies into small fragments. Furthermore, fixed- and live-cell imaging showed that stably expressed CPSF6-358 forms cytoplasmic puncta upon WT HIV-1 infection and leads to capsid permeabilization. These events did not occur when the HIV-1 capsid contained a mutation known to prevent CPSF6 binding, nor did they occur in the presence of a small-molecule inhibitor of capsid binding to CPSF6-358. Together, our in vitro biochemical and transmission electron microscopy data and in vivo intracellular imaging results provide the first direct evidence for an oligomeric nature of CPSF6-358 and suggest a plausible mechanism for restriction of HIV-1 infection by CPSF6-358.IMPORTANCE After entry into cells, the HIV-1 capsid, which contains the viral genome, interacts with numerous host cell factors to facilitate crucial events required for replication, including uncoating. One such host cell factor, called CPSF6, is predominantly located in the cell nucleus and interacts with HIV-1 capsid. The interaction between CA and CPSF6 is critical during HIV-1 replication in vivo Truncation of CPSF6 leads to its localization to the cell cytoplasm and inhibition of HIV-1 infection. Here, we determined that truncated CPSF6 protein forms large higher-order complexes that bind directly to HIV-1 capsid, leading to its disruption. Truncated CPSF6 expression in cells leads to premature capsid uncoating that is detrimental to HIV-1 infection. Our study provides the first direct evidence for an oligomeric nature of truncated CPSF6 and insights into the highly regulated process of HIV-1 capsid uncoating.

Impact of fog drip versus fog immersion on the physiology of Bishop pine saplings.

Fog-drip to the soil is the most obvious contribution of fog to the water budget of an ecosystem, but several studies provide convincing evidence that foliar absorption of fog water through leaf wetting events is also possible. The focus of our research was to assess the relative importance of fog drip and fog immersion (foliar wetting) on leaf gas-exchange rates and photosynthetic capacity of a coastal pine species, Bishop pine (Pinus muricata D.Don), a drought-sensitive species restricted to the fog belt of coastal California and offshore islands. In a controlled experiment, we manipulated fog water inputs to potted Bishop pine saplings during a 3 week dry-down period. Ten saplings were randomly assigned one of two fog treatments: (1) fog drip to the soil and canopy fog immersion, or (2) fog immersion alone. Five saplings were assigned the 'control' group and received no fog water inputs. We found that fog immersion alone significantly increased carbon assimilation rates and photosynthetic capacity of saplings as soil moisture declined compared with those that received no fog at all. The highest carbon assimilation rates were observed in saplings that also received fog drip. Soil moisture was 40% higher in the fog immersion compared with the control group during the dry-down, indicating a reduced demand for soil water in saplings that had only leaves wetted by canopy interception of fog. Leaf-level physiology is more strongly enhanced by fog drip compared with fog immersion, although the results of this study provide evidence that foliar absorption is a viable mechanism by which Bishop pines use fog water and that it can enhance instantaneous plant carbon gain and potentially whole plant productivity.

Coastal fog during summer drought improves the water status of sapling trees more than adult trees in a California pine forest.

Fog water inputs can offset seasonal drought in the Mediterranean climate of coastal California and may be critical to the persistence of many endemic plant species. The ability to predict plant species response to potential changes in the fog regime hinges on understanding the ways that fog can impact plant physiological function across life stages. Our study uses a direct metric of water status, namely plant water potential, to understand differential responses of adult versus sapling trees to seasonal drought and fog water inputs. We place these measurements within a water balance framework that incorporates the varying climatic and soil property impacts on water budgets and deficit. We conducted our study at a coastal and an inland site within the largest stand of the regionally endemic bishop pine (Pinus muricata D. Don) on Santa Cruz Island. Our results show conclusively that summer drought negatively affects the water status of sapling more than adult trees and that sapling trees are also more responsive to changes in shallow soil moisture inputs from fog water deposition. Moreover, between the beginning and end of a large, late-season fog drip event, water status increased more for saplings than for adults. Relative to non-foggy conditions, we found that fog water reduces modeled peak water deficit by 80 and 70 % at the inland and coastal sites, respectively. Results from our study inform mechanistically based predictions of how population dynamics of this and other coastal species may be affected by a warmer, drier, and potentially less foggy future.

Respiratory syncytial virus infection enhances Pseudomonas aeruginosa biofilm growth through dysregulation of nutritional immunity.

Clinical observations link respiratory virus infection and Pseudomonas aeruginosa colonization in chronic lung disease, including cystic fibrosis (CF) and chronic obstructive pulmonary disease. The development of P. aeruginosa into highly antibiotic-resistant biofilm communities promotes airway colonization and accounts for disease progression in patients. Although clinical studies show a strong correlation between CF patients' acquisition of chronic P. aeruginosa infections and respiratory virus infection, little is known about the mechanism by which chronic P. aeruginosa infections are initiated in the host. Using a coculture model to study the formation of bacterial biofilm formation associated with the airway epithelium, we show that respiratory viral infections and the induction of antiviral interferons promote robust secondary P. aeruginosa biofilm formation. We report that the induction of antiviral IFN signaling in response to respiratory syncytial virus (RSV) infection induces bacterial biofilm formation through a mechanism of dysregulated iron homeostasis of the airway epithelium. Moreover, increased apical release of the host iron-binding protein transferrin during RSV infection promotes P. aeruginosa biofilm development in vitro and in vivo. Thus, nutritional immunity pathways that are disrupted during respiratory viral infection create an environment that favors secondary bacterial infection and may provide previously unidentified targets to combat bacterial biofilm formation.

Low TCR signal strength induces combined expansion of Th2 and regulatory T cell populations that protect mice from the development of type 1 diabetes.

AIMS/HYPOTHESIS: Weak stimulation of CD4(+) T cells induces expansion of CD4(+) forkhead box P3(+) regulatory T cells (Tregs) and can also promote T helper (Th) 2 responses, which have demonstrable beneficial effects on autoimmune diabetes. This study explored the feasibility of combined Treg/Th2 expansion for immunotherapy of type 1 diabetes in NOD mice. METHODS: We compared Treg and Th responses to dendritic cells (DC) presenting scaled antigen doses to islet-specific NOD CD4(+) T cells. Flow cytometric and Luminex analyses were performed to determine the phenotype and cytokine profile of expanded T cells. The ability of expanded T cells to prevent type 1 diabetes was tested in an adoptive transfer model. RESULTS: In vitro studies revealed a hierarchical, selective expansion of Treg and T effector (Teff) populations at different antigen doses. Thus, a single low dose produced a mixture of Tregs Th2 and type 1 regulatory (Tr1) cells, which prevented diabetes in NOD-SCID mice and increased the ratio of Treg/Teff cells infiltrating the pancreatic islets. Subcutaneous injection of DC, previously shown to prevent diabetes in NOD mice, induced expansion of the same mixture of Tregs Tr1 and Th2 cells. Low-dose expansion of Treg required MHC-T cell receptor interaction and was partly dependent on T cell derived TGF-ß and IL-2. Autocrine IFN-γ was required for the promotion of diabetogenic Th1 cells at high antigen doses. CONCLUSIONS/INTERPRETATION: Weak stimulation of CD4(+) T cells with DC and low-dose antigen expands a combination of antigen-specific Tregs Th2 and Tr1 cells that prevent autoimmunity, without the need to target or purify specific Treg populations.

Decision analysis for designing marine protected areas for multiple species with uncertain fishery status.

Marine protected areas (MPAs) are growing in popularity as a conservation tool, and there are increasing calls for additional MPAs. Meta-analyses indicate that most MPAs successfully meet the minimal goal of increasing biomass inside the MPA, while some do not, leaving open the important question of what makes MPAs successful. An often-overlooked aspect of this problem is that the success of fishery management outside MPA boundaries (i.e., whether a population is overfished) affects how well MPAs meet both conservation goals (e.g., increased biomass) and economic goals (e.g., minimal negative effects on fishery yield). Using a simple example of a system with homogeneous habitat and periodically spaced MPAs, we show that, as area in MPAs increases, (1) conservation value (biomass) may initially be zero, implying no benefit, then at some point increases monotonically; and (2) fishery yield may be zero, then increases monotonically to a maximum beyond which further increase in MPA area causes yield to decline. Importantly, the points at which these changes in slope occur vary among species and depend on management outside MPAs. Decision makers considering the effects of a potential system of MPAs on multiple species are confronted by a number of such cost-benefit curves, and it is usually impossible to maximize benefits and minimize costs for all species. Moreover, the precise shape of each curve is unknown due to uncertainty regarding the fishery status of each species. Here we describe a decision-analytic approach that incorporates existing information on fishery stock status to present decision makers with the range of likely outcomes of MPA implementation. To summarize results from many species whose overfishing status is uncertain, our decision-analysis approach involves weighted averages over both overfishing uncertainty and species. In an example from an MPA decision process in California, USA, an optimistic projection of future fishery management success led to recommendation of fewer and smaller MPAs than that derived from a more pessimistic projection of future management success. This example illustrates how information on fishery status can be used to project potential outcomes of MPA implementation within a decision analysis framework and highlights the need for better population information.

The influence of summertime fog and overcast clouds on the growth of a coastal Californian pine: a tree-ring study.

The coast of California is home to numerous rare, endemic conifers and other plants that are limited in distribution by drought sensitivity and the summer-dry climate that prevails across most of the state. Ecologists have long assumed that some coastal plant populations survived the early Pleistocene transition to a warmer and drier environment because they benefit from frequent fog and stratus clouds that provide water and shade during the rainless summer. One such population is that of Torrey pine (Pinus torreyana ssp. Insularis) on Santa Rosa Island in Channel Islands National Park. Here we report that the tree-ring width record from this population indicates strong growth sensitivities to summer fog drip and cloud shading. We quantified the effects of summer cloud cover by comparing ring-width indices to coastal airport cloud-frequency records (1944-2004). For the first time observed, summertime cloud frequency correlated positively with ring-width indices, regardless of whether the effect of rainfall was first removed from the ring-width record. The effect of ground-level fog was strongest in July early mornings (03:00 PST, R(2) = 0.262, P < 0.0002). The effect of clouds high enough to provide shade but not fog water was also strongest in July, but climbed steadily throughout the day before becoming strongest in late afternoon (16:00-18:00 PST, R(2) = 0.148, P < 0.004). Correlations were substantially stronger in years with higher soil moisture, suggesting that growth response to summer clouds is strongly affected by pre-summer rainfall. A change in the height and/or timing of coastal cloud formation with climate change would likely affect this and other populations of California's coastal vegetation.

Children show highest levels of polybrominated diphenyl ethers in a California family of four: a case study.

Polybrominated diphenyl ethers (PBDEs), a major class of flame retardants, are ubiquitous environmental contaminants with particularly high concentrations in humans from the United States. This study is a first attempt to report and compare PBDE concentrations in blood drawn from a family. Serum samples from family members collected at two sampling occasions 90 days apart were analyzed for PBDE congeners. Concentrations of the lower-brominated PBDEs were similar at the two sampling times for each family member, with children's levels 2- to 5-fold higher than those of their parents. Concentrations of, for example, 2,2',4,4'-tetrabromodiphenyl ether (BDE-47) varied from 32 ng/g lipid weight (lw) in the father to 60, 137, and 245 ng/g lw in the mother, child, and toddler, respectively. Decabromodiphenyl ether (BDE-209) concentrations differed significantly between the two samplings. September concentrations in the father, mother, child, and toddler were 23, 14, 143, and 233 ng/g lw, respectively. December concentrations (duplicate results from the laboratory) were 2 and 3, 4 and 4, 9 and 12, and 19 and 26 ng/g lw, respectively. Parents' summation operatorPBDE concentrations approached U.S. median concentrations, with children's concentrations near the maximum (top 5%) found in U.S. adults. The youngest child had the highest concentrations of all PBDE congeners, suggesting that younger children are more exposed to PBDEs than are adults. Our estimates indicate that house dust contributes to children's higher PBDE levels. BDE-209 levels for all family members were 10-fold lower at the second sampling. The short half-life of BDE-209 (15 days) indicates that BDE-209 levels can decrease rapidly in response to decreased exposures. This case study suggests that children are at higher risk for PBDE exposures and, accordingly, face higher risks of PBDE-related health effects than adults.