Methodological differences between studies confound one-size-fits-all approaches to managing surface waterways for food and water safety.

Even though differences in methodology (e.g., sample volume and detection method) have been shown to affect observed microbial water quality, multiple sampling and laboratory protocols continue to be used for water quality monitoring. Research is needed to determine how these differences impact the comparability of findings to generate best management practices and the ability to perform meta-analyses. This study addresses this knowledge gap by compiling and analyzing a data set representing 2,429,990 unique data points on at least one microbial water quality target (e.g., Salmonella presence and Escherichia coli concentration). Variance partitioning analysis was used to quantify the variance in likelihood of detecting each pathogenic target that was uniquely and jointly attributable to non-methodological versus methodological factors. The strength of the association between microbial water quality and select methodological and non-methodological factors was quantified using conditional forest and regression analysis. Fecal indicator bacteria concentrations were more strongly associated with non-methodological factors than methodological factors based on conditional forest analysis. Variance partitioning analysis could not disentangle non-methodological and methodological signals for pathogenic Escherichia coli, Salmonella, and Listeria. This suggests our current perceptions of foodborne pathogen ecology in water systems are confounded by methodological differences between studies. For example, 31% of total variance in likelihood of Salmonella detection was explained by methodological and/or non-methodological factors, 18% was jointly attributable to both methodological and non-methodological factors. Only 13% of total variance was uniquely attributable to non-methodological factors for Salmonella, highlighting the need for standardization of methods for microbiological water quality testing for comparison across studies.IMPORTANCEThe microbial ecology of water is already complex, without the added complications of methodological differences between studies. This study highlights the difficulty in comparing water quality data from projects that used different sampling or laboratory methods. These findings have direct implications for end users as there is no clear way to generalize findings in order to characterize broad-scale ecological phenomenon and develop science-based guidance. To best support development of risk assessments and guidance for monitoring and managing waters, data collection and methods need to be standardized across studies. A minimum set of data attributes that all studies should collect and report in a standardized way is needed. Given the diversity of methods used within applied and environmental microbiology, similar studies are needed for other microbiology subfields to ensure that guidance and policy are based on a robust interpretation of the literature.

Maternal fish consumption and child neurodevelopment in Nutrition 1 Cohort: Seychelles Child Development Study.

Maternal fish consumption exposes the fetus to beneficial nutrients and potentially adverse neurotoxicants. The current study investigated associations between maternal fish consumption and child neurodevelopmental outcomes. Maternal fish consumption was assessed in the Seychelles Child Development Study Nutrition Cohort 1 (n 229) using 4-day food diaries. Neurodevelopment was evaluated at 9 and 30 months, and 5 and 9 years with test batteries assessing twenty-six endpoints and covering multiple neurodevelopmental domains. Analyses used multiple linear regression with adjustment for covariates known to influence child neurodevelopment. This cohort consumed an average of 8 fish meals/week and the total fish intake during pregnancy was 106·8 (sd 61·9) g/d. Among the twenty-six endpoints evaluated in the primary analysis there was one beneficial association. Children whose mothers consumed larger quantities of fish performed marginally better on the Kaufman Brief Intelligence Test (a test of nonverbal intelligence) at age 5 years (ß 0·003, 95 % CI (0, 0·005)). A secondary analysis dividing fish consumption into tertiles found no significant associations when comparing the highest and lowest consumption groups. In this cohort, where fish consumption is substantially higher than current global recommendations, maternal fish consumption during pregnancy was not beneficially or adversely associated with children's neurodevelopmental outcomes.

Scale of analysis drives the observed ratio of spatial to non-spatial variance in microbial water quality: insights from two decades of citizen science data.

AIMS: While fecal indicator bacteria (FIB) testing is used to monitor surface water for potential health hazards, observed variation in FIB levels may depend on the scale of analysis (SOA). Two decades of citizen science data, coupled with random effects models, were used to quantify the variance in FIB levels attributable to spatial versus temporal factors. METHODS AND RESULTS: Separately, Bayesian models were used to quantify the ratio of spatial to non-spatial variance in FIB levels and identify associations between environmental factors and FIB levels. Separate analyses were performed for three SOA: waterway, watershed, and statewide. As SOA increased (from waterway to watershed to statewide models), variance attributable to spatial sources generally increased and variance attributable to temporal sources generally decreased. While relationships between FIB levels and environmental factors, such as flow conditions (base versus stormflow), were constant across SOA, the effect of land cover was highly dependent on SOA and consistently smaller than the effect of stormwater infrastructure (e.g. outfalls). CONCLUSIONS: This study demonstrates the importance of SOA when developing water quality monitoring programs or designing future studies to inform water management.

Structural Equation Models Suggest That On-Farm Noncrop Vegetation Removal Is Not Associated with Improved Food Safety Outcomes but Is Linked to Impaired Water Quality.

While growers have reported pressures to minimize wildlife intrusion into produce fields through noncrop vegetation (NCV) removal, NCV provides key ecosystem services. To model food safety and environmental tradeoffs associated with NCV removal, published and publicly available food safety and water quality data from the Northeastern United States were obtained. Because data on NCV removal are not widely available, forest-wetland cover was used as a proxy, consistent with previous studies. Structural equation models (SEMs) were used to quantify the effect of forest-wetland cover on (i) food safety outcomes (e.g., detecting pathogens in soil) and (ii) water quality (e.g., nutrient levels). Based on the SEMs, NCV was not associated with or had a protective effect on food safety outcomes (more NCV was associated with a reduced likelihood of pathogen detection). The probabilities of detecting Listeria spp. in soil (effect estimate [EE] = -0.17; P = 0.005) and enterohemorrhagic Escherichia coli in stream samples (EE = -0.27; P < 0.001) were negatively associated with the amount of NCV surrounding the sampling site. Larger amounts of NCV were also associated with lower nutrient, salinity, and sediment levels, and higher dissolved oxygen levels. Total phosphorous levels were negatively associated with the amount of NCV in the upstream watershed (EE = -0.27; P < 0.001). Similar negative associations (P < 0.05) were observed for other physicochemical parameters, such as nitrate (EE = -0.38). Our findings suggest that NCV should not be considered an inherent produce safety risk or result in farm audit demerits. This study also provides a framework for evaluating environmental tradeoffs associated with using specific preharvest food safety strategies. IMPORTANCE Currently, on-farm food safety decisions are typically made independently of conservation considerations, often with detrimental impacts on agroecosystems. Comanaging agricultural environments to simultaneously meet conservation and food safety aims is complicated because farms are closely linked to surrounding environments, and management decisions can have unexpected environmental, economic, and food safety consequences. Thus, there is a need for research on the conservation and food safety tradeoffs associated with implementing specific preharvest food safety practices. Understanding these tradeoffs is critical for developing adaptive comanagement strategies and ensuring the short- and long-term safety, sustainability, and profitability of agricultural systems. This study quantifies tradeoffs and synergies between food safety and environmental aims, and outlines a framework for modeling tradeoffs and synergies between management aims that can be used to support future comanagement research.

Gut microbiome in people living with HIV is associated with impaired thiamine and folate syntheses.

People living with HIV have a high incidence of cardiovascular and neurological diseases as comorbid disorders that are commonly linked to inflammation. While microbial translocation can augment inflammation during HIV infection, functional microbiome shifts that may increase pro-inflammatory responses have not been fully characterized. In addition, defining HIV-induced microbiome changes has been complicated by high variability among individuals. Here we conducted functional annotation of previously-published 16S ribosomal RNA gene sequences of 305 HIV positive and 249 negative individuals, with adjustment for geographic region, sex, sexual behavior, and age. Metagenome profiles were inferred from these individuals' 16S data. HIV infection was associated with impaired microbial vitamin B synthesis; around half of the gene families in thiamine and folate biosynthesis pathways were significantly less abundant in the HIV positive group than the negative control. These results are consistent with the high prevalence of thiamine and folate deficiencies in HIV infections. These HIV-induced microbiota shifts have the potential to influence cardiovascular and neurocognitive diseases, given the documented associations between B-vitamin deficiencies, inflammation, and these diseases. We also observed that most essential amino acid biosynthesis pathways were downregulated in the microbiome of HIV-infected individuals. Microbial vitamin B and amino acid synthesis pathways were not significantly recovered by antiretroviral treatment when we compared 262 ART positive and 184 ART negative individuals. Our meta-analysis provides a new outlook for understanding vitamin B and amino acid deficiencies in HIV patients, suggesting that interventions for reversing HIV-induced microbiome shifts may aid in lessening the burdens of HIV comorbidities.

Developmental Toxicology of Metal Mixtures in Drosophila: Unique Properties of Potency and Interactions of Mercury Isoforms.

Mercury ranks third on the U.S. Agency of Toxic Substances and Disease Registry priority list of hazardous substances, behind only arsenic and lead. We have undertaken uncovering the mechanisms underlying the developmental toxicity of methylmercury (MeHg), inorganic mercury (HgCl2), lead acetate (Pb), and sodium arsenite (As). To probe these differences, we used the Drosophila model, taking advantage of three developmental transitions-pupariation, metamorphosis, and eclosion-to differentiate potentially unique windows of toxicity. We elaborated dose response profiles for each individual metal administered in food and accounted for internal body burden, also extending analyses to evaluate combinatorial metal mixture effects. We observed all four metals producing larval lethality and delayed pupariation, with MeHg being most potent. Compared to other metals, MeHg's potency is caused by a higher body burden with respect to dose. MeHg uniquely caused dose-dependent failure in eclosion that was unexpectedly rescued by titrating in HgCl2. Our results highlight a unique developmental window and toxicokinetic properties where MeHg acts with specificity relative to HgCl2, Pb, and As. These findings will serve to refine future studies aimed at revealing tissue morphogenesis events and cell signaling pathways, potentially conserved in higher organisms, that selectively mediate MeHg toxicity and its antagonism by HgCl2.

Multiple imputation and direct estimation for qPCR data with non-detects.

BACKGROUND: Quantitative real-time PCR (qPCR) is one of the most widely used methods to measure gene expression. An important aspect of qPCR data that has been largely ignored is the presence of non-detects: reactions failing to exceed the quantification threshold and therefore lacking a measurement of expression. While most current software replaces these non-detects with a value representing the limit of detection, this introduces substantial bias in the estimation of both absolute and differential expression. Single imputation procedures, while an improvement on previously used methods, underestimate residual variance, which can lead to anti-conservative inference. RESULTS: We propose to treat non-detects as non-random missing data, model the missing data mechanism, and use this model to impute missing values or obtain direct estimates of model parameters. To account for the uncertainty inherent in the imputation, we propose a multiple imputation procedure, which provides a set of plausible values for each non-detect. We assess the proposed methods via simulation studies and demonstrate the applicability of these methods to three experimental data sets. We compare our methods to mean imputation, single imputation, and a penalized EM algorithm incorporating non-random missingness (PEMM). The developed methods are implemented in the R/Bioconductor package nondetects. CONCLUSIONS: The statistical methods introduced here reduce discrepancies in gene expression values derived from qPCR experiments in the presence of non-detects, providing increased confidence in downstream analyses.

HIITE: HIV-1 incidence and infection time estimator.

Motivation: Around 2.1 million new HIV-1 infections were reported in 2015, alerting that the HIV-1 epidemic remains a significant global health challenge. Precise incidence assessment strengthens epidemic monitoring efforts and guides strategy optimization for prevention programs. Estimating the onset time of HIV-1 infection can facilitate optimal clinical management and identify key populations largely responsible for epidemic spread and thereby infer HIV-1 transmission chains. Our goal is to develop a genomic assay estimating the incidence and infection time in a single cross-sectional survey setting. Results: We created a web-based platform, HIV-1 incidence and infection time estimator (HIITE), which processes envelope gene sequences using hierarchical clustering algorithms and informs the stage of infection, along with time since infection for incident cases. HIITE's performance was evaluated using 585 incident and 305 chronic specimens' envelope gene sequences collected from global cohorts including HIV-1 vaccine trial participants. HIITE precisely identified chronically infected individuals as being chronic with an error less than 1% and correctly classified 94% of recently infected individuals as being incident. Using a mixed-effect model, an incident specimen's time since infection was estimated from its single lineage diversity, showing 14% prediction error for time since infection. HIITE is the first algorithm to inform two key metrics from a single time point sequence sample. HIITE has the capacity for assessing not only population-level epidemic spread but also individual-level transmission events from a single survey, advancing HIV prevention and intervention programs. Availability and implementation: Web-based HIITE and source code of HIITE are available at http://www.hayounlee.org/software.html. Supplementary information: Supplementary data are available at Bioinformatics online.

SPMM: estimating infection duration of multivariant HIV-1 infections.

MOTIVATION: Illustrating how HIV-1 is transmitted and how it evolves in the following weeks is an important step for developing effective vaccination and prevention strategies. It is currently possible through DNA sequencing to account for the diverse array of viral strains within an infected individual. This provides an unprecedented opportunity to pinpoint when each patient was infected and which viruses were transmitted. RESULTS: Here we develop a mathematical tool for early HIV-1 evolution within a subject whose infection originates either from a single or multiple viral variants. The shifted Poisson mixture model (SPMM) provides a quantitative guideline for segregating viral lineages, which in turn enables us to assess when a subject was infected. The infection duration estimated by SPMM showed a statistically significant linear relationship with that by Fiebig laboratory staging (P = 0.00059) among 37 acutely infected subjects. Our tool provides a functional approach to understanding early genetic diversity, one of the most important parameters for deciphering HIV-1 transmission and predicting the rate of disease progression. AVAILABILITY AND IMPLEMENTATION: SPMM, webserver, is available at http://www.hayounlee.org/web-tools.html. CONTACT: hayoun@usc.edu SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Molecular clock of HIV-1 envelope genes under early immune selection.

BACKGROUND: The molecular clock hypothesis that genes or proteins evolve at a constant rate is a key tool to reveal phylogenetic relationships among species. Using the molecular clock, we can trace an infection back to transmission using HIV-1 sequences from a single time point. Whether or not a strict molecular clock applies to HIV-1's early evolution in the presence of immune selection has not yet been fully examined. RESULTS: We identified molecular clock signatures from 1587 previously published HIV-1 full envelope gene sequences obtained since acute infection in 15 subjects. Each subject's sequence diversity linearly increased during the first 150 days post infection, with rates ranging from [Formula: see text] to [Formula: see text] with a mean of [Formula: see text] per base per day. The rate of diversification for 12 out of the 15 subjects was comparable to the neutral evolution rate. While temporal diversification was consistent with evolution patterns in the absence of selection, mutations from the founder virus were highly clustered on statistically identified selection sites, which diversified more than 65 times faster than non-selection sites. By mathematically quantifying deviations from the molecular clock under various selection scenarios, we demonstrate that the deviation from a constant clock becomes negligible as multiple escape lineages emerge. The most recent common ancestor of a virus pair from distinct escape lineages is most likely the transmitted founder virus, indicating that HIV-1 molecular dating is feasible even after the founder viruses are no longer detectable. CONCLUSIONS: The ability of HIV-1 to escape from immune surveillance in many different directions is the driving force of molecular clock persistence. This finding advances our understanding of the robustness of HIV-1's molecular clock under immune selection, implying the potential for molecular dating.

Precision detection of recent HIV infections using high-throughput genomic incidence assay.

HIV incidence is a key measure for tracking disease spread and identifying populations and geographic regions where new infections are most concentrated. The HIV sequence population provides a robust signal for the stage of infection. Large-scale and high-precision HIV sequencing is crucial for effective genomic incidence surveillance. We produced 1,034 full-length envelope gene sequences from a seroconversion cohort by conducting HIV microdrop sequencing and measuring the genomic incidence assay's genome similarity index (GSI) dynamics. The measured dynamics of 9 of 12 individuals aligned with the GSI distribution estimated independently using 417 publicly available incident samples. We enhanced the capacity to identify individuals with recent infections, achieving predicted detection accuracies of 92% (89%-94%) for cases within 6 months and 81% (74%-87%) for cases within 9 months. These accuracy levels agreed with the observed detection accuracy intervals of an independent validation data set. Additionally, we produced 131 full-length envelope gene sequences from eight individuals with chronic HIV infection. This analysis confirmed a false recency rate (FRR) of 0%, which was consistent with 162 publicly available chronic samples. The mean duration of recent infection (MDRI) was 238 (209-267) days, indicating an 83% improvement in performance compared to current recent infection testing algorithms. The shifted Poisson mixture model was then used to estimate the time since infection, and the model estimates showed an 88% consistency with the days post infection derived from HIV RNA test dates and/or seroconversion dates. HIV microdrop sequencing provides unique prospects for large-scale incidence surveillance using high-throughput sequencing. IMPORTANCE Accurate identification of recently infected individuals is vital for prioritizing specific populations for interventions, reducing onward transmission risks, and optimizing public health services. However, current HIV-specific antibody-based methods have not been satisfactory in accurately identifying incident cases, hindering the use of HIV recency testing for prevention efforts and partner protection. Genomic incidence assays offer a promising alternative for identifying recent infections. In our study, we used microdroplet technologies to produce a large number of complete HIV envelope gene sequences, enabling the accurate detection of early infection signs. We assessed the dynamics of the incidence assay's metrics and compared them with statistical models. Our approach demonstrated high accuracy in identifying individuals with recent infections, achieving predicted detection rates exceeding 90% within 6 months and over 80% within 9 months of infection. This high-resolution method holds significant potential for enhancing the effectiveness of HIV incidence screening for case-based surveillance in public health initiatives.

Postnatal methylmercury exposure and neurodevelopmental outcomes at 7 years of age in the Seychelles Child Development Study Nutrition Cohort 2.

BACKGROUND: Consumption of fish yields many nutritional benefits, but also results in exposure to methylmercury (MeHg). The developing brain is known to be particularly susceptible to MeHg toxicity in high doses. However, the potential impact of low-level environmental exposure from fish consumption on children's neurodevelopment remains unclear. METHODS: We investigated postnatal MeHg exposure at 7 years and its association with a battery of 17 neurodevelopmental outcomes in a subset of children (n = 376) from 1535 enrolled mother-child pairs in Nutrition Cohort 2 of the Seychelles Child Development Study (SCDS NC2). Each outcome was modeled in relation to postnatal MeHg exposure using linear regression, adjusting for prenatal MeHg exposure, levels of maternal polyunsaturated fatty acids (PUFA), and several other covariates known to be associated with neurodevelopmental outcomes. RESULTS: Median postnatal MeHg exposure at 7 years was 2.5 ppm, while the median prenatal MeHg exposure was 3.5 ppm. We found no statistically significant associations between postnatal MeHg exposure and any of the 17 neurodevelopmental outcomes after adjusting for prenatal MeHg exposure and other covariates. CONCLUSIONS: These findings are consistent with previous cross-sectional analyses of the SCDS Main Cohort. Continued follow-up of the entire NC2 cohort at later ages with repeated exposure measures is needed to further confirm these findings.

KEAP1 polymorphisms and neurodevelopmental outcomes in children with exposure to prenatal MeHg from the Seychelles Child Development Study Nutrition Cohort 2.

BACKGROUND: Humans differ in the metabolism of the neurotoxicant methyl mercury (MeHg). This variation may be partially due to variation in genes encoding the transcription factor Nuclear factor E2-related factor 2 (NRF2) and its negative regulator Kelch-like ECH-Associated Protein 1 (KEAP1), which regulate glutathione and related transporter and antioxidant proteins that play a role in the metabolism and neurotoxicity of MeHg. AIM: To elucidate a potential risk from genetic variation in NFE2L2 (encoding NRF2) and KEAP1 toward prenatal mercury exposure and child neurodevelopmental outcomes at 20 months and 7 years of age in a population with variable prenatal exposure to MeHg from maternal fish consumption. MATERIAL AND METHODS: Nutrition Cohort 2 is a mother-child cohort in the Republic of Seychelles. Children were genotyped for NFE2L2 (rs2364723, rs13001694) and KEAP1 (rs8113472, rs9676881) polymorphisms (N = 1285 after removing siblings). Total mercury (Hg) was measured in cord blood as a biomarker for prenatal MeHg exposure. Child neurodevelopmental outcomes included the Bayley Scales of Infant Development II administered at 20 months of age, and outcomes across multiple neurodevelopmental domains from 14 tests administered in children and 3 instruments completed by parents when children were 7 years of age. RESULTS: The mean cord blood MeHg concentration was 34 (95% CI 11, 75) µg/L. None of the four polymorphisms had a significant association (p < 0.05) with either cord MeHg or neurodevelopmental test results at 20 months. There were no significant associations between either NFE2L2 polymorphism and any developmental test scores. At 7 years, children carrying KEAP1 rs8113472 CA showed significantly worse performance on psychomotor function than children with the CC variant (finger tapping, dominant hand: ß - 1.19, SE 0.34; finger tapping, non-dominant hand: ß - 0.92, SE 0.31) and worse social communication (SCQ Total: ß 0.65, SE 0.27). Children carrying rs8113472 AA, versus children with CC, showed significantly better performance on social communication (SRS Total: ß - 8.88, SE 3.60). Children carrying KEAP1 rs9676881 AG, versus children with GG, showed significantly worse performance on psychomotor function (trailmaking A time: ß 8.66, SE 3.37) and cognition (KBIT Matrices: ß - 0.96, SE 0.36). CONCLUSION: No associations between NFE2L2 and KEAP1 polymorphisms and MeHg concentration were identified. However, at 7 years, KEAP1 polymorphisms were associated with differences in neurodevelopmental outcomes in children from a population with high fish intake.

Radiation Therapy Exacerbates Tumor-Promoting Innervation and Nerve Signaling in Rectal Cancer.

PURPOSE: Many solid tumors present with perineural invasion (PNI), and innervation correlates with worsened prognosis. The effects that commonly administered therapies such as radiation therapy (RT) have on PNI status remain unknown. We investigated the contribution of RT on the nervous system and elucidated the implications that increased nerve signaling can have on tumor burden using our previously developed orthotopic murine model of rectal cancer (RC) and our targeted and clinically relevant short-course RT (SCRT) regimen. METHODS: Medical charts for patients with RC treated at the Wilmot Cancer Institute were obtained and PNI status was analyzed. Human data were accompanied by an orthotopic murine model of RC. Briefly, luciferase-expressing murine colon-38 (MC38-luc) tumor cells were injected orthotopically into the rectal wall of C57BL6 mice. Targeted SCRT (5 gray (Gy) per fraction for 5 consecutive fractions) was administered to the tumor. Intratumoral innervation was determined by immunohistochemistry (IHC), local norepinephrine (NE) concentration was quantified by enzyme-linked immunosorbent assay (ELISA), and ß2-adrenergic receptor (B2AR) expression was assessed by flow cytometry. Chronic NE signaling was mirrored by daily isoproterenol treatment, and the effect on tumor burden was determined by overall survival, presence of metastatic lesions, and tumor size. Isoproterenol signaling was inhibited by administration of propranolol. RESULTS: Human RC patients with PNI have decreased overall survival compared with patients without PNI. In our mouse model, SCRT induced the expression of genes involved in neurogenesis, increased local NE secretion, and upregulated B2AR expression. Treating mice with isoproterenol resulted in decreased overall survival, increased rate of metastasis, and reduced SCRT efficacy. Interestingly, the isoproterenol-induced decrease in SCRT efficacy could be abrogated by blocking the BAR through the use of propranolol, suggesting a direct role of BAR stimulation on impairing SCRT responses. CONCLUSIONS: Our results indicate that while SCRT is a valuable treatment, it is accompanied by adverse effects on the nervous system that may impede the efficacy of therapy and promote tumor burden. Therefore, we could speculate that therapies aimed at targeting this signaling cascade or impairing nerve growth in combination with SCRT may prove beneficial in future cancer treatment.

New insights into the responder/nonresponder divide in rectal cancer: Damage-induced Type I IFNs dictate treatment efficacy and can be targeted to enhance radiotherapy.

Rectal cancer ranks as the second leading cause of cancer-related deaths. Neoadjuvant therapy for rectal cancer patients often results in individuals that respond well to therapy and those that respond poorly, requiring life-altering excision surgery. It is inadequately understood what dictates this responder/nonresponder divide. Our major aim is to identify what factors in the tumor microenvironment drive a fraction of rectal cancer patients to respond to radiotherapy. We also sought to distinguish potential biomarkers that would indicate a positive response to therapy and design combinatorial therapeutics to enhance radiotherapy efficacy. To address this, we developed an orthotopic murine model of rectal cancer treated with short course radiotherapy that recapitulates the bimodal response observed in the clinic. We utilized a robust combination of transcriptomics and protein analysis to identify differences between responding and nonresponding tumors. Our mouse model recapitulates human disease in which a fraction of tumors respond to radiotherapy (responders) while the majority are nonresponsive. We determined that responding tumors had increased damage-induced cell death, and a unique immune-activation signature associated with tumor-associated macrophages, cancer-associated fibroblasts, and CD8+ T cells. This signature was dependent on radiation-induced increases of Type I Interferons (IFNs). We investigated a therapeutic approach targeting the cGAS/STING pathway and demonstrated improved response rate following radiotherapy. These results suggest that modulating the Type I IFN pathway has the potential to improve radiation therapy efficacy in RC.

Local Delivery of SBRT and IL12 by mRNA Technology Overcomes Immunosuppressive Barriers to Eliminate Pancreatic Cancer.

The immunosuppressive milieu in pancreatic cancer (PC) is a significant hurdle to treatments, resulting in survival statistics that have barely changed in 5 decades. Here we present a combination treatment consisting of stereotactic body radiation therapy (SBRT) and IL-12 mRNA lipid nanoparticles delivered directly to pancreatic murine tumors. This treatment was effective against primary and metastatic models, achieving cures in both settings. IL-12 protein concentrations were transient and localized primarily to the tumor. Depleting CD4 and CD8 T cells abrogated treatment efficacy, confirming they were essential to treatment response. Single cell RNA sequencing from SBRT/IL-12 mRNA treated tumors demonstrated not only a complete loss of T cell exhaustion, but also an abundance of highly proliferative and effector T cell subtypes. SBRT elicited T cell receptor clonal expansion, whereas IL-12 licensed these cells with effector function. This is the first report demonstrating the utility of SBRT and IL-12 mRNA in PC. Statement of significance: This study demonstrates the use of a novel combination treatment consisting of radiation and immunotherapy in murine pancreatic tumors. This treatment could effectively treat local and metastatic disease, suggesting it may have the potential to treat a cancer that has not seen a meaningful increase in survival in 5 decades.

Grafted and Vanishing Random Subspaces.

The Random Subspace Method (RSM) is an ensemble procedure in which each constituent learner is constructed using a randomly chosen subset of the data features. Regression trees are ideal candidate learners in RSM ensembles. By constructing trees upon different feature subsets, RSM reduces correlation between trees resulting in a stronger ensemble. Furthermore, it lessens computational burden by only considering a subset of the features when building each tree. Despite its apparent advantages, RSM has a notable drawback. In some instances a randomly chosen subspace may lack informative features. This is especially true in situations in which the number of truly informative variables is small relative to the total number of variables. Trees that are constructed using feature subsets lacking informative features can be damaging to the ensemble. Here we present Grafted Random Subspaces (GRS) and Vanishing Random Subspaces (VRS), two novel ensemble procedures designed to remedy the aforementioned drawback by reusing information across trees. Both techniques borrow from RSM by growing individual trees on randomly selected feature subsets. For each tree in a GRS ensemble, the most important variable is identified and guaranteed inclusion into the next q feature subsets. This allows GRS to recycle a promising feature from one tree across several successive trees, effectively grafting the variable into the next q active subsets. In the VRS procedure the least important feature is guaranteed exclusion from the next q feature subsets. This creates a more enriched pool of candidate variables from which the successive feature subsets are drawn.

Contribution of child ABC-transporter genetics to prenatal MeHg exposure and neurodevelopment.

BACKGROUND: There is emerging evidence that exposure to prenatal methylmercury (MeHg) from maternal fish consumption during pregnancy can differ between individuals due to genetic variation. In previous studies, we have reported that maternal polymorphisms in ABC-transporter genes were associated with maternal hair MeHg concentrations, and with children's early neurodevelopmental tests. In this study, we add to these findings by evaluating the contribution of genetic variation in children's ABC-transporter genes to prenatal MeHg exposure and early child neurodevelopmental tests. METHODS: We genotyped six polymorphisms (rs2032582, rs10276499 and rs1202169 in ABCB1; rs11075290 and rs215088 in ABCC1; rs717620 in ABCC2) in DNA from cord blood and maternal blood of the Seychelles Child Development Study Nutrition Cohort 2. We determined prenatal MeHg exposure by measuring total mercury (Hg) in cord blood by atomic fluorescence spectrometry. We assessed neurodevelopment in children at approximately 20 months using the Bayley Scales of Infant Development (BSID-II). We used linear regression models to analyze covariate-adjusted associations of child genotype with cord MeHg and BSID-II outcomes (Mental Developmental and Psychomotor Developmental Indexes). We also evaluated interactions between genotypes, cord MeHg, and neurodevelopmental outcomes. All models were run with and without adjustment for maternal genotype. RESULTS: Of the six evaluated polymorphisms, only ABCC1 rs11075290 was associated with cord blood MeHg; children homozygous for the T-allele had on average 29.99 µg/L MeHg in cord blood while those homozygous for the C-allele had on average 38.06 µg/L MeHg in cord blood (p < 0.001). No polymorphisms in the children were associated with either subscale of the BSID. However, the association between cord MeHg and the Mental Developmental Index (MDI) of the BSID differed significantly across the three genotypes of ABCB1 rs10276499 (2df F-test, p = 0.045). With increasing cord MeHg, the MDI decreased (slope=-0.091, p = 0.014) among children homozygous for the rare C-allele. CONCLUSIONS: These findings support the possibility that child ABC genetics might influence prenatal MeHg exposure.