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Background: Adoptive cell therapy (ACT) with tumor-infiltrating lymphocytes (TIL) is a personalized immunotherapy. The efficacy of TIL-ACT has been demonstrated prospectively in patients with advanced melanoma but is not limited to melanoma patients. Many patients are refractory to TIL-ACT, however, or their cancer becomes resistant. Combining anti-programmed cell death protein 1 (anti-PD-1) with TIL-ACT to antagonize the immunosuppressive tumor microenvironment may synergize to enhance the antitumor potential. Material and methods: We set up the BaseTIL trial (NCT04165967), a single-center investigator-initiated phase I trial, to test feasibility and safety of TIL-ACT followed by PD-1 blockade in patients with advanced cutaneous melanoma with disease progression after at least one line of anti-PD-1. TIL-ACT included tumor collection, ex vivo TIL expansion, lymphodepletion with cyclophosphamide and fludarabine, TIL transfer, and in vivo TIL stimulation with interleukin 2 (125 000 IU/kg, 10 days). TIL-ACT was followed by nivolumab treatment for a maximum of 2 years. Nine patients were planned for inclusion. Results: Between 2020 and 2022, we enrolled 11 patients and 9 underwent a TIL transfer (median transfused cell number: 66.25 × 109). Two patients did not start lymphodepletion. Nine patients received at least 1 dose of interleukin 2 (median number: 10; range, 1-10), seven started nivolumab (median number: 5; range, 2-23). All patients had hematologic adverse events (AEs). Most common non-hematologic AEs were fever and cytokine release syndrome. No nivolumab-associated AEs of ≥ grade 2 occurred. The objective response rate to TIL-ACT was 22% (2/9, 2 partial remission). Conclusions: TIL-ACT with nivolumab is feasible and safe. Larger trials are needed to further determine the efficacy of this combination.

Folate is an essential mediator in one-carbon metabolism, which provides methyl groups for DNA synthesis and methylation. The availability of active methyl groups can be influenced by the uptake of folic acid. We conducted a randomized intervention trial to test the influence of folic acid supplementation on DNA methylation in an unfortified population in Germany. A total of 16 healthy male volunteers (age range 23–61 y) were randomized to receive either 400 μg (n = 9) or 800 μg (n = 7) folic acid supplements daily for 8 weeks. Infinium Human Methylation 450K BeadChip Microarrays were used to assay site-specific DNA methylation across the genome. Microarray analyses were conducted on PBL DNA. We estimated several epigenetic clocks and mean DNA methylation across all autosomal probes on the array. AgeAccel was estimated as the residual variation in each metric. In virtually all participants, both serum and red blood cell (RBC) folate increased successively throughout the trial period. Participants with a larger increase in RBC folate had a larger increase in DNAmAge AgeAccel (Spearman Rho: 0.56, p-value = 0.03). No notable changes in the methylome resulting from the folic acid supplementation emerged. In this population with adequate folate levels derived from diet, an increase in RBC folate had a modest impact on the epigenetic clock predicting chronologic age.

Abstract Background Low skeletal muscle mass (myopenia) is common in cancer populations and is associated with functional decline and mortality, but prior oncology studies did not assess total body skeletal muscle mass. Instead, they measured surrogates such as cross‐sectional area (CSA) of skeletal muscle at L3 from computed tomography (CT) or appendicular lean mass (ALM) from dual‐energy X‐ray absorptiometry (DXA). D3‐creatine (D3Cr) dilution is a non‐invasive method to assess total body skeletal muscle mass, which has been examined in a variety of populations but not in cancer. To compare the associations of D3Cr muscle mass, CT CSA, and DXA ALM with myopenia and physical function, we conducted a cross‐sectional study among 119 patients with colon cancer (2018–2022). Methods For each technique (D3Cr, CT and DXA), myopenia was defined as the lowest sex‐specific quartile of its measurement. Physical function was measured by the short physical performance battery and grip strength. We calculated Pearson correlations (r) among three techniques, computed Cohen's kappa coefficients (κ) to assess the agreement of myopenia, and estimated Pearson correlations (r) of three techniques with physical function. All analyses were sex‐specific. Results Sixty‐one (51.3%) participants were male, the mean (standard deviation) age was 56.6 (12.9) years, and most (68.9%) had high physical function (short physical performance battery: ≥11 points). Correlations and myopenia agreement among three techniques were greater in men than women; for example, regarding D3Cr muscle mass versus CT CSA, r was 0.73 (P

Signals are exchanged at all stages of the arbuscular mycorrhizal (AM) symbiosis between fungi and their host plants. Root-exuded strigolactones are well-known early symbiotic cues, but the role of other phytohormones as interkingdom signals has seldom been investigated. Here we focus on ethylene and cytokinins, for which candidate receptors have been identified in the genome of the AM fungus Rhizophagus irregularis. Ethylene is known from the literature to affect asymbiotic development of AM fungi, and in the present study, we found that three cytokinin forms could stimulate spore germination in R. irregularis. Heterologous complementation of a Saccharomyces cerevisiae mutant strain with the candidate ethylene receptor RiHHK6 suggested that this protein can sense and transduce an ethylene signal. Accordingly, its N-terminal domain expressed in Pichia pastoris displayed saturable binding to radiolabeled ethylene. Thus, RiHHK6 displays the expected characteristics of an ethylene receptor. In contrast, the candidate cytokinin receptor RiHHK7 did not complement the S. cerevisiae mutant strain or Medicago truncatula cytokinin receptor mutants and seemed unable to bind cytokinins, suggesting that another receptor is involved in the perception of these phytohormones. Taken together, our results support the hypothesis that AM fungi respond to a range of phytohormones and that these compounds bear multiple functions in the rhizosphere beyond their known roles as internal plant developmental regulators. Our analysis of two phytohormone receptor candidates also sheds new light on the possible perception mechanisms in AM fungi. [Graphic: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

The COVID-19 pandemic has emphasized the importance of designing effective methods for remote teaching. At the University of California, Berkeley, and the University of California, Santa Cruz, instructors adapted to the necessity of remote laboratory instruction by creating choose-your-own-adventure-style video-based online experiments introduced to thousands of students across 11 different courses. These experiments are designed to provide students with the opportunity to make and receive feedback on experimental decisions and learn from common mistakes that they may have encountered in hands-on laboratory instruction. Students' and instructors' impressions of the online experiments and student learning outcomes in both online and traditional laboratory courses were assessed using surveys, focus groups, and interviews via a mixed-methods approach. Though most respondents (79%) did not agree that online laboratory instruction was as effective as in-person instruction, the majority agreed that the online experiments were clear and easy to follow (75%), interesting and engaging (52%), and helpful for learning about lab techniques (70%) and the concepts underlying these techniques (77%). Many also mentioned benefits of online laboratory instruction, including flexibility in scheduling and an increased focus on conceptual learning. Assessments of student learning also suggested that students who took the course online learned as much conceptually as students who had previously completed the course in-person. The results of this study highlight the positive and negative aspects of this type of interactive online laboratory instruction, which could help inform the design of future lab experiences whether they take place in an online, hybrid, or in-person environment.

Biochemical premature adrenarche is defined by elevated serum DHEAS [≥40 μg/dL] before age 8 y in girls. This condition is receiving more attention due to its association with obesity, hyperinsulinemia, dyslipidemia, and polycystic ovary syndrome. Nevertheless, the link between early androgen excess and these risk factors remains unknown. Epigenetic modifications, and specifically DNA methylation, have been associated with the initiation and progression of numerous disorders, including obesity and insulin resistance. The aim of this study was to determine if prepubertal androgen exposure is associated with a different methylation profile in pubertal girls. Eighty-six healthy girls were studied. At age 7 y, anthropometric measurements were begun and DHEAS levels were determined. Girls were classified into Low DHEAS (LD) [

Introduction: High mammographic density is among the strongest and most established predictors for breast cancer risk. Puberty, the period during which breasts undergo exponential mammary growth, is considered one of the critical stages of breast development for environmental exposures. Benzylbutyl phthalate (BBP) and perfluorooctanoic acid (PFOA) are pervasive endocrine disrupting chemicals that may increase hormone-sensitive cancers. Evaluating the potential impact of BBP and PFOA exposure on pubertal breast density is important to our understanding of early-life environmental influences on breast cancer etiology. Objective: To prospectively assess the effect of biomarker concentrations of monobenzyl phthalate (MBzP) and PFOA at specific pubertal window of susceptibility (WOS) on adolescent breast density. Method: This study included 376 Chilean girls from the Growth and Obesity Cohort Study with data collection at four timepoints: Tanner breast stages 1 (B1) and 4 (B4), 1- year post- menarche (1YPM) and 2-years post-menarche (2YPM). Dual-energy X-ray absorptiometry was used to assess the absolute fibroglandular volume (FGV) and percent breast density (%FGV) at 2YPM. We used concentrations of PFOA in serum and MBzP in urine as an index of exposure to PFOA and BBP, respectively. Parametric G-formula was used to estimate the time-specific effects of MBzP and PFOA on breast density. The models included body fat percentage as a time-varying confounder and age, birthweight, age at menarche, and maternal education as fixed covariates. Results: A doubling of serum PFOA concentration at B4 resulted in a non-significant increase in absolute FGV (β:11.25, 95% confidence interval (CI): −0.28, 23.49)), while a doubling of PFOA concentration at 1YPM resulted in a decrease in % FGV (β:-4.61, 95% CI: −7.45, −1.78). We observed no associations between urine MBzP and breast density measures. Conclusion: In this cohort of Latina girls, PFOA serum concentrations corresponded to a decrease in % FGV. No effect was observed between MBzP and breast density measures across pubertal WOS.

Microtubules are a ubiquitous eukaryotic cytoskeletal element typically consisting of 13 protofilaments arranged in a hollow cylinder. Using CryoEM and subvolume averaging, Ferreira and Pražák et al. show that Plasmodium does not adhere to a single microtubule structure. Instead, the cytoskeleton changes substantially to produce a unique, fit for purpose structure and organisation at each stage of its life cycle.

Park et al. show that upon exposure to ethylene gas, CTR1, which normally has a negative effect on ethylene signaling, translocates into the nucleus and enhances the plant’s response to ethylene by stabilizing the EIN3 transcription factor.

A meta-analysis of pre-pregnancy maternal body mass index (ppBMI) and placental DNA methylation from 2631 mother-child pairs identifies 27 CpG sites associated with ppBMI, providing insight into how maternal obesity could be associated with metabolic health outcomes in offspring.

“Star” survey designs have become an increasingly popular alternative to parallel line designs in fisheries-independent sampling of areas with isolated fish aggregations, such as artificial reefs, seamounts, fish aggregating devices, and spawning aggregation sites. In this study, we simulated three scenarios of fish aggregating around a feature of interest with variations in the size and complexity of aggregations as well as their location relative to the habitat feature. Simulated and empirical data representing goliath grouper (Epinephalus itajara) spawning aggregations at artificial reefs were utilized as a case study, and scenarios were generated in relation to both a single habitat feature and a reef complex with multiple structures. Seven variations of survey design using both star and parallel transects were examined and compared by geostatistical and generalized additive models (GAMs) to identify the most robust approach to quantify fish aggregations in each scenario. In most scenarios, precision in the mean and variability of backscatter estimates is not significantly affected by the number of transects passing over the habitat feature as long as at least one pass is made. Estimation error is minimized using the GAM approach, and is further reduced when sampling variance is high, which was better accomplished by parallel designs overall. These results will help inform surveyors on the best overall approach to improve precision in quantifying fish aggregations given basic knowledge of their behavior around an established habitat feature and help them to adapt their survey designs based on common difficulties in sampling these populations simulated below.

Coastal fish communities are under increasing levels of stress associated with climate variation and anthropogenic activities. However, the high degree of behavioral plasticity of many species within these communities allow them to cope with altered environmental conditions to some extent. Here, we combine meteorological information, data from hydroacoustic surveys, and recordings of goliath grouper sound production to examine the response of coastal fish communities to heavy rainfall events in South Florida, USA, that resulted in the release of excess storm water into surrounding estuaries and coastal waters. We observed a nearly 12,000% increase in water column acoustic backscatter following a heavy rainfall event of September 16th, 2015. Interestingly, estimates of school backscatter, a proxy for biomass, increased by 172% with the onset of the perturbation. Schooling fish density also increased by 182%, as did acoustically derived estimates of mean schooling fish length (21%). Following the perturbed period, school backscatter decreased by 406%, along with schooling density (272%), and mean schooling fish length (35%). Hydrophone and hydroacoustic data also revealed that goliath grouper (Epinephelus itajara) spawning aggregations were persistent in the region throughout the duration of the study and continued to exhibit courtship behavior during the perturbed period. Our observations demonstrate the high level of resistance common in coastal species but raises new questions regarding the threshold at which fish communities and reproductive activities are disrupted. As coastal land use continues to increase, and the effects of global climate change become more pronounced, more Before-After Control Impact (BACI) studies will provide improved insight into the overall response of nearshore communities to future perturbations and the cumulative effect of repeated perturbations over extended periods.

Tumor subtype and menopausal status are strong predictors of breast cancer (BC) prognosis. We aimed to find and validate subtype- or menopausal-status-specific changes in tumor DNA methylation (DNAm) associated with all-cause mortality or BC progression. Associations between site-specific tumor DNAm and BC prognosis were estimated among The Cancer Genome Atlas participants (n = 692) with Illumina Infinium HumanMethylation450 BeadChip array data. All-cause mortality and BC progression were modeled using Cox proportional hazards models stratified by tumor subtypes, adjusting for age, race, stage, menopausal status, tumor purity, and cell type proportion. Effect measure modification by subtype and menopausal status were evaluated by incorporating a product term with DNAm. Site-specific inference was used to identify subtype- or menopausal-status-specific differentially methylated regions (DMRs) and functional pathways. The validation of the results was carried out on an independent dataset (GSE72308; n = 180). We identified a total of fifteen unique CpG probes that were significantly associated (P≤1×10−7 with survival outcomes in subtype- or menopausal-status-specific manner. Seven probes were associated with overall survival (OS) or progression-free interval (PFI) for women with luminal A subtype, and four probes were associated with PFI for women with luminal B subtype. Five probes were associated with PFI for post-menopausal women. A majority of significant probes showed a lower risk of OS or BC progression with higher DNAm. We identified subtype- or menopausal-status-specific DMRs and functional pathways of which top associated pathways differed across subtypes or menopausal status. None of significant probes from site-specific analyses met genome-wide significant level in validation analyses while directions and magnitudes of coefficients showed consistent pattern. We have identified subtype- or menopausal-status-specific DNAm biomarkers, DMRs and functional pathways associated with all-cause mortality or BC progression, albeit with limited validation. Future studies with larger independent cohort of non-post-menopausal women with non-luminal A subtypes are warranted for identifying subtype- and menopausal-status-specific DNAm biomarkers for BC prognosis.

ABSTRACT Mature gametocytes of Plasmodium falciparum display a banana (falciform) shape conferred by a complex array of subpellicular microtubules (SPMT) associated with the inner membrane complex (IMC). Microtubule-associated proteins (MAPs) define MT populations and modulate interaction with pellicular components. Several MAPs have been identified in Toxoplasma gondii, and homologues can be found in the genomes of Plasmodium species, but the function of these proteins for asexual and sexual development of malaria parasites is still unknown. Here, we identified a novel subpellicular MAP, termed SPM3, that is conserved within the genus Plasmodium, especially within the subgenus Laverania, but absent in other Apicomplexa. Conditional knockdown and targeted gene disruption of Pfspm3 in Plasmodium falciparum cause severe morphological defects during gametocytogenesis, leading to round, nonfalciform gametocytes with an aberrant SPMT pattern. In contrast, Pbspm3 knockout in Plasmodium berghei, a species with round gametocytes, caused no defect in gametocytogenesis, but sporozoites displayed an aberrant motility and a dramatic defect in invasion of salivary glands, leading to a decreased efficiency in transmission. Electron microscopy revealed a dissociation of the SPMT from the IMC in Pbspm3 knockout parasites, suggesting a function of SPM3 in anchoring MTs to the IMC. Overall, our results highlight SPM3 as a pellicular component with essential functions for malaria parasite transmission. IMPORTANCE A key structural feature driving the transition between different life cycle stages of the malaria parasite is the unique three-membrane pellicle, consisting of the parasite plasma membrane (PPM) and a double membrane structure underlying the PPM termed the inner membrane complex (IMC). Additionally, there are numerous linearly arranged intramembranous particles (IMPs) linked to the IMC, which likely link the IMC to the subpellicular microtubule cytoskeleton. Here, we identified, localized, and characterized a novel subpellicular microtubule-associated protein unique to the genus Plasmodium. The knockout of this protein in the human-pathogenic species P. falciparum resulted in malformed gametocytes and aberrant microtubules. We confirmed the microtubule association in the P. berghei rodent malaria homologue and show that its knockout results in a perturbed microtubule architecture, aberrant sporozoite motility, and decreased transmission efficiency.

During the COVID-19 pandemic, many public schools across the United States shifted from fully in-person learning to alternative learning modalities such as hybrid and fully remote learning. In this study, data from 14,688 unique school districts from August 2020 to June 2021 were collected to track changes in the proportion of schools offering fully in-person, hybrid and fully remote learning over time. These data were provided by Burbio, MCH Strategic Data, the American Enterprise Institute's Return to Learn Tracker and individual state dashboards. Because the modalities reported by these sources were incomplete and occasionally misaligned, a model was needed to combine and deconflict these data to provide a more comprehensive description of modalities nationwide. A hidden Markov model (HMM) was used to infer the most likely learning modality for each district on a weekly basis. This method yielded higher spatiotemporal coverage than any individual data source and higher agreement with three of the four data sources than any other single source. The model output revealed that the percentage of districts offering fully in-person learning rose from 40.3% in September 2020 to 54.7% in June of 2021 with increases across 45 states and in both urban and rural districts. This type of probabilistic model can serve as a tool for fusion of incomplete and contradictory data sources in order to obtain more reliable data in support of public health surveillance and research efforts.

Background: Epidemiologic evidence suggests that environmental factors acting as endocrine disrupting chemicals (EDCs) are associated with mammographic breast density and the risk of breast cancer. Exposure to EDCs during puberty, a period of rapid breast development, may affect susceptibility to breast carcinogenesis. Methods: In a cohort of 366 Chilean adolescents from the Growth and Obesity Cohort Study, we evaluated the relation between urinary concentrations of 15 suspected EDC biomarkers across three pubertal time points (Tanner breast stage 1 (B1), 4 (B4), and 1-year post-menarche) and breast fibroglandular volume (FGV; percent FGV [%FGV] and absolute FGV [aFGV]) and total breast volume (tBV) at 2-years post-menarche. We used linear mixed models to test differences in creatinine-corrected EDC biomarker concentrations at B4 and 1-year post-menarche compared to B1 and calculated intraclass correlation coefficients (ICC) of EDC concentrations across time points to appraise the consistency of measurements. We fit multivariable generalized estimating equations (GEEs) to evaluate windows of susceptibility for the association between log10-transformed EDCs and log10-transformed breast outcomes. GEEs were adjusted for age, body fat percentage, total caloric intake, and maternal education. Results: Urinary EDC biomarker concentrations highly varied across pubertal time points (ICC range 0.01–0.30). For 12 EDCs, biomarker concentrations decreased over time. Triclosan measured at 1-year post-menarche was inversely associated with %FGV at 2-years post-menarche (β = −0.025, 95 % confidence interval = −0.041, −0.008). Mono(2-ethyl-5-carboxypentyl) phthalate and the sum of di(2-ethylhexyl) phthalate metabolite concentrations at B4 were positively associated with aFGV and tBV at 2-years post-menarche. No measured phenols were associated with aFGV and tBV, while no measured parabens were associated with %FGV and aFGV. Conclusions: Our study suggests relatively high variability in EDC biomarker concentrations across the peripubertal time period. We also found evidence to suggest that there may be pubertal windows of susceptibility to select EDCs for the association with adolescent breast density.

Abstract Background Aging and inflammation are important components of Parkinson’s disease (PD) pathogenesis and both are associated with changes in hematopoiesis and blood cell composition. DNA methylation (DNAm) presents a mechanism to investigate inflammation, aging, and hematopoiesis in PD, using epigenetic mitotic aging and aging clocks. Here, we aimed to define the influence of blood cell lineage on epigenetic mitotic age and then investigate mitotic age acceleration with PD, while considering epigenetic age acceleration biomarkers. Results We estimated epigenetic mitotic age using the “epiTOC” epigenetic mitotic clock in 10 different blood cell populations and in a population-based study of PD with whole-blood. Within subject analysis of the flow-sorted purified blood cell types DNAm showed a clear separation of epigenetic mitotic age by cell lineage, with the mitotic age significantly lower in myeloid versus lymphoid cells (p = 2.1e-11). PD status was strongly associated with accelerated epigenetic mitotic aging (AccelEpiTOC) after controlling for cell composition (OR = 2.11, 95 % CI = 1.56, 2.86, p = 1.6e-6). AccelEpiTOC was also positively correlated with extrinsic epigenetic age acceleration, a DNAm aging biomarker related to immune system aging (with cell composition adjustment: R = 0.27, p = 6.5e-14), and both were independently associated with PD. Among PD patients, AccelEpiTOC measured at baseline was also associated with longitudinal motor and cognitive symptom decline. Conclusions The current study presents a first look at epigenetic mitotic aging in PD and our findings suggest accelerated hematopoietic cell mitosis, possibly reflecting immune pathway imbalances, in early PD that may also be related to motor and cognitive progression.

Abstract Rickettsial diseases (RDs) are transmitted to humans by ectoparasites, including ticks and fleas. Symptoms range from mild febrile illness, to severe disease or death. Doxycycline is the treatment of choice for patients of all ages; early treatment based on clinical diagnosis is critical to prevent severe outcomes. We conducted a descriptive analysis using insurance claims data captured by IBM MarketScan® research databases to describe demographics, treatment patterns, and outcomes of patients diagnosed with RDs in the United States during 2005–2017. Overall, 14,830 patients had a RD diagnosis during 2005–2017; 7,517 (50.7%) spotted fever rickettsiosis (SFR), 4,571 ( 30.8%) ehrlichiosis, 1,362 (9.2%) typhus group rickettsiosis (TGR), and 1,193 (8.0%) other rickettsial diseases. Among all patients diagnosed, 53.1% received doxycycline. Prescription rates varied by diagnosis and age; 24.1% of TGR and 61.1% of SFR patients received doxycycline; 23.9% of persons

Hybrid imaging systems, comprising PET and Compton camera modules, have recently gained in interest, due to their capability to simultaneously detect positron annihilation photons and γ-rays from single-photon emitting sources as also used in SPECT. A unique feature of such systems, however, is the capability to also be operated in a so called γ-PET mode. Here, specific β+- emitting radioisotopes (such as 44Sc, 1°C or 14O) are used to detect triple-coincidences between two annihilation γ-rays (in PET imaging) and a third, prompt photon (in Compton imaging), that is emitted by the deexcitation of the decay’s daughter nucleus. Consequently, an intersection between the line-of-response (LOR) and the Compton cone can be determined, which (in principle) allows to localize the photons’ emission vertices on a single decay basis. In practice, however, a few tens of events are required to localize a point source, which still results in a considerable sensitivity improvement compared to conventional PET imaging.For a proof-of-principle study, we used a pixelated GAGG crystal array (16 × 16 crystals; 1.45 × 1.45 × 6 mm3 crystal volume; 25 μm SPAD SiPMs as readout) as Compton camera scatterer and PET detectors, and a three-layered LYSO crystal array (1.2 × 1.2 × 6.66 mm3 crystal volume; 50 μm SPAD SiPMs as readout) as Compton camera absorber. We characterized the individual detector components with regard to their energy resolution and the capability to identify the various scintillator array’s individual crystals. Our first γ-PET prototype was tested in PET-only and Compton-only imaging mode, in which spatial resolutions of 3.2–3.5 mm FWHM (PET-only mode) and 14.4–19.3 mm FWHM (Compton-only mode at 1,274 keV) were achieved, respectively, using a22Na point source and 10 iterations of an ML-EM reconstruction algorithm. By using triple-coincidences in a γ-PET mode (event-wise intersection of the LOR and the Compton cone), we could demonstrate the capability of the prototype to perform a full 3D point source reconstruction using only 77 events.

Abstract Background Biological aging estimators derived from DNA methylation data are heritable and correlate with morbidity and mortality. Consequently, identification of genetic and environmental contributors to the variation in these measures in populations has become a major goal in the field. Results Leveraging DNA methylation and SNP data from more than 40,000 individuals, we identify 137 genome-wide significant loci, of which 113 are novel, from genome-wide association study (GWAS) meta-analyses of four epigenetic clocks and epigenetic surrogate markers for granulocyte proportions and plasminogen activator inhibitor 1 levels, respectively. We find evidence for shared genetic loci associated with the Horvath clock and expression of transcripts encoding genes linked to lipid metabolism and immune function. Notably, these loci are independent of those reported to regulate DNA methylation levels at constituent clock CpGs. A polygenic score for GrimAge acceleration showed strong associations with adiposity-related traits, educational attainment, parental longevity, and C-reactive protein levels. Conclusion This study illuminates the genetic architecture underlying epigenetic aging and its shared genetic contributions with lifestyle factors and longevity.

Abstract Background Systemic inflammation may play a role in shaping breast composition, one of the strongest risk factors for breast cancer. Pubertal development presents a critical window of breast tissue susceptibility to exogenous and endogenous factors, including pro-inflammatory markers. However, little is known about the role of systemic inflammation on adolescent breast composition and pubertal development among girls. Methods We investigated associations between circulating levels of inflammatory markers (e.g., interleukin-6 (IL-6), tumor necrosis factor receptor 2 (TNFR2), and C-reactive protein (CRP)) at Tanner stages 2 and 4 and breast composition at Tanner stage 4 in a cohort of 397 adolescent girls in Santiago, Chile (Growth and Obesity Cohort Study, 2006–2018). Multivariable linear models were used to examine the association between breast composition and each inflammatory marker, stratifying by Tanner stage at inflammatory marker measurement. Accelerated failure time models were used to evaluate the association between inflammatory markers concentrations at each Tanner stage and time to menarche. Results In age-adjusted linear regression models, a doubling of TNFR2 at Tanner 2 was associated with a 26% (95% CI 7–48%) increase in total breast volume at Tanner 4 and a 22% (95% CI 10–32%) decrease of fibroglandular volume at Tanner 4. In multivariable models further adjusted for body fatness and other covariates, these associations were attenuated to the null. The time to menarche was 3% (95% CI 1–5%) shorter among those in the highest quartile of IL-6 at Tanner 2 relative to those in the lowest quartile in fully adjusted models. Compared to those in the lowest quartile of CRP at Tanner 4, those in the highest quartile experienced 2% (95% CI 0–3%) longer time to menarche in multivariable models. Conclusions Systemic inflammation during puberty was not associated with breast volume or breast density at the conclusion of breast development among pubertal girls after adjusting for body fatness; however, these circulating inflammation biomarkers, specifically CRP and IL-6, may affect the timing of menarche onset.

The ever-increasing demand for phosphorus fertilisers for securing global food production, coupled with finite phosphate rock reserves, is one of the emerging problems in the world. Indeed, phosphate rock is listed as an EU critical raw material, triggering attention to find an alternative source to substitute the use of this limited resource. Cheese whey, characterized by a high content of organic matter and phosphorus, represents a promising feedstock for phosphorus recovery and recycling. An innovative application of a membrane system coupled with freeze concentration was assessed to recover phosphorus from cheese whey. The performances of a microfiltration membrane (0.2 µm) and an ultrafiltration (200 kDa) membrane were evaluated and optimized under different transmembrane pressures and crossflow velocities. Once the optimal operating conditions were determined, a pre-treatment including lactic acid acidification and centrifugation was applied to increase the permeate recovery. Finally, the efficiency of progressive freeze concentration for the treatment of the permeate obtained from the optimum conditions (UF 200 kDa with TMP of 3 bar, CFV of 1 m/s and lactic acid acidification) was evaluated at specific operating conditions (−5 °C and 600 rpm of stirring speed). Finally, 70% of phosphorus could be recovered from cheese whey using the coupled technology of the membrane system and freeze concentration. A phosphorus-rich product was obtained with high agronomic value, which constitutes a further step towards establishing a broader circular economy framework.

Background In the United States, national ecological studies suggest a positive impact of COVID-19 vaccination coverage on outcomes in adults. However, the national impact of the vaccination program on COVID-19 in children remains unknown. To determine the association of COVID-19 vaccination with U.S. case incidence, emergency department visits, and hospital admissions for pediatric populations during the Delta and Omicron periods. Methods We conducted an ecological analysis among children aged 5–17 and compared incidence rate ratios (RRs) of COVID-19 cases, emergency department visits, and hospital admissions by pediatric vaccine coverage, with jurisdictions in the highest vaccine coverage quartile as the reference. Results RRs comparing states with lowest pediatric vaccination coverage to the highest pediatric vaccination coverage were 2.00 and 0.64 for cases, 2.96 and 1.11 for emergency department visits, and 2.76 and 1.01 for hospital admissions among all children during the Delta and Omicron periods, respectively. During the 3-week peak period of the Omicron wave, only children aged 12–15 and 16–17 years in the states with the lowest versus highest coverage, had a significantly higher rate of emergency department visits (RR = 1.39 and RR = 1.34, respectively). Conclusions COVID-19 vaccines were associated with lower case incidence, emergency department visits and hospital admissions among children during the Delta period but the association was weaker during the Omicron period. Pediatric COVID-19 vaccination should be promoted as part of a program to decrease COVID-19 impact among children; however, vaccine effectiveness may be limited when available vaccines do not match circulating viral variants.

Abstract Background The influence of genetics on variation in DNA methylation (DNAme) is well documented. Yet confounding from population stratification is often unaccounted for in DNAme association studies. Existing approaches to address confounding by population stratification using DNAme data may not generalize to populations or tissues outside those in which they were developed. To aid future placental DNAme studies in assessing population stratification, we developed an ethnicity classifier, PlaNET (Placental DNAme Elastic Net Ethnicity Tool), using five cohorts with Infinium Human Methylation 450k BeadChip array (HM450k) data from placental samples that is also compatible with the newer EPIC platform. Results Data from 509 placental samples were used to develop PlaNET and show that it accurately predicts (accuracy = 0.938, kappa = 0.823) major classes of self-reported ethnicity/race (African: n = 58, Asian: n = 53, Caucasian: n = 389), and produces ethnicity probabilities that are highly correlated with genetic ancestry inferred from genome-wide SNP arrays (> 2.5 million SNP) and ancestry informative markers (n = 50 SNPs). PlaNET’s ethnicity classification relies on 1860 HM450K microarray sites, and over half of these were linked to nearby genetic polymorphisms (n = 955). Our placental-optimized method outperforms existing approaches in assessing population stratification in placental samples from individuals of Asian, African, and Caucasian ethnicities. Conclusion PlaNET provides an improved approach to address population stratification in placental DNAme association studies. The method can be applied to predict ethnicity as a discrete or continuous variable and will be especially useful when self-reported ethnicity information is missing and genotyping markers are unavailable.

Middle East respiratory syndrome coronavirus (MERS-CoV) shedding and antibody responses are not fully understood, particularly in relation to underlying medical conditions, clinical manifestations, and mortality. We enrolled MERS-CoV–positive patients at a hospital in Saudi Arabia and periodically collected specimens from multiple sites for real-time reverse transcription PCR and serologic testing. We conducted interviews and chart abstractions to collect clinical, epidemiologic, and laboratory information. We found that diabetes mellitus among survivors was associated with prolonged MERS-CoV RNA detection in the respiratory tract. Among case-patients who died, development of robust neutralizing serum antibody responses during the second and third week of illness was not sufficient for patient recovery or virus clearance. Fever and cough among mildly ill patients typically aligned with RNA detection in the upper respiratory tract; RNA levels peaked during the first week of illness. These findings should be considered in the development of infection control policies, vaccines, and antibody therapeutics.

The formation of fish spawning aggregations (FSAs) is an essential part of the life history of many economically important fish species; however, their status are often poorly described in the literature either due to their occurrence in remote locations, during seasons with unsafe ocean conditions, or because they move on space and time scales that are difficult to predict and validate. Even in areas that are relatively accessible and heavily fished, such as southeast Florida, regionally relevant information describing FSA dynamics is generally absent from the literature and unaccounted for in existing management plans. We propose that this can be attributed to the fact that information is often held by stakeholders or found in unpublished manuscripts and reports. These sources are not widely disseminated and are therefore difficult to locate and integrate into fisheries management decisions. In this paper, we present a case study demonstrating the value of regional data syntheses as a tool to improve management activities in southeast Florida. Specifically, we engaged with local stakeholders to collect reports of FSA occurrence, and used Web of Science queries to collate information describing the reproductive dynamics of locally occurring snapper and grouper species. Reports were combined with regional FSA literature and provided to managers as a support tool to anticipate FSA occurrence, and to guide policy development and future FSA research. Resource users identified 13 potential aggregations from five species, but Web of Science queries revealed a paucity of information. Echosounder, camera, and fisheries dependent surveys were then used to corroborate reportedly active cubera snapper (Lutjanus cyanopterus), hogfish (Lachnolaimus maximus), and gag grouper (Mycteroperca microlepis) aggregations. Variability in the spatiotemporal aspects of FSA occurrence make them difficult to study, but this may also explain how certain species have avoided detrimental impacts from aggregation fishing. These data represent a first step toward describing FSAs that have historically occurred in the Southeast Florida Coral Reef Ecosystem Conservation Area and can be used by managers to prioritize future research efforts focused on species or hotspots of multispecies activity along the northern extent of the Florida Reef Tract.

Abstract Management of manure and cereal rye (Secale cereale L.) has implications for crop production and nutrient cycling. This 1‐yr, full‐factorial experiment conducted in Pennsylvania quantified the effects of three management factors—(a) rye management (RyeM; early‐terminated cover crop [CC] vs. double crop harvested a week later [DC]), (b) manure application method (ManM; unincorporated broadcast manure [BM] vs. shallow disk injected manure [IM]), and (c) fall field operation prioritization (priority; manure priority [MP] manure application in late September with rye planting in mid‐October vs. rye priority [RP], rye planting in late September with manure application in early November)—on rye biomass, nutrient recovery, and forage yield and the effect of ManM and priority on DC forage nutritive value in a rye–corn (Zea mays L.) cropping sequence. This experiment was intended to be a repeated 2‐yr study, but due to MP treatment rye crop failure in the second year, the resulting 1‐yr dataset was analyzed to assess priority effects. Prioritizing rye planting in the fall with DC increased total forage production and apparent N recovery (ANR) when manure was broadcast. These results highlight the value of prioritizing fall rye planting in DC systems to increase rye spring biomass and nutrient recovery when manure is broadcast. More experiments should be conducted on fall field operation timing to develop reliable recommendations.

Observing multiple size classes of organisms, along with oceanographic properties and water mass origins, can improve our understanding of the drivers of aggregations, yet acquiring these measurements remains a fundamental challenge in biological oceanography. By deploying multiple biological sampling systems, from conventional bottle and net sampling to in situ imaging and acoustics, we describe the spatial patterns of different size classes of marine organisms (several microns to ∼10 cm) in relation to local and regional (m to km) physical oceanographic conditions on the Delaware continental shelf. The imaging and acoustic systems deployed included (in ascending order of target organism size) an imaging flow cytometer (CytoSense), a digital holographic imaging system (HOLOCAM), an In Situ Ichthyoplankton Imaging System (ISIIS, 2 cameras with different pixel resolutions), and multi-frequency acoustics (SIMRAD, 18 and 38 kHz). Spatial patterns generated by the different systems showed size-dependent aggregations and differing connections to horizontal and vertical salinity and temperature gradients that would not have been detected with traditional station-based sampling (∼9-km resolution). A direct comparison of the two ISIIS cameras showed composition and spatial patchiness changes that depended on the organism size, morphology, and camera pixel resolution. Large zooplankton near the surface, primarily composed of appendicularians and gelatinous organisms, tended to be more abundant offshore near the shelf break. This region was also associated with high phytoplankton biomass and higher overall organism abundances in the ISIIS, acoustics, and targeted net sampling. In contrast, the inshore region was dominated by hard-bodied zooplankton and had relatively low acoustic backscatter. The nets showed a community dominated by copepods, but they also showed high relative abundances of soft-bodied organisms in the offshore region where these organisms were quantified by the ISIIS. The HOLOCAM detected dense patches of ciliates that were too small to be captured in the nets or ISIIS imagery. This near-simultaneous deployment of different systems enables the description of the spatial patterns of different organism size classes, their spatial relation to potential prey and predators, and their association with specific oceanographic conditions. These datasets can also be used to evaluate the efficacy of sampling techniques, ultimately aiding in the design of efficient, hypothesis-driven sampling programs that incorporate these complementary technologies.

Abstract Background Estrogen receptor-α (ER-α) is a transcriptional regulator, which mediates estrogen-dependent breast development, as well as breast tumorigenesis. The influence of epigenetic regulation of ER-α on adolescent breast composition has not been previously studied and could serve as a marker of pubertal health and susceptibility to breast cancer. We investigated the association between ER-α DNA methylation in leukocytes and breast composition in adolescent Chilean girls enrolled in the Growth and Obesity Cohort Study (GOCS) in Santiago, Chile. Breast composition (total breast volume (BV; cm3), fibroglandular volume (FGV; cm3), and percent fibroglandular volume (%FGV)) was measured at breast Tanner stage 4 (B4). ER-α promoter DNA methylation was assessed by pyrosequencing in blood samples collected at breast Tanner stages 2 (B2; n = 256) and B4 (n = 338). Results After adjusting for fat percentage at breast density measurement, ER-α methylation at B2, and cellular heterogeneity, we observed an inverse association between B4 average ER-α DNA methylation and BV and FGV. Geometric mean BV was 15% lower (95% CI: − 28%, − 1%) among girls in the highest quartile of B4 ER-α methylation (6.96–23.60%) relative to the lowest (0.78–3.37%). Similarly, FGV was 19% lower (95% CI: − 33%, − 2%) among girls in the highest quartile of B4 ER-α methylation relative to the lowest. The association between ER-α methylation and breast composition was not significantly modified by body fat percentage and was not influenced by pubertal timing. Conclusions These findings suggest that the methylation profile of ER-α may modulate adolescent response to estrogen and breast composition, which may influence breast cancer risk in adulthood.

Abstract In humans the expression of lactase changes during post-natal development, leading to phenotypes known as lactase persistence and non-persistence. Polymorphisms within the lactase gene (LCT) enhancer, in particular the −13910C > T, but also others, are linked to these phenotypes. We were interested in identifying dynamic mediators of LCT regulation, beyond the genotype at −13910C > T. To this end, we investigated two levels of lactase regulation in human intestinal samples obtained from New England children and adolescents of mixed European ancestry: differential expression of transcriptional regulators of LCT, and variations in DNA methylation, and their relation to phenotype. Variations in expression of CDX2, POU2F1, GATA4, GATA6, and HNF1α did not correlate with phenotype. However, an epigenome-wide approach using the Illumina Infinium HM450 bead chip identified a differentially methylated position in the LCT promoter where methylation levels are associated with the genotype at −13910C > T, the persistence/non-persistence phenotype and lactase enzymatic activity. DNA methylation levels at this promoter site and CpGs in the LCT enhancer are associated with genotype. Indeed, taken together they have a higher power to predict lactase phenotypes than the genotype alone.

Abstract Background The age of menarche has been associated with metabolic and cardiovascular disease, as well as cancer risk. The decline in menarcheal age over the past century may be partially attributable to increased exposure to endocrine disrupting chemicals (EDCs). Methods We assessed the influence of 26 phenol and phthalate biomarkers on the timing of menarche in a longitudinal cohort of Chilean girls. These EDCs were quantified in urine collected prior to the onset of breast development (Tanner 1; B1), and during adolescence (Tanner 4; B4). Multivariable accelerated failure time (AFT) models were used to analyze associations between biomarker concentrations and the age of menarche adjusting for body mass index (BMI) Z-score and maternal education, accounting for within-subject correlation. Results Several biomarkers were significantly associated with the age at menarche; however, these associations were dependent on the timing of biomarker assessment. A log(ng/ml) increase in B1 concentrations of di(2-ethylhexyl) phthalate biomarkers was associated with later menarche (hazard ratio (HR): 0.77; 95% CI: 0.60, 0.98), whereas higher B1 concentrations of 2,5-dichlorophenol and benzophenone-3 were associated with earlier menarche (HR: 1.13; 95% CI: 1.01, 1.27; HR: 1.17; 95% CI: 1.06, 1.29, respectively). Elevated B4 concentrations of monomethyl phthalate were similarly associated with earlier menarche (HR: 1.30; 95% CI: 1.10, 1.53). The impact of monoethyl phthalate and triclosan concentrations on pubertal timing were significantly modified by BMI Z-score. Higher monoethyl phthalate and triclosan concentrations were associated with earlier menarche among overweight or obese girls, but not among those that were normal weight. Conclusions This study identifies modulation of sexual maturation by specific EDC biomarkers in Latina girls.

Epigenetic age is an indicator of biological aging, capturing the impact of environmental and behavioral influences across time on cellular function. Deviance between epigenetic age and chronological age (AgeAccel) is a predictor of health. Pubertal timing has similarly been associated with cancer risk and mortality rate among females. We examined the association between AgeAccel and pubertal timing and adolescent breast composition in the longitudinal Growth and Obesity Cohort Study. AgeAccel was estimated in whole blood using the Horvath method at breast Tanner 2 (B2) and 4 (B4). Total breast volume, absolute fibro-glandular volume (FGV), and %FGV were evaluated at B4 using dual X-ray absorptiometry. The impact of AgeAccel (mean: 0; SD: 3.78) across puberty on the time to breast development (thelarche), menarche, and pubertal tempo (thelarche to menarche) was estimated using accelerated failure time models; generalized estimating equations were used to evaluate associations with breast density. A five-year increase in average adolescent AgeAccel was associated with a significant decrease in time to menarche [hazard ratio (HR): 1.37; 95% confidence interval (CI): 1.04, 1.80] adjusting for birth weight, maternal pre-pregnancy body mass index, maternal height, maternal education, B2 height, fat percentage, and cell composition. AgeAccel displayed a stronger inverse association with pubertal tempo (HR: 1.48; 95% CI: 1.10, 1.99). A five-year increase in AgeAccel was associated with 5% greater %FGV, adjusting for B4 percent body fat, and maternal traits (95% CI: 1.01, 1.10). Our study provides unique insight into the influence of AgeAccel on pubertal development in girls, which may have implications for adult health.

We investigated the impact of maternal smoking during pregnancy on placental DNA methylation and how this may mediate the association between maternal smoking and pro-inflammatory proteins in cord blood. The study population consisted of 27 individuals exposed to maternal smoking throughout pregnancy, 32 individuals exposed during a proportion of the pregnancy, and 61 unexposed individuals. Methylation of 11 regions within 6 genes in placenta tissue was assessed by pyrosequencing. Levels of 7 pro-inflammatory proteins in cord blood were assessed by electrochemiluminescence. Differential methylation was observed in the CYP1A1 promoter and AHRR gene body regions between women who smoked throughout pregnancy and non-smokers on the fetal-side of the placenta and in the GFI1 promoter between women who quit smoking while pregnant and non-smokers on the maternal-side of the placenta. Maternal smoking resulted in elevated levels of IL-8 protein in cord blood, which was not mediated by DNA methylation of our candidate regions at either the maternal or the fetal side of the placenta. Placental DNA methylation was associated with levels of inflammatory proteins in cord blood. Our observations suggest that maternal smoking during pregnancy affects both placental DNA methylation and the neonate's immune response.

Epigenetics: choose your DNA methylation probing tool wisely! Researchers who study human epigenetics need to carefully consider the platform used to measure genome-wide patterns of DNA methylation. A team led by Karin Michels and Andrea Baccarelli from Harvard University in Boston, Massachusetts, USA, empirically examined the strengths and weaknesses of two methylation profiling tools: Illumina’s Infinium BeadChip, which uses a microarray system to interrogate hundreds of thousands of methylation sites across the genome at single-nucleotide resolution; and a high-throughput sequencing-based approach known as rapid multiplexed reduced representation bisulfite sequencing, or rmRRBS. The former did a better job at reading methylation in protein-coding and mitochondrial-related genes, while the latter required less input DNA and covered more methylation sites across the genome. The authors conclude that a scientist’s platform preference should depend on the nature of his or her investigation.

Ethylene regulates fruit ripening and several plant functions (germination, plant growth, plant-microbe interactions). Protein quantification of ethylene receptors (ETRs) is essential to study their functions, but is impaired by low resolution tools such as antibodies that are mostly nonspecific, or the lack of sensitivity of shotgun proteomic approaches. We developed a targeted proteomic method, to quantify low-abundance proteins such as ETRs, and coupled this to mRNAs analyses, in two tomato lines: Wild Type (WT) and Never-Ripe (NR) which is insensitive to ethylene because of a gain-of-function mutation in ETR3. We obtained mRNA and protein abundance profiles for each ETR over the fruit development period. Despite a limiting number of replicates, we propose Pearson correlations between mRNA and protein profiles as interesting indicators to discriminate the two genotypes: such correlations are mostly positive in the WT and are affected by the NR mutation. The influence of putative post-transcriptional and post-translational changes are discussed. In NR fruits, the observed accumulation of the mutated ETR3 protein between ripening stages (Mature Green and Breaker + 8 days) may be a cause of NR tomatoes to stay orange. The label-free quantitative proteomics analysis of membrane proteins, concomitant to Parallel Reaction Monitoring analysis, may be a resource to study changes over tomato fruit development. These results could lead to studies about ETR subfunctions and interconnections over fruit development. Variations of RNA-protein correlations may open new fields of research in ETR regulation. Finally, similar approaches may be developed to study ETRs in whole plant development and plant-microorganism interactions.

Ethylene is a gas that has long been known to act as a plant hormone. We recently showed that a cyanobacterium, Synechocystis sp. PCC 6803 (Synechocystis) contains an ethylene receptor (SynEtr1) that regulates cell surface and extracellular components leading to altered phototaxis and biofilm formation. To determine whether other cyanobacteria respond to ethylene, we examined the effects of exogenous ethylene on phototaxis of the filamentous cyanobacterium, Geitlerinema sp. PCC 7105 (Geitlerinema). A search of the Geitlerinema genome suggests that two genes encode proteins that contain an ethylene binding domain and Geitlerinema cells have previously been shown to bind ethylene. We call these genes GeiEtr1 and GeiEtr2 and show that in air both are expressed. Treatment with ethylene decreases the abundance of GeiEtr1 transcripts. Treatment of Geitlerinema with 1000 nL L–1 ethylene affected the phototaxis response to white light as well as monochromatic red light, but not blue or green light. This is in contrast to Synechocystis where we previously found ethylene affected phototaxis to all three colors. We also demonstrate that application of ethylene down to 8 nL L–1 stimulates phototaxis of both cyanobacteria as well as biofilm formation of Synechocystis. We formerly demonstrated that the transcript levels of slr1214 and CsiR1 in Synechocystis are reduced by treatment with 1000 nL L–1 ethylene. Here we show that application of ethylene down to 1 nL L–1 causes a reduction in CsiR1 abundance. This is below the threshold for most ethylene responses documented in plants. By contrast, slr1214 is unaffected by this low level of ethylene and only shows a reduction in transcript abundance at the highest ethylene level used. Thus, cyanobacteria are very sensitive to ethylene. However, the dose-binding characteristics of ethylene binding to Geitlerinema and Synechocystis cells as well as to the ethylene binding domain of SynEtr1 heterologously expressed in yeast, are similar to what has been reported for plants and exogenously expressed ethylene receptors from plants. These data are consistent with a model where signal amplification is occurring at the level of the receptors.

Cytokinins (CKs) and ethylene (ET) are among the most ancient organic chemicals on Earth. A wide range of organisms including plants, algae, fungi, amoebae, and bacteria use these substances as signaling molecules to regulate cellular processes. Because of their ancestral origin and ubiquitous occurrence, CKs and ET are also considered to be ideal molecules for inter-kingdom communication. Their signal transduction pathways were first historically deciphered in plants and are related to the two-component systems, using histidine kinases as primary sensors. Paradoxically, although CKs and ET serve as signaling molecules in different kingdoms, it has been supposed for a long time that the canonical CK and ET signaling pathways are restricted to terrestrial plants. These considerations have now been called into question following the identification over recent years of genes encoding CK and ET receptor homologs in many other lineages within the tree of life. These advances shed new light on the dissemination and evolution of these hormones as both intra- and inter-specific communication molecules in prokaryotic and eukaryotic organisms.