Prevalence, patterns and predictors of paranormal beliefs in The Netherlands: a several-analysts approach.

Paranormal beliefs encompass a wide variety of phenomena, including the existence of supernatural entities such as ghosts and witches, as well as extraordinary human abilities such as telepathy and clairvoyance. In the current study, we used a nationally representative sample ( N = 2534 ) to investigate the presence and correlates of paranormal beliefs among the secular Dutch population. The results indicated that most single paranormal phenomena (e.g. belief in clairvoyance) are endorsed by 10-20% of Dutch respondents; however, 55.6% of respondents qualify as paranormal believers based on the preregistered criterion that they believe in at least one phenomenon with considerable certainty. In addition, we invited four analysis teams with different methodological expertise to assess the structure of paranormal beliefs using traditional factor analysis, network analysis, Bayesian network analysis and latent class analysis (LCA). The teams' analyses indicated adequate fit of a four-factor structure reported in a 1985 study, but also emphasized different conclusions across techniques; network analyses showed evidence against strong connectedness within most clusters, and suggested a five-cluster structure. The application of various analytic techniques painted a nuanced picture of paranormal beliefs and believers in The Netherlands and suggests that despite increased secularization, subgroups of the general population still believe in paranormal phenomena.

Erratum. Consensus Guidance for Monitoring Individuals With Islet Autoantibody-Positive Pre-Stage 3 Type 1 Diabetes. Diabetes Care 2024;47:1276-1298.

In the article cited above, incorrect affiliation information was given for author Linda A. DiMeglio. The correct affiliation for Linda A. DiMeglio is Department of Pediatrics, Indiana University School of Medicine, Indianapolis, IN. The online version of the article (https://doi.org/10.2337/dci24-0042) has been updated to correct the error.

The effects of underwater diving on the lung function of divers in coastal areas of tourism destinations: A cross-sectional study in North Sulawesi Province, Indonesia.

North Sulawesi is renowned for its captivating underwater tourism, attracting SCUBA divers and traditional divers. Given the potential impact of diving activities on lung function, this study aims to explore differences in lung function among self-contained breathing apparatus (SCUBA) diving tourism workers, traditional divers utilizing breathing air compressors and air lines, and non-divers within the coastal areas of tourism destinations in North Sulawesi. Employing a descriptive comparative study with a cross-sectional approach, 90 subjects were categorized into three groups, and selected through simple random sampling. Spirometry was utilized for data collection, and Analysis of Variance (ANOVA) was employed for comparisons, with post hoc analysis for specific differences. The ANOVA analysis revealed significant variations in FVC Predicted, FVC Measured, FEV1, and obstruction and restriction values among SCUBA divers, traditional divers, and non-divers with a level of significance of p < 0.05. Post hoc analysis exhibited noteworthy distinctions in FVC Predicted, FVC Measured, and FEV1 between traditional divers and non-divers, and between SCUBA divers and traditional divers (p < 0.00). Obstruction values showed differences between SCUBA divers and traditional divers (p = 0.012), while restriction values indicated disparities between traditional divers and non-divers (p = 0.019). In conclusion, there were significant variations in lung function among SCUBA diving tourism workers, traditional divers using compressors and air lines, and non-divers within the coastal areas of tourism destinations in North Sulawesi.

Resveratrol attenuates doxorubicin-induced toxicity during in vitro culture of mouse-isolated preantral follicles.

The aims of this study were to evaluate the doxorubicin concentration that induces toxic effects on in vitro culture of isolated mouse secondary follicles and to investigate whether resveratrol can inhibit or reduce this toxicity. Secondary follicles were isolated and cultured for 12 days in control medium (α-MEM+) or in α-MEM+ supplemented with doxorubicin (0.1 µg/ml) or different concentrations of resveratrol (0.5, 2, or 5 µM) associated with doxorubicin (0.1 µg/ml) (experiment 1). For experiment 2, follicles were cultured in α-MEM+ alone or supplemented with doxorubicin (0.3 µg/ml) or different concentrations of resveratrol (5 or 10 µM) associated or not with doxorubicin (0.3 µg/ml) (experiment 2). The endpoints analyzed were morphology (survival), antrum formation, follicular diameter, mitochondrial activity, glutathione (GSH) levels and DNA fragmentation. In the first experiment, doxorubicin (0.1 µg/ml) maintained survival and antrum formation similar to the control, while 5 µM resveratrol showed increased parameters, maintained mitochondrial activity and increased GSH levels compared to the control. In the second experiment, doxorubicin (0.3 µg/ml) reduced survival, antrum formation and follicular diameter compared to the control. Resveratrol at a concentration of 10 µM attenuated the damage caused by doxorubicin by improving follicular survival and did not present DNA fragmentation. In conclusion, supplementation of the in vitro culture medium with 0.3 µg/ml doxorubicin reduced the survival and impaired the development of mouse-isolated preantral follicles. Resveratrol at 10 µM reduced doxorubicin-induced follicular atresia, without DNA fragmentation in the follicles.

Detailed Analysis of Prebiotic Fructo- and Galacto-Oligosaccharides in the Human Small Intestine.

Galacto-oligosaccharides (GOS) and fructo-oligosaccharides (FOS) are food ingredients that improve human health, but their degradation throughout the human small intestine is not well understood. We studied the breakdown kinetics of FOS and GOS in the intestines of seven healthy Dutch adults. Subjects were equipped with a catheter in the distal ileum or proximal colon and consumed 5 g of chicory-derived FOS (degree of polymerization (DP) DP2-10), and 5 g of GOS (DP2-6). Postprandially, intestinal content was frequently collected until 350 min and analyzed for mono-, di-, and oligosaccharides. FOS and GOS had recoveries of 96 ± 25% and 76 ± 28%, respectively. FOS DP ≥ 2 and GOS DP ≥ 3 abundances in the distal small intestine or proximal colon matched the consumed doses, while GOS dimers (DP2) had lower recoveries, namely 22.8 ± 11.1% for ß-D-gal-(1↔1)-α-D-glc+ß-D-gal-(1↔1)-ß-D-glc, 19.3 ± 19.1% for ß-D-gal-(1 → 2)-D-glc+ß-D-gal-(1 → 3)-D-glc, 43.7 ± 24.6% for ß-D-gal-(1 → 6)-D-gal, and 68.0 ± 38.5% for ß-D-gal-(1 → 4)-D-gal. Lactose was still present in the distal small intestine of all of the participants. To conclude, FOS DP ≥ 2 and GOS DP ≥ 3 were not degraded in the small intestine of healthy adults, while most prebiotic GOS DP2 was hydrolyzed in a structure-dependent manner. We provide evidence on the resistances of GOS with specific ß-linkages in the human intestine, supporting the development of GOS prebiotics that resist small intestine digestion.

Kidney Transplant Outcome Following Donation After Euthanasia.

Importance: In the Netherlands, organ donation after euthanasia (donation after circulatory death type V [DCD-V]) has been increasingly performed since 2012. However, the outcomes of DCD-V kidney grafts have not been thoroughly investigated. It is critical to assess the outcomes of these kidney grafts to ascertain whether DCD-V is a safe and valuable way to increase the kidney donor pool. Objectives: To investigate the outcomes of DCD-V kidney transplantation and compare them with outcomes of kidney transplantation after circulatory death after withdrawal of life-sustaining therapies (DCD type III [DCD-III]) and donation after brain death (DBD). Design, Setting, and Participants: A retrospective cohort study was conducted using the database from the Dutch Transplant Foundation. All kidney transplants in the Netherlands between January 2012 (start of the euthanasia program) and July 2023 were included. Follow-up was obtained through 5 years after transplantation. Data analysis was performed from November 2023 until February 2024. Exposures: Kidney transplantation with a DCD-V graft compared with DCD-III and DBD grafts. Main Outcomes and Measures: The primary outcome was death-censored graft survival until 5 years after transplantation. Secondary outcomes were the incidence of delayed graft function (DGF), permanent nonfunction (PNF), serum creatinine concentration, and patient survival until 5 years after kidney transplantation. Results: A total of 145 DCD-V kidney transplants were compared with 1936 DCD-III and 1255 DBD kidney transplants. Median (IQR) recipient age was 59 (46-66) years in the DCD-V cohort, compared with 61 (50-68) years in the DCD-III cohort and 61 (50-68) years in the DBD cohort. The incidence of DGF with DCD-V kidney transplants (26%) was significantly less than that with DCD-III kidney transplants (49%; P < .001) and similar to that with DBD kidney transplants (22%; P = .46). PNF occurrence with DCD-V kidneys (6%) was similar to that with DCD-III kidneys (6%; P = .79) and higher than in DBD kidneys (4%; P < .001). There was no difference in 5-year death-censored graft survival between DCD-V grafts (82%) and DCD-III (86%; P = .99) or DBD (84%; P = .99) grafts. There was no difference in 5-year patient survival between DCD-V kidney transplants (69%) and DCD-III (76%; P = .45) or DBD (73%; P = .74) kidney transplants. A propensity score analysis was performed to match the DCD-V and DCD-III cohort, showing results similar to those of the unmatched cohort. Conclusions and Relevance: This study found that DCD-V kidney transplantation yielded a lower incidence of DGF compared with DCD-III kidney transplantation and yielded long-term results similar to those of DCD-III and DBD kidney transplantation. The findings suggest that DCD-V is a safe and valuable way to increase the kidney donor pool.

SATURN: assessing the feasibility of utilising existing registries for real-world evidence data collection to meet patients, regulatory, health technology assessment and payer requirements.

BACKGROUND: SATURN (Systematic Accumulation of Treatment practices and Utilisation, Real world evidence, and Natural history data) for the rare condition osteogenesis imperfecta (OI) has the objective to create a common core dataset by utilising existing, well-established data sources to meet the needs of the various stakeholders (physicians, registry/dataset owners, patients and patient associations, OI community leaders, European [EU] policymakers, regulators, health technology assessments [HTA]s, and healthcare systems including payers). This paper describes the steps taken to assess the feasibility of one existing OI registry (i.e., the Registry of OI [ROI]) as a candidate for SATURN. The same methodology will be applied to other existing OI registries in the future and this same concept could be utilised for other rare disease registries. METHODS: The approach to assessing the feasibility of the ROI registry consisted of three steps: (1) an assessment of the registry characteristics using the Registry Evaluation and Quality Standards Tool (REQueST); (2) a gap analysis comparing SATURN required Core Variables to those being captured in the registry's Case Report Form (CRF); and (3) a compliance check on the data exchange process following the Title 21 of Code of Federal Regulations (CFR) Part 11/EudraLex Annex 11 Compliance Checklist. The first registry that SATURN has assessed is the ROI database at the Istituto Ortopedico Rizzoli (IOR) in Italy. RESULTS: The results from the ROI REQueST have demonstrated satisfactory complete responses in terms of methodology, essential standards, interpretability, and interoperability-readiness for data linkage, data sources, and ethics to meet the needs of data customers. However, the ROI data is from a tertiary referral centre which may limit the ability to understand the full patient journey. The gap analysis has revealed that an exact or logical match between SATURN requested variables and the ROI current variables exists for the following items: patient characteristics, treatment of OI (medical and surgical) and treatment of pain (with the exception of frequency of treatment and reasons for discontinuation), fracture history and bone density. However, data on safety was missing. The compliance check has implied that the ROI implemented appropriate controls for the web-based platform (i.e., Genotype-phenotype Data Integration Platform [GeDI]) that is involved in processing the electronic patient data, and GeDI is a validated/compliant application that follows relevant 21 CFR Part 11/EudraLex Annex 11 regulations. CONCLUSIONS: This robust feasibility process highlights potential limitations and opportunities to develop and to refine the collaboration with the ROI as the SATURN programme moves forward. It also ensures that the existing datasets in the rare condition OI are being maximised to respond to the needs of patients, data customers and decision-makers. This feasibility process has allowed SATURN to build a compliant methodology that aligns with the requirements from the European Medicines Agency (EMA) and HTAs. More data variables will continue to be developed and refined along the way with more registries participating in SATURN. As a result, SATURN will become a meaningful and truly collaborative core dataset, which will also contribute to advancing understanding of OI diagnosis, treatment, and care.

Association of osteotomy, age, and component fixation with the outcomes of total hip arthroplasty in patients with hip dysplasia: a Dutch population-based registry study.

BACKGROUND AND PURPOSE:  Hip dysplasia can present challenges for total hip arthroplasty (THA) due to anatomic abnormalities. We aimed to assess the association of age, sex, osteotomies prior to THA, and fixation method on 5- and 10-year revision-free implant survival and patient-reported outcome measures (PROMs) of THAs in patients with hip dysplasia. METHODS: Using Dutch Arthroplasty Register data, we studied hip dysplasia patients receiving primary THAs in 2007-2021 (n = 7,465). THAs were categorized by age, pelvic osteotomy prior to THA (yes/no), and fixation (cemented, uncemented, hybrid, reverse hybrid). Kaplan-Meier and multivariable Cox models were used to determine 5- and 10-year revision-free implant survival and adjusted hazard ratios including 95% confidence intervals (CIs). Reasons for revision and PROMs were compared within the categories. RESULTS:  We found a 10-year revision-free implant survival of 94.9% (CI 94.3-95.5). Patients younger than 50 years had a 10-year implant survival of 93.3% (CI 91.9-94.7), Patients with prior pelvic osteotomy had a 10-year implant survival of 92.0% (CI 89.8-94.2). Fixation method and sex were not associated with implant survival. Patients with a prior pelvic osteotomy had more revisions due to cup loosening and reported lower PROM scores than patients without earlier osteotomy. CONCLUSION:  5- and 10-year revision-free implant survival rates of THA for hip dysplasia are 96.4% and 94.9%. Age and prior osteotomies were associated with decreased implant survival rates in patients with hip dysplasia, while fixation method was not. Prior osteotomies were also associated with reduced PROM scores.

Blocking tumor-intrinsic MNK1 kinase restricts metabolic adaptation and diminishes liver metastasis.

Dysregulation of the mitogen-activated protein kinase interacting kinases 1/2 (MNK1/2)-eukaryotic initiation factor 4E (eIF4E) signaling axis promotes breast cancer progression. MNK1 is known to influence cancer stem cells (CSCs); self-renewing populations that support metastasis, recurrence, and chemotherapeutic resistance, making them a clinically relevant target. The precise function of MNK1 in regulating CSCs, however, remains unexplored. Here, we generated MNK1 knockout cancer cell lines, resulting in diminished CSC properties in vitro and slowed tumor growth in vivo. Using a multiomics approach, we functionally demonstrated that loss of MNK1 restricts tumor cell metabolic adaptation by reducing glycolysis and increasing dependence on oxidative phosphorylation. Furthermore, MNK1-null breast and pancreatic tumor cells demonstrated suppressed metastasis to the liver, but not the lung. Analysis of The Cancer Genome Atlas (TCGA) data from breast cancer patients validated the positive correlation between MNK1 and glycolytic enzyme protein expression. This study defines metabolic perturbations as a previously unknown consequence of targeting MNK1/2, which may be therapeutically exploited.

Elucidating sequence-function relationships in a template-independent polymerase to enable novel DNA recording applications.

Harnessing DNA as a high-density storage medium for information storage and molecular recording of signals has been of increasing interest in the biotechnology field. Recently, progress in enzymatic DNA synthesis, DNA digital data storage, and DNA-based molecular recording has been made by leveraging the activity of the template-independent DNA polymerase, terminal deoxynucleotidyl transferase (TdT). TdT adds deoxyribonucleotides to the 3' end of single-stranded DNA, generating random sequences of single-stranded DNA. TdT can use several divalent cations for its enzymatic activity and exhibits shifts in deoxyribonucleotide incorporation frequencies in response to changes in its reaction environment. However, there is limited understanding of sequence-structure-function relationships regarding these properties, which in turn limits our ability to modulate TdT to further advance TdT-based tools. Most TdT literature to-date explores the activity of murine, bovine or human TdTs; studies probing TdT sequence and structure largely focus on strictly conserved residues that are functionally critical to TdT activity. Here, we explore non-conserved TdT sequence space by surveying the natural diversity of TdT. We characterize a diverse set of TdT homologs from different organisms and identify several TdT residues/regions that confer differences in TdT behavior between homologs. The observations in this study can design rules for targeted TdT libraries, in tandem with a screening assay, to modulate TdT properties. Moreover, the data can be useful in guiding further studies of potential residues of interest. Overall, we characterize TdTs that have not been previously studied in the literature, and we provide new insights into TdT sequence-function relationships.

Maternal Supplementation with Dietary Betaine during Late Gestation Increased Ewe Plasma Creatine and Lamb Thermoregulation under Field Conditions.

Twin lamb mortality is a significant economic problem impacting the Australian sheep industry. Maternal betaine supplementation improved lamb vigour and early post-natal survival when ewes and lambs were housed indoors, suggesting that betaine may be beneficial to feed under extensive pasture systems. This study investigated whether maternal betaine supplementation during late gestation would improve Merino twin lamb live weight, thermoregulation, vigour and survival to weaning under field conditions. Ewes received dietary betaine at either 0 g/day (CTL; n = 115) or 4 g/day from day 110 of gestation (dG 110) until ~49 days post-partum (pp) (BET; n = 115). Measures indicative of lamb viability and survival were collected within 4-24 h of birth and at ~49 days pp and ~93 days pp. BET ewes had higher creatine and creatinine concentrations at dG 130 than CTL ewes (p < 0.05). BET lambs had a higher rectal temperature within 4-24 h following birth than CTL lambs (p < 0.05). CTL lambs were heavier at ~49 days pp and grew faster from birth to ~49 days pp than BET lambs (both p < 0.05). The time taken after release from the researcher to first suckling was quicker in the CTL lambs than BET lambs (p < 0.05). This study demonstrated that supplementing betaine increased creatine concentration in twin-bearing ewes and thermoregulatory capacity in neonatal lambs under extensive grazing systems.

AI-guided identification of risk variants for adrenocortical tumours in TP53 p.R337H carrier children: a genetic association study.

Background: Adrenocortical tumours (ACT) in children are part of the Li-Fraumeni cancer spectrum and are frequently associated with a germline TP53 pathogenic variant. TP53 p.R337H is highly prevalent in the south and southeast of Brazil and predisposes to ACT with low penetrance. Thus, we aimed to investigate whether genetic variants exist which are associated with an increased risk of developing ACT in TP53 p.R337H carrier children. Methods: A genetic association study was conducted in trios of children (14 girls, 7 boys) from southern Brazil carriers of TP53 p.R337H with (n = 18) or without (n = 3) ACT and their parents, one of whom also carries this pathogenic variant (discovery cohort). Results were confirmed in a validation cohort of TP53 p.R337H carriers with (n = 90; 68 girls, 22 boys) or without ACT (n = 302; 165 women, 137 men). Findings: We analysed genomic data from whole exome sequencing of blood DNA from the trios. Using deep learning algorithms, according to a model where the affected child inherits from the non-carrier parent variant(s) increasing the risk of developing ACT, we found a significantly enriched representation of non-coding variants in genes involved in the cyclic AMP (cAMP) pathway known to be involved in adrenocortical tumorigenesis. One among those variants (rs2278986 in the SCARB1 gene) was confirmed to be significantly enriched in the validation cohort of TP53 p.R337H carriers with ACT compared to carriers without ACT (OR 1.858; 95% CI 1.146, 3.042, p = 0.01). Interpretation: Profiling of the variant rs2278986 is a candidate for future confirmation and possible use as a tool for ACT risk stratification in TP53 p.R337H carriers. Funding: Centre National de la Recherche Scientifique (CNRS), Behring Foundation, Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq).

Hormone-free versus FSH-primed infertility treatment of women with polycystic ovary syndrome using biphasic in vitro maturation: a randomized controlled trial.

OBJECTIVE: To compare oocyte maturation rates and pregnancy outcomes in women with polycystic ovary syndrome (PCOS) undergoing biphasic in vitro maturation (capacitation [CAPA]-IVM) with versus without follicle-stimulating hormone (FSH) priming. DESIGN: Randomized, controlled, assessor-blinded trial SUBJECTS: Women aged 18-37 years with PCOS and an indication for CAPA-IVM. INTERVENTION: Participants were randomized (1:1) to undergo CAPA-IVM with or without FSH priming. The FSH priming group had two days of FSH injections before oocyte pick-up; no FSH was given in the non-FSH group. After CAPA-IVM, day-5 embryos were vitrified for transfer in a subsequent cycle. MAIN OUTCOME MEASURE(S): The primary endpoint was number of matured oocytes. Secondary outcomes included rates of live birth, implantation, clinical pregnancy, ongoing pregnancy, pregnancy complications, obstetric and perinatal complications, and neonatal complications. RESULTS: The number [interquartile range] of matured oocytes did not differ significantly in the non-FSH versus FSH group (13 [9-18] vs. 14 [7-8]; absolute difference -1 [95% confidence interval (CI) -5, 4]); other oocyte and embryology outcomes did not differ between groups. Rates of ongoing pregnancy and live birth were both 38.3% in the non-FSH group and both 31.7% in the FSH group (risk ratio for both outcomes: 1.21, 95% CI 0.74-1.98). Maternal complications were infrequent and occurred at a similar rate in the two groups; there were no preterm deliveries before 32 weeks' gestation. CONCLUSION: These findings open the possibility of a new, hormone-free approach to infertility treatment of women with PCOS.

Advancements in supervised deep learning for metal artifact reduction in computed tomography: A systematic review.

BACKGROUND: Metallic artefacts caused by metal implants, are a common problem in computed tomography (CT) imaging, degrading image quality and diagnostic accuracy. With advancements in artificial intelligence, novel deep learning (DL)-based metal artefact reduction (MAR) algorithms are entering clinical practice. OBJECTIVE: This systematic review provides an overview of the performance of the current supervised DL-based MAR algorithms for CT, focusing on three different domains: sinogram, image, and dual domain. METHODS: A literature search was conducted in PubMed, EMBASE, Web of Science, and Scopus. Outcomes were assessed using peak signal-to-noise ratio (PSNR) and structural similarity index measure (SSIM) or any other objective measure comparing MAR performance to uncorrected images. RESULTS: After screening, fourteen studies were selected that compared DL-based MAR-algorithms with uncorrected images. MAR-algorithms were categorised into the three domains. Thirteen MAR-algorithms showed a higher PSNR and SSIM value compared to the uncorrected images and to non-DL MAR-algorithms. One study showed statistically significant better MAR performance on clinical data compared to the uncorrected images and non-DL MAR-algorithms based on Hounsfield unit calculations. CONCLUSION: DL MAR-algorithms show promising results in reducing metal artefacts, but standardised methodologies are needed to evaluate DL-based MAR-algorithms on clinical data to improve comparability between algorithms. CLINICAL RELEVANCE STATEMENT: Recent studies highlight the effectiveness of supervised Deep Learning-based MAR-algorithms in improving CT image quality by reducing metal artefacts in the sinogram, image and dual domain. A systematic review is needed to provide an overview of newly developed algorithms.

Maternal Complications After Laser Surgery for Twin-to-Twin Transfusion Syndrome, a Cohort Study.

OBJECTIVES: To assess maternal complications after fetoscopic laser surgery (FLS) for the twin-to-twin transfusion syndrome (TTTS). METHODS: All consecutive cases treated with FLS for TTTS between 2008 and 2021 at the Leiden University Medical Center (LUMC) were included. We allocated complications in three timeframes: "Admission for laser surgery," "pregnancy after laser," and "delivery and third stage of labor." Maternal complications were graded according to the Maternal and Fetal Adverse Event Terminology (MFAET) and for intra-abdominal hemorrhage, the Common Terminology Criteria for Adverse Events (CTCAE). RESULTS: In the study period, 637 mothers were treated for TTTS with FLS. There were 1559 occurrences of maternal complications. The rate of severe maternal complications (grade 3 or 4) was 8.0%. Severe complications consisted of six cases of severe intra-abdominal hemorrhage, nine cases of severe hemorrhage in pregnancy, one with severe chorioamnionitis, 10 with severe preeclampsia/HELLP syndrome, and 25 with a severe postpartum hemorrhage. CONCLUSIONS: Even though it is the gold standard for treating TTTS, FLS comes at a risk to the mother which should not be neglected. And even though not all complications have serious consequences to the mother, the severe maternal complication rate of 8.0% should be added to the inherent risks for the fetus, and should be discussed with patients eligible for surgery in order to make an informed decision on treatment options.

Visuell snø ­ forstyrrende, men ufarlig.

Visual snow is a persistent visual disturbance in which small dots of light appear across the visual field of both eyes, like snowfall. The condition is not widely known, but an increasing number of cases have been described in the last 5 - 6 years. Many patients have consulted a number of specialists without receiving an explanation for their symptoms, and in this clinical review article we therefore wish to make the condition known to more clinicians.

Fixational eye movements enhance the precision of visual information transmitted by the primate retina.

Fixational eye movements alter the number and timing of spikes transmitted from the retina to the brain, but whether these changes enhance or degrade the retinal signal is unclear. To quantify this, we developed a Bayesian method for reconstructing natural images from the recorded spikes of hundreds of retinal ganglion cells (RGCs) in the macaque retina (male), combining a likelihood model for RGC light responses with the natural image prior implicitly embedded in an artificial neural network optimized for denoising. The method matched or surpassed the performance of previous reconstruction algorithms, and provides an interpretable framework for characterizing the retinal signal. Reconstructions were improved with artificial stimulus jitter that emulated fixational eye movements, even when the eye movement trajectory was assumed to be unknown and had to be inferred from retinal spikes. Reconstructions were degraded by small artificial perturbations of spike times, revealing more precise temporal encoding than suggested by previous studies. Finally, reconstructions were substantially degraded when derived from a model that ignored cell-to-cell interactions, indicating the importance of stimulus-evoked correlations. Thus, fixational eye movements enhance the precision of the retinal representation.

A Massively Parallel In Vivo Assay of TdT Mutants Yields Variants with Altered Nucleotide Insertion Biases.

Terminal deoxynucleotidyl transferase (TdT) is a unique DNA polymerase capable of template-independent extension of DNA. TdT's de novo DNA synthesis ability has found utility in DNA recording, DNA data storage, oligonucleotide synthesis, and nucleic acid labeling, but TdT's intrinsic nucleotide biases limit its versatility in such applications. Here, we describe a multiplexed assay for profiling and engineering the bias and overall activity of TdT variants with high throughput. In our assay, a library of TdTs is encoded next to a CRISPR-Cas9 target site in HEK293T cells. Upon transfection of Cas9 and sgRNA, the target site is cut, allowing TdT to intercept the double-strand break and add nucleotides. Each resulting insertion is sequenced alongside the identity of the TdT variant that generated it. Using this assay, 25,623 unique TdT variants, constructed by site-saturation mutagenesis at strategic positions, were profiled. This resulted in the isolation of several altered-bias TdTs that expanded the capabilities of our TdT-based DNA recording system, Cell HistorY Recording by Ordered InsertioN (CHYRON), by increasing the information density of recording through an unbiased TdT and achieving dual-channel recording of two distinct inducers (hypoxia and Wnt) through two differently biased TdTs. Select TdT variants were also tested in vitro, revealing concordance between each variant's in vitro bias and the in vivo bias determined from the multiplexed high throughput assay. Overall, our work and the multiplex assay it features should support the continued development of TdT-based DNA recorders, in vitro applications of TdT, and further study of the biology of TdT.