Semiconductor Deposition via Laser Printing of a Bespoke Toner Containing Metal Xanthate Complexes.

A methodology to use laser printing, a form of electrophotography, to print metal chalcogenide complexes on paper, is described. After fusing the toner to paper, a heating step is used to cause the printed metal xanthate complexes to thermolyze within the toner and form three target metal chalcogenides: CuS, SnS, and ZnS. To achieve this, we synthesize a poly(styrene-co-n-butyl acrylate) thermopolymer that emulates the thermal properties of a commercial toner and is also solution processable with the metal xanthate complexes used: [Zn(S2COEt)2], [Cu(S2COEt)·(PPh3)2], and [Sn(S2COEt)2]. We demonstrate through energy dispersive X-ray mapping that the toner is deposited following printing and that thermolysis of the metal xanthate complexes occurs in the fused toner, demonstrating the first example of laser printing of inorganic complexes and, in turn, semiconductors.

Building momentum through networks: Bioimaging across the Americas.

In September 2023, the two largest bioimaging networks in the Americas, Latin America Bioimaging (LABI) and BioImaging North America (BINA), came together during a 1-week meeting in Mexico. This meeting provided opportunities for participants to interact closely with decision-makers from imaging core facilities across the Americas. The meeting was held in a hybrid format and attended in-person by imaging scientists from across the Americas, including Canada, the United States, Mexico, Colombia, Peru, Argentina, Chile, Brazil and Uruguay. The aims of the meeting were to discuss progress achieved over the past year, to foster networking and collaborative efforts among members of both communities, to bring together key members of the international imaging community to promote the exchange of experience and expertise, to engage with industry partners, and to establish future directions within each individual network, as well as common goals. This meeting report summarises the discussions exchanged, the achievements shared, and the goals set during the LABIxBINA2023: Bioimaging across the Americas meeting.

Concurrent Outbreaks of Hepatitis A, Invasive Meningococcal Disease, and Mpox, Florida, USA, 2021-2022.

In 2022, concurrent outbreaks of hepatitis A, invasive meningococcal disease (IMD), and mpox were identified in Florida, USA, primarily among men who have sex with men. The hepatitis A outbreak (153 cases) was associated with hepatitis A virus genotype IA. The IMD outbreak (44 cases) was associated with Neisseria meningitidis serogroup C, sequence type 11, clonal complex 11. The mpox outbreak in Florida (2,845 cases) was part of a global epidemic. The hepatitis A and IMD outbreaks were concentrated in Central Florida and peaked during March--June, whereas mpox cases were more heavily concentrated in South Florida and had peak incidence in August. HIV infection was more common (52%) among mpox cases than among hepatitis A (21%) or IMD (34%) cases. Where feasible, vaccination against hepatitis A, meningococcal disease, and mpox should be encouraged among at-risk groups and offered along with program services that target those groups.

Characterization of endoplasmic reticulum-associated degradation in the human fungal pathogen Candida albicans.

Background: Candida albicans is the most prevalent human fungal pathogen. In immunocompromised individuals, C. albicans can cause serious systemic disease, and patients infected with drug-resistant isolates have few treatment options. The ubiquitin-proteasome system has not been thoroughly characterized in C. albicans. Research from other organisms has shown ubiquitination is important for protein quality control and regulated protein degradation at the endoplasmic reticulum (ER) via ER-associated protein degradation (ERAD). Methods: Here we perform the first characterization, to our knowledge, of ERAD in a human fungal pathogen. We generated functional knockouts of C. albicans genes encoding three proteins predicted to play roles in ERAD, the ubiquitin ligases Hrd1 and Doa10 and the ubiquitin-conjugating enzyme Ubc7. We assessed the fitness of each mutant in the presence of proteotoxic stress, and we used quantitative tandem mass tag mass spectrometry to characterize proteomic alterations in yeast lacking each gene. Results: Consistent with a role in protein quality control, yeast lacking proteins thought to contribute to ERAD displayed hypersensitivity to proteotoxic stress. Furthermore, each mutant displayed distinct proteomic profiles, revealing potential physiological ERAD substrates, co-factors, and compensatory stress response factors. Among candidate ERAD substrates are enzymes contributing to ergosterol synthesis, a known therapeutic vulnerability of C. albicans. Together, our results provide the first description of ERAD function in C. albicans, and, to our knowledge, any pathogenic fungus.

Toward scalable reuse of vEM data: OME-Zarr to the rescue.

The growing size of EM volumes is a significant barrier to findable, accessible, interoperable, and reusable (FAIR) sharing. Storage, sharing, visualization and processing are challenging for large datasets. Here we discuss a recent development toward the standardized storage of volume electron microscopy (vEM) data which addresses many of the issues that researchers face. The OME-Zarr format splits data into more manageable, performant chunks enabling streaming-based access, and unifies important metadata such as multiresolution pyramid descriptions. The file format is designed for centralized and remote storage (e.g., cloud storage or file system) and is therefore ideal for sharing large data. By coalescing on a common, community-wide format, these benefits will expand as ever more data is made available to the scientific community.

Improving the Accuracy of Biologically Effective Dose Estimates, from a Previously Published Study, After Radiosurgery for Acoustic Neuromas.

OBJECTIVE: To recalculate biological effective dose values (BED) for radio-surgical treatments of acoustic neuroma from a previous study. BEDs values were previously overestimated by only using beam-on times in calculations, so excluding the important beam-off-times (when deoxyribonucleic acid repair continues) which contribute to the overall treatment time. Simple BED estimations using a mono-exponential approximation may not always be appropriate but if used should include overall treatment time. METHODS: Time intervals between isocenters were estimated. These were especially important for the Gamma Knife Model 4C cases since manual changes significantly increase overall treatment times. Individual treatment parameters, such as iso-center number, beam-on-time, and beam-off-time, were then used to calculate BED values using a more appropriate bi-exponential model that includes fast and slow components of DNA damage repair over a wider time range. RESULTS: The revised BED estimates differed significantly from previously published values. The overestimates of BED, obtained using beam-on-time only, varied from 0%-40.3%. BED subclasses, each with a BED range of 5 Gy2.47, indicated that revised values were consistently reduced when compared with originally quoted values, especially for 4C compared with Perfexion cases. Furthermore, subdivision of 4C cases by collimator number further emphasized the impact of scheduled gap times on BED. Further analysis demonstrated important limitations of the mono-exponential model. Target volume was a major confounding factor in the interpretation of the results of this study. CONCLUSIONS: BED values should be estimated by including beam-on and beam-off times. Suggestions are provided for more accurate BED estimations in future studies.

Staphylococcus Aureus Swabbing and Decolonization Before Neuromodulation Procedures: A Systematic Review and Meta-Analysis.

INTRODUCTION: Staphylococcus aureus (S aureus) is the foremost bacterial cause of surgical-site infection (SSI) and is a common source of neuromodulation SSI. Endogenous colonization is an independent risk factor for SSI; however, this risk has been shown to diminish with screening and decolonization. MATERIALS AND METHODS: A systematic review was performed according to Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines using the PubMed, Cochrane Library, and Embase data bases from inception to January 1, 2022, for the purposes of identifying all studies reporting on the use of S aureus swabbing and/or decolonization before neuromodulation procedures. A random-effects meta-analysis was performed using the metaphor package in R to calculate odds ratios (OR). RESULTS: Five observational cohort studies were included after applying the inclusion and exclusion criteria. The average study duration was 6.6 ± 3.8 years. Three studies included nasal screening as a prerequisite for subsequent decolonization. Type of neuromodulation included spinal cord stimulation in two studies, deep brain stimulation in two studies, intrathecal baclofen in one study, and sacral neuromodulation in one study. Overall, 860 and 1054 patients were included in a control or intervention (ie, screening and/or decolonization) group, respectively. A combination of nasal mupirocin ointment and a body wash, most commonly chlorhexidine gluconate soap, was used to decolonize throughout. Overall infection rates were observed at 59 of 860 (6.86%) and ten of 1054 (0.95%) in the control and intervention groups, respectively. Four studies reported a significant difference. The OR for intervention (screen and/or decolonization) vs no intervention was 0.19 (95% CI, 0.09-0.37; p < 0.001). Heterogeneity between studies was nonsignificant (I2 = 0.43%, τ2 = 0.00). CONCLUSIONS: Preoperative S aureus swabbing and decolonization resulted in significantly decreased odds of infection in neuromodulation procedures. This measure may represent a worthwhile tool to reduce neuromodulation SSI, warranting further investigation.

Loss of WNT4 in the gubernaculum causes unilateral cryptorchidism and fertility defects.

Undescended testis (UDT) affects 6% of male births. Despite surgical correction, some men with unilateral UDT may experience infertility with the contralateral descended testis (CDT) showing no A-dark spermatogonia. To improve our understanding of the etiology of infertility in UDT, we generated a novel murine model of left unilateral UDT. Gubernaculum-specific Wnt4 knockout (KO) mice (Wnt4-cKO) were generated using retinoic acid receptor ß2-cre mice and were found to have a smaller left-unilateral UDT. Wnt4-cKO mice with abdominal UDT had an increase in serum follicle-stimulating hormone and luteinizing hormone and an absence of germ cells in the undescended testicle. Wnt4-cKO mice with inguinal UDT had normal hormonal profiles, and 50% of these mice had no sperm in the left epididymis. Wnt4-cKO mice had fertility defects and produced 52% fewer litters and 78% fewer pups than control mice. Wnt4-cKO testes demonstrated increased expression of estrogen receptor α and SOX9, upregulation of female gonadal genes, and a decrease in male gonadal genes in both CDT and UDT. Several WNT4 variants were identified in boys with UDT. The presence of UDT and fertility defects in Wnt4-cKO mice highlights the crucial role of WNT4 in testicular development.

Gene dosage changes in KCTD13 result in penile and testicular anomalies via diminished androgen receptor function.

Despite the high prevalence of hypospadias and cryptorchidism, the genetic basis for these conditions is only beginning to be understood. Using array-comparative-genomic-hybridization (aCGH), potassium-channel-tetramerization-domain-containing-13 (KCTD13) encoded at 16p11.2 was identified as a candidate gene involved in hypospadias, cryptorchidism and other genitourinary (GU) tract anomalies. Copy number variants (CNVs) at 16p11.2 are among the most common syndromic genomic variants identified to date. Many patients with CNVs at this locus exhibit GU and/or neurodevelopmental phenotypes. KCTD13 encodes a substrate-specific adapter of a BCR (BTB-CUL3-RBX1) E3-ubiquitin-protein-ligase complex (BCR (BTB-CUL3-RBX1) E3-ubiquitin-protein-ligase complex (B-cell receptor (BCR) [BTB (the BTB domain is a conserved motif involved in protein-protein interactions) Cullin3 complex RING protein Rbx1] E3-ubiqutin-protein-ligase complex), which has essential roles in the regulation of cellular cytoskeleton, migration, proliferation, and neurodevelopment; yet its role in GU development is unknown. The prevalence of KCTD13 CNVs in patients with GU anomalies (2.58%) is significantly elevated when compared with patients without GU anomalies or in the general population (0.10%). KCTD13 is robustly expressed in the developing GU tract. Loss of KCTD13 in cell lines results in significantly decreased levels of nuclear androgen receptor (AR), suggesting that loss of KCTD13 affects AR sub-cellular localization. Kctd13 haploinsufficiency and homozygous deletion in mice cause a significant increase in the incidence of cryptorchidism and micropenis. KCTD13-deficient mice exhibit testicular and penile abnormalities together with significantly reduced levels of nuclear AR and SOX9. In conclusion, gene-dosage changes of murine Kctd13 diminish nuclear AR sub-cellular localization, as well as decrease SOX9 expression levels which likely contribute in part to the abnormal GU tract development in Kctd13 mouse models and in patients with CNVs in KCTD13.

Expanding the eukaryotic genetic code with a biosynthesized 21st amino acid.

Genetic code expansion technology allows for the use of noncanonical amino acids (ncAAs) to create semisynthetic organisms for both biochemical and biomedical applications. However, exogenous feeding of chemically synthesized ncAAs at high concentrations is required to compensate for the inefficient cellular uptake and incorporation of these components into proteins, especially in the case of eukaryotic cells and multicellular organisms. To generate organisms capable of autonomously biosynthesizing an ncAA and incorporating it into proteins, we have engineered a metabolic pathway for the synthesis of O-methyltyrosine (OMeY). Specifically, we endowed organisms with a marformycins biosynthetic pathway-derived methyltransferase that efficiently converts tyrosine to OMeY in the presence of the co-factor S-adenosylmethionine. The resulting cells can produce and site-specifically incorporate OMeY into proteins at much higher levels than cells exogenously fed OMeY. To understand the structural basis for the substrate selectivity of the transferase, we solved the X-ray crystal structures of the ligand-free and tyrosine-bound enzymes. Most importantly, we have extended this OMeY biosynthetic system to both mammalian cells and the zebrafish model to enhance the utility of genetic code expansion. The creation of autonomous eukaryotes using a 21st amino acid will make genetic code expansion technology more applicable to multicellular organisms, providing valuable vertebrate models for biological and biomedical research.

Hamsters Expressing Human Angiotensin-Converting Enzyme 2 Develop Severe Disease following Exposure to SARS-CoV-2.

The rapid emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has created a global health emergency. While most human disease is mild to moderate, some infections lead to a severe disease characterized by acute respiratory distress, hypoxia, anosmia, ageusia, and, in some instances, neurological involvement. Small-animal models reproducing severe disease, including neurological sequela, are needed to characterize the pathophysiological mechanism(s) of disease and to identify medical countermeasures. Transgenic mice expressing the human angiotensin-converting enzyme 2 (hACE2) viral receptor under the control of the K18 promoter develop severe and lethal respiratory disease subsequent to SARS-CoV-2 intranasal challenge when high viral doses are used. Here, we report on SARS-CoV-2 infection of hamsters engineered to express the hACE2 receptor under the control of the K18 promoter. K18-hACE2 hamsters infected with a relatively low dose of 100 or 1,000 PFU of SARS-CoV-2 developed a severe and lethal disease, with most animals succumbing by day 5 postinfection. Hamsters developed severe lesions and inflammation within the upper and lower respiratory system, including infection of the nasal cavities causing marked destruction of the olfactory epithelium as well as severe bronchopneumonia that extended deep into the alveoli. Additionally, SARS-CoV-2 infection spread to the central nervous system (CNS), including the brain stem and spinal cord. Wild-type (WT) hamsters naturally support SARS-CoV-2 infection, with the primary lesions present in the respiratory tract and nasal cavity. Overall, infection in the K18-hACE2 hamsters is more extensive than that in WT hamsters, with more CNS involvement and a lethal outcome. These findings demonstrate the K18-hACE2 hamster model will be valuable for studying SARS-CoV-2. IMPORTANCE The rapid emergence of SARS-CoV-2 has created a global health emergency. While most human SARS-CoV-2 disease is mild, some people develop severe, life-threatening disease. Small-animal models mimicking the severe aspects of human disease are needed to more clearly understand the pathophysiological processes driving this progression. Here, we studied SARS-CoV-2 infection in hamsters engineered to express the human angiotensin-converting enzyme 2 viral receptor under the control of the K18 promoter. SARS-CoV-2 produces a severe and lethal infection in transgenic hamsters that mirrors the most severe aspects of COVID-19 in humans, including respiratory and neurological injury. In contrast to other animal systems, hamsters manifest disease with levels of input virus more consistent with natural human infection. This system will be useful for the study of SARS-CoV-2 disease and the development of drugs targeting this virus.

A new approach for quantifying epithelial and stromal thickness changes after orthokeratology contact lens wear.

The aim of the study was to develop an automatic segmentation approach to optical coherence tomography (OCT) images and to investigate the changes in epithelial and stromal thickness profile and radius of curvature after the use of orthokeratology (Ortho-K) contact lenses. A total of 45 right eyes from 52 participants were monitored before, and after one month of, uninterrupted overnight Ortho-K lens wear. The tomography of their right eyes was obtained using optical OCT and rotating Scheimpflug imaging (OCULUS Pentacam). A custom-built MATLAB code for automatic segmentation of corneal OCT images was created and used to assess changes in epithelial thickness, stromal thickness, corneal and stromal profiles and radii of curvature before, and after one month of, uninterrupted overnight wear of Ortho-K lenses. In the central area (0-2 mm diameter), the epithelium thinned by 12.8 ± 6.0 µm (23.8% on average, p < 0.01) after one month of Ortho-K lens wear. In the paracentral area (2-5 mm diameter), the epithelium thinned nasally and temporally (by 2.4 ± 5.9 µm, 4.5% on average, p = 0.031). The stroma thickness increased in the central area (by 4.8 ± 16.1 µm, p = 0.005). The radius of curvature of the central corneal anterior surface increased by 0.24 ± 0.26 mm (3.1%, p < 0.01) along the horizontal meridian and by 0.34 ± 0.18 mm (4.2%, p < 0.01) along the vertical meridian. There were no significant changes in the anterior and posterior stromal radius of curvature. This study introduced a new method to automatically detect the anterior corneal surface, the epithelial posterior surface and the posterior corneal surface in OCT scans. Overnight wear of Ortho-K lenses caused thinning of the central corneal epithelium. The anterior corneal surface became flattered while the anterior and posterior surfaces of the stroma did not undergo significant changes. The results are consistent with the changes reported in previous studies. The reduction in myopic refractive error caused by Ortho-K lens wear was mainly due to changes in corneal epithelium thickness profile.

Further development of spinal cord retreatment dose estimation: including radiotherapy with protons and light ions.

PURPOSE: A graphical user interface (GUI) was developed to aid in the assessment of changes in the radiation tolerance of spinal cord/similar central nervous system tissues with time between two individual treatment courses. METHODS: The GUI allows any combination of photons, protons (or ions) to be used as the initial, or retreatment, radiotherapy courses. Allowances for clinical circumstances, of reduced tolerance, can also be made. The radiobiological model was published previously and has been incorporated with additional checks and safety features, to be as safe to use as possible. The proton option includes use of a fixed RBE of 1.1 (set as the default), or a variable RBE, the latter depending on the proton linear energy transfer (LET) for organs at risk. This second LET-based approach can also be used for ions, by changing the LET parameters. RESULTS: GUI screenshots are used to show the input and output parameters for different clinical situations used in worked examples. The results from the GUI are in agreement with manual calculations, but the results are now rapidly available without tedious and error-prone manual computations. The software outputs provide a maximum dose limit boundary, which should not be exceeded. Clinicians may also choose to further lower the number of treatment fractions, whilst using the same dose per fraction (or conversely a lower dose per fraction but with the same number of fractions) in order to achieve the intended clinical benefit as safely as possible. CONCLUSIONS: The new GUI will allow scientific-based estimations of time related radiation tolerance changes in the spinal cord and similar central nervous tissues (optic chiasm, brainstem), which can be used to guide the choice of retreatment dose fractionation schedules, with either photons, protons or ions.

A rich gallery of carbon dots based photoluminescent suspensions and powders derived by citric acid/urea.

In this study we demonstrate simple guidelines to generate a diverse range of fluorescent materials in both liquid and solid state by focusing on the most popular C-dots precursors, i.e. the binary systems of citric acid and urea. The pyrolytic treatment of those precursors combined with standard size separation techniques (dialysis and filtration), leads to four distinct families of photoluminescent materials in which the emissive signal predominantly arises from C-dots with embedded fluorophores, cyanuric acid-rich C-dots, a blend of molecular fluorophores and a mixture of C-dots with unbound molecular fluorophores, respectively. Within each one of those families the chemical composition and the optical properties of their members can be fine-tuned by adjusting the molar ratio of the reactants. Apart from generating a variety of aqueous dispersions, our approach leads to highly fluorescent powders derived from precursors comprising excessive amounts of urea that is consumed for the build-up of the carbogenic cores, the molecular fluorophores and the solid diluent matrix that suppresses self-quenching effects.

Human convalescent plasma protects K18-hACE2 mice against severe respiratory disease.

SARS-CoV-2 is the causative agent of COVID-19 and human infections have resulted in a global health emergency. Small animal models that reproduce key elements of SARS-CoV-2 human infections are needed to rigorously screen candidate drugs to mitigate severe disease and prevent the spread of SARS-CoV-2. We and others have reported that transgenic mice expressing the human angiotensin-converting enzyme 2 (hACE2) viral receptor under the control of the Keratin 18 (K18) promoter develop severe and lethal respiratory disease subsequent to SARS-CoV-2 intranasal challenge. Here we report that some infected mice that survive challenge have residual pulmonary damages and persistent brain infection on day 28 post-infection despite the presence of anti-SARS-COV-2 neutralizing antibodies. Because of the hypersensitivity of K18-hACE2 mice to SARS-CoV-2 and the propensity of virus to infect the brain, we sought to determine if anti-infective biologics could protect against disease in this model system. We demonstrate that anti-SARS-CoV-2 human convalescent plasma protects K18-hACE2 against severe disease. All control mice succumbed to disease by day 7; however, all treated mice survived infection without observable signs of disease. In marked contrast to control mice, viral antigen and lesions were reduced or absent from lungs and absent in brains of antibody-treated mice. Our findings support the use of K18-hACE2 mice for protective efficacy studies of anti-SARS-CoV-2 medical countermeasures (MCMs). They also support the use of this system to study SARS-CoV-2 persistence and host recovery.

PRSS50 is a testis protease responsible for proper sperm tail formation and function.

Multiple morphological abnormalities of the sperm flagella (MMAF) are a major cause of asthenoteratozoospermia. We have identified protease serine 50 (PRSS50) as having a crucial role in sperm development, because Prss50-null mice presented with impaired fertility and sperm tail abnormalities. PRSS50 could also be involved in centrosome function because these mice showed a threefold increase in acephalic sperm (head-tail junction defect), sperm with multiple heads (spermatid division defect) and sperm with multiple tails, including novel two conjoined sperm (complete or partial parts of several flagellum on the same plasma membrane). Our data support that, in the testis, as in tumorigenesis, PRSS50 activates NFκB target genes, such as the centromere protein leucine-rich repeats and WD repeat domain-containing protein 1 (LRWD1), which is required for heterochromatin maintenance. Prss50-null testes have increased IκκB, and reduced LRWD1 and histone expression. Low levels of de-repressed histone markers, such as H3K9me3, in the Prss50-null mouse testis may cause increases in post-meiosis proteins, such as AKAP4, affecting sperm formation. We provide important insights into the complex mechanisms of sperm development, the importance of testis proteases in fertility and a novel mechanism for MMAF.

Covid-19 transmission modelling of students returning home from university.

We provide an open-source model to estimate the number of secondary Covid-19 infections caused by potentially infectious students returning from university to private homes with other occupants. Using a Monte-Carlo method and data derived from UK sources, we predict that an infectious student would, on average, infect 0.94 other household members. Or, as a rule of thumb, each infected student would generate (just less than) one secondary within-household infection. The total number of secondary cases for all returning students is dependent on the virus prevalence within each student population at the time of their departure from campus back home. Although the proposed estimation method is general and robust, the results are sensitive to the input data. We provide Matlab code and a helpful online app (http://bit.ly/Secondary_infections_app) that can be used to estimate numbers of secondary infections based on local parameter values. This can be used worldwide to support policy making.

Development of a coronavirus disease 2019 nonhuman primate model using airborne exposure.

Airborne transmission is predicted to be a prevalent route of human exposure with SARS-CoV-2. Aside from African green monkeys, nonhuman primate models that replicate airborne transmission of SARS-CoV-2 have not been investigated. A comparative evaluation of COVID-19 in African green monkeys, rhesus macaques, and cynomolgus macaques following airborne exposure to SARS-CoV-2 was performed to determine critical disease parameters associated with disease progression, and establish correlations between primate and human COVID-19. Respiratory abnormalities and viral shedding were noted for all animals, indicating successful infection. Cynomolgus macaques developed fever, and thrombocytopenia was measured for African green monkeys and rhesus macaques. Type II pneumocyte hyperplasia and alveolar fibrosis were more frequently observed in lung tissue from cynomolgus macaques and African green monkeys. The data indicate that, in addition to African green monkeys, macaques can be successfully infected by airborne SARS-CoV-2, providing viable macaque natural transmission models for medical countermeasure evaluation.

An atlas of neural crest lineages along the posterior developing zebrafish at single-cell resolution.

Neural crest cells (NCCs) are vertebrate stem cells that give rise to various cell types throughout the developing body in early life. Here, we utilized single-cell transcriptomic analyses to delineate NCC-derivatives along the posterior developing vertebrate, zebrafish, during the late embryonic to early larval stage, a period when NCCs are actively differentiating into distinct cellular lineages. We identified several major NCC/NCC-derived cell-types including mesenchyme, neural crest, neural, neuronal, glial, and pigment, from which we resolved over three dozen cellular subtypes. We dissected gene expression signatures of pigment progenitors delineating into chromatophore lineages, mesenchyme cells, and enteric NCCs transforming into enteric neurons. Global analysis of NCC derivatives revealed they were demarcated by combinatorial hox gene codes, with distinct profiles within neuronal cells. From these analyses, we present a comprehensive cell-type atlas that can be utilized as a valuable resource for further mechanistic and evolutionary investigations of NCC differentiation.