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Telomeric G-quadruplexes (G4s) are non-canonical DNA structures composed of TTAGGG repeats. They are extensively studied both as biomolecules key for genome stability and as promising building blocks and functional elements in synthetic biology and nanotechnology. This is why it is extremely important to understand how the interaction between G4s is affected by their topology. We used small-angle x-ray scattering to investigate the end-to-end stacking of antiparallel telomeric G-quadruplexes formed by the sequence AG3(T2AG3)3. To represent the experimental data, we developed a highly efficient coarse-grained fitting tool, which successfully described the samples as an equilibrium mixture of monomeric and dimeric G4 species. Our findings indicate that the antiparallel topology prevents the formation of long multimeric structures under self-crowding conditions, unlike the hybrid/parallel structures formed by the same DNA sequence. This result supports the idea that the stacking of monomeric G-quadruplexes is strongly affected by the presence of diagonal loops. [ABSTRACT FROM AUTHOR]

With the help of the one-dimensional random Potts-like model, we study the origins of fine structures observed on differential melting profiles of double-stranded DNA. We theoretically assess the effects of sequence arrangement on DNA melting curves through the comparison of results for random, correlated, and block sequences. Our results re-confirm the smearing out of the fine structure with the increase in chain length for all types of sequence arrangements and suggest that the fine structure is a finite-size effect. We have found that the fine structures on melting curves of chains comprised of blocks with correlations in sequence are more persistent, probably because of increased sequence disorder the blocks introduce. Many natural DNAs show a well-expressed fine structure of melting profiles. Our results for block sequences may suggest the existence of such sequence motifs in natural DNA sequences. [ABSTRACT FROM AUTHOR]

Enteroviruses are responsible for [approximately equal to] 10-15 million symptomatic illnesses in the United States annually; however, epidemiologic surveillance and genetic characterization of many enterovirus subspecies is limited (1-3). Coxsackievirus [...]

We report a prolonged and diffuse outbreak of listeriosis in Switzerland that involved 34 patients. The first case was reported in April 2022, and the outbreak peak occurred in 2023 [...]

As a kind of mycotoxin, aflatoxin B1 (AFB1), which is often found in agricultural products, poses a threat to human health. Developing a simple sensitive method for AFB1 detection is in great demand. Here, we reported an aptamer-based fluorescence assay for AFB1 detection by using DNAzyme to generate and amplify a signal. We redesigned a pair of DNA sequences, which originated from the anti-AFB1 aptamer and RNA-cleaving DNAzyme 10–23. In the absence of AFB1, the aptamer hybridized with the region of the substrate-binding arm of the DNAzyme, inhibiting the activity of the DNAzyme. In the presence of AFB1, the binding of AFB1 to the aptamer led to the displacement of the DNAzyme from the aptamer. The substrate-binding arm was unblocked, and the activity of the DNAzyme was restored for the hydrolysis of the fluorophore and quencher-labeled substrate, causing a significant fluorescence increase. This assay could detect AFB1 in the dynamic range from 0.98 to 2000 nmol/L with high selectivity, and the detection limit was 0.98 nmol/L. Moreover, the assay was able to detect AFB1 in a complex sample matrix. This work provides a useful tool for the analysis of AFB1. [ABSTRACT FROM AUTHOR]

In this study, a novel electrochemical biosensor was developed for the sensitive and selective detection of the Acinetobacter baumannii gene sequence. The biosensor was created by immobilizing a capture probe specific to the A. baumannii gene on the surface of chitosan-gold modified pencil graphite electrodes. Following solid-state hybridization on the Chit-Au/PGE surface, the target DNA sequence of the A. baumannii was detected by measuring the guanine signal using square wave voltammetry (SWV). All experimental parameters impacting sensor response are examined in order to improve hybridization efficacy, and the electrochemical biosensor's performance. The limit of detection (LOD) for the A. baumannii gene sequence was calculated and found to be 1.93 nM. Three different non-complementary DNA sequences were used to evaluate the assay selectivity, but no interference effect was obtained. Additionally, the potential applicability of the biosensor to real samples was tested in artificial serum media. The suggested electrochemical test procedure is simple, approachable, and quick, making it a convenient approach for the screening of DNA sequence. [Display omitted] • Pencil graphite electrodes were modified with chitosan-Au composite. • CHIT-Au/PGEs were used for electrochemical biosensing of A. baumannii gene • The [Fe(CN) 6 ]3-/4- oxidation signal was measured by SWV technique. • The proposed electrochemical assay is simple, rapid, and accessible. [ABSTRACT FROM AUTHOR]

Identifying distinct noncanonical structures in pathogenic genomes is crucial for developing new diagnostic tools. This study uncovers stable G-quadruplex (GQ) structures in conserved DNA sequences unique to the monkeypox virus (MPV). Furthermore, we developed a method for the detection of target GQ using a fluorogenic probe. [ABSTRACT FROM AUTHOR]

Cancer evolution is a multifaceted process leading to dysregulation of cellular expansion and differentiation through somatic mutations and epigenetic dysfunction. Clonal expansion and evolution is driven by cell-intrinsic and -extrinsic selective pressures, which can be captured with increasing resolution by single-cell and bulk DNA sequencing. Despite the extensive genomic alterations revealed in profiling studies, there remain limited experimental systems to model and perturb evolutionary processes. Here, we integrate multi-recombinase tools for reversible, sequential mutagenesis from premalignancy to leukemia. We demonstrate that inducible Flt3 mutations differentially cooperate with Dnmt3a , Idh2 , and Npm1 mutant alleles, and that changing the order of mutations influences cellular and transcriptional landscapes. We next use a generalizable, reversible approach to demonstrate that mutation reversion results in rapid leukemic regression with distinct differentiation patterns depending upon co-occurring mutations. These studies provide a path to experimentally model sequential mutagenesis, investigate mechanisms of transformation and probe oncogenic dependency in disease evolution. [Display omitted] • Somatic Flt3 mutations induce distinct acute and delayed transcriptional kinetics • Dnmt3a and Npm1 mutations differentially license mutant- Flt3 for transformation • Triple-recombinase models reveal that mutation order influences differentiation • Reversible Flt3 -mutant alleles possess divergent responses from chemical inhibition Bowman et al. develop orthogonal DNA recombinase tools to sequentially and reversibly control somatic mutations in leukemic evolution. These approaches identify how mutation timing and order influence transcriptional states in leukemia, and implicate cooperating mutations in dictating cell fate following oncogene reversion. These tools provide a path toward modeling clonal evolution in cancer. [ABSTRACT FROM AUTHOR]

The complexities encountered in microbial metabolic engineering continue to elude prediction and design. Unravelling these complexities requires strategies that go beyond conventional genetics. Using multiplex mutagenesis with double stranded (ds) DNA, we extend the multiplex repertoire previously pioneered using single strand (ss) oligonucleotides. We present ReaL-MGE (Recombineering and Linear CRISPR/Cas9 assisted Multiplex Genome Engineering). ReaL-MGE enables precise manipulation of numerous large DNA sequences as demonstrated by the simultaneous insertion of multiple kilobase-scale sequences into E. coli, Schlegelella brevitalea and Pseudomonas putida genomes without any off-target errors. ReaL-MGE applications to enhance intracellular malonyl-CoA levels in these three genomes achieved 26-, 20-, and 13.5-fold elevations respectively, thereby promoting target polyketide yields by more than an order of magnitude. In a further round of ReaL-MGE, we adapt S. brevitalea to malonyl-CoA elevation utilizing a restricted carbon source (lignocellulose from straw) to realize production of the anti-cancer secondary metabolite, epothilone from lignocellulose. Multiplex mutagenesis with dsDNA enables the incorporation of lengthy segments that can fully encode additional functions. Additionally, the utilization of PCR to generate the dsDNAs brings flexible design advantages. ReaL-MGE presents strategic options in microbial metabolic engineering. ReaL-MGE (Recombineering and Linear CRISPR/Cas9 assisted Multiplex Genome Engineering) enables precise insertion of large DNA sequences in multiple species. Here, authors demonstrate its utility in enhancing malonyl-CoA levels and producing valuable compounds like epothilone from lignocellulose. [ABSTRACT FROM AUTHOR]

Hereditary spherocytosis (HS) is the most prevalent form of congenital hemolytic anemia, being caused by genetic mutations in genes encoding red blood cell cytoskeletal proteins. Mutations in the ANK1 and SPTB genes are the most common causes of HS.; however, pathogenicity analyses of these mutations remain limited. This study identified three novel heterozygous mutations in 3 HS patients: c.1994 C > A in ANK1, c.5692 C > T, and c.3823delG in SPTB by whole-exome sequencing (WES) and validated by Sanger sequencing. To investigate the functional consequences of these mutations, we studied their pathogenicity using in vitro culture erythroblast derived from CD34 + stem cells. All three mutations lead to the generation of a premature stop codon. Real-time PCR assay revealed that the two SPTB mutations resulted in reduced SPTB mRNA expression, suggesting a potential role for the nonsense-mediated mRNA degradation pathway. For the ANK1 mutation, gene expression was not reduced but was predicted to produce a truncated version of the ANK1 protein. Flow cytometry analysis of red blood cell-derived microparticles (MPs) revealed that HS patients had higher MP levels compared to normal subjects. This study contributes to the current understanding of the molecular mechanisms underlying mutations in the ANK1 and SPTB genes in HS. [ABSTRACT FROM AUTHOR]

This study evaluates long-read and short-read sequencing for mitochondrial DNA (mtDNA) heteroplasmy detection. 592,315 bootstrapped datasets generated from two single-nucleotide mismatched ultra-deep sequenced mtDNA samples were used to assess basecalling error and accuracy, limit of heteroplasmy detection, and heteroplasmy detection across various coverage depths. Results showed high Phred scores of data with GC-rich sequence bias for long reads. Limit of detection of 12% heteroplasmy was identified, showing strong correlation (R2 ≥ 0.955) with expected heteroplasmy but underreporting tendency of high-level variants. Nanopore sequencing shows potential for direct applicability in mitochondrial diseases diagnostics, but stringent validation processes to ensure diagnostic result quality are required. [ABSTRACT FROM AUTHOR]

AbstractThis study aimed to analyze the clinical phenotype of the HBA2: c.95G>A mutation in the Chinese population and to provide guidance for clinical diagnosis and genetic counseling. Peripheral blood samples were collected from 16 patients, including 6 newborns, 2 children, and 8 adults. Hematological parameters and hemoglobin electrophoresis were analyzed, and genotypes were identified using methods such as PCR combined with reverse dot blot (RDB), nested PCR, gap polymerase chain reaction (Gap-PCR), and DNA sequencing. The results showed that 10 patients had mild anemia, 2 had moderate anemia, and 12 exhibited microcytic hypochromic features with MCV values ranging from 53 to 74.7 fl and MCH values from 16.2 to 25.4 pg. Additionally, 3 cases displayed obvious HbH + HbBarts bands (>15%). Among the 16 cases, various combinations of the HBA2: c.95G>A mutation were observed: one case had –α3.7 combined with HBA2: c.95G>A, another had –α4.2 combined with HBA2: c.95G>A, and five had –SEA combined with HBA2: c.95G>A, while the remaining cases were HBA2: c.95G>A heterozygotes. The study concludes that the HBA2: c.95G>A mutation in the α2 globin gene causes α+ thalassemia. When this mutation is combined with the Southeast Asian deletion (–SEA), it results in HbH disease, characterized by moderate microcytic hypochromic anemia and the presence of HbH + HbBarts bands. [ABSTRACT FROM AUTHOR]

Koelerioid grasses (subtribe Aveninae, tribe Poeae; Pooideae) resolve into two major clades, here called Koelerioid Clades A and B. Phylogenetic relationships among koelerioid grasses are investigated using plastid DNA sequences of rpl32‐trnL, rps16‐trnK, rps16 intron, and ITS regions, focusing on Trisetum, Acrospelion, and some annual species (Rostraria p.p. and Trisetaria p.p.) closely related to Trisetum flavescens in Koelerioid Clade A. Phylogenetic analyses of several selected data sets performed for 80 taxa and using maximum likelihood and Bayesian methods, revealed mostly congruent topologies in the nuclear and plastid trees, but also reticulation affecting several lineages. Trisetum is restricted to one species, T. flavescens, which is a sister to the clade formed by Trisetum gracile and Trisetaria aurea. The latter two species are classified here in the genus Graciliotrisetum gen. nov. The sister clade includes three species of Rostraria and Trisetaria lapalmae, all of which are classified here in a resurrected genus, Aegialina, which includes four species. Acrospelion is enlarged to include 13 species after the addition of other species formerly classified in Trisetum sect. Trisetum and T. sect. Acrospelion. We also transfer Trisetum ambiguum, Trisetum longiglume, and Koeleria mendocinensis to Graphephorum; and Helictotrichon delavayi to Tzveleviochloa, expanding these genera to eight and six species, respectively. We evaluate cases of reticulate evolution between Koelerioid Clades A and B and within Koelerioid Clade A, which probably gave rise to Graphephorum, Rostraria cristata, and Rostraria obtusiflora. Finally, we comment on polyploidy and biogeographic patterns in koelerioid grasses. We propose the following 26 new combinations: Acrospelion alpestre, Acrospelion altaicum, Acrospelion argenteum, Acrospelion bertolonii, Acrospelion buschianum, Acrospelion buschianum subsp. transcaucasicum, Acrospelion fuscum, Acrospelion laconicum, Acrospelion macrotrichum, Acrospelion rigidum, Acrospelion rigidum subsp. teberdense, Acrospelion tamonanteae, Acrospelion velutinum, Aegialina lapalmae, Aegialina pubescens, Aegialina pumila, Aegialina pumila subsp. fuscescens, Aegialina salzmannii, Aegialina salzmannii subsp. cossoniana, Graciliotrisetum aureum, Graciliotrisetum gracile, Graphephorum ambiguum, Graphephorum longiglume, Graphephorum mendocinense, Graphephorum orthochaetum, and Tzveleviochloa delavayi. Lectotypes are designated for the names Aegialitis tenuis, Aira melicoides, Avena aspera var. parviflora, Avena delavayi, Koeleria grisebachii var. mendocinensis, Koeleria pubescens subsp. cossoniana, Koeleria pumila, Koeleria salzmannii, Phalaris pubescens, Trisetum aureum, Trisetum cernuum, Trisetum fuscescens, Trisetum longiglume, and Trisetum wolfii; and we designate one neotype for Alopecurus litoreus. [ABSTRACT FROM AUTHOR]

Lithothamnion mirabile is currently considered a heterotypic synonym of L. muelleri. Three of the 14 specimens and additional fragments comprising the holotype of L. mirabile were selected for DNA sequencing as they best matched the protologue. The three 263-base pair rbcL sequences were distinct from each other, and were classified in the order Hapalidiales, but none were resolved in Lithothamnion. One sequenced specimen belongs in Roseolithon and was selected as the lectotype; thus we propose R. mirabile (Foslie) comb. nov. Another sequenced specimen groups with Phymatolithopsis and the third is well resolved in an undescribed genus of Hapalidiales sister to Boreolithothamnion. [ABSTRACT FROM AUTHOR]

Understanding of non-geniculate coralline algal (NGCA) diversity, prevalence, and distribution in the tropical western North Atlantic is currently limited by reliance on morphological and anatomical features that are inadequate for species identifications. Reef surveys from two study sites, the Florida Keys (Florida, USA) and Roatán (Honduras), provided prevalence data for the main benthic organisms as well as relative abundance for NGCA genera. NGCA collected during these surveys and from additional sites and depths were identified using DNA sequences from two plastid genes, psbA and rbcL, and two nuclear genes, SSU and LSU, and supported by morphological and anatomical descriptions. Bayesian and Maximum Likelihood phylogenetic analyses, along with species delimitation and genetic distance methods, resolved 11 genera within two orders. Within the Corallinales, the genera Dawsoniolithon, Porolithon and Spongites each had one species, Harveylithon and Lithophyllum had two, Titanoderma three, Hydrolithon four, Neogoniolithon eight, and an unknown genus within the subfamily Neogoniolithoideae had 11 putative species. Within the Hapalidiales, the genera Phymatolithon and Roseolithon each had one species. This study reinforces the inability of morphological and anatomical features to distinguish NGCA species in the tropical western North Atlantic, resulting in substantial underestimations of species richness and diversity. [ABSTRACT FROM AUTHOR]

Cytosine base editing is a powerful tool for making precise single nucleotide changes in cells and model organisms like zebrafish, which are valuable for studying human diseases. However, current base editors struggle to edit cytosines in certain DNA contexts, particularly those with GC and CC pairs, limiting their use in modelling disease-related mutations. Here we show the development of zevoCDA1, an optimized cytosine base editor for zebrafish that improves editing efficiency across various DNA contexts and reduces restrictions imposed by the protospacer adjacent motif. We also create zevoCDA1-198, a more precise editor with a narrower editing window of five nucleotides, minimizing off-target effects. Using these advanced tools, we successfully generate zebrafish models of diseases that were previously challenging to create due to sequence limitations. This work enhances the ability to introduce human pathogenic mutations in zebrafish, broadening the scope for genomic research with improved precision and efficiency. Cytosine base editing is crucial for modeling human diseases in zebrafish. Here, the authors present zevoCDA1 and zevoCDA1-198, optimized editors that improve editing efficiency and precision, allowing zebrafish modeling for disease-related mutations which were previously limited by DNA sequence contexts. [ABSTRACT FROM AUTHOR]

Purpose: Upper tract urothelial carcinoma (UTUC) and urothelial carcinoma of the bladder (UCB) share histomorphological and therapeutic features but distinct epidemiologic and clinicopathologic characteristics. We examined alterations of chromatin regulatory genes in molecular subtypes, clonal relatedness, and T‐cell receptor (TCR) diversity in UTUC and UCB. Materials and Methods: Targeted next‐generation sequencing or whole‐exome DNA sequencing and TCR sequencing were conducted with 34 UTUC and 49 UCB specimens from 63 patients. Tumors were subtyped based on the expression of CK5 and GATA3. Results of tissue microarray of 78 muscle‐invasive bladder cancer (MIBC) samples were used as prognostic factors of different subtypes of MIBC. Results: Chromatin regulatory genes were frequently mutated in both UTUC and UCB. Rapid relapse and progression of non‐MIBC are correlated with alterations of KMT2C and EP300. Frequency of alterations in chromatin regulatory genes is higher in UTUC patients with SBS22 and SBS2 signatures and lower in UCB patients with SBS2 and SBS6 signatures. GATA3 and CK5 double‐positive patients with higher frequencies of SMARCA4, ARID1A, and EP300 mutations have better prognoses than patients with basal subtypes. Although UTUC and UCB in the same patient can be either clonally related or developed independently, mutated genes in chromatin pathway were enriched in the related clones. Compared to UTUC, UCB had more deleterious mutations in DNA damage repair (DDR) genes, higher levels of tumor mutation burden (TMB) and copy number variations (CNVs), as well as higher TCR clonality and lower TCR diversity. Conclusions: Since genetic alterations of the chromatin pathway genes are important in both UTUC and UCB, they could serve as potential biomarkers for predicting disease progression and therapeutic targets. Differences in mutation frequencies of DDR pathway, TMB, CNV, and TCR might be the contributing factors for the distinct responses to immune checkpoint inhibitor (ICI) between UTUC and UCB. [ABSTRACT FROM AUTHOR]

Enhancers are cis-regulatory DNA sequences that are widely distributed throughout the genome. They can precisely regulate the expression of target genes. Since the features of enhancer segments are difficult to detect, we propose DeepDualEnhancer, a DNABert-based method using a multi-scale convolutional neural network, BiLSTM, for enhancer identification. We first designed the DeepDualEnhancer method based only on the DNA sequence input. It mainly consists of a multi-scale Convolutional Neural Network, and BiLSTM to extract features by DNABert and embedding, respectively. Meanwhile, we collected new datasets from the enhancer–promoter interaction field and designed the method DeepDualEnhancer-genomic for inputting DNA sequences and genomic signals, which consists of the transformer sequence attention. Extensive comparisons of our method with 20 other excellent methods through 5-fold cross validation, ablation experiments, and an independent test demonstrated that DeepDualEnhancer achieves the best performance. It is also found that the inclusion of genomic signals helps the enhancer recognition task to be performed better. [ABSTRACT FROM AUTHOR]

Forkhead box O4 (FOXO4), a human transcription factor, recognizes target DNA through its forkhead domain (FHD) while maintaining comparable binding affinity to non-target DNA. The conserved region 3 (CR3), a transactivation domain, modulates DNA binding kinetics to FHD and contributes to target DNA selection, but the underlying mechanism of this selection remains elusive. Using paramagnetic relaxation enhancement analysis, we observed a minor state of CR3 close to FHD in the presence of non-target DNA, a state absent when FHD interacts with target DNA. This minor state suggests that CR3 effectively masks the non-target DNA-binding interface on FHD. The interaction weakens significantly under high salt concentration, implying that CR3 or high salt concentrations can modulate electrostatic interactions with non-target DNA. Our 15N relaxation measurements revealed FHD's flexibility with non-target DNA and increased rigidity with target DNA binding. Our findings offer insights into the role of FOXO4 as a transcription initiator. FOXO4 CR3 masks the non-target DNA binding interface of FHD. FHD increases flexibility with non-target DNA and rigidity with target DNA, releasing CR3. Salt modulates the rate of FHD-DNA binding, discriminating between target and non-target DNA. [ABSTRACT FROM AUTHOR]

Serine integrases are phage- (or mobile element-) encoded enzymes that catalyse site-specific recombination reactions between a short DNA sequence on the phage genome (attP) and a corresponding host genome sequence (attB), thereby integrating the phage DNA into the host genome. Each integrase has its unique pair of attP and attB sites, a feature that allows them to be used as orthogonal tools for genome modification applications. In the presence of a second protein, the Recombination Directionality Factor (RDF), integrase catalyses the reverse excisive reaction, generating new recombination sites, attR and attL. In addition to promoting attR x attL reaction, the RDF inhibits attP x attB recombination. This feature makes the directionality of integrase reactions programmable, allowing them to be useful for building synthetic biology devices. In this report, we describe the degree of orthogonality of both integrative and excisive reactions for three related integrases (ϕC31, ϕBT1, and TG1) and their RDFs. Among these, TG1 integrase is the most active, showing near complete recombination in both attP x attB and attR x attL reactions, and the most directional in the presence of its RDF. Our findings show that there is varying orthogonality among these three integrases – RDF pairs. ϕC31 integrase was the least selective, with all three RDFs activating it for attR x attL recombination. Similarly, ϕC31 RDF was the least effective among the three RDFs in promoting the excisive activities of the integrases, including its cognate ϕC31 integrase. ϕBT1 and TG1 RDFs were noticeably more effective than ϕC31 RDF at inhibiting attP x attB recombination by their respective integrases, making them more suitable for building reversible genetic switches. AlphaFold-Multimer predicts very similar structural interactions between each cognate integrase – RDF pair. The binding surface on the RDF is much more conserved than the binding surface on the integrase, an indication that specificity is determined more by the integrase than the RDF. Overall, the observed weak integrase/RDF orthogonality across the three enzymes emphasizes the need for identifying and characterizing more integrase – RDF pairs. Additionally, the ability of a particular integrase's preferred reaction direction to be controlled to varying degrees by non-cognate RDFs provides a path to tunable, non-binary genetic switches. [ABSTRACT FROM AUTHOR]

Objective: To perform a collaborative review of the literature exploring the microsatellite instability/deficient mismatch repair (MSI/dMMR) phenotype in patients with upper tract urothelial carcinoma (UTUC). Method: A collaborative review of the literature available on Medline was conducted by the Cancer Committee of the French Association of Urology to report studies describing the genetic mechanisms, investigation, prevalence and impact of the MSI/dMMR phenotype in UTUC patients. Results: The predominant genetic mechanism leading to the MSI/dMMR phenotype in UTUC patients is related to the constitutional mutation of one allele of the MMR genes MLH1, MSH2, MSH6 and PMS2 within Lynch syndrome. Indications for its investigation currently remain limited to patients with a clinical suspicion for sporadic UTUC to refer only those with a positive testing for germline DNA sequencing to screen for this syndrome. With regard to technical aspects, despite the interest of MSIsensor, only PCR and immunohistochemistry are routinely used to somatically investigate the MSI and dMMR phenotypes, respectively. The prevalence of the MSI/dMMR phenotype in UTUC patients ranges from 1.7% to 57%, depending on the study population, investigation method and definition of a positive test. Younger age and a more balanced male to female ratio at initial diagnosis are the main specific clinical characteristics of UTUC patients with an MSI/dMMR phenotype. Despite the conflicting results available in the literature, these patients may have a better prognosis, potentially related to more favourable pathological features. Finally, they may also have lower sensitivity to chemotherapy but greater sensitivity to immunotherapy. Conclusion: Our collaborative review summarises the available data from published studies exploring the MSI/dMMR phenotype in UTUC patients, the majority of which are limited by a low level of evidence. [ABSTRACT FROM AUTHOR]

Purpose: We report two cases of fungal keratitis due to rare melanized fungal pathogens in ocular infection, one is the first case report of keratitis due to Pseudopithomyces maydicus and the second is keratitis due to rare fungal pathogen in ocular infections Phialophora chinensis. Method: Conventional mycology during routine diagnostics helped in identifying these rare fungal isolates, following which we proceeded for the confirmation of identification by DNA sequencing and did in-vitro antifungal susceptibility test to understand their susceptibility pattern. The clinical information for these two patients were collected from hospital electronic medical records. Results: We discuss the clinical presentation, treatment given, and clinical outcome in these patients and correlate these with the conventional microbiology and sequencing techniques, which helped in identifying the pathogen and the in-vitro antifungal susceptibility of these rare isolates. We also do a brief literature review for these two rare fungal pathogens. Conclusions: Pseudopithomyces maydicus and Phialophora chinensis are rare causes of fungal keratitis due to melanized fungi. Both of this fungal keratitis respond well to medical therapy alone if diagnosed and treated early. [ABSTRACT FROM AUTHOR]

Societal Impact Statement: The Jersey Daffodil Project, a secondary school initiative, boosted students' aspirations in science careers. Fieldwork, DNA sequencing and discussions with scientists enriched their learning and aspirations. Surveys revealed a positive shift in students' views towards STEM careers. The project also influenced an entire cohort to pursue biological sciences in 2021. The use of cutting‐edge technology reduced costs and time, showing promise for broader school collaboration. This project not only transforms science education but also inspires students, shaping their futures and fostering a new generation of scientists. [ABSTRACT FROM AUTHOR]

Meningioangiomatosis (MAM) remains a poorly understood lesion responsible for epileptic disease. In the past, MAM was primarily described in the context of neurofibromatosis type 2 before being mainly reported sporadically. Moreover, the malformative or tumoral nature is still debated. Because a subset of MAM are associated with meningiomas, some authors argue that MAM corresponds to an infiltration pattern of these tumors. For these reasons, MAM has not been added to the World Health Organization (WHO) Classification of Central Nervous System Tumors as a specific entity. In the present study, we characterized a series of pure MAM (n = 7) and MAM associated with meningiomas (n = 4) using histopathology, immunohistochemistry, genetic (fluorescent in situ and DNA sequencing analyses), and epigenetic (DNA‐methylation profiling) data. We evidenced two distinct morphological patterns: MAM with a fibroblastic‐like pattern having few lesional cells, and MAM with a more cellular pattern. A subset was associated with the genetic alterations previously reported in meningiomas (such as a KMT2C mutation and a hemizygous deletion of chromosome 22q including the NF2 gene). The DNA‐methylation profile, using a t‐distributed stochastic neighbor embedding analysis, evidenced that MAM (pure or associated with meningiomas) clustered in a separate group from pediatric meningiomas. The present results seem to suggest that MAM represents a neoplastic lesion and encourage the further study of similar additional series so that it may be included in a future WHO classification. [ABSTRACT FROM AUTHOR]

The use of environmental DNA (eDNA) is becoming prevalent as a novel method of ecological monitoring. Although eDNA can provide critical information on the distribution and biomass of particular taxa, the DNA sequences of an organism remain unaltered throughout its existence, which complicates the accurate identification of crucial events, including spawning. Therefore, we examined DNA methylation as a novel source of information from eDNA, considering that the methylation patterns in eggs and sperm released during spawning differ from those of somatic tissues. Despite its potential applications, little is known about eDNA methylation, including its stability and methods for detection and quantification. Therefore, we conducted tank experiments and performed methylation analysis targeting 18S rDNA through bisulphite amplicon sequencing. In the target region, eDNA methylation was not affected by degradation and was equivalent to the methylation rate of genomic DNA from somatic tissues. Unmethylated DNA, abundant in the ovaries, was detected in the eDNA released during fish spawning. These results indicate that eDNA methylation is a stable signal reflecting targeted gene methylation and further demonstrate that germ cell‐specific methylation patterns can be used as markers for detecting fish spawning. [ABSTRACT FROM AUTHOR]

Thrips (Thysanoptera), being eminent pests with significant agricultural implications on a global scale, cause considerable economic harm to an extensive array of ornamental, horticultural, and fruit crops. While morphological identification of thrips is often sufficient, the exploration of molecular methodologies can provide additional accuracy in species identification when needed. In this study, we used sequencing methods targeting the partial mitochondrial cytochrome oxidase subunit I (COI) gene to identify ten thrips species collected from 19 plant species in Brazil. Species-specific primers were developed based on COI and 28S sequences for precise identification of Caliothrips phaseoli and Frankliniella occidentalis. Bayesian phylogenetic analyses showed the absence of species clustering within their respective genus clades, while pairwise analyses demonstrated significant intraspecific variation in F. schultzei. [ABSTRACT FROM AUTHOR]

The dysfunction of the ABO glycosyltransferase (GT) enzyme, which is caused by mutations in the ABO gene, can lead to weak ABO phenotypes. In this study, we have discovered a novel weak ABO subgroup allele and investigated the underlying mechanism to causing its A weak phenotype. The ABO phenotyping and genotyping were performed by serological studies and direct DNA sequencing of ABO gene. The role of the novel single nucleotide polymorphism (SNP) was evaluated by 3D model, predicting protein structure changes, and in vitro expression assay. The total glycosyltransferase transfer capacity in supernatant of transfected cells was examined. The results of serological showed the subject was A weak phenotype. A novel SNP c.424A > G (p. M142V) based on ABO∗A1.02 was identified, and the genotype of the subject was AW-var/O.01 according to the gene analysis. In silico analysis showed that the SNP c.424A > G on the A allele may change the local conformation by damaging the hydrogen bonds and reduce the stability of GT. In vitro expression study showed that SNP p.M142V impaired H to A antigen conversion, although it did not affect the generation of A glycosyltransferase (GTA). One novel AW allele was identified and the SNP c.424A > G (p.M142V) can cause the A weak phenotype through damaging the hydrogen bonds and reducing stability of the GTA. [ABSTRACT FROM AUTHOR]

Purpose: To perform a molecular profiling of the metastases from papillary thyroid carcinomas (PTCs) and poorly differentiated thyroid carcinomas (PDTCs). Methods: We retrieved and analyzed the molecular and clinical features of 136 metastases from PTCs and 35 metastases from PDTCs subjected to targeted DNA sequencing, from cBioPortal. The clinicopathological data included the number and location of the metastases, and genomic data included mutations, translocations, copy number alterations and fraction of the genome altered (FGA). Results: Bone metastases from PTCs had a lower frequency of BRAF mutations than the lymph node metastases (LNMs) (43% vs 88%, p < 0.01), and a higher frequency of RBM10 and NRAS mutations than the LNMs (21% vs 3% for both, p < 0.05). The FGA of the bone metastases was higher than the FGA of the lung metastases (5.6% vs 1.3%, p < 0.05). The frequency of RET translocations was higher in the lung metastases from PTCs than the LNMs (15% vs 3%, p < 0.05). The LNMs from PTC patients harboring 4 or more distant metastases (DMs) had a higher frequency of TERT promoter mutations than the LNMs from patients harboring less than 4 DMs (96% vs 65%, p < 0.001). SDHA gene amplifications were enriched in the bone metastases from PDTCs and absent in the LNMs (38% vs 0%, p < 0.05). Conclusion: Metastases from PTCs and PDTCs harbor clinically relevant alterations affecting distinct body locations, such as NRAS and RBM10 mutations, RET translocations and SDHA amplifications that may be explored therapeutically. [ABSTRACT FROM AUTHOR]

In this study, from the perspective of image processing, we propose the iDNA-ITLM model, using a novel data enhance strategy by continuously self-replicating a short DNA sequence into a longer DNA sequence and then embedding it into a high-dimensional matrix to enlarge the receptive field, for identifying DNA methylation sites. Our model consistently outperforms the current state-of-the-art sequence-based DNA methylation site recognition methods when evaluated on 17 benchmark datasets that cover multiple species and include three DNA methylation modifications (4mC, 5hmC, and 6mA). The experimental results demonstrate the robustness and superior performance of our model across these datasets. In addition, our model can transfer learning to RNA methylation sequences and produce good results without modifying the hyperparameters in the model. The proposed iDNA-ITLM model can be considered a universal predictor across DNA and RNA methylation species. [ABSTRACT FROM AUTHOR]

Introduction: Thiamine-responsive megaloblastic anemia syndrome (TRMA) is a rare autosomal recessive disease with a homozygous or compound-heterozygous mutation in the SLC19A2 gene characterized by megaloblastic anemia, diabetes mellitus (DM), and sensorineural hearing loss with onset in childhood. Folic acid and vitamin B12 in serum are normal with dysplastic erythropoiesis in the bone marrow often mimicking myelodysplastic neoplasms (MDS) as a potential differential diagnosis. Thiamine substitution leads to normalization of anemia, without effects on hearing loss or DM. Case Presentation: We report about a 38-year-old male patient, presented with a 12-year history of anemia, insulin dependent DM, optic neuropathy, and a cataract since early childhood. The laboratory showed megaloblastic anemia. Other values were normal. The bone marrow smear showed dysplastic erythropoiesis with megaloblastic changes, and normal findings in cytogenetic and molecular genetic examinations. Next-generation sequencing-based diagnostics revealed a heterozygous missense variant in the SLC19A2 gene on the maternal allele and a 3.4 Mb inversion in the chromosomal region 1q24.2 with breaking points in FAM78B and SLC19A2 on the paternal allele. Treatment with oral thiamine 100 mg daily was initiated, and 12 weeks later hemoglobin levels and bone marrow morphology had normalized. Conclusion: Late-onset TRMA should be considered in adult patients with indicative comorbidities and a typical phenotype, which may mimic features of MDS. [ABSTRACT FROM AUTHOR]

Southwest Spain’s Río Tinto is a stressful acidic microbial habitat with a noticeably high concentration of toxic heavy metals. Nevertheless, it has an unexpected degree of eukaryotic diversity in its basin, with a high diversity of fungal saprotrophs. Although some studies on the eukaryotic diversity in Rio Tinto have been published, none of them used molecular methodologies to describe the fungal diversity and taxonomic affiliations that emerge along the river in different seasons. The aim of the present study was to isolate and describe the seasonal diversity of the fungal community in the Río Tinto basin and its correlation with the physicochemical parameters existing along the river’s course. The taxonomic affiliation of 359 fungal isolates, based on the complete internal transcribed spacer DNA sequences, revealed a high degree of diversity, identifying species belonging primarily to the phylum Ascomycota, but representatives of the Basidiomycota and Mucoromycota phyla were also present. In total, 40 representative isolates along the river were evaluated for their tolerance to toxic heavy metals. Some of the isolates were able to grow in the presence of 1000 mM of Cu2+, 750 mM of As5+ and Cd2+, and 100 mM of Co2+, Ni2+, and Pb2+ [ABSTRACT FROM AUTHOR]

Several Amaranthus hybridus L. biotypes present single and triple amino acid substitutions in EPSPS (5‐enolypyruvylshikimate‐3‐phosphate synthase). In resistant and tolerant species to glyphosate, differential susceptibility and alterations in the EPSPS sequence could cause modifications in their morphological–structural characteristics, an issue that is unknown for biotypes of A. hybridus. The aim of the work was to determine the susceptibility to glyphosate, the presence of mutations in EPSPS and the growth form in four biotypes of this weed from north‐central Argentina. To carry out this, a dose–response assay was conducted, to determine the presence of mutations in the EPSPS, DNA was extracted, amplified and sequenced, and finally, to evaluate the morphological–structural variations, the growth form was analysed. Our findings revealed that Ah03, Ah07 and Ah36 biotypes had resistance factors of 22 to 118. The EPSPS sequence remains unaltered in the Ah39 biotype, leading it to be considered a susceptible biotype. Biotypes Ah03 and Ah36 had heterozygous changes in the DNA sequence resulting in amino acid substitutions at positions 102, 103 and 106 being considered resistant biotypes, whereas in Ah07 some individuals showed unaltered DNA sequences and others exhibited heterozygous changes. The growth form was quantitatively different between biotypes, standing the most susceptible biotype that produced more seeds and required less thermal time for the appearance of the terminal inflorescence. The present investigation provides new evidence that resistant biotypes with the triple DNA mutation exhibit variations in the growth form compared to susceptible biotypes of this weed. [ABSTRACT FROM AUTHOR]

Hypermutation and malignant transformation are potential complications arising from temozolomide treatment of IDH‐mutant gliomas. However, the natural history of IDH‐mutant low‐grade gliomas without temozolomide treatment is actually under‐studied. We retrieved retrospectively from our hospitals paired tumors from 19 patients with IDH‐mutant, 1p19q non‐codeleted Grade 2 astrocytomas where no interim adjuvant treatment with either temozolomide or radiotherapy was given between primary resections and first recurrences. Tissues from multiple recurrences were available from two patients and radiotherapy but not temozolomide was given before the last specimens were resected. We studied the natural molecular history of these low‐grade IDH‐mutant astrocytomas without pressure of temozolomide with DNA methylation profiling and copy number variation (CNV) analyses, targeted DNA sequencing, TERTp sequencing, FISH for ALT and selected biomarkers. Recurrences were mostly higher grades (15/19 patients) and characterized by new CNVs not present in the primary tumors (17/19 cases). Few novel mutations were identified in recurrences. Tumors from 17/19 (89.5%) patients showed either CDKN2A homozygous deletion, MYC or PDGFRA focal and non‐focal gains at recurrences. There was no case of hypermutation. Phylogenetic trees constructed for tumors for the two patients with multiple recurrences suggested a lack of subclone development in their evolution when under no pressure from temozolomide. In summary, our studies demonstrated, in contrast to the phenomenon of temozolomide‐induced hypermutation, IDH‐mutant, 1p19q non‐codeleted Grade 2 astrocytomas which had not been treated by temozolomide, acquired new CNVs at tumor recurrences. These findings improve our understanding of the molecular life history of IDH‐mutant astrocytomas. [ABSTRACT FROM AUTHOR]

DNA lesions are linked to cancer, aging, and various diseases. The recognition and sequencing of special DNA lesions are of great interest but highly challenging. In this paper, an unnatural‐base‐pair‐promoting method for sequencing highly mutagenic ethenodeoxycytidine (εC) DNA lesions that occurred frequently is developed. First, a promising unnatural base pair of dεC–dNaM to recognize εC lesions is identified, and then a conversion PCR is developed to site‐precise transfer dεC–dNaM to dTPT3–dNaM for convenient Sanger sequencing. The low sequence dependence of this method and its capacity for the enrichment of dεC in the abundance of as low as 1.6 × 10–6 nucleotides is also validated. Importantly, the current method can be smoothly applied for recognition, amplification, enrichment, and sequencing of the real biological samples in which εC lesions are generated in vitro or in vivo, thus offering the first sequencing methodology of εC lesions at single‐base resolution. Owing to its simple operations and no destruction of inherent structures of DNA, the unnatural‐base‐pair strategy may provide a new platform to produce general tools for the sequencing of DNA lesions that are hardly sequenced by traditional strategies. [ABSTRACT FROM AUTHOR]

The jumping spider Marpissa marina (Goyen, 1892) is endemic to Aotearoa New Zealand. In this paper, we redescribe the species. Marpissa marina was placed in the Northern Hemisphere genus Marpissa due to its subjective similarity to spiders in that genus. The original description lacked modern morphological diagnoses and useful illustrations. The description was based largely on the female, and therefore lacked the taxonomically important characters seen in the male pedipalp. Previous literature has suggested M. marina may be related to the Australian genus, Maratus Karsch, 1878. We applied an integrative taxonomic approach based on diagnostic morphological characters and molecular analysis for a more detailed species description and assignment into an appropriate genus. Morphological features indicate that M. marina belongs to Maratus. This is supported by phylogenetic analyses using DNA sequences from the nuclear gene Actin 5C and the mitochondrial gene 16S ribosomal RNA. We transfer Marpissa marina into the genus Maratus, as Maratus marinus (Goyen, 1892), new combination. [ABSTRACT FROM AUTHOR]

A 50‐year‐old male with a pancreatic tail tumor underwent distal pancreatectomy. At 14 and 27 months after the primary surgery, metachronous liver metastases were identified and partial hepatectomies were performed for each. Pathologic findings of the primary pancreatic tumor were heterogeneous, but they essentially categorized into two components based on their cytologic features: (i) clear cell component and (ii) epithelioid cell component. The metastatic hepatic tumor was entirely composed of the epithelioid cell component. SMARCB1 expression was lost by immunohistochemistry and heterozygous deletion of SMARCB1 was identified by fluorescence in situ hybridization for both the primary and metastatic tumors. Targeted DNA sequencing of a metastatic hepatic tumor sample was performed and SMARCB1 loss was identified. Based on the morphologic, immunohistochemical, and molecular analyzes, the present case was difficult to classify into any of the existing entities. SMARCB1 deficiency might play a key role in the tumorigenesis. [ABSTRACT FROM AUTHOR]

Single nucleotide polymorphisms (SNPs) in homologous regions play a critical role in the field of genetics. However, genotyping these SNPs is challenging due to the presence of repetitive sequences within genome, which demand specific method. We introduce a new, mid-throughput method that simplifies SNP genotyping in homologous DNA sequences by utilizing a combination of multiplex kb level PCR (PCR size 2.5k-3.5 kb) for capturing targeted regions and multiplex nested PCR library construction for next-generation sequencing (Multi-kb level capture-seq). First of all, we randomly selected 7 SNPs in homologous regions and successfully captured 6-plex kb level amplicons (one of segments contains 2 SNPs, while the remaining segments each have only one SNP) in a single tube. And then, the amplification products were subjected to multiplex nested PCR for library construction and sequenced on Illumina platform. We tested this strategy using 600 amplicons from 100 samples and accurately genotyped 96.8% of target SNPs with a coverage depth of ≥ 15×. For the uniformity within the samples, over 66.7% (4/6) of the amplicons had a coverage depth above 0.2-fold of average sequencing depth. To validate the accuracy of this approach, we performed Ligase detection reaction PCR for genotyping the 100 samples, and found that the genotyping data was 97.71% consistent with our NGS results. In conclusion, we have developed a highly efficient and accurate method for SNP genotyping in homologous regions, which offers researchers a new strategy to explore the complex regions of genome. [ABSTRACT FROM AUTHOR]

Thyroid cancer (TC) represents one of the most prevalent endocrine malignancies, with a rising incidence worldwide. Epigenetic alterations, which modify gene expression without altering the underlying DNA sequence, have garnered significant attention in recent years. Increasing evidence underscores the pivotal role of epigenetic modifications, including DNA methylation, RNA methylation, and histone methylation, in the pathogenesis of TC. This review provides a comprehensive overview of these reversible and environmentally influenced epigenetic modifications, highlighting their molecular mechanisms and functional roles in TC. Additionally, the clinical implications, challenges associated with studying these epigenetic modifications, and potential future research directions are explored. [ABSTRACT FROM AUTHOR]

Variants in mitochondrial genomes (mtDNA) can cause various neurological and mitochondrial diseases such as mitochondrial myopathy, encephalopathy, lactic acidosis, stroke-like episodes (MELAS). Given the 16 kb length of mtDNA, continuous sequencing is feasible using long-read sequencing (LRS). Herein, we aimed to show a simple and accessible method for comprehensive mtDNA sequencing with potential diagnostic applications for mitochondrial diseases using the compact and affordable LRS flow cell "Flongle." Whole mtDNA amplification (WMA) was performed using genomic DNA samples derived from four patients with mitochondrial diseases, followed by LRS using Flongle. We compared these results to those obtained using Cas9 enrichment. Additionally, the accuracy of heteroplasmy rates was assessed by incorporating mtDNA variants at equimolar levels. Finally, mtDNA from 19 patients with Parkinson's disease (PD) was sequenced using Flongle to identify disease risk-associated variants. mtDNA variants were detected in all four patients with mitochondrial diseases, with results comparable to those obtained from Cas9 enrichment. Heteroplasmy levels were accurately detected (r2 > 0.99) via WMA using Flongle. A reported variant was identified in three patients with PD. In conclusion, Flongle can simplify the traditionally cumbersome and expensive mtDNA sequencing process, offering a streamlined and accessible approach to diagnosing mitochondrial diseases. [ABSTRACT FROM AUTHOR]

ABSTRACT One essential initial step in the analysis of ancient DNA is to authenticate that the DNA sequencing reads are actually from ancient DNA. This is done by assessing if the reads exhibit typical characteristics of post‐mortem damage (PMD), including cytosine deamination and nicks. We present a novel statistical method implemented in a fast multithreaded programme, ngsBriggs that enables rapid quantification of PMD by estimation of the Briggs ancient damage model parameters (Briggs parameters). Using a multinomial model with maximum likelihood fit, ngsBriggs accurately estimates the parameters of the Briggs model, quantifying the PMD signal from single and double‐stranded DNA regions. We extend the original Briggs model to capture PMD signals for contemporary sequencing platforms and show that ngsBriggs accurately estimates the Briggs parameters across a variety of contamination levels. Classification of reads into ancient or modern reads, for the purpose of decontamination, is significantly more accurate using ngsBriggs than using other methods available. Furthermore, ngsBriggs is substantially faster than other state‐of‐the‐art methods. ngsBriggs offers a practical and accurate method for researchers seeking to authenticate ancient DNA and improve the quality of their data. [ABSTRACT FROM AUTHOR]

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Probe-based nucleic acid enrichment represents an effective route to enhance the detection capacity of next-generation sequencing (NGS) in a set of clinically diverse and relevant microbial species. In this study, we assessed the effect of the enrichment-based sequencing on identifying respiratory infections using tiling RNA probes targeting 76 respiratory pathogens and sequenced using both Illumina and Oxford Nanopore platforms. Forty respiratory swab samples pre-tested for a panel of respiratory pathogens by qPCR were used to benchmark the sequencing data. We observed a general improvement in sensitivity after enrichment. The overall detection rate increased from 73 to 85% after probe capture detected by Illumina. Moreover, enrichment with probe sets boosted the frequency of unique pathogen reads by 34.6 and 37.8-fold for Illumina DNA and cDNA sequencing, respectively. This also resulted in significant improvements on genome coverage especially in viruses. Despite these advantages, we found that library pooling may cause reads mis-assignment, probably due to crosstalk issues arise from post-capture PCR and from pooled sequencing, thus increasing the risk of bleed-through signal. Taken together, an overall improvement in the breadth and depth of pathogen coverage is achieved using enrichment-based sequencing method. For future applications, automated library processing and pooling-free sequencing could enhance the precision and timeliness of probe enrichment-based clinical metagenomics. [ABSTRACT FROM AUTHOR]

Understanding the effects of microorganisms on coffee fermentation is crucial to ensure sensory quality and food security. The analysis of the dynamics of the microbial community during fermentation can contribute to a better understanding of the beneficial and harmful effects of microorganisms and help select starter cultures to improve coffee quality. Furthermore, the anaerobic environment produced by carbonic maceration of the coffee fruits inhibits aerobic respiratory processes and stimulates fermentative metabolism, modulating the microbial community during coffee fermentation. This study evaluated the effects of carbonic maceration in the fungal community dynamics during the fermentation of Coffea arabica fruits at 18, 28, and 38 °C for 24, 48, 72, 96, and 120 h. Fungal diversity was accompanied by high-throughput sequencing (NGS) of the Internal Transcribed Spacer (ITS) region. During the coffee fermentation, the fungal community changed over time, with the most significant changes occurring at 18 and 28 °C after 72 h. However, at 38 °C, there were greater variations in fungal composition and fungal diversity was highest after 120 h. The yeast Pichia cephalocereana was predominant in the fermentations. These results indicated that temperature and fermentation conditions influence the fungal community during coffee fermentation. Lower temperatures might favor a more stable microbial environment, while higher temperatures lead to more intense changes. Thus, our data from NGS can help in the identification, isolation, and metabolic characterization of fungi for the fermentation of coffee fruits. [ABSTRACT FROM AUTHOR]

Precise and efficient identification of single‐stranded nucleic acids is crucial for both pathological research and early diagnosis of diseases, such as cancers. Therefore, we have devised a novel biosensor that utilizes an elegantly designed handgrip‐ring structure sensing probe to enhance the detection sensitivity and reduce background signals. The handgrip‐ring structure sensing probe combines ring padlock‐based target recognition and hairpin structure probe‐based signal amplification. The target sequences form a binding interaction with the ring padlock in the sensing probe, leading to the elongation of the single‐stranded chain with the assistance of polymerase. This elongation step results in the release of the hairpin probe, triggering a signal amplification process. This design significantly minimized the potential discrepancies that may occur during the signal amplification process, hence bestowing the approach with a low level of background signals. By utilizing this innovative design, the current biosensor demonstrates a remarkable ability to detect miRNA with a limit as low as 376 aM and single‐stranded DNA sequences with a limit as low as 45.3 aM. In addition, it possesses exceptional discrimination capabilities. The efficacy of this approach in diagnosing targets was also effectively proved by the rational redesign of the ring padlock. [ABSTRACT FROM AUTHOR]

Noonan Syndrome (NS) is associated with an increased risk of low-grade central nervous system tumours in children but only very rarely associated with high-grade gliomas. Here we describe the first reported case of a spinal high-grade astrocytoma with piloid features (HGAP) in a child with NS. This case was a diagnostic and treatment dilemma, prior to whole-genome germline and tumour sequencing, tumour transcriptome sequencing and DNA methylation analysis. The methylation profile matched strongly with HGAP and sequencing identified somatic FGFR1 and NF1 variants and a PTPN11 germline pathogenic variant. Therapeutic targets were identified but also alterations novel to HGAP such as differential expression of VEGFA and PD-L1. The germline PTPN11 finding has not been previously described in individuals with HGAP. This case underscores the power of precision medicine from a diagnostic, therapeutic and clinical management perspective, and describes an association between HGAP and NS which has not previously been reported. [ABSTRACT FROM AUTHOR]

CRISPR/Cas9 is a popular genome editing technology, yet its clinical application is hindered by off-target effects. Many deep learning-based methods are available for off-target prediction. However, few can predict off-target activities with insertions or deletions (indels) between single guide RNA and DNA sequence pairs. Additionally, the analysis of off-target data is challenged due to a data imbalance issue. Moreover, the prediction accuracy and interpretability remain to be improved. Here, we introduce a deep learning-based framework, named Crispr-SGRU, to predict off-target activities with mismatches and indels. This model is based on Inception and stacked BiGRU. It adopts a dice loss function to solve the inherent imbalance issue. Experimental results show our model outperforms existing methods for off-target prediction in terms of accuracy and robustness. Finally, we study the interpretability of this model through Deep SHAP and teacher–student-based knowledge distillation, and find it can provide meaningful explanations for sequence patterns regarding off-target activity. [ABSTRACT FROM AUTHOR]

CRISPR/Cas9 has been applied to edit the genome of various organisms, but our understanding of editing outcomes at specific sites after Cas9-mediated DNA cleavage is still limited. Several deep learning-based methods have been proposed for repair outcome prediction; however, there is still room for improvement in terms of performance regarding frameshifts and model interpretability. Here, we present DeepIndel, an end-to-end multi-label regression model for predicting repair outcomes based on the BERT-base module. We demonstrate that our model outperforms existing methods in terms of accuracy and generalizability across various metrics. Furthermore, we utilized Deep SHAP to visualize the importance of nucleotides at various positions for DNA sequence and found that mononucleotides and trinucleotides in DNA sequences surrounding the cut site play a significant role in repair outcome prediction. [ABSTRACT FROM AUTHOR]

Coral microbiomes play crucial roles in holobiont homeostasis and adaptation. The host's ability to populate broad ecological niches and to cope with environmental changes seems to be related to the flexibility of the coral microbiome. By means of high-throughput DNA sequencing we characterized simultaneously both bacterial (16S rRNA) and Symbiodiniaceae (ITS2) communities of four reef-building coral species (Mussismilia braziliensis, Mussismilia harttii, Montastraea cavernosa, and Favia gravida) that differ in geographic distribution and niche specificity. Samples were collected in a marginal reef system (Abrolhos, Brazil) in four sites of contrasting irradiance and turbidity. Biological filters governed by the host are important in shaping corals' microbiome structure. More structured associated microbial communities by reef site tend to occur in coral species with broader geographic and depth ranges, especially for Symbiodiniaceae, whereas the endemic and habitat-specialist host, M. braziliensis, has relatively more homogenous bacterial communities with more exclusive members. Our findings lend credence to the hypothesis that higher microbiome flexibility renders corals more adaptable to diverse environments, a trend that should be investigated in more hosts and reef areas. [ABSTRACT FROM AUTHOR]

Prediction of chromatin interactions from DNA sequence has been a significant research challenge in the last couple of years. Several solutions have been proposed, most of which are based on encoder-decoder architecture, where 1D sequence is convoluted, encoded into the latent representation, and then decoded using 2D convolutions into the Hi-C pairwise chromatin spatial proximity matrix. Those methods, while obtaining high correlation scores and improved metrics, produce Hi-C matrices that are artificial - they are blurred due to the deep learning model architecture. In our study, we propose the HiCDiffusion, sequence-only model that addresses this problem. We first train the encoder-decoder neural network and then use it as a component of the diffusion model - where we guide the diffusion using a latent representation of the sequence, as well as the final output from the encoder-decoder. That way, we obtain the high-resolution Hi-C matrices that not only better resemble the experimental results - improving the Fréchet inception distance by an average of 11 times, with the highest improvement of 56 times - but also obtain similar classic metrics to current state-of-the-art encoder-decoder architectures used for the task. [ABSTRACT FROM AUTHOR]

Background: Transcription factors (TFs) bind to different parts of the genome in different types of cells, but it is usually assumed that the inherent DNA-binding preferences of a TF are invariant to cell type. Yet, there are several known examples of TFs that switch their DNA-binding preferences in different cell types, and yet more examples of other mechanisms, such as steric hindrance or cooperative binding, that may result in a "DNA signature" of differential binding. Results: To survey this phenomenon systematically, we developed a deep learning method we call SigTFB (Signatures of TF Binding) to detect and quantify cell-type specificity in a TF's known genomic binding sites. We used ENCODE ChIP-seq data to conduct a wide scale investigation of 169 distinct TFs in up to 14 distinct cell types. SigTFB detected statistically significant DNA binding signatures in approximately two-thirds of TFs, far more than might have been expected from the relatively sparse evidence in prior literature. We found that the presence or absence of a cell-type specific DNA binding signature is distinct from, and indeed largely uncorrelated to, the degree of overlap between ChIP-seq peaks in different cell types, and tended to arise by two mechanisms: using established motifs in different frequencies, and by selective inclusion of motifs for distint TFs. Conclusions: While recent results have highlighted cell state features such as chromatin accessibility and gene expression in predicting TF binding, our results emphasize that, for some TFs, the DNA sequences of the binding sites contain substantial cell-type specific motifs. [ABSTRACT FROM AUTHOR]