Non-coding Research Articles

MicroRNA‑24 alleviates colorectal cancer progression via a rs28382740 single nucleotide polymorphism in the long noncoding region of X‑linked inhibitor of apoptosis protein.

Colorectal cancer (CRC) is one of the most prevalent malignant diseases worldwide. Recurrence is associated with the poor survival of patients with CRC. Targeted therapy and precision medicine for recurrent CRC may improve the clinical outcome. Therefore, finding biomarkers that can detect CRC early, assess its prognosis and survival, and predict its treatment response is key to improving the clinical prognosis. The aim of this study was to assess CRC recurrence by analyzing molecular differences using postoperative specimens. Whole-exome sequencing was first used to evaluate the molecular differences in CRC tissues from patients with recurrent disease, and the results were then verified with tissue array methods. The regulation of single nucleotide polymorphisms (SNPs) in long noncoding regions of interest was analyzed in the presence of target microRNAs (miRs) using luciferase assays. The results demonstrated that in patients with recurrent CRC, the G allele was mainly detected at the rs28382740 SNP in the 3'-untranslated region of the X-linked inhibitor of apoptosis (XIAP)-encoding gene. From the tissue arrays, 60% (3/5) of patients with the G allele of the rs28382740 SNP were diagnosed with CRC recurrence, whilst only 10% (1/10) of patients without the G allele had recurrent CRC (P=0.077). Furthermore, XIAP levels were high in non-CRC (50%; 2/4) and CRC (75%; 3/4) tissues of patients with recurrent disease and CRC (54.5%; 6/11) tissues of patients without recurrent disease. However, but only 9.1% (1/11) of non-CRC tissues of nonrecurrent patients had significantly high XIAP expression levels (P=0.022). Using a luciferase assay, it was demonstrated that miR-24s (miR-24-1-5p and miR-24-2-5p) targeting the rs28382740 SNP reduced XIAP levels in CRC cells with rs28382740 SNP genotype G. These results indicate that apoptosis-related proteins, such as XIAP, may be therapeutic targets or biomarkers for tumor development. The data from the present study support an inhibitory effect of miR-24s on XIAP expression. However, this inhibitory potency depends on the rs28382740 SNP genotype and may alleviate CRC progression by regulating the expression of XIAP.

Ribosomal computing: implementation of the computational method

BackgroundSeveral computational and mathematical models of protein synthesis have been explored to accomplish the quantitative analysis of protein synthesis components and polysome structure. The effect of gene sequence (coding and non-coding region) in protein synthesis, mutation in gene sequence, and functional model of ribosome needs to be explored to investigate the relationship among protein synthesis components further. Ribosomal computing is implemented by imitating the functional property of protein synthesis.ResultIn the proposed work, a general framework of ribosomal computing is demonstrated by developing a computational model to present the relationship between biological details of protein synthesis and computing principles. Here, mathematical abstractions are chosen carefully without probing into intricate chemical details of the micro-operations of protein synthesis for ease of understanding. This model demonstrates the cause and effect of ribosome stalling during protein synthesis and the relationship between functional protein and gene sequence. Moreover, it also reveals the computing nature of ribosome molecules and other protein synthesis components. The effect of gene mutation on protein synthesis is also explored in this model.ConclusionThe computational model for ribosomal computing is implemented in this work. The proposed model demonstrates the relationship among gene sequences and protein synthesis components. This model also helps to implement a simulation environment (a simulator) for generating protein chains from gene sequences and can spot the problem during protein synthesis. Thus, this simulator can identify a disease that can happen due to a protein synthesis problem and suggest precautions for it.

The complete mitochondrial genome of the black-breasted thrush Turdus dissimilis (passeriformes: Turdidae)

The Back-breasted Thrush (Turdus dissimilis Blyth 1847), a medium-sized Turdus bird in the Turdidae family, is widely distributed in montane areas from northeastern India and Myanmar to southern China. The mitochondrial DNA of T. dissimilis is packaged in a compact 16,761-basepair (bp) circular molecule with A + T content of 52.50%. It contains 37 typical mitochondrial genes, including 13 protein-coding genes, 2 rRNAs and 22 tRNAs, and 1 noncoding region. We reconstructed a phylogenetic tree based on the mitogenome sequences of 10 Turdidae species and one outgroup. Phylogenetic analysis indicated that T. dissimilis is a sister taxon to T. unicolor. The new mitogenome data would provide useful information for application in conservation.

The complete mitochondrial genome of Triplophysa erythraea (Huang et al. 2019) (Cypriniformes, Nemacheilidae)

In this study, the complete mitochondrial genome of Triplophysa erythraea was determined for the first time. Results showed the mitogenome was 16 565 bp, including 2 ribosomal RNA genes, 22 transfer RNA genes, 13 protein-coding genes and 2 non-coding regions. The overall base composition of its mitochondrial genome was 28.9% A, 25.2% T, 28.7% C and 17.2% G. Phylogenetic tree revealed that Triplophysa erythraea had the closest relationship with Triplophysa xiangxiensis, which was also found in Xiangxi Tujia and Miao Nationality Autonomous Prefecture, Hunan province. In general, this study provided valuable information for conservation genetics analyses of Triplophysa erythraea and further displayed the evolution of species within the genus Triplophysa.

The complete mitochondrial genome of the shining leaf chafer Mimela junii (Duftschmidt, 1805) (Coleoptera: Scarabaeidae)

The complete mitochondrial genome of the shining leaf chafer Mimela junii was sequenced and is herein described. The mitogenome consists of a circular molecule of 16,805 bp, with an overall AT content of 75.7%. It encodes for 13 protein-coding genes (PCGs), 22 transfer RNA genes (tRNAs), two ribosomal RNA genes (rRNAs) and contains a non-coding Control Region (CR) characterized by the presence of tandem repeats. The gene order corresponds to the ancestral Pancrustacea model and mitogenome characteristics are congruous with those of hexapods. In the phylogenetic analysis, M. junii is nested within a paraphyletic Anomala with high support, and is herein associated with Anomala corpulenta with medium/low support.

Leveraging non-coding regions to guarantee the accuracy of small-sized panel-based tumor mutational burden estimates.

Accurate estimation of tumor mutational burden (TMB) as a predictor of responsiveness to immune checkpoint inhibitors in gene panel assays requires an adequate panel size. The current calculations of TMB only consider coding regions, while most of gene panel assays interrogate non-coding regions. Leveraging the non-coding regions is a potential solution to address this panel size limitation. However, the impact of including non-coding regions on the accuracy of TMB estimates remains unclear. This study investigated the validity of leveraging non-coding regions to supplement panel size using the OncoGuide NCC Oncopanel System (NOP). The aim of this study was to evaluate test performance against orthogonal assays and the association with responsiveness to immune checkpoint inhibitors was not included in the evaluation. We compared TMB status and values between TMB calculated only from coding regions (NOP-coding) and from both coding and non-coding regions (NOP-overall) using whole exome sequencing (WES) and FoundationOne®CDx (F1CDx) assay. Our findings revealed that NOP-overall significantly improved the overall percent agreement (OPA) with TMB status compared with NOP-coding for both WES (OPA: 96.7% vs. 73.3%, n = 30) and F1CDx (OPA: 90.0% vs. 73.3%). Additionally, the mean difference in TMB values compared with WES was lower for NOP-overall (3.55 [95% CI: 0.98-6.13]) than for NOP-coding (6.22 [95% CI: 3.73-8.70]). These results exemplify the utility of incorporating non-coding regions to maintain accurate TMB estimates in small-sized panels.

Characterization and phylogenetic analysis of the mitochondrial genome of Cylicostephanus longibursatus.

Cylicostephanus longibursatus is a common parasite in equine animals. Hosts infected by these nematodes might face disease or death. This study utilized next-generation sequencing technology to sequence the complete mitochondrial genome (mt genome) of C. longibursatus. Through bioinformatics techniques, the genomic base composition, codon usage, tRNA secondary structures, evolutionary relationships, and taxonomic status were analyzed. The results revealed that the mitochondrial genome of C. longibursatus is a double-stranded, 13,807-bp closed circular molecule with an AT content of 76.0%, indicating a clear preference for AT bases. The mitochondrial genome consisted of a total of 12 protein-coding genes, 22 tRNA genes, 2 rRNA genes, and 2 non-coding regions. Among the 12 protein-coding genes, TTG and ATT were the common start codons. TAA was the predominant termination codon, except for the ND3 and ND6 coding genes, and the COШ genes used TAG and "T" as termination codons, respectively. All tRNAs exhibited atypical clover-leaf secondary structures, except for tRNALys and tRNALeu2, where two tRNASer genes lacked DHU arms and DHU loops, tRNAmet lacked the TΨC-arm, tRNAIle lacked the TΨC-loop, and the remaining 16 tRNAs lacked the TΨC-arm and TΨC loop, which were substituted by the "TV-replacement loop". Phylogenetic analyses, based on the 12 protein-coding genes and utilizing maximum likelihood (ML) and Bayesian inference (BI) analyses, indicated that C. longibursatus did not form a monophyletic group with other Cylicostephanus but was instead more closely related to Cyathostomum. These research findings provide fundamental data for exploring the population classification and phylogeny of strongylid nematodes.

Comparative study on chloroplast genome of Tamarix species.

Tamaricaceae comprises about 120 species and has a long evolutionary history, Tamarix Linn accounts for approximately 75% of the total species in this family. It is the most widely distributed and diverse genus in the family. They have important ecological significance for transforming deserts and improving climate conditions. However, Tamarix is the most poorly classified genera among flowering plants owing to its large variability and high susceptibility to interspecific hybridization. In this study, the complete chloroplast genomes of three Tamarix species and one draft chloroplast genome were obtained in this study. Combined with eight chloroplast genomes deposited in GenBank, complete chloroplast sequences of 12 Tamarix species were used for further analysis. There are 176 non-SSR-related indels and 681 non-indel-related SSRs in the 12 Tamarix chloroplast genomes. The mononucleotide SSRs are the most prevalent among all types of SSRs. The mVISTA results indicate high sequence similarities across the chloroplast genome, suggesting that the chloroplast genomes are highly conserved, except for sample Tamarix androssowii (ENC850343). The IR regions and the coding regions are more conserved than the single-copy and noncoding regions. The trnF-ndhJ, ndhC-trnM-CAU, ycf1, and trnL-UAG-ndhF regions are the most variable and have higher variability than those of the universal DNA markers. Finally, the first phylogenetic tree of Tamaricaceae was constructed which confirmed the monophyly of Tamarix in Tamaricaceae. The first phylogenetic tree of Tamarix was based on the complete chloroplast genome to date, the changes in branch length and support rate can potentially help us clarify the phylogenetic relationships of Tamarix. All the obtained genetic resources will facilitate future studies in population genetics, species identification, and conservation biology of Tamarix.

Characterization of the Complete Mitochondrial Genome of Pleurogenoides japonicus (Digenea, Pleurogenidae): Comparison With the Members of Microphalloidea and Phylogenetic Implications.

Pleurogenoides japonicus (Trematoda: Microphalloidea) is an important parasite in wood frogs with high infection rates and significant ecological, economic, and societal importance. The scarcity of molecular data for these parasites severely limits population genetics and phylogenetic studies. In the present study, for the first time, we determined and described the entire mitochondrial (mt) genome of P. japonicus as the first representative of the family Pleurogenidae. The entire mt genome of P. japonicus was circular, with 15,043 bp (GenBank accession number OR900118), containing 36 genes, comprising 12 protein-coding genes (cox1-3, nad1-6, nad4L, cytb, and atp6), two ribosomal RNA genes, 22 transfer RNA genes, and two non-coding regions. There were 23 intergenic spacers, ranging from 2 to 162 bp, and only one 40 bp overlap between nad4L and nad4 genes in the P. japonicus mt genome. The nucleotide composition of P. japonicus mt genome exhibited a strong AT bias with a 63.75% A + T content, while the AT- and GC-skews were - 0.435 and 0.407, respectively. Comparative analysis demonstrated that the P. japonicus mt genome shared the most common characteristics with Microphalloidea trematodes, and the cox1 gene was the longest and most conserved gene in Microphalloidea trematodes. The gene arrangements of Xiphidiata trematodes were of the same order based on protein-coding genes and rRNA genes, except for tRNA. More than two gene arrangement types exist in Echinostomata and Xiphidiata, and the gene rearrangement events mainly occurred in "trnE-trnG" and "trnG-trnE". Phylogenetic analysis suggested that trematodes of the family Pleurogenidae clustered more with Prosthogonimidae than Eucotylidae. The mt genome data of P. japonicus provide an accurate genetic marker for further studies of Xiphidiata trematodes.

SAKit: An all-in-one analysis pipeline for identifying novel proteins resulting from variant events at both large and small scales.

Background: Genetic mutations that cause the inactivation or aberrant activation of essential proteins may trigger alterations or even dysfunctions in cellular signaling pathways, culminating in the development of precancerous lesions and cancer. Mutations and such dysfunctions can result in the generation of "novel proteins" that are not part of the conventional human proteome. Identification of these proteins carries a profound potential for unraveling promising drug targets and designing innovative therapeutic models. Despite the emergence of diverse tools for detecting DNA or RNA variants, facilitated by the widespread adoption of nucleotide sequencing technology, these methods primarily target point mutations and exhibit suboptimal performance in detecting large-scale and combinatorial mutations. Additionally, the outcomes of these tools are confined to the genome and transcriptome levels, and do not provide the corresponding protein information resulting from genetic alterations. Results: We present the development of Sequencing Analysis Kit (SAKit), a bioinformatics pipeline for hybrid sequencing analysis integrating long-read and short-read RNA sequencing data. Long reads are utilized for detecting large-scale variations such as gene fusions, exon skipping, intron retention, and aberrant expression in non-coding regions, owing to their excellent coverage capabilities. Short reads serve to validate these findings at breakpoints and splice junctions. Conversely, short reads are employed for identifying small-scale variations, including single nucleotide variants, deletions, and insertions, due to their superior sequencing depth, with long reads providing additional validation. SAKit is designed to perform analyses using inter-species configuration files comprising genome references and annotation data, making it applicable to both human and mouse studies. Furthermore, SAKit implements a hierarchical filtering approach to eliminate low-confidence variants and employs open reading frame (ORF) analysis to translate identified variants into protein sequences. Conclusion: SAKit is a robust and versatile bioinformatics tool designed for the comprehensive identification of both large-scale and small-scale variants from RNA-seq data, facilitating the discovery of novel proteins. This pipeline integrates analysis of long-read and short-read sequencing data, offering a powerful solution for researchers in genomics and transcriptomics. SAKit is freely accessible and open-source, available through GitHub (https://github.com/therarna/SAKit) and as a Docker image https://hub.docker.com/repository/docker/therarna). Implemented primarily within a Snakemake framework using Python, SAKit ensures reproducibility, scalability, and ease of use for the scientific community.

Widespread Impact of Natural Genetic Variations in CRISPR-Cas9 Outcomes.

The clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR-associated protein 9 (Cas9) is a genome editing tool widely used in biological research and clinical therapeutics. Natural human genetic variations, through altering the sequence context of CRISPR-Cas9 target regions, can significantly affect its DNA repair outcomes and ultimately lead to different editing efficiencies. However, these effects have not been systematically studied, even as CRISPR-Cas9 is broadly applied to primary cells and patient samples that harbor such genetic diversity. Here, we present comprehensive investigations of natural genetic variations on CRISPR-Cas9 outcomes across the human genome. The utility of our analysis is illustrated in two case studies, on both preclinical discoveries of CD33 knockout in chimeric antigen receptor-T cell therapy and clinical applications of transthyretin (TTR) inactivation for treating TTR amyloidosis. We further expand our analysis to genome-scale, population-stratified common variants that may lead to gene editing disparity. Our analyses demonstrate pitfalls of failing to account for the widespread genetic variations in Cas9 target selection and how they can be effectively examined and avoided using our method. To facilitate broad access to our analysis, a web platform CROTONdb is developed, which provides predictions for all possible CRISPR-Cas9 target sites in the coding and noncoding regulatory regions, spanning over 5.38 million guide RNA targets and 90.82 million estimated variant effects. We anticipate CROTONdb having broad clinical utilities in gene and cellular therapies.

Complete chloroplast genome sequence of Artemisia littoricola (Asteraceae) from Dokdo Island Korea: genome structure, phylogenetic analysis, and biogeography study.

The Asteraceae family, particularly the Artemisia genus, presents taxonomic challenges due to limited morphological characteristics and frequent natural hybridization. Molecular tools, such as chloroplast genome analysis, offer solutions for accurate species identification. In this study, we sequenced and annotated the chloroplast genome of Artemisia littoricola sourced from Dokdo Island, employing comparative analyses across six diverse Artemisia species. Our findings reveal conserved genome structures with variations in repeat sequences and junction boundaries. Notably, the chloroplast genome of A. littoricola spans 150,985bp, consistent with other Artemisia species, and comprises 131 genes, including 86 protein-coding, 37 tRNA, and 8 rRNA genes. Among these genes, 16 possess a single intron, while clpP and ycf3 exhibit two introns each. Furthermore, 18 genes display duplicated copies within the IR regions. Moreover, the genome possesses 42 Simple Sequence Repeats (SSRs), predominantly abundant in A/T content and located within intergenic spacer regions. The analysis of codon usage revealed that the codons for leucine were the most frequent, with a preference for ending with A/U. While the chloroplast genome exhibited conservation overall, non-coding regions showed lower conservation compared to coding regions, with the Inverted Repeat (IR) region displaying higher conservation than single-copy regions. Phylogenetic analyses position A. littoricola within subgenus Dracunculus, indicating a close relationship with A. scoparia and A. desertorum. Additionally, biogeographic reconstructions suggest ancestral origins in East Asia, emphasizing Mongolia, China (North East and North Central and South Central China), and Korea. This study underscores the importance of chloroplast genomics in understanding Artemisia diversity and evolution, offering valuable insights into taxonomy, evolutionary patterns, and biogeographic history. These findings not only enhance our understanding of Artemisia's intricate biology but also contribute to conservation efforts and facilitate the development of molecular markers for further research and applications in medicine and agriculture.

Genotypic Analysis of ABCA4 Coding Sequence in Thai Patients with Stargardt Disease

Objective: To study the mutational spectrum of the ABCA4 gene in Thai patients with Stargardt disease. Materials and Methods: DNA sequencing of all 50 exons of the ABCA4 gene was performed in nine Thai patients with clinically diagnosed Stargardt disease. Results: Amino acid sequence variation in the ABCA4 gene was found in five patients. Six missense mutations, c.71G>A, c.635G>A, c.1268A>G, c.3626T>C, c.4283C>T, and c.5761G>A, previously associated with Stargardt disease, were identified in our cohort. The variant c.1268A>G was the most prevalent in our study. Conclusion: In this cohort, only 56% of Thai Stargardt patients had missense mutations in the ABCA4 gene. Mutations in the non-coding regions of the ABCA4 or mutations in other genes may be responsible for Stargardt phenotypes in the remaining patients. Our findings are the first to reveal the mutational spectrum of ABCA4 leading to Stargardt disease in the Thai population and demonstrate a potential for ABCA4 screening as well as the importance of genetic variability in Thai patients with clinically suspected Stargardt disease.

An innovative CRISPR/Cas9 mouse model of human isolated microtia indicates the potential contribution of CNVs near HMX1 gene

BackgroundMicrotia is a prevalent congenital malformation, the precise etiology and pathogenesis of which remain elusive. Mutations in the non-coding region of the HMX1 gene have been implicated in isolated cases of microtia, emerging as a significant focus of contemporary research. Several pathogenic copy number variations (CNVs) proximal to the HMX1 gene have been documented in wild animal populations, whereas only a single large segmental duplication in this region has been identified in humans. However, the absence of a gene-edited animal model has impeded the investigation of the unclear gene function associated with HMX1 mutations in human isolated microtia. In this study, we sought to precisely identify the pathogenic mutation by analyzing three pedigrees alongside population controls. Subsequently, our objective was to develop a CRISPR/Cas9 gene-edited mouse model to elucidate the functional implications of the identified mutation. MethodsGenomic DNA was collected from 32 affected individuals across three pedigrees, as well as from 2,000 control subjects. Comprehensive genomic analyses, including genome-wide linkage analysis, targeted capture, second-generation sequencing, and copy number analysis, were conducted to identify potential mutations associated with congenital auricle malformation. CRISPR/Cas9 gene-edited murine models were generated in response to the identified mutation. The auricular phenotypes of these gene-edited mice were systematically monitored. Small-animal Micro-CT scanning was employed to identify potential craniofacial or skeletal abnormalities. Furthermore, the expression of the HMX1 gene in the PA2 region of mouse embryos was quantified using RT-qPCR. ResultsA co-segregated 600 base pair duplication located on chromosome 4 (chr4:8701900-8702500, hg19) was identified in affected individuals across three pedigrees, but was absent in healthy controls. Two types of CRISPR/Cas9 gene-edited mice were subsequently generated. The knock-in (KI) mouse model was engineered by inserting one copy of the duplicated sequence directly adjacent to the mutated site, whereas the knockout (KO) mouse model was created by excising the mutation sequence. The phenotypes of different group of CRISPR/Cas9 gene-edited mice demonstrated distinct auricular deformities. Furthermore, an increase in the copy number of the mutated sequence was associated with elevated expression levels of HMX1 in the gene-edited mouse model. ConclusionsIn this study, we further narrowed down and identified a 600 base pair copy number variation (CNV) located at chr4:8701900-8702500 (hg19), which is implicated in human bilateral, isolated microtia. Utilizing CRISPR/Cas9 technology, we developed novel mouse models harboring the identified mutation. These models serve as a robust platform for the comprehensive investigation of the underlying mechanisms of the disease.

HuR (ELAVL1) regulates the CCHFV minigenome and HAZV replication by associating with viral genomic RNA.

Crimean-Congo Hemorrhagic Fever virus (CCHFV) is a tick-borne pathogen that causes severe acute fever disease in humans and requires a biosafety level 4 laboratory for handling. Hazara virus (HAZV), belonging to the same virus genus as CCHFV, does not exhibit pathogenesis in humans. To investigate host RNA-binding proteins (RBPs) that regulate CCHFV replication, we generated a series of mutant RAW264.7 cells by CRISPR/Cas9 system and these cells were infected with HAZV. The viral titers in the supernatant of these cells was investigated, and HuR (ELAVL1) was identified. HuR KO RAW264.7 cells reduced HAZV replication. HuR is an RBP that enhances mRNA stability by binding to adenyl-uridine (AU)-rich regions in their 3' non-coding region (NCR). HuR regulates innate immune response by binding to host mRNAs of signaling molecules. The expression of cytokine genes such as Ifnb, Il6, and Tnf was reduced in HuR KO cells after HAZV infection. Although HuR supports the innate immune response during HAZV infection, we found that innate immune activation by HAZV infection did not affect its replication. We then investigated whether HuR regulates HAZV genome RNA stability. HAZV RNA genome was precipitated with an anti-HuR antibody, and HAZV genome RNA stability was lowered in HuR KO cells. We found that HuR associated with HAZV RNA and stabilized it to enhance HAZV replication. Furthermore, HuR-deficiency reduced CCHFV minigenome replication. CCHFV is a negative-strand RNA virus and positive-strand RNA is produced during replication. HuR was associated with positive-strand RNA rather than negative-strand RNA, and AU-rich region in 3'-NCR of S segment was responsible for immunoprecipitation with anti-HuR antibody and minigenome replication. Additionally, HuR inhibitor treatment reduced CCHFV minigenome replication. Our results indicate that HuR aids replication of the CCHFV minigenome by associating with the AU-rich region in the 3'-NCR.

Unraveling the Mitogenomic Characteristics and Phylogenetic Implications of Leuciscus merzbacheri (Zugmayer, 1912), an Endangered Fish in the Junggar Basin of Xinjiang, Northwest China.

Leuciscus merzbacheri is a rare and endangered fish in Xinjiang, China. As a representative species of the fauna in the Junggar Basin, it is of high economic and scientific value. The genetic data are still limited, and the mitochondrial genomic characteristics remain unexplored. A high-throughput sequencing method was used to obtain the complete mitogenome of L. merzbacheri. The full length of the circular DNA was 16,609 bp, and it consisted of 13 protein-coding genes (PCGs), 22 tRNAs, 2 rRNAs and 2 non-coding regions. The overall nucleotide compositions of both the mitogenome and PCGs showed an obvious AT preference with percentages of 54.20% and 53.60%, respectively. Three commonly used amino acids were Leu (16.43%), Ala (8.95%) and Thr (7.85%) in turn. All tRNAs could form the typical clover structures excluding tRNA-Ser AGY. The presumed secondary structures of two rRNAs contained several stem-loop domains, and the structure of 12S rRNA seemed to be more stable than that of 16S rRNA. Extended termination sequence regions (ETASs), central conserved regions (CSB-F, CSB-E and CSB-D), and conserved sequence regions (CSB-1, CSB-2 and CSB-3) were identified in the control region. The phylogenetic tree showed that L. merzbacheri was recovered with strong supports as a sister to the other members of the genus. The location in the outermost branch implied that it might be a relatively ancient species among its congeners. This study would complement the genetic data on L. merzbacheri and contribute to a better understanding of molecular evolution in Leuciscus as well.

Investigating the potential impact of ELF3-associated SNPs on chondrocyte inflammation

Objective: Chondrocyte inflammation is a critical factor in degenerative joint diseases, such as osteoarthritis (OA), significantly impairing quality of life through chronic pain and limited mobility. Genetic predisposition is recognized as a critical factor in the progression of chondrocyte inflammation and OA, with particular focus on the role of genetic variants in the expression and regulation of inflammatory mediators. This study aimed to obtain information about the potential roles of a few genes containing ELF3-associated SNPs in the pathogenesis of chondrocyte inflammation. Materials and Methods: GVAT Database was used to select top candidate SNPs associated with ELF3, a cardinal transcription factor in chondrocyte inflammation. Inflammation was induced by IL-1β treatment in differentiated chondrocytes to analyze gene expression patterns. Transcriptome analysis was done by RNA sequencing. Results: The most important SNPs that could potentially affect the binding affinity of ELF3 transcription factor were analyzed. As a result of the analysis, 52% of ELF3-associated SNPs were found in protein-coding regions, 40% in gene-free intergenic regions, and 8% in non-coding RNA sequences. Some of these SNPs are located in regulatory regions (enhancers). A significant increase in expression levels in the ELF3 gene was detected after IL-1β administration, indicating that IL-1β promotes the activity of this transcription factor. mRNA expressions of TLN2, BABAM2, PEPD, and NUDT5 were also increased after IL-1β stimulation. Conclusion: The presence of ELF3-associated SNPs in the enhancer sequences of TLN2 and BABAM2 genes, in addition to the increased expression of these two genes upon IL-1β stimulation, suggested that TLN2 and BABAM2 genes may be associated with the severity of chondrocyte inflammation and OA.

In-silico identification of simple sequence repeat (SSR) markers and phylogenetic analysis from chloroplast genomes of the genus Bambusa

Microsatellites or simple sequence repeats (SSRs), are short DNA sequences composed of repetitive units typically 1 to 6 base pairs long. These highly variable sequences are found in both protein-coding and noncoding DNA regions and exhibit a high degree of polymorphism. In the present study, chloroplast genomes of five species of Bambusa viz. B. bambos, B. multiplex, B. teres, B. vulgaris, and B. tulda, along with three outgroup species viz. Sorghum timorense, Hordeum vulgare, and Triticum aestivum were retrieved from NCBI in FASTA and GenBank formats and screened for chloroplast SSRs (cpSSRs) using MISA Perl script. The sequence analysis of the chloroplast genomes of different species of Bambusa confirmed the presence of 144 cpSSRs. The number of cpSSRs varied among the five bamboo species, with B. vulgaris exhibiting the highest count (31) and B. multiplex showing the lowest (27). Tetranucleotides were the most observed repeats in Bambusa, followed by mono-, di- and trinucleotides. Of the 144 SSR markers identified, only 16 (11.11 %) were suitable for primer design since the remaining sequences failed to produce acceptable primer pairs. An unweighted pair-group method with an arithmetic mean (UPGMA) dendrogram was constructed to analyze the genetic relationships among the genotypes. This analysis revealed three distinct clusters among the genotypes. SSR markers were found to be informative in differentiating the species of Bambusa and were likely to find potential applications in genetic fingerprinting, population genetics, conservation, and phylogenetics.

Storytelling of Hypertrophic Cardiomyopathy Discovery.

The discovery of hypertrophic cardiomyopathy (HCM) dates back to 1958, when the pathologist Donald Teare of the St. George's Hospital in London performed autopsies in eight cases with asymmetric hypertrophy of the ventricular septum and bizarre disorganization (disarray) at histology, first interpreted as hamartoma. Seven had died suddenly. The cardiac specimens were cut along the long axis, similar to the 2D echo. In the same year, at the National Institute of Health U.S.A., Eugene Braunwald, a hemodynamist, and Andrew Glenn Morrow, a cardiac surgeon, clinically faced a patient with an apparently similar morbid entity, with a systolic murmur and subaortic valve gradient. "Discrete" subaortic stenosis was postulated. However, at surgery, Dr. Morrow observed only hypertrophy and performed myectomy to relieve the obstruction. This first Braunwald-Morrow patient underwent a successful cardiac transplant later at the disease end stage. The same Dr. Morrow was found to be affected by the familial HCM and died suddenly in 1992. The term "functional subaortic stenosis" was used in 1959 and "idiopathic hypertrophic subaortic stenosis" in 1960. Years before, in 1957, Lord Brock, a cardiac surgeon at the Guy's Hospital in London, during alleged aortic valve surgery in extracorporeal circulation, did not find any valvular or discrete subaortic stenoses. In 1980, John F. Goodwin of the Westminster Hospital in London, the head of an international WHO committee, put forward the first classification of heart muscle diseases, introducing the term cardiomyopathy (dilated, hypertrophic, and endomyocardial restrictive). In 1995, the WHO classification was revisited, with the addition of two new entities, namely arrhythmogenic and purely myocardial restrictive, the latter a paradox of a small heart accounting for severe congestive heart failure by ventricular diastolic impairment. A familial occurrence was noticed earlier in HCM and published by Teare and Goodwin in 1960. In 1989-1990, the same family underwent molecular genetics investigation by the Seidman team in Boston, and a missense mutation of the β-cardiac myosin heavy chain in chromosome 14 was found. Thus, 21 years elapsed from HCM gross discovery to molecular discoveries. The same original family was the source of both the gross and genetic explanations of HCM, which is now named sarcomere disease. Restrictive cardiomyopathy, characterized grossly without hypertrophy and histologically by myocardial disarray, was found to also have a sarcomeric genetic mutation, labeled "HCM without hypertrophy". Sarcomere missense mutations have also been reported in dilated cardiomyopathy (DCM) and non-compaction cardiomyopathy. Moreover, sarcomeric gene defects have been detected in some DNA non-coding regions of HCM patients. The same mutation in the family may express different phenotypes (HCM, DCM, and RCM). Large ischemic scars have been reported by pathologists and are nowadays easily detectable in vivo by cardiac magnetic resonance with gadolinium. The ischemic arrhythmic substrate enhances the risk of sudden death.

Wide circulation of type 1 vaccine-derived poliovirus strains in clinical specimens from suspected cases of poliomyelitis, their contacts and in wastewater in Madagascar since late 2020

Madagascar has faced three major outbreaks of vaccine-derived polioviruses (VDPVs) in recent decades, with VDPV type 1 reemerging in late 2020. Here, we report the molecular characterization of these cVDPV1 strains. WHO protocols were used for poliovirus detection in stool and wastewater samples. Molecular genotyping was based on the 5′ non-coding (5′NC), VP1, and 3Dpol regions. From 2020 to 2022, 92 of 5690 stool samples and 129 of 1046 wastewater samples tested positive for cVDPV1. Genetic analysis of the VP1 gene revealed 1.3%–6.1% variability compared to the Sabin strain. Most sequences showed mutations at neurovirulence attenuation sites. Phylogenetic analysis distributed strains into four genogroups originating from Southern Madagascar. All analyzed cVDPV1 strains were recombinant, containing mutated oral polio vaccine sequences in VP1 and type C enterovirus sequences in other regions. This study demonstrated that all strains were closely related during this epidemic.