Search Results (426)

Background/Objectives: Autosomal recessive primary microcephaly is a rare and genetically heterogeneous disorder characterized by congenital non-syndromic microcephaly, with at least 28 causative genes identified to date. Biallelic variants in the CDK5RAP2 gene, an ultra-rare cause of autosomal recessive primary microcephaly, lead to Primary Autosomal Recessive Microcephaly 3 (MCPH3). Methods: We present seven patients from six families diagnosed with MCPH3 in light of clinical and molecular findings using whole-exome sequencing (WES). Furthermore, we investigated the effects of the identified intronic variants on splicing through RNA analysis. Results: Almost all patients had severe microcephaly, mild to moderate intellectual disability, speech delay, and cutaneous pigmentary abnormalities. Four patients presented with postnatal short stature, and two showed weight deficiency. Dysmorphic evaluation revealed that the most prominent features included brachycephaly, hypertelorism, epicanthus, high-arched eyebrows, prominent nasal bridge, and micrognathia. We identified five distinct homozygous CDK5RAP2 variants in our patients, including four novel variants. Segregation analysis verified that the parents were carriers. Two of these variants were intronic (c.3148+5G>C and c.383+4dupA), two were frameshift (c.3168del), and one was a nonsense variant (c.1591C>T). Both intronic variants disrupted splicing, generating a premature stop codon and resulting in a truncated protein. Conclusions: This study broadens the mutational landscape of CDK5RAP2. We also sought to demonstrate the functional consequences of the CDK5RAP2 intronic variants on gene function using RNA analysis. The identification of four novel variants underscores the importance of molecular diagnostics in patients with primary microcephaly and provides valuable data for genetic counseling and future functional studies. Full article

White-lipped tree frogs, Polypedates braueri, are currently included in the list of terrestrial wildlife with important ecological, scientific, and social value in China. Understanding the structure and characteristics of the mitochondrial genome provides essential information for resource conservation and phylogenetic analyses of P. braueri. While the complete mitochondrial genomes serve as important molecular markers for phylogenetic and genetic studies, the mitochondrial genome of P. braueri has received little attention. In this paper, we analyzed the characterization of the mitochondrial genome of P. braueri and investigated the phylogenetic relationships of Rhacophoridae. The complete mitochondrial genome of P. braueri was 20,254 bp in length, containing thirty-six genes (twelve protein-coding genes (PCGs), two ribosomal RNA genes (rRNAs), twenty-two transfer RNA genes (tRNAs)), three non-coding regions for the origin of light strand (O_L), and two control regions (CR). There were six overlapping regions and seventeen intergenic spacer regions in the mitogenome. The mitogenome also showed a bias towards A + T content (61.87%) and had negative AT-skew (−0.039) and GC-skew (−0.209). All the PCGs employed the ATG, ATA, or ATT as the start codon and TAA, TAG, AGG, or single T as the stop codon. Additionally, all of the tRNAs displayed a typical cloverleaf secondary structure, except trnS1, which lacked the D arm. The phylogenetic analysis, based on the maximum likelihood (ML) and Bayesian inference (BI) methods, revealed that Rhacophoridae could be classified into four monophyletic genera. The phylogenetic status of P. braueri was closely related to that of Polypedates megacephalus and Polypedates leucomystax. Additionally, selective pressure analysis suggested that COX1 and ND1 were highly efficient for discriminating closely related species in the genus Polypedates, while ND4L was the most appropriate marker for population-level genetic analyses. The diversification of the Polypedates commenced during the Late Oligocene and extended into the Miocene. The present study provides valuable genomic information on P. braueri and new insights into the phylogenetic relationships of Rhacophoridae. Full article

Listeria monocytogenes (L. monocytogenes) is an important foodborne pathogen that poses great risks to food safety and public health, and knowledge about its presence and diversity in potential sources is crucial for effectively tracking and controlling it in the food chain. In this study, we investigated the prevalence, antimicrobial susceptibility, and genomic characteristics of Listeria monocytogenes (L. monocytogenes) collected from retail chicken meat samples in Qingdao, China, in 2022. A total of 38 (10.6%, 38/360) L. monocytogenes isolates were recovered from 360 retail chickens. All 38 isolates were classified into two lineages (I and II), three serogroups (IIa, IIb, IIc), eight sequence types (STs), eight clonal complexes (CCs), eight Sublineages (SLs) and nine cgMLSTs (CTs). ST121 and ST9 were the most prevalent STs in this study. The ST121 strains from China had heterogeneity with those from other countries, while the Chinese ST9 strains had homogeneity with those from other countries. One resistance cassette tet(M)-entS-msr(D) was identified in eight L2-SL121-ST121-CT13265 isolates, the genetic structure of which was identical to that of three reference genomes. All isolates carried the L. monocytogenes pathogenic island (LIPI)-1, with only one carrying LIPI-3 and three carrying LIPI-4. In addition, 11 isolates subtyped as L2-SL121-ST121-CT13265 were found to have a premature stop codon (PMSC) in the inlA gene in this study. Our data revealed the antimicrobial susceptibility, genomic characteristics and evolutionary relationships of L. monocytogenes in retail chicken in Qingdao, China. The characterization of genotypes, virulence, stress and antimicrobial markers of strains circulating in retail chicken in Qingdao, as described in this study, provides the opportunity to improve risk assessments of L. monocytogenes exposure. Full article

Eryngium foetidum L. belongs to the Apiaceae family and is a perennial herb. The entire plant is rich in essential oils, which have a distinctive aroma similar to cilantro. This plant exhibits significant biological activity and possesses characteristics such as disease resistance and antimicrobial properties, showing great potential in medical and food applications. Additionally, its essential oil has substantial commercial value. Mitochondria play a crucial role as organelles within plant cells; however, the mitochondrial genome of E. foetidum remains underexplored. To fill this research gap, we conducted sequencing and assembly of the mitochondrial genome of E. foetidum, aiming to uncover its genetic mechanisms and evolutionary trajectories. Our investigation reveals that the mitochondrial genome of E. foetidum is a circular structure, similar to that of other species, with a length of 241,660 bp and a GC content of 45.35%, which is within the range observed in other organisms. This genome encodes 59 genes, comprising 37 protein-coding sequences, 18 tRNA genes, and 4 rRNA genes. Comparative analysis highlighted 16 homologous regions between the mitochondrial and chloroplast genomes, with the longest segment spanning 992 bp. By analyzing 37 protein-coding genes (PCGs), we identified 479 potential RNA editing sites, which induce the formation of stop codons in the nad3 and atp6 genes, as well as start codons in the ccmFC, atp8, nad4L, cox2, cox1, and nad7 genes. Meanwhile, the genome shows a preference for A/T bases and A/T-ending codons, with 32 codons having a relative synonymous codon usage (RSCU) value greater than 1. The codon usage bias is relatively weak and mainly influenced by natural selection. Most PCGs are under purifying selection (Ka/Ks < 1), while only a few genes, such as rps7 and matR, may be under positive selection. Phylogenetic analysis of mitochondrial PCGs from 21 species showed E. foetidum at the basal node of Apiaceae, consistent with the latest APG angiosperm classification and chloroplast genome-based phylogenetic relationships. In summary, our comprehensive characterization of the E. foetidum mitochondrial genome not only provides novel insights into its evolutionary history and genetic regulation but also establishes a critical genomic resource for future molecular breeding efforts targeting mitochondrial-associated traits in this economically important species. Full article

The Naozhou stock of large yellow croakers (Larimichthys crocea) exhibits unique phenotypic traits and high genetic diversity, making it a valuable resource for selective breeding and genetic conservation in aquaculture. Despite its importance, simple sequence repeat (SSR) markers have not been developed for this stock, which limits efforts in genetic evaluation, breeding optimization, and sustainable utilization of this commercially important species. In this study, 195,263 SSRs were identified from the genome of the Naozhou stock of large yellow croaker, covering a total length of 16,578,990 bp with a density of 288 bp/Mb. Dinucleotide repeats were the most common, with the AC motif being the most prevalent. The frequency of SSR markers ranged from 245.63 to 346.60 per Mb. A total of 30 primer pairs were synthesized, of which 28 pairs (93.3%) successfully amplified clear and reproducible bands in PCR assays. Among these, 28 SSR markers exhibited distinct and reproducible bands following gel electrophoresis. For eight SSR loci, the number of alleles (Na) ranged from 4 to 22 (mean = 11.375), while the effective number of alleles (Ne) ranged from 1.5401 to 10.4727 (mean = 5.6475). The assembled mitochondrial genome (mtDNA) was 16,467 bp in length and comprised 37 genes, including 13 protein-coding genes (PCGs), 22 tRNA genes, and 2 rRNA genes. The total sequence length of the PCGs was 11,431 bp, accounting for 69.4% of the mtDNA. A large portion of the PCGs (5) used incomplete stop codons (e.g., nad2, nad3, cox2), while others used TAA stop codons (e.g., nad6, nad5, TrnT). The mtDNA encoded a total of 3808 codons, with UAA showing the highest relative synonymous codon usage value. The SSR markers and mtDNA data generated in this study provide valuable tools for future genetic breeding and genomic research on the Naozhou stock of large yellow croakers. Full article

Listeria monocytogenes is a Gram-positive bacterial species that causes listeriosis, a major foodborne disease worldwide. The virulence factor inlA facilitates the invasion of L. monocytogenes into intestinal epithelial cells expressing E-cadherin receptors. Naturally occurring premature stop codon (PMSC) mutations in inlA have been shown to result in the production of truncated proteins associated with attenuated virulence. Moreover, different L. monocytogenes strains contain distinct inlA variants. In this study, we first characterized inlA in 546 L. monocytogenes strains isolated from various foods in Shanghai. The results showed that 36.1% (95% Confidence Interval: 32.0~40.2%) of the food isolates harbored inlA with PMSC, which was found to be associated with clonal complex (CC) types, with the highest proportions observed in CC9 and CC121. To investigate the function of inlA, we first used the dominant CC87 isolated from patients as the test strain and constructed an inlA-deleted strain via homologous recombination. Resistance tests and virulence tests showed that while inlA did not affect the resistance of L. monocytogenes, it significantly influenced cell adhesion and invasiveness. To further explore the function of inlA, we performed virulence tests on five CC-type strains carrying inlA with PMSC and their corresponding strains with intact inlA. We found that the virulence of L. monocytogenes strains carrying inlA or inlA with PMSC was associated with their CC type. Our preliminary results showed that premature termination of inlA did not significantly affect the adhesion and invasion abilities of low-virulence CC-type L. monocytogenes strains in Caco-2 cells, but substantially promoted those of high-virulence strains such as CC8 and CC7. In summary, this study preliminarily evaluated the effects of inlA integrity and PMSC mutation variation on the virulence of L. monocytogenes, providing a foundation for further research on inlA-related pathogenic mechanisms. Full article

Sulforaphane (SFN) is a bioactive compound belonging to the isothiocyanate family, known for its neuroprotective properties. While transcriptomic studies have highlighted SFN’s role in regulating gene expression, its impact on alternative splicing (AS), a key regulatory mechanism in neuronal metabolism, remains underexplored. In this study, we investigated whether SFN pre-treatment influences mRNA splicing patterns in an in vitro neuronal model using retinoic acid (RA)-differentiated SH-SY5Y cells. Using a dedicated RNA-seq-based splicing analysis pipeline, we identified 194 differential alternative splicing events (DASEs) associated with SFN treatment. Gene Ontology enrichment revealed significant over-representation of DNA repair processes. To better understand the functional implications, we integrated in silico predictions of premature stop codons, DASE/miRNA hybridizations, and DASE/RNA-binding protein (RBP) motif occurrences. Our findings suggest that SFN may modulate splicing of key DNA repair genes, contributing to protecting neurons against DNA damage. These preliminary results underscore a novel layer of SFN’s molecular effects and propose it as a valuable adjuvant in physiological conditions to enhance cellular health. Further studies are warranted to dissect the mechanistic underpinnings of SFN-mediated AS and its relevance in DNA-damage-related disorders. Full article

Bacteriophages of the order Crassvirales are highly abundant and near-universal members of the human gut microbiome worldwide. Zeta crAss-like phages comprise a separate group in the order Crassvirales, and their genomes exhibit greater variability than genomes of crAss-like phages from other families within the order. Zeta crAss-like phages employ multiple adaptation mechanisms, ensuring their survival despite host defenses and environmental pressure. Some Zeta crAss-like phages use alternative genetic coding and exploit diversity-generating retroelements (DGRs). These features suggest complex evolutionary relationships with their bacterial hosts, sustaining parasitic coexistence. Mutations in tail fiber proteins introduced by DGR can contribute to their adaptation to changes in the host cell surface and even expand the range of their hosts. In addition, the exchange of DNA polymerases via recombination makes it possible to overcome the bacterial anti-phage protection directed at these enzymes. Zeta crAss-like phages continuously adapt due to genetic diversification, host interaction tweaks, and counter-defense innovations, driving an evolutionary arms race with hosts. Based on the genome characteristics of the Zeta crAss-like phages, we propose to separate them into the Echekviridae family (“эчәк”—“intestines” in Tatar) following the tradition of using the word “intestines” in different languages, suggested previously. Full article

Zinc finger proteins are frequently implicated in a wide range of neurodevelopmental disorders (NDDs). In this study, we report a case of mild intellectual disability (ID), global developmental delay (GDD), and developmental coordination disorder (DCD) in an individual with unaffected parents. Trio-based whole-exome sequencing (WES) identified a de novo variant (c.1530dup, p.Glu511ArgfsTer16) in the ZNF496 gene of the proband. According to ACMG guidelines, this novel variant is classified as pathogenic. It creates a frameshift that introduces a premature stop codon, resulting in a truncated protein of 525 amino acids (compared to the wild-type 587 residues). Notably, NMDEscPredictor analysis predicted that the transcript escapes nonsense-mediated decay (NMD) despite the frameshift. Computational analyses suggest the potential pathogenetic effects of the identified variant. As documented, ZNF496 interacts with JARID2, a gene associated with NDDs, ID and facial dysmorphism (MIM: #620098). In silico analyses suggest that the identified mutation disrupts this interaction by deleting ZNF496’s C2H2 domain, potentially dysregulating JARID2 target genes. To our knowledge, this is the first reported association between ZNF496 and NDDs, and the variant has been submitted to the ClinVar database (SCV006100880). Functional studies are imperative to validate ZNF496’s role in NDDs and confirm the mutation’s impact on ZNF496-JARID2 interactions. Full article

Chionea is classified within the Tipuloidea superfamily and predominantly inhabits cold regions. However, its phylogenetic relationships remain contentious. In this study, the first three mitogenomes of Chionea (Diptera: Limoniidae) sampled in northeastern China (Jilin region) were sequenced, and their phylogenetic relationships were reconstructed by integrating these sequences with 30 additional Tipuloidea mitogenomes retrieved from NCBI. Unlike other Tipuloidea species, which are predominantly distributed in relatively warmer regions, this research investigates whether positive selection has acted on the mitogenomes of these three Chionea species due to environmental pressures, thereby elucidating key evolutionary drivers for Chionea. The three mitogenomes of Chionea exhibit characteristic features typical of insect mitogenomes, comprising 13 protein-coding genes (PCGs), 2 ribosomal RNA genes (16S rRNA and 12S rRNA), 22 transfer RNA genes (tRNA), and a single non-coding control region (D-loop). Notably, the secondary structure of trnS1 lacks the DHU arm in all three samples, and UUA (Leu) emerges as the most frequently utilized codon. Furthermore, the COX2 and ND5 genes utilize incomplete stop codons “T”. Utilizing these 13 PCGs, we reconstructed the internal phylogenetic relationships within Tipuloidea, revealing that Chionea tianhuashana and C. sphaerae form sister branches, while (C. tianhuashana + C. sphaerae) constitutes a sister branch to C. crassipes. Moreover, our analysis confirms the monophyly of Tipulidae, Tipula, and Nephrotoma as well as the polyphyly of Tipulinae, Chioneinae, and Limoniidae. In the branch site model analysis, three positively selected sites were detected when Chionea was designated as the foreground branches: COX3 (at position 242), ND5 (at position 535), and ND6 (at position 138). Full article

HIV-1 proviral landscapes were investigated using near-full-length HIV single-genome sequencing on blood samples from five children with vertically acquired infection and on ART for ~7–9 years. Proviral structures were compared to published datasets in children prior to ART, children on short-term ART, and adults on ART. We found a strong selection for large internal proviral deletions in children, especially deletions of the env gene. Only 2.5% of the proviruses were sequence-intact, lower than in the comparative datasets from adults. Of the proviruses that retained the env gene, >80% contained two or more defects, most commonly stop codons and/or gag start mutations. Significantly fewer defects in the major splice donor site (MSD) and packaging signal were found in the children on short or long-term ART compared to the adults, and tat was more frequently defective in children. These results suggest that different selection pressures may shape the proviral landscape in children compared to adults and reveal potentially different genetic regions to target for measuring the intact HIV reservoir and for achieving HIV remission in children. Full article

The SARS-CoV-2 ORF8 protein is a unique accessory viral protein among human coronaviruses, characterized by recurrent deletions and mutations with functional consequences. In this short report, we demonstrate that several dominant SARS-CoV-2 strains, despite encoding ORF8, fail to secrete the protein, revealing a recurring pattern of ORF8 functional impairment that cannot be detected by sequence analysis alone. In agreement with other studies, several high-frequency mutations were identified using the Nextstrain/augur pipeline, including G8Stop, Q27Stop, D119-/F120- double deletions, and nucleotide substitution C27889U, which occurred in XBB.1.5, Alpha, Delta, and BA.5.2 variants, respectively. Notably, the D119-/F120- deletions and C27889U substitution do not introduce premature stop codons, yet ORF8 secretion was lost in Delta and BA.5.2 virus-infected cultures. This indicates that the extracellular ORF8 function is impaired in these variants, resulting in ORF8 deficiency. Our findings highlight that the impairment of ORF8 secretion arises not only from premature stop codons but also from other mutations. Therefore, the functional validation of ORF8 secretion and activity is essential following sequence analysis to accurately assess ORF8’s role in SARS-CoV-2 infection. Full article

Listeria monocytogenes causes listeriosis, a severe foodborne disease with high mortality. Contamination with it poses significant risks to food safety and public health. Notably, genetic characteristic differences exist between strains causing human infections and those found in routine food inspections. This study examined the genotypic factors influencing the pathogenicity of L. monocytogenes, focusing on virulence gene profiles and key integrity genes like inlA to explain these divergences. The dataset included 958 strains isolated from human, food, and environmental samples. Whole-genome sequencing identified virulence genes, and principal component analysis (PCA) examined 92 virulence genes and inlA integrity to uncover potentially pathogenic patterns. The results highlight differences in virulence characteristics between strains of different origins. The integrity of inlA and genes such as inlD, inlG, and inlL were pivotal to pathogenicity. Strains with premature stop codons (PMSCs) in inlA, associated with reduced virulence, accounted for a low percentage of human cases but over 30% of food isolates. Sequence types (STs) like ST121, ST580, and ST199 showed unique profiles, while ST9, dominant in food, occasionally caused human cases, posing risks to vulnerable individuals. This research highlights the complexity of the pathogenicity of L. monocytogenes and emphasizes the importance of genomic surveillance for effective risk assessment. Full article

Enteric pathogens remain a health threat for children in low-income countries. A single nucleotide polymorphism (SNP) in the FUT2 gene that precludes the expression of fucosyltransferase 2 has been reported to influence the susceptibility to rotavirus and norovirus infections. The aim of this study was to investigate the association between G428A at rs601338 (stop codon variant) in the FUT2 gene and a range of enteric pathogens in children under 5 years of age. Rectal swab samples from 668 children (median age 13.6 months, 51% males, 93% rotavirus vaccinated, 468 with diarrhea) from Rwanda were analyzed via PCR for pathogen detection and SNP genotyping. A FUT2 stop codon (‘non-secretor’ status) was found in 19% of all children. Rotavirus was detected in 5.3% of non-secretors compared with in 13% of secretors (OR = 0.39, p = 0.019). Rotavirus P[8] was the predominant genotype and was found in 2.3% of non-secretors compared with 8.8% of secretors (p = 0.009). There was no association with any other pathogen, including noroviruses, of which 2 of 14 GII.4 infections were detected among non-secretors. Thus, the FUT2 stop codon variant was associated with rotavirus but not with any other pathogen. Full article

Background: Three-dimensional skeletal muscle organoids (3D SkMO) are becoming of increasing interest for preclinical studies in Duchenne muscular dystrophy (DMD), provided that the used platform demonstrates the possibility to form functional and reproducible 3D SkMOs, to investigate on potential patient-related phenotypic differences. Methods: In this study, we employed fibrin-based 3D skeletal muscle organoids derived from immortalized myogenic precursors of DMD patients carrying either a stop codon mutation in exon 59 or a 48–50 deletion. We compared dystrophic lines with a healthy wild-type control (HWT) by assessing microtissue formation ability, contractile function at multiple timepoints along with intracellular calcium dynamics via calcium imaging, as well as expression of myogenic markers. Results: We found patient-specific structural and functional differences in the early stages of 3D SkMO development. Contractile force, measured as both single twitch and tetanic responses, was significantly lower in dystrophic 3D SkMOs compared to HWT, with the most pronounced differences observed at day 7 of differentiation. However, these disparities diminished over time under similar culturing conditions and in the absence of continuous nerve-like stimulation, suggesting that the primary deficit lies in delayed myogenic maturation, as also supported by gene expression analysis. Conclusions: Our results underline that, despite the initial maturation delay, DMD muscle precursors retain the capacity to form functional 3D SkMOs once this intrinsic lag is overcome. This suggests a critical role of dystrophin in early myogenic development, while contraction-induced stress and/or an inflammatory microenvironment are essential to fully recapitulate dystrophic phenotypes in 3D SkMOs. Full article