Search Results (5,103)

Background: Coverage uniformity is pivotal in whole genome sequencing (WGS), as uneven read distributions can obscure clinically relevant variants and compromise downstream analyses. While enzyme-based fragmentation methods for WGS library preparation are widely used, they can introduce sequence-specific biases that disproportionately affect high-GC or low-GC regions. Here, we compare four PCR-free WGS library preparation workflows—one employing mechanical fragmentation and three based on enzymatic fragmentation—to assess their impact on coverage uniformity and variant detection. Results: Libraries were generated with Coriell NA12878 and DNA isolated from DNA blood, saliva, and formalin-fixed paraffin-embedded (FFPE) samples. Sequencing was performed on an Illumina NovaSeq 6000, followed by alignment to the human reference genome (GRCh38/hg38) and local realignment. We assessed coverage at both chromosomal and gene levels, including 504 clinically relevant genes detected in the TruSight™ Oncology 500 (TSO500) panel. Additionally, we examined the relationship between GC content and normalized coverage, as well as variant detection across high- and low-GC regions. Conclusions: Our findings show that mechanical fragmentation yields a more uniform coverage profile across different sample types and across the GC spectrum. Enzymatic workflows, on the other hand, demonstrated more pronounced coverage imbalances, particularly in high-GC regions, potentially affecting the sensitivity of variant detection. This effect was evident in analyses focusing on the TSO500 gene set, where uniform coverage is critical for accurate identification of disease-associated variants and for minimizing false negatives. Downsampling experiments further revealed that mechanical fragmentation maintained lower Single Nucleotide Polymorphism (SNPs) false-negative and false-positive rates at reduced sequencing depths, thereby highlighting the advantages of consistent coverage for resource-efficient WGS. This study introduces a novel framework for evaluating WGS coverage uniformity, providing guidance for optimizing library preparation protocols in clinical and translational research. By quantifying how fragmentation strategies influence coverage depth and variant calling accuracy, laboratories can refine their sequencing workflows to ensure more reliable detection of clinically actionable variants—especially in high-GC regions often implicated in hereditary disease and oncology. Full article

The genus Manglietia Blume is an important group of Magnoliaceae that has high economic and ornamental value. Owing to the small morphological differences among most Manglietia species and the limited sample sizes in previous molecular-level studies, its infrageneric classification remains unclear, and interspecific relationships for some species are still contentious. Clarifying the phylogenetic relationships within the genus Manglietia is crucial for species classification, genetic diversity assessment, and evolutionary developmental studies. This study sequenced, assembled, and annotated the chloroplast (cp) genomes of Manglietia guangnanica, Manglietia hookeri, and Manglietia longirostrata. The results indicated that these cp genomes are canonical quadripartite structures with total lengths of 160,067 bp, 160,067 bp, and 160,076 bp, respectively. All three cp genomes were annotated with 133 genes, comprising 88 protein-coding genes, 37 tRNAs, and 8 rRNAs. A total of 31, 30, and 30 dispersed repeats and 53, 53, and 56 SSRs were detected, respectively. ENC plot, neutrality plot, and PR2 plot analyses indicated that codon usage bias was influenced primarily by natural selection. Nucleotide diversity analysis revealed 8 highly variable regions in the cp genomes, among which petA-psbJ, rpl32-trnL, and ccsA-ndhD are recommended as candidate molecular markers for Manglietia species. Phylogenetic analysis revealed four highly supported clades: Clade I (18 species), Clade II (M. decidua only), Clade III (9 species), and Clade IV (M. caveana only). Among these clades, Clade IV is a newly discovered monotypic clade in this study, which differs from the results of all previous studies. Further investigations of Clades I and III, which include multiple Manglietia species, revealed that the presence or absence of hairs on Twigs, Stipules, and the abaxial surface of the leaf are important morphological characteristics for distinguishing species between these two clades. Furthermore, the results revealed that M. guangnanica and M. calcarea are two distinct species, and the treatment of M. longirostrata as a variety of M. hookeri was not supported by our study. This study enriches the cp genome data of Manglietia, provides new insights into infrageneric classification, and lays a foundation for further phylogenetic and evolutionary studies of Manglietia. Full article

Background: Biotinidase deficiency (BD) is a common autosomal recessive metabolic disorder in Qatar and the Arab world. It is treatable if detected early, making it essential to understand the genetic variants involved. This study aimed to investigate the carrier frequency of BD-related variants in a healthy Qatari population, reflecting the genetic landscape of the broader Middle Eastern region; classify them using bioinformatics tools; and compare findings with global datasets. Methods: Whole-genome sequencing data from 14,669 participants in the Qatar Genome Program (QGP), a multiethnic cohort including Qatari nationals and long-term residents (≥15 years), were analyzed to identify BTD variants. A total of 723, including 653 single-nucleotide polymorphisms (SNPs) and 70 structural variants (SVs) in BTD associated with BD, were screened against the Qatari cohort and compared with international data. In silico tools were used to assess variant pathogenicity, conservation, and protein stability. Molecular dynamics (MD) simulations were performed to evaluate structural and functional changes in the BTD. Results: A total of 80 SNPs and 3 SVs were identified, among which 21 variants (19 SNPs and 2 SVs) were classified as pathogenic or likely pathogenic, according to ClinVar. The carrier frequency of BTD-related variants in Qatar was 1:20, primarily driven by rs13078881 (D444H). Molecular dynamics (MD) simulations revealed significant conformational changes with H323R, D444H, and P497S, which demonstrated increased flexibility (higher RMSD/RMSF and PCA trace values). Additionally, R209C and D444H showed reduced compactness (higher Rg) and distinct energy minima, suggesting altered conformational states. Conclusions: This study demonstrates a high carrier frequency of pathogenic BTD variants in the Qatari population, underscoring the need to integrate these SNPs and SVs into the national genomic neonatal screening program (gNBS) for enhanced early detection and treatment strategies. The mild structural deviations observed in the D444H mutant through MD simulations may explain its association with milder clinical phenotypes of BTD, offering valuable insights for personalized therapeutic approaches. Full article

Bacillus paranthracis, a species of the genus Bacillus, is a Gram-positive bacterium classified as an opportunistic pathogen that can cause foodborne diarrhea and other intestinal diseases in humans and various animals. To date, there has been limited research on B. paranthracis, and there are few records of this bacterium being isolated from animal intestines. In this study, a strain named Qf-1 was isolated and purified from faecal samples of mink. Through culturomics, 16S rRNA gene sequencing, whole-genome sequencing, and average nucleotide identity (ANI) analysis, the strain was confirmed to be B. paranthracis. Whole-genome sequencing revealed that the strain has a genome size of 5.27 Mb, comprising one chromosome (5,224,739 bp) and one plasmid (51,964 bp). Functional annotation of its genome identified multiple potential pathogenic factors associated with pneumonia, including the key genes AsbD and AsbF, which facilitate bacterial colonisation of the lungs and trigger inflammatory responses, as well as EsxB and EsxL, which exacerbate lung inflammation and promote infection spread. Comparative genomics analysis revealed that this strain shares a close evolutionary relationship with previously reported B. paranthracis strains. The structure and function of the bacterial genes were analyzed in depth using multi-omics methods. Through mouse pathogenicity experiments, it was found that this bacterium may cause pneumonia and enteritis in mice. We predict that it may also pose a threat to the health of the mink. These research findings contribute to the establishment of a stable experimental model between pathogens and mink hosts, laying the foundation for further elucidating their pathogenicity and pathogenic mechanisms. This is of great significance for the diagnosis and prevention of bacterial diseases in mink in the future. Full article

The Atlas cedar Cedrus atlantica is a relict and endemic conifer from Morocco and Algeria, although plantations may be found in several locations aside from its natural range. Recurrent droughts have been widely related to Atlas cedar dieback, growth decline, and mortality, but the genetic basis of potential adaptive capacity is unknown. We used the double digest restriction-site associated DNA sequencing technique (ddRAD-seq) to describe the genetic structure and variability of Atlas cedar along an aridity gradient in Morocco. Furthermore, we investigated the potential genetic origin of three Spanish plantations, also along an aridity gradient. The obtained single nucleotide polymorphisms (SNPs) were used to perform genotype–environment associations (GEAs) to define SNPs related to bioclimatic variables of temperature and precipitation. The vulnerability of this species to environmental variations was also estimated by its risk of non-adaptedness (RONA). Population structure showed a divergence between the Moroccan natural stands and some of the Spanish plantations, with each Moroccan nucleus being genetically distinct. The genetic variability was significantly lower in plantations than in natural populations. The drier Spanish plantations (easternmost) were genetically very similar to the driest Moroccan population (southernmost), suggesting that as its origin. A total of 41 loci under selection were obtained with the Moroccan dataset. In relation to temperature and precipitation variables, isothermality showed the highest number of associated loci (10) in GEA studies, and genotype–phenotype associations (GPAs) showed one locus associated with the Specific Leaf Area. RONA value was higher in the southernmost High Atlas population, where rising temperature was the main driver of expected genetic offset by allele frequency changes under the worst emissions scenario. In contrast, Spanish plantations would need smaller genetic changes to cope with the expected climate change. Likely gene flow from southern to northern areas suggests a latitudinal heading, where Spanish plantations might operate as an assisted migration. Moreover, one locus showed a northern/southern pattern in saplings but not in adults, suggesting a potential latitudinal pattern of selection. Our results are discussed on the basis of their management and conservation. Full article

Background/Objectives: Pasteurella multocida is responsible for a wide variety of animal diseases worldwide, causing major economic losses. These infections are usually treated with antibiotics; however, the emergence of multidrug-resistant (MDR) strains is increasingly hindering. Understanding antibiotic resistance in P. multocida is important for effective treatment strategies and public health, as it impacts both animal and human welfare. Methods: In this study, the antibiotic susceptibility of 80 P. multocida isolates was evaluated by phenotypic (disk diffusion and broth microdilution) and genotypic analysis via whole-genome sequencing, with particular attention to the occurrence of MDR strains. The strains were tested against antibiotics from nine antimicrobial classes (penicillins, cephalosporins, aminoglycosides, tetracyclines, macrolides, fluoroquinolones, lincosamides, phenicols, and sulfonamides). Antimicrobial resistance gene (ARG) sequences and single-nucleotide polymorphisms (SNPs) were evaluated in paired reads using the Bacterial and Viral Bioinformatics Resource Center (BV-BRC) and the Comprehensive Antibiotic Resistance Database (CARD) via Resistance Gene Identifier (RGI), respectively. Results: Phenotypic results indicated that cephalosporins and phenicols were the most effective drugs against P. multocida; however, the majority of strains were also susceptible to fluoroquinolones and tetracyclines. In contrast, high resistance rates were observed to sulfamethoxazole and clindamycin. The most prevalent resistance genes were strA, sul2, and tetH, while none of the strains harbored the bla-_TEM or erm (42) genes. Conclusions: Of the two phenotypic methods, MIC values showed a stronger positive correlation with genotypic results, making it a more suitable method for determining antibiotic susceptibility. The phenotypic results for phenicols, tetracyclines, and fluoroquinolones showed a strong correlation with the detected resistance genes. In contrast, resistance to sulfamethoxazole, β-lactams, and macrolides remained genetically unexplained, suggesting the existence of additional resistance mechanisms to be explored. Full article

The mitochondrial genome and 5S rDNA are powerful tools for exploring and confirming species identity and understanding evolutionary trajectories. In addition, evaluating the nutritional value of Cambaroides dauricus by determining and analyzing the nutritional components of its muscles could provide essential data for relevant artificial breeding and food processing. In this study, for the first time, we characterized the mitochondrial genome of Cambaroides dauricus using Illumina sequencing technology. The results showed that the mitochondrial genome was a circular genome of 16,215 bp in length. The average sequencing depth of the mitochondrial genome was 100X. The nucleotide composition of the mitochondrial genome was 33.4% A, 39.1% T, 11.0% C and 16.5% G. Phylogenetic analyses showed that Cambaroides dauricus has a very close relationship with Cambaroides wladiwostokiensis. The results of 5S rDNA showed that the genetic relationship between Procambarus clarkii and Cambaroides dauricus is quite close. After determining the nutritional components of Cambaroides dauricus’ muscles with normal analysis methods, the results revealed that it has a crude protein content of 18.47 ± 0.87 per 100 milligrams, a crude fat content of 0.83 ± 0.12 per 100 milligrams, a crude ash content of 0.63 ± 0.06 per 100 milligrams and a moisture content of 79.8 ± 0.72 per 100 milligrams. There are eight essential amino acids in its protein, accounting for 41.59% of the total amino acids, and the proportion of umami amino acids is 36.27%. This study will provide a valuable basis for further studies of taxonomy, phylogenetic analyses and artificial breeding in Cambaroides dauricus. Full article

Sheep pox, caused by sheep pox virus (SPV), is a transboundary disease that threatens sheep production and trade. This study aimed to identify genetic, immunological, and biochemical markers associated with susceptibility to SPV in Barki ewes. A total of 100 adult ewes were examined, including 50 clinically healthy and 50 naturally infected animals. PCR detected SPV DNA in 60% of suspected scab samples, highlighting diagnostic challenges in field investigations. Blood samples were analyzed for hematological indices, cytokine profiles, acute phase proteins, oxidative stress biomarkers, iron metabolism, and hormonal parameters. Expression profiles and single-nucleotide polymorphisms (SNPs) in 15 immune and antioxidant genes were characterized from cDNA-derived sequences. Infected animals exhibited microcytic hypochromic anemia, leukocytosis, elevated proinflammatory cytokines, and reduced IL-10. Acute phase proteins, oxidative stress markers, and cortisol were increased, whereas antioxidant capacity and transferrin were reduced. Twenty-three SNPs were identified, including non-synonymous variants, which showed promising but unvalidated associations with disease status. These findings highlight immune, oxidative, and genetic alterations in SPV-infected sheep, but further longitudinal and cross-validated studies are needed to establish their diagnostic or breeding utility. Full article

The genus Corynebacterium is commonly isolated from camel uteri, yet it is rarely identified to the species level. During our routine clinical examination of she-camels brought to the hospital with history of reproductive and systemic health issues, four isolates from the uterus and one isolate from blood could not be assigned to any valid Corynebacterium species. Therefore, we aim to identify these isolates, determine any potential virulence factors, and describe how gene turnover contributed to the evolution of these species. Genome-based and phenotypic identification, along with resistome, mobilome, virulome and phylogenomics analysis, was used to characterize the isolates. The isolates were Gram stain-positive, catalase-positive, and rod-shaped. The isolates were assigned to the genus Corynebacterium based on 16S rRNA gene sequence similarity and phylogenetic analysis. The isolates 3274 and ayman were classified as two new Corynebacterium species based on the average nucleotide identity (ANI) values of 78.46% and 68.88% and digital DNA–DNA hybridization (dDDH) values of 20.9% and 22.4%. The isolates 2581A, 2583C, and 4168A constitute a single Corynebacterium species based on their pairwise ANI value of 99% and dDDH value of more than 90%. In addition, isolates 2581A, 2583C, and 4168A showed ANI values of 75.99%, 75.86%, and 76.04% and dDDH values of 23.1%, 23%, and 22.5% with closely related species, and were designated as single new Corynebacterium species. Genes for mycolic acid and menaquinone biosynthesis were detected in all isolates. The isolates were susceptible to ceftiofur, linezolid, penicillin, erythromycin, and tetracycline. All isolates harbored the antiseptic resistance gene qacA. Moreover, virulence factors involved in cell adhesion and iron acquisition were detected. The evolution of these species is dominated by gene gain rather than gene loss. The majority of these genes are acquired through horizontal gene transfer, mediated by prophages and genomic islands. In summary, we characterized three new Corynebacterium species, expanding the number of new Corynebacterium species from animals. Moreover, we described the mechanism underlying the genome evolution of these new species. The clinical findings and detection of virulence genes highlight the significance of these isolates as possible pathogens, contributing to the development of endometritis in camels. Full article

As a globally significant fruit crop, litchi (Litchi chinensis Sonn.) exhibits substantial variation in seed size, which is a key determinant of fruit quality. However, the lack of molecular markers closely associated with seed-related traits has hindered targeted breeding efforts. In this study, we systematically evaluated six critical traits—single fruit weight, seed weight, seed length, seed width, edible rate, and seed-to-fruit weight ratio—across 131 early-maturing litchi accessions. Hierarchical clustering analysis (HCA) and principal component analysis (PCA) revealed a clear bifurcation of these accessions into two distinct groups based on seed size-related traits. Using bulked segregant analysis sequencing (BSA-seq), we identified a candidate genomic region (24.93–25.69 Mb) on chromosome 5, potentially regulating litchi seed size. Within this region, 1600 single-nucleotide polymorphisms (SNPs) and 314 insertion/deletion mutations (InDels) exhibited significant divergences between the extreme pools. To validate these findings, we performed PCR-based screening on 87 litchi accessions. Two InDel markers demonstrated strong phenotypic associations: Chr5_25610680_InDel showed highly significant correlations with seed weight, edible rate, seed length, seed width, and seed-to-fruit weight ratio, explaining 22.60–35.54% of phenotypic variation. Meanwhile, Chr5_25585686_InDel was significantly associated with seed weight and edible rate, accounting for 18.66% and 18.94% of the phenotypic variation, respectively. These findings provide valuable molecular markers for marker-assisted breeding of litchi seed size, offering a promising avenue to advance precision breeding in this economically important crop. Full article

Rhizoctonia solani root rot (RSRR) is a major disease that significantly reduces soybean yields, causing substantial economic losses to global soybean production. To elucidate the genetic basis of RSRR resistance, 310 soybean germplasm accessions were evaluated using the disease severity index (DSI) following inoculation with R. solani. Among these accessions, 46.13% were susceptible, and only 2.26% exhibited high resistance. Utilizing resequencing data consisting of 738,561 Single Nucleotide Polymorphism (SNP) loci, a genome-wide association study (GWAS) was performed by integrating both general linear model (GLM) and mixed linear model (MLM) approaches, resulting in the identification of 21 SNPs significantly associated with resistance on chromosomes 3, 13, 15, 16, 17, and 18, and six candidate genes. RT-qPCR expression analysis revealed that four genes, including Glyma.03G166300, Glyma.03G168100, Glyma.13G212700, and Glyma.13G212300, were significantly upregulated in resistant genotypes after inoculation. Furthermore, Cleaved Amplified Polymorphic Sequences (CAPS) and Kompetitive Allele Specific PCR (KASP) molecular markers were successfully developed based on the RSRR-associated SNPs S3_38086892, S3_38247290, and S13_32595026, providing effective tools for marker-assisted selection (MAS). The findings strengthen our genetic knowledge concerning RSRR resistance and contribute to the molecular breeding of resistant soybean cultivars. Full article

Increasing the litter size in sheep is a primary objective in breeding programs, driving sustained interest in identifying candidate functional genes and molecular markers associated with fecundity. The known FecD mutation in the LEPR gene has been shown to regulate reproductive traits by influencing the ovulation rate of Davisdale sheep. However, the relationship between single-nucleotide polymorphisms (SNPs) in the LEPR and litter size remains unknown in Gobi short tail sheep and Sonid sheep. In this research, we found one novel variant and thirteen known variants through direct sequencing in Sonid sheep and Gobi short tail sheep and performed an association study in a large-scale population. The association analysis identified two SNPs (c.240C>T (rs159694506) and c.279C>T (rs159694508)) with significant litter size associations in Gobi short tail sheep (p < 0.01 ). In Sonid sheep, the c.240C>T (rs159694506), c.279C>T (rs159694508), g.41249772C>T (rs412130067), g.41249873A>C (rs425490800), g.41250357T>C (rs424307284), and g.41250358T>C (rs404651806) SNPs were significantly associated with litter size (p < 0.05 ). In addition, in GB sheep, the frequency of the litter-size-associated C allele at the c.240C>T (rs159694506) and c.279C>T (rs159694508) variants were significantly lower than that in Sonid sheep (SN), Mongolia sheep (MG), Ujimqin sheep (UM), Tan sheep (Tan), Hu sheep (Hu), and Small-tailed Han sheep (STH) populations. In SN sheep, the frequency of the litter-size-associated C allele at the c.240C>T (rs159694506) and c.279C>T (rs159694508) variant was significantly lower than that in the DPU. These findings provided valuable molecular markers pertinent to the fecundity of sheep, offering scientific evidence for the genetic improvement of these breeds. Full article

A comprehensive comparative analysis was conducted on the nucleotide and amino acid sequences of intact phiLM21-like prophages (phiLM21-LPhs), which currently represent the most prevalent prophages in Sinorhizobium meliloti—a symbiotic partner of Fabaceae plants. Remarkably, the nucleotide sequences of 25 phiLM21-LPhs, identified across 36 geographically dispersed S. meliloti strains, covered no more than 34% of the phiLM21 phage genome. All prophages were integrated into specific isoacceptor tRNA genes and carried a tyrosine-type integrase gene; however, this integration did not exhibit features of tRNA-dependent lysogeny. Only one-fifth of phiLM21-LPhs encoded the minimal set of regulators for lysogenic/lytic cycle transitions, while the remainder contained either uncharacterized regulatory elements or appeared to be undergoing genomic “anchoring” within the host bacterium. The phiLM21-LPhs harbored open reading frames (ORFs) of diverse origins (phage-derived, bacterial, and unknown), yet over half of these ORFs had undeterminable functions, representing genetic “dark matter”. The observed diversification of intact phiLM21-like prophages likely stems from recombination events involving both virulent/temperate phages and phylogenetically remote bacterial taxa. The evolutionary and biological significance of the substantial genetic “dark matter” within these prophages in soil saprophytic bacteria remains an unresolved question. Full article

Background/Objectives: Malassezia pachydermatis is a lipophilic yeast frequently associated with otitis externa and dermatological disorders in companion animals. This study aimed to evaluate the antifungal susceptibility of M. pachydermatis isolates from dogs and cats and to investigate the genomic determinants of reduced susceptibility. Methods: Susceptibility testing of 87 clinical isolates was performed using a modified CLSI broth microdilution method in Sabouraud dextrose broth supplemented with 1% Tween 80. The whole genome of ten representative isolates was sequenced and the genetic factors that are involved in drug resistance were investigated. Results: Ketoconazole, itraconazole, and terbinafine exhibited the highest efficacy, while miconazole and clotrimazole showed reduced activity. Whole genome sequencing revealed single nucleotide polymorphisms (SNPs) in genes that play a key role in the ergosterol biosynthesis pathway, particularly in ERG11 and ERG1. While some specific amino acid substitutions (e.g., K446R in ERG11) were found only in isolates with elevated MIC values, no direct correlation with resistance could be unequivocally established. Conclusions: Genomic analyses also uncovered chromosomal mutations and the heterozygosity of certain isolates, suggesting that complex, multifactorial mechanisms may drive the development of drug resistance. These findings highlight the importance of standardized susceptibility testing and further genomic investigations to promote effective antifungal therapy in veterinary medicine. Full article

Common vetch (Vicia sativa L.) is a globally green manure and forage crop, cultivated extensively worldwide. Its seeds serve as an important concentrated feed. Due to the late release of the reference genome, few studies were conducted to analyze the genetic mechanisms of grain yield, which hindered the progress of common vetch breeding. Marker-assisted selection (MAS) is the best and most effective way to accelerate the genetic improvement of grain yield-related traits in common vetch. In this study, we performed a genome-wide association study (GWAS) using the high-density single nucleotide polymorphism (SNP) data obtained through re-sequencing to better understand the genetic basis of grain yield-related traits. In total, six grain yield-related traits were evaluated in 172 accessions mainly sourced from China and Russia, across four environments, including branches per plant (NB), pod length (PL), number of pods per plant (NP), number of grains per pod (NG), hundred-grain weight (HGW), and grain yield (GY). Population structure analysis of the 172 accessions revealed four distinct subpopulations, exhibiting strong geographical correlation. In total, 38 loci have been identified as significantly associated with six grain yield-related traits, accounting for 13.3–31.7% of the phenotypic variances. Among them, qGY1.1 and qNG1.1, qNG2.2 and qPL2.1, qNG3.2 and qGY3.2, qNG4.1 and qPL4.1, qGY4.1 and qHGW4.1, qNG6.1 and qPL6.1, and qNB6.2 and qGY6.2 exhibit overlapping regions, suggesting that these regions are pleiotropic and should be prioritized for further research and breeding. In total, 12 candidate genes encoding auxin response factor, F-box repeat protein, gibberellin receptor, serine/threonine-protein kinase-like protein, and cellulose synthase-like protein were identified. Furthermore, we successfully developed and verified a kompetitive allele-specific PCR (KASP) marker (Kasp-NB6.2) for the number of branches. These findings provide molecular insights into grain yield-related traits in common vetch and offer valuable loci and molecular tools for MAS breeding. Full article