Search Results (10)

The recently described genus Candolleomyces (Basidiomycota, Agaricales, Psathyrellaceae) is now recognized as a distinct taxonomic group separate from Psathyrella. Currently, no fully assembled and accurately annotated genomes of Candolleomyces species are available, limiting our understanding of their physiological traits and biotechnological potential. Numerous tools exist for fungal genome assembly and annotation, each using different algorithms, resulting in substantial variation in gene content and distribution within the same genome. In this work, a hybrid assembly and annotation of the genome of strain CMU-8613 were performed using pipelines that combine different assembly and annotation tools. Phylogenetic analysis showed that the analyzed strain CMU-8613 belongs to Candolleomyces candolleanus. The assembled genome size ranged from 46.8 Mb (NECAT + Racon) to 59.3 Mb (Canu + Coprinellus micaceus genome assembly), depending on the assembly and polishing strategy. The analysis identified 15–25 secondary metabolite gene clusters (BGCs), depending on the genome assembly and the tools used for BGC prediction. In strain CMU-8613, CAZyme-encoding genes varied across assemblies: 494 genes were detected in the Flye assembly and 453 in NECAT; in both cases, the AA (Auxiliary Activities) and GH (Glycoside Hydrolases) families were the most represented. The diversity of CAZymes observed among Candolleomyces species suggests differences in their saprophytic capacities. Analysis of the MAT-A/MAT-B loci revealed that C. candolleanus possesses a tetrapolar mating system. This study provides the first annotated genome of C. candolleanus, highlighting its enzymatic potential to degrade plant biomass and its capacity to synthesize diverse secondary metabolites. The combination of assembly and annotation tools employed here offers robust alternative strategies for characterizing non-model fungi or species lacking high-quality reference genomes. Full article

Colletotrichum lini is a fungal pathogen of flax that can cause significant yield and quality losses. In this work, we obtained the first complete annotated genome assembly of the highly virulent C. lini strain #394-2. The nuclear genome consisted of ten core and two accessory chromosomes and had a length of 53.7 Mb. The mitochondrial genome was 39.1 kb. The assembly was obtained by the Canu–Racon ×2–Medaka–Polca algorithm using Oxford Nanopore Technologies and Illumina data. As a result of the annotation with the Illumina RNA-Seq data, 12,449 genes were identified. Potential signaling proteins were tested for effector functions and 550 effector proteins were predicted using EffectorP. The visualization of the effector protein localization revealed that the presence of effector proteins was associated with repeat-rich regions. A comparison of the genomic structure of C. lini with chromosome-level and complete assemblies of the genus Colletotrichum representatives revealed that the genomes of Colletotrichum species differed by the presence of chromosomal rearrangements. The obtained assembly expands the knowledge of the genomic structure of Colletotrichum species and provides the basis for further studies of C. lini, which will help to understand the virulence mechanisms and protect flax from anthracnose. Full article

Background: Eukaryotes’ whole-genome sequencing is crucial for species identification, gene detection, and protein annotation. Oxford Nanopore Technology (ONT) is an affordable and rapid platform for sequencing eukaryotes; however, the relatively higher error rates require computational and bioinformatic efforts to produce more accurate genome assemblies. Here, we evaluated the effect of read correction tools on eukaryote genome completeness, gene detection and protein annotation. Methods: Reads generated by ONT of four eukaryotes, C. albicans, C. gattii, S. cerevisiae, and P. falciparum, were assembled using minimap2 and underwent three rounds of read correction using flye, medaka and racon. The generates consensus FASTA files were compared for total length (bp), genome completeness, gene detection, and protein-annotation by QUAST, BUSCO, BRAKER1 and InterProScan, respectively. Results: Genome completeness was dependent on the assembly method rather than on the read correction tool; however, medaka performed better than flye and racon. Racon significantly performed better than flye and medaka in gene detection, while both racon and medaka significantly performed better than flye in protein-annotation. Conclusion: We show that three rounds of read correction significantly affect gene detection and protein annotation, which are dependent on assembly quality in preference to assembly completeness. Full article

Colletotrichum lini is a flax fungal pathogen. The genus comprises differently virulent strains, leading to significant yield losses. However, there were no attempts to investigate the molecular mechanisms of C. lini pathogenicity from high-quality genome assemblies until this study. In this work, we sequenced the genomes of three C. lini strains of high (#390-1), medium (#757), and low (#771) virulence. We obtained more than 100× genome coverage with Oxford Nanopore Technologies reads (N50 = 12.1, 6.1, 5.0 kb) and more than 50× genome coverage with Illumina data (150 + 150 bp). Several assembly strategies were tested. The final assemblies were obtained using the Canu–Racon ×2–Medaka–Polca scheme. The assembled genomes had a size of 54.0–55.3 Mb, 26–32 contigs, N50 values > 5 Mb, and BUSCO completeness > 96%. A comparative genomic analysis showed high similarity among mitochondrial and nuclear genomes. However, a rearrangement event and the loss of a 0.7 Mb contig were revealed. After genome annotation with Funannotate, secreting proteins were selected using SignalP, and candidate effectors were predicted among them using EffectorP. The analysis of the InterPro annotations of predicted effectors revealed unique protein categories in each strain. The assembled genomes and the conducted comparative analysis extend the knowledge of the genetic diversity of C. lini and form the basis for establishing the molecular mechanisms of its pathogenicity. Full article

Exponential advances in computational power have fueled advances in many disciplines, and biology is no exception. High-Performance Computing (HPC) is gaining traction as one of the essential tools in scientific research. Further advances to exascale capabilities will necessitate more energy-efficient hardware. In this article, we present our efforts to improve the efficiency of genome assembly on ARM-based HPC systems. We use vectorization to optimize the popular genome assembly pipeline of minimap2, miniasm, and Racon. We compare different implementations using the Scalable Vector Extension (SVE) instruction set architecture and evaluate their performance in different aspects. Additionally, we compare the performance of autovectorization to hand-tuned code with intrinsics. Lastly, we present the design of a CPU dispatcher included in the Racon consensus module that enables the automatic selection of the fastest instruction set supported by the utilized CPU. Our findings provide a promising direction for further optimization of genome assembly on ARM-based HPC systems. Full article

Antimicrobial resistance is increasing despite new treatments being employed, so novel strategies are required to ensure that bacterial infections remain treatable. Bacteriophages (phages; bacteria viruses) have the potential to be used as natural antimicrobial methods to control bacterial pathogens such as Salmonella spp. A Salmonella phage, Wara, was isolated from environmental water samples at the Subaé River Basin, Salvador de Bahia, Brazil. The basin has environmental impacts in its main watercourses arising from the dumping of domestic and industrial effluents and agricultural and anthropological activities. The phage genome sequence was determined by Oxford Nanopore Technologies (ONT) MinION and Illumina HiSeq sequencing, and assembly was carried out by Racon (MinION) and Unicycler (Illumina, Illumina + MinION). The genome was annotated and compared to other Salmonella phages using various bioinformatics approaches. MinION DNA sequencing combined with Racon assembly gave the best complete genome sequence. Phylogenetic analysis revealed that Wara is a member of the Tequintavirus genus. A lack of lysogeny genes, antimicrobial resistance, and virulence genes indicated that Wara has therapeutic and biocontrol potential against Salmonella species in healthcare and agriculture. Full article

High-quality genome sequences help to elucidate the genetic basis of numerous biological processes and track species evolution. For flax (Linum usitatissimum L.)—a multifunctional crop, high-quality assemblies from Oxford Nanopore Technologies (ONT) data were unavailable, largely due to the difficulty of isolating pure high-molecular-weight DNA. This article proposes a scheme for gaining a contiguous L. usitatissimum assembly using Nanopore data. We developed a protocol for flax nuclei isolation with subsequent DNA extraction, which allows obtaining about 5 μg of pure high-molecular-weight DNA from 0.5 g of leaves. Such an amount of material can be collected even from a single plant and yields more than 30 Gb of ONT data in two MinION runs. We performed a comparative analysis of different genome assemblers and polishers on the gained data and obtained the final 447.1-Mb assembly of L. usitatissimum line 3896 genome using the Canu—Racon (two iterations)—Medaka combination. The genome comprised 1695 contigs and had an N50 of 6.2 Mb and a completeness of 93.8% of BUSCOs from eudicots_odb10. Our study highlights the impact of the chosen genome construction strategy on the resulting assembly parameters and its eligibility for future genomic studies. Full article

While plant genome analysis is gaining speed worldwide, few plant genomes have been sequenced and analyzed on the African continent. Yet, this information holds the potential to transform diverse industries as it unlocks medicinally and industrially relevant biosynthesis pathways for bioprospecting. Considering that South Africa is home to the highly diverse Cape Floristic Region, local establishment of methods for plant genome analysis is essential. Long-read sequencing is becoming standard procedure for plant genome research, as these reads can span repetitive regions of the DNA, substantially facilitating reassembly of a contiguous genome. With the MinION, Oxford Nanopore offers a cost-efficient sequencing method to generate long reads; however, DNA purification protocols must be adapted for each plant species to generate ultra-pure DNA, essential for these analyses. Here, we describe a cost-effective procedure for the extraction and purification of plant DNA and evaluate diverse genome assembly approaches for the reconstruction of the genome of rooibos (Aspalathus linearis), an endemic South African medicinal plant widely used for tea production. We discuss the pros and cons of nine tested assembly programs, specifically Redbean and NextDenovo, which generated the most contiguous assemblies, and Flye, which produced an assembly closest to the predicted genome size. Full article

Oxford Nanopore sequencing can be used to achieve complete bacterial genomes. However, the error rates of Oxford Nanopore long reads are greater compared to Illumina short reads. Long-read assemblers using a variety of assembly algorithms have been developed to overcome this deficiency, which have not been benchmarked for genomic analyses of bacterial pathogens using Oxford Nanopore long reads. In this study, long-read assemblers, namely Canu, Flye, Miniasm/Racon, Raven, Redbean, and Shasta, were thus benchmarked using Oxford Nanopore long reads of bacterial pathogens. Ten species were tested for mediocre- and low-quality simulated reads, and 10 species were tested for real reads. Raven was the most robust assembler, obtaining complete and accurate genomes. All Miniasm/Racon and Raven assemblies of mediocre-quality reads provided accurate antimicrobial resistance (AMR) profiles, while the Raven assembly of Klebsiella variicola with low-quality reads was the only assembly with an accurate AMR profile among all assemblers and species. All assemblers functioned well for predicting virulence genes using mediocre-quality and real reads, whereas only the Raven assemblies of low-quality reads had accurate numbers of virulence genes. Regarding multilocus sequence typing (MLST), Miniasm/Racon was the most effective assembler for mediocre-quality reads, while only the Raven assemblies of Escherichia coli O157:H7 and K. variicola with low-quality reads showed positive MLST results. Miniasm/Racon and Raven were the best performers for MLST using real reads. The Miniasm/Racon and Raven assemblies showed accurate phylogenetic inference. For the pan-genome analyses, Raven was the strongest assembler for simulated reads, whereas Miniasm/Racon and Raven performed the best for real reads. Overall, the most robust and accurate assembler was Raven, closely followed by Miniasm/Racon. Full article

The whole genome sequencing (WGS) has become a crucial tool in understanding genome structure and genetic variation. The MinION sequencing of Oxford Nanopore Technologies (ONT) is an excellent approach for performing WGS and it has advantages in comparison with other Next-Generation Sequencing (NGS): It is relatively inexpensive, portable, has simple library preparation, can be monitored in real-time, and has no theoretical limits on reading length. Sorghum bicolor (L.) Moench is diploid (2n = 2x = 20) with a genome size of about 730 Mb, and its genome sequence information is released in the Phytozome database. Therefore, sorghum can be used as a good reference. However, plant species have complex and large genomes when compared to animals or microorganisms. As a result, complete genome sequencing is difficult for plant species. MinION sequencing that produces long-reads can be an excellent tool for overcoming the weak assembly of short-reads generated from NGS by minimizing the generation of gaps or covering the repetitive sequence that appears on the plant genome. Here, we conducted the genome sequencing for S. bicolor cv. BTx623 while using the MinION platform and obtained 895,678 reads and 17.9 gigabytes (Gb) (ca. 25× coverage of reference) from long-read sequence data. A total of 6124 contigs (covering 45.9%) were generated from Canu, and a total of 2661 contigs (covering 50%) were generated from Minimap and Miniasm with a Racon through a de novo assembly using two different tools and mapped assembled contigs against the sorghum reference genome. Our results provide an optimal series of long-read sequencing analysis for plant species while using the MinION platform and a clue to determine the total sequencing scale for optimal coverage that is based on various genome sizes. Full article