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bioRxiv2022

The largest haplotype-phased and chromosome-scale genome assembly of the fungal pathogen, Puccinia polysora, by HiFi reads and Hi-C data

Rust fungi are characterized by large genomes with high repeat content and have two haploid nuclei in most life stages, which makes achieving high-quality genome assemblies challenging. Here, we described a pipeline using HiFi reads and Hi-C data to assemble a gigabase-sized fungal pathogen, Puccinia polysora f.sp. zeae, to haplotype-phased and chromosome-scale. The final assembled genome is 1.71 Gbp, with ~850 Mbp and 18 chromosomes in each haplotype, being currently one of the two giga-scale fungi assembled to chromosome level. Transcript-based annotation identified 47,512 genes for dikaryotic genome with a similar number for each haplotype. A high level of interhaplotype variation was found with 10% haplotype-specific BUSCO genes, 5.8 SNPs/kbp and structural variation accounting for 3% of the genome size. The P. polysora genome displayed over 85% repeat contents, with genome-size expansion and copy number increasing of species-specific orthogroups. Interestingly, these features did not affect overall synteny with other Puccinia species having smaller genomes. Fine-time-point transcriptomics revealed seven clusters of co-expressed secreted proteins that are conserved between two haplotypes. The fact that candidate effectors interspersed with all genes indicated the absence of a "two-speed genome" evolution in P. polysora. Genome resequencing of 79 additional isolates revealed a clonal population structure of P. polysora in China with low geographic differentiation. Nevertheless, a minor population differentiated from the major population by having mutations on secreted proteins including AvrRppC, indicating the ongoing virulence to evade recognition by RppC, a major resistance gene in Chinese corn cultivars. The high-quality assembly provides valuable genomic resources for future studies on disease management and the evolution of P. polysora.

genomics↗

bioRxiv2022

Genomic erosion in the assessment of species extinction risk and recovery potential

Many species are undergoing rapid population declines and environmental deterioration, leading to genomic erosion, the loss of genetic diversity, accumulation of deleterious mutations, maladaptation, and introgression, that undermines fitness and long-term viability. Critically, this process continues even after demographic recovery due to a time-lagged genetic burden known as drift debt. Current conservation assessments, such as the IUCN Red List, focus on short-term extinction risk and do not capture the long-term consequences of genomic erosion. Likewise, the longer-term assessments of the IUCN Green Status may overestimate population recovery by failing to account for the lasting effects of genomic erosion. As genome sequencing becomes increasingly accessible, there is a growing opportunity to quantify genomic erosion and integrate it into conservation planning. Here, we use genomic simulations to illustrate how how different genomic metrics are sensitive to the drift debt. We test how ancestral effective population size (Ne) and bottleneck history influence the tempo and severity of genomic erosion, and demonstrate how these dynamics shape genetic load and additive genetic variation, key indicators of long-term evolutionary potential. Finally, we present a proof-of-concept for a Genomic Green Status framework that aligns genomic metrics with conservation impact assessments, laying the foundation for genomics-informed strategies to support species recovery.

genomics↗

bioRxiv2022

Genomic deletions and rearrangements in monkeypox virus from the 2022 outbreak, USA

Genomic surveillance of monkeypox virus (MPXV) during the 2022 outbreak has been mainly focused on single nucleotide polymorphism (SNP) changes. DNA viruses, including MPXV, have a lower SNP mutation rate than RNA viruses due to higher fidelity replication machinery. We identified a large genomic rearrangement in a MPXV sequence from a 2022 case in the state of Minnesota (MN), USA, from an abnormal, uneven MPXV read mapping coverage profile in whole-genome sequencing (WGS) data. We further screened WGS data of 206 U.S. MPXV samples and found seven (3.4 percent) sequenced genomes contained similar abnormal read coverage profiles that suggested putative large deletions or genomic rearrangements. Here, we present three MPXV genomes containing deletions ranging from 2.3 to 15 kb and four genomes containing more complex rearrangements. Five genomic changes were each only seen in one sample, but two sequences from linked cases shared an identical 2.3 kb deletion in the 3 terminal region. All samples were positive using VAC1 and Clade II (formerly West African)-specific MPXV diagnostic tests; however, large deletions and genomic rearrangements like the ones reported here have the potential to result in viruses in which the target of a PCR diagnostic test is deleted. The emergence of genomic rearrangements during the outbreak may have public health implications and highlight the importance of continued genomic surveillance.

genomics↗

bioRxiv2022

Haploid-resolved and chromosome-scale genome assembly in hexa-autoploid sweetpotato (Ipomoea batatas (L.) Lam)

Sweetpotato (Ipomoea batatas (L.) Lam) is the worlds seventh most important food crop by production quantity. Cultivated sweetpotato is a hexaploid (2n = 6x = 90), and its genome (B1B1B2B2B2B2) is quite complex due to polyploidy, self-incompatibility, and high heterozygosity. Here we established a haploid-resolved and chromosome-scale de novo assembly of autohexaploid sweetpotato genome sequences. Before constructing the genome, we created chromosome-scale genome sequences in I. trifida using a highly homozygous accession, Mx23Hm, with PacBio RSII and Hi-C reads. Haploid-resolved genome assembly was performed for a sweetpotato cultivar, Xushu18 by hybrid assembly with Illumina paired-end (PE) and mate-pair (MP) reads, 10X genomics reads, and PacBio RSII reads. Then, 90 chromosome-scale pseudomolecules were generated by aligning the scaffolds onto a sweetpotato linkage map. De novo assemblies were also performed for chloroplast and mitochondrial genomes in I. trifida and sweetpotato. In total, 34,386 and 175,633 genes were identified on the assembled nucleic genomes of I. trifida and sweetpotato, respectively. Functional gene annotation and RNA-Seq analysis revealed locations of starch, anthocyanin, and carotenoid pathway genes on the sweetpotato genome. This is the first report of chromosome-scale de novo assembly of the sweetpotato genome. The results are expected to contribute to genomic and genetic analyses of sweetpotato.

genomics↗

bioRxiv2023

De novo genome assemblies reveal structural variations between laboratory and natural isolates of C. elegans

Genomic structural variations (SVs) and transposable elements (TEs) can be significant contributors to genome evolution, altered gene expression, and risk of genetic diseases. Recent advancements in long-read sequencing have greatly improved the quality of de novo genome assemblies and enhanced the detection of sequence variants at the scale of hundreds or thousands of bases. Comparisons between two diverged wild isolates of Caenorhabditis elegans, the Bristol and Hawaiian strains, have been widely utilized in the analysis of small genetic variations. Genetic drift, including SVs and rearrangements of repeated sequences such as TEs, can occur over time from long-term maintenance of wild type isolates within the laboratory. To comprehensively detect both large and small structural variations as well as TEs due to genetic drift, we generated de novo genome assemblies and annotations for each strain from our lab collection using both long- and short-read sequencing and compared our assemblies and annotations with that of other lab wild type strains. Within our lab assemblies, we annotate over 3.1Mb of sequence divergence between the Bristol and Hawaiian isolates: 337,584 SNPs, 94,503 small insertion-deletions (<50bp), and 4,334 structural variations (>50bp). Further, we define the location and movement of specific DNA TEs between N2 Bristol and CB4856 Hawaiian wild type isolates. Specifically, we find the N2 Bristol genome has 20.6% more TEs from the Tc1/mariner family than the CB4856 Hawaiian genome. Moreover, we identified Zator elements as the most abundant and mobile TE family in the genome. Using specific TE sequences with unique SNPs, we also identify 38 TEs that moved intrachromosomally and 9 TEs that moved interchromosomally between the N2 Bristol and CB4856 Hawaiian genomes. By comparing the de novo genome assembly of our lab collection Bristol isolate to the VC2010 Bristol assembly, we also reveal that lab lineages display over 2 Mb of total variation: 1,162 SNPs, 1,528 indels, and 897 SVs with 95% of the variation due to SVs. Overall, our work demonstrates the unique contribution of SVs and TEs to variation and genetic drift between wild type laboratory strains assumed to be isogenic despite growing evidence of genetic drift and phenotypic variation. Author SummaryFor multiple model organisms, propagation of wild type strains in independent labs can lead to multiple phenotypic differences over time. To assess recombination, map mutations, and understand genomic changes during speciation, Caenorhabditis elegans researchers primarily use the wild type isolates Bristol and Hawaiian. Here, we map structural variations, transposable elements, and sequence divergence between the Bristol and Hawaiian natural isolates and between genomes of different lab lineages of these same strains.

genomics↗

bioRxiv2023

Range-wide differential adaptation and genomic vulnerability in critically endangered Asian rosewoods

In the billion-dollar global illegal wildlife trade, rosewoods have been the worlds most trafficked wild product since 20051. Dalbergia cochinchinensis and D. oliveri are the most sought-after rosewoods in the Greater Mekong Subregion2. They are exposed to significant genetic risks and the lack of knowledge on their adaptability limits the effectiveness of conservation efforts. Here we present genome assemblies and range-wide genomic scans of adaptive variation, together with predictions of genomic vulnerability to climate change. Adaptive genomic variation was differentially associated with temperature and precipitation-related variables between the species, although their natural ranges overlap. The findings are consistent with differences in pioneering ability and in drought tolerance3. We predict their genomic offsets will increase over time and with increasing carbon emission pathway but at a faster pace in D. cochinchinensis than in D. oliveri. These results and the distinct gene-environment association in the eastern coastal edge suggest species-specific conservation actions: germplasm representation across the range in D. cochinchinensis and focused on vulnerability hotspots in D. oliveri. We translated our genomic models into a seed source matching application, seedeR, to rapidly inform restoration efforts. Our ecological genomic research uncovering contrasting selection forces acting in sympatric rosewoods is of relevance to conserving tropical trees globally and combating risks from climate change. Significant statementIn the billion-dollar global illegal wildlife trade, rosewoods have been the worlds most trafficked wild product since 2005, with Dalbergia cochinchinensis and D. oliveri being the most sought-after and endangered species in Southeast Asia. Emerging efforts for their restoration have lacked a suitable evidence base on adaptability and adaptive potential. We integrated range-wide genomic data and climate models to detect the differential adaptation between D. cochinchinensis and D. oliveri in relevance to temperature- and precipitation-related variables and projected their vulnerability until 2100. We highlighted the stronger local adaptation in the coastal edge of the species ranges suggesting conservation priority. We developed genomic resources including chromosome-level genome assemblies and a web-based application seedeR for genomic model-enabled assisted migration and restoration.

genomics↗

bioRxiv2023

A comparative genomics approach reveals a local genetic signature of Leishmania tropica in Morocco

In Morocco, cutaneous leishmaniasis (CL) caused by Leishmania (L.) tropica is an important health problem. Despite its high incidence in the country, the genomic heterogeneity of these parasites is still incompletely understood. In this study, we sequenced the genomes of 14 Moroccan isolates of L. tropica collected from confirmed cases of CL to investigate their genomic heterogeneity. Comparative genomics analyses were conducted by applying the recently established Genome Instability Pipeline (GIP), which allowed us to conduct phylogenomic and PCA analyses, and to assess genomic variations at the levels of the karyotype, gene copy number, and single nucleotide polymorphisms (SNPs). The results identified a core group of 12 isolates that were genetically highly related but evolutionarily distant to the reference genome as judged by the presence of over 100,000 SNPs, 75% of which were shared inside this core group. In addition, we identified two highly divergent strains, M3015 and Ltr_16, that were phylogenetically distinct between each other as well as to the core group and the reference genome. Read-depth analysis revealed important karyotypic variations across all isolates and uncovered important differences in gene copy number between the isolates of the core group and the L. tropica reference genome, as well as between the core group and M3015. In conclusion, our NGS results suggest the presence of a local SNP signature that distinguishes Moroccan L. tropica from other endemic regions and from the reference genome. These results pave the way for future research with a larger number of strains that will allow to correlate diverse phenotypes (resistance to treatments, virulence) and origins (geography, host species, year of isolation) to defined genomic signals that may represent interesting biomarker candidates.

genomics↗

bioRxiv2023

Spatial and temporal characterization of the rich fraction of plastid DNA present in the nuclear genome of Moringa oleifera reveals unanticipated complexity in NUPTs' formation

Beyond the massive amounts of DNA and genes transferred from the protoorganelle genome to the nucleus during the endosymbiotic event that gave rise to the plastids, stretches of plastid DNA of varying size are still being copied and relocated to the nuclear genome in a process that is ongoing and does not result in the concomitant shrinking of the plastid genome. As a result, plant nuclear genomes are featured by a small, but variable, fraction of their genomes of plastid origin, the so-called nuclear plastid DNA sequences (NUPTs). However, research on the topic was mostly focused on a limited number of species and of plastid DNA. Here, we leveraged a chromosome-scale version of the Moringa oleifera genome, featured by the largest fraction of plastid DNA in any plant nuclear genome, to examine the chromosomal distribution and arrangement of NUPTs, to explicitly model and test the correlation between their age and size distribution, as well as to characterize their sites of origin at the chloroplast genome and their sites of insertion at the nuclear one. We found a bimodal distribution of NUPT relative ages, which implies NUPTs in moringa were formed through two separate events. Furthermore, NUPTs from every event showed markedly distinctive features, suggesting they originated through distinct mechanisms. Our results reveal an unanticipated complexity of the mechanisms at the origin of NUPTs and of the evolutionary forces behind their fixation and highlight moringa species as an exceptional model to study the mechanisms of origin and evolutionary fate of NUPTs. Significance statementSequencing of plant genomes has revealed the presence of a variable, but significant, content of DNA arising from their organelles, i.e., plastids and mitochondria. However, the mechanisms underlying their origin and fixation are not yet fully elucidated. Here, by thoroughly examining the spatial and temporal distribution of the large fraction of plastid DNA present in the nuclear genome of the orphan crop Moringa oleifera, we identified their episodic origin trough seemingly distinct mechanisms, highlighting the complexity of the molecular and evolutionary dynamics underlying their origin and fixation and revealing moringa as an outstanding model to assess the impact of plastid DNA in the evolution of the architecture and function of plant nuclear genomes.

genomics↗

bioRxiv2023

Generating reference-quality de novo genome assemblies from parasitic nematodes using the Oxford Nanopore Technologies MinION sequencing platform

Parasitic nematode infections represent a significant burden of disease in impoverished populations. Genomic studies of parasitic nematodes have revealed novel drug and vaccine targets and provided unprecedented insights into parasite biology. A key component of these studies is the availability of high-quality reference genomes (i.e., genomes that are contiguous, complete, and accurate) that capture the biological diversity contained within species. However, relatively few genomic resources exist for parasitic nematodes and few species are represented by more than a single reference genome. Streamlined laboratory and computational workflows to generate high-quality reference genomes from individual specimens using a single data source have the potential to increase the availability of genomic data from parasitic nematodes. The Oxford Nanopore Technologies (ONT) MinION is an accessible sequencing platform capable of generating ultra-long read data ideal for assembling genomes. However, lower read-level accuracy of ONT data has previously required assemblies to be error-corrected with more accurate short-read data. In this study, we assessed the quality of de novo genome assemblies for three species of parasitic nematodes (Brugia malayi, Trichuris trichiura, and Ancylostoma caninum) generated using only ONT MinION data. Assemblies were benchmarked against current reference genomes and against additional assemblies that were supplemented with short-read Illumina data through polishing or hybrid assembly approaches. For each species, assemblies generated using only MinION data had similar or superior measures of contiguity, completeness, and gene content. In terms of gene composition, depending on the species, between 88.9-97.6% of complete coding sequences predicted in MinION data only assemblies were identical to those predicted in assemblies polished with Illumina data. Polishing MinION data only assemblies with Illumina data therefore improved gene-level accuracy to a degree. Furthermore, modified DNA extraction and library preparation protocols produced sufficient genomic DNA from B. malayi and T. trichiura to generate de novo assemblies from individual specimens. Data Availability StatementQuality-controlled MinION and Illumina data for each species are deposited in the NCBI Sequence Read Archive (SRA) under the BioProject accession ID PRJNA1074771 under BioSample accession nos. SAMN39888962 (Brugia malayi), SAMN39888963 (Trichuris trichiura) and SAMN39888964 (Ancylostoma caninum). Final assemblies for each species are publicly available on GenBank.

genomics↗

bioRxiv2023

The history of chromosomal instability in genome doubled tumors

Tumors frequently display high chromosomal instability (CIN) and contain multiple copies of genomic regions. Here, we describe GRITIC, a generic method for timing genomic gains leading to complex copy number states, using single-sample bulk whole-genome sequencing data. By applying GRITIC to 5,656 tumors, we found that non-parsimonious evolution is frequent in the formation of complex copy number states in genome-duplicated tumors. We measured CIN before and after genome duplication in human tumors and found that late genome doubling was followed by an increase in the rate of copy number gain. Copy number gains often accumulate as punctuated bursts, commonly after genome duplication. We infer that genome duplications typically affect the selection landscape of copy number losses, while only minimally impacting copy number gains. In summary, GRITIC is a novel copy number gain timing framework that permits the analysis of copy number evolution in chromosomally unstable tumors. Statement of significanceComplex genomic gains are associated with whole-genome duplications, which are frequent across tumors, span a large fraction of their genomes, and are linked to poorer outcomes. GRITIC infers when these gains occur during tumor development, which will help to identify the genetic events that drive tumor evolution.

genomics↗

bioRxiv2023

Sequence-based machine learning reveals 3D genome differences between bonobos and chimpanzees

Phenotypic divergence between closely related species, including bonobos and chimpanzees (genus Pan), is largely driven by variation in gene regulation. The 3D structure of the genome mediates gene expression; however, genome folding differences in Pan are not well understood. Here, we apply machine learning to predict genome-wide 3D genome contact maps from DNA sequence for 56 bonobos and chimpanzees, encompassing all five extant lineages. We use a pairwise approach to estimate 3D divergence between individuals from the resulting contact maps in 4,420 1 Mb genomic windows. While most pairs were similar,[~] 17% were predicted to be substantially divergent in genome folding. The most dissimilar maps were largely driven by single individuals with rare variants that pro-duce unique 3D genome folding in a region. We also identified 89 genomic windows where bonobo and chimpanzee contact maps substantially diverged, including several windows harboring genes associated with traits implicated in Pan phenotypic divergence. We used in silico mutagenesis to identify 51 3D-modifying variants in these bonobo-chimpanzee diver-gent windows, finding that 34 or 66.67% induce genome folding changes via CTCF binding motif disruption. Our results reveal 3D genome variation at the population-level and identify genomic regions where changes in 3D folding may contribute to phenotypic differences in our closest living relatives.

genomics↗

bioRxiv2023

Comprehensive Survey of Conserved RNA Secondary Structures in Full-Genome Alignment of Hepatitis C Virus

Hepatitis C virus (HCV) is a plus-stranded RNA virus that often chronically infects liver hepatocytes and causes liver cirrhosis and cancer. These viruses replicate their genomes employing error-prone replicases. Thereby, they routinely generate a large "cloud" of RNA genomes which - by trial and error - comprehensively explore the sequence space available for functional RNA genomes that maintain the ability for efficient replication and immune escape. In this context, it is important to identify which RNA secondary structures in the sequence space of the HCV genome are conserved, likely due to functional requirements. Here, we provide the first genome-wide multiple sequence alignment (MSA) with the prediction of RNA secondary structures throughout all representative full-length HCV genomes. We selected 57 representative genomes by clustering all complete HCV genomes from the BV-BRC database based on k-mer distributions and dimension reduction and adding RefSeq sequences. We include annotations of previously recognized features for easy comparison to other studies. Our results indicate that mainly the core coding region, the C-terminal NS5A region, and the NS5B region contain secondary structure elements that are conserved beyond coding sequence requirements, indicating functionality on the RNA level. In contrast, the genome regions in between contain less highly conserved structures. The results provide a complete description of all conserved RNA secondary structures and make clear that functionally important RNA secondary structures are present in certain HCV genome regions but are largely absent from other regions. Full-genome alignments of all branches of Hepacivirus C are provided in the supplement.

genomics↗

bioRxiv2023

Genome mining of Streptomyces for the discovery of low-resistance antibiotics

2.Antimicrobial resistance is considered one of the top ten global health crises, which requires discovering and developing antibiotics with low resistance. Historically, Streptomyces bacteria are well-known for adapting to their complex environment by accumulating numerous clusters of specialized metabolites, some of which remain inactive in laboratory settings. The genomic revolution significantly increased their potential, complementing laboratory practices, and allowing for the discovery of antimicrobials associated with this biosynthetic machinery that are not constitutively produced. In the current study, we used the latest bioinformatics analyses to improve upon these predictions for Streptomyces genomes, and to identify low-resistance novel antimicrobial candidates. Our integrated pipeline used antiSMASH, BiG- SCAPE/EFI-EST, BiG-FAM, and additional tools and identified 326 novel BGCs and 67 peptides with antimicrobial potential. Further exploration of ribosomally synthesized and post-translationally modified peptides (RiPPs) revealed diverse chemical structures and suggested new mechanisms of action. Artificial intelligence platforms, such as MACREL, predicted the antimicrobial activity of the identified peptides, offering a comprehensive strategy for discovering bioactive compounds. Their low-resistance potential was estimated on a case-by-case basis. This study showcases the extensive genomic potential of Streptomyces, providing valuable insights for future antibiotic discovery efforts. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=118 SRC="FIGDIR/small/573565v1_ufig1.gif" ALT="Figure 1"> View larger version (33K): org.highwire.dtl.DTLVardef@1d8e6c6org.highwire.dtl.DTLVardef@116fa1eorg.highwire.dtl.DTLVardef@17edca5org.highwire.dtl.DTLVardef@1c7e6d7_HPS_FORMAT_FIGEXP M_FIG C_FIG 3. Impact statementExploring the potential applications of microorganisms is essential in fields like medicine, ecology, and biotechnology. Genomics is an invaluable tool that can uncover secondary metabolites with various applications in these fields. To this end, we conducted a detailed analysis of 388 complete genomes of Streptomyces, with a focus on identifying new biosynthetic gene clusters (BGCs) and antimicrobial peptides. Our analysis involved the use of various tools, including AntiSMASH, BiG-SCAPE, and reference databases like MiBIG and BiG-FAM, to unravel the diversity of these biosynthetic systems. We established a manually curated database and a robust pipeline for identifying valuable compounds, which allowed us to prioritize 326 BGCs with novel and diverse biosynthetic machinery. These unique insights into the genomic richness of Streptomyces will serve as a valuable guide for future antibiotic discovery efforts, facilitating the selection of strains that can produce new natural products. Moreover, our approach also helped us identify 67 potential antimicrobial peptides, including some belonging to the newly discovered Class V lanthipeptides, highlighting the diversity and promise of these compounds. Our work offers a glimpse into the potential of genomics in identifying important molecules and provides a framework for future studies in this field. Data summaryBig tables and special files can be found in Zenodo, under the following link In there, the following objects are included: O_LISupplementary Table 1: a spreadsheet with nine tables: (Table S1) Accession numbers and genomic characteristics of the genomes used in this study; (Table S2) summary of predicted Biosynthetic Gene Clusters (BGCs) for each genome; (Table S3) summary of the distribution of BGC types present in Streptomyces genomes; (Table S4) IDs and strains of MiBIG BGCs; (Table S5) All information regarding the database of peptides with antimicrobial activity; (Table S6) Data of BGCs recovered after the analysis of MiBIG and BiG-FAM; (Table S7) Complete and potentially novel BGC data used for the creation of the SSN with EFI-EST; (Table S8) Data predicting potential antimicrobial activity using MACREL v1.2.0; (Table S9) Data of predicted peptides with positive antimicrobial activity. These data were used to create the graph showing the distribution of RiPPs classes within these peptides. C_LIO_LISupplementary Data 1: a folder containing all .gbk files for the BGCs predicted by antiSMASH and MiBIG. These files were used as input for BiG-SCAPE (https://drive.google.com/file/d/1VGWJo9rsF429IUFf2g-tqs_NfBicNX91/view?usp=drive_link). C_LIO_LISupplementary Data 2: amino acid sequences of the mined and reference peptides (from the database built into this work) in .fasta format. C_LIO_LISupplementary Table 2: a spreadsheet with five tables: Data from the evaluation of the prediction of antimicrobial activity with five different AIs through five comparisons of the created database (Table S1-Macrel); (Table S2-CAMPR3); (Table S3-AMP Scanner); (Table S4-AmPEP); (Table S5-AL4AMP). C_LIO_LISupplementary Figure 1: results of the predictive performance analysis (script: https://github.com/Bert-IQ/ripps-classification). C_LI

genomics↗

bioRxiv2023

Whole genome sequencing of newly emerged fungal pathogen Aspergillus lentulus and its Azole resistance gene prediction

Aspergillus lentulus is an important newly recorded species in the A. fumigatus complex and its resistance to azole drugs and the high mortality rate of infected individuals have emerged as problems. Comprehensive understanding of the A. lentulus is limited due to lack of genome-wide fine mapping data. The aim of this study was to investigate the A. lentulus signature at the molecular level, analyze the genome-wide profile of this strain and predict its possible genes that execute azole resistance. In this study, a whole genome sequencing of a clinically isolated A. lentulus strain (named A. lentulus PWCAL1) was studied by PacBio Sequel sequencing platform. Azole resistance genes were predicted based on whole-genome sequencing data analysis, gene function annotation, comparative genomics analysis, and blastp alignment using the Mycology Antifungal Resistance Database (MARDy) to comprehensively understanding the genome-wide features, pathogenicity, and resistance mechanisms of A. lentulus. The results of whole genome sequencing demonstrated that the total length of A. lentulus PWCAL1 genome was 31255105 bp, the GC content was 49.24%, and 6883 coding genes were predicted. A total of 4565, 1824 and 6405 genes were annotated in GO, KOG and KEGG databases, respectively. In the PHI and DFVF databases, 949 and 259 interacting virulence factors were identified, respectively, with the main virulence factor-Mutant virulence phenotype was enriched in reduced virulence. Comparative genomics analysis showed that there were 5456 consensus Core Genes in this strain and four closely related strains of A. fumigatus complex, which were mainly involved in Human Diseases, Metabolism, Organismal Systems, etc. Among the three aligned A. lentulus strains, the number of unique genes of this bacterium was the highest with number of 171, and these genes were mainly associated with Carbohydrate metabolism, Cell growth and death. Resistance gene prediction demonstrated that the A5653 gene of this bacterium had F46Y/N248T double point mutations on the CYP51A gene, but no tandem repeat TR mutations in the promoter region were detected. Further more, twelve genes belonging to the fungal multidrug resistance ABC transporters were identified based on the complete genome sequence and phylogenetic analysis of A. lentulus, which belonged to the ALDp subfamily, the PDR subfamily (AtrB, CDR1, and CDR2), and the MDR subfamily (MDR1), respectively, and there were four genes that are annotated to the MFS multidrug transporter. Further phylogenetic tree classification of the ABC transporter subfamilies predicted in the nine selected A. fumigatus complex strains showed that these putative ABC proteins were divided into two main clusters, which belonged to PDR (CDR1, CDR2, AtrB, and AtrF) and MDR subfamilies (MDR1, MDR2, and MDR3). The distribution of these ABC proteins varies among different species of the A. fumigatus complex. The main result obtained from this study for the whole genome of A. lentulus provide new insights into better understanding the biological characteristics, pathogenicity and resistance mechanisms of this bacterium. In this study, two resistance mechanisms which include CYP51A gene mutation and multidrug-resistant ABC transporter were predicted in a single isolate. Based on the predicted CYP51A-F46Y/N248T site mutation combination, we speculate that the CYP51A gene of A. lentulus may be partially responsible for azole resistance. Based on the predicted ABC transporter family genes, we hypothesize that resistance to multiple azoles in A. lentulus is mediated, at least in part, by these ABC transporters with resistance.

genomics↗

bioRxiv2024

Genome-AC-GAN: Enhancing Synthetic Genotype Generationthrough Auxiliary Classification

In recent years, there have been increasing attempts to develop computational methods for generating synthetic genomic data that aim to mimic real genomic datasets. Artificial genomes (AGs) generated by these methods have emerged as a promising potential solution for privacy concerns raised by public genomic datasets and as means to provide adequate representation of under-sampled populations. However, existing methods for generating AGs provide a very limited capability for faithfully capturing features of different sub-populations within a larger cohort. In this study, we propose a novel method called the Genome Auxiliary Classifier Generative Adversarial Network (Genome-AC-GAN), which generates AGs tailored to specific sub-populations. We conducted experiments to evaluate the performance of the Genome-AC-GAN and compare the AGs it generates with real genomic data as well as with AGs generated by previously published methods. The Genome-AC-GAN outperforms other methods and faithfully models population structure, which is not adequately captured by existing methods. We also demonstrate the use of AGs generated by the Genome-AC-GAN in augmentation of datasets used as training sets for classifying genomes into populations. These experiments demonstrate the benefits of AGs in enhancing classification accuracy, especially when dealing with under-sampled and closely related populations.

genomics↗

bioRxiv2024

First whole genome assembly of Vaccinium floribundum Kunth, an emblematic Andean species

BackgroundVaccinium floribundum Kunth, known as "mortino," is an endemic shrub species of the Andean region adapted to harsh conditions in high-altitude ecosystems. It plays an important ecological role as a pioneer species in the aftermath of deforestation and human-induced fires within paramo ecosystems, emphasizing its conservation value. While previous studies have offered insights into the genetic diversity of mortino, comprehensive genomic studies are still missing to fully understand the unique adaptations of this species and its population status, highlighting the importance of generating a reference genome for this plant. ResultsONT and Illumina sequencing were used to establish a reference genome for this species. Three different de novo genome assemblies were generated and compared for quality, continuity and completeness. The Flye assembly was selected as the best and refined by filtering out short ONT reads, screening for contaminants and genome scaffolding. The final assembly has a genome size of 529 MB, containing 1,317 contigs and 97% complete BUSCOs, indicating a high level of integrity of the genome. Additionally, the LAI Index of 12.93, further categorizes this assembly as a reference genome. ConclusionsThe genome of V. floribundum reported in this study is the first reference genome generated for this species, providing a valuable tool for further studies. This high-quality genome, based on the quality and completeness parameters obtained, will not only help uncover the genetic mechanisms responsible for its unique traits and adaptations to high-altitude ecosystems, but will also contribute to conservation strategies for a species endemic to the Andes.

genomics↗

bioRxiv2024

Chromosome-scale Salvia hispanica L. (Chia) genome assembly reveals rampant Salvia interspecies introgression

Salvia hispanica L. (Chia), a member of the Lamiaceae, is an economically important crop in Mesoamerica, with health benefits associated with its seed fatty acid composition. Chia varieties are distinguished based on seed color including mixed white and black (Chia pinta) and black (Chia negra). To facilitate research on Chia and expand on comparative analyses within the Lamiaceae, we generated a chromosome-scale assembly of a Chia pinta accession and performed comparative genome analyses with a previously published Chia negra genome assembly. The Chia pinta and negra genome sequences were highly similar as shown by a limited number of single nucleotide polymorphisms and extensive shared orthologous gene membership. There is an enrichment of terpene synthases in the Chia pinta genome relative to the Chia negra genome. We sequenced and analyzed the genomes of 20 Chia accessions with differing seed color and geographic origin revealing population structure within S. hispanica and interspecific introgressions of Salvia species. As the genus Salvia is polyphyletic, its evolutionary history remains unclear. Using large-scale synteny analysis within the Lamiaceae and orthologous group membership, we resolved the phylogeny of Salvia species. This study and its collective resources further our understanding of genomic diversity in this food crop and the extent of inter-species hybridizations in Salvia. PLAIN LANGUAGE SUMMARYChia pinta is an economically important crop due to the high fatty acid present in the seeds. There are multiple types of Chia based on the seeds color including mixed which and black (Chia pinta), black (Chia negra), and white (Chia blanca). We generated a genome assembly of Chia pinta and compared it to existing genome assemblies. While the assemblies are highly similar there are key differences in terpene synthase composition between Chia pinta and Chia negra. We also sequenced 20 other Chia accessions with different seed color and geographic origin to determine a population structure within Chia. We generated genomic resources to further our understanding of this food crop. ABBREVIATIONSBGC Biosynthetic gene cluster BUSCO Benchmarking Universal Single Copy Orthologs GO Gene ontology SNP Single nucleotide polymorphism TIR Terminal inverted repeat TPS Terpene synthase WGS Whole genome shotgun

genomics↗

bioRxiv2024

Comparative genomics reveals diversity and taxonomic relationships among Clostridioides difficile phages

Clostridioides difficile is associated with life-threatening antibiotic-associated diarrhea, colitis, and toxin-mediated infections. While antibiotics are the primary treatment against C. difficile infections, increasing resistance necessitates alternatives. Bacteriophages and bacteriophage-derived proteins such as endolysins hold promise as potential solutions. Understanding phage biology at the genomic level is crucial for their therapeutic use. We conducted a comparative genomic analysis of 44 C. difficile phage genomes from public databases, examining both whole genome and proteome levels and grouping them by shared protein content. Relationships within each group were observed, and core and highly conserved genes were identified. Using genome and proteome phylogeny, average nucleotide identity, and core gene identification, we proposed an updated taxonomic classification. Nine distinct clusters were identified, without any singleton. Cluster members exhibited similar genome architecture, genome sizes, GC content, number of coding sequences, presence of core genes, and high nucleotide identity. Additionally, we propose 23 new genera, three families and the elevation of currently assigned genera to subfamilies. The lytic module proteins, endolysins, and holins were also characterized, revealing four distinct endolysin organizations with diverse domain architectures. Notably, the amidase_3 and LysM domains were highly conserved and subjected to purifying selection within the C. difficile phage genomes. This is the first comprehensive comparative study regarding C. difficile phage genomes. Our study provides valuable genomic insights that add to the current understanding of the phages. Our taxonomic analysis may improve the classification scheme of C. difficile phages and aid in the future classification of newly isolated C. difficile phage genomes.

genomics↗