Complete Reference Research Articles

Phenotypic and Complete Reference Whole Genome Sequence Analyses of Two Paenibacillus spp. Isolates from a Gray Wolf (Canis lupus) Gastrointestinal Tract

Inflammatory bowel disease (IBD) is increasing among mammals around the world, and domestic dogs are no exception. There is no approved cure for canine IBD with limited treatment options. Novel probiotic bacteria discovery from free-ranging animals for the treatment of IBD in domestic pets can likely yield promising probiotic candidates. Consequently, the overall aim was to isolate bacteria from free-ranging animals that could potentially be utilized as novel probiotics. Two bacteria identified as unique Paenibacillus spp. strains by small ribosomal RNA (16S) gene sequencing were isolated from the gastrointestinal tract of a North American Gray Wolf (Canis lupus). The bacteria were typed as Gram-variable, and both were catalase/oxidase positive as well as sensitive to commonly used antibiotics. The bacteria digested complex carbohydrates and lipids by standard assays. The isolated bacteria also inhibited the growth of Staphylococcus aureus and Micrococcus luteus. The whole genome sequence (WGS) length of bacterial isolate ClWae17B was 6,939,193 bp, while ClWae19 was 7,032,512 bp, both similar in size to other Paenibacillus spp. The genomes of both bacteria encoded enzymes involved with the metabolism of complex starches and lipids, such as lyases and pectinases, along with encoding antimicrobials such as lanthipeptides, lasso peptides, and cyclic-lactone-autoinducers. No pernicious virulence genes were identified in the WGS of either bacterial isolate. Phylogenetically, the most closely related bacteria based on 16S gene sequences and WGS were P. taichungensis for ClWae17B and P. amylolyticus for ClWae19. WGS analyses and phenotypic assays supported the hypothesis that the isolates described constitute two novel candidate probiotic bacteria for potential use in dogs.

Multiple introductions of NRCS-A Staphylococcus capitis to the neonatal intensive care unit drive neonatal bloodstream infections: a case-control and environmental genomic survey.

Background. The Staphylococcus capitis NRCS-A strain has emerged as a global cause of late-onset sepsis associated with outbreaks in neonatal intensive care units (NICUs) whose transmission is incompletely understood.Methods. Demographic and clinical data for 45 neonates with S. capitis and 90 with other coagulase-negative staphylococci (CoNS) isolated from sterile sites were reviewed, and clinical significance was determined. S. capitis isolated from 27 neonates at 2 hospitals between 2017 and 2022 underwent long-read (ONT) (n=27) and short-read (Illumina) sequencing (n=18). These sequences were compared with S. capitis sequenced from blood culture isolates from other adult and paediatric patients in the same hospitals (n=6), S. capitis isolated from surface swabs (found in 5/150 samples), rectal swabs (in 2/69 samples) in NICU patients and NICU environmental samples (in 5/114 samples). Reads from all samples were mapped to a hybrid assembly of a local sterile site strain, forming a complete UK NRCS-A reference genome, for outbreak analysis and comparison with 826 other S. capitis from the UK and Germany.Results. S. capitis bacteraemia was associated with increased length of NICU stay at sampling (median day 22 vs day 12 for other CoNS isolated; P=0.05). A phylogeny of sequenced S. capitis revealed a cluster comprised of 25/27 neonatal sterile site isolates and 3/5 superficial, 2/2 rectal and 1/5 environmental isolates. No isolates from other wards belonged to this cluster. Phylogenetic comparison with published sequences confirmed that the cluster was NRCS-A outbreak strain but found a relatively high genomic diversity (mean pairwise distance of 84.9 SNPs) and an estimated NRCS-A S. capitis molecular clock of 5.1 SNPs/genome/year (95% credibility interval 4.3-5.9). The presence of S. capitis in superficial cultures did not correlate with neonatal bacteraemia, but both neonates with rectal NRCS-A S. capitis carriage identified also experienced S. capitis bacteraemia.Conclusions. S. capitis bacteraemia occurred in patients with longer NICU admission than other CoNS. Genomic analysis confirms clinically significant infections with the NRCS-A S. capitis strain, distinct from non-NICU clinical samples. Multiple introductions of S. capitis, rather than prolonged environmental persistence, were seen over 5 years of infections.

VClean: assessing virus sequence contamination in viral genomes.

Recent advancements in viral metagenomics and single-virus genomics have improved our ability to obtain the draft genomes of environmental viruses. However, these methods can introduce virus sequence contaminations into viral genomes when short, fragmented partial sequences are present in the assembled contigs. These contaminations can lead to incorrect analyses;however, practical detection tools are lacking. In this study, we introduce vClean, a novel automated tool that detects contaminations in viral genomes. By applying machine learning to the nucleotide sequence features and gene patterns of the input viral genome, vClean could identify contaminations. Specifically, for tailed double-stranded DNA phages, we attempted accurate predictions by defining single-copy-like genes and counting their duplications. We evaluated the performance of vClean usingsimulated datasets derived from complete reference genomes, achieving a binary accuracy of 0.932. When vClean was applied to 4693 genomes of medium or higher quality derived from public ocean metagenomic data, 1604 genomes (34.2%) were identified as contaminated. We also demonstrated that vClean can detect contamination in single-virus genome data obtained from river water. vClean provides a new benchmark for quality control of environmental viral genomes and has the potential to become an essential tool for environmental viral genome analysis.

Annotated Checklist of Rhopalocera of Himachal Pradesh, India (Insecta: Lepidoptera)

The checklist includes all known Rhopalocera species ever recorded in the state of Himachal Pradesh. All previously published literature, research papers, and reference material were referred and a complete bibliographic reference of all the species mentioned in the checklist is incorporated. This checklist covers 437 species, belonging to 184 genera under 19 subfamilies and 6 families. Moreover, a list of doubtful or misidentified Rhopalocera species reported from Himachal Pradesh is also provided with relevant details in the discussion section. Comments on taxonomic insights and distribution are provided for some species and a list of expected Rhopalocera species is also provided in the discussion section.

Concepts and Methods of Tadabbur al-Qur'an for Early Childhood: A Review of the Book Tadabbur al-Qur’an for Kids

Islam places great importance on the education of children starting from an early age. If children lack education at the beginning of their lives, some of them will grow up with corrupt morals, like lying, envy, stealing, criticizing, and interfering in other people's affairs. The Qur'anic surah that alludes to the importance of the existence of children themselves is QS. al-Kahf verse 46, Allah says: “Treasures and children are ornaments in the life of the world”. This study aims to discuss the concept and method of tadabbur al-Qur'an through analyzing the work of the Ummul Qura' editorial team, namely the book Tadabbur Al-Qur’an for Kids. By using qualitative methodology, literature study, this research focuses on two main aspects: the understanding of the editorial team of Ummul Qura' about the concept and method of tadabbur al-Qur'an for early childhood, and how it is implemented in everyday life. The results showed that the editorial team of Ummul Qura' was able to provide solutions for early childhood education by presenting an applicable and meaningful understanding, including presenting convenience for children in tadabbur al-Qur'an, making it easy to memorize the Qur'an, presenting the translation of each verse, introducing the naming of the surah of the Qur'an, tadabbur and understanding the essence of the Qur'an's science, a guide to practicing the contents of the Qur'an, complete references, inviting children to dive further, and providing motivation to become experts in the Qur'an.

Neo-Sex Chromosome Evolution in Treehoppers Despite Long-Term X Chromosome Conservation.

Sex chromosomes follow distinct evolutionary trajectories compared to the rest of the genome. In many cases, sex chromosomes (X and Y or Z and W) significantly differentiate from one another resulting in heteromorphic sex chromosome systems. Such heteromorphic systems are thought to act as an evolutionary trap that prevents subsequent turnover of the sex chromosome system. For old, degenerated sex chromosome systems, chromosomal fusion with an autosome may be one way that sex chromosomes can "refresh" their sequence content. We investigated these dynamics using treehoppers (hemipteran insects of the family Membracidae), which ancestrally have XX/X0 sex chromosomes. We assembled the most complete reference assembly for treehoppers to date for Umbonia crassicornis and employed comparative genomic analyses of 12 additional treehopper species to analyze X chromosome variation across different evolutionary timescales. We find that the X chromosome is largely conserved, with one exception being an X-autosome fusion in Calloconophora caliginosa. We also compare the ancestral treehopper X with other X chromosomes in Auchenorrhyncha (the clade containing treehoppers, leafhoppers, spittlebugs, cicadas, and planthoppers), revealing X conservation across more than 300 million years. These findings shed light on chromosomal evolution dynamics in treehoppers and the role of chromosomal rearrangements in sex chromosome evolution.

Applying phylogenetic methods for species delimitation to distinguish B-cell clonal families.

The adaptive immune system generates a diverse array of B-cell receptors through the processes of V(D)J recombination and somatic hypermutation. B-cell receptors that bind to an antigen will undergo clonal expansion, creating a Darwinian evolutionary dynamic within individuals. A key step in studying these dynamics is to identify sequences derived from the same ancestral V(D)J recombination event (i.e. a clonal family). There are a number of widely used methods for accomplishing this task but a major limitation of all of them is that they rely, at least in part, on the ability to map sequences to a germline reference set. This requirement is particularly problematic in non-model systems where we often know little about the germline allelic diversity in the study population. Recognizing that delimiting B-cell clonal families is analogous to delimiting species from single locus data, we propose a novel strategy of reconstructing the phylogenetic tree of all B-cell sequences in a sample and using a popular species delimitation method, multi-rate Poisson Tree Processes (mPTP), to delimit clonal families. Using extensive simulations, we show that not only does this phylogenetically explicit approach perform well for the purpose of delimiting clonal families when no reference allele set is available, it performs similarly to state-of-the-art techniques developed specifically for B-cell data even when we have a complete reference allele set. Additionally, our analysis of an empirical dataset shows that mPTP performs similarly to leading methods in the field. These findings demonstrate the utility of using off-the-shelf phylogenetic techniques for analyzing B-cell clonal dynamics in non-model systems, and suggests that phylogenetic inference techniques may be potentially combined with mapping based approaches for even more robust inferences, even in model systems.

IMPLEMENTASI ZAKAT SEBAGAI PENGURANG PAJAK DI NANGROE ACEH DARUSSALAM, INDONESIA

This study aims to analyze the implementation of zakat as a deduction for income tax payable on tax revenue in Aceh in the context of Individual and Corporate Taxpayers, Comparing the application of zakat as a tax deduction in other countries and the Aceh government and identifying the role of Amil Zakat Institutions and the Directorate General of Taxes that are harmonious in managing zakat in Aceh. The research method uses a qualitative method with literature study, this research is conducted by analyzing various documents, books, journals, scientific articles, laws and regulations, relevant reports and other supporting sources by collecting various complete references. The results showed that (1) There are several problems between the application of tax deductible zakat in Aceh between Individual and Corporate Taxpayers (2) Comparison between countries and Aceh in the implementation of zakat as a deduction for payable income tax can be a pilot for the national scale and (3) There needs to be a seriousness of the government represented by the DGT and BAZNAS from the zakat organizer side to create an integrated system that makes it easier for taxpayers to pay zakat and updated real time in the tax return.

NAMERS: a purpose-built reference DNA sequence database to support applied eDNA metabarcoding

Applied eDNA metabarcoding is increasingly being considered as a tool to inform management decisions, regulations, or policy development. Because these downstream considerations are coming to the forefront of eDNA applications, optimizing workflow elements is essential to increasing standardization, efficiency, and competency of metabarcoding results. Reference DNA sequences are critical workflow elements that currently lack consistent approaches to generating, curating, or publishing. We present a complete mitochondrial genome and nuclear ribosomal DNA cistron reference DNA sequence library for 92% of the freshwater fish species of British Columbia, Canada. This resource is published as the Novel Applied eDNA Metabarcoding Reference Sequences (NAMERS) repository (https://namers.ca), a user-friendly and interactive website for specialists and non-specialists alike to explore and generate custom reference libraries for taxa and genes of interest. We demonstrate the power of NAMERS for optimization of applied eDNA metabarcoding study design by analyzing the number of primer mismatches and species resolution power of existing metabarcoding markers. NAMERS demonstrates that high quality curated genomic information is within a reasonable reach to meet the increasing demand for actionable eDNA metabarcoding applications. The framework used here incorporating the pillars of accuracy, completeness and accessibility can be applied for new iterations of other reference sequence databases to bring DNA-based monitoring into a new era.

Abstract A037: Improved liquid biopsy assay performance using novel highly accurate sequencing-by-binding (SBB) technology

Abstract Liquid biopsy is revolutionizing the field of early cancer detection research through non-invasive detection of tumor DNA in the blood. However, existing liquid biopsy assays are limited in their sensitivity for ctDNA detection at low variant allele frequencies (VAFs), with most relying on extreme sequencing depth and computational error correction to separate the true ctDNA signal from background errors. This limitation is particularly problematic in the area of early cancer detection and minimal residual disease monitoring, in which expected ctDNA allele frequencies are extremely low. Novel strategies are therefore needed to help improve liquid biopsy assay sensitivity and reduce per-sample sequencing requirements. Here we describe PacBio’s application of the Onso short-read sequencing system to enable detection of ctDNA at low VAFs using the SeraCare Complete ctDNA Mutation Mix reference standard. The Onso system makes use of a novel sequencing by binding (SBB) method to achieve 10- to 100-fold greater quality scores, with ≥90% of reads at Q40 or above. We performed targeted capture and sequencing of libraries prepared from the SeraCare reference mix diluted into WT human DNA at the following VAFs: 0.00% (WT), 0.05%, 0.10%, and 0.25%, and compared the sensitivity at each VAF for SBB compared to a competitor method using sequencing by synthesis (SBS) at varying sequencing depths. At equivalent sequencing depth (12,000X) we observed superior recall for low frequency variants (VAF = 0.05%, 0.1%) in the reference sample for Onso compared to SBS. Furthermore, SBB was able to achieve comparable sensitivity results to SBS using four- fold less sequencing (6,000X vs. 24,000X), and without the use of computational error correction. Combining SBB with error correction methods boosted sensitivity even further, suggesting an additive value for these technologies. Comparison of false positive variant calls in the WT samples showed a dramatic decrease in false positives for Onso (n=2) compared to SBS (n=49) across 10 subsampling replicates, and observed allele frequencies for Onso were closer to the expected values. In conclusion, Onso achieved superior sensitivity for ctDNA detection at compared to SBS at equivalent sequencing depths, and achieved similar performance with four- fold less sequencing and without the use of error correction. This improvement was due in part to the reduction in false positive variant calling in the WT sample, which lowers the limit of detection to maximize variant detection while maintaining high specificity. Taken together, our results demonstrate the potential of SBB to improve upon existing methods of liquid biopsy and better enable research on early cancer detection and minimal residual disease. Citation Format: Alexandra Sockell, Dan Nasko, Young Kim, Kristi Kim, Jonas Korlach. Improved liquid biopsy assay performance using novel highly accurate sequencing-by-binding (SBB) technology [abstract]. In: Proceedings of the AACR Special Conference: Liquid Biopsy: From Discovery to Clinical Implementation; 2024 Nov 13-16; San Diego, CA. Philadelphia (PA): AACR; Clin Cancer Res 2024;30(21_Suppl):Abstract nr A037.

Microsatellite density landscapes illustrate short tandem repeats aggregation in the complete reference human genome

BackgroundMicrosatellites are increasingly realized to have biological significance in human genome and health in past decades, the assembled complete reference sequence of human genome T2T-CHM13 brought great help for a comprehensive study of short tandem repeats in the human genome.ResultsMicrosatellites density landscapes of all 24 chromosomes were built here for the first complete reference sequence of human genome T2T-CHM13. These landscapes showed that short tandem repeats (STRs) are prone to aggregate characteristically to form a large number of STRs density peaks. We classified 8,823 High Microsatellites Density Peaks (HMDPs), 35,257 Middle Microsatellites Density Peaks (MMDPs) and 199, 649 Low Microsatellites Density Peaks (LMDPs) on the 24 chromosomes; and also classified the motif types of every microsatellites density peak. These STRs density aggregation peaks are mainly composing of a single motif, and AT is the most dominant motif, followed by AATGG and CCATT motifs. And 514 genomic regions were characterized by microsatellite density feature in the full T2T-CHM13 genome.ConclusionsThese landscape maps exhibited that microsatellites aggregate in many genomic positions to form a large number of microsatellite density peaks with composing of mainly single motif type in the complete reference genome, indicating that the local microsatellites density varies enormously along the every chromosome of T2T-CHM13.

Full-field three-dimensional system calibration for composite surfaces reconstruction

The accurate calibration of composite surface measurement systems for specular and diffused surface is of great importance in a number of industrial applications. This paper addresses the challenges in calibrating composite surface reconstruction by proposing a comprehensive full-field calibration method. The principal innovation is the construction of a comprehensive reference phase and the pixel-by-pixel calibration of system parameters within the same coordinate system. The details are as follows: (1) construct a complete reference phase for the composite surfaces through external parameter constraints phase fusion and interpolation; (2) calibrate the mapping relationship coefficients, compensate the accuracy degradation of the partial reflection. Experimental results validate the accuracy of the full-field calibration data, demonstrating the effectiveness and feasibility.

Three decades of rat genomics: approaching the finish(ed) line.

The rat, Rattus norvegicus, has provided an important model for investigation of a range of characteristics of biomedical importance. Here we survey the origins of this species, its introduction into laboratory research, and the emergence of genetic and genomic methods that utilize this model organism. Genomic studies have yielded important progress and provided new insight into several biologically important traits. However, some studies have been impeded by the lack of a complete and accurate reference genome for this species. New sequencing and genome assembly methods applied to the rat have resulted in a new reference genome assembly, GRCr8, which is a near telomere-to-telomere assembly of high base-level accuracy that incorporates several elements not captured in prior assemblies. As genome assembly methods continue to advance and production costs become a less significant obstacle, genome assemblies for multiple inbred rat strains are emerging. These assemblies will allow a rat pangenome assembly to be constructed that captures all the genetic variations in strains selected for their utility in research and will overcome reference bias, a limitation associated with reliance on a single reference assembly. By this means, the full utility of this model organism to genomic studies will begin to be revealed.

A telomere-to-telomere Eucalyptus regnans genome: unveiling haplotype variance in structure and genes within one of the world’s tallest trees

BackgroundEucalyptus regnans (Mountain Ash) is an Australian native giant tree species which form forests that are among the highest known carbon-dense biomasses in the world. To enhance genomic studies in this ecologically important species, we assembled a high-quality, mostly telomere-to-telomere complete, chromosome-level, haplotype-resolved reference genome. We sampled a single tree, the Centurion, which is currently a contender for the world’s tallest flowering plant.ResultsUsing long-read sequencing data (PacBio HiFi, Oxford Nanopore ultra-long reads) and chromosome conformation capture data (Hi-C), we assembled the most contiguous and complete Eucalyptus reference genome to date. For each haplotype, we observed contig N50s exceeding 36 Mbp, scaffold N50s exceeding 43 Mbp, and genome BUSCO completeness exceeding 99%. The assembled genome revealed extensive structural variations between the two haplotypes, consisting mostly of insertions, deletions, duplications and translocations. Analysis of gene content revealed haplotype-specific genes, which were enriched in functional categories related to transcription, energy production and conservation. Additionally, many genes reside within structurally rearranged regions, particularly duplications, suggesting that haplotype-specific variation may contribute to environmental adaptation in the species.ConclusionsOur study provides a foundation for future research into E. regnans environmental adaptation, and the high-quality genome will be a powerful resource for conservation of carbon-dense giant tree forests.

Comparative Analysis of Plyometric Training Protocols in Volleyball: A Meta-Analysis

Purpose: This meta-analysis synthesizes findings from 40 research studies to comprehensively examine the impact of plyometric training protocols on overall performance outcomes in volleyball athletes. Method: A systematic search identified relevant studies on plyometrics and volleyball performance. Data on training programs, performance measures, and outcomes were extracted. Statistical analysis assessed the overall effect size and addressed potential variation between studies. Findings: The constant-effects version revealed a giant wonderful common impact (Fisher r-to-z converted correlation coefficient = 0.7531, 95% CI: 0.6952 to 0.8110), affirming the fine relationship between plyometrics and volleyball overall performance. However, the random-effects model indicated giant heterogeneity among authentic effects (I² = 78.53%), prompting a radical exploration of ability sources of variability. Despite diagnosed limitations, inclusive of heterogeneity and ability publication bias, this meta-analysis underscores the want for standardized protocols and in addition studies to refine training strategies. Conclusion: The importance of this analysis lies in its contribution to proof-primarily based practices, providing valuable insights to coaches and practitioners in search of to optimize plyometric training for volleyball athletes. As a complete reference, it courses destiny research endeavors and advances know-how within the nuanced courting between plyometrics and volleyball.

Basis-Set Limit CCSD(T) Energies for Large Molecules with Local Natural Orbitals and Reduced-Scaling Basis-Set Corrections.

The calculation of density-based basis-set correction (DBBSC), which remedies the basis-set incompleteness (BSI) error of the correlation energy, is combined with local approximations. Aiming at large-scale applications, the procedure is implemented in our efficient local natural orbital-based coupled-cluster singles and doubles with perturbative triples [LNO-CCSD(T)] scheme. To this end, the range-separation function, which characterizes the one-electron BSI in space, is decomposed into the sum of contributions from individual localized molecular orbitals (LMOs). A compact domain is constructed around each LMO, and the corresponding contributions are evaluated only within these restricted domains. Furthermore, for the calculation of the complementary auxiliary basis set (CABS) correction, which significantly improves the Hartree-Fock (HF) energy, the local density fitting approximation is utilized. The errors arising from the local approximations are examined in detail, efficient prescreening techniques are introduced to compress the numerical quadrature used for DBBSC, and conservative default thresholds are selected for the truncation parameters. The efficiency of the DBBSC-LNO-CCSD(T) method is demonstrated through representative examples of up to 1000 atoms. Based on the numerical results, we conclude that the corrections drastically reduce the BSI error using double-ζ basis sets, often to below 1 kcal/mol compared to the reliable LNO-CCSD(T) complete basis set references, while significant improvements are also achieved with triple-ζ basis sets. Considering that the calculation of the DBBSC and CABS corrections only moderately increases the wall-clock time required for the post-HF steps in practical applications, the proposed DBBSC-LNO-CCSD(T) method offers a highly efficient and robust tool for large-scale calculations.

EDNAqua-Plan—Standardisation Overview for eDNA Sequencing of Aquatic Organisms and the Downstream Data Ecosystem

The eDNAqua-Plan project stands as a beacon of innovation in the biomonitoring of marine and freshwater ecosystems, propelled by the urgent need to integrate DNA-based approaches in aquatic bioassessment and monitoring frameworks. The broad utilisation of cutting-edge environmental DNA (eDNA) and DNA barcoding methodologies is dependent on complete, reliable, and accessible reference DNA sequence data (Rimet et al. 2021). Complete and interoperable metadata is crucial to allow a broad reuse of (e)DNA data and analysis outputs, and for a broader uptake of results by end users. The eDNAqua-Plan project aims to address key limitations to the routine implementation of eDNA-based monitoring methods in Europe by developing plans for federated DNA barcode reference libraries and eDNA data repositories to support DNA-based environmental monitoring. This will ensure a sustainable and reliable infrastructure to underpin its broad use, thereby paving the way for more effective conservation and management strategies. The project is working towards creating a comprehensive overview of standardisation efforts and data workflows, through collaborations with other projects, initiatives and infrastructures for aquatic monitoring across the European Union (EU) and associated countries. We are analysing existing archives (e.g., International Nucleotide Sequence Database Collaboration (INSDC), Barcode of Life Data System (BOLD), Global Biodiversity Information Facility (GBIF), Ocean Biodiversity Information System (OBIS)), portals, and papers to determine current and best practices through the use of questionnaires, manual evaluation of repositories, and machine learning methods (LLMs). This includes an overview of the usage of existing metadata and data standards (e.g., Minimum Information about any (X) Sequence Specifications from the Genomics Standards Consortium (GSC), Darwin Core standard). The results are being integrated by a team of experts in marine and freshwater biomonitoring. With a diverse consortium comprising 18 partner institutions from 11 countries and one international institute, eDNAqua-Plan brings together experts in marine and freshwater monitoring, eDNA analysis, and data science. The collective effort by this consortium will lay the groundwork for the creation of a digital ecosystem of eDNA repositories and an integrated reference library of marine and freshwater species, adhering to FAIR (Findable, Accessible, Interoperable, and Reusable) principles.

Hybrid assembly and comparative genomics unveil insights into the evolution and biology of the red-legged partridge

The red-legged partridge Alectoris rufa plays a crucial role in the ecosystem of southwestern Europe, and understanding its genetics is vital for conservation and management. Here we sequence, assemble, and annotate a highly contiguous and nearly complete version of its genome. This assembly encompasses 96.9% of the avian genes flagged as essential in the BUSCO aves_odb10 dataset. Moreover, we pinpointed RNA and protein-coding genes, 95% of which had functional annotations. Notably, we observed significant chromosome rearrangements in comparison to quail (Coturnix japonica) and chicken (Gallus gallus). In addition, a comparative phylogenetic analysis of these genomes suggests that A. rufa and C. japonica diverged roughly 20 million years ago and that their common ancestor diverged from G. gallus 35 million years ago. Our assembly represents a significant advancement towards a complete reference genome for A. rufa, facilitating comparative avian genomics, and providing a valuable resource for future research and conservation efforts for the red-legged partridge.

De-novo assembly of a reference genome for a critically threatened Aotearoa New Zealand tree species, Syzygium maire (Myrtaceae)

Aotearoa New Zealand’s swamp forests have experienced significant habitat loss in fewer than two hundred years. Many of the country’s tree species are endemic with sparse to no genetic information available to underpin conservation strategies. Syzygium maire, Aotearoa’s only endemic Syzygium species, is a culturally and ecologically important component of swamp forest habitats. Unfortunately, populations of S. maire have been greatly reduced, heavily fragmented and are susceptible to the emergent pathogen Austropuccinia psidii (myrtle rust), posing eminent danger of a further decline of the species. We sought to develop genomic resources to inform conservation management of S. maire. To this end, we used long read, high accuracy sequencing technology to produce a highly complete reference quality genome for S. maire. The genome sequence was named ‘Ngā Hua o te Ia Whenua’ by the local Māori tribe where the tree used for genome sequencing grows. We assess whether genome-level divergence with other Myrtaceae may have followed geographic isolation of the species. We detect conservation of large scale synteny between three Syzygium species and Eucalyptus grandis, providing support for the stability of Syzygium genomes across evolutionary time. We annotate genes implicated in fungal pathogen defence, identifying several hundred putative NLR genes, including putative homologs of previously identified Austropuccinia psidii resistance genes. Finally, we evaluate the genetic relationships of individuals of a small, isolated population of trees. We find evidence of high levels of kinship and inbreeding within small and isolated S. maire populations, informing local-scale conservation strategies for the species. Our findings enable practical conservation actions and provide resources for larger scale studies of S. maire and other Syzygium species in the future.

Development and implementation of a core genome multilocus sequence typing scheme for Haemophilus influenzae.

Haemophilus influenzae is part of the human nasopharyngeal microbiota and a pathogen causing invasive disease. The extensive genetic diversity observed in H. influenzae necessitates discriminatory analytical approaches to evaluate its population structure. This study developed a core genome multilocus sequence typing (cgMLST) scheme for H. influenzae using pangenome analysis tools and validated the cgMLST scheme using datasets consisting of complete reference genomes (N = 14) and high-quality draft H. influenzae genomes (N = 2297). The draft genome dataset was divided into a development dataset (N = 921) and a validation dataset (N = 1376). The development dataset was used to identify potential core genes, and the validation dataset was used to refine the final core gene list to ensure the reliability of the proposed cgMLST scheme. Functional classifications were made for all the resulting core genes. Phylogenetic analyses were performed using both allelic profiles and nucleotide sequence alignments of the core genome to test congruence, as assessed by Spearman's correlation and ordinary least square linear regression tests. Preliminary analyses using the development dataset identified 1067 core genes, which were refined to 1037 with the validation dataset. More than 70% of core genes were predicted to encode proteins essential for metabolism or genetic information processing. Phylogenetic and statistical analyses indicated that the core genome allelic profile accurately represented phylogenetic relatedness among the isolates (R 2 = 0.945). We used this cgMLST scheme to define a high-resolution population structure for H. influenzae, which enhances the genomic analysis of this clinically relevant human pathogen.