The NGS Magic Pudding: A Nanopore-Led Long-Read Genome Assembly for the Commercial Australian Freshwater Crayfish, Cherax destructor.

Cherax destructor, the yabby, is an iconic Australian freshwater crayfish species, which, similar to other major invertebrate groups, is grossly under-represented in genomic databases. The yabby is also the principal commercial freshwater crustacean species in Australia subject to explotation via inland fisheries and aquaculture. To address the genomics knowledge gap for this species and explore cost effective and efficient methods for genome assembly, we generated 106.8 gb of Nanopore reads and performed a long-read only assembly of the Cherax destructor genome. On a mini-server configured with an ultra-fast swap space, the de novo assembly took 131 h (∼5.5 days). Genome polishing with 126.3 gb of PCR-Free Illumina reads generated an assembled genome size of 3.3 gb (74.6% BUSCO completeness) with a contig N50 of 80,900 bp, making it the most contiguous for freshwater crayfish genome assemblies. We found an unusually large number of cellulase genes within the yabby genome which is relevant to understanding the nutritional biology, commercial feed development, and ecological role of this species and crayfish more generally. These resources will be useful for genomic research on freshwater crayfish and our methods for rapid and super-efficient genome assembly will have wide application.

Genotypic characterization and genome comparison reveal insights into potential vaccine coverage and genealogy of Neisseria meningitidis in military camps in Vietnam.

BACKGROUND: Neisseria meningitidis remains the main cause of sporadic meningitis and sepsis in military camps in Vietnam. Yet, very limited molecular data of their genotypic and epidemiological characteristics are available from Vietnam, and particularly the military environment. Whole genome sequencing (WGS) has proven useful for meningococcal disease surveillance and guiding preventative vaccination programs. Previously, we characterized key genetic and epidemiological features of an invasive N. meningitidis B isolate from a military unit in Vietnam. Here, we extend these findings by sequencing two additional invasive N. meningitidis B isolated from cerebrospinal fluid (CSF) of two meningitis cases at another military unit and compared their genomic sequences and features. We also report the sequence types and antigenic profiles of 25 historical and more recently emerged N. meningitidis isolates from these units and other units in proximity. METHODS: Strains were sequenced using the Illumina HiSeq platform, de novo assembled and annotated. Genomes were compared within and between military units, as well as against the global N. meningitidis collection and other isolates from the Southeast Asia region using PubMLST. Variations at the nucleotide level were determined, and phylogenetic relationships were estimated. Antigenic genotypes and vaccine coverage were analyzed using gMATS and PubMLST. Susceptibility of isolates against commonly used antibiotic agents was examined using E-test. RESULTS: Genome comparison revealed a high level of similarity among isolates both within and between units. All isolates showed resistance to chloramphenicol and carried identical catP gene with other Southeast Asian isolates, suggesting a common lineage. Their antigenic genotypes predicted no coverage by either Bexsero®or Trumenba®, and nucleotide variation analysis revealed diverse new, unassigned alleles at multiple virulence loci of all strains. Groups of singleton and unique novel sequence types extending beyond individual camps were found from epidemiological data of 25 other isolates. Our results add to the sparse published molecular data of N. meningitidis in the military units in Vietnam, highlight their diversity, distinct genetic features and antibiotic resistance pattern, and emphasize the need for further studies on the molecular characteristics of N. meningitidis in Vietnam.

Dataset for genome sequencing and de novo assembly of the Vietnamese bighead catfish (Clarias macrocephalus Günther, 1864).

Freshwater catfish of the genus Clarias, known as the airbreathing catfish, are widespread and important for food security through small scale inland fisheries and aquaculture. Limited genomic data are available for this important group of fishes. The bighead catfish (Clarias macrocephalus) is a commercial aquaculture species in southeast Asia used for aquaculture and threatened in its natural environment through habitat destruction, over-exploitation and competition from other introduced species of Clarias. Despite its commercial importance and threats to natural populations, public databases do not include any genomic data for C. macrocephalus. We present the first genomic data for the bighead catfish from Illumina sequencing. A total of 128 Gb of sequence data in paired-end 150 bp reads were assembled de novo, generating a final assembly of 883 Mbp contained in 27,833 scaffolds (N50 length: 80.8 kbp) with BUSCO completeness assessments of 96.3% and 87.6% based on metazoan and Actinopterygii ortholog datasets, respectively. Annotation of the genome predicted 21,124 gene sequences, which were assigned putative functions based on homology to existing protein sequences in public databases. Raw fastq reads and the final version of the genome assembly have been deposited in the NCBI (BioProject: PRJNA604477, WGS: JAAGKR000000000, SRA: SRR11188453). The complete C. macrocephalus mitochondrial genome was also recovered from the same sequence read dataset and is available on NCBI (accession: MT109097), representing the first mitogenome for this species. Lastly, we find an expansion of the mb and ora1 genes thought to be associated with adaptations to air-breathing and a semi-terrestrial life style in this genus of catfish.

Dataset for sequencing and de novo assembly of the European endangered white-clawed crayfish (Austropotamobius pallipes) abdominal muscle transcriptome.

The white-clawed crayfish (Austropotamobius pallipes) is an endangered species in Europe with limited genomic information. Despite its conservation status there is no transcriptomic data available for A. pallipes in public databases. The data here represents the first transcriptome profile of the white-clawed crayfish generated using Illumina stranded RNA sequencing. Pair-end reads were assembled de novo with three separate transcriptome assemblers (Trinity, RNABloom, and RNASpades) followed by transcript assembly reduction and gene reconstruction using the EvidentialGene pipeline. The transcriptome was functionally annotated using InterProScan and genes coding for carbohydrate-active enzymes were identified through the dbCAN2 server. Raw fastq reads and the final version of the transcriptome assembly have been deposited in the NCBI-SRA (SRR10549898) and NCBI-TSA (GICG01) databases.

Improved genomic resources for the black tiger prawn (Penaeus monodon).

World production of farmed crustaceans was 7.8 million tons in 2016. While only making up approximately 10% of world aquaculture production, crustaceans are generally high-value species and can earn significant export income for producing countries. Viet Nam is a major seafood producing country earning USD 7.3 billion in 2016 in export income with shrimp as a major commodity. However, there is a general lack of genomic resources available for shrimp species, which is challenging to obtain due to the need to deal with large repetitive genomes, which characterize many decapod crustaceans. The first tiger prawn (P. monodon) genome assembly was assembled in 2016 using the standard Illumina PCR-based pair-end reads and a computationally-efficient but relatively suboptimal assembler, SOAPdenovo v2. As a result, the current P. monodon draft genome is highly fragmented (> 2 million scaffolds with N50 length of <1000 bp), exhibiting only moderate genome completeness (< 35% BUSCO complete single-copy genes). We sought to improve upon the recently published P. monodon genome assembly and completeness by generating Illumina PCR-free pair-end sequencing reads to eliminate genomic gaps associated with PCR-bias and performing de novo assembly using the updated MaSurCA de novo assembler. Furthermore, we scaffolded the assembly with low coverage Nanopore long reads and several recently published deep Illumina transcriptome paired-end sequencing data, producing a final genome assembly of 1.6 Gbp (1,211,364 scaffolds; N50 length of 1982 bp) with an Arthropod BUSCO completeness of 96.8%. Compared to the previously published P. monodon genome assembly from China (NCBI Accession Code: NIUS01), this represents an almost 20% increase in the overall BUSCO genome completeness that now consists of more than 90% of Arthropod BUSCO single-copy genes. The revised P. monodon genome assembly (NCBI Accession Code: VIGR01) will be a valuable resource to support ongoing functional genomics and molecular-based breeding studies in Vietnam.

The Pontastacus leptodactylus (Astacidae) Repeatome Provides Insight Into Genome Evolution and Reveals Remarkable Diversity of Satellite DNA.

Pontastacus leptodactylus is a native European crayfish species found in both freshwater and brackish environments. It has commercial importance for fisheries and aquaculture industries. Up till now, most studies concerning P. leptodactylus have focused onto gaining knowledge about its phylogeny and population genetics. However, little is known about the chromosomal evolution and genome organization of this species. Therefore, we performed clustering analysis of a low coverage genomic dataset to identify and characterize repetitive DNA in the P. leptodactylus genome. In addition, the karyogram of P. leptodactylus (2n = 180) is presented here for the first time consisting of 75 metacentric, 14 submetacentric, and a submetacentric/metacentric heteromorphic chromosome pair. We determined the genome size to be at ~18.7 gigabase pairs. Repetitive DNA represents about 54.85% of the genome. Satellite DNA repeats are the most abundant type of repetitive DNA, making up to ~28% of the total amount of repetitive elements, followed by the Ty3/Gypsy retroelements (~15%). Our study established a surprisingly high diversity of satellite repeats in P. leptodactylus. The genome of P. leptodactylus is by far the most satellite-rich genome discovered to date with 258 satellite families described. Of the five mapped satellite DNA families on chromosomes, PlSAT3-411 co-localizes with the AT-rich DAPI positive probable (peri)centromeric heterochromatin on all chromosomes, while PlSAT14-79 co-localizes with the AT-rich DAPI positive (peri)centromeric heterochromatin on one chromosome and is also located subterminally and intercalary on some chromosomes. PlSAT1-21 is located intercalary in the vicinity of the (peri)centromeric heterochromatin on some chromosomes, while PlSAT6-70 and PlSAT7-134 are located intercalary on some P. leptodactylus chromosomes. The FISH results reveal amplification of interstitial telomeric repeats (ITRs) in P. leptodactylus. The prevalence of repetitive elements, especially the satellite DNA repeats, may have provided a driving force for the evolution of the P. leptodactylus genome.

Genomic characterization of Vibrio parahaemolyticus from Pacific white shrimp and rearing water in Malaysia reveals novel sequence types and structural variation in genomic regions containing the Photorhabdus insect-related (Pir) toxin-like genes.

The Malaysian and global shrimp aquaculture production has been significantly impacted by acute hepatopancreatic necrosis disease (AHPND) typically caused by Vibrio parahaemolyticus harboring the pVA plasmid containing the pirAVp and pirBVp genes, which code for Photorhabdus insect-related (Pir) toxin. The limited genomic resource for V. parahaemolyticus strains from Malaysian aquaculture farms precludes an in-depth understanding of their diversity and evolutionary relationships. In this study, we isolated shrimp-associated and environmental (rearing water) V. parahaemolyticus from three aquaculture farms located in Northern and Central Malaysia followed by whole-genome sequencing of 40 randomly selected isolates on the Illumina MiSeq. Phylogenomic analysis and multilocus sequence typing (MLST) reveal distinct lineages of V. parahaemolyticus that harbor the pirABVp genes. The recovery of pVA plasmid backbone devoid of pirAVp or pirABVp in some V. parahaemolyticus isolates suggests that the toxin genes are prone to deletion. The new insight gained from phylogenomic analysis of Asian V. parahaemolyticus, in addition to the observed genomic instability of pVa plasmid, will have implications for improvements in aquaculture practices to diagnose, treat or limit the impacts of this disease.

Genomic evidence of neo-sex chromosomes in the eastern yellow robin.

BACKGROUND: Understanding sex-biased natural selection can be enhanced by access to well-annotated chromosomes including ones inherited in sex-specific fashion. The eastern yellow robin (EYR) is an endemic Australian songbird inferred to have experienced climate-driven sex-biased selection and is a prominent model for studying mitochondrial-nuclear interactions in the wild. However, the lack of an EYR reference genome containing both sex chromosomes (in birds, a female bearing Z and W chromosomes) limits efforts to understand the mechanisms of these processes. Here, we assemble the genome for a female EYR and use low-depth (10×) genome resequencing data from 19 individuals of known sex to identify chromosome fragments with sex-specific inheritance. FINDINGS: MaSuRCA hybrid assembly using Nanopore and Illumina reads generated a 1.22-Gb EYR genome in 20,702 scaffolds (94.2% BUSCO completeness). Scaffolds were tested for W-linked (female-only) inheritance using a k-mer approach, and for Z-linked inheritance using median read-depth test in male and female reads (read-depths must indicate haploid female and diploid male representation). This resulted in 2,372 W-linked scaffolds (total length: 97,872,282 bp, N50: 81,931 bp) and 586 Z-linked scaffolds (total length: 121,817,358 bp, N50: 551,641 bp). Anchoring of the sex-linked EYR scaffolds to the reference genome of a female zebra finch revealed 2 categories of sex-linked genomic regions. First, 653 W-linked scaffolds (25.7 Mb) were anchored to the W sex chromosome and 215 Z-linked scaffolds (74.4 Mb) to the Z. Second, 1,138 W-linked scaffolds (70.9 Mb) and 179 Z-linked scaffolds (51.0 Mb) were anchored to a large section (coordinates ∼5 to ∼60 Mb) of zebra finch chromosome 1A. The first ∼5 Mb and last ∼14 Mb of the reference chromosome 1A had only autosomally behaving EYR scaffolds mapping to them. CONCLUSIONS: We report a female (W chromosome-containing) EYR genome and provide genomic evidence for a neo-sex (neo-W and neo-Z) chromosome system in the EYR, involving most of a large chromosome (1A) previously only reported to be autosomal in passerines.

Comparative mitogenomics of the Decapoda reveals evolutionary heterogeneity in architecture and composition.

The emergence of cost-effective and rapid sequencing approaches has resulted in an exponential rise in the number of mitogenomes on public databases in recent years, providing greater opportunity for undertaking large-scale comparative genomic and systematic research. Nonetheless, current datasets predominately come from small and disconnected studies on a limited number of related species, introducing sampling biases and impeding research of broad taxonomic relevance. This study contributes 21 crustacean mitogenomes from several under-represented decapod infraorders including Polychelida and Stenopodidea, which are used in combination with 225 mitogenomes available on NCBI to investigate decapod mitogenome diversity and phylogeny. An overview of mitochondrial gene orders (MGOs) reveals a high level of genomic variability within the Decapoda, with a large number of MGOs deviating from the ancestral arthropod ground pattern and unevenly distributed among infraorders. Despite the substantial morphological and ecological variation among decapods, there was limited evidence for correlations between gene rearrangement events and species ecology or lineage specific nucleotide substitution rates. Within a phylogenetic context, predicted scenarios of rearrangements show some MGOs to be informative synapomorphies for some taxonomic groups providing strong independent support for phylogenetic relationships. Additional comparisons for a range of mitogenomic features including nucleotide composition, strand asymmetry, unassigned regions and codon usage indicate several clade-specific trends that are of evolutionary and ecological interest.

Whole-genome sequencing and characterization of an antibiotic resistant Neisseria meningitidis B isolate from a military unit in Vietnam.

BACKGROUND: Invasive meningococcal disease (IMD) persists in military units in Vietnam despite the availability of antibiotics and vaccines. A hindrance to reducing the incidence of IMD in Vietnam is a lack of molecular data from isolates of the causative agent, Neisseria meningitidis from this country. Here, we characterized key genetic and epidemiological features of an invasive N. meningitidis isolate from a military unit in Vietnam using whole-genome sequencing. METHODS: Neisseria meningitidis was isolated from a conscript admitted for meningitis and tested against seven antibiotics. DNA from the isolate was extracted and sequenced using the Illumina HiSeq platform. Denovo assembly and scaffolding were performed to construct a draft genome assembly, from which genes were predicted and functionally annotated. Genome analysis included epidemiological characterization, genomic composition and identification of antibiotic resistance genes. RESULTS: Susceptibility testing of the isolate showed high levels of resistance to chloramphenicol and diminished susceptibility to ampicillin and rifampicin. A draft genome of ~ 2.1 Mb was assembled, containing 2451 protein coding sequences, 49 tRNAs and 3 rRNAs. Fifteen coding sequences sharing ≥ 84% identity with known antibiotic resistance genes were identified. Genome analysis revealed abundant repetitive DNAs and two prophages. Epidemiological typing revealed newly described sequence type, antigenic finetype and Bexsero® Antigen Sequence Typing (BAST). The BAST profile showed no coverage by either Bexsero® or Trumenba®. CONCLUSIONS: Our results present the first genome assembly of an invasive N. meningitidis isolate from a military unit in Vietnam. This study illustrates the usefulness of whole genome sequencing (WGS) analysis for epidemiological and antibiotic resistance studies and surveillance of IMD in Vietnam.

Two reads to rule them all: Nanopore long read-guided assembly of the iconic Christmas Island red crab, Gecarcoidea natalis (Pocock, 1888), mitochondrial genome and the challenges of AT-rich mitogenomes.

Despite recent advances in sequencing technology, a complete mitogenome assembly is still unavailable for the gecarcinid land crabs that include the iconic Christmas Island red crab (Gecarcoidea natalis) which is known for its high population density, annual mass breeding migration and ecological significance in maintaining rainforest structure. Using sequences generated from Nanopore and Illumina platforms, we assembled the complete mitogenome for G. natalis, the first for the genus and only second for the family Gecarcinidae. Nine Nanopore long reads representing 0.15% of the sequencing output from an overnight MinION Nanopore run were aligned to the mitogenome. Two of them were >10 kb and combined are sufficient to span the entire G. natalis mitogenome. The use of Illumina genome skimming data only resulted in a fragmented assembly that can be attributed to low to zero sequencing coverage in multiple high AT-regions including the mitochondrial protein-coding genes (NAD4 and NAD5), 16S ribosomal rRNA and non-coding control region. Supplementing the mitogenome assembly with previously acquired transcriptome dataset containing high abundance of mitochondrial transcripts improved mitogenome sequence coverage and assembly reliability. We then inferred the phylogeny of the Eubrachyura using Maximum Likelihood and Bayesian approaches, confirming the phylogenetic placement of G. natalis within the family Gecarcinidae based on whole mitogenome alignment. Given the substantial impact of AT-content on mitogenome assembly and the value of complete mitogenomes in phylogenetic and comparative studies, we recommend that future mitogenome sequencing projects consider generating a modest amount of Nanopore long reads to facilitate the closing of problematic and fragmented mitogenome assemblies.

Microbiome analysis of Pacific white shrimp gut and rearing water from Malaysia and Vietnam: implications for aquaculture research and management.

Aquaculture production of the Pacific white shrimp is the largest in the world for crustacean species. Crucial to the sustainable global production of this important seafood species is a fundamental understanding of the shrimp gut microbiota and its relationship to the microbial ecology of shrimp pond. This is especially true, given the recently recognized role of beneficial microbes in promoting shrimp nutrient intake and in conferring resistance against pathogens. Unfortunately, aquaculture-related microbiome studies are scarce in Southeast Asia countries despite the severe impact of early mortality syndrome outbreaks on shrimp production in the region. In this study, we employed the 16S rRNA amplicon (V3-V4 region) sequencing and amplicon sequence variants (ASV) method to investigate the microbial diversity of shrimp guts and pond water samples collected from aquaculture farms located in Malaysia and Vietnam. Substantial differences in the pond microbiota were observed between countries with the presence and absence of several taxa extending to the family level. Microbial diversity of the shrimp gut was found to be generally lower than that of the pond environments with a few ubiquitous genera representing a majority of the shrimp gut microbial diversity such as Vibrio and Photobacterium, indicating host-specific selection of microbial species. Given the high sequence conservation of the 16S rRNA gene, we assessed its veracity at distinguishing Vibrio species based on nucleotide alignment against type strain reference sequences and demonstrated the utility of ASV approach in uncovering a wider diversity of Vibrio species compared to the conventional OTU clustering approach.

ORDER within the chaos: Insights into phylogenetic relationships within the Anomura (Crustacea: Decapoda) from mitochondrial sequences and gene order rearrangements.

The infraorder Anomura consists of a morphologically and ecologically heterogeneous group of decapod crustaceans, and has attracted interest from taxonomists for decades attempting to find some order out of the seemingly chaotic diversity within the group. Species-level diversity within the Anomura runs the gamut from the "hairy" spindly-legged yeti crab found in deep-sea hydrothermal vent environments to the largest known terrestrial invertebrate, the robust coconut or robber crab. Owing to a well-developed capacity for parallel evolution, as evidenced by the occurrence of multiple independent carcinization events, Anomura has long tested the patience and skill of both taxonomists attempting to find order, and phylogeneticists trying to establish stable hypotheses of evolutionary inter-relationships. In this study, we performed genome skimming to recover the mitogenome sequences of 12 anomuran species including the world's largest extant invertebrate, the robber crab (Birgus latro), thereby over doubling these resources for this group, together with 8 new brachyuran mitogenomes. Maximum-likelihood (ML) and Bayesian-inferred (BI) phylogenetic reconstructions based on amino acid sequences from mitogenome protein-coding genes provided strong support for the monophyly of the Anomura and Brachyura and their sister relationship, consistent with previous studies. The majority of relationships within families were supported and were largely consistent with current taxonomic classifications, whereas many relationships at higher taxonomic levels were unresolved. Nevertheless, we have strong support for a polyphyletic Paguroidea and recovered a well-supported clade of a subset of paguroids (Diogenidae + Coenobitidae) basal to all other anomurans, though this requires further testing with greater taxonomic sampling. We also introduce a new feature to the MitoPhAST bioinformatics pipeline (https://github.com/mht85/MitoPhAST) that enables the extraction of mitochondrial gene order (MGO) information directly from GenBank files and clusters groups based on common MGOs. Using this tool, we compared MGOs across the Anomura and Brachyura, identifying Anomura as a taxonomic "hot spot" with high variability in MGOs among congeneric species from multiple families while noting the broad association of highly-rearranged MGOs with several anomuran lineages inhabiting extreme niches. We also demonstrate the value of MGOs as a source of novel synapomorphies for independently reinforcing tree-based relationships and for shedding light on relationships among challenging groups such as the Aegloidea and Lomisoidea that were unresolved in phylogenetic reconstructions. Overall, this study contributes a substantial amount of new genetic material for Anomura and attempts to further resolve anomuran evolutionary relationships where possible based on a combination of sequence and MGO information. The new feature in MitoPhAST adds to the relatively limited number of bioinformatics tools available for MGO analyses, which can be utilized widely across animal groups.

Finding Nemo: hybrid assembly with Oxford Nanopore and Illumina reads greatly improves the clownfish (Amphiprion ocellaris) genome assembly.

Background: Some of the most widely recognized coral reef fishes are clownfish or anemonefish, members of the family Pomacentridae (subfamily: Amphiprioninae). They are popular aquarium species due to their bright colours, adaptability to captivity, and fascinating behavior. Their breeding biology (sequential hermaphrodites) and symbiotic mutualism with sea anemones have attracted much scientific interest. Moreover, there are some curious geographic-based phenotypes that warrant investigation. Leveraging on the advancement in Nanopore long read technology, we report the first hybrid assembly of the clown anemonefish (Amphiprion ocellaris) genome utilizing Illumina and Nanopore reads, further demonstrating the substantial impact of modest long read sequencing data sets on improving genome assembly statistics. Results: We generated 43 Gb of short Illumina reads and 9 Gb of long Nanopore reads, representing approximate genome coverage of 54× and 11×, respectively, based on the range of estimated k-mer-predicted genome sizes of between 791 and 967 Mbp. The final assembled genome is contained in 6404 scaffolds with an accumulated length of 880 Mb (96.3% BUSCO-calculated genome completeness). Compared with the Illumina-only assembly, the hybrid approach generated 94% fewer scaffolds with an 18-fold increase in N50 length (401 kb) and increased the genome completeness by an additional 16%. A total of 27 240 high-quality protein-coding genes were predicted from the clown anemonefish, 26 211 (96%) of which were annotated functionally with information from either sequence homology or protein signature searches. Conclusions: We present the first genome of any anemonefish and demonstrate the value of low coverage (∼11×) long Nanopore read sequencing in improving both genome assembly contiguity and completeness. The near-complete assembly of the A. ocellaris genome will be an invaluable molecular resource for supporting a range of genetic, genomic, and phylogenetic studies specifically for clownfish and more generally for other related fish species of the family Pomacentridae.

More evolution underground: Accelerated mitochondrial substitution rate in Australian burrowing freshwater crayfishes (Decapoda: Parastacidae).

To further understand the evolutionary history and mitogenomic features of Australia's highly distinctive freshwater crayfish fauna, we utilized a recently described rapid mitogenome sequencing pipeline to generate 24 new crayfish mitogenomes including a diversity of burrowing crayfish species and the first for Astacopsis gouldi, the world's largest freshwater invertebrate. Whole mitogenome-based phylogeny estimates using both Bayesian and Maximum Likelihood methods substantially strengthen existing hypotheses for systematic relationships among Australian freshwater crayfish with evidence of pervasive diversifying selection and accelerated mitochondrial substitution rate among the members of the clade representing strongly burrowing crayfish that may reflect selection pressures for increased energy requirement for adaptation to terrestrial environment and a burrowing lifestyle. Further, gene rearrangements are prevalent in the burrowing crayfish mitogenomes involving both tRNA and protein coding genes. In addition, duplicated control regions were observed in two closely related Engaeus species, together with evidence for concerted evolution. This study significantly adds to the understanding of Australian freshwater crayfish evolutionary relationships and suggests a link between mitogenome evolution and adaptation to terrestrial environments and a burrowing lifestyle in freshwater crayfish.

Population mitogenomics provides insights into evolutionary history, source of invasions and diversifying selection in the House Crow (Corvus splendens).

The House Crow (Corvus splendens) is a useful study system for investigating the genetic basis of adaptations underpinning successful range expansion. The species originates from the Indian subcontinent, but has successfully spread through a variety of thermal environments across Asia, Africa and Europe. Here, population mitogenomics was used to investigate the colonisation history and to test for signals of molecular selection on the mitochondrial genome. We sequenced the mitogenomes of 89 House Crows spanning four native and five invasive populations. A Bayesian dated phylogeny, based on the 13 mitochondrial protein-coding genes, supports a mid-Pleistocene (~630,000 years ago) divergence between the most distant genetic lineages. Phylogeographic patterns suggest that northern South Asia is the likely centre of origin for the species. Codon-based analyses of selection and assessments of changes in amino acid properties provide evidence of positive selection on the ND2 and ND5 genes against a background of purifying selection across the mitogenome. Protein homology modelling suggests that four amino acid substitutions inferred to be under positive selection may modulate coupling efficiency and proton translocation mediated by OXPHOS complex I. The identified substitutions are found within native House Crow lineages and ecological niche modelling predicts suitable climatic areas for the establishment of crow populations within the invasive range. Mitogenomic patterns in the invasive range of the species are more strongly associated with introduction history than climate. We speculate that invasions of the House Crow have been facilitated by standing genetic variation that accumulated due to diversifying selection within the native range.