Elucidating the ecophysiology of soybean pod-sucking stinkbug Riptortus pedestris (Hemiptera: Alydidae) based on de novo genome assembly and transcriptome analysis.

Food security is important for the ever-growing global population. Soybean, Glycine max (L.) Merr., is cultivated worldwide providing a key source of food, protein and oil. Hence, it is imperative to maintain or to increase its yield under different conditions including challenges caused by abiotic and biotic stresses. In recent years, the soybean pod-sucking stinkbug Riptortus pedestris has emerged as an important agricultural insect pest in East, South and Southeast Asia. Here, we present a genomics resource for R. pedestris including its genome assembly, messenger RNA (mRNA) and microRNA (miRNA) transcriptomes at different developmental stages and from different organs. As insect hormone biosynthesis genes (genes involved in metamorphosis) and their regulators such as miRNAs are potential targets for pest control, we analyzed the sesquiterpenoid (juvenile) and ecdysteroid (molting) hormone biosynthesis pathway genes including their miRNAs and relevant neuropeptides. Temporal gene expression changes of these insect hormone biosynthesis pathways were observed at different developmental stages. Similarly, a diet-specific response in gene expression was also observed in both head and salivary glands. Furthermore, we observed that microRNAs (bantam, miR-14, miR-316, and miR-263) of R. pedestris fed with different types of soybeans were differentially expressed in the salivary glands indicating a diet-specific response. Interestingly, the opposite arms of miR-281 (-5p and -3p), a miRNA involved in regulating development, were predicted to target Hmgs genes of R. pedestris and soybean, respectively. These observations among others highlight stinkbug's responses as a function of its interaction with soybean. In brief, the results of this study not only present salient findings that could be of potential use in pest management and mitigation but also provide an invaluable resource for R. pedestris as an insect model to facilitate studies on plant-pest interactions.

The genome and sex-dependent responses to temperature in the common yellow butterfly, Eurema hecabe.

BACKGROUND: Lepidoptera (butterflies and moths) is one of the most geographically widespread insect orders in the world, and its species play important and diverse ecological and applied roles. Climate change is one of the biggest challenges to biodiversity this century, and lepidopterans are vulnerable to climate change. Temperature-dependent gene expression differences are of relevance under the ongoing climate crisis. However, little is known about how climate affects gene expression in lepidopterans and the ecological consequences of this, particularly with respect to genes with biased expression in one of the sexes. The common yellow butterfly, Eurema hecabe (Family Pieridae), is one of the most geographically widespread lepidopterans that can be found in Asia, Africa, and Australia. Nevertheless, what temperature-dependent effects there may be and whether the effects differ between the sexes remain largely unexplored. RESULTS: Here, we generated high-quality genomic resources for E. hecabe along with transcriptomes from eight developmental stages. Male and female butterflies were subjected to varying temperatures to assess sex-specific gene expression responses through mRNA and microRNA transcriptomics. We find that there are more temperature-dependent sex-biased genes in females than males, including genes that are involved in a range of biologically important functions, highlighting potential ecological impacts of increased temperatures. Further, by considering available butterfly data on sex-biased gene expression in a comparative genomic framework, we find that the pattern of sex-biased gene expression identified in E. hecabe is highly species-specific, rather than conserved across butterfly species, suggesting that sex-biased gene expression responses to climate change are complex in butterflies. CONCLUSIONS: Our study lays the foundation for further understanding of differential responses to environmental stress in a widespread lepidopteran model and demonstrates the potential complexity of sex-specific responses of lepidopterans to climate change.

Millipede genomes reveal unique adaptations during myriapod evolution.

The Myriapoda, composed of millipedes and centipedes, is a fascinating but poorly understood branch of life, including species with a highly unusual body plan and a range of unique adaptations to their environment. Here, we sequenced and assembled 2 chromosomal-level genomes of the millipedes Helicorthomorpha holstii (assembly size = 182 Mb; shortest scaffold/contig length needed to cover 50% of the genome [N50] = 18.11 Mb mainly on 8 pseudomolecules) and Trigoniulus corallinus (assembly size = 449 Mb, N50 = 26.78 Mb mainly on 17 pseudomolecules). Unique genomic features, patterns of gene regulation, and defence systems in millipedes, not observed in other arthropods, are revealed. Both repeat content and intron size are major contributors to the observed differences in millipede genome size. Tight Hox and the first loose ecdysozoan ParaHox homeobox clusters are identified, and a myriapod-specific genomic rearrangement including Hox3 is also observed. The Argonaute (AGO) proteins for loading small RNAs are duplicated in both millipedes, but unlike in insects, an AGO duplicate has become a pseudogene. Evidence of post-transcriptional modification in small RNAs-including species-specific microRNA arm switching-providing differential gene regulation is also obtained. Millipedes possesses a unique ozadene defensive gland unlike the venomous forcipules found in centipedes. We identify sets of genes associated with the ozadene that play roles in chemical defence as well as antimicrobial activity. Macro-synteny analyses revealed highly conserved genomic blocks between the 2 millipedes and deuterostomes. Collectively, our analyses of millipede genomes reveal that a series of unique adaptations have occurred in this major lineage of arthropod diversity. The 2 high-quality millipede genomes provided here shed new light on the conserved and lineage-specific features of millipedes and centipedes. These findings demonstrate the importance of the consideration of both centipede and millipede genomes-and in particular the reconstruction of the myriapod ancestral situation-for future research to improve understanding of arthropod evolution, and animal evolutionary genomics more widely.

Chromosomal-level reference genome of the moth Heortia vitessoides (Lepidoptera: Crambidae), a major pest of agarwood-producing trees.

The moth Heortia vitessoides Moore (Lepidoptera: Crambidae) is a major pest of ecologically, commercially and culturally important agarwood-producing trees in the genus Aquilaria. In particular, H. vitessoides is one of the most destructive defoliating pests of the incense tree Aquilaria sinesis, which produces a valuable fragrant wood used as incense and in traditional Chinese medicine [33]. Nevertheless, a genomic resource for H. vitessoides is lacking. Here, we present a chromosomal-level assembly for H. vitessoides, consisting of a 517 megabase (Mb) genome assembly with high physical contiguity (scaffold N50 of 18.2 Mb) and high completeness (97.9% complete BUSCO score). To aid gene annotation, 8 messenger RNA transcriptomes from different developmental stages were generated, and a total of 16,421 gene models were predicted. Expansion of gene families involved in xenobiotic metabolism and development were detected, including duplications of cytosolic sulfotransferase (SULT) genes shared among lepidopterans. In addition, small RNA sequencing of 5 developmental stages of H. vitessoides facilitated the identification of 85 lepidopteran conserved microRNAs, 94 lineage-specific microRNAs, as well as several microRNA clusters. A large proportion of the H. vitessoides genome consists of repeats, with a 29.12% total genomic contribution from transposable elements, of which long interspersed nuclear elements (LINEs) are the dominant component (17.41%). A sharp decrease in the genome-wide percentage of LINEs with lower levels of genetic distance to family consensus sequences suggests that LINE activity has peaked in H. vitessoides. In contrast, opposing patterns suggest a substantial recent increase in DNA and LTR element activity. Together with annotations of essential sesquiterpenoid hormonal pathways, neuropeptides, microRNAs and transposable elements, the high-quality genomic and transcriptomic resources we provide for the economically important moth H. vitessoides provide a platform for the development of genomic approaches to pest management, and contribute to addressing fundamental research questions in Lepidoptera.

Status Quo and Future Perspectives of Molecular and Genomic Studies on the Genus Biomphalaria-The Intermediate Snail Host of Schistosoma mansoni.

Schistosomiasis, or also generally known as bilharzia or snail fever, is a parasitic disease that is caused by trematode flatworms of the genus Schistosoma. It is considered by the World Health Organisation as the second most prevalent parasitic disease after malaria and affects more than 230 million people in over 70 countries. People are infected via a variety of activities ranging from agricultural, domestic, occupational to recreational activities, where the freshwater snails Biomphalaria release Schistosoma cercariae larvae that penetrate the skin of humans when exposed in water. Understanding the biology of the intermediate host snail Biomphalaria is thus important to reveal the potential spread of schistosomiasis. In this article, we present an overview of the latest molecular studies focused on the snail Biomphalaria, including its ecology, evolution, and immune response; and propose using genomics as a foundation to further understand and control this disease vector and thus the transmission of schistosomiasis.

Differential microRNA expression, microRNA arm switching, and microRNA:long noncoding RNA interaction in response to salinity stress in soybean.

BACKGROUND: Soybean is a major legume crop with high nutritional and environmental values suitable for sustainable agriculture. Noncoding RNAs (ncRNAs), including microRNAs (miRNAs) and long noncoding RNAs (lncRNAs), are important regulators of gene functions in eukaryotes. However, the interactions between these two types of ncRNAs in the context of plant physiology, especially in response to salinity stress, are poorly understood. RESULTS: Here, we challenged a cultivated soybean accession (C08) and a wild one (W05) with salt treatment and obtained their small RNA transcriptomes at six time points from both root and leaf tissues. In addition to thoroughly analyzing the differentially expressed miRNAs, we also documented the first case of miRNA arm-switching (miR166m), the swapping of dominant miRNA arm expression, in soybean in different tissues. Two arms of miR166m target different genes related to salinity stress (chloroplastic beta-amylase 1 targeted by miR166m-5p and calcium-dependent protein kinase 1 targeted by miR166m-3p), suggesting arm-switching of miR166m play roles in soybean in response to salinity stress. Furthermore, two pairs of miRNA:lncRNA interacting partners (miR166i-5p and lncRNA Gmax_MSTRG.35921.1; and miR394a-3p and lncRNA Gmax_MSTRG.18616.1) were also discovered in reaction to salinity stress. CONCLUSIONS: This study demonstrates how ncRNA involves in salinity stress responses in soybean by miRNA arm switching and miRNA:lncRNA interactions. The behaviors of ncRNAs revealed in this study will shed new light on molecular regulatory mechanisms of stress responses in plants, and hence provide potential new strategies for crop improvement.

Rethinking Sesquiterpenoids: A Widespread Hormone in Animals.

The sesquiterpenoid hormone juvenile hormone (JH) controls development, reproduction, and metamorphosis in insects, and has long been thought to be confined to the Insecta. While it remains true that juvenile hormone is specifically synthesized in insects, other types or forms of sesquiterpenoids have also been discovered in distantly related animals, such as the jellyfish. Here, we combine the latest literature and annotate the sesquiterpenoid biosynthetic pathway genes in different animal genomes. We hypothesize that the sesquiterpenoid hormonal system is an ancestral system established in an animal ancestor and remains widespread in many animals. Different animal lineages have adapted different enzymatic routes from a common pathway, with cnidarians producing farnesoic acid (FA); non-insect protostomes and non-vertebrate deuterostomes such as cephalochordate and echinoderm synthesizing FA and methyl farnesoate (MF); and insects producing FA, MF, and JH. Our hypothesis revolutionizes the current view on the sesquiterpenoids in the metazoans, and forms a foundation for a re-investigation of the roles of this important and yet neglected type of hormone in different animals.

Micro-RNA Clusters Integrate Evolutionary Constraints on Expression and Target Affinities: The miR-6/5/4/286/3/309 Cluster in Drosophila.

A striking feature of micro-RNAs is that they are often clustered in the genomes of animals. The functional and evolutionary consequences of this clustering remain obscure. Here, we investigated a micro-RNA cluster miR-6/5/4/286/3/309 that is conserved across drosophilid lineages. Small RNA sequencing revealed expression of this micro-RNA cluster in Drosophila melanogaster leg discs, and conditional overexpression of the whole cluster resulted in leg appendage shortening. Transgenic overexpression lines expressing different combinations of micro-RNA cluster members were also constructed. Expression of individual micro-RNAs from the cluster resulted in a normal wild-type phenotype, but either the expression of several ancient micro-RNAs together (miR-5/4/286/3/309) or more recently evolved clustered micro-RNAs (miR-6-1/2/3) can recapitulate the phenotypes generated by the whole-cluster overexpression. Screening of transgenic fly lines revealed downregulation of leg-patterning gene cassettes in generation of the leg-shortening phenotype. Furthermore, cell transfection with different combinations of micro-RNA cluster members revealed a suite of downstream genes targeted by all cluster members, as well as complements of targets that are unique for distinct micro-RNAs. Considered together, the micro-RNA targets and the evolutionary ages of each micro-RNA in the cluster demonstrate the importance of micro-RNA clustering, where new members can reinforce and modify the selection forces on both the cluster regulation and the gene regulatory network of existing micro-RNAs. Key words: micro-RNA, cluster, evolution.

Genome of the four-finger threadfin Eleutheronema tetradactylum (Perciforms: Polynemidae).

BACKGROUND: Teleost fish play important roles in aquatic ecosystems and aquaculture. Threadfins (Perciformes: Polynemidae) show a range of interesting biology, and are of considerable importance for both wild fisheries and aquaculture. Additionally, the four-finger threadfin Eleutheronema tetradactylum is of conservation relevance since its populations are considered to be in rapid decline and it is classified as endangered. However, no genomic resources are currently available for the threadfin family Polynemidae. RESULTS: We sequenced and assembled the first threadfin fish genome, the four-finger threadfin E. tetradactylum. We provide a genome assembly for E. tetradactylum with high contiguity (scaffold N50 = 56.3 kb) and high BUSCO completeness at 96.5%. The assembled genome size of E. tetradactylum is just 610.5 Mb, making it the second smallest perciform genome assembled to date. Just 9.07-10.91% of the genome sequence was found to consist of repetitive elements (standard RepeatMasker analysis vs custom analysis), making this the lowest repeat content identified to date for any perciform fish. A total of 37,683 protein-coding genes were annotated, and we include analyses of developmental transcription factors, including the Hox, ParaHox, and Sox families. MicroRNA genes were also annotated and compared with other chordate lineages, elucidating the gains and losses of chordate microRNAs. CONCLUSIONS: The four-finger threadfin E. tetradactylum genome presented here represents the first available genome sequence for the ecologically, biologically, and commercially important clade of threadfin fish. Our findings provide a useful genomic resource for future research into the interesting biology and evolution of this valuable group of food fish.

Reconstruction of ancient homeobox gene linkages inferred from a new high-quality assembly of the Hong Kong oyster (Magallana hongkongensis) genome.

BACKGROUND: Homeobox-containing genes encode crucial transcription factors involved in animal, plant and fungal development, and changes to homeobox genes have been linked to the evolution of novel body plans and morphologies. In animals, some homeobox genes are clustered together in the genome, either as remnants from ancestral genomic arrangements, or due to coordinated gene regulation. Consequently, analyses of homeobox gene organization across animal phylogeny provide important insights into the evolution of genome organization and developmental gene control, and their interaction. However, homeobox gene organization remains to be fully elucidated in several key animal ancestors, including those of molluscs, lophotrochozoans and bilaterians. RESULTS: Here, we present a high-quality chromosome-level genome assembly of the Hong Kong oyster, Magallana hongkongensis (2n = 20), for which 93.2% of the genomic sequences are contained on 10 pseudomolecules (~ 758 Mb, scaffold N50 = 72.3 Mb). Our genome assembly was scaffolded using Hi-C reads, facilitating a larger scaffold size compared to the recently published M. hongkongensis genome of Peng et al. (Mol Ecol Resources, 2020), which was scaffolded using the Crassostrea gigas assembly. A total of 46,963 predicted gene models (45,308 protein coding genes) were incorporated in our genome, and genome completeness estimated by BUSCO was 94.6%. Homeobox gene linkages were analysed in detail relative to available data for other mollusc lineages. CONCLUSIONS: The analyses performed in this study and the accompanying genome sequence provide important genetic resources for this economically and culturally valuable oyster species, and offer a platform to improve understanding of animal biology and evolution more generally. Transposable element content is comparable to that found in other mollusc species, contrary to the conclusion of another recent analysis. Also, our chromosome-level assembly allows the inference of ancient gene linkages (synteny) for the homeobox-containing genes, even though a number of the homeobox gene clusters, like the Hox/ParaHox clusters, are undergoing dispersal in molluscs such as this oyster.

A crustacean annotated transcriptome (CAT) database.

BACKGROUND: Decapods are an order of crustaceans which includes shrimps, crabs, lobsters and crayfish. They occur worldwide and are of great scientific interest as well as being of ecological and economic importance in fisheries and aquaculture. However, our knowledge of their biology mainly comes from the group which is most closely related to crustaceans - insects. Here we produce a de novo transcriptome database, crustacean annotated transcriptome (CAT) database, spanning multiple tissues and the life stages of seven crustaceans. DESCRIPTION: A total of 71 transcriptome assemblies from six decapod species and a stomatopod species, including the coral shrimp Stenopus hispidus, the cherry shrimp Neocaridina davidi, the redclaw crayfish Cherax quadricarinatus, the spiny lobster Panulirus ornatus, the red king crab Paralithodes camtschaticus, the coconut crab Birgus latro, and the zebra mantis shrimp Lysiosquillina maculata, were generated. Differential gene expression analyses within species were generated as a reference and included in a graphical user interface database at http://cat.sls.cuhk.edu.hk/. Users can carry out gene name searches and also access gene sequences based on a sequence query using the BLAST search function. CONCLUSIONS: The data generated and deposited in this database offers a valuable resource for the further study of these crustaceans, as well as being of use in aquaculture development.

Neuropeptide and microRNA regulators of juvenile hormone production.

The sesquiterpenoid juvenile hormone(s) (JHs) of insects are the primary regulators of growth, metamorphosis, and reproduction in most insect species. As a consequence, it is essential that JH production be precisely regulated so that it is present only during appropriate periods necessary for the control of these processes. The presence of JH at inappropriate times results in disruption to metamorphosis and development and, in some cases, to disturbances in female reproduction. Neuropeptides regulate the timing and production of JH by the corpora allata. Allatostatin and allatotropin were the names coined for neuropeptides that serve as inhibitors or stimulators of JH biosynthesis, respectively. Three different allatostatin neuropeptide families are capable of inhibiting juvenile hormone but only one family is utilized for that purpose dependent on the insect studied. The function of allatotropin also varies in different insects. These neuropeptides are pleiotropic in function acting on diverse physiological processes in different insects such as muscle contraction, sleep and neuromodulation. Genome projects and expression studies have assigned individual neuropeptide families to their respective receptors. An understanding of the localization of these receptors is providing clues as to how numerous peptide families might be integrated in regulating physiological functions. In recent years microRNAs have been identified that down-regulate enzymes and transcription factors that are involved in the biosynthesis and action of juvenile hormone.

Proteomic Analysis of the Venom of Jellyfishes Rhopilema esculentum and Sanderia malayensis.

Venomics, the study of biological venoms, could potentially provide a new source of therapeutic compounds, yet information on the venoms from marine organisms, including cnidarians (sea anemones, corals, and jellyfish), is limited. This study identified the putative toxins of two species of jellyfish-edible jellyfish Rhopilema esculentum Kishinouye, 1891, also known as flame jellyfish, and Amuska jellyfish Sanderia malayensis Goette, 1886. Utilizing nano-flow liquid chromatography tandem mass spectrometry (nLC-MS/MS), 3000 proteins were identified from the nematocysts in each of the above two jellyfish species. Forty and fifty-one putative toxins were identified in R. esculentum and S. malayensis, respectively, which were further classified into eight toxin families according to their predicted functions. Amongst the identified putative toxins, hemostasis-impairing toxins and proteases were found to be the most dominant members (>60%). The present study demonstrates the first proteomes of nematocysts from two jellyfish species with economic and environmental importance, and expands the foundation and understanding of cnidarian toxins.

Terpenes and Terpenoids in Plants: Interactions with Environment and Insects.

The interactions of plants with environment and insects are bi-directional and dynamic. Consequently, a myriad of mechanisms has evolved to engage organisms in different types of interactions. These interactions can be mediated by allelochemicals known as volatile organic compounds (VOCs) which include volatile terpenes (VTs). The emission of VTs provides a way for plants to communicate with the environment, including neighboring plants, beneficiaries (e.g., pollinators, seed dispersers), predators, parasitoids, and herbivores, by sending enticing or deterring signals. Understanding terpenoid distribution, biogenesis, and function provides an opportunity for the design and implementation of effective and efficient environmental calamity and pest management strategies. This review provides an overview of plant-environment and plant-insect interactions in the context of terpenes and terpenoids as important chemical mediators of these abiotic and biotic interactions.

miRNA-Mediated Interactions in and between Plants and Insects.

Our understanding of microRNA (miRNA) regulation of gene expression and protein translation, as a critical area of cellular regulation, has blossomed in the last two decades. Recently, it has become apparent that in plant-insect interactions, both plants and insects use miRNAs to regulate their biological processes, as well as co-opting each others' miRNA systems. In this review article, we discuss the current paradigms of miRNA-mediated cellular regulation and provide examples of plant-insect interactions that utilize this regulation. Lastly, we discuss the potential biotechnological applications of utilizing miRNAs in agriculture.

The first myriapod genome sequence reveals conservative arthropod gene content and genome organisation in the centipede Strigamia maritima.

Myriapods (e.g., centipedes and millipedes) display a simple homonomous body plan relative to other arthropods. All members of the class are terrestrial, but they attained terrestriality independently of insects. Myriapoda is the only arthropod class not represented by a sequenced genome. We present an analysis of the genome of the centipede Strigamia maritima. It retains a compact genome that has undergone less gene loss and shuffling than previously sequenced arthropods, and many orthologues of genes conserved from the bilaterian ancestor that have been lost in insects. Our analysis locates many genes in conserved macro-synteny contexts, and many small-scale examples of gene clustering. We describe several examples where S. maritima shows different solutions from insects to similar problems. The insect olfactory receptor gene family is absent from S. maritima, and olfaction in air is likely effected by expansion of other receptor gene families. For some genes S. maritima has evolved paralogues to generate coding sequence diversity, where insects use alternate splicing. This is most striking for the Dscam gene, which in Drosophila generates more than 100,000 alternate splice forms, but in S. maritima is encoded by over 100 paralogues. We see an intriguing linkage between the absence of any known photosensory proteins in a blind organism and the additional absence of canonical circadian clock genes. The phylogenetic position of myriapods allows us to identify where in arthropod phylogeny several particular molecular mechanisms and traits emerged. For example, we conclude that juvenile hormone signalling evolved with the emergence of the exoskeleton in the arthropods and that RR-1 containing cuticle proteins evolved in the lineage leading to Mandibulata. We also identify when various gene expansions and losses occurred. The genome of S. maritima offers us a unique glimpse into the ancestral arthropod genome, while also displaying many adaptations to its specific life history.

A Burst of miRNA Innovation in the Early Evolution of Butterflies and Moths.

MicroRNAs (miRNAs) are involved in posttranscriptional regulation of gene expression. Because several miRNAs are known to affect the stability or translation of developmental regulatory genes, the origin of novel miRNAs may have contributed to the evolution of developmental processes and morphology. Lepidoptera (butterflies and moths) is a species-rich clade with a well-established phylogeny and abundant genomic resources, thereby representing an ideal system in which to study miRNA evolution. We sequenced small RNA libraries from developmental stages of two divergent lepidopterans, Cameraria ohridella (Horse chestnut Leafminer) and Pararge aegeria (Speckled Wood butterfly), discovering 90 and 81 conserved miRNAs, respectively, and many species-specific miRNA sequences. Mapping miRNAs onto the lepidopteran phylogeny reveals rapid miRNA turnover and an episode of miRNA fixation early in lepidopteran evolution, implying that miRNA acquisition accompanied the early radiation of the Lepidoptera. One lepidopteran-specific miRNA gene, miR-2768, is located within an intron of the homeobox gene invected, involved in insect segmental and wing patterning. We identified cubitus interruptus (ci) as a likely direct target of miR-2768, and validated this suppression using a luciferase assay system. We propose a model by which miR-2768 modulates expression of ci in the segmentation pathway and in patterning of lepidopteran wing primordia.

The phylogenetic utility and functional constraint of microRNA flanking sequences.

MicroRNAs (miRNAs) have recently risen to prominence as novel factors responsible for post-transcriptional regulation of gene expression. miRNA genes have been posited as highly conserved in the clades in which they exist. Consequently, miRNAs have been used as rare genome change characters to estimate phylogeny by tracking their gain and loss. However, their short length (21-23 bp) has limited their perceived utility in sequenced-based phylogenetic inference. Here, using reference taxa with established phylogenetic relationships, we demonstrate that miRNA sequences are of high utility in quantitative, rather than in qualitative, phylogenetic analysis. The clear orthology among miRNA genes from different species makes it straightforward to identify and align these sequences from even fragmentary datasets. We also identify significant sequence conservation in the regions directly flanking miRNA genes, and show that this too is of utility in phylogenetic analysis, as well as highlighting conserved regions that will be of interest to other fields. Employing miRNA sequences from 12 sequenced drosophilid genomes, together with a Tribolium castaneum outgroup, we demonstrate that this approach is robust using Bayesian and maximum-likelihood methods. The utility of these characters is further demonstrated in the rhabditid nematodes and primates. As next-generation sequencing makes it more cost-effective to sequence genomes and small RNA libraries, this methodology provides an alternative data source for phylogenetic analysis. The approach allows rapid resolution of relationships between both closely related and rapidly evolving species, and provides an additional tool for investigation of relationships within the tree of life.

Identification of putative ecdysteroid and juvenile hormone pathway genes in the shrimp Neocaridina denticulata.

Although the sesquiterpenoid juvenile hormone (JH) and the steroidal ecdysteroids are of vital importance to the development and reproduction of insects, our understanding of the evolution of these crucial hormonal regulators in other arthropods is limited. To better understand arthropod hormone evolution and regulation, here we describe the hormonal pathway genes (e.g. those involved in hormone biosynthesis, degradation, regulation and signal transduction) of a new decapod model, the shrimp Neocaridina denticulata. The majority of known insect sesquiterpenoid and ecdysteroid pathway genes and their regulators are contained in the N. denticulata genome. In the sesquiterpenoid pathway, these include biosynthetic pathway components: juvenile hormone acid methyltransferase (JHAMT); hormone binding protein: juvenile hormone binding protein (JHBP); and degradation pathway components: juvenile hormone esterase (JHE), juvenile hormone esterase binding protein (JHEBP) and juvenile hormone epoxide hydrolase (JHEH), with the JHBP, JHEBP and JHEH genes being discovered in a crustacean for the first time here. Ecdysteroid biosynthetic pathway genes identified include spook, phantom, disembodied, shadow and CYP18. Potential hormonal regulators and signal transducers such as allatostatins (ASTs), Methoprene-tolerant (Met), Retinoid X receptor (RXR), Ecdysone receptor (EcR), calponin-like protein Chd64, FK509-binding protein (FKBP39), Broad-complex (Br-c), and crustacean hyperglycemic hormone/molt-inhibiting hormone/gonad-inhibiting hormone (CHH/MIH/GIH) genes are all present in the shrimp N. denticulata. To our knowledge, this is the first report of these hormonal pathways and their regulatory genes together in a single decapod, providing a vital resource for further research into development, reproduction, endocrinology and evolution of crustaceans, and arthropods in general.