Diversity and evolution of transposable elements in the plant-parasitic nematodes.

BACKGROUND: Transposable elements (TEs) are mobile DNA sequences that propagate within genomes, occupying a significant portion of eukaryotic genomes and serving as a source of genetic variation and innovation. TEs can impact genome dynamics through their repetitive nature and mobility. Nematodes are incredibly versatile organisms, capable of thriving in a wide range of environments. The plant-parasitic nematodes are able to infect nearly all vascular plants, leading to significant crop losses and management expenses worldwide. It is worth noting that plant parasitism has evolved independently at least three times within this nematode group. Furthermore, the genome size of plant-parasitic nematodes can vary substantially, spanning from 41.5 Mbp to 235 Mbp. To investigate genome size variation and evolution in plant-parasitic nematodes, TE composition, diversity, and evolution were analysed in 26 plant-parasitic nematodes from 9 distinct genera in Clade IV. RESULTS: Interestingly, despite certain species lacking specific types of DNA transposons or retrotransposon superfamilies, they still exhibit a diverse range of TE content. Identification of species-specific TE repertoire in nematode genomes provides a deeper understanding of genome evolution in plant-parasitic nematodes. An intriguing observation is that plant-parasitic nematodes possess extensive DNA transposons and retrotransposon insertions, including recent sightings of LTR/Gypsy and LTR/Pao superfamilies. Among them, the Gypsy superfamilies were found to encode Aspartic proteases in the plant-parasitic nematodes. CONCLUSIONS: The study of the transposable element (TE) composition in plant-parasitic nematodes has yielded insightful discoveries. The findings revealed that certain species exhibit lineage-specific variations in their TE makeup. Discovering the species-specific TE repertoire in nematode genomes is a crucial element in understanding the evolution of genomes in plant-parasitic nematodes. It allows us to gain a deeper insight into the intricate workings of these organisms and their genetic makeup. With this knowledge, we are gaining a fundamental piece in the puzzle of understanding the evolution of these parasites. Moreover, recent transpositions have led to the acquisition of new TE superfamilies, especially Gypsy and Pao retrotransposons, further expanding the diversity of TEs in these nematodes. Significantly, the widely distributed Gypsy superfamily possesses proteases that are exclusively associated with parasitism during nematode-host interactions. These discoveries provide a deeper understanding of the TE landscape within plant-parasitic nematodes.

Evolution of parasitism genes in the plant parasitic nematodes.

The plant-parasitic nematodes are considered as one of the most destructive pests, from which the migratory and sedentary endoparasitic plant parasitic nematodes infect more than 4000 plant species and cause over $100 billion crop losses annually worldwide. These nematodes use multiple strategies to infect their host and to establish a successful parasitism inside the host such as cell-wall degradation enzymes, inhibition of host defense proteins, and molecular mimicry. In the present study, the main parasitism-associated gene families were identified and compared between the migratory and sedentary endoparasitic nematodes. The results showed that the migratory and sedentary endoparasitic nematodes share a core conserved parasitism mechanism established throughout the evolution of parasitism. However, genes involved in pectin degradation and hydrolase activity are rapidly evolving in the migratory endoparasitic nematodes. Additionally, cell-wall degrading enzymes such as GH45 cellulases and pectate lyase and peptidase and peptidase inhibitors were expanded in the migratory endoparasitic nematodes. The molecular mimicry mechanism was another key finding that differs between the endoparasitic and sedentary parasitic nematodes. The PL22 gene family, which is believed to play a significant role in the molecular mechanisms of nematode parasitism, has been found to be present exclusively in migratory endoparasitic nematodes. Phylogenetic analysis has suggested that it was de novo born in these nematodes. This discovery sheds new light on the molecular evolution of these parasites and has significant implications for our understanding of their biology and pathogenicity. This study contributes to our understanding of core parasitism mechanisms conserved throughout the nematodes and provides unique clues on the evolution of parasitism and the direction shaped by the host.

Differentially Expressed Genes Associated with Body Size Changes and Transposable Element Insertions between Caenorhabditis elegans and Its Sister Species, Caenorhabditis inopinata.

Why the recently discovered nematode Caenorhabditis inopinata differs so greatly from its sibling species Caenorhabditis elegans remains unknown. A previous study showed that C. inopinata has more transposable elements (TEs), sequences that replicate and move autonomously throughout the genome, potentially altering the expression of neighboring genes. In this study, we focused on how the body size of this species has evolved and whether TEs could affect the expression of genes related to species-specific traits such as body size. First, we compared gene expression levels between C. inopinata and C. elegans in the L4 larval and young adult stages-when growth rates differ most prominently between these species-to identify candidate genes contributing to their differences. The results showed that the expression levels of collagen genes were consistently higher in C. inopinata than in C. elegans and that some genes related to cell size were differentially expressed between the species. Then, we examined whether genes with TE insertions are differentially expressed between species. Indeed, the genes featuring C. inopinata-specific TE insertions had higher expression levels in C. inopinata than in C. elegans. These upregulated genes included those related to body size, suggesting that these genes could be candidates for artificial TE insertion to examine the role of TEs in the body size evolution of C. inopinata.

The compact genome of Caenorhabditis niphades n. sp., isolated from a wood-boring weevil, Niphades variegatus.

BACKGROUND: The first metazoan genome sequenced, that of Caenorhabditis elegans, has motivated animal genome evolution studies. To date > 50 species from the genus Caenorhabditis have been sequenced, allowing research on genome variation. RESULTS: In the present study, we describe a new gonochoristic species, Caenorhabditis niphades n. sp., previously referred as C. sp. 36, isolated from adult weevils (Niphades variegatus), with whom they appear to be tightly associated during its life cycle. Along with a species description, we sequenced the genome of C. niphades n. sp. and produced a chromosome-level assembly. A genome comparison highlighted that C. niphades n. sp. has the smallest genome (59 Mbp) so far sequenced in the Elegans supergroup, despite being closely related to a species with an exceptionally large genome, C. japonica. CONCLUSIONS: The compact genome of C. niphades n. sp. can serve as a key resource for comparative evolutionary studies of genome and gene number expansions in Caenorhabditis species.

piRNA-like small RNAs target transposable elements in a Clade IV parasitic nematode.

The small RNA (sRNA) pathways identified in the model organism Caenorhabditis elegans are not widely conserved across nematodes. For example, the PIWI pathway and PIWI-interacting RNAs (piRNAs) are involved in regulating and silencing transposable elements (TE) in most animals but have been lost in nematodes outside of the C. elegans group (Clade V), and little is known about how nematodes regulate TEs in the absence of the PIWI pathway. Here, we investigated the role of sRNAs in the Clade IV parasitic nematode Strongyloides ratti by comparing two genetically identical adult stages (the parasitic female and free-living female). We identified putative small-interfering RNAs, microRNAs and tRNA-derived sRNA fragments that are differentially expressed between the two adult stages. Two classes of sRNAs were predicted to regulate TE activity including (i) a parasite-associated class of 21-22 nt long sRNAs with a 5' uridine (21-22Us) and a 5' monophosphate, and (ii) 27 nt long sRNAs with a 5' guanine/adenine (27GAs) and a 5' modification. The 21-22Us show striking resemblance to the 21U PIWI-interacting RNAs found in C. elegans, including an AT rich upstream sequence, overlapping loci and physical clustering in the genome. Overall, we have shown that an alternative class of sRNAs compensate for the loss of piRNAs and regulate TE activity in nematodes outside of Clade V.

Possible stochastic sex determination in Bursaphelenchus nematodes.

Sex determination mechanisms evolve surprisingly rapidly, yet little is known in the large nematode phylum other than for Caenorhabditis elegans, which relies on chromosomal XX-XO sex determination and a dosage compensation mechanism. Here we analyze by sex-specific genome sequencing and genetic analysis sex determination in two fungal feeding/plant-parasitic Bursaphelenchus nematodes and find that their sex differentiation is more likely triggered by random, epigenetic regulation than by more well-known mechanisms of chromosomal or environmental sex determination. There is no detectable difference in male and female chromosomes, nor any linkage to sexual phenotype. Moreover, the protein sets of these nematodes lack genes involved in X chromosome dosage counting or compensation. By contrast, our genetic screen for sex differentiation mutants identifies a Bursaphelenchus ortholog of tra-1, the major output of the C. elegans sex determination cascade. Nematode sex determination pathways might have evolved by "bottom-up" accretion from the most downstream regulator, tra-1.

Genome of the fatal tapeworm Sparganum proliferum uncovers mechanisms for cryptic life cycle and aberrant larval proliferation.

The cryptic parasite Sparganum proliferum proliferates in humans and invades tissues and organs. Only scattered cases have been reported, but S. proliferum infection is always fatal. However, S. proliferum's phylogeny and life cycle remain enigmatic. To investigate the phylogenetic relationships between S. proliferum and other cestode species, and to examine the mechanisms underlying pathogenicity, we sequenced the entire genomes of S. proliferum and a closely related non-life-threatening tapeworm Spirometra erinaceieuropaei. Additionally, we performed larvae transcriptome analyses of S. proliferum plerocercoid to identify genes involved in asexual reproduction in the host. The genome sequences confirmed that the S. proliferum has experienced a clearly distinct evolutionary history from S. erinaceieuropaei. Moreover, we found that nonordinal extracellular matrix coordination allows asexual reproduction in the host, and loss of sexual maturity in S. proliferum are responsible for its fatal pathogenicity to humans. Our high-quality reference genome sequences should be valuable for future studies of pseudophyllidean tapeworm biology and parasitism.

Additional description and genome analyses of Caenorhabditis auriculariae representing the basal lineage of genus Caenorhabditis.

Caenorhabditis auriculariae, which was morphologically described in 1999, was re-isolated from a Platydema mushroom-associated beetle. Based on the re-isolated materials, some morphological characteristics were re-examined and ascribed to the species. In addition, to clarify phylogenetic relationships with other Caenorhabditis species and biological features of the nematode, the whole genome was sequenced and assembled into 109.5 Mb with 16,279 predicted protein-coding genes. Molecular phylogenetic analyses based on ribosomal RNA and 269 single-copy genes revealed the species is closely related to C. sonorae and C. monodelphis placing them at the most basal clade of the genus. C. auriculariae has morphological characteristics clearly differed from those two species and harbours a number of species-specific gene families, indicating its usefulness as a new outgroup species for Caenorhabditis evolutionary studies. A comparison of carbohydrate-active enzyme (CAZy) repertoires in genomes, which we found useful to speculate about the lifestyle of Caenorhabditis nematodes, suggested that C. auriculariae likely has a life-cycle with tight-association with insects.

Nearly Complete Genome Assembly of the Pinewood Nematode Bursaphelenchus xylophilus Strain Ka4C1.

Bursaphelenchus xylophilus has been destroying pine forests in East Asia and western Europe. Here, we report its nearly complete genomic sequence containing five ∼12-Mb scaffolds and one ∼15-Mb scaffold representing six chromosomes. Large repeat regions that were previously unidentified are now reasonably integrated, particularly in the ∼15-Mb scaffold.

Telomere-to-Telomere Genome Assembly of Bursaphelenchus okinawaensis Strain SH1.

Bursaphelenchus okinawaensis is a self-fertilizing, hermaphroditic, fungus-feeding nematode used as a laboratory model for the genus Bursaphelenchus, which includes the important pathogen Bursaphelenchus xylophilus Here, we report the nearly complete genome sequence of B. okinawaensis The 70-Mbp assembly contained six scaffolds (>11 Mbp each) with telomere repeats on their ends, indicating complete chromosomes.

Reduced Genome of the Gut Symbiotic Bacterium "Candidatus Benitsuchiphilus tojoi" Provides Insight Into Its Possible Roles in Ecology and Adaptation of the Host Insect.

Diverse animals, including insects, harbor microbial symbionts within their gut, body cavity, or cells. The subsocial parastrachiid stinkbug Parastrachia japonensis is well-known for its peculiar ecological and behavioral traits, including its prolonged non-feeding diapause period and maternal care of eggs/nymphs in an underground nest. P. japonensis harbors a specific bacterial symbiont within the gut cavity extracellularly, which is vertically inherited through maternal excretion of symbiont-containing white mucus. Thus far, biological roles of the symbiont in the host lifecycle has been little understood. Here we sequenced the genome of the uncultivable gut symbiont "Candidatus Benitsuchiphilus tojoi." The symbiont has an 804 kb circular chromosome encoding 606 proteins and a 14.5 kb plasmid encoding 13 proteins. Phylogenetic analysis indicated that the bacterium is closely related to other obligate insect symbionts belonging to the Gammaproteobacteria, including Buchnera of aphids and Blochmannia of ants, and the most closely related to Ishikawaella, an extracellular gut symbiont of plataspid stinkbugs. These data suggested that the symbiont genome has evolved like highly reduced gamma-proteobacterial symbiont genomes reported from a variety of insects. The presence of genes involved in biosynthesis pathways for amino acids, vitamins, and cofactors in the genome implicated the symbiont as a nutritional mutualist, supplementing essential nutrients to the host. Interestingly, the symbiont's plasmid encoded genes for thiamine and carotenoid synthesis pathways, suggesting the possibility of additional functions of the symbiont for protecting the host against oxidative stress and DNA damage. Finally, possible involvement of the symbiont in uric acid metabolism during diapause is discussed.

First record of Aphelenchoides stammeri (Nematoda: Aphelenchoididae) from Turkey.

In previous surveys for the investigation of Bursaphelenchus xylophilus in Turkey, several Bursaphelenchus species, except B. xylophilus, were detected. To determine the insect vectors of these previously reported Bursaphelenchus species, trap trees of each pine species (Pinus brutia and P. nigra) were cut and kept in the field from April to September in 2011. Aphelenchoides stammeri was isolated from wood chip samples of P. nigra from Isparta (Sütçüler). A. stammeri was first identified by using morphological features and allometric criteria of females and males. In addition, molecular analysis using sequences of 18S and 28S ribosomal (rRNA) genes of A. stammeri was performed. This is the first report of A. stammeri from Turkey.In previous surveys for the investigation of Bursaphelenchus xylophilus in Turkey, several Bursaphelenchus species, except B. xylophilus, were detected. To determine the insect vectors of these previously reported Bursaphelenchus species, trap trees of each pine species (Pinus brutia and P. nigra) were cut and kept in the field from April to September in 2011. Aphelenchoides stammeri was isolated from wood chip samples of P. nigra from Isparta (Sütçüler). A. stammeri was first identified by using morphological features and allometric criteria of females and males. In addition, molecular analysis using sequences of 18S and 28S ribosomal (rRNA) genes of A. stammeri was performed. This is the first report of A. stammeri from Turkey.

Stage-specific transcriptome of Bursaphelenchus xylophilus reveals temporal regulation of effector genes and roles of the dauer-like stages in the lifecycle.

The pine wood nematode Bursaphelenchus xylophilus is the causal agent of pine wilt disease, one of the most devastating forest diseases in East Asian and West European countries. The lifecycle of B. xylophilus includes four propagative larval stages and gonochoristic adults which are involved in the pathogenicity, and two stages of dispersal larvae involved in the spread of the disease. To elucidate the ecological roles of each developmental stage in the pathogenic life cycle, we performed a comprehensive transcriptome analysis using RNA-seq generated from all developmental stages of B. xylophilus and compared transcriptomes between stages. We found more than 9000 genes are differentially expressed in at least one stage of the life cycle including genes involved in general nematode biology such as reproduction and moulting but also effector genes likely to be involved in parasitism. The dispersal-stage transcriptome revealed its analogy to C. elegans dauer and the distinct roles of the two larval stages from each other regarding survival and transmission. This study provides important insights and resources to understand B. xylophilus parasitic biology.

Assessment of the behaviour and survival of nematodes under low oxygen concentrations.

Oxygen is required for the completion of almost all known metazoan lifecycles, but many metazoans harbour abilities to withstand varying degrees and periods of hypoxia. Caenorhabditis elegans, one of the most popular model organism is extensively used as a model for the study of hypoxia and anoxia biology and it has been found that this nematode is capable of tolerance to varying degrees of hypoxia. Considering the extremely high diversity of nematodes, the effects of low oxygen concentration and mechanisms of adaptation to oxygen depletion differ among species. In this study, we used a simple assay to examine anoxia tolerance in four nematode species, including three free-living and one plant parasitic nematode. We found that the plant parasitic nematode Bursaphelenchus xylophilus can survive more than 14 days under anoxic conditions. Comparisons of behaviour during anoxia induction and the repertoire of oxygen sensation genes among the tested species suggested the existence of different oxygen sensation systems between B. xylophilus and C. elegans, which quickly introduce suspended animation in response to oxygen depletion to survive long-term anoxia.