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Large-scale genome-structural evolution is common in various organisms. Recent developments in speciation genomics revealed the importance of inversions, whereas the role of other genome-structural rearrangements, including chromosome fusions, have not been well characterized. We study genomic divergence and reproductive isolation of closely related nematodes: the androdioecious (hermaphroditic) model Pristionchus pacificus and its dioecious sister species Pristionchus exspectatus. A chromosome-level genome assembly of P. exspectatus using single-molecule and Hi-C sequencing revealed a chromosome-wide rearrangement relative to P. pacificus. Strikingly, genomic characterization and cytogenetic studies including outgroup species Pristionchus occultus indicated two independent fusions involving the same chromosome, ChrIR, between these related species. Genetic linkage analysis indicated that these fusions altered the chromosome-wide pattern of recombination, resulting in large low-recombination regions that probably facilitated the coevolution between some of the ~14.8% of genes across the entire genomes. Quantitative trait locus analyses for hybrid sterility in all three sexes revealed that major quantitative trait loci mapped to the fused chromosome ChrIR. While abnormal chromosome segregations of the fused chromosome partially explain hybrid female sterility, hybrid-specific recombination that breaks linkage of genes in the low-recombination region was associated with hybrid male sterility. Thus, recent chromosome fusions repatterned recombination rate and drove reproductive isolation during Pristionchus speciation.

Nematodes such as Caenorhabditis elegans and Pristionchus pacificus are extremely successful model organisms for comparative biology. Several studies have shown that phenotypic novelty but also conserved processes are controlled by taxon-restricted genes. To trace back the evolution of such new or rapidly evolving genes, a robust phylogenomic framework is indispensable. Here, we present an improved version of the genome of Parapristionchus giblindavisi which is the only known member of the sister group of Pristionchus. Relative to the previous short-read assembly, the new genome is based on long reads and displays higher levels of contiguity, completeness, and correctness. Specifically, the number of contigs dropped from over 7,303 to 735 resulting in an N50 increase from 112 to 791 kb. We made use of the new genome to revisit the evolution of multiple gene families. This revealed Pristionchus-specific expansions of several environmentally responsive gene families and a Pristionchus-specific loss of the de novo purine biosynthesis pathway. Focusing on the evolution of sulfatases and sulfotransferases, which control the mouth form plasticity in P. pacificus, reveals differences in copy number and genomic configurations between the genera Pristionchus and Parapristionchus. Altogether, this demonstrates the utility of the P. giblindavisi genome to date and polarizes lineage-specific patterns.

Although the microbiota is known to affect host development, metabolism, and immunity, its impact on host behavior is only beginning to be understood. In order to better characterize behavior modulation by host-associated microorganisms, we investigated how bacteria modulate complex behaviors in the nematode model organism Pristionchus pacificus. This nematode is a predator that feeds on the larvae of other nematodes, including Caenorhabditis elegans. By growing P. pacificus on different bacteria and testing their ability to kill C. elegans, we reveal large differences in killing efficiencies, with a Novosphingobium species showing the strongest enhancement. This enhanced killing was not accompanied by an increase in feeding, which is a phenomenon known as surplus killing, whereby predators kill more prey than necessary for sustenance. Our RNA-seq data demonstrate widespread metabolic rewiring upon exposure to Novosphingobium, which facilitated screening of bacterial mutants with altered transcriptional responses. We identified bacterial production of vitamin B12 as an important cause of such enhanced predatory behavior. Although vitamin B12 is an essential cofactor for detoxification and metabolite biosynthesis, shown previously to accelerate development in C. elegans, supplementation with this enzyme cofactor amplified surplus killing in P. pacificus, whereas mutants in vitamin B12-dependent pathways reduced surplus killing. By demonstrating that production of vitamin B12 by host-associated microbiota can affect complex host behaviors, we reveal new connections between animal diet, microbiota, and nervous system.

The model organism Pristionchus pacificus and the genus Pristionchus, Kreis, 1932 have been intensively studied in the last decade with contemporary work focusing on the development, evolution, ecology, behavior, neurobiology, and genomics of this group of organisms. In particular, mechanistic studies on the development and evolution of mouth-form plasticity, predation and associated self-recognition processes enabled unique insight into life history strategies and the evolution of novelty. These studies include a comparative research agenda making use of the 39 available species of Pristionchus, all of which can be studied in living cultures. Sampling efforts revealed that Asia represents a biodiversity hotspot for Pristionchus worms. However, previous samplings have a bias towards northern and island areas, largely for logistic reasons. Here, we report on two extensive sampling trips to the Yunnan and Shaanxi provinces in Mainland China. We report the isolation of nine new Pristionchus species by morphology, morphometrics, mating experiments and genome-wide sequence analysis.

During aging, proteostasis capacity declines and aggregation-prone proteins become instable, accumulating in protein aggregates both inside and outside cells. Both in disease and during aging, proteins selectively aggregate in certain tissues and not others. Yet, tissue-specific regulation of protein aggregation remains poorly understood. Surprisingly, we found that the inhibition of three core protein quality control systems, i.e. chaperones, proteasome and macroautophagy, leads to lower levels of age-dependent protein aggregation in C. elegans pharyngeal muscles, but higher levels in body-wall muscles. We describe a novel safety mechanism called SAPA that selectively targets newly synthesized aggregation-prone proteins to suppress aggregation and proteotoxicity. vha-12 and scav-3 mutants reveal that SAPA relies on macroautophagy-independent lysosomal degradation. Furthermore, SAPA involves several previously uncharacterized components of the intracellular pathogen response. We propose that SAPA represents an anti-aggregation machinery targeting aggregation-prone proteins for lysosomal degradation.

Animals are associated with a diverse bacterial community that impacts host physiology. It is well known that nutrients and enzymes synthesized by bacteria largely expand host metabolic capacity. Bacteria also impact a wide range of animal physiology that solely depends on host genetics through direct interaction. However, studying the synergistic effects of the bacterial community remains challenging due to its complexity. The omnivorous nematode Pristionchus pacificus has limited digestive efficiency on bacteria. Therefore, we established a bacterial collection that represents the natural gut microbiota that are resistant to digestion. Using this collection, we show that the bacterium Lysinibacillus xylanilyticus by itself provides limited nutritional value, but in combination with Escherichia coli, it significantly promotes life-history traits of P. pacificus by regulating the neuroendocrine peptide in sensory neurons. This gut-to-brain communication depends on undigested L. xylanilyticus providing Pristionchus nematodes a specific fitness advantage to compete with nematodes that rupture bacteria efficiently. Using RNA-seq and CRISPR-induced mutants, we show that 1-h exposure to L. xylanilyticus is sufficient to stimulate the expression of daf-7-type TGF-β signaling ligands, which induce a global transcriptome change. In addition, several effects of L. xylanilyticus depend on TGF-β signaling, including olfaction, body size regulation, and a switch of energy allocation from lipid storage to reproduction. Our results reveal the beneficial effects of a gut bacterium to modify life-history traits and maximize nematode survival in natural habitats.

The evolution of island systems is characterized by processes that result in extreme morphological diversity, high endemism and high extinction rates. These dynamics can make phylogenetic reconstruction difficult, i.e. the extinct flightless Dodo from Mauritius was assigned to the family of doves only through DNA analysis of subfossils. Many insect species on islands have gone extinct through habitat loss, and face similar challenges to decipher their evolutionary history, however historical specimens have not yet been harnessed for phylogenomic reconstructions. Here, we employed historical museum specimens from the Mascarene Islands to generate the first whole-genome based phylogeny of three presumably extinct species of the rhinoceros beetle genusOryctes. We compared their genomes with those of an extantOryctesspecies from the island of Réunion, as well as a flightless Réunion-based species previously placed into the supposedly unrelated genusMarronus. We found thatMarronus borbonicusbelongs instead to the genusOryctesand that the two Réunion-based species (O. borbonicusandM. borbonicus) are not sister taxa, suggesting two independent colonizations. The divergence time between them (M. borbonicusbecame flightlessin situ. Our study showcases the power of genomes from insect museum specimens to address evolutionary questions in light of increasing extinction rates.

In metazoans, the secreted proteome participates in intercellular signalling and innate immunity, and builds the extracellular matrix scaffold around cells. Compared with the relatively constant intracellular environment, conditions for proteins in the extracellular space are harsher, and low concentrations of ATP prevent the activity of intracellular components of the protein quality-control machinery. Until now, only a few bona fide extracellular chaperones and proteases have been shown to limit the aggregation of extracellular proteins1–5. Here we performed a systematic analysis of the extracellular proteostasis network in Caenorhabditis elegans with an RNA interference screen that targets genes that encode the secreted proteome. We discovered 57 regulators of extracellular protein aggregation, including several proteins related to innate immunity. Because intracellular proteostasis is upregulated in response to pathogens6–9, we investigated whether pathogens also stimulate extracellular proteostasis. Using a pore-forming toxin to mimic a pathogenic attack, we found that C. elegans responded by increasing the expression of components of extracellular proteostasis and by limiting aggregation of extracellular proteins. The activation of extracellular proteostasis was dependent on stress-activated MAP kinase signalling. Notably, the overexpression of components of extracellular proteostasis delayed ageing and rendered worms resistant to intoxication. We propose that enhanced extracellular proteostasis contributes to systemic host defence by maintaining a functional secreted proteome and avoiding proteotoxicity. A systematic analysis of the proteostasis network of secreted proteins in Caenorhabditis elegans identifies numerous regulators of protein homeostasis outside the cell, and highlights the contribution of extracellular proteostasis to host defence.

Although the microbiota is known to affect host development, metabolism, and immunity, its impact on host behavior is only beginning to be understood. In order to better characterize behavior modulation by host-associated microorganisms, we investigated how bacteria modulate complex behaviors in the nematode model organism Pristionchus pacificus. This nematode is a predator that feeds on the larvae of other nematodes, including Caenorhabditis elegans. By growing P. pacificus on different bacteria and testing their ability to kill C. elegans, we reveal large differences in killing efficiencies, with a Novosphingobium species showing the strongest enhancement. This enhanced killing was not accompanied by an increase in feeding, which is a phenomenon known as surplus killing, whereby predators kill more prey than necessary for sustenance. Our RNA-seq data demonstrate widespread metabolic rewiring upon exposure to Novosphingobium, which facilitated screening of bacterial mutants with altered transcriptional responses. We identified bacterial production of vitamin B

Phenotypic plasticity has been suggested to act through developmental switches, but little is known about associated molecular mechanisms. In the nematode Pristionchus pacificus, the sulfatase eud-1 was identified as part of a developmental switch controlling mouth-form plasticity governing a predatory versus bacteriovorous mouth-form decision. Here we show that mutations in the conserved histone-acetyltransferase Ppa-lsy-12 and the methyl-binding-protein Ppa-mbd-2 mimic the eud-1 phenotype, resulting in the absence of one mouth-form. Mutations in both genes cause histone modification defects and reduced eud-1 expression. Surprisingly, Ppa-lsy-12 mutants also result in the down-regulation of an antisense-eud-1 RNA. eud-1 and antisense-eud-1 are co-expressed and further experiments suggest that antisense-eud-1 acts through eud-1 itself. Indeed, overexpression of the antisense-eud-1 RNA increases the eud-1-sensitive mouth-form and extends eud-1 expression. In contrast, this effect is absent in eud-1 mutants indicating that antisense-eud-1 positively regulates eud-1. Thus, chromatin remodelling and antisense-mediated up-regulation of eud-1 control feeding plasticity in Pristionchus., In the nematode Pristionchus pacificus, a developmental switch, the sulfatase eud-1, controls mouth-form plasticity. Here, the authors show that mutations in two conserved histone modifying enzymes mimic the eud-1 phenotype, in part mediated by an antisense eud-1 RNA, resulting in the absence of one mouth-form

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Summary Mutation and recombination are main drivers of phenotypic diversity, but the ability to create new allelic combinations is strongly dependent on the mode of reproduction. While most animals are dioecious (i.e., separated male and female sexes), in a number of evolutionary lineages females have gained the ability to self-fertilize [ 1 , 2 ], with drastic consequences on effective recombination rate, genetic diversity, and the efficacy of selection [ 3 ]. In the genus Caenorhabditis, such hermaphroditic or androdioecious lineages, including C. briggsae and C. tropicalis, display a genome shrinkage relative to their dioecious sister species C. nigoni and C. brenneri, respectively [ 4 , 5 ]. However, common consequences of reproductive modes on nematode genomes remain unknown, because most taxa contain single or few androdioecious species. One exception is the genus Pristionchus, with seven androdioecious species. Pristionchus worms are found in association with scarab beetles in worldwide samplings, resulting in deep taxon sampling and currently 39 culturable and available species. Here, we use phylotranscriptomics of all 39 Pristionchus species to provide a robust phylogeny based on an alignment of more than 2,000 orthologous clusters, which indicates that the seven androdioecious species represent six independent lineages. We show that gene loss is more prevalent in all hermaphroditic lineages than in dioecious relatives and that the majority of lost genes evolved recently in the Pristionchus genus. Further, we provide evidence that genes with male-biased expression are preferentially lost in hermaphroditic lineages. This supports a contribution of adaptive gene loss to shaping nematode genomes following the evolution of hermaphroditism.

Pristionchus pacificus Sommer, Carta, Kim, and Sternberg, 1996 is an important model organism in evolutionary biology that integrates developmental biology with ecology and population genetics. This species is part of a sub-complex of the genus Pristionchus that is considered to have originated in East Asia. Here, we describe two new species of Pristionchus, P. maxplancki and P. quartusdecimus, which were isolated from beetles in Japan, supporting the hypothesis that a region including Japan is the origin of diversification of the P. pacificus species complex. Phytogeny inferred from a partial small subunit rRNA gene and 25 ribosomal protein genes shows P. maxplancki to be the closest known outgroup to a triad of sibling species, including P. pacificus. Pristionchus quartusdecimus is a putative outgroup to the P. pacificus species complex, supporting a more ancient origin of Pristionchus species in the region. Species diagnoses are based on morphological and molecular characters, in addition to reproductive isolation for P. maxplancki. Members of the P. pacificus species complex as well as P. quartusdecimus are distinguished by stegostomatal structures, male genital papilla arrangement, and gubernaculum shape. The discovery of a new member of the P. pacificus species complex allows greater precision in polarizing and reconstructing ancestral states in the comparative model system centering on P. pacificus. Together with previous reports, these findings support an important biogeographic role of Japan in the evolution of the genus Pristionchus and the P. pacificus species complex, especially the associated phenotypic evolution of mouth morphology.

Developmental plasticity is often correlated with diversity and has been proposed as a facilitator of phenotypic novelty. Yet how a dimorphism arises or how additional morphs are added is not understood, and few systems provide experimental insight into the evolution of polyphenisms. Because plasticity correlates with structural diversity in Pristionchus nematodes, studies in this group can test the role of plasticity in facilitating novelty. Here, we describe three new species, Pristionchus fukushimae sp. nov., Pristionchus hoplostomus sp. nov., and the hermaphroditic Pristionchus triformis sp. nov., which are characterized by a novel polymorphism in their mouthparts. In addition to showing the canonical mouth dimorphism of diplogastrid nematodes, comprising a stenostomatous (‘narrow-mouthed’) and a eurystomatous (‘wide-mouthed’) form, the new species exhibit forms with six, 12, or intermediate numbers of cheilostomatal plates. Correlated with this polymorphism is another trait that varies among species: whereas divisions between plates are complete in P. triformis sp. nov., which is biased towards a novel ‘megastomatous’ form comprising 12 complete plates, the homologous divisions in the other new species are partial and of variable length. In a reconstruction of character evolution, a phylogeny inferred from 26 ribosomal protein genes and a partial small subunit rRNA gene supported the megastomatous form of P. triformis sp. nov. as the derived end of a series of split-plate forms. Although split-plate forms were normally only observed in eurystomatous nematodes, a single 12-plated stenostomatous individual of P. hoplostomus sp. nov. was also observed, suggesting independence of the two types of mouth plasticity. By introducing these new species to the Pristionchus model system, this study provides further insight into the evolution of polymorphisms and their evolutionary intermediates. © 2013 The Linnean Society of London

Summary Switching between alternative complex phenotypes is often regulated by "supergenes," polymorphic clusters of linked genes such as in butterfly mimicry. In contrast, phenotypic plasticity results in alternative complex phenotypes controlled by environmental influences rather than polymorphisms. Here, we show that the developmental switch gene regulating predatory versus non-predatory mouth-form plasticity in the nematode Pristionchus pacificus is part of a multi-gene locus containing two sulfatases and two α-N-acetylglucosaminidases ( nag ). We provide functional characterization of all four genes, using CRISPR-Cas9-based reverse genetics, and show that nag genes and the previously identified eud-1 /sulfatase have opposing influences. Members of the multi-gene locus show non-overlapping neuronal expression and epistatic relationships. The locus architecture is conserved in the entire genus Pristionchus . Interestingly, divergence between paralogs is counteracted by gene conversion, as inferred from phylogenies and genotypes of CRISPR-Cas9-induced mutants. Thus, we found that physical linkage accompanies regulatory linkage between switch genes controlling plasticity in P. pacificus .

A polyphenism discovered to consist of five discrete morphotypes shows that extreme diversification is possible within single species., Diversification is commonly understood to be the divergence of phenotypes accompanying that of lineages. In contrast, alternative phenotypes arising from a single genotype are almost exclusively limited to dimorphism in nature. We report a remarkable case of macroevolutionary-scale diversification without genetic divergence. Upon colonizing the island-like microecosystem of individual figs, symbiotic nematodes of the genus Pristionchus accumulated a polyphenism with up to five discrete adult morphotypes per species. By integrating laboratory and field experiments with extensive genotyping of individuals, including the analysis of 49 genomes from a single species, we show that rapid filling of potential ecological niches is possible without diversifying selection on genotypes. This uncoupling of morphological diversification and speciation in fig-associated nematodes has resulted from a remarkable expansion of discontinuous developmental plasticity.

Summary Background Nematode vulva formation provides a paradigm to study the evolution of pattern formation and cell-fate specification. The Caenorhabditis elegans vulva is generated from three of six equipotent cells that form the so-called vulva equivalence group. During evolution, the size of the vulva equivalence group has changed: Panagrellus redivivus has eight, C. elegans six, and Pristionchus pacificus only three cells that are competent to form vulval tissue. In P. pacificus, programmed cell death of individual vulval precursor cells alters the size of the vulva equivalence group. Results We have identified the genes controlling this cell-death event and the molecular mechanism of the reduction of the vulva equivalence group. Mutations in Ppa-hairy , a gene that is unknown from C. elegans , result in the survival of two precursor cells, which expands the vulva equivalence group. Mutations in Ppa-groucho cause a similar phenotype. Ppa -HAIRY and Ppa -GROUCHO form a molecular module that represses the Hox gene Ppa-lin-39 and thereby reduces the size of the vulva equivalence group. The C. elegans genome does not encode a similar hairy -like gene, and no typical HAIRY/GROUCHO module exists. Conclusions We conclude that the vulva equivalence group in Pristionchus is patterned by a HAIRY/GROUCHO module, which is absent in Caenorhabditis . Thus, changes in the number, structure, and function of nematode hairy -like transcription factors are involved in the evolutionary alteration of this equivalence group.