Search

Abstract A diplogastrid nematode was isolated from a dung beetle, Onthophagus sp., collected from a rotten mushroom in Kyoto, Japan. The species is characterised by its cheilostomatal shape, separated into 12 narrow plates (rugae), deep stegostom, large ellipsoidal amphids, conical female tail and characteristic receptaculum seminis in the female. Based on its phylogenetic status and stomatal composition, the species is typologically similar to two other diplogastrid genera, Neodiplogaster and Mononchoides. The species can be distinguished from these two genera by the size and shape of the amphid (small pore in Neodiplogaster), female tail shape (long and filiform in Mononchoides) and presence of receptaculum seminis (absence in the two nominal genera), and is described as a monotypic member of a new genus, Onthodiplogaster japonica n. gen., n. sp. Observation of feeding behaviour suggested that O. japonica n. gen., n. sp. does not show clear stomatal dimorphism or polymorphism, which is found in its close relatives, but the species can feed on nematodes (predation), fungi and bacteria. This monomorphic omnivory possibly represents its habitat of dung and other rotten materials, where the environment is biologically divergent, and its condition changes rapidly.

Three Stigmatomyces species were detected on five drosophilid species from Japan. We report Stigmatomyces majewskii on Drosophila rufa and Drosophila suzukii, Stigmatomyces scaptodrosophilae on Scaptodrosophila coracina and Scaptodrosophila subtilis, and Stigmatomyces sacaptomyzae on Scaptomyza graminum. Except for Scaptomyza graminum, each of these species is a newly identified Stigmatomyces host. Our discovery that D. suzukii is a host of S. majewskii may provide new pest management approaches for this global agricultural pest insect.

Abstract 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.

Viviparity, a reproductive form that supplies nutrients to the embryo during gestation, has repeatedly and independently occurred in multiple lineages of animals. During the convergent evolution of viviparity, various modifications of development, structure, and physiology emerged. A new species of nematode, Tokorhabditis tufae, was discovered in the alkaline, hypersaline, and arsenic-rich environment of Mono lake. Its reproductive form is viviparity because it is obligately live-bearing and the embryo increases in size during development. However, the magnitude of the increase in size and nutrient provisioning are unclear. We measured egg and embryo sizes at three developmental stages in T. tufae. Eggs and embryos of T. tufae at the threefold stage were respectively 2.6- and 3.6-fold larger than at the single-cell stage. We then obtained T. tufae embryos at the single-cell, lima bean, and threefold developmental stages and investigated the egg hatching frequency at three different concentrations of egg salt buffer. Removal of embryos from the uterus halted embryonic development at the single-cell and lima bean stages in T. tufae irrespective of the solution used for incubation, indicating the provision of nutrients within the uterus. Ultrastructural and permeability evaluation showed that the permeability barrier did not form during embryonic development, resulting in increased molecular permeability. This high permeability caused by the absence of the permeability barrier likely enables supply of nutrients from the mother. The structural and physiological modifications in T. tufae are like those in other viviparous animals. We conclude that T. tufae is a viviparous rather than an ovoviviparous nematode. T. tufae will facilitate investigation of the evolution of viviparity in animals.

Abstract Life in extreme environments is typically studied as a physiological problem, although the existence of extremophilic animals suggests that developmental and behavioral traits might also be adaptive in such environments. Here, we describe a new species of nematode, Tokorhabditis tufae, n. gen., n. sp., which was discovered from the alkaline, hypersaline, and arsenic-rich locale of Mono Lake, California. The new species, which offers a tractable model for studying animal-specific adaptations to extremophilic life, shows a combination of unusual reproductive and developmental traits. Like the recently described sister group Auanema, the species has a trioecious mating system comprising males, females, and self-fertilizing hermaphrodites. Our description of the new genus thus reveals that the origin of this uncommon reproductive mode is even more ancient than previously assumed, and it presents a new comparator for the study of mating-system transitions. However, unlike Auanema and almost all other known rhabditid nematodes, the new species is obligately live-bearing, with embryos that grow in utero, suggesting maternal provisioning during development. Finally, our isolation of two additional, molecularly distinct strains of the new genus—specifically from non-extreme locales—establishes a comparative system for the study of extremophilic traits in this model.

Abstract 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.

Abstract A survey for the presence of nematodes on the skin of the native Florida manatee, Trichechus manatus latirostris from Crystal River, Florida was conducted during annual manatee health assessments. A putative isolate of Cutidiplogaster manati (Diplogastridae) and two other nematodes belonging to the same family were recovered from mid-dorsal tail skin-scrapings from all sampled winter-collected healthy wild adult manatees during two successive years (2018–2019). Qualitative abundance estimates of these three species of diplogastrid nematodes suggest that an average wild Florida manatee adult might possess between 30,000 and 120,000 nematodes on its tail dorsum and that the entire body dorsum including the tail might possess 160,000–640,000 nematodes in roughly equal ratios. Attempts to culture these nematodes on a variety of different culture media were unsuccessful but examination of the mouth (stomatal) morphology suggests specialized feeding on microbes such as diatoms or predation on other nematodes. No skin lesions were observed during the 2018–2019 samplings suggesting that under normal conditions these nematodes are highly specialized free-living epibionts of the skin that are tightly bound to this niche and horizontally transferred between individual manatees in an analogous fashion to human skin mites (Demodex folliculorum and D. brevis). Molecular phylogenetic inferences using sequences of near full length SSU and D2–D3 expansion segments of LSU rRNA genes revealed a putative new morphospecies in Cutidiplogaster sister to C. manati that was monophyletic with several named Mononchoides species, and another putative new morphospecies that formed a clade with several undescribed species similar in appearance to Mononchoides as well as Tylopharyx, Eudiplogasterium, Paroigolaimella and Sachsia.

Abstract To show the importance of vector switching of nematodes in the evolution of the Bursaphelenchus xylophilus group, we tested a hypothesis that “Bursaphelenchus doui (or its ancestor) was transferred by Acalolepta fraudatrix, Acalolepta sejuncta, and/or Monochamus subfasciatus (or their ancestral species) from broad‐leaved trees to conifers, switched vectors from these cerambycid beetles to Monochamus beetles in conifers, and then evolved into the common ancestor of Bursaphelenchus mucronatus and B. xylophilus.” We used a simple nematode‐loading method to beetles and produced 20 binary combinations of five B. xylophilus group species and four cerambycid beetle species in the tribe Lamiini. The affinity of the nematodes for the beetles was examined based on phoretic stage formation of the nematodes. Phoretic stages of B. doui appeared in all beetle species examined, namely Acalolepta luxuriosa, Psacothea hilaris, A. fraudatrix, and Monochamus alternatus, although the affinity of the nematode for M. alternatus was weak. This finding indicates that B. doui could switch vectors to conifer‐using Monochamus beetles after transfer by A. fraudatrix from broad‐leaved trees to conifers. We conclude that vector switching of nematodes could have potentially happened during the evolutionary history of the B. xylophilus group.

The parasitism of two groups of host-manipulating parasites of hornets was examined in Kyoto, Japan. Vespa mandarinia (661 individuals), V. simillima (303), V. analis (457), V. ducalis (158), V. crabro (57), and V. dybowskii (4) were collected either by bait trap or hand collection with an insect net, and examined for their parasites. An endoparasitic nematode, Sphaerularia vespae was isolated from three overwintered gynes of V. mandarinia and a gyne of V. ducalis. While endoparasitic insects, Xenos spp., were recovered from 13 V. mandarinia, 77 V. analis, two V. ducalis, and three V. crabro, and those recovered from V. analis and others were molecularly identified as X. oxyodontes and X. moutoni, respectively. Comparing Xenos parasitism level and capturing methods, the parasitism level was significantly higher in trapped hosts than in hand-collected ones, suggesting that stylopized hosts are more strongly attracted to the food source (bait trap) compared with unparasitized hosts. The genotypes of S. vespae were identical to each other, and near identical to its type population. While each of the two Xenos spp. showed four mitochondrial DNA haplotypes. A phylogenetic comparison suggested that Xenos haplotypes found in the present study are close to those previously reported from Japan and other Asian countries.

The nematode genus Bursaphelenchus is a highly divergent group. This genus mainly consists of mycophagous entomophilic species, but some species have specialized as obligate or facultative plant parasites, facultative insect parasites, or exhibit feeding dimorphism (phenotypic plasticity) leading to mycophagous and predatory forms. In the present study, a new Bursaphelenchus species, B. suri n. sp. was isolated from fresh syconia (figs) of Ficus sur and is described and illustrated based on its typological characters and molecular phylogenetic status. The new species is characterized by its highly derived feeding structures found in obligate plant parasites, lip possessing a labial disc and a long and thick stylet with a long conus and extremely well-developed basal swellings. In addition, slender body of both sexes is characteristic of the species. The new species is phylogenetically and typologically closely related to B. sycophilus, i.e., these two species share the characteristic feeding structures and form a well-supported clade within the B. fungivorus group in the genus. Biologically, these two species are both isolated from fresh figs of the section Sycomorus. However, the new species differs from B. sycophilus by the length of the female post-uterine sac and the shape of the male spicule, i.e., the new species has a long post-uterine sac and spicule condylus without dorsal recurvature. Thus, the new species is the second obligate fig parasite of the genus, and the evolutionary relationship between the B. suri n. sp. and B. sycophilus clade and section Sycomorus figs is hypothesized as an example of adaptive radiation with more species to be discovered.

A preliminary survey of Seinura spp. was conducted in the Kyoto area, Western Japan. The survey yielded four new strains of Seinura spp., including two strains of S. caverna, a strain of S. italiensis, and a strain of an undescribed species. Molecularly, the two strains of S. caverna were nearly identical to the type strain but showed some minor variations, particularly in the mitochondrial cytochrome oxidase subunit I gene. The small subunit and D2-D3 large subunit sequences of the Japanese strain of S. italiensis were nearly identical and identical to its original description, respectively, and the difference in the small subunit was due to mis-reading of the sequences. The new species, S. shigaensis n. sp., was phylogenetically close to S. caverna and S. persica, although these three species were clearly different phylogenetically. The new species was typologically similar or nearly identical to several other Seinura spp., including S. chertkovi, S. christiei, S. italiensis, S. steineri, and S. tenuicaudata, but it can be distinguished from those species by the morphometric values. Because the new species is phylogenetically very close to S. caverna, it could be a good comparative system for S. caverna as a potential satellite model for the predatory nematode.

Nematodes of figs and fig wasps have received limited attention in Africa since their discovery in 1973. Sixteen of the 25 species of native South African figs were sampled for nematode associates using molecular barcoding with three loci (SSU, LSU D2-D3 and mtCOI) and fourteen (93%) were positive for at least one nematode species. Thirty-three putative species of nematodes were identified and classified according to the loci that were amplified and successfully sequenced. Fourteen putative nematode species were classified as Aphelenchoididae, of which nine were identified as Ficophagus from four species of Ficus from the section Galoglychia (i.e., five ex F. burkei including one shared with F. natalensis, one ex F. glumosa, one ex F. lutea, and one ex F. stuhlmannii) and one species ex F. sur from the section Sycomorus. In addition, there were four nematode species classified as Schistonchus s.s. from section Galoglychia figs (i.e., one ex F. burkei, two ex F. trichopoda, and one ex F. glumosa). There was also one species of Bursaphelenchus nematode recovered from F. sur from the section Sycomorus. Sixteen putative nematode species were classified as Diplogastridae, of which eight occurred in two clades of what is currently called Parasitodiplogaster with one (P. salicifoliae) being recovered from two Ficus species in the section Urostigma (F. salicifolia and F. ingens) and seven diplogastrids being associated with six species of Ficus from the section Galoglychia (i.e., two ex F. burkei including P. sycophilon, one ex F. stuhlmannii, one ex F. burtt-davyi, one ex F. trichopoda, one ex F. abutilifolia and one ex F. sansibarica). Three Acrostichus spp., a Teratodiplogaster and a Pristionchus species were recovered from F. sur and two Teratodiplogaster spp. and Pristionchus sycomori were recovered from F. sycomorus with both Ficus species belonging to the subgenus and section Sycomorus. The identities of the previously described T. martini and Parasitodiplogaster doliostoma (= Pristionchus sp. 35) are discussed. Lastly, there was a panagrolaimid identified from F. petersii.

Caenorhabditis nematodes are important model organisms. Here, the authors report the biology and genome of Caenorhabditis inopinata, a first sibling species of C. elegans, and develop genetic and molecular techniques for C. inopinata.

Nematodes can grow within the inflorescences of many fig trees (Ficus spp., Moraceae); however, the feeding behaviour of most nematodes is not known. Fig pollinating wasps (Hymenoptera: Agaonidae) transfer nematodes into young figs upon the wasps’ entry into the figs to deposit their eggs. Most Asian fig trees, however, are functionally dioecious, and the pollinating wasps that enter female figs are unable to reproduce. They fail to produce the offspring required to carry the new generations of nematodes. We examined whether female figs of F. hispida can nonetheless support the development of phoretic nematode populations. Nematodes were extracted from male and female figs sampled in Sumatra, Indonesia, to compare the growth of their populations within the figs. We found three species of nematodes that grew within figs of male and female trees of F. hispida: Ficophagus cf. centerae (Aphelenchoididae), Martininema baculum (Aphelenchoididae) and Caenorhabditis sp (Rhabditidae). The latter species (Caenorhabditis sp.) has never been reported to be associated with F. hispida before. Nematode populations peaked at around 120–140 individuals in both sexes of figs, at the time when a succeeding generation of adult fig wasps appeared within male figs. The female figs could support the growth and reproduction of the three nematodes species; however, the absence of vectors meant that female figs remained as traps from which there could be no escape.

A new nematode species of the genus Parasitodiplogaster was recovered from syconia of Ficus religiosa at the Guangxiao Temple, Guangzhou, China. It is described herein as P. religiosae n. sp. and is characterised by possessing the longest and thinnest spicule of all currently described males in the genus, an elongated laterally "ε-shaped" and ventrally rhomboid-like gubernaculum, a stoma without teeth, consisting of a ring-like cheilostom with indistinct anteriolateral projections, a tube-like gymnostom and a funnel-like stegostom, monodelphic with a mean vulval position of 66%. There are three pre-cloacal and six post-cloacal male genital papillae with the arrangement P1, P2, P3, (C, P4), P5, P6d, P7, P8, P9d, Ph. This new species was easily differentiated from other members of the genus by DNA sequences of partial small subunit rRNA gene (SSU) and the D2-D3 expansion segments of the large subunit rRNA gene (LSU). Phylogenetic analysis also corroborated its reasonable placement within a well-supported monophyletic clade with other Parasitodiplogaster species and within the australis-group that includes P. australis and P. salicifoliae that are all associates of fig wasp pollinators (Platyscapa sp.) of figs of the subsection Urostigma.

Recent inferences of phylogeny from molecular characters, as well as a reexamination of morphological and biological characters, reject the monophyly of the nematode genus Koerneria Meyl, 1960 (Diplogastridae). Here, Koerneria sensu lato is revised. The genus, which previously consisted of 40 species, is separated into three genera. Almost all of the transferred species are moved to the resurrected genus Allodiplogaster Paramonov & Sobolev in Skrjabin et al. (1954). Koerneria and Allodiplogaster are distinguished from each other by a weakly vs. clearly striated body surface, an undivided vs. divided stomatal cheilostom, and arrangement of the terminal ventral triplet of male genital papillae, namely in that v5 and v6 are paired and separated from v7 vs. v5–v7 being close to each other. Allodiplogaster is further divided into two groups of species, herein called the henrichae and striata groups, based on both morphological and life-history traits. The henrichae group is characterized by papilliform labial sensilla and male genital papillae, a conical tail in both males and females, and an association with terrestrial habitats and insects, whereas the striata group is characterized by setiform labial sensilla and male genital papillae, an elongated conical tail in both sexes, and an association with aquatic habitats. A second genus, Anchidiplogaster Paramonov, 1952, is resurrected to include a single species that is characterized by its miniscule stoma and teeth, unreflexed testis, and a distinct lack of male genital papillae or stomatal apodemes. Lastly, one further species that was previously included in Koerneria sensu lato is transferred to the genus Pristionchus Kreis, 1932. The revision of Koerneria sensu lato is necessitated by the great variability in its subordinate taxa, which occupy a variety of habitats, in addition to the increased attention to Diplogastridae as a model system for comparative mechanistic biology.

Using transmission electron microscopy, we examined the body cuticle ultrastructures of phoretic and parasitic stages of the parasitaphelenchid nematodes Bursaphelenchus xylophilus, B. conicaudatus, B. luxuriosae, B. rainulfi; an unidentified Bursaphelenchus species, and an unidentified Parasitaphelenchus species. Nematode body cuticles usually consist of three zones, a cortical zone, a median zone, and a basal zone. The phoretic stages of Bursaphelenchus spp., isolated from the tracheal systems of longhorn beetles or the elytra of bark beetles, have a thick and radially striated basal zone. In contrast, the parasitic stage of Parasitaphelenchus sp., isolated from bark beetle hemocoel, has no radial striations in the basal zone. This difference probably reflects the peculiar ecological characteristics of the phoretic stage. A well-developed basal radially striated zone, composed of very closely linked proteins, is the zone closest to the body wall muscle. Therefore, the striation is necessary for the phoretic species to be able to seek, enter, and depart from host/carrier insects, but is not essential for internal parasites in parasitaphelenchid nematodes. Phylogenetic relationships inferred from near-full-length small subunit ribosomal RNA sequences suggest that the cuticle structures of parasitic species have apomorphic characters, e.g., lack of striation in the basal zone, concurrent with the evolution of insect parasitism from a phoretic life history.

The pine wood nematode, Bursaphelenchus xylophilus, is the causal agent of pine wilt disease. This nematode has two developmental forms in its life cycle; i.e., the propagative and dispersal forms. The former is the form that builds up its population inside the host pine. The latter is specialized for transport by the vector. This form is separated into two dispersal stages (third and fourth); the third-stage dispersal juvenile (JIII) is specialized for survival under unfavorable conditions, whereas the fourth-stage juvenile (JIV), which is induced by a chemical signal from the carrier Monochamus beetle, is transported to new host pines and invades them. Because of its importance in the disease cycle, molecular and chemical aspects of the JIV have been investigated, while the mechanism of JIII induction has not been sufficiently investigated. In an effort to clarify the JIII induction process, we established inbred lines of B. xylophilus and compared their biological features. We found that the total number of nematodes (propagation proportion) was negatively correlated with the JIII emergence proportion, likely because nematode development was arrested at JIII; i.e., they could not develop to adults via the reproductive stage. In addition, JIII induction seemed to be regulated by a small number of genes because the JIII induction proportion varied among inbred lines despite the high homozygosity of the parental line. We also demonstrated that JIII can be artificially induced by the nematode's secreted substances. This is the first report of artificial induction of JIII in B. xylophilus. The dauer (dispersal) juvenile of the model organism Caenorhabditis elegans corresponds functionally to JIII of B. xylophilus, and this stage is known to be induced by a chemical signal referred to as daumone, derived from the nematodes' secretion. The artificial induction of JIII suggests the presence of daumone-like material in B. xylophilus.

The internal armature of the hindgut of Pericapritermes nitobei was examined under a light microscope. Termites were obtained from Iriomote Island, Okinawa Prefecture, Japan, identified based on the typological characters of soldiers and 12S (mitochondrial) ribosomal RNA sequencing, and dissected to examine hindgut morphology. The hindgut of P. nitobei could be separated into four parts, P1-P4, and P1-P3 had cuticular spines on the inner surface. P1 bore small triangular spines. P2 formed an enteric valve, composed of six finger-shaped enteric pads with triangular spines, and six marginal regions with small dot-like spines. P3 bore three different types of spines: relatively large star-shaped spines; thorn-like small spines, and long curved spines with brush-like tips.

Several genera belonging to the nematode family Diplogastridae show characteristic dimorphism in their feeding structures; specifically, they have microbial feeding stenostomatous and predatory eurystomatous morphs. A diplogastrid satellite model species, Pristionchus pacificus, and its close relatives have become a model system for studying this phenotypic plasticity, with intensive physiological and structural studies having been undertaken. However, the many other species that are morphologically and phylogenetically divergent from P. pacificus have not been examined to date. In the present study, the detailed stomatal structure and induction of dimorphism in Neodiplogaster acaloleptae were examined. N. acaloleptae has a fungal feeding stenostomatous morph and a predatory eurystomatous morph. The predatory morph was induced by starvation, high population density, and co-culturing with its potential prey, Caenorhabditis elegans. The feeding behavior of the stenostomatous and eurystomatous morphs of N. acaloleptae was confirmed, demonstrating that 1) the stomatal and pharyngeal movements of the two morphs were basically identical, and 2) the stomatal elements were protracted to cut open the hyphae and/or prey to feed when a N. acaloleptae flips its dorsal movable tooth dorsally and tilts its subventral stegostomatal cylinder ventrally, forming a pair of scissors to cut the food source. The stoma morphology of N. acaloleptae with a single movable tooth and a long stoma is markedly different from that of Pristionchus, which has two movable teeth and a short stoma. It is, however, similar to that of Mononchoides, tentatively a sister to Neodiplogaster.

Developmental plasticity has been proposed to facilitate phenotypic diversification in plants and animals, but the macroevolutionary potential of plastic traits remains to be objectively tested. We studied the evolution of feeding structures in a group of 90 nematodes, including Caenorhabditis elegans, some species of which have evolved a mouthpart polyphenism, moveable teeth, and predatory feeding. Comparative analyses of shape and form, using geometric morphometrics, and of structural complexity revealed a rapid process of diversification associated with developmental plasticity. First, dimorphism was associated with a sharp increase in complexity and elevated evolutionary rates, represented by a radiation of feeding-forms with structural novelties. Second, the subsequent assimilation of a single phenotype coincided with a decrease in mouthpart complexity but an even stronger increase in evolutionary rates. Our results suggest that a macroevolutionary ‘pulse’ of plasticity promotes novelties and, even after the secondary fixation of phenotypes, permits sustained rapid exploration of morphospace.

A new nematode species, Bursaphelenchus sycophilus n. sp. is described. The species was found in syconia of a fig species, Ficus variegata during a field survey of fig-associated nematodes in Japan. Because it has a well-developed stylet and pharyngeal glands, the species is considered an obligate plant parasite, and is easily distinguished from all other fungal-feeding species in the genus based upon these characters. Although B. sycophilus n. sp. shares an important typological character, male spicule possessing a strongly recurved condylus, with the "B. eremus group" and the "B. leoni group" of the genus, it was inferred to be monophyletic with the "B. fungivorus group". The uniquely shaped stylet and well-developed pharyngeal glands is reminiscent of the fig-floret parasitic but paraphyletic assemblage of "Schistonchus". Thus, these morphological characters appear to be an extreme example of convergent evolution in the nematode family, Aphelenchoididae, inside figs. Other characters shared by the new species and its close relatives, i.e., lack of ventral P1 male genital papilla, female vulval flap, and papilla-shaped P4 genital papillae in males, corroborate the molecular phylogenetic inference. The unique biological character of obligate plant parasitism and highly derived appearance of the ingestive organs of Bursaphelenchus sycophilus n. sp. expands our knowledge of the potential morphological, physiological and developmental plasticity of the genus Bursaphelenchus.

BACKGROUND: The molecular operational taxonomic unit (MOTU) has recently been applied to microbial and microscopic animal biodiversity surveys. However, in many cases, some of the MOTUs cannot be definitively tied to any of the taxonomic groups in current databases. To surmount these limitations, the concept of "reverse taxonomy" has been proposed, i.e. to primarily list the MOTUs with morphological information, and then identify and/or describe them at genus/species level using subsamples or by re-isolating the target organisms. Nevertheless, the application of "reverse taxonomy" has not been sufficiently evaluated. Therefore, the practical applicability of "reverse taxonomy" is tested using termite-associated nematodes as a model system for phoretic/parasitic organisms which have high habitat specificity and a potential handle (their termite host species) for re-isolation attempts. METHODOLOGY: Forty-eight species (from 298 colonies) of termites collected from the American tropics and subtropics were examined for their nematode associates using the reverse taxonomy method and culturing attempts (morphological identification and further sequencing efforts). The survey yielded 51 sequence types ( = MOTUs) belonging to 19 tentatively identified genera. Within these, four were identified based on molecular data with preliminary morphological observation, and an additional seven were identified or characterized from successful culturing, leaving eight genera unidentified. CONCLUSIONS: That 1/3 of the genera were not successfully identified suggests deficiencies in the depth of available sequences in the database and biological characters, i.e. usually isolated as phoretic/parasitic stages which are not available for morphological identification, and too many undiscovered lineages of nematodes. Although there still is the issue of culturability of nematodes, culturing attempts could help to make reverse taxonomy methods more effective. However, expansion of the database, i.e., production of more DNA barcodes tied to biological information by finding and characterizing additional new and known lineages, is necessary for analyzing functional diversity.

Bursaphelenchus xylophilus is the nematode responsible for a devastating epidemic of pine wilt disease in Asia and Europe, and represents a recent, independent origin of plant parasitism in nematodes, ecologically and taxonomically distinct from other nematodes for which genomic data is available. As well as being an important pathogen, the B. xylophilus genome thus provides a unique opportunity to study the evolution and mechanism of plant parasitism. Here, we present a high-quality draft genome sequence from an inbred line of B. xylophilus, and use this to investigate the biological basis of its complex ecology which combines fungal feeding, plant parasitic and insect-associated stages. We focus particularly on putative parasitism genes as well as those linked to other key biological processes and demonstrate that B. xylophilus is well endowed with RNA interference effectors, peptidergic neurotransmitters (including the first description of ins genes in a parasite) stress response and developmental genes and has a contracted set of chemosensory receptors. B. xylophilus has the largest number of digestive proteases known for any nematode and displays expanded families of lysosome pathway genes, ABC transporters and cytochrome P450 pathway genes. This expansion in digestive and detoxification proteins may reflect the unusual diversity in foods it exploits and environments it encounters during its life cycle. In addition, B. xylophilus possesses a unique complement of plant cell wall modifying proteins acquired by horizontal gene transfer, underscoring the impact of this process on the evolution of plant parasitism by nematodes. Together with the lack of proteins homologous to effectors from other plant parasitic nematodes, this confirms the distinctive molecular basis of plant parasitism in the Bursaphelenchus lineage. The genome sequence of B. xylophilus adds to the diversity of genomic data for nematodes, and will be an important resource in understanding the biology of this unusual parasite.