Introduction
The genus Cranifera Kloss, Reference Kloss1960 belongs to a group of arthropod pinworms (superfamily Thelastomatoidea), a heterogeneous assemblage of Clade 3 nematodes found in the digestive system of diplopods, cockroaches, mole crickets, adult hydrophilid and passalid beetles, larvae of scarab beetles and crane flies (Carreno Reference Carreno2014). Its type species was originally described under the name Leidynema cranifera Chitwood, Reference Chitwood1932 from the hindgut of the cockroaches Blaberus craniifer Burmeister, 1838 and B. atropos (Stoll, 1813) from Florida, USA (Chitwood Reference Chitwood1932), and was later transferred into a genus of its own based on the absence of a diverticulum in the anterior portion of the intestine (Kloss Reference Kloss1960). Females of Cranifera are characterised by the presence of a short first cephalic annule, unarmed cervical cuticle, corpus with a cylindrical anterior half and a sub-cylindrical, wider posterior half, and a didelphic genital tract (Adamson and Van Waerebeke Reference Adamson and Van Waerebeke1992; Morffe et al. Reference Morffe, García, Véliz, Hasegawa and Carreno2022). Males possess a spine-like caudal appendage, arising dorsally; three pairs of copulatory papillae arranged as one pre-cloacal and two post-cloacal (one adanal pair and another near the base of the tail appendage) and a single spicule (Chitwood Reference Chitwood1932). Besides the type species of the genus, three additional species have been described: (a) C. chitwoodi Kloss, Reference Kloss1966 from the cockroach Hormetica scrobiculata Burmeister, 1838 from Brazil, (b) C. mexicana Coy & García, Reference Coy and García1995 from the death’s head cockroach B. craniifer from Mexico, and (c) C. robustum Camino & Achinelly, Reference Camino and Achinelly2012 from the larvae of an Argentinean scarab Cyclocephala signaticollis Burmeister, 1847 from Argentina (Camino and Achinelly Reference Camino and Achinelly2012; Coy and García Reference Coy and García1995; Kloss Reference Kloss1966).
Since Chitwood’s (Reference Chitwood1932) description, C. cranifera has been recorded in several wild and laboratory-reared blaberid cockroaches from Europe and the Americas, namely captive colonies of death’s head cockroach B. craniifer and the dubia roach Blaptica dubia Serville, 1838 from Germany (Leibersperger Reference Leibersperger1960), laboratory colonies of B. dubia from Russia (Spiridonov and Guzeeva Reference Spiridonov and Guzeeva2009), as well as wild populations of the peppered cockroach Archimandrita tessellata Rehn, 1903 from Costa Rica (Carreno and Tuhela Reference Carreno and Tuhela2011), and the discoid cockroach Blaberus discoidalis Serville, 1838 from Cuba (Morffe et al. Reference Morffe, García, Véliz, Hasegawa and Carreno2022). Males of C. cranifera were found and described only by Chitwood (Reference Chitwood1932) and Leibersperger (Reference Leibersperger1960); however, some taxonomically valuable features of male morphology were not sufficiently described. In this publication a population of C. cranifera from laboratory-reared colonies of the dubia roach was used to provide a detailed redescription of the males of the species as well as to generate the nuclear and mitochondrial genomes using long-read sequencing data. We use this opportunity to propose a new term ‘morphogenomic species description’, in order to highlight/distinguish papers combining full morphology-based species descriptions together with detailed description of the genome of a species (already published examples include Pettrich et al. Reference Pettrich, Suwanngam, Guiglielmoni, Ledoux, Villegas, Treonis, Stevens, Kieninger, Paulini, Blaxter, Waldvogel, Holovachov and Schiffer2026; Slos et al. Reference Slos, Sudhaus, Stevens, Bert and Blaxter2017; Stevens et al. Reference Stevens, Félix, Beltran, Braendle, Caurcel, Fausett, Fitch, Frézal, Gosse, Kaur, Kiontke, Newton, Noble, Richaud, Rockman, Sudhaus and Blaxter2019) and contrast them with: (1) traditional morphology-based (sometimes but not always accompanied by molecular data represented by few loci) species descriptions; (2) genome notes focusing solely on descriptions of genomes of known species and providing the briefest morphological data in the shape of voucher pictures, if any at all (examples include Röseler et al. Reference Röseler, Yoshida and Rödelsperger2025; https://www.darwintreeoflife.org/genomes/genome-notes/); and (3) genome-based species descriptions that include minimal or partial morphological information, often relegated to supplementary data (Devi et al. Reference Devi, Haryoso, Rais, Karuniawan, Yahya, Richaud, Wang, Rockman, Tarno and Félix2025; Dulovic et al. Reference Dulovic, Renahan, Röseler, Rödelsperger, Rose and Streit2020; Launay et al. Reference Launay, Félix, Dieng and Delattre2020).
Material and methods
Processing of the hosts and nematodes
Specimens of Blaptica dubia Serville, 1838 (Blattodea: Blaberidae) from a captive colony were killed by decapitation and immediately dissected using longitudinal incisions in both abdominal pleural membranes. Intestines were withdrawn from the body and dissected in dishes with 0.9% NaCl physiological solution. Nematodes for morphological studies were killed with hot 0.9% NaCl (70°C) and fixed in 4% phosphate-buffered formalin. For light microscopy, the nematodes were transferred to anhydrous glycerin via the slow evaporation method (Seinhorst Reference Seinhorst1959) and mounted on permanent slides in glycerin with paraffin wax as support for the coverslip. The edges of the coverslips were sealed with nail polish. Several specimens were pooled for genomic studies and directly fixed in 1× DNA/RNA Shield™ (Zymo Research, California, USA) and stored at −20°C until DNA extraction.
Morphological and morphometric studies
Nematodes were measured with the aid of a drawing tube attached to a Nikon Eclipse Ni microscope (Nikon, Tokyo, Japan). All curved structures were measured along the curved median line. De Man’s indices a, b, c, and V% were calculated. Variables are shown as the range followed by the mean plus standard deviation in parentheses, and the number of measurements is also given when it is different from the total number of measured specimens. Micrographs were generated with a Sony ɑ6400 digital camera attached to a Nikon Eclipse 80i compound microscope (Nikon, Tokyo, Japan). Line drawings were made with the aid of a drawing tube. Scale bars of all figures are given in micrometers. In total, 19 females (SMNH 231508–231516) and 5 males (SMNH 231517–231518) were examined for morphology.
Scanning Electron Microscopy studies
Nematodes were post-fixed overnight with 2% glutaraldehyde in 0.1 M phosphate buffer (pH 7.0). They were dehydrated through a graded ethanol series (30%, 50%, 70%, 90%, 95%, 100% × 2, 30 min in each); transferred to hexamethyldisilazane (HMDS) through a graded series of 25%, 50%, and 75% HMDS in 100% ethanol (30 min in each); washed in pure HMDS for 30 min; and finally were kept overnight in a fume hood until total evaporation of the HMDS. Nematodes were then mounted on double-sided conductive adhesive carbon tabs on a stub, sputter-coated with gold using an AGB7234 high-resolution sputter coater (Agar Scientific, Rotherham, UK), and observed with a Quanta FEG 650 scanning electron microscope (Thermo Fisher Scientific, MA, USA) at 10 kV accelerating voltage.
Genome sequencing
Nematodes preserved in Zymo DNA/RNA Shield solution were disrupted manually with a pestle while frozen and high-molecular-weight DNA was extracted using the Zymo Quick-DNA HMW MagBead kit (Zymo Research, California, USA) following the manufacturer’s protocol. Due to the low concentration of extracted DNA, an ultra-low-input (ULI) library was prepared with the SMRTbell prep 3.0 kit (https://www.pacb.com/wp-content/uploads/Procedure-checklist-Amplifying-genomic-DNA-for-SMRTbell-library-preparation-and-HiFi-sequencing.pdf, Pacific Biosciences, CA, USA). Whole genome amplification was performed with the Universal Adapter System (Twist Bioscience, CA, USA) and the KOD Xtreme™ Hot Start DNA polymerase (Merck, Germany). The library underwent quality control (Fragment Analyzer, Agilent, CA, USA) and size selection (Pippin HT, Sage Science, MA, USA) in order to quantify and eliminate shorter DNA fragments, with the final target fragment size of 10–20 kilobases. The library was pooled and sequenced on a Revio system (Pacific Biosciences, CA, USA). PacBio-generated BAM files were converted into FASTQ format using samtools (Danecek et al. Reference Danecek, Bonfield, Liddle, Marshall, Ohan, Pollard, Whitwham, Keane, McCarthy, Davies and Li2021). usegalaxy.eu (The Galaxy Community 2026) installation of fastplong (Chen Reference Chen2023) was used to remove sequencing adapters, trim 5′ and 3′ sequence ends, and remove sequences with average phred quality less than 20. usegalaxy.eu (The Galaxy Community 2026) installation of NanoPlot (De Coster and Rademakers Reference De Coster and Rademakers2023) was used to assess the quality of the filtered raw read dataset.
Nuclear genome assembly
Draft genomes were assembled with Flye (Kolmogorov et al. Reference Kolmogorov, Yuan, Lin and Pevzner2019), Hifiasm (Cheng et al. Reference Cheng, Asri, Lucas, Koren and Li2024), and PECAT (Nie et al. Reference Nie, Ni, Huang, Zhang, Wang, Xiao, Luo and Wang2024) using usegalaxy.eu (The Galaxy Community 2026). Assembly mode (--pacbio-corr, --pacbio-raw, --pacbio-hifi), assembly type (regular or metagenomic), and coverage for initial assembly were varied in case of assembly with Flye. For Hifiasm, genome size estimation was left blank or pre-set to 100 Mb, while for PECAT both PacBio sequencing platforms (hifi and clr) were tested. Quantitative completeness of draft assemblies was assessed using BUSCO v.6.0.0 comparison against the nematoda_odb12 reference dataset consisting of 596 universal single-copy orthologous genes (Tegenfeldt et al. Reference Tegenfeldt, Kuznetsov, Manni, Berkeley, Zdobnov and Kriventseva2025). barrnap (Galaxy version 1.2.2) on the usegalaxy.eu server (equivalent to standalone version 0.9, https://github.com/tseemann/barrnap) was used to mine both assemblies for Eukaryotic rRNA in order to confirm the identification of a target species and to check for presence of contaminants, in particular for the host DNA. Two draft assemblies were selected for genome reconciliation: standard assembly generated with Flye using ‘–pacbio-raw’ mode under offered highest N50, while metagenomic assembly generated with Flye using –pacbio-hifi mode had the highest BUSCO completeness, although the differences were minor. Completeness and contamination level of both draft assemblies were further evaluated using BlobToolKit (Challis et al. Reference Challis, Richards, Rajan, Cochrane and Blaxter2020; https://www.sanger.ac.uk/tool/blobtoolkit/), and contigs identified as ‘chordata’ and ‘arthropoda’, as well as all non-metazoan contigs, were removed. quickmerge (Chakraborty et al. Reference Chakraborty, Baldwin-Brown, Long and Emerson2016) was used to reconcile both assemblies using either one as a query or a reference. Haplotigs (variants of the same gene originating from different alleles in the population) were purged from both reconciled assemblies using redundans (Pryszcz and Gabaldón Reference Pryszcz and Gabaldón2016), since purge_dups (Guan et al. Reference Guan, McCarthy, Wood, Howe, Wang and Durbin2020) was not able to handle population-level heterozygosity. Purged assemblies were again evaluated with BUSCO v.6.0.0 by comparing them against the four most current reference datasets: nematoda_odb12 (596 single copy-orthologs), nematoda_odb12.1 (1477 single-copy orthologs), chromadorea_odb12 (1126 single-copy orthologs), and chromadorea_odb12.1 (2034 single-copy orthologs) (Tegenfeldt et al. Reference Tegenfeldt, Kuznetsov, Manni, Berkeley, Zdobnov and Kriventseva2025) and the final chosen assembly was visualised using BlobToolKit (Challis et al. Reference Challis, Richards, Rajan, Cochrane and Blaxter2020). Annotation of protein-coding genes was not performed due to the absence of RNAseq data.
Mitochondrial genome assembly
A first attempt to assemble a mitochondrial genome was done with MitoHifi pipeline (Allio et al. Reference Allio, Schomaker-Bastos, Romiguier, Prosdocimi, Nabholz and Delsuc2020; Laslett and Canbäck Reference Laslett and Canbäck2004; Uliano-Silva et al. Reference Uliano-Silva, Ferreira, Krasheninnikova, Formenti, Abueg, Torrance, Myers, Durbin, Blaxter and McCarthy2023) using the mitochondrial genome of Hammerschmidtiella sp. (KY399989.1) as a reference and two alternative Hifiasm-based assemblies as input. This produced two consensus contigs, 24645 and 35654 bases long. The discrepancies in length were caused by mis-assembly and not by collapsed/uncollapsed repetitive regions, requiring the use of the following alternative approach. Nucleotide sequences of nematode mitochondrial genes were downloaded from the https://github.com/WormsEtAl/Nematode-Mitochondrial-Database (Gendron et al. Reference Gendron, Sevigny, Byiringiro, Thomas, Powers and Porazinska2023) and converted into a BLAST-compatible nucleotide mitochondrial reference database with the usegalaxy.eu (The Galaxy Community 2026) installation of an NCBI BLAST+ makeblastdb (Altschul et al. Reference Altschul, Gish, Miller, Myers and Lipman1990). Amino acid sequences of protein-coding genes were extracted from the mitochondrial genomes of 52 nematode species across the entire phylum downloaded from the NCBI GenBank, and converted into a Diamond-compatible amino-acid mitochondrial reference database with the usegalaxy.eu (The Galaxy Community 2026) installation of Diamond makedb (Buchfink et al. Reference Buchfink, Reuter and Drost2021). blastn (Altschul et al. Reference Altschul, Gish, Miller, Myers and Lipman1990) with an E-value cutoff of 0.001 was used to search the filtered raw reads against the nucleotide mitochondrial reference database, while Diamond blastx (Buchfink et al. Reference Buchfink, Reuter and Drost2021) set to the invertebrate mitochondrial code and E-value cutoff 0.001 was used to search the filtered raw reads against the amino acid mitochondrial reference database. BBedit text editor was used to remove duplicate hits and create a list of read sequence headers for seqkit (Shen et al. Reference Shen, Sipos and Zhao2024), which was used to pull relevant reads from the filtered raw reads file. Three input datasets, one based on blastn hits, one based on diamond blastx hits, and one using combined hits, were used as an input for the mitogenome assembly with usegalaxy.eu (The Galaxy Community 2026) installation of Canu (Koren et al. Reference Koren, Walenz, Berlin, Miller, Bergman and Phillippy2017). Multiple assemblies were generated using the following constant settings: technology = PacBio; maximum corrected overlap mismatch = 0.2; target coverage for corrected reads = 10; low coverage depth = 5; estimated genome size = 25k. Four combinations of minimum read length/minimum overlap were used in parallel: 1000/500, 2000/1000, 3000/1500, and 4000/2000. Mitos2 (Bernt et al. Reference Bernt, Donath, Jühling, Externbrink, Florentz, Fritzsch, Pütz, Middendorf and Stadler2013) was used to extract COX1 from several randomly selected assemblies with the following settings: genetic code = Invertebrate (5), reference data = refseq63m. This COX1 sequence was then identified using NCBI blastn against GenBank core nucleotide database (excluding uncultured and environmental sample sequences). The same COX1 sequence was used to positively identify remaining assembled contigs as belonging to the same target species, orient them in the same direction, and align and circularise using AliView (Larsson Reference Larsson2014). Draft consensus assembly was created by combining all aligned contigs (circularised and partial) using a majority-rule approach. minimap2 (Li Reference Li2021) was used to map raw reads against draft assemblies using map-hifi setting while pilon (Walker et al. Reference Walker, Abeel, Shea, Priest, Abouelliel, Sakthikumar, Cuomo, Zeng, Wortman, Young and Earl2014) was used to improve draft consensus assembly based on the minimap2 output. Tandem Repeats Finder (Benson Reference Benson1999) and Claude AI by Anthropic were both used to identify repetitive regions and assembly artefacts in the final assembly. usegalaxy.eu (The Galaxy Community 2026) installations of Mitos2 (Bernt et al. Reference Bernt, Donath, Jühling, Externbrink, Florentz, Fritzsch, Pütz, Middendorf and Stadler2013) and Aragorn (Laslett and Canbäck Reference Laslett and Canbäck2004) were used to annotate final mitochondrial genome assembly, with additional manual checks aided by Claude AI by Anthropic using the following genomes as references: Aspiculuris tetraptera (KT764937), Enterobius vermicularis (NC056632), Hammerschmidtiella sp. (KY399989), Oxyuris equi (NC027190), Passalurus ambiguus (NC028345), Ruizia karukerae (MF509850 and MF509851), Rhigonema thysanophora (NC024020), Spauligodon atlanticus (PX608399), Syphacia muris (AP017697), S. obvelata (KT900946), Travassosinema sp. (PX431499 and PX431500), Wellcomia compar (MW059037), and W. siamensis (NC016129). samtools (Danecek et al. Reference Danecek, Bonfield, Liddle, Marshall, Ohan, Pollard, Whitwham, Keane, McCarthy, Davies and Li2021) was used to estimate the read coverage from the minimap2 output while a custom script (written by Claude AI by Anthropic on 24 May 2026) was used to normalise read depth values to get scores between 0 and 1. Proksee (Grant et al. Reference Grant, Enns, Marinier, Mandal, Herman, Chen, Graham, Van Domselaar and Stothard2023) was used to visualise the final assembly.
Intragenomic variation of rRNA
barrnap (Galaxy version 1.2.2) on the usegalaxy.eu server (equivalent to standalone version 0.9, https://github.com/tseemann/barrnap) was used to mine ribosomal RNA genes from the final assembly. Several 28S rRNA and 18S rRNA sequences (partial fragments) were retrieved from GenBank and used in comparative/phylogenetic analysis. Severianoia blapticola Guzeeva, 2009 and Travassosinema viatorum Morffe, García & Hasegawa, 2023 were included as outgroups for the 28S rRNA and 18S rRNA analyses, respectively. The former species was selected for the 28S rRNA analysis since it is the closest sister group of Cranifera, despite having been considered a species inquirenda by Malysheva et al. (Reference Malysheva, Shmatko and Spiridonov2020). Multiple sequence alignments were made using MUSCLE (Edgar Reference Edgar2004) with the default parameters as implemented in MEGA12 (Kumar et al. Reference Kumar, Stecher, Suleski, Sanderford, Sharma and Tamura2024). MEGA12 was also used to identify the optimal model of evolution for the data set (GTR+I for the 28S rRNA and GTR+G+I for the 18S rRNA datasets) following the corrected Akaike Information Criterion (AICc) and to construct phylogenetic trees based on the Maximum Likelihood (ML) method. Nodal support was inferred by bootstrap analysis using 1,000 iterations. Pairwise distances were calculated for the 28S rRNA and 18S rRNA sequences of C. cranifera with MEGA12 using the Kimura 2-parameter model (Kimura Reference Kimura1980).
Use of artificial intelligence tools
Claude AI by Anthropic was used to search and summarise published reference data; it was not used for writing the manuscript or for data analysis except for cases specifically described above.
Results
Additional data on male morphology
Body smaller and more slender than that of females (Table 1), slightly robust and fusiform, widening from the anterior end with maximum width at ca. the first third of the body, then narrowing towards cloaca (Figure 1C). Anterior end of body truncated, posterior end ventrally curved, blunt with a dorsally located tail appendage. Lateral alae well-developed from the level of the last third of the isthmus to a very short distance (ca. 2 μm) before the level of the first pre-cloacal pair of papilliform sensilla (Figure 2D). Cephalic cap ca. 10 μm in diameter, 3 μm in length, truncated, with smooth cuticle (Figure 2E). Cervical cuticle unarmed, presenting annuli from base of cephalic cap that extend towards posterior end of body, ending ventrally and laterally at level of the pre-cloacal pair of papilliform sensilla, then continuing dorsally to the level of the second post-cloacal pair of papilliform sensilla. The annuli from the cervical region are fine and at level of the isthmus become gradually wider towards the posterior end of body. Annuli of the last two-thirds of the body ornamented with tubercles that form a single row across each annulus (Figure 2C). Tubercles more prominent towards the posterior part of body. Mouth surrounded by a fine, wrinkled cuticular cordon followed by a loose, veliform labial ring anchored bilaterally to the cuticle via a frenulum-like structure (Figure 2F). Amphids lateral, slit-like, located in prominences of the labial ring (Figure 2F). Buccal capsule very short. Oesophagus consisting of a comparatively short, muscular, clavate corpus, well set off from the long isthmus (Figure 1A). Anterior part of isthmus wider at the joint with the base of corpus then its diameter decreases towards the spherical basal bulb. Valve-plate of basal bulb well-developed. Intestine simple, sub-rectilinear. Nerve ring located at level of posterior third of isthmus. Excretory pore ventral, located at ca. 1.5–2 corresponding body-widths posterior to the basal bulb (Figure 1A). Monorchic. Testis ventral, reflexed just posterior to the excretory pore, flexure ca. one corresponding body-width long. Drop-like spermatids, ca. 8 μm length. Cloaca located in a prominent cloacal cone. Spicule single, sclerotized, almost straight, shaft spindle-shaped, capitulum rounded, ventrally bent, and distal tip sharp (Figure 1D). Eight copulatory papilliform sensilla present, two pre-cloacal and six post-cloacal (Figure 1B, Figure 2D). Pre-cloacal pair sub-ventral, consisting of large, prominent, mammiform papillae, located at ca. 10 μm anterior to the level of cloaca. First pair of post-cloacal papillae smaller, sub-ventral, ad-cloacal. Second pair of post-cloacal papillae large and mammiform, sub-dorsal, near the base of the tail appendage. Third pair of post-cloacal papillae ventral, consisting of minute papillae located at the level of proximal fifth of the tail appendage.
Comparison of the morphometrics of Cranifera cranifera (Chitwood, Reference Chitwood1932) from captive-reared Blaptica dubia Serville, 1838 from Sweden with previous records of the same species. All the measurements are in the form of the range followed by mean plus standard deviation in parentheses; the number of measurements is also given if different from the total number of studied individuals
Table 1. Long description
The table compares morphometric data for Cranifera cranifera from five localities.
Header Information:
* Sweden: Host is captive colony of Blaptica dubia; Authors: Current study; Columns for females (n=18) and males (n=5).
* Florida, USA: Host is wild populations of Blaberus craniifer and B. atropos; Author: Chitwood Reference Chitwood1932; Columns for females and males.
* Germany: Host is captive colony of Blaptica dubia; Author: Leibersperger Reference Leibersperger1960; Columns for females and males.
* Costa Rica: Host is wild populations of Archimandrita tessellata; Authors: Carreno and Tuhela 2003; Column for females (n=25).
* Cuba: Host is wild populations of Blaberus discoidalis; Authors: Morffe et al. 2022; Column for females (n=15).
Key Morphometric Data (Sweden Current Study):
* Total length (mm): Females 2.9 to 4.2 (3.8 plus or minus 0.3); Males 0.87 to 0.96 (0.91 plus or minus 0.03).
* Maximum width (mu m): Females 200 to 338 (289 plus or minus 41); Males 64 to 76 (69 plus or minus 5).
* Oesophagus length (mu m): Females 404 to 462 (431 plus or minus 16); Males 107 to 117 (113 plus or minus 4).
* Tail length (mu m): Females 547 to 880 (725 plus or minus 79); Males 38 to 44 (41 plus or minus 3).
* Spicule length (mu m): Males 25 to 30 (28 plus or minus 2).
Other characters listed include de Man's indices (a, b, c), V percent, first cephalic annule height and width, stoma length, corpus length, isthmus length, basal bulb diameter, nerve ring-anterior end, excretory pore-anterior end, vulva-anterior end, and egg dimensions. Data is presented as range followed by mean plus or minus standard deviation.
Line drawings of a male of Cranifera cranifera (Chitwood, Reference Chitwood1932). A: Anterior body end showing pharyngeal region and excretory pore (ventral side to the left); B: posterior body end showing cloaca, spicule, genital sensilla, and tail (ventral side to the left); C: entire male (ventral side to the left); D: spicule (ventral side to the right).
Figure 1. Long description
A set of four line drawings labeled A through D.
Panel A shows the anterior body end. It features a long, narrow pharyngeal region with a distinct bulbous structure at the base. A scale bar indicates 50 micrometers. The excretory pore is visible on the left side.
Panel B provides a close-up of the posterior body end. It displays the cloaca, a pointed spicule, and genital sensilla. The tail tapers into a sharp point. A scale bar indicates 10 micrometers.
Panel C depicts the entire male nematode in a curved posture. The internal organs, including the pharynx and reproductive system, are visible through the body wall. A scale bar indicates 100 micrometers.
Panel D shows a high-magnification view of a single spicule, which is curved with a hooked proximal end and a sharp distal tip. A scale bar indicates 10 micrometers.
SEM micrographs of a male of Cranifera cranifera (Chitwood, Reference Chitwood1932). A: Entire male (ventral side to the bottom); B: anterior body end showing beginning of lateral alae (ventral side to the left); C: posterior region showing the end of lateral alae and the ornamentations of the cuticle (ventral side to the bottom); D: posterior body end showing cloaca, end of lateral alae, genital sensilla, and tail (ventral side to the left); E: labial region; F: labial region, en face view. Scale bars: A = 100 μm. B, D = 20 μm. C, E = 5 μm. F = 3 μm.
Figure 2. Long description
A collection of six S E M micrographs labeled A through F.
* Panel A: A wide view of the entire male nematode, curved in a C-shape with the ventral side at the bottom. A vertical scale bar at the top right represents 100 micrometers.
* Panel B: A close-up of the anterior end showing the start of the lateral alae on the left side. Scale bar is 20 micrometers.
* Panel C: A high-magnification view of the posterior region cuticle, showing block-like ornamentations and the termination of the lateral alae. Scale bar is 5 micrometers.
* Panel D: The posterior body end featuring the cloaca, genital sensilla, and a thin, curved tail. Scale bar is 20 micrometers.
* Panel E: A lateral view of the labial region showing thick, segmented cuticular folds. Scale bar is 5 micrometers.
* Panel F: An en face view of the labial region, looking directly into the circular oral opening surrounded by textured labial structures. Scale bar is 3 micrometers.
Males of the present population are longer (0.873–0.956 mm vs. 0.790 mm vs. 0.790 mm) and wider (64–76 μm vs. 40 μm vs. 40 μm) than those reported by Chitwood (Reference Chitwood1932) and Leibersperger (Reference Leibersperger1960), respectively (Table 1). Their tail is also longer (38–44 μm vs. 35 μm vs. 35 μm). Compared with the record of Leibersperger (Reference Leibersperger1960), the body of the present male specimens is more robust (a = 11.9–14.3 vs. 19.8) and the relative length of the oesophagus is shorter (b = 7.7–8.5 vs. 6.9). Again, these differences can be equally attributed to both intraspecific variability of distant populations and measuring errors.
Notes on female morphology
Females from the currently studied population (Figure 3) are morphologically identical to the specimens from the original description (Chitwood Reference Chitwood1932). Most of the morphometrics also overlap (Table 1), and only the oesophagus length is slightly shorter in the present specimens (404–462 μm vs. 470–540 μm). The morphology and measurements of the individuals from the population described by Leibersperger (Reference Leibersperger1960) from B. dubia are also similar, except for the length of the eggs, which are slightly shorter in current specimens (62–72 μm vs. 75–80 μm). The excretory pore is more anterior in the specimens recorded by Carreno and Tuhela (Reference Carreno and Tuhela2011) from Costa Rican A. tessellata (396–500 μm vs. 620–800 μm). The body of the specimens found in B. discoidalis from Cuba (Morffe et al. Reference Morffe, García, Véliz, Hasegawa and Carreno2022) is slightly less robust (a = 15.0–20.5 vs. 11.5–14.8). All these differences can be equally attributed to both intraspecific variability of distant populations and measurement errors.
Light micrographs of a female (A–D) and a male (E) of Cranifera cranifera (Chitwood, Reference Chitwood1932). A: Anterior body end showing pharyngeal region (ventral side to the right); B: tail (ventral side to the left); C: intra-uterine egg; D: labial region; E: caudal region (ventral side to the left), white arrows point to genital sensilla pairs, black arrow points to cloaca. Scale bars: A, B = 200 μm. C, D, E = 25 μm.
Figure 3. Long description
The micrograph consists of five panels labeled A through E.
* Panel A: A vertical view of the anterior end of a female. The pharyngeal region is visible with the ventral side oriented to the right. A white scale bar in the top right represents 200 micrometers.
* Panel B: The long, slender tail of a female with the ventral side to the left. A white scale bar in the top right represents 200 micrometers.
* Panel C: A high-magnification view of an oval-shaped intra-uterine egg with a granular internal texture. A white scale bar in the top right represents 25 micrometers.
* Panel D: A close-up of the female labial region showing the rounded head structure and striated cuticle. A white scale bar in the top right represents 25 micrometers.
* Panel E: The caudal region of a male with the ventral side to the left. A black arrow points to the cloaca. Four white arrows point to pairs of genital sensilla located along the posterior end and the base of the spicule-like tail. A white scale bar in the top right represents 25 micrometers.
Nuclear genome assembly
The final decontaminated genome assembly has a total length of 242 Mb, consisting of 7562 contigs, with an N50 = 43 kb and the longest contig being 1.10 Mb (Table 2). BUSCO v.6.0.0 (Tegenfeldt et al. Reference Tegenfeldt, Kuznetsov, Manni, Berkeley, Zdobnov and Kriventseva2025) analysis using four latest reference datasets: nematoda_odb12 (596 single copy-orthologs), nematoda_odb12.1 (1477 single-copy orthologs), chromadorea_odb12 (1126 single-copy orthologs), and chromadorea_odb12.1 (2034 single-copy orthologs) identified 90.5–94% of the expected genes (Table 2), depending on the reference database used. The snail plot in Figure 4A summarises the scaffold length distribution for the final assembly, while the blob plot in Figure 4B shows the distribution of contigs by GC proportion and coverage. High fragmentation of the final assembly and high rate of duplications after purging can be explained by population-level heterozygosity of the input sample. Annotation of protein-coding genes was not attempted due to the absence of RNAseq data.
Nuclear genome assembly statistics
Table 2. Long description
The table is divided into two main sections. The top section provides general assembly metadata for Accession P R J E B 1 1 5 7 8 2 and Biosample S A M E A 1 2 2 9 7 8 8 7 5. The assembly level is contig, with a span of 242.73 M b, 7562 contigs, and a contig N 50 of 43 k b.
The bottom section details B U S C O reference database results across four categories: nematoda odb 12 (596 orthologs), nematoda odb 12.1 (1477 orthologs), chromadorea odb 12 (1126 orthologs), and chromadorea odb 12.1 (2034 orthologs).
Key data points for these four databases respectively include:
* Complete (all): 542 (90.9 percent), 1314 (89.0 percent), 987 (87.7 percent), and 1763 (86.7 percent).
* Complete (single-copy): 460 (77.2 percent), 1089 (73.7 percent), 827 (73.4 percent), and 1491 (73.3 percent).
* Complete (duplicated): 82 (13.8 percent), 225 (15.2 percent), 160 (14.2 percent), and 272 (13.4 percent).
* Fragmented: 18 (3.0 percent), 47 (3.2 percent), 42 (3.7 percent), and 78 (3.8 percent).
* Missing: 36 (6.0 percent), 116 (7.9 percent), 97 (8.6 percent), and 193 (9.5 percent).
Nuclear (A–B) and mitochondrial (C) genomes of Cranifera cranifera (Chitwood, Reference Chitwood1932). A: The BlobToolKit snail plot visualising the assembly metrics, the circumference represents the length of the whole genome sequence, the outermost blue tracks display the distribution of GC, AT, and N percentages, scaffolds are arranged clockwise from longest to shortest and are depicted in dark grey, the longest scaffold is indicated by the red arc, and the deeper orange and pale orange arcs represent the N50 and N90 lengths; B: the BlobToolKit blob plot visualising sequence coverage (vertical axis) and GC content (horizontal axis) with the circles representing scaffolds, with the size proportional to scaffold length and the colour representing phylum while the histograms along the axes display the total length of sequences distributed across different levels of coverage and GC content; C: annotated mitogenome showing protein-coding genes (blue), ribosomal RNA genes (green), transfer RNA genes (red), read coverage (brown), GC content (black), and repetitive region (yellow).
Figure 4. Long description
Panel A is a BlobToolKit snail plot. At the center, a grey spiral represents scaffolds arranged by length. Moving outward, a dark orange arc indicates the N 50 length of 43 k and a pale orange arc indicates the N 90 length of 15.3 k. The outermost blue tracks show G C content at 38.8 percent and A T content at 61.2 percent. A red line marks the longest scaffold at 1.10 M.
Panel B is a BlobToolKit blob plot. The horizontal x-axis represents G C content from 0.25 to 0.7. The vertical y-axis represents sequence coverage from 1 to 1 k. Circles represent scaffolds, colored by phylum (mostly blue for Nematoda) with size proportional to length. Histograms on the top and right margins show the distribution of sequence lengths across G C content and coverage levels.
Panel C is a circular mitogenome map. From the innermost layer outward:
- A scale bar in k b p from 0 to 24.
- A black G C content track with a yellow highlighted repetitive region between 14 and 22 k b p.
- A brown read coverage track.
- A circular track of genes: blue blocks for protein-coding genes (such as n a d 5, c o x 1, n a d 4), green blocks for r R N A genes (r r n L, r r n S), and red lines for t R N A genes (such as t R N A-G l n, t R N A-T r p). Labels with red and blue text point to specific gene locations around the circumference.
Mitochondrial genome assembly
The complete mitochondrial genome has a total length of 24644 bases (Figure 4C), including a region with high repeat content between 12653 and 22383 bases: repeat #1 is the largest duplicated block 2975–2977 bases long (12653–15626 and 19407–22383 positions); repeats #2 and #3 sit inside that same overall region; first copy of a 1318-base-long repeat #2 (15109–16426 positions) overlaps the tail of repeat #1’s first copy while the second copy of repeat #2 (17261–18578 positions) falls in the gap between repeat #1’s two copies; first copy of a 688-base repeat #3 (17091–17778 positions) falls in the gap between repeat #1’s two copies while the second copy of repeat #3 (19237–19924 positions) partially overlaps with the second copy of repeat #1 (Figure 3C).
Annotation using both Mitos2 (Bernt et al. Reference Bernt, Donath, Jühling, Externbrink, Florentz, Fritzsch, Pütz, Middendorf and Stadler2013) and Aragorn (Laslett et al. Reference Laslett and Canbäck2004) identified a complete set of 12 protein-coding genes (atp6, cob, cox1-3, nad1-6, and nad4L) with atp8 being absent, as expected for nematodes, two rRNA genes and 22 tRNA genes, some of which were annotated several times by both Mitos2 and Aragorn. Specifically, original annotation included eight tRNA -Lys (ttt) and four tRNA -Asn (gtt) copies, duplicates of tRNA -Ser (tct) and tRNA -Leu (taa), and a spurious tRNA -Tyr (gta) nested inside tRNA -Gly (tcc); both rRNAs were truncated (rrnS at 386 nt and rrnL at 666 nt), while several protein-coding genes had various annotation errors. The curated mitogenome includes the standard set of 22 tRNAs, with one functional copy kept for each tRNA duplicated in the repeat array, and rRNAs are re-annotated to 677 nt in rrnS and 961 nt in rrnL, in agreement with conserved motifs and gene lengths in the reference genomes. All protein-coding genes have all start and stop codons in frame with no internal stops and their gene boundaries agree within a few amino acids with 13 published reference mitogenomes.
Intragenomic variation of rRNA
Since the 28S rRNA gene is the most commonly used barcode for species delimitation in Thelastomatidae, the newly assembled nuclear genome was mined for rRNA amplicon sequences in order to access intragenomic and intraspecific variation. Two amplicon variants of the 28S rDNA were mined from the newly assembled genome (Figure 5A). In the Maximum Likelihood phylogram based on the 753-bp-long alignment of the D2-D3 expansion domains of the 28S rDNA, one of these sequences (QB046965) forms a clade with the other three sequences of C. cranifera available in NCBI GenBank, although bootstrap support for this clade is moderate, and the internal topology is not resolved. The second copy of the gene (QB046966) was recovered as a sister clade to all other sequences. Based on the same alignment, the sequence of C. cranifera from B. discoidalis from Cuba (ON616698) was identical to QB046965. The sequence QB046966 differs in eight homologous positions from QB046965 (0.67% genetic distance) and in five and eight homologous positions with the sequences from C. cranifera from the Cuban Blaberus discoidalis and from captive Blaptica dubia from Russia, respectively (0.67% and 1.08% genetic distance, respectively). Seven of the eight variable bases between QB046965 and QB046966 are located within a variable segment 30 bases long and two of these correspond to indels (Figure 5A).
Maximum-likelihood (ML) trees inferred from the D2-D3 28S rDNA for several sequences of Cranifera cranifera (Chitwood, Reference Chitwood1932) with Severianoia blapticola Guzeeva, 2009 used as an outgroup (A) and the 18S rDNA for several sequences of C. cranifera and species of the family Thelastomatidae with Travassosinema viatorum Morffe, García & Hasegawa, 2023 used as an outgroup (B). Pairwise comparison of the C. cranifera sequences on the right. Sequences from the present study are highlighted in bold.
Figure 5. Long description
The image consists of two panels, A and B.
Panel A (Top):
* Left: A Maximum-likelihood M L tree based on D 2-D 3 28S r D N A. The tree shows a cluster of Cranifera cranifera sequences including Blaptica dubia (Sweden) Q B 0 4 6 9 6 5, Blaptica dubia (Russia) E U 3 6 5 6 3 2, Blaberus discoidalis (Cuba) O N 6 1 6 6 9 8, Archimandrita tessellata (Costa Rica) O N 6 1 6 6 9 7, and Blaptica dubia (Sweden) Q B 0 4 6 9 6 6. Severianoia blapticola E U 3 6 5 6 3 1 is the outgroup. A scale bar indicates 0.02 substitutions per site.
* Right: A pairwise comparison table for the five sequences listed above. Values range from 0.00 to 1.08.
Panel B (Bottom):
* Left: A Maximum-likelihood M L tree based on 18S r D N A. It includes various species from the family Thelastomatidae. C. cranifera sequences highlighted in bold are Blaptica dubia (Sweden) Q B 0 4 2 1 5 1 and Blaptica dubia (Sweden) Q B 0 4 2 1 5 2. Other species include Hammerschmidtiella, Suifunema, Desmicola, Buzionema, Severianoia, Aorurus, Archimandrita, Blaberus, and Cephalobellus. Travassosinema viatorum O P 7 9 3 5 0 5 is the outgroup. A scale bar indicates 0.01 substitutions per site.
* Right: A pairwise comparison table for four sequences: O N 6 1 6 6 9 3, Q B 0 4 2 1 5 1, O N 6 1 6 6 9 4, and Q B 0 4 2 1 5 2. Values range from 0.00 to 1.83.
Despite the 18S rDNA gene being more useful at the supraspecific level than as a species delimitation marker, it was included in the analyses. Two amplicon variants of the 18S rDNA were also recovered from the newly assembled genome (Figure 5B) and used for the phylogenetic analysis together with the available sequences of C. cranifera and selected species of the family Thelastomatidae. In the Maximum Likelihood phylogram based on the 1508-bp-long alignment, all the sequences of C. cranifera form a strongly supported clade, with the sequence from A. tessellata from Costa Rica (ON616693) located in a basal position. The internal topology of this clade is not resolved, with the current sequences (QB042151 and QB042152) forming a clade with moderate support with the sequence of C. cranifera from B. discoidalis from Cuba (ON616698). Based on the same alignment, one of the current sequences (QB042152) was identical to the one from the Cuban B. discoidalis, similar to 28S (see above). The sequence QB042151 differs in 24 homologous positions (1.62% of genetic distance) with QB042152 and in 27 and 24 homologous positions with the sequences from A. tessellata from Costa Rica and B. discoidalis from Cuba (1.83% and 1.62% genetic distance, respectively) (Figure 5B).
Discussion
Cranifera cranifera is a common intestinal endobiont of several blaberid cockroaches, three of which are reared in captivity as pets (Archimandrita tessellata and Blaberus craniifer) or food sources for other animals (Blaptica dubia), which makes this species easily accessible for both morphological and molecular studies. Despite this, many aspects of its biology remain poorly understood or completely unknown. For example, in addition to the present work, males of C. cranifera have only been previously recorded twice: the first time in the original description of the species by Chitwood (Reference Chitwood1932) and the second time in subsequent redescription by Leibersperger (Reference Leibersperger1960). These authors only included drawings of the posterior end of the males and not from the anterior region and oesophagus, notwithstanding remarkable sexual dimorphism in this species (and in all Thelastomatoidea). Here, the anterior and oesophageal regions of the male are illustrated for the first time. The morphology of the oesophagus was described differently by Chitwood (Reference Chitwood1932) and Leibersperger (Reference Leibersperger1960). Both considered the corpus as sub-divided into an anterior region (‘a weak pseudobulb’ according to Leibersperger (Reference Leibersperger1960)) and a posterior region but did not describe their shape. Also, they considered the isthmus as short. In the present work we observed that this anterior region is the proper corpus and the posterior one constitutes a long isthmus. The arrangement of the copulatory papilliform sensilla observed by Chitwood (Reference Chitwood1932) and Leibersperger (Reference Leibersperger1960) coincides with the current specimens: a pre-cloacal pair near the cloaca, and three post-cloacal pairs, one ad-anal pair, one sub-dorsal near the base of the tail appendage, and one ventral, minute pair along the tail appendage. However, the original description by Chitwood (Reference Chitwood1932) mentioned the copulatory papillae pattern as ‘caudal papillae consisting of one pair of preanal papillae, one pair of ad-anal or postanal papillae, and a median papilla near the base of the spinelike tail’, while the line drawing of the tail region showed both the ad-anal and the post-anal pairs of papillae. The latter points to a possible lapsus calami by the author.
The nuclear genomic resources for arthropod pinworms of the superfamily Thelastomatoidea are extremely limited (Lü et al. Reference Lü, Zhang, Gu, Zhang, Hugot, Gibson, Chen, Zhang, Xu and Li2026), despite these species often being considered important ecological (not phylogenetic) intermediates between free-living nematodes and obligate endoparasites inhabiting some of the earliest terrestrial macrofauna (Sudhaus Reference Sudhaus2010). Phylogenetically, they are nested within the more specialised groups of parasites (Lü et al. Reference Lü, Zhang, Gu, Zhang, Hugot, Gibson, Chen, Zhang, Xu and Li2026; Sudhaus Reference Sudhaus2010) which makes the origin and evolution of parasitism in Clade 3 nematodes an interesting puzzle. The nuclear genome of C. cranifera is the third for Thelastomatoidea arthropod pinworms, in addition to Thelastoma shijiazhuangense Zeng, Yuan, Liu & Meng, 2024 and Travassosinema thyropygi Hunt, 1996 (Lü et al. Reference Lü, Zhang, Gu, Zhang, Hugot, Gibson, Chen, Zhang, Xu and Li2026), the latter two being based on Illumina short-read sequencing with no assembly or annotation files being publicly available. As such, building a comprehensive and dependable phylogenetic framework for early branching Clade 3 nematodes remains nearly impossible, due to huge gaps in taxonomic sampling for sequencing projects.
While there are many more mitochondrial genomes available for Clade 3 nematodes, including Thelastomatoidea (Lü et al. Reference Lü, Zhang, Gu, Zhang, Hugot, Gibson, Chen, Zhang, Xu and Li2026; Xu et al. Reference Xu, Zhang, Zhang and Li2026), their utility for phylogenetic analysis remains questionable, mainly due to topological discrepancies with phylogenies based on nuclear genomes in the interrelationships between Clade 3 (Spirurina) and Clade 4 (Tylenchina) nematodes (Zhan et al. Reference Zhan, Shen, Mundock, Ren, Li, Xue and Guo2026). Gene arrangements in mitochondrial genomes also vary considerably across the entire Nematoda, being relatively conserved in some lineages and showing no conserved pattern at all in other groups (Du et al. Reference Du, Guo, Gao, Wang, Qing and Li2022; Lü et al. Reference Lü, Zhang, Gu, Zhang, Hugot, Gibson, Chen, Zhang, Xu and Li2026; Nikolaeva et al. Reference Nikolaeva, Ovcharenko, Khorkhordina, Miroliubova, Sadovskaya, Scobeyeva, Sanamyan, Panina, Mikhailov, Rusin, Tchesunov and Aleoshin2025; Tomé et al. Reference Tomé, Harris, Jorge, de Sousa, Archer, Muñoz-Merida, Rato, Perera and Mulder2026), which can be partially explained by sequencing bias: conserved patterns are often observed in closely related species, members of the same genus, while more diverse arrangements of protein-coding genes are more commonly observed in sparsely sequenced lineages. As such, we are not discussing the variations of gene order in mitochondrial genomes of nematodes, awaiting a much larger body of information. Instead, we would like to highlight the impact of repetitive regions on the assembly and annotation of nematode mitochondrial genomes. The mitochondrial genome of C. cranifera is 24644 bases long including a region with complex repeat content spanning 9729 bases, with the largest repeat fragment being almost 3000 bases long. This region was assembled both by MitoHifi and our custom pipeline. We cannot completely exclude the possibility that it is an assembly artefact, since alternative assemblies 17888, 22497 and 31115 bases long were also recovered. However, mapping raw reads against all four alternative assemblies revealed assembly collapse in both 17888- and 22497-base-long assemblies, and coverage drop in 31115-base-long assembly. Similarly sized non-coding regions were identified in mitogenomes of other nematode species as well, specifically in Hoplolaimus columbus (7661 bp and 3157 bp long), Pratylenchus vulnus (6847 bp), Meloidogyne chitwoodi (5404 bp), M. graminicola (5063 bp), and Meloidogyne incognita (4097 bp) (Humphreys-Pereira and Elling Reference Humphreys-Pereira and Elling2014; Ma et al. Reference Ma, Agudelo, Richards and Baeza2020; Sultana et al. Reference Sultana, Kim, Lee, Han, Kim, Min, Nadler and Park2013; Sun et al. Reference Sun, Zhuo, Lin, Wang and Liao2014). Therefore, presence of duplicated regions in mitogenome assemblies based on short-read data (where duplications are likely to be collapsed during assembly) may not be recovered, unless read coverage is used for verification.
Close match between recovered sequences of 28S rDNA and 18S rDNA (especially QB046965 and QB042152, respectively) and reference D2-D3 domains of the 28S rDNA and 18S rDNA available from the NCBI GenBank confirms the identity of the studied population as C. cranifera and complements the morphological and morphometric data. The basal position of QB046966 in the 28S rDNA phylogram compared to all other sequences of C. cranifera in our opinion represents intragenomic variation, a less common sequence variant. Despite the fact that QB042151 and QB042152 are located in the same clade of the 18S rDNA tree, the divergence rate of QB042151 also points to the presence of intragenomic variation in this gene. Intragenomic variation in rRNA sequences has been previously documented in several groups of free-living and parasitic nematodes by several authors (i.e. Bik et al. Reference Bik, Fournier, Sung, Bergeron and Thomas2013; Qing et al. Reference Qing, Bik, Yergalyev, Gu, Fonderie, Brown-Miyara, Szitenberg and Bert2019); one of the sequence variants can be more divergent from the other, creating a polyphyletic topology (Figure 5A) and supporting the hypothesis that such divergent variants are less common copies, negatively selected during regular polymerase chain reaction amplification, but readily identifiable in whole-genome datasets, and should be accepted as a fact of intragenomic heterogeneity of the rRNA operon and not a signal of interspecific divergence of cryptic speciation.
Last but not least, the actual impact of thelastomatid nematodes on their hosts remains poorly understood. Studies suggest that thelastomatid nematodes specialise in feeding on the gut content of detritivorous hosts (Hominick and Davey Reference Hominick and Davey1973) and may positively affect the diversity of the host microbiome (Vicente et al. Reference Vicente, Ozawa and Hasegawa2016), but little is known about how this affects host health (McCallister Reference McCallister1988), and nothing about whether the host provides a response. Being relatively easy to identify compared with other Thelastomatoidea, having a simple life cycle, inhabiting commercially accessible and easy-to-maintain hosts that are currently completely exempt from ethical review processes, being more closely related to many parasites of veterinary and human health importance than C. elegans, and having genomic resources available now, C. cranifera has great potential to be used as a model system to study not only the evolution of parasitism in general but also the acquisition of anthelmintic resistance and how to counteract it. Other species from the same family have already been used to study anthelmintic response in the past (Holoman Reference Holoman1980; Yu and Crites Reference Yu and Crites1990).
Data availability
Morphological vouchers on permanent glass slides are deposited in the invertebrate collection of the Department of Zoology, Swedish Museum of Natural History (SMNH), Stockholm, Sweden (accession numbers SMNH 231508–231518). Raw data and final nuclear genome assembly are available from the European Nucleotide Archive under the accession number PRJEB115782. Raw reads are available under BioSample number SAMEA122978875 and accession number ERR17501394, respectively. The curated nuclear genome is available under the accession number ERZ30225045 and at https://www.ebi.ac.uk/ena/browser/view/GCA_986284995. The mitochondrial genome is available from the NCBI GenBank under the accession number QB100924.
Author contribution
Jans Morffe performed collection and preservation of nematodes and DNA extraction and morphological analysis; Nadège Guiglielmoni, Nadim Wassey, Karim Gueddach, Anja Schuster, and Kerstin Becker generated genomic data; Jens Bast and Philipp Schiffer acquired funding and oversaw generation of genomic data; Oleksandr Holovachov conceptualised and supervised the study in general as well as performed genome assemblies. All authors contributed to writing and approved the final version of the manuscript.
Acknowledgements
The authors are grateful to Niklas Apelqvist (Swedish Museum of Natural History) for providing the Blaptica dubia colony. Computational support and infrastructure were provided by the ‘Centre for Information and Media Technology’ (ZIM) at the University of Düsseldorf (Germany).
Financial support
Specimen collection, morphological study and data analysis were funded by the project ‘Diversity and phylogeny of insect associated nematodes in Sweden’ (SLU. dha.2024.4.3-192) granted to Jans Morffe and Oleksandr Holovachov by the Swedish Taxonomy Initiative, Artdatabanken, Swedish University of Agricultural Sciences. Sequencing was funded by the DFG through the grant BA 58004-1 to Jens Bast and Philipp Schiffer within the DFG Sequencing call 2021.
Competing interests
The authors declare no competing interests.
Ethical standards
Not applicable. No ethics approval is necessary.
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