Untangling the Derogenes varicus species complex in Scandinavian waters and the Arctic: description of Derogenes abba n. sp. (Trematoda, Derogenidae) from Hippoglossoides platessoides and new host records for D. varicus (Müller, 1784) sensu stricto

Several studies have shown that the euryxenic trematode Derogenes varicus (Müller, 1784) represents a species complex. Four lineages have been designated (DV1–4) with the DV1 clade corresponding to D. varicus sensu stricto. Herein, we investigate newly collected specimens of D. varicus sensu lato from Scandinavian and Arctic waters using integrative taxonomy. The trematodes were collected from Melanogrammus aeglefinus, Eutrigla gurnardus, Trachinus draco, and Merluccius merluccius off the Atlantic coast of Sweden and from Hippoglossoides platessoides from Arctic Svalbard. 28S sequences of derogenids from Sweden were identical to D. varicus sensu stricto, confirming its euryxeny. The 28S sequences of Derogenes sp. from H. platessoides were identical to Derogenes DV2 and differed from D. varicus sensu stricto by 3% and from Derogenes DV3 by 2%. The 28S sequence divergences of Derogenes sp. from H. platessoides with D. ruber and D. lacustris were 3 and 10%, respectively. ITS2 and cox1 divergences between Derogenes sp. from H. platessoides and other Derogenes species/lineages were at levels of interspecific differences. The species from H. platessoides is described here as D. abba n. sp. We also examined the type material of Progonus muelleri (Levinsen, 1881), the type and only species of the genus Progonus, with redescription and designations of paralectotypes. Based on specimens from Theodor Odhner’s collections at the Swedish Museum of Natural History, SMNH, Stockholm, we provide novel morphological and anatomical data for D. varicus sensu lato species complex. Lastly, we investigated Arthur Looss’s “lost collection” of Trematodes at the SMNH and characterised a putative species Derogenes sp. “limula”.


Introduction
A common problem in taxonomy and biodiversity assessment is the recognition of cryptic species.Although morphologically indistinguishable, cryptic species are genetically divergent, which may lead to unclear species boundaries between taxa.Additionally, the identification and study of cryptic species, along with the classification of organisms into nominal species, has importance beyond mere biodiversity assessment.This is particularly significant in the context of helminth parasites that impact human and veterinary health, as the presence of cryptic species can influence important medical and epidemiological factors such as pathogenicity and drug resistance [11].
The frequency of cryptic species varies among helminth groups [11].Within the Trematoda, the richest known fauna of all the major metazoan taxa of fishes [6], cryptic species are frequently recognised with reports of them from at least 20 families [8].Previous solid studies have demonstrated that encountering cryptic trematode species has become so frequent that their presence can almost be anticipated and should definitely be considered [12,16].In trematodes, firstly, there is the issue of phenotypic variation, which can mask common characteristics, potentially hindering identification.Secondly, there is the challenge of cryptic species, where species may appear similar externally but are genetically distinct [12].Trematode species with similar morphology, but reported from wide host and geographic ranges, are often revealed as complexes with cryptic species [21].Molecular techniques are indispensable for identifying cryptic species, as demonstrated in various trematode families [12].The records of the derogenid Derogenes varicus (Müller, 1784), from 69 fish species across 33 families worldwide [3], displaying an exceptional euryxeny, hint that cryptic species may be present, and it has frequently been suggested that this species represents a species complex [19,28].Krupenko et al. [30] used molecular evidence to recognise four genetic lineages and infer that it is a species complex (designated as "DV1-4").By analysing 28S sequences of D. varicus from the type-host, the Atlantic salmon Salmo salar from Norway, Bouguerche et al. [3] demonstrated that the DV1 clade is D. varicus sensu stricto (s.s.).
Herein, we investigate specimens of D. varicus sensu lato (s.l.) collected from several unrelated marine fish species, from Scandinavian and Arctic waters.
Based on DNA sequences of the second internal transcribed spacer (ITS2), the large subunit ribosomal DNA (28S) and the subunit I (cox1) in mtDNA and the examination of newly collected specimens, we provide a formal description of a new species in the genus Derogenes Lühe, 1900, recognised and described from the American plaice Hippoglossoides platessoides.We also identify D. varicus s. s. using 28S DNA sequences from additional host species.Examination of additional specimens of D. varicus s. l. from Sweden and Norway, Northeast Atlantic, from Theodor Odhner's collections at the Swedish Museum of Natural History (SMNH), Stockholm, enabled us to provide novel morphological and anatomical data for D. varicus s. l. from various hosts.We also examined the type material of Progonus muelleri (Levinsen, 1881), the type and only species of the genus Progonus Looss, 1899 from the shorthorn sculpin Myoxocephalus scorpius from Aasiaat [Egedesminde], Greenland.Finally, we further investigated Arthur Looss's "lost collection" of trematodes, bought by T. Odhner and preserved at the SMNH and we characterise a putative species Derogenes sp., an unpublished species referred to by A. Looss as "Derogenes limula".The importance of integrative taxonomy is stressed here and demonstrated for species delimitations within the D. varicus species complex and in the genus Derogenes.

Host and parasite collection
Fishes were collected from off Sweden, Northeast Atlantic (Skagerrak, Kattegat, and Gullmarsfjorden) and off Svalbard, Arctic Norway (Table 1).
Specimens from Skagerrak and Kattegat were collected by the SLU Aqua team as part of the biannual International Bottom Trawl Survey, within the scope of their research projects and permits.Specimens from Gullmarsfjorden were collected in the vicinity of Kristineberg Center for Marine Research and Innovation, outside of the borders of the Gullmarns nature reserve, and within the scope of the permit for animal research from the Swedish Board of Agriculture (Enheten för försöksdjur och sällskapsdjur, Jordbruksverket, Dnr.5.2.18-5483/18) and ethical approval for animal research from the Uppsala animal ethics committee (Uppsala djurförsöksetiska nämnd, Jordbruksverket, Dnr.5.8.18-17209/2021) issued to the Swedish Museum of Natural History.Specimens from the Arctic Ocean were collected during the HHUMTL22 cruise by the Arctic University Museum of Norway, within the scope of the fieldwork sampling permit issued by the governor of Svalbard (RiS-1D12021Al) and the permission to trawl from the Norwegian Directorate of Fisheries (21/16250).Fishes were euthanised and made available for examination.
Several specimens of greater weever Trachinus draco, grey gurnard Eutrigla gurnardus, European hake Merluccius merluccius, and haddock Melanogrammus aeglefinus from Skagerrak and Kattegat were collected during the biannual International Bottom Trawl Survey by the SLU along the Swedish coast (    [68]. Digeneans were collected from freshly killed fishes.The gastrointestinal tract was removed and examined for trematodes using the gut wash method [14,26].Digeneans were fixed in near-boiled saline without pressure and preserved immediately in 80% ethanol for morphological and molecular studies.Nine specimens were processed as hologenophores (sensu Pleijel et al. [53]).Type specimens of P. muelleri were requested from the Natural History Museum of Denmark (SNM) and nine specimens of P. muelleri from M. scorpius from Aasiaat [Egedesminde] Greenland preserved in 70% ethanol and marked as holotype (old catalogue number ZMUC-TRE-000032) were received.Two of the specimens were stained in acetocarmine and studied by microscopy.

Morphological methods
Preserved specimens of Derogenes including hologenophores were stained in iron acetocarmine, destained with acid-alcohol (1% HCl in 70% ethanol), dehydrated in an ethanol series (70-100%), cleared in clove oil, and mounted in Canada balsam.Two specimens of the type material of P. muelleri were stained according to the same methods and mounted on two separate slides (NHMD-114950).
Drawings were made through a Nikon Eclipse i80 microscope with DIC (differential interference contrast) and a drawing tube.Drawings were scanned and redrawn on a computer with Adobe Illustrator 2023.Stained specimens were measured by ImageJ ver.1.53K [57].Measurements are in micrometres and indicated as the range followed by the number of measurements in parentheses.Types and vouchers were deposited at the Swedish Museum of Natural History (SMNH), Stockholm, Sweden; the Natural History Museum of Denmark (SNM), Copenhagen, Denmark and the Arctic University Museum of Norway (UiT), Tromsø, Norway.

Molecular methods
Genomic DNA was extracted from seven hologenophores of D. varicus s. l. from T. draco, E. gurnardus, M. merluccius, M. aeglefinus, and two hologenophores of Derogenes sp. from H. platessoides.Genetic data were generated for three markers: cox1, ITS2, and the 28S rDNA.Small fragments of each hologenophore (posterior third) were placed in a 1.5 mL microcentrifuge tube containing 20 lL buffer ATL (QIAGEN, Hilden, Germany).For extraction of genomic DNA (gDNA), 20 lL buffer ATL and 20 lL proteinase K were added to each sample, followed by vortexing and incubation in an incubating microplate shaker at 56 °C and 300 rpm overnight.The lysed samples were processed to obtain gDNA, following the manufacturer's instructions for gDNA extraction using a QIAGEN QiAmp DNA Microkit.The PCR reaction was performed following Bouguerche et al. [3].

Trees and distances
Phylogenetic analyses were performed using the newly generated sequences of Derogenes spp.and those of related species available in GenBank (Table 2), mainly the D. varicus complex and P. muelleri complex provided by Krupenko et al. [30] and by Bouguerche et al. [3].Alignments for each gene region were constructed separately in AliView [32], then trimmed to the shortest sequence.Phylogenetic tree inference was carried out by the maximum likelihood (ML) method using MEGA11 [66].Nucleotide substitution models for phylogenetic  analyses using the ML method were selected using MEGA11 [66].The Hasegawa-Kishino-Yano with Gamma Distributed (HKY+ G) model [23] was selected for the 28S, the Kimura 2-parameter (K2) model [27] for ITS2, and the Tamura-Nei model with Gamma Distributed with Invariant sites (TN93+ G+I) [65] for cox1.The probabilities were computed by the bootstrap analysis of 500 replications.We also constructed phylogenetic trees of respective regions for the same data sets using the neighbour-joining (NJ) method [56] with MEGA11, with 2,000 bootstraps computed for cox1, ITS2, and 28S.The p-distances [27] were computed from the same datasets with MEGA11.urn:lsid:zoobank.org:act:7CB281BD-05E0-4BAF-A717-A1E4E1477AE1
Paratypes with molecular information: anterior parts of specimens mounted on a slide, posterior part used for molecular analysis: slide SMNH-Type-9563; slide SMNH-Type-9564.
Additional material examined for comparison: Whole mounts: ( 1 Etymology: Named after ABBA, the Swedish pop supergroup renowned for hits like "Dancing Queen", "Chiquitita" and "Money, Money, Money" which served as a source of entertainment for the first author during the creation of the illustrations.The group's name is an acronym of the first letters of their first names arranged as a palindrome.Invariable, treated as a noun in apposition. Description: Measurements and comparisons in Tables 3-5.Body stocky (Figs.1A and 1B), nearly sausage-shaped; anterior and posterior ends rounded.Pre-oral lobe short.Oral sucker rounded.Pre-pharynx absent.Pharynx muscular.Oesophagus short.Intestines bifurcating anterior to sinus-organ.Intestinal caeca extending posteriorly to vitelline masses and terminating blindly.Ventral sucker rounded.
Ovary globular, voluminous, post-testicular, sometimes overlapped by right vitelline mass.Laurer's canal not observed.Vitelline masses in hindbody, round to oval, paired, situated on each side of body.Vitelline ducts fuse antero-medial to ovary.Seminal receptacle not observed.Uterus convoluted, uterine coils extending from near posterior extremity to sinus sac.Eggs oval.Excretory vesicle Y-shaped; branches reuniting dorsal to pharynx.
The morphology of the cercaria was described by Krupenko et al. [30].Archival documents: in addition to a single slide mounted by A. Looss (SMNH-208361) (Fig. 2), the archives include two unpublished line drawings (Figs.3A and 3B), combined and reproduced in Figures 4A-4E.
Remarks: Derogenes sp. that we described above was found in A. Looss's collection, labelled as "Derogenes limula".As there are no published records of a species under that name, A. Looss probably intended to describe this Derogenes specimen from P. tentacularis as a new species, with the name "D. limula".The eggs of this Derogenes sp. that we described above are over 40 lm and the species is thus consistent with "the large eggs group".We compared the single specimen of Derogenes sp.(or "D.limula" as initially labeled by A. Looss) ex P. tentacularis to the Mediterranean congeneric Derogenes species.The present specimen Derogenes sp.ex P. tentacularis differs from D. minor, D. robustus, D. affine, and D. latus by its larger eggs.It resembles D. ruber in egg size (61 Â 39 in Derogenes sp. vs. 62 Â 39 in D. ruber) and in having lobed, tear-shaped vitelline masses.However, Derogenes sp.ex P. tentacularis can be readily distinguished from D. ruber by being smaller in all body measurements including the body (855 Â 253 vs. 7869 Â 1847).Derogenes sp.ex P. tentacularis can be easily distinguished from D. varicus s. s. and D. abba n. sp. by having lobed vitelline masses.

Remarks:
The type-material received from Natural History Museum of Denmark (SNM), Copenhagen, Denmark includes nine specimens preserved in 70% ethanol from M. scorpius off Aasiaat [Egedesminde], Greenland, and were indicated as the Holotype.Two specimens were stained in acetocarmine, mounted on slides, and designated herein as Paralectotypes.Comparative terminal genitalia of D. varicus s. s. shown in Figure 5C.

Molecular characterisation
The NJ and ML methods led to similar tree topologies and thus only the ML trees are shown (Figures 6-8).
The 28S dataset included 53 nucleotide sequences of the Derogenidae Nicoll, 1910.The trimmed matrix comprised 642 positions.The newly generated sequences of D. varicus ex M. aeglefinus, E. gurnardus, T. draco, and M. merluccius from Sweden were identical and clustered within the D. varicus s. s. clade without any host-related structuring.
The two newly generated 28S sequences for D. abba n. sp.ex H. platessoides were identical to each other and to sequences of rediae of D. varicus DV2 of Krupenko et al. [30] ex the gastropod intermediate mollusc hosts Euspira pallida and Amauropsis islandica from the White Sea and the Barents Sea; they were also identical to adults ex B. scalariforme from the White Sea and from the definitive host H. platessoides from the North Sea.All sequences divergence of D. abba n. sp.ex H. platessoides from D. varicus s. s. (= DV1 [26]) from these hosts from the White Sea, Barents Sea (Russia) and Table 6.Measurements of Derogenes sp."limula".from Parablennius tentacularis off Trieste, Italy and Derogenes spp.first described from the Mediterranean.*Diameter. 1We included measurements from the redescription of Rudolphi's specimen by Lühe (1901) as the original description did not provide any measurements.The ITS2 tree was constructed using 36 sequences of the Derogenidae (Fig. 7).The trimmed matrix included 428 positions.The newly generated sequences of D. abba n. sp.from Svalbard (ex H.The cox1 sequences of D. abba n. sp.from Svalbard (ex H. platessoides) were aligned with 31 other derogenid sequences, all relating to the genera Derogenes and Progonus.The trimmed matrix included 783 positions.Two of them were identical, while these sequences differed by 1% from those of D. varicus DV2 from the White and Barents seas off Russia (ex intermediate hosts).The divergence was 21-31% between the sequences of D. abba n. sp. and those of D. varicus s. s. [3,30] and was 21-24% between the former and those of D. ruber from Algerian C. lastoviza [18].The highest divergence to a congener was 38-39% from D. lacustris from Argentinean freshwater fishes [67].In the phylogenetic tree of cox1 (Fig. 8  In the following section, we compare the new species D. abba n. sp. with five species of the large egg group first described from the Atlantic (Table 3), five species first described from the Mediterranean (Table 4), and seven species first described from the Pacific, Antarctic, and Indian Oceans (Table 5).analysis of the 28S sequences of D. varicus from the type-host, the Atlantic salmon S. salar.Krupenko et al. [30] did not find any adult stages of DV2 in fish and hence could not provide further morphological comparison of DV1 and DV2.Additionally, the only available sequences of D. varicus s. s. from the type-host had been made available only recently [3].Herein, the sequences of D. abba n. sp.from Arctic H. platessoides were identical to those of D. varicus s. l. from the same host in the North Sea [50] and those of D. varicus DV2 as rediae from gastropods in the White and Barents seas [30].They were also identical to D. varicus of The lineage D. varicus DV4 corresponds to an 18S rDNA sequence (AJ287511) of D. varicus of Littlewood and Olson [37] from the same host as D. abba n. sp., H. platessoides from the North Sea.DV4 was labeled only based on the genetic divergence of 0.13% (p-distance) from D. abba n. sp.(DV2) in the 18S rDNA sequence [30].In light of the available data, it is premature to consider D. varicus DV4 as a distinct species from D. abba n. p. considering that our study does not include the 18S rDNA analysis.Hence, we cannot estimate the taxonomical position of DV4 further.Meanwhile, the possibility that D. varicus (s.s.) also occurs in H. platessoides is not ruled out.

Comparison with Atlantic species (
Overall, the evolution of understanding of this genus is such that while most combinations of species can be distinguished on the basis of morphology, some are presently morphologically cryptic with respect to each other.On this basis, it remains critical that further study is based on both morphological and molecular data.A key area of uncertainty relates to patterns of host-specificity.Some species, e.g.D. varicus s.s., appear to be genuinely euryxenic, whereas several others apparently have highly restricted host ranges.The extent to which this is a genuine reflection of the true nature of these species is uncertain.Certainly, for clearly closely related species, it is not obvious why host-specificity patterns should differ so dramatically.
The German parasitologist Prof. Arthur Looss (1861Looss ( -1923) ) was among one of the most prolific parasitologists and taxonomists and was known for his "enthusiasm and energy as a researcher that have probably seldom been surpassed, especially a painstaking attention to detail that is unfortunately rare" [33].After his death, his collection was divided between numerous institutions: Smithsonian National Museum of Natural History in Washington (USA), the Natural History Museum in Berlin and the Natural History Museum in Leipzig (Germany), Gothenburg Museum of Natural History and the Swedish Museum of Natural History (Sweden) (see Kuzmina and Holovachov [31]).A part of Looss's collection including his archives was sold to the Swedish Museum of Natural History (Naturhistoriska riksmuseet) in Stockholm by his widow, Elise Looss in 1924 [31] and includes slides of which some are actually type material and vials containing trematodes preserved in ethanol, along with several publication-ready drawings and original line drawings.
One of the intriguing derogenids that we encountered in this collection, is "D.limula" which we described above as Derogenes sp., ex P. tentacularis, collected from off Trieste, Italy, Central Mediterranean.Curiously, we also found line drawings by A. Looss (see Figs. 3A and 3B) labeled as "Derogenes limula", which suggests that he intended to describe Derogenes from P. tentacularis as a new species, with the name "D. limula".We found one specimen (SMNH 208361) for which the measurements are presented in Table 6.The eggs of this "D.limula" that we described above as Derogenes sp. are over 40 lm and the species is thus consistent with the "large eggs group".We compared the measurements of the sole specimen of Derogenes ex P. tentacularis to those of congeneric species from the Mediterranean (Table 6).The single specimen of Derogenes sp.(or "D.limula" as initially referred to by A. Looss on the illustrations) ex P. tentacularis differs from D. minor, D. robustus, D. affine, and D. latus by its larger eggs.It resembles D. ruber in egg size (61 Â 39 in Derogenes sp. vs. 62 Â 39 in D. ruber) and in having lobed, tear-shaped vitelline masses.However, Derogenes sp.ex P. tentacularis differ from D. ruber by being smaller in all body measurements including the body (855 Â 253 vs. 7869 Â 1847).We note though that body size is not a sufficient differentiating character and can vary with the age of the worm and the suitability of the host.
Lebour [34] described some D. varicus adults with a similar "spiny" appearance and stated ". . .curious fact noticed is that all these larval D. varicus are beset with small spines, whereas it is a characteristic of the adult that although it has sometimes a wrinkling of the skin, it is unarmed and usually smooth.It is possible that these wrinkles may be the remains of the spines  fused together.The spines are especially distinct in the younger specimens."Interestingly, some of her specimens also contained eggs (i.e., progenetic).She also showed this spination in her figures [34].Hence, this "D.limula" that we described as Derogenes sp., is arguably an immature D. ruber.We could not find any published records for Derogenes in Blenniidae in the Mediterranean, but D. varicus was recorded from the butterfly blenny Blennius ocellaris in the Atlantic [47].Besides the localities being distant (Trieste, Mediterranean for "D.limula" from P. tentacularis vs. Plymouth, Atlantic for D. varicus of Nicoll [47]), "D.limula" from P. tentacularis can be readily distinguished from D. varicus by having lobed vitelline masses.Hence, "D.limula" of A. Looss is clearly not D. varicus.
Herein, we described it as Derogenes sp.pending further examination based on more specimens.
Progonus muelleri (Levinsen, 1881) and Derogenes varicus sensu lato The monotypic genus Progonus was erected by Looss [38] for P. muelleri, first described from waters off Greenland [35].The two genera Derogenes and Progonus are very similar with the only difference being the presence of a cyclocoel in P. muelleri vs. blindly ending caeca in species of Derogenes [19,35,49].We examined body sections of representatives of both genera, mainly D. varicus s. s. from Limanda limanda (Fig. 5C) and P. muelleri from M. scorpius and the two species can also be readily distinguished by the seminal vesicle, located in the anterior third of the midbody of D. varicus s. s. vs. in the posterior third of the midbody in P. muelleri.Additionally, the pars prostatica is far shorter in P. muelleri (see Fig. 5).
We examined additional specimens of Derogenes spp.from T. Odhner's collections at the Invertebrates collection in the SMNH (Figs. 9 and 10), all identified and labelled as D. varicus, which we consider herein as representative of the D. varicus species complex or D. varicus s. l.The hosts were Argentina sphyraena, Brosme brosme, Molva molva, and Platichthys flesus.Corresponding morphometrical data are indicated in Table 7. Overall, specimens from the previously mentioned hosts share the same anatomical features, concerning the organisation of gonads and genital terminalia (Figs.9B, 9D, 9F, 9H, 10B).Derogenes varicus s. l. from the previously mentioned hosts share the sausage-shaped appearance, except D. varicus s. l. from B. brosme that has a stockier appearance.The testes in D. varicus s. l. from A. sphyraena and the one from H. hippoglossus differ by having more longitudinally elongated testes.The most striking one is D. varicus s. l. from A. sphyraena that also differed slightly by the organisation of the genital terminalia (Fig. 9B), the shape of vitelline masses being larger and elongated, by having a smaller body and smaller organs but the measurements of eggs overlapped.However, the number of specimens measured from T. Odhner's collection is low and thus we refrained from comparing morphometrical values.Since we are attempting to delineate species within the D. varicus species complex using integrative taxonomy, and as molecular sequence data are lacking for Derogenes from A. sphyraena, B. brosme, M. molva, and P. flesus, we consider specimens from the previously mentioned hosts as D. aff.varicus or D. varicus s. l., pending further examinations.
Site in host: Stomach.Deposited examined material: Whole mounts: Two specimens of P. muelleri ex Myoxocephalus scorpius from Aasiaat [Egedesminde], Greenland, deposited at the Natural History Museum of Denmark (SNM), Copenhagen, Denmark (NHMD-114950/ old catalogue number ZMUC-TRE-000032), Paralectotypes.Transverse sections: T. Odhner's collections in the Invertebrates collection of the Swedish Museum of Natural History

C.
Bouguerche et al.: Parasite 2024, 31, 26  Scandinavian waters of the North Sea (Sweden and Norway) was 3%.There was 2% divergence between the sequences of D. abba n. sp. and those of D. varicus DV3[30] recovered from E. fedorovi in the North Pacific[63].They differed from those of D. ruber ex Chelidonichthys lastoviza off the coast of Algeria by 3%.The largest divergence was shown between the sequences of D. abba n. sp. and those of D. lacustris from Argentinean freshwater fishes[67], reaching up to 10%.The divergence between D. abba n. sp. and P. muelleri from different White Sea fishes[30] was 8%.In the phylogenetic tree of 28S rDNA sequences (Fig.6), D. abba n. sp.from Svalbard (ex H. platessoides) clustered as a well-supported clade with D. varicus DV2 ex the same host from the North Sea[46] and ex B. scalariforme from the White Sea[30].This clade was well separated from the D. varicus s. s. clade ex multiple hosts from the White Sea, Barents Sea (Russia) and Scandinavian waters of the North Sea (Sweden and Norway), the DV3 clade (ex E. fedorovi), the D. lacustris clade (ex G. maculatus), and the D. ruber clade (ex C. lastoviza).
platessoides) were identical to those of D. varicus DV2 ex intermediate hosts from the White Sea and Barents Sea (Russia).They differed by 4% from the sequences of D. varicus s. s. from the White Sea, Barents Sea (Russia), and Scandinavian waters of the North Sea (Sweden and Norway) and those of D. ruber off Algeria (ex C. lastoviza).The sequences of D. abba n. sp.from Svalbard (ex H. platessoides) formed a well-supported clade with those of D. varicus DV2.This clade separated from the D. varicus s. s. clade and the D. ruber clade.

14 C. 26 Figure 6 .
Figure 6.Tree inferred using the maximum likelihood method based on the 28S rDNA sequence data; only bootstrap values higher than 70 are indicated.The newly generated sequences of Derogenes varicus sensu stricto from Sweden, Northeast Atlantic are indicated by*.Other D. varicus sensu stricto are those of Krupenko et al. [30] from the Barents Sea and White Sea; and those of Bouguerche et al. [3] from Northeast Atlantic, off Sweden and Norway and from Svalbard, Norway, Arctic Ocean.

Figure 7 .
Figure 7. Tree inferred using the maximum likelihood method based on the ITS2 sequence data; only bootstrap values higher than 70 are indicated.

Figure 8 .
Figure 8. Tree inferred using the maximum likelihood method based on the cox1 sequences; only bootstrap values higher than 70 are indicated.

Table
).Eight specimens of M. aeglefinus from 2 C. Bouguerche et al.: Parasite 2024, 31, 26 Gullmarsfjorden were collected in the vicinity of the Kristineberg Center for Marine Research and Innovation.The American plaice H. platessoides from the Arctic Ocean was collected by bottom trawl, at 79 59.860558 N, 15 27.879328 E, and 170 m depth

Table 1 .
Fishes examined from Scandinavian waters of the North Sea, Northeast Atlantic and the Arctic Ocean during this study.

Table 2 .
[30]ection data for sequences analysed in this study.aTwosequences by Krupenko et al.[30]are wrongly annotated on GenBank: OM761977 and OM762017 and these two Derogenes varicus complex sp.DV1 isolates are in fact DV2.

Table 3 .
Measurements of Derogenes abba n. sp.from Hippoglossoides platessoides off Svalbard and Derogenes spp.first described from the Atlantic.*Diameter.1Note that D. lacustris is a freshwater species.

Table 5 .
Measurements of Derogenes abba n. sp.from Hippoglossoides platessoides off Svalbard and Derogenes spp.first described from the Pacific, Antarctic, and Indian Ocean.*Diameter.