Спросить
Войти
Russian Journal of Nematology, 2018, 26 (1), 77 - 85
Angiostoma meets Phasmarhabditis: a case of Angiostoma kimmeriense Korol Spiridonov, 1991
Elena S. Ivanova and Sergei E. Spiridonov
Centre of Parasitology, A.N. Severtsov Institute of Ecology and Evolution, Russian Academy of Sciences,
Leninskii Prospect 33, 119071, Moscow, Russia e-mail: elena_s_ivanova@rambler.ru
Accepted for publication 28 June 2018
Summary. Angiostoma kimmeriense (= A. kimmeriensis) Korol Spiridonov, 1991 was re-isolated from the snail Oxyhilus sp. in the West Caucasus (Adygea Republic) and characterised morphologically and molecularly. The morphology of the genus Angiostoma Dujardin, 1845 was discussed and vertebrate-associated species suggested to be considered as species insertae sedis based on the head end structure (3 vs 6 lips). Phylogenetic analysis based on partial sequences of three RNA domains (D2-D3 segment of LSU rDNA and ITS rDNA) did not resolve the relationships of A. kimmeriense, as the most similar sequences of these loci were found between members of another gastropod associated genus, Phasmarhabditis Andrassy, 1976. However, such biological traits of A. kimmeriense as its large size, limited number of parasites within the host and the site of infection, point to a parasitic rather than pathogenic/necromenic way of life typical for Phasmarhabditis.
The family Angiostomatidae comprises two genera, Angiostoma Dujardin, 1845 with its 18 species, and monotypic Aulacnema Pham Van Luc, Spiridonov and Wilson, 2005. The representatives of the family are generally associated with terrestrial gastropods but four species of Angiostoma have been described from vertebrate (amphibian and reptile) hosts. It had been shown that the gastropod-associated angiostomatids belong to the clade V of Nematoda together with gastropod-associated representatives of Agfa Dougherty, 1955 and Phasmarhabditis Andrassy, 1976 (Ross et al., 2010). The position of the Angiostoma species from vertebrate hosts remains unclear as no molecular data for them are yet available. As to the gastropod-associated species, only half of species have been molecularly characterised so far. Below we present the molecular-phylogenetic analysis for angiostomatids from gastropods based on the recent discovery of A. kimmeriense. Originally, A. kimmeriense Korol Spiridonov, 1991 (= A. kimmeriensis in original description, spelled here according to the gender of the genus following Ivanova Wilson, 2009) was recovered from the oesophagus of an endemic snail Oxychilus deilus (Bourguignat, 1857) in Crimea (Korol Spiridonov, 1991). The surveys carried in Crimea in 2008, 2011
and 2014 did not reveal the presence of the nematode in this or other gastropods examined (Vorobjeva et al., 2008; Ivanova et al., 2013). Several species of snails collected in Adygea (West Caucasus) were examined for the presence of nematodes. Oxychilus sp. about 20 mm in diam. (most probably, O. difficilis) was found infected by Angiostoma sp. together with two other nematode species, Phasmarhabditis sp. and Alloionema sp. Morphological examination of Angiostoma sp. showed that it represents A. kimmeriense. Three RNA domains were examined and partial sequences obtained hereby giving the molecular characteristics of this species. The description of A. kimmeriense from the West Caucasus is given together with illustrations.
MATERIAL AND METHODS
Nematode material. Snails, Oxychilus sp., were collected at the field research station of the South Federal University in Adygea, North Caucasus, Russian Federation in June 2016 and April 2018. Each snail collected was found to contain a mixed parasite infection. The mixed infection of Phasmarhabditis sp. with Angiostoma sp., Phasmarhabditis sp. and Alloionema sp.,
© Russian Society of Nematologists, 2018; doi:10.24411/0869-6918-2018-10007
Phasmarhabditis sp. and unidentified trematodes was observed as well as other combinations including more than two parasites.
Angiostoma kimmeriense were isolated from the snail oesophagus and anterior portions of intestine and preserved as follows: for morphological studies, in hot 4-5% formalin, and for molecular studies, by freezing at -20°C. Formalin-preserved nematodes were then processed to glycerin following the method of Seinhorst (1959) and mounted on permanent slides using the wax ring method. Measurements and drawings were obtained with Nikon Eclipse E200 microscopes with drawing attachments. Illustrations were finalised with a WACOM Intuos A4 USB drawing tablet and Adobe Illustrator CS5. Abbreviations in the text are: a, b, c, c& - de Man indices; L - body length; V% -distance from anterior to vulva as the percentage of the body length; GP - male genital papillae. For the SEM studies, formalin-preserved material was dehydrated and dried out at critical point in a Hitachi Critical Point Dryer HCP-1 and coated with gold in a S150A sputter coater. Images were taken on a Tescan Cam Scan MV 2300.
Molecular characterisation and phylogenetic analysis. Individual nematodes identified under light microscope were used for DNA extraction according to the protocol of Holterman et al. (2006). PCR amplification was performed as described by Ivanova Spiridonov (2017). The partial sequence of A. kimmeriense D2-D3 expansion segment of LSU rDNA was deposited in GenBank under MH627383, and partial ITS rDNA as MH627382. BLAST-search was used to identify sequences similar to these in NCBI GenBank, which were then used for comparison in phylogenetic analysis. The sequences were aligned using the Clustal_X program (Thompson et al., 1997). Subsequently, the sequences were edited using the Genedoc 2.7 program (Nicholas et al., 1997), to prepare a file for the analysis in MEGA5 (Tamura et al., 2011). Phylogenetic trees were obtained using three different methods (MP - maximum parsimony, NJ -neighbour joining and ML - maximum likelihood) and pairwise nucleotide differences were calculated.
The prevalence of Angiostoma infection was 83% (10 out of 12) and the mean intensity 6 (3-16) specimens. Morphological examination using light and scanning microscopy has shown that the discovery represents A. kimmeriense as the specimens from Adygea were similar to that in the original description in metric characteristics and general morphological traits.
DESCRIPTION
Angiostoma kimmeriense Korol Spiridonov, 1991 (Figs 1 2)
General. Body long, cylindrical, tapering to both ends, tail short. Cuticle thinly annulated. Lateral field expressed as simple thin band less than 1 ^m wide (Fig. 2G).
Anterior portion of head ca 13 ^m long and ca 25-30 ^m wide, hemi-spherical, lacking annulations, slightly offset from body contour. Tissue layer just beneath cuticle thickened (ca 3-4 ^m thick) and enforced by fine, densely packed fibres arranged perpendicularly to the body surface. Six elevations situated at the start of annulated region just beneath the margin of the non-annulated part. Each elevation bearing a salient papilla of internal circle and smaller one just posterior to it of external circle. Amphids tiny, located slightly farther back. Mouth aperture triangular, moderately sized. Stoma 4-5 ^m wide, ca 15-18 ^m long, stoma walls not sclerotised, cylindrical at anterior and expanding at bottom. Each stoma sector at bottom bearing 3 prominent cuticular denticles, one ca 3 ^m long and other two ca 1.5 ^m long. Pharynx long, finely muscular, club-like. Procorpus two thirds of pharynx length; isthmus moderately narrow, poorly marked; basal bulb as wide as procorpus. The anterior part of procorpus slightly narrower than middle and posterior parts forming a kind of a collar surrounding the posterior half of stoma. Valves in the basal bulb very fine, haustrulum present. Cardia large, conical, protruding into intestine lumen.
The position of a nerve ring varies from anterior part of isthmus to anterior part of bulb and the position of an excretory pore from bulb base to two body diameters posterior to bulb base. The distance between the nerve ring and the excretory pore is nearly constant and the more anterior position of the former corresponds with the similar position for the latter. Excretory duct ca 35-45 ^m long leading to ampoule-like chamber. Intestine with thick walls forming diverticula in some specmens.
Male. n = 4. L = 3643 ± 593 (3180-4460) ^m; a = 38.3 ± 3.7 (35.2-43.3); b = 11.5 ± 0.7 (11.1-12.4); c = 60.4 ± 6.9 (51.9-66.1); c& = 1 ± 0.2 (0.7-1.3); pharynx length = 316 ± 75 (257-400) ^m; distance from anterior extremity to nerve ring = 230 ± 34 (208-270) ^m; distance from anterior extremity to excretory pore = 305 ± 65 (262-380) ^m; tail length = 62 ± 17 (49-86) ^m; spicule length measured on arc = 202 ± 8 (191-211) ^m; spicule length measured on chord = 186 ± 13 (175-198) ^m; gubernaculum length = 59 ± 5 (54-66) ^m. Stoma
Fig. 1. Line drawings of Angiostoma kimmeriense from Adygea. A-C: head (A, female; B, male; C, juvenile female); D, E: anterior end of female with different position of excretory pore; F: female tail; G, H: male tail; I: tail of juvenile female; J: female gonad. Except I, all in lateral position. Scale bars in ^m.
Fig. 2. SEM images of Angiostoma kimmeriense from Adygea. A-C: male head; D: male tail; E: male tail, posterior part; F: posteriormost GP and phasmid; G: cuticle of mid-body region of female; H, I: tail tip of female. Arrow indicating opening of pharyngeal gland (B); reduced lip (C); phasmids (E); GP and phasmid (F); lateral field (H). Scale bars in ^m.
Bursa narrow, leptoderan. GP formula as follows 1 + 1 + 3/2 + 1 + ph, with papillae in pairs 4 5 and 6 7 very close. GP 4 5 not staggered.
Except GP 5 7, all papillae reaching margin of bursa. Both GP5 and GP7 opened on dorsal side of bursa. GP8 not incorporated into bursa and situated closely to tail tip in subventral position. Close to GP8, a pair of slightly smaller than GP8 phasmids situated in ventral position observed in SEM only (Fig. 2E F). The precloacal unpaired papillae and a pair of circumcloacal papillae present. Spicule distal tips sharply pointed with a narrow ala surrounded. Spicules strong, arcuate, manubria hardly demarcated. Spicular velum short, extending to mid-spicule length. A variation was observed in the
_ Rhabditis terrícola EF417152
. Angiostoma gland ¡cola GQ167724
-Phasmarhabditis hermaphroditei KM510193
I Phasmarhabditis sp. KXO 17483 -Phasmarhabditis sp. EU 195967
Phasmarhabditis huizhouensis KPO17253 - Phasmarhabditis sp. KP877242 Phasmarhabditis sp. KM510197 Phasmarhabditis sp. MF806606
Phasmarhabditis sp. KX267674 Phasmarhabditis sp. KM510200
-Phasmarhabditis apuliae KX017482
Angiostoma I i macis GQ 167725 Angiostoma kimmeriense MH627383
Angiostoma sp. MF192962 Angiostoma dentiferum GQ 167726 Angiostoma norvegicum KU712561 Angiostoma milacis 0949063 Angiostoma margaretae MF ! 92963 Angiostoma margaretae KU712562
Pellioditis mediterránea AM399068
i Oscheius ruga o ens is KT884891
& Oscheius necromenus KY 366264
Fig. 3. Phylogenetic relationships of Angiostoma kimmeriense based on D2-D3 LSU rDNA. Bootstrap support values are presented near nodes as MP/NJ/ML. ML analysis (500 bootstrap replications), K2 + G model. For MP and NJ - 1000 bootstrap replications.
number of precloacal papillae in a single specimen (see Fig. 1G) where papillae 2 and 3 were merged retaining two sensillar endings. Denticles on tail tip absent.
Female. n = 10. L = 5936 ± 1231 (3450-7530) ^m; a = 35 ± 3.3 (30.3-41.8); b = 14.8 ± 2.5 (9.117.9); c = 27.4 ± 3.2 (21.3-31.4); c& = 3.8 ± 0.3 (3.44.3); pharynx length = 403 ± 53 (330-490) ^m; distance from anterior extremity to nerve ring = 320 ± 56 (260-413) ^m; distance from anterior extremity to excretory pore = 415 ± 85 (317-540) ^m; V% = 48 ± 0 (42.7-53.3); tail length= 216 ± 31 (162-260) ^m. Lateral field extending to tail extremity. Stoma 18 ± 1 (17-21) ^m long and 4.3 ± 0.8 (3-5) ^m wide. Cardia ca 13 ^m long. Ovaries paired, opposed, distal parts not reversed. Developing
oocytes arranged in one row. Spermathecae located between each ovary and uterus and filled with sperm. Sperm cells in spermathecae and uteri 5-7 ^m in diam. Uteri long, filled with widely oval, thin-walled eggs on different stages of development; 64 ± 4 (57-70) ^m long and 37 ± 4 (30-42) ^m wide. Juveniles in uteri not observed. Vulva transverse slit, vulval lips not or very slightly inflated, vagina about two-third body diameter long or 134 ± 26 (87-176) ^m long. Copulatory plug present in fertilised specimens. Musculature in vulval and anal regions well developed. Rectum 77 ± 10 (64-90) ^m long, sometimes inflated; lining strongly cuticularised. Anal glands large. Phasmids papilliform, located at 35 ± 4 (30-40) ^m from tail tip. Two-four denticles on tail tip present.
Young female. n = 2. Body 2009 ^m and 2070 ^m long and 61 ^m and 78 ^m wide. Lip region less reflected than in adult female, 4 ^m and 6 ^m long. Stoma retaining rhabditoid plan with separate rhabdions, 15 ^m and 18 ^m long and 3 ^m wide; denticles in gymnostom indistinct. Pharynx 223 ^m and 280 ^m long, nerve ring at 157 ^m and 213 ^m from apex, excretory pore at 184 ^m and 291 ^m from apex. Cardia 11 ^m and 7 long ^m. Spermatheca region demarcated by morphologically different tissue connecting distal portions of ovary and uterus with distinct duct inside. Vulva at 52.2% of body length. Tail 98 ^m and 130 ^m long, phasmids at 30 ^m and 35 ^m from tail tip. Tail tip pointed without denticles.
Remarks. We observed the variations in the position of a nerve ring and excretory pore position and the correlation between a nerve ring position and an excretory pore position in our material which was not mentioned in the original description. We did not find whether such variations were correlated with the size and age of the nematode.
Specimens from Crimea and Caucasus are very similar in general body proportions and metric characteristics except for the longer spicules (average 186 ^m vs 124 ^m measured on chord) and gubernaculum (average 59 ^m vs 44 ^m) of males in the material from Caucasus. As the host preferences are similar in both cases we tend to
believe that specimens from Adygea represent A. kimmeriense. Molecular characteristics of A. kimmeriense from the type locality are yet to be obtained and compared with the Caucasian ones.
Molecular analysis and phylogenetic position of Angiostoma kimmeriense. A matrix of nucleotide data with the length of 493 bp was obtained for D2-D3 LSU rDNA with 136 characters being parsimony-informative. A matrix of nucleotide data with the length of 783 bp was obtained for ITS rDNA with 209 parsimony-informative characters. The results inferred from the analysis of D2-D3 LSU rDNA sequences revealed that EM434 strain of Phasmarhabditis sp. (deposited as EU195967) was the most similar form with the difference in only 16 bp, while the nucleotide differences with all other nematodes (including species of Angiostoma) were above 21 bp. The relationships between species of Phasmarhabditis and Angiostoma were not resolved in D2-D3 LSU rDNA analysis (Fig. 3). Similarly, the phylogenetic relationships were not resolved in the ITS rDNA analysis (Fig. 4). The lowest level of nucleotide difference in ITS rDNA was found between A. kimmeriense and two unidentified strains of Phasmarhabditis from Czech Republic (VP &185& and VP &CH1& deposited as KX017487 and KX017486, respectively).
, AncyJostoma canin urn KC755029
■ Phasmarhabditis californien KM510204 Phasmarhabditis apuliae KXO17488 ] 00/99/100 phasmarhabditis sp. KXO 17487 Phasmarhabditis sp. KXO 17486 _| Phasmarhabditis sp. KX267675
PhasmarhabditispapiUosa KM5 !0205
Angiostoma margaretae MF 192968
-Angiostoma kimmeriense MH627382
-Phasmarhabditis neopapillosa FJ516760
Phasmarhabditis hcrmaphrodita FJ516761 Phasmarhabditis hcrmaphrodita KM 510202 Phasmarhabditis hcrmaphrodita MG551718
J Phasmarhabditis bohcmica KXO 17489 Phasmarhabditis bohcmica KX017490
Fig. 4. Phylogenetic relationships of Angiostoma kimmeriense based on ITS rDNA. Bootstrap support values are presented near nodes as MP/NJ/ML. ML analysis (500 bootstrap replications), T92 + G model. For MP and NJ - 1000 bootstrap replications.
DISCUSSION
Four out of 19 species of Angiostomatidae were described from amphibian and reptile hosts: A. plethodontis Chitwood, 1933, A. onychodactyla Bursey Goldberg, 2000, A. carettae Bursey Manire, 2001 and A. lamotheargumedoi Falcon-Ordaz et al., 2008. In the absence of molecular data on these and many gastropod-associated angiostomatids we have to recourse to the comparative analysis of their morphology.
The main morphological trait differentiating these four species from the rest of the genus is the head end structure with its three distinct lips. Here, each lip bears a pair of cephalic papillae. By contrast, the head end of the majority of Angiostoma species associated with gastropods is not differentiated into lips but has 6 elevations situated closely to a mouth aperture. The elevations are reduced, flattened lips.
Among gastropod-associated species, three retain the possession of 6 lip-like structures at the head end but differ in its organisation. These are A. glandicola Ivanova Spiridonov, 2010 with 6 probolae apically bearing head sensillae and A. kimmeriense and A. zonitidis Ivanova Wison, 2009 with 6 elevations displaced from the mouth aperture posterior to the margin of a cap-like structure of a head end (Ivanova Wison, 2009; Ivanova Spiridonov, 2010). Such general 6-elements scheme of a head end is characteristic for all known gastropod-associated members of the genus. Thus, the three-lipped structure of a head of vertebrate-associated Angiostoma spp. indicates that they may belong to a different genus (or genera, considering the variable vertebrate hosts, from amphibians to reptiles). Genetic distance between two parts of the genus (or between species) can be determined only with the use of the molecular data, which are not yet available. For the time being, we suggest that A. plethodontis, A. onychodactyla, A. carettae and A. lamotheargumedoi are to be considered as species insertae sedis (Chitwood, 1933; Bursey Goldberg, 2000; Bursey Manire, 2001; Falcon-Ordaz et al., 2008).
Surprisingly, the D2-D3 28S rDNA analysis has placed A. kimmeriense close to Phasmarhabditis sp. EU 195967. Another sequence of Phasmarhabditis sp. was the closest one to A. kimmeriense in the ITS rDNA analysis. It seems that two loci used are not informative enough to resolve the relationships between these two genera. In LSU rDNA phylogram (Fig. 3) the sequences of Phasmarhabditis and Angiostoma did not form separate clades but were resolved as a united clade with the morphologically
and molecularly deviant A. glandicola from tropical gastropods as a basal form. This united Phasmarhabditis + Angiostoma clade was characterised by strong support.
Affinity of Phasmarhabditis, Angiostoma and Agfa (a parasite of salivary glands of gastropods) was shown by Ross et al. (2010) based on the analysis of 18S rDNA gene. The latter two genera have evidently undergone morphological transformations due to the parasitic way of life while Phasmarhabditis still retains the general morphological traits of free-living Rhabditidae. According to Sudhaus (2011) "There is hardly any apomorphic morphological character that can substantiate this relationship apart from GP4 and GP5 standing very close and staggered, and perhaps GP5 and GP9 opening on the dorsal surface of the bursa velum".
Considering the affinity of A. kimmeriense and, perhaps, A. zonitidis to Phasmarhabditis the following common traits should be noted: cylindrical stoma; denticles in stegostom; narrowed anterior part of corpus enveloping posterior of stoma similarly to pharyngeal collar of Phasmarhabditis, male phasmids situated close to the last pair of GP. The dissimilarities are more numerous and include the large size, head end papillae displaced, leptoderan bursa, outstretched ovaries and many more.
While the number of genital papillae in males of Phasmarhabditis is stable, the same feature in Angiostoma in general is variable (7 to 10) and differs even in the closest species of A. kimmeriense and A. zonitidis (8 vs 10). Whether the rule of &GP4 and GP5 standing very close and staggered& within Angiostoma is usually observed, the other (&GP5 and GP9 opening on the dorsal surface of the bursa velum&) is not.
The position of male phasmids within Angiostoma (Morand, 1986, 1992; Ivanova Wilson, 2009; Ivanova Spiridonov, 2010) was posterior to the last pair of genital papillae, which conforms to observations by Kiontke (1999) on Angiostoma spp. and supports the affinity with Phasmarhabditis.
On the other hand, such traits of A. kimmeriense as the large size, parasitising of adult nematodes in a host and the site of infection points to a parasitic rather than pathogenic way of life. At present, we refrain from transferring A. kimmeriense to Phasmarhabditis. Whether A. kimmeriense represents the case of emergence of parasitism within necromenic/pathogenic genus requires further research.
ACKNOWLEDGEMENTS
All SEM studies were carried out at the Joint Usage Center "Instrumental Methods in Ecology" at A.N. Severtsov Institute of Ecology and Evolution. The authors thank B. Efeykin for the help in the organisation of the collection trip. The financial support of a grant 17-04-00095a from the Russian Fund for Basic Research is appreciated.
REFERENCES
Bursey, C.R. Goldberg, S.R. 2000. Angiostoma onychodactyla n. sp. (Nematoda: Angiostomatidae) and other intestinal helminthes of the Japanese clawed salamander, Onychodactylus japonicus (Caudata: Hynobiidae), from Japan. Comparative Parasitology 67: 60-65.
Bursey, C.R. Manire, C.A. 2006. Angiostoma carettae n. sp. (Nematoda: Angiostomatidae) from the loggerhead sea turtle Caretta caretta (Testudines: Cheloniidae), Florida, U.S.A. Comparative Parasitology 73: 253-256. Chitwood, B.G. 1933. On some nematodes of the superfamily Rhabditoidea and their status as parasites of reptiles and amphibians. Journal of the Washington Academy of Science 23: 508-520. Falcón-Ordaz, J., Mendoza-Garfias, B., Windfield-Pérez, J. C., Parra-Olea, G. Pérez Ponce de Leon, G. 2008. Angiostoma lamotheargumedoi n. sp. (Nematoda: Angiostomatidae) from the intestine of Pseudoeurycea mixteca (Caudata: Plethodontidae) in central Mexico. Revista Mexicana de Biodiversidad 79: 107-112.
Holterman, M., van der Wurff, A., van den Elsen, S., van Megen, H., Bongers, T., Holovachov, O., Bakker, J. Helder, J. 2006. Phylum-wide analysis of SSU rDNA reveals deep phylogenetic relationships among nematodes and accelerated evolution towards crown clades. Molecular Biology and Evolution 23: 1792-1800.
Ivanova, E. Wilson, M. 2009. Two new species of Angiostoma (Angiostomatidae, Rhabditidae) from British terrestrial mollusks. Systematic Parasitology 74: 113-124.
Ivanova, E.S. Spiridonov, S.E. 2010. Angiostoma glandicola sp. n. (Nematoda: Angiostomatidae): a parasite in the land snail Megaustenia sp. from the Cat Tien Forest, Vietnam. Journal of Helminthology 84: 297-304.
Ivanova, E.S. Spiridonov, S.E. 2017. Phasmarhabditis meridionalis sp. n. (Nematoda: Rhabditidae) from a land snail Quantula striata (Gastropoda: Dyakiidae) from south of Vietnam.
Russian Journal of hematology 25: 129-140.
Ivanova, E.S., Panayotova-Pencheva, M.S. Spiridonov, S.E. 2013. Observations on the nematode fauna of terrestrial molluscs of the Sofia area (Bulgaria) and the Crimea peninsula (Ukraine). Russian Journal of Nematology 21: 41-49.
Kiontke, K. 1999. The nematode fauna of rotting cactus and phasmids in male Secernentea. Ph.D. Dissertation, Freie Universität Berlin, Berlin, Germany, 264 pp.
Korol, E.N. Spiridonov, S.E. 1991. Angiostoma kimmeriensis sp. n. and Agfa taurica sp. n. - parasitic Rhabditida (Nematoda) from Crimean terrestrial mollusks. Helminthologia 28: 179-182.
Morand, S. 1986. Angiostoma aspersae n. sp. (Nematoda: Angiostomatidae) parasite de Helix aspersa Müller (Gastropoda, Helicidae). Bulletin du Museum National d&Historie Naturelle 8: 111-115.
Morand, S. 1992. Angiostoma spiridonovi n. sp. (Nematoda: Angiostomatidae) from Limax flavus (Gastropoda: Limacidae). Journal of the Helminthological Society of Washington 59: 212-217.
Nicholas, K.B., Nicholas Jr., H.B. Deerfield, D.W. 1997. Multiple Sequence Alignment Editor and Shading Utility, Version 2.7.000: URL: http://www.psc.edu/ biomed /genedoc (accessed: May 21, 2018).
Pham Van Luc, Spiridonov, S.E. Wilson, M.J. 2005. Aulachnema monodelphis n. g., n. sp. and Angiostoma coloaense n. sp. (Nematoda: Rhabditida: Angiostomatidae) from terrestrial molluscs of Vietnam. Systematic Parasitology 60: 91-97.
Ross, J.L., Ivanova, E.S., Spiridonov, S.E., Waeyenberge, L., Moens, M., Nicol, G.W. Wilson, M.J. 2010. Molecular phylogeny of slug-parasitic nematodes inferred from 18S rRNA gene sequences. Molecular Phylogenetics and Evolution 55: 738-743.
Ross, J.L., Haukeland, S., Hatteland, B.A. Ivanova, E.S. 2017. Angiostoma norvegicum n. sp. (Nematoda: Angiostomatidae) a parasite of arionid slugs in Norway. Systematic Parasitology 94: 51-63.
Ross, J.L., Pieterse, A., Malan, A.P. Ivanova, E.S. 2018. Phasmarhabditis safricana n. sp. (Nematoda: Rhabditidae), a parasite of the slug Deroceras reticulatum from South Africa. Zootaxa 4420: 391-404.
Seinhorst, J.W. 1959. A rapid method for the transfer of nematodes from fixative to anhydrous glycerin. Nematologica 4: 67-69.
Sudhaus, W. 2011. Phylogenetic systematisation and catalogue of paraphyletic "Rhabditidae" (Secernentea, Nematoda). Journal of Nematode Morphology and Systematics 14: 113-178.
Tamura, K., Peterson, D., Peterson, N., Stecher, G., Nei, M. Kumar, S. 2011. MEGA5: molecular evolutionary genetics analysis using maximum
likelihood, evolutionary distance, and maximum parsimony methods. Molecular Biology and Evolution 28: 2731-2739.
Thompson, J.D., Gibson, T.J., Plewniak, F., Jeanmougin, F. Higgins, D.G. 1997. The Clustal_X windows interface: flexible strategies for multiple sequence alignment aided by quality analysis tools. Nucleic Acids Research 24: 4876-4882.
Vorobjeva, O.V., Ivanova, E.S. Leonov, S.V. 2008. [Survey of nematode fauna of terrestrial molluscs in Crimea: Preliminary results]. In: Biodiversity and Ecology of Parasites in Terrestrial and Water Coenoses. Materials of International Conference in Honour of 130th Anniversary of Academician K.I. Skrjabin. pp. 66-68, Moscow, Russia (in Russian).
Е.С. Иванова и С.Э. Спиридонов. Angiostoma или Phasmarhabditis: случай Angiostoma kimmeriense Korol Spiridonov, 1991.
Резюме. Angiostoma kimmeriense (= A. kimmeriensis) Korol Spiridonov, 1991, впервые описанную из Крыма, выделили из наземной улитки Oxyhilus sp. на западном Кавказе в Республике Адыгея и получили морфологические и молекулярные характеристики вида. В результате обсуждения морфологии видов Angiostoma Dujardin, 1845 пришли к решению рассматривать те из видов, которые ассоциированы с позвоночными, в качестве insertae sedis на основании структуры головного конца (3 vs 6 губ). Проведен филогенетический анализ на основе частичных последовательностей трех участков РНК (18S, 28S и ITS), который не смог определить взаимоотношения A. kimmeriense и других ангиостом, так как наиболее близкие последовательности по этим участкам РНК были найдены среди членов другого ассоциированного с гастроподами рода - Phasmarhabditis Andrassy, 1976. Однако такие биологические особенности A. kimmeriense как большие размеры тела, ограниченное число паразитов в хозяине и место локализации паразитов указывают скорее на паразитический образ жизни этих нематод, чем на типичные для Phasmarhabditis некромению или патогенное влияние.
