Journal of Nematology 39(3):263–274. 2007. © The Society of Nematologists 2007. of the Order from Andalucía Oriental, Spain. The Genera (Osche, 1952) Dougherty, 1953 and Cobb, 1913, with Description of P. spiculocrestata sp. n. and a Species Protorhabditis Key J. Abolafia,R.Pen˜a-Santiago Abstract: A new species of the genus Protorhabditis is described from natural areas in the SE Iberian Peninsula. Protorhabditis spiculocrestata sp. n. is distinguished by its body length 387–707 µm in females and 375–546 µm in males, lip very low and flattened, stoma 14–22 µm long, female tail conical-elongate (48–100 µm, c = 6.4–8.3, cЈ = 4.8–7.5), phasmid near anus, male tail conical (20–27 µm, c = 18.3–22.3, cЈ = 1.4–1.5), bursa peloderan closed anteriorly and bears eight papillae (1+2+1+1+3), spicules 23–26 µm long, and gubernaculum 10–16 µm long. Diploscapter coronatus is also presented. Description, measurements and illustrations, including SEM photographs, are provided. A key to species of Protorhabditis is also given as well a compendium of their measure- ments. Key words: description, Diploscapter, key, morphology, new species, Protorhabditis, Rhabditids, SE Spain, SEM, .

Rhabditid nematodes are an interesting zoological mis (Bütschli, 1873) Sudhaus, 1976, P. tristis (Hirsch- taxon. They are very abundant in all types of soil and mann, 1952) Dougherty, 1955 and D. coronatus (Cobb, sediments of freshwater bodies and play important eco- 1893) Cobb, 1913. Nevertheless, no relevant morpho- logical roles mainly as primary consumers—their free- logical or taxonomical information on them was pro- living forms display saprophagous or bacteriophagous vided in the corresponding publications. In this contri- feeding habits—but also as parasites, in particu- bution, part of the series on rhabditid species from lar enthomopathogenic forms. From a systematic point Andalucía Oriental (SE Iberian Peninsula, Spain), D. of view, rhabdits are a difficult group whose coronatus and an undescribed species of Protorhabditis classification has been matter of long and strong dis- are reported. cussions and whose diversity is far from being well known. Materials and Methods Several years ago, a research project was planned to Nematodes were extracted from soil samples by study the Iberian rhabditid fauna, focused on both its Flegg’s (1967) method and a somewhat modified Baer- morphological characterization and its taxonomy, since mann’s (1917) funnel technique. Nematodes obtained no monographic study on this matter was available. were later relaxed and killed by heat, fixed in 4% form- The genera Protorhabditis (Osche, 1952) Dougherty, aldehyde, and processed to anhydrous glycerine ac- 1953 and Diploscapter Cobb, 1913 are two rhabditid taxa cording to Siddiqi (1964). Measurements were taken characterized by having a long stoma whose stegostom using an ocular micrometer, and drawings were made lacks glottoid apparatus. They were classified by An- using a drawing tube attached to a Leica microscope; drássy (1983) in separate families ( Örley, LM pictures were made using a Nikon Eclipse 80i mi- 1880 and Diploscapteridae Micoletzky, 1922, respec- croscope provided with a digital video camera Nikon tively) of the superfamily Rhabditoidea Örley, 1880, Digital Sight DS-5M. For SEM studies, fixed specimens mainly due to morphology of lip region: six rounded were hydrated in distilled water, dehydrated in a graded lips with similar aspect in Protorhabditis; subdorsal and ethanol and acetone series, critical point dried and subventral lips hook-like and lateral lips membranous coated with gold (Abolafia and Peña-Santiago, 2005), and rounded with dentate margin in Diploscapter. How- and observed with a JEOL JSM-5800 microscope. The ever, more recent evidence based on both morphologi- terminology used for morphology of stoma and spicules cal and molecular studies demonstrated a close rela- follows the proposals by De Ley et al. (1995) and Abo- tionship between both genera (Fitch, 2000; Sudhaus lafia and Peña-Santiago (2006), respectively. and Fitch, 2001). In addition, the D. coronatus specimens examined Three species belonging to these two genera have were cultured in vitro in petri dishes (2% agar and been recorded from several localities in the Iberian 1.5% glucose) and maintained under laboratory condi- Peninsula (Abolafia and Peña-Santiago, 2001): P. filifor- tions for several months. This culture is no longer avail- able.

Received for publication July 25, 2007. Description Departamento de Biología Animal, Biología Vegetal y Ecología, Universidad de Jaén. Campus “Las Lagunillas” s/n. 23071-Jaén, Spain. The authors thank Dr. W. Sudhaus for his examination of type material of Protorhabditis spiculocrestata sp. n. Protorhabditis spiculocrestata, Research Technical Services of University of Jaén (Figs. 1–3) (Spain) for the assistance in SEM study of the material, and the financial support received from the project entitled “Fauna Ibérica VIII” (CGL2004– The specific epithet refers to the presence of a sub- 04680-C10–09 / BOS). E-mail: abolafi[email protected] dorsal crest in the spicules, a remarkable morphologi- This paper was edited by Zafar Handoo. cal feature of this species. 263 264 Journal of Nematology, Volume 39, No. 3, September 2007

mus. Excretory pore at 82–92% of neck length, at level of isthmus base or basal bulb, behind the hemi- zonid. Deirid at basal bulb level, at 90–97% of neck length. Reproductive system didelphic-amphidelphic, with anterior branch dextral and posterior branch sinistral. Ovaries having a flexure. Uterus two times the corresponding body diameter long, having thicker walls at proximal portion and distinct lumen at distal one. Eggs 15–23 × 37–39 µm. Vagina extend- ing inwards to one-third of body diameter, with thicker walls at proximal portion. Rectum 1.1–1.3 times as long as anal body diameter. Tail conical- elongate, and terminus with acute mucro. Phasmid near anus. Male (n = 3): Similar to female in general morphol- ogy. Habitus ventrad curved after fixation. Reproduc- tive system monorchic. Testis reflexed ventrally anteri- orly. Tail conical. Bursa pelodera, closed anteriorly, and with eight papillae (1+2+1+1+3). Spicules free: ma- nubrium rounded; lamina bent at its middle, with well developed dorsal hump and ventral wing at its middle; terminus ventrad curved, with acute tip and a subdorsal crest. Gubernaculum sigmoid.

Other material examined (see Table 1) Very similar to type population, but showing varia- tions in body length, shorter in the population from Almijara Mountain (387 µm in females and 395–405 µm in males), and longer in the populations from Al- FIG.1. Protorhabditis spiculocrestata sp. n. A: Neck. B: Anterior end. margen and Sierra de las Nieves (634 µm and 644–707 C: Female reproductive system. D: Entire male. E: Female posterior µm in females, respectively). end. F: Entire female. G, H: Male posterior end (lateral side). J: Idem (ventral side). I: Spicule. Diagnosis Measurements: Listed in micrometers in Table 1. Protorhabditis spiculocrestata sp. n. is distinguished by Type population collected near road from Fuente de Piedra its body length 387–707 µm in females and 375–546 µm to Alameda, province of Málaga in males, lips very low and flattened, stoma 14–22 µm, Female (n = 7): Small nematodes, 441–561 µm long. female tail conical-elongate (48–100 µm, c = 6.4–8.3, Body cylindrical, tapering towards both ends, more cЈ = 4.8–7.5), phasmid near to anus, male tail conical so posteriad. Habitus rather sigmoid after fixation. Cu- (20–27 µm, c = 18.3–22.3, cЈ = 1.4–1.5), bursa ticle with distinct transverse striations or annuli, about peloderan closed anteriorly and bears eight papillae 1 µm wide at midbody. Lateral field with five inci- (1+2+1+1+3), spicules 23–26 µm long, and gubernacu- sures or four ridged wings at midbody, two at posterior lum 10–16 µm long. region which fade out near anus. Lip region with six small lips, more or less rounded; papillae visible. Stoma Relationships rhabditoid, lacking glottoid apparatus. Cheilostom The new species resembles Protorhabditis oxyuroides with weakly refractive walls. Buccal ring (membrane Sudhaus, 1974, although it can be distinguished from it wedge ring) lower in longitudinal section. Buccal prism by its having lips lower and reduced (vs. more promi- with straight walls, very long and narrow. Pharyngeal nent), stoma shorter (14–22 µm vs. 21–27 µm), and collar very short. Procorpus cylindrical. Metacorpus spicules with different morphology (vs. terminus lack- swollen, almost ovoid, with its inner walls more re- ing subdorsal crest). fractive. Isthmus almost as long as procorpus. Basal bulb small, ovoid or pyriform. Cardia conoid to hemi- Type locality and habitat spherical, surrounded by intestinal tissue. Intestine lacking distinct specializations, but a cardiac portion Los Carvajales, near the road from Alameda to Fu- is sometimes differentiated at its anterior part. Nerve ente de Piedra, province of Málaga, Spain; GPS coor- ring at 71–80% of neck length, surrounding the isth- dinates: N 37°8.0Ј, E 4°43.0Ј. Soil in a Mediterranean Spanish Rhabditida: Abolafia and Peña-Santiago 265

FIG.2. Protorhabditis spiculocrestata sp. n. (LM). A: Neck. B: Female reproductive system (anterior branch). C: Female tail. D. Male posterior end (bursa view). E. Male posterior end (spicule view). plant community whose dominant species are Olea eu- carpa (L.) Boiss., Phlomis purpurea L., Thymus sp., Quer- ropaea L. var. sylvestris Brot., Daphne gnidium L., Rosmari- cus coccifera L., and Ulex parviflorus Pourret. Collected nus officinalis L., Cistus monspeliensis L., Retama sphaero- on 19 October 1991. 266 Journal of Nematology, Volume 39, No. 3, September 2007

FIG.3. Protorhabditis spiculocrestata sp. n. (SEM). A: Lip region (arrow indicates amphid opening). B: Lateral field at middle body. C: idem at posterior end. D: idem near to anus. E: Phasmid. F, G: Male posterior end.

Other localities and habitats road to Canillas de Aceituno, Periana, road from Ar- chidona to Villanueva del Trabuco, Sierra de Mijas, and The species has also been collected from several lo- Sierra de las Nieves Natural Park (province of Málaga). calities in the SE Iberian Peninsula: Sierra de Guilli- It has been found associated with the rhizosphere of mona (province of Granada); Sierras de Cazorla, Se- several wild plant communities whose dominant species gura and Las Villas Natural Park (province of Jaén); were Pinus nigra Arn. var. salzmannii (Dunal) Franco, Spanish Rhabditida: Abolafia and Peña-Santiago 267

TABLE 1. Measurements (in µm) of females (n = 17) and males (n = 11) of Protorhabditis spiculocrestata sp. n. presented as mean ± s.d. and (range).

Road from Archidona Road from Fuente de Road to Canillas to Villanueva Sierra de Locality Piedra to Alameda de Aceituno del Trabuco Periana Almargen las Nieves Ma´laga Ma´laga Ma´laga Ma´laga Ma´laga Ma´laga Province Brushwood Holm oak tree Walnut tree Eucalyptus Pine Brushwood Habitat n Holotype & Paratypes 6&& Paratypes 3(( & 2(( 4&& &(& 4(( 3&& (

Body length 540 511.0 ± 43.8 502.3 ± 52.6 387 395, 405 528.3 ± 47.6 562 375 634 450.5 ± 37.8 644–707 518 (441–561) (444–546) (488–597) (421–506) a 21.6 23.7 ± 3.0 20.9 ± 1.0 20.4 21.4, 24.5 26.4 ± 1.4 22.5 20.3 ? 22.4 ± 1.5 22.3–24.8 24.7 (19.2–27.9) (20.2–21.7) (25.3–28.4) (20.2–23.4) b 4.5 4.4 ± 0.1 4.2 ± 0.5 3.6 3.3, 3.9 4.6 ± 0.6 4.6 ? 5.1 4.2 ± 0.1 4.4–5.2 4.4 (4.3–4.4) (3.9–4.6) (4.2–5.5) (4.0–4.3) c 6.5 7.1 ± 0.4 20.8 ± 2.1 8.1 19.8, 19.8 7.2 ± 0.5 8.0 18.3 8.3 21.1 ± 0.7 6.9–8.0 21.1 (6.4–7.6) (19.3–22.3) (6.5–7.7) (20.5–22.0) cЈ 6.0 5.9 ± 0.4 1.4 ± 0.1 4.8 1.4, 1.5 6.7 ± 0.5 5.6 1.5 5.4 1.4 ± 0.1 5.9–6.6 1.4 (5.3–6.4) (1.4) (6.3–7.5) (1.4–1.5) V 53 54.1 ± 1.8 — 56 — 53.4 ± 4.3 53 — 56 — 52–54 — (51–56) (47–56) Lip region 8 7.6 ± 0.9 7.0 ± 0.0 7 7 7.0 ± 0.0 8 7 7 7.0 ± 0.7 8–9 7 width (7–9) (7) (7) (7–8) Stoma 18 18.6 ± 0.6 18.5 ± 0.9 18 19, 20 18.1 ± 0.6 17 15 14 16.5 ± 1.1 20–22 19 (18–20) (18–20) (18–19) (15–18) Pharyngeal 52 53.3 ± 2.6 50.3 ± 2.3 48 48, 42 48.9 ± 1.3 51 ? 52 48.3 ± 1.4 63–65 50 corpus (51–56) (48–53) (48–50) (48–50) Isthmus 31 30.1 ± 3.3 31.0 ± 0.9 30 32, 29 29.3 ± 1.9 30 ? 31 27.7 ± 3.1 32–39 29 (29–35) (30–32) (27–32) (25–31) Basal bulb 20 18.8 ± 1.1 18.7 ± 0.6 17 18 18.9 ± 1.2 21 17 18 16.0 ± 1.1 22–25 21 (18–21) (18–19) (18–21) (15–18) Pharynx length 120 121.3 ± 8.1 117.7 ± 3.2 108 120, 104 114.3 ± 7.5 122 ? 125 109.7 ± 6.7 135–146 119 (114–130) (114–120) (109–125) (104–117) Nerve ring-ant. 92 88.3 ± 5.0 86.5 ± 4.3 73 80, 79 81.4 ± 2.9 87 72 81 78.9 ± 4.3 105–110 86 end (84–96) (82–91) (78–85) (74–85) Excretory pore- 110 99.7 ± 0.6 102.0 ± 8.5 84 93, 87 ? 96 78 ? 95.3 ± 11.0 120–126 ? ant. end (99–100) (96–108) (88–103) Deirid-ant. end 116 108 116.0 ? 106, ? ? 108 ? ? 90.0 ? ? (n=11) (n=1) (n=1) Annuli width 1 1 ± 0.0 1.0 ± 0.0 1 1 1 ± 0.0 1 1 1 1 ± 0.0 ? ? (1) (1) (1) (1) Cuticle thickness 1 1.0 ± 0.0 1.0 ± 0.0 1 1 1 ± 0.0 1 1 2 1.0 ± 0.0 1 2 (1) (1) (1) (1) Body width; neck base 23 20.8 ± 2.1 21.3 ± 1.9 18 18, 16 18.8 ± 1.0 23 18 24 19.0 ± 2.0 25–27 22 (18–24) (20–24) (18–20) (18–22) midbody 25 21.8 ± 2.5 23.2 ± 3.4 19 19, 17 20.0 ± 1.1 25 19 ? 20.3 ± 3.2 26–31 21 (18–24) (21–27) (19–21) (18–25) anus 14 12.3 ± 0.9 16.8 ± 1.0 10 14 11.0 ± 0.8 13 14 14 14.8 ± 1.0 14–15 18 (11–14) (16–18) (10–12) (14–16) Vagina 10 8.6 ± 0.8 — 7 — 7.4 ± 0.8 8 — ? — 10 — (8–10) (7–8) Anterior ovary/ 51 47.3 ± 5.6 77.5 ± 22.3 40 74, 93 39.7 ± 5.0 65 80 38 82.1 ± 9.2 62–66 95 testis (42–58) (62–103) (34–44) (70–91) Anterior genital 110 100.3 ± 16.9 307.0 ± 52.0 67 205, 222 98.0 ± 7.0 110 205 82 271.8 ± 31.7 138–157 308 branch (85–125) (255–359) (93–106) (241–315) Posterior ovary 56 46.0 ± 5.8 — 35 — 40.0 ± 5.7 56 — 43 — 44–56 — (35–50) (36–44) Posterior genital 110 98.3 ± 16.6 — 63 — 87.0 ± 19.8 105 — 132 — 115–145 — branch (77–121) (73–101) Rectum 15 15.5 ± 1.0 ? 15 ? 12.9 ± 1.4 15 ? 18 ? 18–20 ? (14–17) (12–15) Tail 84 72.3 ± 5.9 24.8 ± 2.0 48 20, 21 74.0 ± 6.3 70 21 76 21.4 ± 2.1 85–100 25 (66–80) (23–27) (67–82) (20–25) Vulva-anterior 286 277.2 ± 30.7 — 216 — 280.8 ± 8.6 298 — 353 — 348–379 — end (224–311) (270–291) Vulva-anus/tail 2.0 2.2 ± 0.1 — 2.6 — 2.3 ± 0.4 2.8 — 2.7 — 2.3–2.8 — (2.1–2.4) (1.9–2.9) Spicules — — 24.7 ± 1.5 — 23, 24 — — 23 — 23.8 ± 1.0 — 25 (23–26) (23–25) Gubernaculum — — 11.8 ± 0.8 — 10, 12 — — 11 — 10 ± 0.0 — 16 (11–13) (10) 268 Journal of Nematology, Volume 39, No. 3, September 2007

Pinus pinaster Aiton, Quercus rotundifolia Lam., Q. coccif- Population from agar culture comes from specimens collected era L., Juglans regia L., Retama sphaerocarpa (L.) Boiss., in Carchuna, province of Granada Juniperus oxycedrus L., Ulex sp., Cistus sp., Nerium oleander L. and Eucalyptus sp. Female (n = 20): Small nematodes, 358–504 µm long. Habitus ventrad curved after fixation. Cuticle annu- Type material lated; annuli 1–2 µm wide at midbody. Lateral field occupying 14–22% of the midbody diameter, with four Six females (holotype and paratypes) and three incisures under LM and two separated wings under males (paratypes) deposited in Departamento de Bi- SEM which fade out before to anus level. Lip region ología Animal, Biología Vegetal y Ecología, Universi- with six lips: the subdorsal and the subventral are very dad de Jaén, Spain; and one female (paratype) depos- refractive and hook-like (4–5 µm high); the lateral lips ited in the nematode collection of the Swedish Museum are membranous (termed laciniae), high and with den- of Natural History, Stockholm (Sweden). tate margin, not easily observed under LM since only the central triangular part is more refractive. Stoma Diploscapter coronatus (Cobb, 1893) Cobb, 1913 rhabditoid. Buccal prism with straight walls, very long syn. coronata Cobb, 1893; Rhabditis bicornis and narrow. Stegostom lacking glottoid apparatus. Pha- Zimmermann, 1898; Diploscapter bicornis ryngeal corpus well demarcated, slightly longer than (Zimmermann, 1898) Goodey, 1963; Rhabditis isthmus and basal bulb together; procorpus cylindrical cephaloides Stefan´ski, 1922; Acrobeles armatus Kreis, and metacorpus swollen, almost as long as wide. Basal 1929. bulb spheroid, with well developed valves at its middle. (Figs. 4–6) Cardia hemispherical, surrounded by intestinal tissue. Intestine lacking any peculiar differentiation. Nerve Measurements: Listed in micrometers in Table 2. ring at isthmus level, at 70–77% of neck length. Excre- tory pore at 77–92% of neck length, at level of posterior portion of isthmus or basal bulb. Deirid at basal bulb level, at 81–97% of neck length. Reproductive system didelphic-amphidelphic. Anterior genital branch dex- tral (on the right side of intestine) and posterior branch sinistral (on the left). Ovaries with a flexure, rarely straight. Uterus short, less than the correspond- ing body diameter long. Vagina with thin walls, extend- ing inwards one-third of body diameter. Rectum 0.8– 2.0 times anal body diameter long. Tail conical- elongate to filiform, ending in acute tip. Phasmid at 11–12% of tail length. Male: Not found.

Other material examined (see Table 2)

Very similar to population from Carchuna, although having smaller body (273 µm long) and pharyngeal corpus shorter than isthmus-basal bulb.

Distribution

The species has been collected in six localities: (i) Vélez-Málaga, province of Málaga, in association with Persea americana Miller; (ii) La Vidriera, Guillimona Mountain, province of Granada, in association with Quercus rotundifolia Lam. and Pinus sp.; (iii) Santa Fé, province of Granada, in association with Populus alba L.; (iv) Desfiladero de los Gaitanes, province of Málaga, in association with Citrus limon (L.) Burm. fil.; (v) Car- chuna, province of Granada, in association with Lyco- persicum esculentum Miller.; and (vi) La Caldera pool, FIG.4. Diploscapter coronatus (Cobb, 1893) Cobb, 1913 (female). A: Neck. B: Anterior end. C: Entire female. D: Reproductive system. E: Sierra Nevada National Park (3,000 m height), prov- Posterior end. ince of Granada. Spanish Rhabditida: Abolafia and Peña-Santiago 269

FIG.5. Diploscapter coronatus (Cobb, 1893) Cobb, 1913 (LM). A: Neck. B: Female reproductive system. C: Lip region (lacinia level). D: Female tail. E. Lateral field.

Remarks scriptions by Paetzold (1958), Loof (1964), Andrássy The material examined agrees well with the re- (1968) and Bongers (1988), although the specimen col- description by Eyualem et al. (1998) and previous de- lected in Sierra de Guillimona is smaller (273 vs. 317– 270 Journal of Nematology, Volume 39, No. 3, September 2007

FIG.6. Diploscapter coronatus (Cobb, 1893) Cobb, 1913 (SEM). A–C: Lip region (A: lateral view. B: frontal view. C: ventral view). D: Lateral field. E–F: Female tail (E: lateral view. F: ventral view).

480 µm). Our material is also very similar to popula- greater tail width range, c value, and cЈ value (54–80 tions studied by Tahseen et al. (2002), although having µm, c = 5.1–7.4, cЈ = 4.4–6.8 vs. 42–65 µm, c = 5–8, cЈ = greater body length range (273–504 vs. 300–491 µm), 5–8, respectively). Spanish Rhabditida: Abolafia and Peña-Santiago 271

TABLE 2. Measurements (in µm) of females (n = 21) of Diploscapter Comments on the genus Protorhabditis (Osche, coronatus (Cobb, 1893) Cobb, 1913 presented as mean ± s.d. and (range). 1952) Dougherty, 1953 syn. Rhabditis (Protorhabditis) Osche, 1952 Carchuna Granada Guillimona Mountain Granada Tomato Holm oak tree Locality Province Paradoxorhabditis Khera, 1971 op. Andrássy (2005) Habitat n 20&& &

Body length 437.6 ± 39.5 273 Andrássy (2005) has provided an emended diagnosis (358–504) of Protorhabditis as well a list of its species. Sudhaus a 16.7 ± 1.0 22.8 (1991) transferred the type and only species of the ge- (14.9–18.0) nus Paradoxorhabditis Khera 1971, P. paradoxa Khera, b 4.7 ± 0.3 4.0 (4.0–5.3) 1971, to Rhabditis, but did not formally propose the c 6.4 ± 0.5 5.1 synonymy of both genera, although very recently, An- (5.7–7.4) drássy (2005) proposed its synonymy with Protorhabditis. Ј c 5.8 ± 0.7 6.8 Paradoxorhabditis is characterized by, among other fea- (4.4–6.8) V/T 50.7 ± 1.9 53 tures, the absence of glottoid apparatus and fits well the (48–57) diagnosis of Protorhabditis, a fact that justifies the pro- Lip region: high 4.0 ± 0.0 4 posed synonymy by Andrássy. The presence of pseudo- (4) peloderan bursa in P. paradoxa is a peculiar feature of width 9.7 ± 0.5 9 (9–11) this species, which is herein regarded as intrageneric Stoma 18.3 ± 1.4 18 variability, but that should be incorporated in the diag- (16–22) nosis of Protorhabditis. Pharyngeal corpus 40.0 ± 2.6 25 (35–47) The genus includes hitherto 13 species plus two spe- Isthmus 18.1 ± 1.7 14 cies inquirendae or incertae sedis (see Table 3): (15–22) Basal bulb 17.0 ± 1.0 12 Type species: (15–19) Protorhabditis xylocola (Körner in Osche, 1952) Dough- Pharynx length 95.3 ± 5.0 68 erty, 1953 (90–109) Nerve ring-ant. end 70.5 ± 4.0 52 syn. Rhabditis (Protorhabditis) xylocola Körner in Osche, (66–85) 1952 Excretory pore-ant. 78.0 ± 9.0 ? end (69–100) Other species: Deirid-ant. end 81.3 ± 4.6 ? P. cervi (Andrássy, 1985) Sudhaus, 1991 (75–89) Annuli width 1.2 ± 0.3 ? syn. cervi Andrássy, 1985 (1–2) P. elaphri (Hirschmann, 1952) Dougherty, 1955 Cuticle thickness 1.0 ± 0.1 1 syn. Rhabditis (Protorhabditis) elaphri Hirschmann, (1–2) 1952 Body width: neck base 24.3 ± 3.1 14 P. filiformis (Bütschli, 1873) Sudhaus, 1976 (20–30) syn. Rhabditis filiformis Bütschli, 1873 midbody 26.3 ± 3.1 12 Rhabditis (Choriorhabditis) filiformis (Bütschli, 1873) (20–32) anus 12.0 ± 1.7 8 Osche, 1952 (9–17) Rhabditis agilis von Linstow, 1876 Lateral field 4.8 ± 0.7 ? Protorhabditis lengerkeni Paezotld, 1958 (4–6) P. macrovelata Sudhaus, 1974 Vagina 7.3 ± 0.8 ? (7–9) P. oxyuroides Sudhaus, 1974 Anterior ovary 59.1 ± 11.5 ? syn. Rhabditis oxyuris apud Bütschli, 1873, nec Claus, (37–80) 1862 Anterior genital 83.3 ± 14.1 31 P. paradoxa (Khera, 1971) Andrássy, 2005 branch (57–106) Posterior ovary 60.1 ± 15.3 ? syn. Paradoxorhabditis paradoxa Khera, 1971 (37–83) Rhabditis (Rhabditis) paradoxa (Khera, 1971) Posterior genital 84.5 ± 18.6 33 Sudhaus, 1991 branch (59–130) P. parvovelata (Körner in Osche, 1952) Dougherty, 1955 Rectum 14.9 ± 3.4 9 (10–24) syn. Rhabditis (Protorhabditis) parvovelata Körner in Tail 68.8 ± 5.9 54 Osche, 1952 (55–80) P. postneri (Körner in Osche, 1952) Dougherty, 1955 Vulva-anterior end 221.7 ± 21.7 144 syn. Rhabditis (Protorhabditis) postneri Körner in Osche, (182–260) Phasmid-anus 8.3 ± 0.4 ? 1952 distance (8–9) P. ruehmi (Körner in Osche, 1952) Dougherty, 1955 Vulva-anus/tail 2.1 ± 0.3 1.4 syn. Rhabditis (Protorhabditis) ruehmi Körner in Osche, (1.6–2.6) 1952 272 Journal of Nematology, Volume 39, No. 3, September 2007

P. spiculocrestata sp. n.

(µm) P. tristis (Hirschmann, 1952) Dougherty, 1955 Spicules syn. Rhabditis (Protorhabditis) tristis Hirschmann, 1952 P. virgo (Körner in Osche, 1952) Dougherty, 1955 syn. Rhabditis (Protorhabditis) virgo Körner in Osche, 1952 arrangement Bursal papillae Species inquirendae and/or incertae sedis: P. lepida (Kreis, 1930) Sudhaus, 1976 syn. Rhabditis lepida Kreis, 1930 Rhabditis (Choriorhabditis) lepida Kreis, 1930 (Osche, 1952) Rhabditis elegans apud Kreis, 1929, nec Maupas, 1899 P. minuta (Cobb, 1893) Dougherty, 1955 syn. Rhabditis minuta Cobb, 1893

Notes on some species

(µm) Bursa The material identified and described as Rhabditis Female tail filiformis by Cobb (1893) certainly does not belong to this species because of its rounded lips with acute setae, monodelphic-prodelphic female reproductive system and vulva very posterior close to anus. It better fits the Pharyngeal medial bulb diagnosis of (Osche, 1952) Dougherty, 1953, but further studies are needed to clarify its iden- (µm)

Stoma tity. length Some doubts persist on the true identity of Rhabditis agilis von Linstow, 1876. It was considered (Sudhaus, 1976; Andrássy, 1983) to be a junior synonym of P. filiformis (Bütschli, 1873) Sudhaus, 1976. However,

Ј there are some remarkable differences between both c taxa, among others, shorter female tail (c = 5.5–6.0 vs. (female) V c = 3.2–4.0) and spicules (26 vs. 10 µm long), and they might not be identical. c P. lepida (Kreis, 1930) Sudhaus, 1976 (= Rhabditis el- (female) egans apud Kreis, 1929) is herein regarded as species inquirendae (cf. Andrássy, 1983), since the original de-

b scription lacks many morphological details. Very re-

(female) cently, Andrássy (2005) considered it to be a valid spe- cies, but it is better to maintain it as species inquiren- dae. a The original description of Rhabditis minuta Cobb, (female) species. 1893 is very poor in diagnostic features, and its true identity might not be established with accuracy. Thus, it should be maintained as species incertae sedis (cf. An- Male (mm) drássy, 1983). body length Protorhabditis

Key to Protorhabditis species identification: 1. Female tail longer (cЈ = 10–20) 2 — 0.49–0.57 — — — 3.0–4.0 ? 22–25 Yes — Pelodera (open)------8 (2+4+2) 16–21 0.46 — 30 4.8 5.5 7.5* 59 13 Yes ? — — — (mm) Female Female tail shorter (cЈ less than 8) 5

0.56–0.680.63–0.92 0.57–0.600.44–0.96 0.54–0.69 22–25 0.35–0.59 20–280.59–0.87 4.5–4.7 22–350.90–0.97 5.7–7.1 0.32–0.72 8.3–9.00.58–0.87 4.2–7.1 0.64–0.67 4.0–9.00.63–0.87 17–21 5.0–5.4 0.52–0.64 3.2–4.30.60–0.75 0.63–0.72 10–140.38–0.71 54–56 24 4.3–5.9 20–24 0.54–0.66 10–130.45–0.55 20–23 43–51 16–22 0.37–0.55 6.7–10.80.43–0.53 4.0–5.9 42–48 17–21 5.0–5.1 0.27–0.30 20–220.52–0.68 4.3–5.3 19–28 5.0 15–17 11–30 3.7–4.0 Yes 4.4–4.9 22–27 0.40–0.58 7.8–13.5 — 3.6–5.5 No 8.0–9.0 1.5–4.0 53–56 15–20 3.0–4.0 4.4–6.2 Yes 19–22 6.4–8.3 21–27 68–76 56–62 56–63 3.0–4.0 3.3–4.5 22–26 4.6–5.7 42 4.8–7.5 86–175 54–58 18 18* 125–233 Pelodera (closed) 9.6–13.2 12–14 4.3–5.5 47–56 Yes 18–20 Pelodera (open) 21 Pelodera 3.0–4.0 (closed) 14–22 46–48 6–9 Yes 54–58 Yes 17–20 Yes 18–20 Yes Yes ? 4.0–5.0 7 8 or (1+4+3) 9 (3+3+3) Yes 53–57 8 Yes (1+4+3) ? ? Pelodera 48–100 22–24 (closed) 15 17–21 ? Pelodera 10–16 (open) Pelodera ? (open) Pelodera (open) Pelodera (open) ? Yes Pseudopelodera (open) Pelodera 8 (open) (1+3+1+3) Pelodera (open) 13–28 7 8 (3+3+1) (1+3+1+3) ? 8 (2+3+3) 8------(2+2+2+2) 8 23–26 (2+1+2+3) 25–27 23–30 22–34 24–29 Pelodera 9 (open) (2+4+3) 8 (2+1+3+2) 21–29 13 — — body length 2. Pharyngeal collar very long, occupying most of the length of the stoma; pharyngeal corpus cy- lindrical (metacorpus not swollen) ------elaphri 3. Morphometrics of Pharyngeal collar short or absent (shorter than Species ABLE stoma width); pharyngeal corpus with metacor- T * Measurements from drawings.

P. cervi P. elaphri P. filiformis P. lepida P. macrovelata P. oxyuroides P. paradoxa P. parvovelata P. postneri P. ruehmi P. spiculocrestata P. tristis P. virgo P. xylocola pus swollen ------3 Spanish Rhabditida: Abolafia and Peña-Santiago 273

3. Bursa bearing 9 pairs of papillae ------tristis Baermann G. 1917. Eine einfache Methode zur Auffindung von Bursa bearing 7–8 pairs of papillae 4 Ankylostomum (Nematoden) Larven in Erdproben. Geneeskunding ------Tijdschrift voor Nederlandsch-Indië 57:131–137. 4. Cuticle lacking longitudinal striae; female tail Ј Bongers, T. 1988. De nematoden van Nederland. Stichting Uit- shorter (c = 10–12); bursa anteriorly closed, geverij Koninklijke Nederlandse Natuurhistorische Vereniging. pelodera ------filiformis Bütschli, O. 1873. Beiträge zur Kenntnis der freilebenden Nema- Cuticle longitudinally striated; female tail toden. Nova Acta der Kaiserlich-Leopoldinisch-Carolinische Deut- longer (cЈ = 15–20); bursa anteriorly open, schen Akademie der Naturforscher 36:1–124. Claus, C. 1862. Über einige im Humus lebende Anguillulinen. pseudopelodera, with a short and fine terminal Zeitschrift für Wissenschaftliche Zoologie 12:354–359. filament ------paradoxa Cobb, N. A. 1893. Nematode worms found attacking sugar-cane. 5. Bursa anteriorly open------6 Agricultural Gazette of New South Wales 4:808–833. Bursa anteriorly closed ------11 Cobb, N. A. 1913. New nematode genera found inhabiting fresh 6. Body length less than 500 µm ------virgo water and non-brackish soils. Journal of Washington Academy of Sci- Body length more than 500 µm 7 ences 3:432–433. ------De Ley, P., Van de Velde, M. C., Mounport, D., Baujard, P., and 7. Bursa with scarcely developed velum, in lateral Coomans, A. 1995. Ultrastructure of the stoma in , view not exceeding the ventral margin of body - and Rhaditidae, with a proposal for a revised stoma ------parvovelata terminology in Rhabditida (Nematoda). Nematologica 41:153–182. Bursa with well developed velum, in lateral view Dougherty, E. C. 1953. The genera of the subfamily Rhabditinae Micoletzky, 1922 (Nematoda). Thaper Commemoration Volume:69– exceeding the ventral margin of body ------8 76. 8. Lips having acute tines ------ruehmi Dougherty, E. C. 1955. The genera and species of the subfamily Lips lacking acute tines ------9 Rhabditinae Micoletzky, 1922 (Nematoda). A nomenclatorial analy- 9. Spicule length 16–21 µm ------macrovelata sis–including an addendum on the composition of the family Rhab- ditidae Örley, 1880. Journal of Helminthology 29:105–152. Spicule length 21–34 µm ------10 Eyualem, A., Karegar, A., Nabil, H. and De Ley, P. 1998. A re- 10. Inner (perioral) region of lips peculiarly el- description and ultrastructural study of Diploscapter coronatus evated; cheilorhabdia divergent anteriorly ------(Cobb, 1893) Cobb, 1913 from Ethiopia and Iran. Russian Journal of ------postneri Nematology 6:17–22. Inner (perioral) region of lips as usual, not pe- Fitch, D. H. A. 2000. Evolution of “Rhabditidae” and the male tail. culiarly elevated; cheilorhabdia not divergent Journal of Nematology 32:235–244. Flegg, J. J. M. 1967. Extraction of Xiphinema and Longidorus spe- anteriorly ------xylocola cies from soil by a modification of Cobb’s decanting and sieving 11. Pharyngeal collar well developed, occupying technique. Annals of applied Biology 60:429–437. more than half of the stoma length ------cervi Goodey, T. 1963. Soil and freshwater nematodes. 2nd ed. rewritten Pharyngeal collar very short or absent (shorter by J. B. Goodey. London: Methuen & Co. than stoma width) Hirschmann, H. 1952. Die Nematoden der Wassergrenze mittel- fränkischer Gewässer. Zoologische Jahrbücher - Abteilung für Sys- 12. Lip region narrower than adjacent part of neck; tematik, Ökologie und Geographie der Tiere 81:313–364. stoma 14–22 µm long; spicules bearing subdor- Khera, S. 1971. Nematodes from the banks of still and running sal crest ------spiculocrestata waters. XI. Subfamily Rhabditinae. Indian Journal of Nematology 1: Lip region wider than adjacent part of neck; 237–243. stoma 21–27 µm long; spicules lacking subdor- Kreis, H. A. 1929. Freilebende terrestrische Nematoden aus der Umgebung von Peking (China). I. Zoologischer Anzeiger 84:283– sal crest ------oxyuroides 294. Kreis, H. A. 1930. Freilebende terrestrische Nematoden aus der Literature Cited Umgebung von Peking (China). II. Zoologischer Anzeiger 87:67–87. Abolafia, J., and Peña Santiago, R. 2001. Rhabditid species (Nema- Linstow, O. F. B. von 1876. Helminthologische Beobachtungen. toda, Rhabditida) recorded in Peninsular Spain and Balearic Islands. Archiv für Naturgeschichte 42:1–18. Graellsia 57:113–131. Loof, P. A. A. 1964. Free-living and plant-parasitic nematodes from Abolafia, J., and Peña-Santiago, R. 2005. Nematodes of the order Venezuela. Nematologica 10:201–300. Rhabditida from Andalucía Oriental: Pseudacrobeles elongatus (de Man, Maupas, E. F. 1899. La mue et l’enkystement chez les nématodes. 1880) comb. n. Nematology 7:917–926. Archive de Zoologie Expérimental and Génetique 7:563–628. Abolafia, J., and Peña-Santiago, R. 2006. 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Siddiqi, M. R. 1964. Studies on Discolaimus spp. (Nematoda: Dory- (Nematoda: Rhabditidae) discovered between 1976 and 1986. Nema- laimidae) from India. Zeitschrift für Zoologische Systematik und Evolu- tologica 37:229–236. tionsforschung 2:174–184. Sudhaus, W., and Fitch, D. 2001. Comparative studies on the phy- Sudhaus, W. 1974. Zur Systematik, Verbreitung, Ökologie und Bi- logeny and systematics of the Rhabditidae (Nematoda). Journal of ologie neuer und wenig bekannter Rhabditiden (Nematoda). 1. Teil. Nematology 33:1–70. Zoologische Jahrbücher (Systematik) 101:173–212. Tahseen, Q., Siddiqi, M. R., and Rowe, J. 2002. Study on species of Sudhaus, W. 1976. Nomenklatorische Bemerkungen über Arten Diploscapter Cobb (Nematoda: Rhabditida) from India including D. und Gattungen der Unterfamilie Rhabditinae sensu lato (Rhabditidae, indicus sp. n. International Journal of Nematology 12:183–188. Nematoda). Nematologica 22:49–61. Zimmermann, A. W. P. 1898. De nematoden der koffiewortels. Sudhaus, W. 1991. Check list of species of Rhabditis sensu lato Deel I. Mededelingen’s Lands Plantentuin Buitenzorg 27:1–64.