J. Helminthol. Soc. Wash. 59(1), 1992, pp. 96-110 The Genus Bitylenchus Filip'ev, 1934 (Nematoda: Tylenchida) with Descriptions of Two New Species from Spain A. GOMEZ-BARCiNA,1 M. R. SiDDiQi,2 AND P. CASTILLO3 1 Centre de Investigation y Desarrollo Agrario, Camino de Purchil s/n, Apartado 2027, 18080 Granada, Spain, 2 CAB International Institute of Parasitology, 395 A Hatfield Road, St. Albans, England AL4 OXU, and 3 Institute de Agronomia y Protection Vegetal, Apartado 3048, 14080 Cordoba, Spain ABSTRACT: The taxonomic position of the genus Bitylenchus Filip'ev, 1934, is discussed and its diagnosis emended. Two new species, B. serranus sp. n. and B. pratensis sp. n., from southeastern Spain are described and illustrated using light and scanning electron microscopy (SEM). Bitylenchus serranus sp. n. is characterized by a hemispherical lip region with 6-8 annuli, stylet 21 /^m long, lateral field with regularly areolated outer bands, a short postanal intestinal sac extending up to one-fourth of the tail length, and a broadly conoid tail usually ventrally curved (about 90% of specimens studied). Bitylenchus pratensis sp. n. is characterized by a subcylindrical female tail regularly tapering to a rounded and striated terminus and a basal esophageal bulb as long as or longer than the isthmus. Several populations of B. maximus (Allen, 1955) Siddiqi, 1986, from Scotland, Spain, Trinidad, and U.S.A. are described and compared. The Spanish population is characterized by the presence of males and inseminated females; SEM observations are provided. A population of B. huesingi (Paetzold, 1958) Siddiqi, 1986, is described. A table with the morphometric data of each species is included. KEY WORDS: Bitylenchus huesingi, Bitylenchus maximus, Bitylenchus pratensis sp. n., Bitylenchus serranus sp. n., plant-parasitic nematodes, SEM observations, Spain, taxonomy. During a nematode survey on several plant Description communities in southeastern Spain, 2 new and 2 known species of the genus Bitylenchus Filip'- Bitylenchus serranus sp. n. ev, 1934 (Siddiqi, 1986), were collected. Females (Table 1, Figs. 1-14) and males of the 4 species are described herein HOLOTYPE (female in glycerine): L = 883 /xm; as B. serranus sp. n., B. pratensis sp. n., B. maxi- a = 32.7; b = 6.1; b, = 10.3; V = 53; c = 17.3; mus (Allen, 1955) Siddiqi, 1986, and B. huesingi c' = 2.4; stylet = 22 jtm; m = 53; S = 1.5; O = (Paetzold, 1958) Siddiqi, 1986. Morphology of 9; tail annuli = 40. 3 female populations of B. maximus from Trin- PARATYPE FEMALES (N= 36): Morphometrics idad, U.S.A., and Scotland is reported. Mor- are given in Table 1. Body moderately thin, phometry and morphology using SEM obser- slightly curved ventrally upon relaxation. Body vations are also given. annuli opposite mid-esophagus 1-2 and opposite mid-body 1 wide. Lateral field with 4 lines, (8- Materials and Methods 11) wide at mid-body, and outer bands areolated. Lip region hemispherical, anteriorly flattened, set Nematodes were killed by gentle heat and fixed in a off by a slight constriction from the body; 4 high 4% solution of formaldehyde, then dehydrated and processed to glycerine according to Seinhorst's method. and 7-9 wide, bearing 6-8 annuli. Labial plate Body length and curved structures were measured with fused with labial sectors (Figs. 10, 11). Longi- the aid of a precision curvimeter; straight structures, tudinal indentations on lip annuli behind am- such as maximum body width, stylet, anal body width, phids present (Figs. 10, 11). Cephalic framework etc., were measured using a micrometer-scale in the lightly sclerotized. Stylet 2.5-2.7 times longer than eyepiece of a high-power (x 1,250) microscope. All measurements are in micrometers unless otherwise lip region width, conus 11-12 long. Basal knobs stated. rounded, anterior surface posteriorly sloping, For examination under the scanning electron micro- flattened, or slightly indented, 4-5 across. Dorsal scope (SEM), specimens were processed with Spurr's esophageal gland orifice 2 from stylet base. Pro- low viscosity epoxy resin, coated with gold (De Grisse, corpus slender, 1.1-1.6 times longer than isth- 1973), and examined with a Zeiss DSM 950 scanning electron microscope at accelerating voltages of 10 and mus. Median esophageal bulb round-oval, 15 kV. strongly muscular, 13-17 long x 10—14 wide, 96 Copyright © 2011, The Helminthological Society of Washington 97 Table 1. Morphometric data for Bitylenchus serranus sp. n. female and male paratypes (measurements in /tin). Females (N = 36) Males (N= 18) x± SD Range X± SD Range L 824.0 ± 47.3 734.0-922.0 756.0 ± 61.4 650.0-906.0 a 30.0 ± 1.5 26.0-33.0 32.0 ± 2.1 28.0-36.0 b 6.0 ± 0.5 5.0-7.0 5.0 ± 0.4 5.0-6.0 b, 10.0 ± 0.8 8.0-12.0 10.0 ± 0.5 8.0-10.0 — V% 52.0 ± 1.5 49.0-54.0 — T% — — 48.0 ± 5.7 39.0-58.0 G, 25.0 ± 3.4 18.0-32.0 — — G2 27.0 ± 3.5 19.0-34.0 — — c 17.0 ± 1.5 14.0-20.0 15.0 ± 1.4 13.0-19.0 c' 2.6 ± 0.2 2.0-3.0 2.7 ± 0.2 2.5-3.0 Stylet length 21.0 ± 0.6 19.0-22.0 20.0 ± 0.6 19.0-21.0 m 52.0 ± 0.9 50.0-55.0 52.0 ± 1.2 50.0-53.0 o 10.0 ± 1.1 9.0-13.0 11.0 ± 1.1 10.0-13.0 S 1.4 ± 0.3 1.0-2.0 1.5 ± 4.2 1.4-1.5 MB 51.0 ± 1.6 49.0-55.0 52.0 ± 1.7 49.0-55.0 Nerve ring 96.0 ± 7.5 79.0-113.0 91.0 ± 7.2 79.0-104.0 Excretory pore 121.0 ± 9.2 97.0-137.0 113.0 ± 8.0 101.0-128.0 Esophagus length 145.0 ± 10.9 116.0-161.0 137.0 ± 9.8 124.0-156.0 Maximum body width 28.0 ± 1.4 25.0-31.0 24.0 ± 1.9 19.0-27.0 Anal body width 19.0 ± 1.5 16.0-22.0 19.0 ± 1.0 17.0-20.0 Tail length 50.0 ± 5.4 40.0-62.0 52.0 ± 4.1 45.0-57.0 — Tail annuli 38.0 ± 4.3 31.0-46.0 — Spicule length - - 28.2 ± 1.4 26.0-31.0 Gubernaculum length — — 14.2 ± 0.7 13.0-16.0 with refractive cuticular valve 4 long. Basal bulb stylet = 19 Mm; m = 52; S = 1.5; O = 10; spicules pyriform, 21-29 long, 12-15 wide. Excretory pore = 30 Mm; gubernaculum = 14 Mm. located opposite anterior half of basal bulb level. PARATYPE MALES (TV = 18): Morphometrics Hemizonid occupying 3-4 annuli, located 2-4 are given in Table 1. Similar to female in mea- annuli anterior to excretory pore. Esophago-in- surements and morphology, except for the sexual testinal junction with 2 cardial cells. Fasciculi dimorphism of tail region. Testis well developed, (lateral canal, serpentine canal) present, occa- with 2 rows of spermagonia, 301^99 long. Spic- sionally obscure. ules ventrally curved, with pointed tip. Guber- Vulva transverse, flush with body surface or naculum bent, not protruding. Tail conoid, en- slightly raised; body cuticle just anterior to vulva veloped by a crenate bursa 80-103 long. Phasmid slightly thickened; body just behind vulva slight- located 16-26 from anus. ly constricted. Vagina straight, 10-12 long. Sper- TYPE HABITAT AND LOCALITY: Specimens col- matheca round, 12-18 wide, filled with round lected around the roots of pinus (Pinus nigra sperm, 2-3 wide. Egg in uterus oval, 18-19 x salzmanni) at Nava del Espino, Sierra de Ca- 60—61. Ovaries outstretched, posterior usually zorla, Jaen, Spain. longer than anterior, with a single row of oocytes. TYPE SPECIMENS: Holotype female, allotype Tail subcylindroid, with rounded terminus, and male, 29 female and 15 male paratypes (slides usually ventrally curved (about 90% of speci- ETC 1-BTC 28) at nematode collection of Cen- mens studied), rarely straight. Terminus coarsely tro de Investigation y Desarrollo Agrario, Gra- annulated, terminal cuticle 5-9 thick. Short post- nada, Spain; 6 female and 2 male paratypes (slides anal intestinal sac 10-19 long, extending up to BTC 24-BTC 28) at CAB International Institute one-fourth of tail length. Phasmids porelike, lo- of Parasitology (CIP), St. Albans, Herts., En- cated anterior to middle of tail, and at 7-17 an- gland. nuli posterior to anus level, seldom opposite. DIAGNOSIS AND RELATIONSHIPS: Bitylenchus ALLOTYPE (male in glycerine): L = 805 nm; serranus sp. n. is characterized by a hemispher- a = 31.0; b = 5.8; T = 52; c = 14.6; c' = 2.7; ical lip region with 6-8 annuli, stylet 19-23 long, Copyright © 2011, The Helminthological Society of Washington 98 JOURNAL OF THE HELMINTHOLOGICAL SOCIETY Figures 1-8. Bitylenchus serranus sp. n. 1. Female anterior region. 2. Lip region. 3-5. Female tails. 6. Male tail. 7, 8. Tail termini of females. lateral field with regularly areolated outer bands, coarsely annulated. It differs from B. wilskii in a short postanal intestinal sac extending up to finer body annuli: 1 vs. 2-3 wide, in stylet length: one-fourth of tail length, and tail subcylindroid, 19-23 vs. 24-27, larger stylet knobs, and female usually ventrally curved, rarely straight. tail terminus being coarsely annulated. Bitylenchus serranus is most similar to B.