Xiphinema parasimile Barsi et Lamberti, 2004

Lazarova, Stela, De Luca, Francesca & Peneva, Vlada, 2008, On two closely related species of Xiphinema americanum-group: X. simile Lamberti, Choleva & Agostinelli, 1983 and X. parasimile Barsi & Lamberti, 2004 (Longidoridae), with a description of the male of X. parasimile, ZooKeys 3 (3), pp. 29-50 : 31-41

Xiphinema parasimile Barsi et Lamberti, 2004

( Figs 1 ; 2A, D, F, I-K, N, P, T; 3 A-C; 4 A-E; 5 A-C; 6-8 )

Measurements. See Tables 1-3 ;

Description. Females. Body C to spiral shaped. Labial region 4-5 µm high, setoff from the rest of the body. Amphids hardly visible, its opening 4 µm in a paratype specimen (40 % of the corresponding body width); 5-5.5 µm wide in specimens from Vinogradets and Orlyane (50-64 %). Odontostyle with poorly developed basal collar. Pharyngeal characters presented in Table 2 . Reproductive system amphidelphic, symbiont bacteria present in the ovaries. Uteri short, ovejector and separate uteri present ( Table 3 ); vagina 41.5 -52.2% of the corresponding body width (averages 48.4±1.4, n=5, paratype material; 46.4±3.4, n=17, Vi; 46.8±1.4, n=7, Or). Sperm cells observed in some of the females from Vinogradets. Tail conical, dorsally convex, ventrally almost straight or slightly concave with pointed terminus.

Male. One specimen was found in Vinogradets population. Male similar to the female with posterior region more strongly curved. Lip region and tail shape as in females, a difference was observed within anal body width, which reflect the lower c’ value. Spicules slightly curved, lateral guiding piece 4 µm long. Adanal supplement pair preceded by a row of five irregularly spaced supplements. Tail conoid, ventrally straight, dorsally convex with pointed terminus.

Juveniles. The scatter diagram based on functional and replacement odontostyle, and body length revealed presence of four juvenile stages ( Fig. 7 ).

PCR-RFLP analysis. The amplification of the D1-D2 expansion domains of X. parasimile generated a unique band of about 0.8 kb. Th e restriction fragments of the amplified region of X. parasimile population from Vinogradets locality using several enzymes are presented in Fig. 8 .

Remarks. Metric data are within the ranges reported in the original description, with slightly lower average values for the female body length (1.78-1.82 vs 1.93-1.99

mm). odontophore length (37.5-38.0 vs 40.7-41.6 µm), and body width (26.5-27.1 vs 27.9-28.3 µm) for the Bulgarian populations.

Male of X. parasimile compared to X. simile males from Serbia and Bulgaria had posterior part of the body less curved and a shorter body (1.84 vs 1.98-2.13 mm), narrower lip region (8.5 vs 10-10.5 µm), longer odontostyle (72 vs 63-69 µm) and shorter lateral guiding piece (4 vs 6 µm).

The RFLP results showed that the Bulgarian population of X. parasimile has the same restriction profiles as that from Serbia ( Barsi and De Luca 2008). The enzyme Ava II produced in the Bulgarian population two extra bands that were absent in X. parasimile from Serbia, suggesting the existence of differences in restriction sites in D1-D2 sequences. Furthermore, the enzyme Alu I always showed a band of 0.8 kb, corresponding to the undigested product, along with the expected restriction fragments suggesting microheterogeneity of the D1-D2 region of Bulgarian materials of X. parasimile ( Powers et al. 1997, Hugall et al. 1999, Hung et al. 1999, Subbotin et al. 2000, Morales-Hojas et al. 2001, Elbadri et al. 2002, Otranto et al. 2003, De Luca et al. 2004).

Figures

Fig. 1. Xiphinema parasimile Barsi et Lamberti, 2004. Female: A-C, Anterior region; D, Part of posterior genital branch; E, Posterior ovary; J, K, Vaginal region – different optical sections; L-O, Variation in tail shape; Male: F, Anterior end; G, Testis with sperm; H, Posterior end – muscles and supplements; I, Tail and copulatory apparatus; P, Tail. Scale bar: 20 µm.

Fig. 3. Variability of vulval region. A-C: Xiphinema parasimile, A1-A5: Vi; B1-B3: Or; C1-C3: paratypes; D-H: Xiphinema simile, D1-D5: KB; E1-E6: Or; F: Ka; G1-G3: SR; H: Kb. Scale bar: A-H=50 µm.

Fig. 4. Xiphinema parasimile.A-E: Females. A, B: Anterior genital branch, Vi: C: Anterior genital branch, Or; D: Posterior genital branch, Or; E:Posterior genital branch, paratype; Xiphinema simile. F-K.Females.Posterior genital branches. F: Or; J: Ka; K: Ka; Anterior genital branches. G: Or; H: SR; I: KB; Scale bar: A-K=50 µm.

Fig. 5. Variability of female tale shape in studied populations. A-C: Xiphinema parasimile, A1-A8: Vi; B1- B9: Or; C1-C3: paratypes; D-H: Xiphinema simile, D1-D5: SR; E1-E8: Or; F1-F2: Kb; G1: Ka; H1-H5: KB. Scale bar: A-H=50µm.

Fig. 6. Xiphinema parasimile. A-D, Anterior region of first, second, third and forth juvenile stages; E, Female anterior end; F-I, Tail of first, second, third and forth juvenile stages; J, Female tail. Scale bar: A-J, 20 µm.

Fig. 7. Scatter plot of odontostyle (■) and replacement odontostyle (□) against body length of Xiphinema parasimile juveniles and females from Vinogradets population.

Fig. 8. Restriction fragments of amplified D1-D2 expansion domains of Xiphinema parasimile. A: Alu I, Av: Ava II, D: Dde I, N: Nde II, P: Pst I, R: Rsa I and M: 100bp ladder.

Tables

Table 2. Pharyngeal characters of Xipinema parasimile and X. simile females from different localities.

		Xipinema parasimile			Xiphinema simile
Locality Character	Orlyane	Vinogradets	Ralja , Serbia Paratypes	Srebarna Reserve	Kalimok-Brashlen Orlyane
	n=6	n=14	n=5	n=8	n=12	n=11
Pharynx length (µm)	266.7±28.9 (245-288)	250.5±21.0 (233-311) n=11	282.2±2.1 (260-310)	303.1±9.6 (293-320)	289.9±10.4 (273-310)	293.9±15.5 (273-316)
Bulbus length (µm)	62.0±2.1 (58-65)	59.8±3.5 (55.5-68)	61.6±4.8 (56-63)	80.2±1.3 (78-82)	83.0±3.4 (78-92)	84.2±5.7 (76-92)
Bulbus width (µm)	11.75±0.4 (11-12)	12.0±0.6 (11-13)	14.4±0.5 (14-15)	13.1±0.4 (12-14)	13.3±1.1 (12-15)	13.1±0.8 (12-14)
Bulbus length/ Pharynx length (%)	23.4±1.9 (20.3-25.6)	24.2±2.3 (19-28) n=11	21.8±0.7 (20.7-22.6)	27.5±1.1 (24.3-28.0)	28.6±1.4 (27.1-31.4)	28.9±1.1 (27.7-30.5)

Table 3. Measurements of uteri, ovejector and vaginal parts. All measurements in micrometres presented as mean ± standard deviation (range).

	Characters	Anterior		Posterior		Ovejector		Vagina length	Pars distalis		Pars proximalis
	Locality	uterus		uterus					vaginae		vaginae
	Ralja, Trešna	40.0±11.3						14.5±1.05	7.8±0.8		8.75±0.3
Xiphinema parasimile	Serbia,	(27-46)		-		-		(13-16)	(7-8.5)		(8.5-9)
	paratypes	n=3						n=5	n=3		n=4
	Vinogradets	33.1±0.4		31.2±0.7		29.4±4.3		14.5±1.7	7.4±0.5		7.4±0.5
		(30-38)		(24-39)		(26-33.5)		(13-15)	(7-8)		(7-9)
		n=13		n=13		n=10		n=17	n=15		n=19
	Orlyane	32.3±4.2		30.5±5.7		26.8±2.1		13.9±0.9	7.0±1.4		7.6±1.4
		(24-43)		(24-38)		(23-30)		(13-15)	(6-8)		(7-8.5)
		n=6		n=6		n=4		n=7	n=6		n=7
	Srebarna	18.8±2.8		18.5±2.4		36.3±6.4		14.8±1.3	5.8±0.4		9.5±0.9
		(14-21)		(15-20)		(29-41)		(13-16)	(5.5-6)		(8.5-11)
		n=6		n=6		n=3		n=5	n=8		n=7
simile	Kalimok- Brashlen 21.8±1.9			21.5±1.8		43.1±3.1		16.8±0.8	6.4±0.65		8.6±0.5
		(16.5-24)		(19-24)		(36.5-48)		(15-18)	(5.5-7)		(8-10)
Xiphinema		n=14		n=14		n=12		n=15	n=17		n=17
	Orlyane	21.75±2.2		22.1±2.3		43.8±4.2		16.9±1.1	6.05±0.6		9.05±0.8
		(17-24)		(19-26)		(36-50)		(15-18)	(5.5-7)		(8-10)
		n=7		n=7		n=7		n=8	n=11		n=10
	Kamen bryag	23.0±4.8		24.2±4.15		47.2±8.9		15.9±1.8	6.4±0.6		9.8±0.8
		(18-30)		(19-30)		(37-60)		(13-17)	(6-7)		(9-10)
		n=5		n=5		n=5		n=5	n=5		n=5