European Journal of Taxonomy 171: 1-20

http://dx.doi.org/10.5852/ejt.2016.171

This work is licensed under a Creative Commons Attribution 3.0 License.

ISSN 2118-9773

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2016 • Nguyen T.A.D. et al.

Research article

urn:lsid:zoobank.org:pub:2AE15A24-33C0-42F9-BC47-5A44DF36AE9F

Two atypical new species of the genus Sectonema Thorne, 1930

(Nematoda, Dorylaimida, Aporcelaimidae) from Vietnam

Thi Anh Duong NGUYENJoaquin ABOLAFIA 2 , Michael BONKOWSKI 3 ,

Reyes PENA-S ANTI AGO 4 & Sergio ALVAREZ-ORTEGA 5 -*

13 Department of Terrestrial Ecology, Zoological Institute, University of Cologne,

Ziilpicher Strafie 47b, 50674 Cologne, Germany.

1 Institute of Ecology and Biological Resources, Vietnam Academy of Science and Technology,

18 Hoang Quoc Viet, Hanoi, Vietnam.

124 5 Departamento de Biologia Animal, Biologia Vegetal y Ecologia, Universidad de Jaen,

Campus ‘Las Lagunillas’ s/n, Edificio B3, 23071 Jaen, Spain.

* Corresponding author: [email protected]

1 um:lsid:zoobank.org:author:22D081FF-7764-4ECF-8C48-AF2EC20014FD

2 um:lsid:zoobank.org:author:D7E30782-F6F2-48CA-B26F-9CA512FEC078

3 urn:lsid:zoobank.org:author:6C0B083D-8A7E-4E0E-B03E-C00529DC4FC9

4 urn:lsid:zoobank.org:author:0B263323-3533-48A6-9D5C-BFF39D87BD47

5 um:lsid:zoobank.org: author: 68794F1F-07DB-4880-9D28-8D50028B32C7

Abstract. Two new species of the genus Sectonema from natural habitats of northern Vietnam are studied.

This paper includes their descriptions, measurements, line illustrations, and light microscope (LM) and

scanning electron microscope (SEM) pictures. Sectonema tropicum sp. nov. is characterized by a 2.56-

3.24 mm long body, 19-21 pm broad lip region, odontostyle 20-21 pm long at its ventral side, 730-

834 pm long neck, pharyngeal expansion occupying 52-59% of total neck length, uterus a simple tube-like

structure 150-242 pm long or 1.2-2.5 tunes the body diameter, pars refringens vaginae present, V= 48-52,

short (31—40 pm, c = 70-91, c ' = 0.5-0.6) and rounded tail, 91-97 pm long spicules, and only one wealdy

developed ventromedian supplement. Sectonema vietnamense sp. nov. is characterized by its slender (<a =

33M9) and 2.71-4.25 mm long body, 14-16 pm broad lip region, odontostyle 8-9 pm long at its ventral

side, 716-918 pm long neck, pharyngeal expansion occupying 63-67% of total neck length, uterus simple

and 209-242 pm long or 2.5-2.9 times the corresponding body diameter, para refringens vaginae absent,

V= 54, short (34-39 pm, c = 70-115, c ’ = 0.6-0.8) and rounded tail, 59-75 pm long spicules, and three or

four irregularly spaced ventromedian supplements bearing hiatus. Both species are also characterized by

their nearly continuous lip region, an atypical feature in this genus. Molecular analysis of S. tropicum sp.

nov. confirms that Sectonema is a natural (monophyletic) taxon, very close to Metaporcelaimus.

Keywords. D2-D3 expansion segments of LSU ribosomal DNA, morphology, morphometries,

taxonomy, SEM.

Nguyen T.A.D., Abolafia J., Bonkowski M., Pena-Santiago R. & Alvarez-Ortega S. 2016. Two atypical new

species of the genus Sectonema Thorne, 1930 (Nematoda, Dorylaimida, Aporcelaimidae) from Vietnam. European

Journal of Taxonomy 171: 1-20. http://dx.doi.org/10.5852/ejt.2016.171

European Journal of Taxonomy 171: 1-20 (2016)

Introduction

This contribution presents partial results of two nematological projects that overlap in their objectives.

On the one hand, it is the second in a series of papers devoted to studying the nematode diversity of

aporcelaims (family Aporcelaimidae Heyns, 1965) from natural areas of Vietnam, the first one (Alvarez-

Ortega et al. 2015) dealing with the representatives of the genus Aporcelaimoides Heyns, 1965. On the

other hand, it is part of a general revision of the taxonomy and systematics of this dorylaimid group,

in which the genera Aporcelaimellus Heyns, 1965, Metaporcelaimus Lordello, 1965 and Sectonema

_ r

Thorne, 1930 have already received some attention (see for instance, and respectively, Alvarez-Ortega &

r r

Pena-Santiago 2013; Alvarez-Ortega et al. 2013a; Pena-Santiago & Alvarez-Ortega 2014a).

Interesting material belonging to the genus Sectonema was collected by the first author during several

nematological surveys conducted in natural areas of Vietnam over the last years. The specimens are

characterized by a nearly continuous lip region, indeed a rather atypical feature in Sectonema. Its detailed

study revealed it to belong to two unknown forms, which are herein described.

Material and methods

Nematodes

Nematodes were collected from pristine areas in Northern Vietnam, extracted from soil samples using the

methods by Baermann (1917) and Flegg (1967), relaxed and killed by heat, fixed in 4% formaldehyde,

and processed to anhydrous glycerine following Siddiqi’s (1964) technique. Finally, the specimens were

mounted on permanent glass slides to allow handling and observation under TM.

Light microscopy

Specimens were measured using a light Olympus BH-2 microscope equipped with differential

interference contrast (DIC). Morphometries included de Man’s indices and the usual measurements.

The location of the pharyngeal gland nuclei is expressed according to Loof & Coomans (1970) and

spicule terminology follows Pena-Santiago et al. (2014). Some of the best preserved specimens were

photographed with a Nikon Eclipse 80i microscope and a Nikon DS digital camera. Raw photographs

were edited using Adobe® Photoshop® CS version 8.0.1. Drawings were made using a camera lucida.

Scanning electron microscopy

After examination and identification, a few of the preserved specimens were selected for observation

under SEM following the protocol by Abolafia & Pena-Santiago (2005). The nematodes were hydrated

in distilled water, dehydrated in a graded ethanol and acetone series, critical point dried, coated with

gold, and observed with a Zeiss Merlin microscope.

DNA extraction, PCR and sequencing

DNA was extracted from individuals using the proteinase K protocol. Nematode material was transferred

to an 0.5 ml Eppendorf tube containing 18 pi of Worm Lysis Buffer (WLB) (50 mM KCL, 10 mM Tris pH

8.3, 2.5 mM MgCl 0 , 0.45% NP 40, and 0.45% Tween 20) and 2 pi proteinase K (600 pg ml 1 ) (Thermo

Scientific). The tubes were incubated at 65°C (1 h) and then at 95°C (15 min). The PCR was performed in

a 30 pi final volume containing 24.9 pi of sterile water, 0.6 pi of each PCR primer, 0.6 pi dNTP mixture,

0.3 pi Taq poly erase, 3 pi Buffer lOx Thermo Scientific Green and 1 pi of DNA extracted solution. The

PCR amplification profile consisted of four min at 94°C, 35 cycles of 1 min at 94°C, 1.5 min at 55°C and

2 min at 72°C, followed by a final step of 10 min at 72°C. The primers used for amplification were D2A

(5 -ACAAGTACCGTGAGGGAAAGTTG-3 ) and D3B (5 -TCGGAAGGAACCAGCTACTA-3') for

amplification of D2-D3 region of 28S (Subbotin et al. 2006).

NGUYEN T.A.D. et al, Two new Sectonema species from Vietnam

The PCR products were purified using Exo-SAP PCR cleanup containing 7 pi of PCR product, 0.15 pi

Exonuclease I (Exo), 0.9 pi Shrimps Phosphatase Alkali (SAP) and 1.95 pi of sterile water. They were

incubated at 37°C for 20 min and then heated up to 85°C for 15 min; then the purified solution was

diluted ten times.

The sequencing reaction was performed with 1 pi of DNA purified template, 0.25 pi BigDye v3.1,

2.25 pi 5x BigDye sequencing buffer, 0.25 pi of one primer. The mixture was heated up for 10 s at 96°C,

5s at 55°C repeated for 32 cycles followed by 4 min at 60°C. The sequencing was performed at the

Cologne Center for Genomics (CCG). The sequences obtained were submitted to the GenBank database

under accession numbers KT868957 and KT868958.

Phylogenetic analyses

The newly obtained sequences were aligned with other forty two D2-D3 expansion segments of 28S

rRNA gene sequences available in GenBank using ClustalX 1.83 (Thompson et al. 1997). Outgroup

taxa were chosen according to the results of previously published data (Holterman et al. 2008; Alvarez-

Ortega et al. 2013b). Sequence alignments were manually edited using GenDoc 2.6.002 (Nicholas et al.

1997). The sequence dataset was analysed with Bayesian inference (BI) and Maximum Likelihood (ML)

using MrBayes 3.1.2 (Huelsenbeck & Ronquist 2001; Ronquist & Huelsenbeck 2003) and MEGA 6

(Tamura et al. 2013), respectively. The best fit model of DNA evolution for BI was obtained using the

program MrModeltest 2.2 (Nylander 2002) with the Akailce Information Criterion in conjunction with

PAUP* 4b 10 (Swofford 2003). BI analysis under the GTR + G + I model was initiated with a random

starting tree and run with the four Metropolis-coupled Markov chain Monte Carlo (MCMC) for 10 6

generations. ML analysis was implemented under the best-fitting evolutionary model (GTR + I + G),

obtained using the program MEGA 6, and 1000 bootstrap replications. The topologies were used to

generate a 50% majority rule consensus tree. Posterior probabilities (PP) are given on appropriate

clades. The trees were visualised with the program FigTree vl.4.0 and drawn with Adobe Acrobat XI

Pro 11.0.1.

List of abbreviations

a = body length / greatest body diameter

b = body length / distance from anterior end to pharyngo-intestinal junction

c = body length / tail length

c’ = tail length / tail diameter at anus or cloaca

DN = distance from body anterior end to the nucleus of pharyngeal dorsal gland expressed as percentage

(%) of total neck length

DO = distance from body anterior end to the outlet of pharyngeal dorsal gland expressed as percentage

(%) of total neck length

L = overall body length

n = number of specimens on which measurements are based

S jNj = distance from body anterior end to the anterior nucleus of first pair of ventro-sublateral pharyngeal

glands expressed as percentage (%) of total neck length

SjN 2 = distance from body anterior end to the posterior nucleus of the first pair of ventro-sublateral

pharyngeal glands expressed as percentage (%) of total neck length

S 0 N = distance from body anterior end to the nuclei of the second pair of ventro-sublateral pharyngeal

glands expressed as percentage (%) of total neck length

V = distance from body anterior end to vulva expressed as percentage (%) of the body length.

European Journal of Taxonomy 171: 1-20 (2016)

Results

Phylum Nematoda Cobb, 1932

Class Enoplea Inglis, 1983

Order Dorylaimida Pearse, 1942

Family Aporcelaimidae Heyns, 1965

Genus Sectonema Thorne, 1930

Sectonema tropicum sp. nov.

um:lsid:zoobank.org:act:014CA9CB-57EE-43BB-BBFF-0ClA7EBD522D

Figs 1—4; Table 1

Diagnosis

The new species is characterized by a 2.56-3.24 mm long body, lip region hardly offset by very weak

depression and 19-21 pm broad, odontostyle 20-21 pm long at its ventral side and 6.6-7.1 times as long

as wide, 730-834 pm long neck, pharyngeal expansion 403M70 pm long or occupying 52-59% of total

neck length, uterus a simple tube-like structure 150-242 pm long or 1.2-2.5 times the body diameter,

pars refringens vaginae present, V = 48-52, short (32-39 pm, c = 70-88, c ’ = 0.5-0.6) and rounded

female tail, male tail similar to that of female (31—40 pm, c = 73-91, c ’ = 0.5-0.6), 91-97 pm long

spicules, and only one weakly developed ventromedian supplement.

Etymology

The specific epithet refers to the tropical area where the new species was collected.

Type material examined

Holotype

VIETNAM: §, in acceptable state of preservation, Northern Vietnam, Cao Bang Province, Pia Oac

Natural Reserve, 22° 36’28” N, 105° 52’15” E, tropical evergreen forest soil associated with Machilus

sp. and Dimocarpus sp., deposited in the nematode collection of the University of Jaen, Spain.

Paratypes

VIETNAM: 4 3 <$<$, in acceptable state of preservation, same data as holotype; 1 $, 1 in

acceptable state of preservation, same locality, deposited in the nematode collection of the Institute of

Ecology and Biological Resources, Hanoi, Vietnam; 1 S, same locality, used for SEM.

Description

Adult

Moderately slender to slender nematodes of medium to big size, 2.56-3.24 mm long. Body cylindrical,

distinctly tapering towards the anterior end, less so towards the posterior end as the caudal region is

short and rounded. Habitus curved ventrad after fixation, especially in posterior body region, C-, G- or

spiral-shaped. Cuticle 3.0—4.5 pm thick at anterior region, 5-6 pm in mid-body and 8-10 pm on tail;

consisting of three layers, especially distinguishable at caudal region: thinner outer layer bearing very

fine transverse striation across the entire body, thicker intermediate layer with radial striation, and thin

inner layer. Lateral chords 13-21 pm wide at mid-body, occupying about one-sixth (13-18%) of mid¬

body diameter. Two ventral and two dorsal body pores often present at level of odontostyle-odontophore.

Lip region hardly offset from the adjacent body by weak, but perceptible depression, 2.8-3.0 times

wider than high and up to one-fourth (17-25%) of body diameter at neck base; lips (under SEM) mostly

amalgamated, but their perioral part distinctly separated by the existence of six radial, interlabial, deep

incisures delimiting six perceptible liplets; button-like labial papillae, the inner ones located at the margin

of the oral field and surrounded by two or three concentric annuli, whereas the outer papillae, located

NGUYEN T.A.D. et al ., Two new Sectonema species from Vietnam

Fig. 1 . Sectonema tropicum sp. nov. (Line). A. Lip region in lateral surface view. B. Anterior region in

mid-optical level, lateral view. C. Neck region. D. 5 , anterior genital branch. E. Vagina. F. S, posterior

body region. G. posterior body region. H. entire. I. S, entire. J. Lateral guiding piece. K. Spicule.

European Journal of Taxonomy 171: 1-20 (2016)

Fig. 2. Sectonema tropicum sp. nov. (LM). A, C, E. Anterior region in lateral, median view. B. Lip region

in lateral, surface view. D. Neck region. F. Pharyngo-intestinal junction. G. Vagina. H. 5, anterior

genital branch. I. Vagina and anterior uterus containing sperm cells inside. J. Uterine egg. Scale bars:

A-B, E, G = 10 pm; C, F, I-J = 20 pm; D, H = 50 pm.

NGUYEN T.A.D. et al ., Two new Sectonema species from Vietnam

a little behind the inner papillae, are surrounded by only one annulus; cephalic papillae pore- rather

than button-like, also surrounded by only one ring-like annulus; oral aperture a dorso-ventral, slightly

hexagonal orifice, the lip region hence showing a bi-radial symmetry. Amphid fovea cup-shaped, its

opening occupying 9-10 pm or less than one-half (43—49%) of lip region diameter. Cheilostom nearly

cylindrical, without any differentiation. Stomatal protruding structure apparently a reduced odontostyle

6.6-7.1 times longer than wide (see: Remarks), its ventral side 1.0-1.1 times longer than lip region

diameter and 0.65-0.83% of body length. Guiding ring simple, plicate, at 0.8-0.9 lip region diameters

from the anterior end. Odontophore linear, rod-like, 1.7-1.9 times the odontostyle length, somewhat

irregular at its base and with (in lateral view) the ventral side slightly longer than the dorsal one (Fig.

2A). Anterior region of pharynx enlarging very gradually; basal expansion 7.1-10.7 times longer than

wide, 3.6-5.6 times as long as body diameter, and occupying 52-59% of total neck length; gland nuclei

Fig. 3. Sectonema tropicum sp. nov. (LM). A. <$, posterior body region. B. §, posterior body region.

C. 5 , caudal region. D. S, caudal region. E-G. Spicules, E-F also showing the lateral guiding piece.

Scale bars: A-B = 20 pm; C-G =10 pm.

European Journal of Taxonomy 171: 1-20 (2016)

often obscure, located as follows: DN = 60-62 (n = 4); SjN) = 75 (n = 2); SjN 2 = obscure; S 0 N = obscure.

Nerve ring located at 182-216 pm from anterior end or 25-26% of total neck length. Cardia rounded

conoid, 13-17 x 13-17 jam; a weak ring-like structure is present surrounding its junction to pharyngeal

base.

Female

Genital system didelphic-amphidelphic, with almost equally and well developed branches, the anterior

320^142 pm or 12-15% of body length and the posterior 348, 440 pm (n = 2) or 14, 16% of body

length (447, 509 pm or 15, 17% bearing one uterine egg inside). Reflexed ovaries well developed,

often surpassing the sphincter level, the anterior 182-319 pm, the posterior 207-380 pm long; oocytes

arranged first in two or more rows, then in a single row. Oviduct 124-174 pm long or 1.1-1.8 times the

■

Fig. 4. Sectonema tropicum sp. nov. (SEM, S). A. Lip region in ventral view. B. Lip region in frontal

view. C. Lip region in lateral view. D. Lip region in sublateral view. E. Posterior body region in ventral

view. F. Posterior body region in lateral view. G. Caudal region in sublateral view. Scale bars: A-D =

5 pm; E-F = 20 pm; G = 10 pm

NGUYEN T.A.D. et al.. Two new Sectonema species from Vietnam

corresponding body diameter, and consisting of a slender part made of prismatic cells and a moderately

developed pars dilatata, with visible lumen and often containing sperm cells inside. Oviduct-uterus

junction marked by a sphincter. Uterus a simple, tube-like structure 150-190 pm long or 1.2-1.7 times

the corresponding body diameter [223, 242 pm (n = 2) long or 2.2, 2.5 times the corresponding body

diameter, with one uterine egg inside; and 231, 240 pm (n = 1) long, with two uterine eggs], always

containing abundant sperm cells inside. Vagina extending inwards 52-57 pm or one-half to four-

sevenths (48-57%) of body diameter: pars proximalis 33^13 x 23-28 pm, with somewhat sigmoid walls

surrounded by weak musculature; pars refiringens with two trapezoidal pieces measuring (in optical

section) 13-16 x 7-9 pm and with a combined width of24-28 pm; pars distalis 5.5-7.0 pm long. Vulva

a nearly equatorial transverse slit. Prerectum 2.0-2.4, rectum 0.7-1.0 anal body diameters long. Tail

short and rounded; two pairs of caudal pores, sublateral and close together.

Male

Genital system diorchic, with opposite testes. In addition to the ad-cloacal pair, situated at 18-21 pm

from cloacal aperture, one wealdy developed ventromedian supplement, lying out the range of retracted

spicules and located at 80-131 pm from the ad-cloacal pair. Spicules distinctly robust and massive,

especially in its posterior half, 3.6—4.2 times its maximum width, 1.4-1.6 times the body diameter at

level of the cloacal aperture: dorsal contour regularly convex, ventral contour slightly concave, with

distinct hump and hollow; curvature 121-124°; head occupying 11-16% of spicule total length, its

dorsal side visibly curved at its anterior end and longer than the ventral one, which is shorter and almost

straight; median piece 5.4-6.2 times longer than wide, occupying 51-59% of spicule maximum width,

reaching the posterior tip; posterior end 7-8 pm wide. Lateral guiding pieces 23-29 pm long, 5.3-

7.1 times longer than wide. Prerectum 3.2—4 6, cloaca 0.9-1.3 times the corresponding body width long.

Cloacal aperture, as seen under SEM, a curved anteriad, transverse slit. Tail similar to that of female;

caudal pores two pairs, one lateral, another subdorsal.

Relationships

In having a long stomatal protruding structure (20-21 pm at its ventral side) and short (c ’ up to 1.0) and

rounded tail, S. tropicum sp. nov. is morphologically very similar to S. demani Altherr, 1965, S. heynsi

Altherr, 1968, S. rotundicauda Goodey, 1951, S. septentrionale Pena-Santiago & Alvarez-Ortega,

2015 and S. sica Clark, 1964, but it can be distinguished from all of them in its smaller general size

(body length 2.56-3.24 mm, neck 730-834 pm long vs more than 5 mm, more than 1000 pm long,

respectively), nearly continuous lip region (vs offset by a more or less deep constriction) and stomatal

protruding structure of different nature (a reduced odontostyle vs a mural tooth; see Remarks). Besides,

the new species differs from S. demani (see recent re-description by Pena-Santiago & Alvarez-Ortega

2014b) in its less slender (a = 21-36 vs a = 48-56) body, narrower (19-21 vs 27-28 pm broad) lip

region, shorter pharyngeal expansion (occupying 52-59 vs 68-69% of total neck length), shorter female

tail (32—39 vs 51-55 pm, c’ = 0.5-0.6 vs c’= 0.7-0.8), and male present (vs absent). It differs from S.

heynsi (see recent re-description by Pena-Santiago & Alvarez-Ortega 2014b) in its narrower (19-21

vs 28 pm broad) lip region, circumoral area lacking (vs bearing) cilia- or setae-like structures, shorter

pharyngeal expansion (occupying 52-59 vs 68% of total neck length), shorter (32-39 vs 54 pm, c ’ =

0.5-0.6 vs c’= 0.7) female tail, and male present (vs absent). It differs from S. rotundicauda in its

less slender (a = 21-36 vs a = 80) body, narrower (19-21 vs about 28 pm broad) lip region, shorter

(31-40 vs 64 pm) male tail, fewer (one vs four) ventromedian supplements, and shorter (91-97 vs about

155 pm) spicules. It differs from S. septentrionale in its less slender (a = 21-36 vs a = 47-64) body,

narrower (19-21 vs 25-29 pm broad) lip region, shorter (occupying 52-59 vs 60-73% of total neck

length) pharyngeal expansion, uterus simple (vs tripartite) and shorter (150-242 pm or 1.2-2.5 times

the body diameter vs 370—493 pm or 2.9—4.3 times the corresponding body diameter), shorter (31—40 vs

40-62 pm, c ’ = 0.5-0.6 vs c ’ = 0.6-0.8) tail, fewer (one vs 5-11) ventromedian supplements, and shorter

(91-97 vs 100-145 pm) spicules. Finally, it differs from S. sica in its narrower (19-21 vs about 24 pm

European Journal of Taxonomy 171: 1-20 (2016)

Table 1. Morphometries of Sectonema tropicum sp. nov. Measurements in pm (except L, in mm), and in

the form: mean ± standard deviation (range).

Population

Character

Cao Bang Province (Pia Oac Natural Reserve)

Holotype

5?$

Paratypes

5<?<?

3.04

2.81 ±0.19(2.57-3.05)

2.87 ±0.31 (2.56-3.24)

24.9 ±4.7 (21-31)

29.0 ± 5.9 (23-36)

3.7 ±0.2 (3.4-3.9)

3.7 ±0.3 (3.4-3.9)

79.4 ±6.5 (70-88)

81.3 ±7.0 (73-91)

c’

0.6

0.6 ± 0.0 (0.5-0.6)

0.6 ±0.1 (0.5-0.6)

50.6 ± 1.5 (48-52)

—

Lip region diam.

20.1 ±0.6(20-21)

20.1 ±0.6(19-21)

Odontostyle length at ventral side

20.6 ±0.6 (20-21)

20.7 ±0.5 (20-21)

Odontostyle length at dorsal side

22.5 ± 0.5 (22-23)

22.5 ± 0.4 (22-23)

Odontophore length

36.0 ± 1.0 (35-38)

35.3 ±0.7 (34-36)

Guiding ring from ant. end

17.4 ±0.6 (17-18)

18.1 ±0.1 (18-18)

Neck length

764 ± 23 (730-790)

801 ±49 (745-834)

Pharyngeal expansion length

427 ± 29 (403-468)

448 ± 19 (436-470)

Diam. at neck base

103 ±12 (91—116)

95 ± 11 (84-111)

at midbody

114 ± 14(98—130)

101 ± 12(90-118)

at anus

64.4 ±4.5 (61-72)

61.2 ±2.1 (60-65)

Prerectum length

143

136 ±9 (126-150)

228 ±37 (189-277)

Rectum/cloacla length

53.8 ±3.0 (52-59)

66.3 ± 8.8 (53-77)

Tail length

35.5 ±3.3 (32-39)

35.4 ± 3.0 (31M0)

Spicule length

—

92.5 ±2.6 (91-97)

Ventromedian supplements

—

1 ±0(1-1)

broad) lip region, shorter (occupying 52-59% vs two-thirds of total neck length) pharyngeal expansion,

more posterior (V= 48-52 vs V= 40) vulva, shorter (32-39 vs 54 pm, c ’ = 0.5-0.6 vsc’ = 0.7) female

tail, and male present (vs absent).

Moreover, in having a lip region nearly continuous with the adjacent body, the new species resembles

S. mucrodens Siddiqi, 1984 and S. truxum Siddiqi, 1984, but it can be easily distinguished from these by the

more anterior position of the stomatal protruding structure (its anterior tip in front of the level of guiding ring,

just behind the base of lip region vs its anterior tip distinctly behind the level of guiding ring, far from the lip

region base) and pars refringens vaginae well (vs faintly) developed. Moreover, it differs from S. mucrodens

by its shorter (730-834 vs 1086-1157 pm) neck, shorter (occupying 52-59 vs 69% of total neck length)

pharyngeal expansion, shorter (32-39 pm, c ’ = 0.5-0.6 vs 44—50 pm, c ’ = 0.7) female tail, and male present

(vs absent). It differs from S. truxum in its narrower (19-21 vs 24 pm broad) lip region, shorter (730-834 vs

837-1070 pm) neck, more anterior (V= 48-52 vs V= 54-56) vulva, and male present (vs absent).

Molecular characterisation

Two sequences of the D2-D3 28S rRNA gene were obtained from one female and one male. Both

sequences are very similar (99%) and differ by two nucleotides only. The evolutionary relationships

of the new species with representatives of the order Dorylaimida are presented in Figs 5-6. The two

S. tropicum sp. nov. sequences are clustered together with other available sequences of Sectonema

representatives and form a highly supported clade together with members of the genus Metaporcelaimus ,

thus confirming previous results (Alvarez-Ortega et al. 2013a, b).

NGUYEN T.A.D. et al. Two new Sectonema species from Vietnam

100

TOO

Remarks

The nature of the stomatal protruding structure in the new species is difficult to interpret and deserves

further explanation. The specimens examined bear a reduced odontostyle (comparable to that described

in the type species S. ventrale Thone, 1930; see recent description by Pena-Santiago & Alvarez-Ortega

- Crassolabium sp. (AY593041)

Crassolabium circidifer (AY593039)

Crassolabium circulifer (AY593038)

Crassolabium sp. (AY593040)

Epidorylaimus lugdunensis (AY593035)

Epidoiylaimus lugdunensis (AY593036)

—Eudorylaimus sp. (AY593037)

-Enchodelus sp. (EF207240)

Longidorella sp. (AY593043)

Longidorella cf. macramphis (AY593042)

100

—Longidorella sp. (AY593044)

— Microdorylaimus miser (AY593046)

— Eudorylaimus minutus (AY593048)

• Microdorylaimus modestus (AY593049)

Pungentus silvestris (AY593052)

Pungentus silvestris (AY593053)

— Pungentus engadmensis (AY593050)

iooi— Tylencholaimus cf. teres (EF207243)

100

— Tylencholaimus mirabilis (AY593027)

- Tylencholaimus mirabilis (EF207242)

Tylencholaimus sp. (AY593028)

100

Aporcella simplex (JQ778274)

Aporcella simplex (JQ778273)

Discolaimus cf. major (EF207239)

■Discolaimus major (AY593025)

Discolaimus major (AY593026)

Carcharodiscus banaticus (AY593024)

Carcharodiscus banaticus (AY593023)

- Discolaimoides symmetricus (EF207238)

100

Sectonema harbatoides (AY593030)

Sectonema harbatoides (AY593031)

Sectonema harbatoides (AY593032)

Sectonema sp. (AY593033)

ioo Sectonema tropicum sp, nov. (KT868958)

Sectonema tropicum sp. nov. (KT868957)

- Metaporcelaimus marinensis (JQ9274.39)

Metaporcelaimus ovogranulosus (JQ927441)

Metaporcelaimus ovogranulosus (JQ927440)

- Metaporcelaimus capitatus (JQ927438)

100

Clax-icaudoides trophums (EF207236)

■ Clavicaudoides trophums (EF207237)

ioo Clavicaudoides elavicaudatus (EF207235)

Clavicaudoides elavicaudatus (EF207234)

- Nygolaimus cf. brachyuris (AY593061)

Fig. 5. Bayesian 50% majority rule consensus trees as inferred from D2-D3 expansion segments of 28S

rRNA gene sequence alignments under the GTR + I + G model. Posterior probabilities are given for

appropriate clades. Newly obtained sequences are indicated by bold letters.

European Journal of Taxonomy 171: 1-20 (2016)

2014a) rather than a mural tooth as the base of its dorsal side is visibly sclerotized and seems to join the

dorsal stomatal wall (vs dorsal side not scletorized at the base, somewhat sigmoid and not joining the

dorsal stomatal wall). Nonetheless, some doubt persists in this matter.

100

— Tylencholaimus cf. teres (EF207243)

— Tylencholaimus mirabilis (AY593027)

— Tylencholaimus mirabilis (EF207242)

— Tylencholaimus sp. (AY593028)

100

Aporcella simplex (JQ778274)

Aporcella simplex (JQ778273)

Discolaimus cf. major (EF207239)

■Discolaimus major (AY593025)

100

L Discolaimus major (AY593026)

- Carcharodiscus banaticus (AY593024)

100

L Carcharodiscus banaticus (AY593023)

- Discolaimoides symmetricus (EF207238)

100

Sectonema barbatoides (AY593030)

Sectonema barbatoides (AY593031)

Sectonema barbatoides (AY593032)

L Sectonema sp. (AY593033)

Sectonema tropicum sp. nov. (KT868958)

Sectonema tropicum sp. nov. (KT868957)

— Metaporcelaimus marinensis (JQ927439)

Metaporcelaimus ovogmnulosus (JQ927441)

Metaporcelaimus ovogmnulosus (JQ927440)

- Metaporcelaimus capitatus (JQ927438)

- Crassolabium sp. (AY593041)

■ Crassolabium circulifer (AY593039)

Crassolabium sp. (AY593040)

100

L Crassolabium circulifer (AY593038)

ioo r Epidorylaimus lugdunensis (AY593035)

L Epidorylaimus lugdunensis (AY593036)

—Eudorylaimus sp. (AY593037)

■Enchodelus sp. (EF207240)

—Longidorella sp. (AY593044)

■ Microdorylaimus miser (A Y593046)

-Longidorella sp. (AY593043)

Longidorella cf. macramphis (AY593042)

100

—Eudorylaimus minutus (AY593048)

■ Microdorylaimus modestus (AY593049)

100

ioo Pungentus silvestris (AY593052)

Pungentus silvestris (AY593053)

— Pungentus engadinensis (AY593050)

Clavicaudoides davicaudatus (EF207235)

Clavicaudoides clavicaudatus (EF207234)

100

Clavicaudoides trophums (EF207236)

Clavicaudoides trophums (EF207237)

- Nygolaimus cf. brachyuris (AY593061)

Fig. 6. Maximum Likelihood tree as inferred from D2-D3 expansion segments of 28S rRNA gene

sequence alignments under the GTR + I + G model. Bootstrap values are given for appropriate clades.

Newly obtained sequences are indicated by bold letters.

NGUYEN T.A.D. et al.. Two new Sectonema species from Vietnam

Sectonemci vietnamense sp. nov.

urn:lsid:zoobank.org:act:AF58451E-D7A7-4E49-8A18-9F3998EF4A87

Figs 7-9, Table 2

Diagnosis

The new species is characterized by a slender (a = 33—49) 2.71M.25 mm long body, lip region offset by

depression and 14-16 pm broad, odontostyle 8-9 pm long at its ventral side, 716-918 pm long neck,

pharyngeal expansion 463-618 pm long or 63-67% of total neck length, uterus simple and 209-242 pm

long or 2.5-2.9 times the corresponding body diameter, pars refringens vaginae absent, V = 54, short

(36 pm, c= 105, A = 0.7) and rounded tail in female and slightly more conoid (34-39 pm, c = 70-

115, c’ = 0.6-0.8) in male, 59-75 pm long spicules, and three or four irregularly spaced ventromedian

supplements bearing hiatus.

Etymology

The specific epithet refers to the geographical origin of the species.

Type material examined

Holotype

VIETNAM: §, in good state of preservation, Northern Vietnam, Cue Phuong National Park, pristine

tropical forest, deposited in the nematode collection of the University of Jaen, Spain.

Paratypes

VIETNAM: 4 SS, 3 juvs, in good state of preservation, same data as holotype; 1 3, in good state

of preservation, same locality, deposited in the nematode collection of the Institute of Ecology and

Biological Resources, Hanoi, Vietnam; lc?, 1 juv., same locality, used for SEM.

Other material examined

VIETNAM: 2 33 , in acceptable state of preservation. Northern Vietnam, Thai Binh province, intensively

cultured field, deposited in the nematode collection of the University of Jaen, Spain.

Description (type population)

Adult

Very slender nematodes of big size, 3.13-4.25 mm long. Body cylindrical, distinctly tapering towards

the anterior end, less so towards the posterior one as the caudal region is rounded. Habitus regularly

curved ventrad after fixation, to a more or less open C, occasionally more curved at posterior body

region. Cuticle 2.0-2.5 pm thick at anterior region, 3.0—4.5 pm in mid-body and 8-10 pm on tail;

consisting of three layers especially distinguishable at caudal region: thinner outer layer bearing very

fine transverse striation through the entire body, thicker intermediate layer with radial striation, and

thin inner layer. Lateral chords 6-12 pm wide at mid-body, occupying up to one-sixth (8-15%) of

mid-body diameter. Two ventral and two dorsal body pores often present at level of stoma, and four

distinct lateral pores 6-20 pm apart visible behind the amphid fovea. Lip region nearly continuous (a

shallow depression is, however, often perceptible), weakly angular, visibly narrower than adjacent

body, 2.1-2.3 times wider than high and about one-fifth (18-23%) of body diameter at neck base; it

appears (under SEM) marked by radial (oral field) or longitudinal (remaining lip region) incisures, with

amalgamated lips and protruding, pore-like labial and cephalic papillae, all of them surrounded by a

ring-like annulus, and the inner labial papillae visibly larger than the outer ones; oral field perceptibly

hexagonal, oral aperture a dorso-ventral slit, the lip region hence showing a bi-radial symmetry. Cup¬

shaped amphid fovea, its opening at level of cephalic depression and occupying 10-11 pm or 68-72% of

lip region diameter. Nearly cylindrical cheilostom, without differentiation. Odontostyle comparatively

European Journal of Taxonomy 171: 1-20 (2016)

Fig. 7. Sectonema vietnamense sp. nov. (Line). A. Anterior region in median, lateral view. B. Lip region

in surface, lateral view. C. Neck region. D. Vagina. E. §, anterior genital branch. F. entire. G. $,

entire. H. posterior body region. I. posterior body region. J. Spicule. K. Lateral guiding piece.

NGUYEN T.A.D. et al ., Two new Sectonema species from Vietnam

short and occupying the whole stomatal lumen, its ventral side 0.5-0.6 times longer than lip region

diameter and 0.20-0.29% of body length. Guiding ring simple, somewhat plicate, at 0.5 times the

lip region diameter from anterior end. Odontophore linear, rod-like, 3.7^1.2 times the odontostyle

length. Tripartite pharynx, consisting of an anteriorly thickened section behind the odontophore base,

a perceptible narrower intermediate section with the nerve ring surrounding it, which enlarges very

gradually into the basal expansion 9.9-15.0 times longer than wide, 6.7-7.3 times longer than body

Fig. 8. Sectonema vietnamense sp. nov. (LM, type population). A. entire. B-C. Anterior region in

lateral, median view. D. §, anterior genital branch. E. c?, posterior body region. F. Oviduct-uterus junction.

G. Lip region in lateral, surface view. H. Vagina. I. S, caudal region. J-K. Spicules. L. §, caudal region.

Scale bars: A = 500 pm; B-C, H = 10 pm; D = 100 pm; E-F = 50 pm; G = 5 pm; I-L = 20 pm.

European Journal of Taxonomy 171: 1-20 (2016)

diameter and occupying 63-67% of total neck length; gland nuclei mostly obscure, only S 2 N are

visible, at 77-78% (n = 3) of total neck length. Nerve ring located at 147-178 pm from anterior end or

19-22% of total neck length. Cardia rounded conoid, 15-19 x 12-17 pm; a ring-like structure is present

surrounding its junction to pharyngeal base.

Female

Genital system didelphic-amphidelphic, with almost equally and well developed branches, the anterior

473 pm long or 13% of body length and the posterior 387 pm long or 10% of body length, each branch

bearing one uterine egg. Moderately sized ovaries, not surpassing the sphincter level, the anterior

191 pm, the posterior 141 pm long; oocytes arranged first in two or more rows, then in a single row.

Oviduct 117, 142 pm long or 1.4, 1.7 times the corresponding body diameter, and consisting of a slender

part made of prismatic cells and a well developed pars dilatata with wide lumen and sperm cells inside.

Oviduct-uterus junction marked by a sphincter. Uterus a simple, tube-like structure 209, 242 pm long or

2.5, 2.9 times the corresponding body diameter, containing abundant sperm cells and one uterine egg.

Vagina extending inwards 36 pm or about three-sevenths (44%) of body diameter: pars proximalis 23 x

28 pm, somewhat sigmoid walls and surrounded by weak musculature; pars refringens absent; pars

Fig. 9. Sectonema vietnamense sp. nov. (SEM, A = juvenile and B-J = <f). A. Lip region in ventral

view. B-C. Lip region in lateral view. D. Posterior body region in ventral view. E. Lip region in frontal

view. F. Detail of a ventromedian supplement. G. Caudal region in lateral view. H. Caudal region in

subventral view. I. Detail of the ad-cloacal pair of supplements. J. Caudal region in ventral view. Scale

bars = A-C, E = 2 pm; D = 20 pm; F, I = 1 pm; G-H =10 pm; J = 5 pm.

NGUYEN T.A.D. et al ., Two new Sectonema species from Vietnam

Table 2. Morphometries of Sectonema vietnamense sp. nov. Measurements in pm (except L, in mm),

and in the form: mean ± standard deviation (range).

Population

Character

Cue Phuong National Park

Thai Binh Province

Holotype

Paratypes

6 88

2 88

3.79

3.57 ±0.49 (3.13-4.25)

3.02,2.71

45.9 ± 3.0 (41—49)

35, 33

4.6

4.5 ±0.2 (4.3M.7)

105

96.9 ± 11.4 (86-115)

83, 70

c’

0.7

0.7 ±0.1 (0.6-0.8)

0.8, 0.7

—

Lip region diam.

14.9 ±0.5 (14-16)

16 (n = 2)

Odontostyle length at ventral side

8.8 ±0.3 (8-9)

9 (n = 2)

Odontostyle length at dorsal side

9.5 ±0.2 (9-10)

10 (n = 2)

Odontophore length

34.2 ± 1.3 (33-36)

33 (n = 2)

Guiding ring from ant. end

7.3 ±0.6 (7-8)

9 (n = 2)

Neck length

820

790 ±84 (716-918)

Pharyngeal expansion length

534

521 ±65 (463-618)

Diam. at neck base

72.0 ± 8.6 (63-85)

80, 74

at midbody

77.7 ± 8.3 (68-89)

87, 82

at anus

50.6 ±4.3 (46-58)

48, 55

Prerectum length

149

237 ±40 (210-297)

199,?

Rectum/cloaca length

58.8 ±9.3 (47-71)

66, 62

Tail length

36.8 ± 1.8 (34-39)

36,38

Spicule length

—

67.7 ± 5.3 (59-75)

73, 70

Ventromedian supplements

—

3.7 ±0.5 (3-4)

4 (n - 2)

distalis well developed, 15 pm long. Vulva a post-equatorial transverse slit. Prerectum 3.0, rectum 0.9

anal body diameters long. Short and rounded tail, its inner core with a finger-like projection at tail end;

two pairs of caudal pores, one sub-lateral, another sub-dorsal.

Male

Genital system diorchic, with opposite testes. In addition to the ad-cloacal pair, situated at 14-19 pm

from cloacal aperture, there is a series of three or four irregularly spaced, 15M8 pm apart, ventromedian

supplements, the posteriormost of which lying out the range of retracted spicules, at 76-91 pm from the

ad-cloacal pair. Spicule robust and massive, especially in its posterior half, 3.6M.6 times its maximum

width, 1.3-1.4 times the body diameter at level of the cloacal aperture: dorsal contour regularly convex,

ventral contour slightly concave, with distinct hump and hollow; curvature 125-136°; head occupying

13-19% of spicule total length, its dorsal contour distinct curved at its anterior end and longer than the

ventral one, which is short and almost straight; median piece 5.9-7.1 times longer than wide, occupying

52-65% of spicule maximum width, reaching the posterior tip; posterior end 6-7 pm wide. Lateral

guiding pieces 15-17 pm long, 5.3-6.5 times longer than wide. Prerectum 4.3-5.7, cloaca 1.0-1.3 times

the corresponding body width long. Rectal glands and their ducts ( cf. Coomans & Loof 1986) very well

perceptible (Fig. 8K). Cloacal aperture, as seen under SEM, a somewhat curved anteriad, transverse

slit; the two supplements of the precloacal pair distinctly separated. Tail a little more conoid than that of

female; two pairs of caudal pores, one almost lateral, another sub-dorsal.

European Journal of Taxonomy 171: 1-20 (2016)

Relationships

The new species is easily distinguishable from its congeners in the unique combination of these features:

narrow and nearly continuous lip region, very short odontostyle, absence of pars refringens vaginae and

short and rounded tail. Nonetheless, it resembles S. barbatum Heyns, 1965 in its general morphology,

but it can be distinguished from this because of its oral field lacking any setae-like structure (vs bearing

numerous short forwardly pointing setae), narrower (14-16 vs about 22 pm broad) lip region, shorter

(8-9 vs 16 pm) odontostyle, pars refringens vaginae absent (vs present), shorter (36 pm, c = 105, c ’ =

0.7 vs 48 pm, c = 81, c *= 1.0) female tail, and male present (vs absent). In having a comparatively small

body size and nearly continuous lip region, S. vietnamense sp. nov. is close to S. mucrodens and S. truxum ,

from which it differs in its narrower (14-16 vs more than 20 pm broad) lip region, shorter stomatal

protruding structure (8-9 vs more than 20 pm) located as usual (vs the anterior tip distinctly behind the

level of guiding ring), and pars refringens vaginae absent (vs consisting of two small pieces). Sectonema

vietnamense sp. nov. is also similar to S. pseudoventrale Heyns, 1965 in having a short odontostyle, but can

be distinguished from this by its narrower (14-16 vs 21-23 pm broad) lip region, shorter (209-242 pm or

2.5-2.9 times the corresponding body diameter vs 123 pm or 1.3 times the corresponding body diameter)

uterus, shorter (36 pm, c = 105, c ’ = 0.7 vs 46-63 pm, c = 82-130, c ’= 0.8-1.0) female tail, shorter (59-

75 vs 90-95 pm) spicules, and fewer (three or four vs six or seven) ventromedian supplements. Finally,

the new species also resembles S. tropicum sp. nov., but it can be distinguished from this by its narrower

(14-16 vs 19-21 pm broad) lip region, shorter (8-9 vs 20-21 pm at its ventral side) odontostyle, longer

(occupying 63-67 vs 52-59% of total neck length) pharyngeal expansion, more posterior vulva (V= 54

vs V =48-52), pars refringens vaginae absent (vs present), shorter (59-75 vs 91-97 pm) spicules and

higher number of ventromedian supplements (three or four vs one).

Remarks

The two males from Thai Binh are very similar to those of the type population, but some minor

morphometric differences have been also noted: somewhat smaller (2.71, 3.02 vs 3.13—4.25 mm long)

and more obese (a = 33, 35 vs a = 41—49) body. Such differences have been regarded as instraspecific

variation as the number of specimens examined is low and the main morphological and morphometric

features are coincident.

Acknowledgements

We would like to thank the directors of Cue Phuong National Park and Cao Bang Nature Reserve

for issuing relevant permits. This research was supported by the project entitled Aporcelaimidae

Mundi: Revision de la familia Aporcelaimidae Heyns, 1965 (Nematoda, Dorylaimida) (ref. CGL2012-

33239; co-financed FEDER), the German Academic Exchange Service (DAAD) (PKZ 91540366), the

Zoological Institute (University of Cologne) and IDEAL-WILD (United State). We thank Dr. Tam Vu

for providing the specimens from Thai Binh and Idea-Wild (United States of America) for supporting

the first author during her master thesis. SEM pictures were obtained with the assistance of technical

staff and equipment of “Centro de Instrumentacion Cientifico-Tecnica (CICT)”, University of Jaen.

References

Abolafia J. & Pena-Santiago R. 2005. Nematodes of the order Rhabditida from Andalucia Oriental:

Pseudacrobeles elongatus (de Man, 1880) comb. n. Nematology 7 (6): 917-926. http://dx.doi.

org/10.1163/156854105776186415

r ___ _

Alvarez-Ortega S., Nguyen T.A.D., Abolafia J., Vu T.T.T. & Pena-Santiago R. 2015. Three new species of

the genus Aporcelaimoides Heyns, 1965 from Vietnam (Nematoda, Dorylaimida, Aporcelaimidae), and

an updated taxonomy of the genus. ZooKeys 516: 1-26. http://dx.doi.Org/10.3897/zookeys.516.10087

NGUYEN T.A.D. et al.. Two new Sectonema species from Vietnam

r _

Alvarez-OrtegaS. & Pena-Santiago R. 2013. Taxonomy of the genus Aporcelaimellus Heyns, 1965 (Nematoda,

Dorylaimida, Aporcelaimidae). Zootcoca 3669: 243-260. http://dx.doi.Org/10.11646/zootaxa.3669.3.3

Alvarez-OrtegaS., Subbotin S. A. & Pena-Santiago R. 2013a. Morphological andmolecular characterization

of Californian species of the genus Metaporcelaimus Lordello, 1965 (Dorylaimida, Aporcelaimidae), with

a new concept of the genus. Nematology 15 (3): 251-278. http://dx.doi.org/10.1163/15685411-000Q2674

Alvarez-Ortega S., Subbotin S.A. & Pena-Santiago R. 2013b. Morphological and molecular

characterization of Californian species of the genus Aporcelaimellus Heyns, 1965 (Dorylaimida:

Aporcelaimidae). Nematology 15 (4): 431-439. http://dx.doi.org/10.1163/15685411-000Q2691

Bnerma nn G. 1917. Eine einfache Methode zur Auffindung von Ankylostomum (Nematoden) Larven in

Erdproben. Geneeskunding Tijdschrift voor Nederlandsch-Indie 57: 131-137.

Coomans A. & Loof PA.A. 1986. Observations on the glands of the male reproductive system in

dorylaims and its phylogenetic importance. Revue de Nematologie 9 (3): 261-265.

Flegg J.J.M. 1967. Extraction of Xiphinema and Longidorus species from soil by a modification of

Cobb’s decanting and sieving technique. Annals of Applied Biology 60 (3): 429-437. http://dx.doi.

org/10.1111/i. 1744-7348.1967.tb04497.x

Holterman M., Rybarczyk K., van den Elsen S., van Megen H., Mooyman P, Pena-Santiago R., Bongers

T., Bakker J. & Helder J. 2008. Aribosomal DNA-based framework for the detection and quantification

of stress-sensitive nematode families in terrestrial habitats. Molecular Ecology Resources 8 (1): 23-34.

http://dx.doi.Org/10.llll/i.1471-8286.2007.01963.x

Huelsenbeck J.P & Ronquist F. 2001. MRBAYES: Bayesian inference of phylogenetic trees.

Bioinformatics 17 (8): 754-755. http://dx. doi.org/10.1093/bioinformatics/17.8.754

Loof PA. A. & Coomans A. 1970. On the development and location of the oesophageal gland nuclei in

the Dorylaimina. Proceedings of the IX th International Nematology Symposium (Warsaw, Poland, 1967)\

79-161.

Nicholas K.B., Nicholas Jr H.B. & Deerfield II D.W. 1997. GeneDoc: analysis and visualization of

genetic variation. EMBnet News 4: 1-14.

Nylander J.A.A. 2002. MrModeltest vl.Ob. Department of Systematic Zoology, Uppsala University.

Pena-Santiago R., Abolafia J. & Alvarez-Ortega S. 2014. New proposal for a detailed description

of the dorylaim spicule (Nematoda: Dorylaimida). Nematology 16 (9): 1091-1095. http://dx.doi.

org/10,1163/15685411-00002834

Pena-Santiago R. & Alvarez-Ortega S. 2014a. Studies on the genus Sectonema Thorne, 1930

(Dorylaimida: Aporcelaimidae). Re-description of S. ventrale , the type species of the genus. Nematology

16 (9): 1097-1104. http://dx.doi.org/10.1163/15685411-000Q2837

Pena-Santiago R. & Alvarez-Ortega S. 2014b. Re-description of three species of the genus Sectonema

Thorne, 1930 (Nematoda: Dorylaimida: Aporcelaimidae) originally studied by E. Altherr. Zootaxa 3881:

63-74. http://dx.doi.Org/10.11646/zootaxa.3881.l.5

Ronquist F. & Huelsenbeck J. 2003. MrBayes 3: Bayesian phylogenetic inference under mixed models.

Bioinformatics 19 (12): 1572-1574. http://dx.doi.org/10.1093/bioinformatics/btg 180

Siddiqi M.R. 1964. Studies on Discolaimus spp. (Nematoda: Dorylaimidae) from India. Zeitschrift fur

Zoologische Systematik und Evolutionsforschung 2: 174-184. http://dx.doi.org/10. Ill 1/i. 1439-0469.1964.

tb00720.x

European Journal of Taxonomy 171: 1-20 (2016)

Subbotin S.A., Sturhan D., Chizhov V.N., Vovlas N. & Baldwin J.G. 2006. Phylogenetic analysis of

Tylenchida Thome, 1949 as inferred from D2 and D3 expansion fragments of the 28S rRNA gene

sequences. Nematology 8 (3): 455M74. http://dx.doi.org/10.1163/156854106778493420

Swofford D.L. 2003. PAUP*: phylogenetic analysis using parsimony (*and other methods). Version

4.0b 10. Sunderland, MA, USA, Sinauer Associates.

Tamura K., Stecher G., Peterson D., Filipski A. & Kumar S. 2013. MEGA6: Molecular Evolutionary

Genetics Analysis version 6.0. Molecular Biology / and Evolution 30: 2725-2729. http://dx.doi.

org/10.1093/molbev/mstl97

Thompson J.D., Gibson T.J., Plewniak F., Jeamnougin F. & Higgins D.G. 1997. The ClustalX windows

interface: flexible strategies for multiple sequence alignment aided by quality analysis tools. Nucleic

Acids Research 25 (24): 4876^1882. http://dx.doi.org/10.1093/nar/25.24.4876

Manuscript received: 2 September 2015

Manuscript accepted: 22 October 2015

Published on: 15 January 2016

Topic editor: Rudy Jocque

Desk editor: Charlotte Thionois

Printed versions of all papers are also deposited in the libraries of the institutes that are members of the

EJT consortium: Museum national d’Histoire naturelle, Paris, France; Botanic Garden Meise, Belgium;

Royal Museum for Central Africa, Tervuren, Belgium; Natural History Museum, London, United

Kingdom; Royal Belgian Institute of Natural Sciences, Brussels, Belgium; Natural History Museum of

Denmark, Copenhagen, De nm ark