A compendium of Aphelenchoides (Fischer, 1894) (Nematoda: Tylenchina: Aphelenchoidea) nematodes with the description of a new species from Manipur, India

: The present compendium is based on the findings of a research work on the survey of nematodes belonging to the family Aphelenchoidea in the northeastern states of India and the literature available on this particular species, mainly from Manipur. During the study, a total of 12 Aphelenchoides spp. were found, among which six species were reported for the first time from Manipur. A new species, Aphelenchoides oryzae is also described in the present article. The present study will help in making us understand the biodiversity status of Aphelenchoides nematodes in the region. Diagnosis of the species and illustrations along with dichotomous keys are provided in the manuscript.


INTRODUCTION
Forest conservation improves ecosystem functions and will help to protect natural biodiversity.Plant communities are the critical indicators for forest restoration.Below-ground diversity relates very closely with above-ground biodiversity.Plant parasitic nematodes are found in every soil of varied ecosystems.Nematodes are often sensitive to habitat disturbance, showing the characteristic sequence of recolonization after disturbance.Furthermore, they are represented in a wide array of trophic groups as herbivores (Tylenchid), bacterivores & fungivores (Aphelenchid), omnivores (Dorylaimid), and predators (Mononchid) reflecting resource availability and changes of environmental conditions in the soil providing information on succession and changes in decomposition pathways in the soil food -web, nutrient status & soil fertility, acidity, and the effects of soil contaminants (Yeates & Bongers 1999).Bacterial as well as fungal feeding nematodes like aphelenchids have a high carbon: nitrogen (C:N) ratio (±5.9) than their substrate (±4.1),consuming bacteria, they take in more N than necessary for their body structure.The excess nitrogen is excreted as ammonia.The bacterial and fungal feeding nematodes' community in the top 15 cm of the field soil mineralizes N at rates increasing to 1.01µg -N g -soil -1d -1 in the rhizosphere (Ferris et al. 1995;1996;1997).
Aphelenchid nematodes have diverse habitats.Several aphelenchids are associated with insects, some spending a part of their life -cycle in insects besides being phytophagous, while others are mycetophagous.Some forms of aphelenchids are true plant parasites and are, therefore, economically significant.Of the available aphelenchid nematodes, three species are major pests of agricultural and horticultural crops, i.e., white-tip nematode Aphelenchoides besseyi; Red-ring nematode of coconut Rhadinaphelenchus cocophilus, and pine-wilt nematode Bursaphelenchus xylophilus respectively in the world.So far, 138 species of Aphelenchoides (Fischer, 1894) have been identified, of which India contributed more than 12 species.

Study site
Manipur has a geographical area of 22,327 km 2 , situated at an altitude of 790 m with a longitude of 24.733333333333334 and a latitude of 93.9666666666667 (Figure 1).Soil samples were collected from different localities of Manipur.

Methodology
For collection of soil samples around the rhizospheric region of a particular host, 500 g of soil around the plant from 8 different sides were taken.The soil was mixed together thoroughly.From the thoroughly mixed soil, again 500 g were taken, serving as the sample soil for a particular host plant or tree.The samples were processed for extraction of nematodes through the Cobb's (1918) sieving and decanting method and Baermann's funnel technique.Collected nematodes through the process were fixed with warm formalin alcohol (F.A) (4:1) for 24 hours and afterward, dehydrated under the Seinhorst (1952) dehydration techniques.Dehydrated nematodes were mounted on clean non-greased slides with dehydrated glycerin as mountants.The specimens were studied, measurements taken and diagrams were drawn using a drawing tube attached to a microscope.Remark: The morphometric details of the present species conform well with those described by Chanu et al. (2015).Male: Not found.Remarks: The morphometric details of the present species conform well with those described by Chanu et al., (2015).

Diagnosis
Female: Body contour slightly ventrally curved.Fine cuticularisation with four incisures throughout body length.Cephalic region set off flattened.Spear 10.2─12.8µm long with a small basal knob.Oesophagous typical of aphelenchoid.Mono-prodelphic reproductive system and ovary outstretched.Vulva is a transverse slit with slightly protruding lips.Oocytes in single row with a uterine sac.Tail 38.4-46.2µm long, about 4─5 times anal body width long, terminus with a small ventral mucro which is multi-papillate almost to its tips.
Male: Not found.Remarks: The morphometric details of the present species conform well with those described by Chanu et al. (2015).

Diagnosis
Female: Body contour gradually tapering near extremities.Cuticle with fine striations and lateral fields marked by four fine lines.Cephalic region set-off.Spear 10.2─17 µm long.Oesophagous typical aphelenchoid with massive valvular apparatus.Nerve ring is behind the oesophageal bulb.Excretory pore at level of the nerve ring.Hemizonid posterior to excretory pore.
Vulva with protuberant labia and vagina directed forward.Uterus spheroid-shaped, filled with sperm.Ovary outstretched and post uterine sac collapsed.Tail 20.4-25.3µm long without a mucro.
Male: Not found.Remarks: The morphometric details of the present species conform well with those described by Chanu et al. (2015).

Diagnosis
Female: Body contour slender ventrally curved.Lateral filed marked by three incisures.The body cuticle is fine.The cephalic framework is high.Spear 13.6-15.3µm long, slender with indistinct basal knobs.Oesophagous typical with tamarind seed-shaped median bulb.Nerve ring behind the median bulb, 59.5-69.0Oesophageal gland lying dorsally along the intestine.Monoprodelphic reproductive system and oocytes arranged in a single row with uterine sac.Vulva protrudes in some species.Tail 62.9-98.0µm long, highly curved ventrally tapering into a pointed terminus.
Male: Not found.Remarks: The morphometric details of the present species conform well with those described by Chanu et al. (2012).

Diagnosis
Female: Body contour slightly curved with fine annulation.Lateral fields with four longitudinal lines merge into two lines at around the tail region.The cephalic region indistinctly set off with six equal lips.Spear 24.22 µm long with indistinct basal knobs.Procorpus straight, median bulb spherical to pyriform in shape.Excretory pore at the base of the median bulb.The vulva is a transverse slit.The monoprodelphic reproductive system and oocytes are arranged in a single row.Uterine sac well developed.The tail gradually tapers into a cylindrical tube, terminating in a ventral prong tip.
Remarks: The morphometric details of the present species conform well with those described by Chanu et al. (2014).

Diagnosis
Female: Body contour cylindrical, ventrally arcuate, the cephalic framework set off and flat.Cuticle marked by fine annulus.Lateral fields with two incisures.Spear short, 10.33-13.84µm long with small rounded basal knobs.The median bulb is spherical and basal bulb bifurcated.Excretory pore at 51.9-76.12µm from anterior end of body.Nerve ring behind median bulb,     Male: Not found.Remarks: The morphometric details of the present species conform well with those described by Chanu et al. (2018).

Diagnosis
Female: Body contour is slender with three lateral incisures.The cephalic region is slightly set off.Stylet slender 11.9 µm long without basal swellings.Oesophagous typical, median bulb elongated pearshaped.Nerve ring 66.3─68.0µm long.Excretory pore at 62.9-64.4µm, at the level of the nerve ring.The oesophageal gland was dorsal to the intestine.Reproductive system monoprodelphic and oocytes arranged in a single row, and uterine sac well developed.Tail 45.9-54.4µm long, short, and pointed with a ventral mucro.
Male: not found Remarks: The morphometric details of the present species conform well with those described by Chanu et al. (2012).(Figure 8, Table 2)

Diagnosis
Female: Body contour straight, tapering towards both extremities with four incisures in lateral field.Cephalic framework smooth and set-off.Spear 8.65 µm long with distinct stylet knobs.Procorpus zig-zag, coiled, strongly rounded corpus with sclerotized plates and elongated gland lobe, dorsal to the intestine.The excretory pore is close to the nerve ring.Vulval lips protrude with an inclined vagina.Monoprodelphic reproductive system, oocytes are arranged in a single row reaching up to the oesophageal bulb.Spermatheca large elongated and uterine sac empty.The anterior lip of the anus protrudes, the tail bluntly rounded, 12.11-25.95µm in length, with a small hair-like mucro.
Male: Not found.Remarks: The morphometric details of the present species conform well with those described by Chanu & Mohilal (2014).

Descriptions
Holotype female: Body straight, cylindrical, and robust upon fixation, 685.08 µm long.Lip region offset with rounded sides & flattened anteriorly, 5.19 µm wide and 1.7 µm high, and smooth in appearance.Body elongate with fine transverse annulations, 0.8 µm at mid body region.Lateral lines extend almost to tail tip with two ridges having four evenly spaced lines in the middle of the body.
Stylet is slender, 17.3 µm long, the conus slightly shorter than the shaft with indistinct swellings.Median oesophageal bulb rounded to slightly oval with the refractive thickenings usually placed centrally, 13.84µm rostrum and apex are moderately developed; a tangent drawn from the apex to the spicule tips is separated from the tangent from the apex through rostrum.Etymology: The species name is derived from the host plant.

Diagnosis and Relationships
Aphelenchoides oryzae sp.nov. is characterized by narrow cylindrical body, adults being 569.17─750.82(675.04±53.74)µmlong with a stylet about 17.3 µm long with indistinct basal swellings, a prominent median bulb and short end bulb, 4 lateral lines throughout the body, and tail straight, convex-conoid with a simple terminal mucro.
Males are common and functional with prominal spicules with dorsal limb flattened to indent in its distal half and the tip curled ventrally.Lateral fields of adults usually with four lines.
In view of its association with paddy plants, Aphelenchoides oryzae sp.nov.might be confused with Aphelenchoides besseyi Christie, 1942 the rice nematode.However, Aphelenchoides oryzae sp.nov. is separated from A. besseyi Christie, 1942 in having a single, simple, tail mucro instead of three─four processes as in A. besseyi.The present species also has longer oesophagus and stylet than A. besseyi Christie, 1942 whereas oesophagus ranges from 64─68 µm and stylet 10.0-12.5 µm in A. besseyi Christie, 1942.Aphelenchoides oryzae sp.nov.differed from other species of Aphelenchoides in having a female body length of 569.17─750.82(675.04 ± 58.74) µm, with a slender stylet length of 17.3 µm and a convex conoid tail with a simple terminal mucro with four lateral lines along the whole body length.
The differences of characters between closely related species of Aphelenchoides is provided in Table 6.Based on these morphometric differences the present species is reported as new to science.
The Shannon -Wiener species diversity index and Evenness for all the mentioned species are given in the table 7.

CONCLUSIONS
The richness of the species in the region may be due to warm climatic conditions, suitable habitats, and hosts as well as due to the absence of drastic changes in the climatic conditions during the past few years.But the present work could not cover all the varied ecosystems of Manipur.Since, nematodes are soil dwelling living around the roots of plants as well as plant parasitic

Figure 2 .
Figure 2. Aphelenchoides aerialis Chanu et al., 2013: A-Anterior part of Female body | B-Entire body of Male | C-Entire body of Female | D-Anterior body part of male | E-Posterior end of body (enlarged) | F-Tail region of Female.
contour straight to slightly curved ventrally.Lateral fields with three incisures.Cephalic framework set off without annulation.Spear 4.8-8.2µm long with indistinct basal thickenings.Oesophagous is aphelenchoid type with squarish muscular median bulb with a flat base and crescentric wave.Excretory pore at level of nerve ring.Vulva, a transverse slit with prominent lips.Oocytes are arranged in single row with a uterus.Tail 22.4-30.8µm long, rounded with ventral mucro.

The median Journal of Threatened Taxa | www.threatenedtaxa.org | 26 October 2023 | 15(10): 24063-24078 24065 J TT oesophageal
bulb is oblong with centered valvular apparatus.Oesophagous overlap intestine.Nerve ring behind the esophagus-intestinal junction.Excretory pore above nerve ring.Vagina with sphincter and raised vulval lips.Gonads monoprodelphic and oocytes are arranged in single rows.Spermatheca is filled with sperms and with uterine sac.Tail 42.5─48.2µm long,conical, and with small single mucro.Male: Body more curved at tail region.Testis long and outstretched.Spicule is typical of the genus.Dorsal limb without knob.The capitulum and rostrum very well developed.Post anal genital papillae, one pair situated above the tail terminus.Tail terminating into a long spine-like mucro.

Siddiqi et al., 1967 (Table 1 )
Diagnosis Female: Body contour slightly curved.Cuticle fine, striations about 1.7 µm at mid-body.Lip region set-off round.Lateral fields with four incisures.Stylet slender 10.2 µm long with weakly developed basal knobs.Oesophagous typical of the genus.Excretory pore at level of the nerve ring.Ovary outstretched and oocytes arranged in single row.Spermatheca oblong with

26 October 2023 | 15(10): 24063-24078 24069 J TT ovary
Chanu & Mohilal (2018)and oocytes arranged in single row reaching the basal bulb.Spermatheca elongated oval with sperms.Uterine sac filled with sperms and ventral rounded tip.Vulva protuberant and vagina at right angle to the body.Tail curved ventrally with rounded tip with a small mucro at tip.Male: Body ventrally curved.Tail conoid with mucronated lip.Testis 138.4-190.3µm.spermatocytes in single row, spicules simply arcuate, rostrum rounded, and prominent apex.Three pairs of sub ventral papillae present towards tip of spicule.Bursa and gubernaculum absent.Remarks: The morphometric details of the present species conform well with those described byChanu & Mohilal (2018).