Surface features in the of species

Donald C,M. CORBETTand Sybil A. CLARK Agricultural Research Council, 160 Great Portland Street, London, and Rothanzsted Ezperirnental Station, Harpenden, Herts.

SUMMARY Surfacefeatures of eighteenspecies of Prafylenchus wereexamined by scanningelectron microscopy. Head shape and annulation were found to begood taxonomic characters, and the specieswere separable into three groups by the pattern formed by the first head annule and oral disc (the face n). The lateral field was found to be Loo variable for distinguishing species as were the depth and distance apart of transverse striae, except is one or two species. Shape of female tail and striations around the tail tipwere stable, and helped to distinguish between species, but number of annules on the ventral surfaceof the female tail was similarfor nearly al1 species examined.

RÉSUMB Les caractères de la cuticule dans la taxonomie des esphces de Pratylenchus .Les carackèrescuticulaires de dix-huit espèces de Pratylenchus ont été examines au microscopeélectronique à balayage. Il a été observé que la forme et l’annélation de la tête étaient de bon carac,tères taxonomiques et les espèces peuvent être séparées en trois groupes d’après la configuration du premier anneau de la tête et du disque labial (la s face )>). Excepté dans un ou deux cas, les champs latéraux, ainsi que la profondeur et l’écartement des stries transverses, étaient trop variables pour caractériser les espèces. La forme de la queue chez les femelles et les striations qui entourent l’extrémité de la queue sont stableset peuvent être utilisées pourdifférencier les especes ; mais le nombre d’anneauxsur la face ventrale de la queue de la femelle étaitle même chez presque toutesles espèces examinées.

Loof (1978) inthe most recent comprehensive Materials and methods review of Pratylelzchus liststwelve diagnostic characiers that have proved reliable for distinguishing species! Of these six are surface features and there- Most of the examined inOur study were foreamenable to investigalion with the scanning culturedaxenically on excised roots in agar eleclronmicroscope (SEM). These are : number of (Mountain, 1955) or on lucerne callus in agar (Krus- lipregion annules, shape of lipregion, depth and berg &. Blickenstaff, 1964). Many of the populations dislance of transversestriae, structure of lateral were derived from single females, but in one or two field, shape of female tail and number of tail annules cases the specimenscollected from field samples inthe Iemale.Corbett and Clark(1973) were the were used. first tostudy the fronl; surface of thehead (or For examination under the SEM they were killed ‘face’) in Pratylerzchus which can be seen only with by heat andfixed in TAF (Courtney, Polley & Miller, the SEM but since their report this feature lzas not 1955) or killed and fixed in hot FP 4 : 1 (Netscher been used systematically, although oneor two species & Seinhorst,1969). Specimens were processed to have been illustrated. We bring together and examine Spurr’s low viscosity epoxy resin, coated with gold for eighteen species (Tab. 1) the surfacefeatures (fordetails cf. Clark & Stone,1975) and examined that we thinkare most useful in distinguishing with a Cambridge Instruments Stereoscan MK. IIa between Pratylenchus species. microscope at accelerating voltages of 5 or 10 MV.

Revue Néinatol. 6 (1) : 85-98 (1983) 85 D.C.M. Corbett di- S.A. Clark

Results

Thetechniques used toprepare nematodes for the SEM frequently cause some distortion ; in this study we have selectedthose which showed least collapse.

LIP REGION

The shape of the lip region showed well in SEM photographs and was characteristic for some species (Figs. 2, 3 and4) but heads were moredomed in profile thaninprepared glycerine mounts seen withthe light microscope ; however they differ Fig. 1. Diagram of ‘face’ showing oral disc. and varia- sufficiently between species ta form a useful aid to tions of first. headannule. aa = amphidaperture ; identification. P. brachyurus (Fig.2d) closelyre- ils = innerlabial sensilla ; 1s = lateralsegment ; sembles P. neglectus (Fig. 3e) inshape but has a oa = oral aperture ; od = oral disc ; sds = sub-dorsal different, face pattern. P. coffeae and P. loosi are very segment ; svs = sub-ventral segment. similar with heads continuous wit,h the body contour and resemble no otherspecies (Fig. 2e,f). P. crenatus, Table 1 P. zeae and P. sefeansis (Fig. 2j-l),P. n. sp. 1 (Fig. 3s) and P. fallaz (Fig. 4j) resemble each other in having Species of Pratylenchus examinedand their origins dome shaped heads set off from the body ; P. thornei and P. n. sp. 2 have high heads almost continuous with the body cont,our (Fig. 3k, 1) and P. penetrans, Pratylenchus Originand Host P. andinui and P. pinguicaudatus (Fig.4f, k, 1) species have the same almost cap-like head outline set on top of a wider body. P. n. sp.3 (Fig. 3j) and P. uulnus (Fig.4e) have high rounded heads and P. goodeyi andinus , Bolivia (Fig. 3d) and P. pratensis (Fig. 4d) are flattened in n. sp. 1 Oats, Rolivia * front with rounded edges ta the head outline. brachyurus Maize soil, Bolivia The number of lip region annules varied between cofleae Cit.rus, Florida species and somewhat within species. For example, P. neglectus (Fig. 3e) normally with two head annules Barley, England (two populations) crenatus was found occasionally with a partial third annule ; fallax Barley, England P. crenatus (Fig. 2j) usually with three head annules, goodeyi Bananas, England occasionally with a fourth ; P. thornei (Fig. 31~)with loosi Tea, Ceylon breaks and folds in head annules to give threeor four n. sp. 2 Alexandrian clover, Israel * depending on the viewing position or P. pinguicau- neglectus Barley, England ; spinach, Bolivia datzzs (Fig. 41), normally with three annules rkduced penetrans Arctaea,England ; apple soil, New to twoat one point on the head,or on other specimens York, USA having four head annules. The most variable species pinguicaudatus Wheat., England studiedwas P. n.sp. 3 (Fig. 3j) ; mosthad four pratensis Meadow grass, England annules but many had three on one or both sides se faensis Maize, Nigeria of ’thehead, seen by light microscopy. Variation of this kind was found in most species studied, but thornei Wheat, England ; wheat, Bolivia variability was not so large as to invalidate number vulnus Glasshouse , England of lip region annules as a good distinguishing charac- n. sp. 3 Cordyline, Scotland * ter:however it didemphasise t,he need to look at zeae Maize soil, Bolivia ; maize, Malawi ; more than onespecimen in a population to be certain maize, Zimbabwe of identifying the species. When combined, number of lipregion annules and shape of thelip region constitute good taxonomic characters with the light * Type locality and type host microscope or the SEM.

86 Reuue Nématol. 6 (1) : 85-98 (1.983) Surface fealurcs in taxonolny of Pratylenchus

~~

Fig. 2. Pratylenchus spp. heads. a-c, g-i : face view ; d-f, j-1 : profile ; a, d : P. brachyurus ; b, e : P. loosi ; c, f : P. coffeae ; g, j : P. crenatus ; h, k : P. zeae ; i, 1 : P. sefaensis. Scale bar = 2 Pm.

Revue Nèmatol. 6 (1) : 85-98 (1983) 87 D.C.M. Corbett & S.A. Clark

Fig. 3. Prafylenchus spp. heads a-c, g-i : face view ; d-f, j-1 : profile ; a, d : P. goodeyi ; h, e : P. neglecfus ; c., f' : P. n. sp. 1 ; g, j : P. n. sp. 3 ; h, k : P. thornei, arrows in h point to possible 1ocat.ion of cephalic sensilla ; i, 1 : n. sp. 2. Scale bar = 2 Fm. a8 Revue Némafol. 6 (1) : 85-98 (1983) Surface fcatures i72 faxonomg of Pratylenchus

Fig. 4. Pratylenchus spp. heads. a-c, g-i : face view ; d-f, j-1 : profile ; a, d : P. pratensis ; b, e : P. vulnus, arrows in b point to possible location of cephalic sensilla ; c, f : P. penetrans ; g, j : P. fallax ; h, k : P. andinus ; i, 1 : P. pinguicaudatus. Scale bar = 2 Pm.

Revue Nètnatol. 6 (1) : 85-98 (1983) 89 D.C.M. Corbeft LE. S.A. Clark

Only five authors indude enface views of Praty- from UK and Bolivia, P. zeae from Bolivia, Malawi lenchus indescriptions (Allen Sc Jensen, 1951 ; and Zimbabwe. Corbett,1969 ; Fortuner, 1973 ; Sher Sc Allen, Three species have incisures in some parl; of the 1953 ; Thorne,1949). The head is verysmall, less face pattern. P. n. sp. 3 mayhave a diagonal than 8 pm across atthe front, and when excised incisure across the outer part of the lateral segment. headsare viewed in transmittedlight the heavy (Fig.3g) ; in P. n. sp. 2 theventral sub-median refractive head skeleton preyents easy discernment segments are divided by an inc.isure running almost of feaf.,ures, especially at the surface. Wi‘c.11 the SEM to the oral disc (Fig. 3i) and in P. pratensis therc is ‘the front, of tbe head or ‘face’ is seen to be formed anincisure in bothsub-median segments parallel by the oral disc. and first head annule (Fig. 1). Al1 tothe outer edge and midway between t.he oral species of Pratylenchus havesix small pores very apertureand the edge of the face(Fig. 4a). The dose to the oval oral apert,ure : these are probably signific.anc.e of these ineisures is at present unclear. t,heinner labial sensilla(Coomans, 1979). On each Inaddition, t.he head annulesin P. pratensis are side of the oral aperture, on t.he inner edge of the wavy in outline. first head annule t>here is an amphid opening varying in size according to species, usuallyslightly dorsal to a linebisecting the oral aperture laterally. In DEPTHOF TRANSVERSE STRIAE AND WIDTH OF ANNULES many species the amphid opening is oblique, slanting inwards towards the dorsal surface. Insome speci- menssmall depressions can be seen on t,he outer The body annules are separated from each other by portion of the sub-mediansegments of the first tsansverse striae, t,he depth of which may appear to headannule ; thesemay indicate the posit,ion of Vary widely. Theirdistance apart, corresponds to the c,ephalic. sensilla (Fig. 3 hl 4 b) (Coomans, 1979). i.he width of the annules. Four species were examined under the light microscope : P. crerzafus, apparent,ly The ‘face’ is undivided, or divided intosub- coarsely annulatehad striae 0.50 to 0.67 Fm deep medianand lateral segments, and the, different. and 1.3 to 1.4 pm apart. ; P. goodegi, finely annulate, arrangements faIl intothree groups. In Group 1, had striae 1.2 to 1.5 pm apart. and 0.33 pm deep ; the nematodes have a plain undivided face with no P. neglpcfus, one of t.he mostwidely dist.ributed division betweensub-median and lateral segments, and variable species, had striae 1.2, to 1.4 pm apart presumablydue to fusion of the first headannule and0.33 pm deep ; and P. fhornei, probablythe to t,he oral disc.. These are : P. brachyurus, P. coffeae, most finely annulate, sometimes appearing ho have crenatus, goodeyi, loosi, sefaensis and P. P. P. P. a smooth cut,icle,1.3 to 1.6 pm apart and 0.22t.o P. zeae (Figs. 2a-c, g-i, 3a). In these nematodes there 0.33 pm deep. were differences in shape, size and orientation of the Annulation varies as shown in Figures 5, 6 and 7 ; amphid apertures, but,ot.herwise the st.ructureof the while thedistance between striae can easily be ‘face’ was similar. measuredin SEM phot,ographs, the depth of striae In other species examined, the area immediat,ely cannot. Table 2 lisk the width of annules of most surroundingthe oral apert,ure was similar, but the ofOur species measuredon SEM photographs at outer part, of the face was sub-dividedinto sub- mapifications from 2 500 x to 12 200 x. The SEM medianand lateral sectors. Group 2, consistingof photographs also mnke clear that differences in P. n. sp. 1, P. n. sp. 2,, P. nrglectus, P. thornei and form of annulationoccur, some e.g. P. crenatus P. n. sp. 3 has sub-median segments that are fused and P. zeae Ilavingrounded annules wit-h deep tothe oral disc. andare narrower at, th&inner striae between (Fig. Gd, e) and others e.g. P. thornei extremitywidening towards the outer edge of the and P. pinguicaudafus being alnlost flat with shallow face : theyare separated from t.he larger lateral st,riae bdwecn tjhem (Fig. 6j, p). segments that complete t.he circularface, wit,h amphiclopenings on their inner edges (Fig.3b, c, g-i). Group 3, whic.h includes P. andinus, P. fallas, P. penetrans, P. pingnicaudafus, P. prafensis and STRUCTUREOF THE LATERAL FIELD P. vulnus, has a more distinctive dumb-bel1 shaped pattern of thesub-median segments with slightly This is a character much used to distinguish species smallerlateral segments to complete the circle. of Pratylenchus. It is dific.ult to see c,learly in trans- The amphid apertures are again on the inner edges mitted light,, despite the useof specialised techniques of thelateral segments (Fig. 4a-c,g-i). Theseface (De Grisse,1961). The SEM, however, shows the formations were consistent for diflerent populations lateral field up so clearlyt3hat it isnow possible of severalspecies : P. penetrans from the UK and for the first time to evaluate its worth as a character the USA, P. thornei from UK and Bolivia,P. neglecfus ta be used in identification.

90 Reuue NCmatol. 6 (1) : 85-98 (1983) Fig. 5. Pratylenchus spp. lateral fields. a : P. zeae, showing origin of lateral field ; b : P. brachyurus ; G, d : P. lOOSi ; e : P. neglecfus, showing origin of lateral field ; f : P. coffeae ; g : P. goodeyi ; h, j, lc : P. neglectus ; i : P. thornei, showing origin of lateral field ; 1 : P. fhornei ; m : P. n. sp. 2, showing origin of lateral field ; n : P. n. sp. 2 ; O : P. prafensis ; p : P. vulnus ; q : P. penefrans, showing origin of lateral field ; r : P. penetrans ; s : P. fallax ; t : P. pinguicaudafus.

R evue Nématol. Revue 6 (1) : 85-98 (1983) 91 Fig. 6. Pratglenchus spp. tails. a : P. brachyurus ; b : P. loosi ; c : P. cofieae ; d : P. crenafus ; e : P. ;eue ; f : P. sefaen- sis ; g : P. goodeyi ; h : P. neglectus ; i : P. n. sp. 1 ; j : P. fhornei;li : P. pratensis ; 1 : P. vulnus ; m : P. penefrans ; n : P. fa1la.r: ; O : P. andipus ; p : P. pinguiraudafus. Scale bar = 2 pm.

92 Nèmatol. Revue 6 (1) : 85-98 (1983) Fig. 7. a-g : Pratylenchus spp. tails. a : P. loosi ; b : P. zeae ; G : P. goodeyi ; d : P. n, sp. 1 ; e : P. n. sp. 3 ; f : P. pratensis ; g : P. penetrans ; h, i : Pratylenchus spp. tail tips, showing tiling on upper surface ; h : P. coffeae ; i : P. pinguicaudatus ;j : P. thornei from Bolivia and k : P. thornei from Britain, showingpores near lip region ; 1 : P. fallax spicules, inset showing poresat tip.

Revue Nèmatol. 6 (1) : 85-98 (1983) 93 D.C.M. Corbett & S.A. Clark

Table 2 especiallyon thetail (Fig. 6) thanin the middle third of the . Width of annules in mid-body of species of Prafy- lenchus, measuredfrom SEM photographs at point Fewdescriptions of species contain a sufficiently where annules join lateral field detailed description of the lateral field to enable it to beused as a differentialcharacter. Loof (1960) has discussed t>hisin some detail. Pratylenchus Annules width Annules widfh, species range (Fm) mean(Fm) TAILSHAPE AND ANNULATION andinus 1.4-1.9 1.6 The shape and amount of annulation or crenation brachyurus 0.9-1.5 1.4 of the female tail is includedin al1 descriptions of Prafylenchus species.The SEM picturesshow, for cofteae 1.2,-1.5 1.4 the most part, that descriptions have been accurate. fallax 1.3-1.9 1.4 Theyalso show that there is some variability but goodeyi 1.0-1.3 1.2 that the shape of the tail and presence or lack of loosi 0.9-1.4 1.1 annulationand its form around the tail tip are n. sp. 2 1.3-1.6 1.4 suffhientlycharacteristic foreach species tobe a neglecfus 1.0-1.8 1.6 good taxonomiccharacter. Fig. 6 shows thetail penefrans 1.4-1.8 1.5 tips of al1 the spec,ies we shdied except P. n. sp. 3 pinguicauda fus 1.2-1.6 1.4 and P. n. sp. 2 which are illustrated in Corbett (1982). praiensis 0.9-1.2 1.1 Somewrinkling of thetail tip has occured in al1 specimens but it ispossible todistinguish this sefaensis 0.8-0.9 0.8 processing artefactfrom the ' crenation'around thornei 0.9-1.9 1.4 thetip characteristic of P. crenafus, P. pratensis zeae 0.7-1.1 0.9 and P. fallax (Fig. 6d, k, n). The tail shape deswibed ascharacteristic for each speciesshows up well in SEM photographs,generally agreeing with the original authors' descriptions.The lateral field is particularly clear in most specimens ; it extends to the tail tip in manyspecies and seems to continue In Prafylenchus sp. the lateral field starts on the round the tail tip inP. brachyurus, P. lossi, P. thornei seventh to ninth body annule (Fig. 5 a, el il ml q) and P. pinguicaudafus (Fig.6a, b, j, p) and stops and for the greater part of its length has four lines, short of the tail in P. crenatus (Fig. 6d). Where it i.e. consists of three bands (Fig. 5 b-dl f-h, j-1, n-p, is visible, the phasmid opens asa pore about midtail, r-t). It ends on the tail, in many species continuing and slightly to the ventral side ofLhe lateral field. to the tail tip (Fig. 6). Across or lvithin the bands It is very conspicuous in some species, e.g. P. vulnus theremay be further lines whic,h aresaid to be (Fig. 61). characteristic for each species, but which we found to Vary greatly,possibly depending on age of the specim ens examined. specimens .A NUMBER OF TAIL ANNULES We found thatal1 species we studied had areolat,ion of theouter bands (Fig. 5). Severalspecies had Number of annules on the ventral surface of the part,ial or complet,e areolationof the middle band tailbetween the anus and the tail tip is the final also : P. brachyurus, P. loosi, P. neglectus and P. feature noted by Loof (1978) as having some value pinguicaudatus (Fig. 5b, c,hl t).Two species (P.loosi inidentifying Pratylenchus spp.This annulation is and P. uulnus) are illustrated with oblique striae in easily counted under the compound optical micros- the middle band (,Fig. 5d, p) : we also found these cope and very easilyseen in the SEM (Fig.7a-g). in P. crenafus, P. n. sp. 1, P. n. sp. 3 and P. zeae. Wefound (Tab. 3) that t,ail annulenumber varied Theform of lateral field variedgreatly within widely within species and there was some variation species, especially P. neglecfus, and similarities were between species.Mean number of annulesvaried foundbetween species ; P. neglectus and P. fallax from 15 for P. pseudoprafensis (range 12-19) to a showed thesame complex ornamentation of the mean of 27annules for P. sefaensis (range 24-34) middle band (Fig. 5j, s). Areolation across the lateral at the extremes of the range, therefore tail annules field ismore frequenton the anterior part of the canaid in distinguishing species. However,most field (Fig. 5a, el il ml q) andbehind the vulva, species have a meannumber of tailannules that

94 Revue Nèmatol. 6 (1) : 85-98 (1983) Surface features in taxonomy of Pratylenchus fallswithin the range 18-24 annules,with ranges ‘face’ patternand ornamentation of thelateral that overlapped much. It is impossibleto differentiate field in the fore and mid-body regions as was found between these species using number of tail annules in females of thesame species. Thebursa appears as a character. It was found during the examination to be a rearward and ventrad extensionof the lateral thatthe width of tailannules varied widely field. The phasmid appears as a small pore opening (Fig. 7a-g). on the external surface of the bursa connecting to thebody through a short tube within the bursa (Fig. 71). Wefound two pores at the extremity of Table 3 the spicule(Fig. 71, inset),mentioned, but not Number of annules fromthe tail tipon which phasmids illustrated,by Wen and Ghen (1976). are found, and number of annules on ventral surface of tails of Pratylenchus spp. OTHER SURFACE FEATURES

Phasmidsshow clearly in SEM photographs and Pratylenchus Phasmid Number lMean areusually found in the middle of thelateral field species annule species of annules about half way along the tailof females. The number of annules frim thevphasmid to the tail tip counted along the ventral surface is given for a number of alleni - .15-19 * 17 * species in Table 3. As this tended to Vary approxi- andinus 8-12 17-20 18 matelyas the number of tailannules, we do not n. sp. 1 9-15 13-24 18 think it any better as a distinguishing character. brachyurus 8-10 15-21 18 Thereis uncertainty about deirids, small pore cofleae 9-13 17-22 20 likestructures usually on the lateral field inthe convallariae - 16-20 19 ’ oesophageal region. Sher and Allen (1953) described crenatus - 20-24 22 * the genus as having deirids, but Roman and Hirsch- fallax - 16-26 * 21 * mann (1969) couId not find them on six species they flakkensis - 18-24 * 21 * studied. Loof (1978) doubts their presence in Praty- lenchus, and we did not find them in the eighteen goodeyi 10-14 20-27 * ; 19-24 24 * 22 species we studied. It is probable therefore that they loosi 11-15 19-26 23 do not occur in Pratylenchus. n. sp. 2 15-22 18 We found in oneor two species, notably P. thornei, neglectus 8-12 16-19 ; 11-19 17 * 16 that there werepores on or near the head. In the penetrans 8-12 15-27 ; 17-25 20 * 21 Bolivian population of P. thornei a large pore opened pinguicaudatus 8-12 18-25 21 usually in the basal head annule on the ventral side. pratensis _- 20-26 * 24 A similar pore was found in the UK population of pseudopratensis - 12-19 15 * thisspecies, but it moreusually occurred on the scribneri - 21-25 * 23 * first or second body annule than on the head (Fig. 7j, se faensis 12-18 24-34 27 k). We do not know what function they serve nor do we know if they connect with structuresor organs thornei - 19-26 * 23 * in the nematode. - vulnus 22-26 * 24 * We also foundin two species, P. coffeae and P. n. sp. 3 12-15 16-20 18 pinguicaudatus, thatthe body annulation was zeae - 25-27 * 26 * interrupted near the tail tip (Fig. 7h & i) by some longitudinalincisures that gavea tiled appearance to this region. It was an inconstant feature and not * Data from Seinhorst (1968). noted in other species.

Discussion MALES Lip region annulation as a character for separating We studiedmales in the following species in specieswas first introduced by Allen andJensen which they occur commonly : P. coffeae, P. fallax, (1951) andhas become important in diagnosis. P. goodeyi, P. lossi, P. n.sp. 2, P. penetrans and Wehave confirmed andillustrated the variability P. vulnus. In al1 species the males hadthe same found within the species but considerthat the number

Revue Nématol. 6 (1) : 85-98 (1983) 95 D.C.M. Corbett & S.A. Clark of lipannules is constant enough to form a good However, a basic pattern was a determinable charac- specific taxonomiccharacter. However, the varia- teristic for each species, and could be used reliably bility we found does emphasise the need to examine ta differentiatethem. This character is so reliable several in a population to be certain of identifying thatmixtures of closely similarspecies could be the species. distinguished using the SEM before light microscopy Shape of the lip region we also found ta be charac- revealed that more than one species was present. teristicfor certain species, e.g. P. brachyurus, P. Anderson and Townshend (1980) examined popu- thornei, althoughthis feature calls for very well lations of P.penetrans obtained from several different prepared specinlens for SEM photography ; it may hostsand related differences inface patternsto be betterappreciated under the light microscope host differences. Wefound atthe outset of Our wherethe head is seen in profile intransnlitted study that processing for SEM viewing after speci- light and the head skeleton, whichgives the head its mens had been processed to glycerine for permanent shape, can also be seen. Taken together with number mountingon slidesled to collapse of thehead of lip region annules this makes a very good taxo- cuticlearound the framework, and the consequent nomic feature. stretching of the cuticledistorted the pattern, In SEM photographs the front of the head (face) frequentlygiving rounded margins tothe sub- differs completely from earlier descriptions made in median segments. Our results show that the variation transmittedlight and provides an interesting new betweenpopulations is not as great as Anderson character.The specieswe haveexamined fa11 at and Townshend(1980) found withtheir glycerine present into three groups on the form of the ‘face’, processedspecimens, andthe similarity between althoughexamination of more species mayshow thepatterns we foundfor P. corfeae, P. fallas, there to be only two groups, with no discontinuity in P. neglectus, P.’penetrans, P.thornei and P.oulnus face pattern between the types present in Groups 2 andthose recorded by other researchers working and 3. Workin other nematode groups (Hooper with populations collected elsewhere (Momota, 1979 ; & Clark,1980 ; Stone,1975) suggests that form of Sher & Bell,1975 ; Townshend & Anderson,1976) the head provides dues to phylogeny. InPratylenchus shows that the patterns are invariant enough to be morphological similarity has been the main criterion good taxonomicfeatures. However, we believe. a of closeness of relationships. In the species we have study of the influence of hostand other external looked at it is difficult torelate face patternswe conditions on head pattern should be carried out. on havefound ta presumed relationships. In Group 1 several species todetermine the amount of varia- P. coffeae and P. loosi aremorphologically very bility that can be expected. similar butare unrelated to any of theothers in It is dificult to postulate the anatomical features that group.There is no other evidence torelate of the ‘face’. De Grisse’s (1977)photographs and P. crenatus to P. coffeae or P. loosi, and P. brachyurus drawings of P. penefrans showeddeep divisions and P. zeae aremorphologically as dissimilar as between an oral disc and lateral plates. Loof (1978) any two species in the genus : P. crenatus (Group 1) duplicated his findings, including the assertion that more closely resembles females of P. fallaz (Group 3) only four of the six inner labial sensilla are on the than any other species. In Group 2 also there ‘seems front surface of the oral disc. We believe DeGrisse little in common in the morphology of the species. (1977) misinterpreted folds caused by collapse O€ the InGroup 3 P. fallax and P. penetrans aresimilar headcuticle around the head skeleton as divisions morphologically. Workby Tarte and Mai (1976) of the face. Our photograph of the uncollapsed suggests that the features used to distinguish these head of P. penetrans shows anoral aperture with species, in particular tail shape and annulation are six adjoining sensilla ; we found these to be common unreliable and one species may bea morphological to al1 the species of Pratylenchus studied. We show variant of the other. However, Perry, Plowright and photographs of P. fhornei and P. aulnus that suggest Webb(1980) failed toobtain fertile offspring €rom that the four cephalic sensilla lie on the sub-medlan crosses of P. penetrans and P. fallax and concluded segmentsjust outside the circle of the oral disc thatthey wereseparat.e species. P. andinus and but do not terminate in pores like the inner labial P. pinguicaudalus closelyresemble each other in sensilla.Stone (1975) called the sub-median fused this group, but differ slightly in ‘face’ pattern and segments of Heterodera juvenilessub-median lips inother respects. The others in this group are andthe structure bearing the amphids the lateral unrelatedon morphological grounds. With obser- lips. Hooperand Clark (1980) spoke of a cephalic vations of ‘face’ patternsin more Pratylenchus platepattern in their studies of Aphelenchoidea. species true pkiylogenetic relationships may emerge. For convenience anduntil it isproven otherwise, We found some variability in pattern within popu- we prefer to regard the oral disc as that part in the lations of the same species and between populations. centre of the ‘face’ extending out to near the amphids

96 Revue Nèmatol. 6 (1) : 85-98 (1983) Surface features in taxonomy of Pratylenchus andto which are fused to form different patterns preferablywith the aid of theSEM, and consider the sub-medianand lateral segments of thefirst that because of the intraspecific variability we found head annule, the whole forming a cephalic plate. in the lateral field this is too variable to be of use Mean depth of striaevaried from 0.22 Fm to as a good taxonorniccharacter in differentiating 0.67 pm in four species, a very small amount despite Pratylenchus. theapparent large difference betweenspecies in Tai1 shape varies between species quite consider- depth of striae,which is usually associated with ablyand although there is variation within al1 annuleshape. This difference, less than 0.5 Fm, species it ispossible to recognise a shape of tail is smaller than the probable error in measuring. It characteristicfor most species (Fig.6). Some have was not possible to measure it in SEM photographs. striationsaround the tail tip (Fig. 6d) or more It is unrealistic to use this difference quantitatively irregular crenations (Fig.Gn), that are distinguishable but it hasusefulness as a subjective character. from the wrinkling that occurs as a processing arte- Width of annuleswas easily measured on SEM fact. As far as it was possible to tell with the speci- photographs at magnifications of 2 500 x to 12200 >i mens we photographedtail shape is a relatively andshould be more accurate than measurements stablecharacter. However, Tarte and Mai (1976) made under the light microscope. Annule width in and Townshend, Tarte and Mai (1978) have shown fourteen species was unexpectedlyvariable. This thatthis is notsuficiently stable in P. penetrans wasfound to bedue to the presence in several to enable distinction between it and P. fallax. This species of doubleannules - annules of normal characterneeds to be re-examined in specimens width witlz an extra line in the middle (Fig. 5j, r). raised in controlled conditions for these two species, Thistogether with the almost complete overlap and also in others where tail crenation is an important of the range of annule widths in al1 species (except character distinguishing closely similar species. P. sefaensis, verynarrow annules) makes this a In at least two species (P. coffeae and P. pingui- character of little use indistinguishing species. caudatus; Fig. 7h & i) an unusual form of break in Structure of thelateral field hasreceived much annulation just above the tail tip occurred to produce emphasisin Pratylerzchus taxonomy,despite the a ‘tiled’appearance. This feature may be confined dificulty of observation and the form of the. lateral to these species, in which case it could form a useful field hasbeen used as theprincipal differentiating accessory character. Other species should be examin- featurein P. hexincisus. Our worksuggests that ed for its presence. thelateral field in Pratylenchus basicallyconsists Number of annules on the ventral surface of tails of four lines and that a variety of ornamentation of has been used by Seinhorst (1968), and endorsed by thelateral field isfound in so many species as to Loof (1978), to differentiate species of Pratylenchus. make its form a poor distinguishing character. For Wefound much intraspecific variability in this example,oblique striae in the lateral field have character with many species having between thirteen beenused as supplementarydifferentiating charac- and 24 annuleson the ventral surface of thetail ters and Loof (1960) found P. neglectus, P. perzetrans, (Tab. 3). Although one or two species differed from P. thornei, P. loosi and P. coffeae to havethem. the norm in the number of annules on the tail, we Otherworkers have discovered this character in think that itis not a usefuI character in differentiat- P. n. sp. 1, P. n. sp. 3, P. neglectus and P. heximisus ing species. (Corbett,1982 ; Fortuner,1973 ; Sher & Allen, 1953 ; Taylor & Jenkins, 1957 ; Townshend & Anderson,1976). We foundother ornementation, and especiallyfound areolation of the outer bands of the lateral field to be common.Examination in thelight microscope showed P. rzeglectus to have four,six or sevenlines in the lateral field inmid- body, and P. crerzatus was often found to have six lines. We were able to examine paratype specimens ACKNOWLEDGEMENTS of P. hexincisus and found them to have basically four lines in the lateral field, in common with other We thank Dr. J. O’Bannon for P. cofieae cultures ; species,with oblique striae in mid band in some Mr. D.N. Greet for samples of P. Zoosi; Dr. D. Orion specimens, and in othersa broken line in mid band to for P. n.sp. 2 ; Prof. W.F. Mai for P. penetrans ; Dr. F. Caveness for P. sefaensis; Mr. J. Shepherd for give five lines : we did not find six Jines in the para- P. zeae from Zimbabwe ; Dr. A. M. Golden for paratype types, which was one of the main features on which slides of P. hexincisus and Mr. R.M. Webb for main- the species was erected. We consider therefore, that taining and supplying axenic cultures of al1 the nema- the status of P. hexincisus has to bere-examined, todes we used.

Revue Nkmatol, 6 (1) : 85-98 (1983) 97 D.C.M. Corbett & S.A. Clark

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