Fine structure of the posterior cone of females of Cactodera cacti Filip’ev & Schuurmans Stekhoven (Nemata-: ) Diogenes A. CORDEROC.*, James G. BALDWIN**and Manuel MUNDO-~CAMPO** *University of Panama, Facultad Ciencas Agropecuarias,Apartado ZB, David, Chiriqui, Republic of Pananza, and **Department of Nenzatology, University of California, Riverside, CA 92521, USA.

SUMMARY

Detailed development of the posterior cone of females of Cactodera cactigives insight into phylogenetic characters which differ from previously described Heterodera schachtii. Cone growth continues after the final molt in H. schaclztii, but not in C. cacti. Although a gelatinous matrix is produced in both species, H. schachtii deposits eggs whereas C. cacti does not. Contrary to H. schachtii, the body Wall cuticle of the cone in C. cacti includes a D layer but lacks andE layer and bullae.The fenestral region of C. cacti lacks the cuticular fibers presentin the same region of H. schachtii. Unlike H. schachtii, the cone of C. cacti has a short vagina with no underbridge, andthe vaginal musculature is greatly reduced. These vestigial vaginal muscles are closely associated with the cyst Wall as denticles.

~WUMÉ

Structure du cône postérieur des femelles de Cactodera cacti Filip’ev & Schuunnans Stekhoven (Nemata :Heteroderinae) L’étude détaillée du développement du cône postérieur des femelles deCactodera cacti apporte des élémentssur des caractères phylogéniques différant de ceux décrits précédemment Heterodera chez schachtii.La croissance du cône se poursuit après la dernière mue chez H. schachtii, mais non chez C. cacti. Encore qu’une gelée soit produite par les deux espèces,H. schachtii pond des œufs à l’extérieur, ce qui n’est pas le cas de C. cacti. A l’inverse de H. schachtii, la cuticule du cône de C. cacti possède une coucheD, mais il n’y existe ni couche E, ni bullae. La région des fenêtres de C. cacti est dépourvue des fibres cuticulaires que l’on observe dans la région homologue deH. schachtii. A l’inverse de H. schachtii, C. cacti présente un vagin court, dépourvu de sous-pont, et la musculature vaginale est très réduite. Ces muscles vaginaux vestigiaux sont étroitement associésà la paroi du kyste, en tant que denticules.

Cactodera cacti (Filip’ev & Schuurmans Stekhoven, reported as rare in C. cacti, consequently, measurements 1941) Krall & Krall, 1978, first described from orne- of Adam (1932) have been repeated, apparently without mental cacti in Holland, is distributed worldwide and consideringadditional specimens (Filip’ev & Schuur- considered indigenous to Mexico(Adam, 1932; Cooper, mansStekhoven, 1941; Franklin, 1951; Mulvey & 1955~; Minz,1956; Southey, 1957; Kumar,1964; Golden, 1983). There is concern that putative Cactodera Golden & Raski, 1977; Scognamiglio, Caputi & Ciancio, males have been misidentified because mixed popula- 1985; Ebsary, 1986). Cactodera damagescertain cacti tions of Cactodera and Punctodera Mulvey & Stone, grown as foodcrops in Mexico(Mundo-Ocampo, 1976, or Meloidogyne (Goeldi, 1887) Chitwood, 1949, unpubl.) as well as ornamental cacti (Langdon & Esser, are frequentlyfound in thefield (Spears,1956; Southey, 1969; O’Bannon & Esser, 1970; Hamlen, 1975; Mit- 1957; Schneider, 1961). Cooper (1955~)~noting cyst chell, 1985). Since its host range also includes families variability, considered his collections of putative C. cacti other than Cactaceae, C. cacti may damage a variety of to be mixed, and therefore, excluded the species from other crops (Scognamiglio, Caputi & Ciancio, 1985). the key to British Heterodera Schmidt, 1871. Despite the parasitic potentialof C. cacti, little atten- The genus Cactodera was erected by Krall and Krall tionhas been given toits biology. The life cycle of (1978) to accommodate cyst-forming, circumfenestrate, C. cacti has not beendocumented, andthe relative lemon-shaped Heteroderinae with C. cacti as the type abundance of males is unclear. Adam (1932) regarded species. Diagnosis of Cactodera is based primarily on the six males of the original descriptionas inadequate for vulval-cone relatedstructures; ,an emendationadds obtaining reliable measurements. Males continuedto be presence of vulval denticles, but excludes presence of

Revue Nématol. 14 (3) :455-465 (1991) 455 D. A. Cordero, J. G. Baldwin & M. Mundo-Ocampo

~ bullae and underbridge (Krall & Krall, 1978; Mulvey& stored at 4 "C for a period of 2 months to 3 years. To Golden, 1983). More recentlyemendations include induce hatching, cysts were incubated for 10 days at presence of a D layer inthe femalecuticle (Luc, 28 "C in a sterile BPI dish (containing fourto five drops Maggenti & Fortuner, 1988). Many proposed diagnostic of sterilewater) in a sterile Petridish sealed with characters of Cactodera also occur either in Heterodera Parafilm 0, Second stagejuveniles and cysts were added (e.g. vulval cone) or Globodera Skarbilovich, 1959 (Beh- close to roots, using a Pasteur pipette. rens, 1975) (e.g. vulvaldenticles, circurnfenestration, Second stagejuvenile infection was monitored using a D layer, absence of bullae and underbridge). Thorough Nikoninverted microscope equipped with Hoffman understanding of traditional and new characters is re- interference opticsand a Garr time-lapse high resolution quired, so that the phylogenetic relationships of Cacto- videorecorder as previously described(Cordero & dera to other cyst canbe established asa basis Baldwin, 1990). Development of a specimen wasre- for an increasingly meaningful classification(Baldwin & corded and studied or stopped at particular points and Schouest, 1990). the specimen was excised for detailed examination with Recently, Cordero and Baldwin (1990, 1991) described LM, SEM, or TEM. detailed developmental morphology of the cone of He- Transmission electronmicroscopy and LM were used terodera schachtii Schmidt, 1871, and elucidated charac- to study interna1 cone structuresat the following points ters useful in taxonomy and phylogeny of Heteroderi- of development : immediately afterthe fourthmolt, two nae. Findings in H. schachtii will be of greater phylo- and four weeks after the final molt, and in the Young genetic significance when theycan beinterpreted in the cyst. The samestages were examined with SEM to context of knowledge of the cone and posterior terminus follow surface pattern developmentof the posterior end. in additional Heteroderinae. Presently,we report on the Specimenpreparation for LM and TEM was as detailedstructure of the cone of C. cacti. Unlike previouslyreported (Cordero & Baldwin, 1990). Ex- H. schachtii, the cyst of C. cacti has a reduced cone with cised cones were fiied for 8 h in modified Karnovsky's one circurnfenestra, al1 eggs are retained and thefemale (1965) fiiative,8 h in glutaraldehyde-hydrogen peroxide has a pronounced D layer (Cliff & Baldwin, 1985). solution (Byard, Sigurdson & Woods, 1986), and 3 h in Development of the cone of C. cacti is investigated for osmium tetroxide fumes. Dehydration was done in a comparison with H. schachtii using a combination of graduatedethanol series andinfiltration in Spurr's light (LM), scanning (SEM) and transmission (TEM) epoxy. Thin (silver grey) sections for TEM and semi- electron microscopy. thick (0.28 Pm) sections forLM were stained as reported by Baldwin (1983). Specimens were prepared for SEM by glycedn infiltration (Sher & Bell, 1975). Materials and methods To visualize actin, and thus thepresence of muscles inthe cone, females were treatedwith fluorescence Cactodera cacti was established on Schlurnbergera sp. phalloidin (FITC labeled) following the protocol out- Lem, 1858 (Christmas ) by in vitro culture utiliz- lined by Wulf et al. (1979). Cones labeled with FITC ing sterile techniques. Christmas cactus cladodes (leaf- were prepared as describedby Cooper (1955 b) and like modified stems) were washed brieflyin commercial observedwith a Zeissphotomicroscope III equipped detergent and disinfestedin 10 O/o commercial bleach for with epifluorescence optics. 4 min.After rinsing in sterile water, cladodes were transferred to Petri dishes containing Gamborg's B-5 medium, pH 6.5 plus White's organics (Gamborg et al., Results 1976; White, 1943). Gelrite0 (0.25 "O) was used as a gelling agent. Monoxenicculture of C. cacti is simple, allowing Rooting of Christmas cactus cladodeswas induced by ample reproduction, and permitting continuous obser- incubating cultures at 25 "C under 16 h illumination per vation of the life history. Postembryogenesis of C. cacti day (3 O00 lux, GroLuxB light) for a month. Subcul- fromJ2 to adult female is completed 12 daysafter tures of axenically produced cladodes produced roots establishment of a feeding site and resembles that of 10 days after incubation. H. schachtii (Raski, 1950), except that no males were Schlumbergera cladodes were inoculated with 25 se- observed in C. cacti. cond stagejuveniles (J2) or onemonoxenically produced The final molt begins three to four days after shed- cyst. Secondstage juveniles were disinfested by four ding the thirdstage cuticle.The newly molted adult has rinses withsterile water followed by incubation for a shallow broad cone(Figs 1 A, 2 A). The vagina moves 10 min with saturated aqueousRifampicin0 solution. A sinuously in conjunction with contraction of the entire final rinse with sterile water was given to J2s prior to cone. A clear gelatinous matrix is secreted through the inoculation.Incubation of culturesproceeded as de- vulva. Body volume of the increases rapidly scribed above. after molting, particularly at the midbody as eggs de- Cultureswith monoxenically produced cysts were velop. Thus, the previously flask-shaped body becomes

456 Revue Nématol. 14 (3) :455-465 (1991) Posterior cone of Cactodera cacti

A,E 100pm- B-D lOpm F 1Opm -

Fig. 1. Schematic representationof cone of newly emerged female and cyst ofCuctoderu cucti.A : Outline of newly molted female. Box area corresponds to area depicted inB; B : Longitudinal section through vaginaof newly emerged conein lateral orientation; C : Cross section near cone terminusof young female;D : Cross section at level of sphincter vaginae(SV) muscles of young female; E : Outline of cyst. Box area corresponds to area depicted in F; F : Longitudinal section through posterior terminus of cyst in dorsoventral orientation. (AnD = anal depressor muscle, DV = dilutores vaginue muscles, Eg = egg, MR = muscle remnant, R = rectum, SV = sphincter vuginue muscle, U = uterus, Va = vagina). D. A. Cordero, J. G. Baldwin & M. Mundo-Ocampo

Fig. 2. Cactodera cacti.A : SEM of surface pattern of vulval cone of young female;An = anus, VC = vulval crescent,VL = vulval lip; B : SEM of surface pattern of vulval cone of cyst (An = anus, V = vulva, VC = vulval crescent); C : TEM of cross section of body Wall cuticle near base (anterior end) of cone of young female (letters A, B, C, D indicate corresponding cuticular layers, H = hypodermis); D : TEM of cross section of body Wall cuticle near base of cone of yovng female, labels as in C; E : TEM of cross section of body Wall cuticle near cone terminus of mature female showing altemating bands of layersD and modified bands of B (A3 = zone of A layer, VS = vulval slit); F : TEM of cross section of bodyWall cuticle showing enlargement of layerD; G : TEM of cross section near cone terminus showing vulval bridge (VB) and vulval slit (VS).

458 Revue Nématol. 14 (3) :455-465 (1991) Posterior cone of Cactodera cacti

Fig. 3. Cactodera cacti. A : TEM of longitudinal section through vagina (Va)of newly emerged cone in lateral view (letters A, B, C, D indicate corresponding cuticular layers, DV = dilutores vaginue muscles, H = hypodermis, U = uterus); B : TEM of longitudinal section through rectum(R) of newly emerged cone in lateralview (arrowhead = anal depressor muscle, lettersA, B, C, D indicate corresponding cuticular layers, DV = dilutores vuginae muscles, EL = electron lucent matrix, H = hypodermis, I = intestine, Nu = hypodermal nucleus); C:LM of female cone incubated FITCin labeled phalloidin; lateral orientation showing positive fluorescence as an indicator of actin (An = anus, DV = dilutores vuginae muscles, Eg = egg, V = vulva; D : TEM of longitudinal sectionof vaginal lining and musculature in lateral view, DV= diZutores vuginae muscles, He = hemidesmosomes, SV = sphincter vaginue muscles, VaC = vaginal cuticle); E :TEM of longitudinal section throughsphincter vuginae muscles (SV) showing filaments. D. A. Cordero, J. G. Baldwin & M. Mundo-Ocampo

Fig. 4. Cactodera cacti. A : TEM of longitudinal sectionin lateral view through terminusat junction of vagina (Va) with uterus (U) in Young female (SV = sphincter vaginae muscles); B :TEM of longitudinal sectionof female and cross sectionof uterus (v), lining of uterus composed of large cells with distinct nuclei (Nu); C : LM of longitudinal sectionof cyst cone terminus in lateral view (arrowheads correspond to modifed cuticle in region of fenestrae) (A3 = corresponding zone of cuticle); D : TEiM of cross section at base of cyst cone (A3 and C correspond to layers and zoneof cuticle, MR = muscle remnant); E : LM of cross section at base of cyst cone (MR = muscle remnant, R = rectum); F : TE" of muscle remnant in cyst showing persistent filaments.

460 Nématol. Revue 14 (3) :455-465 (1991) Posterior cone of Cactodera cacti lemon-shaped including a distinct small posterior cone beneath the BW cuticle, from the vagina near the cone (Figs 1 E, 2 B). Afterthe final molt, there is little terminus anteriorly tothe Wall near the baseof the cone additional growth of the cone. The mature female is (Figs 1 B, 3 A). A heart-shaped sphincter vaginae (SV) filled with eggs which are retained within the body. Al1 muscle encircles the vagina approximately 16 pm ante- eggs within cysts contain juveniles and thesurface of the rior tothe vulva (Figs 1 B, D; 3 D; 4 A). Vaginal eggs has punctations. Young eggs within females that musculature is characterized by finefilaments and contain developing embryos lack punctations. presence of actin was confirmed by staining with fluor- Four weeks after thefinal molt, growthat themiddle escent phalloidin (Figs 3 C, E). Muscles attach across of the bodycontinues and the frequency of vaginal the hypodermis to the cuticle of the vagina and BW by movement decreases, stopping about 9 weeks after the distinctjunctional complexes and hemidesmosomes finalmolt. At thistime the vulval area of the cone (Fig. 3 D). In thecone of the cyst, muscle remnants with terminus collapses (Fig. 2 B). The resulting depression distinct fibers persist (Figs 1 F, 4 D-F). excludes the vulval “ domed ” crescents sensu Green In additionto the vaginal muscles, a set of anal (1975). The surface of the posterior terminus of the depressormuscles attaches tothe rectum and runs young cone, including the crescents,is relatively smooth obliquely beneath the BW cuticle anteriorly tothe cone (Fig. 2 A). As the female matures, small ridges on the base (Fig. 3 B). The BW cuticle is thin at the shallow crescents surround the sunken vulval lips; these ridges depression surrounding the anus; layer C and zone A3 are retained in the cyst (Fig. 2 B). In aging females, the areparticularly reduced and D is predominant. The regionsurrounding the anus becomes progressively cuticle lining of the rectum is thin and continuous with sunken, and, in the cysts, the anus is encircled by fine layer A of the BW cuticle. ridges (Fig. 2 B). The body Wall (BW) cuticle of the cone of females of C. cacti generally resembles that of the midbody with Discussion typical A, B, C and D layers and zones (Figs 2 C, D) sensu Cliff and Baldwin (1985). However, in the area Monoxenic cultureof Cactoderahas not beenpreviously corresponding to the circumfenestra, the BW cuticle is reported, and the method discussed herein is simpler modifiedwith zone A3 and layer D predominating; than publishedapproaches for other Heteroderinae layer C is progressively reduced in thickness posteriorly (Widdowson et al., 1958; Brown, 1974; Reversat, 1975; and is absent at the terminus (Figs2 E, 3 A). Zone A 3 Lauritis,Rebois & Graney, 1982, 1983;Cordero & includes large irregular electron dense inclusions and Baldwin, 1990,1991). Monoxenicculture, combined layer D is crossed by laterally oriented bridgesof layer B. with video recording, permitsidentifying particular However, in the cyst, the BW cuticle of the terminus is points in development of the cone of C. cacti for more further reduced. Layer D diminishes in theregion from detailed SEM and TEM comparison with other Hetero- the outer boundaryof the crescents to the cone terminus derinae, and specifically with H. schachtii (Cordero & so that eventually an electron dense zone, A3, predomi- Baldwin, 1990, 1991). nates (Figs 2 Dy4 C, D). Video recording revealed a gelatinousmatrix The vagina is about 20 pm long. Near the vulva, the previously reported to be absent in C. cacti by Southey vagina is dorsoventrally flattened, and connected to the (1954, 1957), and present but rudimentary by Cooper BW cuticle at its lateral ends (Figs 1 C, 2 G) forming a (19554. Contrary to the gelatinous matrix present in vulval bridge which persists until circumfenestration. At Heterodera spp., that of Cactodera is clear, less copious, the cone terminus, the lumen of the vagina is narrow lacks eggs, and thus, is easily overlooked in field speci- (Fig. 2 G), but closer to the uterus it doubles in width mens. As in H. schachtii, cells of the uterus are dense forming a broad chamber (Figs 1 B, 3 A, 4 A). The with rough endoplasmic reticulum, and these cells are vagina is lined by cuticle layers A and D (Fig. 3 A), but likely to be the source of the gelatinous matrix (Mack- zone A3 is not resolved and layer D is modified near the intosh, 1960). uterus with disoriented fibers and irregular boundaries. Males have been reported as rare in C. cacti (Adam, The vaginal lining is thinnest in the middle and thicker 1932; Southey, 1954; Shmal’ko, 1959; Golden & Raski, at both ends, becoming particularly broad and diffuse 1977), and some other Cactodera spp. (Golden & Raski, near the uterus (Figs 3 A, 4 A). Near the uterus, the 1977). The present study demonstrated a high rate of vagina branches dorsolaterally (Figs3 A, 4 A). In young reproduction in the absenceof males, further suggesting females, the uterus epitheliumlacks muscular fibers and that C. cacti is parthenogenetic. It is not known if males is composed of large cells with electron dense cytoplasm develop in C. cacti under specific conditions. Forexam- and large nuclei (Figs 1 B, 4 B). ple, temperature and/or CO, concentrations influence Although somatic musculature is lacking in thecone, development of males in monoxenic cultures of Aphe- a diminutive musculature is associated with the vagina Zenchus avenue Bastian, 1865 (Hansen,Buecher & of C. cacti (Figs 1 B, D; 3; 4 D-F). A prominent set of Yanvood, 1972, 1973). Nevertheless, variation in mode dilatores vaginae (DV) muscles radiate obliquely, just or reproduction among Heteroderinae, andeven within

Revue Nématol. 14 (3) :455-465 (1991) 46 1 D. A. Cordero, J. G. Baldwin & M. Mundo-Ocanlpo

Cactodera, suggeststhat the presence or absence of & Baldwin, 1990, 1991). The BW. cuticle of the anal males may be a useful character for phylogeneticanaly- region in C. cacti, in contrast to H. schachtii, is very thin sis (Triantaphyllou, 1970, 1983; Triantaphyllou & with only a narrow portion of layer A persisting in the Hirschmann, 1980). cyst. We suggest that a transformation series exists in Interspecific differences and taxonomic valueof sur- which the thin perineal region of Cactodera is inter- face patterns at theposterior terminus of Heteroderinae mediate between the thick perineal cuticleof Heterodera have beendocumented (Mulvey, 1972,1973, 1974; and the perineal fenestrae of Punctodera. Green, 1975; Othman, 1985). However,Othman (1985), In both the vulval and anal region of C. cacti, layers andOthman, Baldwin andMundo-Ocampo (1988) of the BWcuticle are lostas the female ages and thecyst suggested that some intraspecific morphological vari- develops. This loss contrasts sharply to H. schachtii in ationmay be attributed to ontogeny, and that these which additional layers arelaid down inthe aging changes mustbe identifiedfor reliable phylogenetic female. A D layer occurs in the BW cuticle of both interpretation.Surface ornamentations of the vulval C. cacti and H. schachtii (Cordero & Baldwin,1990). cone of C. cacti Vary with age. In young females, vulval However, in the former, the D layer occurs at both the lipsprotrude and the surface of the lipsand vulval midbody and cone in young females (Cliff & Baldwin, crescents is smooth. However, as the femaleages and the 1985), whereas in thelatter, the D layer is relatively thin. cyst forms, the vulva becomes sunken, the vulval lips It occurs only in aging females and it does not occur in becomeindistinct, and the vulval crescentsbecome the cone. In H. schachtii there is a distinct E layer and patternedwith short ridges. Green (1975) reported associated bullae in aging female and cysts, but in C. domed vulval crescents which were finely or coarsely cacti there is no E layer or bullae. It is important to tuberculated in C. cacti, but he did not report differ- investigate additionalHeteroderinae to determine to ences between young females and cysts. whatextent the persistenceor loss of BW layers is Homology of crescents in semifenestrate and circum- associated with circumfenestrae or semifenestrae. Such fenestrateHeteroderinae is not clearly established. information could helpto determine homologiesof cone Green (1975) considered the two semifenestral regions characters throughout Heteroderinae for more reliable of Heterodera as " crescents " whereas Mulvey (1973) phylogenetic analysis. and Green (1975) used " crescents '' to describe the In females, zone A3 is widest at the cone terminus in tuberculate areas on eitherside of the circumfenestra of the region which corresponds to the vulval crescents.In Globodera. However, the region ruptured with fenestrae addition, laterally oriented bands of layer B bridge the in C. cacti and Globodera does not include the crescents circumfenestra parallel with the vulval slit, alternating sensu Green (1975). Conversely, in H. schachtii, nearly with bands of layer D. This pattern corresponds to the the entire region may be ruptured. In other Heterodera opticalpatterns on the cone top of Globodera cysts species, the outer portion of the " crescent " persists designated by Hesling and Ellis (1974) as " fenestral afterfenestration. Further comparisonis needed to shelves ". Similar bridges of layer B were observed establish homology of thesestructures, to identifya alternating with layer Cin the cone of H. schachtii possible transformation series in patterns of fenestra- (Cordero & Baldwin, 1990). Future fine structural in- tion, and to verify that terminology is applied consist- vestigations of the terminal region of Globodera may ently. substantiate homology of these structures. The sunken vulva with inconspicuous vulval lips in In the cyst of C. cacti, the circumfenestralarea is cysts of Cactodera is also typical of Globodera and Punc- electron-lucent and composed only of the outer portion todera, in which similar vulval circumfenestrae develop. of the A layer. Contrary to the semifenestral region of Although the three genera shareseveral unique charac- H. schachtii there is no mesh-likenetwork of fibers ters, Cactodera shares other characters,such as the cone, (Cordero & Baldwin, 1990, 1991). Furthermore, the with Heterodera. Cactodera is interpreted as phylogene- vaginal lining and vulval bridge are thin and do not tically intermediatebetween the round and lemon- persist incysts of C. cacti(Kiryan0va & Terenteva, 196 1; shaped cyst nematodes (Baldwin & Schouest, 1990). Green, 1975; Krall & Krall, 1978; Cordero & Baldwin, The anal region in C. cacti is similar to the vulval 1990,1991). Cornparisons of the fenestral region of region in that it also varies with age. In young females, additional Heteroderinaewill provide insight into factors the anus opens into a shallow cavity, but in cysts the contributing toloss of the vulval bridgeand thusforma- cavity becomes a deep depression. Green (1975) noted tion of circumfenestrae. From comparative development that the size and shape of the anal area differed within and from arguments of parsimony, we can test hypo- and between species of Heterodera, but species of Cacto- theses of homology of types of fenestrae for greater dera were not considered. However, Golden and Raski insight into phylogeny of Heteroderinae. Specifically, it (1977) reported that the anal region of C. cacti lacks a is important to determine if circumfenestrae are homo- specific pattern. logous in Cactodera,Globodera, Punctodera and Doli- Differences betweenthe anal region of Cactodera and chodera Mulvey & Ebsary, 1980. Heterodera are suggestedby Our investigations (Cordero The cuticle of the fenestral regionin youngfemales of

462 Revue Néwatol. 14 (3) : 455-465 (1991) Posterior cone of Cactodera cacti

C. cacti is continuous with the lining of the apparently of variation of cone characters.In addition, examination nonfunctional vagina. LayerD is prominent proximalto of noncyst-formingHeteroderinae such as Atalodera the cone terminus, however, a modificationof this layer and Sarisoderu, which have cones but lack fenestrae,will extends from approximately the midregionof the vagina allow one to recognizehomologies and the fullrange of to the uterus.A similar modificationin thelining of the cone character States throughout the subfamily. vagina, involving the Clayer, was reported forH. schach- tii (Cordero & Baldwin, 1991). Contrary to H. schachtii, the cuticle lining of the vagina of C. cacti is not greatly ACKNOWLEDGEMENTS expanded near the uterus,however, the surface adjacent A portion ofthe senior author’s Ph.D. thesis. This studywas to the hypodermis is highly irregular with numerous supported in part by LASPAU (Latin American Scholarship projections. These projectionsmight correspond to a Program of American Universities). vestigial underbridge (Green, 1975). Since the under- bridge varies greatly in length among species (Mulvey, 1957, 1972, 1974), a vestigial underbridge could define the limits of a phylogenetically important transform- ation series. REFERENCES Diminutive DV musculature attaches to thevagina in ADAM,W. (1932).Note surHeterodera schachtiischrn. parasite the cone of C. cacti. In contrast to the extensive DV des cactus.Bulletin du MuséumRoyal d’Histoire naturelle de musculature with fourmuscle sets in H. schachtii, there Belgique, 7 : 1-10. is only one prominent DV muscle set in C. cacti. These BALDWIN,J. G. (1983). Fine structure of body Wall cuticle of DV muscles attach to theposterior region of the vagina females of Meloidodera charis, Atalodera lonicerae, and Sari- and extendparallel to theinner surface of the BW sodera hydrophila (). Jounral of Nenratology, cuticle. The SV musculature is also reduced in C. cacti 15 : 370-381. relative to H. schachtii. Although vaginal musculature BALDWIN,J. G. & SCHOUEST,L. P. JR. (1990). Comparative presumably is associated with egglaying, it is interesting detailedmorphology Heteroderinaeof Filip’ev and that muscles persist in a species of Cactodera in which SchuurmansStekhoven, 1941, sensu Luc et al., 1988 : eggs are not laid. Timelapse video recording of C. cacti Phylogeneticsystematics and revised classification. Sys- suggest that muscles rhythmically contract the vagina tenzatic Parasitology, 51 : 81-106. during extrusion of the gelatinous matrix, but we have BROWN,J. A. M. (1974). Test tube reproductionof Heterodera noted thatthe function of thismatrix is unknown. avenue on resistant and susceptible wheats. Nematologica, Differencesbetween H. schachtii and C. cacti with 20 : 192-203. respect to size, number, and points of attachment of vaginal musculature suggest that investigationof these BYARD,E. H., SIGURDSON, J.W. & WOODS,R. A. (1986). A hot aldehyde-peroxide fixation method for electron microscopy characters throughout Heteroderinae will provide im- of the free-living nematode. Caenorhabditis elegans. Stain portant new characters for phylogenetic analysis. Technology, 61 : 33-38. Light microscopy and TEM of the cyst cone in C. cacti revealed remnants of muscles including bundlesof CLIFF,G. M. & BALDWIN,J. G. (1985). Fine structure of body filaments near the inner surfaceof the BW cuticle about Wall cuticle of females of eight genera of Heteroderidae. 20 pm below the circumfenestra. The muscle remnants Journal of Nenzatology, 17 : 286-296. occur singly and in clusters and apparently correspond COOPER, B. A.(1955~). A preliminary key to British species of to previouslydescribed vulval denticles(Golden & Heterodera for use in soi1 examination. In : Kevan, D. K. Raski, 1977). Our investigations support the hypothesis McE. (Ed.) Soil Zoology. London, Buttenvorths Scientific that denticlesare not homologous with bullaesince Publications : 269-280. bullaeare of cuticularorigin (Cordero & Baldwin, COOPER, B. A. (1955b). Mounting technique for identification 1990, 1991). Muscle remnants are also retained in the of Heterodera eelworm cysts. In : Kevan, D. K. McE. (Ed.) cyst of H. schachtii. However, due to the position of Soil Zoology,London; Buttenvorths Scientific Publications: vaginal muscles and persistence of the vagina in this 419-420. genus, they are more closely associated with the vagina CORDERO,D. A. 81 BALDWIN,J. G. (1990). Effect of age on than the BW cuticle. Furthemore, bullae, the vagina, body Wall cuticle of Heterodera schachtii Schmidt females. andthe underbridge visually obscure the vestigial Journal of Nenzatology, 22 : 356-361. musculaturein whole conemounts of H. schachtii, CORDERO,D. À: & BALDWIN,J. G. (1991). Fine structure of whereas muscle remnants persist alone in C. cacti. the posterior cone Heteroderaof schaclztiiSchmidt (Hetero- Comparison of the structure anddevelopment of the derinae) with emphasis on musculature and fenestration. cone of H. schachtii and C. cacti with other Hetero- Jounzal of Nenzatology, 23 : 110-121. derinae will provide new insight into cone charactersand EBSARY,B. A. (1986). Species and distribution ofHetero- their phylogenetic significance. Specifically, comparison deridae and Meloidogynidae (Nematode : ) in with Globodera and Punctodera will elucidate the range Canada. CanadianJounzal of Plant Pathology, 8 : 170-184.

Revue Nénzatol. 14 (3) :455-465 (1991) 463 D. A. Cordero, J. G. Baldwin & M. Mundo-Ocampo

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Accepté pour publication le 30 juillet 1990.

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