Aleutian Heteroderoidea: Bernard 513 cultures were increased in a growth cham- 5. Seinhorst, J. W. 1959. A rapid method for ber at 15 C. No galling was observed on the transfer of nematodes from fixative to anhy- drous glycerin. Nematologica 4:67-69. oat, rescue, or tomato, but these cultures 6. Taylor, A. L., and J. N. Sasser. 1978. Biology, were kept in the greenhouse and thus may identification and control of root-knot nematodes have been grown at unsuitable tempera- (Meloidogyne species). North Carolina State Uni- tures. versity, U.S.A,I.D. 7. Triantaphyllou, A. C. 1979. Cytogenetics of root-knot nematodes. Pp. 85-109 in F. Lamberti LITERATURE CITED and C. E. Taylor, eds. Root-knot nematodes (Mcloidogyne species). Systematics, biology and I. Eroshenko, A. S. 1978. (A new species of control. New York: Academic Press, Meloidodera [Nematoda, Heteroderidae] from the 8. Whitehead, A. G. 1968. Taxonomy of Primorsk territory.) Parazitologiya 12:456-459 (In Meloidogyne (Nematodea: Heteroderidae) with de- Russian). scriptions of four new species. Trans. Zool. Soc. 2. Jenkins, W, R. 1964. A rapid centrifugal- Loud. 31:263-401. flotation technique for separating nematodes from 9. Wouts, W. M. 1973. A revision of the family soil. Plant Dis. Rep. 48:692. Heteroderidae (Nematoda: Tylenchoidea). II. The 3. Mulvey, R. H., and R. V. Anderson. 1980. subfamily Meloidoderinae. Nematologica 19:218- Description and relationships of a new root-knot 235. nematode, Meloidogyne sewelli n. sp. (Nematoda: 10. Young, L. D. 1975. A Meloidogyne sp. on Meloidogynidae) from Canada and a new host American beachgrass in North Carolina. Unpub- record for the genus. Can, J. Zool. 58-1551-1556. lished M.S. thesis. North Carolina State University. 4. Robbins, R. T. 1978. A new Ataloderinae 11. Young, L. D., and L. T. Lucas. 1977. Hosts (Nematoda: Heteroderidae), Thecavermiculatus of Meloidogyne sp. on American beachgrass. Plant gracililancea n. gen., n. sp. J. Nematol. 10:250-254. Dis. Rep. 61:776-777. Identification of Meloidogyne Species on the Basis of Head Shape and Stylet Morphology of the Male 1 J. D. Eisenback and Hedwig Hirschmann x Abstract: Head shape and stylet morphology of males of 90 populations of M. arenaria, M. hapla, M. incognita, and M. javanica from geographic regions of the world were compared by light microscopy (LM). In addition, stylets of one population each of M. arenaria, M. incognita, and M. javanica and three different chromosomal forms of M. hapla race A and two of race B were excised and examitted with a scanning electron microscope (SEM). Differences among species occurred in both head and stylet morphology. Head morphology differed in size and shape of the head cap, annulation of the head region, and width of the head region relative to the first body annule. Differences in stylets occurred in size and shape of the cone, shaft, and knobs. All populations of M. hapla, except one, had similar head morphology, but stylet mor- phology was different between cytological races A and B. Populations of M. javanica varied with respect to the presence of head annulations. Head shape and stylet morphology of males are recommended as additional characters useful in the identification of root-knot nematodes. Key words: Meloidogyne arenaria, M. hapla, M. incognita, M. ]avanica, root-knot nematodes, cytological races, intersexes, scanning electron microscopy, taxonomy. Identification of the four common spe- arenaria (Neal) Chitwood, M. incognito cies of root-knot nematodes--Meloidogyne (Kofoid and White) Chitwood, M. ]avanica (Treub) Chitwood, and M. hapla Chitwood Received for publication l0 February 1981. 1Paper No. 6785 of the Journal Series of the North --is often difficult and time consuming. Carolina Agricultural Research Service, Raleigh. North Caro- Various approaches to Meloidogyne tax- lina. This study was supported, in part, by National Sci- ence Foundation Grant DEB-7917386 to A. C. Triantaphyllou onomy include host response (12,14,17), and U.S. Agency for International Development Contract cytology (17,18,19), biochemistry (3,4,11), to-C-t234 to J. N. Sasser. "-Research Associate and Professor, respectively, Depart- and morphology (1,2,5,6,7,8,9,12,13,15,20, ment of Plant Pathology, North Carolina State University, 21,22). Host-response tests require adequate Raleigh, NC 27650. We thank Dr. J. N. Sasser, Department of Plant Pathology, North Carolina State University. for amounts of nematode inoculum, and the conducting host differential tests and Dr. A. C. Tri- results may not be known for several weeks. antaphyllou, Department of Genetics, North Carolina State University, for providing cytological information. Cytological and biochemical studies require 514 Journal o] Nematology, Volume 13, No. 4, October 1981 specific techniques and sophisticated equip- the respective species (14,16), and the pop- ment. Identification based on morphology ulations o1: M. hapla represented the two can be rapid and practical if reliable dif- cytological races (18). ferentiating characters can be determined. Males were obtained by incubating Generally, Meloidogyne species are identi- washed infected root systems in a moist fied by the perineal pattern morphology of chamber at room temperature. The males the adult female (9,10,16). In populations were killed with hot TAF and mounted in of the four common species, however, the fixative for LM observations. Some perineal patterns are variable and definite specimens were killed, fixed, and mounted identification is not always possible (5,9,10, in glutaraldehyde (7) for additional mor- 12,13). Additional characters that are less phological studies, and others were ex- variable are needed to supplement perineal amined alive for comparison with fixed ma- pattern morphology. terial. For diagnostic studies the males must In a recent scanning electron microscope be viewed in exact lateral position. Camera (SEM) study, we reported that the head lucida drawings and LM photographs of morphology of males of M. arenaria, M. the anterior body region including the incognita, M. javanica, and M. hapla was stylet were made of representative popula- distinct for each species (7). Observed dif- tions. At least 30 specimens from each pop- ferences among species, and in M. hapIa ulation were examined. Eight of the popu- between races, were striking. Subsequent lations used in this study were the same studies revealed that some of these differ- populations used in previous LM and SEM ences, especially in head shape, could also observations of second-stage juveniles, be detected in the light microscope (LM). males, and females (6,7,8). When examining root-knot nematode fe- Stylets of males of one population each males (8), we shouted that differences occur of M. arenaria, M. incognita, and M. javan- in stylet morphology among the four com- ica and five different chromosomal popula- mon species. tions of M. hapla were observed by SEM. The present study compares by LM the The following populations were examined: head shape of males of several populations M. arenaria, 351-Fla; M. incognita, 68-NC; of each of the four common species of root- M. iavanica, 76-GA; M. hapla race A, 6-NC, knot nematodes. It also compares by LM 42-Can, and 86-NC; and M. hapla race B, and SEM the morphology of stylets of males 48-NC and 230-Chile. The technique orig- of one population each of M. arenaria, M. inally developed for the removal of stylets incognita, and M. javanica and five differ- of females (8) was adapted to males. Five ent chromosomal populations of M. hapla. males were transferred into a drop of 45% lactic acid on a glass coverslip. The speci- mens were cut behind the median bulb with MATERIALS AND METHODS a dental root canal file and processed for Ninety populations of the four common SEM observations as previously described root-knot nematode species from geographic (8). At least g0 stylets from each population regions of the world were selected from the were observed. The LM observations of the Meloidogyne collection at North Carolina stylet morphology of all populations were State University. Thirteen populations compared with the SEM observations of the were M. arenaria; 12, M. hapla; 44, M. in- eight populations previously listed. cognita; and 31, M. favanica. All popula- tions were propagated on tomato (Ly- copersicon esculentum Mill. 'Rutgers') in a OBSERVATIONS greenhouse maintained at 22-28 C. Each Head morphology: The basic head mor- population had been characterized by its phology of Meloidogyne males is derived ability to reproduce on five host differ- from SEM and LM observations (Figs. 1, entials (16), and many populations had 2). Labial disc and medial lips as observed been identified on a cytological basis. The in the SEM (7) appear in the LM as a populations of M. arenaria and M. in- single structure, the "head cap." The an- cognita belonged to several host races of nulations in the head region, so distinct in Meloidogyne Male Head Shapes: Eisenback, Hirschmann 515 and the lateral field begins near the level of ba.....__L1 the stylet knobs. The four common species differ in their head morphology with respect to shape and s[[ff( size of the head cap, presence or absence of annttlation in the head region, and the way the head region fuses with the body region. In M. arenaria (Figs. 3, 9) the head cap is low, slopes posteriorly, and is nearly as wide as the head region. Usually the head region 2 is marked by 1-2 incomplete annulations and is not set off from the body region. M. incognita males (Figs. 4, 10) possess a high head cap formed by a large round labial V disc that is raised above the medial lips b! and centrally concave. The head cap is as c wide as tile head region. The head region generally bears 2-4 incomplete annulations, s~ although some populations may have com- sl plete annulations.