Life History of Microtetrameres centuri Barus, 1966 (Nematoda: Tetrameridae) I. Juveniles CHARLES J. ELLIS t Abstract: Anatomical details of experimentally reared juveniles of Microtetrameres centuri are reported. These details are compared to results of similar studies of other Microtetrameres species. Little information concerning anatomical living juveniles were examined under phase- characteristics of juvenile Microtetrameres contrast and dark-field microscopy, fixed by spp. exists. Seurat (12, 13, 14), one of few various methods and mounted in either glyc- investigators to describe Microtetrameres erine jelly or CMC-10 (a General Biological juveniles, included the following features in Supply House mountant). Living and fixed his descriptions of third-stage and what he juveniles were measured using a calibrated termed fourth-stage juveniles of Tropido- ocular micrometer or microprojection tech- cerca (= Microtetrameres ) spiralis recovered niques. All drawings, except those noted, from naturally infected hosts: length of tail, were made with the aid of a microprojector. distance from head to nerve ring and to excretory pore, body length and width, RESULTS length of buccal capsule, lengths of the two EGG: Embryonated eggs were oval, pharyngeal portions as well as the ratio of slightly flattened on one surface (Fig. 1) the entire pharynx to the body length. Schell with a boss on at least one end. Unem- (11) and Cram (5) have also reported on bryonated eggs were collapsed and crescent- juveniles stages of Microtetrameres spp. shaped in cross-section. The egg shell This study was initiated after finding that averaged 2 to 3 ~ in thickness. Filaments 53.7% of the 173 meadowlarks (Sturnella were lacking. Cephalic projections of the negIecta and S. magna) (7) collected in juvenile (Figs. 2, 4, 5) sometimes were seen Iowa were infected with Microtetrameres while the juvenile was still within the shell. spp. The high percentage of infection, the Measurements of 10 eggs from female M. wide (115 species) host range of Micro- centuri raised experimentally in a pigeon tetrameres, and the stomach-content study averaged 50.7 ~ long by 35.3 t~ wide. Similar indicated that meadowlarks were suitable measurements of 82 eggs of female M. hosts for this nematode. centuri recovered from naturally infected meadowlarks averaged 50.5 ~ long by 35.2 METHODS AND MATERIALS wide. Female Microtetrameres centuri gravid FIRST-STAGE JUVENILES : Juveniles with embryonated eggs were fed to grass- emerged within a few hours after eggs were hopper nymphs (Melanoplus spp.). The ingested by a grasshopper nymph. Some nymphs were examined for juvenile nema- juveniles recovered from the grasshopper's todes at various post-feeding times. The fore-gut were seen partially emerged (Figs. 2, 11). First-stage juveniles escaped from Received for publication 26 August 1968. the egg with either end emerging first. Both i Department of Zoology and Entomology, Iowa State Uni- versity, Ames, Iowa 50010. The author gratefully acknowl- methods have been reported for M. inermis edges the counsel and time given by Dr. M. J. Ulmer in the preparation of this paper and the thesis from which it (12). In 13 eggs nine juveniles had emerged is based. This research was supported partially by National Science Foundation Grants G-23597 and GB-5465X. anterior first (Figs. 2, 11 ) and four juveniles 84 LIFE HISTORY OF Microtetrameres centuri BARUS • Ellis 85 tail first. In one instance, a juvenile had four refractile bodies which moved slightly broken the shell but both of its ends were as the nematode moved. Nuclei were seen still within the egg shell. Thirty minutes throughout the living body. later this juvenile had escaped tail first. At the anterior end of some living speci- First-stage juveniles possessed blunt ante- mens the area destined to become the riors (Figs. 3, 11) and pointed tails (Fig. pharynx in older juveniles extended pos- 3). The anterior end of a living first-stage teriorly about 20 ~. Granules were visible juvenile changed from convex to concave within this region in some living individuals. as the animal moved (Figs. 4, 5). In this As the nematode moved, the granules moved aspect, M. centuri first-stage juveniles re- rhythmically, as did the three or four re- sembled juveniles of what appeared to be fractile bodies near the anal pore. No con- Tetrameres, as illustrated by Lieberkuhn nection between the anal pore and the (9). A group of three minute refractile anterior region was seen under bright-field cephalic projections (Figs. 4, 5) appeared or phase-contrast microscopy. Because of anteriorly. These projections do not re- their synchronized movements, however, the semble cephalic hooks (10). The position anterior and posterior groups of refractile of this group of projections was modified bodies appeared to be related. somewhat in accordance with the movement Early first-stage juveniles were distin- of the anterior region of its body. The rapid guished from late first-stage juveniles by movement of living juveniles and the minute their size and cuticular striations. Two types size of these projections made observations of striae were noted on first-stage juveniles. difficult. Movement, together with the Discontinuous striations appearing as rings action of the cephalic projections, may have of tiny dots instead of continuous lines (Figs. added measurably in the migration of 2, 3, 4, 5) occurred only along the anterior juveniles within the tissue of their inter- 16 to 20 ~ of the body of the early first- mediate host. These projections were similar stage juveniles. A second type of striation, to the "aiguillon cephalique" mentioned by characteristic of older first-stage juveniles, Chabaud (2) who considered them aids in was continuous. Such striae were located permitting first-stage juveniles to escape posterior to the discontinuous ones. from their eggs and to penetrate the inter- From a point slightly anterior to the pos- mediate host's gut wall. terior discontinuous striation, two slender The resemblance between a first-stage lines, one on either side of the body, ex- juvenile of M. centuri and a microfilaria is tended nearly to the posterior end of the remarkable. About 50 ~ from the anterior juvenile (Fig. 2). These were seen in nu- end of a living first-stage juvenile a clear merous living and fixed specimens and area similar to the excretory pore of a micro- probably were lateral chords. filaria was seen. A spot resembling the anal Living juveniles displaying characteristics pore of a microfilaria was seen approxi- of both first-stage and second-stage were mately 50 ~ from the tail. Between these recovered 16 days after feeding eggs of M. two areas but closer to the posterior one centuri to a grasshopper nymph. These was a structure resembling a short length of juveniles (Figs. 6, 7, 8, 9) were similar to intestine. Presumably the intestine was con- second-stage juveniles in size and in charac- nected to the anal pore, but it did not appear teristic cephalic projections. They were to extend the length of the juvenile. Within classified, consequently, as first-stage juve- the "lumen" of this structure were three or niles under-going ecdysis to the second stage. 86 Journal o/ Nematology, Vol. 1, No. 1, January 1969 Transverse cuticular striations were pre- approximately 8 t~ in diameter. It opened sent along the length of most specimens. In internally into a nearly spherical chamber others such transverse striations were limited (Fig. 7). Unidentified material was seen to the anterior region. A well-developed protruding from the anus of some juveniles. sheath, slightly raised from the cuticle, and No genital primordium was visible under bearing the cephalic projections of first-stage dark-field or phase-contrast microscopy. juveniles covered the rounded anterior end Juveniles developed from eggs fed to of each specimen. A clear area, consisting grasshoppers in 40 days. The following of two separate translucent spots with dimensions are of living and fixed first-stage definite boundaries and approximately 3-5 t~ juveniles: in diameter, was visible within 13 ~ of the anterior end. Perhaps these spots represent 2 days old (fixed) (10 specimens); length glands associated with histolytic activity. 190 ~ (148-216), width 12.1 tx (10-16) The excretory pore on many of these juve- 4 days old (living) (10 specimens); length niles was easily located slightly more than 264 ~ (235-289), width 20.8 ~ (19.3- 80 ~ from the anterior end. No evidence 21.4) of a mouth was seen. The first 40 ~ of the 5 days old (living) (10 specimens); length digestive tract appeared as a fine line and 318 ~ (300-326), width 15 t~ (11.0- no lumen was visible. Posteriorly, the gut 21.4) expanded, reaching its maximum width at 6 days old (living) (9 specimens); length the pharyngointestinal junction. This junc- 265 ~ (245-286), width 18 t~ (18) tion was plainly visible 130-145 ~ posterior 8 days old (living) (8 specimens); length to the anterior end. The lining of the 336 t~ (321-353), width 21.4 t* (21.4) digestive tract could be traced easily under 8 days old (fixed) (4 specimens); length dark-field microscopy. Body musculature 270 t~ (257-300), width 21 t~ (21) surrounding the tract was seen under phase- SECOND-STAGE JUVENILES : Juveniles contrast microscopy. In some juveniles, molted from first to second-stage between foreign material was observed within the 8 and 16 days after challenge (Table 1). gut. The anus was apparent and measured The length of development time appeared to ...) Fro. 1. Egg of M. centuri containing first-stage juvenile (in mm). FIG. 2. First-stage juvenile emerging from egg (in mm). FIG. 3. Living four day old first-stage juvenile from experimentally infected grasshopper nymph (in mm). FIG. 4. First-stage juvenile, anterior end extended showing cephalic projections. Free hand drawing (in mm).