A laboratory technique is described in which "soft” radiographs are used to study wireworm, Melanotus communis Gyllenhal, movement patterns within a soil matrix. Advantages of this technique as compared with destructive sampling are discussed and examples of several possible applications are give

Gould, F.

Villani, M. G.

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Use of Radiographs for Movement Analysis of theCorn Wireworm, Melanotus communis(Coleoptera: Elateridae)M. G. VILLANI' ANDF. GOULDDepartment of Entomology, North Carolina State University,Raleigh, North Carolina 27695Environ.Entomol.15: 462-464 (1986)ABSTRACT A laboratory technique is described in which "soft" radiographs are used tostudy wireworm, Melanotu8 communis Gyllenhal, movement patterns within a soil matrix.Advantages of this technique as compared with destructive sampling are discussed andexamples of several possible applications are given.DETAILED STUDIES on the movement of soil insectshave been largely ignored by population biologists.There is a general perception that such research,if not impossible, is difficult, tedious, and cost-in-effective when compared with similar studies ofaboveground insects. Historically, agriculturalentomologists have also shied away from this areaof study although a number of studies on themovement of wireworms have been conducted inan effort to control members of this economicallyimportant soil insect family.R. L. Rabb (personal communication) noted that:the greatest problem with studies of soil insects isnot logistics, but rather with the alteration of thesystem through its study. Laboratory studies ofmovement and feeding were often done outsidethe soil (Fulton 1928), or between thin sandwichesof glass plates and sterile, homogeneous sand(Doane et al. 1975), which makes 3-D movementby wireworms impossible. Radioactive tagging ofwireworms with strong 'Y emitters such as 60CO(Amason et al. 1950, Fuller et al. 1950, Frederick-sen and Lilly 1955) proved hazardous to the insectsand too imprecise for monitoring burrow forma-tion accurately. In some cases, feeding behaviorresearch involved destructive sampling of the soilmatrix (Kring 1955, Keaster et al. 1975), whichmay have resulted in alteration of the wireworm'stypical behavior. Other studies of feeding focusedon the results, rather than the process of feeding(i.e., feeding damage) (Stone 1935).In an effort to gain a more realistic picture ofthe events that occur within the soil matrix, wehave adapted an x-ray technique that had beenused to study seed insects, parasitized cocoons, andwood-boring insects in their gallleries (see refer-ences in Yates and Belcher 1968) for the study ofcorn wireworm, Melanotus communis Gyllenhal,movement and feeding behavior. (For a detailedexample in which this technique has been used toICurrentaddress:Dep.ofEntomology,NYSAES,CornellUniv.,Geneva,NY14456.monitor behavior and response of wireworms tonatural antifeedants, see Villani and Gould 1985.)Materials and MethodsThe majority of the experiments were run insoil-tilled (soil moisture 6 + 2%) plexiglass cham-bers (30 by 15 by 4 cm). These dimensions allowedthe wireworms to move freely in all directions un-til the constraints of the chamber walls were met.Readable radiographs in chambers up to 43 by 36by 10 cm were obtained, but as the thickness of asample to be radiographed was increased, therewas a corresponding increase in radiographic dis-tortion. Some studies were done in smaller cham-bers such as bacterial culture tubes.Wireworms used in this study ranged in lengthfrom 2 to 3 em. Tiny lead foil chips (ca. 1 mmdiam) were fixed to the ninth abdominal segmentby coating the dorsal plate of the segment with athin layer of clear nail polish, placing the chip onthe tacky plate, and then coating the chip andplate with two additional layers of nail polish. Carewas taken not to cement the posterior abdominalsegments together through the use of nail polish.(The individual wireworms were cooled to 6°C forseveral minutes to facilitate ease of handling forthis procedure.) The chip, being the densest objectin the soil matrix, stood out as a white dot in theradiograph. The soil appeared medium gray andthe least dense, air-filled burrows appeared darkgray to black in the radiographs.Radiographs were taken with an x-ray system(Faxitron 43805N) using a milliamperage (mA) ofca. 2.8, and a kilovoltage (kVp) of ca. 80. The lengthof exposure was automatically controlled (expo-sure time depended on the thickness of material,material density, moisture content, desired detail,film-to-focal point distance, and kVp). The exposedx-ray film was processed in a 190-liter developingtank (Bar-Ray Products, Brooklyn, N.Y.) with Ko-dak-X-Ray Fixer and Developer and hung to air-dry. Finished radiographs were studied by view-462June 1986 VILLANI & GOULD: RADIOGRAPH OF MOVEMENT 463Fig. 1. Radiograph taken 2 h after the wirewormwas placed in the middle of the chamber. The corn seed(dark area on right) was 4 days after germination. Theapparent directed movement of the wireworm to thecorn seed (dark line) is typical of active wireworms.ing with backlighting or by photographing thebacklighted radiograph and studying the enlarged,projected image.Radiographs are shown in Fig. 1-5. In Fig. 1-4, wireworms were released, head down, in thetop center area of the arena (30 by 15 by 4 cm).Each radiograph shown is one in a time series ofradiographs taken of that arena over a 2-monthperiod. Fig. 1 is representative of radiographs thatindicate that wireworms have directed movementtoward food. Fig. 2 demonstrates that wirewormscan move quickly and freely through a noncom-pacted soil matrix. In Fig. 3, a lead chip placedon the wireworm is visible within the pupal cell.Although this radiograph was taken 45 days afterintroducing the wireworm to the arena, the wire-worm never entered a feeding phase during thattime and pupated early in that period. Fig. 4 rep-resents our attempt to simulate wireworm re-sponse to a heterogeneously compacted soil or onewith a plow layer. Wireworms tended to movealong the compaction interface. Fig. 5 representsresults of a test to determine wireworm responseto a moisture gradient.DiscussionThis technique makes it possible to trace theburrow of wireworms within the soil matrix. It alsoshows physical properties of the soil matrix: par-ticle size, extent of compaction, soil moisture, andheterogeneity (cracks, gaps, gravel, etc.). The 10-Fig. 2. Radiograph of chamber with wirewormwithout food, taken 12 h after placement. Note the com-plex matrix of tunnels that were apparently made dur-ing a search for food.Fig. 3. Radiograph taken 45 days after wirewormplacement. The burrows that are not oriented to thefood bait are typical of prepupal movement patterns.The pupal cell, constructed over several days, is clearlyvisible in this radiograph. The triangular lead chip isvisible within the cell.. cation and identity of target plants and food baitswithin the soil are easily noted with radiographs,and their condition and growth are often appar-ent. By attaching a tiny lead chip to the ninthabdominal segment of the wireworm, one can studyboth the patterns of the wireworm movement(burrows and pupal cells) and the location of thewireworm within its burrow network. Through theuse of an irregularly shaped chip, the orientationof the wireworm in its burrow can be determined(unique shapes at the four compass points allowone to determine the wireworm's anterior-poste-rior and dorsal-ventral position in the test cham-ber). The movement of the wireworm after a molt,indicated by additional burrows and a stationarylead chip, can be used to assay pre- versus post-molting feeding behavior and movement patterns.There was no indication in our research that theuse of "soft" radiographs adversely affected wire-worm feeding or longevity when compared withthose of insects not studied. Soft x-rays are ca. 10A in wavelength, whereas in medicine, the wave-lengths used range from 0.5 to 0.001 A. These areknown as "hard" x-rays. Soft x-rays have less en-ergy and are more useful in investigating materialsof a smaller size than is common in medicine. Spe-cific effects of soft radiographs on the test subjectsmust be evaluated on a case-by-case basis. Studiesover a number of generations in which geneticFig. 4. Burrow pattern of a wireworm presentedwith a stratified soil is shown in this radiograph. Themore compacted lower strata are more dense than theupper plow layer and show as a lighter shade of gray inthe radiograph. Wireworm burrows at the soil interfaceare clearly visible.464 ENVIRONMENT AL ENTOMOLOGY Vol. 15, no. 3Fig. 5. Test tubes with moisture stratification. Onewireworm placed in each tube.changes would be critical must be carefully mon-itored when using any type of radiation source.In conclusion, radiography is a technique thatcan be used to enhance the understanding of insectmovement that occurs within the soil matrix. Usedin conjunction with destructive techniques in thelaboratory and in the field, both the process andresult of soil insect behavior can be studied andanalyzed.We have outlined only one possible use of thistechnique as it applied to our research on cornwireworm feeding behavior. Clearly, adaptationsof this procedure to facilitate its use with soil or-ganisms of a variety of sizes and for experimentswith diverse research objectives can be madethrough empirical modification of this technique.AcknowledgmentThis manuscript is Paper No. 9836 of the Journal Se-ries of the N.C. Agric. Res. Serv., Raleigh.References CitedArnason, A. P., R. A. Fuller, and J. W. T. Spinks.1950. An electronic method of tracing the move-ments of soil-inhabiting insects. Science 3: 5-6.Doane, J. F., Y. W. Lee, J. Klinger, and N. D. Westcolt.1975. The orientation response of Ctenicera de-structor and other wireworms (Coleoptera: Elateri-dae) to germinating grain and to carbon dioxide. Can.Entomol. 107: 1233-1252.Fredericksen, C. F., and J. H. Lilly. 1955. Measuringwireworm reactions to soil insecticides by taggingwith radioactive cobalt. J. Econ. Entomol. 48: 438-442.Fuller, R. A., J. W. T. Spinks, A. P. Arnason, and H.McDonald. 1950. Use of radio-active tracers ininvestigations on soil-inhabiting insects, p. 715. In81st Annual Report of the Entomological Society ofOntario.Fulton, B. B. 1928. Some temperature relations ofMelanotus (Coleoptera: Elateridae). J. Econ. Ento-mol. 21: 889-897.Keaster, A. J., G. M. Chippendale, and B. A. Pill.1975. Feeding behavior and growth of the wire-worms Melanotus depressus and Limonius dubi-tans: effect of host plants, temperature, photoperiod,and artificial diets. Environ. Entomol. 4: 591-595.Kring, J. B. 1955. A chamber for studies of site-selection by Elateridae. Entomol. News 61: 188-189.Stone, M. W. 1935. Technique for life-history studiesof wireworms. J. Econ. Entomol. 28: 817-824.Villani, M. G., and F. Gould. 1985. Screening of crudeplant extracts as feeding deterrents of the wireworm,Melanotus communis. Entomol. Exp. Appl. 37: 69-75.Yates, H., and E. Belcher. 1968. Tree Improvementand Nursery Management Branch, Southeastern Areaof State and Private Forestry. In Proceedings, South-east Forestry Radiography Workshop, January 9-11,1968, Atlanta.Received for publication 2 July 1984; accepted 30December 1985.

Use of Radiographs for Movement Analysis of the Corn Wireworm, Melanotus communis (Coleoptera: Elateridae)

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