——I —._——— _——‘- ’— _———— ’— —-‘—— _———— —— __———— —_——— _——_—— ’— ——— ——— ’- SOME EFFECTS OF RADIANT ENERGY ON THE BEETLES, TRIBOUUM CONFUSUM DUV. SITOPHILUS GRANARIUS (L) , AND I ACANTHOSCELIDES Joa'nscws (SAY) Thesis for the Degree of M. S. MICHIGAN STATE COLLEGE Oscar Taboada 1953 u i‘ i \ I 7' {s ' ) S “X g." “5‘30 gf's.‘ ' ‘ 4': SOME EFFECTS OF RADIANT ENERGY ON THE BmmTLas, TRIBGLIUM CONFUSUM DUV., SITOPEILUS GRANARIUS (L), AND ACANTHOSCELIDES OBTECTUS (SAY) By Oscar iapoada AN ABSTRACT Submitted to the School of Graduate Studies of Michigan State College of Agriculture and Applied Science in partial fulfillment of the requirements for the degree of MASTER OF SCIENCE Department of Entomology 1955 ,J c.,‘ ng Oscar Taboada THESIS ABSTRACT The purpose of this work was to investigate some effects of radiant energy on certain insects which infest stored pro- ducts. Infrared, ultraviolet and x-rays, as well as accelerated electrons were used on the confused flour beetle. The granary weevil was subjected to infrared rays, ultraviolet rays and accelerated electrons and the common bean weevil was limited to only one test of accelerated electrons. The beetles were ,1 reared under laboratory conditions of ad” 2°F. and a relative humidity of about 66 percent. The manner of rearing was so arranged that a constant supply of adult beetles of a known age were produced. The infrared energy tests indicated that both temperature and exposure time were important factors in obtaining effective results. A temperature of at least 138°F. and a maximum ex- posure time of 2.5 minutes was necessary to obtain lethal effects on the confused flour beetle, granary weevil and their eggs. The ultraviolet tests gave no clear indication of its effects on the beetles since the type of lamp used emitted four fifths of its energy as infrared energy. 'No exposure time in the x-ray tests showed any effects on the confused flour beetle. After the tests the adults were not sterile and were able to reproduce normal progeny. However, the results of the accelerated electron tests were more promis— ing. A dose of at least 500,000 rep would be necessary to kill completely a mixed pOpulation of adults and larvae of the con— Oscar Taboada fused flour beetle, the adults of the granary weevil and common bean weevil. A dose of 10,000 rep prevented the eggs of the confused flour beetle and the granary weevil from hatching. This same dose sterilized the adult granary weevil, the adult confused flour beetle and prevented the larvae of the flour beetle, after reaching the adult stage, from reproducing. SOME EFFECTS OF RADIANT ENERGY ON THE BEETLES, TRIBOLIUM CONFUSUM DUV., SITOPHILUS GRANARIUS (L), AND ACANTHOSCELIDES OBTECTUS (SAY) By Oscar Taboada A THESIS Submitted to the School of Graduate Studies of Michigan State College of Agriculture and Applied Science in partial fulfillment of the requirements for the degree of MASTER OF SCIENCE Department of Entomology 1953 ACANOWLLDGMLNTS Particular acknowledgment is made to Professor Ray Hutson, of Michigan State College for his help in selecting a thesis tOpic and for making the preliminary arrangements with the Agricultural Engineering Department and to Dr. Herman King under whose inspiration and guidance this thesis was compiled. Appreciation is expressed to Dr. Julius R. Hoffman of Michigan State College for his valuable criticism of this manuscript and for his help in taking some of the pictures. The author is also grateful to other members of the Department of Entomology of Michigan State College for their helpful suggestions and assistance. Grateful acknowledgment is due to his colleague Vernon H. Baker of the Agricultural Engineering Department, Michigan State College, for his superlative c00peration throughout this investigation. The author extends his sincere thanks to Mr. 0. R. Woods and Mr. N. A. Drake of the Physics Department of the Upjohn Company for their assistance and suggestions, and to the Upjohn Company for the use of their Van de Graaff generator. Acknowledgment is also due Dr. A. C. Wheeler of the School of Veterinary Medicine, Michigan State College, for his assistance with the X-ray tests. The author desires to express his sincere appreciation to his wife, Jeane, for her endless encouragement during the investigation and for the initial typing of some of the sections of this thesis. 3» 7513 I. II. III. IV. VI. . VII. VIII. TABLE OF CONTENTS INTRODUCTION.................................. REVIEW OF LITEHATUHE.......................... Rearing.................................... Infrared................................... Ultraviolet................................ X-Ray...................................... Electrons.................................. PROCEDURES IN BEARING AND HANDLING THE TEST INSECTS..................................... TESTING EQUIPMENT AND PROCEDURES.............. Infrared................................... Ultraviolet................................ X-Ray...................................... Accelerated Electrons...................... Mortality Counts........................... EXPERIMENTAL RESULTS.......................... DISCUSSION OF RESULTS......................... Infrared................................... Ultraviolet................................ X-Ray...................................... Accelerated Electrons...................... SUMMARY....................................... LITERATURE CITEDOOOOOOOOOOOOOOOOOOOOOOOOOOOOO. I INTRODUCTION Insect pests of stored grain and milled cereal products have been a problem to man ever since he learned to keep grain for food or seed. Many of the inSect pests of grains of ancient times are prevalent today and have been distributed throughout the world through commerce. Cotton (6) states, "Evidence indicates that many of the insects that trouble stores of grain today were prevalent in ancient times. Supplies of grain placed in the tombs of ancient Egyptians have been found destroyed by the same species with which we are familiar." Experts have estimated that the annual insect damage in the United States, is about four billion dollars and about one fourth of this sum is caused by insect pests of grain in stored grain and grain products. According to Cotton (6), methods used to combat the pests of stored products, and their feeding activities cause an annual cost of at least $500,000,000 in this country. many methods have been used and are being used to control insects. Some of these methods include the use of dusts, sprays, fumigants and heat. Some of the chemicals which are now widely used for the control of insects may have a residue so high as to be poisonous when used on grain products used as food. An ideal method to control insects without any residual effect would be the use of radiant energy. Before a method is used to any de- gree of success in the field, a series of extensive experiments must be performed in the laboratory. Some methods return to the laboratory for further experimentation and others remain in the laboratory for long periods of time before they can be put to practice. The use of radiant energy for the control of insects has been limited, and, unfortunately, much of this work has re- mained in the experimental stages. With this in mind, this paper deals with tests conducted with radiant energy on insects. An infrared lamp, an ultraviolet lamp, x-ray machines and a Van de Graaff accelerated electron generator were used as sources of energy for the tests. The insects used in the tests were the granary weevil, SitOphilus granarius L., which is a serious pest of stored grain, the confused flour beetle, Talbolium confusum Duval, which is a serious pest of flour mills or where milled grain is stored, and the common bean weevil, Acanthoscelides obtectus (Say), which infests various kinds of beans in the field and in storage. II REVIEW OF LITERATURE Rearing Pyenson and Menusan (l7) reared cultures of bean weevils of known ages by placing 500 - 1000 eggs, at weekly intervals, in pint jars containing red kidney beans. The eggs were taken from special oviposition cages made of pint cardboard boxes with the bottoms replaced by a 20 - mesh wire screening. This allowed the eggs from the adults to drOp through to a collect- ing tray. A few beans in the cage provided a stimulus for oviposition. The cages and the culture jars were kept in an incubator at 25°C and 75 — 80% R. H. The time taken for deveIOpment from egg to egg stage was about 40 days. Lindgren (12) used large-mouthed gallon capacity glass containers filled about one third full of wheat of a moisture content of 14%. He obtained grain weevils of a known age by placing adults in the wheat jars to oviposit for three or four days. Then the adults were sifted through a No. 10 wire sieve and placed in another jar of wheat. Adults from the eggs appeared in about five weeks when the jars were held at a temperature of 25°to 26°C. Robinson (19) did not consider age or temperature in rearing grain weevils for his experiments. Collins (5) maintained a large colony of Tribolium confusum of known ages in an air-circulated incubator at 78°- 83°F. and a relative humidity of 69%. He established the colony by placing adult beetles to ovipOSit in flour jars. After four days the beetles were sifted out with a No. 20 mesh sieve, and placed in another jar. Vorking with the same species, Park (15) used a mechanical device to separate the beetles. The automatic shaker consisted of a motor and a sieve connected to a belt wheel. The beetles were retained in the sieve and the flour fell through to a collecting tray. Infrared Headlee (lO) testing a 260W incandescent therapeutic lamp, a white light Mazda lamp and a 450W quartz - mercury lamp, all at the same distance, found that the therapeutic lamp killed American roaches almost instantly. The other two lamps did not have the same promptness. He concluded that no matter the source, the insects were not killed until a lethal heat was reached. Wigglesworth (25) gives the fatal temperature for small insects at about 110 F. Cotton (6) states that a temperature of 140°F. for 10 minutes is fatal to all grain insects. Blazer (3) using hot air blasts of 140°F. for 30 minutes did not get a satisfactory kill on several insects. The treatment raised the temperature of the rice to 122°F. only, which he stated was not high enough to kill the insects. Dean (7) obtained better results when he increased the temper- ature of a room in a flour mill to 153.5°F. All insects found in the Open, and down to three inches in depth of flour sacks, were killed. Ultraviolet Ray (18) conducted tests on the eggs of MelanOplus differentialis, with an air-cooled Quartz mercury - vapor lamp at 110-115 V.A.C. and 5.7 amps. at 15 cm. distance. The irradiations varied from 5 seconds to 4 hours. There was no noticeable effect at 5 seconds, but effectiveness was obtained at 1 minute exposure by having 18% hatch only, and no egg hatch at 15 minutes eXposure. MacLeod (15) reported that the adults of the bean weevil, Ascanthocelides obtectus, showed no effects from light of wavelengths less than 5126A. However, the eggs and first instars were killed. Ellis and Wells (9) eXposed the eggs of the Ascaris roundworms to ultraviolet rays from six to eight hours. The test did not produce immediate kill. X-Ray Experimenting on the fruit flies, Dacus cucurbitae Cog., and Q, dorsalis Hendel, Koidsumi (11) reported that these insects can be killed by certain amounts of x-ray radiations in all stages of their life cycle. Using a Coolidge tube without filtration, he showed, as Mavor (14) showed for Drosophila, that the resistance of the insects to x-rays becomes greater as deveIOpment proceeds from egg to full grown larva. Whiting (24) made studies on the effects of x-rays on. the parasitic wasp, Habrobracon juglans, Ashmead, Dunning (8) working with the same species, reported that the loss of fertil- ity and viability of individuals receiving the maximum dosage of 8000r. units was transmitted for several generations. How- ever, he was unable to determine whether this loss of fertility was due to less eggs laid, or failure of the eggs to hatch, since the number of eggs laid was not counted. Electrons Trump, Van de Graaff (25) and Burrill (4) were among the first to develop equipment for accelerated electrons. Proctor and Goldblith (16) were among the first investigators to treat various food products with accelerated electrons. Probably the first to report on the use of electrons on insects is Yeomens (26), who reports on the use of a capacitron treatment on various insects. The electron dosages ranged from 180,000 rep to 900,000 rep. He obtained 100% kill from a dose of 800,000 rep on mosquito eggs. The same results were ob- tained from a dose of 510,000 rep on adult confused flour beetles, 48 hours after treatment. A higher dose was required to get 100% kill of adult bean weevils. III PROCEDURES IN HEARING AND HANDLING THE TEST INSECTS Two incubators, as shown in Fig. 1 and 2, in which a relatively constant temperature could be maintained, were used to rear the confused flour beetle, Tgibolium confusum Duval., the granary weevil, Sitophilus granarius L., and the common bean weevil, Ascanthoscelides obtectus (Say). The same incubators were used as recovery cabinets. Various investi- gators have found that a temperature of about 80°F. and a relative humidity of around 75% shortens the life cycles of most of the pests of stored products. The temperature in the incubators was set at 80°F. A periodic check by a thermograph (Fig. 2) indicated fluctuations between 78°- 82°F. Wexler and Brombacker (80) suggest the use of sodium chloride solution made up as a slushy mixture, to produce a relative humidity of around 75%. A commercial salt composed of 99.5 percentsodium chloride and 0.5 percent tri-calcium phosphate was readily available. The salt was used to make up the salt solutions and was mixed in two large enamel pans. A pan was placed at the bottom of each incubator. Every two or three days water had to be added to each pan to replenish the loss of water and thereby keep the salt at a constant slushy mixture. At the onset of the project, the incubators were kept in a refrigerated room, but later were moved to a laboratory. It was found that the humidity was not being controlled in the incubators. An investigation revealed that the humidity in 7 the refrigerated room was about 97 percent, which more than likely, caused the abnormal humidity in the incubators. The humidity in the laboratory, during the winter months of 1952 - 55, was around 55 percent, and relatively higher during the following spring. The humidity readings made for the laboratory and the incubators were recorded from a wall-type hygrometer. However, towards the end of the rearing and testing project, a recording hygrothermograph was used to record the temperature and the relative humidity of the incubators. The recording was for a period of two weeks, and the readings indicated that the temperature fluctuated between 78°- 82°F. and the relative humidity was around 66 percent. According to Shepard (20) and others, food materials that have been in storage are too low in moisture content to rear insects successfully. Whole wheat flour of fine grind, Cor— nell 595 wheat and Michigan navy beans were used as culture media for the confused flour beetle, the granary weevil and the common bean weevil reSpectively. A successful attempt was made to increase the moisture content of the materials as follows. The wheat and beans were spread over an area 50 x 18 inches and one layer in depth on pieces of paper. The flour was spread over a smaller area in boxes and one eighth of an inch in depth. The samples were placed in the refrigerated room previously mentioned. The moisture content of the wheat before being placed in the room was about 10 percent. At the end of three weeks the moisture in the wheat had increased to Fig. 1. General view of incubators used for rearing the test insects. Fig. 2. Interior view of incubators used for rearing the test insects, and as recovery cabinets. 10 about 16 percent. Using the wheat as an index, it was assumed that the other materials had had an increase in moisture also. Then the practice was established whereby the food materials were placed in the refrigerated room three weeks before being used. Four hundred adult confused flour beetles were obtained as a breeding colony. Basically, the procedure used by Collins (5) was employed, butwith certain modifications. Flour was put in pint jars to a depth of about an inch. One hundred beetles were placed in each of four jars which were marked A1, A2, A 3, and A4. The date, which was September 29, 1952, for the first series of jars, was also recorded on each jar. A week after the adults had been placed in the jars to oviposit were sifted from the flour. Beetles from jar Al were placed in jar B1 and beetles from jar A2 were placed in jar B2 etc. The jars containing the flour and the eggs were returned to the cabinet along with the next jars that contained the adult beetles. The same procedure was used at weekly intervals and each time the date was recorded and a successive letter of the alphabet was used. By the mentioned method described, a rough estimate of the age of the adults was made and the use of old adults was avoided. A twenty—mesh sieve was used to separate the beetles from the flour (Figs. 5 & 5). The sieve was held in a vertical position over the jar and a camel's hair brush was used to push the beetles in the jar. Approxi- mately thirty-nine days after incubation, beetles from series A jars appeared. Some of the new adult beetles were used to 11 Fig. 5. Background; constant temperature box used to trans- port insects between buildings and to the Upjohn Co., Kalamazoo, Michigan. Foreground; at left is a ringstand with funnel used to place beetles in recovery vials. At the extreme right is an oviposition cage for the common bean weevils, and in the center are the sieves used to separate the test insects. Fig. 4. Shown are; Petri dish carriers, recovery vial, Petri dish sample, breeding jars and lids. 13 maintain the breeding stock while the old breeding adults were discarded. Essentially the same method was employed with the granary weevils, except that an eight—mesh screen (Figs. 5 & 6) was used to separate the weevils. The screen retained the wheat and the weevils would fall through to a glass vessel (Fig. 6). The vessel was held in a vertical position and the weevils were brushed off into the jar. Unlike the confused flour beetle, the granary weevil is able to crawl up the side of glass. This required the use of cloth covers for the jars in order to keep the weevils confined. A large colony of the granary weevil and the confused flour beetle was soon built up, however, the establishment of the common bean weevil colony proved more difficult. The same marking procedure used for the confused flour beetle and the granary weevil was used for the bean weevil. Small glass jars and four oviposition cages (Fig. 5), as that used by Pyenson and Menusan (17), were used. The eggs were collected in a petri dish at the botton of each cage. At weekly intervals, the eggs were reclaimed and placed in the jars which contained beans. According to Back (1), the usual length of life of the common bean weevil is about two weeks during the active season. It was noted that death occurred in about a wbek among the newly emerged adults. An investigation was made to find the cause of such high mortality, but unfortunately, the cause was not found. Nevertheless, the colony, although weak, was maintained and enough adults were obtained to use in at 14 least one test (Accelerated Electron Test 7). The insects were handled carefully at all times and no injured insects were used in the tests. In preparing test samples of the confused flour beetle, a 50 — mesh sieve was used. The flour from the breeding jars was sifted, with the adults remaining in the sieve. A card was placed in the sieve, and after a large number of beetles had climbed on the card, it was held over an empty petri dish. The beetles were counted as they were brushed off into the dish (Fig. 5). When the desired number was completed the card was placed back in the sieve while a recount was made of the beetles in the dish. The beetles were transferred from the petri dishes to flour samples to be used. This method of handling was re- peated according to the number of samples to be treated. After each test, the beetles, where flour was used were sifted out, and the live ones were brushed off from the sieve into a vial containing fresh flour. A separate vial was used for each sample and was marked with the appropriate sample number. The use of vials, because of their smallness, provided easier handling in mortality counts and provided a means to conserve space in the incubators. In preparing test samples of the granary weevil, an eight- mesh sieve was used. The wheat from the stock jars was put in the sieve and with a few gentle taps on the sieve the weevils would fall through to a large glass vessel (Fig. 6). The wee- vils were counted as they were brushed into small jars. When 15 Fig. 5. Apparatus used to count beetles. 16 the desired number was achieved the jar was immediately covered with a cloth in order to keep the weevils in the jar. Upon the completion of the sample counts, each jar was un- covered long enough to add the desired amount of wheat. Dur- ing the tests the contents of each sample was placed in a petri dish just long enough for the treatment to be performed, then the weevils were placed in a jar. When all the testing was finished the weevils were sifted out, a mortality count was made and the weevils were placed in fresh wheat. The same method, sieve and jar size used for the granary weevil tests was used for the common bean weevil test. Whenever an egg test was made of the confused flour beetle and the granary weevil, the adults were placed in the samples of food materials to oviposit for at least three days (except accelerated Electron Test 1 which was 2; days). The tests, except Infrared Test 5, were made on both the adults and the eggs in the same sample. Immediately after the test, the adults were separated from the samples and placed in fresh material in the incubators. The samples containing the eggs were also placed back in the incubators and observations as to number of emerging adults were recorded. Most of the insects used for the tests were about one month old. None were over a month and one half 01d, nor were they used more than once. However, before enough confused flour beetles were produced from the breeding colony for the tests, adults from a standing culture were used for the in- 17 Fig. 6. Apparatus and method used to prepare weevils for the tests. 18 frared tests. The amounts of food materials used, observations, special handling and the number of samples, including check samples, will be discussed in the tests. 19 IV TESTING EOUlPEENT AND PROCEDURE Only a brief description of the equipment and dosages used will be presented. For a more detailed discussion of this subject, the reader is referred to the writer's colleague, Baker (3). Infrared A series of six tests was conducted and the major equip- ment used in the tests is shown in Fig. 7. A type R—40/250 - watt lamp was used as a source of energy. The lamp height for all tests was measured from the bottom of the lamp to the bottom of the petri dish. The lamp voltage was maintained at 117 volts, unless otherwise indicated. There were slight differences in the procedure of each test. Test 1. The test was in two parts and was of a preliminary nature. The radiant energy was changed by varying the voltage to the lamp with a variable transformer, Fig. 7. A type DW-60 radiation meter was used to measure the energy. The various levels of infrared energy used and data collected for the test are shown in Table l. Twenty adult confused floured beetles placed in each of twenty 9-cm plain petri dishes were used for the first part of the test and twenty-five adults placed in each of 5 petri dishes were used for the second part of the test. Test 2. Twenty adult confused beetles were placed in each 20 Fig. 7. Equipment used for the infrared tests. From left to right; variable transformer, inclined rod with a thermocouple, stOp watch and a DW-60 radiation meter. In the center is the type R-40/z50 W. lamp. of 25 petri dishes. In addition a petri dish sample with the same number of beetles was used as a check. During the test the petri dish containing the insects was set on an aluminum foil insulated block. The energy was measured with a DW-BO radiation meter and the lamp height was measured from the bottom of the petri dish to the bottom of the 250 W type R-40 infrared bulb. The data collected for this test is shown in Table 2, and the apparatus used is shown in Fig. 7. Test 5. This test was designed to determine the effects of infrared energy on confused flour beetle eggs. Fifty adult beetles were placed in each of 50 petri dishes containing 20 grams of whole wheat flour, which was sifted through a bO-mesh sieve before being used. The adults were left to ovi- posit in the samples for three and one half days. During this 76 hour period all the samples were kept in an incubator. Just before the testing period the adults were removed from the flour. The eggs and flour in each of the 25 samples were treated with various amounts of infrared energy. The flour in each sam- ple was leveled evenly so that the depth of the flour was about one-fourth of an inch. Five samples were used as checks. The lamp was turned off at the end of each eXposure time and the thermocouple attached to the inclined rod (Fig. 7) was inserted into the flour. The thermocouple junction touched the bottom of the petri dish and the readings, in millivolts, were measured in the Leeds and Northrup potentiometer (Fig. 8). After the testing period the samples were put back in an incubator for Eb about 56 days. The number of eggs hatched after incubation and other data for this test are shown in Table 5. Test 4. This test was designed to determine the effects of infrared energy on adult confused flour beetles covered with flour. Twenty adult beetles were placed in each of thirty petri dishes containing a0 grams of whole wheat flour, which was sifted through a aO-mesh sieve before being used. Be— fore eXposing each test sample, the insects were covered with the flour and the flour leveled so that the depth was about i inch. Immediately after each exposure, the thermocouple on the inclined rod, Fig. 7, was inserted in the flour so that the element touched the bottom of the petri dish. A Brown potentiometer was used to record the temperatures. After the test, the samples were placed in an incubator and observed at Specified intervals. Mortality counts and other data for this test are shown in Table 4. Test 5. In this test the time of exposure was increased and the adults together with the eggs were tested in the same samples. Forty adults were placed in each of 30 petri dishes containing 10 grams of whole wneat flour. The adults remained in the samples for four days before the testing period in or- der to allow sufficient time for the adults to oviposit. Twen- ty five samples containing the adults and eggs were treated. Five samples were used as checks. Before each test the flour was leveled in each dish, however, no effort was made to cover the insects with flour. At the end of each exposure, the thermocouple, Fig. 7, was inserted into the flour so that the element touched the bottom of the petri dish. Immediately after the test the adults were separated from the samples and placed in fresh flour. The lamp heights, temperature, ex- posure times, mortality counts and eggs hatched are shown in Table 5, and Figs. ll A and B. Test 6. Test 6 was designed to determine the effects of infrared energy on granary weevil adults and eggs. Twenty five adult weevils were placed in each of twelve small Jars contain- ing 20 grams of wheat. The adults remained in the jars for four days before the test in order to allow sufficient time for ovi- position to take place. Ten samples containing adults and eggs were treated in petri dishes. Two samples were used as checks. At the end of each exposure the contents of the dish were poured into the insulated thermocouple cavity (Shown in Fig. 8), and the temperature was recorded by a Brown potentiometer, after which the sample was returned to the jar. When the testing was completed the adults were separated from samples and placed in fresh wheat. The samples containing the adults and the samples containing the eggs were placed in an incubator. Mortality counts and number of eggs hatched, as well as other data are shown in Tables 6 and 7, and Figs. 16A and B. Ultraviolet Two ultraviolet tests were made and the major equipment used in the tests is shown in Fig. 8. A GE type UA-z UVIARC 250 W mercury vapor lamp with a radiation wavelength shorter C d4 Fig. 8. \t , . Equipment used for the ultraviolet tests. From left to right; a Leeds and Northrup potentiometer, variable transformer, thermocouple and cavity, and a parabolic reflector containing the UA—z lamp. Under the lamp is the aluminum foil block used to set the test samples. than 2880A was used as a source of ultraviolet energy. The energy distribution of this lamp was computed by Baker (2). The lamp, mounted under a parabolic reflector, was used at different heights in the tests. The lamp was measured from the point of maximum curvature of the reflector to the bottom of the petri dish. The voltage was maintained at 11? throughout the tests. Tests 1A and 1B. Forty adult confused flour beetles were placed in each of 18 petri dishes containing 5 grams of whole wheat flour to oviposit for four days before the test. Fifteen of the samples were tested and three samples were used as checks. At the end of each eXposure a thermocouple was placed under the layer of flour and the temperature was recorded. Upon the com— pletion of the test a mortality count was made and the live adults were placed in fresh flour. The samples containing the adults and the samples containing the eggs were placed in an incubator. The results and data for this test are shown in Tables 8 and 9. Test 2A and 2B. Twenty—five adult granary weevils were placed in 12 small jars containing 20 grams of wheat. In order to allow enough time for ovipositing, the adults remained in the samples for four days before the test. Before each treat- ment the contents of the jar were placed in a petri dish. After the exposure the contents of the petri dish were poured into the thermocouple cavity (Fig. 8). The maximum temperature was recorded and the contents poured in the jar. At the com- pletion of the testing period the weevils were separated, a mortality count was made and the live adults were placed in fresh wheat. The samples containing the live adults and the samples containing the eggs were placed in an incubator. The results and data for this test are shown in Tables 10 and ll. X-Ray Two x—ray tests were made. The test insect for these tests was the confused flour beetle only. A Hilger machine type 50 KV. was used for Test 1 and a GE Maximar 250 III, 250 KV was used for Test 2. Test 1. The x-ray unit used for this test was a 50 KV Hilger machine of the Physics Department of Michigan State College (Figs. 9A and 9B). Forty adult confused flour beetles were placed in each of 15 petri dishes containing 15.5 grams of whole wheat flour. In order to allow sufficient time for ovipositing the adults remained in the flour for three days before the test. Just before the testing began, the samples were each placed in a cardboard box, 1.5 inches in diameter and .75 inch thick. The test samples were mounted 15.5 inches from the target in order to utilize the maximum beam area. At the time of this test, the dosimeter for the x-ray unit was out of order, therefore no dosages were calculated for this test. After the testing period the adults were separated from the samples and placed in fresh flour. One week later the adults were separated from these samples and again placed in 27 Fig. 9A. General view of the 50 KV Hilger machine used for the x—ray tests. Fig. 9B. Close up view of controls for the Hilger machine. A test sample may be seen in the upper right part of the picture. fresh flour. The flour was saved in order to check on the fertility of the adults irradiated with x—rays. A fertility check was made of the egg test adults also. Approximately 15 days after the appearance of the first adult in the irradiated egg samples, including the check samples, a count was made and the adults were placed in fresh flour. A week later the adults were discarded and the flour was saved for further observations. The exposure time and other observations are in Table 12. Test 2. The x-ray unit used for this test was a GE Maxi- mar 250 III, 250 KV of the School of Veterinary Medicine, Michigan State College. Thirty adult confused flour beetles were placed in each of 15 petri dishes containing 10 grams of whole wheat flour. The adults remained in the samples for three days in order to allow enough time for ovipositing. Twelve samples were irradiated and three samples were used as checks. Unlike Test 1 the treatment was made in the petri dishes. After the test the adults were separated from the sam- ples and placed in fresh flour. The same methods used in Test 1 in checking the fertility of the treated adults and adults from the treated eggs, including all check samples, were used in this test. Observations, dosages and specifications of the x-ray unit used are shown in Table 15. ‘Accelerated Electrons A series of seven tests were conducted with the Van de Graaff generator (Figs. 10A and 10B) of the Upjohn Company at Kalamazoo, Michigan. These tests include irradiations made on the confused flour beetle larva and adults of the common bean weevil. The insects were transported to and from Kala— mazoo by car in an insulated box. (Fig. 5) Tests 1, 2A, 234‘3294. Since this procedure for each of these tests was similar a general description of the pro- cedure is presented under one heading. Forty adult confused flour beetles were placed in each of twenty-four petri dishes containing 15 grams of whole wheat flour for Test 1, (Table 14). The adults remained in the flour for two and one-half days before the test, in order to allow time for the adults to oviposit. In Test 2, one hundred adults were placed in each of la petri dishes containing 50 grams of whole wheat flour. The adults remained in the flour for the three days in order to allow time for the adults to oviposit. In Test 4 fifty adults were placed in 24 petri dishes without flour. The samples were placed on the Van de Graaff generator conveyor belt and treated. The number of replicates, dosages and the number of check samples are shown in Tables 14, 15, 16, and 18. After the tests, the adults were separated from the samples and placed in fresh flour. The beetles from Test 4 were also placed in fresh flour. Observations were made at various intervals as shown in the tables. At the end of one week a final observation was made and the adults were dis- carded, but the flour was saved in order that a fertility check 50 Fig. 10A. View of conveyor belt, shielding blocks and vacuum pump for the Van de Graaff generator. Picture courtesy of the Upjohn Company. Fig. 10B. General view of The Van de Graaff generator and controls. Picture courtesy of The Upjohn Company. 31 COMPRESSED NITROGEN CHARGED HIGH INSULATION VOLTAGE TERMINAL “R FILAMENT ELECTRON SOURCE IN EVACUATED ACCELERATION CHARGE” TUBE ~ REMOVED FROM BELT BELT CHARGED BY CORONA UNSCANNED ELECTRON BEAM ELECTRON BEAM SCANNING SYSTEM THIN ALUMINUM / WINDOW IRRADIATED PRODUCT Fig 11 Schematic sketch Of the 'Van de Graaff electron accelerator. 32. could be made of the adults. A fertility check was made Of the egg test adults also. About two weeks after the appear- ance of the first adult in the irradiated and check samples, the beetles were placed in fresh flour. A week later, the adults were discarded and the flour was saved for further observations. The dosages, observations, and the number of check sam- ples used for each test are shown in Tables 14, 15, 16, 17, and Figs. l7AB and 18AB. Test 5. This test was designed to determine the effects of accelerated electrons on the larvae of the confused flour beetle. Since the method of handling for this test was not discussed under the section, "Procedures In Rearing And Hand- ling The Test Insects", a specific method is presented here. Flour from the stock culture of the confused flour beetle was sifted through a 20-mesh wire screen. The screen separated the adults, the pupae and some of what appeared to be the last instar stage of the larvae. The flour then was sifted through a 40-mesh wire screen as shown in Fig. 3. The flour contain- ing eggs, and possibly the first two instars of the larvae, fell through the sieve. The sieve retained the other instar stages and larger particles of flour. The wire screen with the larvae was inverted on a piece of paper and the method was repeated until it was assumed that enough larvae had been collected. The larvae were divided among twenty-four petri dishes, and flour was added to the petri dishes so that each 5§ contained 13.5 grams of flour. The confused flour beetle larva is very delicate and any repeated handling may injure the insect. In order to avoid the use of an injured larvae no attempt was made to count them. A visual criterion (Fig. 21) was used to determine the activity of the larvae. Several hours before the activity check was made, each sample was gently tapped on the side of the dish. The tunnels made at the bottom of the petri dish by the larvae would collapse and the flour was leveled. The activity was determined several hours later by the amount of tunneling made by the crawling larvae on the bottom of the petri dish. The activity check was made at weekly intervals as shown in Table 19. A sterility observation was made on the emerged adults. Tests 3, 6 and 7. Since the procedure for each of these tests was similar, a general description of the procedure is presented under one heading. Forty adult granary weevils were placed in each of 25 small jars containing 30 grams of wheat for Test 3. The adults remained in the wheat three days before the test in order to allow enough time for the weevils to oviposit. No egg tests were made in Tests 6 and 7. In Test 6, fifty adult granary weevils were placed in each Of 24 empty small jars and in Test 7, fifteen adult common bean weevils were placed in each of ten small jars containing 20 grams of beans. The samples for tests 6 and 7 were prepared several hours before testing period. Before each test was performed the contents of each jar were placed in petri dishes. Each dish was covered with a piece of aluminum foil in order to Keep the weevils confined. The samples were then treated with various dosages. No wheat was used for Test 6. The number of replicates, dosages, and the number of check samples are shown in Tables 17, 20, and 21. The treated and untreated samples were observed immediately. The weevils were placed in jars containing fresh food materials. The wheat containing the eggs of Test 3 was saved and observations as to the number of hatches were made. A sterility check was made on the adults of Test 6 and the adults of Test 3, including the adults from the irradiated eggs. No sterility check was made on the adults of the common bean weevils. The results of Tests 6 and 7 are shown in Tables 20, 21 and the results of Test 3 are shown in Table 17 and Figs. 20 AB. Mortality Counts The criterion for death was the inability of the adult beetle to crawl on a flat surface. This criterion was used by Collins (5). When mortality counts were made the beetles were sifted from the food material and placed on a piece of paper. The adults of the granary weevil were given longer time, because these weevils feign death whenever jarred or handled. The dead or moribund beetles, after given enough time to recover, were discarded. The live beetles were put back in the containers and returned to the incubators. Mor- 55 tality counts were made of each sample at various intervals as shown throughout the tables. However, only one mortality count was made of the confused flour beetle larvae (Test 5). The criterion for death in this test was easily established. 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A Percent Tri helium eggs hatched for infrared test 3. {‘3 l6 . “O 2 2 I4 econds (s) a .5 a 342 3 l .. check c: I. --. d) . 2» ‘ at " ~. 60f * I. 80 90 too llO Temperature Deg F Fleisa’ Percent Tribolium eggs hatched bar chart for , infrared test 3. PERCENT ADULTS KILLE o 2 3 4 5 - GR. CAL/SQ CM/MIN. l0 8 6 ‘ 4 2 LAMP- HEIGHT IN INCHES ‘ Fig It A Pecent adult Tribolium killed for infrared test 3 l P ERCE NT ADgLTS KILLED N o .o ' llO l20 < I30 I40 .ISO 160 I70 l90 2lO 220 ' TEMP DEG F Fig M- 8 Partial barchart for percent Tribolium adults killed for infrared test 4. 115 fi (D O T PERCENT ADULTS KILLEQ 3 8 f I N O I GR. CAL] SQCM/ MIN. IO 8 6 4 2 LAMP HEIGHT IN INCHES FigISA Parcent Tribolium killed for infrared test 5. 2.5 I5 2.0 2.0 25 g (I) O 1 PERCENT ADULTS KILLED 4) CD 0 <2 I N Q 1 90 I00 IIO I20 I30 I40 I50 I60 I70 l80 TEMP DEG F FigISB Partial bar chart for percent Tribolium adults killed for infrared test 5. 45 A O N (D PERCENT EGGS HATCHED 8 ~2.0‘I.5 \I\0 \5 time of exposure in \::\ minutes \\ \\ t\‘ \ \\\ ‘~- °\_~\ \‘\1 GI I I #5? 3"I 2 3 4 5 GR. CAL/SQCM/MIN. I0 8 6 4 2 LAMP HEIGHT IN INCHES FIG I“ A Percent Tribolium eggs hatched for infrared test 5. o In :I: 280 Ex time of exposure in minutes 8 I360 ‘ I- average of I45 eggs hatched in A 34, check samples 0 m 7 0 m a. 20 0 [not I I I 90 I00 IIO l20 I30 I40 I50 I60 I70 I80 * TEMP DEG F ~ FIG 068‘ Partial bar chart for percent Tribolium eggs hatched for infrared test 5. 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COMOOOHE UOHGLOHOOO‘ 61 (n o PERCENT ADULTS KILLED O 2."! In g m 0 PERCENT EGGS HATCHED N vb 9 O O) O .5 O N O 18 TRIBOLIUM ADULT 720 '72 reps x l000 (Flour Beetle) CHECK 46 eggs average hatch in check samples « N0 EGGS HATCHED » TRIBOLIUM EGGS IO reps x l000 62 72 1 1 720 L l .l450 Fig l8 A-B Percent Tribolium confusum adultskllled and eggs hatched for accelerated electron test I using Van de Graaff generator at UPJOHN CO. Feb 5, l953 - Data in table l00 ADULT Sit0philus granarius L. 0 m8 .1 Es" (Granary Weevil) 0,60 5 3 240 I‘- z 23 0:20 In a. O L l l000 500 250 l00 50 CHECK Fig A reps x IOOO loo 8 IO 3 eggs average hatch incheck samples :I: 080 I- <1: :1: 0,60 0 3 h NO EGGS HATCHED q 4 g S it0philus granarius eggs 322 m a. l J l ‘ l l l J l I 1 IOOO 500 250 l00 50 CHECK reps x IOOO ‘ Fig 8 Fig 9.0.A-B Percent Sitophilus granarius adults killed and eggs hatched for accelerated electron test 3 using the Van de Graaff generator at UPJOHN CO. 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VI DISCUSSION OF RESULTS Infrared The procedure for this series of tests is described in sections III and IV. The results of these tests are recorded in Tables 1, 2, 5, 4, 5, 6 and 7. The results of Tests 2, 5, 4, 5, and 6, appear in graphic form in Figs. 12, 15, 14, 15, 16, and 17. As can be seen from Table 1, first part of Test 1, only one beetle was dead 48 hours after the test. However, better results were obtained in the second part of the test. Thirteen out of a total of twenty five adults were dead 48 hours after the test with a lamp height of 10 inches. In test 5, no treat- ment used indicated 100 percent kill of eggs. However, in Test 4, a 100 percent kill of adults was obtained one week after treatment using an exposure time of 105 seconds and a maximum temperature of 215°F., with the height of the lamp at two inches. During this test a number of insects came to the tOp and there does not appear to be any relationship between the number of insects killed and the number that came to the tOp of the sample. The results of Test 5 and 6 were more promising. In Test 5, the minimum time required to kill 100 percent adult confused flour beetles, one hour after the test, was 2.5 minutes at a lamp height of eight inches. The maximum temperature at the end of the test was 127OF. The minimum time required to kill 100 per- cent of the eggs was 2.5 minutes at a lamp height of 6 inches 68 with a maximum temperature of 1580F. This temperature more or less conforms to that of Cotton (6). He states that a temperature of 14OOF. is fatal to all grain insects in 10 min- utes time. In test 6, 100 percent kill of adult granary weevils and eggs was obtained with an exposure time of 1.5 minutes at a lamp height of 4 inches and a maximum temperature of 128°F. at the end of the test. In reviewing the exposure times in the tables of these two tests, it was noted that a longer ex- posure time was required to increase the temperature of the wheat. The granary weevil lays its eggs inside the wheat ker- nel and the confused flour beetle deposits its eggs at random in the flour. Apparently the wheat provided a better insup lation for the granary weevil eggs. Ultraviolet The results of these tests are recorded in Tables 8, 9, 10 and 11. The procedure for these tests is described in sections III and IV. One hundred percent kill of the adult confused flour beetles was obtained at an eXposure time of 8 minutes with a lamp height of 10 inches one hour after treatment. The maxi— mum temperature recorded was 108OF. At this same temperature, lamp height and exposure time, only 1.6 percent of the eggs hatched, and no eggs hatched at an eXposure time of 4 minutes with a lamp height of 7 inches. No adults or eggs survived 69 one hour after treatment with the lamp height of 7 inches and an eXposure time of 4 minutes. The maximum temperature re- corded was 140°F. The confused flour beetle eggs, which are laid at random in the flour, were effected at a lower temperature than the eggs of the granary weevil. Since ultraviolet energy has poor penetration into the wheat kernel, and the fact that the gran— ary weevil deposits its eggs inside the wheat kernel, more energy would be required to kill the egg. However, a temper- ature of 155°F. was fatal to the adults and eggs. The ultraviolet energy radiated from the lamp used in these tests was about one fifth of the total energy, and the other four fifths was infrared energy. Under these conditions it would be difficult to separate the effects of ultraviolet energy from the effects of infrared energy. X-ray The data recorded for these tests are shown in Tables 12 and 15. The procedure, including the sterility checking method, is described in sections III and IV. No eXposure time used in Test 1 for the confused flour beetle adults showed any effects at one hour, or at one week after treatment. The eggs in the egg samples hatched normally as compared to the check samples. In Test 2, one adult died with a dose of l48r units, two adults died with a dose of 70 296r units and two adults died with a dose of 5szr units, two weeks after the treatment. These results may be considered negligible since no adults died with the higher treatments and one died in the cheCk sample two weeks after the test. There was no noticeable external effects on the progeny of the irradi— ated adults nor on the progeny of the irradiated eggs. Accelerated Electrons The results for these tests are presented in Tables 14 through 21 inclusive. Bar charts for Tests 1, 2 and 5, are presented in Figs. l8, l9 and 20. The procedure is given in sections III and IV. Table 14, and bar chart Fig. 18, show that an electron dose of 7so,ooo rep* will kill 100 percent of the adult flour beetles in flour after treatment, whereas as electron dose of 500,000 rep will kill 100 percent of the adult granary weevils in wheat after treatment (Table 17, bar chart Fig. 20). The results for these two Tables (14 and 17) indicate that with a dose of 100,000 rep about 90 percent of the granary weevil adults were killed one week after treatment and a dose of 72,000 rep about 10 percent of the confused flour beetles were killed one week after treatment. In Tests 4 and 6 (Tables 18 and 20) no flour or wheat was used. The results indicate that a dose of 200,000 rep *One rep represents a very minute quantity of energy. One rep (Roentgen—equivalent~physical) = l roentgen: l roentgen = 95 ergs/gram water or tissue. 71 will kill 100 percent of the adult flour beetle after treat- 'ment, whereas a dose of 250,000 rep will kill 100 percent of the adult granary weevils after treatment. A dose of 100,000 rep will kill about 95 percent of the adult flour beetle and a dose of 50,000 will kill about 40 percent of the adult granary weevils. An electron dose of 10,000 rep was lethal to 100 percent of the adult bean weevils one week after treat- ment (Test 7, Table 21). A dose of 75,000 rep killed 100 per- cent of the confused flour beetle larvae one week after treat- ment (Test 5, Table 20). The results shown in Tables 14 and 17, indicate that an electron dose of 10,000 rep prevented the hatching of eggs of the flour beetles and the eggs of the granary weevils. This same dose prevented the adult granary weevil, the adult con~ fused flour beetle and the larvae of the flour beetles, after reaching the adult stage, from reproducing. There were no noticeable external effects on the progeny of the irradiated adults, larvae or eggs. VII SUMMARY Infrared, ultraviolet and x-rays, as well as accelerated electrons were used on the confused flour beetle. The granary weevil was subjected to infrared rays, ultraviolet rays and accelerated electrons and the common bean weevil was limited to only one test of accelerated electrons. The beetles were reared under laboratory conditions of 8005 2°F. and a relative humidity of about 66 percent. The manner of rearing was so arranged that a constant supply of adult beetles of a known age were produced. The infrared energy tests indicated that both temperature and exposure time were important factors in obtaining effective results. A temperature of at least lBB‘F. and a maximum ex— posure time of 2.5 minutes was necessary to obtain lethal effects on the confused flour beetle, granary weevil and their eggs. The ultraviolet tests gave no clear indication of its effects on the beetles since the type of lamp used emitted four fifths of its energy as infrared energy. No exposure time in the x-ray tests showed any effects on the confused flour beetle. After the tests the adults were not sterilized and were able to reproduce normal progeny. However, the results of the accelerated electron tests were more promis- ing. A dose of at least 500,000 rep would be necessary to kill completely a mixed pOpulation of adults and larvae of the con- fused flour beetle, the adults of the granary weevil and common bean weevil. A dose of 10,000 rep prevented the eggs of the '73 confused flour beetle and the granary weevil from hatching. This same dose sterilized the adult granary weevil, the adult confused flour beetle and prevented the larvae of the flour beetle, after reaching the adult stage, from reproducing. 74 1. 2. 10. ll. 12. 13. VIII LITERATURE CITED Back, E.A., 1922. Weevils in Beans and Peas, USDA Farmers Bulletin No. 1295. Baker, V.H., 1955. Some effects of electromagnetic energy and subatomic particles on certain insects which infest wheat, flour and beans. Dissertation for degree of Ph. D., Dept. of Agr. Engin., Mich. State Coll., (unpublished). Blazer, A.I., 1942. "Heating and Drying", Insect Pests g; Stored Rice and Their Control, USDA Farmers Bulletin No. 1906. Burrill, A.E., and Gale, A.J., 1952 (Nov.). "Electron Beam Sterilizes Food and Drugs", Electronics, Nov. Collins, E.W., 1952. The Extension and Refinement of a Laboratory Technique for Evaluating Contact Insecticides, Thesis for M.S. degree, Dept. of Ent. Mich. State Coll., (Unpublished). Cotton, R.T., 1952. Insect Pests gf_§tored Grain and Grain Products. Burgess Pub. Co. Minneapolis, p. l, 226. Dean, G.A., 1916. Heat as a means of controlling mill insects. Jour. Econ. Ent. Vol. 4, pp. 142—158. Dunning, W.F., 1951. A study of the effect of x-ray radi- ation on occurrence of abnormal individuals, mutation rate, viability and fertility of the parasitic wasp Habrobracon jgglandis, Ashmead. Genetics, Vol. 16, pp. 505-551. Ellis, C., and Wells, A.A., 1941. The Chemical Action 2:_ Ultraviolet Rays. Reinhold Pub. Co. N.Y. pp. 692-752. Headlee, T.J., and Joblins, D.M., 1958. Progress to Date on Studies of Radio Waves and Related Forms of Energy for Insect Control. Jour. Ec. Ent., 51, pp. 559-565. Koidsumi, K., 1950. Quantitative studies on the lethal action of x—rays upon certain insects. Jour. Soc. Trop. AgI‘., V01. 2’ NO. 5, pp. 245-2620 Lindgren, D.w., 1957. "Grain Weevils", Rearing Stored Food Insects for Experimental use in_Culture Methods for Invertebrate Animals, Comstock Pub. Co. Inc. Ithaca, N.Y. pp. 481-485. MacLeod, G.F., 1955. Effects of Ultraviolet Radiations on the Bean Weevil, Bruchus obtectus Say. Ann. Ent. Soc. Amer. 26, 605-615. 75 14. 15. 16. 17. 18. 19. Movar, J.H., 1927. A comparison of the susceptibility to x-rays of Drosophila melanogaster at various stages of its life cycle, g, Exp. Z001., Vol. 47, No. 1, pp. 65-85. Park, T. Observations on the general biology of the flour beetle, Tribolium confusum Quart. Rev. Biol. 9:56, 1954. Proctor, B.E., and Goldblith, S.A. Electromagnetic radi- ation fundamentals and their application in food technology. Advances in Food Research 5:160-196. Pyenson, L. and Menusan Jr., H., 1957. Culture Methods for Invertebrate Animals. Comstock Pub. Co. Inc. Ithaca, N.Y. 5?. 478-480. Ray, 0.M., and Bodine, J.H., 1958. Ultraviolet radiation of grasshOpper eggs. Physiol Zool. 11:267-77, J1. Robinson, W., 1926. Low temperature and moisture as factors in the Ecology of the rice weevil, Sitophilus oryza L., and the granary weevil, Sitophilus granarius L., Univ. 9§_Ndnn. Tech. Bul. No. 41, pp. 4-5. Shepard, H., 1945. Bearing Insects That Attack Stored Pro- ducts. Laboratory Procedures in Studies of the Chemical ~m’~ Control of Insects, A. A. A. S. No. 20, Washington, D. C., p. 56. , 1959. The Chemistry and Toxicology 9§_Insecti- cides. Burgess Pub. Co. Minneapolis, Ndnn., pp. 52-55. Trump, J.G., and Van de Graaff, R. J., 1948. Irradiation of Biological Materials by High-Energy Roentgen Rays and Cathod Rays. Jour. 2; Applied Physics, Vol. 19, No. 7, July. Wexler, A., and Brombacker, W.G., 1951. MethOds of Measur- ing Humidity and Testing Hygrometers. N.B. of 8., Cir. 52, p. 15. Whiting, P.W., 1929. X-rays and parasitic wasps. Jour. Heredo, V01. 30, pp. 268-276. Wigglesworth, V.B. 1940. Principles 9§_Insect Physiology. E.P. Dutton Co. New York, N.Y., pp. 560-562. Yeomans, A.H., and Rogers, B.E., 1950. The second eXperi- ment using the capacitron treatment on various insects. Spec. Bept. IN2-21. U.S. Dept. of Agr. Bur. of Ent. Wash- ington, D.C., 9pp. 76 -J VIVVIVI -.. “M“~—..u_. UOL ‘u--— .0 INTER-HERA?! LGAN ' DEC 2 1 '55 M ay 13 .’57 4". v -.I.., at? ,: , ......i.n.ln.... » .t :3 n. {It}. [filly HI 7 .lrL MICHIGAN STATE UNIVERSITY LIBRARIES II II I ll llllllllll 3 1193 03174 8431