Reprinted from "THE AQUARIUM JOURNAL," (San Francisco) Vol. XXV, nos. 6 and 7, (June and July 1954) (Published May 25 and June 23, 1954) W. L. Minckwill. CULTURING FRUIT FLIES (Drosophila), in-- AND OTHER LIVE FISH FOOD Part One of Two Parts

By CLARK HUBBS and KIRK STRAWN University of Texas

osz AQuAruszs agree that live food is Previous Use of Fruit Flies to Feed Fish good for fishes. In the experi- and Other Animals mental laboratories at the University of Texas we culture Drosophila, Daphnia Information on feeding Drosophila to magna, microworms, infusorians, and fish is scanty in aquarium literature. Houston white worms. Detailed culture Feeding vestigial winged ( flightless ) methods, using a banana mash medium, adults, a mutant strain of D. melano- have been worked out for the first three garter, has been recommended by Pop- forms and are presented in the follow- pel ( 1951 ). Mutant strains usually do ing paragraphs. not culture as well as normal or wild type flies. Axelrod ( 1952) briefly men- Fruit Flies tions using Drosophila for aquarium feeding. Neither Innes ( 1952 ) nor Stoye Fruit flies (Drosophila) form one of ( 1935) mention Drosophila. the few live foods that can be raised in Likewise, there is little published in- quantity by the aquarist. Their culture formation on feeding Drosophila to can be carried out in glass containers other animals. Burger ( 1937 ) recom- that occupy very little space. With rea- mends feeding adults to black widow sonable care the cultures will maintain spiders. Hoffman ( 1924 ) mentions the themselves and produce large volumes ease of raising adults and the difficulty of living food. of capturing them for feeding waterbugs. . In our original work we found that Breland (1949) was successful in raising Drosophila served as an excellent aqua- carnivorous mosquito larvae on a com- rium fish food. We used excess fly larvae bination of larval and adult Drosophila. from the Genetics Laboratory at the Making use of the facilities at the University of Texas. As our investiga- Genetics Laboratory of the University tions widened, we found that this excess of Texas, and the kindness of the geneti- supply was not sufficient to meet our cists, other laboratory animals have been needs, and experiments were initiated raised on fruit flies. Dr. John M. Cairns, to determine methods for raising larger then on the staff of the University of quantities. We have received much as- Texas Department of Zoology, fed sala- sistance from the genetics workers and manders on Drosophila larvae. Gradu- others during the investigations.' ate students working under Dr. W. Frank Blair have raised other amphibi- Dr. Marshal R. Wheeler, Assistant Professor of ans on adult Drosophila, as follows: Dr. Zoology at the University of Texas read and Wilmot A. Thornton, toads of the genus criticized the Drosophila discussion. Dr. John Bufo; Aaron 0. Wasserman, spadefoots M. Cairns, Assistant Professor at the University of the genus F. of Oklahoma Medical School aided the early Scaphiopus; William Py- development of the feeding techniques. burn, cricket frogs of the genus Acris;

JUNE 1954 135 and Hague L. Lindsay and John S. e. After cooling ( heat kills yeast) in- Mecham, tree frogs of the genus Pseu- noculate with a pinch of living yeast clacris. and plug tightly with sterile cotton. The Culture Medium Do not remove plug except when flies or larvae are being transferred Although there are many media used (keeps out undesirable mold, bac- for raising and feeding Drosophila, we teria, flies, and ants. Keeps in Droso- employ only the one used by the genetics phila. Keeps medium moist.) Re- laboratory at the University of Texas. frigerated, the medium will last in As the cultured non-gas-forming yeast first class condition for two weeks, used in the laboratory is not available and can be used during the subse- to aquarists, a simplified method of quent two weeks. preparation is given below. f. Place in refrigerator until about three Ingredients for one liter' of medium: days before using. It is ready for use 1) 90 cc water when a heavy growth of yeast is on surface. The yeast forms a more or 2) 20 grams agar ( can be obtained from any biological or chemical supply less pasty covering over the medium. It grows very close to the medium, house such as Turtox Laboratories or seldom becoming high enough to be E. H. Sargent & Co., 4647 West Foster seen from the side. ( Be careful of Avenue, Chicago 30, Illinois). mold. Do not use moldy cultures. 3) 25 cc malt extract ( Blue Ribbon) Molds form a more or less powdery 4) 25 cc syrup ( Karo) covering over the medium. It often 5) 30 grams brewer's yeast ( usually forms a fine thread-like covering over found in any health food store). the medium that usually can be seen 6) 100 cc water easily from the side. Yeast is usually 7) 2 lbs. bananas ( green to overripe) white, molds often dark colored. Ex- 8) living yeast ( Fleischman's fresh- perience is the best teacher. If the active yeast) larvae flourish the growth is yeast, if To prepare the medium: not it may be mold. Mold may be a. Heat and stir 1 and 2 to dissolve prevented by chemicals such as 5 cc agar. of propionic acid added before b. Heat 5 and 6 in pressure cooker for step c.) 45 minutes. Other methods and media for cultur- c. Mix all but 8 and boil gently for 15 ing Drosophila may be found in Sturte- minutes in covered container. vant (1937), Spencer ( 1950 ), and d. Place in sterile' (boiled) bottle to Fletcher (1953). We feel that the depth of about %" ( less depth permits method employed at the University of drying and greater depth is a waste Texas is the most satisfactory because it of medium) and cover with sterile does not leave particles of the medium cloth. The metric system of measurements is used in ' A sterile item is one that has no living organ- this article. It is more convenient and logical ism on it. Non-sterile items are sterilized than the better known Engli4h system. The when all of the living animals and plants on following are the equivalents of the metric them are lcilled. Heating in a pressure cooker units used here: for 20 to 30 minutes is a simple method of 454 grams = 1 pound sterilization. One must be careful not to con- 1057 cc = 1 quart taminate the sterile items when they are re- 1000 cc = 1 liter moved. When removing bottles, do not touch Many of the bottles and measuring devices the inside or the outside near the opening. for preparing baby formulas are marked in cc. Sterile cotton may be purchased in drug stores.

136 AQUARIUM JOURNAL with the larvae after washing. Difco Laboratories, 920 Henry Street, Detroit 1, Michigan, is experimenting with a commercial Drosophila medium.

The Culture Bottles Many kinds of bottles may be used to raise Drosophila. Most laboratories studying Drosophila use half pint milk bottles or smaller vials. The small con- tainers are less satisfactory because too few Drosophila can be raised in each container. It is possible to raise fruit flies in larger containers, but efficiency is lost with the increased size of the con- tainer. As the surface area exposed is a Fig. 1 — Adult males of (a) Drosophila major limiting factor, an increase of melanogaster, (b) Drosophila hydei, and (c) height is of no significant value. In- Drosophila virilis, all ten times natural size. creased diameter is a handicap when the The small fly at the bottom is a natural size flies are shaken from bottle to bottle, as D. melanogaster. Redrawn from Patterson the jelly-like medium is more likely to and Mainland (1944) by Grace Hewitt. come loose if the bottle diameter is large. Moreover, the cotton plug must be larger They not only will tolerate moderately ( and more expensive) with the in- high temperatures, but will also grow creased diameter of the mouth of the bot- well on a banana mash medium. The so- tle. The increased amount of cotton is called vestigial-winged fruit fly is a vari- far greater than the increased production ant of D. melanogaster. of Drosophila. Bottles with necks nar- rower than the base use less cotton and If particular, aquarists may get fruit the medium is less likely to fall out of the fly "starters" from reputable biological bottle during fly changing. The half pint supply houses such as Turtox Labora- milk bottle is not only satisfactory but tories, 761-763 East 69th Place, Chicago also inexpensive. 37, Illinois. If not particular, the aquar- ist can collect wild flies in a Drosophila In the half pint milk bottle is not avail- culture bottle ( see section on culture able almost any bottle will suffice. The bottles ). Many types of bait have been culture techniques can be used for any recommended in the literature. That bottle type. proposed by Dobzhansky ( 1936 ) is relatively simple: "Fermenting banana Kinds and Sources of Fruit Flies mash is most satisfactory as bait. Ripe Only a few of the 613 species of bananas are mashed with the aid of a Drosophila listed by Patterson and Stone spoon or a fork; some drops of a fresh ( 1952 ) are suitable to culture for fish yeast solution is added, and the bottle food. According to W. S. Stone ( per- is left standing for about 24 hours be- sonal communication, 1953), Drosophila fore use. The bait remains good for at melanogaster Meigen, D. simulans least four or five days after first used. Sturtevant, D. virilis Sturtevant, and D. The traps ( culture bottles) are exposed hydei Sturtevant are among the species in such a way as to be readily accessible. most likely to survive the variable con- and left undisturbed for a few hours; ditions found in homes. ( See Fig. 1.) no useful purpose whatever is accom-

JUNE 1954 137 pfished by leaving them exposed for The "worked" part is easily recognized days." We feel that the use of a bottle by being darker than the unused part. containing one-half inch of the culture medium described above will serve to After a few days the larvae will have capture wild Drosophila. After the flies reached maximum size and will begin have entered the bottle, a cotton plug to form pupae. Unlike the larvae all the can be inserted to keep them from es- pupae are motionless. Pupae are darker caping. than the larvae. Figure 2 illustrates pupae of three species. The wild flies can be captured in al- most any outdoor locality, both in cities and in rural regions. Large numbers - of flies are to be found around fruit as-_-- markets. Those flies are more likely to be forms that are easy to culture. It is difficult to capture flies at temperature extremes. Temperatures between 60 and 80° Farenheit are best. When the out- door temperatures are colder, flies might be captured during warm afternoons; and when it is warmer, dawn and dusk are the better collecting times.

Growing the Flies Fig. 2 — Pupae of (a) Drosophila melano- The new flies are placed on fresh gaster, (b) Drosophila hylei, and (c) Droso- medium. After one or two days the phila virilis, all ten times natural size. adults may begin to lay eggs, either on Redrawn from Patterson (1943) by Grace top of the medium or on the side of the Hewitt. bottle immediately above the medium, depending upon the species. Of the flies used in our work only D. virilis lays eggs on the bottle. The elongate eggs are nearly white. If a black pepper grain were elongate it would be about the same size as a Drosophila egg. Fre- quently the eggs are concentrated in one small area, and give a whitish cast to that area. When the eggs are discov- ered the transferral of parents should Fig. 3 — Large larva of Drosophila hydei, be started. ten times natural size. Drawn by Grace Hewitt. When one or two days have passed the eggs hatch. The young Drosophila is When the adult emerges from the a small maggot and when first seen is pupal case, the animal for the first time feeding on the surface of the medium or looks like a fly. Normally the males on the yeast growing on the bottle ad- emerge before the females. If one looks jacent to the medium. Figure 3 is an en- closely the sexes can be recognized by larged drawing of a large larva. As the the larger abdomen of the female. Ordi- larvae grow they begin to "work" or nary room lighting is satisfactory for all soften the upper layer of the medium. developmental stages of Drosophila.

138 AQUARIUM JOURNAL Transferring the Flies guppies and to native Texas darters of about one-half inch in length. Drosophila For the adults to lay many eggs they is not very satisfactory for feeding to must be fed well. To achieve best re- young fish that need a constant supply the adults should be transferred to sults of live food as the larvae do not live fresh media with a heavy growth of long enough under water. The maximum yeast. Daily changes are recommended yield of larvae is from the larger sizes. and D. hydei and every- for D. virilis Thus feeding small larvae to young fish D. simu- other-day-changes are best for is less efficient than feeding large larvae lans and D. melanogaster. to medium sized fishes. To transfer adults, invert the bottle The larvae from two D. virilis bottles containing the flies tightly over the new per day ( one for morning and one for bottle and shake the flies into the new night feedings) should be more than bottle. An up and down shaking motion adequate for most aquarists if the raising is better than side to side. A sudden techniques described below are fol- stop at the end of a downward plunge lowed. Four dozen half-pint milk bot- often succeeds when the flies do not tles should be all that most aquarists move otherwise. If the medium is not need. Each aquarist, however, must firm one should not jar the bottles dur- check to see if his Drosophila cultures ing shaking as the old medium will fall are adequate for his particular needs. into the new bottle. Cold flies transfer easier than warm flies. Harvesting Larvae Harvesting larvae just prior to pupa- Larval Stage is Best to Feed to Fish tion gives the maximum yield. We har- As stated above, during its life history vest the cultures after the first pupae Drosophila passes through four stages• appear. If a culture is refrigerated, de- developing egg, growing larva, metamor- velopment ceases; death eventually re- phosing pupa, and breeding adult. sults from prolonged refrigeration. Thus cultures may be maintained in a har- We use fruit fly larvae (not adults) to vestable stage for about one week if feed native Texas darters and minnows. placed in a refrigerator immediately All of the minnows and most of the after the first pupae appear. Care must darters tested take larvae readily. Young also be taken not to let the flies get too largemouth bass grow well on them. An- warm. A Farenheit temperature above gel fish and bettas are fond of the larvae the high 80's results in sterilization of — in fact, bettas prefer them to daphnia. the flies, and a higher temperature kills Local aquarists find that most tropicals them. will feed eagerly on Drosophila larvae. When the first pupae appear or the Drosophila is most satisfactory as a larvae have reached maximum size, the food for fish of guppy size or larger. De- top of the mash will be semi-liquid. pending upon the food habits of the fish This semi-liquid medium with all of the they will continue growing on larvae for larvae is freed from the firm remainder some time. Large angel fish flourish on by a stream of water. This mixture is Drosophila while bass in excess of three poured through a fine meshed net (nylon inches do not grow on such small food nets last longer) to separate the larvae items, and above five inches ignore them. from the liquid mash. The larvae should be rinsed under a faucet to remove all Drosophila larvae can be fed to young mash remnants. If some particles of the fish. We have fed them to newly born medium get into the net it must be

JUNE 1954 139 picked out or broken up by hand. These than the fish will eat. Scavengers should cautions are necessary to prevent fouling be present to eat the left over larvae. aquaria. If several bottles of larvae are Mystery snails, Corydoras, and Hoplo- used, many larvae will work their way sternum are examples, but like all through the net and fall into the wash- scavengers tested, they also feed on live ings. Recovery can be achieved by pour- larvae. A living supply of Drosophila ing the washings through a net. These can be made available to the fish by larvae are usually smaller; they make placing the larvae on a mat of floating better food for small fish. vegetation such as Riccia. The larvae The washed larvae are ready to be fed will live for several days and the fish to the fish. As they do not live under can eat them when hungry. A "raft" made water for more than an hour or so, care by tightly covering a feeding ring with should be taken not to feed more larvae nylon nettings serves the same purpose.

140 AQUARIUM JOURNAL CULTURING FRUIT FLIES (Drosophila), AND OTHER LIVE FISH FOOD

Part 2 (Conclusion)

Saving Brood Stock larly. The adults survive better and lay more eggs if this is done. If the adults Larvae to produce a breeding stock are left in the bottle, they eat most of should be removed from the net and the yeast. The adults need a constant placed on fresh medium ( about every food supply for high egg production. 10 days for D. hydei and D. virilis and Not only is egg production lowered weekly for the two smaller species. ) It while the food supply is insufficient, but is possible to rear adults of D. simulans the adults do not lay as many eggs dur- and D. melanogaster by setting aside the ing the next day or two, even if given desired number of cultures containing sufficient yeast. Moreover, if the num- harvestable larvae. Larval development ber of flies recommended is in a culture of the other two species makes the medi- bottle, they will lay more eggs than the um so liquid that adult transfer is nearly bottle will support of larvae in one day i mpossible. In general it is safer to trans- (D. virilis and D. hydei) or two days fer breeding stock larvae to a fresh (D. simulans and D. melanogaster). medium prior to pupation. The few bottles at room temperature Between 200 and 500 flies ( the larger which do not contain larvae three days number for smaller species) will produce after the eggs have been laid may be sufficient eggs for a half-pint bottle. One considered barren. Failure of the eggs bottle of larvae should produce enough to hatch may be caused from several adults to supply breeding stock for at factors: ( a ) it may be that the brood least one and probably two bottles, fol- stock has been sterilized by too high lowing our instructions for transfering temperatures; ( b ) the culture may be the flies. If one does not intend to regu- moldy; (c) the first eggs laid by the flies larly transfer flies to new bottles a much might not hatch, or ( d ) the culture smaller breeding stock should be used might be all females. One must be care- as the crowded flies will die. ful when transferring the newly emerged adults as the males emerge before the The adult flies cannot produce eggs females. It is possible to set up culture i mmediately after emerging from the bottles in which most of the individuals pupal case. While the adults are becom- are the same sex. ing ripe for egg laying, they must have an adequate supply of yeast for food. Relative Usefulness of Different Adequate food supply may best be given Fruit Flies by transferring the flies to fresh bottles of media every other day. Our experiments were restricted to four species: D. melanogaster, D. simu- As stated above, after eggs first appear lans, D. virilis, and D. hydei. Wildtype the adults should be transferred regu - D. melanogaster and D. simulans are

162 AQUARIUM JOURNAL similar in size, appearance, and value larvae. Occasionally D. hydei individu- as fish food. About two to four cc (4.9 als pupate in the medium. If a culture cc=one teaspoonful) of damp and of this species is harvested after many packed larvae can be raised on banana pupae form, it is difficult to separate mash in an upright half-pint milk bottle. the larvae from the pupae. If the bottle is laid on its side, 3 to 6 cc The duration of the life cycle also af- of larvae can be raised. The amount of fects the relative value of the four spe- surface exposed is one limiting factor. cies. At room temperature, usually be- Still another is the yeast to feed the tween 76 and 82° F., individuals of D. larvae. If brewer's yeast is sprinkled in melanogaster required a minimum of the bottle daily after larval growth is about nine days from egg to egg D. noticed, "upright" production is raised ; simulans 11 days; D. virilis 16 days; and from 5 to 9 cc. Six to 10 cc of larvae are D. hydei 18 days. Cultures with large raised when the bottle is on its side. numbers of larvae grow more slowly than unproductive cultures. Lower tem- More volume can be raised using either peratures will slow down growth; higher D. virilis or D. hydei as both flies and temperatures may sterilize or kill. The larvae are larger. As the larvae need to temperatures cited above are higher than breathe, only as much medium is eaten the 70 to 75° F. recommended by the as the larvae can reach without going geneticists. Thus one generation of D. below the surface for extensive periods. virilis and D. hydei takes about as long They can "hold their breath" for short as two generations of D. simulans and periods, but this does not appreciably D. melanogaster. One must be especial- increase the amount of medium used. ly careful with D. melanogaster and D. If held under for long, they drown as in simulans as the short larval stage may water. D. virilis in a half-pint milk bot- end before the aquarist has noticed that tle produces: ( about) 5 cc upright with- the culture is ready for harvesting. out yeast, 6 cc on side without yeast, 8.5 If small sized larvae are more desired cc upright with yeast, and 9.5 cc on than volume, D. melanogaster and D. side with yeast. simulans, which never reach large size, should be selected. Of course, D. hydei Even more volume can be produced and D. virilis larvae can be fed at a by D. hydei. Not only is the larva large, small size. but also the female lays more eggs per Many aquarists will dislike the odor day, i.e., about 150 vs. 50 for D. virilis. produced by yeast-fed D. hydei cultures. Other factors may apply as D. virilis If the yeast is added only after the cul- production has never exceeded the vol- ture is well advanced, the aroma is not ume of larvae produced by D. hydei so unpleasant. Non-yeast-fed D. hydei from one day's supply of eggs, even when and the other species do not have as females remain in the cultures D. virilis noticeable an odor. for two or three days. D. hydei produc- tion in a half-pint milk bottle: ( ap- Vestigial-Winged Fruit Flies' proximately ) 7 cc on side without yeast, 16 cc upright with yeast, and 18 cc on We have also experimented with ves- side with yeast. ( One sample contained tigial-winged D. melanogaster. First of 24 cc. ) As dried yeast has a detrimental all, the number of larvae produced is effect on young D. hydei larvae, the yeast must be moistened before being Le Bron's Biologicals, 600 N. Kenilworth, Oak Park, supplies vestigial winged flies to aquarists. — placed in the culture bottle with young The Editors. JULY 1954 163 less than in wild type flies. Moreover, worms) in each new culture bottle. The the flies do not transfer well with our culture bottles should be tightly stop- culturing techniques. While wild type pered with cotton except when worms flies produce better after several trans- are being harvested. fers (until old age), vestigial-winged fly populations produce fewer larvae after For harvesting, pour a few drops of each transfer. Because they cannot fly, water in the bottle and then pour the vestigial-winged flies must spend much water into the aquarium. Put the bottle time on the medium. Frequently, they back on the shelf until a new crop of get stuck and die, especially if the larvae microworms develops. have "worked" the medium. A fold of paper towel is recommended for ves- We have found that the technique de- tigial-winged flies to climb on, but this scribed below produces a larger volume interferes with washing the larvae. of microworms for amount of time spent tending the culture. We now use 9 x 15 The only advantage of vestigial- inch enamel pans that are two inches winged flies — they cannot fly from the deep. The culture medium is placed in surface of the water — disappears when the bottom to a depth of about one larvae are fed. A single culture of Droso- eighth inch. Two or three smooth phila probably has the greatest amount wooden strips 13 x 1 x 3/16 inches are of food material immediately before placed in the pan. The small worms pupation. Prior to pupation, growth in- crawl up on the sides of the pan or onto creases the size of the larvae. Much food the strips and can then be scraped off must be expended by the Drosophila for feeding to the fish. The pans must during the metamorphosis which occurs be tightly covered to keep out fruit flies. in the pupa. Moreover, not all individuals survive pupation and most fish will not We have successfully used two culture feed on pupae. Obviously, each fly is of media ( the oatmeal and brewers' yeast less food value than a full grown larva. described by Innes and a combination of Feeding larvae also shortens the culture tortilla mix' and brewers' yeast) in the period. (If desired, winged adults may enamel pans. We mix one part of brew- be heat killed and fed to the fish. ) ers' yeast with about 10 parts of cooked oatmeal or tortilla mix. We have found Microworms the tortilla mix and brewers' yeast me- dium most satisfactory as it does not go Care must be taken to keep micro- bad so fast. It frequently lasts long worms out of Drosophila cultures. They enough so that if worm production do well on banana mash and compete drops, it pays to sprinkle on more yeast. with the fruit flies. For those who dis- like the odors which are given off by the Daphnia usual microworm cultures, the little We have had success raising Daphnia nematodes grow with little odor on a magna indoors roughly following meth- banana mash medium. When the cul- ods outlined by Gordon ( 1950 ). The ture medium is ready for use in raising pessimistic view of Innes (1952) has not Drosophila, it can also be used for micro- been substantiated by our work. Strong worm cultures. A series of new micro- aeration together with frequent and worm culture bottles can be started by putting a few drops of water into a thriving culture and then pouring two We use Tamalina, a product of B. Martinez and Sons Company, San Antonio, Texas, for or three drops (now containing many the tortilla mix.

164 AQUARIUM JOURNAL heavy feeding is necessary for high pro- duced appears to be in direct propor- duction. If sufficient food is added to tion to the volume of water contained enable rapid daphnia production with- in the barrel. The left over banana mash out aeration, the water will become toxic and wash water from one Drosophila to daphnia. As daphnia are susceptible culture bottle will serve to feed a 55 gal- to low oxygen and high carbon dioxide lon daphnia barrel twice daily. If only concentrations, aeration is necessary if one feeding is made, the mash from two large quantities of food are used. If no Drosophila culture bottles may be used. aeration is available, daphnia may be Proportionally smaller amounts should raised by careful feeding. ( The airstones be used for smaller barrels. The indi- may become clogged. If so, soaking in vidual aquarist can experiment with dif- Clorox will clear them.) Both the left- ferent feedings and find out what quan- over solid banana mash and the wash- tities maintain highest daphnia popu- ings from fruit-fly cultures are superb lations. food for daphnia. If yeast is added to the Drosophila cultures, the wash water Containers with a greater length (not is likely to kill daphnia unless used very height) are superior to barrels (we use sparingly. Small daily or twice daily left over specimen barrels). The extra feedings are better than large infrequent length permits a settling region at one feedings. Not only are many daphnia end of the container. Aeration causes a produced, but only waste food material circulation of small particles that are from Drosophila cultures need be used. continually carried about in the water if there is no still area. Not only do the The daphnia should be harvested with daphnia do better without the particles, a fine meshed net. If a coarse meshed but the particles are caught with the net is used only the larger ones will be daphnia. When barrels are used, the harvested. Harvesting only the breeding aeration cannot be turned up to optimum stock lowers production. A more serious for the daphnia as the particles will not difficulty with harvesting only large in- settle out. Keeping the airstone two or dividuals occurs when other animals such three inches off the bottom of the barrel as copepods are in the culture. After a helps the settling. Strong aeration does short time copepods become abundant not appear to harm daphnia if no par- and the daphnia do not prosper unless ticles are in the water. a fine meshed net is used.

When winter eggs (they look like Our Daphnia magna cultures, similar black spots on the body) appear in the to those of Drosophila, have done best culture, adverse conditions are present. at temperatures near 70° F. We had If this is a result of overcrowding, feed poor results when the temperature ex- more daphnia to the fish. If not, partial ceeded 85° F. drainage of bottom sediments is recom- mended by Gordon, 1950. ( These sedi- Although we have had success with ments, and those from vigorous cultures, daphnia in the two-inch deep enamel contain large concentrations of infusoria pans we use to raise microworms, it is which may be used for small fish.) Re- better to use deeper containers as less fill with new water, preferably old air is needed to stir the water. Currently aquarium water; however, if none is we are using wooden barrels for daph- available, tap water may be used. Un- nia. Sizes range from 10 gallons to 55 less more than half of the water has been gallons. The volume of daphnia pro- removed, Austin tap water ( 0.8 parts

JULY 1954 165 per million of residual chlorine) is safe FLETCHER, ALAN M. "Two new and simple as the organic material in the culture culture media for vestigial-winged fruit ffies ( Drosophila)." In: The Aquarium, volume renders chlorine harmless. Daphnia is 22, no. 1, page 18. very susceptible to chlorine and serves as an excellent indicator of whether tap GORDON, MYRON. 1950. "Fishes as laboratory water is dangerous for fish. animals." In: The Care and Breeding of Laboratory Animals, edited by Edmund J. Farris, pages 345-449. When setting up a new culture, it is " best to siphon half of the contents of a HOFFMAN, WILLIAM E. 1924. Winter food for waterbugs in aquaria." In: Bulletin of the vigorous culture into the new container, Brooklyn Entomological Society, volume 19, and fill with water. Also, the addition pages 149-150. of a few pond snails is advantageous as they eat a slimy substance which traps INNES, Wm. T. 1952. "Exotic aquarium fishes." Ed. 14, Innes Publishing Co., Philadelphia. and kills many daphnia. The slime often

develops on the sides of the cultures. PATTERSON, T., and STONE, W. S. "Evo- J. 1952. As we have found that daphnia cultures lution in the genus Drosophila." Macmillan do much better exposed to light, place Co., New York. them in well-lighted locations. More than " POPPEL, JOSEPH. 1951. An improved culture one culture is recommended, for despite medium for flightless flies." In: Aquarium the best care, population fluctuations oc- Highlights, by William T. Innes, page 195. cur. By the use of several cultures, a more constant supply is probable. For SPENCER, WARREN P. 1950. "Drosophila." In: The care and breeding of laboratory animals, maximum production, a large breeding edited by Edmund J. Farris, pages 450-477. stock is essential. Once a high popu- lation density is attained, only the daily STURTEVANT, A. H. 1937. "Culture methods for increase should be used, as harvesting Drosophila." In: Culture Methods for Inver- breeding stock results in decreased fu- tebrate Animals, Comstock Publishing Co., Ithaca, New York, pages 437-446. ture yield.

Caution must be observed in feeding daphnia in unaerated aquaria, as the daphnia may produce sufficient carbon dioxide to kill the fish.

References

BRELAND, OSMOND P. 1949. "The biology and immature stages of the mosquito, Megarhinus septenirionalis Dyar & Knab." In: Annals of the Entomological Society of America, vol- ume 42, pages 38-47.

" BURGER, ELIZABETH. 1937. Culture of Latro- dectus mactans, the black widow spider." In: Culture Methods for Invertebrate Animals, Comstock Publishing Co., Ithaca, New York, pages 244-245.

DOBZHANSKY, TH. 1936. "Collecting, transport- ing, and shipping wild species of Dro- sophila." In: Drosophila Information Service, No. 6, pages 28-29.

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