AXE 3L?» .flfiih‘efi. 5C!“ ‘z-N 555.55 $51321 ”:35 E HGHTNC FISH 53.54 ‘5 E‘WLENQEN" U ‘Fiwsis 53:1? 5219 Elevgt'ae :1? M. 33. M5525? ‘El’fii‘ STAN: 5954953913373’ fix‘xicti‘mefi Cir. GEMS 1‘23?)va ms w HWWNHIWIWTUW 155 L 23 01062 4876 ‘ LIBRARY . Michigan State University PLACED! mum IOXbmmfiflnWMywmcord. TO AVOID FINESthonorbdm-mm DATE DUE DATE DUE DATE DUE MSU “mm-av. Action/Email Oppoflunlty lmthtflon W1 ABSTRACT SIZE DISCRIMINATION IN THE SIAMESE FIGHTING FISH BETTA SPLENDENS By Michael O. Childs The experiments reported here were designed to test the ability of the Siamese fighting fish, Betta splendens Regan, to discriminate size differences of floating discs. These were presented to the fish in pairs, whose members differed in area, and the area of bubbles deposited under each disc was measured. An effort was made to determine whether size discrimination is relative or absolute in this species, and to identify the maximum area above which no discrimination occurs. It should be noted that unlike discrimination studies of the usual type, conditioning was not employed in these experiments. This avoided the situation in which animals learn gg£_to discriminate. Also, a more reliable sample was obtained since all males of this species are natural nest- builders, while the ability to learn varies widely among individuals. A total of twenty-five males was tested. Each was placed in six different test combinations of discs as 2 follows: 100 and 125 cm2, 100 and 175 cm , 100 and 275 cm2, Michael O. Childs 100 and 500 cm2, 450 and 500 cm2, and 400 and 500 cm2. The results were tested both by chi-square analysis and the method of matched observations. No preference was shown for either the left or right position of the discs. Wherever discrimination occurred, the larger disc was preferred. In the combination of 100 and 125 cm2 discs, a statistically significant preference was not demonstrated. This contrasted with the results obtained by Braddock, Braddodk, and Kowalk (1960) who found that a difference as small as 10 cm2 was discriminated when the larger disc did not exceed 89 cm2 in area. This result suggests that a relative, rather than an absolute, value is reacted to by the fish. Discrimination was clearly demonstrated in the disc pairings where the areas were 100 and 175 cm2 and 100 and 275 cm2. None of the pairings in which the larger disc had an area of 500 cm2 gave significant results. Thus, no evidence was obtained that a preference existed on this level under the conditions of the experiments and it appears that pref- 2 erence is lost with the larger disc between 275 and 500 cm in area. SIZE DISCRIMINATION IN THE SIAMESE FIGHTING FISH BETTA SPLENDENS BY Michael O. Childs A THESIS Submitted to Michigan State University in partial fulfillment of the requirements for the degree of MASTER OF SCIENCE Department of Zoology 1964 G 935*?) (8/13le ACKNOWLEDGEMENTS The author wishes to express his sincere thanks to Dr. James C. Braddodk, of the Department of Zoology, for his guidance and constructive criticism during the course of the experimental work and the preparation of this manuscript. Special thanks is also due to Dr. Philip J. Clark, of the Department of Zoology, for his assistance in the statis- tical analysis, Dr. Herman M: Slatis of the Department of Zoology, and Dr. Roland L. Fischer of the Department of Entomology for their help and advice while serving as com- mittee members. The author would also like to thank Mrs. Bernadette McCarthy Henderson, secretary of the Zoology Department, for her many favors; the author's wife Gale for the typing of the thesis; and especially the author's parents for making his education possible. ii INTRODUCTION METHODS AND MATERIALS RESULTS . . DISCUSSION SUMMARY . . BIBLIOGRAPHY TABLE OF CONTENTS iii Page 13 26 32 33 Table Table Table Table Table Table Table 2a. 2b. 2c. 2d. 2e. 2f. Disc Disc Disc Disc Disc Disc Disc Prefere Pairing Pairing Pairing Pairing Pairing Pairing LIST OF TABLES nces . . . . . . . 2 I (100—125 cm ) . 2 II (100—175 cm ) . 2 III (100-275 cm ) 2 IV (100-500 cm ) . 2 V (450-500 cm ) . 2 VI (400-500 cm ) . iv Page 13 l7 19 20 22 23 25 LIST OF FIGURES Page Figure l. Stimulus and Test Aquaria and Lamp . . 9 Figure 2. Positions of Discs in Aquaria . . . . . lO INTRODUCTION B§E£§_splendens Regan is a well known member of the family Anabantidae. There is diversity of reproductive behavior within this family in that some species are bubble-nest builders, while others are oral incubators (Forselius, 1957). All species possess a labyrinth con— sisting of a pair of cavities lined with vascular epithelium and located in the sides of the head above the gills. This constitutes the air breathing mechanism. The fish gulp air from the surface of the water into this labyrinth (Smith, 1945; Forselius, 1957). It is probable that bubble-nest building has evolved from this air breathing habit. In turn oral incubation as exhibited by B, brederi Myers or B, pigga Cuvier and Valenciennes, species which retain the eggs in the buccal cavity until hatching, has probably evolved from bubble-nest building. Since B, splendens is a bubble-nest builder, it is considered to be a comparatively primitive species with regards to reproductive behavior. Although Forselius (1957) reported that female B, splendens build bubble-nests and may even care for the young, the nests are usually constructed by the males. Each nest consists of a group of mucous-coated air bubbles depoSited on the surface of the water. In nature, these are built under leaves or other floating objects (Smith, 1945). The shape of Anabantid nests depends upon species-specific fac- tors, e.g., the availability of nesting material, and the intensity of the nest—building activity (Forselius, 1957). In our laboratory the nests were usually circular, although not necessarily of the same size as the forms under which they were built. They varied in size from 1 cm2 to 110 cm2. The nests chiefly function as territory markers, to attract females to the territories (Braddock and Braddodk, 1959), >and to protect the eggs and young (Forselius, 1957). Even though nests attract females to the territories, in nature the males will not build in the presence of fe- males and will even drive them away from their territories before the nests are complete. In the laboratory, however, males wi;;_construct nests while in visual contact with females. They will also build when alone or in visual con- tact with males (Braddock and Braddock, 1959). After mating, the males deposit the eggs in the nest, among the bubbles. This involves taking the fertilized eggs into the mouth and "blowing" them into the bubble mass. During the period of incubation (30 to 40 hours), males constantly repair the nests by replacing burst bubbles. This replacement of bubbles serves two purposes: to increase the firmness of the nest and to raise the eggs slightly above the surface of the water Where oxygen is more plentiful (Forselius, 1957). After the young have hatched, the males will retrieve and replace any that have fallen out of the nest; after the free—swimming stage has been reached (three or four days), the nest is abandoned. Many studies prior to this have been made with regard to discrimination in animals. Tinbergen (1957) reported studies of the Stickleback fish, Gasterosteus aculeatus Linnaeus, based upon conditioned reflex techniques. These were concerned with colors and posture as abstract qualities. This approach has two weaknesses in that the animals can learn both to discriminate and also Egg_to discriminate. Furthermore, the ability to learn varies greatly among indi- viduals and genetic strains, and this could result in an inadequate sample. The present study takes advantage of the known natural ‘behavior of all males of the species, namely bubble-nest building, thus assuming an adequate population sample. The adequacy of the procedure is probable since it has long been used by fish breeders. Fish naive to the situation are used. Thus one is reasonably sure of obtaining a response that is innate in the animal and not learned. No training period is needed, and one is able to obtain a clear picture of the animal's discriminatory abilities. Bubble—nests were used here because they are relatively easy to measure and are good quantitative indicators of discriminatory activity. Braddodk, Braddock, and Kowalk (1960) reported that when two wax paper discs were floated on the surface of the water, male B, splendens tended to build bubble-nests under them and almost invariably chose the larger disc. It was also found in tests with the larger disc as large as 80 cm2 that the fish were able to discriminate 10 cm2 differences in total area. Forselius (1957) described the nesting and reproductive behavior of this and other Anabantid species. No informa— tion with regard to size discrimination was presented. A general account of the behavior and general ecology of the species was presented by Smith (1937, 1945). No mention was made of discriminatory abilities. Braddock and Braddodk (1959) discussed the development of nesting behavior in young B, splendens, and the nests themselves were studied with regard to shape, size, and duration. This study is an attempt to determine whether the fish discriminate on a relative or absolute basis, i.e., whether they react to actual differences in disc size or to propor- tional differences. It also is an attempt to determine the upper and lower limits above and below which differences are not discriminated and whether or not an optimum range exists within which differences may be more readily distinguished. METHODS AND MATERIALS The fish used in the experiments reported here were mature males purchased at a local pet shop. Originally, they numbered 40, and of these, five were selected for use as stimulus fish and 25 were chosen for use as test fish. Two died during the course of the experiments and were replaced by fish of approximately the same size and color taken from the group of ten held in reserve. The experimental fish were housed in wide-mouthed jars (3.78 L) while the stimulus individuals were kept in stimulus aquaria (Fig. l). The jars were filled to a depth of 12.5 cm at all times. No gravel or plants were used, and waste products and uneaten food were siphoned out weekly. The water used in the jars and aquaria was tap water aged for not less than five days. Distilled water was added to compensate for loss through evaporation. On two occasions, water directly from the tap was added in small amounts to all jars and aquaria to "freshen" the water present. Throughout the course of the experiments, the fish were isolated physically, and with the exception of the time in the test aquaria, were also visually isolated by opaque plastic bands 12.5 cm in height placed around the wide- mouthed jars. During the experiments, the fish were fed finely ground commercial shrimp daily. This was supplemented with live brine shrimp nauplii once a week. The laboratory in which the experiments were conducted is located on the third floor of the Natural Science build- ing on the Michigan State University campus. Three large windows, which face north, were covered with venetian blinds allowing little natural light to enter the laboratory. This was done to keep light conditions as constant as possible. The laboratory itself was lighted with sixteen 40 watt fluorescent bulbs which were controlled by an automatic timer set to provide twelve hours of light during the normal daylight hours and twelve hours of darkness. In addition, gooseneck lamps, each containing a 25 watt bulb, were cen- trally placed on one side of the test aquaria (Fig. 1). These provided illumination 24 hours a day. Their purpose was to provide enough light for the fish to see their nests, since they have a tendency to destroy them in total darkness. Other environmental conditions were kept as nearly con- stant as possible throughout the experiments. The labora- tory was equipped with an air conditioner and steam heat. A fan kept the air circulating with the result that the temperature was quite uniform (80-820 F). The wide-mouthed jars were arranged in rows of nine on a series of three racks facing the windows. The test aquaria were on a long bench adjacent to the windows and separated from the shelves by a narrow aisle. Five test aquaria were used in the experiments. Each had a capacity of 75.6 L and measured 76.2 cm x 33 cm x 35 cm. These were filled to a depth of 17.8 cm, thus contain- ing approximately 40.8 L of water. Fine gravel was placed on the bottoms of the aquaria to a depth of .64 cm and no plants were present. The sides of the aquaria were covered with one layer of .001" white plastic sheeting which was nearly opaque. The tops were covered with one layer of wax paper set in wooden frames. The purpose of the coverings was to prevent any disturbances to the fish while allowing some light to enter the aquaria. A stimulus aquarium was centrally placed against one of the long sides of each test aquarium (Fig. l) in such a man- ner that the experimental male could see the stimulus male thus promoting nesting activity.1 The stimulus aquaria had 1It is known that when two males confront one another and are unable to make any physical contact, a bubble nest will be constructed as a displacement reaction. a capacity of 9.5 L and measured 25 cm x 20.3 cm x 20.3 cm. They were filled to the same level as the test aquaria and contained approximately 8.3 L. Each stimulus aquarium con- tained .64 cm of fine gravel with no plants and was covered with the same material as the test aquaria. The area where the two aquaria met was left open to form a window. The tops of the stimulus aquaria were also covered by one layer of wax paper set in a wooden frame. a’ Fig. l. Stimulus and Test Aquaria and Lamp Discs were suspended in the test aquaria in contact with the water surface. These were constructed of .001" translu- cent laminating vinyl the color of standard white wax paper. Each was supported by two lengths of fine copper wire at- tached to tabs on the disc itself and taped to the glass curvering the top of the aquarium. These discs were located 5.1 cm from the long side of the aquarium nearest the 10 stimulus aquarium and 10.8 cm from a line bisecting the aquarium at right angles to its long axis (Fig. 2). fifij;-lo.8 cm 5.1.cm1"\q1;) LEFT RIGHT Fig. 2. Positions of Discs in Aquaria Six pairings of discs were presented to the fish. In all but two of these, a 100 cm2 disc was used as one member of the pair. The combinations are tabulated below. Twenty- five replications of each pairing were recorded. PAIRINGS I II III IV V VI DISCS (cmz) 100-125 100-175 100-275 100-500 450-500 400-500 NUMBER 25 25 25 25 25 25 A coin was tossed to determine the side of the aquarium on which the smaller disc would be placed in order to avoid possible left-right bias. The individual fish were origi- nally assigned identification numbers and later were assigned random numbers from a table. The random numbers were then 11 arranged in order of increasing magnitude and each fish number was placed with its corresponding random number. The first five fish were then assigned to aquarium "A," the second five to aquarium "B" and so on. This procedure was repeated for each of the six sets of disc pairings. Immediately before being placed in the test situation, the fish were fed ground shrimp. The temperature of the aquaria and bottles was then noted and recorded. Necessary adjustments were made by placing hot or cold wet paper towels around the jars until the temperature in the test aquaria and jars became equal. The fish were then gently netted and placed in the test aquaria. This was done in such a manner as to cause a minimum of disturbance to the fish and usually required no more than fifteen seconds. The tops of the aquaria were then set in place and the gooseneck lamps turned on. Twenty-four hours later the tops were removed and the position and size of the bubble nests recorded. Since the nests could not be examined directly without damaging them, a set of cardboard discs of known area were compared to the nests and matched as closely as possible with regard to total area. The thickness of the 2 Done under the direction of Dr. P. J. Clark. 12 nest was taken into account when measuring since it is obvious that if two nests having the same diameter are of unequal thickness, the thicker nest or the nest having more bubble layers, will have a greater total area than the thinner nest. The temperatures of the aquaria and jars were again taken and any necessary adjustments were made before the fish were returned to the jars. The bubbles were then removed from the discs, thus preparing the aquaria for another set of observations. The tests were conducted daily, Monday through Friday, from April 1 to June 7, 1963. RESULTS The results of these experiments may be analyzed by two different methods. The first, and simplest, is the determination of the number of trials where preference was indicated by depositing more bubbles under one of the two discs offered. This method simply indicates whether the fish discriminated or not, and if so, whether the larger or smaller disc was preferred. If more bubbles were placed under the larger disc, this was recorded as a preference for that disc and vice versa. The results thus recorded are shown in Table 1. When these results (Table l) were subjected to chi-square tests to determine their signifi- cance,3 a definite preference was shown for the larger disc in pairings I, II, III: 100 and 125, 100 and 175, and 100 and 275 cm2, respectively. Table 1. Disc preferences. No. of Fish Showing Preference Large Small No Pref. Pairing I (100-125) 16 5 4 Pairing II (100-175) 17 4 4 Pairing III (100-275) 19 4 2 Pairing IV (100-500) 8 10 7 Pairing V (450-500) 11 10 4 Pairing VI (400-500) 12 8 5 3Done under the direction of Dr. P. J. Clark. 13 14 When the results from these first three pairings were statistically analyzed, significance was found at the 1% level (X2 = 45.06, 6 d.f., P < .01) indicating that the differences of 25, 75, and 175 cm2 between the discs were discriminated and that the larger discs, up to 275 cm2, were preferred. The results of pairings IV, V, and VI, 100 and 500, 450 and 500, and 400 and 500, respectively, were not found to be statistically significant (X2 = 7.15, 6 d.f., .30 < P < .50) indicating that the fish did not respond to differences above the 275 cm2 level of 400, 50, and 100 cm2 respectively, or, if any possibility exists that they could be discrimi- nated, no preference was demonstrated under the conditions of these experiments. As a result of the above analysis, three conclusions may be drawn: 1) differences as small as 25 cm2 were detected when the larger of the two discs was 125 cm2; 2) differences as small as 75 cm2 were detected when the larger of the two discs was 175 cm2; 3) differences as small as 175 cm2 were detected when the larger of the two discs was 275 cm2. When recorded as percentages of the larger disc, these dif- ferences are respectively: 20%, 43%, and 64%. The percentage differences of pairings IV, V, and VI 15 where the larger disc was 500 cm2, are 80%, 10%, and 20%, respectively. Since the 80% difference is greater than any where the smaller discs gave significant results, the re- sults suggest that 500 cm2 is above the absolute size where preference is shown, and that the absolute maximum size for the larger disc, as regards discrimination or preference, must.lie between 275 and 500 cm2 in this particular situation. The second method of analysis treats the bubble area deposited under each disc. This method is more sensitive and more quantitative than the first in that it indicates the extent, as well as the mere presence, of discrimination. The data were tested for significance using the method of matched observations4 (mean of the differences signifi- cantly different from zero) and when the differences in bub- ble area under each disc were found to be significantly dif- ferent from zero, this was taken as an indication that a pref- erence existed, since the fish were actually discriminating. It will be noted that in all of the tables the fish identification numbers are recorded in order of increasing magnitude. This was done in the interest of clarity. In the actual tests, the numbers were randomized as described earlier and did not appear in the order shown. * IDone under the direction of Dr. P. J. Clark. 16 The experiments treated here are those discussed pre— viously; only the method of analysis has been changed. In none of the six types of pairings was there any significant difference between the means of the total bubble areas blown under the larger disc in the right as compared with the left position. Thus, no preference was shown for either position. The first pairing matched a 100 cm2 disc with a 125 cm2 disc. The results are presented in Table 2a. Table 2 indicates the position of the smaller disc as regards right or left (see also Fig. 2) and the total bubble area ex- pressed in square centimeters under each disc. The raw data would seem to indicate that a preference was shown for the larger disc. When subjected to statis- tical analysis (mean of differences significantly different from zero), however, the results were significant only at the 10% level. Thus there is no clear evidence that the fish were able to discriminate between the two discs of this pairing and the conclusion drawn from the raw data is un- warranted. It will be noted that according to the first method of analysis (chi-square), the results of this pairing were significant at the 1% (.01) level. This discrepancy l7 . in statistical analysis will be treated in the "Discussion" portion of this thesis. 100 and 125 cm2 discs and bubble areas deposited under each. Table 2a. Bubble Area in cm2 Fish Position of Differences in No. Small Disc 100 cm2 125 cm2 Bubble Area* A500 Right 20 21 + 1 A503 Right 10 30 +20 A505 Left 20 40 +20 A506 Right 20 20 O A507 Right 10 2 -8 A510 Right 5 5 0 A511 Right 2 20 +18 A512 Left 40 50 +10 A514 Left 40 20 -20 A516 Right 4 10 + 6 A518 Left 0 0 O A519 Left 10 50 +40 A520 Left 36 4 -32 A521 Right 5 20 +15 A522 Right 7 40 +33 A523 Left 20 30 +10 A524 Right 10 40 +30 A525 Right 30 10 -20 A526 Left! 20 40 +20 A527 Left 0 14 +14 A530 Right 0 30 +30 A531 Right 1 2 + 1 A535 Right 30 0 -30 A537 Right 10 40 +30 A539, Left 0 0 0 Totals 350 537 +188 * + indicates large disc favored - indicates small disc favored 18 The original intention was to increase the size of the larger disc by 25 cm2 at each consecutive pairing while maintaining the size of the smaller one at 100 cm2. Since no significant difference was indicated in the case of the first pairing (100 cm2 and 125 cmz), to save time, the larger disc was then enlarged by 50 cm2. Thus, the second pairing consisted of discs 100 and 175 cm2 in area. The results of this pairing are presented in Table 2b. Statistical analysis of these results indicates that they are significant as suggested by the raw data. Thus, using the method of "matched observations," the observed "t" value of 6.23 is significant at the 1% (.01) level. This method of analysis supports the conclusions reached by the first-tried method (chi-square), namely that the fish were able to discriminate the 75 cm2 difference in the two discs of this pairing and showed a preference for the larger disc. The third pairing consisted of discs with areas of 100 cm2 and 275 cm2. The first two pairings (100 cm2 with 125 cm2 and 175 cm2 respectively) had indicated that the smallest difference that the fish could detect lay between 25 cm2 and 75 cm2 when the smaller disc had an area of 100 cm2. It was now necessary to identify the largest detectable difference. The results of this pairing are presented in Table 2c. 19 100 and 175 cm2 discs and bubble areas deposited under each. Table 2b. Bubble Area in cm2 Fish Position of Differences in No. Small Disc 2 2 Bubble Area* 100 cm ‘ 125 cm A500 Left 0 85 +85 A503 Right 30 40 +10 A505 Left 20 0 -20 A506 Right 10 20 +10 A507 Left 3 60 +57 A510 Left 3 5 +2 A511 Right 20 0 -20 A512 Left 0 40 +40 A514 Left 0 3 +3 A516 Left 0 0 O A518 Right 20 40 +20 A519 Right 40 30 -10 A520 Right 0 50 +50 A521 Left 0 9 +9 A522 Left 30 50 +20 A523 Left 0 0 0 A524 Left 5 7 +2 A525 Left 0 0 0 A526 Right 5 10 +5 A527 Right 3 10 +7 A530 Right 3 30 +27 A531 Left 0 40 +40 A535 Left 20 10 -10 A537 Left 0 70 +70 A539 Right 0 0 0 Totals 212 609 +397 * + indicates large disc favored - indicates small disc favored 20 100 and 275 cm2 discs and bubble areas deposited under each. Table 2c. Bubble Area in cm Fish Position of Differences in No. Small Disc 100 cm2 275 cm2 Bubble Area* A500 Right 0 60 +60 A503 Left 0 10 +10 A505 Right 0 20 +20 A506 Left 0 0 0 A507 Left 10 40 +30 A510 Left 20 40 +20 A511 Left 40 30 ~10 A512 Left 30 90 +60 A514 Left 10 20 +10 A516 Right 0 30 +30 A518 Left 10 30 +20 A519 Left 40 50 +10 A520 Left 0 10 +10 A521 Right 10 20 +10 A522 Left 20 0 ~20 A523 Left 0 0 0 A524 Left 5 30 +25 A525 Right 10 30 +20 A526 Left 20 40 +20 A527 Right 10 0 ~10 A530 Right 10 40 +30 A531 Right 0 5 +5 A535 Right 10 30 +20 A537 Left 50 110 +60 A539 Right 30 10 ~20 Totals 335 745 +410 * + indicates large disc favored ~ indicates small disc favored When the results obtained from this pairing were sub- jected to statistical analysis (method of matched observa- tions), they were found to be significantly different at 21 the 1% (.01) level. This agrees with the indication sug- gested by the first method of analysis (chi-square) and indicates the fish are able to distinguish differences of 175 cm2 between the two discs and still prefer the larger disc, although there is no evidence that this lies near the upper limit of the fish's discriminatory abilities where the smaller disc has an area of 100 cm2. Discs of 100 cm2 and 500 cm2 constituted the fourth pairing. The results are presented in Table 2d. It will be noted that another large increment has been added when comparison is made with the 275 cm2 larger disc of the previous pairing. This represents an attempt to determine the vicinity of the "upper limit" of discrimination with as few unnecessary intermediate pairings as possible. As suggested by the raw data, the results from this pairing did not prove to be statistically significant. This agrees with the first method of analysis (chi-square) and is evidence that the fish did not discriminate the 400 cm2 between the two discs of this pairing. The fifth pairing matched discs with areas of 450 cm2 and 500 cm2. This was done to determine whether or not the fish were able to discriminate a 50 cm2 difference where 2 both discs were considerably larger than 100 cm . This was 22 100 and 500 cm2 discs and bubble areas deposited under each. Table 2d. Bubble Area in cm2 Fish Position of Differences in No. Small Disc 100 cm2 500 cm2 Bubble Area* A500 Right 10 30 +20 A503 Left 0 30 +30 A505 Left 0 0 0 A506 Left 20 40 +20 A507 Left 10 40 +30 A510 Left 30 50 +20 A511 Left 40 30 ~10 A512 Right 0 40 +40 A514 Left 30 30 0 A516 Left 40 0 ~40 A518 Left 30 10 ~20 A519 Right 0 10 +10 A520 Left 27 0 ~27 A521 Left 50 20 ~30 A522 Left 20 20 0 A523 Left 20 40 +20 A524 Left 10 5 -5 A525 Left 10 5 ~5 A526 Left 20 20 0 A527 Left 30 40 +10 A530 Left 0 0 0 A531 Right 0 0 0 A535 Left 20 5 ~15 A537 Left 30 20 ~10 A539 Right 20 0 ~20 * + indicates large disc favored - indicates small disc favored Totals 467 485 +18 the approximate difference discriminated where the smaller disc had an area of 100 cm2. tained from this test, If similar results were ob~ it would seem to indicate that the 23 differences recognized are absolute rather than propor- tional within the size range where discrimination occurs at all. The results are presented in Table 2e. Table 2e. 450 and 500 cm2 discs and bubble areas deposited under each. Fish Position of Bubble Area in cm Differences in No. Small Disc 450 cm2 500 cm2 Bubble Area* A500 Left 40 20 ~20 A503 Left 30 20 ~10 A505 Left 0 O 0 A506 Right 0 20 +20 A507 Left 2 30 +28 A510 Right 40 0 ~40 A511 Left - 0 30 +30 A512 Left 70 40 ~30 A514 Left 0 0 0 A516 Right 70 0 ~70 A518 Right 50 30 ~20 A519 Left 10 30 +20 A520 Left 30 0 ~30 A521 Right 30 40 +10 A522 Left 20 0 ~20 A523 Left 10 20 +10 A524 Left 30 10 ~20 A525 Right 20 40 +20 A526 Right 0 0 0 A527 Right 0 0 O A530 Left 10 20 +10 A531 Left 10 40 +30 A535 Right 0 10 +10 A537 Right 20 50 +30 A539 Left 5 3 ~2 Totals 497 453 ~44 * + indicates large disc favored ~ indicates small disc favored 24 Statistical analysis (method of matched observations) failed to reveal significance at any level, and this was in agreement with the method of analysis previously used (chi— square). Thus the fish were apparently unable to discrimi- nate the 50 cm2 difference in the two discs of this pairing. This suggests that once a certain critical area (between 275 and 500 cm2) is exceeded, an absolute difference easily discriminated where both areas are smaller is no longer recognized. Discs 400 cm2 and 500 cm2 in area constituted the last pairing. This pairing was made in order to determine whether or not the fish would discriminate a difference of 100 cm2 in the two discs. This absolute difference exceeded that (75 cm2) discriminated where the discs were smaller (100-175 cm2) and the proportional difference was the same as the proportional difference of the first set of discs (100-125 cm2). The results are presented in Table 2f. The results of this pairing when analyzed statistically (method of matdhed observations) showed no significance which again was in accord with the first method of analysis (chi-square) and indicates that the fish did not discrimi— nate between the two discs. This result suggests that the 25 absolute area above which neither absolute nor proportional differences are recognized lies below 500 cm2. Table 2f. 400 and 500 cm2 discs and bubble areas deposited under each. Bubble Area in cm2 Fish Position of Differences in No. Small Disc 400 cm2 500 cm2 Bubble Area* A500 Left 30 50 +20 A503 Right 14 0 ~14 A505 Right 5 20 +15 A506 Left 0 20 +20 A507 Left 5 20 +15 A510 Right 20 0 ~20 A511 Right 0 0 0 A512 Right 20 50 +30 A514 Right 0 30 +30 A516 Left 30 14 ~16 A518 Left 20 0 ~20 A519 Left 0 30 +30 A520 Right 10 30 +20 A521 Right 0 0 0 A522 Left 5 10 +5 A523 Right 20 10 ~10 A524 Right 50 30 ~20 A525 Right 0 O 0 A526 Left 0 O 0 A527 Left 14 0 ~14 A530 Right 5 20 +15 A531 Right 0 14 +14 A535 Right 0 0 0 A537 Left 10 40 +30 A539 Right 5 0 ~5 Totals 263 388 +125 * + indicates large disc favored - indicates small disc favored DISCUSSION The original purpose of this study was to determine whether Betta splendens recognizes actual or proportional differences when discriminating between two discs of un- equal area. It was hoped to determine the range within which the best discrimination occurs and where the upper limit of the fish's discriminatory abilities lies. At this point reference should be made to the work of Braddock, Braddock, and Kowalk (1960), This particular study was the first attempt to measure the extent of size discrimination in B, splendens. The work was conducted in the same labora- tory and under the same conditions as was the present study. It was found that male B, splendens can distinguish between discs of various areas between 80 cm2 and 20 cm2 provided the difference between them is 10 cm2 or greater. Thus the practical lower limits of discrimination have been estab- lished since a discrimination of 10 cm2 was found to exist at areas which would be too small (20 cm2) to be practical as nests. No work was done to determine whether the dif- ferences distinguished are absolute or relative- In this study, a disc of 100 cm2 (larger than Kowalk's largest) was matched with a disc having an area of 125 cm2. 26 27 By one method of statistical analysis the 25 cm2 difference was not found to be distinguishable to the fish. This dif- ference is over twice as great as that distinguished in the work of Braddock, _£_§1, Thus if the fish were discrimi- nating on an absolute basis, the 25 cm2 difference should have been discriminated. Since it was not, the conclusion may be drawn that the fish discriminate on a relative or proportional basis. As further evidence, a 175 cm2 disc was matched with the 100 cm2 disc and this proved to be distinguishable to the fish as evidenced by statistical analysis using the method of "matched observations." Here the proportional difference in the two discs was large enough for the fish to discriminate. A matter that requires explanation of this point is the fact that the first method of statistical analysis (chi~ square) showed the results of the first pairing of discs to be significant, i.e., indicated that the fish discriminated the 25 cm2 difference between their two areas. When this same pairing was analyzed by the method of "matched obser- vations," the results were not found to be significant, implying that the fish did not discriminate the 25 cm2 difference. This discrepancy may be explained as follows: in the chi-square analysis, a fish was recorded as shoWing 28 a preference if it deposited more bubbles under either of the discs, even if the difference was as small as one cm2. Thus, a case where 50 cm2 was deposited under the smaller disc and 51 cm2 under the larger was recorded as a prefer- ence for the larger disc. This affected the results of the statistical analysis as shown. The method of "matched observations" was preferred in this case since it is a more sensitive test than the "chi-square." However, it should be pointed out that the results §£§_consistent in that the larger disc was preferred, however, the "matched observa— tions" method of analysis did not show this preference to be significant. The analysis by "matched observations" gave an observed t value of 1.99 for the first set of discs. With 24 degrees of freedom this was below the value for significance (2.064) at the 5% level. For this particular test situation it must be concluded that the evidence is inadequate to show whether the fish were able to discriminate the 25 cm2 difference between the two discs. A third pairing was assembled in order to provide in- formation relevant to the upper limit of the fishes' dis- criminatory abilities. Discs with areas of 100 and 275 cm 2 were used, and, since the 175 cm difference was 29 discriminated, it was concluded that the upper limit must be greater than 275 cm2. The size of the larger disc was then increased to 500 cm2 and even a difference as large as 400 cm2 gave no evi- dence of preference by the fish (Table 2d). Approximately equal amounts of bubbles were deposited under each disc, and analysis of the results showed no statistical signifi- cance. We may thus conclude that the fish probably do not discriminate between 100 and 500 cm2 discs, and it seems that the latter size exceeds the upper limit of discrimi- nation. It will be recalled that an increase in the size of'the larger disc was made from 275 cm2 to 500 cm2 without testing areas intermediate in size. Thus in order to determine the precise upper limit of discrimination, a series of discs intermediate between 275 and 500 cm2 would have to be tested. On the basis of the evidence presented here we may merely conclude that the upper limit lies somewhere between 275 and 500 cm2. The fact that even a difference as large as 400 cm2 was not discriminated when the larger disc had an area of 500 cm2 can possibly be explained on the basis that this larger disc was not recognized as such. In an aquarium of the size used in these experiments, a disc with this area 30 extends almost across the full width and leaves only narrow margins. The fish then, may have reacted to it as a "roof" rather than a floating disc. In this case, use of a larger aquarium might produce different results than those reported here. Doubt is cast upon the above hypothesis by the fact that equal amounts of bubbles were deposited under the 500 cm2 disc and those of other areas paired with it. This seems to indicate that the fish actually reacted to the larger disc as a suitable nest site. Perhaps its large size was simply recognized as "large enough" rather than "larger than." Thus, since the discs paired with it were also "large enough," no preference was shown. The actual method of discrimination used by the fish is not known although it is very likely that it is either visual and/or tactile. Evidence for visual discrimination lies in the fact that nest building individuals attend to their nests inter- mittently and frequently return to them from a distance. They then go to work without prolonged contact with the nest or the object under which it is being built. Tactile con- tact with bubbles already deposited may accelerate the process. 31 It seems probable that in nature B, splendens has a narrow choice of plant species with leaves providing suit- able nest sites. Choice of leaves too small to contain the entire nest might cause the loss of all, or part of, a brood. Choice of a leaf that might be too large would be less critical, perhaps such leaves do not exist in the natural environment-of the species. Thus, ability to select the larger leaf when the critical lower size limit was in- volved, would have adaptive significance. S UMMARY 1. Male Siamese fighting fish were presented with pairs of floating plastic discs with different areas under which they deposited nest bubbles. 2. The bubble area in cmzdeposited under each of the two discs of each pairing was recorded, and, where one size was consistently preferred to another, it was taken as evidence that the fish discriminated between the two discs. 3. Where discrimination occurred the larger disc was the one preferred. 4. Differences as large as 25 cm2 were not discriminated when the larger disc had an area of 125 cm2. 5. Differences of 75 cm2 were discriminated when the larger disc had an area of 175 cm2. 6. Differences of 175 cm2 were discriminated when the larger disc had an area of 275 cm2. 7. In cases where the area of the larger disc was 500 cm2, differences as great as 400 cm2 were not discriminated. It appears that under the conditions of these experiments, 500 cm2 is above the size limit where preference is shOwn. 8. The fish utilize proportional rather than absolute dif- ferences when discriminating between two discs of unequal size. 32 BI BLIOGRAPHY Braddock, J. C., and Z. I. Braddodk. 1959. The develop- ment of nesting behavior in the Siamese fighting fish, Betta splendens. Animal Behavior 1,222~232. Braddodk, J. C., Z. I. Braddodk, and G. Kowalk. 1960. Size discrimination in the Siamese fighting fish, Betta splendens. Bull. of the Ecological Society of America _4_]_._:82. Forselius, S. 1957. Studies of Anabantid fishes. Zoologiska Bidrag Fran Uppsala, Almqvist and Wiksells Boktryckeri Aktiebolag, Uppsala. pp. 93-597. Smith, H. M. 1937. The fighting fish of Siam. Nat. Hist. l;2:265~271. . 1945. The fresh water fishes of Siam or Thailand. U.S. Nat. Mus. Bull. 188:456~461. Tinbergen, N. 1958. The Study 9§_Instinct. University Press, Oxford. 33 R0055 [155 055W nICHIan STATE UNIV. LIBRARIES 5|IWWWIm)”ll)WW“)“WI”“WI 31293010624876