III/E-2
STUDIES ON THE RATE OF FOOD PASSAGE IN THE INTESTINE OF LABEO ROHITA (HAM.), CIRRHINA MRIGALA (HAM.), AND CATLA CATLA (HAM.)

by

S.S. RANADE and H.G. KEWALRAMANI
Taraporevala Marine Biological Research Station,
Taraporevala Aquarium
Bombay-2, India

Abstract

‘Major carps’ of India, comprising mainly Labeo rohita, Cirrhina mrigala, and Catla catla, constitute a group of most important fresh-water food fishes cultured on a large scale in the country. One of the problems in their culture is the determination of suitable diets to obtain maximum growth rates. Their natural food has been determined by several workers by examining gut contents and it has been shown that they are capable of ingesting a wide range of food materials. To what extent these are utilized is, however, not known. In the present study the rate of passage of various items of food normally fed upon in nature, in the intestine of these species, was investigated. Their amyloclastic and proteolytic activities were also determined so as to explore the relationship between rate of passage of food and efficiency of digestion. This investigation has provided an index of the extent of utilization of different food items by ‘major carps’.

ETUDE SUR LE TAUX DE PASSAGE DES ALIMENTS DANS LES INTESTINS DE LABEO ROHITA (HAM.), CIRRHINA MRIGALA (HAM.) ET CATLA CATLA (HAM.)

Résumé

Les “major” carpes indiennes, notamment Labeo rohita, Cirrhina mrigala et Catla catla, constituent un très important groupe de poissons d'eau douce élevés sur une grande échelle dans le pays. L'un des problèmes que pose leur élevage consiste à déterminer le régime alimentaire approprié pour obtenir le taux de croissance maximum. Leur alimentation naturelle est connue par les travaux de plusieurs chercheurs qui ont examiné le contenu des intestins, et il a été constaté que ces espèces sont capables d'ingérer les aliments les plus divers. Toutefois, on ne sait pas dans quelle mesure ces aliments sont assimilés. Des recherches ont été faits sur le taux de passage des différents aliments naturels dans les intestins de poissons des espèces considérées. Leurs fonctions amyloclastiques et protéolytiques ont également été étudiées afin d'établir le rapport entre le taux de passage des aliments et l'efficacité de la digestion. A la suite de ces études il a été établi un index du degré d'utilisation des différents aliments par les “major” carpes indiennes.

ESTUDIOS SOBRE EL PASO DE ALIMENTOS EN EL INTESTINO DE LABEO ROHITA (HAM.) CIRRHINA MRYGALA (HAM.) Y CATLA CATLA (HAM.)

Extracto

Las carpas principales de la India, formadas principalmente por Labeo rohita, Cirrhina mrygala y Catla catla, constituyen un grupo de peces comestibles de agua dulce muy importante, cultivado en gran escala en el país. Uno de los problemas de su cultivo es la determinación de la alimentación adecuada para obtener los máximos coeficientes de crecimiento. Su alimentación natural ha sido determinada por varios especialistas examinando los contenidos intestinales, y se ha comprobado que son capaces de ingerir una gran variedad de alimentos. En el presente estudio se ha investigado la cantidad de los varios tipos de alimentos proporcionados normalmente en la naturaleza que pasa al intestino de estas especies. Se han determinado también sus actividades amiloclásicas y proteolíticas a fin de explorar la relación entre la cantidad de alimentos que pasan al intestino y la eficacia de la digestión. Esta investigación ha proporcionado un índice del grado de utilización de los distintos tipos de alimentos por estas carpas principales.

1 INTRODUCTION

In India, three species of carps, Catla catla, Labeo rohita, and Cirrhina mrigala, belonging to the group popularly known as ‘major carps’, are of prime importance in fish culture. They are likely to be cultured on a much larger scale in the future, now that breeding can be induced by hormone injection. The great potentialities of major carp culture suggest that they should be subjected to the same kinds of studies as farm animals; to find, for example, which of their natural foods are utilized most efficiently for flesh production.

In fishes, although a considerable amount of work has been done on the anatomy and histology of the alimentary canal, comparatively little is known about the physiological aspects of digestion. The literature bearing on the enzyme equipment of different species has been reviewed by Yung (1899), Sullivan (1907), Yonge (1937), Vonk (1937, 1941), and Barrington (1942; 1957). However, the study of the rate of digestion “in vivo” seems to be a new approach to the nutritional physiology of fishes. Moreover, the previous observations on the rate of passage of food in fishes are on species having a stomach (Weinland, 1901; van Slyke and White, 1911; Dobreff, 1927; Karpevitch and Bokoff, 1937; Menon and Kewalramani, 1959; Hirao et al., 1960; Molnar and Tolg, 1962). Stomachless species have as yet been little studied, with the exception of work by Maltzan (1935) and Sokolcv and Chvaliova (1936).

Studies were therefore undertaken on the rate of passage of various foods, normally eaten in nature, through the intestine of Labeo rohita, Cirrhina mrigala and Catla catla. The amyloclastic and proteolytic activities of intestinal mucosa and the associated digestive glands were also to be determined, to explore a relationship, if any, between the rate of food passage and the efficiency of digestion. Previous work on the anatomy and histology of the alimentary canal of these species (Sarbahi, 1940; Ahsan-ul-Islam, 1949; Kapoor, 1953), had shown the absence of a stomach and presence of an unorganized and intrahepatic pancreas. The examination of their gut contents (Mookerjee, 1945; Mookerjee and Ghosh, 1945; Mookerjee and Das, 1945; Chacko and Kurian, 1950–51; Begum, 1959; George, 1963) had shown that they are capable of ingesting a variety of plankters and higher aquatic plants. To what extent these food items could be digested was, however, not known.

2 MATERIALS

Live specimens of Labeo rohita, Cirrhina mrigala, and Catla catla, were obtained from a fresh-water pond near Bombay and transported to large aquarium tanks (3.3 × 1.3 × 1.3 m) with fresh-water circulation at the Taraporevala Aquarium, attached to the Taraporevala Marine Biological Research Station, Bombay. Specimens of about 45 cm length, nearly one year old, were selected both for the study of rate of food passage and for determining the activity of digestive enzymes.

Live plankters, required as food for the fishes, were collected from stagnant pools around Bombay, with as far as possible, near monoculture blooms of the respective plankters.

3 METHODS

3.1 Determination of rate of food passage

Fishes were first kept in a large aquarium tank (3.3 × 1.3 × 1.3 m) for four or five days to acclimatize them to aquarium conditions. They were then transferred to a large experimental tank (3.0 × 0.7 × 0.7 m) containing filtered fresh water in circulation, and starved there for a week to ensure the complete evacuation of the intestine.

For the determination of the rate of passage of different food items, starved fishes were force-fed on nearly equal quantities of each food item separately, and the passage and condition of the food was examined after a lapse of different intervals. Fishes were anaesthetized before force-feeding, so as to be able to introduce a known quantity of each food item into the intestine. Preliminary experiments with M.S.222 (tricaine methanesulphonate, Sandoz) for anaesthetizing fishes for handling and force-feeding, showed that 50 ml of 1 percent M.S.222 solution in 25 litres of water was necessary. A known quantity of a food item was then forced directly into the intestinal bulb of the fish by a soft plastic tube attached to a 60 ml syringe. The whole process of anaesthetization and feeding was over within about five minutes, and recovery was immediate with no after-effects. In the case of higher aquatic plants, a weighed quantity was placed by forceps near the gullet and pushed into the intestinal bulb by a soft plastic tube. Chopped prawn was force-fed in a similar manner. Vomiting seldom occurred; if it did, then such specimens were rejected. The fed fish were returned to the experimental tank. The temperature of the tank water varied from 28°C to 30°C, throughout the investigation. After the requisite period, the fish was killed by pithing, and the intestine was immediately ligatured at various places so as to be able to locate exactly the part where the food was lying. The distance traversed by the food was noted, and the recovered food was also examined microscopically to note the changes in its condition, if any, due to the action of the digestive enzymes of the fish.

Table I

The retention of various food items in the intestine of Labeo rohita, Cirrhina mrigala and Catla catla, of about 45 cm length and nearly one year old.*

* Three readings were taken in each experiment

3.2 Concentrations of amyloclastic and proteolytic enzymes in the three species

For the preparation of enzyme extracts of intestinal mucosa and liver, the freshly killed one-year old specimens of nearly 45 cm length were used. For extracting amylase and lipase, iced distilled water was used as a medium, whereas alcohol was used for the extraction of proteases, in the proportion of 10 parts by volume of extracting solvent to one part by weight of mucosa. The amyloclastic and proteolytic activities of the extracts were studied using one percent starch solution and 10 percent alkaline casein solution, respectively, at a temperature of 37°C, and at a pH 7.0 for amylases and pH 8.5 for proteases. The possibility of the presence of cellulase was investigated, using one percent carboxymethyl cellulose solution, as a substrate.

4 OBSERVATIONS

The times taken by different food items to reach the hindmost end of the intestine are given in Table I and the amyloclastic and proteolytic activities per unit weight of mucosa of different parts of intestine and associated digestive glands are given in Table II. The analysis of the data shows that in individuals of nearly the same size of the same species, and under similar experimental conditions, the rate of passage of different food items varies considerably with the kind of food. The algae, Microcystis, Anabaena, Scendesmus and Chlorella, were retained in the intestine for a longer time than the filamentous algae, Spirogyra and Ulothrix. It is interesting to note that the former were always recovered in the hindmost part of the intestine, in a totally unchanged condition, while the latter were appreciably digested; their cell walls were broken and the cell contents were not seen, probably being consumed due to the action of the digestive enzymes.

Higher aquatic plants were also retained for varying periods in the intestine of each species, and were digested to a different extent. Anacharis, Hydrilla, Ottelia, and Vallisneria, were retained for a considerably longer time in the intestine of each species, than Lemna. The major portion of all these higher aquatic plants consists of thin-walled chlorenchymatous cells. Most of these cells were found to be acted upon by the digestive enzymes. Lemna, which is very thin and the softest among all the plants studied, was digested almost completely. But in the case of other plants, which are comparatively coarse, some cells were always recovered even in the hindmost part of the intestine in an undigested condition, probably because the cell walls were covered with some material not amenable to the digestive process.

Since the major portion of the food of these species consists of plant-matter, the possibility of a cellulase being secreted by the alimentary canal or digestive glands was investigated, but no cellulase was found.

Table II

Time (in hr) required to digest 5 ml starch solution (1 percent) by extracts from equal weights of intestinal bulb, intestine and liver tissue by Indian major carps

Table III

Weight (in mg) of amino acids released from 10 percent casein solution in 24 hours by enzyme extracts from various parts of the digestive tracts of Indian major carps

Microcystis when given fresh passed out undigested, but decayed Microcystis was digested to a considerable extent, since most of the cells were found acted upon by the digestive enzymes, and this passed at a faster rate than fresh Microcystis. The digestion of Anacharis was also more, as well as faster, when it was given in a decayed condition. The decomposition probably breaks down some resistant material in the cell wall and so exposes the cell-contents to the action of the digestive enzymes. In the absence of cellulase, the breaking of the cell walls of fresh as well as decayed plant matter may possibly be due to the bacterial action.

Crustacean zoo-plankters Daphnia, Cyclops, Diaptomus, and Cypris, were digested rapidly except for the outer thin chitinous shell, but chopped prawn and mosquito larvae, with a more thickly chitinized exoskeleton, were retained for a longer time in the intestine. The chitinous exoskeleton of all these animal foods was always recovered, even in the hindmost part of the intestine, in a totally unchanged condition, probably due to lack of chitinase. This is supported by the observation (Table I) that when prawn was given after removing the exoskeleton, it was retained for a shorter time in the intestine.

Comparing the rate of food passage in these three species, it was found that in individuals of about 45 cm length, under similar experimental conditions, the passage of partially digestible plant matter was fastest in Cirrhina mrigala, slower in Labeo rohita, and slowest in Catla catla. The totally indigestible plant matter (Table I), however, passed at a more or less similar rate in the three species. Thus, the indigestible plant matter was retained for the longest time in Cirrhina mrigala which has the longest intestine, and for the least time in Catla catla, which has the shortest intestine. Thus under similar experimental conditions, the passage of a given food item would be expected to be directly proportional to the length of the intestine. This would mean that the food passage should be slowest in Cirrhina mrigala. However, the results show that the passage of digestible plant matter is fastest in Cirrhina mrigala. Obviously some other factor, other than the length of the intestine, is affecting the food passage. This factor is probably the difference in the concentration of amyloclastic enzymes in these species.

Tables II and III give the concentration of digestive enzymes per unit weight of mucosa of different parts of intestine and of digestive glands. This shows that the concentration of amyloclastic enzymes in the intestinal mucosa and liver would be maximum in Cirrhina mrigala, since these enzyme extracts digest 5 ml of 1 percent starch solution in the shortest time. In Labeo rohita and Cirrhina mrigala, although the intestinal bulb and liver show nearly the same concentration of amylase, the rest of the intestine is considerably more amyloclastic in Labeo rohita than in Catla catla, so that the overall amylase secreting capacity would be more in Labeo rohita than in Catla catla. These results correlate well with the data on the rate of passage of digestible plant matter.

The rate of passage of several animal foods was found to be fastest in Catla catla, slower in Cirrhina mrigala, and slowest in Labeo rohita (Table I). It can be seen from Table II that although the bile of Catla catla shows less proteolysis, its intestinal mucosa and liver are much more proteolytic than that of Labeo rohita and Cirrhina mrigala, so that the overall concentration of proteolytic enzymes would be maximum in Catla catla. The intestinal bulb and anterior half of intestine of Labeo rohita have a very weak concentration of proteolytic enzymes. Therefore, although the hind half of intestine and especially the liver show more proteolysis, the overall concentration of proteolytic enzymes is likely to be less in Labeo rohita than in Cirrhina mrigala. These results, therefore, correlate well with the data on the rate of passage of animal foods, described earlier. It is of great interest that these differences in the passage of animal foods are not marked, probably because the differences in the concentration of proteolytic enzymes are slight. This therefore furnishes another clear-cut instance of correlation between rate of food passage and efficiency of digestion.

5 DISCUSSION

When the data on rate of passage of different food items are cross-checked with the data on the concentration of different digestive enzymes, it is seen that the rate of food passage is related to the efficiency of digestion. Thus, food items biochemically so constituted as to be more amenable to the digestive enzymes of a particular species, passed out at a faster rate than others. This is probably correlated with the absence of cellulase, chitinase and other higher carbohydrases. Thus coarse plants and animal foods with thick chitinous exoskeleton, were retained for a longer time than others. Moreover, while the totally indigestible algae passed at a more or less similar rate in the intestine, the passage of digestible plant matter was fastest in Cirrhina mrigala and that of animal foods fastest in Catla catla, which showed maximum amyloclastic and proteolytic activities, respectively.

The relationship between the rate of food passage and the concentration of the digestive enzymes is of great significance in fish biology. It indicates that food items which pass rapidly through the intestine are more efficiently used, since rapid food passage is associated with high digestibility due to high concentration of the requisite enzymes. Moreover, a rapid food passage may also mean a similar increase in the rate of feeding.

A relationship of this kind has so far been sought only in farm mammals. Thus Ewing and Smith (1917) found that in the steer, a rapid passage was associated with high digestibility. Balch (1951) found that the ability to digest crude protein and ether extract fractions of hay and mangolds was greatest in cows that retained hay for the shortest time in the reticulo-rumen. He, however, found that variations in the digestibility of the crude fibre were not related to the rate of passage. Blaxter et al. (1956) obtained results supporting those of earlier workers. They found that those diets which are intrinsically highly digestible pass more rapidly through the guts of sheep.

Considering now the results of the present work in relation to ‘major carp’ culture, it may be suggested that the digestible plant matter would be utilized with maximum intensity by Cirrhina mrigala, less by Labeo rohita, and least by Catla catla, while the animal foods would be utilized most intensively by Catla catla, less by Cirrhina mrigala, and least by Labeo rohita. The difference in efficiency of utilization would, however, be considerably more marked in the case of plant matter than in the case of animal foods.

Several workers have determined the food of these species on the basis of examination of gut contents (Mookerjee, 1945; Mookerjee and Ghosh, 1945; Mookerjee and Das, 1945; Chacko and Kurian 1950–51; Begum, 1959; George, 1963). All these authors have shown that although these species ingest a variety of phytoplankters, zooplankters and higher aquatic plants, Catla catla prefers to feed on plankton, mainly zooplankton, while Labeo rohita and Cirrhina mrigala prefer to feed on plant matter, including decaying vegetation. The present investigation confirms that Catla catla can utilize animal food better than Cirrhina mrigala and Labeo rohita, and shows that the latter is less adapted for it than even Cirrhina mrigala. Utilization of plant matter, however, is considerably better in Cirrhina mrigala and Labeo rohita than in Catla catla. Thus it appears from the present work that Catla catla is a “planktivore” with a bias towards zooplankton, while Labeo rohita and Cirrhina mrigala are “herbivores”.

Within the confines of their preferences, the comparative utility of different food items would depend upon their rate of digestibility, for example, Labeo rohita and Cirrhina mrigala, though both mainly “herbivores”, do not utilize all plants equally. The fact that many fishes cannot digest certain algae is also already known (Fish, 1951; George, 1963). The present work while confirming this, has further shown that these indigestible items of food are retained for a longer time, nearly two days, in the intestine. It may, therefore, be stated that fishes living in ponds where such indigestible foods are abundant, may suffer a considerable loss of time and energy in having their guts full of these foods, resulting in retardation of their growth. This investigation has revealed that filamentous algae, such as Spirogyra and Ulolthrix, higher aquatic plants such as Lemna, decayed vegetation, and zooplankters such as Daphnia, Cyclops, Diaptomus, and the rotifer Brachionus, are more rapidly digested and would, therefore, be more intensively utilized, affording comparatively faster rate of growth.

6 ACKNOWLEDGEMENT

We wish to express our gratitude to Dr. C.V. Kulkarni, Director of Fisheries, Maharashtra State, Bombay, for making available the facilities for this work, and for kindly making necessary corrections in the manuscript.

7 REFERENCES

Ahsan-ul-Islam, 1949 The comparative histology of the alimentary tract of certain fresh water teleost fishes. Proc.Indian Acad.Sci.(B), 33:297–821

Balch, C.C., 1951 Factors affecting the utilization of food by dairy cows. Rate of passage of food through the digestive tract. Brit.J.Nutr., 4:4–361

Barrington, E.J.W., 1942 Gastric digestion in lower vertebrates. Biol.Rev., 17:1–27

Barrington, E.J.W., 1957 The alimentary canal and digestion. In The physiology of fishes ed. by M. Brown. New York, Academic press, pp. 109–61

Blaxter, K.L., N. McGraham and F.W. Wainmann, 1956 Some observations on the digestibility of food by sheep and on related problems. Brit.J.Nutr., 10:2–69

Begum, A., 1959 Observations of the food of major carps. Agric.Pakist., 10:539–54

Chacko, P.I. and G.K. Kurian, 1950–51 The bionomics of carp, Catla catla, in South Indian waters. Proc.zool.Soc.Lond., 120(1):31–42

Dobreff, M., 1927 Magenverdauung der Haifische: eine Bemerkung über die Hungerausdauer derselben Fische. Pflügers Arch.ges.Physiol., 217:221–34

Ewing, P.V. and F.H. Smith, 1917 A study of the rate of passage of food residues through the steer and its influence on digestion coefficients. J.Agric.Res., 10:2

Fish, G.R., 1951 Digestion in Tilapia esculanta. Nature, Lond., 167:901–2

George, M.G., 1963 Selective digestion in the major carps of India. Curr.Sci., 32:2–79

Hirao, S., J. Yamada and R. Kikuchi, 1960 On improving the efficiency of feed for fish culture. Transit and digestibility of diet in eel and rainbow trout, observed by use of P32. Bull.Tokai Fish.Res.Lab., 27:67–72

Karpevich, A.E. and E. Bokoff, 1937 The rate of digestion in marine fishes. Zool.Zh., 16:28–44

Kapoor, B.G., 1953 The histology of the alimentary canal of Catla catla, a herbivorous fish. Proc.Indian Sci.Congr., 40:209–10

Maltzan, Grafin von M., 1935 Zur Ernährungsbiologie und Physilogie des Karpfens. Zool.Zbl., 3(55):191–218

Menon, M. and H.G. Kewalramani, 1959 Studies on some physiological aspects of digestion in three species of elasmobranchs. Proc.Indian Acad.Sci. (B), 50(1):26–39

Molnar, G., and I. Tolg, 1962 Relation between water temperature and gastric digestion in largemouth bass (Micropterus salmoides). J.Fish.Res.Bd Can., 19:1005–12

Mookerjee, H.K., 1945 Life history of some major carps of Bengal. Sci.and Cult., 10(9)

Mookerjee, H.K. and B.K. Das, 1945 Gut of carnivorous and herbivorous fishes in relation to their food. Part 3. Proc.Indian Sci.Congr., 32:109 p

Mookerjee, H.K. and S.S. Ghosh, 1945 Food of major carps. Part 3. Proc.Indian Sci.Congr., 32:110–1

Sarbahi, D.S., 1940 The alimentary canal of Labeo rohita (Ham.). J.Asiat.Soc., 5:2–87

Sokolov, N.P. and M.A. Chvaliova, 1936 Nutrition of Gambusia affinis on the rice fields of Turkestan. J.Anim.Ecol., 5:390

Sullivan, M.X., 1907 The physiology of the digestive tract of elasmobranchs. Bull.U.S.Bur.Fish., (27):1–27

van Slyke, D.D. and F.G. White, 1911 Digestion of protein in the stomach and intestine of the dogfish. J.biol.Chem., 9:209–17

Vonk, H.J., 1937 The specificity and collaboration of digestive enzymes in Metazoa. Biol.Rev., 12:245–84

Vonk, H.J., 1941 Advances in enzymology. Nord and Werkman, eds., New York, Interscience, vol.1:371 p.

Weinland, E., 1901 Zur Magenverdauung der Haifische. 1 und 2. Z.Biol., 41:35–275

Yonge, C.M., 1937 Evolution and adaptation in the digestive system of the Metazoa. Biol.Rev., 12:87–115

Yung, E., 1899 Recherche sur la digestion des poissons. Arch.Zool.exp.gén., 3:7–121