Vomitoxin in diets for rainbow trout (Salmo gairdneri)

Aquaculture, 35 (1983) 93-101 Elsevier Science Publishers B.V., Amsterdam - Printed in The Netherlands

VOMITOXIN

IN DIETS FOR RAINBOW TROUT (SALMO GAIRDNERI)

BILL WOODWARD’$3 L.G. YOUNG’

’ Department

of Nutrition,

and A.K. LUN’

University of Guelph, Guelph, Ont. Nl G 8Wl (Canada)

‘Department of Animal and Poultry Science, Nl G 2 Wl (Canada) 3To whom correspondence (Accepted

93

University of Guelph, Guelph, Ont.

should be addressed.

23 November 1982)

ABSTRACT Woodward, Bill, Young, L.G. and Lun, A.K., 1983. trout (Salmo gairdneri). Aquaculture, 35: 93-101.

Vomitoxin

in diets for rainbow

The present study was conducted to obtain information as to the response and sensitivity of rainbow trout (Salmo gairdneri) to diets containing vomitoxin-contaminated corn. Feed refusal occurred when diets contained 20 pg/g vomitoxin or more, but the trout recovered rapidly when subsequently fed a diet containing no detectable toxin, Diets containing graded levels of vomitoxin, increasing from 1.0 to 13.0 fig/g, caused progressively greater depression in 4-week liveweight gains of juvenile trout. The depression in weight gain ranged from 12% to 92% of the control value and resulted from an adverse effect on both feed intake and feed conversion efficiency. Emesis was not observed in this work. The results demonstrate that rainbow trout are highly sensitive to dietary vomitoxin.

INTRODUCTION

Fusarium molds produce a variety of toxic compounds. Among the naturally-occurring trichothecene mycotoxins (12, 13epoxytrichothec9-enes) is 3, 7, 15-trihydroxy-12, 13_epoxytrichothec-9-en&one which was first isolated by Morooka et al. (1972) and given the trivial names “Rd toxin” and “deoxynivalenol”. Independently, Vesonder et al. (1973) isolated the same compound and named it “vomitoxin”. This toxin has since been implicated in many species as both an emetic (Morooka et al., 1972; Vesonder et al., 1973; Ishii et al., 1975; Forsyth et al., 1977; Yoshizawa and Morooka, 1977; Young et al., 1983) and a feed refusal factor (Vesonder et al., 1976; Forsyth et al., 1977; Yoshizawa and Morooka, 1977; Young et al., 1981; Young et al., 1983). We are unaware;however, of any information concerning the response or sensitivity of the rainbow trout (Sdmo gairdneri), or other fish species, to vomitoxin. Contamination of natural-product trout diets with Fusarium toxins,

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Q 1983 Elaevier Science Publishers B.V.

94

including vomitoxin, is a potential practical problem. Many cereals readily become infected with Fusarium molds, and vomitoxin has been identified in samples from naturally-occurring infestations of corn (Vesonder et al., 1973; Forsyth et al., 1977), barley (Yoshizawa and Morooka, 1977) and wheat (Trenholm et al., 1981). Since wheat products are commonly used in formulating diets for trout, it is important to investigate the influence of mycotoxins such as vomitoxin on the health and performance of these LXlimdS.

MATERIALS AND METHODS

Two experiments were performed. In both cases fish were obtained as fry from Goossens Trout Farm Ltd., Otterville, Ontario, and were maintained in a non-recirculating system of 60-l rectangular tanks supplied with a mixture of University of Guelph well water and Guelph city water at a rate of approximately 2 l/min. The water was continuously aerated and temperature was controlled thermostatically at 15 + 0.5”C. Photoperiod was maintained on a 12L : 12D schedule. Corn from an artificially infected crop was used as the source of dietary vomitoxin. The material comprised a mixture of ground husks, cobs and kernels, and contained approximately 4 fig zearalenone/g, a trace of 7-deoxyvomitoxin, but undetectable levels of numerous other mycotoxins including nivalenol, 3-acetyldeoxynivalenol, fusarenone-X, diacetoxyscirpenol, T-2 toxin, neosolaniol and HT-2 toxin (Young et al., 1983; P.W. Scott, personal communication, 1982). At the levels of corn used in the present experiments, therefore, vomitoxin was the only known Fusarium toxin present in sig-, nificant quantity in the diets. All diets were steam pelleted under conditions similar to those described by Hilton et al. (1981). Statistical analyses were performed according to Sokal and Rohlf (1969), and differences at the 5% level of probability were considered significant.

Experiment 1 The first experiment was a preliminary trial to indicate the nature of the response of the trout to consumption of a diet containing vomitoxincontaminated corn. The basal diet used is shown in Table I. Six diets were formulated to contain graded levels of mycotoxin by replacing good quality ground shelled corn with an equal weight of vomitoxin-contaminated material such that the diets contained 0, 20, 40, 60, 80 or 100 g contaminated corn/kg of diet. By this procedure all diets contained 10% corn. Fry were randomly distributed into six tanks, 125 animals per tank, and each group of fish was fed one of the six diets to near satiety four times daily for 8 weeks. Live weights and feed intakes were recorded fortnightly throughout the experiment. Both the design of the study and the vomitoxin level of each diet measured, according to Young et al. (1983), are

95

TABLE I Basal diet for experiment la Ingredient

g/kg Diet

Herring meal (65% crude protein) Soybean meal (49% crude protein) Wheat middlings (18% crude protein) Corn Vitamin-mineral premixb DL-methionine Capelin oil=

430 200 170 100 28 2 70

aProximate analysis according to the methods of the AOAC (1970) on a % air-dry basis: dry matter, 96.3; crude protein (% N X 6.25), 42.0;ether extract, 13.3; crude fiber, 2.1; ash, 9.6; gross energy, 21.1 kJ/g. bThe supplement supplied the following levels of nutrients/kg of diet: menadione sodium bisulfite 30 mg; thiamin HCl 30 mg; D-calcium pantothenate 150 mg; biotin 0.4 mg; folic acid 10 mg;vitamin B,, 0.03 mg; niacin 200 mg;riboflavin, 10 mg; pyridoxine HC130 mg; choline chloride (50%) 4 g; ascorbic acid 350 mg; ferrous sulfate (21% Fe) 300 mg; cupric sulfate (25% Cu) 100 mg; zinc sulfate (36% Zn) 400 mg. The supplement was brought to appropriate weight (2.8% of the diet) by addition of wheat middlings. cStabilized with ethoxyquin and supplemented with retinyl acetate, cholecalciferol and all-rat-cY-tocopherol acetate to supply (per kg diet) 5000 IU, 2000 IU and 200 IU, respectively . TABLE II Design of experiment 1 Fish group designation

1 2 3 4 5 6

Dieta O-4 Weeks

4-8

Weeks

l(O) 2 (19.4) 3 (40.4) 4 (55.3) 5 (84.3) 6 (109.6)

2 (19.4) I(0) 3 (40.4) l(O) 5 (84.3) l(O)

aDiet No. (vomitoxin level in rg/g, air-dry basis. Limit of detection was 0.2 fig/g).

shown in Table II. After 4 weeks, the diets presented to groups 1, 2, 4 and 6 were switched, as indicated in Table II, so as to d&err&e the ab& ity of fish to recover from exposure to vomitoxin-contaminated diets.

Experiment 2 Fish obtained as fry were maintained, until used as juveniles (approximately 50 g/fish) on an extruded diet similar to. that of Hilton et al. (1981)

96

except that 60-120 g/kg wheat bran replaced an equal weight of wheat middlings. The basal diet for experiment 2 was similar to that of the first trial (Table I) except that it contained 403 g herring meal, 285 g wheat middlings and 12 g corn/kg. By proximate analysis (AOAC, 1970) the air-dry basal diet contained 92.2% dry matter, 37.7% crude protein (% N X 6.25), 12.8% ether extract, 1.8% crude fiber, 8.7% ash and 20.0 kJ/g gross energy. Six diets were formulated from the basal according to the same principles as described for experiment 1. The diets contained 0, 1, 3, 6, 9 and 12 g contaminated corn/kg and, by analysis, 0, 1.0, 2.0, 4.9, 7.5 and 12.9 pg vomitoxin/g (diets 1-6, respectively, air-dry basis). All diets contained 1.2% corn. Fish were randomly distributed into 12 tanks, 50 animals/tank, such that each dietary treatment was represented twice in a randomized complete block design. The fish were fed three times daily to near satiety. Live weights and feed intakes were measured fortnightly, and the experiment was terminated after 4 weeks. A lo_ l Group

1

g_ oGroup2 aGrOUp 8_

oGroup6

Weight

0

1

2

3

0

1

2

3

4 Weeks

5

6

7

a

4

5

6

7

6

Weeks

Fig. 1. Experiment 1 live weights. A, Groups 1, 2, 4 and 6. Arrow indicates point at which diets were switched (cf. Table II). B, Groups 3 and 5 which were fed diets containing a high level of vomitoxin throughout the S-week experiment.

97 RESULTS

Experiment

1

Fish groups 2-6 exhibited feed refusal within 5 days. Although these animals occasionally attempted to ingest a pellet of food, they invariably released the pellet from their mouths without swallowing. This behavior was maintained by groups 3 and 5 throughout the S-week experiment. When offered the non-contaminated diet (diet l), fish in groups 2 and 4 ingested food within 2 days and fed vigorously after 8 and 13 days, respectively. Group 6 animals, when subjected to the same procedure, began to feed after 4 days and exhibited vigorous feeding behavior after 13 days. In contrast, group 1 animals refused diet 2 (19.4 rig/g toxin) within 7 days. The live weights of each group of fish are plotted against time in Fig. 1. During the first 4 weeks of the trial, group 1 fish exhibited a feed conversion efficiency of 1.1 g air-dry feed/g liveweight gain. Similarly, between weeks 4 and 8 groups 2, 4 and 6 exhibited feed efficiencies of 1.0, 1.0 and 1.1, respectively. 50_ -a

Y =47.0-4.9X

101

I 0

1

1 2

Dietary

1

1 4

1

1 6

Vomitoxin

+0.2X2

I

I

I

8 Level

I 10

1

1 12

I

1 14

( )IQ /Q)

Fig. 2. Regression curve of 4-week feed intakes vs. dietary vomitoxin ment 2. Results for each tank of fish are shown (X).

level in experi-

98

Experiment 2 Fish fed diets 1, 2 and 3 exhibited vigorous feeding behavior throughout the experiment, while those given diets 4, 5 and 6 displayed perceptibly less interest in feeding within 2 days of the initiation of the trial. Regression analyses produced the prediction curves shown in Figs. 2, 3 and 4 relating 4-week feed intakes, liveweight gains and feed conversion efficiencies, respectively, to dietary vomitoxin content. It is of interest that, at the a-week time point, the weight gains of fish fed 1.0 pg vomitoxin/g diet (diet 2) were significantly below the gains of fish given diet 1 (18.5 + 0.3 g/fish vs. 21.5 r 0.6 g/fish, respectively; Turkey’s honestly significant difference = 2.3 g/fish). Neither vomiting nor mortality was observed in experiment 1 or in experiment 2. -0

Weight

_

Gain (g/fish)

_

01

’ 0

’

’ 2

Dietary

’

’ 4

’

’ 6

Vomitoxin

’

’ 8

’

’ 10

’

’ 12

’

I 14

Level (pg/g)

Fig. 3. Regression curve of 4-week liveweight gains vs. dietary vomitoxin level in experiment 2. Results for each tank of fish are shown (X). Initial live weights among the 12 tanks of fish ranged from 46.5 to 48.0 g/fish and did not differ among the six dietary treatments (Tukey’s honestly significant difference = 2.5 g/fish).

99 4.0

l-

0 V=e(0.086X+0.02) 3.0, Feed

_

Efficiency

_

(gfeedl

_

ggain)

2 R = 0.91

_

2.0_

0

2

4

Dietary

Fig. 4. Regression

6

Vomitoxin

8

10

12

14

Level (pg /g )

curve of feed conversion

efficiency

experiment2. Results for each tank of fish are shown (X).

vs. dietary

vomitoxin level in

DISCUSSION

The present results show that diets containing vomitoxin depress feed acceptance by rainbow trout. In experiment 2 feed intake was depressed by approximately 9% while weight gain was depressed by about 11% with each additional pg/g of vomitoxin up to a level of 5 pg/g. At least at the juvenile stage, therefore, the rainbow trout appears to be comparable to the pig (cf. Forsyth et al., 1977 and Young et al., 1983) in sensitivity to vomitoxin-contaminated material. Emesis was not observed in the trout, however, so that the response of this species is not identical to the reaction of pigs when high levels (approximately 20 pg/g) of toxin are fed (cf. Young et al., 1983). It is relevant to note that rainbow trout possess a reflex which resembles the mammalian vomiting reflex and will exhibit emesis, for example, when force-fed capsules of food (McLaren et al., 1946; D.G. Dixon, personal communication, 1982) and occasionally when anesthetized with the drug MS-222 (J.W. Hilton, personal communication, 1982). Prolonged exposure of trout to diets containing high levels of vomitoxin exerts no lasting adverse effect on feed intakes, liveweight gains or feed conversion efficiencies (experiment 1). Further work is required in order to determine the ability of trout to recover following diets containing vomitoxin at levels which permit long-term ingestion of the toxin. With respect to this point, however, Moran et al. (1982) found that broiler

100

chickens exhibit rapid, apparently complete recovery following continued ingestion of diets containing high levels of vomitoxin. Diets containing vomitoxin appear to depress the weight gains of rainbow trout through an adverse effect on both feed intake and feed conversion efficiency. Since it is difficult to avoid overfeeding fish which exhibit near-total feed refusal (e.g., diet 6, experiment Z), conversion efficiencies, in such cases, may reflect little more than imperfect feeding practice. In the present study, however, trout exhibited depressed conversion efficiencies after 4 weeks when fed diets which permitted consumption at rates of 40% of control intake (diet 5, experiment 2), or better. Exclusion of diet 6 from the regression analyses of experiment 2 made little difference to the equations obtained for weight gain and feed intake, but this procedure suggested a linear relationship between feed efficiency and dietary vomitoxin content such that each increase of 1 pg/g in toxin level (up to 7.5 pg/g) would result in a 6% decrease in conversion efficiency. Further studies including longer feeding periods are required in order to determine the influence of low levels of vomitoxin on feed conversion efficiency in the trout. Poston et al. (1982) described the response of rainbow trout fry to dietary T-2 toxin which, like vomitoxin, is a trichothecene compound produced by Fuscarium molds. Effects of crystalline T-2 toxin on feeding behavior, food consumption, liveweight increase and feed conversion efficiency were similar in kind and degree to the effects of vomitoxin-conlaminated corn in the present study. In addition, however, T-2 toxin caused transitory ascites, intestinal hemorrhaging and mortality, none of which occurred from feeding vomitoxin-containing material devoid of detectable T-2 toxin in the present experiments. Taken together these results suggest that rainbow trout are highly sensitive to Fusurium toxins of the trichothecene type, and that important production parameters are adversely affected by the presence of these compounds in trout diets. ACKNOWLEDGEMENTS

This work was supported by the Ontario Ministry of Agriculture and Food and by the Natural Sciences and Engineering Research Council, Canada. The technical assistance of Mr. Martin Hodgson is gratefully acknowledged. REFERENCES AOAC, 1970. W. Horowitz (Editor), Official Methods of Analysis of the Association of Official Analytical Chemists, 11th edn. Association of Official Agricultural Chemists, Washington, DC. Forsyth, D-M., Yoshiaawa, T., Morooka, N. and Tuite, J., 1977. Emetic and refusal activity of deoxynivalenol in swine. Appl. Environ. Microbial., 34: 547-552.

101 Hilton, J.W., Cho, C.Y. and Slinger, S.J., 1981. Effect of extrusion processing and steam pelleting diets on pellet durability, pellet water absorption, and the physiological response of rainbow trout (Salmo gairdneri R.). Aquaculture, 25: 185-194. Ishii, K., Ando, Y. and Ueno, Y., 1975. Toxicological approaches to the metabolites of Fusaria. IX. Isolation of vomiting factor from moldy corn infected with Fusarium species. Chem. Pharm. Bull., 23: 2162-2164. McLaren, B.A., Herman, E.F. and Elvehjem, C.A., 1946. Nutrition of rainbow trout; studies with purified rations. Arch. Biochem., 10: 433-441. Moran, Jr., ET., Hunter, B., Ferket, P., Young, L.G. and McGirr, L.G., 1982. High tolerance of broilers to vomitoxin from corn infected with Fusarium graminearum. Poult. Sci., 61: 1828-1831. Morooka, N., Uratsuji, N., Yoshizawa, T. and Yamamoto, H., 1972. Studies on the toxic substances in barley infected with Fusarium spp. J. Food Hyg. Sot. Jpn., 13: 368-375. Poston, H.A., Coffin, J.L. and Combs, Jr., G.F., 1982. Biological effects of dietary T-2 toxin on rainbow trout, Salmo gairdneri. Aquat. Toxicol., 2: 79-88. Sokal, R.R. and Rohlf, F.J., 1969. Biometry. W.H. Freeman and Co., San Francisco, CA, 776 pp. Trenholm, H.L., Cochrane, W.P., Cohen, H., Elliot, J.I., Farnworth, E.R., Friend, D-W., Hamilton, R.M.G., Neish, G.A. and Standish, J.F., 1981. Survey of vomitoxin contamination of the 1980 white winter wheat crop in Ontario, Canada. J. Am. Oil Chem. Sot., 58: 992A. Vesonder, R.F., Ciegler, A. and Jensen, A.H., 1973. Isolation of the emetic principle from Fusarium infected corn. Appl. Microbial., 26: 1008-1010. Vesonder, R.F., Ciegler, A., Jensen, A.H., Rohwedder, W.K. and Weisleder, D., 1976. Co-identity of the refusal and emetic principle from Fusarium -infected corn. Appl. Environ. Microbial., 31: 280-285. Yoshizawa, T. and Morooka, N., 1977. Trichothecenes from mold-infested cereals in Japan. In: J.V. Rodricks, C.W. Hesseltine and MA. Mehlman (Editors), Mycotoxins in Human and Animal Health. Pathotox Publishers, Inc., Park Forest South, IL, pp. 309-321. Young, L.G., Vesonder, R.F., Funnell, H.S., Simons, I. and Wilcock, B., 1981. Moldy corn in diets of swine. J. Anim. Sci., 52: 1312-1318. Young, L.G., McGirr, L.G., Valli, V.E., Lumsden, J.H. and Lun, A.K., 1983. Vomitoxin in corn fed to young pigs. J. Anim. Sci., 57 (3): in press.