Influence of Ochratoxin A and Diacetoxyscirpenol Singly and in Combination on Broiler Chickens1

Influence of Ochratoxin A and Diacetoxyscirpenol Singly and in Combination on Broiler Chickens1 L. F. KUBENA,2 R. B. HARVEY, and T. S. EDPJNGTON USDA, Agricultural Research Service, Food Animal Protection Research Laboratory, Route 5, Box 810, College Station, Texas 77845 G. E. ROTTINGHAUS Veterinary Medical Diagnostic Laboratory, College of Veterinary Medicine, University of Missouri, Columbia, Missouri 65211

1994 Poultry Science 73:408-415

been observed as a natural contaminant of feedstuffs in several countries (Chu, 1974; Ochratoxins, of which ochratoxin A Krogh et al, 1976; Prior, 1976), and field (OA) is the most prevalent, are com- outbreaks of ochratoxicosis have been pounds produced as secondary metabo- documented by Hamilton et al. (1982). lites by certain species of fungi. Van der Experimental feeding of OA has been Merwe et al. (1965) isolated OA as a shown to have detrimental effects on secondary metabolite of Aspergillus growing chicks (Huff et al, 1974, 1975, ochraceus. Additional species of Aspergillus 1992; Prior et al, 1980; Huff and Doerr, and Penicillium have been shown to produce OA (Ciegler, 1972; Ciegler et al, 1972; 1981; Kubena et al, 1983, 1985, 1986, 1988, Hesseltine et al., 1972). Ochratoxin A has 1989a). As OA is a natural contaminant of potential poultry feedstuffs and has a harmful effect on growing chicks, this mycotoxin poses a potential problem to poultry production. Received for publication June 28, 1993. Accepted for publication November 9, 1993. 4,15-Diacetoxyscirpenol (DAS) is a nat1 Mention of a trade name, proprietary product, or urally occurring toxic secondary metabospecific equipment does not constitute a guarantee or warranty by the USDA and does not imply its approval lite predominantly associated with species to the exclusion of other products that may be suitable. of Fusarium (Pathre and Mirocha, 1977). It 2 To whom correspondence should be addressed. has been implicated in reduced feed INTRODUCTION

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ABSTRACT The effects of feeding 2 mg ochratoxin A (OA) and 6 mg 4,15-diacetoxyscirpenol (DAS)/kg of diet singly and in combination were characterized in male broiler chicks from 1 to 19 d of age. Body weights were depressed by OA, DAS, and the OA-DAS combination. There was a significant antagonistic interaction between OA and DAS for uric acid and cholesterol. The efficiency of feed utilization was reduced by DAS alone and by the OA-DAS combination. When compared with controls, additive toxicity was exhibited for reduced efficiency of feed utilization, increased relative weights of the liver and gizzard, and decreased concentration of serum total protein, mean corpuscular volume, and mean corpuscular hemoglobin. All chicks were scored for oral lesions using a scale of 1 to 4 (0 = no visible lesions; 3 = severe lesions). Oral lesions (average score = 2.6) were present in over 90% of the chicks receiving the DAS diet with or without OA. These data demonstrate that both OA and DAS alone and the OA-DAS combination can adversely affect broiler performance and health. (Key words: ochratoxin A, diacetoxyscirpenol, mycotoxins, chicken, interactions)

TOXICITY OF OCHRATOXIN A AND DIACETOXYSCIRPENOL

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nivalenol, zearalenone, or cyclopiazonic acid as established by the methods given by Clement and Phillips (1985). The OA was produced, extracted, and purified by methods previously described by Huff et al. (1974). The OA was determined to be greater than 95% pure using an OA authentic standard by reverse phase HPLC. Through nuclear magnetic resonance and mass spectrometry, the DAS 3 was determined to be greater than 99% pure. The OA and DAS were incorporated into the basal diet by dissolving each of the toxins in 95% ethanol and then mixing the appropriate quantities with 2 kg of the diet. After drying, these 2-kg quantities of diet were mixed with the rest of the basal diet to produce the three treatments containing added toxins. Broilers were weighed individually on a weekly basis, feed consumption was recorded weekly, and mortality was MATERIALS AND METHODS recorded as it occurred. When the chicks Day-old male broiler Peterson x Hub- reached 19 d of age, the feeding trial was bard chicks were obtained from a com- terminated and 24 broilers (eight replimercial h a t c h e r y a n d i n d i v i d u a l l y cates of three chicks each) from each weighed and wing-banded. Chicks were treatment were bled by cardiac puncture placed into electrically heated batteries for serum biochemical analyses. Sixteen blood samples from these same chicks under fluorescent lighting with feed and (eight replicates of two chicks each) from water provided for ad libitum consumpeach treatment were used for hematologition. Based on published data on the cal determinations. Twenty-four chicks effects of OA (Kubena et al, 1989a) and (three chicks from each replicate) from DAS (Kubena et al, 1993), the following each treatment were killed by cervical four dietary treatments were selected: 1) dislocation, and the liver, kidney, spleen, Control, with 0 mg OA and 0 mg DAS/kg pancreas, proventriculus, gizzard, and of diet; 2) 2 mg O A / k g of diet; 3) 6 mg bursa of Fabricius were removed and DAS/kg of diet; and 4) 2 mg OA plus 6 weighed. mg DAS/kg of diet. There were eight At the termination of the study, heads replicates of 7 broilers per dietary treat- were removed from all chicks and visually ment, for a total of 224 chicks. Broilers scored for oral lesions (using a four-point were maintained on these treatments to 19 scoring system ranging from 1 to 4) by the d of age. The chicks were fed an unmedi- same individual without knowledge as to cated corn and soybean meal basal diet of treatment groups. A lesion score of 1 commercial type that contained or ex- indicated no visible lesions; a score of 2 ceeded the levels of critical nutrients was seen as one or two mouth lesions recommended by the National Research clearly visible on either the lower or upper Council (1984). The basal diet was ana- mandible; a lesion score of 4 was seen as lyzed for mycotoxins and was below the large lesions occurring at several sites detection limits for aflatoxin, deoxy- within the mouth, principally on the upper and lower mandibles, the corners of the mouth, and the back of the tongue; lesions scored as 3 were intermediate in 3 Kindly provided by G. H. Rottinghaus, Veterinary Medical Diagnostic Laboratory, College of appearance to lesions scored 2 or 4. Serum concentrations of total protein, Veterinary Medicine, University of Missouri, Columalbumin, glucose, cholesterol, triglycerbia, MO 65202.

intake, feed refusal, or toxicity in ducklings (Steyn et al, 1978) and chickens (Chi and Mirocha, 1978; Hoerr et al, 1981a,b, 1982a,b; Allen et al, 1982; Burditt et al, 1983; Ademoyero and Hamilton, 1989; Richardson and Hamilton, 1990; Sigmon et al, 1990; Kubena et al, 1993). To the authors' knowledge, OA and DAS have not been observed occurring simultaneously in a single potential feed grain source. However, the use of multiple-grain sources in poultry and livestock diets is common; thus the possibility exists for feeding diets cocontaminated with OA and DAS. The purpose of this research was to investigate and describe the major effects of feeding male broiler chicks diets containing OA and DAS singly and in combination from 1 to 19 d of age.

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RESULTS The effects of OA and DAS singly and in combination are shown in Table 1. Ochratoxin A and DAS significantly reduced body weights when fed alone. The body weights of the chicks fed the combination of OA and DAS were almost identical to the chicks fed the diet containing DAS alone. There was a significant OA by DAS interaction for body weights which could best be characterized as less than additive. The efficiency of feed utilization was reduced by DAS alone and the OA-DAS combination. Mortality ranged from 0 to 9% with the 9% mortality occurring in the OA alone treatment. Over

4 Gilford Impact, 400E, Ciba Corning Diagnostics Corp., Gilford Systems, Oberlin, OH 44774. 5 Counter Electronics, Hialeah, FL 33012. 6 SAS Institute Inc., Cary, NC 27512.

90% of the chicks fed diets containing DAS with or without OA had oral lesions with an average lesion score of 2.6 (1 = no lesions; 4 = severe lesions). When compared with controls, relative weights of the liver and kidney were increased by feeding OA singly or in combination with DAS (Table 2). No changes in the relative weights of the liver or kidney were observed when DAS was fed alone. The relative weights of the proventriculus and gizzard were increased by feeding DAS alone, whereas feeding the OA-DAS combination resulted in an increase in the relative weight of the gizzard but not the proventriculus. The relative weights of the spleen, pancreas, and bursa of Fabricius were not significantly different from those of the control in any of the treatments (data not shown). Data presented in Table 3 show the effects of dietary treatments on serum biochemical values, serum enzyme activities, and hematology values. When compared with controls, serum total protein and albumin concentrations were reduced by OA when present alone or in combination with DAS. A significant interaction, best characterized as less than additive, between OA and DAS occurred for decreased serum albumin. There was a significant increase in serum uric acid concentrations in chicks fed the diet containing OA alone, whereas uric acid concentrations were not increased in the chicks receiving DAS alone or the OADAS combination. In fact, there was a significant antagonistic interaction between OA and DAS, with the uric acid concentrations of the OA-DAS combination approaching those of the controls. Serum concentrations of triglycerides were significantly increased, whereas concentrations of cholesterol were decreased in chicks fed the diet containing OA alone. The serum concentrations of triglycerides or cholesterol were not altered in the chicks receiving the diets containing DAS alone or in combination with OA. A significant antagonistic interaction occ u r r e d b e t w e e n OA a n d DAS for cholesterol concentrations. When compared with controls, the activities of creatine kinase were significantly reduced in the serum of the chicks receiving DAS

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ides, uric acid, and blood nitrogen and activities of lactate dehydrogenase, alkaline phosphatase, y glutamyltransferase, aspartate aminotransferase, and creatine kinase were determined on a clinical chemistry analyzer 4 according to the manufacturer's recommended procedure. Hemoglobin was measured as cyanmethemoglobin. 5 Erythrocyte count, mean corpuscular volume, and hematocrit were determined with a Coulter 5 Model ZBI counter equipped with a mean corpuscular volume and hematocrit computer and channelizer. The mean corpuscular hemoglobin and mean corpuscular hemoglobin concentrations were calculated. Data (pen x) analyses for all response variables were conducted on PCSAS® Version 6.02.6 The data were analyzed as a 2 x 2 factorial design to determine main and interactive effects. In addition, data were subjected to ANOVA (Snedecor and Cochran, 1967) using the General Linear Models procedure to establish differences between treatment means. Means showing significant differences in ANOVA were compared using the Fisher's protected least significant difference procedure (Snedecor and Cochran, 1967). All statements of significance are based on the .05 level of probability.

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TOXICITY OF OCHRATOXIN A AND DIACETOXYSCIRPENOL TABLE 1. Effects of dietary ochratoxin A (OA), diacetoxyscirpenol (DAS), and the OA-DAS combination on body weights, feed utilization, mortality, and oral lesions (19 d)1 Treatment OA

DAS (mg/kg)

0 2 0 2 LSD4

Change from control

Body weight 2

(%)

(g) 622-> 549 b 450c 448<4 59

0 0 6 6

0 -12 -28 -28

Feed:gain

Mortality

Oral lesions 3

(kg:kg) 1.61« 1.73bc 1.79* 1.87»

(%) 4 9 2 0

(% of chicks) 0 0 97 91

.13

(Scores) 1.0b 1.0b 2.6» 2.6a .4

a_c

Means within a column with no common superscript differ significantly (P < .05). Values represent the x of eight groups of seven broilers each per treatment minus mortality. Significant OA x DAS interaction (P < .05), best described as less than additive. 3 Visually scored for oral lesions (1 = no lesions; 4 = severe lesions) by the same individual. 4 LSD = least significant difference as determined by Fisher's protected LSD procedure. 1

values did not differ from those of controls (data not shown). DISCUSSION Various mycotoxin combinations have been studied in poultry. These include aflatoxin and OA (Huff and Doerr, 1981; Campbell et al, 1983; Huff et al, 1984, 1988b, 1992), T-2 toxin and DAS (Hoerr et al, 1981a), OA and citrinin (Doerr et al, 1982; Glahn et al, 1988, 1989), OA and penicillic acid (Kubena et al, 1984), aflatoxin and deoxynivalenol (Huff et al, 1986), and OA and T-2 toxin (Kubena et al, 1989a). Other mycotoxin combinations studied in poultry include aflatoxin and T-

TABLE 2. Effects of dietary ochratoxin A (OA), diacetoxyscirpenol (DAS), and the OA-DAS combination on relative organ weights (19 d)1 Treatment OA

DAS

Liver

Kidney

0 0 6 6

3.33b 3.86a 3.19b 3.66a

.54b .67a .55b .63a

.24

.05

- (mg/kg) 0 2 0 2 LSD2 a b

Provenfriculus (g/100 g BW) .60b .67ab .69a .67ab .08

< Means within a column with no common superscript differ significantly (P < .05). Values represent the x of eight groups of three broilers each per treatment. 2 LSD = least significant difference as determined by Fisher's protected LSD procedure. J

Gizzard 2.87b 3.09ab 3.20a 3.33a .30

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alone or in combination with OA. When compared with controls, concentrations of glucose and blood nitrogen and activities of lactate dehydrogenase, alkaline phosphatase, 7 glutamyltransferase, and aspartate aminotransferase were not altered by treatment (data not shown). When compared with values for controls, mean corpuscular volumes were significantly decreased in the chicks receiving diets containing OA alone, DAS alone, or the OA-DAS combination. Mean corpuscular hemoglobin values were reduced in chicks receiving the diets containing DAS alone or in combination with OA. Hemoglobin, erythrocyte count, hematocrit, and mean corpuscular hemoglobin concentration

412

KUBENA ET AL.

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2 toxin (Huff et a/., 1988a; Kubena et al, 1990), OA and deoxynivalenol (Kubena et al, 1988), deoxynivalenol and T-2 toxin (Kubena et al, 1989b), aflatoxin and kojic acid (Girior et al, 1991), aflatoxin and cyclopiazonic acid (Smith et al, 1992), and aflatoxin and DAS (Kubena et al, 1993). Ochratoxin A and DAS are important mycotoxins due to their toxicity and occurrence in feedstuffs that are used in poultry and livestock diets (Pathre and Mirocha, 1977; Chi and Mirocha, 1978; Hamilton et al, 1982). In the present study, the individual toxic effects of OA were expressed as reduced weight gains, increased relative liver and kidney weights, increased serum uric acid concentrations, decreased concentrations of serum total protein and albumin, and alterations in hematological values. These data on the effects of OA alone are consistent with several earlier reports (Huff et al, 1974, 1975, 1992; Prior et al, 1980; Huff and Doerr, 1981; Kubena et al, 1983, 1985, 1986, 1988, 1989a). The individual toxicity of DAS has been demonstrated by several researchers (Chi and Mirocha, 1978; Hoerr et al, 1981a,b, 1982a,b; Allen et al, 1982; Burditt et al, 1983; Ademoyero and Hamilton, 1989; Richardson and Hamilton, 1990; Sigmon et al, 1990; Kubena et al, 1993). The individual toxic effects, including reduced body weight gains, oral lesions, and increased relative weights of the proventriculus and gizzard, are in agreement with previous reports. The current study defined the toxicity of the combination OA and DAS in growing broiler chicks. This toxicity can be characterized by reduced weight gains, decreased efficiency of feed utilization, oral lesions (caused by DAS), increased relative weights of the liver and kidney (caused by OA), and increased relative weight of the gizzard (caused by DAS), decreased serum concentrations of total protein and albumin (caused by OA), decreased creatine kinase activity mean corpuscular volume and mean corpuscular hemoglobin concentration (caused by DAS), and mean corpuscular hemoglobin volume.

TOXICITY OF OCHRATOXIN A AND DIACETOXYSCIRPENOL

the mycotoxins, typically described as focally reddened mucosa (Hoerr et ah, 1982a). These data indicate that under the current experimental conditions, 2 mg OA and 6 mg DAS/kg of diet appear to exert their effects in an additive, less than additive (but not significantly antagonistic), or in an antagonistic manner. Additive toxicity was exhibited for reduced efficiency of feed utilization, increased liver and gizzard weights, and decreased concentration of serum total protein, mean corpuscular volume, and hemoglobin. Variables exhibiting less than additive toxicity but not significantly antagonistic were reduced body weight gains, increased relative weights of the kidney and proventriculus, decreased albumin and triglyceride concentrations, decreased mean corpuscular hemoglobin, and decreased creatine kinase activity. Variables exhibiting a significant antagonistic interaction were decreased serum concentrations of uric acid and cholesterol. These variables were considered antagonistic because when fed together, DAS interfered with the action of OA, which meets the criteria stated by Klaassen and Eaton (1991). These data are in agreement with earlier research with the OA and deoxynivalenol combination (Kubena et ah, 1988). Surprisingly, the current data differ from the OA and T-2 toxin combination research (Kubena et al., 1989b), in which most of the variables, including body weight gains, were additive in nature. The current findings suggest that in some instances OA and DAS may be competing for absorption from the gastrointestinal tract or possibly for an active site on a common enzyme, as previously suggested for OA and deoxynivalenol (Kubena et al., 1988). The difference in response of DAS and T-2 toxin might be associated with the carbon chain moiety at Carbon 8 found on T-2 toxin but not DAS. This carbon moiety, lacking in DAS, possibly had a specific effect on target organs or perhaps altered the biotransformation and tissue kinetics and thereby increased the toxicity, as Hoerr et al. (1982a) found T-2 toxin to be more destructive for lymphoid and hematopoietic tissues than DAS. The sub-

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28% by the OA and DAS combination, indicating a significant interaction at these levels of toxins. The reduction represents the effects of DAS that could not be added to by OA at these toxin concentrations. Mortality ranged from 0% in the OA-DAS combination group to 9% in the OA alone group. The average oral lesion score for chicks receiving DAS with or without OA was 2.6 (1 = no lesions; 4 = severe lesions) with over 90% of the chicks having lesions. These data indicate the reduced body weight gains and the oral lesions were caused by DAS and were not influenced by OA. The reduced growth rates, decreased efficiency of feed utilization, decreased concentrations of serum total proteins and albumen, decreased creatine kinase activity, and decreased hematology values are most likely associated with inhibition of protein synthesis by the mycotoxins (Tung et al, 1975). The OA may compete with phenylalanine for binding sites on the phenylalanine-transfer RNA synthetase enzyme, thus inhibiting protein synthesis (Creppey et ah, 1980). The DAS may block protein synthesis by the inhibition of peptidyltransferase and interfering with initiation and termination reactions (Ueno, 1991). The increased relative weights of the kidney and increased serum concentrations or uric acid indicate impaired renal function due to the ingestion of OA. The increased kidney weights, indicating the nephrotoxicity of OA, have been demonstrated many times, being reported as early as 1974 (Huff et ah, 1974). There was a significant antagonistic effect of DAS on serum concentrations of uric acid but not on the relative weights of the kidney. The increased relative weight of the liver and the increased triglyceride concentrations might be associated with alterations in lipid metabolism, primarily as a result of impaired lipid transport. The reduction in serum cholesterol is most likely due to the inhibition of cholesterol biosynthesis, with liver involvement and perhaps a shift of concentration from the blood to the liver as suggested by Kubena et al. (1989a). The increased relative weights of the proventriculus and gizzard can most likely be attributed to overall irritative properties of

413

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KUBENA ET AL.

stituent groups at Carbons 3, 4, 7, 8, and 15 of the trichothecene nucleus greatly influence the association of the nucleus with the ribosomes. In the cells of yeast, the inhibition was found to be reversible for DAS, being removable from its target site by washing, but irreversible for T-2 toxin (Ueno, 1991). Poultry and livestock may be more susceptible to OA and DAS as well as other mycotoxins and other toxicants if nutritional, environmental, health, or other stress factors are involved.

The authors gratefully acknowledge the excellent technical assistance of Maurice Connell, Albert Blanks, Ellen Moore, James Snodgrass, and Craig Sweatt.

REFERENCES Ademoyero, A. A., and P. B. Hamilton, 1989. Research note: Influence of degree of acetylation of scirpenol mycotoxins on feed refusal by chickens. Poultry Sci. 68:854-856. Allen, N. K., R. L. Jevne, C. J. Mirocha, and Y. W. Lee, 1982. The effect of Fusarium roseum culture and diacetoxyscirpenol on reproduction of White Leghorn females. Poultry Sci. 61: 2172-2175. Burditt, S. J., W. M. Hagler, Jr., and P. B. Hamilton, 1983. Survey of molds and mycotoxins for their ability to cause feed refusal in chickens. Poultry Sci. 62:2187-2191. Campbell, M. L., J. D. May, W. E. Huff, and J. A. Doerr, 1983. Evaluation of immunity of young broiler chickens during simultaneous aflatoxicosis and ochratoxicosis. Poultry Sci. 62:2138-2144. Chi, M. S., and C. J. Mirocha, 1978. Necrotic oral lesions in chickens fed diacetoxyscirpenol, T-2 toxin and crocotin. Poultry Sci. 57:807-808. Chu, F. S., 1974. Studies on ochratoxicosis. Crit. Rev. Toxicol. 3:499-524. Ciegler, A., 1972. Bioproduction of ochratoxin A and penicillic acid by members of the Aspergillus ochraceus group. Can. J. Microbiol. 18:631-636. Ciegler, A., D. J. Fennell, H. J. Mintzlaff, and L. Leistner, 1972. Ochratoxin synthesis by Penicillium species. Naturwissenschaften 59:365-366. Clement, B. A., and T. D. Phillips, 1985. Advances in the detection and determination of mycotoxins via capillary GC/quadrupole mass spectrometry. Toxicologist 5(1):232. Creppy, E. E., M. Schlegel, R. Roschenthaler, and G. Dirheimer, 1980. Phenylalanine prevents acute poisoning by ochratoxin A in mice. Toxicol. Lett. 60:77-80. Doerr, J. A., M. L. Campbell, and W. E. Huff, 1982. Interaction between dietary citrinin and ochratoxin A in broiler chickens. Poultry Sci. 61: 1453.(Abstr.)

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ACKNOWLEDGMENTS

Giroir, L. E., W. E. Huff, L. F. Kubena, R. B. Harvey, M. H. Elissalde, D. A. Witzel, A. G. Yersin, and G. W. Ivie, 1991. The individual and combined toxicity of kojic acid and aflatoxin in broiler chickens. Poultry Sci. 70:1351-1356. Glahn, R. P., R. S. Sharpiro, V. E. Vena, R. F. Wideman, and W. E. Huff, 1989. Effects of chronic ochratoxin A and citrinin toxicosis on kidney function of Single Comb White Leghorn pullets. Poultry Sci. 68:1205-1212. Glahn, R. P., R. F. Wideman, J. W. Evangelisti, and W. E. Huff, 1988. Effects of ochratoxin A in combination with citrinin on kidney function of Single Comb White Leghorn pullets. Poultry Sci. 67:1034-1042. Hamilton, P. B., W. E. Huff, J. R. Harris, and R. D. Wyatt, 1982. Natural occurrences of ochratoxicosis in poultry. Poultry Sci. 61:1832-1841. Hesseltine, C. W., E. E. Vandegraft, D. J. Fennell, M. L. Smith, and O. L. Shotwell, 1972. Aspergilli as ochratoxin producers. Mycologia 64:539-550. Hoerr, F. J., W. W. Carlton, and B. Yagen, 1981a. The toxicity of T-2 toxin and diacetoxyscirpenol in combination for broiler chickens. Food Cosmet. Toxicol. 19:185-188. Hoerr, F. J., W. W. Carlton, and B. Yagen, 1981b. Mycotoxicosis caused by single doses of T-2 toxin or diacetoxyscirpenol in broiler chickens. Vet. Pathol. 18:652-664. Hoerr, F. J., W. W. Carlton, B. Yagen, and A. Z. Joffe, 1982a. Mycotoxicosis produced in broiler chickens by multiple doses of either T-2 toxin or diacetoxyscirpenol. Avian Pathol. 11:369-383. Hoerr, F. J., W. W. Carlton, B. Yagen, and A. Z. Joffe, 1982b. Mycotoxicosis caused by either T-2 toxin or diacetoxyscirpenol in the diet of broiler chickens. Fundam. Appl. Toxicol. 2:121-124. Huff, W. E., and J. A. Doerr, 1981. Synergism between aflatoxin and ochratoxin A in broiler chickens. Poultry Sci. 60:550-555. Huff, W. E., J. A. Doerr, C. J. Wabeck, G. W. Chaloupka, J. D. May, and J. W. Merkley, 1984. The individual and combined effects of aflatoxin and o c h r a t o x i n o n v a r i o u s p r o c e s s i n g parameters of broiler chickens. Poultry Sci. 63: 2153-2161. Huff, W. E., R. B. Harvey, L. F. Kubena, and G. E. Rottinghaus, 1988a. Toxic synergism between aflatoxin and T-2 toxin in broiler chickens. Poultry Sci. 67:1418-1423. Huff, W. E., L. F. Kubena, R. B. Harvey, and J. A. Doerr, 1988b. Mycotoxin interactions in poultry and swine. J. Anim. Sci. 66:2351-2355. Huff, W. E., L. F. Kubena, R. B. Harvey, W. M. Hagler, Jr., S. P. Swanson, T. D. Phillips, and C. R. Creger, 1986. Individual and combined effects of aflatoxin and deoxynivalenol (DON, vomitoxin) in broiler chickens. Poultry Sci. 65: 1291-1298. Huff, W. E., L. F. Kubena, R. B. Harvey, and T. D. Phillips, 1992. Efficacy of hydrated sodium calcium aluminosilicate to reduce the individual and combined toxicity of aflatoxin and ochratoxin A. Poultry Sci. 71:64-69. Huff, W. E., R. D. Wyatt, and P. B. Hamilton, 1975. Nephrotoxicity of dietary ochratoxin A in broiler chickens. Appl. Microbiol. 30:48-51.

TOXICITY OF OCHRATOXIN A AND DIACETOXYSCIRPENOL

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