Fumonisins

Chapter 71

Fumonisins Geof W. Smith

INTRODUCTION

BACKGROUND

Fumonisins are a group of naturally occurring mycotoxins produced by the fungi Fusarium verticillioides (formerly F. moniliforme), Fusarium proliferatum and other Fusarium species. These toxic metabolites of corn have been implicated in field cases of porcine pulmonary edema (PPE) (Colvin et al., 1993; Harrison et al., 1990; Osweiler et al., 1992) and equine leukoencephalomalacia (ELEM) (Jovanovi´c et al., 2015; Wilson et al., 1990a). Experimentally, fumonisin has been shown to cause liver damage in multiple species including pigs, horses, cattle, rabbits, and primates (Gumprecht et al., 1995; Haschek et al., 1992; Jaskiewicz et al., 1987; Osweiler et al., 1993; Ross et al., 1993; Voss et al., 1989) as well as species-specific target organ toxicity, such as lung in pigs (Haschek et al., 1992), brain in horses (Ross et al., 1993), kidney in rats, rabbits, and sheep (Edrington et al., 1995; Gumprecht et al., 1995; Voss et al., 1989), and esophagus in rats and pigs (Casteel et al., 1993; Lim et al., 1996). Many grainbased human foods have been shown to be contaminated with fumonisin metabolites (Scott, 2012) and epidemiologic data has linked ingestion of corn contaminated with F. verticillioides to human esophageal cancer (Rheeder et al., 1992) and infant neural tube defects (Voss et al., 2011). Fumonisins have been shown to be hepatocarcinogenic in rats and mice (Gelderblom et al., 1988; Howard et al., 2001) and the International Agency for Research on Cancer (IARC) has designated fumonisin B1 in Group 2B meaning “possibly carcinogenic to humans.” More recently the fungus Aspergillus niger has also been shown to produce some fumonisin metabolites, which have been found in grapes, raisins, wine, and coffee (Scott, 2012).

Chemical Structure

Veterinary Toxicology. DOI: http://dx.doi.org/10.1016/B978-0-12-811410-0.00071-4 Copyright © 2018 Elsevier Inc. All rights reserved.

First isolated in 1988, the fumonisins are a group of structurally related compounds with the terminal carboxy group composed of propane-1,2,3-tricarboxylic acid involved in ester formation with the C-14 and C-15 hydroxy groups. The 20C chain base carries either 2-acetylamino or 2-amino-12,16-dimethyl-3,5,10,14,15, pentahydroxyicosane (Fig. 71.1). The structures of FB1 and FB2 have been shown to have the empirical formulas of C34H59NO15 and C34H59NO14, respectively, with the only difference being the hydroxyl group present at the C-10 position in FB1 (Bezuidenhout et al., 1988). Additional fumonisin metabolites have been isolated (including B3, B4, B5, B6, A1, and A2), but appear to occur in much lower concentrations than FB1 or FB2, and are considered less important at this time (Gelderblom et al., 1992).

Occurrence and Distribution Although fumonisin metabolites have been found in many different grains including barley, millet, oats, and wheat (Scott, 2012), they are most commonly detected in corn and corn-based foods. ELEM has long been associated with the consumption of moldy corn, and has been reported in most of the world’s continents. Specific cases of ELEM that were directly associated with fumonisincontaminated feed have been reported in South Africa and Egypt (Thiel et al., 1991), the United States (Ross et al., 1991; Wilson et al., 1990a), Brazil (Sydenham et al., 1992), Hungary (Bela and Endre, 1996), Spain (Cerrillo et al., 1996), New Caldonia (Bailly et al., 1996), Mexico (Rosiles et al., 1998), Iran (Raoofi et al., 2003),

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1004 SECTION | XV Mycotoxins

OH

OR

OH CH3 Fumonisin B1

CH3 OR

CH3 OH

NH2

OH

OR

OH CH3 Fumonisin B2

CH3 OR

CH3

NH2

OH CH3OH Sphingosine NH2

OH CH3OH Sphinganine NH2 R=

C O

COOH COOH

FIGURE 71.1 The structure of fumonisin B1, fumonisin B2, sphingosine, and sphinganine.

Argentina (Giannitti et al., 2011) and Serbia (Jovanovi´c et al., 2015). Additionally cases of PPE have been associated with fumonisin-contaminated feeds in the United States (Harrison et al., 1990; Osweiler et al., 1992), Brazil (Sydenham et al., 1992), Hungary (Fazekas et al., 1998), and Thailand (Patchimasiri et al., 1998).

PHARMCOKINETICS/TOXICOKINETICS The pharmacokinetics of fumonisin B1 have been examined in several species including rats, pigs, cattle, laying hens, and primates (Martinez-Larranaga et al., 1999; Prelusky et al., 1994; Prelusky et al., 1995; Richard et al., 1996; Shephard et al., 1995; Vudathala et al., 1994). In general, fumonisin is rapidly absorbed following intravenous or intraperitoneal administration and is eliminated in both the feces and urine. Levels are undetectable by 24 h after dosing in virtually all species and significant concentrations (residues) have not been found in muscle, milk, or eggs. Following oral dosing, very little fumonisin B1 is typically found in the serum of animals indicating low bioavailability.

More specifically, the toxicokinetics of radiolabeled fumonisin B1 were examined after intragastric (0.5 mg fumonisin B1/kg) or intravenous (0.4 mg fumonisin B1/kg) administration to bile-cannulated and noncannulated pigs (Prelusky et al., 1994). Fumonisin-derived radioactivity was not detected in the plasma of pigs dosed intravenously after 180 min in the noncannulated group, or after 90 min in the cannulated group. Urinary excretion began within 3 h of administration and virtually ended after 8 h, accounting for only a small amount of administered toxin. Fecal excretion of fumonisin persisted for 48 h. The excretion in the intravenously dosed group occurred primarily via the bile, with biliary excretion greatest during the first 4 h, but persisting for 24 36 h. Plasma radioactivity in intragastrically dosed pigs was first detected 30 45 min after dosing, with maximal activity present between 60 and 90 min. As reflected in plasma and elimination data, systemic bioavailability of the dose ranged from 3% to 6%. Excretion of the fumonisin occurred primarily via feces, with only trace amounts excreted via urine or bile. At 72 h after administration, tissue radioactivity was highest in the liver, kidney, and large intestine in all

Fumonisins Chapter | 71

groups. Intragastrically dosed groups had 10 20 fold lower tissue concentrations than did intravenously dosed groups, and only intravenously dosed groups had measurable radioactivity in brain, lung, and adrenal. Thus, it seems that liver and kidney are the primary organs of fumonisin metabolism and excretion in the pig, and that enterohepatic circulation prolongs the persistence of fumonisin in the body. The toxicokinetics of fumonisin B1 in horses has not been evaluated. When pigs were fed fumonisin at daily concentrations between 50 and 500 μg of fumonisin B1 per kg of body weight for the last 5 months before slaughter, no muscle or kidney residues were detected (Liguoro et al., 2004). Fumonisin B1 was not detected in the eggs from laying hens following either intravenous or oral administration (Vudathala et al., 1994). Although negligible concentrations have been shown to cross the mammary barrier (Spotti et al., 2001), the toxin was not detected in milk from cattle that consumed a diet containing fumonisins (Richard et al., 1996). Therefore, it appears that fumonisin residues in meat, milk, or eggs do not represent a hazard or food safety concern for humans consuming these products.

MECHANISM OF ACTION Sphingolipid Alterations Fumonisins are structurally related to sphingosine, the major long-chain base backbone of cellular sphingolipids (Fig. 71.1). They are competitive inhibitors of sphinganine and sphingosine N-acyltransferase (also known as ceramide synthase), key enzymes in the de novo sphingolipid biosynthetic pathway (Fig. 71.2). These N-acyltransferase enzymes are responsible for catalyzing the acylation of sphinganine and the reutilization of sphingosine derived from sphingolipid turnover. This inhibition by fumonisin has been characterized in vitro using liver and brain microsomes, as well as in intact mammalian cells in culture (hepatocytes, neurons, renal cells, and macrophages) (Merrill et al., 1995). Fumonisin B1 blocks the incorporation of radiolabeled serine into the sphingoid base backbone of ceramides and complex sphingolipids and prevents the conversion of sphinganine to sphingosine via addition of the 4,5 trans double bond, which occurs after acylation of sphinganine. Fumonisin also blocks reacylation of sphingoid bases (primarily sphingosine) released by hydrolysis of more complex sphingolipids (Merrill et al., 1995). Sphingolipids are located in cellular membranes, lipoproteins (especially low-density lipoproteins), and other lipid-rich structures. Complex sphingolipids are critical for the maintenance of membrane structure, particularly microdomains such as caveolae. They also serve as

1005

Palmitoyl-CoA + Serine

3-ketosphinganine 3-ketosphinganine reductase

Sphinganine Acyl-CoA

Dihydroceramide

CoA

Sphingosine

Ceramide (sphingosine + fatty acid) CoA

Sphingomyelins

Acyl-CoA

Glycosphingolipids (cell membrane turn over)

: Pathway blocked by fumonisins FIGURE 71.2 The effects of fumonisin on the sphingolipid biosynthetic pathway.

binding sites for extracellular matrix proteins as well as for some microorganisms, microbial toxins, and viruses, and regulate the behavior of growth factor receptors (Merrill and Sweeley, 1996). Complex sphingolipids function as precursors for second messengers that mediate cell responses to growth factors, cytokines (including tumor necrosis factor-α), differentiation factors, and 1,25-dihydroxy-vitamin D3. Therefore, sphingolipids are involved in the regulation of cell growth, cell to cell communication, differentiation, and neoplastic transformation (Hannun and Bell, 1989). This enzyme inhibition by fumonisin produces a disruption of sphingolipid metabolism resulting in increased sphinganine and sphingosine along with a decrease in complex sphingolipids in the serum and tissues of animals (Wang et al., 1991). These elevations in concentrations of sphinganine and sphingosine have also been observed in vivo in several species including pigs, horses, and calves (Goel et al., 1996; Mathur et al., 2001; Riley et al., 1993; Smith et al., 1999; Smith et al., 2000). This disruption of sphingolipid metabolism is generally accepted as the probably mechanism of fumonisin toxicity; however, only in pigs has the pathophysiology been definitively determined. PPE has been shown to be a direct result of acute leftsided heart failure related to an increase in plasma and myocardial sphinganine and sphingosine concentrations (Constable et al., 2000; Smith et al., 1999, 2000). Sphingosine is an important intracellular second

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messenger that inhibits L-type calcium channels in myocardial cells, thereby decreasing sarcoplasmic reticulum Ca21-induced Ca21-release and cardiac contractility (McDonough et al., 1994; Webster et al., 1994). As sphingosine concentrations begin to increase in pigs that consume fumonisin, myocardial calcium channels are blocked and contractility begins to decrease (Constable et al., 2000; Smith et al., 2000). Ultimately this decrease in cardiac contractility causes acute left ventricular failure and pulmonary edema (Fig. 71.3). The mechanism of ELEM may also be a direct result of fumonisin-induced increases in sphingosine concentrations. Cardiovascular dysfunction following fumonisin administration in horses has been demonstrated (Smith et al., 2002). This study reported an association between neurologic signs, increased serum and myocardial sphingosine concentrations, and cardiovascular depression in fumonisin-treated horses. At necropsy, horses with

Fumonism consumption

Inhibits sphingosine N-acyltransferase

↑ tissue (sphingosine)

Inhibition of L-type Ca2+ channel

Myocardium

↓ sarcoplasmic reticulum Ca2+-induced Ca2+-release

↓ Myocardial contractility

Left-sided heart failure

Pulmonary edema FIGURE 71.3 Mechanism of fumonisin-induced pulmonary edema in swine. Fumonisin inhibition results in increased tissue sphingosine and sphinganine concentrations. The increased sphingosine concentrations inhibit the L-type Ca21 channels of cardiac myocytes resulting in decreased myocardial contractility. This decrease in contractility results in acute left-sided heart failure and pulmonary edema.

leukoencephalomalacia have histologic evidence of cerebral edema in the brain. Another study reported that fumonisin-treated horses also have elevated protein, albumin, and IgG levels in cerebrospinal fluid samples (Foreman et al., 2004). Taken together, these findings indicate that fumonisin toxicity in horses is associated with the development of vasogenic cerebral edema as a direct result of increased blood brain barrier permeability. Horses are dependent on autoregulation of cerebral blood flow when they lower their head to graze. Because of gravitational forces, distal carotid artery pressure can increase tremendously when the animal bends to eat or drink. However, this rise in carotid pressure does not create a significant increase in cerebral blood flow due the constriction and dilation of cerebral arterioles, which maintain normal cerebral blood pressures (Faraci and Heistad, 1990). It has been shown that L-type calcium channels are the primary mediators of vascular tone in these cerebral arterioles (Michelakis et al., 1994). Therefore, it has been hypothesized that fumonisininduced increases in sphingosine concentrations inhibit the calcium channels in cerebral arterioles leading to the inability to maintain normal cerebral blood pressure and vasogenic cerebral edema. This hypothesis requires further research to be proven definitively.

TOXICITY Fumonisin has been shown to cause liver damage in multiple species including pigs, horses, cattle, rabbits, and primates (Gumprecht et al., 1995; Haschek et al., 1992; Jaskiewicz et al., 1987; Osweiler et al., 1993; Ross et al., 1993; Voss et al., 1989) as well as species-specific target organ toxicity, such as lung in pigs (Haschek et al., 1992), brain in horses (Ross et al., 1993), kidney in rats, rabbits, and sheep (Edrington et al., 1995; Gumprecht et al., 1995; Voss et al., 1989), and esophagus in rats and pigs (Casteel et al., 1993; Lim et al., 1996). This chapter will focus on fumonisin toxicity in pigs and horses since they exhibit the most common clinical poisonings dealt with in veterinary medicine, however cattle and poultry will be briefly discussed.

Spontaneous and Experimental Fumonisin Toxicosis in Swine In early research prior to the initial isolation and characterization of fumonisins, F. verticillioides culture material was reported toxic to swine (Kriek et al., 1981). In that experiment, three pigs were fed F. verticillioides culture material grown on corn. Two of the three pigs fed the culture material in this study died within 5 days of pulmonary edema. The third pig was fed culture material for 89 days and was then killed following a period of feed refusal.

Fumonisins Chapter | 71

The 1989, corn crop in many Midwestern and Southeastern parts of the United States was heavily infected with F. verticillioides, and contaminated screenings fed to animals led to fatal outbreaks of PPE (Harrison et al., 1990; Osweiler et al., 1992). This syndrome was also reproduced experimentally with contaminated corn screenings and purified fumonisin B1 (Harrison et al., 1990; Osweiler et al., 1992). Lung and liver are the major target organs of fumonisin toxicosis in pigs; however, other organs have been reported to be affected. Pigs that ingest fumonisin at concentrations high enough to cause pulmonary edema usually die after about 4 days in field cases (Osweiler, 1992) and after 3 6 days of fumonisin exposure experimentally (Gumprecht et al., 1998; Haschek et al., 1992; Motelin et al., 1994). Pigs that survive chronic exposure to high doses of fumonisin without developing pulmonary edema typically demonstrate hepatic disease with anorexia, weight loss, and generalized icterus (Colvin et al., 1993; Osweiler et al., 1992). Hepatic toxicity occurs at doses significantly lower than those necessary to cause pulmonary edema (Colvin et al., 1993; Motelin et al., 1994).

Fumonisins in Swine-Pulmonary Effects Pulmonary edema (Fig. 71.4) has been reported in pigs fed naturally contaminated fumonisins-containing food, fumonisin-containing culture material or following IV administration of fumonisin (Harrison et al., 1990; Haschek et al., 1992; Motelin et al., 1994; Osweiler et al., 1992). Reported concentrations of fumonisin required to produce pulmonary edema have been variable, presumably due to variability in susceptibility among exposed animals (Table 71.1). However, other constituents in the diet and analytical detection related to the ability to

FIGURE 71.4 Lung from a pig fed fumonisin-containing culture material at a dose of 20 mg fumonisin B1 per kg of body weight for 4 days. Pulmonary edema is characterized by severe widening of the interlobular septa.

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extract fumonisin from different matrices could account for some of the variability. In addition, some reports have reported only the concentration of fumonisin B1 associated with the development of pulmonary edema, while others have reported both fumonisin B1 and fumonisin B2. Fumonisin B2 usually occurs at about 30% of fumonisin B1 in naturally-contaminated corn, and is generally considered to be equitoxic to fumonisin B1 (Ross et al., 1994). Reported doses that induced pulmonary edema in swine include 100 ppm of fumonisin B1 and fumonisin B2 in naturally-contaminated corn (Motelin et al., 1994), 16 mg fumonisin B1/kg/day as fumonisin-containing culture material (Colvin et al., 1993), and 20 mg fumonisin B1/kg/day as culture material (Gumprecht et al., 1998). Fumonisin-induced pulmonary edema has also been reported with naturally-contaminated corn (330 mg of fumonisin B1 per kg of feed) in Hungary (Fazekas et al., 1998), Brazil (Sydenham et al., 1992), and Thailand (Patchimasiri et al., 1998). Another study has suggested that even lower concentrations of fumonisins may be able to induce pulmonary edema in swine (Zomborszky et al., 2000). Fumonisin B1 was fed added to the feed of weaned pigs at doses of 0, 10, 20, and 40 ppm for 4 weeks as fumonisin-containing culture material (five pigs per group). Computed tomography (CT) of the lungs and magnetic resonance imaging of the brains were performed prior to the study and at 2 and 4 weeks of fumonisin feeding. Histopathology was also done at the time of necropsy (4 weeks). The results of this study showed that all five pigs fed fumonisin B1 at 40 ppm developed “severe” pulmonary edema as assessed by CT and histopathology. Two of the five pigs fed fumonisin B1 at 20 ppm had “severe” pulmonary edema while two other pigs in the group had “mild” edema. In the 10 ppm group, three of the five pigs were reported to have “mild” pulmonary edema. Magnetic resonance studies of the brain were not able to identify any significant changes during the course of the study in any group. Clinical signs associated with the development of pulmonary edema consistently begin 3 6 days after initiation of exposure to a high concentration of fumonisins. These include dyspnea and open mouthed breathing, increased respiratory rate, cyanosis of skin and mucous membranes, inactivity and sudden death (Osweiler et al., 1992). Pigs usually die within a few hours after the onset of definitive respiratory distress. Histologically, pulmonary edema is present by day three of fumonisin exposure (Gumprecht et al., 1998) and is characterized by interstitial edema around airways and vessels, in interlobular and subpleural connective tissues, and in alveolar interstitium (Gumprecht et al., 1998; Harrison et al., 1990; Haschek et al., 1992; Osweiler et al., 1992). Lymphatics are dilated and alveolar edema is often present. Fluid is also present within the thoracic cavity.

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TABLE 71.1 Effect of Fumonisin in Pigs Number of Animals

Dose & Route

Duration

Toxic Effects

Reference

Experimental Studies Using Purified Fumonisin 4 pigs

0.174 0.4 mg FB1 per kg of BW/day, IV

4 8 days

2 high dose pigs developed pulmonary edema

Harrison et al. (1990)

2 pigs

0.88 1.15 mg FB1 per kg of BW/day, IV

1 pig for 4 days (0.88 mg/kg/ day)—2nd pig for 9 days (1.15 mg/kg/day)

Mild interstitial pulmonary edema histologically in 1 pig; hepatic lesions; pancreatic lesions

Haschek et al. (1992)

3 pigs

4.5 6.6 mg FB1 per kg of BW/day in feed

5 15 days

2 of the 3 pigs developed severe pulmonary edema; hepatic lesions and mild pancreatic lesions noted

Haschek et al. (1992)

2 pigs

0.4 mg FB1 per kg of BW/day, IV

12 14 days

Elevated AST, GGT, bilirubin; liver lesions

Osweiler et al. (1992)

8 pigs

10 ppm FB1 added to the diet-fed ad libitum

8 weeks

Decreased weight gain

Rotter et al. (1996)

1 mg FB1 per kg of BW/day, IV

4 days

6 pigs

Elevated cholesterol Decrease in left ventricular contractility and mild pulmonary edema

Smith et al. (2000)

Experimental Studies Using Fumonisin-Containing Culture Material 6 pigs

Feeds containing 100 and 190 ppm FB1 fed ad libitum

100 ppm diet-fed ad libitum for 7 days followed by 190 ppm diet for 83 days

Elevated AST, ALP, GGT; nodular hyperplasia of the liver; histologic esophageal lesions

Casteel et al., 1993

4 pigs

Feed containing 200 ppm of FB1 fed ad libitum

Up to 43 days

Elevated bilirubin, AST, cholesterol; hepatic lesions; decreased weight gain

Colvin et al. (1993)

7 pigs

16 64 mg FB1 per kg of BW/day as oral gavage

3 5 days

All pigs developed pulmonary edema within 5 days

Colvin et al. (1993)

3 pigs

4 16 mg FB1 per kg of BW/day as oral gavage

up to 45 days

Severe hepatic disease; icterus; elevated liver enzymes; no pulmonary edema

Colvin et al. (1993)

11 pigs

Feeds containing 100, 160, and 190 ppm FB1 fed ad libitum

6 pigs were fed 100 ppm for 10 days then 190 ppm for up to 83 days; 5 pigs were fed 100 ppm for 5 days and then 160 ppm for up to 205 days

Nodular hyperplasia of the liver; Elevated AST, ALP, GGT, bilirubin

Casteel et al. (1994)

Right ventricular hypertrophy; medial hypertrophy of the small pulmonary arteries

6 pigs

feed containing 47 ppm FB1 fed ad libitum

28 days

Decreased feed consumption; Elevated AST, GGT, ALP, and creatinine; hepatic and renal lesions; medial hypertrophy of the pulmonary arteries

Harvey et al. (1996)

10 pigs

20 mg of FB1 per kg of BW/day, in feed

7 days

Pulmonary edema and cardiovascular abnormalities

Smith et al. (1996a,b)

2 pigs

14.5 and 16 mg of FB1 per kg of BW/day, in feed

4 days

Severe pulmonary edema; renal lesions (Continued )

Fumonisins Chapter | 71

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TABLE 71.1 (Continued) Number of Animals

Dose & Route

Duration

Toxic Effects

Reference

24 pigs

20 mg of FB1 per kg of BW/day, in feed

Up to 5 days-some pigs were euthanized each day of the study

All 12 pigs euthanized on days 0, 1 and 2 had no lesions

Gumprecht et al. (1998)

2/5 day 3 pigs and all of the day 4 and 5 pigs had pulmonary edema Elevations in liver enzymes and hepatic lesions seen on day 2—bile acids first liver parameter to increase

6 pigs

20 mg of FB1 per kg of BW/day, in feed

5 6 days pigs were euthanized as they developed pulmonary edema

Increased pulmonary artery pressure, pulmonary artery wedge pressure and decreased cardiac output, heart rate, and mean arterial pressure

Smith et al. (1999)

5 pigs

Feed containing 40 ppm FB1 fed ad libitum

4 weeks

No clinical signs-gross pulmonary edema evident at necropsy

Zomborszky et al. (2000)

5 pigs

Feed containing 20 ppm FB1 fed ad libitum

4 weeks

2 of the 5 pigs had gross pulmonary edema—2 additional pigs had mild evidence of edema present histologically

Zomborszky et al. (2000)

4 pigs

Feed containing 20 ppm FB1 fed ad libitum

28 days

3 of the 4 pigs had mild pulmonary edema evident histologically

Zomborszky et al. (2000)

7 pigs

20 mg of FB1 per kg of BW/day, in feed

3 days

Decreased left ventricular contractility; heart rate, cardiac output, and mechanical efficiency of the left ventricle

Constable et al. (2000)

Experimental Studies Using Naturally-Contaminated Corn Screenings 6 pigs

105 155 ppm FB1 in corn screenings fed ad libitum

Up to 28 days

3 of 6 pigs developed pulmonary edema; liver and pancreatic lesions

Harrison et al. (1990)

8 pigs

92 ppm FB1 in corn screenings fed ad libitum

Up to 21 days

6 of 8 pigs developed pulmonary edema; the remaining 2 pigs were icteric with increased liver enzymes

Osweiler et al. (1992)

5 pigs

175 ppm (FB1 1 FB2) as corn screenings mixed in a complete ration

Up to 14 days

3 of 5 pigs developed pulmonary edema; hepatotoxicity; decreased weight gain

Motelin et al. (1994)

5 pigs

101 ppm (FB1 1 FB2) as corn screenings mixed in a complete ration

Up to 14 days

Elevated GGT, ALT, AST, ALP, and bilirubin; liver lesions; decreased weight gain

Motelin et al. (1994)

5 pigs

39 ppm (FB1 1 FB2) as corn screenings mixed in a complete ration

14 days

Histologic liver lesions

Motelin et al. (1994)

5 pigs

39 ppm (FB1 1 FB2) as corn screenings mixed in a complete ration

14 days

Histologic liver lesions

Motelin et al. (1994) (Continued )

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TABLE 71.1 (Continued) Number of Animals

Dose & Route

Duration

Toxic Effects

Reference

Reported Fumonisin Concentrations From Naturally-Occurring Outbreaks 34 pigs from 2 farms

105 155 ppm FB1

Unknown

Lethal pulmonary edema

Harrison et al. (1990)

16 pigs from 9 farms

All feed samples associated with pulmonary edema contained $20 ppm FB1 to a maximum of 330 ppm FB1

Unknown

Lethal pulmonary edema

Osweiler et al. (1992)

In ultrastructural studies using immersion fixed lungs, the endothelium was found to be swollen, vacuolated, and sometimes missing in pigs with pulmonary edema (Haschek et al., 1992). Additional studies using intravascularly perfused lungs (to allow better examination of the vascular system) demonstrated accumulations of fragmented membranous material in the cytocavitary region of endothelial cells (Gumprecht et al., 1998).

Fumonisins in Swine-Hepatic Effects Hepatic changes in pigs exposed to fumonisins include elevation of liver associated enzyme activities, altered clinical chemistries, changes in sphingolipid parameters, and morphological alterations. In pigs, hepatic toxicity occurs prior to the development of pulmonary edema, and alterations are time and dose-dependent (Motelin et al., 1994). Increased activities of serum enzymes such as alkaline phosphatase (ALP), aspartate aminotransferase (AST) and gamma glutamyl transpeptidase (GGT) and concentrations of total bilirubin, bile acids, and cholesterol have been reported as early as 1 day after the initiation of fumonisin exposure (Colvin et al., 1993; Gumprecht et al., 1998; Harrison et al., 1990; Haschek et al., 1992; Motelin et al., 1994; Osweiler et al., 1992). These alterations reflect hepatocyte damage as well as altered hepatic function. Morphologic alterations are dose related and progressive with continued ingestion of fumonisins. Following short term exposure, changes include hepatic cord disorganization, cytoplasmic vacuolation, apoptosis, scattered necrosis, and increased cell proliferation (Gumprecht et al., 1998; Harrison et al., 1990; Motelin et al., 1994; Osweiler et al., 1992). Histologic alterations were

observed as early as 2 days after the initiation of treatment with a lethal dose (Gumprecht et al., 1998), and at a concentration as low as 23 ppm when fed for 14 days (Motelin et al., 1994). Long-term fumonisin exposure can result in fibrosis or development of hyperplastic nodules in the liver (Casteel et al., 1993; Harrison et al., 1990). Ultrastructurally, large accumulations of proteinaceous and membranous material were observed in the space of Disse in pigs that developed fumonisin-induced pulmonary edema (Haschek et al., 1992). Hepatocytes lost microvilli from their sinusoidal face while numerous Kupffer cells contained multilamellar bodies.

Fumonisins in Swine-Cardiovascular Effects Fumonisins have been shown to decrease left ventricular contractility, heart rate, cardiac output, mean arterial pressure, arterial and mixed venous blood O2 tensions, and systemic oxygen delivery, while increasing mean pulmonary artery pressure, oxygen consumption, and oxygen extraction ratio in swine (Constable et al., 2000; Smith et al., 1996a,b, 1999, 2000). The decrease in cardiac contractility leads to acute left ventricular failure and pulmonary edema in pigs exposed to high concentrations of fumonisin in feed. Chronic exposure to lower levels of fumonisin leads to the development of right ventricular hypertrophy and medial hypertrophy of the small pulmonary arteries in pigs, likely a result of pulmonary hypertension (Casteel et al., 1994).

Fumonisins in Swine-Immunologic Effects Fumonisins have also been shown to predispose pigs to respiratory disease. In one case-control study, swine farms

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with $ 20 ppm of fumonisin in the feed were at significantly greater risk for pneumonia as compared to farms with low fumonisin concentrations (Bane et al., 1992). As the concentration of fumonisin in the feed increased, the risk of respiratory disease continued to increase. Later it was shown that exposure to fumonisins depressed pulmonary intravascular macrophage function, and pigs exposed to this toxin had decreased pulmonary clearance of blood borne particulates and bacteria when compared to control animals (Smith et al., 1996c). In a more recent study, exposure to fumonisin exacerbated respiratory disease in a Pasteurella multocida challenge model (Halloy et al., 2005). Pigs that were fed 0.5 mg of FB1 per kg of body weight/day for 7 days had delayed growth, increased coughing and more severe lung lesions that control pigs. In another study, pigs fed FB1 at 20 ppm and exposed to Mycoplasma hyopneumoniae had more severe lung lesions as assessed by CT and histopathology as compared to pigs exposed to Mycoplasma but diets without FB1 (Po´sa et al., 2013). Therefore at levels well below those needed to cause hepatic lesions or pulmonary edema, fumonisins are likely to reduce growth rates and increase disease in pigs.

Fumonisin Toxicosis in Horses-Historical Several outbreaks of a neurologic disease in horses occurred in the United States in the early 1900s with thousands of deaths reported in several states. The earliest citation of neurologic deaths associated with the feeding of contaminated corn was from Maryland (MacCallum and Buckley, 1902). The condition was commonly referred to as “cerebrospinal meningitis” and presented with fairly characteristic signs. The duration of disease varied from a few hours to a week, and the brains from affected horses had “softened” areas in the cerebrum involving only the white matter. Additional outbreaks were subsequently reported from Kansas, Iowa, Mississippi, and North Carolina. A similar disease, described as “epizootic cerebritis” had been encountered in 1891, however it is not known whether this was associated with corn (Butler, 1902). When feed from an outbreak of “leucoencephalitis” in Kansas was fed to a horse, it died after developing neurologic signs (Butler, 1902). At necropsy, the left cerebral hemisphere was “soft to the touch, and when cut through, the white matter was broken down extensively, nearly the entire hemisphere being involved.” Several attempts were made to identify an infectious agent in the brain of affected horses and all were negative (MacCallum and Buckley, 1902). The authors concluded that a toxic etiology was likely. In Central Illinois, more than 5,000 horses died during the winter of 1934 35 from a syndrome referred to as “cornstalk disease” (Graham, 1935). Brain tissue

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suspensions and filtrates were inoculated into laboratory animals, however no infectious etiology could be identified. Graham then placed 8 horses into a field containing cornstalks in Rantoul, Illinois. Two of these animals died, 23 and 26 days after being placed in the field (Graham, 1936). These outbreaks were similar to those reported in 1893 and 1914, which had occurred following a summer drought (Graham, 1936). Neurologic deaths with similar lesions were also reported in Iowa during the winter of 1914 and in the spring of 1935 (Schwarte et al., 1937). Histologic examination of the brain revealed no evidence of infectious agents. This “syndrome” was then subsequently reproduced by feeding moldy corn and corn fodder to five horses (Schwarte et al., 1937). A similar disease syndrome was identified and confirmed by feeding trials in Egypt (Badiali et al., 1968; Wilson and Maronpot, 1971) and in South Africa (Marasas et al., 1976). At necropsy, these studies were able to consistently demonstrate swelling of the cerebral hemispheres and flattening of the overlying gyri. On coronal sections, there were cavities of varying sizes with liquefactive necrosis of subcortical white matter in one or both cerebral hemispheres (Fig. 71.5). There was also scattered multifocal hemorrhages in the surrounding white matter (Marasas et al., 1976; Haliburton et al., 1979). Based on these findings, Marasas et al. (1976) coined the term “ELEM” as a distinct clinical and morphologic syndrome in horses associated with the feeding of corn. F. verticillioides was later isolated from corn collected from field outbreaks in Egypt, and leukoencephalomalacia was subsequently reproduced in donkeys fed corn inoculated with the fungus (Wilson and Maronpot, 1971). In South Africa however, samples of corn inoculated with F. verticillioides produced liver damage and icterus in several horses and donkeys, but not brain lesions

FIGURE 71.5 A cross section of a cerebral hemisphere from a horse demonstrating liquefactive necrosis of the white matter typical of equine leukoencephalomalacia.

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(Kellerman et al., 1972). Marasas et al. (1976) then produced a batch of culture material using a strain of F. verticillioides from an outbreak of leukoencephalomalacia to produce liver damage and neurologic disease in a horse. It was then concluded that both hepatic disease and ELEM were manifestations of the same toxicosis, with different clinical syndromes occurring depending on toxin dose and length of exposure. Following the isolation and purification of FB1 in the late 1980s, ELEM was experimentally induced by the administration of purified toxin (Kellerman et al., 1990; Marasas et al., 1988).

Spontaneous and Experimental Fumonisin Toxicosis in Horses Since the discovery of fumonisin as the causative agent of ELEM, many more disease outbreaks associated with the feeding of corn have been reported on virtually all continents of the world (Bailly et al., 1996; Bela and Endre, 1996; Binkerd et al., 1993; Cerrillo et al., 1996; Christley et al., 1993; Giannitti et al., 2011; Jovanovi´c et al., 2015; Rosiles et al., 1998; Ross et al., 1991, 1993; Uhlinger, 1991; Wilkins et al., 1994; Wilson et al., 1990a). Purified fumonisin B1 has induced ELEM when administered orally (Kellerman et al., 1990) and intravenously (Foreman et al., 2004; Marasas et al., 1988; Laurent et al., 1989). Purified fumonisin B2 has also induced ELEM when given orally (Ross et al., 1994). Fumonisin B1 is considered to be the primary cause of ELEM however, as fumonisin B2 is usually present in concentrations that are 20% 40% of fumonisin B1 (Ross et al., 1991). Although ELEM has occurred in horses eating commercial feedstuffs (Ross et al., 1991; Wilson et al., 1990b), the feeding of corn screenings has been more frequently associated with ELEM, because fumonisin concentrations are much higher in screenings than in whole kernels of corn (Binkerd et al., 1993). Fumonisin B1 also appears to survive the pelleting process for equine feeds (Ross et al., 1991). Leukoencephalomalacia has been reproduced with intravenous administration of fumonisin B1 in three separate studies (Foreman et al., 2004; Laurent et al., 1989; Marasas et al., 1988). Marasas et al. (1988) administered 0.125 mg fumonisin B1/kg of body weight, IV, q24h, which produced ELEM in 9 days. Laurent et al. (1989) administered 0.1 mg fumonisin B1/kg of body weight, IV, q24h for 16 days followed by 0.2 mg/kg/day for 2 additional days. Leukoencephalomalacia was induced in 18 days. Foreman et al. (2004) administered 0.05, 0.1 or 0.2 mg fumonisin B1/kg of body weight IV q24h to 10 horses and all developed neurologic signs and were euthanized between days 4 and 12 of the study. In contrast, horses dosed with 0.01 mg fumonisin B1/kg of body

weight for 28 days in this study did not develop neurologic signs. Purified fumonisin B1 has also been administered orally in other studies (1.25 or 2.5 mg fumonisin B1/kg of body weight, PO, q24h), producing mild edema of the brain stem and hepatic disease in 11 12 days in two horses. In a subsequent study, animals were fed 0.6 4.0 mg fumonisin B1/kg of body weight, PO, q 24 h for 33 or 35 days, producing hepatotoxicity and neurologic signs starting on days 22 and 24 in two weanling horses (Kellerman et al., 1990). Doses of fumonisin reported from naturally occurring cases of fumonisin have varied (Table 71.2). One field report calculated that the ingestion of 0.6 2.1 mg fumonisin B1/kg of body weight would induced ELEM in 24 28 days (Wilson et al., 1990b). Another study found that leukoencephalomalacia was associated with ingestion of feed containing fumonisin B1 concentrations greater than 10 ppm, and concluded that feed with fumonisin B1 concentrations greater than 10 ppm was not safe to be fed to horses (Ross et al., 1991).

Neurologic and Hepatic Effects in Horses Several reports have considered ELEM and hepatotoxicity to be two separate syndromes associated with fumonisin toxicity in horses, with the terms “classic neurotoxic syndrome” and “hepatic syndrome” being used (McCue, 1989). However it appears more likely these are not true “distinct” syndromes but are related to the concentration of fumonisin in the feed, the duration of toxin consumption, and the tolerance of the individual horse to fumonisin. In some outbreaks, horses have died from ELEM while other horses have died from hepatotoxicity, and occasionally individual horses exhibiting both neurologic and hepatic signs have been described. Reported clinical signs associated with hepatic disease include icterus, mucous membrane petechiae, and swelling of the lips or muzzle (Ross et al., 1993; Uhlinger, 1991). Ross et al. (1993) described an experimental study where one horse died acutely with “mild encephalopathy and hepatic necrosis” after 9 days of fumonisin exposure whereas two other horses died after 75 and 78 days of ELEM. The horse that died on day 9 showed neurologic signs prior to death (“visual impairment, mild ataxia, and slight head tremors”) and had histologic evidence of leukoencephalomalacia at necropsy, however his death was primarily attributed to hepatotoxicity. This study lead to a common generalization that high doses of fumonisin were likely to induce hepatotoxicity, whereas lower doses of toxin over a longer period of time were necessary to induce ELEM (McCue, 1989; Plumlee and Galey, 1994). However in other experimental studies, intravenous administration of fumonisin B1 induced ELEM in 9 days (Marasas et al., 1988) and 18 days (Laurent et al., 1989).

Fumonisins Chapter | 71

1013

TABLE 71.2 Effect of Fumonisin in Horses Number of Animals

Dose & Route

Duration

Toxic Effects

Reference

Leukoencephalomalacia

Marasas et al. (1988)

Experimental Studies Using Purified Fumonisin One horse

0.125 mg/kg/day, IV

7 treatments over 9 days

Elevated AST, GGT

Marasas et al. (1988)

One horse

2.5 mg/kg/day, PO (by gavage)

6 doses over 11 days

Severe hepatosis

One horse

1.25 mg/kg/day, PO (by gavage)

6 doses over 11 days

Mild hepatosis

One horse

0.1 mg/kg/day, IV and 0.2 mg/kg/day, IV

0.1 mg/kg dose given for 16 days then 0.2 mg/kg dose given for 2 additional days

Leukoencephalomalacia

Laurent et al. (1989)

One horse

1.25 mg/kg to 4.0 mg/kg, PO

20 doses given over 35 days

Leukoencephalomalacia

Kellerman et al. (1990)

One horse

1.0 mg/kg to 4.0 mg/kg, PO

20 doses given over 33 days

Leukoencephalomalacia

13 horses

0.01 to 0.20 mg/kg, IV

Up to 10 days

Leukoencephalomalacia at 0.10 and 0.20 mg/kg—hepatic toxicity only at lower doses

Smith et al. (2002); Foreman et al. (2004)

Elevated AST, GGT

Elevated AST, GGT

Elevated AST

Elevated AST, GGT

Marasas et al. (1988)

Kellerman et al. (1990)

Experimental Studies Using Fumonisin-Containing Culture Material 2 horses

Diet contained 19 ppm FB1 and was fed ad libitum

27 days

None

Schumacher et al. (1995)

2 horses

Diet contained 200 ppm FB1 and was fed ad libitum

12 and 16 days

Leukoencephalomalacia

Schumacher et al. (1995)

2 diets containing 65 and 130 ppm FB1 and was fed ad libitum

265 ppm fed for 10 days

Leukoencephalomalacia

2130 ppm fed for an additional 17 days

Increased GGT

2 horses

Increased GGT

Schumacher et al. (1995)

Experimental Studies Using Naturally-Contaminated Corn Screenings 4 ponies

Diet contained 44 ppm FB1 and was fed ad libitum

10 97 days

2 horses died with ELEM on days 10 and 45 2 horses were normal after 97 days

Wang et al. (1992)

4 ponies

Diets contained between 1 and 22 ppm FB1 and were fed ad libitum

238 326 days

ELEM in 2 of 5 horses

Wilson et al. (1992)

5 ponies

Diet contained 8 ppm FB1 and was fed ad libitum

180 days

Mild histologic lesions in the brain and liver at necropsy

Wilson et al. (1992)

4 ponies

Diets contained between 1 and 88 ppm FB1 and were fed ad libitum

9 120 days

ELEM in all 4 ponies (day 9 120)—also severe hepatosis in 2 ponies

Ross et al. (1993)

Moderate to mild hepatosis in all 5 horses

(Continued )

1014 SECTION | XV Mycotoxins

TABLE 71.2 (Continued) Number of Animals

Dose & Route

Duration

Toxic Effects

Reference

Reported Fumonisin Concentrations From Naturally-Occurring Outbreaks 18 horses

37 122 ppm FB1

Unknown

ELEM confirmed in 14 horses

Wilson et al. (1990)

45 horses

8 126 ppm FB1

7 35 days

All cases had confirmed leukoencephalomalacia

Ross et al. (1991)

6 horses

370 ppm FB1 & 105 ppm FB2

Unknown

4 horses died with ELEM; 2 horses with neurologic signs apparently recovered

Wilkins et al. (1994)

1001 donkeys

4 29 ppm FB1

Unknown

many donkeys died of neurologic disease-ELEM confirmed in 3 cases

Rosiles et al. (1998)

7 horses

12.5 ppm FB1 and 5.3 ppm FB2

10 days

At least 7 horses died with ELEM—at least one other horse with neurologic signs recovered with feed was removed

Giannitti et al. (2011)

1001 horses

6.0 ppm FB1 and 2.4 ppm FB2

,30 days

21 horses developed neurologic signs—15 died within a month period

Jovanovi´c et al. (2015)

In a large study with varying doses of fumonisin, horses treated with higher doses developed leukoencephalomalacia (in 5 8 days), whereas horses that received lower concentrations developed primarily hepatic lesions without any evidence of neurotoxicity (Foreman et al., 2004). Therefore it can be concluded ELEM results from an acute exposure to feed containing high concentrations of fumonisin B1, while hepatotoxicity occurs with chronic ingestion of lower levels. Serum biochemical changes associated with fumonisin toxicity in horses have been predominantly related to hepatotoxicity (increased AST, Wang et al., 1992; increased AST and GGT, Laurent et al., 1989; Kellerman et al., 1990; increased GGT and SDH, Schumacher et al., 1995; increased AST, GGT, and ALP, Ross et al., 1993; increased AST, GGT, ALP, total bilirubin, and bile acids, Wilson et al., 1992; and “elevated liver enzymes,” Ross et al., 1994). The neurologic signs are usually summarized as sudden onset of one or more of the following: frenzy, aimless circling, head pressing, paresis, ataxia, blindness, depression, and hyperexcitability (Ross et al., 1991; Wilson et al., 1992). Other reports have stated that “the disease started with lack of appetite, followed by the disturbance of swallowing and chewing indicating the paralysis of cephalic and pharyngeal muscles. Paralysis of cephalic and cervical muscles spread to the muscles of the extremities and trunk. The animals moved with difficulties, tottering and ataxia developed. Signs of ‘blindness’ developed in one animal. At the final stage of disease, the affected animals lied down and died. In a comprehensive study, early neurologic signs included mild proprioceptive abnormalities, including hindlimb ataxia, delayed forelimb placing reactions, and decreased tongue tone and

movement (Foreman et al., 2004). These signs progressed over 12 48 h to become more readily apparent. Hindlimb and trunkal ataxia in particular became more apparent with time. A variety of behavioral changes were observed including depression, hyperesthesia, and intermittent dementia. All horses had intact menace and pupillary light responses at the time of death. Cerebrospinal fluid findings from horses with ELEM include elevations in protein concentration, albumin, and IgG concentrations and increased albumin quotients (Foreman et al., 2004). Cerebrospinal fluid red blood cell, leukocyte, and glucose concentrations along with creatine kinase activity are not altered in horses with neurologic disease. Along with the histopathologic findings, these cerebrospinal fluid changes indicate the presence of a vasogenic cerebral edema in horses with leukoencephalomalacia.

Fumonisin Toxicity in Cattle Adult beef cattle appear relatively resistant to fumonisin. Feeder calves fed a diet containing fumonisin concentrations up to 148 ppm for 31 days had only mild hepatotoxicity (Osweiler et al., 1993). Although it is tempting to speculate that cattle are able to break down the toxin, it has been shown that fumonisin is poorly metabolized by the rumen. Instead it is though that cattle have an increased tolerance to fumonisin because of differences in the mechanism of action. In milk-fed calves treated with purified fumonisin B1, the kidney was the target organ of toxicity (Mathur et al., 2001). However this study also demonstrated that sphingosine and sphinganine concentrations did not increase

Fumonisins Chapter | 71

in the serum and tissues of calves to the same degree that has been shown in pigs and horses. When a group of 26 dairy cattle were fed a ration containing 100 ppm of fumonisin for the first 70 days of their lactation period, they had a significant decrease in dry matter intake and a lower milk yield as compared to the control group (Diaz et al., 2000). Milk production averaged 7 kg lower in the group fed the ration containing fumonisin and there was a 13% decrease in feed intake over the duration of the study period. Therefore it has been recommended to avoid fumonisin concentrations higher than 30 ppm in the total ration of dairy cattle.

Fumonisin Toxicity in Poultry Fumonisins can be toxic to both chickens and turkeys with concentrations in the feed as low as 100 mg/kg causing decreased body weight gain, diarrhea, and hepatotoxicity (Bermudez et al., 1997; Ledoux et al., 1992). There has also been an association between F. verticillioides (the fungus that produces fumonisin) and an acute death syndrome recognized in young chicks called spiking mortality syndrome. It was initially hypothesized that fumonisins were directly cardiotoxic to poultry and were the cause of this syndrome; however, more recent research has suggested moniliformin (another F. verticillioides produced mycotoxin) is primarily responsible.

DIAGNOSIS AND TREATMENT In addition to pathologic findings in animals, diagnosis of fumonisin toxicosis typically relies on detecting the actual toxin in feed samples. Fungal culture of feeds has little value in diagnosing fumonisin toxicosis because some corn

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samples contain very high concentrations of toxin with low levels of fungus, while others samples have heavy growths of Fusarium fungus with little to no detectable fumonisin. This is partly because the fungus that produced fumonisin also produces other mycotoxins. Therefore, the definitive diagnosis of fumonisin toxicosis in animals must involve analyzing the feed for the presence of the actual toxin. Many diagnostic laboratories across the world offer assays to detect both fumonisin B1 and B2 in corn and feed samples. The two most commonly used methods for toxin detection are chromatography (HPLC) and immunologic (ELISA) assays. To date, there are no commercially available assays that detect fumonisin in serum or tissues of animals. Another assay that may be used more commonly in the future to diagnose fumonisin toxicosis in animals is the sphinganine-to-sphingosine ratio (Sa:So ratio). Because of the fumonisin-induced disruption of sphingolipid biosynthesis (Wang et al., 1992), the Sa:So ratio increases in the serum and tissues of pigs and horses exposed to fumonisin. It has been suggested that this assay could be used to diagnose fumonisin toxicosis when feed analysis is not possible. Sphinganine and sphingosine data may be available from enough pigs and horses in the near future to provide values for normal and affected animals. To date there have been no treatments described for either ELEM or PPE. Generally the onset of clinical signs is acute and the progression of disease is rapid for both syndromes. The most important treatment is to identify and remove the source of contaminated feed to prevent other animals from developing clinical signs. Guidelines for the maximum recommended levels of fumonisins in animal feeds have been published by the FDA Center for Veterinary Medicine (Table 71.3). It is

TABLE 71.3 Recommended Levels for Total Fumonisins (B1 and B2) in Animal Feeds Animal

Recommended Maximum Level of Total Fumonisins in Corn to be Used for Feed (ppm)

Recommended Maximum Level of Total Fumonisin in the Ration (ppm)

Horsea

5

1

20

10

60

30

100

50

Ruminant and poultry breeding stock

30

15

Catfish

20

10

Other animalse

10

5

Swine Ruminants

b

c

Poultry

d

a

Includes donkeys, asses, and zebras. Cattle, sheep, goats, and other ruminants that are .3 months of age and are being fed for slaughter. Turkeys, chickens, ducklings, and other poultry being fed for slaughter. d Includes lactating dairy cows, bulls, laying hens, and roosters. e Includes dogs and cats. Source: From the United States Food and Drug Administration, Center for Veterinary Medicine. b c

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important that livestock producers be aware of these guidelines and have their corn periodically tested for mycotoxins. Research has not yet found effective ways to decrease fumonisin concentrations in animal feedstuffs through processing or feed additives (i.e., binding agents). Corn containing significant levels of fumonisin should be discarded, diluted with corn containing lower concentrations of fumonisin, or fed to a less-sensitive species (i.e., ruminants or poultry interested for slaughter).

CONCLUDING REMARKS AND FUTURE DIRECTIONS Fumonisins remain an active research topic however most studies in the last few years have focused on the potential effects of this mycotoxin in humans. More animal studies are needed to further define the mechanism of neurotoxicity in horses. Studies also need to be done examining the long-term cardiovascular effects associated with lower doses of fumonisin exposure in swine and humans. High fumonisin concentrations seem to appear in the corn crop from the United States every 3 4 years depending on weather, so they will continue to be a toxin of high regulatory concern in the future. Veterinarians and toxicologists must be familiar with this mycotoxin and should be able to quickly recognize clinical signs and gross lesions associated with fumonisin toxicity in animals.

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Haliburton, J.C., Vesonder, R.F., Lock, T.F., Buck, W.B., 1979. Equine leukoencephalomalacia (ELEM): a study of Fusarium moniliforme as an etiologic agent. Vet. Hum. Toxicol. 21, 348 351. Halloy, D.J., Gustin, P.G., Bouhet, S., Oswald, I.P., 2005. Oral exposure to culture material extract containing fumonisins predisposes to the development of pneumonitis caused by Pasteurella multocida. Toxicology. 213, 34 44. Hannun, Y.A., Bell, R.M., 1989. Functions of sphingolipids and sphingolipid breakdown products in cellular regulation. Science 243, 500 507. Harrison, L.R., Colvin, B.M., Greene, J.T., et al., 1990. Pulmonary edema and hydrothorax in swine produced by fumonisin B1, a toxic metabolite of Fusarium moniliforme. J. Vet. Diagn. Invest. 2, 217 221. Harvey, R.B., Edrington, T.S., Kubena, L.F., et al., 1996. Effects of dietary fumonisin B1-containing culture material, deoxynivalenolcontaminated wheat, or their combination on growing barrows. Am. J. Vet. Res. 57, 1790 1794. Haschek, W.M., Motelin, G., Ness, D.K., et al., 1992. Characterization of fumonisin toxicity in orally and intravenously dosed swine. Mycopathology 117, 83 96. Howard, P.C., Eppley, R.M., Stack, M.E., et al., 2001. Fumonisin B1 carcinogenicity in a two-year feeding study using F344 rats and B6C3F1 mice. Environ. Health Perspec. 109 (Suppl. 2), 277 282. Jaskiewicz, K., Marasas, W.F.O., Taljaard, J.J.F., 1987. Hepatitis in vervet monkeys caused by Fusarium monliforme. J. Comp. Path. 97, 281 291. Jovanovi´c, M., Trailovic, D., Kukolj, V., et al., 2015. An outbreak of fumonisin toxicosis in horses in Serbia. World Mycotoxin J. 8, 387 391. Kellerman, T.S., Marasas, W.F.O., Pienaar, J.G., Naude, T.W., 1972. A mycotoxicosis of equidae caused by Fusarium moniliforme sheldon: a preliminary communication. Onderstepoort J. Vet. Res. 39, 205 208. Kellerman, T.S., Marasas, W.F.O., Thiel, P.G., et al., 1990. Leukoencephalomalacia in two horses induced by oral dosing of fumonisin B1. Onderstepoort J. Vet. Res. 57, 269 275. Kriek, N.P.J., Kellerman, T.S., Marasas, W.F.O., 1981. A comparative study of the toxicity of Fusarium verticillioides (5F. moniliforme) to horses, primates, pigs, sheep and rats. Onderstepoort J. Vet. Res. 48, 129 131. Laurent, D., Pellegrin, F., Kohler, F., et al., 1989. Fumonisin B1 in equine leucoencephalomalacia pathogenesis. Microbiologie Aliments Nutr. 7, 285 291. Ledoux, D.R., Brown, T.P., Weibking, T.S., Rottinghaus, G.E., 1992. Fumonisin toxicity in broiler chicks. J. Vet. Diagn. Invest. 4, 330 333. Liguoro, M., Petterino, C., Mezzalira, G., et al., 2004. Vet. Hum. Toxicol. 46, 303 305. Lim, C.W., Parker, H.M., Vesonder, R.F., Haschek, W.M., 1996. Intravenous fumonisin B1 induces cell proliferation and apoptosis in the rat. Nat. Toxins 4, 33 41. MacCallum, W.G., Buckley, S.S., 1902. Acute epizootic leucoencephalitis in horses. Am. Vet. Rev. 26, 21 36. Marasas, W.F.O., Kellerman, T.S., Pienaar, J.G., Naude, T.W., 1976. Leukoencephalomalacia: a mycotoxicosis of equidae caused by Fusarium moniliforme sheldon. Onderstepoort J. Vet. Res. 43, 113 122.

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Marasas, W.F.O., Kellerman, T.S., Gelderblom, W.C.A., et al., 1988. Leukoencepalomalacia in a horse induced by fumonisin B1 isolated from Fusarium moniliforme. Onderstepoort J. Vet. Res. 55, 197 203. Martinez-Larranaga, M.R., Anadon, A., Diaz, M.J., et al., 1999. Toxicokinetics and oral bioavailability of fumonisin B1. Vet. Hum. Toxicol. 41, 357 362. Mathur, S., Constable, P.D., Eppley, R.M., et al., 2001. Fumonisin B1 increases serum sphinganine concentration but does not alter serum sphingosine concentration or induced cardiovascular changes in milk-fed calves. Toxicol. Sci. 60, 379 384. McCue, P.M., 1989. Equine leukoencephalomalacia. Comp. Contin. Educ. Pract. Vet. 11, 646 651. McDonough, P.M., Yasui, K., Betto, R., et al., 1994. Control of cardiac Ca21 levels: inhibitory actions on sphingosine on Ca21 transients and L-type Ca21 channel conductance. Circ. Res. 75, 981 989. Merrill Jr., A.H., Sweeley, C.C., 1996. Sphingolipids metabolism and cell signaling. In: Vance, D.E., Vance, J.E. (Eds.), Biochemistry of Lipids, Lipoproteins, and Membranes. Elsevier, New York, pp. 43 73. Merrill Jr., A.H., Wang, E., Schroeder, J.J., et al., 1995. In: Elklund, M., Richards, M., Mise, K. (Eds.), Molecular Approaches to Food Safety. Issues Involving Toxic Microorganisms. Alaken Press, Fort Collins CO, pp. 429 443. Michelakis, E., Tewari, K., Simard, J.M., 1994. Calcium channels in smooth muscle cells from cerebral precapillary arterioles activate at more negative potentials than those from basilar artery. Pflugers Arch. 426, 459 461. Motelin, G.K., Haschek, W.M., Ness, D.K., et al., 1994. Temporal and dose-response features in swine fed corn screenings contaminated with fumonisin mycotoxins. Mycopathology 126, 27 40. Osweiler, G.D., Ross, P.F., Wilson, T.M., et al., 1992. Characterization of an epizootic of pulmonary edema in swine associated with fumonisins in corn screenings. J. Vet. Diagn. Invest. 4, 53 59. Osweiler, G.D., Kehrli, M.E., Stabel, J.R., et al., 1993. Effects of fumonisin-contaminated corn screenings on growth and health of feeder calves. J. Anim. Sci. 71, 459 466. Patchimasiri, T., Sailasuta, A., Kawtheerakul, K., 1998. Pathological findings in swine in association with fumonisin contaminated feed. Thai J. Vet. Med. 28, 71 82. Plumlee, K.H., Galey, F.G., 1994. Neurotoxic mycotoxins: a review of fungal toxins that cause neurological disease in large animals. J. Vet. Int. Med. 8, 49 54. Po´sa, R., Magyar, T., Stoev, S.D., et al., 2013. Use of computed tomography and histopathologic review for lung lesions produced by the interaction between Mycoplasma hyopneumoniae and fumonisin mycotoxins in pigs. Vet. Pathol. 50, 971 979. Prelusky, D.B., Trenholm, H.L., Savard, M.E., 1994. Pharmacokinetic fate of 14C-labelled fumonisin B1 in swine. Nat. Toxins 2, 73 80. Prelusky, D.B., Savard, M.E., Trenholm, H.L., 1995. Pilot study on the plasma pharmacokinetics of fumonisin B1 in cows following a single dose by oral gavage or intravenous administration. Nat. Toxins 3, 384 394. Raoofi, A., Mardjanmehr, S.H., Khosravi, A.R., et al., 2003. J. Eq. Vet. Sci. 23, 469 470. Rheeder, J.P., Marasas, W.F.O., Thiel, P.G., et al., 1992. Fusarium moniliforme and fumonisins in corn in relation to human esophageal cancer in Transkei. Phytopathology 82, 353 357.

1018 SECTION | XV Mycotoxins

Richard, J.L., Meerdink, G., Maragos, C.M., et al., 1996. Absence of detectable fumonisins in the milk of cows fed Fusarium proliferatum (Matusushima) Nirenberg culture material. Mycopathology 133, 123 126. Riley, R.T., An, N.H., Showker, J.L., et al., 1993. Alteration of tissue and serum sphinganine to sphingosine ratio: an early biomarker of exposure to fumonisin-containing feeds in pigs. Toxicol. Appl. Pharmacol. 118, 105 112. Rosiles, M.R., Bautista, J., Fuentes, V.O., Ross, F., 1998. An outbreak of equine leukoencephalomalacia at Oaxaca, Mexico, associated with fumonisin B1. J. Vet. Med. A. 45, 299 302. Ross, P.F., Rice, L.G., Reagor, J.C., et al., 1991. Fumonisin B1 concentrations in feeds from 45 confirmed equine leukoencephalomalacia cases. J. Vet. Diagn. Invest. 3, 238 241. Ross, P.F., Ledet, A.E., Owens, D.L., et al., 1993. Experimental equine leukoencephalomalacia, toxic hepatitis, and encephalopathy caused by corn naturally contaminated with fumonisins. J. Vet. Diagn. Invest. 5, 69 74. Ross, P.F., Nelson, P.E., Owens, D.L., et al., 1994. Fumonisin B2 in cultured Fusarium proliferatum, M-6104, causes equine leukoencephalomalacia. J. Vet. Diagn. Invest. 6, 263 265. Rotter, B.A., Prelusky, D.B., Fortin, A., et al., 1996. Response of growing swine to dietary exposure to fumonisin B1 during an eight-week period: growth and clinical parameters. Nat. Toxins 4, 42 50. Schumacher, J., Mullen, J., Shelby, R., et al., 1995. An investigation of the role of Fusarium moniliforme in duodenitis/proximal jejunitis of horses. Vet. Human Toxicol. 37, 39 45. Schwarte, L.H., Biester, H.E., Murray, C., 1937. A disease of horses caused by feeding moldy corn. J. Am. Vet. Med. Assoc. 43, 76 85. Shephard, G.S., Thiel, P.G., Sydenham, E.W., Savard, M.E., 1995. Fate of a single dose of 14C-labelled fumonisin B1 in Vervet monkeys. Nat. Toxins 3, 145 150. Scott, P.M., 2012. Recent research on fumonisins: a review. Food Addit. Contam. 29, 242 248. Smith, G.W., Constable, P.D., Bacon, C.W., et al., 1996a. Cardiovascular effects of fumonisins in swine. Fundam. Appl. Toxicol. 31, 169 172. Smith, G.W., Constable, P.D., Haschek, W.M., 1996b. Cardiovascular responses to short-term fumonisin exposure in swine. Fundam. Appl. Toxicol. 33, 140 148. Smith, G.W., Constable, P.D., Smith, A.R., et al., 1996c. Effects of fumonisin-containing culture material on pulmonary clearance in swine. Am. J. Vet. Res. 57, 1233 1238. Smith, G.W., Constable, P.D., Tumbleson, M.E., et al., 1999. Sequence of cardiovascular changes leading to pulmonary edema in swine fed culture material containing fumonisin. Am. J. Vet. Res. 60, 1292 1299. Smith, G.W., Constable, P.D., Eppley, R.M., et al., 2000. Purified fumonisin B1 decreases cardiovascular function but does not alter pulmonary capillary permeablity in swine. Toxicol. Sci. 56, 240 249. Smith, G.W., Constable, P.D., Foreman, J.H., et al., 2002. Cardiovascular changes associated with intravenous administration of fumonisin B1 in horses. Am. J. Vet. Res. 63, 538 545. Spotti, M., Caloni, F., Fracchiolla, L., et al., 2001. Fumonisin B1 carryover into milk in the isolated perfused bovine udder. Vet. Hum. Toxicol. 43, 109 111.

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FURTHER READING Diaz, G.J., Boermans, H.J., 1994. Fumonisin toxicosis in domestic animals: a review. Vet. Human Toxicol. 36, 548 555. Smith, G.W., Constable, P.D., 2004. Fumonisin. In: Plumlee, K.H. (Ed.), Clinical Veterinary Toxicology. Mosby, St. Louis, MO, pp. 250 254.