Mycotoxins in feed – do we know enough?

Ewa Wielogorska and Chris ElliottInstitute for Global Food Security

Queen’s University, Belfast 

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Mycotoxins• Natural toxic secondary metabolites produced by microscopic filamentous 

fungi such as Fusarium, Aspergillus, Penicillium, Alternaria

• Fungi can grow on agricultural products on the field and/or during storage, transport and processing

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Main groups of mycotoxins• Aflatoxins• Fumonisins• Zearalenone• Ochratoxin A• Trichothecenes• Patulin• Ergot alkaloids• Alternaria mycotoxins

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Mycotoxins in feed – summary of the regulations

Mycotoxin Mycotoxin content in feed [mg/kg=ppm]

Type

Aflatoxin B1 0.005 ‐ 0.02 maximum contentDeoxynivalenol 0.9 ‐ 12 guidanceZearalenone 0.1 ‐ 3 guidanceOchratoxin A 0.05 ‐ 0.25 guidanceFumonisin B1 + B2 5 ‐ 60 guidanceT‐2 + HT‐2 0.25 ‐ 2 indicative levels

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Emerging mycotoxins...AurofusarinMoniliforminApicidinEquisetinEmodinSterigmatocystinCurvularinPenitrem ARoquefortine CSkyrinPhomopsins...

EFSA focus (Scientific Opinions published recently)

• Nivalenol (June 2013)• Sterigmatocystin (June 2013)• Beauvericin and enniatins (September 

2014)• Moniliformin (September 2014)• Diacetoxyscirpenol (September 2014)

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fumonisins covalenlty bound to proteins and carbohydrates

EFFECTS ON SWINE 

T-2, DON, Ergots• Dermal and oral lesions• Feed refusal • Vomiting• Impaired growth

T-2, Ergots• Necrosis

T-2, DON, AFB1, OTA, FUM, • Damage of the kidneys• Pale and fatty liver• Porcine pulmonary edema (PPE)• Increased water consumption•Intestinal hemorrhages

ZON, T-2, DON, Ergots• Abortion• Pseudo pregnancy• Low conception rates• Ovarian cysts• Embryonic Loss• Tail necrosis• Stillbirths

AFB1, T-2, OTA• Diarrhea• Blood in faeces and urine

T-2, DON, AFB1, OTA, FUM• Decreased performance• Immune-suppression• Pancreatic necrosis

DIFFERENTIAL DIAGNOSIS

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EFFECTS ON POULTRY

arti

AFLATOXINSOCHRATOXIN A

AFLATOXINSTRICHOTECENES

TRICHOTHECENES

TRICHOTHECENES

TRICHOTHECENESZEARALENONE

ERGOT ALKALOIDS

OCHRATOXIN A

AFLATOXINSOCHRATOXIN ATRICHOTHECENES

FUSARIUM TOXINS

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EFFECTS ON RUMINANTS

Vulvovaginitis

Prolepsis

Embryo mortality

Gastro enteritis

Intestinal haemorrhage Immunodepression

Ovarian cysts Feed refusal

vomiting

Milk contamination

Decline in milk production

Increase in somatic cells

Mycotoxins co‐occurrence

• Fungi can produce multiple mycotoxins

• These can have synergistic and additive effects

• Frequent mycotoxins co‐occurrence ‐ need to test for a number of mycotoxins to asses the safety and quality of feed

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«…Synergistic effects can occur at low levels when the combined effects of two or more mycotoxins are much greater than the individual effects of each toxin alone…» (Source: Biomin Mycotoxin Compendium)

‐ ‐ ‐ ‐ ‐ ‐ ‐ Additive effect

Synergistic effect

(Grenier and Oswald, INRA 2011. In World Mycotoxin Journal Aug 2011, 4 (3) 285-313)

Synergisms among mycotoxins

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HGCA (Home‐Grown Cereals Authority) project at QUB (2013‐ 15)

Assess the safety of the DDGS from bioethanol, distilling and brewing industries as a feed ingredient 

with regard to mycotoxin contamination

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HGCA (Home‐Grown Cereals Authority) project at QUB

Transfer and validate UPLC‐MS/MS for the quantification of 77 

mycotoxins 

Analyse approx. 200 DDGS samples and 

identify most commonly occurring 

contaminants

Evaluate LFD for mycotoxin detection in collaboration with leading diagnostic 

companies

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Dried distillers grain with solubles(DDGS)

• Co‐product of ethanol production from starch cereals (maize, wheat, barley, but also rye, triticale, sorghum and oat)

• Contain valuable amounts of protein, fat and fibre• Used as feed ingredient in diets of livestock, poultry and fish

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0

2000

4000

6000

8000

10000

12000

2000 2001 2002 2003 2004 2005 2006 2007 2008 2009 2010 2011

Thou

sand

s of lite

rs per day

Fuel ethanol production in Europe

United Kingdom

Sweden

Spain

Slovakia

Poland

Norway

Netherlands

Italy

Ireland

Hungary

Germany

France

Finland

Denmark

Czech Republic

Belgium

Austria

1 tonne of grain processed for ethanol production will generate 1/3 tonne of feed co‐products.

Source: Cooper & Weber. 2012 An outlook on world biofuel production and its implications for the animal feed industry .

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Mycotoxin Percentage of contaminated 

samples

Concentration range [µg/kg]

Beauvericin 100% 46 ‐ 561Enniatin A1 100% 0.2 ‐ 16Enniatin B 100% 1.2 ‐ 183Enniatin B1 100% 0.1 ‐ 50Fumonisin B1 100% 81 ‐ 4,470Fumonisin B2 100% 23 ‐ 1,444Fumonisin B3 100% 11 ‐ 647Fusaric acid 100% 400 ‐ 3,500Equisetin 98% 2.4 ‐ 32Mycophenolic acid 96% 3.2 ‐ 67Aflatoxin B1 95% 0.2 ‐ 16Cyclopiazonic acid 86% 0.6 ‐ 17T2 toxin 83% 4.0 ‐ 102Aflatoxin B2 79% 0.6 ‐ 4.2Meleagrin 79% 0.4 ‐ 3.3Ochratoxin A 74% 1.2 ‐ 46Alternariol 71% 4.9 ‐ 47

DDGS maize samples (n = 42)

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Mycotoxin Percentage of contaminated samples

Concentration range [µg/kg]

Beauvericin 100% 3.1 ‐ 108Enniatin A 100% 4.8 ‐ 31Enniatin A1 100% 26 ‐ 172Enniatin B 100% 234 – 1,486Enniatin B1 100% 118 ‐ 533Ergocristine 100% 1.5 ‐ 23Ergocristinine 100% 0.3 ‐ 4.3Ergocryptinine 100% 0.2 ‐ 5.4Ergosine 100% 0.3 ‐ 22Ergotamine 100% 0.6 ‐ 30Deoxynivalenol 100% 39 – 1,122Ergotaminine 98% 0.2 ‐ 4.6Mycophenolic acid 95% 0.1 ‐ 50Ochratoxin A 95% 0.5 ‐ 3.5Ergocryptine 93% 0.1 ‐ 8.1Emodin 61% 2.9 ‐ 33

DDGS wheat samples (n = 41)

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Conclusions to Study

• Multiple mycotoxins are present in DDGS. 

• Pattern consists of regulated and emerging mycotoxins

• Even at 10 – 20% DDGS inclusion rates mycotoxins may have negative impact of animal performance 

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Impacts of mycotoxins in Animal Performance  

Institute have recently initiated  a study on the concentrations of mycotoxins present in animal feed materials in ‘problem herds/flocks.  

Where herds/flocks are showing underperformance in terms of growth, poor feed conversion etc samples of feed materials are being sent for full mycotoxin profiling (80 mycotoxins). 

If available samples of feed from similar herds/flocks without production problems are being analysed as controls. 

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Mycotoxin [ug/kg]Soya Control Farm 

Soya Problem Farm

Beauvericin 3.2DiacetoxyscripenolEnniatin AEnniatin A1Enniatin B 1.6Enniatin B1 0.4NeosolaniolPenicillic acid3‐ and 15‐Acetyldeoxynivalenol3‐AcetyldeoxynivalenolApicidinAurofusarinDeoxynivalenolFumonisin B1 3.2 1.5Fumonisin B2 4.1 2.3Fumonisin B3 1.2 0.7Fusaric acid 55HT2Mycophenolic acidT2 toxinTentoxin 9.2 2.3ZearalenoneTotal nr of mycotoxins detected 6 6

Case 1: Soya Issues? 

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fusaric acid acts synergistically with deoxynivlenol when given to immature pigs causing depression in weight gain and feed intake

fusaric acid acts synergistically with fumonisins ‐ enhanced toxicity was observed when fusaric acid and FB1 were administered together to developing chicken embryo

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Mycotoxin [ug/kg]Maize control Farm 

Maize Problem Farm 1

Maize Problem Farm 2 

Beauvericin 8.3 29.4 44.6Diacetoxyscripenol 0.8 3.3Enniatin AEnniatin A1 0.6 0.2Enniatin B 0.5 7.0 3.7Enniatin B1 2.1 0.4Neosolaniol 2.7Penicillic acid 0.73‐ and 15‐Acetyldeoxynivalenol 198 1713‐Acetyldeoxynivalenol 37.4 30.7Apicidin 11.7Aurofusarin 16.8 811 470Deoxynivalenol 3.9 1,019 402Fumonisin B1 65.8 108 53.5Fumonisin B2 9.0 24.0 11.3Fumonisin B3 4.1 17.1 5.6Fusaric acid 27.9 57 17.1HT2 55Mycophenolic acid 21.7T2 toxin 18.7 4.3TentoxinZearalenone 6.2 106 36.6

Total nr of mycotoxins detected 15 14 15

Case 2: Maize Issues? 

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decrease of protein and fat content in chicken meat quail eggs: 26.4 mg/kg in feed caused decrease in vitamin E, vitamin A, total

carotenoid, lutein, zeaxanthin breeding chickens: 26.4 mg/kg in feed negatively affected the immune system of the

progeny

Aurofusarin

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Sample type

Nr of samplesanalyzed

Nr of mycotoxinsper sample

Main mycotoxinsfound

Sum of mycotoxincontent [µg/kg]

Barley 2 9‐10

Cyclopiazonic acid, Enniatin A,Enniatin A1, Enniatin B, Enniatin B1, Mycophenolic acid, Sterigmatocystin, Tenuazonic acid

277 ‐ 667

Case 3: Barley Issues? 

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intermediate in aflatoxin biosynthesis ‐ structure is similar to that of aflatoxins. genotoxic and carcinogenic (liver and lungs) sterigmatocyction alone or in combination with ochratoxin A and/or citrinin share a 

cytotoxic and cytogenetic potential even at picoMolar concentrations on human hepatoma cells in vitro

Sterigmatocystin

Tenuazonic acid

toxic effect on oesophageal mucosa of mice resulting in weight loss and lower feed consumption

chicken: 1.25 ‐ 1.50 mg/kg bw/day caused lesions in various tissues and weight loss

Summary  

• Where production issues are being found mycotoxins (often unregulated, emerging mycotoxins) are being found. 

• As datasets are built up a correlation between of problems in terms of production and mycotoxin presence will be developed

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Conclusions

• As analytical methods have improved the information about the range and levels of mycotoxin presence in animal feed material has greatly increased 

• Farm animal exposure to mycotoxins is much greater than previously understood 

• The impact complex mixtures of mycotoxins have on animal performance is still very much an unknown but may be a major contributor to issues in animal performance 

• A better understanding of the effects of mycotoxin cocktails and how to deal with them is required to improve profitability and sustainabilty of livestock production