Scientific Opinion on the risks for animal and public health related to the presence of T-2 and HT-2 toxin in food and feed

T-2 toxin and HT-2 toxin are mycotoxins produced by various Fusarium species. The European Commission asked EFSA for a scientific opinion on the risk to human and animal health related to the presence of T-2 and HT-2 toxin in food and feed. A total of 20,519 results for the sum of T-2 and HT-2 toxins in food, feed and unprocessed grains, collected in 2005-2010 from 22 European countries, were used in the evaluation. The highest mean concentrations for the sum of T-2 and HT-2 toxins were observed in grains and grain milling products, notably in oats and oat products. Grains and grain-based foods, in particular bread, fine bakery wares, grain milling products, and breakfast cereals, made the largest contribution to the sum of T-2 and HT-2 toxin exposure for humans. T-2 toxin is rapidly metabolised to a large number of products, HT-2 toxin being a major metabolite. Pigs are amongst the most sensitive animals towards the effects of T-2 toxin, the most sensitive endpoints being immunological or haematological effects. Using these data and a benchmark dose analysis the Panel on Contaminants in the Food Chain established a group tolerable daily intake (TDI) of 100 ng/kg b.w. for the sum of T-2 and HT-2 toxins. Estimates of chronic human dietary exposure to the sum of T-2 and HT-2 toxins based on the available occurrence data are below the TDI for populations of all age groups, and thus not a health concern. For ruminants, rabbits and farmed fish the estimated exposures to the sum of these toxins based on the available occurrence data are considered unlikely to be a health concern, while for pigs, poultry, dogs and horses the risk of adverse health effects is low. For cats the health risk from the exposure to T-2 and HT-2 toxins cannot be assessed.

© European Food Safety Authority, 2011

T-2 toxin and HT-2 toxin are mycotoxins and are members of the large group of fungal sesquiterpenes, commonly denoted as trichothecenes. They are produced by various Fusarium species. Generally, the Fusarium species grow and invade crops under moist cool conditions. T-2 toxin and HT-2 toxin and other trichothecenes are found in cereal grains and products thereof.

Trichothecenes share a common structure with a tetracyclic ring system containing a stable epoxide group between C12 and C13, which seems to account for many of the typical toxic effects of trichothecenes. The structures of T-2 and HT-2 toxins differ only in one functional group, T-2 toxin being acetylated at C-4 whereas HT-2 toxin is not acetylated.

The Scientific Committee on Food (SCF) issued in 2001 an opinion on Fusarium toxins Part 5: T-2 and HT-2 toxin. The SCF concluded that the general toxicity, haematotoxicity and immunotoxicity of T-2 toxin are the critical effects and established a combined temporary tolerable daily intake (t-TDI) for the sum of T-2 toxin and HT-2 toxin of 0.06 µg T-2 toxin/kg body weight (b.w.). This was in line with the provisional maximum tolerable daily intake (PMTDI) established for T-2 and HT-2 toxin by the Joint Food and Agriculture Organization (FAO)/World Health Organization (WHO) Expert Committee on Food Additives (JECFA).

The European Commission (EC) has asked the European Food Safety Authority (EFSA) for a scientific opinion on the risk to human and animal health related to the presence of T-2 and HT-2 toxin in food and feed. In particular the opinion should consider any new results of toxicological studies published since the assessment by the SCF in 2001, in order to assess if the combined t-TDI of 0.06 µg/kg b.w. for T-2 and HT-2 toxin is still appropriate. Additionally, the opinion should include an updated human dietary exposure assessment of T-2 and HT-2 toxins, and a determination of the daily exposure levels of T-2 and HT-2 toxins for different animal species above which signs of toxicity can be observed. It should also include a determination of the daily exposure levels of T-2 and HT-2 toxins above which the level of transfer/carry over of T-2 and HT-2 toxins from the feed to products of animal origin for human consumption results in unacceptable T-2 and HT-2 toxin levels. The EC request also asked for identification of the feed materials which could be considered as sources of T-2 and HT-2 toxins and the characterisation of the distribution of T-2 and HT-2 toxin levels in different feed materials.

Methods for analysis of T-2 toxin are well established and can be applied for the analysis of cereals, food, feed and biological samples. Accurate quantification of T-2 and HT-2 toxins is mostly carried out by liquid chromatography coupled with (multi-stage) mass spectrometry often within a multianalyte approach. For rapid screening several immunochemical methods have become available but they may suffer from undesired cross reactivity. None of the applied methods have been formally validated in interlaboratory validation studies.

Following a call for data by EFSA in July 2010, a total of 17,683 analytical results for T-2 toxin, 16,536 for HT-2 toxin and 20,519 for the sum of T-2 and HT-2 toxins in food, feed and unprocessed grains, collected between 2005 and 2010 from 22 European countries, were received. Overall, 65 % of the results were below the limit of detection (LOD) or limit of quantification (LOQ). In the quantified results, HT-2 toxin concentration represents about two thirds of the sum of T-2 and HT-2 toxin concentration. The highest mean concentrations for the sum of T-2 and HT-2 toxins were observed in grains and grain milling products, notably in oats and oat products. Levels in unprocessed grains were higher than in grain products for human consumption, suggesting that processing applied to grains results in lower T-2 toxin and HT-2 toxin concentrations. During the milling process T-2 and HT-2 toxins are not destroyed but unevenly redistributed between fractions. Because T-2 and HT-2 toxins are mostly attached to the outer hull of the grain, cleaning, sorting, sieving and de-hulling of grains lead to marked increases in T-2 and HT-2 toxins in cereal by-products, e.g. bran. T-2 and HT-2 toxins are relatively stable compounds during baking and cooking.

The Panel on Contaminants in the food chain (CONTAM Panel) estimated total chronic dietary exposures to the sum of T-2 and HT-2 toxins across 14 European countries, using lower bound (LB) and upper bound (UB) mean concentrations of the sum of T-2 and HT-2 toxins in foods, and consumption data for different age groups. For adults the minimum LB to maximum UB was 3.4 to 18 ng/kg b.w. per day for average consumers, and 7.2 to 39 ng/kg b.w. for high consumers (95th percentile consumption in total population). In elderly and very elderly populations, the chronic dietary exposure to the sum of T-2 and HT-2 toxins was slightly lower compared to other adults. The highest chronic dietary exposure estimates are for toddlers (age ≥ 12 months to < 36 months), at 12 to 43 ng/kg b.w. per day for average consumers, and 23 to 91 ng/kg b.w. for 95th percentile consumers.

Grains and grain-based foods, in particular bread, fine bakery wares, grain milling products, and breakfast cereals, made the largest contribution to the sum of T-2 and HT-2 toxin exposure. For infants, the highest contributors were in the food group ‘Foods for infants and small children’, mainly cereal-based foods. No significant difference in the dietary exposure to the sum of T 2 and HT-2 toxins was found between vegetarians and the general population, although the data were limited.

The available information on the toxicokinetics of T-2 and HT-2 toxins is incomplete. T-2 toxin is rapidly metabolised to a large number of products, HT-2 toxin being a major metabolite. The metabolic pathways include hydrolysis, hydroxylation, de-epoxidation, glucuronidation and acetylation. Distribution and excretion of T-2 toxin and its metabolites are rapid. There are no significant data available on the toxicity of most metabolites. De-epoxidation is believed to be a detoxification process.

T-2 toxin inhibits protein, RNA and DNA synthesis. Recent data also indicate that T-2 toxin induces apoptosis, and in some cell types necrosis, as well as lipid peroxidation affecting cell membrane integrity. T-2 toxin induces haematotoxicity and myelotoxicity associated with impairment of haematopoiesis in bone marrow. The published investigations demonstrate that pigs are among the most sensitive animals towards the effects of T-2 toxin, the most sensitive endpoints being immunological or haematological effects which occur from doses of 29 µg/kg b.w. per day. Since the SCF evaluation in 2001 there is no evidence that the other toxic effects including dermal toxicity, developmental and reproductive toxicity and neurotoxicity occur at doses lower than those causing immunotoxicity and haematotoxicity in pigs. Although cats have been shown to be a very sensitive species their particular sensitivity to T-2 toxin is likely to be associated with their inability to excrete T-2 toxin and its metabolites via glucuronide conjugation. Because of this difference in metabolic pathway with that of humans, data for cats are not suitable for human risk assessment.

The assessment by the SCF of the genotoxicity of T-2 toxin indicated a positive effect in several conventional tests for genotoxicity in vitro and in rodents in vivo, in particular for clastogenic effects, but these effects were observed primarily at concentrations also known to inhibit protein and DNA synthesis and produce cytotoxicity. No new reports on cytogenetic damage caused by T-2 toxin have been identified since then. The SCF reported limited evidence for tumourigenicity of T-2 toxin in experimental animals (induction of hepatocellular- and pulmonary adenomas in male mice). No new data are available on T-2 toxin carcinogenicity in experimental animals or on the carcinogenicity to humans of toxins derived from Fusarium sporotrichioides. Also there are no other new epidemiological data in the context of dietary exposure to T-2 and HT-2 toxins and human diseases.

The CONTAM Panel noted that the currently available toxicological studies have some uncertainties. Only one new dose-response study suitable for risk assessment has become available since 2001. The short term study in pigs, used by the SCF as the basis for establishing a t-TDI, was considered by CONTAM Panel to be still the most appropriate study for this purpose, and the immunotoxicological endpoints in this study were found to be the most important biological effects for risk assessment. The lowest-observed-adverse-effect-level (LOAEL) was 29 µg T-2 toxin/kg b.w. per day in this experiment, but no no-observed-adverse-effect-level (NOAEL) was identified. The CONTAM Panel concluded that a reduction in specific antibody response in pigs is the critical effect for human risk assessment.

The data on anti-horse globulin response from this study were used for a benchmark dose (BMD) analysis to identify a reference point for T-2 and HT-2 toxins. The default value for continuous data recommended by EFSA is a benchmark response (BMR) of 5 %. In the absence of statistical or toxicological considerations supporting deviation from the default value the CONTAM Panel chose a BMR of 5 % when applying the BMD approach on the dose-(antibody) response data available. The 95 % lower confidence limit for the benchmark dose response of 5 % (BMDL₀₅) calculated for anti-horse globulin titre values was 10 µg T-2 toxin/kg b.w. per day.

In view of the rapid metabolism of T-2 toxin to HT-2 toxin, and the fact that the toxicity of T-2 toxin might at least partly be attributed to HT-2 toxin, a group TDI was established for the sum of T-2 and HT-2 toxins. An uncertainty factor of 100 was applied to the BMDL₀₅, to establish a group TDI of 100 ng/kg b.w. for the sum of T-2 and HT-2 toxins. As new relevant evidence has become available since the previous t-TDI was established by the SCF in 2001, and as the present assessment was based on a BMDL₀₅, the CONTAM Panel concluded that a full TDI of 100 ng/kg b.w. can now be established.

Estimates of chronic dietary exposure for populations of all age groups to the sum of T-2 and HT-2 toxins based on the available occurrence data are below this group TDI of 100 ng/kg b.w., and therefore there is no health concern.

Animal exposure to the sum of T-2 and HT-2 toxins is primarily from consuming cereal grains and cereal by-products; levels in forages and oilseed meals are generally low. The animals considered were dairy cows, beef cattle, sheep and goats, pigs and piglets, hens, broiler chickens, turkeys, ducks, rabbits, fish, dogs, cats and horses. The highest UB exposure based on the available occurrence data in feed was for milking goats at 3.3 µg/kg b.w. per day and the lowest was for farmed fish at 0.19 µg/kg b.w. per day.

Information on LOAELs and NOAELs for farm and companion animals is limited. In young ruminants, exposure to 300 µg T-2 toxin/kg b.w. per day or more may result in gastrointestinal lesions, altered serum proteins and haematological alterations. In ruminants the effects observed in nutritionally challenged heifers and ewes give rise to the assumption that rumen detoxification of T-2 toxin may not always be complete. For pigs, the published investigations demonstrate that they are among the most susceptible animals towards the effects of T-2 toxin, the most sensitive endpoints being immunological or haematological effects which occur from doses of 29 µg/kg b.w. per day. In poultry, first effects (e.g. mucosal damage in oral cavity) occur at a dose of 40 µg/kg b.w. per day and 48 µg/kg b.w. per day for broiler chickens and fattening turkeys, respectively. In fattening ducks a dose of 40 µg/kg b.w. per day caused a significant reduction in body weight gain. Infertility of eggs and/or reduction of egg production were seen at doses of 120 µg/kg b.w. per day for laying hens.

For rabbits, doses ranging from 500-2000 µg of T-2 toxin/kg b.w. per day generate decrease of body weight gain and mucosal damage. Only moderate signs including haematological and hormonal effects have been observed for doses ranging from 200-500 µg of T-2 toxin/kg b.w. per day. A NOAEL of 100 µg T-2 toxin/kg b.w. per day was identified. Reduced feed intake, growth and haematocrit values as well as an increased mortality have been reported for fish. The lowest NOAEL of 13 µg T-2 toxin/kg b.w. per day has been identified for catfish. Cats are amongst the most sensitive animal species. This particular sensitivity of cats to T-2 toxin is likely to be associated with their inability to excrete T-2 toxin and its metabolites via glucuronide conjugation. Due to the limited data and the severe effects i.e. mortality observed for cats at the low dose levels the available data can not be used to identify a NOAEL or a LOAEL. For dogs no toxicity data are available and a NOAEL or LOAEL cannot be identified. The available data do not provide a NOAEL or a LOAEL for horses.

With regard to animal health risk characterisation, because of the limited knowledge on the effects of T-2 and HT-2 toxins on farm and companion animals, and the absence of a comprehensive database on feed consumption by livestock in the EU, it has not been possible to properly assess the risks of these toxins for animal health. However, the exposure values for the LB and UB concentrations for the sum of T-2 and HT-2 toxins in diets have been estimated for a number of farm livestock and companion animal categories, based on expected feed intakes and example diets, and these have been compared with identified NOAELs/LOAELs or with the calculated BMDL₀₅ for pigs.

The CONTAM Panel used the BMDL₀₅ for pigs as a reference point for risk characterisation for both pigs and poultry. The latter was considered acceptable as there was no indication from identified LOAELs that poultry are more sensitive than pigs. In the absence of NOAELs or LOAELs for horses and dogs, the CONTAM Panel also decided to use the same reference point as that derived for pigs to give an indication on the possible risk, since toxicokinetics of T-2 and HT-2 toxins in horses and dogs are not substantially different to that of pigs. However, due to the differences in oral bioavailability and metabolism in ruminants and fish, the BMDL₀₅ for pigs was not used for the risk characterisation for these species. The identified NOAELs or LOAELs were used for risk characterisation for ruminants, fish and rabbits. For cats the health risk from the exposure to the sum of T-2 and HT-2 toxins could not be assessed as no NOAEL or LOAEL has been identified, and as there is a lack of sufficient data on the feline-specific biotransformation and toxicodynamics. However, cats seem to be amongst the most sensitive animal species to T-2 toxin and HT-2 toxin intoxication.

Based on estimates of feed intake and the available occurrence data on feedingstuffs, the exposures to the sum of T-2 and HT-2 toxins for ruminants are substantially lower than the LOAELs identified, and are therefore considered unlikely to be a health concern. For pigs and poultry, comparison of the estimates of exposure based on the reported levels of the sum of T-2 and HT-2 toxins in feeds to the BMDL₀₅ for pigs indicate that the risk of adverse health effects of feed containing T-2 and HT-2 toxins is low for these species.

The limited data available for rabbits and farmed fish suggest that the estimated exposures to the sum of T-2 and HT-2 toxins in feed at the currently reported concentrations is well below the identified NOAELs, and therefore considered unlikely to be a health concern.

For cats the health risk from the exposure to the sum of T-2 and HT-2 toxins could not be assessed due to the lack of sufficient data. For dogs and horses, the estimates of exposure based on the reported levels of the sum of T-2 and HT-2 toxins in feeds indicate that the risk of adverse health effects as a result of consuming feed containing T-2 and HT-2 toxins is low for these species.

The available data describing possible effects of combined exposure to T-2 and HT-2 toxins with other mycotoxins are too limited to draw any conclusions.

Mycotoxins, Fusarium, HT-2 toxin, T-2 toxin, food, feed, analysis, occurrence, human dietary exposure, animal dietary exposure, risk assessment, toxicity, tolerable daily intake (TDI)