Review of results published by Mesnage et al. (2015) in PLoS ONE

SCIENTIFIC REPORT

APPROVED: 2 October 2015 PUBLISHED: 6 October 2015

doi:10.2903/j.efsa.2015.4258

www.efsa.europa.eu/efsajournal EFSA Journal 2015;13(10):4258

Review of results published by Mesnage et al. (2015) in PLoS ONE and the laboratory findings communicated

by Dr Samsel to Farm Wars

European Food Safety Authority

Abstract

In a paper published in PLoS One and entitled ‘Laboratory rodent diets contain toxic levels of

environmental contaminants: Implications for regulatory tests’, Mesnage et al. (2015) analysed commercial laboratory rodent diets for environmental contaminants and genetically modified

organisms (GMOs). In samples from 13 different commercial rodent diets obtained from five

continents, the authors of the study report the presence of pesticides, heavy metals, polychlorinated dibenzo-p-dioxins and dibenzofurans, and GMOs. The paper by Mesnage et al. (2015) provides a

useful addition to the already existing knowledge in the field. However, there are several limitations with the methodological approach used by the authors, including insufficient information about the

test material and methodology used, incomplete reporting of the data, and inappropriate

interpretation of legislation and results. The vast majority of pesticides were absent (below the limit of detection), and where detected, the levels of pesticides, heavy metals and dioxins were only just

above the limit of detection in the feed samples but below regulatory levels for feed and foodstuffs. Only in a limited number of feed diets did the authors report levels of lead that exceeded the

maximum levels specified by legislation for foodstuffs. The application of the ADI concept to claim the existence of a health risk in rodents or to demonstrate background levels of diseases or disorders in

rodents has no scientific justification. In an interview conducted with Dr Samsel, the farmwars.info

website reports on the presence of glyphosate in three different rodent diets. The information reported on the website is not supported by sufficient detail or a reference to permit a full scientific

review. In conclusion, no new scientific elements were provided that would impact on the validity of regulatory feeding tests in the EU.

© European Food Safety Authority, 2015

Keywords: rodent, feed, pesticides, heavy metals, dioxins, GMO, tumours

Requestor: European Commission

Question number: EFSA-Q-2015-00486

Correspondence: [email protected]

Review of results published by Mesnage et al. (2015) and the laboratory findings by Samsel

www.efsa.europa.eu/efsajournal 2 EFSA Journal 2015;13(10):4258

Acknowledgements: EFSA wishes to thank Daniel Doerge (NTP, USA), Kettil Svensson (National Food Agency, SE) and Jean-Marc Vidal (EMA, UK) as well as EFSA staff members Marco Binaglia,

Daniele Court Marques, George Kass, Anna Lanzoni, Hermine Reich and Hans Steinkellner for the

support provided to this scientific output.

Suggested citation: EFSA (European Food Safety Authority), 2015. Scientific Report: Review of

results published by Mesnage et al. (2015) in PLoS ONE and the laboratory findings communicated by Dr Samsel to Farm Wars. EFSA Journal 2015;13(10):4258, 12 pp. doi:10.2903/j.efsa.2015.4258

ISSN: 1831-4732

© European Food Safety Authority, 2015

Reproduction is authorised provided the source is acknowledged.

The EFSA Journal is a publication of the European Food Safety Authority, an agency of the European Union.



Table of contents

Abstract ......................................................................................................................................... 1 1. Introduction ........................................................................................................................ 4

Background and Terms of Reference as provided by the requestor ........................................ 4 1.1.

2. Assessment ........................................................................................................................ 4 Review of the Paper Published in PLoS One (Mesnage et al., 2015) ........................................ 4 2.1.

2.1.1. Methodologies .................................................................................................................... 4 2.1.2. Results ............................................................................................................................... 5

Review of the study by Samsel ............................................................................................ 6 2.2.

3. Impact of the paper published in PLoS One (Mesnage et al., 2015) ........................................ 7 4. Conclusions ........................................................................................................................ 8 References ..................................................................................................................................... 9 Abbreviations ............................................................................................................................... 11 Appendix A – Analysis of the Pesticide Residues Identified by Mesnage et al. (2015) in the Rodent

Diets and Comparison with Identified NOAELs .................................................................... 12



1. Introduction

In a paper published in PLoS One and entitled ‘Laboratory rodent diets contain toxic levels of environmental contaminants: Implications for regulatory tests’, Mesnage et al. (2015) analysed

commercial laboratory rodent diets for environmental contaminants and genetically modified organisms (GMOs). In samples from 13 different commercial rodent diets obtained from five

continents, the authors of the study report the presence of pesticides, heavy metals, polychlorinated dibenzo-p-dioxins and dibenzofurans, and GMOs. Based on the predicted daily intakes for each

contaminant from the levels found in the rodent diets and the sums of the computed hazard

quotients, the authors of the study considered that a health risk was associated with the chronic consumption of these diets by laboratory rodents and that the diets could contribute to the chronic

pathologies reported in laboratory rodents. This led the authors to the conclusion that efforts towards safer rodent diets should be made to improve the reliability of toxicity tests performed on laboratory

rodents.

The laboratory findings communicated by Dr Samsel to Farm Wars are on the contamination of laboratory rodent diets with glyphosate. On the basis of these claims, Dr Samsel recommended to ban

the use of historical controls which may be biased by toxicologically relevant contamination.

Background and Terms of Reference as provided by the requestor 1.1.

DG SANTE has been made aware of a research paper recently published by Prof Seralini1 in the PLoS

ONE Journal and laboratory studies done by Dr Samsel,2 both testing animal feed diets used in regulatory toxicology tests.

In the first study entitled ‘Laboratory rodent diets contain toxic levels of environmental contaminants: Implications for Regulatory Tests’, the team of Prof Seralini analysed 13 samples of laboratory rat

diets and found traces of pesticides, heavy metals, dioxins, PCBs and GMOs. In the second study, Dr

Samsel tested 3 animal feed diets for the presence of glyphosate. Based on the results, both authors put into question the regulatory toxicology tests conducted to date.

On the basis of Article 31 of Regulation (EC) No 178/2002, EFSA is requested to analyse both studies and indicate whether the results contain new scientific elements that could impact the regulatory

feeding tests in the EU.

2. Assessment

Review of the paper published in PLoS One (Mesnage et al., 2015) 2.1.

The paper by Mesnage et al. (2015) reports on the levels of pesticides, heavy metals, dioxins and dioxin-like compounds and GMOs found in commercial rodent diets. Samples from 13 different

commercial rodent diets obtained from the USA, Brazil, France, Italy, Germany, UK, Kenya, China and

New Zealand were analysed. Overall, this paper provides a useful contribution to existing data on the constituents of feeds used for laboratory animals.

2.1.1. Methodologies

Overall, EFSA noted several limitations with the methodological approach used by the authors,

including insufficient information about the test material and methods used, incomplete reporting of

the data and inappropriate interpretation of legislation and results. The ADIs reported by Mesnage et al. (2015) are quoted only as ‘EFSA or FAO/WHO’ with no further reference to which evaluation was

used by the authors. The ADIs used also appear derived from Provisional Tolerable Monthly Intakes (PTMIs) and Provisional Tolerable Weekly Intakes (PTWIs) established by FAO/WHO; in the case of

lead and arsenic, the PTDI established by FAO/WHO have been withdrawn and should consequently

not be used for derivation of safe levels (EFSA Panel on Contaminants in the Food Chain (CONTAM), 2009, 2010). Likewise, the source of the Maximum Residue Limits (MRLs) cited for rodent diets is not

clear. Finally, the contaminant levels found in the laboratory animal feeds were used to compute

1 Mesnage R, Defarge N, Rocque L-M, de Vendomois J S and Seralini G-E, 2015. Laboratory Rodent Diets Contain Toxic Levels

of Environmental Contaminants: Implications for Regulatory Tests. Plos One, 10(7). 2 http://farmwars.info/?p=14095



potential exposure scenarios for the animals which in turn were compared by the author with the ADIs derived for humans. ADIs take into consideration uncertainty factors to account for inter- and intra-

species variability and are usually at least 100-fold lower than the level at which no adverse effect in

rodents is expected. Because the paper by Mesnage et al. (2015) addresses health risks in rats, these uncertainties do not apply and the comparison is conceptually flawed.

The rationale used by Mesnage et al. (2015) to select the diets analysed in this study lacks clarity and is insufficiently described. According to the authors, the diets have been selected to represent many

geographical sources of laboratory rodent chows (13 suppliers from nine countries) and their various

uses (academic research and regulatory assessment). However, no details were provided on the specific formulation of the diets, and EFSA noted that most of the diets selected for the study were

closed-formula diets for which information on the precise composition of the diet (in term of precise list and amount of ingredients) is proprietary to the supplier (Barnard et al., 2009). Furthermore, the

context of use (regulatory or research) is known only for a few of the diets (e.g., LabDiet 5002 or Teklad, which are commonly used in regulatory studies). Typically, diets used in regulatory studies on

laboratory animals are ‘GLP certified’, which implies that these batches which are controlled by the

supplier for contaminants (including heavy metals, pesticides, PCBs and mycotoxins) and provided with a certificate of analysis, are checked in conformity with standard diet specifications. EFSA noted

some samples were obtained from diets originating from laboratories where these were used and for which storage conditions were not reported.

Although all reported analytical measurements were described as having been conducted by

laboratories accredited by the Comité français d’accréditation (Cofrac), there is lack of clarity regarding the number and details of the batches sampled for each commercial diet and the use of a

‘one-shot’ analysis while reference to sampling in triplicate was made. The composition of feed may vary over time, in particular with regard to the type and the origin of the ingredients. Thus, the results

cannot be generalised but give a snap shot of the samples analysed. The paper does not specify whether the method used for pesticide residue analysis (QuEChERS-method EN 15662) has been

adapted for handling dry matrices. Furthermore, it is noted that the samples should have been

analysed for metabolites relevant for risk assessment [e.g., malathion-oxon, additional metabolites of glyphosate that are of particular relevance for genetically modified crops, i.e., not only

(aminomethyl)phosphonic acid (AMPA)] if the results are to be used for a dietary risk assessment.

2.1.2. Results

Out of 262 pesticides analysed, residues for pirimiphos-methyl, deltamethrin, glyphosate (+ AMPA),

chlorpyrifos methyl and chlorpyrifos, metalaxyl and malathion were found, together with the synergist piperonyl butoxide used in pesticide formulations. The most frequently detected pesticides were

glyphosate [or its metabolite (aminomethyl)phosphonic acid (AMPA)] and pirimiphos-methyl (8 out of the 13 feeds). Piperonyl butoxide was detected in eight diet samples. Malathion and chlorpyriphos-

ethyl (chlorpyriphos) were found in three samples, chlorpyriphos methyl in two samples and

deltamethrin and metalaxyl in a single sample. The maximum levels of the pesticides reported in this study were all below the concentrations legally permitted in barley, maize, oats, rye, sorghum, wheat,

soya beans and a number of other oilseeds, which are the products most likely used as ingredients in feed. For instance, the highest level of glyphosate reported was 0.140 mg/kg, whereas the legal limits

for these commodities range from 1 to 20 mg/kg.3

Mesnage et al. (2015) analysed the same rodent diets for heavy metals and reported the presence of lead in 12 out of 13 diets (range 0.13–0.58 mg/kg). The lead content in five diet samples was above

0.2 mg/kg, and hence above the Maximum Level (ML) of 0.2 mg/kg specified for lead in cereals, legumes and pulses in Commission Regulation (EC) No 1881/2006 setting maximum levels for

contaminants in foodstuffs.4 Cadmium was found in all samples (range 0.03–0.10 mg/kg) but none of the measured levels exceeded the MLs of 0.1–0.2 mg/kg for cereals, wheat and soya beans. Mercury

was detected in only two samples at 0.01 mg/kg each, which is well below the MLs for (total) mercury

3 Commission Regulation (EU) No 293/2013 of 20 March 2013 amending Annexes II and III to Regulation (EC) No 396/2005 of

the European Parliament and of the Council as regards maximum residue levels for emamectin benzoate, etofenprox, etoxazole, flutriafol, glyphosate, phosmet, pyraclostrobin, spinosad and spirotetramat in or on certain products. OJ, L 96, p. 1–30

4 Commission Regulation (EC) No 1881/2006 of 19 December 2006 setting maximum levels for certain contaminants in foodstuffs. OJ, L 364/5, p. 1–20.



of 0.5–1.0 mg/kg.5 Arsenic was found in only one sample at a level of 0.2 mg/kg. No ML is specified in Commission Regulation (EC) No 1881/2006 for arsenic in foodstuffs. EFSA noted that all the levels for

heavy metals reported by Mesnage et al. (2015) were below the lowest MLs for contaminants in feed

materials and complete feed (at 12% maximum moisture) as specified in Directive 2002/32/EC of the European Parliament and of the Council of 7 May 2002 on undesirable substances in animal feed6 as

amended by Commission Regulation (EU) No 744/2012:7 lead, 5 mg/kg; cadmium, 0.5 mg/kg; arsenic, 2 mg/kg; mercury, 0.1 mg/kg. Likewise, it was noted that all the levels for heavy metals

reported by Mesnage et al. (2015) were below the specifications for maximum contaminant levels in

feed and vehicles for test animals recommended by US-EPA: lead, 1.5 mg/kg; cadmium, 0.16 mg/kg; arsenic, 1 mg/kg; mercury, 0.1 mg/kg.8

Analysis of the commercial rodent diets furthermore revealed the presence of dioxins (range 0.067–0.13 ng/kg wet weight), dioxin-like PCBs (range 0.034–0.15 ng/kg wet weight) and six indicator PCBs

(range 0.530-1.950 ng/kg wet weight for the sum). EFSA noted that all the levels for dioxins and dioxin-like PCBs reported by Mesnage et al. (2015) were below the MLs for contaminants in feed

materials (at 12% maximum moisture) specified in Directive 2002/32/EC as amended by Commission

Regulation (EU) No 744/2012 for dioxins (WHO-PCDD/F-TEQ) (0.75–5 ng/kg) and for the sum of dioxins and dioxin-like PCBs (WHO-PCDD/F-PCB-TEQ) (1–20 ng/kg).

The approach used in this paper to detect and quantify GMOs in diets was considered appropriate; however, EFSA noted that no information on the sampling of the material to be analysed in the diets

or the methods used for the analysis were reported. The authors showed that 11 out of 13 diet

samples contained GMO-derived ingredients, mainly GM soya and GM maize, with a total of 12 GM events identified. All the events are authorised in EU. According to the authors, when present, the

relative amounts of GM ingredients varied from 0.47% to 48% for GM soya and from 1.8% to 35.6% for GM maize. The authors provided relative quantitative data in support of the narrative results on

GMO detection and quantification in the diets tested (Mesnage et al. 2015, Fig. 2B and supplementary data), i.e., the percentage of GM crop (or by-product) on the total crop (or by-product) in the feed.

Therefore, in the absence of detailed quantitative information, no correlation between the amount of

glyphosate-resistant GMOs in the diets and the levels of glyphosate residues could be established (see Fig. 3, Mesnage et al. 2015. p. 10).

The authors specifically focused on the six GM crops resistant to glyphosate (RRS and RRS2 soya bean; GA21, MON88017 and NK603 maize; GT73 oilseed rape). The other GMOs identified were three

glufosinate-resistant crops (DAS1507 and T25 maize; MS8Rf oilseed rape); and five crops expressing

insecticidal Bt proteins (DAS1507, MIR162, MON810, MON863, and MON88017 maize). These were not further discussed by the authors, as regards the presence of herbicides other than glyphosate or

Bt insecticidal proteins, and the interpretation of toxicological studies.

Review of the study by Samsel 2.2.

EFSA noted that the source of one of the studies (‘laboratory studies done by Dr Samsel’) is an

interview provided by Dr Samsel and published on a personal website (farmwars.info).9 Three different rodent diets from Purina were reported to have been tested and levels of glyphosate

reported on the Farm Wars webpage ranged from 0.37 to 0.57 mg/kg. The accuracy of the information on the presence of glyphosate and its metabolite AMPA reported on the farmwars.info

website cannot be confirmed at present nor is the information supported by sufficient detail or a

reference to permit a full scientific review.

5 These levels are limited to fishery products because levels of mercury found in foods, other than fish and seafood, were

judged to be of lower concern (Commission Regulation (EC) No 1881/2006). 6 Directive 2002/32/EC of the European Parliament and of the Council of 7 May 2002 on undesirable substances in animal feed.

OJ L 140, 30.5.2002, p.10. 7 Commission Regulation (EU) No 744/2012 of 16 August 2012 amending Annexes I and II to Directive 2002/32/EC of the

European Parliament and of the Council as regards maximum levels for arsenic, fluorine, lead, mercury, endosulfan, dioxins, Ambrosia spp., diclazuril and lasalocid A sodium and action thresholds for dioxins. OJ, L 219/5.

8 EPA, 1979. Good laboratory practice standards for health effects. Fed. Reg. 44, No. 91, 27535-27354. 9 http://farmwars.info/?p=14095



3. Impact of the paper published in PLoS One (Mesnage et al., 2015)

From the maximum residue levels found in the 13 commercial rodent diets, Mesnage et al. (2015) computed maximum daily intake values for rats for each contaminant using the conversion factor for

the dietary administration of test substances proposed by EFSA (2012). The authors compared the derived daily intakes with the ADI for the substance of interest as a reference dose for computing a

corresponding Hazard Quotient (HQ) and then stated that the ‘sum of the hazard quotients of the pollutants in the diets (…) varied from 15.8 to 40.5. Thus the chronic consumption of these diets can

be considered at risk.’ (Mesnage et al., 2015; p 1). They furthermore concluded that ‘these data may

challenge the use of external control groups in regulatory chronic health risk assessments, because differential diet contaminations artificially enhance background effects and hide significant effects’

(Mesnage et al., 2015; p 12) and challenged the use of historical controls.

Mesnage et al. (2015) made a direct comparison between dietary intake values for rats with ADIs

which are derived for humans. An ADI is usually at least 100-fold lower than the level at which no

adverse effect in rodents is expected. Therefore, the application of the ADI concept to claim the existence of a health risk in rodents or to demonstrate background levels of diseases or disorders in

rodents has no justification. For deriving HQs, it would have been more appropriate to use the No Observed Adverse Effect Level (NOAEL) from long-term animal toxicology studies which represent the

maximum amount given to rodents on a daily basis that does not induce any toxicity. The hazard quotients as calculated in this paper will substantially overestimate the cumulative risk associated with

the consumption of the feed.

The approach of determining hazard quotients for single chemicals and hazard indexes is a pragmatic approach developed by US EPA for the assessment of hazardous waste sites to rank a cumulative risk

(US EPA, 1989; US EPA, 2000) that should not be used to assess the risks related to combined exposure to chemical mixtures. For this purpose, more elaborate procedures should be considered,

taking into account the effect targets and mode of actions of the single chemicals (e.g., cumulative

risk assessment for pesticides) (EFSA, 2014).

A main issue related to GMOs highlighted by Mesnage et al. (2015) was the variable but frequent

presence of GMOs in most of the laboratory rodent chows tested, without appropriate or consistent labelling. Under EU law, the placing on the market of products containing, consisting of or produced

from GMOs is subject to an authorisation procedure laid down by Directive 2001/18/EC10 or Regulation

(EC) No 1829/2003, implemented by Implementing Regulation (EU) No 503/2013.11 EFSA is responsible for carrying out scientific risk assessments of GMOs with regard to human and animal

health and the environment, according to rules set in the above mentioned legal acts and in EFSA guidance documents. As specifically regards the GMO events detected and quantified by Mesnage et

al. (2015), all these have been fully assessed by EFSA and found to be safe and as nutritious as their conventional non GM counterparts; on this basis, these were authorised for import and processing and

for food and feed uses in the EU.12 Therefore, the presence of GMOs in animal feed would not

constitute a safety issue. In addition, the threshold identified by the EU legislator (0.9% per ingredient) for labelling is not a safety threshold and is unrelated to any safety assessment

considerations.

In accordance with OECD protocols (OECD, 2002), and in line with best practices for laboratory animal

diet formulation (National Research Council (US) Subcommittee on Laboratory Animal Nutrition, 1995)

and Directive 2010/63/EU,13 the quality of laboratory rodent chows and their control are considered necessary for all studies on laboratory animals both for nutritional value and limitation of

contamination levels with pesticides, heavy metals and mycotoxins. The results of these analyses are detailed in typical regulatory toxicological study reports. Furthermore, in the case of long-term studies

10 Directive 2001/18/EC of the European Parliament and of the Council of 12 March 2001 on the deliberate release into the

environment of genetically modified organisms and repealing Council Directive 90/220/EEC. OJ, L 106, p. 1–38. 11 Commission Implementing Regulation (EU) No 503/2013 of 3 April 2013 on applications for authorisation of genetically

modified food and feed in accordance with Regulation (EC) No 1829/2003 of the European Parliament and of the Council and amending Commission Regulations (EC) No 641/2004 and (EC) No 1981/2006. OJ, L 157, p. 1–48.

12 http://ec.europa.eu/food/dyna/gm_register/index_en.cfm 13 Directive 2010/63/EU of the European Parliament and of the Council of 22 September 2010 on the protection of animals used

for scientific purposes.



(chronic, carcinogenicity or chronic/carcinogenicity), the relevant OECD protocols specify the need for repeated analysis when feed batch changes occur during studies.

The existence of background neoplastic and non-neoplastic lesions in all the rat and mice strains

routinely used for regulatory long-term toxicity studies has been known for at least five decades (e.g. Prejean et al., 1973; Durbin et al., 1966). The difference in the incidence of such lesions between

strains, their contributing factors and their trends have been studied extensively and are well documented in the literature (e.g., Carlus et al., 2013; Kuroiwa et al., 2013; Ando et al., 2008;

Baldrick, 2005; Brix et al., 2005; Tennekes et al., 2004; Keenan et al., 2002; Eiben and Bomhard,

1999; Haseman et al., 1998). The overall consensus is that different genetic predispositions underlie the development of these lesions for each strain. However, environmental factors such as diet,

housing (individual versus group housing), intrinsic factors such as body weight and survival and, in mice, presence of Helicobacter hepaticus can have an important effect on the incidence of some of

these lesions, including neoplastic lesions. Outbred rat strains such as Sprague-Dawley and Wistar rats often show marked differences in the incidence of different neoplastic lesions depending on their

origin, because being derived from different colonies they will be genetically different (Kacew and

Festing, 1996). Even among inbred strains there is a substantial degree of substrain genetic variability that has recently been revealed through whole genome sequencing. For example, 5,854 single

nucleotide variations have been identified in a genetic variability analysis performed on four substrains of the inbred F344 strain (Hermsen et al., 2015).

While the incidence of most tumour types shows little variability with time, remaining essentially

unchanged over the past 20 years, a time-related drift in the incidence of some tumours such as pituitary and mammary tumours is well documented for some strains (e.g. Carlus et al., 2013;

Kuroiwa et al., 2013; Ando et al., 2008; Baldrick, 2005; Brix et al., 2005; Tennekes et al., 2004; Keenan et al., 2002; Eiben and Bomhard, 1999; Haseman et al., 1998). In general, such genetic drift

is more common with outbred Wistar and Sprague-Dawley rats. Taking into consideration that (i) combinations of residues of pesticides, dioxins and heavy metal change over time and location and (ii)

the genetic drift in tumour incidence varies among rat strains and sub-strains, the proposal by

Mesnage et al. (2015) that contaminants found in commercial rodent diets are responsible for the spontaneous diseases is highly speculative. To experimentally verify the proposal by Mesnage et al.

(2015) that contaminants found in commercial rodent diets are responsible for the spontaneous diseases in rats, would require a full study with controlled levels of the suspected contaminants.

Concurrent controls are always used in regulatory research to establish biological significance in

regulatory studies. The importance of internal historical controls is in monitoring genetic drifts and trends in important parameters such as mortality and body weight within a particular strain (Keenan

et al., 2002). Both the National Toxicology Program (NTP) of the Department of Health and Human Services in the U.S. and the European Agency for Medicines (EMA) recommend the use for this

purpose of historical control data compiled from studies of the last seven and last five years,

respectively.

4. Conclusions

Taking into account the agricultural origin of the feed used for laboratory animals, the presence of GMOs and detectable levels of heavy metals, dioxins and pesticides in the feeds analysed in the paper

by Mesnage et al. (2015) is not unexpected. However, the vast majority of pesticides were absent

(below the limit of detection), and where detected, the levels of pesticides, heavy metals and dioxins were only just above the limit of detection in the feed samples but below regulatory levels.

Furthermore, the GMOs detected have all been authorised in EU. It would be necessary to know the composition of the feed samples and their origin to be able to conclude on possible infringements to

the EU pesticide residue legislation.

In conclusion, the paper by Mesnage et al. (2015) provides a useful addition to the already existing

knowledge in the field. However, no new scientific elements are provided that would impact on the

validity of regulatory feeding tests in the EU.



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Abbreviations

ADI acceptable daily intakes

FAO/WHO Food and Agriculture Organization of the United Nations/World Health Organization

GMO genetically modified organisms

HQ hazard quotient

ML maximum level

MRL maximum residue limits

PTDI provisional tolerable daily intakes

PTMI provisional tolerable monthly intakes

PTWI provisional tolerable weekly intakes



Appendix – Analysis of the pesticide residues identified by Mesnage et al. (2015) in the rodent diets and comparison with identified NOAELs

Mesnage et al. (2015) make a direct comparison between dietary intake values for rats with ADIs

which are derived for humans. An ADI is usually at least 100-fold lower than the level at which no adverse effect in rodents is expected. When comparing the highest reported residue levels for

pesticides with the respective relevant long term NOAELs used to set reference values for the eight individual pesticide active substances detected in the diet samples, the computed intake values were

below the NOAELs of the respective active substances, often by several orders of magnitude.

For pirimiphos methyl, the highest residue level was 1.8 mg/kg diet sample, this is considered

equivalent to 0.090 to 0.162 mg/kg body weight (b.w.) per day substance intake for rats pending on

their age (EFSA, 2012). The relevant long term NOAEL identified in this species was 0.4 mg/kg b.w. per day in rat (EFSA, 2005). For deltamethrin, the highest dietary level of 0.141 mg/kg reported would

yield an intake of 0.007 to 0.013 mg/kg b.w. per day in rats which is well below the overall NOAEL of 1 mg/kg b.w. per day set in the legislation (EC, 2002a) and confirmed by EFSA based on its potential

developmental neurotoxicity (EFSA, 2009a). The authors refer to the presence of piperonyl butoxide,

which is added to insecticides as a synergist to prolong the effects of insecticides by inhibiting their metabolism and detoxification. The substance is not considered a pesticide active substance, but the

lowest NOAEL in the rat was 30 mg/kg b.w. per day (IPCS INCHEM, 1995). Therefore, its highest residual level at 1 mg/kg (equivalent to an intake of 0.05 to 0.09 mg/kg b.w. per day in rats) does not

raise a health concern to rodents, even in the presence of other insecticide active substances. The residue levels of glyphosate and of glyphosate’s metabolite AMPA were summed to a maximum of

0.37 mg/kg equivalent to 0.019 to 0.033 mg/kg b.w. per day in rats when an overall long term NOAEL

was set at 31 mg/kg b.w. per day (EC, 2002b) to establish the ADI for glyphosate. The overall NOAEL for chlorpyrifos was revised in 2014 to 0.1 mg/kg b.w. per day (EFSA, 2014), while the NOAEL for

chlorpyrifos-methyl is set in the legislation at 1 mg/kg b.w. per day (EC, 2015); no health concerns are therefore expected from residual intakes of 0.001–0.002 mg/kg b.w. per day for chlorpyrifos (from

the highest residue level of 0.023 mg/kg) and 0.003–0.005 mg/kg b.w. per day for chlorpyrifos-methyl

(from the highest residue level of 0.059 mg/kg). For malathion, a relevant NOAEL of 29 mg/kg b.w. per day was identified (EFSA, 2009b) versus a residual intake of 0.009–0.015 mg/kg b.w. per day

(from the highest residue level of 0.17 mg/kg). For metalaxyl, the relevant NOAEL in rats was 9.4 mg/kg b.w. per day (EC, 2010; EFSA, 2015) and the maximum residues detected (0.02 mg/kg) are

equivalent to 0.001–0.002 mg/kg b.w. per day. Accordingly, the highest levels of pesticide residues

would not have an impact on the health of the animals fed these diets.

Documents

Review of results published by Mesnage et al. (2015) in PLoS ONE