Scientific Opinion of the Panel on Additives and Products or Substances used in Animal Feed. Adopted on 19 September 2007

Transcription

1 The EFSA Journal (2007) 549, 1-11 Safety and efficacy of Toyocerin (Bacillus cereus var. Toyoi) as a feed additive for turkeys 1 Scientific Opinion of the Panel on Additives and Products or Substances used in Animal Feed (Question No EFSA-Q ) Adopted on 19 September 2007 PANEL MEMBERS Georges Bories, Paul Brantom, Joaquim Brufau de Barberà, Andrew Chesson, Pier Sandro Cocconcelli, Bogdan Debski, Noël Dierick, Anders Franklin, Jürgen Gropp, Ingrid Halle, Christer Hogstrand, Joop de Knecht, Lubomir Leng, Anne-Katrine Lundebye Haldorsen, Alberto Mantovani, Miklós Mézes, Carlo Nebbia, Walter Rambeck, Guido Rychen, Atte von Wright and Pieter Wester SUMMARY Toyocerin is a zootechnical additive (functional group: gut flora stabilisers) composed of spores of a strain of Bacillus cereus. The product has already been granted permanent authorisation for use in piglets up to four months of age, in pigs for fattening and in sows from one week prior to farrowing until weaning. It is also permanently authorised for use in cattle, chickens and rabbits for fattening and is provisionally authorised for use in laying hens, calves and breeding does. The applicant is now seeking authorisation for the use of Toyocerin in diets for turkeys from one day of age to slaughter at an inclusion level of x 10 9 CFU kg -1 complete feedingstuff. The safety of Toyocerin for consumers of food products of animals fed diets containing Toyocerin, for users of the product and for the wider environment has been established as part of the previous authorisations. The FEEDAP Panel is not aware of any additional data which would require a reconsideration of these assessments. Consequently, in this opinion, only the issues of safety and efficacy introduced by the extension of use to turkeys for fattening are considered. Efficacy at the highest recommended dose (1 x 10 9 CFU kg -1 ) has been demonstrated in all four studies in which the dose was tested. Four studies also incorporated the lowest recommended dose (0.2 x 10 9 CFU kg -1 ) with significant improvements in relevant parameters in three of the trials. However, two of the three studies in which significant benefits were seen involved the slow-growing Bronze turkey and only one trial a breed and growth rate more typical of European production systems. The degree to which results obtained with the slow-growing Bronze turkey can be extrapolated to other breeds and production systems in practice is open to 1 For citation purposes: Scientific Opinion of the Panel on Additives and Products or Substances used in Animal Feed (FEEDAP) on the safety and efficacy of Toyocerin (Bacillus cereus var. Toyoi) as a feed additive for turkeys. The EFSA Journal (2007) 549, 1-11 European Food Safety Authority, 2007

2 question. However, the FEEDAP Panel considers that, on balance, the minimum recommended dose (0.2 x 10 9 ) is effective in turkeys for fattening. No specific data on the compatibility of Toyocerin with authorised coccidiostats in turkeys was presented. However, the FEEDAP Panel considers it is reasonable to assume that compatibilities established in chickens for fattening would also apply to turkeys for fattening. Based on the two tolerance studies provided in which turkeys tolerated up to a ten-fold overdose of the product, and the known tolerance to substantially higher doses shown by chickens for fattening, the FEEDAP Panel considers that Toyocerin is safe for turkeys when used at the maximum recommended dose. Key words: zootechnical additive, gut flora stabiliser, Toyocerin, Bacillus cereus, probiotic, micro-organism, turkeys for fattening, efficacy, safety, compatibility with coccidiostats The EFSA Journal (2007) 549, 2-11

3 TABLE OF CONTENTS Panel Members...1 Summary...1 Table of Contents...3 Background...4 Terms of reference...4 Assessment Introduction Stability of the additive in relation to the feeding of turkeys Evaluation of the analytical methods by the Community Reference Laboratory (CRL) Efficacy Compatibility with coccidiostats Safety for turkeys Post-market monitoring...9 Conclusions and Recommendations...9 Documentation provided to EFSA...9 References...9 The EFSA Journal (2007) 549, 3-11

4 BACKGROUND Regulation (EC) No 1831/ establishes the rules governing the Community authorisation of additives for use in animal nutrition. In particular, Article 4(1) of that Regulation lays down that any person seeking an authorisation for a feed additive or for a new use of a feed additive shall submit an application in accordance with Article 7. The European Commission received a request from the company Rubinum SA 3 for authorisation of the product Toyocerin, to be used as a feed additive for turkeys for fattening, (category: zootechnical additives; functional group: gut flora stabilizers) under the conditions mentioned under Table 1. According to Article 7(1) of Regulation (EC) No 1831/2003, the Commission forwarded the application to the European Food Safety Authority (EFSA) as an application under Article 4.1 (authorisation of a feed additive or new use of a feed additive). EFSA received directly from the applicant the technical dossier in support of this application. According to Article 8 of that Regulation, EFSA, after verifying the particulars and documents submitted by the applicant, shall undertake an assessment in order to determine whether the feed additive complies with the conditions laid down in Article 5. The particulars and documents in support of the application were considered valid by EFSA as of 15 June The additive Toyocerin is a preparation of Bacillus cereus var. Toyoi (NCIMB 40112/CNCM I-1012). This product is already permanently authorised for its use in piglets up to four months, pigs for fattening, sows from one week prior to farrowing until weaning, cattle for fattening, chickens for fattening and rabbits for fattening (E 1701), and was provisionally authorised (No 1) for laying hens, calves, and breeding does. The Scientific Committee on Animal Nutrition (SCAN) issued an opinion on the use of Toyocerin on toxin production and resistance to antibiotics on 5 December 2001 (EC, 2001). EFSA issued an opinion on the efficacy of this product in feeds for pigs for fattening (EFSA, 2004), one on the modification of terms of authorisation of this additive to allow its use in chicken feed with the coccidiostats diclazuril, narasin-nicarbazin and maduramicin ammonium (EFSA, 2005) and another opinion on the safety and efficacy for sows from service to weaning (EFSA, 2007). TERMS OF REFERENCE According to Article 8 of Regulation (EC) No 1831/2003, EFSA shall determine whether the feed additive complies with the conditions laid down in Article 5. Therefore, EFSA shall deliver an opinion on the efficacy and the safety for the target animals, consumer, user and the environment of the product Toyocerin, which is a preparation of Bacillus cereus var. Toyoi (NCIMB 40112/CNCM I-1012) when used under the conditions described in Table 1. ACKNOWLEDGEMENTS The European Food Safety Authority wishes to thank the members of the Working Group on Micro-organisms for the preparation of this opinion. 2 OJ L 268, , p.29 3 Rubinum SA. Avda. De La Llana, 123. E Rubí Barcelona, Spain The EFSA Journal (2007) 549, 4-11

5 Table 1. Register entry as proposed by the applicant Additive Bacillus cereus var. toyoi NCIMB 40112/CNCM I-1012 Registration number/ec No/No Category of additive Functional group of additive E 1701 Zootechnical additive Gut flora stabiliser Description Composition, description Bacillus cereus var. toyoi NCIMB 40112/CNCM I Chemical formula Preparation of viable spores of Bacillus cereus var. toyoi containing a minimum of 1 x CFU g -1 additive Purity criteria Method of analysis Trade name Toyocerin Name of the holder of authorisation Conditions of use Species or category of animal Maximum Age Minimum content Maximum content CFU kg -1 of complete feedingstuffs Withdrawal period Turkeys From 1 day old until slaughtering 0.2 x x 10 9 Specific conditions or restrictions for use Specific conditions or restrictions for handling Post market monitoring Specific conditions for use in complementary feedingstuffs Other provisions and additional requirements for the labelling In the directions for use of the additive and premixture indicate the storage temperature, storage life and stability to pelleting. May be used in compound feed containing the permitted coccidiostats: monensin sodium, lasalocid sodium, salinomycin sodium, decoquinate, robenidine, narasin, halofuginone, diclazuril, narasin/nicarbazin, maduramicin ammonium. When handling use protective mask and goggles Every lot of Toyocerin placed onto the market is registered in order to be able to trace its destination until the final customer (feed manufacturer or premixture manufacturer) Maximum Residue Limit (MRL) Marker residue Species or category of animal Target tissue(s) or food products Maximum content in tissues The EFSA Journal (2007) 549, 5-11

6 ASSESSMENT 1. Introduction Toyocerin is a zootechnical additive (functional group: gut flora stabilisers) composed of spores of a strain of Bacillus cereus mixed with maize flour and calcium carbonate in an approximate ratio of 6:4:90 by weight. The strain has been deposited in four recognised culture collections, initially in Japan (FERM-P No 1214) and subsequently in Spain (CECT 876), the UK (NCIMB 40112) and France (CNCM ). It is not genetically modified. The product has already been granted permanent authorisation for use in piglets up to four months of age, in pigs for fattening and in sows from one week prior to farrowing until weaning. It is also permanently authorised for use in cattle, chickens and rabbits for fattening and has been provisionally authorised for use in laying hens, calves, and breeding does. The applicant is now seeking authorisation for the use of Toyocerin in diets for turkeys from one day of age to slaughter at an inclusion level of x 10 9 CFU kg -1 complete feedingstuff. The safety of Toyocerin for consumers of food products of animals fed diets containing Toyocerin, for users of the product and for the wider environment has been established as part of the previous authorisations (see background). The FEEDAP Panel is not aware of any additional data which would require a reconsideration of these safety assessments. Consequently, in this opinion, only the issues of safety and efficacy introduced by the extension of use to turkeys for fattening are considered. 2. Stability of the additive in relation to the feeding of turkeys Product stability during storage, in premixes and in a variety of complete feeds together with data on the ease of homogeneous mixing, has been extensively described and assessed in the context of the previous requests for authorisation. Spores of Bacillus species are inherently resistant to heat, light and desiccation, and previous studies have shown that they retain viability for long periods under the conditions encountered in the preparation and storage of feedingstuffs. This was confirmed for turkey feeds in two studies provided by the applicant, the first measuring viability after pelleting and the second viability after three-month storage of the pelleted feed. In both cases, virtually 100 % of the initial activity was recovered. Although no specific data on survival in premixes for turkeys was provided, these are very similar in composition to premixes for other animal categories. Consequently, results previously obtained with other premixes showing essentially 100 % recovery after extended periods can be safely extrapolated to premixes for turkeys. A single study with four batches of three feed formulations confirmed that the product is able to form homogeneous mixtures in feeds for turkeys. 3. Evaluation of the analytical methods by the Community Reference Laboratory (CRL) EFSA has verified the CRL report as it relates to the methods used for the control of the active agent in animal feed. The Executive Summary of the CRL report can be found in the Appendix. 4. Efficacy Data from a total of five trials with turkeys for fattening was made available by the applicant. The first three studies were made at experimental stations at different locations in Europe and the final two at commercial farms in southern Europe under what was described as normal The EFSA Journal (2007) 549, 6-11

7 production conditions. The choice of breeds in the first three studies reflected those commonly used in the more intensive production systems (male BIG 6, female BUT 9 and male BUT 9, respectively) in Europe. In contrast, the two farm-based trials were made with Bronze turkeys, a slow maturing breed often selected by organic producers for the quality of the meat and normally marketed at around 6-9 kg live weight. The studies followed a similar design with diets based on cereals/soybeans, formulated to match needs at different phases of growth. The concentration of the additive in test diets was confirmed by analysis. In each case, day-old birds were randomly assigned to the experimental groups. Trial 2 was slightly unusual since numbers (31)/pen were reduced to 25 after 28 days by removal of those individuals whose bodyweight most differed from the pen average. Coccidiostats were used in trials 1 and 3 (halofuginone and monensin). No mention is made of the use of a coccidiostat in trial 2 but all birds were vaccinated (against rhinotracheitis and fowl pox). Feed intake and bodyweight was measured at the end of each fattening stage and from this data, weight gain and feed:gain ratio calculated (Table 2). In addition, water consumption was monitored in the two farm-based studies and carcass weight measured. Table 2. Trial (Duration in weeks) 1 4 (22) 2 5 (12) 3 6 (18) 4 7 (~6 months) 5 8 (~6 months) Summary of the production results seen in the five efficacy trials Total number of birds (replicates/ treatment x birds/replicate) 540 (9 x 15) Dose (CFU kg -1 ) Weight gain (kg) Final body weight (kg) Feed intake (kg) Feed:gain a a 0.2 x ab a 0.5 x ab a 1.0 x b b a 2.17 a 744 (12 x 31) 1.0 x b 2.13 b 396 (4 x33) 300 (10 x 10) a a a 2.61 a 0.2 x b b b 2.56 b 1.0 x c c a 2.53 c a 4.86 a a 0.2 x b 6.03 b b 1.0 x c 6.14 c b a 4.73 a a 5.64 a 200 (5 x 20) 0.2 x b 5.25 b b 3.90 b a, b, c : Means in a column within a trial not sharing a common superscript are significantly different (P < 0.05) Mortality was low in most cases, although the 12.4 % overall mortality seen in trial 1 was exceptional. Losses appeared randomly distributed across the experimental groups and were ascribed to cannibalism, wing fracture and leg damage. The lowest mortality was seen in trial 2 (1.8 %) although it is unclear whether this was measured only after the homogenisation of the birds at 28 days. 4 Technical dossier/section III/Annex 95 5 Technical dossier/section III/Annex 96 6 Technical dossier/section III/Annex 97 7 Technical dossier/section III/Annex Technical dossier/section III/Annex 127 The EFSA Journal (2007) 549, 7-11

8 Carcass weight (and percentage of live weight) was significantly improved in all treatment groups compared to control groups in both of the farm-based studies (trials 4 and 5). With the highest recommended dose (used in four of the five studies) weight gain and feed intake was significantly improved in three studies and the efficacy of feed utilisation was improved in all four. The lowest recommend dose was also used in four studies with significant improvements in measured parameters in three of the trials. However, two of the three studies in which significant benefits were seen involved the Bronze turkey and only one trial involved a breed and growth rate more typical of European production systems. The degree to which results obtained with the slow-growing Bronze turkey can be extrapolated to other breeds and production systems in practice is open to question. However, the FEEDAP Panel considers that, on balance, the minimum recommended dose (0.2 x 10 9 ) is effective in turkeys for fattening. 5. Compatibility with coccidiostats The proposed registry entry includes the provision for use of the additive in compound feed containing the permitted coccidiostats: monensin sodium, lasalocid sodium, salinomycin sodium, decoquinate, robenidine, narasin, halofuginone, diclazuril, narasin/nicarbazin, maduramicin ammonium. No evidence was provided to support the compatibility of these coccidiostats with the additive in turkeys. However, the FEEDAP Panel considers it is reasonable to assume the compatibilities seen in chickens for fattening would also apply to turkeys for fattening. 6. Safety for turkeys Three tolerance tests were reported, all studying the effect of an intended ten-fold overdose of Toyocerin (1 x CFU kg -1 ). However, one study lasted for only two weeks and is not further considered. In the remaining two studies, BUT9 turkeys were allocated to one of two groups, an untreated control group fed a basal diet and a treatment group fed the basal diet supplemented with Toyocerin at 1 x CFU kg -1 complete feed (confirmed by analysis). The first 9 of the two retained studies involved 400 male birds (eight replicates of 25 turkeys per treatment) and lasted for a period of 16 weeks; the second, birds (ten replicates of four turkeys per treatment) for 12 weeks. Observations in both cases were restricted to regular measurement of performance characteristics (body weight, weight gain, feed intake and mortality) supported by daily checks on health. Samples of intestinal contents were taken from eight birds per treatment for microbiological analysis (Escherichia coli numbers) after 4, 33 and 76 days and at the end of the first study. No adverse effects on the performance parameters measured were seen in any study compared to the negative control. Mortality was not significantly affected by treatment, although the control group in the second study had a high rate of mortality (12.5 %) compared to the treatment group (2.5 %). Minor differences were seen on E. coli counts in caecum or jejunum during the 16 weeks but counts were virtually identical at the end of the study. Conclusions on target species safety Based on the two tolerance studies provided in which turkeys tolerated up to a ten-fold overdose of the product, and the known tolerance to substantially higher doses shown in 9 Technical dossier/section IV/Annex Technical dossier/section IV/Annex 100 The EFSA Journal (2007) 549, 8-11

9 chickens for fattening in which a greater range of end-points were measured, the FEEDAP Panel considers that Toyocerin is safe for turkeys at the maximum recommended dose. 7. Post-market monitoring No specific risks associated with the use of the product are foreseen, and therefore the FEEDAP Panel does not see the need for specific requirements for a post-market monitoring plan other than those detailed in the Feed Hygiene Regulation [Regulation (EC) No 183/2005] and Good Manufacturing Practice. CONCLUSIONS AND RECOMMENDATIONS CONCLUSIONS Efficacy at the highest recommended dose (1 x 10 9 CFU kg -1 ) has been demonstrated in all four studies in which the dose was tested. Four studies also incorporated the lowest recommended dose (0.2 x 10 9 CFU kg -1 ) with significant improvements in relevant parameters in three of the trials. However, two of the three studies in which significant benefits were seen involved the Bronze turkey and only one trial a breed and growth rate more typical of European production systems. On balance, the FEEDAP Panel considers that the minimum recommended dose is effective in turkeys for fattening. The FEEDAP Panel considers it is reasonable to assume that the compatibilities with coccidiostats established in chickens for fattening would also apply to turkeys for fattening. Based on the two tolerance studies provided, in which turkeys tolerated up to a ten-fold overdose of the product, and the known tolerance to substantially higher doses shown in chickens for fattening, the FEEDAP Panel considers that Toyocerin is safe for turkeys when used at the maximum recommended dose. DOCUMENTATION PROVIDED TO EFSA 1. Application dossier seeking an authorisation for a new use of Toyocerin in turkeys for fattening. March Submitted by Rubinum SA. 2. Evaluation report of the Community Reference Laboratory feed additives authorisation on the methods(s) of analysis for Toyocerin for turkeys for fattening. 3. Comments from Member States received through the EFSAnet. REFERENCES EC (European Commission), Scientific Committee on Animal Nutrition. Report of the Scientific Committee on animal Nutrition on product Toyocerin for use as feed additive. < EFSA (European Food Safety Authority), Opinion of the Scientific Panel on additives and products or substances used in animal feed (FEEDAP) on the efficacy of product Toyocerin for pigs for fattening. < EFSA (European Food Safety Authority), Opinion of the Scientific Panel on additives and products or substances used in animal feed (FEEDAP) on the modification of terms of authorisation of the micro-organism preparation of Bacillus cereus var. toyoi (NCIMB 40112/CNCM I-1012) (Toyocerin) authorised as a feed additive in accordance with Council The EFSA Journal (2007) 549, 9-11

10 Directive 70/524/EEC. < EFSA (European Food Safety Authority), Opinion of the Panel on additives and products or substances used in animal feed (FEEDAP) on the safety and efficacy of the product Toyocerin (Bacillus cereus var. Toyoi) as a feed additive for sows from service to weaning, in accordance with Regulation (EC) No 1831/2003. < The EFSA Journal (2007) 549, 10-11

11 APPENDIX Executive Summary of the Evaluation Report of the Community Reference Laboratory Feed Additives Authorisation on the Method(s) of Analysis for Toyocerin for turkeys for fattening. In the current application authorisation is sought for the microbial feed additive Toyocerin under the category 'zootechnical additives', functional group 'gut flora stabiliser' according to Annex I of Regulation (EC) No 1831/2003. Specifically, the use of Toyocerin for turkeys for fattening from day one until slaughtering is requested. Toyocerin contains a minimum of 1 x colony forming units (c.f.u.) of viable spores of Bacillus cereus var. toyoi NCIMB 40112/CNCM I-1012 per gram (g). The feed additive is intended to be mixed into complete feedingstuffs at a final concentration of 0.2 to 1 x 10 9 c.f.u./kg. For the quantification of the active agent (Bacillus cereus var. toyoi NCIMB 40112/CNCM I- 1012) of Toyocerin in the feed additive, premixtures and feedingstuffs appropriate enumeration methods were proposed by the applicant. Analysis data confirmed an appropriate method performance in a second laboratory. For official controls regarding the quantitative determination of the active agent in the feed additive, premixtures and feedingstuffs, another surface plate count enumeration method is recommended which includes a heat-treatment of the initial sample suspension to inactivate vegetative cells and uses subsequently a non-selective agar. This method has been fully ringtrial validated (J.AOAC Int. 2003, 86, ). The method s performance characteristics revealed standard deviations for repeatability (s r ) and reproducibility (s R ) of around log 10 and log 10 calculated from the base 10 logarithms of the measured c.f.u./g premixture or feedingstuff, respectively. The limits of quantification (LOQ) of this method are 100 c.f.u./g feed additive or premixture and 10 7 c.f.u./kg feedingstuff which is well below the minimum anticipated target level of application. The identity of the bacterial strain, Bacillus cereus var. toyoi NCIMB 40112/CNCM I-1012, was analysed by a range of techniques including biochemistry, phage-typing, molecular methods and pyrolysis mass spectrometry. Pulsed field gel electrophoresis (PFGE) is a generally recognised standard methodology for microbial identification and is therefore considered suitable for official controls in the frame of the authorisation. On the basis of the supplied documentation, no supplementary experimental work (testing or method validation) is required. The EFSA Journal (2007) 549, 11-11