Conclusion on the peer review of the pesticide risk assessment of the active substance Pseudomonas sp. strain DSMZ

Transcription

1 EFSA Journal 2012;10(12):2954 CONCLUSION ON PESTICIDE PEER REVIEW Conclusion on the peer review of the pesticide risk assessment of the active substance Pseudomonas sp. strain 1 ABSTRACT European Food Safety Authority 2 European Food Safety Authority (EFSA), Parma, Italy The conclusions of the European Food Safety Authority (EFSA) following the peer review of the initial risk assessments carried out by the competent authority of the rapporteur Member State the Netherlands, for the pesticide active substance Pseudomonas sp. strain are reported. The context of the peer review was that required by Commission Regulation (EU) No 188/2011. The conclusions were reached on the basis of the evaluation of the representative uses of Pseudomonas sp. strain as a fungicide on seed potatoes, flowers, tomatoes, cucumbers, peppers, eggplant, lettuce and cabbage. The reliable endpoints concluded as being appropriate for use in regulatory risk assessment, derived from the available studies and literature in the dossier peer reviewed, are presented. Missing information identified as being required by the regulatory framework is listed. Concerns are identified. European Food Safety Authority, 2012 KEY WORDS Pseudomonas sp. strain, peer review, risk assessment, pesticide, fungicide 1 On request from the European Commission, Question No EFSA-Q , approved on 12 November Correspondence: pesticides.peerreview@efsa.europa.eu Suggested citation: European Food Safety Authority; Conclusion on the peer review of the pesticide risk assessment of the active substance Pseudomonas sp. strain. EFSA Journal 2012;10(12):2954. [32 pp.] doi: /j.efsa Available online: European Food Safety Authority, 2012

2 SUMMARY Pseudomonas sp. strain is a new active substance for which in accordance with Article 6(2) of Council Directive 91/414/EEC the Netherlands (hereinafter referred to as the RMS ) received an application from Sourcon-Padena GmbH & Co.KG for approval. Complying with Article 6(3) of Directive 91/414/EEC, the completeness of the dossier was checked by the RMS. The European Commission recognised in principle the completeness of the dossier by Commission Decision Commission Decision 2008/599/EC of 4 July The RMS provided its initial evaluation of the dossier on Pseudomonas sp. strain in the Draft Assessment Report (DAR), which was received by the EFSA on 3 November The peer review was initiated on 5 October 2011 by dispatching the DAR for consultation of the Member States and the applicant Sourcon-Padena GmbH & Co.KG. Following consideration of the comments received on the DAR, it was concluded that EFSA should conduct an expert consultation in the areas of mammalian toxicology and environmental fate and behaviour and EFSA should adopt a conclusion on whether Pseudomonas sp. strain can be expected to meet the conditions provided for in Article 5 of Directive 91/414/EEC, in accordance with Article 8 of Commission Regulation (EU) No 188/2011. The conclusions laid down in this report were reached on the basis of the evaluation of the representative uses of Pseudomonas sp. strain (Proradix) as a fungicide on seed potatoes, flowers, tomatoes, cucumbers, peppers, eggplant, lettuce and cabbage as proposed by the applicant. Full details of the representative uses can be found in Appendix A to this report. A data gap was identified in the section identity. No data gaps were identified for the section physical and chemical properties and analytical methods. A data gap has been identified for data to prove that no interferences will be produced with methods for the determination of microbiological contamination of water given in directive 98/83/EC, in particular with the method for the analysis of Pseudomonas aeruginosa in drinking water (pren ISO 12780). The available data package in not sufficient to characterize the toxicity, infectivity and pathogenicity of Pseudomonas sp. strain by the inhalation and intraperitoneal routes or to set an AOEL, and consequently the operator and worker risk assessment to Pseudomonas sp. strain DSMZ could not be conducted. The worker risk assessment to metabolites could not be finalised, pending the finalisation of the environmental risk assessment of metabolites produced by Pseudomonas sp strain and the conclusion of their potential toxicity. Additionally, information to show that genetic transfer from Pseudomonas sp. strain will not lead to adverse effects on human and animal health, including resistance to known therapeutics, and will not lead to unacceptable effects on the environment needs to be provided. The consumer risk assessment could not be finalised, pending the submission of further information on the presence in plants of the metabolites produced by Pseudomonas sp. strain after application and on their the toxicity. A data gap for data for information to address persistence and multiplication in bulk soil of Pseudomonas sp strain to fulfil the data requirement specified in Annex II B, Soil of directive 91/414/EEC has been identified. The risk to aquatic organisms can be characterized as low and the potential for groundwater contamination can be considered low only on the basis of the assumption that exposure can be precluded for the representative uses proposed (potato seed treatment and glasshouse uses). Management measures tailored to local practice and legislation would need to be put in place to control the waste disposal of spent application solution and to prevent accidental spillage entering sewers or surface water drains. Further data to assess exposure to operators and EFSA Journal 2012;10(12):2954 2

3 workers through air may be needed, pending the outcome of the toxicological assessment. Metabolites, produced in the environment by Pseudomonas sp. strain need to be identified and assessed for their toxicological and ecotoxicological relevance, in particular the formation and fate of pyochelin (Pseudomonas aeruginosa grow promoter) needs to be addressed. For exposure to the parent substance, Pseudomonas sp. strain, a low risk to birds, aquatic organisms, bees and earthworms was concluded. Data gaps were identified to address the risk to soil microorganisms and to organisms involved in sewage treatment processes. EFSA Journal 2012;10(12):2954 3

4 TABLE OF CONTENTS Abstract... 1 Summary... 2 Table of contents... 4 Background... 5 The identity of the microorganism and the properties of the formulated product... 7 Conclusions of the evaluation Identity of the microorganism/biological properties/physical and technical properties and methods of analysis Mammalian toxicity Residues Environmental fate and behaviour Fate and behaviour in the environment of the microorganism Fate and behaviour in the environment of any relevant metabolite formed by the microorganism under relevant environmental conditions Ecotoxicology Overview of the risk assessment of compounds listed in residue definitions triggering assessment of effects data for the environmental compartments Soil Ground water Surface water and sediment Air List of studies to be generated, still ongoing or available but not peer reviewed Particular conditions proposed to be taken into account to manage the risk(s) identified Concerns Issues that could not be finalised Critical areas of concern Overview of the concerns identified for each representative use considered References Appendices Abbreviations EFSA Journal 2012;10(12):2954 4

5 BACKGROUND In accordance with Article 80(1)(a) of Regulation (EC) No 1107/2009, 3 Council Directive 91/414/EEC 4 continues to apply with respect to the procedure and conditions for approval for active substances for which a decision recognising in principle the completeness of the dossier was adopted in accordance with Article 6(3) of that Directive before 14 June Commission Regulation (EU) No 188/ (hereinafter referred to as the Regulation ) lays down the detailed rules for the implementation of Council Directive 91/414/EEC as regards the procedure for the assessment of active substances which were not on the market on 26 July This regulates for the European Food Safety Authority (EFSA) the procedure for organising the consultation of Member States and the applicant for comments on the initial evaluation in the Draft Assessment Report (DAR) provided by the rapporteur Member State (RMS), and the organisation of an expert consultation, where appropriate. In accordance with Article 8 of the Regulation, EFSA is required to adopt a conclusion on whether the active substance is expected to meet the conditions provided for in Article 5 of Directive 91/414/EEC within 4 months from the end of the period provided for the submission of written comments, subject to an extension of 2 months where an expert consultation is necessary, and a further extension of upto 8 months where additional information is required to be submitted by the applicant(s) in accordance with Article 8(3). In accordance with Article 6(2) of Council Directive 91/414/EEC the Netherlands (hereinafter referred to as the RMS ) received an application from Sourcon-Padena GmbH & Co.KG for approval of the active substance Pseudomonas sp. strain. Complying with Article 6(3) of Directive 91/414/EEC, the completeness of the dossier was checked by the RMS. The European Commission recognised in principle the completeness of the dossier by Commission Decision 2008/599/EC of 4 July The RMS provided its initial evaluation of the dossier on Pseudomonas sp. strain in the DAR, which was received by the EFSA on 3 November 2009 (Netherlands, 2009). The peer review was initiated on 5 October 2011 by dispatching the DAR to Member States and the applicant Sourcon- Padena GmbH & Co.KG for consultation and comments. In addition, the EFSA conducted a public consultation on the DAR. The comments received were collated by the EFSA and forwarded to the RMS for compilation and evaluation in the format of a Reporting Table. The applicant was invited to respond to the comments in column 3 of the Reporting Table. The comments and the applicant s response were evaluated by the RMS in column 3. The need for expert consultation and the necessity for additional information to be submitted by the applicant in accordance with Article 8(3) of the Regulation were considered in a telephone conference between the EFSA, the RMS, and the European Commission on 30 January On the basis of the comments received, the applicant s response to the comments and the RMS s evaluation thereof it was concluded that additional information should be requested from the applicant and that the EFSA should organise an expert consultation in the areas of mammalian toxicology and environmental fate and behaviour. 3 Regulation (EC) No 1107/2009 of the European Parliament and of the Council of 21 October 2009 concerning the placing of plant protection products on the market and repealing Council Directives 79/117/EEC and 91/414/EEC. OJ No L 309, , p Council Directive 91/414/EEC of 15 July 1991 concerning the placing of plant protection products on the market. OJ L 230, , p. 1-32, as last amended. 5 Commission Regulation (EU) No 188/2011 of 25 February 2011 laying down detailed rules for the implementation of Council Directive 91/414/EEC as regards the procedure for the assessment of active substances which were not on the market 2 years after the date of notification of that Directive. OJ No L 53, , p Commission Decision 2008/599/EC of 4 July 2008 recognising in principle the completeness of the dossier submitted for detailed examination in view of the possible inclusion of Pseudomonas sp. strain in Annex I to Council Directive 91/414/EEC. OJ No L 193, , p EFSA Journal 2012;10(12):2954 5

6 The outcome of the telephone conference, together with EFSA s further consideration of the comments is reflected in the conclusions set out in column 4 of the Reporting Table. All points that were identified as unresolved at the end of the comment evaluation phase and which required further consideration, including those issues to be considered in an expert consultation, and the additional information to be submitted by the applicant, were compiled by the EFSA in the format of an Evaluation Table. The conclusions arising from the consideration by the EFSA, and as appropriate by the RMS, of the points identified in the Evaluation Table, together with the outcome of the expert consultation where this took place, were reported in the final column of the Evaluation Table. A final consultation on the conclusions arising from the peer review of the risk assessment took place with Member States via a written procedure in October This conclusion report summarises the outcome of the peer review of the risk assessment on the active substance and the representative formulation evaluated on the basis of the representative uses as a fungicide on seed potatoes, flowers, tomatoes, cucumbers, peppers, eggplant, lettuce and cabbage, as proposed by the applicant. A list of the relevant end points for the active substance as well as the formulation is provided in Appendix A. In addition, a key supporting document to this conclusion is the Peer Review Report, which is a compilation of the documentation developed to evaluate and address all issues raised in the peer review, from the initial commenting phase to the conclusion. The Peer Review Report (EFSA, 2012) comprises the following documents, in which all views expressed during the course of the peer review, including minority views, can be found: the comments received on the DAR, the Reporting Table (6 February 2012), the Evaluation Table (2 October 2012), the reports of the scientific consultation with Member State experts (where relevant), the comments received on the assessment of the additional information (where relevant), the comments received on the draft EFSA conclusion. Given the importance of the DAR including its addendum (compiled version of September 2012 containing all individually submitted addenda (Netherlands, 2012)) and the Peer Review Report, both documents are considered respectively as background documents A and B to this conclusion. EFSA Journal 2012;10(12):2954 6

7 THE IDENTITY OF THE MICROORGANISM AND THE PROPERTIES OF THE FORMULATED PRODUCT Pseudomonas sp. strain bacteria are deposited at the culture collection of the German Collection of Micro-organisms and Cell Cultures (DSMZ) with the accession number. It is a wild-type strain isolated from a lettuce field in Germany. The representative microbial pest control product (MPCP) for the evaluation was Proradix WP, a wettable powder (WP) containing g/kg (min. 6.6 x CFU/kg) Pseudomonas sp. strain. The representative uses evaluated comprise indoor and field applications as a fungicide on seed potatoes and glasshouse applications by pouring around stems of young plants of flowers, tomatoes, cucumbers, peppers, eggplant, lettuce and cabbage, against soil borne fungi. Full details of the GAP can be found in the list of end points in Appendix A. CONCLUSIONS OF THE EVALUATION 1. Identity of the microorganism/biological properties/physical and technical properties and methods of analysis The following guidance documents were followed in the production of this conclusion: OECD Issue Paper on Microbial Contaminant Limits for Microbial Pest Control Products, Series on Pesticides No. 65, ENV/JM/MONO(2011)43 (OECD, 2011). The active substance content in the material used to produce the plant protection product should be not less than 3 x CFU/kg. It should be noted that the current taxonomic classification may change following scientific progress. Identification of Pseudomonas sp. strain at strain level can be done by a ribotyping method. There is no direct evidence of a relationship to known animal or human pathogens and only distant relationship to some plant pathogens. The toxicological profiles of the metabolites (pyochelin, deferoxamine, salicylic acid methylether, indole-3-acetic acid) produced by Pseudomonas sp. strain are unknown (see section 2). EFSA is aware of published studies concerning the production of different pyochelin isomers by different Pseudomonas species (Schlegel, K. et al., 2006; Hayen, H. and Volmer, D.A and Youard, Z.A. et al., 2007). Consequently a data gap has been identified for information on the stereochemistry of the metabolite pyochelin. The strain is not able to grow at 37 C and above. The assessment of the data package revealed no issues that need to be included as critical areas of concern with respect to the identity, physical, chemical and technical properties of the active substance or the representative formulation, however a data gap was identified for analytical data, generated with internationally accepted methods, on microbial contaminants of the active substance. Acceptable methods are available for the determination of the microorganism in the technical material and formulated product and for the determination of the content of contaminating microorganisms. As the Pesticides Peer Review expert s teleconference on mammalian toxicology and environmental fate and behaviour (TC 73) could not conclude on the inhalation toxicity of Pseudomonas sp. strain, the need for a residue analytical method in air is open (see section 2). Pending on the decision on the toxicity of Pseudomonas sp. strain, a method for body fluids and tissues might be needed. EFSA Journal 2012;10(12):2954 7

8 2. Mammalian toxicity Peer review of the pesticide risk assessment of the active substance Pseudomonas sp. strain Pseudomonas sp. strain was discussed at the Pesticides Peer Review TC 73 expert s teleconference. No detailed analysis of the batches used in the toxicological studies is available. However, further information is not required, provided that adequate quality control is undertaken on the batches produced, certifying that toxicologically relevant pathogenic microbial contaminants are kept below levels internationally recognized for microbial contaminants (see data gap in section 1). Potential transfer of genetic material from Pseudomonas sp. strain to other bacteria is considered possible. Information to show that this transfer will not lead to unacceptable effects to human and animal health, including resistance to known therapeutic substances, would need to be provided and a data gap has been identified. In the literature, Pseudomonas fluorescens strains (claimed to be a closely-related microorganism) have been identified in infections of immunocompromised humans. No clinical cases have been described with Pseudomonas sp. strain. In the absence of a reliable test for sensitisation, as for other microorganisms, the following warning phrase is applicable, related to the potential to provoke allergic reactions by inhalation as well as by dermal exposure: Microorganisms may have the potential to provoke sensitising reactions. An acute oral toxicity study was performed in rats with the active substance. The active substance was not acutely toxic via the oral route (LD 50 > 1 x 10 8 CFU/animal). Pathogenicity and infectivity were not investigated in this study. An in vitro permeability study across Caco-2 monolayers showed no indication of pathogenicity and infectivity of Pseudomonas sp. strain. An acute intraperitoneal toxicity/pathogenicity/infectivity study with the active substance was not provided leading to a data gap. An acute inhalation (intratracheal) toxicity/pathogenicity/infectivity study was performed in rats with the active substance at circa 1 x x 10 8 CFU/animal. The study was conducted using a tiered approach. In the first tier, one out of four animals died after exposure. In the first part of the second tier all animals (34) showed severe clinical signs and were euthanized after 2 days. In the second part of the second tier all animals died after 10 to 16 hours. The cause of the death was uncertain and the study design showed some drawbacks. The results of the study are considered inconclusive leading to a data gap. An in vitro permeability study across Calu-3 monolayers showed no cytotoxic effects. Pseudomonas sp. strain is not pathogenic in the in vitro assay with the nematode Caenorhabditis elegans Skin and eye irritation studies were provided for the formulated product (i.e. Proradix WP ). The results indicated that Proradix WP is neither a skin nor eye irritant. No acute oral, dermal and inhalation toxicity studies were provided for Proradix WP. Pseudomonas sp. strain produces metabolites including the siderophore pyochelin (see section 1). The metabolites have been identified in the culture medium and it is unlikely that Proradix WP will contain these metabolites because of the removal of the bacteria from the broth culture. However it cannot be excluded that workers and consumers might be exposed to Pseudomonas sp. strain and its metabolites (see section 4). There is not enough information in the dossier to assess the potential toxicity of metabolites to workers and consumers. The need for further toxicological assessment will have to be considered if metabolites are confirmed to be produced after application leading to a human exposure (see data gap in section 4). There is not enough information EFSA Journal 2012;10(12):2954 8

9 to assess the role of metabolites including the siderophore pyochelin in the potential pathogenesis of Pseudomonas sp. strain, leading to a data gap 7. In conclusion, Pseudomonas sp. strain is not toxic and not expected to be pathogenic by the oral route. Therefore, the derivation of consumer reference values for the microorganism is not needed. Although Pseudomonas sp. strain does not grow in vitro at 37 C and above (see section 1), the available data package does not allow the toxicity, infectivity and pathogenicity of Pseudomonas sp. strain to be characterized by the inhalation and intraperitoneal routes. The derivation of an AOEL for the microorganism is not possible either. The operator and worker exposure risk assessment to Pseudomonas sp. strain cannot be finalized. In addition the worker exposure risk assessment to metabolites cannot finalize. For bystanders, exposure to the active substance for indoor scenarios is likely to be negligible. 3. Residues Pseudomonas sp. strain is not toxic and not expected to be pathogenic to humans by the oral route. However, Pseudomonas sp. strain was shown to produce in the culture medium metabolites of unknown toxicity and no information was provided to confirm whether or not these metabolites may be produced and present in plants after the application of the active substance (see sections 2 & 4). Since no information is available to conclude on the presence on the metabolites after application and on their toxicity, the consumer risk assessment cannot be finalised. 4. Environmental fate and behaviour No information has been provided in relation to potential interference of Pseudomonas sp. strain with the analytical systems for the control of the quality of drinking water provided for in Directive 98/83/EC 8 (see specific Annex VI decision making criteria in Directive 2005/25/EC 9 ). Therefore, a data gap has been identified for evidence to demonstrate that no interferences will be produced with methods for the determination of microbiological contamination of water given in Directive 98/83/EC, in particular with the method for the analysis of Pseudomonas aeruginosa in drinking water (pren ISO 12780). Potential transfer of genetic material from Pseudomonas sp. strain to other bacteria is considered possible. Information to show that this transfer will not lead to unacceptable effects on the environment would need to be provided and a data gap has been identified Fate and behaviour in the environment of the microorganism Persistence and mobility of Pseudomonas sp. strain in soil was investigated in a single soil. The study has deficiencies in the reporting of soil properties and its handling prior to the experiment. Most importantly the soil microbial activity was not reported. In this study it was seen that microorganisms can migrate from the treated potato to the bulk soil, but how relevant this is to agricultural soil cannot be concluded on the basis of this study. The study did not allow an estimation of whether there was decline of the organism in bulk soil. In addition it is not clear if this study has been performed in a laboratory fulfilling the requirements established in the directive (point 2.4 of introduction to Annex II and points 2.2 & 2.3 of introduction to Annex III of Directive 91/414/EEC). Additional information provided from scientific peer reviewed literature investigated only the fate of Pseudomonas sp. strain in the barley plant rhizosphere. Experts in the Pesticides Peer Review TC 73 teleconference agreed that data to address persistence and multiplication in bulk soil of 7 Note after the expert meeting: Data available in the literature indicate that siderophore production is considered an important virulence mechanism in some pathogenic bacteria such as Pseudomonas aeuroginosa. It appears that the siderophore pyochelin has a role on the virulence of some Pseudomonas species (e.g. Sokol and Woods, 1988; Britigan, et al., 1994). However, these data have not been available in the dossier and are not peer reviewed. 8 Council Directive 98/83/EC of 3 November 1998 on the quality of water intended for human consumption. OJ L 330, , p Council Directive 2005/25/EC of 14 March 2005 amending Annex VI to Directive 91/414/EEC as regards plant protection products containing micro-organisms. OJ L 90, , p EFSA Journal 2012;10(12):2954 9

10 Pseudomonas sp strain to fulfil the data requirement as specified in Annex II B, Soil, of Directive 91/414/EEC are not available and a data gap was identified. In relation to the fate and behaviour of Pseudomonas sp. strain in water, the experiments available cannot be considered representative of natural water/sediment systems. Methods used to determine bacterial population in these experiments were not specific to Pseudomonas sp. strain. An UV sterilization experiment showed that when irradiated at wavelengths of 254 nm a reduction in the concentration of bacteria was observed. The study is not relevant to assess environmental exposure (λ > 290 nm), however it may inform on potential methods for decontamination of water exposed to Pseudomonas sp. strain. The risk to aquatic organisms can be characterized as low and the potential for groundwater contamination can be considered low only on the basis of the assumption that any significant exposure can be precluded for the representative uses proposed (potato seed treatment and glasshouse uses). Management measures tailored to local practice and legislation would need to be put in place to control the waste disposal of spent application solution and to prevent accidental spillage entering sewers or surface water drains. Pending the outcome of the toxicological assessment in relation to the infectivity, toxicity and pathogenicity of Pseudomonas sp. strain through inhalation by humans and animals, further data to assess exposure to operators and workers through air may be needed Fate and behaviour in the environment of any relevant metabolite formed by the microorganism under relevant environmental conditions Pseudomonas sp. strain is stated to produce metabolites however the quantity and identity of metabolites is not investigated in the environmental fate and behaviour studies presented in the dossier. Some of these metabolites are expected to be responsible for the pesticide properties of the microorganism (e.g. a very potent siderophore is claimed to be produced by the microorganism in soil). A data gap has been identified for the identification and assessment of the toxicological and ecotoxicological relevance of the Pseudomonas sp. strain metabolites produced in the environment. In particular, information should be provided on the formation in the environment of pyochelin, a known growth promoter of Pseudomonas aeruginosa (Cox, C.D. et al., 1981). Further data on the fate and behaviour of Pseudomonas sp. strain metabolites in the environment will be needed if one or more metabolites of Pseudomonas sp. strain are found to be toxicologically or ecotoxicologically relevant. 5. Ecotoxicology No strain specific data on Pseudomonas sp. strain were submitted to address the toxicity, infectiveness or pathogenicity to non-target organisms; with the exception of two studies on aquatic organisms and an open literature study on earthworms provided in the Addendum of 2012 (Netherlands, 2012). The representative uses included glasshouses uses, and indoor and field application to seed potato. As discussed in section 4, exposure to soil could not be excluded. Exposure to birds and mammals through consumption of contaminated soil organisms such as earthworms is therefore possible. A scientific justification was available in the Addendum of 2012, explaining that since Pseudomonas sp. strain is not expected to grow at 37 C and above (see section 1), which is the body temperature for birds, the risk to birds was therefore considered to be low. Pseudomonas sp. strain is not expected to grow at 37 C and above and moreover it was concluded that Pseudomonas sp. strain is not toxic and not expected to be pathogenic by the oral route in the toxicology section (see section 2). Therefore the risk to mammals through consumption of contaminated soil organisms such as earthworms could be concluded as low. Since exposure of natural aquatic systems consequent to the representative uses could be considered negligible for Pseudomonas sp. strain (see section 4.1), the risk to aquatic organisms was assessed as low. EFSA Journal 2012;10(12):

11 Since the representative glasshouse uses of Pseudomonas sp. strain concern application directly to soil, exposure to honeybees and other pollinators that are frequently used as pollinators in glasshouses is considered negligible. Therefore the risk for honeybees and other pollinators was assessed as low. It was considered that there is potential for soil-dwelling invertebrates, earthworms and soil microorganisms to be exposed (see section 4). An acute toxicity study on earthworms lasting 14 days was available in the Addendum of The study was considered sufficient to address to some extent the evidence of infectiveness or pathogenicity on earthworms. There were no symptoms of toxicity, infectiveness or pathogenicity in earthworms in the study. Therefore the risk of Pseudomonas sp. strain for earthworms was indicated as low. Toxicity studies on soil microorganisms were not available therefore a data gap was identified for the applicant to provide data to address the risk of Pseudomonas sp. strain to soil microorganisms. No data on Pseudomonas sp. strain to address the risk to organisms involved in biological methods for sewage treatment were submitted. The exposure to sewage treatment plants from the representative uses of Pseudomonas sp. strain cannot be concluded. Therefore, a data gap was identified to address the risk to organisms involved in biological methods for sewage treatment. Pseudomonas sp. strain produces metabolites of unknown toxicity. It is noted that a data gap for information on toxins/secondary metabolites produced by this strain was identified (see sections 1, 2 and 4), pending on the outcome of these data gaps, the ecotoxicological risk assessment for toxins/secondary metabolites might need to be considered. EFSA Journal 2012;10(12):

12 6. Overview of the risk assessment of compounds listed in residue definitions triggering assessment of effects data for the environmental compartments 6.1. Soil Compound (name and/or code) Pseudomonas sp. strain Persistence Data gap Ecotoxicology Low risk to earthworms. Data gap for soil microorganisms. Pseudomonas sp. strain metabolites. Data gap Data gap Ground water Compound (name and/or code) Mobility in soil >0.1 μg/l 1m depth for the representative uses (at least one FOCUS scenario or relevant lysimeter) Pesticidal activity Toxicological relevance Ecotoxicological activity Pseudomonas sp. strain Pseudomonas sp. strain metabolites. No data available. No data available. Yes No data available. Low risk. No data available. No data available. No data available. No data available. No data available. EFSA Journal 2012;10(12):

13 6.3. Surface water and sediment Compound (name and/or code) Pseudomonas sp. strain Pseudomonas sp. strain metabolites. Ecotoxicology Low risk. Data gap Air Compound (name and/or code) Pseudomonas sp. strain Toxicology No data available. EFSA Journal 2012;10(12):

14 7. List of studies to be generated, still ongoing or available but not peer reviewed This is a complete list of the data gaps identified during the peer review process, including those areas where a study may have been made available during the peer review process but not considered for procedural reasons (without prejudice to the provisions of Article 7 of Directive 91/414/EEC concerning information on potentially harmful effects). Analytical data, generated with internationally accepted methods, on microbial contaminants of the active substance (relevant for all representative uses evaluated; submission date proposed by the applicant: unknown; see section 1). Data on the stereochemistry of the metabolite pyochelin (relevant for all representative uses evaluated; submission date proposed by the applicant: unknown; see section 1, 2, 4). Study to address the toxicity, infectivity and pathogenicity potential of Pseudomonas sp. strain through the intraperitoneal route is needed (relevant for all representative uses evaluated; submission date proposed by the applicant: unknown; see section 2). A new valid study to clarify the toxicity, infectivity and pathogenicity potential of Pseudomonas sp. strain through the inhalation route is needed (relevant for all representative uses evaluated; submission date proposed by the applicant: unknown; see section 2). Studies to address the toxicity potential of Proradix WP through the inhalation route are needed (relevant for all representative uses evaluated; submission date proposed by the applicant: unknown; see section 2). Information to address the role of metabolites including the siderophore pyochelin in the potential pathogenesis of Pseudomonas sp. strain (relevant for all representative uses evaluated; submission date proposed by the applicant: unknown; see section 2). Data or studies to demonstrate that no interferences will be produced with methods for the determination of microbiological contamination of water given in Directive 98/83/EC, in particular with the method for the analysis of Pseudomonas aeruginosa in drinking water (pren ISO 12780) (relevant for all representative uses evaluated; submission date proposed by the applicant: unknown; see section 4) Information to show that potential transfer of genetic material from Pseudomonas sp. strain will not lead to adverse effects on human and animal health, including resistance to known therapeutics, and will not lead to unacceptable effects on the environment (relevant for all representative uses evaluated; submission date proposed by the applicant: unknown; see section 2 and 4). Data or studies to address persistence and multiplication in bulk soil of Pseudomonas sp strain to fulfil the data requirement as specified in Annex II B, Soil, of Directive 91/414/EEC (relevant for all representative uses evaluated; submission date proposed by the applicant: unknown; see section 4). Metabolites, produced in the environment by Pseudomonas sp. strain, need to be identified and assessed for toxicological and ecotoxicological relevance, in particular the formation and fate of pyochelin should be addressed (relevant for all representative uses evaluated; submission date proposed by the applicant: unknown; see section 1, 2, 4 and 5). The risk of Pseudomonas sp. strain to soil microorganisms (relevant for all representative uses evaluated; submission date proposed by the applicant: unknown; see section 5). EFSA Journal 2012;10(12):

15 Information to address the risk to organisms involved in biological methods for sewage treatment is necessary (relevant for representative uses evaluated; submission date proposed by the applicant: unknown; see section 5). 8. Particular conditions proposed to be taken into account to manage the risk(s) identified No aquatic environmental risk assessment is available assuming exposure arising from representative uses (potato seed treatment and glasshouse uses) will be negligible. Therefore, management measures tailored to local practice and legislation need to be put in place to control the waste disposal of spent application solution and to prevent accidental spillage entering sewers or surface water drains (see section 4). 9. Concerns 9.1. Issues that could not be finalised An issue is listed as an issue that could not be finalised where there is not enough information available to perform an assessment, even at the lowest tier level, for the representative uses in line with the Uniform Principles of Annex VI to Directive 91/414/EEC and where the issue is of such importance that it could, when finalised, become a concern (which would also be listed as a critical area of concern if it is of relevance to all representative uses). 1. The available data package does not allow the toxicity, infectivity and pathogenicity of Pseudomonas sp. strain to be characterised by the inhalation and intraperitoneal route or the setting of an AOEL. Therefore the operator and worker risk assessment cannot be conducted. 2. The worker and consumer risk assessment could not be finalised, pending the finalisation of the environmental risk assessment of metabolites produced by Pseudomonas sp strain and the conclusion of their potential toxicity. 3. Persistence and mobility of Pseudomonas sp strain in soil cannot be assessed with the available data. 4. The environmental risk assessment of metabolites produced by Pseudomonas sp strain DSMZ cannot be finalised with available information. 5. The risk assessment for organisms involved in biological methods for sewage treatment and the risk to soil microorganisms could not be concluded with the available data Critical areas of concern An issue is listed as a critical area of concern where there is enough information available to perform an assessment for the representative uses in line with the Uniform Principles of Annex VI to Directive 91/414/EEC, and where this assessment does not permit to conclude that for at least one of the representative uses it may be expected that a plant protection product containing the active substance will not have any harmful effect on human or animal health or on groundwater or any unacceptable influence on the environment. An issue is also listed as a critical area of concern where the assessment at a higher tier level could not be finalised due to a lack of information, and where the assessment performed at the lower tier level does not permit to conclude that for at least one of the representative uses it may be expected that a plant protection product containing the active substance will not have any harmful effect on human or animal health or on groundwater or any unacceptable influence on the environment. None. EFSA Journal 2012;10(12):

16 9.3. Overview of the concerns identified for each representative use considered (If a particular condition proposed to be taken into account to manage an identified risk, as listed in section 8, has been evaluated as being effective, then risk identified is not indicated in this table.) Representative use Seed potatoes Flowers (summer flowers, chrysanthem ums, roses, gerberas etc) Tomatoes Cucumbers Peppers Eggplant Lettuces Cabbage Operator risk Worker risk Bystander risk Consumer risk Risk to wild non target terrestrial vertebrates Risk to wild non target terrestrial organisms other than vertebrates Risk to aquatic organisms Groundwater exposure active substance Groundwater exposure metabolites Comments/Remarks Risk identified Assessment not finalised Risk identified Assessment not finalised Risk identified Assessment not finalised Risk identified Assessment not finalised Risk identified Assessment not finalised Risk identified Assessment not finalised Risk identified Assessment not finalised Legal parametric value breached Assessment not finalised Legal parametric value breached Parametric value of 10µg/L (a) breached Assessment not finalised X 1 X 1 X 1 X 1 X 1 X 1 X 1 X 1 X 1,2 X 1,2 X 1,2 X 1,2 X 1,2 X 1,2 X 1,2 X 1,2 X 2,4 X 2,4 X 2,4 X 2,4 X 2,4 X 2,4 X 2,4 X 5 X 5 X 5 X 5 X 5 X 5 X 5 X 5 X 1,3,4 X 1,3,4 X 1,3,4 X 1,3,4 X 1,3,4 X 1,3,4 X 1,3,4 X 1,3,4 The superscript numbers in this table relate to the numbered points indicated in sections 9.1 and 9.2. Where there is no superscript number see sections 2 to 6 for further information. (a): Value for non-relevant metabolites prescribed in SANCO/221/2000-rev 10-final, European Commission, 2003 EFSA Journal 2012;10(12):

17 REFERENCES ACD/ChemSketch, Advanced Chemistry Development, Inc., ACD/Labs Release: Product version: (Build 29305, 25 Nov 2008). Britigan B.E., Rasmussen G.T., and Cox C.D., Pseudomonas siderophore pyochelin enhances neutrophil-mediated endothelial cell injury. Am J Physiol Feb; 266 (2 Pt 1): L Cox, C. D.; Rinehart, K. L. Jr.; Moore, M. L. and Cook, J. C., Jr., Pyochelin; Novel structure of an iron-chelatin growth promoter for Pseudomonas aeruginosa. Proc. Natl. Acad. Sci. 78, , EFSA (European Food Safety Authority), Peer Review Report to the conclusion regarding the peer review of the pesticide risk assessment of the active substance Pseudomonas sp. strain DSMZ Hayen, H. and Volmer, D.A., Different iron-chelating properties of pyochelin diastereoisomers revealed by LC/MS. Anal Bioanal Chem 385, Netherlands, Draft Assessment Report (DAR) on the active substance Pseudomonas sp. strain prepared by the rapporteur Member State Netherlands in the framework of Directive 91/414/EEC, October Netherlands, Final Addendum to Draft Assessment Report on Pseudomonas sp. strain DSMZ 13134, compiled by EFSA, September OECD (Organisation for Economic Co-operation and Development), Issue Paper on Microbial Contaminant Limits for Microbial Pest Control Products, Series on Pesticides No. 65, ENV/JM/MONO(2011)43, 12 October Schlegel, K., Lex, J., Taraz, K. and Budzikiewicz, H., The X-ray structure of the pyochelin Fe 3+ complex. Z Naturforsch 61, Sokol P.A. and Woods D.E., Effect of pyochelin on Pseudomonas cepacia respiratory infections. Microb Pathog Sep;5(3): Youard, Z.A., Mislin, G.L.A., Majcherczyk, P.A., Schalk, I.J. and Reimmann, C., Pseudomonas fluorescens CHA0 produces enantio-pyochelin, the optical antipode of the Pseudomonas aeruginosa siderophore pyochelin. J Biol Chem 282, EFSA Journal 2012;10(12):

18 APPENDICES APPENDIX A LIST OF END POINTS FOR THE ACTIVE SUBSTANCE AND THE REPRESENTATIVE FORMULATION Chapter 1 Identity, Biological properties, Details of Uses, Further Information Active micro-organism Function (e.g. control of fungi) Pseudomonas sp. strain (Proradix) Fungicide Identity of the micro-organism (Annex IIM 1) Name of the organism Taxonomy Species, subspecies, strain: Pseudomonas sp. strain (Proradix) Division: Bacteria Family: Genus: The taxonomy of this group of Pseudomonadaceae is rapidly developing and the Pseudomonas sp. are now included in the group of γ-proteobacteria. The bacterium is a new species of fluorescent Pseudomonads. Pseudomonas; part of RNA-group I of γ-proteobacteria. Species: The DSMZ has sufficiently identified the bacterium at the strain level, but concluded that it cannot be assigned to an existing named species and therefore must be considered as a new species. Subspecies: not applicable Strain: (Proradix) Identification First identification by DSMZ, Germany, The identification by DSMZ is based on microscopic examination, 16S rrna analysis and biochemical parameters which characterize different Pseudomonas species and provide useful identification tools. Culture collection (Proradix) Minimum and maximum concentration of the MPCA used for manufacturing of the formulated product (cfu; g/kg): Identity and content of relevant impurities, additives, contaminating organisms in the technical grade of MPCA: Is the MCPA genetically modified; if so provide type of modification 3 to 9 x CFU/g There are no impurities Not genetically modified Biological properties of the micro-organism (Annex IIM 2) Origin and natural occurrence Soil (roots) Pseudomonas sp. strain (Proradix) was isolated from a lettuce field at the University of Hohenheim (Germany). P. sp. Proradix is identified as a new species of fluorescent pseudomonas and is closely related to Pseudomonas fluorescens, a fluorescent pseudomonad EFSA Journal 2012;10(12):

19 Target organism(s) Mode of action Host specificity Life cycle Infectivity, dispersal and colonisation ability Relationships to known plant, animal or human pathogens Genetic stability Production of relevant metabolites/toxins Resistance/sensitivity to antibiotics used in human or veterinary medicine ubiquitously present in soil. Soil-borne fungi The mode of action - restricts the growth of pathogens at the plant root and induction of plant-resistance - based on the production of siderophores, which have affinity for ferric iron. The resulting ferric-siderophore complex is unavailable to other organisms. In this way Pseudomonas sp. proradix restricts the growth of pathogens at the plant root. In addition, fluorescent Pseudomonas sp. proradix induces resistance to soilborne pathogens in the treated plants. This is shown by the deposition of callose on the roots and the activation of most PR-genes in the treated plants. Not directed against a specific pathogen, efficacy against R.solani demonstrated Not applicable P. sp. strain (Proradix) has a maximum growth at ph 7 (range: ph 5-9), an optimum growth temperature of around 28 ºC (not able to grow at 37 ºC). Susceptible to UV light. Pseudomonas sp. Proradix is not related to any human pathogen and only distantly related to some plant pathogens. Stable Pseudomonas sp. Proradix produces a number of metabolites. The metabolites detected in the culture filtrate under the conditions applied are naturally occurring compounds and of no toxicological concern. On basis of the available data there are no indications that Pseudomonas sp. Proradix produces toxic metabolites. Not relevant Classification and proposed labelling with regard to the micro-organism: The active substance should be classified as potentially sensitising by inhalation and skin contact. No classification and labelling for the micro-organism regarding the environment is proposed EFSA Journal 2012;10(12):

20 Summary of intended uses Crop and situation (a) Seed potatoes EU Flowers (summer flowers, chrysanthem ums, roses, Member State or country EU Product name Proradix WP Proradix WP gerberas etc) Tomatoes EU Proradix WP Cucumbers EU Proradix WP Peppers EU Proradix WP F, G, or I (b) I (F) G G G G Pests or groups of pests controlled (c) Soil borne fungi Soil borne fungi Soil borne fungi Soil borne fungi Soil borne fungi Formulation Application Application rate per treatment Type (d-f) Conc. of as CFU/g(i) Method kind (f-h) WP 6.6 *10 10 Application with droplet or water system WP 6.6 *10 10 Application on the soil: pouring around stems of young plants WP 6.6 *10 10 Application on the soil: pouring around stems of young plants WP 6.6 *10 10 Application on the soil: pouring around stems of young plants WP 6.6 *10 10 Application on the soil: pouring around stems of young plants growth stage& season (j) No. min, max (k) interval between applications (min) CFU as /L min/max Prior to 1 n.a. 1 2 *10 12 planting potato tubers in the field 4 * *10 11 During emergence, just before transplanting During emergence, just after transplanting or sowing During emergence, just after transplanting or sowing During emergence, just after transplanting or sowing water L/ha min/max 2-4 (droplet system) (water system) CFU as /ha min/max 1 n.a. 6.6 * * * n.a. 6.6 * * * n.a. 6.6 * * * n.a. 6.6 * * *10 12 PHI (l) Remarks (m) 4 *10 12 n.a. 60 g product/ha (4 *10 12 CFU/ha) n.a. n.a. n.a. n.a. 100 g product /ha (6.6*10 12 CFU /ha) 100 g product /ha (6.6*10 12 CFU /ha) 100 g product /ha (6.6*10 12 CFU/ha) 100 g product /ha (6.6*10 12 CFU /ha) EFSA Journal 2012;10(12):

21 Crop and situation (a) Member State or country Product name Eggplant EU Proradix WP Lettuces EU Proradix WP Cabbage EU Proradix WP F, G, or I (b) G G G Pests or groups of pests controlled (c) Soil borne fungi Soil borne fungi Soil borne fungi Formulation Application Application rate per treatment Type (d-f) Conc. of as CFU/g(i) Method kind (f-h) WP 6.6 *10 10 Application on the soil: pouring around stems of young plants WP 6.6 *10 10 Application on the soil: pouring around stems of young plants WP 6.6 *10 10 Application on the soil: pouring around stems of young plants growth stage& season (j) During emergence, just after transplanting or sowing During emergence, just after transplanting or sowing During emergence, just after transplanting or sowing No. min, max (k) interval between applications (min) CFU as /L min/max water L/ha min/max CFU as /ha min/max 1 n.a. 6.6 * * * n.a. 6.6 * * * n.a. 6.6 * * *10 12 PHI (l) n.a. n.a. n.a. Remarks (m) 100 g product /ha (6.6*10 12 CFU /ha) 100 g product /ha (6.6*10 12 CFU /ha) 100 g product /ha (6.6*10 12 CFU /ha) (a) For crops, the EU and Codex classifications (both) should be used; where relevant, the use situation should be described (e.g. fumigation of a structure) (b) Outdoor or field use (F), glasshouse application (G) or indoor application (I) (c) e.g. biting and suckling insects, soil born insects, foliar fungi, weeds (d) e.g. wettable powder (WP), emulsifiable concentrate (EC), granule (GR) (e) GCPF Codes - GIFAP Technical Monograph No 2, 1989 (f) All abbreviations used must be explained (g) Method, e.g. high volume spraying, low volume spraying, spreading, dusting, drench (h) Kind, e.g. overall, broadcast, aerial spraying, row, individual plant, between the plant - type of equipment used must be indicated (i) Cfu=colony forming units and g/kg or g/l (j) Growth stage at last treatment (BBCH Monograph, Growth Stages of Plants, 1997, Blackwell, ISBN ), including where relevant, information on season at time of application (k) Indicate the minimum and maximum number of application possible under practical conditions of use (l) PHI - minimum pre-harvest interval (m) Remarks may include: Extent of use/economic importance/restrictions EFSA Journal 2012;10(12):