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O =X3 M. RUBBIANI Istituto Superiore di Sanit Roma @4 $#$ & M BIOPESTICIDES : EVALUATION PROCEDURES FOLLOWING LIGHT EU DIRECTIVE 91/414 NN  N Most common used  sBacillus thuringiensis Pseudomonas Trichoderma harzianum Beauveria bassiana Virus carpocapsa Cydia pomonella ecc. 6 D    Applicable to biopesticides:    - Directive 91/414/EEC Directive 90/220/EEC .  ) 'New microorganisms on approval (91/414)('$    - Pseudomonas chlororaphis (fungicide) - Coniotirium minitans (fungicide) - Bacillus subtilis ( fungicide,bactericide ) - Gliocladium catenulatum (fungicide) - Paecilomices fumosoroseus (insecticide) - Spodoptera exigua (insecticide) - 6!                In Annex I Dir 91/414/EEC      Ampelomyces quisqualis    '!"$#Identification of the microorganism$#  qDeposited in a culture collection Identified and named at least at species level Characterized at strain level  Q    <Effects on human health (infectivity/pathogenicity/toxicity)(=$6        L Colonisation Causing damages Production of toxins Production of metabolites M  0Human hazard from microorganism should consider:10$       Normal infectivity and pathogenicity Infectivity from high concentrations Toxin production Sensitisation Infectivity to sensitive individualsL      8      Behaviour and Environmental fate!  ! iTesting includes: appropriate choice of the non target organism host specificity mode of action ecology  j P. ` f3"V+Nf` 3f3f3f` /.fRP̙` f"ffH` f3f|` MMM>?" dd@,|?" dd@   " @ ` n?" dd@   @@``@n?" dd@  @@``PR    @ ` ` p>>0  (    H,$D 0   BCPE(F"2@ 8ApP XP@  `@   0e0e     ?BCE<F& 2@ 5% 8c8c     ?1 d0u0@Ty2 NP'p<'p@A)BCD|E?||FP P  - ffFF@    `@F   BCETF.@' EuOE[: :Pf@      `   0e0e     ?BCE`F2 @ 5% 8c8c     ?1 d0u0@Ty2 NP'p<'p@A)BCD|E?||hD[Drmq>uJ^_! 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E<F&@ ! v" bN'\h4ht-2.#; [! ! @    `   0e0e     ?BCETF. @ 5% 8c8c     ?1 d0u0@Ty2 NP'p<'p@A)BCD|E?||$ )  _s0|d T A% :$@      `y    0e0e     ?BdCHEHF* @ 5% 8c8c     ?1 d0u0@Ty2 NP'p<'p @A)BCD|E?||A @<PdT H"k=_ E;W@     `Y   s *G p  o9Fare clic per modificare lo stile del titolo dello schema: :    c $tG  `    t>Fare clic per modificare lo stile del sottotitolo dello schema? ?    c $ԖG ``  _* 2     c $4G `   a*(2    c $G `   a*(2  Z  B޽h))? ? RP̙ 0 @0a( @  ` 0 0 0TP+ P    ]*    0 0P+     _*   d 0 c $ ?  9 0 0Q+  @  uFare clic per modificare gli stili del testo dello schema Secondo livello Terzo livello Quarto livello Quinto livello: v 0 6tQ+ `P   ]*    0 6Q+ `   _*   H 0 0޽h ? ̙33d 4,0(    0& P    a*"      0D'     c*"      6O+ `P   a*"      6O+ `   c*"    H  0޽h ? ̙33T    |(     S $& PP<$D  0     S & P<$ 0  H  0޽h ? ̙33H  `,p( w , , C tT+ P<$D  0  +  , C T+ <$ 0 + H , 0޽h ? /.fRP̙H  P(p( Ow@ ( ( C S+ P<$D  0    ( C T+ <$ 0  H ( 0޽h ? 3f3f3fn P( z  P^ P 6A1?- H P 0޽h ? RP̙d     (  T   s *4YGx ALLEGATO II (quarta lista) Tutte le sostanze attive (incluse loro varianti quali sali,esteri o ammine)soggette a notificazione completa per la quartafase del programma di lavoro di cui all'articolo 8,paragrafo 2,della direttiva Sostanze attive (incluse le loro varianti)presenti sul mercato prima del 25 luglio 1993 che: 1)sono microorganismi (virus inclusi),comprendenti: Aschersonia aleyrodis Agrotis segetum granulosis virus Bacillus sphaericus Bacillus thuringiensis comprendente (*):  sottospecie aizawai  sottospecie israelensis  sottospecie kurstaki  sottospecie tenebrionis Beauveria bassiana Beauveria brongniartii (sinonimo:B.tenella) Cydia pomonella granulosis virus Mamestra brassica nuclear polyhedrosis virus Metarhizium anisopliae Neodiprion sertifer nuclear polyhedrosis virus Phlebiopsis gigantea Streptomyces griseoviridis Virus del mosaico del pomodoro Trichoderma harzianum Trichoderma polysporum Trichoderma viride Verticillium dahliae Kleb. Verticillium lecanii&                  7                        1              A             +     0  s   H  0޽h ? RP̙H  pDp( Ow@ D D C 4U+ P<$D  0  +  D C U+  <$ 0 + H D 0޽h ? RP̙  HD( ( Hl H C W+ P  +  H C tW+ <$ 0 + H H 0޽h ? RP̙R \z( Pd \B \ s *+0 Commission Directive 2001/36/EC of 16 May 2001 amending Council Directive 91/414/EEC concerning the placing of plant protection products on the market "`(  H \ 0޽h ? RP̙ |tX (  X X s *d+  z&MINISTERO DELLA SALUTE DECRETO 1 febbraio 2002 Attuazione della direttiva 2001/36/CE della Commissione del 16 maggio 2001, che modifica la direttiva 91/414/CEE del Consiglio relativa all'immissione in commercio di prodotti fitosanitari. Gazzetta Ufficiale N. 220 del 19 Settembre 20024`8`8                   &  #         H X 0޽h ? RP̙ l(  lr l 6A ?? H l 0޽h ? RP̙ p( ;Z  pr p 6A ?? H p 0޽h ? RP̙ t( z tr t 6A  ??u  H t 0޽h ? RP̙ x(  xr x 6A ?? H x 0޽h ? RP̙ XP (    s *+_ V4. ANALYTICAL METHODS Descriptions of methods must be provided as well as data on specifity, linearity, accuracy and repeatability for micro-organisms and impurities, metabolites, residues . 4.1. Methods for the analysis of the micro-organism as manufactured - Methods for the identification of the micro-organism. - Methods for providing information on possible variability - Methods to differentiate a mutant of the micro-organism from the parent wild. - Methods for the establishment of purity - Methods to determine the content of the micro-organism in the manufactured material - Methods for the determination of relevant impurities in the manufactured material. - Methods to control the absence and to quantify the possible presence of any human and mammalian pathogens. - Methods to determine storage stability, shelf-life of the micro-organism, if appropriate. 4.2. Methods to determine and quantify residues (viable or non-viable) of: - the active micro-organism(s), - relevant metabolites (especially toxins), on and/or in crop, in foodstuffs and feeding stuffs, in animal and human body tissues and fluids, in soil, in water (including drinking water, ground water and surface water) and in air where relevant. 46Q     Q H  0޽h ? RP̙  02(    s *$+  5. EFFECTS ON HUMAN HEALTH (i) Available information based on the properties of the micro-organism and corresponding organisms (ii) The information provided, taken together with that provided for one or more preparations must be sufficient to permit an .The information provided must be sufficient to: - permit a decision to be made as to whether, or not, the micro-organism can be included in Annex I, - specify appropriate conditions or restrictions to be associated with any inclusion in Annex I, - specify risk and safety phrases (once introduced) for the protection of man, animals and the environment to be included on packaging (containers), - identify relevant first aid measures as well as appropriate diagnostic and therapeutic measures to be followed in the event of infection or another adverse effect in man. (iii) All effects found during investigations should be reported. Investigations which may be necessary in order to evaluate the probable mechanism involved, and to assess the significance of these effects, must also be performed. (iv) For all studies actual achieved dose in colony forming units per kg body weight (cfu/kg), as well as in other appropriate units, must be reported. (v) Evaluation of the micro-organism should be carried out in a tier-wise manner.Rc w   H  0޽h ? RP̙ 91@(    s *+ 7TIER I 5.1. Basic information 5.1.1. Medical data 5.1.2. Medical surveillance on manufacturing plant personnel 5.1.3. Sensitisation/allergenicity observations, if appropriate 5.1.4. Direct observation, e.g. clinical cases 5.2. Basic studies 5.2.1. Sensitisation(6) 5.2.2. Acute toxicity, pathogenicity and infectiveness 5.2.2.1. Acute oral toxicity, pathogenicity and infectiveness 5.2.2.2. Acute inhalation toxicity, pathogenicity and infectiveness 5.2.2.3. Intraperitoneal/subcutaneous single dose 5.2.3. Genotoxicity testing for 5.2.3.1. In vitro studies 5.2.4. Cell culture study 5.2.5. Information on short-term toxicity and pathogenicity 5.2.5.1. Health effects after repeated inhalatory exposure 5.2.6. Proposed treatment: first aid measures, medical treatment TIER II 5.3. Specific toxicity, pathogenicity and infectiveness studies 5.4. In vivo studies in somatic cells 5.5. Genotoxicity - In vivo studies in germ cells >           %       !   q  (   n    5   ! H  0޽h ? RP̙ ;( ;Z    s *A I5.2.1. Sensitisation The test will provide sufficient information to assess the potential of the micro-organism to provoke sensitisation reactions by inhalation as well as with dermal exposure. A maximised test has to be performed. Available information on the sensitisation and allergenic response of workers, including workers in manufacturing plants, agricultural and research workers and others exposed to the micro-organism must be provided, and include, where relevant, details of any incidences of hypersensitivity and chronic sensitisation. The information provided should include details of frequency, level and duration of exposure, symptoms observed and other relevant clinical observation. Information should be given about whether workers have been subjected to any allergy tests or interviewed about allergenic symptoms.  J`4D  H  0޽h ? RP̙ c[P( ;Z    s *A a6.RESIDUES IN OR ON TREATED PRODUCTS, FOOD AND FEED (i)The information provided, taken together with that for one or more preparations containing the micro-organism, must be sufficient to permit an evaluation to be made as to the risk for man and/or animals, arising from exposure to the micro-organism and its residual traces and metabolites (toxins) remaining in or on plants or plant products. (ii)In addition, the information provided must be sufficient to: - permit a decision to be made as to whether or not the micro-organism can be included in Annex I to Directive 91/414/EEC, - specify appropriate conditions or restrictions to be associated with any inclusion in Annex I to Directive 91/414/EEC, - where relevant, set maximum residue levels, pre-harvest intervals to protect consumers and waiting periods, to protect workers handling the treated crops and products. (iii) For the evaluation of risk arising from residues, experimental data on levels of exposure to the residue may not be required where it can be justified, that the micro-organism and its metabolites are not hazardous to humans in the concentrations that could occur as a result of authorised use. 6.1. Persistence and likelihood of multiplication in or on crops, feedingstuffs or foodstuffs 6.2. Further information required 6.2.1. Non-viable residues 6.2.2. Viable residues 6.3. Summary and evaluation of residue behaviour resulting from data submitted under points 6.1 and 6.2 Fc3dB      H  0޽h ? RP̙ y`( x   s *A 7. FATE AND BEHAVIOUR IN THE ENVIRONMENT (i) Information on the origin, the properties, and the survival of the microorganism and its residual metabolites (ii) Information to permit an assessment of its fate and behaviour, its residual traces and toxins. (iii) In particular, the information provided should be sufficient to: - decide whether, or not, the micro-organism can be included in Annex I, - specify appropriate conditions or restrictions to be associated with any inclusion in Annex I, - specify the hazard symbols (once introduced), the indications of danger, and relevant risk and safety phrases for the protection of the environment, which are to be included on packaging (containers), - predict the distribution, fate, and behaviour in the environment of the micro-organism and its metabolites as well as the time courses involved, - identify measures necessary to minimise contamination of the environment and impact on non-target species. (iv) Any relevant metabolites (i.e. of concern for human health and/or the environment (v) Available information on the relationship with naturally occurring wild type relatives . (vi) Before performing studies as referred to below, the applicant shall seek agreement of the competent authorities. 7.1. Persistence and multiplication - competitiveness under the environmental conditions prevailing at and after the intended use, and - population dynamics in seasonally or regionally extreme climates 7.1.1. Soil 7.1.2. Water 7.1.3. Air Jcc6q     ? H  0޽h ? RP̙  p<(    s *dA | 7. EFFECTS ON NON-TARGET ORGANISMS (i) Identity, biological properties and informations to assess the impact on non-target species (ii)The choice of the appropriate non-target organisms based on the identity of the micro-organism (iii)The information must be sufficient to permit an assessment of the impact on non-target species to: - decide whether, or not, the micro-organism can be included in Annex I, - specify appropriate conditions or restrictions to be associated with any inclusion in Annex I, - permit an evaluation of short- and long-term risks for non-target species, - classify the micro-organism as to biological hazard, - specify the precautions necessary for the protection of non-target species, - specify the hazard symbols (i) Report all potentially adverse effects found during routine investigations (ii) Average dose in cfu/kg bw as well as in other appropriate units must be reported. (iii)It may be necessary to conduct separate studies for relevant metabolites (iv)The same strain of each relevant species should be used in the various tests specified. (v)Tests must be performed unless non-target organism will not be exposed. 8.1 Effects on birds 8.2. Effects on aquatic organisms 8.2.1. Effects on fish 8.2.2. Effects on freshwater invertebrates 8.2.3 Effects on algae growth 8.2.4. Effects on plants other than algae 8.3. Effects on bees 8.4. Effects on arthropods other than bees 8.5. Effects on earthworms 8.6. Effects on non-target soil micro-organisms 8.7. Additional studies n#mcmbK     )     H  0޽h ? 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