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UDC 574:608.3
APPRAISAL OF ENVIRONMENTAL RISK OF MANUFACTURING OF THE INSECTICIDE GENETICALLY MODIFIED PLANTS (BT-GMР)
Sokolov M.S., Academician of Russian Academy of Agrarian Sciences Department of protection and biogeotechnology of plants of Russian Academy of Agrarian
Sciences, Moscow
Marchenko A.I., Researcher Research centre of toxicology and a hygienic regulation of biological preparations of Federal
Medical-Biological Aagency, Serpukhov
E-mail: sokolov34@mail.ru
ABSTRACT
The implementation of the state food programme can be accelerated essentially, using the complex of up-to-date biogeotechnology and agrotechnics. According to the experience of some countries the genetically modified plants (GMP) - resistant to herbicides, phytophagans, phytopathogenes, abiotic stress factors - the relevant component of this complex. GMP are of certain concern as well as for a forestry, an amenity planting, landscape ecology, the ornamental floriculture and a perspective, renewable radiant of engine fuel. Annually in the XXI-st century the world acreage under GMP, manufactured in concerns of crop protection and a crop, increase by ~10 %. In Russia these plants are not manufactured yet. Principal causes - a deficiency of the information on biosafety GMP and the imperfect gear of state regulation of their manufacturing.
KEY WORDS
Bt-GMR; CrylAb-protein; Environmental assessment.
The subjects of this report (following the results of perennial probes [1]) are: a) methodology of the experimental and analytical appraisal of environmental safety of manufacturing Bt-GMP, b) a justification of a minimum of the basic measure indispensable for an ecological evaluation of manufacturing of these transgenes.
PROBLEM URGENCY, THE PURPOSE OF RESEARCH
It was of interest scientifically to justify methodology of an ecological evaluation of manufacturing Bt-GMP and to tender a minimum of obligatory (basic) measures for its realization. The up-to-date range of GM-plants is over 70 species which have already passed field probations, and it is known over 120 species of GMP.
The seeds of GMP in 2011 in 27 countries of the world have reached 164 million hectares; for the 16-year-old period (1995-2010) the cumulative square of their cultivation has essentially exceeded 1 billion hectares! The basic transgenic crops are a soya bean, a cotton plant, corn and a rape (canola). In 2010 the transgenic soya bean occupied 81 % of all world plantings of this crop, a cotton plant - 64 %, corn - 29 % and a transgenic rape - 23 %.
From total area of GMP 60 % is under soya bean, 23 % is under corn, 11 % is under cotton plant and ~6 % under other crops. According to the prognoses of experts in the nearest future (by 2015) there will be the growth of world planting acreage of GMPto 200 million hectares, the growth (to 30) of the countries performing mercantile cultivationof GMP. Major modification of items of the EU countries concerning manufacturing GMP and the market of appropriate yields is expected. In the world market the emergence of the new species of GMP created in China, India and other countries where the produced direction is the state priority is expected.
In 2007 in the world it has been admitted to manufacturing 168 lines of GMP (43 - corn, 24 - a rape, 22 - a cotton plant, 22 - a potato, 13 soya bean, 9 - rice, 8 -tomato , 7 - wheat, 3 - sugar-beet, 3 - chicory, 2 - melon, 2 - lucerne; 2 - papaya, 2 - tobacco, 2 - pumpkin, 1 - flux, 1 - lentil, 1 - sunflower, 1 - bent grass forthputting ).
From 168 lines of GMP most of all (106 types!) are registered in the USA, 29 - in EU; 17 - in the Russian Federation. GMP registered in Russia are authorised only for consumption; among them 8 species of corn, 4 - potato, 3 - soya bean, 1 - rice, 1 - sugar-beet. For 10 years of cultivation of insecticide GMP it was possible for the USA to spare 36 thousands of insecticides and to profit ~10 billions $ which one significant fraction is necessary on a fraction of farms. Under other data annual manufacturing only Bt-grain-growing allows the USA to spare nsecticides on the square of ~3 million hectares and ensures the benefit of 2,7 billion $. Social-and-ecological aftereffects of decrease ecotoxicant loads on agrovisual environments and a biobubble [1] are not less relevant thus yet spinnable to strict economic evaluation.
Insecticide GMP(or Bt-crops) are the most problem in the ecological ratio. Their fraction in world-wide plantings of GMP (with allowance for stacker types) compounds ~ 40 % (~50 million hectares). The Bt-cotton plant and Bt-corn are predominating. Environmental risk of manufacturing of these crops is essentially aggravated with that they are cross pollened.
Biosafety and usefulness - original conditions of cultivation GM-grade in Russia [2]. The official confirmation of these major characteristics of GMP are:a) the testimony acknowledging genetical correspondence of a transgenic construction stated, b) the experimental acknowledging of nutritional and stern GMP harmlessness,c) demonstrations of environmental GMP safety, d) plugging GM-grade (following the results of a strain testing) in the state catalogue of the selection reachings admitted to utilisation in the Russian Federation [2].
Extending perennial international experience, authors [3] observe concept of biosafety of genetic activity as «... the system of the measures routed on an avoidance or decrease to safety level of adverse effects GMO on health of the person and environment at implementation of genetically-engineered activity». The international frame of biosafety switches on "quartet" of imperatives: 1) "laws" (the legislative baseline regulating genetically-engineered activity), 2) "monitoring" (the system checking an observance of the laws in the sphere of genetically-engineered activity), 3) "experts" (system of professional valuation of hazards and motivated decision makings), 4) "sociability" (the gear of interacting with interested sociability concerning adjustment of genetically-engineered activity).
POTENTIAL FACTORS OF A HARMFULNESS AND ENVIRONMENTAL RISKS OF MANUFACTURING INSECTICIDE GMP
According to the assaying of world publications (tab. 1) main factors of hazard of manufacturing Bt-GMP include: 1) entomocide Cry-fibers - delta-endotoxins Bt, contained in a biomass (and crop yields), emissioned GMP into soil and falling with anther dust in open ponds; 2) a plant succession in an agrocenosis toxine resistant aspects of phytophagans in return to eliminated; 3) a resistance of target phytophagans to insecticidal plants and Bt-MBP.
In the course of ecological evaluation of Bt-GMP it is necessary to observe all postulates of procedure of an experiment work strictly. Besides, the measures of an appraisal should: a) be economically comprehensible, b)not to double, whenever possible, datas of medicobiological and disease and sanitation appraisals,c) be used for an appraisal of all elements, cloths and metabolites Bt-GMP, including exsudates, emissioned into atmosphere and-or into soil, d) be guided on «a transgenesis yield » (product oriented), on general performances corresponding to isogenous analogue,e) to ensure obtaining of played back data, it is time-invariant also sites of negotiations of testing services. The measures of an appraisal should be applied pertaining to each new generically acetylated crop differing from isogenous analogue in one or several transgene insertions (a precautionary principle!).
Table 1 - The Most significant environmental risks manufacturings Bt-GMP [1]
The agro-ecological Ekologically-genetic
1. Transpollination of Bt-GMP with isogenous (parent) quality and hybrids, and also with wild-growing herbs ( a vertical transfer of cry-genes) 7. An induced resistance of target phytophagans to Bt-GMP (and to Bt-specimens)
2. Migration and accumulation of Bt-toksin in agrovisual environment ingredients 8. Pleiotropic effect Bt-GMP
3. Parasite effect of Bt-GMP on a no-purpose biota, decrease of its biodiversity 9*. A plant succession of dominant phytophagans in agrocnosises Bt-GMP
4*. Breaking of operation of triphtorsystem 10*. Inadvertent "panmixia" Bt-GMP with a no-purpose biota -horizontal transfer of cry-genes
5*. Decrease of efficiency Bt-GMP because of decrease of contents Cry-genes in their cloths or of termination of its biosynthesis (because of clamming a cry-gene) 11. Terminator effect of Bt-GMP
6*. Decrease of dimension variety of leading crops because of preferential cultivation of Bt-quality *Note: noted indirect effects of developing process of environmental risk can be determined only in conditions of post-releasemonitoring, i.e. at field (within several years) cultivation Bt-GMP
The experimental ecological evaluation at all stages of testing services of Bt-GMP should be spent with allowance for: a) a principle of single difference,b) a principle actual (on) GMP correspondence to isogenous analogue, c) plugging in the experience scheme of "below zero monitoring” as obligatory alternative at a stage of toxicological testing. Bt-GMP probes at each stage of an ecological evaluation should meet demands not more low, than presented to Bt-microbiological preparations at a stage of their registration testing services. The basic indexes of environmental Bt-GMP safety should be researched step-by-step, is perfectly in order: «Toxico-hygienic testing ^ ecological testing ^ ecological modelling and prediction». The official procedures of ecological Bt-GMP evaluation should be harmonised as much as possible with already operating, international deeds regulating manufacturing and a turn of transgenics (FAO, the World Health Organization, EU, the WTO, EPA, ISO, etc.). It is extremely relevant, that the state acts, technical orders, Governmental decrees (regulating manufacturing and a turn of transgenics), were, whenever possible, as directives of direct acting.
STAGES AND MEASURE OF ECOLOGICAL EVALUATION OF BT-GMP
It was stated, that ecological evaluation of Bt-GMP should include three consecutive stages. On each of the stages particular measures, test-objects and-or sample pieces (tab. 2) are used.
Table 2 - Obligatory stages and the basic measures recommended at appraisal of Bt-GMP
on environmental safety [1]
Evaluation stage Measure, test-objects, appraisal sample pieces
1.Toxico-hygienic testing* chronic toxicity, allergenicity, immunotoxicity, mutagenicity
2. Ecological testing insect-pollinators, aquatic organisms, soil mesofauna, functions of microbiota
3. Ecological modelling and prediction dynamics of a contents and balance of Bt-toxin of system «Bt-GMP ^ soil»; prediction of conditions and periods of developing process of a resistance for target phytophagans to Bt-GMP and its ecological repercussions; prediction of allowed band of spreading of Bt-toxin with anther dust cross pollened transgenics
*Note: Bt-GMP come under to further testing services (stages 2 and 3) only under condition of a below zero appraisal by all four measure of a stage «Toxico-hygienic testing».
According to the generalised data of foreign authors (tab. 3) etnomotoxins Bt-GMP had an ambiguous effect on soil nematodes and micro-organisms, while on all other geobionts (earthworms, satinflowers, polypodies, collembolans and ticks) Bt-GMP did not affect negatively.
THE RESULTS OF APPROBATION OF THE SEPARATE BIOTESTS TENDERED FOR ECOLOGICAL EVALUATION OF BT-GMP
For the verification of the tendered test system there were experimentally gained preparative quantities of insecticide CrylAb-fiber, toxic for Lepidoptera of hexapods. The protein was designated from a culture fluid of strain-producer BKPM B-1226 (Bac. thuringiensis subsp. Kurstaki H3a3b).
Table 3 - Operation Bt-GMP (in situ and ex situ) on the soil population [1]
n/n Geobionts (figure of experiences) Lack of effect Positive effect Negative effect
1. Earthworms (6) 6 0 0
2. Satinflowers (4) 3 0 1
3. Polypodies (1) 1 0 0
4. collembolans, ticks (9) 8 0 1
5. Nematodes ( 12) 7 1 4
6. Micro-organisms, protozoa (22) 13 3 6
Total (54) 38 4 12
In experiences a lyophilized biomass (leaves, caulises) of isogenous quality of corn was used. They spent an appraisal of the chronic toxicity of Cry1Ab-fiber on Sprague Dawley mice (CD-1 line). They researched chronic toxicity of the alternative material (Cry1Ab-fiber + a corn biomass). Within 30 days (inclusive 10 mg/kg the Cry1Ab-squirrel) fed the alternative material to mice of prototype versions ad libitum, i.e. without limitations. Then within 14 days (the reduction period) they fed a test animal a conventional feedstuff. Animals of a reference group (below zero monitoring) throughout only 1,5 month period of experiment gained only a conventional feedstuff. After a 30-day heading of a feeding and 14 daytime reduction period following the results of clinical overseeing by a common status of skilled animals, effects of the hematological assaying of surface-type blood, biochemical analyses of blood serum, pathomorphologic probes of authentic fluctuations for the experimental animals called by operation the Cry1Ab-squirrel, are not determined.
In joint probeswith prof. A.G.Mannapovym (the manager. Stand of beekeeping TAA) they researched operation of the alternative material on honeybee Apis mellifera L. Aftereffects of a feeding of Cry1Ab-fiber (in 50 %-s' sugar syrup) sized up 10 mg/l in an ontogenesis on datas of habitability and productivity of a Carpatho honeybee colonies (tab. 4). It is fixed, that Cry1Ab-squirrels in the density corresponding to its approximate contents in anther dust of Cry1Ab-corn, does not render the negative operations on growth, evolution and productivity of a honey-bee colony on one of 10 major ontogenetic indexes and productivity performances.
Table 4 - Cry1Ab-fiber operation on habitability and productivity Apis mellifera [1]
Index Monitoring Experience
Longevity of a worker-bee, days 18,6±0,51 19,8±0,7
Runout of a printing brood, piece 460±9,6 516±7,9
Uterus egg production, piece 1535±51,5 1972±40,2
Pulp of one worker-bee, mg 92,8±2,6 95,8±1,43
Load of its honey sac (the chief honey harvest), mg 48,7±1,2 50,8±1,50
The same, (a bolstering honey harvest), mg 40,5±1,02 42,0±1,52
Flight activity of a worker-bee (for 3 mines), piece 271±14,5 276±14,1
Runout of documentary honey on one bee colony, kg 24,4±0,51 25,9±0,62
Honeycombs, built up for one bee colony, piece 7,4±0,68 8,0±0,71
Wax, produced for one bee colony, kg 0,89±0,08 0,96±0,08
Agency on aquatic organisms (Daphnia magna) entomocide Cry1Ab-fiber in the density approximately corresponding to its contents in anther dust of Bt-corn (10,3 mkg/G) or essentially exceeding it was sized up. It is fixed, that Cry1Ab-squirrels does not render toxic effect on daphnes. The density of a researched material calling loss of 50 % of test animals,
is not attained. The density which is not calling of visible faulting effects pertaining to of daphnes, compounds 1000 mg/dm3 (NOAEC: 1000 mg/dm3). The minimum operating density of Cry1Ab-fiber exceeds 1000 mg/dm3 (LOAEC> 1000 mg/dm3). Thus, anther dust of corn with insecticide protein Cry1Ab within probed densities (1^1000 mg/l) for daphnes is not toxic.
It is sized up agencies on earthworms (Eisenia fetida) entomocides Cry1Ab-fiber (together with a corn biomass) in a broad band of densities (0,1^1000 soil mg/kg counting on protein). As it is fixed (tab. 5) the Cry1Ab-fiber introduced to a soddy podzolic soil, within 28 days did not render any negative operation on earthworms.
Cry1Ab-fiber operation on emission C02 from a soddy podzolic soil and on intensity of process of nitrification was researched. The density calling decrease of emission C02 on 50 % (in comparison with monitoring) in experience is not attained (EC50> 1000 mg/kg) - tab. 6. The mostof the learnt densities the researched specimen, not calling authentic below zero agency on activity of soil, compounds 1000 mg/kg (NOAEC: 1000 mg/kg).
Table 5 - Cry1Ab-fiber Operation on tolerance Eisenia fetida in a soddy podzolic soil [1]
Contents the Cry1Ab-protein in soil, mg/kg Quantity of animals (pieces) in Tolerance of animals, % to monitoring
7 days 14 days 21 days 28 days
0 (monitoring) 30 30 30 30 100
0,1 30 30 30 30 100
1 30 30 30 30 100
10 30 30 30 30 100
100 30 30 30 30 100
1000 30 30 30 30 100
Table 6 - Cry1Ab-fiber Operation on dynamics of emission C02 from a soddy podzolic soil [1]
Density of Cry1Ab-fiber in soil, mg/kg Dynamics of emission CO2 from soils (mg/kg) in Inhibition (yes/no)
7 days 14 days 21 days 28 days
0 (monitoring) 15,0±0,34 15,0±0,37 15,0±0,36 15,0±0,18 no
0,1 15,1 ±0,31 15,0±0,42 15,0±0,49 14,9±0,25 no
1 15,0±0,14 14,9±0,13 15,1±0,36 14,9±0,27 no
10 15,1 ±0,07 15,0±0,37 15,1 ±0,35 15,0±0,27 no
100 14,9±0,36 15,0±0,20 15,0±0,12 14,9±0,34 no
1000 15,0±0,13 15,0±0,48 15,1±0,29 15,1 ±0,13 no
Table 7 - Cry1Ab-fiber operation on dynamics of a contents of Sodium nitrates in a soddy podzolic soil
[1]
Density of Cry1Ab-fiber in soil, mg/kg Density of a Sodium nitrate-ion in soil (mg/kg) in Inhibition operation (yes/no)
7 days 14 days 21 days 28 days
0 (monitoring) 6,2±0,2 6,3±0,5 6,4±0,4 6,3±0,3 -
0,1 6,6±1,6 6,1±2,6 6,2±1,9 6,4±2,4 no
1 6,3±0,4 6,6±1,3 6,0±1,8 6,4±2,8 no
10 6,9±1,6 6,4±1,7 5,9±2,2 6,2±1,7 no
100 6,4±0,4 6,0±1,5 5,6±0,6 6,7±1,6 no
1000 6,5±2,6 5,3±0,4 6,6±0,6 7,0±1, 1 no
Minimum operating density exceeds 1000 mg/kg (LOAEC> 1000 mg/kg). The density calling inhibition of accumulating of Sodium nitrates in soil on 50 %, is not also attained (EC50> 1000 mg/kg) - tab. 7. The density of a researched specimen which is not influencing negatively on process of nitrification, compounds 1000 mg/kg (NOAEC: 1000 mg/kg). The minimum operating density the Cry1Ab-protein exceeds 1000 mg/kg (LOAEC> 1000 mg/kg). The most of the learnt densities of the specimen, not calling authentic below zero agency on nitrifying activity of soil, compounds 1000 mg/kg (NOAEC: 1000 mg/kg). The minimum operating density exceeds 1000 mg/kg (LOAEC> 1000 mg/kg). Thus, initiating the Cry1Ab-protein in a soddy podzolic soil has not influenced intensity of its breath and nitrification process.
CONCLUSION
World production of Bt-GMP which has attained in 2011 ~50 million hectares is a wide area implementation of biotechnological strategy of a selfdefence of an agrocenosis, the major innovative and high technology unit of the phytosanitary technics, ensuring to the land user essential economic gains and ecological advantages. The perennial experiment of the countries manufacturing GMP, bears that cultivation of Bt-GMP in the ecological ratio is quite safety though it is bundled to their potential environmental risk of straight and-or indirect negative affecting on a no-purpose biota of an agrovisual environment, necessity of correction of conventional agrotechnics and implementation post-release monitoring. At crop protection and crop yields in the ecological ratio ecological repercussions of manufacturing Bt-GMP for an agrocenosis is much less significant, than application of up-to-date chemical baits.
Ecological evaluation of Bt-GMP should be performed sequentially on we rub to units of basic measures («toxic-hygienic testing» ^ «ecological testing» ^ «ecological modelling and prediction»). The obligatory basic measures tendered for a system appraisal of potential environmental risk of manufacturing Bt-GMP at a stage of their preregistration testing services, should be scrapped and perfected periodically with allowance for singularities of new transgenic types, the assaying of their annual effects post - release monitoring and the up-to-date scientific data.
The land users interested in manufacturing GMP, should gain operatively from responsible principals and managers of the Ministry of Agriculture, Federal Service for Veterinarian and Vegetation Sanitary Supervision, other interested ministries and offices answers to following urgent problems: 1) what GMP, in what size and for whom to manufacture them? 2) what agrotechnics in manufacturing GMP should be applied to ensure keeping of indispensable demands of their environmental safety?
REFERENCES
1. Sokolov M.S, Marchenko A.I. Ecological evaluation of transgene Bt-plants - essential condition of their safety manufacturing. M: FSBUS «Rosinforagrotech. 2011. - 129 p.
2. Generically acetylated radiants of nutriment: an appraisal of safety and monitoring / Under the editorship of V.A.Tuteljana. M: Publishing house of the Russian Academy of Medical Science. 2007. 444 p.
3. Blacksmiths V.V., Tsydendambaev V. D. Biosafety of generically acetylated organisms (expertise of food stuffs on biosafety). M. 2008. 252 p.
