CALL: FP7-KBBE-2010-4 Proposal full title: Novel strategies for integrating land snail pests control of agricultural

crops in Europe, with projection to Latin-America

Proposal acronym: Land snail’s life cycle as pest control core

Type of funding scheme: Collaborative Project. CALL: FP7-KBBE-2010-4

Work programme topics addressed:

Food, Agriculture and Fisheries, and Biotechnology Area: 2.1.2 Topic number: KBBE.2010.12-05

Name of the coordinating person: Dr José Castillejo Murillo Departamento de Zoología y Antropología Física. Facultad de Biología. Universidad de Santiago de Compostela. E-15782 Santiago de Compostela. La Coruña. Galicia. España. Mobile Phone: + 34 654 969 784 Tel: + 34 981563100 Fax: + 34 981 596904 E-mail: jose.castillejo@usc.es

List of participants Participant number Participant organitation name Country 1 (Coordinator) Universidad de Santiago de Compostela Spain 2 3 4 5 6 7 8 9 10 11 12 13 14 15

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TITLE: Novel strategies for integrate land snail pests control at agricultural crops in Europe with

projection to Latin-America. 1: Scientific and/or technical quality, relevant to the topics addressed by the call 1.1 Concept and objectives To introduce in a series of crops, new strategies for the integrated land snail pest control. These strategies

are based in the deep knowledge of the pest´s biology and ecology, so that the abundance and the activity periods can be predicted to elaborate an integrated pest control system and take decisions that can be applied in every develop phase (juvenile, adult, senile or eggs). Thanks to this the farmer will know when to apply the traditional molluscicides (to destroy the land snails), when to use the molluscicides ovicidal (to destroy the egg lays), when to apply the biological control through parasite nematodes or when to use the trap-plants. To develop this integrated control methods it will be necessary:

1. To know the biotic and a biotic factor that describe the land snails biological cycle in different areas of Europe and Latin-America, to elaborate the control method.

2. To know the specific land snails´ diet with the objective of finding the most attractive plant species to use them as trap-plants.

3. To understand the activity in function of the environment biotic and a biotic variables, with the aiming of developing an effective abundance and activity statistical prediction method.

4. To search plants with bio pesticide activity to be used as biomolluscicides and bio-ovicides against land snail and its eggs.

5. To know the ovicidal potential of the usual non residual agrochemicals to use them as molluscicides-ovicides in crops.

6. To know the ovicidal potential of cattle’s and swine’s slurries as molluscicides-ovicides in ecological farming.

7. To search in each study areas of Latin-America a parasite nematode (Phasmarhabdities alike) to use it as a biological control method against land snails.

8. To deliver to the horticultural industry an effective integrated crop management strategies, low chemical and biological protecting methods against land snails.

9. To deliver to the ecological farming effective integrated packages methods based on cattle and swine manure and plant traps, to protect the crops against land snail pests.

With these strategies we are settling the basis for an integrated control method, to achieve a more

rentable crop due to; having less plant damages, less pesticides use, and a more respectful farming with the environment and the wild fauna. The land snail pest problem in Europe and Latin-America is increasing every day due the commercial globalization. The most dangerous land snail species in Europe are: Arion lusitancicus, Deroceras retiuclatum, Lehamnnia marginata, Milax gagates, Criptophalus aspersus and Theba pisana among others. The 90% of the land snail species that are pest in Latin-America are introduced species from Europe and other countries, this species proliferate indiscriminately because of the absence of natural predators, such as Acanthina fúlica, Deroceras reticulatum, Milax gagates, Lhemannia marginata, Arion intermedius, Criptophalus aspersus…. In some Caribbean regions native land snail species cause important damages, such is the case of Veronicella genus in Mexico and Cuba.

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S&T objectives detailed description This project requires the application of novel control strategies to control native or introduced land snail

pests in crops that can be used in any agricultural pest. With the strategies proposed here we can anticipate to the damages caused by the land snail pests, due the application of a preventive method even before damages appear on the crops.

Our strategies are based on: 1. To understand the land snails biological cycle in the crops study areas. 2. To understand the land snails activity in function of the climatic variables and the

crop type. 3. To destroy the land snail´s egg-lays thanks to the plant extracts and non residual

standard agrochemicals collateral effect. 4. To rationalize standard molluscicides consumption in standard farming. 5. To introduce cattle and swine manure and trap-plants as control methods in

organic farming. 6. To understand the collateral effects of the products used in this integrated pest

control methods.

The understanding of the biological cycle is very important, because we attempt to use the molluscicides before damages appear. With the knowledge of the biological cycle we will know which time of the year is juvenile, adult or senile, we will know when the egg-lays are done, and in other words, we will know the sizes, structure and dynamics of their populations. The information provided by the biological cycle is important to implement this new strategy, as these molluscicides must be applied when the population density is lower and when there are fewer egg-lays in the soil. With this we obtain an optimal effectiveness destroying the egg-lays through ovicidal and also killing gastropods. By applying less molluscicides we save money and minimize the side effects on the environment.

Knowing the diet of land snails in the study areas will give us information on the possible use of trap-plants, to evaluate and estimate the damage that they actually produce on crops. By studying the stomach contents of a specified number of land snails we can know their preferences, in previous research we found that plants that had a low abundance in the environment, appeared with high frequency in the stomach of land snails, this means that they have positive selection for this type of plants. These plants can be used as trap-plants to protect vegetable crops deterring land snails to eat the trap-plants. It is a very useful strategy in organic farming.

Knowing the activity of terrestrial gastropods in terms of climatic variables and crop phenology is required to develop a statistical for activity prediction. With this model we can predict with 24-48 beforehand the activity, and provide the farmer information that land snails will be active, and thus may apply the traditional molluscicides at the right time, getting a greater effectiveness using smaller amount of molluscicides, leading to saving resources and reducing side effects on plants, soil and wildlife.

So far we have been talking about traditional molluscicides. The tradiotional molluscicides (metaldehyde, carbamate, iron sulphate, phasmarhadities...) are intended to kill the individuals, in other words, kill the terrestrial gastropods leaving intact the land snail´s egg-lays in the soil.

There is a growing body of evidence to suggest that in the past 4-5 decades there has been an excessive

dumping of chemical toxins on the soil. As a result the soil in many places has become barren and the ground water toxic. Contrast with this the organic inputs that are safe, non toxic, and cost much less. 'Biopesticides' are certain types of pesticides derived from natural materials such as animals, plants, bacteria, and certain minerals. Benefits of biopesticides include; effective control of insects pests, plant diseases and weeds, as well as human and environmental safety. Biopesticides also play an important role in providing pest management tools in areas where pesticide resistance, niche markets and environmental concerns, limit the use of chemical pesticide products. The idea of finding plant extracts with molluscicidal and ovicidal activity against land snails and their eggs is original and promising.

In each country, agricultural authorities allow a number of non residual agrochemicals for different uses

and for different purposes, can be fertilizers, herbicides, acaricides, fungicides, etc... These products have gone through a series of tests to be authorized. In previous research we have tested non residual agrochemicals to see

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if they had the potential to destroy the land snail´s egg-lays, and many of them had as side effect the egg-lays killing. Basing on these assumptions each participant must make a test series with non residual agrochemicals approved in their country, to discover which one or ones have a higher ovicidal power at the lowest concentration in the shortest time. In organic farming the use of synthetic chemicals such as fertilizers, pesticides, antibiotics, etc.., it´s forbidden, with the objective of preserving the environment, maintaining or enhancing soil fertility and provide food with all its natural properties. Fertilizers that can be used in these kinds of crops can be of two types, green fertilizers or livestock manure. In previous research we found that certain concentration of swine and cattle manure had ovicidal action on terrestrial gastropod egg-lays. Therefore each participant will have to do tests to discover the type and concentration of manure that has higher ovicidal power against the land snail´s egg-lays in their study areas.

Finding a new parasite-nematode with a biological cycle like Phasmorhadities hermaphrodita is crucial to have a new tool for biological land snail pests control in agriculture in Latin-America, as the European variety has problems at certain soil temperatures. The task to find European zoo parasitic nematode was made in previous 97- UE - Project.

As a consequence the of traditional and organic farmers are going to get a series of useful tools for land

snail pests and, first they are going to have a new control strategy based on the application of the ovicidal-molluscicides at the correct time, determined by the life cycle of the terrestrial gastropods, and not by the crop phenology. It will be explained which chemicals they need to destroy land snail´s egg-lays. Through the predictive model, the farmers will have the information necessary to know which day and a time terrestrial gastropods will be active, thus apply the traditional molluscicides at the right, time, place and amount. This information must be transmitted through scientific meetings, counselling State Agricultural Agencies and through web pages of the State Servers.

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COMPLIANCE WITH THE OBJETIVES OF THE WORK PROGRAMME AND ITS PRIORITIES This project can definitely solve the problem of land snail pests in agriculture that has been globalized by

the reform of the European Common Agricultural Policy and which it is forced to fulfil in the countries from which we import agricultural products.

This project is related to THEME 2 (Food, Agriculture and Fisheries, and Biotechnology), Area 2.1.2 KBBE.2010.1.2-05 "Integrated pest management in farming systems of major importance for Europe" out of the 7th Framework Program of Cooperation (FP7 Cooperation Work Program), and our project fits in perfectly in the topics (issues) of integrated pest control (management) (In the context of Integrated Pest Management - IPM-) in particular (specifically) can be said that:

1. It includes preventive measures such as, molluscicides application in the more labile phases of the life cycle of terrestrial gastropods and when there is less density of population and eggs in the soil, which generally coincides with (periods in which) (times when) there is nothing planted on farms .

2. The design of this strategy is based on the study of the biological cycle of pests and the study of their activity in terms of biotic and abiotic variables of the environment, representing perfect control and more accurate information to take preventive measures.

3. With this strategy control measures are applied at the right time, anticipating the emergence of the pest, which implies less molluscicide applied to achieve a better effect, besides the molluscicide is never next to plants, if the measures control are applied prior to planting. Our strategy is aimed at controlling or eradicating the pest to protect the crop, in other words, we use a preventive strategy.

4. It is an integrated control based on the decision-making through the statistical model prediction. We only use low toxicity chemicals, we use the biological control of nematodes through zooparasites, not to mention the use of trap plants to deter terrestrial gastropods that attack crops or the use of swine and cattle manure as ovicidal all this applied at the time we enter the life cycle of the pest snail and the predictive model.

5. This strategy helped to reduce pesticide use on crops applying the necessary quantity at the right time, not introducing new chemicals in agricultural crops, but taking advantage of the farmers’ standard used non residual agrochemicals just giving it a different use or using the favourable side-effects. Thereby decreasing the amount of toxic agents that may be harmful to humans and to wildlife and soil.

6. This project combines "combine modelling and experimentation" because the entire strategy is based on the study of biological and ecological cycle of the pest, its dynamics, in order to achieve the greatest success of control with the least effort and with the least means.

7. The risks of not succeeding in this project are limited, first all the participants who are part of the research group have demonstrated ability to perform all the tasks outlined in the Work Packages, and also the USC team that coordinates this project has an extensive experience in developing predictive models of activity of the snails and slugs in agricultural crops of Galicia (Spain) and in controlling pests in vegetable crops and vineyards and many others European participants have similar experience in similar fields. Given the globalization of trade, over 90% of land snail species pests in Latin-America are of European origin, in other words, they are introduced species that we have been working with over 20 years.

8. The balance between costs and benefits will always be positive because we will control the final shape of land snail pests in agriculture, and we will bring to the traditional and organic farmers to have a number of tools to obtain a more profitable crop and seeding time management, very respectful with the environment.

9. Finally, all information obtained from this project and the control strategies are available for the farmer, either through meetings, workshops taught by the competent authorities or available through “on line” services were the farmers will resolve questions and provide information of the pest activity.

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1.2 Progress beyond the state-of-the-art

At global scale, the economic damage caused by terrestrial gastropods has increased, thanks to the globalization of the trade market which leads us to talk of land snail species introduced from one continent to another, species that having no specific predators grow their population exponentially. While some land snails can reach pest status even in relatively arid regions, slugs are particularly problematic in temperate and rainy weathers, but even in this case, the extent of damage done to crops varies greatly at Regional and year to year scale (Port and Port, 1986). Many experts agree that the damage caused by gastropods has increased very significantly in the last 2 or 3 decades due to a combination of factors such as simplified cultivation techniques (reduced tillage, seeding direct), the reduction of predatory insects population caused by the insecticide products or by the use of new crop varieties more susceptible to be attacked by gastropods (Hommay, 1995, 2002, Godan, 1999; Speiser, 2002). Furthermore, the rising quality standards, makes the consumers tolerance to damaged products shrink, resulting in an intensification of the pest control measures.

Terrestrial gastropods Pest control is performed, almost exclusively, through the application of bait (pellets) containing between 2% and 8% of metaldehyde or carbamates (Godan, 1983, 1999, South 1992, Garthwaite and Thomas, 1996; Bailey, 2002; Speiser, 2002). The world's largest producer of metaldehyde is the Swiss company Lonza. The most widely used carbamate in land snail pest control is the metilocarbamate, which manufacture licensed is owned by the German company Bayer. Both compounds show similar efficacy in terms of reducing damage to plants (Bailey, 2002), and both have collateral effects on other animal groups populations (South, 1992; Bailey, 2002). Buchs, Heimbach and Czarnecki (1989) have noted the existence of the molluscicides metaldehyde baits have negative effects on some carabid populations; also Bieri, Schweizer, Christensen and Daniel (1989) have documented a reduction in the carabids and staphylinids populations, after the application of molluscicides metiocarbamate baits in meadows. Although currently all molluscicides baits incorporate pigments (usually blue) and other substances to reduce the mammals and birds ingestion risk, the cases of domestic animals poisoning because of molluscicides bait are frequent (Bailey, 2002). In the late 80s, molluscicides carbamate baits were banned in many U.S. states, due to the high frequency of poisoned bird cases were recorded (Sakovich, 1996). Tarrant and Westlake (1988) suggest that the use of molluscicides baits with metiocarbamato is a serious threat to field mice (Apodemus sylvaticus) populations (Linnaeus, 1758). Gibson and Reynolds (1991) reported high acetaldehyde concentrations (resulting from the metaldehyde depolymerization in the gastrointestinal tract) in hedgehogs (Erinaceus europaeus) (Linnaeus, 1758) found dead on the field, and Gemmeke (1997) observed symptoms of poisoning cases and death in hedgehogs fed with slugs that had eaten metiocarbamate baits.

In recent years has appeared on the market a new chemical molluscicide, under the Ferramola trade name manufactured by the German company Neudorff GMBH. This product is also presented in the bait format and contains iron phosphate as active ingredient. The tests conducted to corroborate its effectiveness (Iglesias and Speiser 2001, Speiser and Kistler, 2002) indicate that the iron phosphate it´s comparable to the classic chemical molluscicides such as metaldehyde and metiocarbamate. Unlike these which are totally synthetic, iron phosphate occurs naturally, as part of various minerals, especially strengite (Fe III PO 4 2 (H 2 O) orthorhombic) and metastrengita (Fe III PO 4 2 (H 2 O) monocyclic) (Roberts, Campbell and Rapp, 1990, Clark 1993) and it is a compound with a very low toxicity (EPA, 1998).

The lack of gastropods pest control methods that are authorised in organic farming makes these animals to be considered as the most damaging ones by many professional associations in Britain and Switzerland in this kind of plantations (Peackock and Norton, 1990; Kesper and Imhof, 1998). The only currently marketed biological pest control agent for land snail, is the nematode Phasmarhabditis hermaphrodita (Schneider, 1859), released first in Britain in 1994. Numerous field tests in a wide variety of crops in European countries have shown that P. hermaphrodita can reduce the damage done by land snails in plantations (Wilson, Glen and George, 1993; Wilson, Glen, George and Hughes, 1995, Wilson and Glen Hughes, 1995; Ester & Geelen, 1996; Iglesias, Castillejo & Castro, 2001ab). Its effectiveness against the species D. reticulatum is beyond any doubt (Glen Wilson, Brain and Stroud, 2000), but there are signs that its effectiveness against other species might be low (Wilson et al. 1995a; Coupland, 1995, Glen et al., 1996, Speiser and Andermatt, 1996; Speiser, Zaller and Neudecker, 2001, Iglesias and Speiser, 2001). The treatments with P. hermaphrodita effectiveness is very conditional to the soil moisture and

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temperature, which greatly affects their survival, but has the advantage that the favourable moisture and temperature conditions for the land snails activity, are also the best suited for the nematode development, whereas chemical molluscicides effectiveness (as methaldeide baits) is greatly reduced due to this conditions, in which gastropods cause the most damages to plants (Glen et al., 1996). However, the high economic costs of the nematode pest control treatments makes their use restricted to high value crops such as ornamentals plants and some vegetables (Grunder, 2000).

The application of molluscicides is only a short terms control measure, it is a temporarily protection from the damage that might cause gastropods on plants. Also it doesn´t have a significant and lasting effect on gastropods populations residents in the plantation areas, so the risk of damage is permanent (Hommay, 2002, Port and Ester 2002). This is because applied molluscicides affect only the active portion of the population, as the gastropod egg-lays, which are found in soil, are not affected by conventional molluscicides treatments, resulting in a rapid population recovery (Glen, Wiltshire and Milson, 1988). It has been estimated that treatments based in molluscicides metaldehyde or carbamate baits kill less than 50% of the population of gastropods existing at the time of implementation (Glen & Wiltshire, 1986; Wiltshire and Glen 1989, Glen, Wiltshire and Butler, 1991). On the other hand, it is common that the amount of molluscicide bait ingested by gastropods has only a transient sublethal effects (Kemp & Newell, 1985; Wedgwood and Bailey 1986, Briggs and Henderson, 1987; Bourne, Jones & Bowen 1988), and it has been shown that the fertility of individuals who experience such sublethal poisoning is not affected, so they continue laying eggs once they are recovered (Kemp & Newell, 1985).

In the 60 surges the Integrated Pest Management concept (IPM) (Stern, Smith, van der Bosch and Hagen, 1959), which currently is part of another broader concept, which is the sustainable development. The IPM involves the integration of many fields knowledge (biology, chemistry, agronomy, climatology, economics, etc). In order to develop the most appropriate control strategies, from the economic, environmental and Public Health viewpoint (Dent, 1991). While it is a system based on the combination of different methods in order to minimize the use of chemical pesticides is not excluded, a priori, the use of any control agent (Coombs and Hall, 1998).

Methodologically, the IPM can be described as a "decision making process", based on the analysis of all the available relevant data, deciding what action to take and when, to best control the plagues. Besides, the IPM searches the most effective, economic and less aggressive approach, from the economic and the the environmental viewpoint (Bechinski, Mahler and Homan, 2002).

Currently, a lot of IPMs for many arthropods and fungi pest species are based on the use of prediction methods (forecasting systems) (Dent, 1991; Frahm, John and Volk, 1996). Predicting when a pest can cause significant damages to crops is essential, to know the exact moment to apply molluscicides (Buhler, 1996). Furthermore, depending on the pesticide action mode, its effectiveness can be influenced by the organism life cycle or its activity level (Bailey, 2002). In short, criteria it´s needed to determine the need and desirability of applying pesticides.

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The advances that the proposed project and patents will bring.

To this day, non residual agrochemicals with ovicidal action had not been used to control land snail pests in agriculture. The strategy proposed in this project is innovative, original and this pest control methods are very efficient, and makes it very easy to eradicate pest. It is original because it attempts to control the causative agent pest in the phase of their life cycle [biological cycle] where they are most fragile, in the egg stage. It is an original method that will develop statistical models to predict the abundance and activity of the land snail with which the farmer can act in a preventive manner so as not to damage because it will indicate the moment when which has to apply the ovicidal or the traditional molluscicides. It is original because it doesn´t introduce new pesticides in agriculture, it is based in non residual agrochemicals that the farmer habitually uses, it seeks the best profile to exploit its ovicidal activity. It is new because it puts into the hands of the organic farming a number of tools to control land snails pests using natural fertilizers or deterrent strategies implemented by trap plants deterring land snails from attacking crops.

It is a project that uses current technologies to see how the pests live, which are their weaknesses, and attack them manipulating it that way to minimize side effects on crops, and even on the man. With this project we are going to obtain results that will be patented. As a result we will be able to patent the pest control strategies, establish a patent for the active use of non residual agrochemicals with ovicidal action against the land snail´s egg-lays, and finally be able to patent the Statistical Prediction Activity Model and abundance of land snail that are pests, and most likely be able to patent the use of a zooparasite nematodes for the land snail pests biological control.

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1.3 S/T methodology and associated work plan 1.3.1 Overall strategy of the work plan (1 page). Detailed description of the proposed work The whole strategy is designed to perform integrated land snail pests control in conventional and organic

farming, and it is here to introduce the use of new molluscicides ovicidal action, capable of destroying the egg-lays, the use of trap-plants as a deterrent, and the rational use of traditional molluscicides.

WP. 1 .-To understand the biology of land snail that are pests in a range of vegetable crops, it is necessary to have information of the size, structure and dynamics of their populations.

WP. 2 .-To study food and qualitative and quantitative composition of the diet of terrestrial gastropods that are pests in order to find plants that can be used as trap plants in organic farming and traditional.

WP. 3 .-To develop a statistical model that could explain and predict the abundance and activity of the land snail as a function of environmental variables such biotic and abiotic.

WP. 4 .-To investigate the feasibility of using plant extracts as biomolluscicides and bio-ovicides. To carry

this out it will be necessary: 4.1. Laboratory tests on filter paper (direct contact) and artificial soil (standard soil) to select the plant

extracts with molluscicidal and /or ovicidal activity. 4.2. Mini plots experiments on horticultural soil to evaluate the efficacy of the selected plant stract against lands snails and its eggs.

4.3. Mini plots analysis to know the collateral effect of plant extract selects on invertebrate soil. 4.4. Chemical analysis to find the plant extracts active principle by analytic steps. WP. 5 .-To investigate the feasibility of using commercial agrochemical activity as ovicidals with control land

snail egg-lays for key Conventionally grown horticultural crops. To carry this out it will also be necessary: 5.1. Laboratory tests on filter paper (direct contact) and artificial soil (standard soil) to select the

agrochemical which best suits the egg types of the pest species and soil type in the study area. 5.2. Field experiments on horticultural crops to evaluate the efficacy of the selected agrochemical as Ovicidal-molluscicides for the pest control of key conventional grown horticultural crops.

5.3. Field analysis to know the collateral effect of agrochemical selects on invertebrate soil fauna and border effect on wild land snails in conventional horticultural crops.

WP. 6 .- To investigate the feasibility of using swine and cattle manure of killing land snail eggs and plant-tramp strategy of land snail pest control for key organic horticultural crops grown. To carry this out it will also be necessary:

6.1. Laboratory testing to determine concentrations of swine and cattle manure that are effective against the egg-lays of terrestrial gastropods pest. Discriminant trials will be made on filter paper (direct contact) and artificial soil.

6.2. Field experiments to evaluate the efficacy of swine and cattle manure land snail egg-lays as control for key organic horticultural crops.

6.3. Field analysis to investigate the collateral effect of swine and cattle manure on soil invertebrate fauna and border effect on wild land snails in organic horticultural crops.

6.4. Field experiments to use plants as tramp-deterrent method to protect organic horticultural crops WP. 7 .- Make field trials in traditional crops to compare the effectiveness of the control strategy of pest

land snail proposed by us versus conventional approaches of applying chemical molluscicides when observed damage to the crops. Field experiments in Conventional key horticultural crops to evaluate the efficacy of selected agrochemical ovicidal with activity against land snail egg-lays in relation to other standard commercial low-chemical methods of killing animals land snail. Final trial.

WP. 8 .- Field experiments organics in key horticultural crops to evaluate the efficacy of organic Molluscicides-ovicides and the use of plant-traps as land snails method to control pests.

WP. 9 .- To identify improved strains of nematodes Phasmarhabditis Which are more effective biocontrol agents of larger land snail species in Hispano-America.

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METHODOLOGY AND RESEARCH WORK PACKAGES

Work Package 1 Field research to investigate life cycle of land snail past in horticultural crops Participants 1 (……. Man-month) Participants 2 (……. Man-month) Participants 3 (……. Man-month) Participants 4 (……. Man-month) Participants 5 (……. Man-month) Participants 6 (……. Man-month) Participants 7 (……. Man-month) Participants 8 (……. Man-month) Participants 9 (……. Man-month) OBJETIVES To investigate size, structure and dynamic of land snail for a key horticultural crops Estudiar el tamaño, estructura y dinámica de sus poblaciones. BACKGROUND Methodological review. Many methods have been used to make quantitative studies of land snail

populations. According to South (1992) these methods can be qualified in three categories: A. Absolute methods, expresses the number of individuals per unit area. B. Relative methods, expresses the number of individuals per unit effort or the relation to non-standardized

traps. C. Indirect methods, expresses sizes of population in terms of traces left or the effects produced by land

snails (for example, depending on the damage extension done to the crop, or according to bait consumption). Relative methods are faster and much more comfortable but they have the disadvantage that the estimates

are highly dependent of the land snail activity levels, this can lead to incorrect population sizes because of weather conditions that have an important effect on the land snail activity (Getz, 1959; Hunter, 1968a; South, 1992).

Absolute Methods involve the absolute quantification of individuals per surface area. This can be done on site by applying an irritant substance to a determined area. For years formaldehyde was used for this purpose, but South (1964) rejected this method when he realized that most of the land snails died before they could reach the surface. Högger (1993) proposed a new method. First he determines an area, limiting it with a metal ring of 15 cm of height; then he introduces it in to the ground to a depth of 5 cm. Once done this, he applies mustard oil to the ground and captures the land snails that come out to the surface. Another method, proposed by Ferguson, Barratt & Jones (1989), it is based on the placement of shelter traps located inside of the area determined by the metal ring, which in this occasion is covered with a top that prevents the escape of land snails and helps to keep the humidity (moisture) inside. The shelter traps and the area inside the ring are inspected each day removing the land snails caught, until no new individuals appear.

Another way of obtaining absolute estimates is to take soil samples of known surfaces and transfer them to

the laboratory for further land snail extraction and quantification. The extraction can be done by; the progressive flooding of the soil samples to make land snails surface, or by washing the soil sample on sieves with water. South (1964) compared the last sieving method with absolute methods (flooding with cold or hot water, extraction with chemicals, dry sieving) and relative methods (trapping, Direct observation during night). South concluded that the water sieving is the most reliable method, since it allows the recovery of almost the 100% of the land snails contained in one soil samples. Hunter (1968a) proved the efficiency of this method when he recovered almost all individuals of a known population; South (1964) concludes that the water sieving is the most accurate method; It´s also is the only reliable method to obtain and quantify land snail lays.

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Almost all the authors agree that land snails and it´s lays are located in the uppermost stratum of the soil. According to South (1964), 100% of the lays and land snails of D. reticulatum species is located in the first 2 or 3cm of soil in field areas. In crop areas, Hunter (1966) found out that 83% of individuals of D. reticulatum were located in the soil first 7´5cm, and a 6% were located above 15cm. Also in crop areas, Runham & Hunter (1970) observed that in the first 10cm of soil contained the 97% of D. reticulatum. Rollo & Ellis (1974) pointed out that the 90% of snail lays are also located in the same soil layers. According to Marquet (1985), in standard conditions any land snail appears above the first 5cm of soil, However in exceptionally severe winters up to a 15% of individuals may appear at depths between 10 and 20cm. South (1992) suggests that sampling the soil´s first 10cm it´s enough to quantify land snail populations.

IMPORTANCE: The WP.1 is important because it will provide us with information of the size, structure and

dynamics of the land snail pest populations. This information will be used in the development of a predictive model.

PROGRESS ASSESSMENT AND RESULTS: Through semi-annual and annual reports, and by regular personal controls carried out by the coordinator to each one of the participants in the concerned country.

-------------------------------------------------------------------------------------------------------- Task 1.1 Objetives Studying the land snail population size in crops where the study will be conducted. Participants: all participants Materials and Methods The methodology used is based on absolute estimation methods; land snails quantification of known

surface soil samples. Soil sampling. The choice of the sampling location is randomly selected. To carry this out the plot is divided in a grid

composed at least of 50 frames of 4 x 4 m; each one subdivided in four quadrants. The quadrants are determined using the last two digits of the randomly generated value of the “random” (ran) calculator function. Once selected 20 frames, we proceed to determine which quadrant of each frame, by using again the calculator random number generator following the next code: 0,000-0,249 for the upper left quadrant; 0,250-0,499 for the upper right quadrant; 0,500-0,749 for the lower left quadrant; 0,750-0,999 for the lower right quadrant.

The sample extraction is performed with a rectangular spade mark with the depth to be achieved (10

centimeters). First is selected the area to take the soil sample, then we place on the ground an aluminum frame of 25 x 25 cm, then the spade is stuck in to the ground along its entire contour until the marked depth and extract the sample. Each sample is introduced in a properly labeled opaque plastic bag; these bags are conserved in a cold storage at 4ºC in the dark, for its further laboratory analysis in the next 3 days.

To determine the number of soil samples needed its used an statistical software; The means and variances

of the average land snails and eggs are calculated, to find out all the possible combinations of 2 samples, 3samples, 4samples till the total of 20 samples. For each number of samples the degree of error is calculated dividing the standard deviation by the arithmetic mean. In previous investigations we observed that to obtain a degree of error less or equal to 10%, 18 samples are enough in the case of land snails and 16 in the case of eggs.

The abundance of land snails and eggs is estimated by washing the soil samples precedent from the study plots. On a monthly basis 20 soil samples of 25x25 cm square and 10 cm deep.

Soil samples treatment. Each sample is placed individually on a white plastic tray; in the first place the

sample is thoroughly inspected to capture any snail that might be in the soil surface, once done this the vegetal cover is removed, cutting it with scissors; then the soil sample is washed in sieves, with a decreasing mesh sizes from 4mm to 1mm. The soil samples are crumbled to smaller pieces with the help of the water jet. The thicker roots are cut to smaller pieces. The sieves content is carefully inspected under a 10X magnifying glass and a powerful white light source.

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Larger land snails are retained in the upper sieve, and in the lower sieve the smaller land snails and the

eggs. The collected land snails are kept in a tray and the eggs in a Petri dish, both with a humid filter paper. Once finished separating eggs and land snails from the soil samples we proceed to identify them.

--------------------------------------------------------------------------------------------------------------------- Task 1.2 Objetives Field experiments to investigate the land snails pests populations dynamics and structure. Participant: All participants Materials and Methods To study the population structures and variation over time it´s necessary to determine the maturity state of

the land snails constituting the population along time. Because of this, Bett (1960), Hunter (1968a), and Hunter & Symonds (1971), based their studies exclusively on sperms presence or absence along the genital tract. South (1989a) used the same methodology, but he also obtained the gonad and albumin gland mass, (Hermaphrodite Gland Index, H.G.I. and Albumin Gland Index A.G.I. respectively, of each individual. This index express the % of corporal mass represented by each gland, that have a characteristically variation along the maturation cycle of D. reticulatum.

Duval & Banville (1989) and Barker (1991), in adition to calculating the H.G.I. and A.G.I., incorporated in

their work the gonad cytological analysis of individuals, and determined their maturity level using as reference the previous studies of Runham & Laryea (1968), based on the presence and relative abundance of each cellular type in gonad gametogenisis in D. reticulatum gonad. Haynes et al. (1996) classified the land snails in five categories based on the body mass index H.G.I. A.G.I. instead of using the cytological gonad analysis to describe the population structure. The population dynamic and structure study are only done on the species that really are a pest; in our previous studies (Barrada, 2003) we focused exclusively on Deroceras reticulatum that really constitute a pest in the European crops.

Individual management. Individuals belonging to the pest species are weight on a laboratory balance to the hundredth of milligram, and then they are sacrificed by a brief immersion in water at 50ºC. This method is a modification from the Haynes, Rushton y Port (1996) method, which is to dip them in boiling water. The sacrificed individuals are introduced in properly labeled glass tubes, with preservation 70% alcohol. Then the individuals are dissected and they have their hermaphrodite gland and albumin gland removed, which is used to determine the maturity state of individuals.

Following Barker (1991) approach, individuals with BMI(body mass index) exceeding 20 mg are not

dissected, assuming that they don´t have a differentiated gonad. The hermaphrodite and albumin gland, extracted from individuals with BMI greater or equal to 20 mg, the glands are weighed up to the hundredth of milligram immediately after its extraction. The hermaphrodite gland is fixed with Carnoy for 24 hours and preserved in alcohol 70%.

Determining the maturity degree. It will follow the methodology used by Duval & Banville (1989) and

Barker (1991), the sexual maturity status of individuals at each monthly sampling is determined by gonad cytological analysis, using as a reference the states defined by Runham & Laryea (1968) . To this end, each individual gonad was dehydrated by a series of ethanol baths of progressively higher degree (70%, 96% and 100%), ending with 2 baths of toluene. Next the sample is included in a paraffin block; which is sectioned at 8 μ thick. Obtained sections are rehydrated by reversing the previous dehydration, replacing toluene by xylene and dipping them in distilled water. Then the sections are stained using hematoxylin-eosin staining, and finally dehydratated again.

Each cellular type that appear in the selected land snails gonad and the maturity states are available in

Pelluet & Watts (1951), Watts (1952), Bridgeford & Pelluet

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(1952), Henderson & Pelluet (1960), Smith (1966), Runham & Laryea, (1968), Bailey (1973), Hill & Bowen (1976), Parivar (1978, 1980, 1981), Nicholas (1984), South (1992), Fawcett (1987) and Lutchel et al. (1997) works.

Each captured individual was identified as belonging to one of the following sexual maturity stages: i)

undifferenciated spermatogonia, ii) spermatocyte, iii) spermatid, iv) espermatozoa, v) oocyte and vi) senescent. The first three states correspond to immature land snails, with no reproduction ability; sexually mature land snails are those that are in spermatozoon and oocyte state (Runham y Laryea, 1968; South 1989a).

In other words, each captured individual is characterized by their body mass (mg), by their maturity state and by the mass (mg) of the hermaphrodite and albumin gland. From these values it´s calculated for each individual, the hermaphrodite gland index (H.G.I.) and the albumin gland index (A.G.I.), as follows,

H.G.I. = Hermaphrodite gland mass 100 / individual mass

A.G.I. = Albumin gland mass 100 / individual mass For each sample occasion, individuals who have similar characteristics referring to its sexual maturity state

and body mass index values, H.G.I. and A.G.I., are considered as belonging to the same land snail generation. In this research project we follow the methodology used by Duval & Banville (1989) and Barker (1991) for

the population structure study. As these authors did, we assume that individuals with body mass exceeding 20mg are land snails with completely undifferentiated gonads (from the cytological viewpoint) are not analyzed. In this regard, it should be mentioned that South (1989a) indicates a value of 40 mg body mass as a limit from which, the maturity state of D. reticulatum can be defined by studying the cytology of the testis. Previous obtained results agree with the values set by South (1989a).

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Work Package 2 Field research to investigate the feeding habits and the qualitative and quantitative diet of land snails in

horticultural crops. Objetives To Study land snail feeding and qualitative and quantitative composition of their diet in order to find plant

species that can be used as trap plants in organic farming. Participants 1 (……. Man-month) Participants 2 (……. Man-month) Participants 3 (……. Man-month) Participants 4 (……. Man-month) Participants 5 (……. Man-month) Participants 6 (……. Man-month) Participants 7 (……. Man-month) Participants 8 (……. Man-month) Participants 9 (……. Man-month) Background The study of the material ingested by land snails is carried out through microscopic analysis of the small

plant fragments found in their feces or inside the animal’s digestive tract. This method has been used by Grime, Blythe and Thornton (1970), Mason (1970), Wolda et al. (1971), Chatfield (1975), Richardson (1975), Williamson and Cameron (1976), Szlavecz (1986), Speiser and Rowell-Rahier (1991), Hatziioannou et al. (1994), to study the land snails diet as Cepaea nemoralis, Oxychilus cellarius (Müller, 1774), Oxychilus alliarius (Miller, 1822), Discus rotundatus (Müller 1774), Arianta arbustorum (Linnaeus, 1758), Monadenia hillebrandi (Smith, 1957), Monacha cantiana (Montagu, 1803), Monacha cartusiana (Müller, 1774), Braybaena fructicum (Müller, 1774), Helix lucorum (Linnaeus, 1758), Sandy Xeropicta (Ziegler 1827) and Cepaea vindobonensis (Férussac 1821). Hunter (1968b), Pallant (1969, 1972), and Jennings and Barkham (1975) (it) has been used to study the slugs diet as Deroceras reticulatum, Tandonia budapestensis (Hazay, 1881), Arion hortensis (Férussac, 1819), and Arion ater.

The study of feces is a faster method than the analysis of the stomach contents, as it does not require the slaughter or dissection of the animals. However, the substances present in the feces have passed through all the animal´s digestive tract and are more degraded, making them harder to identify than those extracted from the stomach (Cook and Radford, 1988; Hatziioannou et al., 1994).

The feces study is a suitable method to verify the presence or absence of certain elements in the diet of the animals, but the proportion of unidentified substances in the feces is often so high that it is not an effective method to perform a quantitative characterization of the diet (Williamson and Cameron, 1976; Szlavecz, 1986; Speiser and Rowell-Rahier 1991). Vadas (1977) points out that the most numerous and more easily recognizable substances in the animals feces are metabolically the least used and perhaps, are also eaten in smaller amounts, so the feces analysis leads to an overestimation of the little nutrients consumed; on the other hand, the most consumed nutrients are underestimated because they are ingested in great quantities and therefore become more scarce and difficult to identify in the feces.

The analysis of the animals digestive tract contents is a tougher method, but it provides a more realistic picture of the diet of the animals (Norbury and Sanson, 1992). Pallant (1969, 1972) studied the diet of D. reticulatum in natural populations (forests and grasslands) through the analysis of stomach contents. To minimize gastrointestinal degradation of the food eaten by the captured land snails and slugs and to facilitate their identification, Pallant (1969, 1972) introducing the captured animals directly in 70% alcohol and made their dissection to extract the contents of the digestive tube tract during the following 2 hours after they were caught. Triebskorn and Florschutz (1993) conducted a study on the transit of the food through the digestive tract of D. reticulatum, using a special food preparation (lettuce, corn and milk powder) radioactively marked. Subsequently x-rays were done to the animals at regular intervals; according to their results, the ingested food penetrates immediately in the maw, stomach and anterior intestine, where it remains for a minimum period of at least two hours and a half before starting to move through the intestine; the maw and the stomach are completely emptied in the following thirteen hours after ingestion.

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Importance: the information obtained in this study will be useful to designing the strategy of the plant-traps use as deterrent agents, and also to understand and assess the real damages on crops.

Progress assessment and results. Through semi-annual and annual reports, and through regular personal controls, that the Coordinator will carry out at each participants concerned country.

Work and methodology Description Sampling. The diet study is done by the analysis of the digestive tract content of 20 land snails captured in

monthly samplings. The captures of these animals are randomly done in the course of a run across the whole plot. Only the collection of small land snails is avoided, because their processing is very difficult. The capture of the land snails is done during the next 3 or 4 hours after sunset, which is when it takes place the main feeding activity of land snails and D. reticulatum in particular (Dobson & Bailey, 1982, Rollo, 1988ab; South, 1992; Hommay, Lorvelec & Jacky, 1998). Every plant species, where each land snail is captured, is recorded. The land snails are individually introduced in plastic containers with a piece of damp cotton for its further transport and storage in the laboratory.

The day after the capture of the land snails, the plot´s vegetation composition is determined. The different plant species coverage percentage is estimated using a measuring tape extended along the plot´s principal axis, at intervals of 0.5 meters. The tape´s intersection points with the different plant species are noted, and with this data the coverage percentage for each species is calculated. Graminaceous are considered as a whole.

Individual Processing. The land snails caught for the diet analysis are transported and processed within 2 hours, in order to minimize digestive degradation of their stomach contents (Pallant, 1969, 1972).

Land snails are weighed on a scale up to the hundredth of milligram, before being sacrificed by immersion in hot water (50 º C). Then the individuals are dissected and have their maw removed, which is placed onto a slide. Under a binocular microscope the maw is open along the longitudinal axis and all its contents are collected, using a fine brush. Each land snail´s stomach contents is weighed to the hundredth of milligram and immediately introduced into a small plastic tube, were 2 milliliters of 1N hydrochloric acid is added to eliminate the mucus and epithelial debris from the maw sample (Hatziioannou et al., 1994). Stomach contents are always analyzed in the following two days after its capture.

Qualitative diet determination. The qualitative diet study is based on the food fragments identification, found in the snails´ digestive tract. The plant fragments identification is possible through epithelial formations such as stomata and trachoma. The appearance and distribution of these formations are characteristic for each plant species.

Before starting the samplings, land snails are captured around the study area, they are fed with a monospecifical plant diet, and their feces are used to make an image collection of the fragments found in the land snails´ stomachs. These land snails are kept inside a climate chamber with a photoperiod of 12 hours, 18 º C and 90% relative humidity. All land snails are housed inside transparent plastic boxes 20 × 20 × 10 cm, with the bottom covered with a damp filter paper, and all the plastic boxes are perforated to allow the air renewal. In each case four individuals are kept. After a period of 72 hours without food all land snails will be fed with a diet consisting on a single plant species collected in the study plot. Then after six hours, the land snails are processed as described in the previous section, and the fragments found in their stomachs are photographed under a microscope (Barrada, 2003).

For each plant we have a large reference image collection of each one after being eaten by land snails. The captured land snail´s digestive tract is studied using the same microscope equipment. Fragments found in the land snails´ digestive tract are compared with the reference images collection for their further identification.

Quantitative diet determination. To determine the land snails´ quantitative diet composition, we follow the methodology used by Hatziioannou et al. (1994). A stomach sample content of each land snail is taken; the surface area of each fragment is measured with the help of a software image analysis (SPSS ® Sigma Scan Pro Image Analysis Version 5.0.0.). The surface areas of all the same type fragments are added together and the percentage is calculated and represented in relation with the sum of all the fragments contained in the same sample surface areas. According to this, each plant contribution to the land snail diet is estimated.

Using 0.24 ml sample taken from the stomach contents of each land snail, diluted with 2 ml of hydrochloric acid and then homogenized with the aid of an agitator pressure SBS ® AT-1 or similar. All fragments of food contained in the sample are photographed, identified and measured.

The sample volume used for each digestive tract is previously determined from the land snails´ stomach contents study, whose diet is known. To this purpose four land snails are individually housed in each plastic box.

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After a fasting period of 72 hours, the land boxed land snails are provide with a small piece of 3 different plant species, and 6 hours after they are sacrificed and their stomach contents are removed, as previously described. Their stomach contents are diluted in 2 ml of HCl for their analysis in 6 successive 0.08 ml samples. In previous researches (Barrada, 2003), it was determined that a 0.16 ml sample (8% of total volume) is an accurate representation of the land snails stomach contents composition, equivalent to a 0.48 ml sample (24% of the total volume). To minimize the error degree it is always used a 0.24 ml sample (12% of total volume) for the digestive tract contents representation.

Diversity and selection Indexes. The diversity land snails´ diet and the vegetation variety in the study plots, is calculated through the Shannon-Weaver index, H '= Σ (pi) (log2pi), where pi is the frequency for each component of the diet or vegetation (Margalef, 1982).

The index C is used as a selection index, Pearre (1982), this index reflects the outcome of predator-prey interaction taking into account the abundance in the environment for each prey type. This index has a value ranging from +1 and -1, where C = 0 indicates no selection. It shows if a plant is consumed above (positive values) or below (negative values) it´s expected consumption according to their availability in the nature. This index is based on the χ2 test that allows establishing the significance of the selection degree for any sample size.

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Work Package 3 Objetives To develop a statistical method to explain and predict the land snails´ activity according to atmospheric

conditions. Participants 1 (……. Man-month) Participants 2 (……. Man-month) Participants 3 (……. Man-month) Participants 4 (……. Man-month) Participants 5 (……. Man-month) Participants 6 (……. Man-month) Participants 7 (……. Man-month) Participants 8 (……. Man-month) Participants 9 (……. Man-month) Background

The terrestrial gastropods activity is regulated by complex mechanisms that involve both, external (environmental) and internal (endogenous rhythms) factors (Bailey and Lazaridou-Dimitriadou, 1986; Aupinel 1987, Young & Port, 1989, Cook, 2001).

As a general rule, it is recognized that these animals spend their days idle at their shelters, their activity is mostly nocturne, and the weather conditions largely determine their state (Hommay et al., 1998). However, it recognizes the existence of interspecific differences in regard to their activity patterns and the influence of environmental factors on them (Cook, 2001). It has also hinted at the existence of intraspecific differences in the relative importance of different environmental factors that control the activity, depending on the microclimate or the type of living environment in each population (Lorvelec and Daguzan 1990, Iglesias and Castillejo, 1996 ). The close dependence of terrestrial gastropods have regarding environmental conditions qualities make them ideal models for studying the relationship between animal behavior and climate (Rollo, 1982). His character of agricultural pests is another aspect that helps explain the interest shown by many researchers in the study of their activity in relation to climatic conditions (Dainton, 1954ab; Getz, 1963; Webley, 1964; Newell, 1968, Cook and Ford 1989, Young & Port 1991, Young, Port, Emmet and Green, 1991; Hommay, Lorvelec and Jack, 1998, Grimm and Kaiser, 2000, Barred, 2003).

One way to deal with this study, which is receiving much attention in recent years due to its applicability in the pest control field, is the development of predictive models or expert models (Bohan et al., 1997, Cook, 2001). Predictive models of activity as a matter of pest animals represent a useful tool from the applied point of view, since they allow to predict periods when crops can suffer further damage and to optimize use of the pesticides used to control reducing economic and environmental costs resulting from its application at times when there is no risk of damage to crops (Frahm, John, and Volk, T. 1996; Hommay et al., 1998; Hommay, 2002; Port and Ester, 2002).

Importance: data obtained after this investigation will be used to develop a statistical model to predict abundance and activity.

Progress assessment and results. Through semi-annual and annual reports, and through regular personal controls, that the Coordinator will carry out at each participants concerned country.

Methodology The Barrada(2003)methodology is followed to develop the activity models, this methodology is based on

the ones designed by Young & Port (1989) and Young et al. (1991). These authors identified, for the diverse environmental conditions, the limits that define the high land snail activity nights, without attempting to mathematically describe the limits that define those lines, but only giving the extreme values to define the

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optimal range values for each variable. These models predict that high activity nights will be those with all the variables comprehend between their optimal ranges.

Sampling. The land snail´s activity analysis is done in the same study area used to analyze its feeding habits. The samplings are done during three consecutive nights per month; this means a total of 72 nights during the sampling period. Every night two investigators examine during three hours the study area, searching for active land snails. The tours are done thoroughly, searching the soil and vegetation, but no effort is done to locate those out of the observer´s sight.

At the beginning of each sampling and subsequently, at 1 hour intervals, the soil temperature and the relative humidity are registered at 5 cm above ground. The soil temperature can be registered with an electronic thermometer fixed in the ground at 10 cm deep, and the temperature and the relative humidity are registered with an electronic thermo-higrometer or a similar instrument.

Variables and Statistical analysis. In the model, the activity is divided in three categories (low, average and high) according to the number of active land snails registered during 72 nights in each study area. This activity levels made up the dependent models or response variables, in other words, (the variable whose value was expressed in terms of the values of the independent or predicted variables). Within these environmental variables were considered measures on site during the samplings are considered as independent variables measures on site during the samplings is considered as independent variables: i) the atmospheric conditions registered on site during the samplings; ii) atmospheric conditions corresponding to the sampling day and the days before it, according to the data registered in a near thermo-pluviometric station; iii) time variables (month, season); and iv) related variables with the land snails population dynamics in the sample area. All considered variables are compiled in table 5.1.

Because of the data nature, qualitative response variables with more than two categories (three categories: low, average and high) and a serial of qualitative (factors) and quantitative (co variables) variables as independent variables, the statistical procedure to relate the land snails activity level with the independent variables was the ordinal regression (McCullagh, 1980; McCullagh & Nelder, 1989). To carry out the data analysis it can be used the SPSS, using the PLUM method for the ordinal regression (SPSS documents), or any related statistic programs (Barrada, 2003).

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Work Package 4 To search for a Biopesticide elaborated of plant extracts with molluscicide and ovicide action. Participants 1 (……. Man-month) Participants 2 (……. Man-month) Participants 3 (……. Man-month) Participants 4 (……. Man-month) Participants 5 (……. Man-month) Participants 6 (……. Man-month) Participants 7 (……. Man-month) Participants 8 (……. Man-month) Participants 9 (……. Man-month) Objetives To investigate the feasibility of using plant extracts as biomolluscicides and bio-ovicides. Background Farmers in their traditional knowledge have identified and used a variety of plant products and extracts as

pest control, especially in storage. 2121 plant species are reported to have pest management properties, 1005 species showed insecticide properties, 384 with antifeed properties, 297 with repellant properties, 27 with attractant properties and 31 with growth inhibition properties. The most commonly used plants are neem (Azadirachta indica), pongamia (Pongamia glabra) and mahua (madhuca indica). 2-5 % neem or mahua seed kernel extract has been found effective against rice cutworm, tobacco caterpillar, rice green leafhopper, and several species of aphids and mites. The efficacy of vegetable oils in preventing infestation of stored product pests such as bruchids, rice and maize weevils has been well documented. Root extracts of Tagetes or Asparagus as nematicide and Chenopodium and Bougainvillea as antivirus have also been reported 'Biopesticide' can reduce pesticide risks, because- (a) Biopesticides are the best choice instead of the conventional pesticides, and usually inherently less toxic. (b) Biopesticides generally affect only the target pest and a wide spectrum of closely related organisms, conventional pesticides that may affect organisms as rent as birds, insects and mammals. (c) Biopesticides are often effective in very small quantities and decompose quickly, thereby resulting in lower exposures and widely avoiding the pollution problems caused by conventional pesticides. (d) When used as a fundamental component of Integrated Pest Management (IPM) programs, biopesticides can greatly decrease the use of conventional pesticides, while crop yields remain high. (e) Amenable to small-scale, local production in developing countries and products available in small, niche markets that are typically unaddressed by large agrochemical companies.

The European Commission has published a proposed European Regulation concerning the placing on the market and use of biocidal products. The new European Regulation would replace the current regulatory regime for biocides, which is laid out in the Biocidal Products Directive 98/8/EC and transposed into UK law by the Biocidal Products Regulations 2001. Once in force (scheduled for January 2013(2019?)), the European Regulation would be directly acting on all Member States, including the UK.

Methodology 1.-MATERIAL AND METHODS Extracts preparation for the tests: The plants selection will be made using two parameters: abundance, medicinal and therapeutic

properties. Previously it is necessary to consult references on bibliography. The plants will be collected in the field and identified.

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The extracts will be made with different fresh plant parts, after drying in an oven at 50ºC (Medina and

Woodbury, 1979 & Makkar et al 1991) or at temperature room?(climaticed room). The different plant parts (leaves, steams, fruits, flowers, roots and seeds) will be separated and crushed to powder. The powder of the plant will be put in glass flasks, where they will be spilled to the solvent, leaving it macerate during 24 hours at a temperature room. Passed these 24 hours the samples with acetone/water will be extracted with the help of a Rotary Evaporator machine (Mendes et al, 1993).

The concentrations used will be: 80,000 ppm, 40,000 ppm and 8,000 ppm to the water extracts; 100,000

ppm and 50,000 ppm to the extracts made with acetone/water; and: 100,000 ppm, 50,000 ppm, 10,000 ppm, 1,000 ppm and 100 ppm for both extracts.

Molluscicide activity: The tests will be made using 10 cm diameter glass Petri dishes with a 9 cm diameter filter paper on the

bottom (Albet 400, of 90g/m2 of weight and a thickness of 0.21mm). 1 ml extract will be put in each dish, using a pipette that had a milk filter (Alfa Laval Agri) in the tip to avoid that the pipette gets plugged with the plants´ powder. The dish with the filter will be dried off at a temperature room; once it´s completely dry it will be moistened with 1 ml of distilled water, then the eggs-lays will be deposited on the filter (5 eggs in each dish, ater the presvious eggs selection).- They will stay in the incubation chamber until the death or hatching. To check the effect of the extracts on the embryo the eggs will be observed each 24 hours, with a binocular magnifying glass using an oviscope (glass tube).

The water extract controls will be made with distilled water, while controls for the acetone/water extracts

will be made with an acetone/water mixture (in the proportion: 7:3), then the acetone will be removed with the Rotary Evaporator Machine and the water will be used as controls.

To do the tests, 1 ml of water will be put on the filter papers, when the paper dries off at the temperature

room, another millilitre will be put on to it and then the eggs-lays will be placed on the moist paper. The Petri Dishes will be closed and put in the incubation chamber.

The same process will be made with the standard soil and with the commercial substratum. At this time

plastic Petri dishes of 9 cm of diameter will be used to these purposes. 1 ml of extract will be put on 25g of standard soil with a humidity of 35%, and the same process will be made with the substratum.

Task 4.1. To carry out laboratory experiments to find plant extracts that can be used as biopesticides to control

agricultural land snail pests. Objectives. Laboratory tests on filter paper (direct contact) and standard soil to select plant extracts with molluscicidal

and /or ovicidal action.

Materials and Methods Egg-lays. The land snails are collected in the crops to obtain the egg-lays to make the experiments once in

the laboratory. The individuals are kept in plastic boxes (25 x 25 x 15 cm) with perforated walls and lids and the floor covered with a moist filter paper. Black small polyethylene tube pieces are used as shelters for land snails, using as food, lettuce, carrots, cabbages, runner beans, potatoes, and mushrooms, supplemented with powdered CO3Ca. The cages are placed in a climatic chamber at 17ºC day/15ºC night with a 12D:12L photoperiod and 85% relative humidity. Cleaning and food replacement will be done twice weekly. Since land snail breeding takes place whenever environmental conditions are suitable (Carrick, 1938; South, 1989), the cages are inspected looking for egg-lays every day. The land snails laid their eggs directly on the filter paper, mainly in those places covered by

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pieces of food. The eggs will be collected, cleaned with distilled water and incubated on wet filter paper inside Petri dishes in the darkness at 18ºC. The entire course of the development of the embryo is observable when the egg is immersed in water and viewed under transmitted light (Carrick, 1938). About a week after collection, all the eggs will be inspected under a binocular microscope for the selection of those to be used in the experiments. Only eggs containing a single living embryo and without foreign inclusions will be selected for the tests, and they will be used when the embryo achieved the developmental stage IV according to Carrick (1938), which is recognisable by the differentiation of rudiments of the tentacles and posterior sac. The movement of the embryo in the form of a slow and continuous rotation, and the rhythmical contractions and expansions of the still-small posterior sac were the criteria used to determine whether the embryo was alive. Non-motile embryos were considered to be dead. Carrick (1938) described the structure of the egg and the embryonic development of Deroceras reticulatum, while the histochemistry of the egg was described by Bayne (1966, 1968) and Barrada (2003).

Contact toxicity tests on artificial soil. The tests will be made in glass Petri dishes 9 cm in diameter, containing 40 g of standard artificial soil. The standard soil (OECD, 1998) consisted of 10% peat, 20% kaolin and 70% quartz sand. The pH will be adjusted to six with calcium carbonate and soil moisture was set to 35% (w/w). All tests consisted of a control and five doses of the compound, arranged in a two-fold geometric series, with five replicas each and five eggs per replica. For each compound, the appropriate quantity will be dissolved or suspended in 20 ml of distilled water. The appropriate quantity of compound will be calculated, according to its formulation, to render the desired maximum concentration of a.i. cm-2 when applying two ml of the solution or suspension to the soil surface of one dish. Thereafter, ten millilitres of that solution or suspension will be applied to the soil (2 ml per replica of the highest concentration) and the remain ten millilitres will be diluted to 50% to obtain the next lower concentration. Dilutions and application of the solutions or suspensions will be made under continuous shaking. The different treatments will be applied spraying 2 ml of the solution or suspension on each Petri dish. Two ml of distilled water will be applied to the control dishes. After treatment, the soil in the Petri dishes will be allowed to dry for 24 hours at room temperature. Immediately before the start of the test, the soil was moistened again with distilled water in an amount calculated from the weight loss of the controls. Then, five eggs will be placed on different points of the soil surface of each dish. After the start of the experiments, all the eggs will be inspected under a binocular microscope every 24 hours to assess whether the embryos were still alive. To avoid immersion of the eggs in water to view them under the microscope, which would produce an undesirable wetting of the eggs, the test-eggs will be introduced into a glass tube, 3 mm inner diameter; in this way through the contact zone between the egg and the glass is possible to see the embryo inside the egg producing the same effect as immersion in water. Median lethal doses (LD50), i.e. the calculated doses which produce 50% mortality of the eggs, and 95% confidence limits, will be calculated by probity analysis. For the different compounds, LD50s will be calculated for periods of exposure in which at least one of the doses tested resulted in no mortality and one in 100% mortality (OECD, 1998, Guideline 313 de la OECD (OECD Guidelines for the Testing of Chemicals. Contact toxicity tests on artificial soil/ wet filter paper). (Iglesias, J., Castillejo, J., Parana, R., Mascato, P. and Lombardía, M.J. 2000; Iglesias, J., Castillejo, J. y Ester, A., 2002; Iglesias, J., Castillejo, J., Ester, A. y Lombardia, M.J., 2002)

Task 4.2

To test the plant extracts biomolluscicides activity on non human destined agricultural soil mini plots. Objetives. To make mini plots experiments on horticultural soil to evaluate the efficiency of the selected plant extracts

against lands snails and its egg-lays. Materials and Methods TEST PLOTS In each study plot six areas are defined in squares of 4 x 4 m= six mini plots of 16 m2. Plant extracts with a

specific composition are tested in five mini plots and the sixth acts as the control plot. Between each mini plot there is a security area of 2 m (wide).

In each mini plot there are placed five Bayer trap-shelters and five PVC tubes specific for land snails. These tubes are our invention; they are PVC tubes of 30cm long and 5 cm in diameter and closed at one end. They are

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placed on wooden supports fastened and nailed around the crop plants, with its hole facing down so that the snails can take shelter inside (Córdoba, 2009).

PLANT EXTRACTS CONCENTRATION AND APPLICATION METHODS From each plant extract with ovicidal action we know the LD50 in grams per square meter, we also know

the surface that will be treated (4 x 4 m2), and we know how much depth we can find in the soil the eggs of land snail, so it will be easy to calculate the amount to apply in each mini-plot (to be applied in every mini plot.).

Knowing when to apply the plant extract with ovicidal action, that is, to know the period of the year when to apply them, (in which it is necessary to apply that) is determined by the land snails´ life cycle. Pests that have been studied in the first year of the project will be applied when the land snails population density and egg-lays in the soil is minimum. It will be done as many agrochemical applications will be done as valleys or sinuses that the land snails´ life-cycle curve has in the study area.

Shelter traps are placed after each treatment and are removed during the application of agrichemicals (the agrochemical one.)

SAMPLING The time length: 24 consecutive months (2 years). The sampling frequency: monthly. Before carrying out

the first agrochemical application, the gastropods population density will be estimated in the land parcels object of study. The estimation will be done following the methodology exposed in Work Package 1, Task 1.1.

After the application of the plant extracts with ovicidal action it will be estimated in each area during 2

consecutive years with a monthly periodicity, the land snails population density and egg-lays found under the shelter traps.

Every month the shelter traps will be lifted and the land snails identified, all captured individuals will be

measured and weighted. The eggs found under the traps will be identified and counted (Cordoba, 2009). Every six months a statistical analysis will be done based on the data obtained from the land snails

population density and the egg-lays. After the first year of testing, and once collected the harvest, it will be done another estimation of land snails and egg-lays population density in the soil according to the methodology outlined in Work Package 1, Task 1.1.

CROP DAMAGES EVALUATION After the treatment with plant extracts, land snail damages will be assessed during 3 days and thereafter at

monthly intervals during the crop´s life. Damage will be recorded as the percentage of leaf area eaten (percentage leaf loss) to the nearest 5% and will be assessed separately for each plant. For statistical analyses the recordings of 20 plants growing in the same plot were averaged, because they reflect the activity of the same land snail population, resulting in six independent assessments for every treatment. A random program will be used for 20-plants selection. Data for land snail damage (percentage leaf loss) will be transformed to angles prior to analysis of variance. Data for number of damaged plants and for number of land snails were compared non-parametrically by the Mann-Whitney U-test. (Iglesias, Castillejo and Castro, 2001, 2001b and 2003)

Task 4.3. To carry out Mini plots analysis to know the collateral effect of plant extract selects on

invertebrate soil. Materials and Methods In areas near the test plots, and following the methodology of sampling and analysis presented in the Work

Packcage 1, Task 1.1, soil samples are taken to study side effects. On a sieve of appropriate mesh size worms, mites and springtails are collected. To determine the possible toxicity of the chemicals used in the study plots tests will be conducted recommended by the OECD GUIDELINES FOR TESTING CHEMICALS (1th draft, November, 2007 Proposal for a New Guideline) on earthworms, mites and springtails.

The assessment (valuation) of edge effects on terrestrial gastropods in wild areas near and without plant extracts treatment (untreated) it will be applied the same methodology of the Bayer type trap shelters and PVC pipes has now been exposed (WP. 4, Task 4.2.)

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Task 4.4 Carry out Chemical analysis to find the plant extracts active principle by analytic steps. These kinds of analytic analysis and posterior test on eggs, snails and slugs will be carried out by Syngenta

Company at Switzerland laboratory in collaboration with Participant 1 (Coordinator) for effectiveness as molluscicide and ovicide activity against slugs, snails and eggs and parallels studies on collateral effects on soil fauna. These analyses and the USC collaboration will be out of global work packages of this project

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Work Package 5 Destroy land snail egg-lays with non residual agrochemical compounds. PARTICIPANTS Participants 1 (……. Man-month) Participants 2 (……. Man-month) Participants 3 (……. Man-month) Participants 4 (……. Man-month) Participants 5 (……. Man-month) Participants 6 (……. Man-month) Participants 7 (……. Man-month) Participants 8 (……. Man-month) Participants 9 (……. Man-month) OBJETIVES To investigate the feasibility of using commercial agrochemical with ovicidal activity to kill control land snail

eggs for key conventionally grown horticultural crops BACKGROUND According to Mendis et al. (1996), among the many methods proposed to control gastropod pests in

agriculture, the eggs of the animals have received very little attention. Particularly, there is very few data of the effect of pesticides or other substances on the viability of terrestrial gastropods eggs (Stringer & Morgan, 1969, 1970, 1972, cited by Godan, 1983; Ryder & Bowen, 1977a). The high susceptibility of the eggs of D. reticulatum in contact with low doses of metal salts (Iglesias et al. 2000) and with some pesticides (Iglesias, Castillejo & Ester, 2002) has been demonstrated recently in paper contact-toxicity tests.

Importance: Traditional molluscicides can only kill some land snails, but with the use of ovicidals all the egg-lays will be destroyed, this ovicidals are agrochemical compounds usually managed by the farmer.

Progress Assessment and results. Through the weekly and annual informs, and by regular personal controls carried out by the Coordinator in each of the participants´ concerned country.

WORK AND METHODOLOGY DESCRIPTION See tasks ----------------------------------------------------------------------------------------------------- Task 5.1 To carry out laboratory tests whit the selected commercial non residual agrochemicals with ovicidal action. Objectives To make laboratory tests on filter paper (direct contact) and on standard soil to select the non residual

commercial agrochemicals that are more effective against land snails lays, for their posterior use as ovicides in crops.

Agrochemical selection. To search for non residual agrochemicals compounds with ovicidal action. Agrochemicals used in the tests are the ones approved by the local authorities.

See Work Package 4, Task 4.1. Changing plant extracts for non residual agrochemicals. Task 5.2 Field experiments to investigate the feasibility of using non residual agrochemicals compounds with ovicidal

activity of killing land snail eggs. Objetives Field experiments on horticultural crops to evaluate the efficacy of selected agrochemical as molluscicide-

ovicicde of killing land snail eggs of control for key conventionally grown horticultural crops. Final trial.

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Materials and Methods See Work Package 4, Task 4.2. Changing plant extracts for non residual agrochemicals. Task 5.3 To investigate the collateral effect of agrochemical compounds on soil fauna and wild land snails. Objetives Field analysis to know the collateral effect of agrochemical selects on invertebrate soil fauna and border

effect on wild land snails in conventionally horticultural crops. Materials and Methods See Work Package 4, Task 4.3. Changing plant extracts for non residual agrochemicals.

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Work Package 6 To carry out laboratory and field experiments to investigate the feasibility of using livestock manure and

trap-plants as land snail pest control in non human consumption organic farming. PARTICIPANTS Participants 1 (……. Man-month) Participants 2 (……. Man-month) Participants 3 (……. Man-month) Participants 4 (……. Man-month) Participants 5 (……. Man-month) Participants 6 (……. Man-month) Participants 7 (……. Man-month) Participants 8 (……. Man-month) Participants 9 (……. Man-month) Objetives To investigate the cattle and swine manure feasibility as a land snail eggs-lays killer and the use of trap-

plants as a control strategy against land snail pests for key organic horticultural farms. Background The use of animal manure as organic fertilizers is widely used in agriculture. Knowledge of its toxicity to soil

fauna has also been shown. Participant 1 has proved in previous tests that pig and cow manure at low concentrations has ovicidal capacity

In particular we have seen that in order to remove all the land snails and slugs eggs that are in a hectare of land it is necessary to spread....10.000 kg of cattle manure or 8.000 kg of swine manure. The destruction of the eggs takes place 5 days before the application.

The use of trap-crops is an alternative that is receiving increased attention from researchers. This strategy is based on the coexistence with other crop species of low economic value that serve as food for snails and slugs, and that are more attractive to them than the crop species. The vast majority of research done in this respect are laboratory experiments, in which we seek cultivated species, that are more attractive to snails and slugs than the crop species but the results obtained can hardly be extrapolated to real field situations, where the snails and slugs diet is very strongly conditioned by the availability of different crop plants.

The species used as trap-crops must fulfil its protective function to remain plentiful as they compete with crop species for nutrients.

In the current study we found that D. reticulatum selected the wild species Sonchus Oleraceaus to feed on. This specie made a significant contribution to the slugs and snails diet, despite being a rare one on the plot

study, it is a good candidate to be used as a trap-crop. Its ability to reduce the snails and slugs damage to crop species should be evaluated in field trials.

Swine and cattle manure are generally used as fertilizers on pastures and crops. We consider the manure as any other agrochemical compound and for this reason the methodology to be used will be the same one.

Importance: The use of cattle and swine manure as ovicides to kill the slug and snails eggs is something new, and in this way it is considered to be a very useful tool in organic farming.

Assessment of progress and results: Through the semi-annual and annual reports and regular personal controls carried out by the coordinator to each of the participants in the concerned country.

WORK AND METHODOLOGY DESCRIPTION Plot selection. To carry out this study, an organic farming plot is selected, were no chemical pesticides nor

fertilizers are used. Cattle and Swine manure. The cattle and swine slurries are considered as animal origin agrochemicals

suitable for ecological farming. ---------------------------- Task 6.1. To carry out laboratory tests with cattle and swine manure to determine their ovicidal action against land

snail egg-lays.

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Objetives To carry out laboratory tests to determine the effective concentration of the cattle and swine slurries

against land snails and egg-lays. The linear discriminant analysis done on filter paper (direct contact) and on standard soil, first and second screening.

Material and Methods The methodology set out in Work Package 4, Task 4.1 for agrochemical compounds will be followed. ------------------------------- Task 6.2. Objetives Field experiments to evaluate the cattle and swine manure efficiency as land snail eggs-lays control for key

organic horticultural crops. Final trial. Material and Methods The methodology set out in Work Package 4, Task 4.2 will be followed. Plant’s crops damage evaluation See Work Package 4, Task 4.2. ------------------------------- Task 6.3. Objetives To carry out field analysis to find out the cattle and swine manure collateral effects on invertebrate soil

fauna and the border effect on wild land snails in organic horticultural crops. Material and Methods The methodology set out in Work Package 4, Task 4.3 will be followed. ------------------------------- Task 6.4. Objetives Carry out field experiments to use trap-plants as a deterrent method of protecting organic horticultural

farms. Material and Methods Trap-plants. One of the consequences of the development of WP.2 is to find out which plants that exist in

the study area is more attractive for land snails. These plants have a high selection index and remain low in abundance. The trap-plants have little or no nutritional valor, these plants have to coexist with the crop plants and at the same time be an attractive alternative food source for pests, to reduce the damages done to the principal crop. This plants must also be in a low abundance, so that they don´t compete for the resources with the principal crop.

Methodology. To design the mini plots tests the followed methodology is the same as the one explained in WP.5, Task 4.2. The trap-plants are set around the mini-plots, as a stockade or in between the crop plants. The organic farming damages will be done by estimating the foliage loss or estimating the number of damaged leaves, caused by land snails, to evaluate the effectiveness of the trap-plants.

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Work Package 7 Study the effectiveness of agrochemicals with ovicidal action on conventional horticultural crops. PARTICIPANTS Participants 1 (……. Man-month) Participants 2 (……. Man-month) Participants 3 (……. Man-month) Participants 4 (……. Man-month) Participants 5 (……. Man-month) Participants 6 (……. Man-month) Participants 7 (……. Man-month) Participants 8 (……. Man-month) Participants 9 (……. Man-month) OBJETIVES Field experiments in conventional key horticultural crops to evaluate the efficacy of selected agrochemical

with ovicidal action against land snail egg-lays in relation to other commercial standard low-chemical methods of killing gastropods. Final trial.

BACKGROUND

Integrated pest control and prediction models. Traditionally, the concept of pest control in agriculture posed as a core objective, the total elimination of the causative agent of plague, by applying pesticides, when it was detected in culture. In the 60s, in Europe and the U.S., does the concept of integrated pest control (IPM) (Stern, Smith, van der Bosch and Hagen, 1959), which currently is part of another broader concept, which is the sustainable development. The IPM involves the integration of knowledge from many fields (biology, chemistry, agronomy, climatology, economics, etc) in order to develop control strategies most suitable economically, environmental and public health (Dent, 1991). As a system based on the combination of different methods in order to minimize the use of chemical pesticides, is not excluded, a priori, the use of any control agent (Coombs and Hall, 1998).

Methodologically, the IPM can be described as a "decision making process" is that on the basis of all available relevant information, decide what action to take and when to apply them, to control plague it, besides effective as profitable as possible from the economic standpoint and the least aggressive as possible from the environmental viewpoint (Bechinski Mahler and Homan, 2002).

Predict when a pest can cause significant damage to a crop is essential to make a decision regarding the need for pesticides (Buhler, 1996). Without a system for predicting the risk of damage, the choices faced by a farmer are either not apply pesticides or apply them consistently follow a precautionary approach. The first option may involve a loss of production in the case of a pest occurs. The second option involves economic and environmental costs that will be unnecessary if the pest does not occur. Therefore, for the rational use of pesticides is necessary to have criteria to determine the needs for its implementation. Currently, integrated control programs for many species of arthropods and fungi pests in a variety of crops, based on the use of risk prediction systems (Dent, 1991; Frahm, John and Volk, 1996).

In the case of terrestrial gastropods pest control, in recent years there has been progress, the commercialization of biocontrol agent P. hermaphrodita, aimed at achieving a reduction in use of chemical molluscicides, but the weak point of pest snails and slugs remains the lack of criteria on which to base the decision to apply molluscicides (Hommay, 2002, Port and Ester, 2002). In this situation, farmers choose, usually by the systematic application of molluscicides in their crops (Bohan et al., 1997, Speiser and Kistler, 2002). From the standpoint of the farmer, this is justified because the slugs can attack and severely damage crops in any season (Port and Port, 1986), because many crops, especially horticulture, are extremely sensitive to attack

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slugs from the time of planting to harvest, and the low economic cost of conventional chemical molluscicides (Port and Ester 2002).

Importance: thanks to the predictive model of activity and knowledge of the life cycle of snails and slugs will know when and how to apply molluscicides, thereby achieving high efficiency, leaving benefit farmers, consumers and the environment.

Progress and results assessment. Through the quarterly and annual reports, and through personal checks done regularly by the coordinator to each of the participants in the country concerned

WORK AND METHODOLOGY DESCRIPTION It will follow the same methodology as outlined in Work Package 4, Task 4.2 for each of the tested

molluscicides, and there will be a control plot. The tested products are: non residual agrochemicals with ovicidal action and commercial molluscicides (Methaldehude, Carabamatos, Ferramol…)

Plant’s crops damage evaluation See Work Package 4, Task 4.2.

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Work Package 8 Field experiments in organic key horticultural crops to evaluate the efficacy of organic Molluscicides-

ovicides and the use of plant-traps as pests control method for land snails. PARTICIPANTS Participants 1 (……. Man-month) Participants 2 (……. Man-month) Participants 3 (……. Man-month) Participants 4 (……. Man-month) Participants 5 (……. Man-month) Participants 6 (……. Man-month) Participants 7 (……. Man-month) Participants 8 (……. Man-month) Participants 9 (……. Man-month) OBJETIVES Field experiments in organic key horticultural crops to evaluate the efficacy of selected compounds with

ovicidal activity against land snail and egg-lays. WORK AND METHODOLOGY DESCRIPTION See Work Package 7; only change the kind of compound tested and the crop type.

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Work Package 9 PARTICIPANTS: only Americans participants Participants 1 (……. Man-month) Participants 2 (……. Man-month) Participants 3 (……. Man-month) Participants 4 (……. Man-month) Participants 5 (……. Man-month) Participants 6 (……. Man-month) Participants 7 (……. Man-month) Participants 8 (……. Man-month) Participants 9 (……. Man-month) OBJETIVES To identify improved strains of Phasmarhabditis nematodes which are more effective biocontrol agents of

larger land snail species in Hispano-America. BACKGROUND The use of the rhabtitid nematode Phasmarhabditis hermaphrodita (Schneider) as a biological control agent

for land snails has been proposed (Wilson, Glen & George, 1993) and a commercial product based on P.

hermaphrodita (Nemaslug, MicroBio Ltd, UK) was launched for sale in the UK in spring 1994, and later in other European countries. Phasmarhabditis hermaphrodita has been tested successfully for biocontrol of land snails in a number of field trials, including a range of arable and horticultural crops (Wilson et al., 1994a, 1996; Wilson, Glen, Wiltshire & George, 1994b; Wilson, Glen, George & Hughes 1995a; Wilson, Hughes & Glen, 1995b; Ester & Geelen, 1996; Glen et al., 1996; Iglesias, Castillejo & Castro, 2001). In some experiments, however, nematode application did not reduce land snail damage (Wilson et al., 1995a, 1996; Speiser & Andermatt, 1996).

The strain of the nematode currently used as a biocontrol agent is well adapted to the relatively low temperatures at which land snails are troublesome as pests in north west Europe (Glen et al.,1994a).Thus, this strain is likely to be suitable for biocontrol of land snails in vegetable and fruit crops in northern Europe. However, this strain may not be suitable for use in warmer regions of Latino America because it is unable to survive for more than a few hours at temperatures greater than 25"C (Wilson et al., 1993c). It is likely that strains better adapted to warmer regions of Latin America can be found, because P. hermaphrodita and two closely related species from the same genus have been recorded from southern Europe (Morand, 1988).

However due to the high costs in the treatment with nematodes nowadays, their use is still restricted to high value crops such as ornamental plants and some vegetables.

IMPORTANCE: to be able to control land snail pests with nematode zooparasite and also effective on tropical crops, it will be very beneficial to organic farming.

ASSESSMENT OF PROGRESS AND RESULTS: Through semi-annual and annual reports and regular personal controls carried out by the coordinator to each of the participants in the concerned country.

WORK AND METHODOLOGY DESCRIPTION STANDARD OPERATING PROCEDURE FOR PROJECT PARTICIPANTS ISOLATING POSSIBLE NEMATODE LAND

SNAILS PARASITES 1. All land snail and soil samples collected for the identifying purpose of possible new nematode parasites

will be labeled with the following essential information, which it will be provided for every new strain of nematodes found.

* Nearest place name, nearest main town, and Country * Latitude and longitude * Land snail species (if sample is from a land snail) * Number of each species (if sample is from more than one land snail) * Soil type

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* Crop or habitat * Recollection Date * Recollection method * Recent and current weather * Number of subcultures (if any) since nematodes were isolated from infective land snails 2. At all times, land snails suspected of being parasited with nematodes (swollen mantle symptom) should

be kept in individual Petri dishes lined with moist filter paper at below 20 ºC (recommended 15 ºC). Land snails should be kept like this until nematodes have reproduced within/on the cadaver.

3. Once nematodes are seen crawling on the land snail cadaver (this is usually about 7 days after infection;

at this point adult and some juvenile nematode stages should be present), the land snail re++++++++mains should be transferred onto either an extraction tray or a modified White trap and kept at the same temperature as above for several days.

Extraction tray method (also known as modified Baermann tray or Whitehead tray) (in EU first 6-month

report, previously referred to as double tray/milk filter technique): This method is frequently used for extracting plant-parasitic nematodes from root material Whitehead and

Hemming, 1965; Southey, 1986). It consists of a coarse nylon sieve (1 mm gauze) which has supports fixed to the bottom lifting it about 1 cm up. It is lined with a single layer of tissue paper or milk filter to prevent waste material getting through. The sieve is then placed in a tray with water just reaching the bottom of the sieve. Infected land snails can be placed on the sieve. Nematode stages will migrate out of the land snail cadaver into the Water, with the filter preventing unwanted land snail remains contaminating the water. Trays should be left for several days allowing all nematodes to emerge from the land snail remains. Nematodes collected in the water can be harvested every other day or so, with the tray being refilled with fresh water.

White trap: The modified White trap it is a standard way of extracting and collecting entomopathogenic

nematodes from infected insect larvae (White, 1927; Woodring and Kaya, 1988). It is probably also a better method of collecting the infective juveniles of land snail-parasitic nematodes than using extraction trays. It consists of a small container (9 cm diameter x 5 cm deep) in which an inverted 5 cm diameter, 2 cm deep Petri dish bottom is placed. A shallow layer of water (1 cm deep) is added to the container. A filter paper is then draped on the Petri dish platform so that it comes into contact with the water. Infected land snails are then placed onto the moist filter paper on the platform. Once the nematode has completed its life cycle, infective juveniles will start to emerge from the land snail remains and migrate across the moist filter paper into the water after about 14 days (or longer depending on nematode species, level of infection and temperature). Nematodes can be harvested every other day or so, until nematodes are no longer found in the water. The container should be refilled with fresh water after every harvest.

Ideally, only one infected land snail should be placed in each extraction tray or White trap. If this is

impossible (e.g. because large numbers of infected land snails have been collected at the same time), then each tray should contain land snails of one species, collected from the same site at the same time.

4. Infective juveniles collected should be cleaned before storage. The nematode suspension is poured in a

small (100 ml) beaker and left undisturbed for about 30 min, allowing nematodes to settle onto the bottom. The supernatant is then carefully decanted, leaving behind a concentrated nematode suspension. The beaker should then be refilled with fresh well-oxygenated tap water (or preferably sterile distilled water) and the processes are repeated 3 to 4 times or until the suspension appears clear. Nematodes can then be stored in shallow containers with breathing holes, the suspension being only approximately 0.5 cm in depth. Storing nematodes in a large surface-to-volume ratio ensures sufficient oxygen availability. The suspension should ideally have a concentration of no more than approximately 5000/ml and be stored at 5 ºC. Nematode viability should be checked regularly (after room temperature acclimatization). Nematodes should be hatched again after several months or when a large proportion of nematodes start to die.

5. A live sample of each isolated nematode will be sent by Courier, to Participant 1 for specific

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identification. Nematodes will be sent as infective juveniles in tap water in a part-filled container, with a large ratio of air to water (20 volumes of air to 1 volume of water) for culturing and confirmation of species identity.

Gantt Chart showing the timing of the different WPs and their components.

Work Packge Nº

Task Nº

Work Packages Title

Participant No.

Participant organism

name

First Year Second Year Third Year

WP.1

Land Snail Biological Cycle. the size, structure and dynamics of their populations

All partners 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12

WP.2.

Land Snails diet composition. Trap plants All partners 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12

WP.3.

Statistical Model to predict land snails activity All partners 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12

WP.4.

Task 4.1.

Bio pesticides, Bio Molluscicides, Bio Ovicides Plants Extracts. Laboratory test on paper filter Standard soil

All partners 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12

Task 4.2.

Mini plots tests on horticultural soil All partners 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12

Task 4.3.

Collateral effects on soil fauna All partners 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12

Task 4.4.

Chemicals analysis to search the active principle of plant extracts with mulliscicide and ovicide activity

Sygenta Company and USC (Spain)

1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12

WP.5.

Task 5.1.

Laboratory test to find non residual agrochemicals with ovicidal activity

All partners 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12

Task 5.2.

Field experiments on conventionally horticultural crops to evaluate the efficacy of selected agrochemical as molluscicide-ovicide

All partners 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12

Task 5.3.

Field trials to evaluate the collateral effects on soil Fauna and border effect on wild land snails of ovicide agrochemicals.

All partners 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12

WP.6.

Task 6.1.

Manure Laboratory test on land snail eggs for accurate Ovicidal concentration

All partners 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12

Task 6.2.

Field experiments to evaluate the efficacy of cow and pig manure as slug eggs control for key organic horticultural crops

All partners 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12

Task 6.3.

Field analysis to investigate the collateral effect on soil fauna and border effect on wild land snails of cow and pig manure

All partners 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12

Task 6.4

Field experiments to use tramp-plants as deterrent method to protect organic horticultural crops alone and in combination of cow and pig manure

All partners 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12

WP. 7

Field experiments in conventionally crops to evaluate the efficacy of ovicide agrochemicals alone and in combination with other commercial molluscicides

All partners 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12

WP. 8

Field experiments in organics horticultural crops to evaluate the efficacy of organic molluscicides-ovicides and the use of plant-traps

All partners 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12

WP. 9 To identify improved strains of Phasmarhabditis nematodes which are more effective biocontrol agents of larger slug species in Hispano-America.

Only Latino America Parteners

1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12

1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12

1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12 1 2 3 4 5 6 7 8 9 10 11 12

WP 1 Manager: Participant 1

Participants 1, 2 ,3, 4, 5, 6, 7, 8

WP ???? Manager: Participant 1

Participants 1, 2 ,3, 4, 5, 6, 7, 8

WP 7 Manager: Participant 1

Participants 1,2,3,4, 5, 6, 7, 8

WP 6 Manager: Participant 1

Participants 1, 2 ,3, 4, 5, 6, 7, 8

WP 5 Manager: Participant 1

Participants 1, 2 ,3, 4, 5, 6, 7. 8

WP 3 Manager: Participant 1

Participants 1, 2 ,3, 4, 5, 6, 7, 8

WP 4 Manager: Participant 1

Participants 1, 2 ,3, 4, 5, 6, 7, 8

WP 2 Manager: Participant 1

Participants 1, 2 ,3, 4, 5, 6, 7, 8

DELIVEREBLES

•Statistical model to predic activity • Molluscicides ovicicides • Nematode zooparasitic

• Integrate package for organic crops • Integrate package for convencional crops

COORDINATION Participant 1

PROJECT MANAGEMENT STRUCTURE

OJO. Coordinador de cada WP

WP 1 LAND SNAIL’S BIOLOGICAL

CYCLE

Participant 1, 2 ,3, 4, 5, 6, 7, 8

WP 9 NEMATODO ZOOPARASITIC

Phasmarhadities

Participant: Latino America

WP 7 & 8 FIED EXPERIMENTS CONVENTIONALLY, ORGANICS CROPS

Participant 4, 5, 6, 7, 8

WP 6 PIG AND COW MANURE AS BIO OVICIDES TESTS

Participant 1, 2 ,3, 4, 5, 6, 7,

8

WP 5 BIO OVICIDAS

AGROCHEMICALS TESTS

Participant 1, 2 ,3, 4, 5, 6, 7. 8

WP 3 STATISTICAL ACTIVITY

MODEL

Participant 1, 2 ,3, 4, 5, 6, 7, 8

WP 4 BIOMOLLUSCICIDES TESTS

COLLATERAL EFFECTS

Participant 1, 2 ,3, 4, 5, 6, 7, 8

WP 2 DIET COMPOSITION

TRAP PLANTS

Participant 1, 2 ,3, 4, 5, 6, 7, 8

• Statistical model to predic activity

• Molluscicides ovicicides • Nematode zooparasitic

• Integrate package for organic crops • Integrate package for convencional crops

RELATIONSHIP TASKS and ParticipantS

WP 1 LAND SNAIL’S BIOLOGICAL

CYCLE

Participant 1, 2 ,3, 4, 5, 6, 7, 8

WP 9 NEMATODO ZOOPARASITIC

Phasmarhadities

Participant: Latino America

WP 7 & 8 FIED EXPERIMENTS CONVENTIONALLY, ORGANICS CROPS

Participant 4, 5, 6, 7, 8

WP 6 PIG AND COW MANURE AS BIO OVICIDES TESTS

Participant 1, 2 ,3, 4, 5, 6, 7,

8

WP 5 BIO OVICIDAS

AGROCHEMICALS TESTS

Participant 1, 2 ,3, 4, 5, 6, 7. 8

WP 3 STATISTICAL ACTIVITY

MODEL

Participant 1, 2 ,3, 4, 5, 6, 7, 8

WP 4 BIOMOLLUSCICIDES TESTS

COLLATERAL EFFECTS

Participant 1, 2 ,3, 4, 5, 6, 7, 8

WP 2 DIET COMPOSITION

TRAP PLANTS

Participant 1, 2 ,3, 4, 5, 6, 7, 8

RELATIONSHIP TASKS and Participants

intercommunication

Table 2 Schedule of meeting to be held and reports to be produced

MEETING Due Date

Initial planning meeting, Santiago de Compostela, Spain December 2010

(at the beginning of Project)

Progress review & planning meeting, Norwich

December 2011

Progress review & planning meeting, Lithuania

December 2012

Final review meeting, Argentina, Ireland

December 2013

MEANS OF COORDINATOR VERIFICATION

First travel to verification, Europe/Hispano America

May 2011

Second travel to verification, Europe/Hispano America

October 2011

Third travel to verification, Europe/Hispano America

June 2012

SIX MONTH PROGRESS REPORT

First Six-month Progress Report

June 2011

Second Six-month Progress Report

June 2012

Third Six-month Progress Report

June 2013

ANNUAL PROGRESS REPORT

First Annual Progress Report

December 2012

Second Annual Progress Report

December 2013

Third Annual Progress Report

December 2014

FINAL REPORT March 2014

Brochure(s) and Web page for extension services, consultants and/or growers March 2014

Table 1.3a: Work Package List showing Person-months

Work Packge Nº

Task Nº

Work Packages Title Type activity

Lead participant nº

Lead Participant short name

Total Person-months

Start month End month

WP.1

Land Snail Biological Cycle. the size, structure and dynamics of their populations

RTD 1 USC 34.2 1 24

WP.2.

Land Snails diet composition. Trap plants

RTD 1 USC 41.3 1 24

WP.3.

Statistical Model to predict land snails activity

RTD 1 USC 41.3 13 36

WP.4.

Task 4.1. Bio pesticides, Bio Molluscicides, Bio Ovicides Plants Extracts. Laboratory test on paper filter Standard soil

RTD 1 USC 37.7 1 30

Task 4.2.

Mini plots tests on horticultural soil

RTD 1 USC 27.1 13 36

Task 4.3.

Collateral effects on soil fauna

RTD 1 USC 25.3 19 36

Task 4.4.

Chemicals analysis to search the active principle of plant extracts with mulliscicide and ovicide activity RTD 1 USC 25.3 19 36

WP.5.

Task 5.1.

Laboratory test to find non residual agrochemicals with ovicidal activity RTD 1 USC 34.2 1 24

Task 5.2.

Field experiments on conventionally horticultural crops to evaluate the efficacy of selected agrochemical as molluscicide-ovicide

RTD 1 USC 30.6 13 30

Task 5.3.

Field trials to evaluate the collateral effects on soil Fauna and border effect on wild land snails of ovicide agrochemicals.

RTD 1 USC 9.6

19 36

WP.6.

Task 6.1.

Manure Laboratory test on land snail eggs for accurate Ovicidal concentration RTD 1 USC 8.3 1 18

Task 6.2.

Field experiments to evaluate the efficacy of cow and pig manure as slug eggs control for key organic horticultural crops

RTD 1 USC 10.1 7 30

Task 6.3. Field analysis to investigate the collateral effect on soil fauna and border effect on wild land snails of cow and pig manure

RTD 1 USC 9.6 19 36

Task 6.4

Field experiments to use tramp-plants as deterrent method to protect organic horticultural crops alone and in combination of cow and pig manure

RTD 1 USC 6,7 13 36

WP. 7

Field experiments in conventionally crops to evaluate the efficacy of ovicide agrochemicals alone and in combination with other commercial molluscicides

RTD 1 USC 44.2 13 36

WP. 8

Field experiments in organics horticultural crops to evaluate the efficacy of organic molluscicides-ovicides and the use of plant-traps

RTD 1 USC 44.2 13 36

WP. 9 To identify improved strains of Phasmarhabditis nematodes which are more effective biocontrol agents of larger slug species in Hispano-America.

RTD 1 USC 30.3 1 36

TOTAL 507.7 p/m

Table 1.3 e: SUMMARY OF STAFF EFFORT

Participant no./short name

WP 1 WP 2 WP 3 WP 4 WP 5 WP 6 WP 7 WP 8 WP 9 Total person

month

No. 1 USC-ES

No. 2

No. 3

No. 4

No. 5

No. 6

No. 7

No. 8

No. 9

No. 10

No. 11

No. 12

Milestones The project milestones for the nine individual work tasks are shown in Table 1.3 c Deliverables 1. Contract deliverables At the end of each year a progress report will be produced by each of the individual participants in which research methods and results achieved will be included and related to the project milestones. After 18 months (half the duration of the project) a project mid-term review report will also be produced following a mid-term review meeting. This mid-term report will include input from all partners and will summarise overall progress and needs for the remainder of the project duration. After three years (end of project) a final report will be produced. 2. Technical deliverables (a) The introduction into vegetable and fruit crops of a novel strategy for land snails control, combined where appropriate with novel low-toxicity agrochemicals with bio-ovividal activity, as a replacement for current chemical which give inadequate control, are hazardous to pets, wildlife and beneficial invertebrates and cause concerns because of residues in food crops. (b) Novel methods of bio-molluscicides and bio-ovicides control of slugs in vegetable crops. (c) Effective integrated packages of crop management, biopestices methods and, where appropriate, low-chemical methods of protecting vegetable crops from slug damage. Each year, technologies developed in this project will be transferred via extension services and consultants, to vegetable growers. Further development of results will be undertaken by Partner 2 (a SME), so as to introduce the novel bio-mollusicicide agent into the vegetable growing sector of the European horticultural industry. For plan strects, official registration of the product will be required and appropriate industrial partners will be involved to do this. At the end of the project results will be further developed to provide practical advice for vegetable growers. Table 1.3 c: Annual milestones for the 9 Work Packages proposed.

Milestone number

Milestone name Work Package(s) involved

Espected Data

Means of verification

Dinámica, estructura y tamaño de la población of land snail pest. First year on field sampling

WP. 1 12 Field survey complete and report

Complete field study of land snails populations to establish its dynamic, structure and size.

WP.1 24 Field survey complete and report

Establish best methods to find out the land snails fife cycle.

WP. 1 12 Field survey complete and report

Life cycle, complete field observation and sampling

Wp.1 24 Field survey complete and report

Establish best methods to find out the land snails diet.

WP. 2 12 Field survey complete and report

Complete field study of land snails diet. WP.2 24 Field survey complete and report

First draft of the Statistical Model to predict the land snails activity

WP. 3 12 Complete laboratory processing and report

Second draft of the Statistical Model to predict the land snails activity

WP. 3 24 Complete laboratory processing and report

Definitive Statistical Model to predict the land snails activity in every zone studied

WP. 3 36 Report

Select potential plants for extracting compound as bio-molluscicide

WP.4 12 Laboratory survey complete and report

Select better compound for direct contact on filter paper. Results of laboratory tests

WP.4 12 Laboratory survey complete and report

Select better compound for mini plots testing and better methods of application.

WP.4 24 Field survey complete and report

Field test of best compounds for determinate the effect on non-target soil animals

WP.4 24 Field survey complete and report

Select better agrochemical based on laboratory tests WP. 5 12 Laboratory survey complete and report

Complete small-scale field trials on agrochemicals WP. 5 24 Field survey complete and report

Select cow and pig manure concentration based on laboratory tests

WP. 6 12 Laboratory survey complete and report

Complete small-scale field trials on manure WP. 6 24 Field survey complete and report

Parcial estudio de la actividad de los gasterópodos terrestres plaga. Refine use of best methods.

WP. 3 12 Field survey complete and report

Complete second year of activity WP. 3 24 Field survey complete and report

Complete third year of activity, definitive model WP. 4 36 Field survey complete and report

Evaluar la eficacia de los ovicidas en conventionally crops Complete small-scale field trials

WP. 5 24 Field survey complete and report

Evaluar la eficacia de los purines como ovicidas en organic crops. Complete small-scale field trials

WP. 6 24 Field survey complete and report

Conocer los efectos colaterales de los molusquicidas ovicidas, partial trials.

WP. 5 &6 24 Field survey complete and report

Partial and Complete field test on side effects. Investigate effects on no-target soil fauna.

WP. 5&6 24 and 36 Field survey complete and report

Comprobar la capacidad de las plantas trampa para proteger los cultivos

WP. 7 24 Field survey complete and report

Comparar la eficacia en conventionally crops de los molusquicidas ovicidas frente a los molusquicidas tradionales. Determine best way to combine with others molluscicides.

WP. 7 24 Field survey complete and report

Comparar la eficacia de los ovicidas orgánicos frente a controles biológicos. Determine best way to combine with others molluscicides.

WP. 8 24 Field survey complete and report

Búsqueda de un nematodo Phasmarhadities autóctono para usarlo en el control biológico. Select the best methods based on laboratory tests.

WP. 9 12, 24 and 36

Field survey complete and report

Devise integrated packages for further development on conventional crops

WP. 1,2,3, 4,5,6,7 y 8

36 Report

Devise integrated packages for further development on organic crops

WP. 1,2,3, 4,5,6,7 y 8

36 Report

Table 1.3 b: Deliverables List

Del.nº Deliverable name WP nº Nature Dissemination level

Deliverable date.

Months

1.1

Conocer el ciclo biológico de las especies plaga en función de la fenología del cultivo y su situación geográfica con el objeto de introducir medidas preventivas

WP. 1 R PU Partial: 12 Final: 24

1.2

Conocer por medio del estudio del tamaño y estructura de la población la magnitud de los daños que las babosas y caracoles causan en los cultivos

WP.1 R PU Partial: 12 Final: 24

1.3

Saber por medio de la dinámica de las poblaciones de caracoles y babosas en que fases de su desarrollo son más dañinos para los cultivos

WP.1 R PU Partial: 12 Final: 24

2.1

Conocer las plantas más apetecidas por los caracoles y babosas por medio del estudio de sus contenidos estomacales.

WP.2 R PU Partial: 12 Final: 24

2.2 Obtener la base científica para desarrollar la estrategia del uso de plantas-trampas para proteger los cultivos

WP.2 R PU Final: 24

3.1

Conocer los periodos de actividad en cada una de las zonas de estudio en función de las condiciones medio ambientales y la fenológicas

WP.3 R PU Partial: 12 Final: 24

3.2

Desarrollar un Modelo Estadístico que nos prediga los periodos de actividad de las especies de gasterópodos terrestres plaga

WP.3 R PU Partial: 24 Final: 36

3.3

Tener información de cuál es el momento más adecuado para aplicar un tipo concreto de molusquicida o emplear una estrategia de control idónea

WP.3 R PU

Partial: 12 Partial: 24 Final: 36

3.4

Proporcionarle al agricultor información para que desarrolle una estrategia preventiva con la que se adelante al ataque de la plaga, use menos molusquicidas y consiga una mayor eficacia

WP.3 R PU Partial: 24 Final: 36

4.1

Conocer plantas de las que se puedan obtener biopesticidas con acción molusquicida y ovicida para que en futuro reemplacen a los molusquicidas tradicionales, o bien con ellas se puedan hacer abonos verdes que puedan ser usados en cultivos ecológicos como controladores de las plagas de caracoles y babosas. (WP. 4)

WP.4

R PU Partial: 24 Final: 36

4.2

Poner en mano de la industria de agroquímicos nuevas vías de control de las plagas de caracoles y babosas por medio de Biomolusquicidas y Bio-ovicidas obtenidos a partir de extractos de plantas. The possibility to introduce into key horticultural crops of a novel plant extract as biocontrol agent for land snails control

WP.4

R PU Partial: 24 Final: 36

4.3

Saber que efectos colaterales puede tener el uso de biomolusquicidas y bio-ovicidas sobre la fauna edáfica y de zonas colindantes (WP.4)

R PU Partial: 24 Final: 36

5.1

Development of novel techniques for low-toxicity chemical control of land snail eggs - the stage in the pest life cycle against which no current controls are available

WP.5 R PU Partial: 24 Final: 36

5.2

Proporcionarle al agricultor información de cómo usar los agroquímicos que habitualmente emplea en los cultivos tradicionales le puedan servir para controlar las plagas de los gasterópodos terrestres

WP.5 R PU Partial: 24 Final: 36

5.3

Proporcionarle información al agricultor de los efectos colaterales que sobre la fauna pueden tener los molusquicidas-ovicidas que se proponen usar en cultivos tradicionales

WP.5 R PU Partial: 24 Final: 36

6.1

Proporcionarle al agricultor ecológico información de cómo puede usar los abonos naturales para controlar las plagas de gasterópodos terrestres

WP.6 R PU Partial: 24 Final: 36

6.2

Indicar al agricultor ecológico que tipos de plantas puede usar para disuadir a los gasterópodos terrestres de que no ataquen a los cultivos

WP.6 R PU Partial: 24 Final: 36

6.3

Saber que efectos colaterales puede tener el uso de abonos orgánicos a concentración ovicida sobre la fauna edáfica y de zonas colindantes

WP.6 R PU Partial: 24 Final: 36

7.1

Proporcionarle al agricultor convencional información sobre la forma de usar los distintos molusquicidas solos o en combinación, y sobre su eficacia de estos dependiendo del tipo de cultivo, del tipo suelo y de las especie de gasterópodo terrestre que son plaga en una zona concreta

WP.7 R PU Partial: 24 Final: 36

8.1

Proporcionarle al agricultor ecológico información sobre la forma de usar los distintos biomolusquicidas solos o en combinación, y sobre su eficacia de estos dependiendo del tipo de cultivo, del tipo suelo y de las especie de gasterópodo terrestre que son plaga en una zona concreta

WP.8 R PU Partial: 24 Final: 36

9.1

Conocer si existen posibles nematodos zooparásitos que se puedan ser usados en el control biológico de las plagas de caracoles y basas en cultivos agrícolas

WP.9 R PU Partial: 24 Final: 36

9.2 The possibility to introduct into key horticultural crops of a novel nematode biocontrol agent for land snails

WP.9 R PU Final: 36

5&6.1

Greater uptake of environmentally favourable integrated crop production systems, as a result of reduced risk of land snail damage

WP.5&6 R PU Final: 36

5&6.2 Improved food safety, resulting from reduced levels of WP.5&6 R PU Final: 36

molluscicide residues in harvested produce

5&6.1

Effective integrated packages of control measures for protecting conventionally grown horticultural crops from land snails damage

WP.5&6 R PU Final: 36

4&5&6.1

Improved environmental safety, resulting from reduced reliance on chemical molluscicides

WP. 4&5&6

R PU Final: 36

6&9.1

Effective integrated packages of control measures for protecting organically grown horticultural crops from land snail damage

WP. 6&9

R PU Final: 36

7&8.1

Technology transfer through meetings of growers, practical demonstrations, extension services, trade shows and articles in the horticultural press

(WP. 7&8

R PU Final: 36

All.1

Brochure with two parts, or separate brochures describing integrated packages of control measures for conventional and organic horticultural producers and Web links for specific advices.

(WP. all) R PU Final: 36

Project: Participant 1 – Universidad de Santiago de Compostela, Spain Table – Estimated Breakdown of the Total Allowable Cost

ALLOWABLE COST AMOUNT IN EUROS LABOUR 237.600

TRAVEL, SUSBSISTENCES AND MEETINGS 94.200

DURABLE EQUIPMENT 116.000

CONSUMABLES 40.000

EXTERNAL ASISTANCE 0.00

COMPUTING & OTHER COST 40.000

OVERHEADS 20% 110.360

TOTAL Requested from EU

638.160

Rounded to

638.000 €

a) A detailed list of personnel to the execution of the research (person-month)

Postdoctoral Research Scientist (Group 1) ………………. 36 person/month Graduate Research Scientist (Group 2) ……………………. 36 person/month

Official Salary Scales (See attached documents)

Group 1, Postdoctoral Research Scientist: 144000 Euros/36 months before tax 108000 Euros/36 months after tax

144000 €

Group 2, Graduate Research Scientist: 93600 Euros/year before tax 72000 Euros/year after tax

93600 €

TOTAL 237600 €

b) Idem for “external assistance”

TOTAL

SHARED COST PROJECT

c) A list of meeting for planning and travel and subsistence

COST Euros

3 Meetings for planning & coordination 18.000 €

Land snails sampling in Spain (see below for sampling) 37.800 €

3 Hispano America/Europe extra trip to verification and adviser 44.400 €

TOTAL 94.200 €

Details of Meeting for planning & coordination (3 staff x 7 days x 3 meetings). Ireland, Norwich, and Lithuania Plane return ticket: 1.000 Euros/person x 3 people x 3 trips= 9.000 € Accommodation Hotel: 100 Euros/nigh- person x 5 night x 3 people x 3 trips = 4.500 € Maintenance: 70 Euros/day- person x 7 days x 3 people x 3 trips = 4.500 € TOTAL: 18.000 Euros Details of land snail sampling in Spain

Sampling for land snail density and activity in crops, monthly: 2011, 2012 y 2013. 12 annual trips, three zones visited from 5 characteristic crops in each zone. Each trip will involve three people for a total 3 days per trip. Year 2011: 12 trips x 3 people x 3 days x (60 € hotel + 50 € subsistence) = 11800 € Year 2012: 12600 € Year 2013: 13400 € TOTAL: 37800 €

Extra Hispano American trip for verification and adviser

3 extra trips to Hispano America/Europe for controlling the development of WP project. Trip for 2 people and 20 days lenth. Return Plane ticket: 4000 €/trip/person = 24000 €/3 return ticket/2 people Hotel: 20 nights x 3 trips x 2 people x 100 € Room = 12000 € Subsistence: 20 days x 3 trips x 2 people x 70 € subsistence = 8400 € TOTAL: 44.400 €

d) A list of durable equipment

Two cameras with inside atmosphere controlled. FITOCLIMA D1200PLH with total inside environment controlled: temperature, humidity, air, light, gases... Model ARALAB D1200PLH , Price/u: 27000 €/unit

54000 €

Inverted Microscopy, low magnification, Led light. Mod. Ix51. Tri-ocular with video camera intake and video recorder systems. 30000 €

Inverted Stereo zoom microscope, with viseLed White LED illumination system Digital Camera

10000 €

Digital System to Store activity dates, images and habitat variables 9000 €

Five Digitals AXIS IP cameras to control activity of field land snails with remote control and Wi-Fi connections

13000 €

TOTAL 116.000 €

e) Consumables. Details of others cost

Computing 4000 €

Field sampling facilities 6000 €

Field experiments 10000 €

Laboratory test facilities 10000 €

Laboratory test consumables 10000 €

TOTAL 40.000 €