Implementation of a Microbial Pest Control Product in an Integrated Pest Management Programme

Abstract Key factors to cost-effective implementation of a microbial pest control product are application strategy, compatibility, and knowledge transfer to the user. This requires the design of a comprehensive integrated pest management programme in which the microbial pest control product is to be incorporated. The first element is an optimal application strategy per targeted pest and cropping system. Determinative parameters such as dosage and spray volume are discussed as well as factors that influence these parameters such as the tritrophic plant-host-pathogen relationship. The second element is the incorporation of the microbial pest control product in an IPM system. Compatibility with chemical pesticides and with beneficial organisms must be established and critical aspects are mentioned for each type of pathogen. Microbial pest control products and natural enemies are often simultaneously used in IPM systems to enhance control. Synergistic, additive and negative interactions are reviewed. Combinations of microbial pest control products and combinations with other means are discussed. The risk of resistance will be briefly treated. The third element is the implementation of a microbial pest control product into practice. Adoption of new products needs knowledge transfer in the entire supply chain to the grower, and pivotal considerations including economic aspects will be reviewed. Requirements and recommendations will be provided on how to accomplish a successful implementation. Considerations will be presented with regard to the compatibility profile, the advantages and the disadvantages of biopesticides, the cost of the implementation process, and to the potential of combined use of biopesticides.

1 Introduction

In greenhouse crops, microbial pesticides are generally used as part of an integrated pest management programme (IPM). They are mainly used as an inundative release strategy from which direct effects are expected resulting in a considerable decrease of the pest population within a relatively short period. In greenhouse vegetable crops, the main biocontrol components of the IPM system are natural enemies. Microbial pest control products (MPCPs) are often used as selective and corrective measures, or as additional pest control tools. The same can be seen in ornamental

W.J. Ravensberg, A Roadmap to the Successful Development and Commercialization 235

of Microbial Pest Control Products for Control of Arthropods, Progress in Biological Control 10, DOI 10.1007/978-94-007-0437-4_6, © Springer Science+Business Media B.V. 2011

crops such as roses and chrysanthemum, but in other ornamental crops they are used in a more chemical oriented programme. In such a case, they are used as a stand-alone product against a particular pest, or as an additional control product. MPCPs can be used to control key pests, often in combination with other methods, or they can be used against secondary or minor pests. As justly stated by Hofstein and Fridlender (1994): "Design of a comprehensive disease control program is the main challenge of those who intend to introduce biological pesticides as an attractive alternative or addition to existing control programs". The same is of course true for a pest control programme. The concept of an IPM programme is to have an optimum exploitation of all the pest control options with a strong emphasis on preventative measures and biological control methods. This implies a good understanding of all components and their interactions, in any given cropping system.

The concept of integration of pesticides in IPM programmes in greenhouses has been reviewed by Blumel et al. (1999). They described the importance of selective pesticides, and sequential test methods on detecting side-effects on natural enemies. Furthermore, approaches were suggested on pesticide application methods and on how to minimize negative effects on beneficials. Little attention, however, was given to microbial pesticides. An introduction on compatibility of pesticides, including MPCPs, with biological control agents is given by Hassan and Van de Veire (2004). They briefly reviewed the use of pesticides, including microbials, in IPM systems in greenhouse crops.

Given the entomopathogenic character of MPCPs and their use together with beneficial arthropods, as well as the use with other pesticides, a "two-sided" compatibility profile of a MPCP with these other crop protection components must be established. This is indispensable for successful implementation and commercialization of a MPCP. In the literature compatibility of entomopathogens with other methods of pest control has been studied for many cases. This kind of research is usually aimed at investigating whether there are any adverse effects of chemical pesticides on MPCPs, or whether MPCPs have any adverse effects on natural enemies and pollinators. The latter is sometimes called biocompatibility. An early overview of research results of entomopathogenic bacteria, fungi, viruses, nema-todes and protozoa and compatibility with chemicals and beneficial arthropods is given by Jacques and Morris (1981). They also reviewed the combined use of entomopathogens with pesticides. Examples were given of enhanced as well as of reduced effects. The option to combine a MPCP with other methods may give improved control through additive or synergistic effects. Compatibility studies and combination effects have been investigated by many scientists with all four types of pathogens treated in this book. It is not possible to review all these studies. I will highlight the most significant publications below with the focus on greenhouse crops.

The efficacy of entomopathogens depends on a number of factors such as environmental and agronomic factors. These relationships need to be studied. Some of the factors can be influenced so that entomopathogens can be used in a more effective way. Inglis et al. (2001) reviewed the importance of temperature, humidity, sunlight, soil type, rainfall, and interactions among these environmental factors, and their influence on the activity of fungal entomopathogens. Understanding these influences is vital for developing an effective IPM programme in various cropping systems. The authors illustrated this by examples from glasshouse crops (aphids) and field crops (Colorado potato beetle, European corn borer and whiteflies). Compatibility with chemicals as well as with beneficials is reviewed, and innovative strategies such as using semio-chemicals and trap crops, were discussed. These factors are also relevant for entomopathogenic bacteria, nematodes and baculoviruses, and vary per organism, target pest and cropping system.

In the development of a MPCP, following product development, the first step is to establish the optimal application strategy under commercial conditions. The product should be used in such a way that its pest control features are fully exploited within the given situation. The application strategy needs to take into account factors such as the delivery technology, the biology of the pest, the crop and how it is cultivated, and the environmental conditions. Each factor likely to influence the entomopathogens efficacy needs to be considered and studied, if deemed critical, in the application strategy. Secondly, the use of the MPCP must be implemented in the entire pest and disease control programme. An integrated programme needs to be established, with optimal effects on the pests and minimal harmful effects to all the biological pest control agents. These two aspects are the cornerstones of a successful implementation of a new MPCP in commercial horticultural crops. I will discuss how to consider these pivotal elements in the development and integration of a MPCP in an IPM system.

Furthermore, I will discuss other relevant aspects related to this topic such as the chance of resistance with pathogens, practical aspects of testing compatibility, the transfer of knowledge to the end-users, and the cost considerations concerning implementation of a MPCP within an IPM programme. Finally, I will provide recommendations and conclusions on the critical steps in the implementation of a MPCP in a multi-component pest and disease control system, and on the potential of combinations of control agents to enhance biological pest control.

2 Development of an Application Strategy

In order to achieve effective results with a MPCP against a certain pest, an optimal application strategy needs to be developed. For a "broad-spectrum" MPCP, this may differ per target species. First of all, the level of the pest determines when and how to intervene. Vital factors therein are the application method, the frequency of applications, intervals between applications, the optimal dosage, the spray volume used, and appropriate equipment. Furthermore, the strategy is strongly influenced by the crop, the cultivation methods, the type of greenhouse, the climate, etc. (Fig. 6.1). These parameters influence the target pest as well as the pathogen. The plant-host-pathogen is a tritrophic system that is influenced by a multitude of factors which results in complex study systems. To investigate all these relationships is impossible and it is obvious that the most influential factors should be determined and studied where they are expected to have a relevant effect on the performance of the

Fig. 6.1 Interactions that need to be considered in the development of an application strategy and in the implementation of a microbial insecticide in an integrated pest management system

biopesticide. In the literature, only few examples have been presented where the authors have investigated and established a complete and effective application strategy for a pathogen. Depending on the pathogen and its biology, particular aspects have to be considered, and I will discuss them below.

Factors influencing activity and persistence of Bt and its toxins are reviewed by Glare and O'Callaghan (2000). The fate of Bt is only known in general terms. Persistence can be years in soil, but only days in the phyllosphere. Little is known from greenhouse environments. Ingestion of Bt by the pest organism usually follows application quickly and therefore environmental factors influencing survival of Bt are less critical as with fungi. Generally, insects become less susceptible to Bt as larvae become older. The same is known for entomopathogenic fungi and bac-uloviruses. If possible, this should be taken into account with regard to timing of the treatments. Examples of greenhouse application strategies with Bt are hard to find in literature, most likely because Bt behaves almost like a chemical and is less influenced by crop and environmental parameters.

Entomopathogenic fungi may pose the greatest challenge of the pathogens treated in this book due to their mode of action. This is exemplified by a thorough study of application parameters on efficacy of Beauveria bassiana for control of Frankliniella occidentalis (Ugine et al. 2007a). The objective was to develop a spray application guideline for maximum efficacy of B. bassiana in a greenhouse ornamental crop. They investigated the application rate, the interval between applications, the water volume and the timing of the sprays during the full growing cycle of impatiens. Thrips population reduction was the highest when plants were sprayed multiple times with 5-day intervals with high rate and high volume applications. Early treatments in the plant cycle gave larger suppressive effects than later treatments. But Ugine et al. (2007a) also found great variation in the population reduction between spray programmes. This extensive study illustrates the complexity of establishing an effective spray programme and the variations that may occur. Another example is the use of the entomopathogenic fungus M. anisopliae for thrips control in plant-growing media in combination with chemicals (Ansari et al. 2007). The fungus proved to be a robust biocontrol agent giving good control independent of the growing media, the method of application (pre-mixing resp. drenching) and whether used alone or in combination with insecticides. Earlier work had shown similar results against the black vine weevil (Bruck and Donahue 2007; Shah et al. 2007a).

The use of Spodoptera exigua NPV in ornamental crops and tomato was investigated by Smits et al. (1987). Reduction of feeding damage was important in ornamental crops and virus application timing should be directed for young caterpillars as early as possible. Sprays should be directed to the underside of the leaves since early instars mainly feed there. This is difficult in dense crop canopies as with chrysanthemum and gerbera, but feasible in tomato. The authors also recommended using pheromones for monitoring the population and optimal timing of sprays.

Nematodes are very different from traditional pesticides as well as from micro-bial pesticides due to their size and active behaviour. It is essential to understand the attributes and the limitations of these biocontrol agents. An overview of the crucial application factors is given by Koppenhofer (2007), Wright et al. (2005) and Shapiro-Ilan et al. (2006). An example of developing an effective application strategy is given by Ehlers (2003a) for control of black vine weevil in strawberry. Their study showed that application through the drip-irrigation system did not give an even distribution to all plants. Instead, a dipping method with young plants was developed and this proved successful for the control of black vine weevil. Other interesting examples of the development of an application strategy are the use of nematodes with insecticides to control leafminers in lettuce and Chinese cabbage (Head et al. 2003) and to control fungus gnats with nematodes in poinsettia and impatiens, grown in various potting media (Jagdale et al. 2004). Application strategies with nematodes for glasshouse pests are reviewed by Tomalak et al. (2005).

Above examples illustrate the complexity of an application strategy and which factors need to be taken into account when an optimal application strategy is being developed. I will discuss some of these parameters more in depth.

2.1 Delivery Methods for Microbial Pest Control Products

Efficient delivery of the infective propagules is crucial for a good efficacy. A number of authors have addressed the issue of optimal application of biopesticides (Evans 1999; Matthews 2000; Bateman et al. 2007; Chapple et al. 2007). Because of the particulate and living nature of biopesticides, they pose a very different challenge to efficient delivery through appropriate equipment than chemicals. The available delivery system, however, is usually the existing equipment developed for chemicals. MPCPs should be capable of application through standard available equipment with minimal special requirements. Growers will not readily change or buy equipment just to apply MPCPs. Nor will they easily accept a very different spray regime or more frequent applications than is normal practice. Only minor adjustments are possible and acceptable for growers. As an example, pre-treatment soaking of Mycotal already turned out to be hampering adoption of this product (see Chapter 3 under Section 5.2). Taking out sieves when nematodes are applied or changing nozzle types for products with small particles are about the maximum accepted modifications.

In protected crops, the most used type of technology for foliar treatment is the high pressure hydraulic nozzle system. This ranges from knapsack sprayers to handgun sprayers, up to automatic spraying boom systems. To some extent other equipment is used such as air-blast sprayers, hot and cold foggers, and low volume misting systems (LVM). Biopesticides are generally applied by means of the hydraulic nozzle system. With these systems good coverage can be obtained, and nozzle position can be adjusted to cover the upper side or underside of leaves or even both. Foggers and LVMs are sometimes used, but the deposit hardly reaches the underside of the leaves, and for many pests coverage is inappropriate then and good control is not achieved. Spinning discs are not used in protected crops since it is not possible to penetrate into the dense canopy with this technique. The methods used to deliver biopesticides have been reviewed by Gan-Mor and Matthews (2003), and they argued that often the method of application has not been adequately considered. They recommended more research in the area of formulation and engineering, but the challenge remains getting farmers to buy and use new technology. On the other hand, I have seen growers constructing their own spraying devices or dripping application systems when they are convinced that this will bring a solution to their problems. It depends on their motivation and the results they can achieve. In general, however, one cannot count on this from the outset of the development of a new product or a new programme.

The application of nematodes differs from the application of bacteria, fungi and viruses because they are much bigger. For soil applications, coarse droplets can be used and this has been working well. Critical aspects, also for the microorganisms, but even more for nematodes, are sedimentation in the tank, pump pressure and the size of sieves and nozzle openings. Many studies have highlighted the critical factors of the application technology used for nematodes (Nilsson and Gripwall 1999; Fife et al. 2003; Wright et al. 2005; Laczynski et al. 2006, 2007) and alternatives as spinning discs have been suggested for foliar treatments (Mason et al. 1999). Optimizing the parameters for application of EPNs is still a topic for research after 30 years of using nematodes for pest control (Brusselman et al. 2007).

2.2 Applications and Timing, Frequency, Intervals and Combinations

The timing of an application is essential in respect to controlling the pest and/or keeping the damage acceptable. The biology of the pest, the level of the pest population and the mode of action of the pathogen must be considered, and thus well known. Often, there is a difference in susceptibility between the life stages of a pest and the application should be targeted at the most susceptible life stage. This is possible when the insect population shows distinctive generations, but generally generations start to overlap later in the growing season and then this becomes increasingly difficult. In greenhouse crops with favourable conditions for a pest with a short life cycle, it takes repeated treatments before the pest population starts to decrease considerably. This has been demonstrated with whitefly and thrips (Ravensberg et al. 1990). Intervals between treatments and the frequency are determining factors then. Determination of the time of spraying is very important in MPCPs. The time of the application during the day can be determinative for the result of an application when environmental conditions influence the survival and activity of the pathogen. To avoid high levels of UV, high temperatures or low humidity, it is often advised to spray pathogens early in the day or late in the afternoon. It has even been suggested to spray shortly before shipping plants to customers in order to control insect pests (Osborne et al. 2008). Conditions during shipping allow the fungus Isaria fumosorosea (formerly Paecilomyces fumosoroseus) to infect and kill larval stages of Bemisia tabaci in poinsettia.

Combined treatments are often carried out by growers for reasons of convenience and cost savings. Most of the time, these combinations (tank-mixes) consist of two chemical pesticides and are targeted at different problems, like aphids and powdery mildew, or caterpillars and whitefly. Tank-mixes are not often carried out for the control of a single pest. Combined treatments including a MPCP, or of two MPCPs, has not been common practice, at least not in the Netherlands. Alternate use of products with different mode of actions to control a pest is more commonly applied; including MPCPs. Resistance management is one of the reasons for this. It can also be that older larval stages or adult stages of the pest require a different product. When a microbial pesticide is used in a tank-mix or in an alternating scheme, the compatibility must be known, either of direct effects or of residual effects. The number of combinations is endless and in order to achieve the optimal effect of an ento-mopathogenic product in an integrated programme, it requires a lot of knowledge on many interactions. This is discussed below. Further to using a biopesticide as a blanket treatment, biopesticides can also be used as a spot treatment, or as a band spraying (top or lower bands), creating separation in location and making it compatible in this way. For instance, a product may be sprayed on the top of the plants only, leaving natural enemies lower in the crop unharmed.

2.3 Environmental Effects

Abiotic factors are salient parameters that can influence the results of MPCPs. The most important one is temperature; others are relative humidity or availability of free water, solar radiation and wind. Soil or other growing medium conditions must be considered when pathogens are applied to the growing medium, as with nema-todes and fungi. The environmental conditions can be controlled to some extent in greenhouse crops by computerized environmental control. In temperate regions growers use advanced techniques and modern greenhouse structures to control the environment as much as possible. Completely closed greenhouses are a new trend in which climate control is fully regulated. In Mediterranean greenhouses, climate control is much less advanced and temperature and humidity ranges are large. For an overview of greenhouse structure factors influencing biological control, I refer to Lindquist and Short (2004). Consequently, greenhouse technology in a broad sense has an impact on the activity of entomopathogens. Mediterranean greenhouses are also much more open for reasons of cooling, and wind can have a great effect on relative humidity, even at the microclimate at the leaf-boundary level which is critical for the efficacy of fungi (Fargues et al. 2003a). UV radiation is harmful to many pathogens, but under glass or plastic this effect is greatly reduced. Nevertheless, UV can still be harmful and various types of plastics can have different effects (Costa et al. 2001).

Butt (2002) and Jaronski (2010) reviewed the environmental factors that influence the success of fungi against insects. It has been often mentioned that epizootic effects of pathogens are very valuable and a marked difference from chemicals. I have witnessed wonderful epizootic effects in cucumber and chrysanthemum with L. muscarium on whitefly and with L. longisporum on aphids in the 1980s following treatments of the commercial products. This worked in low greenhouses with cucumbers where the RH was very high at certain periods of the year. A similar situation occurred under the plastic covers used for decreasing the day length period to induce flowering in chrysanthemum. But since these technologies have changed dramatically, such favourable conditions for fungi no longer occur. In using biopesticides, epizootics cannot be counted on, and moreover, they are unlikely to occur. I will come back to this later in this chapter.

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