Abstract

The turbulent velocity field in and above homogeneous forest canopies on flat ground has several universal features. These include a mean velocity profile with a point of inflexion; vertical inhomogeneity in second moments; positive u and negative w skewnesses; and integral length scales of order h, the canopy height. These features scale with u* and h across a wide range of wind tunnel models and real canopies. Turbulence spectra also share common features such as vertical invariance of the position of the spectral peak and departures from classical inertial subrange behaviour. Many of these characteristics are more similar to what is observed in a plane mixing layer than in a boundary layer. The eddy structure of a plane mixing layer owes its origin to the instability of its inflexion point velocity profile, one of the features it shares with canopy flows. Linear perturbation models of this instability yield results that fit well with the eddy structure observed in canopies, strongly suggesting that it is instability of the canopy mean velocity profile, a profile set by the momentum absorption capacity of the canopy as a whole, that controls the universal turbulent structure of plant canopy flow. Turbulence produced directly by eddies shed from plant parts plays a minor role. Turbulent airflow over isolated hills also displays some characteristic features, including a large relative velocity speed-up at low levels above the hill crest. It can be demonstrated that this is caused by the pressure field set up by the whole of the flow field around the hill. In this sense it can be regarded as ‘imposed’ on any canopy growing on the hill.

Introduction

Host specificity screening is the most important step that each potential weed biological control agent (whether insect or pathogen) has to pass prior to its introduction (Schroeder, 1983). Regardless of its control potential, only the demonstrated safety of plants of economic or ecological importance in the release area, will lead to approval for release of the control agent. The use of screening-protocols for weed biological control agents (Wapshere, 1975; Schroeder, 1983; CAB, 1986) are now widely accepted. Even less rigorous tests in the first part of this century have never led to the release of a ‘new pest’ (Crawley, 1989; Howarth, 1991).

The safety record of weed biocontrol using insects is excellent, nonetheless, the growing awareness of the public towards environmental interference by man, has led to legislative regulations such as the ‘Biological Control Act of 1984’ in Australia. Today, information on the environmental impact, alternatives, the relationship between local short-term uses and long-term productivity, and a risk analysis of the proposed action are required. Conflicts of interest over the implementation or safety of a specific biological control programme or agents, may still arise (Delfosse, 1990).

During the last decades especially, the critical objections of nature conservationists who fear for already stricken and endangered plant species related to the target weed, have considerably delayed the progress of programmes (Schroeder and Goeden, 1986). Despite the excellent safety record of weed biocontrol, we have to assess the risks of introducing a foreign species into a new environment.

There are several early nineteenth century observations of disorganized clumps of material in the cortex of orchid roots, but Reissek (1847) may have been the first to realize that these clumps were the remains of fungal hyphae coiled inside the plant cells. He studied the roots of Neottia nidus-avis and observed tissues that were so young that living hyphae were still present. Reissek also reported these coils from roots of other European orchid species belonging to Orchis, Ophrys and Gymnadenia and from several tropical orchids. Some years later, ‘une matière granulaire’ originating from decomposed hyphal coils was observed in the basal cells of orchid protocorms (Prillieux & Rivière, 1856). These observations stimulated interest in orchid mycorrhiza, particularly as regards investigating the species that display marked chlorophyll deficiency (Prillieux, 1856; Johow, 1885, 1889; Groom, 1895; Janse, 1897; MacDougal, 1899a, b; Magnus, 1900).

In a survey of more than 500 cultivated orchid species, Wahrlich (1886) managed to find the characteristic fungal infections in every one of them, and Frank (1891) concluded from observations of the infected tissue that the hyphae became lysed, and that the nuclear modification in the infected cells was a sign of the active participation of the host in this process.

A more complete understanding of the functions of fungi in the life history of orchids arose from Bernard's germination studies during the years 1899 to 1909, and from Burgeff's study that primarily dealt with the fungi (1909). Both of these investigators succeeded in germinating orchids in vitro and in observing the process of infection in embryos and seedlings. Among the early investigators opinions differed as to the nature of the relationship.

Introduction

Biotechnology (genetic engineering) and biological control offer many opportunities to improve agricultural production. Potential benefits include: achieving higher crop and livestock yields; improving nutritional make-up of crops and livestock; substituting biocontrols for pesticide use; as well as controlling some soil and water pollutants. At the same time, the use of biotechnology and release of genetically engineered organisms (for biocontrol and other purposes) into the environment could create serious ecological, social, and economic hazards (Pimentel et al, 1989).

One objective in the implementation of any new technology is to achieve maximum benefits with minimal risks to the environment, economy, and society. The other major objective is to be able to use the technology so that all future benefits are realized. If a serious problem results from the application of just one project, the future development of the entire technology is jeopardized. Note what has happened to nuclear energy in North America because of mismanagement by industry and the government.

In this article, the potential risks of the use of biotechnology and release of genetically engineered crops and biocontrol organisms into the environment are assessed. Various approaches are suggested that could be employed to minimize and reduce the risks of biotechnology to the environment and society.

Ecological issuesCrop resistance to pests

The engineering of crop resistance to insect and plant pathogen pests offers advantages to reduce the use of insecticides and fungicides in crop protection. This will generally reduce problems from pesticides (NAS, 1989; Pimentel et al, 1993) and improve the economics of pest control (Pimentel, 1986).

Isolation

Orchid endophytes can be isolated from naturally occurring protocorms, from roots, and occasionally from rhizomes, tubers or corms. The advantage of extracting a fungus from protocorm tissue is that its role in seedling development may almost be taken for granted, whereas organs of mature plants may house a variety of fungi (Harvais & Hadley, 1967a; Hadley, 1970b). Furthermore, infection is extensive in the basal part of the protocorm, and protocorm cells are often fairly large. In contrast, the cortical cells of roots are long and narrow and the infection tends to be patchy and irregular. The most promising places to search for infection in roots are areas of the cortex below epidermal hairs, especially those close to the root tip (Bernard, 1909). Infected parts of an orchid root tend to become faintly yellowish or opaque. The living pelotons, where present, will be found close to the surface of the organ.

When the rhizome is infected it is usually the outer cortex that is most suitable for isolation purposes. In Liparis lilifolia both the condensed rhizome and the bases of attached leaves are infected. Isolation from tubers of orchidoid species has apparently never been successful, although Fuchs & Ziegenspeck (1925) report that superficial cell layers of globose tubers may be infected as well as the extremities of palmately divided tubers. Corms are usually without infection (Chapter 7).

Care must always be taken to remove the soil organisms that are normally found on any underground plant parts. Fungi that occur in the rhizosphere can include strains of Rhizoctonia that differ from those that form pelotons inside, and their relevance for the orchid mycorrhiza is not clear (Warcup & Talbot, 1967).

Background

Surely no plant group surpasses the orchid family in its appeal to scientific curiosity and imagination. The untiring professional and public interest in the Orchidaceae is reflected in a considerable number of volumes published each year on orchids; in particular, the great structural variety in the orchid flower is well represented in the literature.

Compared with the floristic study of this large and taxonomically complex group, other types of investigation lag behind. Of the many unusual features that can be noted in the orchid family their mode of life in association with fungi has probably attracted the least attention, which is surprising since it is without doubt a fundamental aspect of orchid biology.

The orchid family is the only large group of higher plants that makes consistent use of an alternative nutritional system. Orchid mycorrhiza differs from other major types of mycorrhiza in that the fungus supplies the plant with energy. During some of its life stages the orchid can rely entirely on mycotrophy for nutrition, while during other stages the plant makes use of both mycotrophy and phototrophic nutrition, either alternately or the one supplementing the other.

The great diversity of orchid habitats, which include places where paucity of soil or light precludes most plant life, is best understood in the context of orchid mycotrophy. In fact, many of the outstanding features of the orchids, such as the complexity of floral structures, the diversity of plant–pollinator interactions and the unusual characteristics of the seeds, can all be viewed as functional adaptations in relation to or dependent on the mycotrophic lifeform (Chapter 12). This in itself justifies a treatise on orchids centred on their mycorrhiza.

Introduction

The agro-ecosystems in Fennoscandia are characterized by relatively few and moderate plant protection problems. The number of pests and plant diseases is smaller than in warmer regions. Due to a shorter growing season and lower temperatures, many multivoltine insects develop only one generation a year. The beneficial fauna, primarily the many predaceous bugs (anthocorids and mirids) and mites (phytoseiids), have also proved to be highly efficient as control measures.

Pest control systems have been developed for several agricultural crops, which require minimum inputs of pesticides. The programmes for many greenhouse cultures, based on biocontrol, and those for fruit crops, using integrated approaches, have been especially successful.

Development of IPM in Norwegian fruit orchards

In the 1950s and early 1960s a spray programme that consisted of 6–8 annual applications was commonly used against insects and mites. This heavy routine spray programme was expensive and caused undesirable environmental impact.

An extensive research programme on IPM, which started in the 1960s, provided important information on the biology and ecology of major pests and their natural enemies. Based on this new knowledge, the use of pesticides could be reduced. By 1974, farmers who had adopted IPM in their orchards, used only 0–3 pesticide applications a year without significant damage to the crop (Edland, 1989a). During recent years, common use of scab-warning devices has resulted in further reduction of spray programmes. Thus, in 1988 the average use of insecticides and acaricides was 1.1 sprays and of fungicides 3.5 sprays, for the whole season in many orchards (Edland, 19896). This is less than 20% of what is being used in integrated fruit production in some other European countries (Prinoth, 1990).

Introduction

Many attempts have been made to control soilborne plant pathogens and to improve the growth of plants by inoculation of seed or soil with selected strains of micro-organisms; mainly bacteria. More recently, selected bacteria that are expected to degrade xenobiotics have been suggested for the bioremediation of polluted sites (Short et al 1990). Nowadays, risk assessment studies also need to address the fate of engineered telluric soil and non-telluric soil bacteria (Tiedje et al., 1989; Doyle etal, 1991).

Besides increased crop yields claimed by Russian workers (Schroth and Becker, 1990), most of the tentative applications of micro-organisms in agricultural soils have failed. The widely practised inoculation of legume seeds with Rhizobium spp. is one of the few examples of success of application of micro-organisms to improve crop yield (Stacey and Upchurch, 1984). Application of the strain K84, and more recently of the modified strain K1026, of Agrobacterium radiobacter to control crown gall of plants is one of the few examples of a biological control method commercially applied with some success in several countries (Ryder and Jones, 1990). In most cases, the beneficial effects expected from the microbial inoculation are not consistently reproduced under field conditions. The poor survival of the introduced microorganisms in soil is the main explanation for these failures. In fact, the population density of the introduced micro-organisms decreases to the limit of the carrying capacity of the soil. Moreover, the soil represents very heterogeneous environments in which the introduced micro-organisms must find suitable habitats, some of which are very strain specific (Hattori and Hattori, 1976).

Introduction

The spore-forming, Gram-positive bacterium. Bacillus thuringiensis (Bt), is a ubiquitous soil micro-organism with a world-wide distribution (Martin and Travers, 1989). Many thousands of isolates have been discovered since Ishiwata first isolated Bt from diseased silkworm larvae in 1901 (Nakamura and Dulmage, 1988; Beegle and Yamamoto, 1992; Feitelson et al, 1992), and it is possible to group these isolates into at least 34 serovars (also called subspecies) (de Barjac and Fraction, 1990). However, biomolecular techniques, such as multiplex polymerase chain reaction (PCR), permit a more rapid identification of strains (Bourque et at., 1993), and it is now possible to target specific DNA sequences with the use of this technology.

The delta-endotoxin produced by Bt is the most widely used biological pesticide – 2.3 million kg was used annually according to Rowe and Margaritis (1987) – and it is the ‘front-runner’ in attempts to commercialize biological control agents (BCAs); albeit that Bt is only a BCA in a limited sense, since it is the product of a living organism (the delta-endotoxin) which is utilized, rather than the organism itself.

The precise ecological role of this cosmopolitan invertebrate pathogen in the natural environment remains speculative (Smith and Couche, 1991). Bt reportedly does not produce spores or crystals in infected host cadavers, it is relatively poor at spreading natural infections in the field and rarely causes epizootics (Fuxa, 1989). Its ubiquitous occurrence in the soil (Martin and Travers, 1989), phylloplane microflora (Smith and Couche, 1991), and in man-made ecosystems (Meadows et al., 1992) suggests that it may have a primary function other than as a pathogen.

Time of germination

In field sowing experiments performed in Maryland, USA, I observed the first signs of germination in Goodyera pubescens at the beginning of May, i.e. in spring, and in Galearis spectabilis and Corallorhiza odontorhiza at the beginning of June (Rasmussen & Whigham, 1993). The seeds had been harvested during the previous autumn and sown in November.

All other evidence on germination in situ is based on observations made at various times of the year of seedlings that were assumed to be young, although their exact age could not be determined. Casual observations of single protocorms are almost without value, but more reliable data are available from year-round studies of a given population which allow the size distribution of seedlings at different seasons to be compared. The general impression is that most of the holarctic species germinate in spring (Table 3.1). Ames (1922) found young seedlings of the American species Goodyera pubescens in spring, which agrees with my own observations from field sowings, and seedlings of many European taxa have also been found in spring, exceptions being some species of Dactylorhiza, Orchis, Himantoglossum, Spiranthes, Epipogium and Corallorhiza, the young seedlings of which were dug up in late summer or autumn (Table 3.1; see also Chapter 13).

A very large proportion of the holarctic species, except for those mentioned in Table 3.1, have not been recorded as protocorms in nature. A wider application of the field sowing technique that has recently been described (Rasmussen & Whigham, 1993) should yield more information on when seeds germinate and how rapidly the seedlings develop under near-natural conditions.

Introduction

The fungus Gliocladium virens Miller, Giddens and Foster is an important biological control agent (Papavizas, 1985). A formulation of this fungus (strain Gl-21) was recently registered with the US Environmental Protection Agency (EPA) by W. R. Grace & Co. Conn. The formulation was developed in cooperation with the Biocontrol of Plant Diseases Laboratory (BPDL), US Department of Agriculture (USDA) (Lumsden et al, 1991). It is intended for use against dampingoff diseases of vegetable and ornamental seedlings caused by the soil-borne plant pathogens, Pythium ultimum and Rhizoctonia solani in glasshouse operations (Lumsden and Locke, 1989). This fungus is one of the first to be registered for biocontrol of plant diseases and will soon be available in the US for commercial use in glasshouse applications under the trade name, GliogardTM

Certain criteria were considered important in the early stages of development of biocontrol agents (Lumsden and Lewis, 1989). In the development of a screening method for the selection of an appropriate micro-organism, the following points were considered. The screening would involve: (1) the use of a relatively uniform, commercially available soil-less medium that is used extensively in commercial glasshouses where the disease problem occurs; (2) targeted pathogens were selected that are important in the confines of a glasshouse where use of a biological control agent would probably be most successful because of a relatively uniform environment; (3) microorganisms indigenous to the US were selected because non-indigenous micro-organisms might be conceived as more likely problems for the US environment; (4) a single isolate of a biocontrol agent for control of both pathogens was preferred over a mixture of isolates; and (5) a high value crop, important in the ornamental production industry, was selected to defray the cost of development and registration.

Introduction

The number of plant protection products and active ingredients has been considerably reduced in Germany during the 5 years since the new Plant Protection Act (Gesetz zum Schutz der Kulturpflanzen) of September 1986 came into force. The number of registered plant protection products now amounts to 851 formulations with 216 active ingredients (18 August 1993, Fig. 28.1). The decline is a result of the consistent application of more stringent requirements for product registration.

In the coming years, the number of authorizations is expected to decrease further. Authorizations made before the Plant Protection Act of 1986 have expired and today's requirements for re-registration are tougher. Only about 25% of the plant protection products that were registered in West Germany in 1986, have already become subject to the new Plant Protection Act. In the new Bundeslander (Federal States) in eastern Germany, old registrations are valid until the end of 1995 with the exception of some products banned in Germany and some whose use is not acceptable. It is assumed that about 90% of them do not comply with the requirements for reregistration. Although the range of plant protection products has decreased in Germany, there was no marked effect on their usage until 1989. In the late 1980s consumption of active ingredients in West Germany amounted to about 36000 tonnes p.a., but dropped after 1989. About 33000 tonnes were sprayed in West Germany in 1990. In 1992 33 570 tonnes were sold in united East and West Germany, although the acreage nearly doubled after unification (Table 28.1).

There is a considerable body of observations on the development of orchid seedlings in vitro in relation to various physical and chemical factors. Provided that conditions in vitro support reasonably healthy growth, such observations can give us a fairly accurate idea as to how development is regulated in nature, in various soil types and during the different seasons. However, little has yet been done to supplement the laboratory evidence by setting up explantation experiments in the field.

As regards physical factors, their effect can be studied by subjecting cultures to varying regimes of temperature, light and atmosphere. In vitro cultures are not suitable for observation of the effects of moisture conditions since humidity in vitro is consistently high. The effects of chemical factors have been studied in numerous in vitro experiments with substrates of varying composition, but these observation may or may not be applicable to natural conditions. Some of the requirements of asymbiotic seedlings reflect the absence of both mycotrophy and fully functional photosynthesis, all nourishment being obtained by direct absorption from the substrate. The external requirements of symbiotic seedlings may yield some information on the needs of the orchid/fungus association in a natural substrate.

Seedlings grown in vitro pass through some developmental stages that appear to be critical. The first occurs immediately after germination when many seedlings die before a shoot tip has differentiated (Stoutamire, 1974; Van Waes, 1984).