9 Methods for Assessment of Contaminants of Invertebrate Biological Control Agents and Associated Risks Mark S. Goettel and G. Douglas Inglis Lethbridge Research Centre, Agriculture and Agri-Food Canada, 5403–1st Avenue South, Lethbridge, Alberta T1J 4B1, Canada (email: [email protected]; [email protected]; fax number: +1-403-382-3156) Abstract With the importation or transport of any commodity, there exists the hazard that unwanted organisms or substances (i.e. ‘contaminants’) will be conveyed and introduced. Invertebrate biological control agents (IBCAs) can be contaminated with numerous biotic and abiotic enti- ties such as parasitoids, hyperparasitoids, pathogenic and/or non-pathogenic microorgan- isms, other organisms, pesticide residues, unwanted packaging materials, etc. Therefore, assessment of the risk posed by the contaminant must be addressed in the commerce of IBCAs. In this chapter, we provide an overview of possible contaminants of IBCAs and of the methods used to detect them. We consider two major factors when assessing risk. These are: (i) whether the IBCA is field collected or insectary reared; and (ii) whether the IBCA is exotic, being introduced for classical biological control or is indigenous and to be used for inundative biological control. We conclude that minimal risk is posed by contaminants of commercially mass-produced IBCAs, that are established in the area of use and are to be used inundatively. For such IBCAs, we recommend that the standards established for impor- tation of most commodities, such as many foodstuffs, plants, vegetables, fruits etc. (i.e. qual- ity control assurances by the producers) be adopted. Field-collected IBCAs, on the other hand, have a much higher potential for harbouring unknown contaminants that may repre- sent a risk. We recommend that feral IBCAs to be released outside of the area from which they were collected should be kept for at least one generation under quarantine, if at all pos- sible, and that the appropriate quarantine protocols are applied. This would allow the detec- tion and elimination of biotic contaminants. We stress that the key to the regulation of IBCAs is to address the extent of the possibility that a contaminant could pose a hazard to the com- modity or to the environment of the commodities’ final destination, and, if warranted, to ensure that such harm does not take place. The extent to which measures for prevention of transfer of contaminants are implemented must be weighed in relation to the present transfer of unknown or unwanted substances by other means. If the cautionary approach is strictly implemented for all possible contaminants, then almost certainly the international move- ment of IBCAs would grind to a halt. The ramifications of this must be weighed against the presently known benefits of IBCAs in our agriculture and forestry industries. ©CAB International 2006. Environmental Impact of Invertebrates for Biological Control of Arthropods: Methods and Risk Assessment (eds F. Bigler et al.) 145 146 M.S. Goettel and G.D. Inglis Introduction could affect the IBCA’s efficacy, the health of the user, or which could become estab- In the importation or transport of any com- lished or pollute the new environment. modity, there is always a concern that Possibilities include pathogenic or non- ‘contaminants’ will be conveyed and intro- pathogenic microorganisms, parasitoids, duced. Before we proceed, it is first neces- hyperparasitoids, misidentified inverte- sary to define what constitutes a brates, pesticide residues, unwanted pack- contaminant. According to Merriam- aging materials, etc. In this chapter, we Webster’s Medical Dictionary (2003), a con- characterize possible contaminants as taminant is defined as: ‘a substance that either microorganisms, invertebrates or abi- contaminates; to contaminate is to soil, otic agents. stain or corrupt by contact; to tarnish; to pollute; contamination is the act of conta- minating or polluting including (either Microorganisms intentionally or accidentally) unwanted substances or factors.’ The key word in this Microorganisms are ubiquitous and they definition is ‘unwanted’. Restricting the are always found in association with both definition to unwanted presents a cadre of field-collected and mass-reared inverte- problems. What is ‘unwanted’ as far as an brates, including IBCAs. Their associations invertebrate biological control agent (IBCA) are complex, and their associations with is concerned? Human-pathogenic bacteria IBCAs can be considered as either inciden- associated with IBCAs may be unwanted, tal, mutualistic, pathogenic or commensal- yet the introduction of a small number of istic. It is important to emphasize that cells of a relatively weak human-patho- these categories are not necessarily exclu- genic microorganism does not necessarily sive of each other. A number of arthropods pose a serious threat. Microorganisms are vector mammalian (e.g. West Nile Virus) ubiquitous and no invertebrates are devoid and plant pathogens. Furthermore, micro- of them unless special measures are taken. organisms associated with IBCAs may be How does one determine whether a micro- pathogens of invertebrates, plants or verte- organism falls into the ‘unwanted’ cate- brates, including humans. Groups of micro- gory? To determine this, the definition of organisms dealt with in this chapter contaminant must also address risk, and include the viruses, bacteria, fungi and this paper defines contamination as the protozoa. For convenience, we also include inclusion of any unwanted substance or the nematodes in this section. Numerous factor (i.e. a contaminant) in the commerce examples of pathogens of beneficial arthro- of IBCAs that poses an unacceptable risk. pods are provided by Vinson (1990), and of In defining unacceptable risk, we limit our mass-produced IBCAs by Bjørnson and discussion to the impacts of contaminants Schütte (2003). on the health of the IBCAs or on humans, and their potential impact on ecosystems Viruses (e.g. introduction of non-indigenous micro- organisms). We also compare risk assess- Viruses are obligate, intracellular ments applied to other invertebrates in pathogens that consist of double-stranded some OECD (Organization for Economic or single-stranded nucleic acid (DNA or Cooperation and Development) countries. RNA) encased in a protective coating called a capsid. Collectively, the nucleic acid and capsule are termed a nucleocap- Contaminants Associated with sid. Depending on the virus, some nucleo- Invertebrates capsids are enclosed within a lipid envelope. Virions are the infectious unit of Using the above definitions, a contaminant a virus. In enveloped viruses, the virions could be one of numerous factors that consist of the nucleocapsid (i.e. nucleic Methods for Assessment of Contaminants of Invertebrate BCAs 147 acid and capsid) and the envelope. In non- by the hymenopteran parasitoid, Cotesia enveloped viruses, the virions are com- marginiventris (Cresson) (Hymenoptera: prised only of the nucleocapsid. Viruses do Braconidae), from infected to healthy not possess the ability to replicate them- Spodoptera larvae (Lepidoptera: selves independently of a living host, and Noctuidae) (Hamm et al., 1985). Viral thus cannot be cultured on microbiological pathogens are also present in numerous media. They, in essence, highjack the meta- mite and insect species that are used in bolic machinery of the host cell and trick it biological control (Bjørnson and Schütte, into producing progeny viruses. A number 2003). For instance, cytovirus and nuclear- of entomopathogenic viruses produce polyhedroviruses are known from the occlusion bodies (OBs), in which the viri- aphid predator, Chrysoperla (Neuroptera: ons are embedded within a paracrystalline Chrysopidae) (Martignoni and Iwai, 1986). protein matrix. The OBs protect the virions Unidentified, non-occluded virus particles (i.e. from ultraviolet light) and increase were observed in the yolk of predatory persistence of the virions outside of the Neoseiulus cucumeris (Oudemans) host; their formation has important conse- (Mesostigmata: Phytoseiidae) and quences for their disease-producing poten- Phytoseiulus persimilis Athias-Henriot tial. They may also serve an important (Mesostigmata: Phytoseiidae) mites; how- function in the infection process. ever, the effects of these on predatory effi- Most of the viruses associated with cacy were not established (Bjørnson et al., insects belong to one of 12 viral families, 1997). More information on entomopatho- but many remain unclassified. Of particular genic viruses can be found in Granados concern to IBCAs are viruses in the families and Federici (1986), Adams and Bonami Baculoviridae, Poxviridae, Parvoviridae, (1991a), Tanada and Kaya (1993), Miller Reoviridae and Polydnaviridae. Some of (1997), Hunter-Fujita et al. (1998) and these viruses possess restricted host ranges Miller and Ball (1998). affecting insects in a specific genus, whereas others can affect a variety of hosts Bacteria belonging to different orders. For most viruses of IBCAs, the primary route of The bacteria represent a very large and infection is through the alimentary canal diverse group of prokaryotes. There are two after ingestion. However, other routes of main types of prokaryotes, the archaeabac- infection do occur (e.g. mechanical intro- teria and the eubacteria (collectively duction of virions on infested ovipositors). referred to as bacteria). Although all bacte- In some instances, viruses are restricted to ria lack a nucleus and organelles, they