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Containment testing

Host range testing methods

No-choice tests

The no-choice test design was the first approach used in early weed biocontrol projects (van Driesche and Murray 2004). No-choice tests can be used with adult insects to assess feeding, survival to maturity and the willingness to oviposit. With larvae no-choice tests measure the ability of a plant to support the development of the immature life stages. The accuracy of the data obtained from no-choice tests obviously relies upon providing in non-limiting quantities the required part of the plant, whether it be bud, leaf, flower, seed pod etc, in optimum physiological condition. For species with larvae that do not move between plants, use of the no-choice test is ideal because the only choice available to the larva in the field would also be to either feed or die. The strength of no-choice tests is that negative results are very robust and provide convincing evidence that a test species is not likely to be used as a field host, provided of course that the experimental design includes an environment that permits normal behaviour of the biological control agent, as evidenced by a positive response (or excellent larval survival) to the normal host that is provided as a control. Use of no-choice tests early in the testing sequence provides a strong rationale for classifying un-attacked test species as non-hosts.

Unfortunately no-choice development tests sometimes indicate that herbivorous insect larvae can feed successfully on plant species that adult insects did not find or accept for oviposition. Sometimes this is as a result of time-dependent changes in responsiveness, that can be viewed as a high motivation to feed or oviposit when starvation or death is imminent (Barton-Browne and Withers 2002). Also positive responses within no-choice tests can sometimes be argued to have been artificially induced by confinement. The cage environment may bring the agent into close contact with a test species such that important host finding steps may be skipped (since the insect is literally put on the host or very near to it), allowing oviposition or adult feeding to occur on test species that might not have been approached or utilised in the field. This issue can be particularly important with parasitoids (van Driesche and Murray 2004).

So often no-choice tests alone cannot be used for risk assessment without incorporating other assay methods, or potentially safe and effective agents may be rejected unnecessarily Hill (1999). Often when used on phytophagous larvae to test development, no-choice tests are becoming known as the method for ascertaining the "physiological host range" or "fundamental host range" of an insect (van Klinken 2000). Using this terminology reminds the reader or reviewer that the risk assessment is not complete based on no-choice data alone, and data incorporating other testing methods will also be required to reliably predict the "field host range" (van Klinken 2000). Undoubtedly the most important benefit of no-choice tests is the tendency for them to reveal lower-ranked (lower preference or lower palatability) hosts whose status within the physiological host range can be missed if too much reliance is given to the result of choice tests (Withers 1999).

Often survival from immature larva to adult is not an absolute measure obtained from no-choice tests. Instead more careful monitoring is often conducted to obtain survivorship curves on the range of test plant species. When some survival to pupation or adulthood does occur, a single measure of percentage survival to adult of the cohort is often insufficient to enable accurate interpretation of the data. Some insects feeding on sub-optimal hosts conserve body mass at the expense of time taken to reach full size, others conserve development time at the expense of final body size (Barton-Browne 1995). The repercussions for these two strategies can be significant, so where a degree of physiological suitability for development is revealed for an insect feeding on a non-target plant in a no-choice test, both time taken to develop, as well as final body size, recorded in relation to gender, needs to be recorded and analysed appropriately to aid interpretation of the data.

Positive controls are essential to validate negative responses by showing that the group from which the test agents were taken had the capacity for oviposition or feeding. In parasitoids, negative controls are also required, in which test species are not exposed to parasitoids. These are held to detect mortality of test herbivores that is unrelated to the direct action of parasitism (van Driesche and Murray 2004).

Hill (1999) argued that the size and mobility of the agent should influence the choice of test design. Where agents are large, and mobile females can choose to move freely between plants with relative ease, and with reasonable frequency, choice tests may be more appropriate than no-choice tests. In this situation females can select or reject a host, and respond by arresting movement or dispersing once more. No-choice tests may be a more appropriate test for target species that are small relative to their host and disperse passively, or conversely are relatively immobile. In this case the choices available to an individual are to feed or to not to feed, a scenario that can be effectively tested by no-choice tests.

References

Barton-Browne L. (1995). Ontogenetic changes in feeding behavior. Pp. 307-342 In: Regulatory Mechanisms in Insect Feeding, R.F. Chapman and G. de Boer (Ed.) Chapman and Hall.

Barton-Browne L. and Withers T.M. (2002). Time-dependent changes in the host-acceptance threshold of insects: implications for host specificity testing of candidate biological control agents. Biocontrol Science and Technology 12: 677-693.

Hill R.L. (1999). Minimising uncertainty - in support of no-choice tests. Pp. 1-10 In: Host specificity testing in Australasia: towards improved assays for biological control, W.T.M., L. Barton Browne and J. N. Stanley (Ed.) CRC for Tropical Pest Management, Brisbane, Australia.

Withers T.M. (1999). Examining the hierarchy threshold model in a no-choice feeding assay. Entomologia Experimentalis et Applicata 91: 89-95.

van Driesche R.G. and Murray T.J. (2004). Overview of testing schemes and designs used to estimate host ranges. Pp. 68-89 In: Assessing host ranges for parasitoids and predators used for classical biological control: a guide to best practice, R.G. Van Driesche and R. Reardon (Ed.) USDA Forest Service, Morgantown, West Virginia.

van Klinken R.D. (2000). Host-specificity testing: why do we do it and how we can do it better. Pp. 54-68 In: Host-specificity testing of exotic arthropod biological control agents: the biological basis for improvement in safety, R.G. Van Driesche, T. Heard, A.S. McClay and R. Reardon (Ed.) USDA Forest Service Bulletin, Morgantown, West Virginia, USA.