In a series of laboratory experiments, acclimated pupae of Tuta absoluta were exposed to various constant low temperatures in order to estimate their maximum survival times (Kaplan–Meier, Lt99.99). A Weibull function was fitted to the data points, describing maximum survival time as a function of temperature. In another experiment at −6°C, the progress of mortality increasing with exposure time was identified. These values were fitted by a sigmoidal function converging asymptotically to 100% mortality for very long exposure times. Analysing mortality data from the maximum survival experiment by a generalized linear model showed a significant common slope parameter (p < .001) that reveals parallelism of the survival curves at each temperature if a log time axis is used. These curves appear stretched (time scaled) if plotted with a nonlogarithmic time axis. By combining these mathematical relations, it was possible to calculate a species‐specific ‘mortality surface’ which exhibits mortalities, depending on temperature and duration of exposure. In order to accumulate hourly mortalities for courses of varying temperatures, an algorithm was developed which yields mortality values from that surface taking into account the attained mortality level. In validation experiments, recorded mortalities were compared against modelled mortalities. Prediction of mortality was partially supported by the model, but pupae experiencing intensely fluctuating temperatures showed decreased mortality, probably caused by rapid cold hardening during exposure. Despite this observation, mortality data converged to distinct levels very close to 100% depending on the intensity of temperature fluctuations that were characteristic for different types of experiments. The highest mortality limit occurred at intensely fluctuating temperatures in laboratory experiments. This constituted a benchmark that was not reached under various field conditions. Thus, it was possible to identify temperature limits for the extinction of field populations of Tuta absoluta pupae.
The ectoparasitic mite Varroa destructor poses one of the biggest threats to the western honeybee Apis mellifera. Possibilities to control the mites are limited; therefore, it is important to assess the performance of novel alternatives like the use of biological control agents. The goal of our work was to evaluate the effects of the entomopathogenic fungus Metarhizium anisopliae var. anisopliae BIPESCO 5, which is a potential biological control agent against the bee parasitic mite V. destructor, on the behavior of the parasite. In detail, we investigated whether the presence of fungal spores on the surface of host nurse bees, A. mellifera, affects the host choice behavior of adult female mites. We conducted two behavioral assays to monitor the behavior of the mites towards adult bees inoculated with a solution of fungal spores. Both choice and no-choice experiment revealed that M. anisopliae has a significant repellent influence on V. destructor‘s host selection behavior. The mites preferred, and stayed longer on, nurse bees free from fungal spores over bees carrying fungal spores. Our study provides the first evidence of sub-lethal behavioral effects of a fungal biocontrol agent against Varroa mites.
Purpose The successful implementation of a plant protection product depends on its effectiveness against a target species and its safety for the environment. Risk assessment schemes have therefore been devised to facilitate classification and regulation. These guidelines, however, are directed towards chemical substances and are in many cases less suitable for the assessment of products employing microorganisms. Methods In this study, we developed a protocol for non-target testing of soil-applied entomopathogenic fungi for the biocontrol of insect pests. Using the predatory mite Gaeolaelaps (Hypoaspis) aculeifer as a non-target model organism, our protocol evaluates the lethal and sublethal effects of the fungus in recommended and ten-fold field concentrations. Results The proposed protocol considers fungal biology when setting test duration, endpoints, and quality control measures. To assess its practicability, we performed a trial with Metarhizium brunneum ART2825 as a representative entomopathogenic fungus. The biocontrol agent was able to infect a susceptible host and reproduce, showing that potential hazards can be detected using our approach. No hazard was detected for the non-target species, with no statistically significant differences in 5-week survival and reproductive output between treated and untreated groups. Conclusion Based on our results, the protocol is deemed appropriate for the detection of non-target effects. Subject to further validation, our approach could thus provide the basis for standardized protocols for the evaluation of the environmental safety of biocontrol organisms.
Entomopathogenic fungi (EPF) represent promising control agents against wireworms but success in field experiments is inconsistent. The physiological condition of the targeted insect is crucial for its ability to withstand fungal infection. In particular, nutritional status is among the most important determinants of the insects’ immune defense. In this study, we investigated the effects of diet on the development of the wireworm Agriotes obscurus (L.) (Coleoptera: Elateridae) and its subsequent susceptibility to the fungal pathogen Metarhizium brunneum (Petch) (Hypocreales: Clavicipitaceae) in a pot experiment. After being reared on one of five plant diets for eight weeks, wireworms were exposed to an environment inoculated with the EPF and monitored for their susceptibility to fungal infection. We then performed a field experiment in which three plant diets (clover, radish, and a cover crop mix), selected according to the insects’ performance in the laboratory experiment, were grown as a cover crop with EPF application. Plant diet influenced growth and development of larvae, but there were no strong differences in susceptibility toward fungal infection in the laboratory experiment. Damage levels in EPF-treated plots in the field varied depending on the cover crop. Damage was highest in plots planted with a mix of cover crop species, whereas damage was lowest in plots with clover or radish alone. This agrees with the laboratory results where insect performance was inferior when fed on clover or radish. Cover crop effects on wireworm damage in the subsequent cash crop may thus vary depending on the cover crop species selected.
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