Effect of Norwegian entomopathogenic fungal isolates against <em>Otiorhynchus sulcatus </em>larvae at low temperatures and persistence in strawberry rhizospheres

AbstractBiological control of belowground stages of the black vine weevil Otiorhynchus sulcatus F. (Coleoptera: Curculionidae) in strawberries in cool temperate regions using entomopathogens is challenged by low temperatures during the periods when larvae are vulnerable to infections. In a laboratory study we tested six indigenous Norwegian isolates of entomopathogenic fungi (one Beauveria bassiana, three Beauveria pseudobassiana, and two Metarhizium brunneum; Ascomycota: Hypocreales) for their efficacy against O. sulcatus larvae at 6, 12, and 18°C. At the lowest temperature only Beauveria spp. affected survival of O. sulcatus while all three fungal species reduced larval survival compared to the control treatment at 12 and 18°C. Two of the Norwegian isolates, one B. pseudobassiana and one M. brunneum, were then evaluated for long-term persistence (>1year) in the bulk soil and the rhizosphere soil of strawberries in a semi-field experiment. An exotic isolate of M. brunneum sharing origin with a widespread commercial biocontrol agent (F52/Met52 (Novozymes)) was included for comparison. All three isolates showed significantly higher abundances in the rhizosphere soil compared to bulk soil at 153, 366, and 471days after inoculation, thus indicating rhizosphere competence for B. pseudobassiana. Notably, CFU levels for both Norwegian isolates were much higher than for the exotic M. brunneum isolate. Selection of locally adapted isolates may therefore be of importance when considering biocontrol strategies of belowground pests in strawberry production

Reduced entomopathogen abundance in <i>Myrmica</i> ant nests-testing a possible immunological benefit of myrmecophily using <i>Galleria mellonella</i> as a model

Social insects such as ants have evolved collective rather than individual immune defence strategies against diseases and parasites at the level of their societies (colonies), known as social immunity. Ants frequently host other arthropods, so-called myrmecophiles, in their nests. Here, we tested the hypothesis that myrmecophily may partly arise from selection for exploiting the ants’ social immunity. We used larvae of the wax moth Galleria mellonella as ‘model myrmecophiles’ (baits) to test this hypothesis. We found significantly reduced abundance of entomopathogens in ant nests compared with the surrounding environment. Specific entomopathogen groups (Isaria fumosorosea and nematodes) were also found to be significantly less abundant inside than outside ant nests, whereas one entomopathogen (Beauveria brongniartii) was significantly more abundant inside nests. We therefore hypothesize that immunological benefits of entering ant nests may provide us a new explanation of why natural selection acts in favour of such a life-history strategy

Mortality risk from entomopathogenic fungi affects oviposition behavior in the parasitoid wasp <em>Trybliographa rapae</em>.

AbstractBiological control of pests in agroecosystems could be enhanced by combining multiple natural enemies. However, this approach might also compromise the control efficacy through intraguild predation (IGP) among the natural enemies. Parasitoids may be able to avoid the risk of unidirectional IGP posed by entomopathogenic fungi through selective oviposition behavior during host foraging. Trybliographa rapae is a larval parasitoid of the cabbage root fly, Delia radicum. Here we evaluated the susceptibility of D. radicum and T. rapae to two species of generalist entomopathogenic fungi, Metarhizium brunneum isolate KVL 04-57 and Beauveria bassiana isolate KVL 03-90. Furthermore, T. rapae oviposition behavior was assessed in the presence of these entomopathogenic fungi either as infected hosts or as infective propagules in the environment. Both fungi were pathogenic to D. radicum larvae and T. rapae adults, but with variable virulence. When host patches were inoculated with M. brunneum conidia in a no-choice situation, more eggs were laid by T. rapae in hosts of those patches compared to control and B. bassiana treated patches. Females that later succumbed to mycosis from either fungus laid significantly more eggs than non-mycosed females, indicating that resources were allocated to increased oviposition due to perceived decreased life expectancy. When presented with a choice between healthy and fungal infected hosts, T. rapae females laid more eggs in healthy larvae than in M. brunneum infected larvae. This was less pronounced for B. bassiana. Based on our results we propose that T. rapae can perceive and react towards IGP risk posed by M. brunneum but not B. bassiana to the foraging female herself and her offspring. Thus, M. brunneum has the potential to be used for biological control against D. radicum with a limited risk to T. rapae populations

Establishment Success of the Beetle Tapeworm <i>Hymenolepis diminuta</i> Depends on Dose and Host Body Condition

Parasite effects on host fitness and immunology are often intensity-dependent. Unfortunately, only few experimental studies on insect-parasite interactions attempt to control the level of infection, which may contribute substantial variation to the fitness or immunological parameters of interest. The tapeworm Hymenolepis diminuta—flour beetle Tenebrio molitor model—has been used extensively for ecological and evolutionary host–parasite studies. Successful establishment of H. diminuta cysticercoids in T. molitor relies on ingestion of viable eggs and penetration of the gut wall by the onchosphere. Like in other insect models, there is a lack of standardization of the infection load of cysticercoids in beetles. The aims of this study were to: (1) quantify the relationship between exposure dose and establishment success across several H. diminuta egg concentrations; and (2) test parasite establishment in beetles while experimentally manipulating host body condition and potential immune response to infection. Different egg concentrations of H. diminuta isolated from infected rat feces were fed to individual beetles 7–10 days after eclosion and beetles were exposed to starvation, wounding, or insertion of a nylon filament one hour prior to infection. We found that the establishment of cysticercoids in relation to exposure dose could be accurately predicted using a power function where establishment success was low at three lowest doses and higher at the two highest doses tested. Long-term starvation had a negative effect on cysticercoid establishment success, while insertion of a nylon filament and wounding the beetles did not have any effect compared to control treatment. Thus, our results show that parasite load may be predicted from the exposure dose within the observed range, and that the relationship between dose and parasite establishment success is able to withstand some changes in host body condition

Factors Affecting Vegetable Growers’ Exposure to Fungal Bioaerosols and Airborne Dust

We have quantified vegetable growers’ exposure to fungal bioaerosol components including (1→3)-β-d-glucan (β-glucan), total fungal spores, and culturable fungal units. Furthermore, we have evaluated factors that might affect vegetable growers’ exposure to fungal bioaerosols and airborne dust. Investigated environments included greenhouses producing cucumbers and tomatoes, open fields producing cabbage, broccoli, and celery, and packing facilities. Measurements were performed at different times during the growth season and during execution of different work tasks. Bioaerosols were collected with personal and stationary filter samplers. Selected fungal species (Beauveria spp., Trichoderma spp., Penicillium olsonii, and Penicillium brevicompactum) were identified using different polymerase chain reaction-based methods and sequencing. We found that the factors (i) work task, (ii) crop, including growth stage of handled plant material, and (iii) open field versus greenhouse significantly affected the workers’ exposure to bioaerosols. Packing of vegetables and working in open fields caused significantly lower exposure to bioaerosols, e.g. mesophilic fungi and dust, than harvesting in greenhouses and clearing of senescent greenhouse plants. Also removing strings in cucumber greenhouses caused a lower exposure to bioaerosols than harvest of cucumbers while removal of old plants caused the highest exposure. In general, the exposure was higher in greenhouses than in open fields. The exposures to β-glucan during harvest and clearing of senescent greenhouse plants were very high (median values ranging between 50 and 1500 ng m−3) compared to exposures reported from other occupational environments. In conclusion, vegetable growers’ exposure to bioaerosols was related to the environment, in which they worked, the investigated work tasks, and the vegetable crop

Diversity within the entomopathogenic fungal species <em>Metarhizium flavoviride</em> associated with agricultural crops in Denmark

BACKGROUND: Knowledge of the natural occurrence and community structure of entomopathogenic fungi is important to understand their ecological role. Species of the genus Metarhizium are widespread in soils and have recently been reported to associate with plant roots, but the species M. flavoviride has so far received little attention and intra-specific diversity among isolate collections has never been assessed. In the present study M. flavoviride was found to be abundant among Metarhizium spp. isolates obtained from roots and root-associated soil of winter wheat, winter oilseed rape and neighboring uncultivated pastures at three geographically separated locations in Denmark. The objective was therefore to evaluate molecular diversity and resolve the potential population structure of M. flavoviride. RESULTS: Of the 132 Metarhizium isolates obtained, morphological data and DNA sequencing revealed that 118 belonged to M. flavoviride, 13 to M. brunneum and one to M. majus. Further characterization of intraspecific variability within M. flavoviride was done by using amplified fragment length polymorphisms (AFLP) to evaluate diversity and potential crop and/or locality associations. A high level of diversity among the M. flavoviride isolates was observed, indicating that the isolates were not of the same clonal origin, and that certain haplotypes were shared with M. flavoviride isolates from other countries. However, no population structure in the form of significant haplotype groupings or habitat associations could be determined among the 118 analyzed M. flavoviride isolates. CONCLUSIONS: This study represents the first in-depth analysis of the molecular diversity within a large isolate collection of the species M. flavoviride. The AFLP analysis confirmed a high level of intra-specific diversity within the species and lack of apparent association patterns with crop or location indicates limited ecological specialization. The relatively infrequent isolation of M. flavoviride directly from crop roots suggests low dependence of root association for the species. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s12866-015-0589-z) contains supplementary material, which is available to authorized users

Habitat selection of a parasitoid mediated by volatiles informing on host and intraguild predator densities

To locate and evaluate host patches before oviposition, parasitoids of herbivorous insects utilize plant volatiles and host-derived cues, but also evaluate predator-derived infochemicals to reduce predation risks. When foraging in host habitats infested with entomopathogenic fungi that can infect both a parasitoid and its host, parasitoids may reduce the risk of intraguild predation (IGP) by avoiding such patches. In this study, we examined whether the presence of the entomopathogenic fungi Metarhizium brunneum and Beauveria bassiana in soil habitats of a root herbivore, Delia radicum, affects the behavior of Trybliographa rapae, a parasitoid of D. radicum. Olfactometer bioassays revealed that T. rapae avoided fungal infested host habitats and that this was dependent on fungal species and density. In particular, the parasitoid avoided habitats with high densities of the more virulent fungus, M. brunneum. In addition, host density was found to be important for the attraction of T. rapae. Volatiles collected from host habitats revealed different compound profiles depending on fungal presence and density, which could explain the behavior of T. rapae. We conclude that T. rapae females may use volatile compounds to locate high densities of prey, but also compounds related to fungal presence to reduce the risk of IGP towards themselves and their offspring. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s00442-015-3326-2) contains supplementary material, which is available to authorized users