Imperfect match between radiation exposure times required for conidial viability loss and infective capacity reduction attenuate UV-B impact on Beauveria bassiana.

BACKGROUND: UV-B radiation represents a significant challenge for the widespread use of entomopathogenic fungi in pest management. This study focused on research of the asynchronous response between virulence and conidial viability against Ceratitis capitata adults using specific statistical models. Moreover, it was also investigated whether the observed differences in susceptibility to UV-B radiation in in vitro assays among three selected isolates of Beauveria bassiana were reflected in the above-mentioned asynchrony. RESULTS: While the irradiation of the three isolates of B. bassiana was associated with a significant loss of conidial viability, their virulence was not significantly affected compared to nonirradiated treatments when exposed to 1200 mW m-2 for 6 h before or after the inoculation of C. capitata. In fact, the irradiation time needed to reduce the mortality to 50% compared to the controls was 34.69 h for EABb 10/225-Fil, 16.36 h for EABb 09/20-Fil, and 24.59 h for EABb 09/28-Fil. Meanwhile, the irradiation time necessary to reduce conidial viability to 50% was 9.89 h for EABb 10/225-Fil, 8.74 h for EABb 09/20-Fil, and 4.71 h for EABb 09/28-Fil. CONCLUSION: These results highlight the importance of modeling the response of entomopathogenic fungi virulence and conidial susceptibility when exposed to UV-B radiation for the selection of environmentally competent isolates, regardless of the results obtained in previous in vitro assays on conidial germination. This strategic approach is critical in overcoming the challenges posed by UV-B radiation and holds the key to realizing the full potential of entomopathogenic fungi in pest management. © 2023 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Land-Use Type Drives Soil Population Structures of the Entomopathogenic Fungal Genus Metarhizium.

Species of the fungal genus Metarhizium are globally distributed pathogens of arthropods, and a number of biological control products based on these fungi have been commercialized to control a variety of pest arthropods. In this study, we investigate the abundance and population structure of Metarhizium spp. in three land-use types-arable land, grassland, and forest-to provide detailed information on habitat selection and the factors that drive the occurrence and abundance of Metarhizium spp. in soil. At 10 sites of each land-use type, which are all part of the Swiss national soil-monitoring network (NABO), Metarhizium spp. were present at 8, 10, and 4 sites, respectively. On average, Metarhizium spp. were most abundant in grassland, followed by forest and then arable land; 349 Metarhizium isolates were collected from the 30 sites, and sequence analyses of the nuclear translation elongation factor 1α gene, as well as microsatellite-based genotyping, revealed the presence of 13 Metarhizium brunneum, 6 Metarhizium robertsii, and 3 Metarhizium guizhouense multilocus genotypes (MLGs). With 259 isolates, M. brunneum was the most abundant species, and significant differences were detected in population structures between forested and unforested sites. Among 15 environmental factors assessed, C:N ratio, basal respiration, total carbon, organic carbon, and bulk density significantly explained the variation among the M. brunneum populations. The information gained in this study will support the selection of best-adapted isolates as biological control agents and will provide additional criteria for the adaptation or development of new pest control strategies.

Different strategies to kill the host presented by Metarhizium anisopliae and Beauveria bassiana.

Studies conducted over the last decades have shown the potential of entomopathogenic fungi for the biocontrol of some insect pests. Entomopathogenic fungi infect their host through the cuticle, so they do not need to be ingested to be effective. These fungi also secrete secondary metabolites and proteins that are toxic to insect pests. In this context, we analyzed the pathogenicity of Metarhizium anisopliae (Metschn.) strains IBCB 384 and IBCB 425 and Beauveria bassiana (Bals.-Criv.) Vuill. strains E 1764 and E 3158 against Galleria mellonella (Linn.) larvae, during pre-invasion and post-invasion phases. The results showed M. anisopliae, especially strain IBCB 384, was most virulent in the pre-invasion phase against G. mellonella, whereas B. bassiana, especially strain E 1764, was most virulent in the post-invasion phase. During in vivo development and in the production of toxic serum, B. bassiana E 3158 was the most virulent. Different fungal growth (or toxin) strategies were observed for studied strains. Metarhizium anisopliae IBCB 425 prioritizes the growth strategy, whereas strain IBCB 384 and B. bassiana strains E 1764 and E 3158 have a toxic strategy. All strains have pathogenicity against G. mellonella, indicating their possible use for biocontrol.

UV-B radiation-related effects on conidial inactivation and virulence against Ceratitis capitata (Wiedemann) (Diptera; Tephritidae) of phylloplane and soil Metarhizium sp. strains.

Recent studies have demonstrated the presence of Metarhizium species on the epigeal areas of weeds and woody plants in various Mediterranean ecosystems, and the question arises whether isolates from the phylloplane, which experiences greater exposure to environmental UV-B radiation than soil isolates do, could have better UV-B radiation tolerance. The in vitro response of 18 Metarhizium strains isolated from phylloplane and soil of several Mediterranean ecosystems to UV-B radiation and the in vitro and in vivo effects of UV-B radiation on the viability and virulence of a selected M. brunneum strain against C. capitata were determined. The conidial germination, culturability and colony growth of these strains exposed to 1200mWm-2 for 2, 4 or 6h were evaluated. Germination rates below 30% and poor conidia recovery rates were observed for all strains. However, no relationship between the Metarhizium species or isolation habitat and the effect of UV-B radiation was found. Strain EAMa 01/58-Su, which showed a high tolerance to UV-B inactivation in terms of relative germination, was subsequently selected to investigate the UV-B related effects on virulence toward C. capitata adults. In a series of bioassays, the virulence and viability was determined using pure dry conidia, which were irradiated with 1200mWm-2 for 6h prior or after adult flies were inoculated, which resulted in a significant 84.7-86.4% decrease in conidial viability but only a slightly significant reduction of virulence, with 100.0% and 91.4% adult mortality rates and 4.6 and 5.9days average survival time for the no UV-B and UV-B treatments, respectively. A second series of experiments was performed to determine whether the UV-B effects on strain EAMa 01/58-Su were dose- or exposure time-dependent. Adult flies were inoculated with five doses (1.0×104-1.0×108conidiaml-1) and then irradiated at 1200mWm-2 for 6h, and similar LC50 values, 3.8×107 and 4.3×107conidiaml-1, were determined for the UV-B and no UV-B treatments, respectively. However, the LT50 values for flies inoculated with 1.0×108conidiaml-1 and with1.0×107conidiaml-1 were 15.1% and 30.8% longer for UV-B treatments than no UV-B treatments, respectively. Next, adult flies were treated with 1.0×108conidiaml-1 and then exposed to 1200mWm-2 for 0, 6, 12, 24, 36 and 48h, and the relationships among exposure time and conidia viability and fly mortality losses were determined. The exposure time for adult flies at 1200mWm-2 to achieve a 50% reduction in fly mortality was 47.2h, which was longer than that of 5.6h required for a 50% reduction in conidia viability. Our results show that the UV-B radiation significantly affected the virulence of EAMa 01/58-Su strain against C. capitata adults, with this effect being dependent on the exposure time but not related to fungal dosage.

Responses to abiotic environmental stresses among phylloplane and soil isolates of Beauveria bassiana from two holm oak ecosystems.

The response of entomopathogenic mitosporic ascomycete (EMAs) to abiotic stresses might be adapted to the microhabitats in which they inhabit. In phylloplane, these organisms are more exposed to such stresses than they are in soil, which may have led to adaptation to this environment. In the present work, we investigate whether Beauveria bassiana genotype or isolation habitat, i.e., soil or phylloplane, within the same geographic area influences their responses to key environmental stresses, such as temperature, moisture and ultraviolet radiation (UV-B), which can affect their successful use in microbial control. Twenty isolates of B. bassiana obtained from the soil and phylloplane in two ecosystems from southern Spain (holm oak dehesa and a reforested area) were selected to study the population distribution of these isolates and evaluate their thermal, humidity and UV-B requirements. Molecular characterization was conducted by using elongation factor-1α (EF-1α), the intergenic nuclear region Bloc and 15 microsatellite primers. The cluster analysis based on concatenated EF-1α and Bloc sequences grouped the 20 isolates into five clades within B. basiana, with Clades a, b, d and e containing both soil and phylloplane isolates and Clade c including three phylloplane isolates. The dendrogram and the minimal spanning network generated from the genetic distances among multilocus genotypes showed four divergent groups corresponding to the five clades obtained based on the sequence data (Clades b and d were represented in the same group), with a high degree of shared alleles within groups and few alleles shared among groups. Although no relationship was found between MLG and the habitat (soil or phylloplane) of isolation, isolates grouped into Clade c, all of which were collected from phylloplane, formed a separate group of MLGs. To investigate our hypothesis, the responses to temperature (germination and colony growth evaluated in the range 15-35°C), water activity (conidia germination evaluated against values of aw between 1 and 0.862) and UV-B exposure (conidia exposed to 920 or 1200mWm-2 for 2, 4 or 6h) of the soil and phylloplane isolates from the five clades were investigated. No associations of isolate-specific genetic or physiological characteristics with isolate habitat, i.e., soil or phylloplane, were found. These results provide no support for the hypothesis that EMAs strains from the phylloplane have evolved to resist unfavourable environmental conditions.

Diversity of entomopathogenic Hypocreales in soil and phylloplanes of five Mediterranean cropping systems.

The diversity of entomopathogenic Hypocreales from the soil and phylloplanes in five Mediterranean cropping systems with different degrees of management [organic olive orchard conventional olive orchard, holm oak reforestation, holm oak dehesa (a multifunctional agro-sylvo-pastoral system), and sunflower plantation] was studied during four seasons. A total of 697 entomopathogenic fungal isolates were obtained from 272 soil samples, 1608 crop phylloplane samples and 1368 weed phylloplane samples. The following nine species were identified: Beauveria amorpha, B. bassiana, B. pseudobassiana, B. varroae, Metarhizium brunneum, M. guizhoense, M. robertsii, Paecilomyces marquandii and lilacinum using EF-1α gene sequences. All the fungal entomopathogenic species were found in both the soil and phylloplane samples, with the exception of M. robertsii, which was only isolated from the soil. The species richness, diversity (Shannon-Wiener index) and evenness (Pielou index) were calculated for each cropping system, yielding the following species ranking, which was correlated with the crop management intensity: holm oak reforestation>organic olive orchard>conventional olive orchard>holm oak dehesa>sunflower plantation. The number of fungal species isolated was similar in both phylloplane habitats and dissimilar between the soil and the crop phylloplane habitats. The ISSR analysis revealed high genotypic diversity among the B. bassiana isolates on the neighbourhood scale, and the isolates were clustered according to the habitat. These results suggest that the entomopathogenic Hypocreales in the phylloplane could result from the dispersal of fungal propagules from the soil, which might be their habitat of origin; a few isolates, including EABb 09/28-Fil of Beauveria bassiana, inhabit only the phylloplane.