Akanthomyces diversity in Brazil and their pathogenicity to plant-sucking insects.

Currently, species within the genus Akanthomyces are poorly studied and explored compared to other hypocrealean entomopathogenic fungi employed as commercial biocontrol agents. This study aimed to molecularly identify 23 Brazilian Akanthomyces strains, most originally isolated from aphids and scales (n = 22), and one from the coffee leaf rust, and further investigate their pathogenicity to six plant-sucking insects as a means to better understand their host spectra. We also explored the capacity of A. muscarius CG935 for blastospore production via liquid fermentation. Akanthomyces dipterigenus, A. muscarius, A. lecanii, and two unidentified species were recognized as naturally occurring in Brazil. Akanthomyces dipterigenus CG829 and A. muscarius CG935 were highly virulent to nymphs of Bemisia tabaci (67.5-85.4% confirmed mortality) and the aphid Aphis fabae (74.6-75.3%), but only the first strain was virulent to the mealybug Planococcus sp. (80.9%). Akanthomyces lecanii CG824 was weakly virulent to all tested insects. None of the strains were pathogenic to the thrips Caliothrips phaseoli, and all strains showed low virulence to the wooly whitefly Aleurothrixus floccosus and the scale Duplachionaspis divergens. Submerged liquid fermentation yields varied from 1.72 × 109 (day 2) to 3.90 × 109 (day 5) blastospores mL-1. Blastospores or aerial conidia from A. muscarius CG935, at a single concentration of 1 × 107 viable propagules mL-1, resulted in 67.5-83.1% mortality of B. tabaci nymphs within 8 days post-treatment. Overall, these results encourage additional studies that could lead to the development of new mycopesticides based on Akanthomyces strains.

Diversity of anamorphic Cordyceps (formerly Isaria) isolated from Brazilian agricultural sites.

A total of 53 anamorphic strains of Brazilian Cordyceps species currently maintained in a government-owned culture collection, were reassessed for diversity and species identity using multi-loci-based phylogenetic methods. The strains used in this study were originally obtained from soil samples or were isolated from insects of the orders Hemiptera, Lepidoptera, Coleoptera and Diptera, mostly from agricultural sites. A Bayesian phylogenetic tree was constructed based on a concatenation of five loci (ITS, LSU, RPB1, RPB2 and TEF). In a few cases of ambiguity, morphological traits were also considered for species delimitations. Considerable variability within the set of strains was detected and six Cordyceps species were identified: C. amoenerosea, C. fumosorosea, C. javanica, C. tenuipes and, for the first time, C. brevistroma and C. spegazzinii are reported in Brazil. Four other taxonomically equivocal groups, closely related to other known taxa (C. amoenerosea, C. cateniannulata, C. polyarthra and C. spegazzinii), were also recognized, although further studies will be required to confirm their identifications or their descriptions as new species. Cordyceps javanica was the most common species in our dataset, originally isolated from soil and several different insect orders, and includes 17 strains from the whitefly, Bemisia tabaci. Interestingly, strains previously identified as C. fumosorosea based on morphology and growth characteristics, were shown to be C. javanica, including the active ingredients of some commercial mycoinsecticides. Cordyceps farinosa, usually mentioned in the literature as occurring in Brazil, was not found in our study. Since most strains were from insect crop pests, further studies with hosts from non-agricultural settings or from environmental samples would be advisable for a deeper understanding of the occurrence of anamorphic Cordyceps in Brazil.

On-farm Production of Microbial Entomopathogens for use in Agriculture: Brazil as a Case Study.

In Brazil, the production of beneficial microorganisms by growers exclusively for their own use is a practice known as "on-farm production". Regarding on-farm bioinsecticides, they were initially deployed for pests of perennial and semi-perennial crops in the 1970s but, since 2013, their use has extended to pests of annual crops such as maize, cotton, and soybean. Millions of hectares are currently being treated with these on-farm preparations. Local production reduces costs, meets local needs, and reduces inputs of environmentally damaging chemical pesticides, facilitating establishment of more sustainable agroecosystems. Critics argue that without implementation of stringent quality control measures there is the risk that the on-farm preparations: (1) are contaminated with microbes which may include human pathogens or (2) contain very little active ingredient, impacting on field efficacy. The on-farm fermentation of bacterial insecticides predominates, especially that of Bacillus thuringiensis targeting lepidopteran pests. However, there has been a rapid growth in the past 5 years in the production of entomopathogenic fungi, mostly for the control of sap-sucking insects such as whitefly (Bemisia tabaci (Gennadius)) and the corn leafhopper (Dalbulus maidis (DeLong and Wolcott)). In contrast, on-farm production of insect viruses has seen limited growth. Most of the ca. 5 million rural producers in Brazil own small or medium size properties and, although the vast majority still do not practice on-farm production of biopesticides, the topic has aroused interest among them. Many growers who adopt this practice usually use non-sterile containers as fermenters, resulting in poor-quality preparations, and cases of failure have been reported. On the other hand, some informal reports suggest on-farm preparations may be efficacious even when contaminated, what could be explained, at least partially, by the insecticidal secondary metabolites secreted by the pool of microorganisms in the liquid culture media. Indeed, there is insufficient information on efficacy and mode of action of these microbial biopesticides. It is usually the large farms, some with > 20,000 ha of continuous cultivated lands, that produce biopesticides with low levels of contamination, as many of them possess advanced production facilities and have access to specialized knowledge and trained staff. Uptake of on-farm biopesticides is expected to continue but the rate of adoption will depend on factors such as the selection of safe, virulent microbial strains and implementation of sound quality control measures (compliance with emerging Brazilian regulations and international standards). The challenges and opportunities of on-farm bioinsecticides are presented and discussed.

Potential Impact of Chemical Fungicides on the Efficacy of Metarhizium rileyi and the Occurrence of Pandora gammae on Caterpillars in Soybean Crops.

Entomopathogenic fungi may play a crucial role in the regulation of caterpillar populations in soybean crops, either through natural occurrences or applied as mycopesticides. In the present work, we reported the naturally occurring entomopathogenic fungus Pandora gammae attacking the caterpillar Chrysodeixis includens, with infection rates in field trials ran in two consecutive years in the 10-35% range. As many chemicals are potentially harmful to entomopathogenic fungi, this work aimed to investigate the potential impact of two chemical fungicides (azoxystrobin + benzovindiflupyr and trifloxistrobina + prothioconazole) used to control soybean rust (Phakopsora pachyrhizi) on the natural occurrence of P. gammae and Metarhizium rileyi, as well as the efficacy of the latter fungus applied as different formulations against the soybean caterpillars Anticarsia gemmatalis and C. includens. Under laboratory conditions, fungicides used at field-recommended rates had a considerable negative impact on the germinability of M. riley on the medium surface, and all tested formulations did not protect conidia from damage by these chemicals. This harmful effect also impacted host infectivity, as the larval mortality owing to this fungus was reduced by 30-40% compared to that of the fungicide-free treatments. In field trials conducted in two subsequent years, unformulated and formulated M. rileyi conidia applied to soybean plants produced primary infection sites in caterpillar populations after a single spray. Spraying unformulated or formulated M. rileyi conidia following fungicide application on plants did not affect host infection rates over time. Moreover, the use of M. rileyi-based formulations or chemical fungicide did not interfere with the natural infection rates by P. gammae on its host, C. includens. Although a higher degree of exposure to non-selective fungicides can negatively affect fungal entomopathogens, a single foliar application of fungicides may be harmless to both M. rileyi and P. gammae in soybean fields. Additionally, this work showed that naturally occurring wasps and tachnids also play an important role in the regulation of A. gemmatalis and, notably, C. includens, with parasitism rates above 40-50% in some cases.

Natural occurrence of Beauveria caledonica, pathogenicity to Cosmopolites sordidus and antifungal activity against Fusarium oxysporum f. sp. cubense.

BACKGROUND: Entomopathogenic fungi can provide a set of ecological services, such as suppressing arthropod pests and plant pathogens. In this study, novel indigenous Beauveria caledonica (Bc) strains were isolated from naturally infected banana weevils (Cosmopolites sordidus) occurring in commercial banana plantations in Brazil. RESULTS: The prevalence of infection by Bc strains on field-caught C. sordidus ranged from 1.3% to 12.9%. Similar to the Beauveria bassiana strains tested, none of the Bc strains caused more than 50% weevil mortality at a concentration of 1 × 108 conidia ml-1 . Bc strain CMAA1810 caused the highest mortality in C. sordidus and had enhanced insecticidal activity when formulated with an emulsifiable oil. In paired co-culture assays, this same strain showed a significant growth-inhibitory effect on the causal agent of Fusarium banana wilt (Fusarium oxysporum f. sp. cubense, Foc) of twofold magnitude compared with the control. Cell-free crude filtrates derived from the red-pigmented culture broth of Bc (CMAA1810) strongly reduced Foc conidial viability, and this inhibitory activity was inversely related to the age of the Bc culture. Crude concentrated filtrates from 4-day-old cultures exhibited the strongest antifungal activity (13-fold) compared with untreated Foc conidia. The abundant compound identified in the crude filtrate of Bc was oosporein (1,4-dibenzoquinone) present at a concentration of 0.829 ± 0.018 mg g-1 dry matter, and the antifungal activity of the filtrate was demonstrated. CONCLUSION: These results indicated that Bc strains might have the potential to manage both C. sordidus and Foc, two of the major phytosanitary problems in banana crops worldwide. Further research under field conditions using suitable formulations of virulent Bc strains in combination with the metabolite oosporein is needed to evaluate their efficacy in the management of C. sordidus and Foc in banana plantations. © 2022 Society of Chemical Industry.

Evaluation of key parameters for developing a Metarhizium rileyi-based biopesticide against Spodoptera frugiperda (Lepidoptera: Noctuidae) in maize: laboratory, greenhouse, and field trials.

BACKGROUND: The entomopathogenic fungus Metarhizium rileyi is known to cause natural infections in some caterpillars. In this multiyear study, we carried out laboratory, greenhouse and three field trials with the aim of developing a sustainable option for control of the fall armyworm (FAW) in maize. Association of M. rileyi with Spodoptera frugiperda baculovirus (SfMNPV) and delivery strategies were also investigated. RESULTS: The selected isolate (CG381) was effective in the laboratory at a low concentration (~ 4 × 103 conidia cm-2 ), killing >95% of FAW larvae within 8 days. In the greenhouse assay, applications of conidia suspended in water or as dry powder in maize whorls (~ 6.3 × 106 conidia per plant) produced similar larval mortalities (88%-96%). In the field trials, conventional spraying of unformulated conidia (0.6 and 1.2 × 1012 conidia ha-1 ) caused low larval mortalities (27-31%). Simultaneous application of either unformulated or oil-based formulations of M. rileyi conidia and S. frugiperda baculovirus (SfMNPV) to plant rows caused larval mortalities comparable with each of the pathogens applied alone. However, when a formulation containing both pathogens was sprayed directly into the whorls, the overall mortality of S. frugiperda larvae due to pathogens (mostly to M. rileyi infections) reached ~ 59%, twofold higher than the other treatments with conventional spraying, whereas in the control, the overall mortality was only 1%. CONCLUSION: Optimizing exposure of S. frugiperda larvae to an inoculum of virulent entomopathogens through directed applications to maize whorls is critical to produce satisfactory mortality levels and is promising for integrated pest management.

First report on the natural occurrence of entomopathogenic fungi in populations of the leafhopper Dalbulus maidis (Hemiptera: Cicadellidae): Pathogen identifications and their incidence in maize crops.

The corn leafhopper Dalbulus maidis is one of the most important pests of maize in Latin America. Here we report, for the first time, the natural occurrence of two fungal species infecting the adult stage of this pest. In 2020, insects killed by a pale bluish green fungus in irrigated maize fields located in Northeast Brazil were found attached to the abaxial surface of leaves. Using morphological characters and multigenic phylogeny, it was identified as Metarhizium brasiliense. In the beginning of 2021, the same pathogen was seen on adults in a maize field in the Central-Western region, alongside an entomophthoralean fungus during an epizootic. The latter pathogen was molecularly identified as a species in the genus Batkoa. The number of Batkoa-infected leafhoppers, displaying the typical swollen abdomen and extended wings, reached an average of 1.88 per maize leaf (86.42% of the sampled adults). The incidence of M. brasiliense was higher in plots in the Northeastern region (0.22 and 0.53 adult per leaf) when compared to the Central-Western region (0.04 adult per leaf). The report of D. maidis adults infected by M. brasiliense in agricultural settings located in different geographic regions and over 550 km apart indicates probable widespread occurrence of this pathogen in Brazil. Moreover, this opens the possibility of more applied biological control studies and, perhaps, the development of new tools to manage D. maidis populations.

Field Efficacy of Metarhizium rileyi Applications Against Spodoptera frugiperda (Lepidoptera: Noctuidae) in Maize.

The efficacy of the fungus Metarhizium rileyi (Farlow) Kepler, S. A. Rehner & Humber (Hypocreales: Clavicipitaceae) against the fall armyworm Spodoptera frugiperda (J.E. Smith) (Lepidoptera: Noctuidae) and the impact on the community of epigeans predators as well as the natural occurrence of native fungal strains were evaluated after sequential sprays in maize. In the first season, the infestation was lower than 0.2 larvae per plant throughout the three fungal sprays and no significant difference was observed in the number of infected larvae among treatments. In the second season, insect population reached an average of 0.8 larvae per plant between the second and third sprays and the number of larvae succumbed by M. rileyi was higher in fungal-treated plots in the following weeks. Molecular characterization of the collected isolates on mummified larvae revealed the introduction of a new isolate did not eliminate the native strain on the fungal-treated plots. In both seasons, the diversity and abundance of predator species in the plots were not affected by any of the treatments. We showed that applications of selected strains of M. rileyi early in the season have no significant impact on the occurrence of natural enemies in maize.

Assessment of the diversity of Brazilian entomopathogenic fungi in the genus Beauveria.

We combined matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) along with sequencing of the B locus intergenic region (Bloc) to assess the diversity of Brazilian species within the anamorphic genus Beauveria. A total of 121 strains maintained in a government-owned culture collection and isolated from a range of hosts/substrates over a long time span (1981-2015) were assessed. Strains were collected in five of six Brazilian biomes, mostly in the Atlantic Forest (42.2%) and Cerrado (29.8%), primarily from insect pests of crops. All strains were subjected to MS, and those not accurately identified by this technique were genomically analyzed. Among the outcomes of this study, four taxa from the genus Beauveria were recognized, with the great majority of strains belonging to B. bassiana s.str. (93.4%), followed by B. caledonica (2.5%), B. pseudobassiana (2.5%) and B. amorpha (1.6%). B. bassiana s.str. was found in all biomes and isolated from a wide range of hosts/substrates. Due to low numbers, associations of the remaining Beauveria species with specific hosts or habitats/biomes were not clear, except that all three B. caledonica strains were found only in the Cerrado biome and were associated with adults of the banana weevil, Cosmopolites sordidus (Col.:Curculionidae). B. pseudobassiana is reported for the first time on the South American continent, in a subtropical region and from two insect orders not yet associated with this taxon. We also showed that some strains previously ascribed to B. brongniartii were misidentifications. The biodiversity of Beauveria analyzed in our study was comparatively low. The geographic origins of strains used in our study were biased towards biomes with intense human interventions. Future surveys on more conserved, less environmentally disturbed biomes, such as Caatinga, Pampa, Pantanal, and Amazon are needed for a more comprehensive picture of the diversity of Beauveria and related genera in Brazil.

Laboratory and field evaluation of an autoinoculation device as a tool to manage poultry red mite, Dermanyssus gallinae, infestations with Beauveria bassiana.

The poultry red mite, Dermanyssus gallinae, is a cosmopolitan ectoparasite in hens and has been considered an important threat to the egg production industry. This study evaluated an alternative to manage poultry red mite populations as a complement to conventional chemical treatments and other control strategies in poultry houses. A simple autoinoculation device prepared with corrugated cardboard (CB) or loofah sponge (LS) as inert supports to anchor Beauveria bassiana conidia was used to aggregate and infect mites from infested poultry houses. In the laboratory, mites gathered inside the traps and the average mortalities by the fungus were higher than 70% in CB and LS traps after 5 and 4 days of exposure, respectively. Conidial viability was around 80% in CB and LS traps after 14 and 60 days under unrefrigerated conditions (26 °C), respectively. Both trap types tied to hen cages efficiently captured fed mites after blood meal, and fungal infection was observed in 65-90% of the mites in field tests. Between 5 and 25% of the mites recaptured in monitoring cardboard traps installed immediately after CB and LS removal were infected by B. bassiana. According to our results, the use of B. bassiana in an autoinoculation strategy is a potential alternative method for D. gallinae control.

Within-host interactions of Metarhizium rileyi strains and nucleopolyhedroviruses in Spodoptera frugiperda and Anticarsia gemmatalis (Lepidoptera: Noctuidae).

Members of the family Baculoviridae have been quite successfully used as biocontrol agents against some lepidopterans. Likewise, a number of fungi are important natural enemies of these pests. An interesting approach to increase control efficacy could be the combination of a given nucleopolyhedrovirus (NPV) and a fungus, since they possess distinct modes of action. As a first step towards this goal, we assessed the interaction between NPV (either AgMNPV-79 or SfMNPV-6nd) and the entomopathogenic fungus Metarhizium rileyi (either CG1153 or CG381), using Anticarsia gemmatalis and Spodoptera frugiperda as hosts. In sequential applications of these pathogens, per os inoculation of an NPV (leaf discs with 2.5 × 104 occlusion bodies) either two days before or two days post-spraying of its counterpart fungal strain (5 × 103 conidia.cm-2 sprays) usually resulted in an antagonistic effect. When both pathogens were simultaneously applied at different combined dosages, usually an additive effect was seen. Interestingly, a number of dead larvae showing signs of co-infections (partially with soft integument and partially mummified) were recorded. However, mixes with lower dosages of both pathogens did not cause significantly higher insect mortalities compared to low dosages of the fungus applied alone. The advantages and disadvantages of the simultaneous applications of NPV and M. rileyi aiming at the management of either A. gemmatalis or S. frugiperda were discussed.

Current status and perspectives of fungal entomopathogens used for microbial control of arthropod pests in Brazil.

Entomopathogenic fungi play a central role in Brazil's biopesticide market. Approximately 50% of registered microbial biopesticides comprise mycoinsecticides and/or mycoacaricides consisting of hypocrealean fungi, with most based on Metarhizium anisopliae sensu stricto (s. str.) and Beauveria bassiana s. str. These fungi are mainly used to control spittlebugs in sugarcane fields and whiteflies in row crops, respectively, with annual applications surpassing three million hectares. Research also emphasizes the potential of fungal entomopathogens to manage arthropod vectors of human diseases. Most registered fungal formulations comprise wettable powders or technical (non-formulated) products, with relatively few new developments in formulation technology. Despite the large area treated with mycoinsecticides (i.e., approx. 2 million ha of sugarcane treated with M. anisopliae and 1.5 million ha of soybean treated with B. bassiana), their market share remains small compared with the chemical insecticide market. Nevertheless, several major agricultural companies are investing in fungus-based products with the aim at achieving more sustainable IPM programs for major pests in both organic and conventional crops. Government and private research groups are pursuing innovative technologies for mass production, formulation, product stability and quality control, which will support cost-effective commercial mycoinsecticides. Here, we summarize the status of mycoinsecticides currently available in Brazil and discuss future prospects.

Increased tolerance of Beauveria bassiana and Metarhizium anisopliae conidia to high temperature provided by oil-based formulations.

The influence of the temperature of aqueous conidial sprays on conidial viability and virulence against Diatraea saccharalis was evaluated for pure conidia, rice + fungus (technical concentrates) and oil-based formulations of Beauveria bassiana s.s. and Metarhizium anisopliae s.s. under laboratory conditions. The fungal preparations were suspended in water and maintained at 26 °C, 36 °C and 46 °C for one, four and six hours. Conidial viability was determined by plating aliquots of each suspension onto PDA medium followed by incubation for 20-22 h and observing for viable conidia (germ tubes longer than diameter of conidia). Fungal virulence was determined by spraying suspensions onto third-instar larvae of D. saccharalis. In general, germination and virulence, particularly for unformulated conidia, were negatively affected by increases in water temperature and exposure time in suspension. However, the decrease in conidial viability in the oil-in-water emulsion was less than 7% for both species after 6 h of exposure at 36 °C, in contrast to reductions of 7-21% and 28-60% for the oil-free suspensions of B. bassiana and M. anisopliae, respectively. For the sprays of conidia in an oil-in-water emulsion previously exposed to elevated water temperatures for longer periods, the levels of insect mortality were higher than those of pure conidia or technical concentrates under identical conditions. Our results indicate that emulsifiable oil-based formulations can protect the conidia of both species of fungi from the adverse effects of high water temperatures before spraying in the field.

A natural fungal infection of a sylvatic cockroach with Metarhizium blattodeae sp. nov., a member of the M. flavoviride species complex.

A wild, forest-dwelling cockroach from the subfamily Ectobiidae (order Blattodea) in a nature reserve in Cavalcante, in the state of Goiás, Brazil, was found to be infected by a new, genetically distinct species in the Metarhizium flavoviride species complex that we describe here as Metarhizium blattodeae. The status of this fungus as a new species is supported by both multigenic sequence comparisons and protein profiles generated by MALDI-TOF (matrix-assisted laser desorption/ionization time-of-flight) mass spectrometry. This is one of the first reports of a naturally occurring fungal pathogen affecting any sylvatic (forest-dwelling) cockroach from any part of the world. M. blattodeae caused up to 96 % mortality of Periplaneta americana nymphs (a serious peridomestic cockroach species) after 10 d.

Conidial vigor vs. viability as predictors of virulence of entomopathogenic fungi.

We tested the hypothesis that debilitated conidia exhibiting slow-germination (requiring>16h to germinate) are less virulent than vigorous conidia exhibiting fast germination (requiring⩽16h to germinate). Preparations of Beauveria bassiana s.l. strain CG 1027 with variable ratios of vigorous to debilitated conidia were assayed against third-instar larvae of Spodoptera frugiperda. As the proportion of debilitated conidia in test preparations increased, LC50 expressed in terms of total viable conidia increased, while LC50 expressed solely in terms of vigorous conidia remained constant, indicating that vigorous conidia were responsible for nearly all mortality observed in the assays. Larvae treated with conidia from low-quality batches (with high proportions of debilitated conidia) survived consistently longer than those treated with comparable doses of conidia from high-quality batches. These results confirm our previous hypotheses that inclusion of debilitated conidia in viability assessments can lead to overestimation of the quality (potency) of mycoinsecticide preparations and support our recommendation for use of short incubation periods for assessing viability whenever viability is relied upon as an indicator of product quality.

MALDI-TOF mass spectrometry applied to identifying species of insect-pathogenic fungi from the Metarhizium anisopliae complex.

Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) has proven to be a powerful tool for taxonomic resolution of microorganisms. In this proof-of-concept study, we assessed the effectiveness of this technique to track the current gene sequence-based phylogenetic classification of species in the Metarhizium anisopliae complex. Initially the phylogenetic analysis of 5' strains by sequencing of the 59' end of the TEF-1α gene region revealed seven species within M. anisopliae sensu lato and two varieties outside this complex. Because initial studies on MS profiles from different cell types showed that mycelial fragments or conidia produced on nutrient-poor medium may yield too much background noise, all subsequent spectrometric analyses were performed with acidhydrolyzed conidia from 10-12 d old PDA cultures. The initial MALDI-TOF reference library included protein spectral profiles from nine taxonomically distinct, molecularly identified isolates sharing high genetic homology with the ex-type or ex-epitype isolates of these taxa in Metarhizium. A second reference library added one isolate each for M. anisopliae sensu stricto and M. robertsii. The second, larger reference library (including 11 taxa) allowed nearly perfect MALDI-TOF matching of DNA-based species identification for the 40 remaining isolates molecularly recognized as M. anisopliae sensu stricto (n = 19), M. robertsii (n = 6), M. majus (n = 3), M. lepidiotae (n = 1), M. acridum (n = 3), M. flavoviride var. pemphigi (n = 1), plus seven unidentified strains (six of them phylogenetically close to M. anisopliae sensu stricto and one outside the Metarhizium pingshaense-anisopliae-robertsii-brunneum clade). Due to the increasing frequency of phylogenetically (genomically) based taxonomic revisions of fungi, this approach is especially useful for culture collections, because once the protein profiles of Metarhizium isolates are obtained taxonomic updating of MALDI-TOF library data is easily accomplished by comparing stored profiles with those of newly proposed taxa.

Falsa broca aumenta disseminação de Beauveria bassiana em populações de campo da broca-do-rizoma da bananeira / West Indian Sugarcane Weevil enhances dissemination of Beauveria bassiana on field populations of the banana weevil

A transmissão horizontal de fungos entomopatogênicos tem papel importante na epizootiologia, pois está diretamente associada à disseminação do agente causal da doença de insetos. Em hospedeiros que apresentam restrita mobilidade, a presença de pragas secundárias no agroecossistema pode ajudar na formação de focos secundários da doença e, com isso, aumentar a mortalidade das pragas-chave. Para tanto, a disseminação de Beauveria bassiana, introduzida inoculativamente através de iscas tipo telha, foi avaliada em um bananal infestado por Cosmopolites sordidus (Piracicaba) e outro por C. sordidus e Metamasius hemipterus (Botucatu). Constatou-se que, em Piracicaba, não foram capturados insetos infectados na testemunha, sendo os índices de infecção nas parcelas que receberam B. bassiana de 8 e 15 por cento. Em Botucatu, o índice de infecção dos insetos capturados na testemunha chegou a 17 por cento, enquanto que, nas parcelas tratadas, variou de 28 a 54 por cento. Os resultados obtidos neste trabalho demostram que a presença de M. hemipterus influenciou na disseminação de B. bassiana para a população de C. sordidus, praga-chave dessa cultura.

Horizontal transmission of entomopathogenic fungi plays an important role in the epizootiology, because it is directly associated with the spread of causal agents of insect diseases. In hosts with restricted mobility, the presence of secondary pests in the agroecosystem can help in the formation of secondary disease outbreaks and increase the mortality of the target pest. For this purpose, the dissemination of Beauveria bassiana inoculated using tile-type baits was evaluated in a banana plantation infested with Cosmopolites sordidus (Piracicaba) and one with C. sordidus and Metamasius hemipterus (Botucatu). In Piracicaba, insects contaminated with the pathogen were not found in the control group, and infection rates in the plots that received B. bassiana ranged between 8 and 15 percent. In Botucatu, the infection rate of insects in the control reached 17 percent, whereas in the fungal treatment plots, it varied from 28 to 54 percent. The present results demonstrate the influence of M. hemipterus in the dissemination of B. bassiana in populations of C. sordidus, a target pest of this crop.

Culture media selection for mass production of Isaria fumosorosea and Isaria farinosa

This work investigated the production of the fungi Isaria fumosorosea and Isaria farinosa in biphasic fermentation using agro-industrial products and residues. Combinations of natural liquid substrates, alternative to the complete medium and potato dextrose medium, were evaluated. The best liquid media were sugarcane molasses + rice broth, rice broth + yeast and sugarcane molasses + yeast + rice broth, which resulted in the highest viable propagule concentration. The molasses + rice broth medium was selected for the next phase of the study in which the production of both fungal isolates was evaluated in solid grain substrates. In solid-state fermentation, the best conidia production was achieved with the soybean meal and broken corn for I. farinosa, and whole rice and broken rice for I. fumosorosea. Results demonstrated that the two fungal species could be rapidly produced with higher yield of conidia on agro-industrial resources by using biphasic fermentation techniques.

Estudou-se a produção dos fungos Isaria fumosorosea (ESALQ1296) e Isaria farinosa (ESALQ1205) em fermentação líquida e sólida utilizando produtos e resíduos agroindustriais. Inicialmente, avaliaram-se combinações de substratos líquidos naturais e alternativos aos meios completo e batata-dextrose. Os meios líquidos mais promissores foram: melaço + caldo de arroz, caldo de arroz + levedura e melaço + levedura + caldo de arroz, pois resultaram nas maiores produções de propágulos viáveis. O meio melaço + caldo de arroz foi selecionado para etapa seguinte do trabalho, na qual se avaliou a produção dos fungos em grãos. A fermentação sólida foi mais eficiente quando se utilizaram os meios de farelo de soja e milho quirela para I. farinosa,e arroz inteiro e arroz quirela para I. fumosorosea. A fermentação bifásica para os fungos estudados a partir desses produtos agroindustriais selecionados proporcionou elevada produção de conídios em curto intervalo de tempo, podendo ser implantada em escala industrial.

Beauveria bassiana yeast phase on agar medium and its pathogenicity against Diatraea saccharalis (Lepidoptera: Crambidae) and Tetranychus urticae (Acari: Tetranychidae).

Beauveria bassiana colonizes insect hosts initially through a yeast phase, which is common in some artificial liquid cultures, but not reported on artificial solid media. We describe a yeast-like phase for B. bassiana isolate 447 (ATCC 20872) on MacConkey agar and its virulence toward Diatraea saccharalis and Tetranychus urticae. The yeast-like cells of B. bassiana developed by budding from germinating conidia after 24-h incubation. Cells were typically 5-10 microm and fungal colonies were initially circular and mucoid, but later were covered with mycelia and conidia. Ability to produce yeast-like cells on MacConkey medium was relatively common among different B. bassiana isolates, but growth rate and timing of yeast-like cell production also varied. Metarhizium anisopliae and Paecilomyces spp. isolates did not grow as yeast-like cells on MacConkey medium. Yeast-like cells of B. bassiana 447 were more virulent against D. saccharalis than conidia when 10(7)cells/ml were used. At 10(8)cells/ml, the estimated mean survival time was 5.4 days for the yeast suspension and 7.7 days for the conidial suspension, perhaps due to faster germination. The LC(50) was also lower for yeast than conidial suspensions. Yeast-like cells and conidia had similar virulence against T. urticae; the average mortalities with yeast-like cells and conidia were, respectively, 42.8 and 45.0%, with 10(7)cells/ml, and 77.8 and 74.4%, with 10(8)cells/ml. The estimated mean survival times were 3.6 and 3.9 for yeast and conidial suspensions, respectively. The bioassay results demonstrate the yeast-like structures produced on MacConkey agar are effective as inoculum for B. bassiana applications against arthropod pests, and possibly superior to conidia against some species. Obtaining well-defined yeast phase cultures of entomopathogenic hyphomycetes may be an important step in studies of the biology and nutrition, pathogenesis, and the genetic manipulation of these fungi.