This study was conducted to evaluate the efficacy of mulberry leaf mixed diet on larval growth of Protaetia brevitarsis. As a result of adding mulberry leaves to oak fermented sawdust, the survival rate of P. brevitarsis larvae were higher than those of the control when mass rearing larvae. When fed oak fermented sawdust with 5%, 10% and 20% of mulberry leaves, the cumulative proportion of larvae over 2.5g was 93.1% at 65 days, 95.6% at 55 days, and 93.9% at 55 days after rearing at 25±1℃, respectively. Also, heavy metals were not detected in larvae of P. brevitarsis fed on mulberry leaves.

Insect killing fungus Beauveria bassiana has been widely studied as a biological control agent. However, many studies have been focused on lab or field-based management. Herein this work, comparison of three B. bassiana strains was investigated under a molecular level. The whole genome sequences of ERL836, JEF-007 were analyzed by PacBio (35.5 Mb of ERL836 and 36.5 Mb of JEF-007) and ARSEF2860 referenced from GenBank (33.7 Mb). To compare the three strains, virulence, thermotolerance and chemical resistance were assayed. The transcriptomes of non-infecting B. bassiana and infecting B. bassiana against western flower thrips were analyzed using RNA-seq. This work can provide that genome features, functions, morphology and gene expression could be different under the molecular level, even if in the same species.

Melon thrips, Thrips palmi Karny (Thysanoptera: Thripidae) is a worldwide polyphagous pest. The management of this pest have mainly relied on chemical agents. However, the overuse is harmful to the environment and results in insects resistance. Therefore, alternative eco-friendly control methods that have different mode of action, such as biological control, are necessary to overcome the current issue. In this study, we isolated entomopathogenic fungi from Korean soil, and characterized them via morphological and molecular techniques and pathogenicity assay against Tenebrio molitor larvae. The isolated fungi were screened for virulence against T. palmi under laboratory conditions and the results were used to establish a thrips-pathogenic fungal library. The highly virulent isolates were selected and further characterized for optimum culture conditions and application as biopesticide in the field.

The global biopesticide market was estimated to become about 4% of the total crop protection market in 2015, mainly due to variability of their efficacy, narrow spectrum or difficulties in long-term storage. Therefore, many people focus on overcoming these issues as a big trend. Suggested solutions include the investigation of synergy between microorganisms, the use of genetic engineering, improving the pesticide life shelf, etc. As a result, biopesticides market has grown by more than 17% over the last decade. In this context and aiming to develop new entomopathogenic fungi–based pest management tools, we constructed a fungal library by isolating insect pathogenic fungi from soil. A total of 581 isolates belonging to 35 species were isolated and characterized. Beauveria bassiana was the most abundant, representing 38.55% of the total strains, followed by Metharizium anisopliae (22.55%) and bubillosa (8.6). …% of the total isolates were highly virulent against Tenebrio molitor killing most of the treated insects in 2 to 3 days.

Japanese pine sawyer, Monochamus alternatus, is the main pest that mediates pine wilt nematode, Bursaphelenchus xylophilus, that causes serious damage to pine forests. In this study, we studied the strategy to control M. alternatus using entomopathogenic fungi. The fungi were collected from soil by an insect-baiting method and two fungal isolates (Metarhizium anisopliae JEF-197 and JEF-279) showed high virulence against M. alternatus. The Metarhizium isolates were evaluated for insecticidal activity against M. alternatus by spray treatment on live pine trees and wintering trees, and the M. anisopliae JEF-197 showed high insecticidal activity. In addition, the interaction of fungi and M. alternatus were analyzed by RNA-seq. This result can contribute to the development of insect control agents using entomopathogenic fungi.

Beauveria bassiana is widely studied for its potential as biopesticide. However, little is known about the factors to influence genetic diversity among isolates. In this work, we aimed to study the gene diversity of 42 isolates to figure out the impact of sequences variability on various biological features. The accumulated data showed no correlation between sequence variability and the fungal geographic localization. Meanwhile, weak to strong correlations have been obtained between the sequence diversity of various genes and the studied biological features. Interestingly, Biotrophy-associated gene 2 gene variability was correlated with all the studied biological features, being an important marker to determine isolates of interest for biological control. Therefore, the obtained data would serve as a database to focus on interesting genes related to improving or screening highly virulent isolates as biological control agents.

A species of Beauveria bassiana is widely used for biological pest management in many countries. Many efforts have been given to figure out the clear fungal mode of action to enhance the insecticidal activity. Homologous recombination (knock-out) or hairpin RNA (knock-down) is popularly used in fungal gene function study, but gene cloning and generation of knock-out or -down mutants takes long time or temporarily knock-downed. Here in this work, we used previously generated egfp-expressing B. bassiana strain (Bb-egfp #3) and integrated dsegfp to the Bb-egfp #3 using a protoplast integration method. This work suggests that protoplast integration with dsRNA possibly generate significantly reduced gene expression in B. bassiana and the reduction is quite stable over generations which provide easy of functional study for fungal mode of action.

The serious emergence of chemical-mediated residual toxicity and insect resistance have been enforced the regulation of synthetic pesticides. Future decisions to select more realistic control options probably depend on the speed of technological development in chemical and biological pesticides. Now, a strategic collaboration between synthetic pesticides and biopesticides has been progressed, such as distribution and R&D in collaboration and M&A for obtaining microbial resources. Recently registered microbial pesticides are entomopathogenic fungi in pest management. A concept of e-biopesticide could be properly combined with digital agriculture and accelerate the use of biological control agents in the future farming.

The longhorned tick, Haemaphysalis longicornis, is one of the vectors of severe fever with thrombocytopenia syndrome virus (SFTSV) in human. The use of pyrethroid insecticides induced pest resistance and environmental residual toxicity. Here in this work, our interest was given to the selection of highly virulent fungi against longhorned tick. A total of 101 fungal pathogens were assayed by dipping the nymph stage of ticks into a conidial suspension. Interestingly of the several species, one species showed high virulence and mycosis were observed in 7-15 days. Highly virulent strains were selected, and semi-field experiments were conducted. As a result, the control efficacy of the isolate was over 80% at 30 days of treatment. This work suggests that entomopathogenic fungi could be used to effectively control longhorend ticks.

Silverleaf whitefly, Bemisia tabaci is a worldwide agricultural pest that cause serious damage to crops. However, since this insect developed resistance to variety of chemicals, alternative control strategy needs to be studied. In this study, we aimed to evaluate the potential of entomopathogenic fungi as biological control tools against this pest. We developed a dipping method in laboratory conditions to screen insecticidal activity of different fungal species and isolates against the nymphal stage of whitefly. The highly virulent isolates were characterized and 2 isolates were further selected based on their high conidial productivity and thermotolerance. Finally, several grain substrates were used to confirm the conidial productivity and thermotolerance of the selected isolates, revealing optimum conidial production when the fungi were cultured on millet grains. Both isolates could be used in further millet grain-based formulations to control the whitefly in fields.

Red mite, Dermanyssus gallinae is one of the serious pests, damaging to the egg production in chickens. Moreover, the overuse of chemical insecticides caused pest resistance and environmental residual toxicity. Therefore, this work provides a screening method to select entomopathogenic fungi as a candidate of environmentally safe control agent, having high miticidal activity against D. Gallinae. The virulence test was conducted using a spraying method. Of the several species, Beauveria species showed high virulence and mycosis were observed. Some isolates were produced on eight cereal grains in Petri dish conditions. Millet, perilla seed and barley showed the high conidia production. Conidia of the isolates produced on millet and rice showed high thermal stability, when exposed to 45℃. Based on these results, this work suggests that entomopathogenic fungi could be used to control the D. Gallinae.

In order to develop the D. adamsi breeding technique that is highly likely to be used as an emotional pet insect, the results of the D. adamsi was collected in the large exhibition Jang Tae-san in Daejeon to the indoor breeding began to spawn after 10 days after the training, the egg laying period was 33.6 days, the average number of spawn per female was 27.7, and the period was 11.8 days. D. adamsi larvae bred with food of oak-boiled molted two times, the duration of 1st larvae development was 11.8 days, 2nd larvae 14.5 days, and the third larvae was 29.4, and the larvae were used to build houses using Masato(soil) and in the pupae, and then in April of the following year. D. adamsi has a characteristic to build a house with fallen leaves and spawn one or two eggs in the fallen leaves, and the results of the spawn mat test using fallen leaves, leaf mold and a sawdust, etc., and the number of spawn was high in the Masato was spread about 3 ~ 5cm, and a fallen leaves on a 10 cm or so, and the growth and survival rate of the larvae were also high. Larvae breeding density was 2 ~ 3 ℓ in the container size to raise the object was normal growth, the higher the density mortality was high and the growth was sluggish. Larvae feeding conditions were normally developed in dry fallen leaves and fermented fallen leaves, compared to the sterile fallen leaves, oak sawdust, sterile fallen leaves and oak sawdust was abnormal, the mortality rate was higher than 50%.

Insect-killing fungi have high potential in pest management. A deeper insight into the fungal genes at the whole genome level is necessary to understand the inter-species or intra-species genetic diversity of fungal genes, and to select excellent isolates. In this work, we conducted a whole genome sequencing of Beauveria bassiana (Bb) JEF-007 and characterized pathogenesis-related features and compared with other isolates including Bb ARSEF2860. A large number of Bb JEF-007 genes showed high identity with Bb ARSEF2860, but some genes showed moderate or low identity. The two Bb isolates showed a significant difference in vegetative growth, antibiotic-susceptibility, and virulence against Tenebrio molitor larvae. When highly identical genes between the two Bb isolates were subjected to real-time PCR, their transcription levels were different, particularly in heat shock protein 30 (hsp30) gene which is related to conidial thermotolerance. In several B. bassiana isolates, chitinases and trypsin-like protease genes involved in pathogenesis were highly conserved, but other genes showed noticeable sequence variation within the same species. Given the transcriptional and genetic diversity in B. bassiana, a selection of virulent isolates with industrial advantages is a pre-requisite, and this genetic approach could support the development of excellent biopesticides with intellectual property protection.

Tetranychus urticae and Myzus persicae are one of the most serious insect pests in many crops, vegetables, flowers, and fruit trees worldwide. Many insecticides have been developed to control green peach aphid and two spotted spider mite, but resistance to almost all insecticides has reduced their control effect. Particular groups of plant-beneficial microbials are not only root colonizers that provide plant disease suppression, but in addition are able to infect and kill insect larvae. Antimicrobial compounds produced by biocontrol microbes are effective weapons against a vast diversity of organisms such as fungi, nematodes, and viruses. In this study, we evaluated the effectiveness of mixtures plant extracts and improvement of culture process biocontrol microbials on insecticidal activity. Azadirachta indica and Derris elliptica mixed with micorbials, which are nutrient sources of mung bean extract and lecithin, were more effective than other the mixtures. Leaf spraying with the mixture of Pseudomonas fluorescens significantly showed the highest insecticidal power in vivo for 24 hours after treatment. The effect of spraying mixture was more than 50% at 2000 times dilution, and the spraying concentration of 90% or more showed a dilution of up to 500 times. Our results indicated that the nutrient sources of microbe act as a key antimicrobial metabolite in biocontrol of insect pests, and mixing with plant extracts can provide synergistic effects as an optimal usage of the biocontrol agents.

Longhorned tick, Haemaphysalis longicornis (Ixodida: Ixodidae) is one of the vectors of severe fever with thrombocytopenia syndrome virus (SFTSV) in human. The tick occurs in mostly grass fields, and the use of pyrethroid insecticides induced pest resistance and environmental residual toxicity. Particularly the use of chemicals near residential areas where persons live become a big issue, so environmentally safe control agents need to be considered. Here in this work, our interest was given to the selection of highly virulent fungi against H. longicornis. A total of 101 fungal pathogens collected from mountainous areas were assayed by a dipping the nymph stage of ticks into a conidial suspension (1×107conidia/ml). Interestingly among several species, one species showed high virulence and mycosis were observed in 7-15 days. Most of the selected isolates produced a large amount of conidia in Italian millet, rice and millet with thermotolerance at 40-45°C. Based on these results, we selected a couple of isolates with high virulence against H. longicornis and they could be used for the control of the ticks after the set-up of a practical application strategy in fields by optimizing fungal colonization in soil and phyllosphere.

Entomopathogenic fungi have been known as promising candidates for biological control of insect pests. Recently, researchers consider the fungal thermotolerance in formulations and field applications. In this study, we investigated the production of thermotolerant Isaria javanica and I.fumosorosea conidia through grain-based solid cultures and exposure to light stress. As results, of the ten grain substrates, Italian millet, rice, perilla seed and sesame, rice, sorghum produced highly thermotolerant conidia in the strains. The two strains were exposed to a light stress and showed enhanced thermal stability compared to control, when exposed to 45°C for 2 hours. This work suggests that heatresistant entomopathogenic fungal conidia can be produced by grainbased solid cultures and exposure to light stress.

Thrips is one of the serious insect pests in many economic crops, in which general managements rely mainly on chemical insecticides. However, the overuse of chemicals led to the development of insect resistance and environmental residual issues. Therefore, thrips management requires the consideration of alternative strategies which are less harmful to the environment and working on different target points. In this work, we isolated entomopathogenic fungi from soil, and identified them with morphological and molecular biology methods, followed by a preliminary virulence assay against Tenebrio molitor larvae. Selected fungal isolates were adjusted to 1×107 conidia/ml for an indoor virulence assay against thrips adults, and highly virulent isolates were added to a thrip-pathogenic fungal library. Biological characteristics of efficacious isolates were investigated comparing to a previously commercialized fungal isolate. This entomopathogenic fungal library could be used as a valuable resource for developing effective strains to control thrips in agricultural fields.

Whitefly has a broad host range of more than 600 species, and 40 biotypes have been reported worldwide. Among the various ecological types, B and Q are the most problematic. Especially, Biotype Q, which causes serious damage in the green house, mediates more than 40 kinds of viruses, including Tomato yellow leaf cull virus (TYLCV). The development of resistance to whitefly, adverse effects on non-target organisms, and strengthened legal regulations are making chemical control difficult in application. For these reasons, the use of alternative strategies is needed to suppress the Biotype Q whitefly population. Entomopathogenic fungi were investigated and developed to control hard-control insect pest, such as whitefly. These fungi are natural pathogen of their invertebrate host and contribute to the regulation of their host population in the environment. In addition, these agents are typically harmless to humans and non-targeted organisms and do not adversely impact the environment. Therefore, in this study, we screened pathogenic fungi against whitefly using our entomopathogenic fungal library, and virulence tests were conducted with conoidal suspension (1×107conidia/ml) in laboratory conditions. As a result, 10 isolates showed 80-100% mortality at 5 days after treatment. We will analyze the characteristics of these fungal isolates and evaluate their potential as insecticides against whitefly. The selected isolates against the high mortality of whiteflies can be effectively used in integrated pest management programs for whiteflies.

Japanese pine sawyer beetle, Monochamus alternatus Hope (Coleoptera: Cerambycidae) is considered as a serious pest in pine trees. To develop an eco-friendly strategy to manage this forest insect, we collected entomopathogenic fungi from Korean soil and assessed their virulence against the adults of the insect in laboratory conditions. As a result, two isolates with conidial suspension (1.0×107conidia/ml), showed 87% and 90% mortality 12 days after fungal treatment, respectively. We assessed the potential of the fungi-derived destruxin and protease as additives to the fungal isolates, and they showed insecticidal activity via feeding and spraying treatments. Finally, we produced fungal conidia in massive solid cultures and formulated wettable powders, and now studying optimal conditions of oil-based formulation with two isolates. Based on these results, we are evaluating the control efficacy of the fungal agents against M. alternatus in field conditions.