Isolation and characterization of a native strain of the entomopathogenic fungus Beauveria bassiana for the control of the palm weevil Dynamis borassi (Coleoptera: Curculionidae) in the neotropics.

This study aimed to isolate and characterize a native strain of Beauveria bassiana, coded as Bv065, showcasing its potential as a biological control agent targeting the palm weevil Dynamis borassi. Originating from a naturally infected D. borassi specimen collected in southwestern Colombia, the fungus underwent molecular identification and was identified as B. bassiana, exhibiting high sequence similarity with known reference strains. The physiological characterization revealed that Bv065 thrived within a temperature range of 25 to 30 °C and a pH range of 6 to 9. Moreover, the key carbon sources that allow optimal growth of the strain were identified through metabolic profiling, including sucrose, D-mannose, and γ-amino-butyric acid. These findings offer strategic insights for scalability and formulation methodologies. Additionally, enzymatic analyses unveiled robust protease activity within Bv065, crucial for catalysing insect cuticle degradation and facilitating host penetration, thus accentuating its entomopathogenic potential. Subsequent evaluations exposed Bv065's pathogenicity against D. borassi, causing significant mortality within nine days of exposure, albeit exhibiting limited effectiveness against Rhynchophorus palmarum. This study underscores the importance of understanding optimal growth conditions and metabolic preferences of B. bassiana strains for developing effective biopesticides. The findings suggest Bv065 as a promising candidate for integrated pest management strategies in neotropical regions, particularly for controlling palm weevil infestations in coconut and peach palm cultivation. Future research avenues include refining mass production methodologies, formulating novel delivery systems, and conducting comprehensive field efficacy trials to unlock the full potential of Bv065 in fostering sustainable pest management practices. Overall, this study contributes to the growing body of knowledge on entomopathogenic fungi and their pivotal role in biological control, offering nuanced perspectives on eco-friendly alternatives to conventional insecticidal interventions.

Assessment of Fungal Lytic Enzymatic Extracts Produced Under Submerged Fermentation as Enhancers of Entomopathogens' Biological Activity.

The application of enzymes in agricultural fields has been little explored. One potential application of fungal lytic enzymes (chitinases, lipases, and proteases) is as an additive to current biopesticides to increase their efficacy and reduce the time of mortality. For this, a screening of lytic overproducer fungi under submerged fermentation with a chemical-defined medium was performed. Then, the enzymatic crude extract (ECE) was concentrated and partially characterized. This characterization consisted of measuring the enzymatic activity (lipase, protease and, chitinase) and determining the enzyme stability after storage at temperatures of - 80, - 20 and, 4 °C. And lastly, the application of these concentrated enzymatic crude extracts (C-ECE) as an enhancer of spores-based fungal biopesticide was proven. Beauveria were not as good producers of lytic enzymes as the strains from Trichoderma and Metarhizium. The isolate M. robertsii Mt015 was selected for the co-production of chitinases and proteases; and the isolate T. harzianum Th180 for co-production of chitinases, lipases, and proteases. The C-ECE of Mt015 had a protease activity of 18.6 ± 1.1 U ml-1, chitinase activity of 0.28 ± 0.01 U ml-1, and no lipase activity. Meanwhile, the C-ECE of Th180 reached a chitinase activity of 0.75 U ml-1, lipase activity of 0.32 U ml-1, and protease activity of 0.24 U ml-1. Finally, an enhancing effect of the enzymatic extracts of M. robertsii (66.7%) and T. harzianum (43.5%) on the efficacy of B. bassiana Bv064 against Diatraea saccharalis larvae was observed. This work demonstrates the non-species-specific enhancing effect of enzymatic extracts on the insecticidal activity of conidial-based biopesticides, which constitutes a contribution to the improvement of biological control agents' performance.

Performance of Metarhizium rileyi Nm017: nutritional supplementation to improve production and quality conidia.

This study aimed to analyze the effect of nutritional supplements on improving conidia production of Metarhizium rileyi Nm017 at laboratory scale (yields of conidia/substrate and biomass/substrate, and substrate consumption). Also, the influence on quality parameters were evaluated (germination at 36 and 48 h, enzymatic activity, and insecticidal activity on Helicoverpa zea). Six treatments (T1-T6) were assessed and all of them reached maximum conidia concentration after 7 days fermentation, a feasible production timetable. Yields from treatment T6 (yeast extract + V8 juice) were 1.5-threefold higher than the other treatments. Conidia from T6 reached germinations of 56% and 12% at 36 and 48 h, respectively, higher than T1 (without supplements), which had the lowest values found. M. rileyi conidia obtained from treatment T6 had the highest enzymatic activity (0.45 U chitinase g-1, 0.28 U lipase g-1, and 1.29 U protease g-1). However, treatments with the highest conidia yields and enzymatic activity were not positively correlated to the efficacy against H. zea. When M. rileyi was produced on T5 (yeast hydrolysate + V8 juice), conidia were 35% more virulent than treatment T6. The findings evidenced the noticeable impact of nutritional substrate amended for conidia production and quality. This work showed the relevance of insecticidal activity assessment as a selection criterion in the mass production development of a biocontrol agent.

Use of Trichoderma koningiopsis chitinase to enhance the insecticidal activity of Beauveria bassiana against Diatraea saccharalis.

Trichoderma is a well-known soil-borne fungus, highly efficient producer of extracellular enzymes including chitinases. The aim of this study was to recover a chitinase from fermentation waste after harvesting Trichoderma koningiopsis Th003 conidia and assess its potential as an enhancer of Beauveria bassiana insecticidal activity against Diatraea saccharalis. T. koningiopsis was produced by solid fermentation, conidia were harvested, and a crude extract (CE) was recovered by washing the residual substrate (rice:wheat bran). The partially purified chitinase (PPC) (75 kDa product) with N-acetyl-ß-glucosaminidase activity was obtained by chromatography to 29.3-fold with optimal activity at pH 5 and 55°C. Both the CE and the PPC were mixed with B. bassiana Bv062 conidia and assessed in a bioassay against D. saccharalis larvae. The CE and PPC from T. koningiopsis Th003 did not affect the germination or viability of B. bassiana conidia and enhanced its insecticidal activity when used at 0.06 U/ml enzymatic activity with a 24.5% reduction in B. bassiana lethal time (LT90 ). This study demonstrated the potential of chitinases produced by T. koningiopsis in solid fermentation to be recovered from the waste substrate and used as an additive to enhance B. bassiana, adding value to the main waste from the Trichoderma biopesticide/biofertilizer industries.