A Comparative Analyses of the Complete Mitochondrial Genomes of Fungal Endosymbionts in Sogatella furcifera, White-Backed Planthoppers

Sogatella furcifera Horvath, commonly known as the white-backed planthoppers (WBPH), is an important pest in East Asian rice fields. Fungal endosymbiosis is widespread among planthoppers in the infraorder Fulgoromorpha and suborder Auchenorrhyncha. We successfully obtained complete mitogenome of five WBPH fungal endosymbionts, belonging to the Ophiocordycipitaceae family, from next-generation sequencing (NGS) reads obtained from S. furcifera samples. These five mitogenomes range in length from 55,390 bp to 55,406 bp, which is shorter than the mitogenome of the fungal endosymbiont found in Ricania speculum, black planthoppers. Twenty-eight protein-coding genes (PCGs), 12 tRNAs, and 2 rRNAs were found in the mitogenomes. Two single-nucleotide polymorphisms, two insertions, and three deletions were identified among the five mitogenomes, which were fewer in number than those of four species of Ophiocordycipitaceae, Ophiocordyceps sinensis, Hirsutella thompsonii, Hirsutella rhossiliensis, and Tolypocladium inflatum. Noticeably short lengths (up to 18 bp) of simple sequence repeats were identified in the five WBPH fungal endosymbiont mitogenomes. Phylogenetic analysis based on conserved PCGs across 25 Ophiocordycipitaceae mitogenomes revealed that the five mitogenomes were clustered with that of R. speculum, forming an independent clade. In addition to providing the full mitogenome sequences, obtaining complete mitogenomes of WBPH endosymbionts can provide insights into their phylogenetic positions without needing to isolate the mtDNA from the host. This advantage is of value to future studies involving fungal endosymbiont mitogenomes.

Hirsutella thompsonii and Pochonia chlamydosporia (Syn. Verticillium chlamydosporium) Mycelia Growth and Predation on Panagrellus redivivus

This research aimed to evaluate the nematophagous ability of 4077-Verticillium chlamydosporium var. chlamydosporium and 4466-Hirsutella thompsonii isolates and relate mycelia growth to the influence provoked by movement of nematodes. Each fungus grew in PDA (potato, dextrose, agar) medium end up to pure colonization. Then, ten mycelia plugs of 8 mm diameter were removed from colony borders and transferred to the center of ten Petri plates containing water-agar 2% medium. These plates were previously divided into four quadrants that received a number of 25 individuals of free-living nematodes (Panagrellus redivivus), composing a total of 100 nematodes per plate. Evaluations started after 24 hours of interaction, considering predation percentage and mycelia growth as stimuli of nematodes presence. Results showed growing predation performance to both isolates, being higher for V. chlamydosporium var. chlamydosporium since from first evaluation time, controlling more than 50% of nematode population initially added. Its predation potential was 39.2%, 38.4% and 48.35% higher than H. thompsonii at first, second and third evaluation day, respectively. Generally, nematodes did not stimulate mycelia growth, unless for H. thompsonii at 72 hours of interaction compared to control plates (without nematodes). Stress resulting from isolates transference from PDA to water-agar 2% resulted in sparse mycelia growth and it could have affected the predation performance of H. thompsonii that controlled nematodes in low levels throughout experiment. Independently of predation level, pictures revealed that both isolates has ability to control P. redivivus through hyphae penetration.

Characterization of a new toxin from the entomopathogenic fungus Metarhizium anisopliae: the ribotoxin anisoplin

Metarhizium anisopliae is an entomopathogenic fungus relevant in biotechnology with applications like malaria vector control. Studies of its virulence factors are therefore of great interest. Fungal ribotoxins are toxic ribonucleases with extraordinary efficiency against ribosomes and suggested as potential insecticides. Here we describe this ribotoxin characteristic activity in M. anisopliae cultures. Anisoplin has been obtained as a recombinant protein and further characterized. It is structurally similar to hirsutellin A, the ribotoxin from the entomopathogen Hirsutella thompsonii. Moreover, anisoplin shows the ribonucleolytic activity typical of ribotoxins and cytotoxicity against insect cells. How Metarhizium uses this toxin and possible applications are of interest.

Evaluation of fungal pathogens, Fusarium semitectum Berk. and Ravenel and Hirsutella thompsonii Fisher against red spider mite, Tetranychus neocaledoni-cus (Andre) of okra under laboratory and greenhouse conditions

Efficacy of fungal pathogens, Fusarium semitectum Berk. and Ravenel and Hirsutella thompsonii Fisher was evaluated against red spider mite, Tetranychus neocaledonicus (Andre) of okra under laboratory and greenhouse conditions. Among the treatments evaluated under laboratory conditions, the highest mortality of 78.20 and 88.34 percent adults and immatures, respectively was recorded at 2.10 x 109 spores per ml of F. semitectum, whereas it was 82.05 and 92.31 per cent adults and immatures, respectively in case of H. thompsonii at 4.60 x 108 spores per ml. Among the twelve treatments evaluated under greenhouse conditions, H. thompsonii 4.60 x 108 spores per ml, F. semitectum 2.10 x 109 spores per ml + H. thompsonii 4.60 x 108 spores per ml, H. thompsonii 4.60 x 108 spores per ml + dicofol 0.025% and F. semitectum 2.10 x 109 spores per ml + H. thompsonii 4.60 x 108 spores per ml + dicofol 0.025% were found to be on par with each other with a cumulative percent mortality of 81.20, 81.48, 81.21 and 82.59 percent, respectively. By testing the field efficacy of F. semitectum and H. thompsonii against T. neocaledonicus, these fungal pathogens can be used as potential biocontrol agents for the sustainable management of mites on okra. Hence the present study evaluates the potentiality of F. semitectum for the management of T. neocaledonicus by comparing with the well-known acaricidal fungus, H. thompsonii.

Involvement of loop 5 lysine residues and the N-terminal β-hairpin of the ribotoxin hirsutellin A on its insecticidal activity

Ribotoxins are cytotoxic members of the family of fungal extracellular ribonucleases best represented by RNase T1. They share a high degree of sequence identity and a common structural fold, including the geometric arrangement of their active sites. However, ribotoxins are larger, with a well-defined N-terminal β-hairpin, and display longer and positively charged unstructured loops. These structural differences account for their cytotoxic properties. Unexpectedly, the discovery of hirsutellin A (HtA), a ribotoxin produced by the invertebrate pathogen Hirsutella thompsonii, showed how it was possible to accommodate these features into a shorter amino acid sequence. Examination of HtA N-terminal β-hairpin reveals differences in terms of length, charge, and spatial distribution. Consequently, four different HtA mutants were prepared and characterized. One of them was the result of deleting this hairpin [Δ(8-15)] while the other three affected single Lys residues in its close spatial proximity (K115E, K118E, and K123E). The results obtained support the general conclusion that HtA active site would show a high degree of plasticity, being able to accommodate electrostatic and structural changes not suitable for the other previously known larger ribotoxins, as the variants described here only presented small differences in terms of ribonucleolytic activity and cytotoxicity against cultured insect cells.