Discovery of mosquitocides from fungal extracts through a high-throughput cytotoxicity-screening approach

Background Mosquitoes transmit a variety of diseases. Due to widespread insecticide resistance, new effective pesticides are urgently needed. Entomopathogenic fungi are widely utilized to control pest insects in agriculture. We hypothesized that certain fungal metabolites may be effective insecticides against mosquitoes. Methods A high-throughput cytotoxicity-based screening approach was developed to search for insecticidal compounds in our newly established global fungal extract library. We first determined cell survival rates after adding various fungal extracts. Candidate insecticides were further analyzed using traditional larval and adult survival bioassays. Results Twelve ethyl acetate extracts from a total of 192 fungal extracts displayed > 85% inhibition of cabbage looper ovary cell proliferation. Ten of these 12 candidates were confirmed to be toxic to Anopheles gambiae Sua5B cell line, and six showed > 85% inhibition of Anopheles mosquito cell growth. Further bioassays determined a LC50, the lethal concentration that kills 50% of larval or adult mosquitoes, of 122 µg/mL and 1.7 µg/mosquito, respectively, after 24 h for extract 76F6 from Penicillium toxicarium. Conclusions We established a high-throughput MTT-based cytotoxicity screening approach for the discovery of new mosquitocides from fungal extracts. We discovered a candidate extract from P. toxicarium that exhibited high toxicity to mosquito larvae and adults, and thus were able to demonstrate the value of our recently developed approach. The active fungal extracts discovered here are ideal candidates for further development as mosquitocides. Graphical abstract

Assessing the impact of a viral infection on the expression of transposable elements in the cabbage looper moth (Trichoplusia ni)

Most studies of stress-induced transposable element (TE) expression have so far focused on abiotic sources of stress. Here we analyzed the impact of an infection by the AcMNPV baculovirus on TE expression in a cell line (Tnms42) and midgut tissues of the cabbage looper moth (Trichoplusia ni). We find that a large fraction of TE families (576/636 in Tnms42 cells and 503/612 in midgut) is lowly expressed or not expressed at all (≤ 4 Transcripts Per Million [TPM]) in the uninfected condition (median TPM of 0.37 in Tnms42 and 0.46 in midgut cells). In the infected condition, a total of 62 and 187 TE families were differentially expressed (DE) in midgut and Tnms42 cells, respectively, with more up- (46) than down- (16) regulated TE families in the former and as many up- (91) as down- (96) regulated TEs in the latter. Expression log2 fold changes of DE TE families varied from -4.95 to 9.11 in Tnms42 cells, and from -4.28 to 7.66 in midgut. Large variations in expression profiles of DE TEs were observed depending on the type of cells and on time after infection. Overall, the impact of AcMNPV on TE expression in T. ni is moderate, but potentially sufficient to affect TE activity and genome architecture. Interestingly, one host-derived TE integrated into AcMNPV genomes is highly expressed in infected Tnms42 cells. This result shows that virus-borne TEs can be expressed, further suggesting that they may be able to transpose, and that viruses may act as vectors of horizontal transfer of TEs in insects.

A Tale of Two Transcriptomic Responses in Agricultural Pests via Host Defenses and Viral Replication

Autographa californica Multiple Nucleopolyhedrovirus (AcMNPV) is a baculovirus that causes systemic infections in many arthropod pests. The specific molecular processes underlying the biocidal activity of AcMNPV on its insect hosts are largely unknown. We describe the transcriptional responses in two major pests, Spodoptera frugiperda (fall armyworm) and Trichoplusia ni (cabbage looper), to determine the host–pathogen responses during systemic infection, concurrently with the viral response to the host. We assembled species-specific transcriptomes of the hemolymph to identify host transcriptional responses during systemic infection and assessed the viral transcript abundance in infected hemolymph from both species. We found transcriptional suppression of chitin metabolism and tracheal development in infected hosts. Synergistic transcriptional support was observed to suggest suppression of immune responses and induction of oxidative stress indicating disease progression in the host. The entire AcMNPV core genome was expressed in the infected host hemolymph with a proportional high abundance detected for viral transcripts associated with replication, structure, and movement. Interestingly, several of the host genes that were targeted by AcMNPV as revealed by our study are also targets of chemical insecticides currently used commercially to control arthropod pests. Our results reveal an extensive overlap between biological processes represented by transcriptional responses in both hosts, as well as convergence on highly abundant viral genes expressed in the two hosts, providing an overview of the host–pathogen transcriptomic landscape during systemic infection.

Distinct Arabidopsis Responses to Two Generalist Caterpillar Species Differing in Host Breadth

In general, caterpillar herbivores with a narrow host preference (specialists) have evolved mechanisms to circumvent specific plant defenses. In contrast, caterpillars with a broader host range (generalists) may manipulate phytohormone pathways common to many plant species to attenuate induced defenses. Many studies have compared plant responses to specialist versus generalist caterpillars. In contrast, this study evaluates the induced response of Arabidopsis thaliana to two generalist caterpillar species, the cabbage looper, Trichoplusia ni, and the beet armyworm, Spodoptera exigua. Although both caterpillars are considered generalists, S. exigua has a broader plant host range, whereas T. ni prefers Brassicaceous plants. Our study shows that most responses to caterpillar herbivory, such as the jasmonate burst, are similar in plants attacked by either insect species; however, we do observe dynamic and temporal differences in specific responses. Expression of AtZAT10, a 12-oxo-phytodienoic acid-responsive gene, is only induced in response to T. ni damage. In comparison, only S. exigua herbivory activates the salicylic acid/NPR1-dependent pathway, as observed by the expression of the marker gene AtPR1. Even though both species induce AtPDF1.2 expression, we found caterpillar-specific temporal differences: T. ni herbivory results in sustained expression over time, whereas gene expression is sharply downregulated at 36 h in S. exigua-attacked plants. Although damage by these two caterpillar species induced AtMYB28 and AtMYB34 expression, specific short- and long-chain aliphatic and indolic glucosinolates accumulate only in response to S. exigua herbivory. These species-specific, plant-induced responses likely reflect differences in effectors found in caterpillar oral secretions. [Formula: see text] Copyright © 2020 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license .

Stimulation of Insect Herbivory by Elevated Temperature Outweighs Protection by the Jasmonate Pathway

Rising global temperatures are associated with increases in the geographic range, population size, and feeding voracity of insect herbivores. Although it is well established that the plant hormone jasmonate (JA) promotes durable resistance to many ectothermic herbivores, little is known about how JA-mediated defense is influenced by rising temperatures. Here, we used the Arabidopsis-Trichoplusia ni (cabbage looper) interaction to investigate the relative contribution of JA and elevated temperature to host resistance. Video monitoring of T. ni larval behavior showed that elevated temperature greatly enhanced defoliation by increasing the bite rate and total time spent feeding, whereas loss of resistance in a JA-deficient mutant did not strongly affect these behaviors. The acceleration of insect feeding at elevated temperature was not attributed to decreases in wound-induced JA biosynthesis, expression of JA-responsive genes, or the accumulation of defensive glucosinolates prior to insect challenge. Quantitative proteomic analysis of insect frass, however, provided evidence for a temperature-dependent increase in the production of T. ni digestive enzymes. Our results demonstrate that temperature-driven stimulation of T. ni feeding outweighs the protective effects of JA-mediated resistance in Arabidopsis, thus highlighting a potential threat to plant resilience in a warming world.