The Fungus Metarhizium sp. BCC 4849 Is an Effective and Safe Mycoinsecticide for the Management of Spider Mites and Other Insect Pests

Five isolates of Metarhizium sp. were evaluated for their pathogenicity against the spider mite (Tetranychus truncatus Ehara) (Acari: Tetranychidae) and Metarhizium sp. BCC 4849 resulted in the highest mortality (82%) on the 5th day post-inoculation (DPI). Subsequent insect bioassay data indicated similar high virulence against five other insects: African red mites (Eutetranychus africanus Tucker) (Acari: Tetranychidae), bean aphid (Aphis craccivora Koch) (Hemiptera: Aphididae), cassava mealybug (Phenacoccus manihoti Matile-Ferrero) (Hemiptera: Pseudococcidae), sweet potato weevil (Cylas formicarius Fabricius) (Coleoptera: Brentidae), and oriental fruit fly (Bactrocera dorsalis Hendel) (Diptera: Tephritidae), at mortalities of 92–99%, on 3rd–6th DPI, and in laboratory conditions. The pathogenicity assay against E. africanus in hemp plants under greenhouse conditions indicated 85–100% insect mortality on 10th DPI using the fungus alone or in combination with synthetic acaricide. Genome sequencing of Metarhizium sp. BCC 4849 revealed the high abundance of proteins associated with zinc-, heme-, and iron-binding; oxidation-reduction; and transmembrane transport, implicating its versatile mode of interaction with the environment and adaptation to various ion homeostasis. The light and scanning electron microscopy indicated that at 24 h post inoculation (PI), adhesion and appressorial formation occurred, notably near the setae. Most infected mites had stopped moving and started dying by 48–72 h PI. Elongated hyphal bodies and oval blastospores were detected in the legs. At 96–120 h PI or longer, dense mycelia and conidial mass had colonized the interior and exterior of dead mites, primarily at the bottom than the upper part. The shelf-life study also indicated that conidial formulation combined with an oxygen-moisture absorber markedly enhanced the viability and germination after storage at 35 °C for four months. The fungus was tested as safe for humans and animals, according to our toxicological assays.

Addressing Unusual Assay Variability with Robust Statistics

Well-behaved, in vitro bioassays generally produce normally distributed values in their primary (efficacy) data. Accordingly, the best practices for statistical analysis are well documented and understood. However, assays may occasionally display unusually high variability and fall outside the assumptions inherent in these standard analyses. These assays may still be in the optimization phase, in which the source of variation could be identified and addressed. They might also represent the best available option to address the biological process being examined. In these cases, the use of robust statistical methods may provide a more appropriate set of tools for both data analysis and assay optimization. This article provides guidance on best practices for the use of robust statistical methods for the analysis of bioassay data as an alternative to standard methods. Impacts on experimental design and interpretation will be discussed.

Baseline susceptibility of Philippine Ostrinia furnacalis (Lepidoptera: Crambidae) populations to insecticidal Cry1A.105 and Cry2Ab2 proteins and validation of candidate diagnostic concentration for monitoring resistance

Alcantara EP, Atienza MM, Camacho L, Parimi S. 2021. Baseline susceptibility of Philippine Ostrinia furnacalis (Lepidoptera: Crambidae) populations to insecticidal Cry1A.105 and Cry2Ab2 proteins and validation of candidate diagnostic concentration for monitoring resistance. Biodiversitas 22: 956-960. This study estimated the baseline susceptibility of Ostrinia furnacalis populations from the Philippines, to purified insecticidal Cry1A.105 and Cry2Ab2 proteins and determined a diagnostic concentration (DC) through a validation experiment. The insect populations were collected from separate sites of corn farms in Northern and Central Luzon and in South Cotabato province of the island of Mindanao. Dose-response bioassays using artificial diet surface overlay method were conducted on eight populations. The bioassay results revealed that the LC50 of Cry1A.105 and Cry2Ab2 to O. furnacalis ranged from 0.03 ng/cm2 to 0.18 ng/cm2 and 1.40 ng/cm2 to 9.98 ng/cm2, respectively. The relative susceptibility ratios between the most susceptible and most tolerant populations were 6-fold for Cry1A.105 and about 7-fold for Cry2Ab2. The candidate diagnostic concentrations (DC) based on the LC99 were calculated using the baseline bioassay data for both Cry1A.105 and Cry2Ab2. The validation was performed on populations from the same locations used in the baseline susceptibility assay and a reference strain to produce at least 99% mortality for each protein. Data showed that populations tested with Cry1A.105 produced average mortality of at least 99% for the upper limit, while this was observed in the LC99 estimate for Cry2Ab2. The validated diagnostic concentration can be used for monitoring the resistance development of O. furnacalis exposed to Bt Corn, MON89034, in the Philippines.

Identification of Chemopreventive Components from Halophytes Belonging to Aizoaceae and Cactaceae Through LC/MS—Bioassay Guided Approach

Six halophytes, namely, Aptenia cordifolia var. variegata, Glottiphyllum linguiforme, Carpobrotus edulis, Ferocactus glaucescens, F. pottsii and F. herrerae were investigated for chemopreventive effect. Prioritization of most promising plant for further investigation was carried out through an integrated liquid chromatography–high resolution electrospray ionization mass spectrometry profiling—bioassay guided approach. NAD(P)H: quinone oxidoreductase-1 (NQO-1) induction in cultured murine hepatoma cells (Hepa-1c1c7) and inhibition of nitric oxide (NO) production in lipopolysaccharide-activated macrophages (RAW 264.7) were carried out to investigate chemopreventive effect. Bioassay data revealed that F. herrerae, A. cordifolia, C. edulis and F. glaucescens were the most active with 2-, 1.7-, 1.6- and 1.5-folds induction of NQO-1 activity. Only F. glaucescens exhibited >50% inhibition of NO release. LCMS profiling of the F. glaucescens revealed its high content of flavonoids, a known micheal acceptor with possible NQO-1 induction, as proved by quantitative high-performance liquid chromatography analysis. Thus, the extract of F. glaucescens was subjected to chromatographic fractionation leading to the isolation of four compounds including (i) 2S-naringenin, (ii) trans-dihydrokaempferol (aromadendrin), (iii) 2S-naringenin-7-O-β-d-glucopyranoside and (iv) kaempferol-7-O-β-d-glucopyranoside (populnin). The current study through an LCMS dereplication along with bio guided approach reported the activity of populnin as NO inhibitor and NQO-1 inducer with promising chemopreventive potential.

Application 2D Descriptors and Artificial Neural Networks for Beta-Glucosidase Inhibitors Screening

Beta-glucosidase inhibitors play important medical and biological roles. In this study, simple two-variable artificial neural network (ANN) classification models were developed for beta-glucosidase inhibitors screening. All bioassay data were obtained from the ChEMBL database. The classifiers were generated using 2D molecular descriptors and the data miner tool available in the STATISTICA package (STATISTICA Automated Neural Networks, SANN). In order to evaluate the models’ accuracy and select the best classifiers among automatically generated SANNs, the Matthews correlation coefficient (MCC) was used. The application of the combination of maxHBint3 and SpMax8_Bhs descriptors leads to the highest predicting abilities of SANNs, as evidenced by the averaged test set prediction results (MCC = 0.748) calculated for ten different dataset splits. Additionally, the models were analyzed employing receiver operating characteristics (ROC) and cumulative gain charts. The thirteen final classifiers obtained as a result of the model development procedure were applied for a natural compounds collection available in the BIOFACQUIM database. As a result of this beta-glucosidase inhibitors screening, eight compounds were univocally classified as active by all SANNs.

Climate stress resistance in male Queensland fruit fly varies among populations of diverse geographic origins and changes during domestication

Background The highly polyphagous Queensland fruit fly (Bactrocera tryoni Froggatt) expanded its range substantially during the twentieth century and is now the most economically important insect pest of Australian horticulture, prompting intensive efforts to develop a Sterile Insect Technique (SIT) control program. Using a “common garden” approach, we have screened for natural genetic variation in key environmental fitness traits among populations from across the geographic range of this species and monitored changes in those traits induced during domestication. Results Significant variation was detected between the populations for heat, desiccation and starvation resistance and wing length (as a measure of body size). Desiccation resistance was correlated with both starvation resistance and wing length. Bioassay data for three resampled populations indicate that much of the variation in desiccation resistance reflects persistent, inherited differences among the populations. No latitudinal cline was detected for any of the traits and only weak correlations were found with climatic variables for heat resistance and wing length. All three stress resistance phenotypes and wing length changed significantly in certain populations with ongoing domestication but there was also a strong population by domestication interaction effect for each trait. Conclusions Ecotypic variation in heat, starvation and desiccation resistance was detected in Australian Qfly populations, and these stress resistances diminished rapidly during domestication. Our results indicate a need to select source populations for SIT strains which have relatively high climatic stress resistance and to minimise loss of that resistance during domestication.

Antioviposition and Reduction of Callosobruchus chinensis Pic. 1902 (Coleoptera: Bruchidae) Emergence on Phaseolus vulgaris by Dioscorea sansibarensis Powder and Its Chemical Composition

Callosobruchus chinensis causes damage to the Phaseolus vulgaris seeds. Traditionally, Dioscorea sansibarensis serves as a medicinal plant. Naturally, D. sansibarensis has toxins that protect against herbivores and the surrounding invasive plants in its natural habitat. Phytochemical analysis by thin-layer chromatography (TLC) and laboratory experiments was carried out to determine the activity of D. sansibarensis leaves, bulbils, and yams powders on antioviposition and inhibition of the F1 emergence of C. chinensis. Bioassay data were subjected to nonparametric (Kolmogorov–Smirnov) statistical analysis and a generalized linear model at P ≤ 0.05 . Statistically, the powders had an antioviposition activity of 34.3% (R2 = 0.343). A recommendable activity on antioviposition was displayed by the yams powder; treatment by 0.8 g of yams powder had a Wald Chi-Square value of 1.291, P = 0.26 . Inhibition of F1 emergence was significantly attained by the yams powder; the treatment by 0.6 g of yams powder had a Wald Chi-Square value of 7.72, P = 0.01 . Statistically, the bulbils powder displayed low antioviposition and inhibition of F1 emergence. Observations on the TLC exposed compounds with similar Rf values; saponin with an Rf value of 0.72 was portrayed in the leaves, bulbils, and yams. A terpenoid and a flavonoid with Rf values of 0.37 and 0.71, respectively, were observed in bulbils and yams but absent in leaves. A terpenoid with an Rf value of 0.49 was visualized in leaves and bulbils but not in the yams powder. The study concluded that the D. sansibarensis yams and leaves powders are viable for application by the farmers in the protection of stored legumes against attack by C. chinensis. However, there may be other diverse interests in other storage insects and other methods of phytochemical analysis that have not been investigated.

Short Persistence and Vector Susceptibility to Ficam 80WP (bendiocarb active ingredient) During Pilot Application of Indoor Residual Spraying in Burkina Faso, West Africa

Indoor residual spraying (IRS) was applied in addition to the use of long-lasting insecticidal nets in the South West in Burkina Faso, where Anopheles gambiae s.l. the major malaria vector was resistant to pyrethroids. This study was designed to evaluate the efficacy and residual life of bendiocarb (active ingredient) used for spraying on different wall surfaces (mud and cement). Cone bioassays were done monthly with the susceptible An. gambiae ‘Kisumu’ strain and the local wild populations to determine the duration for which insecticide was effective in killing mosquitoes. Cone bioassay data showed low efficacy and short persistence of bendiocarb applied on mud and cement walls, lasting 2 mo with the susceptible insectary strain and less than 1 mo with An. gambiae wild populations. In addition, WHO tube assays confirmed resistance of An. gambiae wild populations to 0.1% bendiocarb with mortality rates less than 80% in both study sites (sprayed and unsprayed sites). The pilot study of IRS with bendiocarb showed that the residual efficacy of bendiocarb was very short, and resistance to bendiocarb was confirmed in wild populations of An. gambiae s.l. Therefore, Ficam 80 WP was not suitable for IRS in this area due to the short residual duration related mainly to vectors resistance to bendiocarb. While waiting for innovative malaria control tool, alternative insecticide (organophosphate or neonicotinoid classes) or combinations of insecticides have to be used for insecticide resistance management in Burkina Faso.