Biostimulant and antagonistic potential of endophytic fungi against fusarium wilt pathogen of tomato Fusarium oxysporum f. sp. lycopersici.

Endophytic fungal-based biopesticides are sustainable and ecologically-friendly biocontrol agents of several pests and diseases. However, their potential in managing tomato fusarium wilt disease (FWD) remains unexploited. This study therefore evaluated effectiveness of nine fungal isolates against tomato fusarium wilt pathogen, Fusarium oxysporum f. sp. lycopersici (FOL) in vitro using dual culture and co-culture assays. The efficacy of three potent endophytes that inhibited the pathogen in vitro was assessed against FWD incidence, severity, and ability to enhance growth and yield of tomatoes in planta. The ability of endophytically-colonized tomato (Solanum lycopersicum L.) plants to systemically defend themselves upon exposure to FOL were also assessed through defence genes expression using qPCR. In vitro assays showed that endophytes inhibited and suppressed FOL mycelial growth better than entomopathogenic fungi (EPF). Endophytes Trichoderma asperellum M2RT4, Hypocrea lixii F3ST1, Trichoderma harzianum KF2R41, and Trichoderma atroviride ICIPE 710 had the highest (68.84-99.61%) suppression and FOL radial growth inhibition rates compared to EPF which exhibited lowest (27.05-40.63%) inhibition rates. Endophytes T. asperellum M2RT4, H. lixii F3ST1 and T. harzianum KF2R41 colonized all tomato plant parts. During the in planta experiment, endophytically-colonized and FOL-infected tomato plants showed significant reduction of FWD incidence and severity compared to non-inoculated plants. In addition, these endophytes contributed to improved growth promotion parameters and yield. Moreover, there was significantly higher expression of tomato defence genes in T. asperellum M2RT4 colonized than in un-inoculated tomato plants. These findings demonstrated that H. lixii F3ST1 and T. asperellum M2RT4 are effective biocontrol agents against FWD and could sustainably mitigate tomato yield losses associated with fusarium wilt.

Effect of cooking methods on nutritional value and microbial safety of edible rhinoceros beetle grubs (Oryctes sp.).

Although edible rhinoceros beetle (Oryctes sp.) larvae are popularly consumed in many countries worldwide, they are prepared using different methods such as boiling, roasting, toasting, and deep-frying, whose effect on nutritional value and microbial safety is scarcely known. Here we investigated the effect of these methods on the nutritional value and microbial safety of Oryctes sp. larvae. Our hypothesis was that cooking the grubs using the four methods had no effect on their nutritional content and microbial loads and diversity. The grubs were analyzed for proximate composition, and fatty and amino acid profiles using standard chemical procedures; and microbial safety using standard culturing procedures. Deep-frying reduced protein and carbohydrate content, but elevated fat content. Boiling lowered ash content, but increased fibre and carbohydrate composition. Roasting and toasting increased protein and ash contents, respectively. Forty fatty acids were detected in the larvae, of which levels of only five were not significantly affected by cooking method, while the levels of the others were differentially affected by the different cooking methods. Amino acid profiles and levels were largely comparable across treatments, but lysine and arginine were higher in all cooked grubs than raw form. All the cooking methods eliminated Enterobacteriaceae, Shigella sp. and Campylobacter sp. from the grubs. Except boiling, all methods reduced total viable count to safe levels. Salmonella sp. were only eliminated by toasting and roasting; while boiling promoted growth of yeast and moulds. Staphylococcus aureus levels exceeded safety limits in all the cooking methods. These findings offer guidance on the type of method to use in preparing the grubs for desired nutritional and safety outcomes.

Interactions between Bacillus thuringiensis and selected plant extracts for sustainable management of Phthorimaea absoluta.

Phthorimaea absoluta is a global constraint to tomato production and can cause up to 100% yield loss. Farmers heavily rely on synthetic pesticides to manage this pest. However, these pesticides are detrimental to human, animal, and environmental health. Therefore, exploring eco-friendly, sustainable Integrated Pest Management approaches, including biopesticides as potential alternatives, is of paramount importance. In this context, the present study (i) evaluated the efficacy of 10 Bacillus thuringiensis isolates, neem, garlic, and fenugreek; (ii) assessed the interactions between the most potent plant extracts and B. thuringiensis isolates, and (iii) evaluated the gut microbial diversity due to the treatments for the development of novel formulations against P. absoluta. Neem recorded the highest mortality of 93.79 ± 3.12% with an LT50 value of 1.21 ± 0.24 days, Bt HD263 induced 91.3 ± 3.68% mortality with LT50 of 2.63 ± 0.11 days, compared to both Bt 43 and fenugreek that caused < 50% mortality. Larval mortality was further enhanced to 99 ± 1.04% when Bt HD263 and neem were combined. Furthermore, the microbiome analyses showed that Klebsiella, Escherichia and Enterobacter had the highest abundance in all treatments with Klebsiella being the most abundant. In addition, a shift in the abundance of the bacterial genera due to the treatments was observed. Our findings showed that neem, garlic, and Bt HD263 could effectively control P. absoluta and be integrated into IPM programs after validation by field efficacy trials.

Biochemistry and transcriptomic analyses of Phthorimaea absoluta (Lepidoptera: Gelechiidae) response to insecticides.

Phthorimaea absoluta is an invasive solanaceous plant pest with highly devastating effects on tomato plant. Heavy reliance on insecticide use to tackle the pest has been linked to insecticide resistance selection in P. absoluta populations. To underline insights on P. absoluta insecticide resistance mechanisms to diamides and avermectins, we evaluated the transcriptomic profile of parental (field-collected) and F8 (lab-reared) populations. Furthermore, to screen for the presence of organophosphate and pyrethroid resistance, we assessed the gene expression levels of acetylcholinesterase (ace1) and para-type voltage-gated sodium channel (VGSG) genes in the F1 to F8 lab-reared progeny of diamide and avermectin exposed P. absoluta field-collected populations. The VGSG gene showed up-regulation in 12.5% and down-regulation in 87.5% of the screened populations, while ace1 gene showed up-regulation in 37.5% and down-regulation in 62.5% of the screened populations. Gene ontology of the differentially expressed genes from both parental and eighth generations of diamide-sprayed P. absoluta populations revealed three genes involved in the metabolic detoxification of diamides in P. absoluta. Therefore, our study showed that the detoxification enzymes found could be responsible for P. absoluta diamide-based resistance, while behavioural resistance, which is stimulus-dependent, could be attributed to P. absoluta avermectin resistance.

Vendors' handling practices of edible long-horned grasshoppers (Ruspolia differens) products and implications on microbial safety.

Edible grasshopper, Ruspolia ruspolia, has nutritional and cherished cultural and economic importance to people from diverse cultures, particularly in over 20 African countries. It is consumed at home or commercially traded as sautéed, deep-fried, or boiled products. However, there is limited information on the hygiene practices of the vendors and the implications on the microbial safety of the final product. This research aimed at assessing the food safety knowledge, handling practices and shelf life of edible long-horned grasshopper products among vendors and the microbial safety of ready-to-eat products sold in 12 different markets in Uganda. Samples of raw, deep-fried and boiled grasshoppers were randomly collected from 74 vendors (62% street and 38% market vendors) and subjected to microbial analysis. Over 85% of the vendors surveyed had no public health food handler's certificate and >95% had limited post-harvest handling knowledge. Total aerobic bacteria (7.30-10.49 Log10 cfu/g), Enterobacteriaceae (5.53-8.56 Log10 cfu/g), yeasts and molds (4.96-6.01 Log10 cfu/g) total counts were significantly high and above the acceptable Codex Alimentarius Commission and Food Safety Authority of Ireland (FSAI) limits for ready-to-eat food products. Eight key pathogenic bacteria responsible for foodborne diseases were detected and these isolates were characterized as Bacillus cereus, Hafnia alvei, Serratia marcescens, Staphylococcus aureus, S. xylosus, S. scuiri, S. haemolyticus, and Pseudomonas aeruginosa. Findings from this study highlight the urgent need to create local and national food safety policies for the edible grasshopper "nsenene" subsector to regulate and guide street and market vending along the value chain, to prevent the transmission of foodborne diseases to consumers.

Comparative transcriptome analysis of false codling moth, Thaumatotibia leucotreta in response to high and low-temperature treatments.

The false codling moth (FCM), Thaumatotibia leucotreta, is a major quarantine pest native to Africa. Physical postharvest phytosanitary measures such as cold and heat treatments are championed to control its spread to new regions. However, the molecular changes that T. leucotreta undergoes as it attempts to adjust to its surroundings during the treatments and withstand the extreme temperatures remain largely unknown. The current study employs RNA-seq using the next-generation Illumina HiSeq platform to produce transcriptome profiles for differential gene expression analysis of T. leucotreta larvae under thermal stress. The transcriptome assembly analysis revealed 226,067 transcripts, clustering into 127,018 unigenes. In comparison to the 25 °C treated group, 874, 91, 159, and 754 individual differentially expressed genes (DEGs) co-regulated at -10, 0, 40, and 50 °C, respectively were discovered. Annotation of the DEGs by gene ontology (GO) revealed several genes, previously implicated in low and high-temperature stresses, including heat shock proteins, cytochrome P450, cuticle proteins, odorant binding proteins, and immune system genes. Kyoto Encyclopedia of Genes and Genomics (KEGG) classification analysis revealed that substantive DEGs were those involved in metabolic pathways such as thiamine, purine, folate, and glycerolipid metabolism pathways. The RT-qPCR validation of several significantly up- and down-regulated DEGs showed congruence between RNA-seq and qPCR data. This baseline study lays a foundation for future research into the molecular mechanisms underlying T. leucotreta's cold/heat tolerance by providing a thorough differential gene expression analysis that has identified multiple genes that may be associated with the insect's ability to withstand cold and heat.

Efficacy of Metarhizium anisopliae and (E)-2-hexenal combination using autodissemination technology for the management of the adult greenhouse whitefly, Trialeurodes vaporariorum Westwood (Hemiptera: Aleyrodidae).

The efficiency of an autodissemination technique in controlling adult whiteflies, Trialeurodes vaporariorum Westwood (Hemiptera: Aleyrodidae) on tomato, Solunum lycopersicum was investigated with previously identified potent fungal isolates of Metarhizium anisopliae ICIPE 18, ICIPE 62 and ICIPE 69 under screenhouse or semi-field conditions. The autodissemination device was inoculated with dry conidia of the M. anisopliae isolates, while control insects were exposed to a fungus-free device. Sampling for conidia uptake, conidial viability and persistence, and insect mortality was done at 1, 2, 3, 5 and 8 days post-exposure, and collected insects were monitored for mortality over ten days. Overall, mortality was higher in insects exposed to ICIPE 18 (62.8%) and ICIPE 69 (61.8%) than in those exposed to ICIPE 62 (42.6%), with median lethal times, (LT50) ranging between 6.73-8.54 days. The control group recorded the lowest mortality rates (18.9%). A general linear reduction in conidial viability with exposure time was observed, although this was more pronounced with M. anisopliae ICIPE 62. Insects exposed to M. anisopliae ICIPE 69 also recorded the highest conidia uptake, hence selected for further evaluation with a T. vaporariorum attractant volatile organic compound, (E)-2-hexenal. The volatile inhibited fungal germination in laboratory compatibility tests, therefore, spatial separation of M. anisopliae ICIPE 69 and (E)-2-hexenal in the autodissemination device was conducted. The inhibitory effects of the volatile were significantly reduced by spatial separation at a distance of 5 cm between the fungus and the volatile, which was found to be more suitable and chosen for the subsequent experiments. Results showed that (E)-2-hexenal did not influence conidia uptake by the insects, while fungal viability and the subsequent mortality variations were more related to duration of exposure. The fungus-volatile compatibility demonstrated with spatial separation provides a basis for the optimisation of the volatile formulation to achieve better T. vaporariorum suppression with an excellent autodissemination efficiency when used in the management of whiteflies under screenhouse conditions.