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Most geminiviruses are not transmitted by mechanical inoculation. Therefore, pathogenicity and plant-pathogen interaction studies rely on agroinoculation using infectious clones, which involves cloning the geminiviral genome in a binary vector (see previous chapter for details). A suspension containing the infectious clone inserted into Agrobacterium tumefaciens cells is then inoculated into plants, i.e., agroinoculated. Below is a simple protocol for agroinoculation of an infectious geminivirus clone into plants., (© 2024. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Sweetpotato is a staple crop in Brazil presenting a smaller number of pathogens and diseases in comparison with other root and tuberous crops. Root-knot nematodes are among the most serious sweetpotato root pathogens. The impact of these pathogens also extends to succeeding crops. Because in Brazil, it is common to cultivate more than one crop per season, this problem is rapidly disseminated. The aim of this study was to assess the resistance of two sweetpotato clones and four commercial sweetpotato genotypes to three different Meloidogyne species. Assays were performed under greenhouse and field conditions. 'CIP BRS Nuti' and 'Canadense' were resistant to Meloidogyne incognita , M . javanica , and M . enterolobii under greenhouse conditions. This finding was obtained based on the reproduction factor < 0.2, meaning that the initial population of 5,000 nematodes was reduced to <1,000 individuals after 90 days of inoculation. Tomato and sweetpotato cultivar Beauregard, known to be susceptible to the three nematode species, were highly damaged by the pathogens. Similar results were also observed under field conditions. To our knowledge, 'CIP BRS Nuti' and 'Canadense' are the first South American commercial cultivars with triple nematode resistance.

Background: The neurotoxin peptide ω-ACTX-Hv1a, fused to the carrier molecule GNA, presents potential for insect control as a biopesticide, being orally toxic to insect pests from different orders. However, thorough evaluation is required to assure its safety towards non-target invertebrates. Effects of this novel biopesticide on the parasitoid Eulophus pennicornis via its host Lacanobia oleracea are presented., Results: Hv1a/GNA did not cause mortality when injected or fed to fifth-stage L. oleracea, but caused up to 39% reduction in mean larval weight (P < 0.05) and increased developmental time when injected. When fed, GNA, but not Hv1a/GNA, caused ∼35% reduction in larval weight, indicating that host quality was not affected by the fusion protein. Although GNA and Hv1a/GNA were internalised by the hosts following ingestion, and thus were available to higher trophic levels, no significant changes in the rate of E. pennicornis parasitism occurred. Number of parasitoid pupae per host, adult emergence and sex ratio were unaffected by GNA- or Hv1a/GNA-treated hosts (P > 0.05). The fusion protein was degraded by parasitoid larvae, rendering it non-toxic., Conclusion: Hv1a/GNA has negligible effects on the parasitoid, even under worst-case scenarios. This low toxicity to these insects is of interest in terms of biopesticide specificity and safety to non-target organisms., (© 2015 Society of Chemical Industry.)

The coffee berry borer, Hypothenemus hampei (Ferrari), is an important devastating coffee pest worldwide. Both trypsin and chymotrypsin enzyme activities from H. hampei larval midgut can be inactivated by proteinaceous enzyme-inhibitors. A serine proteinase inhibitor belonging to the Bowman-Birk class was purified from a wild accession of Phaseolus coccineus L. seeds. The inhibitor (PcBBI1) is a cysteine-rich protein that is heat-stable at alkaline pH. MALDI-TOF/MS analysis showed that PcBBI1 occurs in seeds as a monomer (8689 Da) or dimer (17,378 Da). Using in vitro inhibition assays, it was found that PcBBI1 has a high inhibitory activity against H. hampei trypsin-like enzymes, bovine pancreatic chymotrypsin, and trypsin. According to this, PcBBI1 could be a promising tool to make genetically modified coffee with resistance to coffee berry borer.

Different isolates of the soil bacterium Bacillus thuringiensis produce multiple crystal (Cry) proteins toxic to a variety of insects, nematodes and protozoans. These insecticidal Cry toxins are known to be active against specific insect orders, being harmless to mammals, birds, amphibians, and reptiles. Due to these characteristics, genes encoding several Cry toxins have been engineered in order to be expressed by a variety of crop plants to control insectpests. The cotton boll weevil, Anthonomus grandis, and the fall armyworm, Spodoptera frugiperda, are the major economically devastating pests of cotton crop in Brazil, causing severe losses, mainly due to their endophytic habit, which results in damages to the cotton boll and floral bud structures. A cry1Ia-type gene, designated cry1Ia12, was isolated and cloned from the Bt S811 strain. Nucleotide sequencing of the cry1Ia12 gene revealed an open reading frame of 2160 bp, encoding a protein of 719 amino acid residues in length, with a predicted molecular mass of 81 kDa. The amino acid sequence of Cry1Ia12 is 99% identical to the known Cry1Ia proteins and differs from them only in one or two amino acid residues positioned along the three domains involved in the insecticidal activity of the toxin. The recombinant Cry1Ia12 protein, corresponding to the cry1Ia12 gene expressed in Escherichia coli cells, showed moderate toxicity towards first instar larvae of both cotton boll weevil and fall armyworm. The highest concentration of the recombinant Cry1Ia12 tested to achieve the maximum toxicities against cotton boll weevil larvae and fall armyworm larvae were 230 microg/mL and 5 microg/mL, respectively. The herein demonstrated insecticidal activity of the recombinant Cry1Ia12 toxin against cotton boll weevil and fall armyworm larvae opens promising perspectives for the genetic engineering of cotton crop resistant to both these devastating pests in Brazil.