RESUMEN

Using the amino acid sequences and analysis of selected known structures of Bt Cry toxins, Cry1Ab, Cry1Ac, Cry1Ah, Cry1B, Cry1C and Cry1F we specifically designed immunogens. After antibodies selection, broad-spectrum polyclonal antibodies (pAbs) and monoclonal antibody (namely 1A0-mAb) were obtained from rabbit and mouse, respectively. The produced pAbs displayed broad spectrum activity by recognizing Cry1 toxin, Cry2Aa, Cry2Ab and Cry3Aa with half maximal inhibitory concentration (IC50) values of 0.12-9.86 µg/mL. Similarly, 1A0-mAb showed broad spectrum activity, recognizing all of the above Cry protein (IC50 values of 4.66-20.46 µg/mL) with the exception of Cry2Aa. Using optimizations studies, 1A10-mAb was used as a capture antibody and pAbs as detection antibody. Double antibody sandwich enzyme-linked immunosorbent assays (DAS-ELISAs) were established for Cry1 toxin, Cry2Ab and Cry3Aa with the limit of detection (LOD) values of 2.36-36.37 ng/mL, respectively. The present DAS-ELISAs had good accuracy and precisions for the determination of Cry toxin spiked tap water, corn, rice, soybeans and soil samples. In conclusion, the present study has successfully obtained broad-spectrum pAbs and mAb. Furthermore, the generated pAbs- and mAb-based DAS-ELISAs protocol can potentially be used for the broad-spectrum monitoring of eight common subtypes of Bt Cry toxins residues in food and environmental samples.

Asunto(s)

RESUMEN

The fall armyworm (Spodoptera frugiperda) is a major global pest causing severe damage to various crops, especially corn. Transgenic corn producing the Cry1F pesticidal protein from the bacterium Bacillus thuringiensis (Cry1F corn) showed effectiveness in controlling this pest until S. frugiperda populations at locations in North and South America evolved practical resistance. The mechanism for practical resistance involved disruptive mutations in an ATP binding cassette transporter subfamily C2 gene (SfABCC2), which serves as a functional Cry1F receptor in the midgut cells of susceptible S. frugiperda. The SfABCC2 protein contains two transmembrane domains (TMD1 and TMD2), each with a cytosolic nucleotide (ATP) binding domain (NBD1 and NBD2, respectively). Previous reports have demonstrated that disruptive mutations in TMD2 were linked with resistance to Cry1F, yet whether the complete SfABCC2 structure is needed for receptor functionality or if a single TMD-NBD protein can serve as functional Cry1F receptor remains unknown. In the present study, we separately expressed TMD1 and TMD2 with their corresponding NBDs in cultured insect cells and tested their Cry1F receptor functionality. Our results show that the complete SfABCC2 structure is required for Cry1F receptor functionality. Moreover, binding competition assays revealed that Cry1F specifically bound to SfABCC2, whereas neither SfTMD1-NBD1 nor SfTMD2-NBD2 exhibited any significant binding. These results provide insights into the molecular mechanism of Cry1F recognition by SfABCC2 in S. frugiperda, which could facilitate the development of more effective insecticidal proteins.

Asunto(s)

RESUMEN

BACKGROUND: The widespread use of chemical herbicides and the growing issue of weed resistance pose significant challenges in agriculture. To address these problems, there is a pressing need to develop biological herbicides based on bacterial metabolites. RESULTS: In this study, we investigated the impact of the cell-free culture filtrate (CFCF) from the ZT isolate, a bacilliform bacterium obtained from diseased wheat seeds, on the germination and seedling growth of various plant species, including wild oat, ryegrass, redroot, wheat, and chickpea. The results revealed that CFCF had a detrimental effect on the fresh and dry weight of stems and roots in most of the studied plants, except chickpeas. The CFCF was further subjected to separation into aqueous and organic phases using chloroform, followed by the division of the aqueous phase into 13 fractions using an alumina column. Notably, both the aqueous phase (20%) and all 13 fractions (ranging from 50% to 83%) displayed the ability to reduce the root length of ryegrass, a monocotyledonous weed. Liquid chromatography-mass spectrometry (LC-MS) analysis identified that fractions 3 and 7, which were effective against ryegrass but not redroot, contained Cry family proteins, including Cry10 Aa, Cry4 Ba, and Cry4 Aa. Additionally, 16s rRNA gene sequencing revealed that the ZT isolate is closely related (98.27%) to Bacillus wiedmannii. CONCLUSION: Conclusively, metabolites from the ZT bacterium hold promise for monocotyledonous weed-targeted herbicides, providing a constructive strategy to confront agricultural issues tied to chemical herbicides and weed resistance. © 2024 Society of Chemical Industry.

Asunto(s)

Asunto(s)

RESUMEN

Bacillus thuringiensis (Bt) Cry toxins have been widely used in the development of genetically modified organisms (GMOs) for pest control. This work aimed to establish more cost effective and broader detection methods for commonly used Cry toxins. Using ligand blot and bio-layer interferometry, we confirmed that a recombinant toxin-binding fragments derived from Helicoverpa armigera cadherin-like protein (HaCad-TBR) could broadly bind Cry1Ab, Cry1Ac, Cry2Aa, and Cry2Ab with the affinity of 0.149, 0.402, 120, and 4.12 nM, respectively. Based on the affinity results, a novel receptor-antibody sandwich assay broadly detecting Cry1A and Cry2 toxins was developed by using HaCad-TBR as capture molecules, and anti-Cry1A/Cry2A polyclonal antibodies (pAbs) as the detection antibodies. The detection limit (LOD) for Cry1Ab, Cry1Ab, Cry2Aa, and Cry2Ab were 5.30, 5.75, 30.83 and 13.70 ng/mL. To distinguish Cry1A and Cry2A toxins in a singular test, anti-Cry1A pAbs and anti-Cry2A pAbs were labelled with different quantum dots (QDs). The LOD for the four toxins by receptor-QDs-pAbs sandwich assay were calculated to be 1.36, 4.71, 17.48, and 7.54 ng/mL, respectively. The two developed methods were validated by spiked rice and corn samples, suggesting they may potentially be used in monitoring and quantifying Cry toxins in food and environment.

Asunto(s)

RESUMEN

Bacillus thuringiensis (Bt) strains produce pore-forming toxins (PFTs) that attack insect pests. Information for pre-pore and pore structures of some of these Bt toxins is available. However, for the three-domain (I-III) crystal (Cry) toxins, the most used Bt toxins in pest control, this crucial information is still missing. In these Cry toxins, biochemical data have shown that 7-helix domain I is involved in insertion in membranes, oligomerization and formation of a channel lined mainly by helix α4, whereas helices α1 to α3 seem to have a dynamic role during insertion. In the case of Cry1Aa, toxic against Manduca sexta larvae, a tetrameric oligomer seems to precede membrane insertion. Given the experimental difficulty in the elucidation of the membrane insertion steps, we used Alphafold-2 (AF2) to shed light on possible oligomeric structural intermediates in the membrane insertion of this toxin. AF2 very accurately (<1 Å RMSD) predicted the crystal monomeric and trimeric structures of Cry1Aa and Cry4Ba. The prediction of a tetramer of Cry1Aa, but not Cry4Ba, produced an 'extended model' where domain I helices α3 and α2b form a continuous helix and where hydrophobic helices α1 and α2 cluster at the tip of the bundle. We hypothesize that this represents an intermediate that binds the membrane and precedes α4/α5 hairpin insertion, together with helices α6 and α7. Another Cry1Aa tetrameric model was predicted after deleting helices α1 to α3, where domain I produced a central cavity consistent with an ion channel, lined by polar and charged residues in helix α4. We propose that this second model corresponds to the 'membrane-inserted' structure. AF2 also predicted larger α4/α5 hairpin n-mers (14 ≤n ≤ 17) with high confidence, which formed even larger (~5 nm) pores. The plausibility of these models is discussed in the context of available experimental data and current paradigms.

Asunto(s)

RESUMEN

The fall armyworm (FAW), Spodoptera frugiperda (J.E. Smith; Lepidoptera: Noctuidae) is an invasive agricultural pest with a global distribution, causing major crop losses annually. Its control strategies largely rely on chemical insecticides and transgenic crops expressing Bacillus thuringiensis insecticidal proteins (Cry and Vip toxins); however, the development of high resistance poses a significant issue. The ATP-binding cassette transporter C2 (ABCC2) has been linked to Cry toxin pore formation, acting as a receptor of some Cry toxins. Recently detected mutations in the SfABCC2 gene in extracellular loop 4 (ECL4) have been associated with Bt toxin resistance in FAW. In the present study, we expressed the SfABCC2 gene in Drosophila melanogaster, a species normally unaffected by the Bt toxins. We demonstrate that susceptibility can be introduced by the ectopic and tissue-specific expression of wildtype SfABCC2. Next, we introduced mutations into ECL4-both individually and in combination-that have been recently described in Brazilian FAW and functionally validated by toxicity bioassays against the foliar Bt product Xentari. Our results provide an efficient demonstration of the suitability of transgenic Drosophila for validating FAW ABCC2 resistance mutations in ECL4 against Bt toxins, and potential cross-resistance issues between closely related proteins that use ABCC2.

Asunto(s)

RESUMEN

The native Cry4Aa δ-endotoxin produced exclusively in Bacillus thuringiensis during sporulation as a ∼130-kDa inactive protoxin is confined within the parasporal crystalline inclusion that dissolves at alkaline pH in the midgut lumen of mosquito larvae. Here, the recombinant Cry4Aa toxin over-expressed in Escherichia coli at 30 °C as an alkaline-solubilizable inclusion was found inevitably lost during isolation from the cell lysate (pH ∼6.5) of which host cells were pre-suspended in distilled water (pH ∼5.5). When 100 mM KH2PO4 (pH 5.0) was used as host cell-suspending buffer, the cell lysate's pH became more acidic (pH 5.5), allowing the expressed protoxin to be entirely retained in the form of crystalline inclusion rather than a soluble form, and thus high-yield recovery of the partially purified inclusion was obtained. Upon dialysis of the alkaline-solubilized protoxin against the KH2PO4 buffer, the protoxin precipitate was efficiently recovered and still exhibited high toxicity to Aedes aegypti mosquito larvae. Additionally, the precipitated protoxin was completely resolubilized in 50 mM Na2CO3 buffer (pH 9.0) and proteolytically processed by trypsin to produce the 65-kDa activated toxin comprising ∼47- and ∼20-kDa fragments. In silico structural analysis suggested that His154, His388, His536 and His572 were involved in a dissolution of the Cry4Aa inclusion at pH 6.5, conceivably through interchain salt bridge breakage. Altogether, such an optimized protocol described herein was effective for the preparation of alkaline-solubilizable inclusions of the recombinant Cry4Aa toxin in large amounts (>25 mg per liter culture) that would pave the way for further structure-function relationship studies of different Cry toxins.

Asunto(s)

RESUMEN

Bacillus thuringiensis (Bt) produces different insecticidal proteins effective for pest control. Among them, Cry insecticidal proteins have been used in transgenic plants for the control of insect pests. However, evolution of resistance by insects endangers this technology. Previous work showed that the lepidopteran insect Plutella xylostella PxHsp90 chaperone enhanced the toxicity of Bt Cry1A protoxins by protecting them from degradation by the larval gut proteases and by enhancing binding of the protoxin to its receptors present in larval midgut cells. In this work, we show that PxHsp70 chaperone also protects Cry1Ab protoxin from gut proteases degradation, enhancing Cry1Ab toxicity. We also show that both PxHsp70 and PxHsp90 chaperones act cooperatively, increasing toxicity and the binding of Cry1Ab439D mutant, affected in binding to midgut receptors, to cadherin receptor. Also, insect chaperones recovered toxicity of Cry1Ac protein to a Cry1Ac-highly resistant P. xylostella population, NO-QAGE, that has a disruptive mutation in an ABCC2 transporter linked to Cry1Ac resistance. These data show that Bt hijacked an important cellular function for enhancing its infection capability, making use of insect cellular chaperones for enhancing Cry toxicity and for lowering the evolution of insect resistance to these toxins.

Asunto(s)

RESUMEN

The sugarcane giant borer, Telchin licus licus, is an insect pest that causes significant losses in sugarcane crops and in the sugar-alcohol sector. Chemical and manual control methods are not effective. As an alternative, in the current study, we have screened Bacillus thuringiensis (Bt) Cry toxins with high toxicity against this insect. Bioassays were conducted to determine the activity of four Cry toxins (Cry1A (a, b, and c) and Cry2Aa) against neonate T. licus licus larvae. Notably, the Cry1A family toxins had the lowest LC50 values, in which Cry1Ac presented 2.1-fold higher activity than Cry1Aa, 1.7-fold larger than Cry1Ab, and 9.7-fold larger than Cry2Aa toxins. In silico analyses were performed as a perspective to understand putative interactions between T. licus licus receptors and Cry1A toxins. The molecular dynamics and docking analyses for three putative aminopeptidase N (APN) receptors (TlAPN1, TlAPN3, and TlAPN4) revealed evidence for the amino acids that may be involved in the toxin-receptor interactions. Notably, the properties of Cry1Ac point to an interaction site that increases the toxin's affinity for the receptor and likely potentiate toxicity. The interacting amino acid residues predicted for Cry1Ac in this work are probably those shared by the other Cry1A toxins for the same region of APNs. Thus, the presented data extend the existing knowledge of the effects of Cry toxins on T. licus licus and should be considered in further development of transgenic sugarcane plants resistant to this major occurring insect pest in sugarcane fields.

Asunto(s)

RESUMEN

Aedes albopictus is a species of mosquito, originally from Southeast Asia, that belongs to the Culicidae family and the Dipteran insect order. The distribution of this vector has rapidly changed over the past decade, making most of the temperate territories in the world vulnerable to important human vector-borne diseases such as dengue, yellow fever, zika or chikungunya. Bacillus thuringiensis var. israeliensis (Bti)-based insecticides represent a realistic alternative to the most common synthetic insecticides for the control of mosquito larvae. However, several studies have revealed emerging resistances to the major Bti Crystal proteins such as Cry4Aa, Cry4Ba and Cry11Aa, making the finding of new toxins necessary to diminish the exposure to the same toxicity factors overtime. Here, we characterized the individual activity of Cyt1Aa, Cry4Aa, Cry4Ba and Cry11Aa against A. albopictus and found a new protein, Cyt1A-like, that increases the activity of Cry11Aa more than 20-fold. Additionally, we demonstrated that Cyt1A-like facilitates the activity three new Bti toxins: Cry53-like, Cry56A-like and Tpp36-like. All in all, these results provide alternatives to the currently available Bti products for the control of mosquito populations and position Cyt proteins as enablers of activity for otherwise non-active crystal proteins.

Asunto(s)

RESUMEN

The gut microbiota of insects plays a vital role in digestion, nutrient acquisition, metabolism of dietary toxins, pathogen immunity and maintenance of gut homeostasis. Bacillus thuringinensis (Bt) poisons target insects through its toxins that are activated in the insect gut. The effects of Bt toxins on gut microbiota of insects and their underlying mechanisms are not well understood. In this study, we found that Cry1Ab/2Ab toxins significantly changed the gut bacterial community's structure and reduced the total load of gut bacteria in the Locusta migratoria. In addition, Cry toxins significantly increased the level of reactive oxygen species (ROS) in the gut of locusts. Our results also showed that Cry1Ab/2Ab toxins induced the host gut's immune response by up-regulating of key genes in the Immune deficiency (IMD) and Toll pathway. RNA interference showed that knocking down Relish could narrow the difference in the load, diversity, and composition in gut bacteria caused by Cry toxins. Our findings suggest that Bt potentially influences the gut bacterial community of L. migratoria through host immune response.

Asunto(s)

RESUMEN

The pine wilt disease is caused by the pinewood nematode Bursaphelenchus xylophilus and it results in serious ecological and economic losses. Therefore, effective prevention and control methods for the pinewood nematode are urgently required. Bacillus thuringiensis (Bt), a widely used microbial insecticide, produces toxins that are toxic to several species of parasitic nematodes, however, its effects on B. xylophilus have not been determined. In this study, Cry5Ba3, App6Aa2, Cry12Aa1, Cry13Aa1, Cry14Aa1, Cry21Aa3, Cry21Fa1, Xpp55Aa1, and Cyt8Aa1 toxins' nematocidal activity against B. xylophilus was evaluated, six toxins with high toxicity were identified: App6Aa2 (LC50 = 49.71 µg/mL), Cry13Aa1 (LC50 = 53.17 µg/mL), Cry12Aa1 (LC50 = 58.88 µg/mL), Cry5Ba3 (LC50 = 63.99 µg/mL), Xpp55Aa1 (LC50 = 65.14 µg/mL), and Cyt8Aa1 (LC50 = 96.50 µg/mL). The six toxins caused shrinkage and thinning of the intestinal cells, contraction of the intestine from the body wall, vacuolization, and degenerated appearance of the pinewood nematodes. The results of this study provide basic information to study the action mechanism of nematocidal toxins on the pinewood nematode and direction for the use of nematocidal toxins in the biological control of B. xylophilus.

Asunto(s)

RESUMEN

Insect midgut cadherins function as receptors and play critical roles as protein receptors of insecticidal Bacillus thuringiensis (Bt) toxins used as biopesticides and in Bt transgenic crops worldwide. Here, we cloned and characterized the full-length midgut cadherin (CmCad) cDNA from the rice leaffolder (Cnaphalocrocis medinalis), a destructive pest of rice in many Asian countries. Expression of recombinant proteins corresponding to the extracellular domain of CmCad allowed testing binding of Cry proteins. Results from in vitro ligand blotting and enzyme-linked immunosorbent assays supported that the extracellular domain of CmCad contains regions recognized by both Cry1Ac and Cry2Aa. Molecular modelling and docking simulations indicated that binding to both Cry1Ac and Cry2Aa is localized primarily within a CmCad motif corresponding to residues T1417-D1435. A recombinant CmCad protein produced without residues T1417-D1435 lacked binding to Cry1Ac and Cry2Aa, confirmed our modelling predictions that CmCad has a shared Cry1Ac and Cry2Aa binding site. The potential existence of a shared binding region in CmCad suggests that caution should be taken when using combinations of Cry1Ac and Cry2Aa in pyramided transgenic rice, as their combined use could speed the evolution of resistance to both toxins.

Asunto(s)

RESUMEN

Bacillus thuringiensis (Bt) is a bacterium capable of producing Cry toxins, which are recognized for their bio-controlling actions against insects. However, a few Bt strains encode proteins lacking insecticidal activity but showing cytotoxic activity against different cancer cell lines and low or no cytotoxicity toward normal human cells. A subset of Cry anticancer proteins, termed parasporins (PSs), has recently arisen as a potential alternative for cancer treatment. However, the molecular receptors that allow the binding of PSs to cells and their cytotoxic mechanisms of action have not been well established. Nonetheless, their selective cytotoxic activity against different types of cancer cell lines places PSs as a promising alternative treatment modality. In this review, we provide an overview of the classification, structures, mechanisms of action, and insights obtained from genetic modification approaches for PS proteins.

Asunto(s)

RESUMEN

Helicoverpa armigera is a major insect pest of several crops worldwide. This insect is susceptible to some Bacillus thuringiensis (Bt) Cry insecticidal proteins expressed in transgenic crops or used in biopesticides. Previously, we identified H. armigera prohibitin (HaPHB) as a Cry1Ac-binding protein. Here, we further analyzed the potential role of PHB as a Cry toxin receptor in comparison to cadherin (CAD), well recognized as a Cry1Ac receptor. HaPHB-2 midgut protein and HaCAD toxin-binding region (TBR) fragment from H. armigera were expressed in Escherichia coli cells, and binding assays with different Cry1 toxins were performed. We demonstrated that Cry1Ab, Cry1Ac, and Cry1Fa toxins bound to HaPHB-2 in a manner similar to that seen with HaCAD-TBR. Different Cry1Ab mutant toxins located in domain II (Cry1AbF371A and Cry1AbG439D) or domain III (Cry1AbL511A and Cry1AbN514A), which were previously characterized and found to be affected in receptor binding, were analyzed regarding their binding interaction with HaPHB-2 and toxicity against H. armigera One ß-16 mutant (Cry1AbN514A) showed increased binding to HaPHB-2 that correlated with 6-fold-higher toxicity against H. armigera, whereas the other ß-16 mutant (Cry1AbL511A) was affected in binding to HaPHB-2 and lost toxicity against H. armigera Our data indicate that ß-16 from domain III of Cry1Ab is involved in interactions with HaPHB-2 and in toxicity. This report identifies a region of Cry1Ab involved in binding to HaPHB-2 from a Lepidoptera insect, suggesting that this protein may participate as a novel receptor in the mechanism of action of the Cry1 toxins in H. armigeraIMPORTANCEHelicoverpa armigera is a polyphagous pest that feeds on important crops worldwide. This insect pest is sensitive to different Cry1 toxins from Bacillus thuringiensis In this study, we analyzed the potential role of PHB-2 as a Cry1 toxin receptor in comparison to CAD. We show that different Cry1 toxins bound to HaPHB-2 and HaCAD-TBR similarly and identify ß-16 from domain III of Cry1Ab as a binding region involved in the interaction with HaPHB-2 and in toxicity. This report characterized HaPHB-Cry1 binding interaction, providing novel insights into potential target sites for improving Cry1 toxicity against H. armigera.

Asunto(s)

RESUMEN

3D-Cry toxins, produced by the entomopathogenic bacterium Bacillus thuringiensis, have been extensively mutated in order to elucidate their elegant and complex mechanism of action necessary to kill susceptible insects. Together with the study of the resistant insects, 3D-Cry toxin mutants represent one of the pillars to understanding how these toxins exert their activity on their host. The principle is simple, if an amino acid is involved and essential in the mechanism of action, when substituted, the activity of the toxin will be diminished. However, some of the constructed 3D-Cry toxin mutants have shown an enhanced activity against their target insects compared to the parental toxins, suggesting that it is possible to produce novel versions of the natural toxins with an improved performance in the laboratory. In this report, all mutants with an enhanced activity obtained by accident in mutagenesis studies, together with all the variants obtained by rational design or by directed mutagenesis, were compiled. A description of the improved mutants was made considering their historical context and the parallel development of the protein engineering techniques that have been used to obtain them. This report demonstrates that artificial 3D-Cry toxins made in laboratories are a real alternative to natural toxins.

Asunto(s)

RESUMEN

Bacillus thuringiensis (Bt) is a natural pathogen of insects and some other groups of invertebrates that produces three-domain Cry (3d-Cry) toxins, which are highly host-specific pesticidal proteins. These proteins represent the most commonly used bioinsecticides in the world and are used for commercial purposes on the market of insecticides, being convergent with the paradigm of sustainable growth and ecological development. Emerging resistance to known toxins in pests stresses the need to expand the list of known toxins to broaden the horizons of insecticidal approaches. For this purpose, we have elaborated a fast and user-friendly tool called CryProcessor, which allows productive and precise mining of 3d-Cry toxins. The only existing tool for mining Cry toxins, called a BtToxin_scanner, has significant limitations such as limited query size, lack of accuracy and an outdated database. In order to find a proper solution to these problems, we have developed a robust pipeline, capable of precise 3d-Cry toxin mining. The unique feature of the pipeline is the ability to search for Cry toxins sequences directly on assembly graphs, providing an opportunity to analyze raw sequencing data and overcoming the problem of fragmented assemblies. Moreover, CryProcessor is able to predict precisely the domain layout in arbitrary sequences, allowing the retrieval of sequences of definite domains beyond the bounds of a limited number of toxins presented in CryGetter. Our algorithm has shown efficiency in all its work modes and outperformed its analogues on large amounts of data. Here, we describe its main features and provide information on its benchmarking against existing analogues. CryProcessor is a novel, fast, convenient, open source (https://github.com/lab7arriam/cry_processor), platform-independent, and precise instrument with a console version and elaborated web interface (https://lab7.arriam.ru/tools/cry_processor). Its major merits could make it possible to carry out massive screening for novel 3d-Cry toxins and obtain sequences of specific domains for further comprehensive in silico experiments in constructing artificial toxins.

Asunto(s)

RESUMEN

Bacillus thuringiensis (Bt) toxins are effective in controlling insect pests either through the spraying of products or when expressed in transgenic crops. The discovery of endophytic Bt strains opened new perspectives for studies aimed at the control of sap-sucking insects, such as the Asian citrus psyllid Diaphorina citri Kuwayama (Hemiptera: Liviidae), a vector of "Candidatus Liberibacter spp.," associated with citrus huanglongbing (HLB). In this study, translocation of endophytic Bt strains in citrus seedlings inoculated with Bt suspension delivered by soil-drench, and their systemic pathogenicity to D. citri nymphs were investigated. The pathogenicity of three wild-type Bt strains against D. citri third-instar nymphs was demonstrated. Among the 10 recombinant strains tested (each of them harboring a single cry or cyt gene), 3 can be highlighted, causing 42%-77% and 66%-90% nymphal mortality at 2 and 5 d after inoculation, respectively. The isolation of Bt cells from young citrus shoots and dead nymphs, and PCR performed with specific primers, confirmed the involvement of the Bt strains in the psyllid mortality. This is the first report showing the translocation of Bt strains from citrus seedling roots to shoots and their potential to control D. citri nymphs that fed on these soil-drench inoculated seedlings. The Bt strains that caused the highest mortality rates have the potential to be used as bioinsecticides to control D. citri and the identified genes can be used for the production of transgenic Bt citrus.

Asunto(s)

RESUMEN

Science is observation. Application of Science is engineering. A 0% error is desired in Science, while a 25% error is usually allowed in Engineering. Technology is engineering with Science where the error rate is considerably reduced to improve precision. Biotechnology is truly interdisciplinary with an optimal mix of physics, chemistry and biology linked by Mathematics. Chemistry evolved into Chemical Engineering and thus Biochemistry into Biochemical Engineering. Biochemical engineering with genetics and molecular biology created Biotechnology. Biotechnology with computer science developed Bioinformatics. Bioinformatics used biological data to glean BIOINFORMATION for Biological Knowledge Discovery (BKD). This helped to accelerate drug discovery and develop other biologics (biomarkers, vaccines, seed developments, bio-fertilizers and bio-pesticides) towards improved service in healthcare, agriculture, food production, food processing and food distribution across international borders as per demand supply in the supply chain. It is joyful to realize the personal experience with the multifaceted features of Biotechnology, Bioinformatics and Bioinformation in a comprehensive manner over a period of three decades. This educational path is truly exciting, engaging and enterprising. This journey provided an opportunity to debate on cry toxins, lipase, ibuprofen, HLA alleles, antigens, peptide vaccines, protein-protein interactions, genomes and biological knowledge discovery models.