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Williams, Trevor

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https://academica-e.unavarra.es/handle/2454/50181

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Williams

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Trevor

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Producción Agraria

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2621

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Now showing 1 - 10 of 55

Open Access
Genetic variability of Chrysodeixis includens nucleopolyhedrovirus (ChinNPV) and the insecticidal characteristics of selected genotypic variants
(MDPI, 2019) Aguirre Sánchez, Eduardo; Beperet Arive, Inés; Williams, Trevor; Caballero Murillo, Primitivo; Institute for Multidisciplinary Research in Applied Biology - IMAB
Genetic variation in baculoviruses is recognized as a key factor, not only due to the influence of such variation on pathogen transmission and virulence traits, but also because genetic variants can form the basis for novel biological insecticides. In this study, we examined the genetic variability of Chrysodeixis includens nucleopolyhedrovirus (ChinNPV) present in field isolates obtained from virus-killed larvae. Different ChinNPV strains were identified by restriction endonuclease analysis, from which genetic variants were isolated by plaque assay. Biological characterization studies were based on pathogenicity, median time to death (MTD), and viral occlusion body (OB) production (OBs/larva). Nine different isolates were obtained from eleven virus-killed larvae collected from fields of soybean in Mexico. An equimolar mixture of these isolates, named ChinNPV-Mex1, showed good insecticidal properties and yielded 23 genetic variants by plaque assay, one of which (ChinNPV-R) caused the highest mortality in second instars of C. includens. Five of these variants were selected: ChinNPV-F, ChinNPV-J, ChinNPV-K, ChinNPV-R, and ChinNPV-V. No differences in median time to death were found between them, while ChinNPV-F, ChinNPV-K, ChinNPV-R and ChinNPV-V were more productive than ChinNPV-J and the original mixture of field isolates ChinNPV-Mex1. These results demonstrate the high variability present in natural populations of this virus and support the use of these new genetic variants as promising active substances for baculovirus-based bioinsecticides.
Open Access
Iflavirus increases its infectivity and physical stability in association with baculovirus
(PeerJ, 2016) Jakubowska, Agata; Murillo Pérez, Rosa; Carballo Palos, Arkaitz; Williams, Trevor; Lent, Jan W. M. van; Caballero Murillo, Primitivo; Herrero, Salvador; Nekazaritza Ekoizpena; Producción Agraria; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua
Virus transmission and the prevalence of infection depend on multiple factors, including the interaction with other viral pathogens infecting the same host. In this study, active replication of an iflavirus, Spodoptera exigua iflavirus 1 (order Picornavirales) was observed in the offspring of insects that survived following inoculation with a pathogenic baculovirus, Spodoptera exigua multiple nucleopolyhedrovirus. Tracking the origin of the iflavirus suggested the association of this virus with the occlusion bodies of the baculovirus. Here we investigated the effect of this association on the stability and infectivity of both viruses. A reduction in baculovirus pathogenicity, without affecting its infectivity and productivity, was observed when associated with the iflavirus. In contrast, viral association increased the infectivity of the iflavirus and its resistance to ultraviolet radiation and high temperature, two of the main factors affecting virus stability in the field. In addition, electron microscopy analysis revealed the presence of particles resembling iflavirus virions inside the occlusion bodies of the baculovirus, suggesting the possible co-occlusion of both viruses. Results reported here are indicative of facultative phoresis of a virus and suggest that virus virus interactions may be more common than currently recognized, and may be influential in the ecology of baculovirus and host populations and in consequence in the use of baculoviruses as biological insecticides.
Open Access
Post-mortem incubation influences occlusion body production in nucleopolyhedrovirus-infected larvae of Spodoptera frugiperda
(Elsevier, 2019) Ramírez-Arias, Fernando G.; Lasa, Rodrigo; Murillo Pérez, Rosa; Navarro de la Fuente, Laura; Mercado, Gabriel; Williams, Trevor; Agronomía, Biotecnología y Alimentación; Agronomia, Bioteknologia eta Elikadura
The inefficient production of nucleopolyhedrovirus occlusion bodies (OBs) can limit the commercialization of virus-based insecticides. The production of OBs in SfMNPV-infected Spodoptera frugiperda fourth instars was compared among groups of larvae that were frozen (−20 °C) immediately following death, or subjected to a 9-day period of post-mortem incubation at 5 °C or 15 °C. Incubation at 15 °C resulted in a ∼40% increase in total OB production and OBs per mg larval weight, compared to the −20 °C and 5 °C treatments. OBs from the 5 °C treatment were ∼10% smaller in cross sectional area and small OBs (<1 µm 2 ) were more abundant than in other treatments, possibly due to reduced post-mortem OB maturation in this treatment. SfMNPV genomic DNA in OB samples was 3.6-fold higher in the −20 °C treatment than the 5 °C treatment and 1.7-fold higher than the 15 °C treatment, possibly due to differences in the exposure of viral genomes to degradative enzymes. However, these differences did not affect the concentration-mortality relationship or speed of kill of OBs from the different treatments. The abundance of aerobic microbes increased from ∼3 × 10 7 CFU/ml in the −20 °C treatment, to approximately 2 × 10 8 and 3 × 10 8 CFU/ml in the 5 °C and 15 °C incubation treatments, respectively, similar to levels seen in other nucleopolyhedroviruses produced in insects. We conclude that post-mortem incubation at 15 °C likely involves continuing processes of virion occlusion and OB maturation that increase overall OB production without loss of insecticidal activity, although the value of this step in commercial virus insecticide production will depend on the cost of the incubation step and the value of the additional OBs produced.
Open Access
Sf29 Gene of Spodoptera frugiperda multiple nucleopolyhedrovirus is a viral factor that determines the number of virions in occlusion bodies
(American Society for Microbiology, 2008) Simón de Goñi, Oihane; Williams, Trevor; Asensio, Aarón C.; Ros Terés, Sarhay; Gaya Cacho, Andrea; Caballero Murillo, Primitivo; Possee, Robert D.; Nekazaritza Ekoizpena; Producción Agraria; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua
The genome of Spodoptera frugiperda multiple nucleopolyhedrovirus (NPV) was inserted into a bacmid (Sfbac) and used to produce a mutant lacking open reading frame 29 (Sf29null). Sf29null bacmid DNA was able to generate an infection in S. frugiperda. Approximately six times less DNA was present in occlusion bodies (OBs) produced by the Sf29null bacmid in comparison to viruses containing this gene. This reduction in DNA content was consistent with fewer virus particles being packaged within Sf29null bacmid OBs, as determined by fractionation of dissolved polyhedra and comparison of occlusion-derived virus (ODV) infectivity in cell culture. DNA from Sfbac, Sf29null, or Sf29null-repair, in which the gene deletion had been repaired, were equally infectious when used to transfect S. frugiperda. All three viruses produced similar numbers of OBs, although those from Sf29null were 10-fold less infectious than viruses with the gene. Insects infected with Sf29null bacmid died 24 h later than positive controls, consistent with the reduced virus particle content of Sf29null OBs. Transcripts from Sf29 were detected in infected insects 12 h prior to those from the polyhedrin gene. Homologs to Sf29 were present in other group II NPVs, and similar sequences were present in entomopoxviruses. Analysis of the Sf29 predicted protein sequence revealed signal peptide and transmembrane domains, but the presence of 12 potential N-glycosylation sites suggest that it is not an ODV envelope protein. Other motifs, including zinc-binding and threonine-rich regions, suggest degradation and adhesion functions. We conclude that Sf29 is a viral factor that determines the number of ODVs occluded in each OB.
Open Access
Spodoptera frugiperda multiple nucleopolyhedrovirus as a potential biological insecticide: genetic and phenotypic comparison of field isolates from Colombia
(Elsevier, 2011-04-24) Barrera Cubillos, Gloria Patricia; Simón de Goñi, Oihane; Villamizar, Laura; Williams, Trevor; Caballero Murillo, Primitivo; Producción Agraria; Nekazaritza Ekoizpena; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua
Thirty-eight isolates of Spodoptera frugiperda multiple nucleopolyhedrovirus (SfMNPV), collected from infected larvae on pastures, maize, and sorghum plants in three different geographical regions of Colombia, were subjected to molecular characterization and were compared with a previously characterized Nicaraguan isolate (SfNIC). Restriction endonuclease analysis (REN) using six different enzymes showed two different patterns among Colombian isolates, one profile was particularly frequent (92%) and was named SfCOL. The physical map of SfCOL was constructed and the genome was estimated to be 133.9 kb, with few differences in terms of number and position of restriction sites between the genomes of SfNIC and SfCOL. The PstI-K and PstI-M fragments were characteristic of SfCOL. These fragments were sequenced to reveal the presence of seven complete and two partial ORFs. This region was collinear with SfMNPV sf20–sf27. However, two ORFs (4 and 5) had no homologies with SfMNPV ORFs, but were homologous with Spodoptera exigua MNPV (se21 and se22/se23) and Spodoptera litura NPV (splt20 and splt21). Biological characterization was performed against two different colonies of S. frugiperda, one originating from Colombia and one from Mexico. Occlusion bodies (OBs) of the SfCOL isolate were as potent (in terms of concentration–mortality metrics) as SfNIC OBs towards the Mexican insect colony. However, SfCOL OBs were 12 times more potent for the Colombian colony than SfNIC OBs and three times more potent for the Colombian colony than for the Mexican colony. SfCOL and SfNIC showed a slower speed of kill (by ∼50 h) in insects from the Colombian colony compared to the Mexican colony, which was correlated with a higher production of OBs/larvae. SfCOL is a new strain of SfMNPV that presents pathogenic characteristics that favor its development as the basis for a biopesticide product in Colombia.
Open Access
Potential of Cry10Aa and Cyt2Ba, two minority δ-endotoxins produced by Bacillus thuringiensis ser. israelensis, for the control of Aedes aegypti larvae
(MDPI, 2020) Valtierra de Luis, Daniel; Villanueva San Martín, Maite; Lai, Liliana; Williams, Trevor; Caballero Murillo, Primitivo; Agronomia, Bioteknologia eta Elikadura; Institute for Multidisciplinary Research in Applied Biology - IMAB; Agronomía, Biotecnología y Alimentación
Bacillus thuringiensis ser. israelensis (Bti) has been widely used as microbial larvicide for the control of many species of mosquitoes and blackflies. The larvicidal activity of Bti resides in Cry and Cyt δ-endotoxins present in the parasporal crystal of this pathogen. The insecticidal activity of the crystal is higher than the activities of the individual toxins, which is likely due to synergistic interactions among the crystal component proteins, particularly those involving Cyt1Aa. In the present study, Cry10Aa and Cyt2Ba were cloned fromthe commercial larvicideVectoBac-12AS® and expressed in the acrystalliferous Bt strain BMB171 under the cyt1Aa strong promoter of the pSTAB vector. The LC50 values for Aedes aegypti second instar larvae estimated at 24 hpi for these two recombinant proteins (Cry10Aa and Cyt2Ba) were 299.62 and 279.37 ng/mL, respectively. Remarkable synergistic mosquitocidal activity was observed between Cry10Aa and Cyt2Ba (synergistic potentiation of 68.6-fold) when spore + crystal preparations, comprising a mixture of both recombinant strains in equal relative concentrations, were ingested by A. aegypti larvae. This synergistic activity is among the most powerful described so far with Bt toxins and is comparable to that reported for Cyt1A when interacting with Cry4Aa, Cry4Ba or Cry11Aa. Synergistic mosquitocidal activity was also observed between the recombinant proteins Cyt2Ba and Cry4Aa, but in this case, the synergistic potentiation was 4.6-fold. In conclusion, although Cry10Aa and Cyt2Ba are rarely detectable or appear as minor components in the crystals of Bti strains, they represent toxicity factors with a high potential for the control of mosquito populations.
Open Access
Expression of a peroral infection factor determines pathogenicity and population structure in an insect virus
(Public Library of Science, 2013) Simón de Goñi, Oihane; Williams, Trevor; Cerutti, Martine; Caballero Murillo, Primitivo; López Ferber, Miguel; Nekazaritza Ekoizpena; Producción Agraria; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua
A Nicaraguan isolate of Spodoptera frugiperda multiple nucleopolyhedrovirus is being studied as a possible biological insecticide. This virus exists as a mixture of complete and deletion genotypes; the latter depend on the former for the production of an essential per os transmission factor (pif1) in coinfected cells. We hypothesized that the virus population was structured to account for the prevalence of pif1 defector genotypes, so that increasing the abundance of pif1 produced by a cooperator genotype in infected cells would favor an increased prevalence of the defector genotype. We tested this hypothesis using recombinant viruses with pif1 expression reprogrammed at its native locus using two exogenous promoters (egt, p10) in the pif2/pif1 intergenic region. Reprogrammed viruses killed their hosts markedly faster than the wild-type and rescue viruses, possibly due to an earlier onset of systemic infection. Group success (transmission) depended on expression of pif1, but overexpression was prejudicial to group-specific transmissibility, both in terms of reduced pathogenicity and reduced production of virus progeny from each infected insect. The presence of pif1-overproducing genotypes in the population was predicted to favor a shift in the prevalence of defector genotypes lacking pif1-expressing capabilities, to compensate for the modification in pif1 availability at the population level. As a result, defectors increased the overall pathogenicity of the virus population by diluting pif1 produced by overexpressing genotypes. These results offer a new and unexpected perspective on cooperative behavior between viral genomes in response to the abundance of an essential public good that is detrimental in excess.
Open Access
Defective or effective?: mutualistic interactions between virus genotypes
(Royal Society, 2003-11-07) López Ferber, Miguel; Simón de Goñi, Oihane; Williams, Trevor; Caballero Murillo, Primitivo; Producción Agraria; Nekazaritza Ekoizpena
Defective viruses lack genes essential for survival but they can co-infect with complete virus genotypes and use gene products from the complete genotype for their replication and transmission. As such, they are detrimental to the fitness of complete genotypes. Here, we describe a mutualistic interaction between genotypes of an insect baculovirus (nucleopolyhedrovirus of Spodoptera frugiperda (Lepidoptera)) that increases the pathogenicity of the viral population. Mixtures of a complete genotype able to be transmitted orally and a deletion mutant unable to be transmitted orally resulted in a phenotype of increased pathogenicity. Because the infectiousness of mixed genotype infections was greater than that of single genotype infections, we predict that the transmissibility of mixed genotype occlusion bodies will be greater than that of any of their single genotype components. Such interactions will be subject to frequency-dependent selection and will influence the impact of these viruses on insect population dynamics and their efficacy as biological insecticides.
Open Access
Quantification of dose-mortality responses in adult Diptera: validation using Ceratitis capitata and Drosophila suzukii responses to spinosad
(Public Library of Science, 2019) Valtierra de Luis, Daniel; Villanueva San Martín, Maite; Caballero Sánchez, Javier; Matas Casado, Isabel María; Williams, Trevor; Caballero Murillo, Primitivo; Agronomia, Bioteknologia eta Elikadura; Institute for Multidisciplinary Research in Applied Biology - IMAB; Agronomía, Biotecnología y Alimentación; Gobierno de Navarra / Nafarroako Gobernua, BTMOL-PI028; Universidad Pública de Navarra / Nafarroako Unibertsitate Publikoa
Quantitative laboratory bioassay methods are required to evaluate the toxicity of novel insecticidal compounds for pest control and to determine the presence of resistance traits. We used a radioactive tracer based on P-32-ATP to estimate the volume of a droplet ingested by two dipteran pests: Ceratitis capitata (Tephritidae) and Drosophila suzukii (Drosophilidae). Using blue food dye it was possible to distinguish between individuals that ingested the solution from those that did not. The average volume ingested by C. capitata adults was 1.968 mu l. Females ingested a similar to 20% greater volume of solution than males. Adults of D. suzukii ingested an average of 0.879 mu l and females ingested similar to 30% greater volume than males. The droplet feeding method was validated using the naturally-derived insecticide spinosad as the active ingredient (a.i.). For C. capitata, the concentration-mortality response did not differ between the sexes or among three different batches of insects. Lethal dose values were calculated based on mean ingested volumes. For C. capitata LD50 values were 1.462 and 1.502 ng a.i./insect for males and females, respectively, equivalent to 0.274 and 0.271 ng a.i./mg for males and females, respectively, when sex-specific variation in body weight was considered. Using the same process for D. suzukii, the LD50 value was estimated at 2.927 ng a.i./insect, or 1.994 ng a.i./mg based on a mean body weight of 1.67 mg for both sexes together. We conclude that this technique could be readily employed for determination of the resistance status and dose-mortality responses of insecticidal compounds in many species of pestiferous Diptera.
Open Access
Functional importance of deletion mutant genotypes in an insect nucleopolyhedrovirus population
(American Society for Microbiology, 2005) Simón de Goñi, Oihane; Williams, Trevor; López Ferber, Miguel; Caballero Murillo, Primitivo; Producción Agraria; Nekazaritza Ekoizpena
A Nicaraguan isolate of a nucleopolyhedrovirus (SfNIC) that attacks the fall armyworm, Spodoptera frugiperda, survives as a mixture of nine genotypes (SfNIC A to I) that all present genomic deletions, except variant B (complete genotype). Sequencing of cloned restriction fragments revealed that genotypic variants lack between 5 and 16 of the open reading frames present in a contiguous sequence of 18 kb of the SfNIC genome. The absence of oral infectivity of SfNIC-C and -D variants is related to the deletion of the pif and/or pif-2 gene, while that of SfNIC-G remains unexplained. The presence of open reading frame 10, homolog of Se030, also appeared to influence pathogenicity in certain variants. Previous studies demonstrated a significant positive interaction between genotypes B and C. We compared the median lethal concentration of single genotypes (A, B, C, D, and F) and co-occluded genotype mixtures (B+A, B+D, B+F, A+C, and F+C in a 3:1 ratio). Mixtures B+A and B+D showed increased pathogenicity, although only B+D restored the activity of the mixture to that of the natural population. Mixtures of two deletion variants (A+C and F+C) did not show interactions in pathogenicity. We conclude that minority genotypes have an important influence on the overall pathogenicity of the population. These results clearly demonstrate the value of retaining genotypic diversity in virus-based bioinsecticides.