Person:
Williams, Trevor

person.page.identifierURI

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

Last Name

Williams

First Name

Trevor

person.page.departamento

Producción Agraria

person.page.upna

2621

Full item page

Search Results

Now showing 1 - 10 of 37

Open Access
Lacanobia oleracea nucleopolyhedrovirus (LaolNPV): a new European species of alphabaculovirus with a narrow host range
(Public Library of Science, 2017) Simón de Goñi, Oihane; Erlandson, Martin A.; Frayssinet, Marie; Williams, Trevor; Theilmann, David A.; Volkoff, Anne Nathalie; Caballero Murillo, Primitivo; Nekazaritza Ekoizpena; Producción Agraria; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua; Gobierno de Navarra / Nafarroako Gobernua, IIQ1406-RI1
During an insect sampling program in alfalfa crops near Montpellier, France in 2011, Lacanobia oleracea larvae were collected that died due to nucleopolyhedrovirus infection (LaolNPV). This virus was subjected to molecular and biological characterization. The virus was a multiple nucleocapsid NPV that showed similar restriction profiles to Mamestra configurata NPV-A (MacoNPV-A) but with significant differences. Polypeptide analysis demonstrated similar proteins in occlusion bodies and occlusion derived virions, to those observed in NPVs from Mamestra spp. Terminal sequencing revealed that the genome organization shared similarity with that of MacoNPV-A. The most homologous virus was MacoNPV-A 90/2 isolate (95.63% identity and 96.47% similarity), followed by MacoNPV-A 90/4 strain (95.37% and 96.26%), MacoNPV-B (89.21% and 93.53%) and M. brassicae MNPV (89.42% and 93.74%). Phylogenetic analysis performed with lef-8, lef-9, polh and a concatenated set of genes showed that LaolNPV and the Mamestra spp. NPVs clustered together with HaMNPV, but with a closer genetic distance to MacoNPV-A strains. The Kimura 2-parameter (K-2-P) distances of the complete genes were greater than 0.05 between LaolNPV and the MbMNPV/MacoNPV-B/HaMNPV complex, which indicates that LaolNPV is a distinct species. K-2-P distances were in the range 0.015±0.050 for comparisons of LaolNPV with MacoNPV-A strains, such that additional biological characteristics should be evaluated to determine species status. While MacoNPV-A was pathogenic to seven lepidopteran species tested, LaolNPV was only pathogenic to Chrysodeixis chalcites. Given these findings, Lacanobia oleracea nucleopolyhedrovirus should be considered as a new species in the Alphabaculovirus genus.
Open Access
Baculovirus expression and functional analysis of Vpa2 proteins from Bacillus thuringiensis
(MDPI, 2020) Simón de Goñi, Oihane; Palma Dovis, Leopoldo; Fernández González, Ana Beatriz; Williams, Trevor; Caballero Murillo, Primitivo; Institute for Multidisciplinary Research in Applied Biology - IMAB
The mode of action underlying the insecticidal activity of the Bacillus thuringiensis (Bt) binary pesticidal protein Vpa1/Vpa2 is uncertain. In this study, three recombinant baculoviruses were constructed using Bac-to-Bac technology to express Vpa2Ac1 and two novel Vpa2-like genes, Vpa2-like1 and Vpa2-like2, under the baculovirus p10 promoter in transfected Sf9 cells. Pairwise amino acid analyses revealed a higher percentage of identity and a lower number of gaps between Vpa2Ac1 and Vpa2-like2 than to Vpa2-like1. Moreover, Vpa2-like1 lacked the conserved Ser-Thr-Ser motif, involved in NAD binding, and the (F/Y)xx(Q/E)xE consensus sequence, characteristic of the ARTT toxin family involved in actin polymerization. Vpa2Ac1, Vpa2-like1 and Vpa2-like2 transcripts and proteins were detected in Sf9 culture cells, but the signals of Vpa2Ac1 and Vpa2-like2 were weak and decreased over time. Sf9 cells infected by a recombinant bacmid expressing Vpa2-like1 showed typical circular morphology and produced viral occlusion bodies (OBs) at the same level as the control virus. However, expression of Vpa2Ac1 and Vpa2-like2 induced cell polarization, similar to that produced by the microfilament-destabilizing agent cytochalasin D and OBs were not produced. The presence of filament disrupting agents, such as nicotinamide and nocodazole, during transfection prevented cell polarization and OB production was observed. We conclude that Vpa2Ac1 and Vpa2-like2 proteins likely possess ADP-ribosyltransferase activity that modulated actin polarization, whereas Vpa2-like1 is not a typical Vpa2 protein. Vpa2-like2 has now been designated Vpa2Ca1 (accession number AAO86513) by the Bacillus thuringiensis delta-endotoxin nomenclature committee.
Open Access
Gender-mediated differences in vertical transmission of a nucleopolyhedrovirus
(Public Library of Science, 2013) Virto Garayoa, Cristina; Zárate Chaves, Carlos Andrés; López Ferber, Miguel; Murillo Pérez, Rosa; Caballero Murillo, Primitivo; Williams, Trevor; Nekazaritza Ekoizpena; Producción Agraria; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua
With the development of sensitive molecular techniques for detection of low levels of asymptomatic pathogens, it becoming clear that vertical transmission is a common feature of some insect pathogenic viruses, and likely to be essential to virus survival when opportunities for horizontal transmission are unfavorable. Vertical transmission of Spodoptera exigua multiple nucleopolyhedrovirus (SeMNPV) is common in natural populations of S. exigua. To assess whether gender affected transgenerational virus transmission, four mating group treatments were performed using healthy and sublethally infected insects: i) healthy males (H=)6healthy females (HR); ii) infected males (I=)6healthy females (HR); iii) healthy males (H=)6infected females (IR) and iv) infected males (I=)6infected females (IR). Experimental adults and their offspring were analyzed by qPCR to determine the prevalence of infection. Both males and females were able to transmit the infection to the next generation, although female-mediated transmission resulted in a higher prevalence of infected offspring. Malemediated venereal transmission was half as efficient as maternally-mediated transmission. Egg surface decontamination studies indicated that the main route of transmission is likely transovarial rather than transovum. Both male and female offspring were infected by their parents in similar proportions. Incorporating vertically-transmitted genotypes into virusbased insecticides could provide moderate levels of transgenerational pest control, thereby extending the periods between bioinsecticide applications.
Open Access
A qPCR assay for the quantification of selected genotypic variants of spodoptera frugiperda multiple nucleopolyhedrovirus (Baculoviridae)
(MDPI, 2024-05-20) Molina-Ruiz, Cindy S.; Zamora-Briseño, Jesús Alejandro; Simón de Goñi, Oihane; Lasa, Rodrigo; Williams, Trevor; Institute for Multidisciplinary Research in Applied Biology - IMAB
Alphabaculoviruses are lethal dsDNA viruses of Lepidoptera that have high genetic diversity and are transmitted in aggregates within proteinaceous occlusion bodies. This mode of transmission has implications for their efficacy as biological insecticides. A Nicaraguan isolate of Spodoptera frugiperda multiple nucleopolyhedrovirus (SfMNPV-NIC) comprising nine genotypic variants has been the subject of considerable study due to the influence of variant interactions on the insecticidal properties of mixed-variant occlusion bodies. As part of a systematic study on the replication and transmission of variant mixtures, a tool for the accurate quantification of a selection of genotypic variants was developed based on the quantitative PCR technique (qPCR). First, primer pairs were designed around a region of high variability in four variants named SfNic-A, SfNic-B, SfNic-C and SfNic-E to produce amplicons of 103–150 bp. Then, using cloned purified amplicons as standards, amplification was demonstrated over a dynamic range of 108–101 copies of each target. The assay was efficient (mean ± SD: 98.5 ± 0.8%), reproducible, as shown by low inter- and intra-assay coefficients of variation (<5%), and specific to the target variants (99.7–100% specificity across variants). The quantification method was validated on mixtures of genotype-specific amplicons and demonstrated accurate quantification. Finally, mixtures of the four variants were quantified based on mixtures of budded virions and mixtures of DNA extracted from occlusion-derived virions. In both cases, mixed-variant preparations compared favorably to total viral genome numbers by quantification of the polyhedrin (polh) gene that is present in all variants. This technique should prove invaluable in elucidating the influence of variant diversity on the transmission and insecticidal characteristics of this pathogen.
Open Access
Coocclusion of Helicoverpa armigera single nucleopolyhedrovirus (HearSNPV) and Helicoverpa armigera multiple nucleopolyhedrovirus (HearMNPV): pathogenicity and stability in homologous and heterologous hosts
(MDPI, 2022) Arrizubieta Celaya, Maite; Simón de Goñi, Oihane; Ricarte Bermejo, Adriana; López Ferber, Miguel; Williams, Trevor; Caballero Murillo, Primitivo; Institute for Multidisciplinary Research in Applied Biology - IMAB; Gobierno de Navarra / Nafarroako Gobernua
Helicoverpa armigera single nucleopolyhedrovirus (HearSNPV) is a virulent pathogen of lepidopterans in the genera Heliothis and Helicoverpa, whereas Helicoverpa armigera multiple nu-cleopolyhedrovirus (HearSNPV) is a different virus species with a broader host range. This study aimed to examine the consequences of coocclusion of HearSNPV and HearMNPV on the patho-genicity, stability and host range of mixed-virus occlusion bodies (OBs). HearSNPV OBs were approximately 6-fold more pathogenic than HearMNPV OBs, showed faster killing by approximately 13 h, and were approximately 45% more productive in terms of OB production per larva. For coocclusion, H. armigera larvae were first inoculated with HearMNPV OBs and subsequently inoculated with HearSNPV OBs at intervals of 0-72 h after the initial inoculation. When the interval between inoculations was 12-24 h, OBs collected from virus-killed insects were found to comprise 41¿57% of HearSNPV genomes, but the prevalence of HearSNPV genomes was greatly reduced (3- 4%) at later time points. Quantitative PCR (qPCR) analysis revealed the presence of HearSNPV genomes in a small fraction of multinucleocapsid ODVs representing 0.47¿0.88% of the genomes quan-tified in ODV samples, indicating that both viruses had replicated in coinfected host cells. End-point dilution assays on ODVs from cooccluded mixed-virus OBs confirmed the presence of both viruses in 41.9¿55.6% of wells that were predicted to have been infected by a single ODV. A control exper-iment indicated that this result was unlikely to be due to the adhesion of HearSNPV ODVs to HearMNPV ODVs or accidental contamination during ODV band extraction. Therefore, the dispar-ity between the qPCR and end-point dilution estimates of the prevalence of mixed-virus ODVs likely reflected virus-specific differences in replication efficiency in cell culture and the higher in-fectivity of pseudotyped ODVs that were produced in coinfected parental cells. Bioassays on H. armigera, Spodoptera frugiperda and Mamestra brassicae larvae revealed that mixed-virus OBs were capable of infecting heterologous hosts, but relative potency values largely reflected the proportion of HearMNPV present in each mixed-virus preparation. The cooccluded mixtures were unstable in serial passage; HearSNPV rapidly dominated during passage in H. armigera whereas HearMNPV rapidly dominated during passage in the heterologous hosts. We conclude that mixed-virus coocclusion technology may be useful for producing precise mixtures of viruses with host range properties suitable for the control of complexes of lepidopteran pests in particular crops, although this requires validation by field testing.
Open Access
Stability of a Spodoptera frugiperda nucleopolyhedrovirus deletion recombinant during serial passage in insects
(American Society for Microbiology, 2009) Simón de Goñi, Oihane; Williams, Trevor; Possee, Robert D.; López Ferber, Miguel; Caballero Murillo, Primitivo; Nekazaritza Ekoizpena; Producción Agraria; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua
The stabilities of the Spodoptera frugiperda multiple nucleopolyhedrovirus (SfMNPV) complete genome bacmid (Sfbac) and a deletion recombinant (Sf29null) in which the Sf29 gene was replaced by a kanamycin resistance cassette were determined during sequential rounds of per os infection in insect larvae. The Sf29 gene is a viral factor that determines the number of virions in occlusion bodies (OBs). The Sf29null bacmid virus was able to recover the Sf29 gene during passage. After the third passage (P3) of Sf29null bacmid OBs, the population was observed to reach an equilibrium involving a mixture of those with a kanamycin resistance cassette and those with the Sf29 gene. The biological activity of Sf29null bacmid OBs at P3 was similar to that of Sfbac OBs. The recovered gene in the Sf29null virus was 98 to 100% homologous to the Sf29 genes of different SfMNPV genotypes. Reverse transcription-PCR analysis of uninoculated S. frugiperda larvae confirmed the expression of the SfMNPV ie-0 and Sf29 genes, indicating that the insect colony harbors a covert SfMNPV infection. Additionally, the nonessential bacterial artificial chromosome vector was spontaneously deleted from both viral genomes upon passage in insects.
Open Access
Iflavirus covert infection increases susceptibility to nucleopolyhedrovirus disease in Spodoptera exigua
(MDPI, 2020) Carballo Palos, Arkaitz; Williams, Trevor; Murillo Pérez, Rosa; Caballero Murillo, Primitivo; Agronomia, Bioteknologia eta Elikadura; Institute for Multidisciplinary Research in Applied Biology - IMAB; Agronomía, Biotecnología y Alimentación
Naturally occurring covert infections in lepidopteran populations can involve multiple viruses with potentially different transmission strategies. In this study, we characterized covert infection by two RNA viruses, Spodoptera exigua iflavirus 1 (SeIV-1) and Spodoptera exigua iflavirus 2 (SeIV-2) (family Iflaviridae) that naturally infect populations of Spodoptera exigua, and examined their influence on susceptibility to patent disease by the nucleopolyhedrovirus Spodoptera exigua multiple nucleopolyhedrovirus (SeMNPV) (family Baculoviridae). The abundance of SeIV-1 genomes increased up to ten-thousand-fold across insect developmental stages after surface contamination of host eggs with a mixture of SeIV-1 and SeIV-2 particles, whereas the abundance of SeIV-2 remained constant across all developmental stages. Low levels of SeIV-2 infection were detected in all groups of insects, including those that hatched from surface-decontaminated egg masses. SeIV-1 infection resulted in reduced larval weight gain, and an unbalanced sex ratio, whereas larval developmental time, pupal weight, and adult emergence and fecundity were not significantly affected in infected adults. The inoculation of S. exigua egg masses with iflavirus, followed by a subsequent infection with SeMNPV, resulted in an additive effect on larval mortality. The 50% lethal concentration (LC50) of SeMNPV was reduced nearly 4-fold and the mean time to death was faster by 12 h in iflavirus-treated insects. These results suggest that inapparent iflavirus infections may be able to modulate the host response to a new pathogen, a finding that has particular relevance to the use of SeMNPV as the basis for biological pest control products.
Open Access
Superinfection exclusion in alphabaculovirus infections is concomitant with actin reorganization
(American Society for Microbiology, 2014) Beperet Arive, Inés; Irons, Sarah L.; Simón de Goñi, Oihane; King, Linda A.; Williams, Trevor; Possee, Robert D.; López Ferber, Miguel; Caballero Murillo, Primitivo; Nekazaritza Ekoizpena; Producción Agraria; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua
Superinfection exclusion is the ability of an established virus to interfere with a second virus infection. This effect was studied in vitro during lepidopteran-specific nucleopolyhedrovirus (genus Alphabaculovirus, family Baculoviridae) infection. Homologous interference was detected in Sf9 cells sequentially infected with two genotypes of Autographa californica multiple nucleopolyhedrovirus (AcMNPV), each one expressing a different fluorescent protein. This was a progressive process in which a sharp decrease in the signs of infection caused by the second virus was observed, affecting not only the number of coinfected cells observed, but also the level of protein expression due to the second virus infection. Superinfection exclusion was concurrent with reorganization of cytoplasmic actin to F-actin in the nucleus, followed by budded virus production (16 to 20 h postinfection). Disruption of actin filaments by cell treatment with cytochalasin D resulted in a successful second infection. Protection against heterologous nucleopolyhedrovirus infection was also demonstrated, as productive infection of Sf9 cells by Spodoptera frugiperda nucleopolyhedrovirus (SfMNPV) was inhibited by prior infection with AcMNPV, and vice versa. Finally, coinfected cells were observed following inoculation with mixtures of these two phylogenetically distant nucleopolyhedroviruses—AcMNPV and SfMNPV—but at a frequency lower than predicted, suggesting interspecific virus interference during infection or replication. The temporal window of infection is likely necessary to maintain genotypic diversity that favors virus survival but also permits dual infection by heterospecific alphabaculoviruses.
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.