Artículos de revista IdAB - IdAB Aldizkari artikuluak

Permanent URI for this collection

http://hdl.handle.net/2454/10687

Browse

Now showing 1 - 20 of 213

Open Access
Complete genome sequence of five Chrysodeixis chalcites nucleopolyhedrovirus genotypes from a Canary Islands isolate
(American Society for Microbiology, 2013-10-24) Bernal Rodríguez, Alexandra; Williams, Trevor; Muñoz Labiano, Delia; Caballero Murillo, Primitivo; Simón de Goñi, Oihane; Producción Agraria; Nekazaritza Ekoizpena; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua; Gobierno de Navarra / Nafarroako Gobernua
The Chrysodeixis chalcites single nucleopolyhedrovirus (ChchSNPV) infects and kills C. chalcites larvae, an important pest of banana crops in the Canary Islands. Five genotypes present in the most prevalent and widespread isolate in the Canary Islands were sequenced, providing genetic data relevant to the genotypic and phenotypic diversity of this virus.
Open Access
Remarkably efficient production of a highly insecticidal Chrysodeixis chalcites nucleopolyhedrovirus (ChchNPV) isolate in its homologous host
(Wiley, 2018-01-03) Bernal Rodríguez, Alexandra; Simón de Goñi, Oihane; Williams, Trevor; Muñoz Labiano, Delia; Caballero Murillo, Primitivo; Producción Agraria; Nekazaritza Ekoizpena; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua
Background: a Chrysodeixis chalcites nucleopolyhedrovirus from the Canary Islands (ChchNPV-TF1) has proved to be effective for control of Chrysodeixis chalcites on banana crops. Commercialization of this virus as a bioinsecticide requires an efficient production system. Results: the sixth instar (L6) was the most suitable for virus production, producing 1.80 × 1011 occlusion bodies (OB)/larva and showed a lower prevalence of cannibalism (5.4%) than fourth (L4) or fifth (L5) instars. Inoculation of L6 at 24 h post molting produced six times more OB (5.72 × 1011 OB/larva) than recently molted L6 larvae (1.00 × 1011 OB/larva). No significant differences were recorded in mean time to death (165–175 h) or OB production per larva (3.75 × 1011 to 5.97 × 1011) or per mg larval weight (1.30 × 1011 to 2.11 × 109), in larvae inoculated with a range of inoculum concentrations (LC50–LC90). Groups of infected L6 larvae reared at a density of 150 larvae/container produced a greater total number of OBs (8.07 × 1013 OB/container) than lower densities (25, 50 and 100 OB/container), and a similar number to containers with 200 inoculated larvae (8.43 × 1013 OB/container). Conclusion: the processes described here allow efficient production of sufficient OBs to treat ∼ 40 ha of banana crops using the insects from a single container.
Open Access
Chrysodeixis chalcites, a pest of banana crops on the Canary Islands: incidence, economic losses and current control measures
(Elsevier, 2018-03-05) Fuentes Barrera, Ernesto Gabriel; Hernández Suárez, Estrella; Simón de Goñi, Oihane; Williams, Trevor; Caballero Murillo, Primitivo; Producción Agraria; Nekazaritza Ekoizpena; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua
Chrysodeixis chalcites is an emergent pest in bananas (Musa acuminata Colla) grown on the Canary Islands. Feeding damage to leaves and fruit and the control measures targeted at this pest were evaluated over a two-year period (2013–2014). The prevalence of infestations (42–100%) on the islands was similar during the two years of the study. Mean foliar damage (1.5–7.3% depending on island) and fruit damage (1.0–5.7%) detected in field surveys varied significantly across islands, plantation aspect (north- or south-facing) and season. Fruit damage was not correlated with foliar damage (P > 0.05). The weight of C. chalcites damaged bananas varied significantly (0.2–4.2% of harvested fruit) across islands, particularly in the spring. Overall, 3155 tonnes of bananas/yr are likely discarded due to C. chalcites damage, representing 1.5% of annual production or 2.68 million €/yr. The most frequently used pesticide was indoxacarb, usually applied on three occasions per crop cycle, for which the cost of control measures would average 240 €/ha per crop cycle. The direct damage that C. chalcites causes to banana fruit results in significant economic losses in addition to the direct costs of pesticide based control measures. Effective and sustainable control strategies are required against this pest.
Open Access
Selection of a nucleopolyhedrovirus isolate from Helicoverpa armigera as the basis for a biological insecticide
(Wiley, 2014-05-01) Arrizubieta Celaya, Maite; Williams, Trevor; Caballero Murillo, Primitivo; Simón de Goñi, Oihane; Producción Agraria; Nekazaritza Ekoizpena; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua; Gobierno de Navarra / Nafarroako Gobernua
Background: the cotton bollworm, Helicoverpa armigera, is an insect that causes damage in a wide range of crops in Spain.Seven isolates of H. armigera single nucleopolyhedrovirus (HearSNPV) from the Iberian Peninsula were subjected to molecularand biological characterization and compared with a Chinese genotype (HearSNPV-G4). Results: the estimated sizes of the Iberian genomes varied between 116.2 and 132.4 kb, compared to 131.4 kb of theHearSNPV-G4 reference genome. Phylogenetic analysis based on the lef-8, lef-9 and polh genes revealed that the Iberianstrains were more closely related to one another than to other HearSNPV isolates. Occlusion body (OB) concentration-mortalityresponses (LC 50 values) did not differ signiﬁcantly among Iberian isolates when tested against a Helicoverpa armigera colonyfrom Oxford (UK). Despite being the fastest killing isolate, HearSNPV-SP1 was as productive as isolates with lower virulence,with an average yield of 3.1 × 109 OBs larva−1 . OBs of HearSNPV-SP1 and HearSNPV-G4 were similarly pathogenic against arecently established colony from southern Spain, although HearSNPV-SP1 was faster killing than HearSNPV-G4 against a rangeof instars. Conclusion: the insecticidal properties of HearSNPV-SP1 mean that this strain is likely to prove useful as the basis for abiological insecticide for control of Helicoverpa armigera in Spain.
Open Access
A native variant of Chrysodeixis chalcites nucleopolyhedrovirus: the basis for a promising bioinsecticide for control of C. chalcites in Canary Islands' banana crops
(Elsevier, 2013-08-13) Bernal Rodríguez, Alexandra; Williams, Trevor; Hernández Suárez, Estrella; Carnero, Aurelio; Caballero Murillo, Primitivo; Simón de Goñi, Oihane; Producción Agraria; Nekazaritza Ekoizpena; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua
Chrysodeixis chalcites (Lepidoptera: Noctuidae) larvae cause up to 30% production loss in banana crops in the Canary Islands. Larvae of this species are susceptible to a nucleopolyhedrovirus (ChchNPV). This study aimed at evaluating the genetic diversity and bioinsecticidal activity of ChchNPV isolates collected from C. chalcites larvae in the Canary Islands. From a total 97 isolates collected in different banana greenhouses, restriction endonuclease analysis identified five genetic variants that differed slightly from ChchNPV isolates from Netherlands (ChchSNPV-NL) and Almería, Spain (ChchNPV-SP1). Physical maps revealed minimal differences at the genome level, mostly due to variation in the position/existence of restriction sites. ChchSNPV-TF1 was the most prevalent variant, representing 78% of isolates examined, and was isolated at all Canary Island sampling sites. This isolate was the most pathogenic isolate against C. chalcites second instars in terms of concentration-mortality metrics, compared to homologous variants or two heterologous viruses Autographa californica multiple nucleopolyhedrovirus (AcMNPV) and Anagrapha falcifera multiple nucleopolyhedrovirus (AnfaMNPV). ChchSNPV-TF1 was also one of the fastest killing variants although no differences were observed in occlusion body production among the different variants in second instars. We conclude that ChchSNPV-TF1 merits further evaluation as the basis for a biological insecticide for control of C. chalcites in banana crops in the Canary Islands.
Open Access
Analysis of a naturally-occurring deletion mutant of Spodoptera frugiperda multiple nucleopolyhedrovirus reveals sf58 as a new per os infectivity factor of lepidopteran-infecting baculoviruses
(Elsevier, 2012-10-21) Simón de Goñi, Oihane; Palma Dovis, Leopoldo; Williams, Trevor; López Ferber, Miguel; Caballero Murillo, Primitivo; Producción Agraria; Nekazaritza Ekoizpena; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua; Gobierno de Navarra / Nafarroako Gobernua
The Nicaraguan population of Spodoptera frugiperda multiple nucleopolyhedrovirus, SfMNPV-NIC, is structured as a mixture of nine genotypes (A–I). Occlusion bodies (OBs) of SfMNPV-C, -D and -G pure genotypes are incapable of oral transmission; a phenotype which in SfMNPV-C and -D is due to the absence of pif1 and pif2 genes. The complete sequence of the SfMNPV-G genome was determined to identify possible factors involved in this phenotype. Deletions of 4860 bp (22,366–27,225) and 60 bp (119,759–119,818) were observed in SfMNPV-G genome compared with that of the predominant complete genotype SfMNPV-B (132,954 bp). However no genes homologous to previously described per os infectivity factors were located within the deleted sequences. Significant differences were detected in the nucleotide sequence in sf58 gene (unknown function) that produced changes in the amino acid sequence and the predicted secondary structure of the corresponding protein. This gene is conserved only in lepidopteran baculoviruses (alpha- and betabaculoviruses). To determine the role of sf58 in peroral infectivity a deletion mutant was constructed using bacmid technology. OBs of the deletion mutant (Sf58null) were not orally infectious for S. frugiperda larvae, whereas Sf58null rescue virus OBs recovered oral infectivity. Sf58null DNA and occlusion derived virions (ODVs) were as infective as SfMNPV bacmid DNA and ODVs in intrahemocelically infected larvae or cell culture, indicating that defects in ODV or OB morphogenesis were not involved in the loss of peroral infectivity. Addition of optical brightener or the presence of the orally infectious SfMNPV-B OBs in mixtures with SfMNPV-G OBs did not recover Sf58null OB infectivity. According to these results sf58 is a new per os infectivity factor present only in lepidopteran baculoviruses.
Open Access
Occlusion body pathogenicity, virulence and productivity traits vary with transmission strategy in a nucleopolyhedrovirus
(Elservier, 2011-10-25) Cabodevilla de Andrés, Oihana; Ibáñez Elosua, Itxaso; Simón de Goñi, Oihane; Murillo Pérez, Rosa; Caballero Murillo, Primitivo; Williams, Trevor; Producción Agraria; Nekazaritza Ekoizpena; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua
The prevalence of sublethal infections of Spodoptera exigua multiple nucleopolyhedrovirus (SeMNPV) was quantified in natural populations of S. exigua in Almería, Spain, during 2006 and 2007. Of 1045 adults collected, 167 (16.1%) proved positive for viral polyhedrin gene transcripts by RT-PCR. The prevalence of covert infection varied significantly according to sex and sample date. Of 1660 progeny of field-collected insects, lethal disease was observed in 10¿33% of offspring of transcript-positive females and 9¿49% of offspring of transcript-negative females. Isolates associated with vertically transmitted infections were characterized by restriction endonuclease analysis using BglII or EcoRV and compared with isolates originating from greenhouse soil-substrate believed to be horizontally transmitted. Insects from a sublethally infected Almerian colony were between 2.3-fold and 4.6-fold more susceptible to infection than healthy insects from a Swiss colony, depending on isolate. Horizontally transmitted isolates were significantly more pathogenic than vertically transmitted isolates in insects from both colonies. Mean speed of kill in second instars (Swiss colony) varied between isolates by >20 h, whereas mean occlusion body (OB) production in fourth instars (Swiss colony) varied by 3.8-fold among isolates. Intriguingly, all three horizontally transmitted isolates were very similar in speed of kill and OB production, whereas all three vertically transmitted isolates differed significantly from one another in both variables, and also differed significantly from the group of horizontally transmitted isolates in speed of kill (one isolate) or both variables (two isolates). We conclude that key pathogenicity and virulence traits of SeMNPV isolates vary according to their principal transmission strategy.
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
Population genetic structure determine the virulence and transmissibility of Spodoptera frugiperda multiple necleopolyhedrovirus
(Elsevier, 2007-12-28) Simón de Goñi, Oihane; Williams, Trevor; López Ferber, Miguel; Taulemesse, Jean-Marie; Caballero Murillo, Primitivo; Producción Agraria; Nekazaritza Ekoizpena; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua
A Nicaraguan isolate of Spodoptera frugiperda multiple nucleopolyhedrovirus (SfNIC) survives as a complex mixture of genotypes (named A to I). The speed of kill, time-mortality distribution, and occlusion body (OB) production of single genotypes (A, B and F) and co-occluded mixtures of genotypes, in a 75% + 25% ratio, were compared to determine the contribution of each genotype to the transmissibility of the viral population. Pure genotypes differed markedly in their speed of kill in second instar S. frugiperda. The speed of kill of SfNIC was attenuated compared to that of the dominant genotype B, indicating that interactions involving two or more genotypes likely determine host killing traits in the virus population. Genotypes A, F and defective genotype C, had no significant effects on the distribution of insect deaths over time when present as minority components in mixtures comprising 75% of genotype B. Similarly, the mortality pattern over time of insects infected by genotype F, the fastest-killing genotype tested, was not affected by the presence of genotypes A or C. Semi-quantitative PCR studies indicated that the genetic composition did not differ significantly between SfNIC-infected insects that died soon (67 h) or late (139 h) after inoculation, suggesting that stability in genotypic composition is important for virus survival. Median OB production per insect was correlated with mean time to death so that attenuated speed of kill of SfNIC resulted in high OB yields. We conclude that (i) minority genotypes play a functional role in determining the timing of mortality of infected hosts and (ii) the genotypic structure of the virus population is stably maintained to maximize the likelihood of survival.
Open Access
Sequence comparison between three geographically distinct Spodoptera frugiperda multiple nucleopolyhedrovirus isolates: detecting positively selected genes
(Elsevier, 2011-01-14) Simón de Goñi, Oihane; Palma Dovis, Leopoldo; Beperet Arive, Inés; Muñoz Labiano, Delia; López Ferber, Miguel; Caballero Murillo, Primitivo; Williams, Trevor; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua
The complete genomic sequence of a Nicaraguan plaque purified Spodoptera frugiperda nucleopolyhedrovirus (SfMNPV) genotype SfMNPV-B was determined and compared to previously sequenced isolates from United States (SfMNPV-3AP2) and Brazil (SfMNPV-19). The genome of SfMNPV-B (132,954 bp) was 1623 bp and 389 bp larger than that of SfMNPV-3AP2 and SfMNPV-19, respectively. Genome size differences were mainly due to a deletion located in the SfMNPV-3AP2 egt region and small deletions and point mutations in SfMNPV-19. Nucleotide sequences were strongly conserved (99.35% identity) and a high degree of predicted amino acid sequence identity was observed. A total of 145 open reading frames (ORFs) were identified in SfMNPV-B, two of them (sf39a and sf110a) had not been previously identified in the SfMNPV-3AP2 and SfMNPV-19 genomes and one (sf57a) was absent in both these genomes. In addition, sf6 was not previously identified in the SfMNPV-19 genome. In contrast, SfMNPV-B and SfMNPV-19 both lacked sf129 that had been reported in SfMNPV-3AP2. In an effort to identify genes potentially involved in virulence or in determining population adaptations, selection pressure analysis was performed. Three ORFs were identified undergoing positive selection: sf49 (pif-3), sf57 (odv-e66b) and sf122 (unknown function). Strong selection for ODV envelope protein genes indicates that the initial infection process in the insect midgut is one critical point at which adaptation acts during the transmission of these viruses in geographically distant populations. The function of ORF sf122 is being examined.
Open Access
Use of recombinant iron-superoxide dismutase as a marker of nitrative stress
(Elservier, 2008-04-20) Larrainzar Rodríguez, Estíbaliz; Urarte Rodríguez, Estíbaliz; Auzmendi, Iñigo; Ariz Arnedo, Idoia; Arrese-Igor Sánchez, César; González García, Esther; Morán Juez, José Fernando; Ciencias del Medio Natural; Natura Ingurunearen Zientziak; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua; Gobierno de Navarra / Nafarroako Gobernua, 57/2007
Superoxide dismutases (SODs; EC 1.15.1.1) are a group of metalloenzymes which are essential to protect cells under aerobic conditions. In biological systems, it has been reported that SODs and other proteins are susceptible to be attacked by peroxynitrite (ONOO-) which can be originated from the reaction of nitric oxide with superoxide radical. ONOO- is a strong oxidant molecule capable of nitrating peptides and proteins at the phenyl side chain of the tyrosine residues. In the present work, bovine serum albumin (BSA) and recombinant iron¿superoxide dismutase from the plant cowpea (Vu_FeSOD) are used as target molecules to estimate ONOO- production. The method employs the compound SIN-1, which simultaneously generates -NO and O2- in aerobic aqueous solutions. First, assay conditions were optimized incubating BSA with different concentrations of SIN-1, and at a later stage, the effect on the tyrosine nitration and catalytic activity of Vu_FeSOD was examined by in-gel activity and spectrophotometric assays. Both BSA and Vu_FeSOD are nitrated in a dose-dependent manner, and, at least in BSA nitration, the reaction seems to be metal catalyzed.
Open Access
Expression and localization of a Rhizobium-derived cambialistic superoxide dismutase in pea (Pisum sativum) nodules subjected to oxidative stress
(The American Phytopathological Society, 2011-09-07) Asensio, Aarón C.; Marino Bilbao, Daniel; James, Euan K.; Ariz Arnedo, Idoia; Arrese-Igor Sánchez, César; Aparicio Tejo, Pedro María; Arredondo-Peter, Raúl; Morán Juez, José Fernando; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua; Ciencias del Medio Natural; Natura Ingurunearen Zientziak
Two phylogenetically unrelated superoxide dismutase (SOD) families, i.e., CuZnSOD (copper and zinc SOD) and FeMn-CamSOD (iron, manganese, or cambialistic SOD), eliminate superoxide radicals in different locations within the plant cell. CuZnSOD are located within the cytosol and plastids, while the second family of SOD, which are considered to be of bacterial origin, are usually located within organelles, such as mitochondria. We have used the reactive oxygen species¿producer methylviologen (MV) to study SOD isozymes in the indeterminate nodules on pea (Pisum sativum). MV caused severe effects on nodule physiology and structure and also resulted in an increase in SOD activity. Purification and N-terminal analysis identified CamSOD from the Rhizobium leguminosarum endosymbiont as one of the most active SOD in response to the oxidative stress. Fractionation of cell extracts and immunogold labeling confirmed that the CamSOD was present in both the bacteroids and the cytosol (including the nuclei, plastids, and mitochondria) of the N-fixing cells, and also within the uninfected cortical and interstitial cells. These findings, together with previous reports of the occurrence of FeSOD in determinate nodules, indicate that FeMnCamSOD have specific functions in legumes, some of which may be related to signaling between plant and bacterial symbionts, but the occurrence of one or more particular isozymes depends upon the nodule type.
Open Access
Foliar application of urea to "Sauvignon Blanc" and "Merlot" vines: doses and time of application
(Springer Nature, 2012-02-19) Lasa Larrea, Berta; Menéndez Villanueva, Sergio; Sagastizabal, Kepa; Calleja Cervantes, María Eréndira; Irigoyen Iriarte, Ignacio; Muro Erreguerena, Julio; Aparicio Tejo, Pedro María; Ariz Arnedo, Idoia; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua; Producción Agraria; Nekazaritza Ekoizpena
A careful control of the N nutritional status of grapevines can have a determining effect on wine characteristics; therefore a suitable management of N fertilization might allow some wine parameters to be modified, thereby improving product quality. The aim of this study was to determine the effect of foliar application of urea at different doses and different times of the growing season on the parameters of Sauvignon Blanc and Merlot grape juice. The research described herein involved Sauvignon Blanc and Merlot grapevines (V. vinifera L.) at a commercial vineyard and was conducted over 2 years. In the first year, N treatment involved a foliar application at a dose of 10 kg N ha−1 during veraison, whereas in the second year it involved a foliar urea application at two doses (10 and 50 kg N ha−1) and at three different times—3 weeks before veraison, during veraison and 3 weeks after veraison. In this second year, the urea applied at a dose of 10 kg N ha−1 was isotopically labelled with 1% 15N. Chemical parameters, yeast assimilable N, amino acid content, amino acid profile and N isotopic composition were determined for all treatments. Grape and grape-juice parameters for Merlot were found to be more affected by N fertilization than for Sauvignon Blanc and were also more affected during the second year than during the first year, thus indicating that the climatic characteristics of each campaign could affect these parameters. The yeast assimilable N in grape juice was found to be higher for late applications of foliar urea, with application of the higher dose of urea during veraison increasing the amino acid and proline contents in both varieties. The isotopic analysis data showed that the urea applied to leaves was transferred to the berries, with the maximum translocation in Sauvignon Blanc occurring for the post-veraison treatment and in Merlot for the veraison treatment. We can therefore conclude that foliar application of urea could modify grape juice quality and could therefore be used as a tool for obtaining quality wines.
Open Access
The physiological implications of urease inhibitors on N metabolism during germination of Pisum sativum and Spinacea oleracea seeds
(Elsevier, 2012-03-08) Ariz Arnedo, Idoia; Cruchaga Moso, Saioa; Lasa Larrea, Berta; Morán Juez, José Fernando; Jáuregui Mosquera, Iván; Aparicio Tejo, Pedro María; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua; Ciencias del Medio Natural; Natura Ingurunearen Zientziak
The development of new nitrogen fertilizers is necessary to optimize crop production whilst improving the environmental aspects arising from the use of nitrogenous fertilization as a cultural practice. The use of urease inhibitors aims to improve the efficiency of urea as a nitrogen fertilizer by preventing its loss from the soil as ammonia. However, although the action of urease inhibitors is aimed at the urease activity in soil, their availability for the plant may affect its urease activity. The aim of this work was therefore to evaluate the effect of two urease inhibitors, namely acetohydroxamic acid (AHA) and N-(n-butyl) thiophosphoric triamide (NBPT), on the germination of pea and spinach seeds. The results obtained show that urease inhibitors do not affect the germination process to any significant degree, with the only process affected being imbibition in spinach, thus also suggesting different urease activities for both plants. Our findings therefore suggest an activity other than the previously reported urolytic activity for urease in spinach. Furthermore, of the two inhibitors tested, NBPT was found to be the most effective at inhibiting urease activity, especially in pea seedlings.
Open Access
High irradiance induces photoprotective mechanisms and a positive effect on NH4+ stress in Pisum sativum L.
(Elsevier, 2010-04-29) Ariz Arnedo, Idoia; Esteban Terradillos, Raquel; García Plazaola, José Ignacio; Becerril, José María; Aparicio Tejo, Pedro María; Morán Juez, José Fernando; Ciencias; Zientziak; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua
Photosynthesis provides plant metabolism with reduced carbon (C) but is also the main source of oxidative stress in plants. Likewise, high doses of NH4+ as sole N source have been reported to be toxic for most plants, resulting in reduced plant growth and restricting C availability. The combination of high photosynthetic photon flux densities (PPFD) and NH4+ nutrition may provide higher C availability but could also have a detrimental effect on the plants, therefore the objective of this study is to evaluate whether NH4+ induces photo-oxidative stress that is exacerbated under high light conditions. Pea plants (Pisum sativum cv. sugar-snap) were grown hydroponically with NH4+ (0.5, 2.5, 5 and 10 mM) under high (750 μmol photons m−2 s−1) or low PPFD conditions (350 μmol photons m−2 s−1). High PPFD contributes to a higher tolerance to ammonium by pea plants, as it originated higher biomass content due to higher photosynthetic rates. However, a deficit of N (0.5 and 2.5 mM NH4+) under high PPFD conditions caused an antioxidant response, as indicated by increased photoprotective pigment and chloroplastic superoxide dismutase contents. Plants grown with higher doses of N and high PPFD showed less need for photoprotection. An increase in the specific leaf weight (SLW) ratio was observed associated not only with high PPFDs but also with the highest NH4+ dose. Overall, these results demonstrate that, despite the activation of some photoprotective responses at high PPFD, there were no photoinhibitory symptoms and a positive effect on NH4+ toxicity, thus suggesting that the harmful effects of NH4+ are not directly related to the generation of photo-oxidative stress.
Open Access
High irradiance increases NH4+ tolerance in Pisum sativum: higher carbon and energy availability improve ion balance but not N assimilation
(Elsevier, 2011-03-02) Ariz Arnedo, Idoia; Artola Rezola, Ekhiñe; Asensio, Aarón C.; Cruchaga Moso, Saioa; Aparicio Tejo, Pedro María; Morán Juez, José Fernando; Ciencias del Medio Natural; Natura Ingurunearen Zientziak; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua; Institute for Multidisciplinary Research in Applied Biology - IMAB
The widespread use of NO3− fertilization has had a major ecological impact. NH4+ nutrition may help to reduce this impact, although high NH4+ concentrations are toxic for most plants. The underlying tolerance mechanisms are not yet fully understood, although they are thought to include the limitation of C, the disruption of ion homeostasis, and a wasteful NH4+ influx/efflux cycle that carries an extra energetic cost for root cells. In this study, high irradiance (HI) was found to induce a notable tolerance to NH4+ in the range 2.5–10 mM in pea plants by inducing higher C availability, as shown by carbohydrate content. This capacity was accompanied by a general lower relative N content, indicating that tolerance is not achieved through higher net N assimilation on C-skeletons, and it was also not attributable to increased GS content or activity in roots or leaves. Moreover, HI plants showed higher ATP content and respiration rates. This extra energy availability is related to the internal NH4+ content regulation (probably NH4+ influx/efflux) and to an improvement of the cell ionic balance. The limited C availability at lower irradiance (LI) and high NH4+ resulted in a series of metabolic imbalances, as reflected in a much higher organic acid content, thereby suggesting that the origin of the toxicity in plants cultured at high NH4+ and LI is related to their inability to avoid large-scale accumulation of the NH4+ ion.
Open Access
Short term physiological implications of NBPT application on the N metabolism of Pisum sativum and Spinacea oleracea
(Elsevier, 2011-03-01) Cruchaga Moso, Saioa; Artola Rezola, Ekhiñe; Lasa Larrea, Berta; Ariz Arnedo, Idoia; Irigoyen Iriarte, Ignacio; Morán Juez, José Fernando; Aparicio Tejo, Pedro María; Ciencias del Medio Natural; Natura Ingurunearen Zientziak; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua; Producción Agraria; Nekazaritza Ekoizpena
The application of urease inhibitors in conjunction with urea fertilizers as a means of reducing N loss due to ammonia volatilization requires an in-depth study of the physiological effects of these inhibitors on plants. The aim of this study was to determine how the urease inhibitor N-(n-butyl) thiophosphoric triamide (NBPT) affects N metabolism in pea and spinach. Plants were cultivated in pure hydroponic culture with urea as the sole N source. After 2 weeks of growth for pea, and 3 weeks for spinach, half of the plants received NBPT in their nutrient solution. Urease activity, urea and ammonium content, free amino acid composition and soluble protein were determined in leaves and roots at days 0, 1, 2, 4, 7 and 9, and the NBPT content in these tissues was determined 48 h after inhibitor application. The results suggest that the effects of NBPT on spinach and pea urease activity differ, with pea being most affected by this treatment, and that the NBPT absorbed by the plant caused a clear inhibition of the urease activity in pea leaf and roots. The high urea concentration observed in leaves was associated with the development of necrotic leaf margins, and was further evidence of NBPT inhibition in these plants. A decrease in the ammonium content in roots, where N assimilation mainly takes place, was also observed. Consequently, total amino acid contents were drastically reduced upon NBPT treatment, indicating a strong alteration of the N metabolism. Furthermore, the amino acid profile showed that amidic amino acids were major components of the reduced pool of amino acids. In contrast, NBPT was absorbed to a much lesser degree by spinach plants than pea plants (35% less) and did not produce a clear inhibition of urease activity in this species.
Open Access
Effect of N-(n-butyl) thiophosphoric triamide on urea metabolism and the assimilation of ammonium by Triticum aestivum L.
(Springer, 2010-08-25) Artola Rezola, Ekhiñe; Cruchaga Moso, Saioa; Ariz Arnedo, Idoia; Morán Juez, José Fernando; Garnica, María; Houdusse, Fabrice; García Mina, José M.; Irigoyen Iriarte, Ignacio; Lasa Larrea, Berta; Aparicio Tejo, Pedro María; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua; Producción Agraria; Nekazaritza Ekoizpena
The use of urea as an N fertilizer has increased to such an extent that it is now the most widely used fertilizer in the world. However, N losses as a result of ammonia volatilization lead to a decrease in its efficiency, therefore different methods have been developed over the years to reduce these losses. One of the most recent involves the use of urea combined with urease inhibitors, such as N-(n-butyl) thiophosphoric triamide (NBPT), in an attempt to delay the hydrolysis of urea in the soil. The aim of this study is to perform an in-depth analysis of the effects that NBPT use has on plant growth and N metabolism. Wheat plants were cultivated in a greenhouse experiment lasting four weeks and fertilized with urea and NBPT at different concentrations (0, 0.012, 0.062, 0.125%). Each treatment was replicated six times. A non-fertilized control was also cultivated. Several parameters related with N metabolism were analysed at harvest. NBPT use was found to have visible effects, such as a transitory yellowing of the leaf tips, at the end of the first week of treatment. At a metabolic level, plants treated with the inhibitor were found to have more urea in their tissues and a lower amino acid content, lower glutamine synthetase activity, and lower urease and glutamine synthetase content at the end of the study period, whereas their urease activity seemed to have recovered by this stage.
Open Access
A self-induction method to produce high quantities of recombinant functional flavo-leghemoglobin reductase
(Elsevier, 2008-01-29) Urarte Rodríguez, Estíbaliz; Auzmendi, Iñigo; Rol, Selene; Ariz Arnedo, Idoia; Aparicio Tejo, Pedro María; Arredondo-Peter, Raúl; Morán Juez, José Fernando; Institute for Multidisciplinary Research in Applied Biology - IMAB; Gobierno de Navarra / Nafarroako Gobernua
Ferric leghemoglobin reductase (FLbR) is able to reduce ferric leghemoglobin (Lb3+) to ferrous (Lb2+) form. This reaction makes Lb functional in performing its role since only reduced hemoglobins bind O2. FLbR contains FAD as prosthetic group to perform its activity. FLbR-1 and FLbR-2 were isolated from soybean root nodules and it has been postulated that they reduce Lb3+. The existence of Lb2+ is essential for the nitrogen fixation process that occurs in legume nodules; thus, the isolation of FLbR for the study of this enzyme in the nodule physiology is of interest. However, previous methods for the production of recombinant FLbR are inefficient as yields are too low. We describe the production of a recombinant FLbR-2 from Escherichia coli BL21(DE3) by using an overexpression method based on the self-induction of the recombinant E. coli. This expression system is four times more efficient than the previous overexpression method. The quality of recombinant FLbR-2 (based on spectroscopy, SDS-PAGE, IEF, and native PAGE) is comparable to that of the previous expression system. Also, FLbR-2 is purified near to homogeneity in only few steps (in a time scale, the full process takes 3 days). The purification method involves affinity chromatography using a Ni-nitrilotriacetic acid column. Resulting rFLbR-2 showed an intense yellow color, and spectral characterization of rFLbR-2 indicated that rFLbR-2 contains flavin. Pure rFLbR-2 was incubated with soybean Lba and NADH, and time drive rates showed that rFLbR-2 efficiently reduces Lb3+.
Open Access
Expression of recombinant proteins lacking methionine as N-terminal amino acid in plastids: human serum albumin as a case study
(Elsevier, 2007) Fernández San Millán, Alicia; Farrán Blanch, Inmaculada; Molina Azcona, Andrea; Mingo Castel, Ángel; Veramendi Charola, Jon; IdAB. Instituto de Agrobiotecnología / Agrobioteknologiako Institutua