BUSCADOR RECOLECTA

The bacterium Bacillus thuringiensis produces insecticidal Vip3 proteins during the vegetative growth phase with activity against several lepidopteran pests. To date, three different Vip3 protein families have been identified based on sequence identity: Vip3A, Vip3B, and Vip3C. In this study, we report the construction of chimeras by exchanging domains between Vip3Aa and Vip3Ca, two proteins with marked specificity differences against lepidopteran pests. We found that some domain combinations made proteins insoluble or prone to degradation by trypsin as most abundant insect gut protease. The soluble and trypsin-stable chimeras, along with the parental proteins Vip3Aa and Vip3Ca, were tested against lepidopteran pests from different continents: Spodoptera exigua, Spodoptera littoralis, Spodoptera frugiperda, Helicoverpa armigera, Mamestra brassicae, Anticarsia gemmatalis, and Ostrinia furnacalis. The exchange of the Nt domain (188 N-terminal amino acids) had little effect on the stability and toxicity (equal or slightly lower) of the resulting chimeric protein against all insects except for S. frugiperda, for which the chimera with the Nt domain from Vip3Aa and the rest of the protein from Vip3Ca showed a significant increase in toxicity compared to the parental Vip3Ca. Chimeras with the C-terminal domain from Vip3Aa (from amino acid 510 of Vip3Aa to the Ct) with the central domain of Vip3Ca (amino acids 189–509 based on the Vip3Aa sequence) made proteins that could not be solubilized. Finally, the chimera including the Ct domain of Vip3Ca and the Nt and central domain from Vip3Aa was unstable. Importantly, an insect species tolerant to Vip3Aa but susceptible to Vip3Ca, such as Ostrinia furnacalis, was also susceptible to chimeras maintaining the Ct domain from Vip3Ca, in agreement with the hypothesis that the Ct region of the protein is the one conferring specificity to Vip3 proteins., This research was supported to JF by the Spanish Ministry of Economy and Competitivity (Grants Ref. AGL2015‐70584‐C02‐1‐R and RTI2018‐095204‐B‐C21), by the Generalitat Valenciana (GVPROMETEOII‐2015‐ 001), and by European FEDER funds. JGC was recipient of a PhD grant from the Spanish Ministry of Economy and Competitivity (grant ref. BES‐2013‐065848 and EEBB‐I‐17‐12367). Support was also provided to JLJ‐F by an Agriculture and Food Research Initiative Foundational Program competitive grant (No. 2018‐67013‐27820) from the USDA National Institute of Food and Agriculture, and to KH by a grant 'Key Project for Breeding Genetically Modified Organisms' from China (grant ref. 2016ZX08003‐001).

Bacillus thuringiensis (Bt) produces insecticidal proteins that are either secreted during the vegetative growth phase or accumulated in the crystal inclusions (Cry proteins) in the stationary phase. Cry1I proteins share the three domain (3D) structure typical of crystal proteins but are secreted to the media early in the stationary growth phase. In the generally accepted mode of action of 3D Cry proteins (sequential binding model), the formation of an oligomer (tetramer) has been described as a major step, necessary for pore formation and subsequent toxicity. To know if this could be extended to Cry1I proteins, the formation of Cry1Ia oligomers was studied by Western blot, after the incubation of trypsin activated Cry1Ia with insect brush border membrane vesicles (BBMV) or insect cultured cells, using Cry1Ab as control. Our results showed that Cry1Ia oligomers were observed only after incubation with susceptible coleopteran BBMV, but not following incubation with susceptible lepidopteran BBMV or non-susceptible Sf21 insect cells, while Cry1Ab oligomers were persistently detected after incubation with all insect tissues tested, regardless of its host susceptibility. The data suggested oligomerization may not necessarily be a requirement for the toxicity of Cry1I proteins., This work was supported by grants from the Spanish Ministry of Science, Innovation and Universities, the State Research Agency of Spain and the European FEDER founds (Refs. AGL2015-70584-C2 and RTI2018-095204-B-C21), and by the Generalitat Valenciana (GVPROMETEOII-2015-001). M. Domínguez received a predoctoral fellowship from the Universidad Pública de Navarra, Spain.

The genome of the Bacillus thuringiensis BM311.1 strain was sequenced and assembled in 359 contigs containing a total of 6,390,221 bp. The plasmidic ORF of a putative cry gene from this strain was identified as a potential novel Cry protein of 1138 amino acid residues with a 98% identity compared to Cry7Aa1 and a predicted molecular mass of 129.4 kDa. The primary structure of Cry7Aa2, which had eight conserved blocks and the classical structure of three domains, differed in 28 amino acid residues from that of Cry7Aa1. The cry7Aa2 gene was amplified by PCR and then expressed in the acrystalliferous strain BMB171. SDS-PAGE analysis confirmed the predicted molecular mass for the Cry7Aa2 protein and revealed that after in vitro trypsin incubation, the protein was degraded to a toxin of 62 kDa. However, when treated with digestive fluids from Leptinotarsa decemlineata larvae, one major proteinase-resistant fragment of slightly smaller size was produced. The spore and crystal mixture produced by the wild-type BM311.1 strain against L. decemlineata neonate larvae resulted in a LC50 value of 18.8 mu g/mL, which was statistically similar to the estimated LC50 of 20.8 mu g/mL for the recombinant BMB17-Cry7Aa2 strain. In addition, when this novel toxin was activated in vitro with commercial trypsin, the LC50 value was reduced 3.8-fold to LC50 = 4.9 mu g/mL. The potential advantages of Cry7Aa2 protoxin compared to Cry7Aa1 protoxin when used in the control of insect pests are discussed., This research was funded by the Programa Nacional de España (No. AGL2015-70584-C2-2-R) and the Gobierno de Navarra (No. IIQ14065: RI1).

Bacillus thuringiensis (Bt) se distingue de otras bacterias del género Bacillus por su
capacidad de sintetizar d-endotoxinas (Cry y Cyt) que característicamente se
agregan formando uno o más cristales paraesporales. Todas estas proteínas han
sido clasificadas en distintas familias las cuales tienen actividad contra insectos
de distintos órdenes, incluido el orden Coleoptera. En esta tesis doctoral se ha
realizado una revisión actualizada de todas las proteínas insecticidas, que
produce Bt, para las cuales se ha descrito actividad contra especies del orden
Coleoptera. El objetivo de la tesis ha sido realizar una búsqueda de nuevos genes
cry con la finalidad de aumentar la batería de toxinas Bt disponibles para el
control de las plagas causadas por coleópteros. Dicha búsqueda se ha centrado
en dos cepas Bt, seleccionadas en un screening previo, para lo cual se abordó la
secuenciación masiva del DNA genómico de cada una de ellas.
El contenido génico de la cepa BM311.1 reveló la presencia un gen nuevo cry7
que codificaba para una proteína que se denominó Cry7Aa2 por compartir una
identidad del 98% con la proteína Cry7Aa1 previamente descrita. A pesar de esta
elevada identidad, estas dos proteínas mostraron diferencias en su actividad
contra larvas de Leptinotarsa decemlineata. Mientras que la proteína Cry7Aa1 solo
es tóxica cuando el cristal ha sido previamente solubilizado in vitro, la proteína
Cry7Aa2 es tóxica directamente tras ser ingerida en forma de cristal. Esto
representa una clara ventaja práctica de Cry7Aa2, con vistas a su utilización como
bioinsecticida, mientras que ambas son igualmente útiles para la obtención de
plantas transgénicas resistentes frente a L. decemlineata.
Los resultados de este trabajo son científicamente relevantes y pueden ser
utilizados para el diseño de nuevas estrategias para el control de plagas de
insectos, ya sea creando nuevos bioinsecticidas o construyendo nuevas plantas
transgénicas., Bacillus thuringiensis (Bt) is distinguished from other bacteria of the genus Bacillus
for its ability to synthesize d-endotoxins (Cry and Cyt) that characteristically
aggregate forming one or more parasporal crystals. All these proteins have been
classified into different families which have activity against insects of different
orders, including the order Coleoptera. In this doctoral thesis, an up-to-date
review of all the insecticidal proteins produced by Bt has been carried out, for
which their activity against coleopteran species has been described. The aim of
the thesis was to initiate a search for new cry genes in order to increase the
number of Bt toxins available for the control of pests caused by Coleoptera. This
study has been focused on two Bt strains, selected in a previous screening, for
which sequencing of their genomic DNA was performed.
The gene content of strain BM311.1 revealed the presence of a new cry7 gene that
was named cry7Aa2 since it shared a 98% identity at the amino acid level with
the previously described Cry7Aa1 protein. Despite this high identity, these two
proteins showed differences in their activity against Leptinotarsa decemlineata
larvae. While the Cry7Aa1 protein was only toxic when the crystal had been
previously solubilized in vitro, the Cry7Aa2 protein showed toxicity directly after
being ingested as a crystal. This represents a clear practical advantage of Cry7Aa2
over Cry7Aa1 towards its use as a bioinsecticide, while both are equally useful
for obtaining transgenic plants resistant to L. decemlineata.
The results of this thesis are scientifically relevant and can be used to design new
strategies for the control of insect pests, either by creating new bioinsecticides or
by engineering new transgenic Bt-plants., Beca de formación de personal investigador de la Universidad Pública de Navarra (autor, adscrito a los proyectos AGL 2015-70 584-C2-2-R y RTI 2018-095 204-B-C22)., Programa de Doctorado en Biotecnología (RD 99/2011), Bioteknologiako Doktoretza Programa (ED 99/2011)

Here, we report the draft genome sequence of Photorhabdus luminescens strain DSPV002N, which consists of 177 contig sequences accounting for 5,518,143 bp, with a G+C content of 42.3% and 4,701 predicted protein-coding genes (CDSs). From these, 27 CDSs exhibited significant similarity with insecticidal toxin proteins from Photorhabdus luminescens subsp. laumondii TT01., This research was supported by the Spanish Ministry of Economy and Competitiveness (grant AGL2015-70584-C2-2-R).

Mindfulness and meditation techniques have proven successful for the reduction of stress and improvement in general health. In addition, meditation is linked to longevity and longer telomere length, a proposed biomarker of human aging. Interestingly, DNA methylation changes have been described at specific subtelomeric regions in long-term meditators compared to controls. However, the molecular basis underlying these beneficial effects of meditation on human health still remains unclear. Here we show that DNA methylation levels, measured by the Infinium HumanMethylation450 BeadChip (Illumina) array, at specific subtelomeric regions containing GPR31 and SERPINB9 genes were associated with telomere length in long-term meditators with a strong statistical trend when correcting for multiple testing. Notably, age showed no association with telomere length in the group of long-term meditators. These results may suggest that long-term meditation could be related to epigenetic mechanisms, in particular gene-specific DNA methylation changes at distinct subtelomeric regions., The project has received funding from the Network for Prevention and Health Promotion in primary Care (RD12/0005 and RD16/0007/0005) grant and a FIS grant (PI16/00962) from the Instituto de Salud Carlos III of the Spanish Ministry of Economy and Competitiveness, co-financed with European Union ERDF funds. MM has received a grant 'intensificación' from Fundación LaCaixa.

The present study compared the effects of traditional resistance training (TRT) and combined power training (PT) and TRT (PTRT) on cognitive parameters and serum brain-derived neurotrophic factor (BDNF) levels in non-demented, well-functioning, community-dwelling older women. Forty-five older women were randomized into one of three experimental groups: TRT, PTRT, and control group (CG). Cognitive tests explored global cognitive function, short-term memory, and dual-task performance. Serum BDNF levels were assessed at baseline and after the intervention. Exercise sessions were performed twice a week over 22 weeks. In TRT, exercise sessions were based on three sets of 8–10 repetitions at 'difficult' intensity. In PTRT, the first session was based on PT (three sets of 8−10 repetitions at 'moderate' intensity), while the second session was similar to the TRT. Our analyses indicated that overall cognitive function, short-term memory, and dual-task performance were similarly improved after TRT and PTRT. Serum BDNF concentrations were not altered by any training protocol. In conclusion, the two RT programs tested in the present trial improved global cognitive function, short-term memory and dual task performance in non-demented, well-functioning, community-dwelling older women. In addition, our findings suggest that mechanisms other than BDNF may be associated with such improvements., This work was supported by Innovative Medicines Initiative–Joint Undertaking [IMI–JU #115621], the nonprofit research foundation 'Centro Studi Achille e Linda Lorenzon', and by a scholarship to H.J.C.-J. from the Coordenação de Aperfeiçoamento de Pessoal de Nível Superior [CAPES; Finance Code 001].

Here, we report the draft genome sequence of Photorhabdus luminescensstrain DSPV002N, which consists of 177 contig sequences
accounting for 5,518,143 bp, with a GC content of 42.3% and 4,701 predicted protein-coding genes (CDSs). From these, 27 CDSs exhibited significant similarity with insecticidal toxin proteins from Photorhabdus luminescenssubsp. laumondii TT01., This research was supported by the Spanish Ministry of Economy and
Competitiveness (grant AGL2015-70584-C2-2-R).
We thank the High-Throughput Genomics Group at the Wellcome
Trust Centre for Human Genetics (funded by Wellcome Trust grant reference 090532/Z/09/Z) for the generation of sequencing data., Peer reviewed