Infectivity decline of an RNA plant virus by increased mutagenesis supports the lethal defection model in vivo

Lethal mutagenesis is a new antiviral therapy based on increasing the mutation rate by using mutagenic base and nucleoside analogues whose molecular mechanisms are not fully understood. Most of the research has been conducted on animal RNA viruses in cell culture and, to a lesser extent, in vivo. There is experimental evidence supporting the model of lethal defection for lethal mutagenesis of RNA viruses. In this model, viral genomes with a low degree of mutation and low specific infectivity, termed "defectors", exert an interfering activity leading to virus loss. Lethal mutagenesis of plant viruses has not been addressed yet despite being excellent in vivo model systems that develop systemic infections, undergo rapid bottlenecks and pose no ethical issues. Here, we address lethal mutagenesis in vivo of Tobacco mosaic virus (TMV), a single-stranded positive RNA virus of 6.4 Kb. Nicotiana tabacum plants cultured in vitro were treated with 25, 50 and 100 µg/ml of the base analogue 5-fluorouracil (FU) and 24 h later were inoculated with 50 lesion forming units (lfu) of TMV. We analyzed the infectivity, viral load and mutant spectra of viral populations after 5 and 10 days of treatment, as well as of populations that went 10 days of treatment followed by 21 days of ex vitro growth in the absence of FU. The results show that TMV infectivity decreases when treated with 50 and 100 µg/ml FU for 10 days. TMV mutagenized populations grown without FU reach infectivity values higher than untreated populations. Predominant mutations in FU-treated populations with decreased infectivity at 10 dpi are U→C, A→G and G→A transitions, which are expected due to the action of FU. TMV replication is not affected by FU at any dose and there are no imbalances of ribonucleotide triphosphate pools measured by HPLC. No differences in mutation frequencies and Shannon Entropies between control and FU-treated populations with declined infectivity were found. However, we did found a dose-dependent decrease of specific infectivity in FU-treated populations, but not in untreated samples, as well as dominance of molecules with a low degree of mutation. Specific infectivity recovered to control levels after 21 days of growth without the analogue. Altogether, our results suggest that TMV defector molecules mediate the decrease in TMV infectivity. This is the first report that addresses the molecular basis of lethal defection in vivo using an RNA plant virus.Junta de Andalucía (P09-CVI-5428 y P10-CVI-6561), Plan Nacional I+D+i (BFU2007-65080 BMC) y Universidad de Málaga (Plan propio

Infectivity decline of an RNA plant virus by increased mutagenesis in vivo

According to the lethal defection model, during lethal mutagenesis viral genomes with a low degree of mutation and low specific infectivity exert an interfering activity that leads to virus loss. Lethal mutagenesis of plant viruses has not been reported to date. We address lethal defection in vivo of Tobacco mosaic virus (TMV), a single-stranded positive RNA virus. Nicotiana tabacum plants cultured in vitro were treated with 25, 50 and 100 µg/ml of the base analogue 5-fluorouracil (FU) and 24h later were inoculated with 50 lesion forming units of TMV. Results show that TMV infectivity decreases when treated with 50 and 100 µg/ml FU for 10 days. TMV mutagenized populations grown without FU reach infectivity values higher than untreated populations. Predominant mutations in FU-treated populations with decreased infectivity at 10 dpi are transitions which are expected due to the action of FU. Viral load is not affected by FU at any dose and there are no imbalances of ribonucleotide triphosphate pools measured by HPLC. No differences in mutation frequencies and Shannon Entropies between control and FU-treated populations were found. However, we found a dose-dependent decrease of specific infectivity in FU-treated populations, but not in untreated samples, as well as dominance of molecules with a low degree of mutation. Specific infectivity recovered to control levels after 21 days of growth without the analogue. Altogether, our results suggest that TMV defector molecules mediate the decrease in TMV infectivity. This is the first report that addresses the molecular basis of lethal defection in vivo using an RNA plant virus.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Soluble Receptor Isoform of IFN-Beta (sIFNAR2) in Multiple Sclerosis Patients and Their Association With the Clinical Response to IFN-Beta Treatment

Alternative splicing; Soluble receptors; IFNAR; Interferon beta; Multiple sclerosisEmpalmament alternatiu; Receptors solubles; IFNAR; Interferó beta; Esclerosi múltipleSplicing alternativo; Receptores solubles; IFNAR; Interferón beta; Esclerosis múltiplePurpose: Interferon beta receptor 2 subunit (IFNAR2) can be produced as a transmembrane protein, but also as a soluble form (sIFNAR2) generated by alternative splicing or proteolytic cleavage, which has both agonist and antagonist activities for IFN-β. However, its role regarding the clinical response to IFN-β for relapsing-remitting multiple sclerosis (RRMS) is unknown. We aim to evaluate the in vitro short-term effects and after 6 and 12 months of IFN-β therapy on sIFNAR2 production and their association with the clinical response in MS patients. Methods: Ninety-four RRMS patients were included and evaluated at baseline, 6 and 12 months from treatment onset. A subset of 41 patients were classified as responders and non-responders to IFN-β therapy. sIFNAR2 serum levels were measured by ELISA. mRNA expression for IFNAR1, IFNAR2 splice variants, MxA and proteases were assessed by RT-PCR. The short-term effect was evaluated in PBMC from RRMS patients after IFN-β stimulation in vitro. Results: Protein and mRNA levels of sIFNAR2 increased after IFN-β treatment. According to the clinical response, only non-responders increased sIFNAR2 significantly at both protein and mRNA levels. sIFNAR2 gene expression correlated with the transmembrane isoform expression and was 2.3-fold higher. While MxA gene expression increased significantly after treatment, IFNAR1 and IFNAR2 only slightly increased. After short-term IFN-β in vitro induction of PBMC, 6/7 patients increased the sIFNAR2 expression. Conclusions: IFN-β administration induces the production of sIFNAR2 in RRMS and higher levels might be associated to the reduction of therapeutic response. Thus, levels of sIFNAR2 could be monitored to optimize an effective response to IFN-β therapy.This research was funded by grants from the Instituto de Salud Carlos III and co-funded by European Regional Development Fund (ERDF), Technological Development Project in health DTS/1800045 to BO-M. BO-M holds a contract from Red Andaluza de Investigacion Clínica y Traslacional en Neurología (Neuro-reca) (RIC-0111-2019). PA-G is supported by Promoción de Empleo Joven e Implantación de la Garantía Juvenil 2018 (PEJ2018-002719-A). JR-B is supported by grants from Red Temática de Investigación Cooperativa, Red Española de Esclerosis Multiple REEM (RD16/0015/0010). LL holds a Nicolás Monardes research contract (RC-002-2019) from the Andalusian Ministry of Health and Family. IB-M holds a pFIS contract (FI19/00139) from the Spanish Science and Innovation Ministry

Group selection and social evolution in domesticated animals

Social interactions, especially those involving competition among individuals, are important in domesticated livestock and in natural populations. The heritability of traits affected by such interactions has two components, one originating in the individual like that of classical traits (direct effects) and the other originating in other group members (indirect effects). The latter type of trait represents a significant source of ‘hidden heritability’ and it requires population structure and knowledge from relatives in order to access it for selective breeding. When ignored, competitive interactions may increase as an indirect response to direct selection, resulting in diminished yields. We illustrate how population genetic structure affects the response to selection of traits with indirect genetic effects using population genetic and quantitative genetic theory. Population genetic theory permits us to connect our results to the existing body of theory on kin and group selection in natural populations. The quantitative genetic perspective allows us to see how breeders have used knowledge from relatives and family selection in the domestication of plants and animals to improve the welfare and production of livestock by incorporating social genetic effects in the breeding program. We illustrate the central features of these models by reviewing empirical studies from domesticated chickens

Genetic parameters for social effects on survival in cannibalistic layers: Combining survival analysis and a linear animal model

<p>Abstract</p> <p>Background</p> <p>Mortality due to cannibalism in laying hens is a difficult trait to improve genetically, because censoring is high (animals still alive at the end of the testing period) and it may depend on both the individual itself and the behaviour of its group members, so-called associative effects (social interactions). To analyse survival data, survival analysis can be used. However, it is not possible to include associative effects in the current software for survival analysis. A solution could be to combine survival analysis and a linear animal model including associative effects. This paper presents a two-step approach (2STEP), combining survival analysis and a linear animal model including associative effects (LAM).</p> <p>Methods</p> <p>Data of three purebred White Leghorn layer lines from Institut de Sélection Animale B.V., a Hendrix Genetics company, were used in this study. For the statistical analysis, survival data on 16,780 hens kept in four-bird cages with intact beaks were used. Genetic parameters for direct and associative effects on survival time were estimated using 2STEP. Cross validation was used to compare 2STEP with LAM. LAM was applied directly to estimate genetic parameters for social effects on observed survival days.</p> <p>Results</p> <p>Using 2STEP, total heritable variance, including both direct and associative genetic effects, expressed as the proportion of phenotypic variance, ranged from 32% to 64%. These results were substantially larger than when using LAM. However, cross validation showed that 2STEP gave approximately the same survival curves and rank correlations as LAM. Furthermore, cross validation showed that selection based on both direct and associative genetic effects, using either 2STEP or LAM, gave the best prediction of survival time.</p> <p>Conclusion</p> <p>It can be concluded that 2STEP can be used to estimate genetic parameters for direct and associative effects on survival time in laying hens. Using 2STEP increased the heritable variance in survival time. Cross validation showed that social genetic effects contribute to a large difference in survival days between two extreme groups. Genetic selection targeting both direct and associative effects is expected to reduce mortality due to cannibalism in laying hens.</p

Newly rare or newly common: evolutionary feedbacks through changes in population density and relative species abundance, and their management implications

Environmental management typically seeks to increase or maintain the population sizes of desirable species and to decrease population sizes of undesirable pests, pathogens, or invaders. With changes in population size come long-recognized changes in ecological processes that act in a density-dependent fashion. While the ecological effects of density dependence have been well studied, the evolutionary effects of changes in population size, via changes in ecological interactions with community members, are underappreciated. Here, we provide examples of changing selective pressures on, or evolution in, species as a result of changes in either density of conspecifics or changes in the frequency of heterospecific versus conspecific interactions. We also discuss the management implications of such evolutionary responses in species that have experienced rapid increases or decreases in density caused by human actions

Natalizumab-immunogenicity evaluation in patients with infusion related events or disease exacerbations

IntroductionNatalizumab is a biologic drug for relapsing-remitting multiple sclerosis that may induce the generation of anti-drug antibodies in some patients. Anti-natalizumab antibodies (ANA) increase the risk of adverse events and reduce efficacy, being useful biomarkers for monitoring treatment response.MethodsRetrospective observational study including MS patients treated with natalizumab that experienced infusion-related events (IRE) or disease exacerbations (DE). ANA were tested by Elisa including a screening and a confirmation assay. Patients were further classified as transient (one positive result) or persistent (two or more positive results) ANA.ResultsA total of 1251 MS patients were included and 153 (12.3%) had ANA with at least one single point determination, which were more frequent among patients with IRE compared to those with DE (21,6% vs.10.8%) during the first six infusions. Two or more determinations ANA were performed in 184 patients, being 31.5% permanently positive and 7.1% transiently positive. Interestingly, 26.1% of patients that experienced DE had persistent ANA, while 2.6% were transient. In contrast, 43% of patients with IRE had persistent ANA, and 9.3% had transient antibodies. Patients with persistent antibodies had more frequently high levels at the first sampling compared to patients with transient ANA.ConclusionReal-world evidence shows that the presence of ANA is behind an important percentage of patients treated with natalizumab that experience IRE, as well as DE but in a lower degree. These findings support the need to systematically evaluate ANA towards a personalized management of these patients to avoid undesired complications

Towards a catalogue of genes transcribed in multiple tissues of Mytilus galloprovincialis.

Mussels are among the most studied marine organisms because of their ecological role, economic importance and long-standing use in coastal pollution biomonitoring. Despite the bulk of data derived from traditional investigation areas, only limited knowledge is available on mussel genes and their expression in ordinary or stressing conditions. We present here the first data obtained by production and sequencing of 3\u2032 end-specific expressed sequence tags (3\u2032ESTs) from multiple tissues of putatively unstressed mussels (Mytilus galloprovincialis). A total of 524 clusters (98 virtual consensuses and 426 singletons) derived from 829 reliable 3\u2032ESTs were searched in the non-redundant (nr) sequence databases (National Centre for Biotechnology Information (Bethesda, USA), NCBI). Most contigs (59%) showed poor or no similarity, thus revealing unknown mussel genes, other contigs recognized some of the few available records of M. galloprovincialis (e.g. actin, cytochrome oxidase III and twitchin) and a number of significant similarities with mitochondrial (mt) or nuclear genes from other organisms were found. Actually, a variety of ribosomal as well as motility- and adhesion-related genes and some genes potentially involved in stress responses (e.g. myticin A, heat shock proteins, methallothionein) were putatively identified. Uniquely, mitochondrial transcripts were primarily represented by a cluster of 123 3\u2032ESTs (1296 base pairs (bp) of the mussel 16S rRNA). In our knowledge, these results provide the first systematic production and annotation of ESTs in M. galloprovincialis. They also represent the first report of a wider project, based on the strategy of 3\u2032EST identification from normal and stressed mussels and intended to define a mussel complementary DNA (cDNA) microarray for genome-wide transcription studies

Soluble Receptor Isoform of IFN-Beta (sIFNAR2) in Multiple Sclerosis Patients and Their Association With the Clinical Response to IFN-Beta Treatment

Purpose: Interferon beta receptor 2 subunit (IFNAR2) can be produced as a transmembrane protein, but also as a soluble form (sIFNAR2) generated by alternative splicing or proteolytic cleavage, which has both agonist and antagonist activities for IFN-b. However, its role regarding the clinical response to IFN-b for relapsing-remitting multiple sclerosis (RRMS) is unknown. We aim to evaluate the in vitro short-term effects and after 6 and 12 months of IFN-b therapy on sIFNAR2 production and their association with the clinical response in MS patients. Methods: Ninety-four RRMS patients were included and evaluated at baseline, 6 and 12 months from treatment onset. A subset of 41 patients were classified as responders and non-responders to IFN-b therapy. sIFNAR2 serum levels were measured by ELISA. mRNA expression for IFNAR1, IFNAR2 splice variants, MxA and proteases were assessed by RT-PCR. The short-term effect was evaluated in PBMC from RRMS patients after IFN-b stimulation in vitro. Results: Protein and mRNA levels of sIFNAR2 increased after IFN-b treatment. According to the clinical response, only non-responders increased sIFNAR2 significantly at both protein and mRNA levels. sIFNAR2 gene expression correlated with the transmembrane isoform expression and was 2.3-fold higher. While MxA gene expression increased significantly after treatment, IFNAR1 and IFNAR2 only slightly increased. After short-term IFN-b in vitro induction of PBMC, 6/7 patients increased the sIFNAR2 expression. Conclusions: IFN-b administration induces the production of sIFNAR2 in RRMS and higher levels might be associated to the reduction of therapeutic response. Thus, levels of sIFNAR2 could be monitored to optimize an effective response to IFN-b therapy.This research was funded by grants from the Instituto de Salud Carlos III and co-funded by European Regional Development Fund (ERDF), Technological Development Project in health DTS/1800045 to BO-M. BO-M holds a contract from Red Andaluza de Investigacion Clınica y Traslacional en Neurología (Neuro-reca) ́ (RIC-0111-2019). PA-G is supported by Promoción de Empleo Joven e Implantación de la Garantıa Juvenil 2018 (PEJ2018-002719- ́A). JR-B is supported by grantsfrom Red Temática de Investigación Cooperativa, Red Española de Esclerosis Multiple REEM (RD16/0015/0010). LL holds a Nicolás Monardes research contract (RC 002-2019) from the Andalusian Ministry of Health and Family. IB M holds a pFIS contract (FI19/00139)from the Spanish Science and Innovation Ministry.Ye