Developmental stages and episode-specific regulatory genes in andromonoecious melon flower development.

BACKGROUND AND AIMS: Given the lack of specific studies on floral development in melon (Cucumis melo L.), we carried out an extensive study involving morphological and transcriptomic analyses to characterize floral development in this species. METHODS: Using an andromonoecious line, we analysed the development of floral buds in male and hermaphrodite flowers with both light microscopy and scanning electron microscopy. Based on flower lengths, we established a correlation between the developmental stages and four main episodes of floral development and conducted an extensive RNA sequencing analysis of these episodes. KEY RESULTS: We identified 12 stages of floral development, from the appearance of the floral meristems to anthesis. The main structural differences between male and hermaphrodite flowers appeared between stages 6 and 7; later stages of development leading to the formation of organs and structures in both types of flowers were also described. We analysed the gene expression patterns of the four episodes in flower development to find the genes that were specific to each given episode. Among others, we identified genes that defined the passage from one episode to the next according to the ABCDE model of floral development. CONCLUSIONS: This work combines a detailed morphological analysis and a comprehensive transcriptomic study to enable characterization of the structural and molecular mechanisms that determine the floral development of an andromonoecious genotype in melon. Taken together, our results provide a first insight into gene regulation networks in melon floral development that are crucial for flowering and pollen formation, highlighting potential targets for genetic manipulation to improve crop yield of melon in the future.

Advances in Understanding the Mechanism of Cap-Independent Cucurbit Aphid-Borne Yellows Virus Protein Synthesis.

Non-canonical translation mechanisms have been described for many viral RNAs. In the case of several plant viruses, their protein synthesis is controlled by RNA elements in their genomic 3'-ends that are able to enhance cap-independent translation (3'-CITE). The proposed general mechanism of 3'-CITEs includes their binding to eukaryotic translation initiation factors (eIFs) that reach the 5'-end and AUG start codon through 5'-3'-UTR-interactions. It was previously shown that cucurbit aphid-borne yellows virus (CABYV) has a 3'-CITE, which varies in sequence and structure depending on the phylogenetic group to which the isolate belongs, possibly as a result of adaptation to the different geographical regions. In this work, the cap-independent translation mechanisms of two CABYV 3'-CITEs belonging to the Mediterranean (CMTE) and Asian (CXTE) groups, respectively, were studied. In vivo cap-independent translation assays show that these 3'-CITEs require the presence of the CABYV short genomic 5'-UTR with at least 40% adenines in cis and an accessible 5'-end for its activity. Additionally, they suggest that the eIF4E-independent CABYV 3'-CITE activities may not require either eIF4A or the eIF4F complex, but may depend on eIF4G and PABP. By pulling down host proteins using RNA baits containing both 5'- and 3'-CABYV-UTRs, 80 RNA binding proteins were identified. These interacted preferentially with either CMTE, CXTE, or both. One of these proteins, specifically interacting with the RNA containing CMTE, was HSP70.2. Preliminary results suggested that HSP70.2 may be involved in CMTE- but not CXTE-mediated cap-independent translation activity.

Characterization of Two Aggressive PepMV Isolates Useful in Breeding Programs.

Pepino mosaic virus (PepMV) causes significant economic losses in tomato crops worldwide. Since its first detection infecting tomato in 1999, aggressive PepMV variants have emerged. This study aimed to characterize two aggressive PepMV isolates, PepMV-H30 and PepMV-KLP2. Both isolates were identified in South-Eastern Spain infecting tomato plants, which showed severe symptoms, including bright yellow mosaics. Full-length infectious clones were generated, and phylogenetic relationships were inferred using their nucleotide sequences and another 35 full-length sequences from isolates representing the five known PepMV strains. Our analysis revealed that PepMV-H30 and PepMV-KLP2 belong to the EU and CH2 strains, respectively. Amino acid sequence comparisons between these and mild isolates identified 8 and 15 amino acid substitutions for PepMV-H30 and PepMV-KLP2, respectively, potentially involved in severe symptom induction. None of the substitutions identified in PepMV-H30 have previously been described as symptom determinants. The E236K substitution, originally present in the PepMV-H30 CP, was introduced into a mild PepMV-EU isolate, resulting in a virus that causes symptoms similar to those induced by the parental PepMV-H30 in Nicotiana benthamiana plants. In silico analyses revealed that this residue is located at the C-terminus of the CP and is solvent-accessible, suggesting its potential involvement in CP-host protein interactions. We also examined the subcellular localization of PepGFPm2E236K in comparison to that of PepGFPm2, focusing on chloroplast affection, but no differences were observed in the GFP subcellular distribution between the two viruses in epidermal cells of N. benthamiana plants. Due to the easily visible symptoms that PepMV-H30 and PepMV-KLP2 induce, these isolates represent valuable tools in programs designed to breed resistance to PepMV in tomato.

A regulatory role for the redox status of the pepino mosaic virus coat protein.

Cysteine oxidations play important regulatory roles during animal virus infections. Despite the importance of redox modifications during plant infections, no plant virus protein has yet been shown to be regulated by cysteine oxidation. The potexvirus pepino mosaic virus (PepMV) is pandemic in tomato crops. Previously we modeled the structure of the PepMV particle and coat protein (CP) by cryo-electron microscopy and identified critical residues of the CP RNA-binding pocket that interact with the viral RNA during particle formation and viral cell-to-cell movement. The PepMV CP has a single cysteine residue (Cys127) central to its RNA binding pocket, which is highly conserved. Here we show that the Cys127Ser replacement diminishes PepMV fitness, and that PepMV CPWT is oxidized in vivo while CPC127S is not. We also show that Cys127 gets spontaneously glutathionylated in vitro, and that S-glutathionylation blocks in vitro the formation of virion-like particles (VLPs). VLPs longer than 200 nm could be formed after in planta CPC127S overexpression, while very short and dispersed VLPs were observed after CPWT overexpression. Our results strongly suggest that the CP redox status regulates CP functions via cysteine oxidation.

The tomato calcium-permeable channel 4.1 (SlOSCA4.1) is a susceptibility factor for pepino mosaic virus.

The hyperosmolality-gated calcium permeable channel 4.1 (OSCA4.1) belongs to an evolutionarily conserved small family of mechano-sensitive channels. OSCA members may represent key players in plant resistance to drought and to pathogen infection but are scarcely studied. After screening for resistance to pepino mosaic virus (PepMV) a collection of 1000 mutagenized tomato families, we identified a mutant showing no symptoms and reduced virus accumulation. Resistance was mapped to chromosome 2 between positions 46 309 531 to 47 044 163, where a missense mutation caused the putative truncation of the OSCA4.1 protein. A CRISPR/Cas9 slosca4.1 mutant was resistant to PepMV, but not to tobacco mosaic virus or potato virus X. Inoculation of mutant and wild type tomato protoplasts showed that resistance was expressed in single cells, suggesting a role for SlOSCA4.1 in early viral function(s); congruently, SlOSCA4.1 re-localized to structures reminiscent of viral replication complexes. We propose that SlOSCA4.1 contributes to the correct regulation of the Ca2+ homeostasis necessary for optimal PepMV infection. PepMV is a pandemic virus that causes significant losses in tomato crops worldwide. In spite of its importance, no tomato-resistant varieties have been deployed yet; the mutant identified here has great potential to breed tomato varieties resistant to PepMV.

Tomato Brown Rugose Fruit Virus Pandemic.

Tomato brown rugose fruit virus (ToBRFV) is an emerging tobamovirus. It was first reported in 2015 in Jordan in greenhouse tomatoes and now threatens tomato and pepper crops around the world. ToBRFV is a stable and highly infectious virus that is easily transmitted by mechanical means and via seeds, which enables it to spread locally and over long distances. The ability of ToBRFV to infect tomato plants harboring the commonly deployed Tm resistance genes, as well as pepper plants harboring the L resistance alleles under certain conditions, limits the ability to prevent damage from the virus. The fruit production and quality of ToBRFV-infected tomato and pepper plants can be drastically affected, thus significantly impacting their market value. Herein, we review the current information and discuss the latest areas of research on this virus, which include its discovery and distribution, epidemiology, detection, and prevention and control measures, that could help mitigate the ToBRFV disease pandemic.

Managing the deluge of newly discovered plant viruses and viroids: an optimized scientific and regulatory framework for their characterization and risk analysis.

The advances in high-throughput sequencing (HTS) technologies and bioinformatic tools have provided new opportunities for virus and viroid discovery and diagnostics. Hence, new sequences of viral origin are being discovered and published at a previously unseen rate. Therefore, a collective effort was undertaken to write and propose a framework for prioritizing the biological characterization steps needed after discovering a new plant virus to evaluate its impact at different levels. Even though the proposed approach was widely used, a revision of these guidelines was prepared to consider virus discovery and characterization trends and integrate novel approaches and tools recently published or under development. This updated framework is more adapted to the current rate of virus discovery and provides an improved prioritization for filling knowledge and data gaps. It consists of four distinct steps adapted to include a multi-stakeholder feedback loop. Key improvements include better prioritization and organization of the various steps, earlier data sharing among researchers and involved stakeholders, public database screening, and exploitation of genomic information to predict biological properties.

4D nanoimaging of early age cement hydration.

Despite a century of research, our understanding of cement dissolution and precipitation processes at early ages is very limited. This is due to the lack of methods that can image these processes with enough spatial resolution, contrast and field of view. Here, we adapt near-field ptychographic nanotomography to in situ visualise the hydration of commercial Portland cement in a record-thick capillary. At 19 h, porous C-S-H gel shell, thickness of 500 nm, covers every alite grain enclosing a water gap. The spatial dissolution rate of small alite grains in the acceleration period, ∼100 nm/h, is approximately four times faster than that of large alite grains in the deceleration stage, ∼25 nm/h. Etch-pit development has also been mapped out. This work is complemented by laboratory and synchrotron microtomographies, allowing to measure the particle size distributions with time. 4D nanoimaging will allow mechanistically study dissolution-precipitation processes including the roles of accelerators and superplasticizers.

Transcriptomic and metabolic profiling of watermelon uncovers the role of salicylic acid and flavonoids in the resistance to cucumber green mottle mosaic virus.

Understanding the mechanisms underlying plant resistance to virus infections is crucial for viral disease management in agriculture. However, the defense mechanism of watermelon (Citrullus lanatus) against cucumber green mottle mosaic virus (CGMMV) infection remains largely unknown. In this study, we performed transcriptomic, metabolomic, and phytohormone analyses of a CGMMV susceptible watermelon cultivar 'Zhengkang No.2' ('ZK') and a CGMMV resistant wild watermelon accession PI 220778 (PI) to identify the key regulatory genes, metabolites, and phytohormones responsible for CGMMV resistance. We then tested several phytohormones and metabolites for their roles in watermelon CGMMV resistance via foliar application, followed by CGMMV inoculation. Several phenylpropanoid metabolism-associated genes and metabolites, especially those involved in the flavonoid biosynthesis pathway, were found to be significantly enriched in the CGMMV-infected PI plants compared with the CGMMV-infected 'ZK' plants. We also identified a gene encoding UDP-glycosyltransferase (UGT) that is involved in kaempferol-3-O-sophoroside biosynthesis and controls disease resistance, as well as plant height. Additionally, salicylic acid (SA) biogenesis increased in the CGMMV-infected 'ZK' plants, resulting in the activation of a downstream signaling cascade. SA levels in the tested watermelon plants correlated with that of total flavonoids, and SA pre-treatment up-regulated the expression of flavonoid biosynthesis genes, thus increasing the total flavonoid content. Furthermore, application of exogenous SA or flavonoids extracted from watermelon leaves suppressed CGMMV infection. In summary, our study demonstrates the role of SA-induced flavonoid biosynthesis in plant development and CGMMV resistance, which could be used to breed for CGMMV resistance in watermelon.

An Outstandingly Rare Occurrence of Mycoviruses in Soil Strains of the Plant-Beneficial Fungi from the Genus Trichoderma and a Novel Polymycoviridae Isolate.

In fungi, viral infections frequently remain cryptic causing little or no phenotypic changes. It can indicate either a long history of coevolution or a strong immune system of the host. Some fungi are outstandingly ubiquitous and can be recovered from a great diversity of habitats. However, the role of viral infection in the emergence of environmental opportunistic species is not known. The genus of filamentous and mycoparasitic fungi Trichoderma (Hypocreales, Ascomycota) consists of more than 400 species, which mainly occur on dead wood, other fungi, or as endo- and epiphytes. However, some species are environmental opportunists because they are cosmopolitan, can establish in a diversity of habitats, and can also become pests on mushroom farms and infect immunocompromised humans. In this study, we investigated the library of 163 Trichoderma strains isolated from grassland soils in Inner Mongolia, China, and found only four strains with signs of the mycoviral nucleic acids, including a strain of T. barbatum infected with a novel strain of the Polymycoviridae and named and characterized here as Trichoderma barbatum polymycovirus 1 (TbPMV1). Phylogenetic analysis suggested that TbPMV1 was evolutionarily distinct from the Polymycoviridae isolated either from Eurotialean fungi or from the order Magnaportales. Although the Polymycoviridae viruses were also known from Hypocrealean Beauveria bassiana, the phylogeny of TbPMV1 did not reflect the phylogeny of the host. Our analysis lays the groundwork for further in-depth characterization of TbPMV1 and the role of mycoviruses in the emergence of environmental opportunism in Trichoderma. IMPORTANCE Although viruses infect all organisms, our knowledge of some groups of eukaryotes remains limited. For instance, the diversity of viruses infecting fungi-mycoviruses-is largely unknown. However, the knowledge of viruses associated with industrially relevant and plant-beneficial fungi, such as Trichoderma spp. (Hypocreales, Ascomycota), may shed light on the stability of their phenotypes and the expression of beneficial traits. In this study, we screened the library of soilborne Trichoderma strains because these isolates may be developed into bioeffectors for plant protection and sustainable agriculture. Notably, the diversity of endophytic viruses in soil Trichoderma was outstandingly low. Only 2% of 163 strains contained traces of dsRNA viruses, including the new Trichoderma barbatum polymycovirus 1 (TbPMV1) characterized in this study. TbPMV1 is the first mycovirus found in Trichoderma. Our results indicate that the limited data prevent the in-depth study of the evolutionary relationship between soilborne fungi and is worth further investigation.

Fluorescently labelled pepino mosaic virus strains share infected tissues and colocalize in cellular aggregates reminiscent of viral replication complexes.

Mixed infections of pepino mosaic virus (PepMV) isolates from the EU and CH2 strains are frequent in tomato crops. An asymmetric antagonistic relationship has been described between these strains, making their in planta interaction worthy of study. The aim of this work was to verify if PepMV isolates labelled with fluorescent proteins recapitulate the interactions described for wild type isolates and, if so, to determine the proportion of cells infected with each isolate in single and mixed infected plants. Infectious clones were prepared for PepMV-CH2-GFP and -EU-TagRFP, and also for their reciprocal combination, PepMV-CH2-TagRFP and -EU-GFP, and used to inoculate Nicotiana benthamiana plants. The accumulation of viral RNA followed trends that differed from wild type viruses, with the PepMV-EU-GFP and -CH2-TagRFP pair reproducing more closely the wild type interaction. Protoplasts were isolated from leaves that were systemically infected with PepMV-EU-GFP, -CH2-TagRFP, or both, and flow cytometry was used to determine the proportion of cells infected with each tagged isolate. A significant proportion (16.6%) of cells were found to be infected with both, without strong evidence of virus exclusion in coinfections, as could have been expected for related viruses; in fact, cellular structures reminiscent of viral replication complexes were found to be labelled with both fluorescent reporters.

Recent Advances in C-S-H Nucleation Seeding for Improving Cement Performances.

Reducing cement CO2 footprint is a societal need. This is being achieved mainly by replacing an increasing amount of Portland clinker by supplementary cementitious materials. However, this comes at a price: lower mechanical strengths at early ages due to slow pozzolanic reaction(s). This is being addressed by using accelerator admixtures. In this context, calcium silicate hydrate nucleation seeding seems to have a promising future, as it can accelerate cement and pozzolanic reactions at early ages, optimising their microstructures, without compromising late strength and durability performances. In fact, these features could even be improved. Moreover, other uses are low temperature concreting, precasting, shotconcrete, etc. Here, we focus on reviewing recent reports on calcium silicate hydrate seeding using commercially available admixtures. Current knowledge on the consequences of nucleation seeding on hydration reactions and on early and late mechanical strengths is discussed. It is noted that other features, in addition to the classic alite hydration acceleration, are covered here including the enhanced ettringite precipitation and the very efficient porosity refinement, which take place in the seeded binders. Finally, because the seeded binders seem to be denser, durability properties could also be enhanced although this remains to be properly established.

Tomato SlGSTU38 interacts with the PepMV coat protein and promotes viral infection.

Pepino mosaic virus (PepMV) is pandemic in tomato crops, causing important economic losses world-wide. No PepMV-resistant varieties have been developed yet. Identification of host factors interacting with PepMV proteins is a promising source of genetic targets to develop PepMV-resistant varieties. The interaction between the PepMV coat protein (CP) and the tomato glutathione S-transferase (GST) SlGSTU38 was identified in a yeast two-hybrid (Y2H) screening and validated by directed Y2H and co-immunoprecipitation assays. SlGSTU38-knocked-out Micro-Tom plants (gstu38) generated by the CRISPR/Cas9 technology together with live-cell imaging were used to understand the role of SlGSTU38 during infection. The transcriptomes of healthy and PepMV-infected wild-type (WT) and gstu38 plants were profiled by RNA-seq analysis. SlGSTU38 functions as a PepMV-specific susceptibility factor in a cell-autonomous manner and relocalizes to the virus replication complexes during infection. Besides, knocking out SlGSTU38 triggers reactive oxygen species accumulation in leaves and the deregulation of stress-responsive genes. SlGSTU38 may play a dual role: On the one hand, SlGSTU38 may exert a proviral function depending on its specific interaction with the PepMV CP; and on the other hand, SlGSTU38 may delay PepMV-infection sensing by participating in the redox intracellular homeostasis in a nonspecific manner.

Cohombrillo-associated virus: a novel virus infecting Ecballium elaterium plants.

We determined the complete genome sequence of a new virus infecting Ecballium elaterium ('cohombrillo amargo') plants, a weed species common on the borders of cultivated fields in the Mediterranean region. The genome of this virus is composed of two molecules of monocistronic positive-sense RNA, 6,934 and 3,501 nucleotides in length, excluding their poly(A) tails. The highest amino acid sequence similarity (50 % identity) in the Pro-Pol core region encoded by RNA 1 was observed in the corresponding protein of strawberry latent ringspot virus. Based on pairwise comparisons and phylogenetic analysis, this virus, tentatively named "cohombrillo-associated virus" (CoAV), appears to be a member of a new species in the genus Stralarivirus (family Secoviridae), for which the name "Stralarivirus elaterii" is proposed. This new virus has different putative cleavage patterns from members of other species belonging to this genus.

The Expanding Menagerie of Prunus-Infecting Luteoviruses.

Members of the genus Luteovirus are responsible for economically destructive plant diseases worldwide. Over the past few years, three luteoviruses infecting Prunus trees have been characterized. However, the biological properties, prevalence, and genetic diversity of those viruses have not yet been studied. High-throughput sequencing of samples of various wild, cultivated, and ornamental Prunus species enabled the identification of four novel species in the genus Luteovirus for which we obtained complete or nearly complete genomes. Additionally, we identified another new putative species recovered from Sequence Read Archive data. Furthermore, we conducted a survey on peach-infecting luteoviruses in eight European countries. Analyses of 350 leaf samples collected from germplasm, production orchards, and private gardens showed that peach-associated luteovirus (PaLV), nectarine stem pitting-associated virus (NSPaV), and a novel luteovirus, peach-associated luteovirus 2 (PaLV2), are present in all countries; the most prevalent virus was NSPaV, followed by PaLV. The genetic diversity of these viruses was also analyzed. Moreover, the biological indexing on GF305 peach indicator plants demonstrated that PaLV and PaLV2, like NSPaV, are transmitted by graft at relatively low rates. No clear viral symptoms have been observed in either graft-inoculated GF305 indicators or different peach tree varieties observed in an orchard. The data generated during this study provide a broader overview of the genetic diversity, geographical distribution, and prevalence of peach-infecting luteoviruses and suggest that these viruses are likely asymptomatic in peach under most circumstances.

Different RNA Elements Control Viral Protein Synthesis in Polerovirus Isolates Evolved in Separate Geographical Regions.

Most plant viruses lack the 5'-cap and 3'-poly(A) structures, which are common in their host mRNAs, and are crucial for translation initiation. Thus, alternative translation initiation mechanisms were identified for viral mRNAs, one of these being controlled by an RNA element in their 3'-ends that is able to enhance mRNA cap-independent translation (3'-CITE). The 3'-CITEs are modular and transferable RNA elements. In the case of poleroviruses, the mechanism of translation initiation of their RNAs in the host cell is still unclear; thus, it was studied for one of its members, cucurbit aphid-borne yellows virus (CABYV). We determined that efficient CABYV RNA translation requires the presence of a 3'-CITE in its 3'-UTR. We showed that this 3'-CITE requires the presence of the 5'-UTR in cis for its eIF4E-independent activity. Efficient virus multiplication depended on 3'-CITE activity. In CABYV isolates belonging to the three phylogenetic groups identified so far, the 3'-CITEs differ, and recombination prediction analyses suggest that these 3'-CITEs have been acquired through recombination with an unknown donor. Since these isolates have evolved in different geographical regions, this may suggest that their respective 3'-CITEs are possibly better adapted to each region. We propose that translation of other polerovirus genomes may also be 3'-CITE-dependent.

Genetic Differentiation and Migration Fluxes of Viruses from Melon Crops and Crop Edge Weeds.

Weeds surrounding crops may act as alternative hosts, playing important epidemiological roles as virus reservoirs and impacting virus evolution. We used high-throughput sequencing to identify viruses in Spanish melon crops and plants belonging to three pluriannual weed species, Ecballium elaterium, Malva sylvestris, and Solanum nigrum, sampled at the edges of the crops. Melon and E. elaterium, both belonging to the family Cucurbitaceae, shared three virus species, whereas there was no virus species overlap between melon and the other two weeds. The diversity of cucurbit aphid-borne yellows virus (CABYV) and tomato leaf curl New Delhi virus (ToLCNDV), both in melon and E. elaterium, was further studied by amplicon sequencing. Phylogenetic and population genetics analyses showed that the CABYV population was structured by the host, identifying three sites in the CABYV RNA-dependent RNA polymerase under positive selection, perhaps reflecting host adaptation. The ToLCNDV population was much less diverse than the CABYV one, likely as a consequence of the relatively recent introduction of ToLCNDV in Spain. In spite of its low diversity, we identified geographical but no host differentiation for ToLCNDV. Potential virus migration fluxes between E. elaterium and melon plants were also analyzed. For CABYV, no evidence of migration between the populations of the two hosts was found, whereas important fluxes were identified between geographically distant subpopulations for each host. For ToLCNDV, in contrast, evidence of migration from melon to E. elaterium was found, but not the other way around. IMPORTANCE It has been reported that about half of the emerging diseases affecting plants are caused by viruses. Alternative hosts often play critical roles in virus emergence as virus reservoirs, bridging host species that are otherwise unconnected and/or favoring virus diversification. In spite of this, the viromes of potential alternative hosts remain largely unexplored. In the case of crops, pluriannual weeds at the crop edges may play these roles. Here, we took advantage of the power of high-throughput sequencing to characterize the viromes of three weed species frequently found at the edges of melon crops. We identified three viruses shared by melon and the cucurbit weed, with two of them being epidemiologically relevant for melon crops. Further genetic analyses showed that these two viruses had contrasting patterns of diversification and migration, providing an interesting example on the role that weeds may play in the ecology and evolution of viruses affecting crops.

Editing melon eIF4E associates with virus resistance and male sterility.

The cap-binding protein eIF4E, through its interaction with eIF4G, constitutes the core of the eIF4F complex, which plays a key role in the circularization of mRNAs and their subsequent cap-dependent translation. In addition to its fundamental role in mRNA translation initiation, other functions have been described or suggested for eIF4E, including acting as a proviral factor and participating in sexual development. We used CRISPR/Cas9 genome editing to generate melon eif4e knockout mutant lines. Editing worked efficiently in melon, as we obtained transformed plants with a single-nucleotide deletion in homozygosis in the first eIF4E exon already in a T0 generation. Edited and non-transgenic plants of a segregating F2 generation were inoculated with Moroccan watermelon mosaic virus (MWMV); homozygous mutant plants showed virus resistance, while heterozygous and non-mutant plants were infected, in agreement with our previous results with plants silenced in eIF4E. Interestingly, all homozygous edited plants of the T0 and F2 generations showed a male sterility phenotype, while crossing with wild-type plants restored fertility, displaying a perfect correlation between the segregation of the male sterility phenotype and the segregation of the eif4e mutation. Morphological comparative analysis of melon male flowers along consecutive developmental stages showed postmeiotic abnormal development for both microsporocytes and tapetum, with clear differences in the timing of tapetum degradation in the mutant versus wild-type. An RNA-Seq analysis identified critical genes in pollen development that were down-regulated in flowers of eif4e/eif4e plants, and suggested that eIF4E-specific mRNA translation initiation is a limiting factor for male gametes formation in melon.

Transcriptome analyses unveiled differential regulation of AGO and DCL genes by pepino mosaic virus strains.

Pepino mosaic virus (PepMV) is a single-stranded (ss), positive-sense (+) RNA potexvirus that affects tomato crops worldwide. We have described an in planta antagonistic interaction between PepMV isolates of two strains in which the EU isolate represses the accumulation of the CH2 isolate during mixed infections. Reports describing transcriptomic responses to mixed infections are scant. We carried out transcriptomic analyses of tomato plants singly and mixed-infected with two PepMV isolates of both strains. Comparison of the transcriptomes of singly infected plants showed that deeper transcriptomic alterations occurred at early infection times, and also that each of the viral strains modulated the host transcriptome differentially. Mixed infections caused transcriptomic alterations similar to those for the sum of single infections at early infection times, but clearly differing at later times postinfection. We next tested the hypothesis that PepMV-EU, in either single or mixed infections, deregulates host gene expression differentially so that virus accumulation of both strains gets repressed. That seemed to be the case for the genes AGO1a, DCL2d, AGO2a, and DCL2b, which are involved in the antiviral silencing pathway and were upregulated by PepMV-EU but not by PepMV-CH2 at early times postinfection. The pattern of AGO2a expression was validated by reverse transcription-quantitative PCR in tomato and Nicotiana benthamiana plants. Using an N. benthamiana ago2 mutant line, we showed that AGO2 indeed plays an important role in the antiviral defence against PepMV, but it is not the primary determinant of the outcome of the antagonistic interaction between the two PepMV strains.

Portland and Belite Cement Hydration Acceleration by C-S-H Seeds with Variable w/c Ratios.

The acceleration of very early age cement hydration by C-S-H seeding is getting attention from scholars and field applications because the enhanced early age features do not compromise later age performances. This acceleration could be beneficial for several low-CO2 cements as a general drawback is usually the low very early age mechanical strengths. However, the mechanistic understanding of this acceleration in commercial cements is not complete. Reported here is a contribution to this understanding from the study of the effects of C-S-H gel seeding in one Portland cement and two belite cements at two widely studied water-cement ratios, 0.50 and 0.40. Two commercially available C-S-H nano-seed-based admixtures, i.e., Master X-Seed 130 and Master X-Seed STE-53, were investigated. A multi-technique approach was adopted by employing calorimetry, thermal analysis, powder diffraction (data analysed by the Rietveld method), mercury intrusion porosimetry, and mechanical strength determination. For instance, the compressive strength at 1 day for the PC (w/c = 0.50) sample increased from 15 MPa for the unseeded mortar to 24 and 22 MPs for the mortars seeded with the XS130 and STE53, respectively. The evolution of the amorphous contents was determined by adding an internal standard before recording the powder patterns. In summary, alite and belite phase hydrations, from the crystalline phase content evolutions, are not significantly accelerated by C-S-H seedings at the studied ages of 1 and 28 d for these cements. Conversely, the hydration rates of tetracalcium alumino-ferrate and tricalcium aluminate were significantly enhanced. It is noted that the degrees of reaction of C4AF for the PC paste (w/c = 0.40) were 10, 30, and 40% at 1, 7, and 28 days. After C-S-H seeding, the values increased to 20, 45, and 60%, respectively. This resulted in larger ettringite contents at very early ages but not at 28 days. At 28 days of hydration, larger amounts of carbonate-containing AFm-type phases were determined. Finally, and importantly, the admixtures yielded larger amounts of amorphous components in the pastes at later hydration ages. This is justified, in part, by the higher content of amorphous iron siliceous hydrogarnet from the enhanced C4AF reactivity.