21 de marzo de 2018

final notes on EUCARPIA Cereals meeting (III)


Wednesday 21st march 2018 (Spring)

Tatsuya IKEDA (NARO, Japan)
Starts by showing allelic variation in glutenin subunits, which form large complexes connected by SS-bonds. Different alleles (A1,B1,D1 & A3,B3,D3) produce different gluten networks, which are reflected on bread body and loaf volume. Combinations of glutenin alleles control dough quality. He refers to results published at http://dx.doi.org/10.1094/CCHEM-01-14-0009-R. He explains the http://www.wheatinitiative.org, which aims at improving wheat quality and safety and to integrate a research platform with shared materials and gene nomenclature (he shows examples of gene name searches in Ensembl Plants with no results due to lack of naming consensus). They already released glutenin master sets at CIMMYT Germplasm Bank. Starch/amylose content is also important for quality. Then he talks about safety and describes Fusarium toxin analysis.

Sebastian MICHEL (U Vienna, Austria)
His talk is about genomic selection for wheat breeding, aiming at simultaneous selection for grain yield and baking quality. He shows data showing a strong anti-correlation between yield and protein content (typical range 12-16%), which is clearly important for quality and trading. Therefore, they focus on outliers with high protein content. The develop yield and protein indexes, which allow selection of lines with excellent yield and good protein content, and lines with high-protein and acceptable yield, respectively.

Suong CU (Flinders U, Australia)
She talks about GWAS of Zn, Fe, Cu, Mn and P in wheat grain and rachis at two developmental stages with a bio-fortification perspective. She mentions partners CIMMYT and http://www.harvestplus.org. The calculate heritability of the uptake of those micronutrients and their correlation in mature grain and in the rachis, end they were low despite the experiments were done with fertilizer contributing enough of these. They perform GWAS correcting by Kinship and covariates and shows associated markers with most additive value. She shows some candidate genes and for Zn, for instance, there is one transporter and another annotated as a Zinc-finger.

Simon GRIFFITHS (JIC, UK)
His talk is about analysis of arabinoxylan (AX) soluble fibre in wheat grain. He mentions that in the UK there have been spectacular yield gains since 1885, but that nutritional quality has never been a breeding target until now. He focuses on fibre intake and fibre content in bread in the UK and explains that AX is the main fibre component of wheat grain endosperm, and is also part of the bran. He mentions that project https://healthgrain.org was concerned about genetic resources for improving wheat nutritional value. Shows that AX content is highly heritable. They have prepared populations with Yumai34 parent to discover QTLs for AX and found one. He explains that AX enrichment does not affect breadmaking functionality.
Peter ROGOWSKY (INRA Lyon, France)
Delivers a general lecture about CRISPR-Cas9 systems in breeding plants. Starts by explaining cellular DNA repair mechanism. The current success of Cas9 is that predicting RNA/DNA is much easier and reliable than protein-DNA binding. Stable transformation of Cas9 can be done with Agrobacterium, direct transfer (explants, protoplasts) or virus. Transient expression can be obtained by direct transfer of ribonucleoproteins. All these have been used successfully in wheat, with the bottleneck being generally plant regeneration after transformation. PAM-altered Cas9 have not yet been ported to plants, so targets must have a neighbor NGG site. Another option is to use CPF1, which recognizes TTTN sites.
Mutation and base edition have been mastered in plants, with frequencies less than 10%; gene editing still has frequencies below 1% in plants. However, currently genes can be edited by stacking base editions to modify coding sequences or promoters, when expression is important for the trait of interest. 

He also presents recent examples of gene inactivation in different crops, both monocot and dicot, still to be tested in the field, with the SDN1 (site directed nuclease) approach. He explains that Cas9 approaches are expect to have even more applications in terms of agronomic and quality traits. However, regulation is still not clearly defined for Cas9 editions at the EU (decision expected in 2018 by the European Court of Justice). He explains that this technique faces different ethical questions and their answers should be delivered case by case. He finishes with a table of promises/claims of Cas9 and the reality in 2018; this tech is still a long way from having an impact in the field and the EU market.

Fred VAN EEUWIJK (Wageningen U, Netherlands)
Presents methods for modelling genotype by environment (GxE) interactions in QTL mapping and genomic prediction, with applications to barley and wheat. How can we reduce the computing time of computing a linear mixed model per SNP when you have 10E5-E7 SNPs? With packages such as TASSEL P3D, GRAMMAR, FaST-LMM. He seems to prefer linear mixed models to Bayesian choices, as they are easier to customize. For multi-trait / environment modelling, he refers to https://dl.sciencesocieties.org/publications/cs/abstracts/56/5/2119 and this book: http://www.springer.com/gp/book/9783319205618. Fitting these multi models requires reducing the original complexity of the data. He mentions Haploview to merge SNPs in the same gene to build haplotypes/alleles and test them instead of individual SNPs. Finally, he mentiones that a software tool with their latest developments as part of the WHEALBI consortium will be available at the end of the year, and there will be training courses, but the modules will be available in CRAN as R packages (https://cran.r-project.org).

Andreas HUND (ETH Zürich, Switzerland)
Talks about GxE and modern field phenotyping techniques. He mentions that in the Swiss central plateau wheat yield (target 6-7T/ha) is limited by excess water, frost and then heat stress. They have set up all year round platform for phenotyping wheat. He shows canopy cover, height and senescence/reflectance data for winter wheat. They calculate heritabilities for their traits and obtain high values (H2>0.9) except for the duration of developmental stages. He shows data from a couple of papers (http://www.publish.csiro.au/fp/fp14226 and https://link.springer.com/article/10.1007/s10681-017-1940-2) showing, for instance, that earliest genotypes are not necessarily the tallest in terms of final height.

Friedrich LONGIN (U Hohenheim, Germany)
Just after lunch, he talks about genomic selection (GS) schemes for line breeding in wheat, are they really worth it? His simulation data suggest that direct phenotypic selection can yield higher genetic gain, but GS yield higher gains per year while having less plant material in the field. These trends depend on the GS accuracy, but with realistic values of 0.6 there seems to be a 16% annual gain advantage for GS if breeding cycle is sped up.

Daniela BUSTOS-KORTS (Wageningen U, Netherlands)
She talks about combining crop growth and statistical genetic modelling to evaluate phenotyping strategies, and shows wheat data. Genotyping is much cheaper than phenotyping, and she evaluates alternative phenotyping strategies with simulations, inspired by http://www.publish.csiro.au/cp/cp14007. She shows drought patterns observed in the Australia wheat belt, published in https://nph.onlinelibrary.wiley.com/doi/abs/10.1111/nph.12192.

Renaud RINCENT (INRA GDEC, France)
His talk is about genomic prediction of GxE interactions by coupling genetic and physiological modelling. He summarizes the challenges and opportunities of GxE in breeding and uses data grain yield from https://breedwheat.fr. His results indicate that considering GxE when doing phenotyping predictions provides more accurate models than simple models with no interaction. His AMMI model, which computed PCA on the residuals of the principal model, seemed promising but the gains were lost in the cross-validations, likely due to small dataset or to weak GxE in the data.

Philipp BOEVEN (Limagrain, France)
His talk discusses ways of breeding for favorable male floral traits, such as anther extrusion, in hybrid wheat. After GWAS he made BLUP genomic predictions (I had to read what Best Linear Unbiased Prediction is, a widely used genomic prediction method, at: https://www.ncbi.nlm.nih.gov/pubmed/23640517).

Funmi LADEJOBI (NIAB, UK)
The last talk of the meeting is about differentially penalized regressions, a modification of BLUP, which improve genomic prediction of wheat flowering time. She uses rrBLUP R package, documented at https://cran.r-project.org/web/packages/rrBLUP/index.html.

20 de marzo de 2018

notes on EUCARPIA Cereals meeting 2018 (II)


Tuesday 20th March 2018

John FOULKES (U Nottingham, UK & CIMMYT collaborators)
Talks about genetic diversity and resource efficiency in wheat. Genetic gains in yield potential over the last decades has increased 0.5-1% per yr, now it is slowing down. Biomass acounts for a large part of that potential, while harvest index (HI, http://plantsinaction.science.uq.edu.au/content/641-harvest-index) is following an inverse trend. Therefore, we are not taking the full benefit of the genetic gains. Data from field assays indicates that the 2nd internode that is competing with spike growth. He reckons that HI values of 0.6 are likely.
Then he moves on to N fertilization and the WISP project (http://www.wheatisp.org/Consortium/WISP.php), and shows some results about diversity on N uptake and also about photosynthetic efficiency published last year by Gaju et al,  https://www.sciencedirect.com/science/article/pii/S0378429016301022.
Finally, he summarizes experiments on root phenomics (shovelomics, extracting top 20 cm of crown root, see http://plantscience.psu.edu/research/labs/roots/methods/field/shovelomics) with the aim of redesigning root architecture. Using a Rialto x Savanah population, they discovered a great deal of variation in the root traits they were tracking. They also found correlation between root angle and root length under rain-fed conditions. They plan to set up a TILLING experiment to validate candidate genes. He shows nice images of root cuts of irrigated and non-irrigated plants that they’re using to train machine learning algorithms for image analysis.

 
Bruno CONTRERAS-MOREIRA (EEAD-CSIC, Spain)
That’s me. I talked about a series of experiments with pooled barleys designed with the aim of testing the agronomic advantage of the presence of flowering control gene PpdH2 in winter barleys proven through a natural selection approach. I took the chance to present the PpdH2 gene as ana accessory gene in the pan-genome context, using the terminology we have used in our papers (barley and A.thaliana & Brachypodium distachyon). I got questions by Frank Ordon and Simon Griffith: i) the number of seeds used to build the pools, and ii) could plant competition explain the results.

Gaëtan TOUZY (Arvalis, France)
He talks about his project on “Improving Water Use Efficiency in Bread Wheat by Multi-trait multi-Environment Genome-Wide Association Studies”.

Eric OBER (NIAB, UK)
The title of his talk is “Implementing large-scale field phenotyping in genomic selection to accelerate wheat breeding” which reports results of project GplusE (http://gtr.rcuk.ac.uk/projects?ref=BB%2FL022141%2F1). They do visual scoring plus drone and manned flight hyperspectral camera shots. They build Bayesian networks with a few traits and yield. They use data from a few years to predict phenotype (yield) with some success, but with different accuracies among sites and years. Average prediction is safer than predicting the best/worst performer genotypes. Best traits are hyperspectral data, development trains and late-measurements data.

Kerstin NEUMANN (IPK, Germany)
She talks about barley phenotyping at IPK, particularly about using high-throughput image analysis to study stress-adaptive and constitutive biomass QTLs in cereals. She shows data and results of spring barleys, published at https://www.ncbi.nlm.nih.gov/labs/articles/28797222, and winter wheats.

Ulrike LOHWASSER (IPK, Germany)
Her talk is about “Searching for Frost Tolerance in Wheat – A genome wide association study”. Frost tolerance is a complex trait, which involves winter survival, desiccation, anoxia, ice-encasement and even disease resistance. Heritability for frost tolerance is low in most locations when field trails are carried out in cold winters, but it is high in controlled conditions.

Heribert HIRT (KAUST, Saudia Arabia)
He talks about beneficial microbial endophytes to enhance abiotic stress tolerance and yield. He works mostly with Arabidopsis and is interested on plants living in deserts as part of project DARWIN 21 (http://www.darwin21.net). The do trials of Arabidopsis, but also wheat, barley, alfalfa and confirm beneficial effects under stress but not in normal conditions. In A. thaliana they actually observe that inoculation changes the stress response of the plant. They also did experiments to mimic microbial inoculation by adding external chemicals in A. thaliana. He shows data for one of their endophytes, Enterobacter spp. SA187, which was found in both monocot and dicot plants. They have no evidence of crop-specific strains, because they looked for generalists.

Ewen MULLINS (Teagasc, Ireland)
He talks about the impact (-30% in the last couple of years) of Septoria tritici blotch (STB) disease in wheat. They are carrying out intensive field phenotyping to support breeding of resistant lines. They did fungicide-free trials in Ireland and the UK, visually scoring plants, and concluded that different wheat genotypes have different latency periods. However, eventually disease progresses in all of them (https://onlinelibrary.wiley.com/doi/abs/10.1111/ppa.12780), so it would seem that a reasonable breeding target might be further extending the latency period.


Yvan MOËNNE-LOCCOZ (U Lyon, France)
Talks about interactions of plant-beneficial rhizosphere bacteria in cereals. Can breeding benefit from microbiome-based approaches? Does the plant genotype matter? Have modern cultivars lost their microbial partners? They have performed 16S rRNA scans of rhizosphere below different crops and wild plant species such as teosinte. They have data that suggest that some modern lines conserve the ability to interact with inoculated bacteria; others do not, perhaps for being counter-selected. They have used Pseudomonas kilonensis F113 to test root colonization in a panel of wheat cultivars and see that modern cultivars are relatively less colonized than old or landraces. He also mentions Azospirillum brasilense Sp245, which stimulates root growth by producing hormone IAA. He has a number of publications on these topics listed at https://scholar.google.fr/citations?hl=fr&user=rF48UsAAAAAJ.


Laetitia WILLOCQUET (INRA, France)
She talks about phenotyping methods for quantitative host plant resistance using simulation modelling and ROC curves. She reports results published at https://www.sciencedirect.com/science/article/pii/S1360138517300237. Phenotyping is now the bottleneck for breeding resistance. Nonetheless, data was produced to feed models of infection and resistance, estimate parameters and make predictions. Details on these simulation models can be found at this document published in 2014: https://goo.gl/VQpqK7


Hermann BUERSTMAYR (BOKU, Austria)
Genomics assisted improvement of Fusarium head blight resistance in bread wheat, durum wheat and triticale


Javier SANCHEZ-MARTIN (U Zürich, Switzerland)
His talk is about performing GWAS to reveal race-specific resistance genes to powdery mildew in wheats from the WHEALBI project (http://www.whealbi.eu). As in related talks, he discusses how exome capture platforms present mapping problems when aligning genes absent from the reference genome. He has published part of these results in 2016:  https://genomebiology.biomedcentral.com/articles/10.1186/s13059-016-1082-1



Pierre-Antoine PRECIGOUT (U Paris-Saclay, France)
Pierre presents numerical, epidemiological models of foliar fungal pathogens in wheat, which have been described in detailed at https://www.ncbi.nlm.nih.gov/pubmed/28453406 . These model the latent period to predict potential evolutionary directions.

19 de marzo de 2018

notes on EUCARPIA Cereals meeting 2018 (I)


Monday, 19th March 2018 (program at https://symposium.inra.fr/eucarpia-cereal2018)

Intro: Gilles Charmet (INRA-UCA), remembers Patrick Schweizer
Intro Eucarpia: Andreas Borner (IPK), EUCARPIA Cereals section conference

Raphäel Dumoin (Bayer Crop Science) Wheat Innovation Strategy at Bayer
There is a need to breed wheat for both high and low productivity areas around the world. In each area, there is a gap between current productivity and potential yield. They expect that the wheat seed market will be soon as large as corn’s [due to correlation between acreage and seed value for corn, soybean, cotton, canola]. BCS now has breeding stations in North America, EU and Australia and they are developing pure lines and hybrids, as well as looking for yield improving traits. The elements that explain higher yield in wheat are yield stability and abioitic stress tolerance, while maintaining quality. They use heterotic pools for breeding hybrids. They work with both spring and winter wheats and take 7yr to develop a new variety with marker-assisted breeding. They also work with targeted genome optimization/Cas9 edition, which can be done in 1-2yr but faces regulatory hurdles in EU. They actively engage in collaborations with public R&D organizations and private companies around the world.

Andreas Graner (IPK) Ex situ germoplasm collections
There is a increased demand of crops and a need for sustainability (Steffen Science 2015). The quest for innovation in plant breeding needs the interface between genomics, metabolomics and phenomics. The breeding methodology is now genomic selection, that increases explained variability by adding minor QTLs. Both doubled haploids and transformation/Cas9 are key enabling technologies. He emphasizes the importance of surveying and exploiting the available genetic resources. He mentions that currently the German federal ex-situ genebank contains 27K wheat and 23K Hordeum accessions, with seed multiplication done on average every 20-30 yr (https://www.nature.com/articles/srep05231). These experiments have allowed estimating heritabilities of 0.89-0.95 and are now the ground for GWAS analyses with very large populations after careful curation of data. At IPK they are taking advantage of a large phenomics facility put together recently to quantitatively characterize traits such as lipid content at large scale (see paper 2014 on Avena lipids).  They have sequenced with GBS 23K barleys, observing that genetic diversity mimics geographic origin. He mentions data management FAIR principles and APIs. They have recently released the BRIDGE barley IPK DB (https://t.co/fLPAkkF7nY). He argues that the Nagoya Protocol on Access and Benefit Sharing (https://www.cbd.int/abs) is against Open Access, as it will restrict, for instance, dissemination of phenotypic data from collections.

Davide Guerra (CREA, Italy)
Presents the WHEALBI collection with 512 barley accessions from 73 countries, including both cultivars and landraces. These were exon-captured and sequenced to yield 403 validates sampled with 64M called variants, which they used to allocate barleys to 6 geography-based subpopulations. A series of common garden experiments were carried out in several latitudes and irrigations regimes. He shows preliminary results on multi-environment GWAS experiments and discusses a few confirmed candidate genes they have found, including VRNH1, PpdH1 or HvCEN. He then goes into some depth to show his results on Copy Number Variation (CNV) at the CBF locus, the frost tolerance experiments carried out to characterize the alleles discovered and the PCR experiments ahead to survey that particular genomic locus.

Ernesto Igartua (EEAD-CSIC, Spain)
Presents the Spanish Barley Core Collection (SBCC, http://www.eead.csic.es/barley/index.php) and explains that Spanish landraces comprise actually 4 subpopulations. These SBCC barleys have been used in the CLIMBAR FACCEJGI project to analyze their association to agro-climatic variables. He presents first the genetic differentiation of the 4 subpopulations (XtX, diversity). Then a table is shown with linkage disequilibrium. First, it is found that cold tolerance and water balance are the main variables explaining the genetic diversity. Second, GWAS experiments with both Bayenv2 and LFMM confirm the CBF locus (+ control) and unveil a candidate amino-oxydase associated to cold/heat responses.

Marco Maccaferri (U Bologna, Italy); Luigi CATTIVELLI (CREA, Italy)
Genome assembly of durum wheat cv Svevo (http://www.tasaco.com/Seed.aspx?cesit=44) and then a tetraploid diversity panel of 1.9K lines. Estimates average LD < 0.2 with dist(SNPs) between 400Kb and 1.9Mb depending on the population.
Luigi talks more about the genome project (https://www.interomics.eu/durum-wheat-genome), assembled with NRGene software. 90% of the genome in 2K scaffolds. 95% scaffolds are mapped and anchored. The same protocol was used by other team to sequence wild emmer cv Zavitan, parent of wheat tetraploids, which was already sequenced (http://science.sciencemag.org/content/357/6346/93) and suggests that there is a lot of CNV, concentrated at the end of chromosome arms. In addition, they found 600 loss-of-function genes in durum compared to Zavitan, due to gained stop codons or frame shifts due to indels%3 > 0. These must have occurred in less than 10K yr.

Helmy M YOUSSEF (IPK, Germany)
Talks about natural diversity of inflorescence in Hordeum vulgare, reporting results published in https://www.nature.com/articles/ng.3717. He explains what two, six-rowed barleys are and describes labile and intermedium spikes as well. They discover and describe gene Vrs2, which affect spike architecture.

Constance LAVERGNE (U Nottingham, UK)
Talks about introducing/introgressing of Aegilops sharonensis cytoplasm into common wheat and production of addition/translocation lines which are often male-sterile. She shows seed pictures of different generations, as well as GISH preparations of introgressed and translocation lines.


 
Scott Allen JACKSON (U Georgia, USA)
Talks about legume genomes (10 references available currently). While annual soybeans are Chinese, there are a few perennials in Australia. Phaseolus is more ancestral and is used to root trees. Breeding is just a series of bottlenecks, and domestication is likely the most important one. However, improvement requires genetic variation. Discusses that reference genomes, while allowing many types of diversity studies, have limitations, as they are just genomic snapshots. He argues that pan-genomes are better tools and he shows the wild Glycine pan-genome, reported at https://www.nature.com/articles/nbt.2979. He mentions that having it allowed to test for genes under selection in G. max, and they found just under seven hundred.
He then talks about transposable elements (TEs) and their role in genome evolution as sources of novel diversity. TEs live for about 2Myr in a typical plant (half-life). There are no subgenomes dominance effect in soybean, and there is large PAV. He talks also about DNA methylation (CG, CHG, CHH, 3 different plant methylases) and how it changes TEs (he cites https://www.nature.com/articles/nrg.2016.139). He says methylation is the preferred mechanism to silence inserted TEs in plant genomes, and how differentially methylated regions (DMRs) in a pan-genome occur, usually because TE move. Most DMRs are inherited stably and behave like SNPs. He also cites a recent paper showing that post-duplication methylation diminishes are evolutionary time passes (https://onlinelibrary.wiley.com/doi/abs/10.1111/pce.13127). Non-syntenic genes tend to be C-methylated. His last statement is that a third of pan-genome genes are in low recombinogenic regions, including TE non-colinear genes.


Caroline JUERY (INRA GDEC, France)
Explains histone marks of euchromatin and heterochromatin and then explains she wants to check whether the wheat epigenome is partitioned according to H4K27me3, H3K36me3, H4K9ac, H3K4me3 marks (or lack of) ascertained by ChIP-seq. She concludes there are clearly epigenetic territories and then looks to triads of homeologous genes to measure the effect of epigenome marks (upstream, ATG, stop, downstream, as in figure 3 of http://www.plantcell.org/content/21/4/1053) on gene expression, not protein expression yet.


Cécile MONAT (IPK, Germany)
She starts by defining the basics of pan-genomes and presents the http://www.10wheatgenomes.com project, which is starting to produce reference-quality assemblies of 10 wheat cultivars combining NRGene assemblies, linked 10x reads (https://community.10xgenomics.com/t5/10x-Blog/A-basic-introduction-to-linked-reads/ba-p/95), POPSEQ and Hi-C data. Cécile has a preprint describing the pan-genome of two African rice species at https://www.biorxiv.org/content/early/2018/01/09/245431.


Maria BUERSTMAYR (BOKU, Austria)
Talks about high-resolution mapping of the pericentromeric region on wheat chromosome arm 5AS harboring the Fusarium head blight resistance QTL Qfhs.ifa-5A. Used gamma-radiation to promote double-breaks in DNA and overcome recombination limitations in the centromere, even with large populations, by building a radiation hybrid map with markers in cR units.


Romain DE OLIVEIRA (INRA GDEC, France)
He defines CNV and then Presence Absence Variation (PAV). He explains his reference-mapping pipeline to identify TE-element-related CNV in wheat. He shows that wheat accessions can be clustered in terms of PAV of TEs.  At least 15% of genes are PAV variable among accessions.