Asheesh K. Singh

Machine Learning for High-Throughput Stress Phenotyping in Plants

Advances in automated and high-throughput imaging technologies have resulted in a deluge of high-resolution images and sensor data of plants. However, extracting patterns and features from this large corpus of data requires the use of machine learning (ML) tools to enable data assimilation and feature identification for stress phenotyping. Four stages of the decision cycle in plant stress phenotyping and plant breeding activities where different ML approaches can be deployed are (i) identification, (ii) classification, (iii) quantification, and (iv) prediction (ICQP). We provide here a comprehensive overview and user-friendly taxonomy of ML tools to enable the plant community to correctly and easily apply the appropriate ML tools and best-practice guidelines for various biotic and abiotic stress traits.

Phytochemicals to suppress Fusarium head blight in wheat–chickpea rotation

Fusarium diseases cause major economic losses in wheat-based crop rotations. Volatile organic compounds (VOC) in wheat and rotation crops, such as chickpea, may negatively impact pathogenic Fusarium. Using the headspace GC-MS method, 16 VOC were found in greenhouse-grown wheat leaves: dimethylamine, 2-methyl-1-propanol, octanoic acid-ethyl ester, acetic acid, 2-ethyl-1-hexanol, nonanoic acid-ethyl ester, nonanol, N-ethyl-benzenamine, naphthalene, butylated hydroxytoluene, dimethoxy methane, phenol, 3-methyl-phenol, 3,4-dimethoxy-phenol, 2,4-bis (1,1-dimethyethyl)-phenol, and 1,4,7,10,13,16-hexaoxacyclooctadecane; and 10 VOC in field-grown chickpea leaves: ethanol, 1-penten-3-ol, 1-hexanol, cis-3-hexen-1-ol, trans-2-hexen-1-ol, trans-2-hexenal, 3-methyl-1-butanol, 3-hydroxy-2-butanone, 3-methyl-benzaldehyde and naphthalene. Also found was 1-penten-3-ol in chickpea roots and in the root nodules of two of the three cultivars tested. Chickpea VOC production pattern was related (P=0.023) to Ascochyta blight severity, suggesting that 1-penten-3-ol and cis-3-hexen-1-ol were induced by Ascochyta rabiei. Bioassays conducted in Petri plates established that chickpea-produced VOC used in isolation were generally more potent against Fusarium graminearum and Fusarium avenaceum than wheat-produced VOC, except for 2-ethyl-1-hexanol, which was rare in wheat and toxic to both Fusarium and tetraploid wheat. Whereas exposure to 1-penten-3-ol and 2-methyl-1-propanol could suppress radial growth by over 50% and octanoic acid-ethyl ester, nonanol, and nonanoic acid-ethyl ester had only weak effects, F. graminearum and F. avenaceum growth was completely inhibited by exposure to trans-2-hexenal, trans-2-hexen-1-ol, cis-3-hexen-1-ol, and 1-hexanol. Among these VOC, trans-2-hexenal and 1-hexanol protected wheat seedlings against F. avenaceum and F. graminearum, respectively, in a controlled condition experiment. Genetic variation in the production of 2-ethyl-1-hexanol, a potent VOC produced in low amount by wheat, suggests the possibility of selecting Fusarium resistance in wheat on the basis of leaf VOC concentration. Results also suggests that the level of Fusarium inoculum in chickpea-wheat rotation systems may be reduced by growing chickpea genotypes with high root and shoot levels of trans-2-hexen-1-ol and 1-hexanol.

Identification and mapping of leaf, stem and stripe rust resistance quantitative trait loci and their interactions in durum wheat.

Leaf rust (Puccinia triticina Eriks.), stripe rust (Puccinia striiformis f. tritici Eriks.) and stem rust (Puccinia graminis f. sp. tritici) cause major production losses in durum wheat (Triticum turgidum L. var. durum). The objective of this research was to identify and map leaf, stripe and stem rust resistance loci from the French cultivar Sachem and Canadian cultivar Strongfield. A doubled haploid population from Sachem/Strongfield and parents were phenotyped for seedling reaction to leaf rust races BBG/BN and BBG/BP and adult plant response was determined in three field rust nurseries near El Batan, Obregon and Toluca, Mexico. Stripe rust response was recorded in 2009 and 2011 nurseries near Toluca and near Njoro, Kenya in 2010. Response to stem rust was recorded in field nurseries near Njoro, Kenya, in 2010 and 2011. Sachem was resistant to leaf, stripe and stem rust. A major leaf rust quantitative trait locus (QTL) was identified on chromosome 7B at Xgwm146 in Sachem. In the same region on 7B, a stripe rust QTL was identified in Strongfield. Leaf and stripe rust QTL around DArT marker wPt3451 were identified on chromosome 1B. On chromosome 2B, a significant leaf rust QTL was detected conferred by Strongfield, and at the same QTL, a Yr gene derived from Sachem conferred resistance. Significant stem rust resistance QTL were detected on chromosome 4B. Consistent interactions among loci for resistance to each rust type across nurseries were detected, especially for leaf rust QTL on 7B. Sachem and Strongfield offer useful sources of rust resistance genes for durum rust breeding.

Genome-wide association and epistasis studies unravel the genetic architecture of sudden death syndrome resistance in soybean.

Soybean [Glycine max (L.) Merr.] is an economically important crop that is grown worldwide. Sudden death syndrome (SDS), caused by Fusarium virguliforme, is one of the top yield-limiting diseases in soybean. However, the genetic basis of SDS resistance, especially with respect to epistatic interactions, is still unclear. To better understand the genetic architecture of soybean SDS resistance, genome-wide association and epistasis studies were performed using a population of 214 germplasm accessions and 31,914 SNPs from the SoySNP50K Illumina Infinium BeadChip. Twelve loci and 12 SNP-SNP interactions associated with SDS resistance were identified at various time points after inoculation. These additive and epistatic loci together explained 24-52% of the phenotypic variance. Disease-resistant, pathogenesis-related and chitin- and wound-responsive genes were identified in the proximity of peak SNPs, including stress-induced receptor-like kinase gene 1 (SIK1), which is pinpointed by a trait-associated SNP and encodes a leucine-rich repeat-containing protein. We report that the proportion of phenotypic variance explained by identified loci may be considerably improved by taking epistatic effects into account. This study shows the necessity of considering epistatic effects in soybean SDS resistance breeding using marker-assisted and genomic selection approaches. Based on our findings, we propose a model for soybean root defense against the SDS pathogen. Our results facilitate identification of the molecular mechanism underlying SDS resistance in soybean, and provide a genetic basis for improvement of soybean SDS resistance through breeding strategies based on additive and epistatic effects.

Developing standardized methods for breeding preharvest sprouting resistant wheat, challenges and successes in Canadian wheat

Preharvest sprouting (PHS) in spring wheat (Triticum aestivum L.) and durum wheat (T. turgidum L. var durum) causes significant economic losses due to a reduction in grain yield, grain functionality and viability of seed for planting. Average annual estimated losses in Canada are about

Stripe rust and leaf rust resistance QTL mapping, epistatic interactions, and co‑localization with stem rust resistance loci in spring wheat evaluated over three continents

100 million. Genetic resistance to PHS reduces these losses. Development of PHS resistant cultivars is complicated by the effects of factors under genetic control, such as spike morphology, seed dormancy, environment, and kernel diseases. Resistance to PHS has been a breeding priority since the late 1960s. Development of RL4137, which is the primary source of PHS resistance in the Canada Western Red Spring market class, has led to cultivar improvements. A white-seeded derivative of RL4137 is the primary source of PHS in the Canada Prairie Spring White and Canada Western Hard White Spring wheat market classes. Procedures to select for PHS resistance vary among breeding programs, market classes and by degree of inbreeding. Methods include artificial sprouting of intact spikes, germination tests, natural weathering in field trials, artificial weathering trials, and indirect assessment of sprouting by measuring Hagberg falling number. Although many genetic loci have been attributed to preharvest sprouting resistance, application of molecular markers is currently limited due to the complex inheritance of the trait. In Canada, cultivars are characterized for their relative level of PHS resistance and the information is made available to producers.

Computer vision and machine learning for robust phenotyping in genome-wide studies

Key messageIn wheat, advantageous gene-rich or pleiotropic regions for stripe, leaf, and stem rust and epistatic interactions between rust resistance loci should be accounted for in plant breeding strategies.AbstractLeaf rust (Puccinia triticina Eriks.) and stripe rust (Puccinia striiformis f. tritici Eriks) contribute to major production losses in many regions worldwide. The objectives of this research were to identify and study epistatic interactions of quantitative trait loci (QTL) for stripe and leaf rust resistance in a doubled haploid (DH) population derived from the cross of Canadian wheat cultivars, AC Cadillac and Carberry. The relationship of leaf and stripe rust resistance QTL that co-located with stem rust resistance QTL previously mapped in this population was also investigated. The Carberry/AC Cadillac population was genotyped with DArT® and simple sequence repeat markers. The parents and population were phenotyped for stripe rust severity and infection response in field rust nurseries in Kenya (Njoro), Canada (Swift Current), and New Zealand (Lincoln); and for leaf rust severity and infection response in field nurseries in Canada (Swift Current) and New Zealand (Lincoln). AC Cadillac was a source of stripe rust resistance QTL on chromosomes 2A, 2B, 3A, 3B, 5B, and 7B; and Carberry was a source of resistance on chromosomes 2B, 4B, and 7A. AC Cadillac contributed QTL for resistance to leaf rust on chromosome 2A and Carberry contributed QTL on chromosomes 2B and 4B. Stripe rust resistance QTL co-localized with previously reported stem rust resistance QTL on 2B, 3B, and 7B, while leaf rust resistance QTL co-localized with 4B stem rust resistance QTL. Several epistatic interactions were identified both for stripe and leaf rust resistance QTL. We have identified useful combinations of genetic loci with main and epistatic effects. Multiple disease resistance regions identified on chromosomes 2A, 2B, 3B, 4B, 5B, and 7B are prime candidates for further investigation and validation of their broad resistance.

Quantification of effects of leaf spotting diseases on grain yield and market quality of durum wheat using near-isogenic lines

Traditional evaluation of crop biotic and abiotic stresses are time-consuming and labor-intensive limiting the ability to dissect the genetic basis of quantitative traits. A machine learning (ML)-enabled image-phenotyping pipeline for the genetic studies of abiotic stress iron deficiency chlorosis (IDC) of soybean is reported. IDC classification and severity for an association panel of 461 diverse plant-introduction accessions was evaluated using an end-to-end phenotyping workflow. The workflow consisted of a multi-stage procedure including: (1) optimized protocols for consistent image capture across plant canopies, (2) canopy identification and registration from cluttered backgrounds, (3) extraction of domain expert informed features from the processed images to accurately represent IDC expression, and (4) supervised ML-based classifiers that linked the automatically extracted features with expert-rating equivalent IDC scores. ML-generated phenotypic data were subsequently utilized for the genome-wide association study and genomic prediction. The results illustrate the reliability and advantage of ML-enabled image-phenotyping pipeline by identifying previously reported locus and a novel locus harboring a gene homolog involved in iron acquisition. This study demonstrates a promising path for integrating the phenotyping pipeline into genomic prediction, and provides a systematic framework enabling robust and quicker phenotyping through ground-based systems.

Eurostar durum wheat

Abstract Leaf spotting diseases (LS) in wheat are widespread in western Canada. The most prevalent LS are tan spot and the septoria leaf blotch complex. Near-isogenic lines (NILs) for LS resistance were used to determine the impact of LS on grain yield and market quality of durum wheat. Sixteen NIL pairs, susceptible and resistant to LS caused mostly by Pyrenophora tritici-repentis (tan spot), were evaluated at the milk stage at two locations in southern Saskatchewan, from 2000 to 2002. Tan spot was the most prevalent LS on the leaves. There were differences in plant growth and disease severity among environments. However, in most cases, the resistant NILs had the lowest LS severity. Overall the resistant NILs had higher yields and lower protein concentration than the susceptible NILs, although differences for individual lines were not always significant. For the four most consistently performing pairs in the most favourable years of 2000 and 2002, a 16.2% LS reduction corresponded to increases of 17.2% for yield and 4.6% for kernel weight at Swift Current, whereas a LS reduction of 5.5% was associated with increases of 26.6% for yield and 2.8% for kernel weight at Indian Head. This indicates that in the semi-arid location of Swift Current, a greater LS reduction was required to obtain yield and quality increases similar to those at the location with the greatest yield potential, Indian Head. Protein was reduced in the resistant NILs by 4.1% at Swift Current and 7.6% at Indian Head, suggesting that improvements in LS resistance must be accompanied by breeding for higher protein concentration or recommendations for nitrogen application to maximize returns.

Potential to breed for mycorrhizal association in durum wheat.

Eurostar durum wheat [Triticum turgidum L. subsp. durum (Desf.) Husn.] is adapted to the durum production area of the Canadian prairies. It combines high grain yield, high grain protein concentration, very strong gluten, and low grain cadmium concentration. Eurostar has similar straw strength to Strongfield, and slightly later maturity and similar disease resistance to other currently registered durum cultivars. Key words: Triticum turgidum L. subsp. durum (Desf.) Husn., durum wheat, cultivar description, yield, protein, disease resistance