The Melon Zym Locus Conferring Resistance to ZYMV: High Resolution Mapping and Candidate Gene Identification

Zucchini yellow mosaic virus (ZYMV; potyviridae) represents a major pathogen of Cucurbitaceae crops. ZYMV resistance in melon PI 414723 is conditioned by a dominant allele at the Zym locus. This resistant accession restricts viral spread and does not develop mosaic symptoms, but necrosis sometimes develops in response to inoculation. In previous studies, Zym has been mapped to linkage group II of the melon genetic map. In the present study, positional cloning of the locus was undertaken, starting from the CM-AG36 SSR marker at approximately 2 cm distance. We utilized five mapping populations that share the same resistant parent, PI 414723, and analyzed a total of 1630 offspring, to construct a high-resolution genetic map of the Zym locus. Two melon BAC libraries were used for chromosome walking and for developing new markers closer to the resistance gene by BAC-end sequencing. A BAC contig was constructed, and we identified a single BAC clone, from the ZYMV susceptible genotype MR-1, that physically encompasses the resistance gene. A second clone was isolated from another susceptible genotype, WMR 29, and the two clones were fully sequenced and annotated. Additional markers derived from the sequenced region delimited the region to 17.6 kb of a sequence that harbors a NAC-like transcription factor and, depending on the genotype, either two or three R-gene homologs with a CC-NBS-LRR structure. Mapping was confirmed by saturating the map with SNP markers using a single mapping population. The same region was amplified and sequenced also in the ZYMV resistant genotype PI 414723. Because numerous polymorphic sites were noted between genotypes, we could not associate resistance with a specific DNA polymorphism; however, this study enables molecular identification of Zym and paves the way to functional studies of this important locus.

Variability and Trait Association Studies for Late Leaf Spot Resistance in a Groundnut MAGIC Population

Globally, late leaf spot (LLS), a foliar fungal disease is one of the most important biotic constraint in groundnut production. Multi-Parent Advanced Generation Inter Cross (MAGIC) groundnut population was developed in a convergent crossing scheme using eight founder parents to develop a mapping population for multiple traits includes LLS. The experiments conducted in light chamber using detached leaf assay, and disease field screening nurseries at two locations (ICRISAT and ARS, Kasbe Digraj) showed significant variability for LLS resistance and component of resistance traits. Total 10 MAGIC lines with longer incubation (>11.0 days) and two MAGIC lines with longer latent period (>27 days) than the resistant parent, GPBD 4 were identified. The MAGIC lines, ICGR 171413, and ICGR 171443 with a lesion diameter of <1 mm and 4.10–5.67% of leaf area damage can be valuable sources for the alleles limiting the pathogen severity. A total of 20 MAGIC lines recorded significantly superior for disease score at 105 DAP_I (5.60–6.89) compared to resistant check, GPDB 4 (6.89). Further studies to determine the type and number of genes controlling the LLS component traits in groundnut will be useful for improvement of resistance to LLS. Genomic selection approach can be valuable in groundnut breeding to harness the minor alleles contributing to the component traits of LLS resistance.

NB-LRR-encoding genes conferring susceptibility to organophosphate pesticides in sorghum

Organophosphate is the commonly used pesticide to control pest outbreak, such as those by aphids in many crops. Despite its wide use, however, necrotic lesion and/or cell death following the application of organophosphate pesticides has been reported to occur in several species. To understand this phenomenon, called organophosphate pesticide sensitivity (OPS) in sorghum, we conducted QTL analysis in a recombinant inbred line derived from the Japanese cultivar NOG, which exhibits OPS. Mapping OPS in this population identified a prominent QTL on chromosome 5, which corresponded to Organophosphate-Sensitive Reaction (OSR) reported previously in other mapping populations. The OSR locus included a cluster of three genes potentially encoding nucleotide-binding leucine-rich repeat (NB-LRR, NLR) proteins, among which NLR-C was considered to be responsible for OPS in a dominant fashion. NLR-C was functional in NOG, whereas the other resistant parent, BTx623, had a null mutation caused by the deletion of promoter sequences. Our finding of OSR as a dominant trait is important not only in understanding the diversified role of NB-LRR proteins in cereals but also in securing sorghum breeding free from OPS.

Characterisation of a 4A QTL for Metribuzin Resistance in Wheat by Developing Near-Isogenic Lines

Wheat (Triticum aestivum L.) production is constantly affected by weeds in the farming system. Chemical-based weed management is widely practiced; broad-spectrum herbicides such as metribuzin have been successfully used to control weeds in Australia and elsewhere of the world. Breeding metribuzin-resistant wheat through genetic improvement is needed for effective control of weeds. Quantitative trait loci (QTLs) mapping efforts identified a major QTL on wheat chromosome 4A, explaining up to 20% of the phenotypic variance for metribuzin resistance. The quantitative nature of inheritance of this QTL signifies the importance of near-isogenic lines (NILs), which can convert a quantitative trait into a Mendelian factor for better resolution of the QTL. In the current study, NILs were developed using a heterogeneous inbred family method combined with a fast generation-cycling system in a population of Chuan Mai 25 (resistant) and Ritchie (susceptible). Seven pairs of NILs targeting the 4A QTL for metribuzin resistance were confirmed with a molecular marker and phenotyping. The resistant allele from the resistant parent increased metribuzin resistance by 63–85% (average 69%) compared with the susceptible allele from the susceptible parent. Segregation analysis in the NIL pairs for thousand grain weight (TGW) (g), biomass per plant (kg), tillers per plant, plant height (cm), yield per plant, and powdery mildew visual score (0–9) indicated that these traits were linked with metribuzin resistance. Similarly, TGW was observed to co-segregate with metribuzin resistance in most confirmed NILs, signifying that the two traits are controlled by closely linked genes. The most contrasting NILs can be further characterised by transcriptomic and proteomic analyses to identify the candidate genes responsible for metribuzin resistance.

Developing high-yielding Striga-resistant maize in sub-Saharan Africa.

Striga hermonthica is a major stress of maize in sub-Saharan Africa (SSA). The International Institute of Tropical Agriculture (IITA), in collaboration with national scientists, have used team approach to investigate how best to solve the problem. Emphasis was on (i) establishing a reliable infestation technique for selecting resistant/tolerant genotypes, (ii) availability of appropriate germplasm and good sources of Striga resistance, (iii) use of appropriate breeding methods for incorporating resistance genes into adapted germplasm, and (iv) extensive multilocational evaluation to identify genotypes with stable performance. Host plant resistance, with additive-dominance model, has been the major control option for S. hermonthica infestation on maize. Recurrent selection, followed by hybridization of inbred lines developed from its products, have been used to identify high-yielding, stable hybrids for commercialization in SSA. In a study involving early-maturing tropical maize inbred lines, 24 single nucleotide polymorphism (SNP) markers significantly associated with grain yield, Striga damage, ears per plant, and ear aspect under Striga infestation were detected. In a quantitative trait loci (QTL) mapping study involving extra-early white BC1S1 families obtained from TZEEI 29 (Striga-resistant) and TZEEI 23 (Striga susceptible) inbreds, 14 QTLs were identified for Striga resistance/tolerance traits. In a second QTL study involving the extra-early yellow F2:3 families derived from the Striga-resistant parent (TZEEI 79) and the susceptible parent (TZdEEI 11), 12 QTL were identified for 4 Striga resistance/tolerance traits. QTL identified in the studies would be invaluable for rapid introgression of Striga resistance genes into maize genotypes using marker-assisted selection approaches after validation of QTL in inbreds.

Discovery of Genomic Regions Associated With Resistance to Late Wilt Disease Caused by Harpophora Maydis (Samra, Sabet and Hing) in Maize (Zea mays L.)

Late wilt disease (LWD) caused by Harpophora maydis (Samra, Sabet and Hing) is emerging as major production constraint in maize across the world. As a prelude to develop maize hybrids resistance to LWD, genetic basis of resistance was investigated. Two F2:3 mapping populations (derived from CV156670 × 414-33 (P-1) and CV156670 × CV143587 (P-2)) were challenged with LWD at two locations (Kallinayakanahalli and Muppadighatta) during 2017 post-rainy season. Wider range of LWD scores were observed at both locations in both the populations. LWD response was influenced by significant Genotype × location interaction. Six and 56 F2:3 progeny families showed resistance level better than resistant parent. 150 and 199 polymorphic SNP markers were used to genotype P-1 and P-2, respectively. Inclusive composite interval mapping was performed to detect significant QTL, QTL × QTL, QTL × Location interaction effects. Three major and four minor QTL controlling LWD resistance were detected on chromosome-1. Position and effect of the QTL varied with the location. Significant di-QTL interactions involving QTL (with significant and/or non-significant effects) located within and between all the ten chromosomes were detected. Five of the seven detected QTL in our study showed significant QTL × location interaction. Though two major QTL (q-lw-1.5 and q-lw-1.6) with lower Q×L interaction effects could be considered as stable, their phenotypic variance is not large enough to deploy them in MAS. Based on these results, strategies to breed maize for resistance to LWD are discussed.

Discovery of a major QTL for root-knot nematode (Meloidogyne incognita) resistance in cultivated sweetpotato (Ipomoea batatas)

Key message Utilizing a high-density integrated genetic linkage map of hexaploid sweetpotato, we discovered a major dominant QTL for root-knot nematode (RKN) resistance and modeled its effects. This discovery is useful for development of a modern sweetpotato breeding program that utilizes marker-assisted selection and genomic selection approaches for faster genetic gain of RKN resistance. Abstract The root-knot nematode [Meloidogyne incognita (Kofoid & White) Chitwood] (RKN) causes significant storage root quality reduction and yields losses in cultivated sweetpotato [Ipomoea batatas (L.) Lam.]. In this study, resistance to RKN was examined in a mapping population consisting of 244 progenies derived from a cross (TB) between ‘Tanzania,’ a predominant African landrace cultivar with resistance to RKN, and ‘Beauregard,’ an RKN susceptible major cultivar in the USA. We performed quantitative trait loci (QTL) analysis using a random-effect QTL mapping model on the TB genetic map. An RKN bioassay incorporating potted cuttings of each genotype was conducted in the greenhouse and replicated five times over a period of 10 weeks. For each replication, each genotype was inoculated with ca. 20,000 RKN eggs, and root-knot galls were counted ~62 days after inoculation. Resistance to RKN in the progeny was highly skewed toward the resistant parent, exhibiting medium to high levels of resistance. We identified one major QTL on linkage group 7, dominant in nature, which explained 58.3% of the phenotypic variation in RKN counts. This work represents a significant step forward in our understanding of the genetic architecture of RKN resistance and sets the stage for future utilization of genomics-assisted breeding in sweetpotato breeding programs.

High resolution genetic dissection of the major QTL for tipburn resistance in lettuce, Lactuca sativa

Tipburn is an important physiological disorder of lettuce, Lactuca sativa L., related to calcium deficiency that can result in leaf necrosis and unmarketable crops. The major quantitative trait locus, qTPB5.2, can account for up to 70% of the phenotypic variance for tipburn incidence in the field. This quantitative trait locus was genetically dissected to identify candidate genes for tipburn by creating lines with recombination events within the quantitative trait locus and assessing their resistance to tipburn. By comparing lines with contrasting haplotypes, the genetic region was narrowed down to ∼877 Kb that was associated with a reduction of tipburn by ∼60%. Analysis of the lettuce reference genome sequence revealed 12 genes in this region, one of which is a calcium transporter with a single nucleotide polymorphism in an exon between haplotypes with contrasting phenotypes. RNA-seq analysis of recombinants revealed two genes that were differentially expressed between contrasting haplotypes consistent with the tipburn phenotype. One encodes a Teosinte branched1/Cycloidea/Proliferating Cell factor transcription factor; however, differential expression of the calcium transporter was not detected. The phenotypic data indicated that there is a second region outside of the ∼877 Kb region but within the quantitative trait locus, at which a haplotype from the susceptible parent decreased tipburn by 10 to 20%. A recombinant line was identified with beneficial haplotypes in each region from both parents that showed greater tipburn resistance than the resistant parent; this line could be used as the foundation for breeding cultivars with more resistance than is currently available.

Mapping and validation of a stable quantitative trait locus conferring maize resistance to Gibberella ear rot

Gibberella ear rot (GER), a prevalent disease caused by Fusarium graminearum (F. graminearum), can result in significant yield loss and carcinogenic mycotoxin contamination in maize worldwide., However, only a few quantitative trait loci (QTLs) for GER resistance have been reported. In this study, we evaluated a Chinese recombinant inbred line (RIL) population comprising of 204 lines, developed from a cross between a resistant parent DH4866 and a susceptible line T877, in three field trial environments under artificial inoculation with F. graminearum. The RIL population and their parents were genotyped with an Affymetrix microarray CGMB56K SNP Array. Based on the genetic linkage map constructed using 1,868 bins as markers, eleven QTLs, including five stable QTLs, were identified by individual environment analysis. Joint multiple environments analysis and epistatic interaction analysis revealed six additive and six epistatic (additive × additive) QTLs, respectively. None of the QTLs could explain more than 10% of phenotypic variation, suggesting that multiple minor-effect QTLs contributed to the genetic component of resistance to GER, and both additive and epistatic effects contributed to the genetic architecture of resistance to GER. A novel QTL, qGER4.09, with largest effect, identified and validated using 588 F2 individuals, was colocalized with genomic regions for FER and Aspergillus ear rot, indicating that this genetic locus likely confers resistance to multiple pathogens, and can be potentially utilized in breeding maize varieties, aimed at improving the resistance not only to GER, but also other ear rot diseases.

An Integrated Approach of QTL Mapping and Genome-Wide Association Analysis Identifies Candidate Genes for Phytophthora Blight Resistance in Sesame (Sesamum indicum L.)

Phytophthora blight (PB) caused by Phytophthora nicotianae is a highly destructive disease in sesame (Sesamum indicum L.). In this study, we used linkage mapping and genome-wide association study (GWAS) to identify quantitative trait loci (QTL) and candidate genes associated with PB resistance. The QTL mapping in 90 RILs of the Goenbaek × Osan cross using genotyping-by-sequencing detected significant QTLs for PB resistance on chromosome 10, explaining 12.79%–13.34% of phenotypic variation. Association of this locus to PB resistance was also revealed through bulked segregant analysis in second RIL population (Goenbaek × Milsung cross) comprising 188 RILs. The GWAS of 87 sesame accessions evaluated against three P. nicotianae isolates identified 29 SNPs on chromosome 10 significantly associated with PB resistance. These SNPs were located within a 0.79 Mb region, which co-located with the QTL intervals identified in RIL populations, and hence scanned for identifying candidate genes. This region contained several defense-related candidate R genes, five of which were selected for quantitative expression analysis. One of these genes, SIN_1019016 was found to show significantly higher expression in the resistant parent compared to that in the susceptible parents and selected RILs. Paired-end sequencing of the gene SIN_1019016 in parental cultivars revealed two synonymous SNPs between Goenbaek and Osan in exon 2 of coding DNA sequence. These results suggested SIN_1019016 as one of the candidate gene conferring PB resistance in sesame. The findings from this study will be useful in the marker-assisted selection as well as the functional analysis of PB resistance candidate gene(s) in sesame.