Most published genome sequences are drafts, and most are dominated by computational gene prediction. Draft genomes typically incorporate considerable sequence data that are not assigned to chromosomes, and predicted genes without quality confidence measures. The current Actinidia chinensis (kiwifruit) ‘Hongyang’ draft genome has 164 Mb of sequences unassigned to pseudo-chromosomes, and omissions have been identified in the gene models. A second genome of an A. chinensis (genotype Red5) was fully sequenced. This new sequence resulted in a 554.0 Mb assembly with all but 6 Mb assigned to pseudo-chromosomes. Pseudo-chromosomal comparisons showed a considerable number of translocation events have occurred following a whole genome duplication (WGD) event some consistent with centromeric Robertsonian-like translocations. RNA sequencing data from 12 tissues and ab initio analysis informed a genome-wide manual annotation, using the WebApollo tool. In total, 33,044 gene loci represented by 33,123 isoforms were identified, named and tagged for quality of evidential support. Of these 3114 (9.4%) were identical to a protein within ‘Hongyang’ The Kiwifruit Information Resource (KIR v2). Some proportion of the differences will be varietal polymorphisms. However, as most computationally predicted Red5 models required manual re-annotation this proportion is expected to be small. The quality of the new gene models was tested by fully sequencing 550 cloned ‘Hort16A’ cDNAs and comparing with the predicted protein models for Red5 and both the original ‘Hongyang’ assembly and the revised annotation from KIR v2. Only 48.9% and 63.5% of the cDNAs had a match with 90% identity or better to the original and revised ‘Hongyang’ annotation, respectively, compared with 90.9% to the Red5 models. Our study highlights the need to take a cautious approach to draft genomes and computationally predicted genes. Our use of the manual annotation tool WebApollo facilitated manual checking and correction of gene models enabling improvement of computational prediction. This utility was especially relevant for certain types of gene families such as the EXPANSIN like genes. Finally, this high quality gene set will supply the kiwifruit and general plant community with a new tool for genomics and other comparative analysis.

Summary: O‐acetylserine (thiol) lyases ( OASTL s) are evolutionarily conserved proteins among many prokaryotes and eukaryotes that perform sulfur acquisition and synthesis of cysteine. A mutation in the cytosolic OASTL‐A1 protein ONSET OF LEAF DEATH3 (OLD3) was previously shown to reduce the OASTL activity of the old3‐1 protein in vitro and cause auto‐necrosis in specific Arabidopsis accessions. Here we investigated why a mutation in this protein causes auto‐necrosis in some but not other accessions. The auto‐necrosis was found to depend on Recognition of Peronospora Parasitica 1 (RPP1)‐like disease resistance R gene(s) from an evolutionarily divergent R gene cluster that is present in Ler‐0 but not the reference accession Col‐0. RPP1‐like gene(s) show a negative epistatic interaction with the old3‐1 mutation that is not linked to reduced cysteine biosynthesis. Metabolic profiling and transcriptional analysis further indicate that an effector triggered‐like immune response and metabolic disorder are associated with auto‐necrosis in old3‐1 mutants, probably activated by an RPP1‐like gene. However, the old3‐1 protein in itself results in largely neutral changes in primary plant metabolism, stress defence and immune responses. Finally, we showed that lack of a functional OASTL‐A1 results in enhanced disease susceptibility against infection with virulent and non‐virulent Pseudomonas syringae pv. tomato DC3000 strains. These results reveal an interaction between the cytosolic OASTL and components of plant immunity.

Pseudomonas syringae pv. actinidiae (Psa) biovar 3, a virulent, canker-inducing pathogen is an economic threat to the kiwifruit (Actinidia spp.) industry worldwide. The commercially grown diploid (2×) A. chinensis var. chinensis is more susceptible to Psa than tetraploid and hexaploid kiwifruit. However information on the genetic loci modulating Psa resistance in kiwifruit is not available. Here we report mapping of quantitative trait loci (QTLs) regulating resistance to Psa in a diploid kiwifruit population, derived from a cross between an elite Psa-susceptible ‘Hort16A’ and a resistant male breeding parent P1. Using high-density genetic maps and intensive phenotyping, we identified a single QTL for Psa resistance on Linkage Group (LG) 27 of ‘Hort16A’ revealing 16–19% phenotypic variance and candidate alleles for susceptibility and resistance at this loci. In addition, six minor QTLs were identified in P1 on distinct LGs, exerting 4–9% variance. Resistance in the F1 population is improved by additive effects from ‘Hort16A’ and P1 QTLs providing evidence that divergent genetic pathways interact to combat the virulent Psa strain. Two different bioassays further identified new QTLs for tissue-specific responses to Psa. The genetic marker at LG27 QTL was further verified for association with Psa resistance in diploid Actinidia chinensis populations. Transcriptome analysis of Psa-resistant and susceptible genotypes in field revealed hallmarks of basal defense and provided candidate RNA-biomarkers for screening for Psa resistance in greenhouse conditions.

Polyploidy is a key driver of significant evolutionary changes in plant species. The genus Actinidia (kiwifruit) exhibits multiple ploidy levels, which contribute to novel fruit traits, high yields and resistance to the canker-causing dieback disease incited by Pseudomonas syringae pv. actinidiae (Psa) biovar 3. However, the genetic mechanism for resistance to Psa observed in polyploid kiwifruit is not yet known. In this study we performed detailed genetic analysis of a tetraploid Actinidia chinensis var. chinensis population derived from a cross between a female parent that exhibits weak tolerance to Psa and a highly Psa-resistant male parent. We used the capture-sequencing approach across the whole kiwifruit genome and generated the first ultra-dense maps in a tetraploid kiwifruit population. We located quantitative trait loci (QTLs) for Psa resistance on these maps. Our approach to QTL mapping is based on the use of identity-by-descent trait mapping, which allowed us to relate the contribution of specific alleles from their respective homologues in the male and female parent, to the control of Psa resistance in the progeny. We identified genes in the diploid reference genome whose function is suggested to be involved in plant defense, which underly the QTLs, including receptor-like kinases. Our study is the first to cast light on the genetics of a polyploid kiwifruit and suggest a plausible mechanism for Psa resistance in this species.

Summary: Testing effector knockout strains of the Pseudomonas syringae pv. actinidiae biovar 3 (Psa3) for reduced in planta growth in their native kiwifruit host revealed a number of nonredundant effectors that contribute to Psa3 virulence. Conversely, complementation in the weak kiwifruit pathogen P. syringae pv. actinidifoliorum (Pfm) for increased growth identified redundant Psa3 effectors. Psa3 effectors hopAZ1a and HopS2b and the entire exchangeable effector locus (ΔEEL; 10 effectors) were significant contributors to bacterial colonisation of the host and were additive in their effects on virulence. Four of the EEL effectors (HopD1a, AvrB2b, HopAW1a and HopD2a) redundantly contribute to virulence through suppression of pattern‐triggered immunity (PTI). Important Psa3 effectors include several redundantly required effectors early in the infection process (HopZ5a, HopH1a, AvrPto1b, AvrRpm1a and HopF1e). These largely target the plant immunity hub, RIN4. This comprehensive effector profiling revealed that Psa3 carries robust effector redundancy for a large portion of its effectors, covering a few functions critical to disease.