Resistance to anthracnose caused by the fungal pathogen Colletotrichum lentis was explored through transcriptome sequencing over a period of 24 to 96 h post-inoculation (hpi) of the partially ...resistant recombinant inbred lines (RIL) LR-66-528 and susceptible LR-66-524 of the crop wild relative Lens ervoides population LR-66. The development of infection vesicles and primary hyphae by C. lentis were significantly higher on susceptible RIL LR-66-524 compared to partially resistant LR-66-528 at 24 and 48 hpi, but exponential trends in fungal growth were observed between 24 to 96 hpi in both RILs. Comparison of inoculated with mock-inoculated samples revealed 3091 disease responsive genes, among which 477 were differentially expressed between the two RILs. These were clustered into six expression clusters with genes that had either high or low expression in one of the RILs. Differentially expressed genes (DEGs) were functionally annotated and included genes coding LRR and NB-ARC domain disease resistance proteins, protein detoxification, LRR receptor-like kinase family proteins, and wall-associated Ser/Thr Kinases. DEGs were compared to genes in previously published anthracnose resistance QTLs mapped in LR-66 and revealed 22 DEGs located in 3 QTLs. Expression of 21 DEGs was validated using RT-qPCR confirming expression trends in RNA-seq.
Stemphylium blight (SB), caused by Stemphylium botryosum, is a devastating disease in lentil production. Although it is known that accessions of Lens ervoides possess superior SB resistance at much ...higher frequency than the cultivated lentil species, very little is known about the molecular basis regulating SB resistance in L. ervoides. Therefore, a comprehensive molecular study of SB resistance in L. ervoides was needed to exploit this wild resource available at genebanks for use by plant breeders in resistance breeding.
Microscopic and qPCR quantification of fungal growth revealed that 48, 96, and 144 h post-inoculation (hpi) were interesting time points for disease development in L. ervoides recombinant inbred lines (RILs) LR-66-637 (resistant to SB) and LR-66-577 (susceptible to SB). Results of transcriptome sequencing at 0, 48, 96 and 144 hpi showed that 8810 genes were disease-responsive genes after challenge by S. botryosum. Among them, 7526 genes displayed a similar expression trend in both RILs, and some of them were likely involved in non-host resistance. The remaining 1284 genes were differentially expressed genes (DEGs) between RILs. Of those, 712 DEGs upregulated in LR-66-637 were mostly enriched in 'carbohydrate metabolic process', 'cell wall organization or biogenesis', and 'polysaccharide metabolic process'. In contrast, there were another 572 DEGs that were upregulated in LR-66-577, and some of them were enriched in 'oxidation-reduction process', 'asparagine metabolic process' and 'asparagine biosynthetic process'. After comparing DEGs to genes identified in previously described quantitative trait loci (QTLs) for resistance to SB, nine genes were common and three of them showed differential gene expression between a resistant and a susceptible bulk consisting of five RILs each. Results showed that two genes encoding calcium-transporting ATPase and glutamate receptor3.2 were candidate resistance genes, whereas one gene with unknown function was a candidate susceptibility gene.
This study provides new insights into the mechanisms of resistance and susceptibility in L. ervoides RILs responding to S. botryosum infection. Furthermore, we identified candidate resistance or susceptibility genes which warrant further gene function analyses, and which could be valuable for resistance breeding, if their role in resistance or susceptibility can be confirmed.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Abstract
Cultivated lentil (
Lens culinaris
Medik.) is susceptible to aphanomyces root rot (
ARR
), whereas partial resistance is present in wild lentil including
Lens ervoides
(Brign.) Grande. ...Approximately six generations of selfing are required to fix a desired trait in a population, which usually requires 2 years in a breeding programme, so the primary objective was to develop a rapid generation cycling (
RGC
) technique that achieves this goal in 1 year. Rapid generation cycling was then tested on an F
2
population (
LR
‐59) derived from a
L. culinaris
×
L. ervoides
cross in combination with a reliable
ARR
screening technique, which generates a wide range of disease severities conducive to selection. Phenotyping of an F
2
population of more than 1,200 plants resulted in scores ranging from 2.4 to 4.0 on a scale from zero to five. Plants with scores lower than 4.0 were selected for advancement for five generations using a modified single‐seed descent method, optimum growing conditions, 20‐hr photoperiod and harvest of immature seeds. Seeds were germinated in a 100 μM gibberellin solution. Average generation length after phenotyping was 56 days resulting in five generations within approximately 300 days. Using a modified inoculation protocol,
ARR
phenotyping of the F
7
population resulted in scores ranging from 1.4 to 4.0. This inexpensive, nonsterile speed breeding protocol saves 1 year in the development of lentil varieties with improved
ARR
resistance.
Lens ervoides, a wild relative of lentil is an important source of allelic diversity for enhancing the genetic resistance of the cultivated species against economically important fungal diseases, ...such as anthracnose and Stemphylium blight caused by Colletotrichum lentis and Stemphylium botryosum, respectively. To unravel the genetic control underlying resistance to these fungal diseases, a recombinant inbred line (RIL) population (n = 94, F
) originating from a cross between two L. ervoides accessions, L01-827A and IG 72815, was genotyped on the Illumina HiSeq 2500 platform. A total of 289.07 million 100 bp paired-end reads were generated, giving an average 7.53-fold genomic coverage to the RILs and identifying 2,180 high-quality SNPs that assembled in 543 unique haplotypes. Seven linkage groups were resolved among haplotypes, equal to the haploid chromosome number in L. ervoides. The genetic map spanned a cumulative distance of 740.94 cM. Composite interval mapping revealed five QTLs with a significant association with resistance to C. lentis race 0, six QTLs for C. lentis race 1 resistance, and three QTLs for S. botryosum resistance. Taken together, the data obtained in the study reveal that the expression of resistance to fungal diseases in L. ervoides is a result of rearrangement of resistant alleles contributed by both parental accessions.
Outbreaks of blossom blight in coriander (Coriandrum sativum) and caraway (Carum carvi) are common in western Canada, particularly when cool, wet conditions prevail during flowering, which can lead ...to total crop loss. Earlier attempts to identify the causal organisms were inconclusive, which has hampered blossom blight management and been a barrier for effective resistance breeding in these two crops. In order to address this knowledge gap, the pathogenicity of organisms isolated from coriander and caraway blossoms collected during blossom blight surveys in Saskatchewan in 2015 and 2016 was tested under controlled environment conditions. Isolates of Didymella cari, an undescribed fungus tentatively identified as a Heterosphaeria sp., Fusarium avenaceum, F. graminearum, Botrytis cinerea, as well as an isolate of Sclerotinia sclerotiorum obtained from the USA, were all shown to be pathogenic to coriander blossoms. An isolate of Septoria carvi was isolated from diseased caraway foliage and identified based on spore morphology, disease symptoms and sequence data. This is the first report of Septoria carvi in Canada. Isolates of Heterosphaeria sp. were shown to be pathogenic on caraway blossoms, whereas Septoria carvi and Aureobasidium pullulans failed to cause disease on caraway blossoms under the study conditions.
Colletotrichum lentis causes anthracnose, which is a serious disease on lentil and can account for up to 70% crop loss. Two pathogenic races, 0 and 1, have been described in the C. lentis population ...from lentil.
To unravel the genetic control of virulence, an isolate of the virulent race 0 was sequenced at 1481-fold genomic coverage. The 56.10-Mb genome assembly consists of 50 scaffolds with N
50 scaffold length of 4.89 Mb. A total of 11 436 protein-coding gene models was predicted in the genome with 237 coding candidate effectors, 43 secondary metabolite biosynthetic enzymes and 229 carbohydrate-active enzymes (CAZymes), suggesting a contraction of the virulence gene repertoire in C. lentis.
Scaffolds were assigned to 10 core and two minichromosomes using a population (race 0 9 race 1, n = 94 progeny isolates) sequencing-based, high-density (14 312 single nucleotide polymorphisms) genetic map. Composite interval mapping revealed a single quantitative trait locus (QTL), qClVIR-11, located on minichromosome 11, explaining 85% of the variability in virulence of the C. lentis population. The QTL covers a physical distance of 0.84 Mb with 98 genes, including seven candidate effector and two secondary metabolite genes.
Taken together, the study provides genetic and physical evidence for the existence of a minichromosome controlling the C. lentis virulence on lentil.
Chickpea fields in Saskatchewan, one of the three Canadian prairie provinces, have suffered from major health issues since 2019, but no definitive cause has been determined. Field surveys were ...conducted in Saskatchewan in 2020 and 2021 in order to develop a better understanding of root rot pathogens associated with chickpea. Root samples were analyzed for the presence of 11 potential chickpea root rot pathogens using end-point PCR.
,
and
were the most prevalent pathogen species detected in both survey years. The cause of Fusarium wilt in chickpea,
f. sp.
, was not detected in either year, nor were
spp. and
.
sp. was detected in one field in each year, and
was detected in several fields sampled in 2021. These two pathogens have not been reported previously on chickpea in Saskatchewan. The prevalence of
species obtained from 2021 root isolations was similar to that determined by molecular tests, with frequent isolation of
,
,
and
. A series of indoor pathogenicity testing compared root disease severity caused by a selection of 16 isolates of six
species and single isolates of
,
sp. and
. Results showed that select isolates of
were the most aggressive of the
isolates on chickpea. Despite relatively low inoculum density, a highly aggressive isolate of
caused severe stunting and more root rot symptoms than single isolates of
,
sp. and
under the test conditions.
Ascochyta lentis
is a foliar pathogen of
Lens
species and is of worldwide importance in cultivated lentil production. High levels of resistance were identified in the wild species
Lens ervoides
. ...This resistance was explored through histopathology, qPCR estimation of fungal biomass and transcriptome sequencing in a susceptible and a resistant recombinant inbred line (RIL) of
L. ervoides
infected with an aggressive isolate of
A. lentis
. Necrotrophic growth was delayed in the resistant RIL compared to accelerated necrotrophy of
A. lentis
in the susceptible RIL. Analysis of the fungal secretome indicated that the early activation of cell wall-degrading enzymes contributed to increased virulence of
A. lentis
. On the host side, gene co-expression analysis revealed that the invasion by
A. lentis
caused mRNA, DNA and protein decay in infected plants regardless of the level of resistance in the host. The resistant RIL exhibited a stronger gene co-expression in lipid localization and sulfur processes, and cellular responses to nutrients and stimuli than the susceptible RIL. In addition, differential gene analysis revealed that the repression of both, gibberellin signaling and cell death associated with the hypersensitive response (HR), were associated with enhanced
A. lentis
resistance.
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