Poster Abstracts
Chickpea Innovation Lab 2016 Meeting


Abstracts are listed in alphabetical order by last name of first author.  

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Abstract Titles - click title to go to the full abstract

  1. Expanding Chickpea in non-traditional Lowlands of Ethiopia
  2. Developing a Resource of Wild x Cultivated Introgression Lines for Crop Improvement of Chickpea
  3. Transpiration rate of chickpea wild accessions and cultivars in Turkish and Indian location
  4. Status of genomic assisted resistance breeding for the development of Fusarium wilt (Fusarium oxysporum fsp.ciceris) resistant Chickpea (Cicer arietinum L.) genotypes in Ethiopia
  5. Effect of inoculation of chickpea and lentil with stress tolerant rhizobia of Morocco
  6. Sequencing chickpea genomes: genotyping and high-quality reference genomes for association studies and breeding
  7. Towards the characterization of adaptive genetic differentiation in wild chickpea
  8. Phenotyping of chickpea (Cicer arietinum L.) germplasm using agro-morphological and phenological traits to screen heat stress tolerant genotypes
  9. Assessment of Genetic Diversity in a Collection of Wild Chickpea Populations in Turkey
  10. Occurrence and distribution of Chickpea Fusarium wilt in Ethiopia
  11. Phylogenetic and Genetic characterization of root nodule bacteria isolated from major growing areas of Ethiopia.
  12. The potential of persistently green-seeded chickpea for an under-utilized fresh vegetable market and production value chain that is inclusive of small-holder farmers.
  13. Developing methodology to evaluate chickpeas for resistance to root-lesion nematode Pratylenchus thornei.
  14. Evaluation of Wild Relatives of Chickpea for Resistance to Pod Borer, Helicoverpa armigera
  15. Identification of DNA markers through Genome Wide Association Studies (GWAS) and in wild (Cicer reticulatum L.) and cultivated chickpea (Cicer arietinum L.)
  16. Genetic Resources for Resistance to Ascochyta Blight of Chickpea
  17. Enhancing Food Security through Improved Productivity, Nutrition and Marketing of Chickpea in Central and Western Ethiopia
  18. Assessing chickpea’s wild relatives for resistance to root-lesion nematodes
  19. Effect of nitrogen availability on chickpea leaf and root morphology
  20. Exploring Genetic Diversity in Landrace Chickpea Accessions in the Secondary Centers of Diversity and in the Fertile Crescent

Expanding Chickpea in non-traditional Lowlands of Ethiopia

Abiy Addisu1, Desta Gibre1, Fisseha Tadesse2, Negussie Tadesse3, Kedir Ebise2, Seid Ahmed3 and Zewdie Bishaw3

1Ethiopian Institute of Agricultural Research; 2Oromiya Agricultural Research Institute; 3International Agricultural Research Institute, Addis Ababa, Ethiopia

Abstract
Frequent drought is affecting the mixed farming and agro-pastoral communities in Ethiopia. In the Agro-pastoral lowlands, maize, sorghum and common bean are the key crops produced by famers when rainfall is favorable. However, during severe drought or erratic rainfall patterns, these crops do not yield enough grain to feed the communities, generate incomes and feed to their livestock. In order to diversify the cropping system in two in irrigated and rainfed lowlands, non-traditional crops (wheat, barley, chickpea, lentil, and cowpea) were evaluated by farmers in 2014/15 cropping season. The season was very dry in the rainfed lowlands, and all crops failed to produce grain for the farming communities. In both rain fed and irrigated sites, the kabuli chickpea cv.
Habru was selected by farmers. The average seed yield of chickpea in the rain fed low land was more than 1.5t/ha compared to total failure of maize and other tradition crops grown in the area during severe drought. The introduction of high yielding and Ascochyta blight resistant chickpea in the rainfed lowlands is one of the options to diversify the farming system and prevent complete crop losses due to drought and erratic rainfall patterns. Besides producing acceptable yield, chickpea and cowpea were not attacked by termite which is a key pest in the low lands. In the irrigated areas, the yield was higher than 2t/ha and can be used to improve land productivity in the cotton-fallow cropping system. In conclusion, adoption of new crops like chickpea to diversify farming system can improve the resilience of farmers in dry areas. The breeding programs of EIAR and ICARDA should further develop chickpea cultivars with high yield (seed and biomass),large seeded resistant to foliar diseases and responsive to supplementary irrigation. 


Developing a Resource of Wild x Cultivated Introgression Lines for Crop Improvement of Chickpea

Lijalem K. Balcha1, Dagnachew B. Besha1, Kassaye N. Dinegde2, Donna Lindsay3, Reyaz Mir Rouf4, Bullo Mamo5, Syed Gul A.S. Sani5, Lisa Vance5, Emily Bergmann5, Bunyamin Tar'an3, Asnake Fikre1, R. Varma Penmesta5, Douglas R. Cook5

1 Ethiopian Institute of Agricultural Research (EIAR), Debre Zeit Agricultural Research Center, Debre Zeit, ETHIOPIA.
2 Ethiopian Institute of Agricultural Research (EIAR), Melkasa Agricultural Research Center, Nazareth, ETHIOPIA.
3 University of Saskatchewan, Department of Plant Sciences, Saskatoon, CANADA.
4 Sher-E-Kashmir University of Science & Technology-Jammu, Division of Plant Breeding and Genetics, INDIA.
5 University of California at Davis, Department of Plant Pathology, Davis, USA.

Abstract

The use of crop wild relatives (CWRs) has been a component of cultivar improvement programs since 1920s and 1930s, after Vavilov recognized their value as a source of increased variation. Reduced genetic diversity in elite varieties of modern crop plants derives from a combination of an early domestication bottleneck and subsequent focus on fewer improved genotypes during modern breeding. As a result, the prospects for sustainable genetic gain from elite germplasm is increasingly limited. By contrast, the wild ancestors of crop plants typically possess high levels of genetic diversity and an expanded range of adaptive traits that may be of agricultural relevance. CWRs have been used in the improvement of several crop species, but rarely in a broad and systematic manner. Chickpea (Cicer arientinum L.) is one of the most valuable global crops and suffers from a lack of genetic diversity. With the goal of increasing genetic diversity in chickpea, we initiated a novel and systematic introgression from wild Cicer species (C. reticulatum and C. echinospermum) into cultivated elite germplasm. The approach combines (1) systematic survey of wild diversity, (2) introgression of a representative set of genotypes, and (3) marker-assisted normalization of phenology among segregating progeny. Twenty diverse wild founders of C. reticulatum were selected from 270 wild accessions based on a combination their genomic sequence information and the ecology of their origin sites. Each of the 20 founders was crossed to elite cultivars of India (ICCV-96029), Ethiopia (Habru and Minjar), Canada (CDC Leader and CDC Consul) and Turkey (Gokce). At the F2 stage, a subset of progeny within each lineage were intercrossed to increase chromosomal recombination and thus genetic power in the resulting populations. Early generation segregant populations (e.g., F3 and F4) will be phenotyped for traits at multiple sites in Ethiopia and India beginning with the fall 2016/winter 2017 crop cycle. This resource of introgressed populations will serve as a platform for gene discovery through genomics, modeling and phenotyping, and for the development of tools for marker-assisted selection for breeding. Here we report the status and progress of chickpea population development in the USAID Feed the Future Chickpea Innovation Lab, with a particular focus on populations with elite cultivars from Ethiopia and India. 


Transpiration rate of chickpea wild accessions and cultivars in Turkish and Indian locations

Fatma Başdemir1, Bekir Bükün1, Mehmet Yıldırım1, Tuba Biçer1, Douglas R. Cook2, Vincent Vadez3, Abdullah Kahraman4, Jens Berger5

1 University of Dicle, Diyarbakır, TURKEY. 2 University of California, Davis, USA.
3 ICRISAT, INDIA.
4 Harran University, Sanliurfa, TURKEY

5 CSIRO Plant Industry, AU.

To phenotype chickpea wild accessions and cultivars for drought tolerance, as a first year research of a Global Crop Diversity Trust project, two experiments were conducted in series at both Dicle University in Turkey and at ICRISAT in India.

Transpiration rate and fraction of transpirable soil water (FTSW) measurements were analyzed at Dicle University from February 2016 until late May 2016. Full assessment of FTSW was not yet complete when this was poster prepared. 26 wild chickpea accessions multiplied at Harran University and 4 commercial cultivars were evaluated in accordance with project aims. Leaf transpiration rates under naturally changing vapor pressure deficits (VPD) were determined by weighing pots at one our intervals, from morning (09:00 am) to late afternoon (04:00 pm) under greenhouse conditions. Temperature and relative humidity were recorded using a data logger to determine atmospheric VPD. Among genotypes “Gökçe (check)”, “Derei-072” and “CudiB-022C” had the lowest transpiration rates suggesting their higher relative drought tolerance. By contrast, “Deste-080” and “Sirnak-060” had the highest recorded transpiration rates.

Similar experiments were conducted with 10 genotypes by Fatma Basdemir during a brief internship in the laboratory of Vincent Vadez at ICRISAT. Transpiration rates gradually increased under rising VPD conditions in all tested genotypes. Relatively tolerant genotypes possessed the lowest transpiration rates at low VPD and continued this trend throughout increasing VPD.

Key words: Transpiration rate, wild chickpea, drought tolerance 


Status of genomic assisted resistance breeding for the development of Fusarium wilt (Fusarium oxysporum fsp.ciceris) resistant Chickpea (Cicer arietinum L.) genotypes in Ethiopia

Dagnachew Bekele.1,2, Kassahun Tesfaye.2, Douglas R. Cook.3, and Asnake Fikre.1

Ethiopian Institute of Agricultural Research, Addis Ababa, Ethiopia1.Addis Ababa University, Institute of Biotechnology, Addis Ababa, Ethiopia2.University of California, Department of Plant Pathology, Davis, USA3.

Abstract

Although the yield potential of present day chickpea cultivars exceeds 5.0t/ha, in Ethiopia the average national yield is below 2t/ha.The wide gap between average yield and potential yield is mainly due to different biotic and abiotic stresses at various crop growth and development stages, leading to low productivity. Recent advances in next generation sequencing and genotyping technologies have enabled generation of significant genomic resources that can support the development ofbiotic and abiotic stresses tolerant genotypes in chickpea. Fusarium wilt caused by soil borne fungus, Fusarium oxysporum f.sp. ciceris (FOC), is the most devastating disease of chickpea in Ethiopia which needs very efficient and robust genomic tools to generate resistant chickpea genotypes. However, only little works have been done on chickpea Fusarium wilt, and limited information is available to generate resistant genotypes in Ethiopia. Tackling such challenge has got the top priority in Ethiopian chickpea improvement program. From 217 farmers fields, samples of diseased plants, seeds and soils were collected and isolated. A total of 209 Fusarium oxysporum isolates were identified based on their morphological characteristics such as abundance of micro and macro conidia, short and unbranched monophialides, white to creamy white colony color on PDA medium. These isolates will be genotypes using a genotyping by sequencing method (RAD-GBS), and the SNP data will be used to calculate standard population genetic and diversity metrics to describe the diversity of FOC in Ethiopia. Besides, based on thepathogenicity test and efficient phenotyping of reverse introgressed lines(reVILs) and advanced backcross introgression lines(ABIs) segregating populations, FOC resistance genes in chickpea will be mapped. Subsequently, marker assisted backcrossing will be used for the introgression of Fusarium Wilt resistant gene into adapted Ethiopian chickpea cultivars.


Effect of inoculation of chickpea and lentil with stress tolerant rhizobia of Morocco.

I. BENJELLOUN1,2, I. THAMI ALAMI1, M. EL KHADIR2, A. DOUIRA2, A. GREENSPAN3, D. R. COOK3, S. M. UDUPA4*

1 Institut Nationale de la Recherche Agronomique (INRA), B.P.415, Rabat, Morocco
2 Laboratoire de Botanique, Biotechnologie et de Protection des Plantes, Département de Biologie, Faculté des Sciences de Kenitra, Université Ibn Tofayl, BP 242, Kénitra, Morocco. 3 University of California, Department of Plant Pathology, 354 Hutchison Hall, One Shields Ave, Davis, CA 95616-8680, USA
4 ICARDA-INRA Cooperative Research Project, International Center for Agricultural Research in the Dry Areas (ICARDA), B.P. 6299, Rabat, Morocco

* presenting author

Abstract

Chickpea and lentil are important pulse crops grown mainly in arid and semi-arid regions where numerous environmental factors including drought, salinity, extreme temperatures and soil pH may dramatically affect nodulation, nitrogen fixation and biomass production. Thus the yield remains very low despite an increasing demand for food. To enhance the productivity and limit the use of chemical fertilizers, it is necessary to support the use of biofertilizers based on stress-tolerant microorganisms for an economic and ecologically- sustainable agriculture.

The aim of this study is to characterize populations of rhizobia nodulating lentil and chickpea in order to select the most tolerant and effective ones for use as inocula by farmers. To achieve this objective 207 and 206 isolates were purified from chickpea and lentil nodules respectively. Isolates were characterized using rep-PCR which showed the presence of 23 groups among lentil rhizobia and 40 groups for chickpea rhizobia. Examination for their tolerance to environmental stresses (pH, salinity, water stress, and extreme temperatures) revealed large phenotypic variability for these conditions. Two highly efficient and tolerant strains were selected for each crop for a field inoculation test. The experiment was carried out at two sites in Morocco, Merchouch (clay-silty) and Ain Sbit ( clay-sandy) in a Randomized Complete Block Design (RCBD) design, with four treatments: Control (no inoculation and no N application), S1 (inoculation with strain 1) and S2 (inoculation with strain 2) and N fertilization (N 120kg/ha), each with four replications.

Results showed that inoculation with the selected strains significantly increased nodules number for chickpea and lentil especially in Merchouch trial. Effect of inoculation on shoot dry weight and nitrogen content at both sites was statistically similar to the effect of nitrogen application. Grain and straw yield were significantly enhanced compared to the control indicating that the used strains were more competitive and effective than native ones except for chickpea in Ain Sbit where the yield was enhanced but not significantly.

Used strains were efficient in both sites with different degree of significance. This suggests that nodulation, nitrogen fixation and grain yield of chickpea and lentil can be improved by inoculation using competitive stress tolerant rhizobia as an inexpensive way to increase the productivity of these crops in arid and semi-arid areas for sustainable agriculture. 


Sequencing chickpea genomes: genotyping and high-quality reference genomes for association studies and breeding.

Peter Chang1, Vasantika Suryawanshi2, Matilde Cordeiro2, Noelia Carrasquilla-Garcia1, Wendy Vu2, Sripada Udupa3, Eric vonWettberg4, R. Varma Penmetsa1, Sergey Nuzhdin2, Douglas R. Cook1

1 University of California, Davis, USA
2 University of Southern California, Los Angeles, USA
3 International Center for Agricultural Research in the Dry Areas (ICARDA), MOROCCO. 4 Florida International University, Miami, USA

Abstract:

Natural populations of species closely related to cultivated chickpea (Cicer arietinum) were identified and collected systematically using ecological principles from their native range in southeastern Turkey. Collection was focused primarily on C. reticulatum (wild progenitor of the cultigen) and C. echinospermum, a sister species, and with limited co-incident collection of the more distantly related species C. bijugum and C. pinnatifidum.

Over 1,000 wild individuals sampled in 2013 were genotyped using Restriction- enzyme Associated DNA Genotyping By Sequencing (RAD-GBS). Based on this sequencing, allele-frequency based population assignment was conducted for all genotypes leading to the choice of the focal genotypes as donor parents for introgression population development.

Both the currently available reference genomes for cultivated chickpea are draft assemblies containing several ambiguous regions, and whole genome assemblies of wild relatives are currently unavailable. To address these limitations, focal genotypes for high- resolution reference genomes for each of the three species were selected based on: i) their use in introgression population development; ii) genetic relationships to genotypes from other sites of the same species; and iii) likelyhood of long-term stability of the collection site for potential future in-situ studies. Genotypes CDCFrontier (C.ari), Besev_079 (C. ret) and S2Drd_065 (C. ech) represent the three species. For each genotype, sequence data from ~60x short-read Illumina and ~30x long-read PacBio are being integrated with BioNano optical mapping data. Assemblies will be assessed via high-density linkage mapping (RAD-GBS) of early generation progenies derived from wild x wild and wild x cultivated crosses.

In addition, 26 wild accessions that represent ecological and molecular variation within the species and serving as potential introgression donors and recipient cultivars were sequenced to ~30x coverage via Illumina short read sequencing, data that allow for analysis of genome-wide signatures of selection and for trait-gene associations. To identify rare alleles among populations and to calculate linkage disequilibrium, and association studies with native site ecological parameters, ~200 genotypes from the same populations were sequenced to medium depth (~10x) via Illumina short-read sequencing.

Together these genome data represent a novel resource for chickpea biology, to improve the accuracy and precision of association mapping, trait-marker discovery and introgression breeding. 


Towards the characterization of adaptive genetic differentiation in wild chickpea

Matilde A. Cordeiro, Vasantika Suryawanshi, Peter L. Chang, Sergey V. Nuzhdin

Low genetic diversity within domesticated chickpea (Cicer arietinum) is reflected on the extremely reduced adaptive potential of the available germplasm. Wild populations of the closely related species C. reticulatum and C. echinospermum were identified and collected along their natural range to help identify novel adaptations.

Local adaptation is one possible result of evolution where genotypes and genes that perform better in their home environment compared to a foreign environment. For local adaptation to evolve, selection must favor different traits in different environments and populations must contain significant standing genetic variation. Demographic factors, such as genetic drift or gene flow, may outpace the strength of selection and hinder local adaptation. Finding relevant genetic variation in natural populations is an ongoing challenge in evolutionary ecology and is key for the identification of useful genetic differentiation in wild chickpea for introgression in cultivated germplasm. Understanding the genetic signatures of selection and demographic processes in populations has been the object of experimental and theoretical studies over the last decades. Quantifying genetic, phenotypic and eco-geographic differentiation and their correlations is key to depict relevant variation that may broaden chickpea agricultural ranges.

A Bayesian approach was used to model pairwise genetic differentiation based on geography and ecology, calculating the relative contribution of geographic and ecological pairwise distances to the covariance of allele frequencies (BEDASSLE; Bradburd et al. 2013). Geological coordinates and altitude data were used to calculate the geographic and ecological distance matrixes, respectively, and a total of 6466 independent loci covered in 21 C. reticulatum and C. echinospermum populations were used to generate the allele frequency matrix. Elevation is more correlated with genome-wide signatures than geographic distance, such that 1 m of elevation is equivalent to 72.4 km of geographical distance (αE:αD 0.011382) when looking at both species, and 1 m of elevation is comparable to 11.2 km of horizontal distance effects genome-wide on C. reticulatum only (αE:αD 0.08933). The observed differences when using different species are consistent with observations that elevation differences are frequently correlated with environmental and soil type differences. It seems like lower genetic diversity between C. reticulatum populations allows for a better resolution of genetic signals and therefore improves depicting altitude from other factors.

To identify the genetic basis of adaptation and enable the distinction of selection from demography, deeper sequencing is required. Identifying such genetic signatures relies on the association of patterns in the DNA sequence that differ from the expected by neutrality, such as Tajima’s D and other variations, that may be used to test for polymorphism frequency variation in a target sequence and potentially infer distinct signals of selection. Other tests rely on the evolution of coding sequences, where the ratio between synonymous and non-synonymous substitutions within and among species is calculated and compared with the candidate allele variations to identify deviations from neutrality. Therefore, identifying less frequent alleles and detecting the relationships between loci are key to further our understanding of the genetic basis of adaptation. Whole genome medium coverage data (10x) was generated for ~200 genotypes from the 21 wild populations. Such data will enable the combination of ecological and genomic methods to characterize adaptive genetic differentiation in wild chickpea. 


Phenotyping of chickpea (Cicer arietinum L.) germplasm using agro-morphological and phenological traits to screen heat stress tolerant genotypes

Tsegaye Getahun1, Masresha Fetene1, Kassahun Tesfaye1, Asnake Fikre2, R. Varma Penmesta3, Douglas R. Cook3

1. Addis Ababa University, Ethiopia, 2. Ethiopian Institute of Agricultural Research, Ethiopia, 3. University of California Davis, USA

Abstract
Heat stress (
30oC) affect plant growth and development and may lead to a drastic reduction in economic yield from 6-30%. The reproductive phase, flowering and podding, is highly vulnerable to heat stress. Thus, developing heat stress tolerant genotypes are very important. One hundred twenty chickpea germplasm, 80 landraces and 40 advanced cross lines, have been screening at three locations namely Kobo, Werer and Debre Zeit. Local and standard checks were sown at each site. Kobo and Werer are heat stressed areas. Debre Zeit is the normal growing environment. The germplasm were sown using alpha lattice design in three replications at each location. Each plot is 1m by 0.6m and contains 14 seeds for each and 10 cm distance between each plant. The primary traits that have been phenotyped are plant height, pod filling duration, physiological maturity, first days of flowering, 50% days of flowering, first days of podding, number of pods per plant, plant biomass, grain yield/ha, harvest index and 100 gm weight. All genotypes are germinated at all locations. Early flowering genotypes (38-40 days) were found in Kobo. The future plan is to study the mechanisms of heat stress tolerance (physiological, biochemical and molecular mechanisms) for the selected heat stress tolerant and sensitive genotypes. The genetic basis (QTLs and associated molecular markers) for heat stress tolerance will be determined using GWAS. Lastly, breeding for heat stress tolerance will be conducted. Crossing of at least four elite genotypes with four heat stress tolerant genotypes will be carried out. They will be developed till the F2 generations and MAS will be carried for the segregating generations. Multi-location trial will be carried out for heat tolerant and high yielder genotypes. 


Assessment of Genetic Diversity in a Collection of Wild Chickpea Populations in Turkey

Abdullah Kahraman1*, Anamika Pandey1, Mohd. Kamran Khan1, Ahmet Cakmak1, Bilal Aydin1, Jens Berger2, Mahmut Gayberi3

1Harran University, Faculty of Agriculture, Department of Field Crops, Sanliurfa, Turkey 2CSIRO Plant Industry, Wembley, WA 6913, Australia
3GAP Agriculture Research Institute (GAPTAEM), Turkey

* corresponding author- kahraman@harran.edu.tr

Chickpea is considered to have been domesticated from its wild progenitor, Cicer reticulatum around 11,000 years ago in central Fertile Crescent that presently belongs to South Eastern part of Turkey and Syria. During the year 2013 and 2014, we have newly collected approximately 502 wild chickpea accessions belonging to 31 distinct populations and planted them under plastic house conditions. These accessions were evaluated for different agro-morphological traits; here we present the variation in days to flowering, days to maturity and seed weight per plant of the germplasm grown in plastic house. Statistical analysis revealed significant variation among the collected populations. In principal component analysis, first two components explained 96% of the morphological variation. Phylogenetic analysis based on three traits revealed two main groups and four sub-groups amongst 31 populations. Coefficient of variation for days to flowering, days to maturity and seed weight per plant was in the range of 1.1-7.7, 0.9-29.3 and 13.8-60.2, respectively. Variability in the studied traits suggests the potential exploitation of these wild germplasm resources in future breeding programs for the genetic improvement of cultivated varieties. Association of these traits with molecular markers can be of much significance for further genetic studies. 


Occurrence and distribution of Chickpea Fusarium wilt in Ethiopia

Sultan Mohammed1 Chemeda Fininsa2, Brendan K.Reily3, Noelia Carrasquilla-Garcia3, Alex

Greenspan3, Negussie Tadese4, Seid Ahmed4, Aladdin Hamwieh4, Douglas Cook3

1. Woldia University, Ethiopia. 2.Haromaya University, Ethiopia, 3. University of California Davis, USA. 4. International Center for Agricultural Research in the Dry Areas (ICARDA).

Abstract
Fusarium wilt caused by Fusarium oxysporum f. sp. ciceris (Foc) is the most serious disease of chickpea throughout the world, causing 100% loss under conditions favorable to the pathogen. Foc is both soil and seed borne and difficult to eradicate from infested fields because fungal chlamydospores survive in soils for many years even in the absence of the host plant. Variation in pathogen populations has been previously inferred based on differential host responses, but such inferences are indirect. To better understand geographic patterns of pathogen virulence and genomic variation of Foc in Ethiopia, a GPS-based survey was conducted in the 2014/15 cropping season. Fusaria were isolated from symptomatic plants in a total of 156 fields, representing 5 regions that encompass 30 major chickpea producing districts. Pure strains were derived from single spore isolation and purified strains were assessed for virulence on susceptible chickpea genotypes. For purposes of genomic analysis, various methods of DNA extraction were tested, resulting an a hybrid approach involving Qiagen kit DNA extraction followed by a single round of isothermal whole genome amplification. Whole genome sequencing libraries were constructed to compare the efficacy of mechanical versus enzymatic shearing of DNA. An initial sequencing and assembly result for these test genomes is presented. At the time of specimen collection, the incidence and severity of chickpea wilt were recorded in each field, revealing differences in rates of disease among the 5 primary geographic regions of chickpea cultivation. Disease incidence (38.28 ± 13.99) and severity (6.01 ± 1.31) were greatest in Gojam, and lowest in Shewa (Disease incidence = 7.95 ± 2.18; severity = 1.64 ± 0.42). More over, fields among districts were divided in to 5 categories based on disease severity. The categories will be used to test the hypothesis that certain Foc genotypes are correlated with severity of Fusarium wilt. High disease incidence likely derives from a combination of the use of non-certified sources of local desi chickpea genotypes and poor agronomic practices adopted by the farmers. Based on a combination of pathogenic and genetic variability of the causal organisms, and the agro- ecology of collection sites, we plan to select a subset of Fusarium isolates to begin systematic testing of popular elite genotypes of chickpea, with the eventual goal of breeding for resistance to the diversity of Foc present in Ethiopia. 


Phylogenetic and Genetic characterization of root nodule bacteria isolated from major growing areas of Ethiopia.

Zehara Mohammed, Alex Greenspan, Douglas Cook, Fassil Assefa and Asnake Fikre

Abstract

Nitrogen is a plant nutrient commonly a major limiting factor for crop production. The use microbial process that converts atmospheric nitrogen into a plant-usable form is the major mechanism of supplying N in those soils lacking native effective rhizobia and nitrogen fixation ability is widely distributed among phylogenetically diverse bacteria. Thus this study investigates the diversity between strains and phylogenetic relationships through multi-locus sequence approaches (16S rRNA, atp D, dnaJ, glnA, gyrB and rec A) genes. Nodules were collected from 140 farmers’ fields at Ethiopia with soil, seed and questioner survey. 106 isolates were screened and genomic DNA extracted; Polymerase chain reaction (PCR) on two genes (16S rRNA and nodC) performed; correspondingly 80 isolates were genomic analyzed; nine major phylogenetic groups of Mesorhizobium were found. Most of the strains clustered with the M. plurifarium within different species category. The rest of the strains distributed in six new clades: M.sp1& 2, M.gobiense, M.ciceri, M.opportunistum, Rhizobium, one unidentified and Tigray collection will be included.  Greenhouse activity at Ethiopia DZARC is underway with treatment component 20 Ethiopian indigenous strain from present work, one imported Tunisian PCH, one Menagesha Biotech CP 029, 0.05% KNO3 (w/v) as positive control and no inoculant as negative control on two chickpea genotypes (Natoli and Arerti) and non-nodulating Chickpea isolines cultivar PM233; followed by field evaluation on 2016 on four indigenous elite strain well performed at greenhouse, Menagesha CP 029, Imported Tunisian PCH, N fertilizer as positive control and no inoculant as negative control will be conducted on two chickpea genotype. This research is useful for improving root nodulation and yields to benefit the subsistence farmers and a baseline to Ethiopian chickpea Mesorhizobium genetic resources.


The potential of persistently green-seeded chickpea for an under-utilized fresh vegetable market and production value chain that is inclusive of small-holder farmers.

R. Varma Penmetsa1, Jana Kholova2, Reyazul Rouf Mir3, Gul Sani1, Kassaye Negash1, Mohammed Rezaei4, Srikanth Mallayee2, Irshad Ahmed3, Noelia Carrasquilla-Garcia1, Peter Chang1, Matilde Cordeiro M5, Eric vonWettberg6, Bunyamin Ta’ran4,Vincent Vadez2, Douglas Cook1

1 University of California, Davis, USA; 2 International Crops Research Institute for the Semi-Arid Tropics, Patancheru, India; 3 Sher-e-Kashmir University of Agricultural Sciences & Technology, Jammu, India; 4 University of Saskatchewan, Saskatoon, Canada; 5 University of Southern California, Los Angeles, USA; 6 Florida International University, Miami, USA

ABSTRACT:
A rare morpho-form of chickpea has green cotyledons and seed coats. Mature, dry seed of such

green-seeded chickpea have been shown to have elevated beta-carotene, to levels that are similar to the first generation of golden rice [Ashokkumar et al (2014) Crop Sc., 54: 2225-2235]. In ongoing work, developing immature, fresh vegetable form of the green-seeded type were also found to have elevated nutrient levels compared to regular tan color chickpea.

Of ~30 green-seeded lines available in genebanks, most also exhibit a delayed-age senescence phenotype wherein many plant organs including leaves and pods exhibit delayed de-greening, reminiscent of 'stay-green' mutants characterized in a range of other crops and plants.

Using a candidate gene approach we identified molecular variation in the chickpea ortholog of the staygreen protein, we designate as CaStGR1, in 25 of 27 green-seeded lines. Sequence variation comprised of changes that are predicted to result in loss-of-function of CaStGR1 in the green-seeded germplasm, and include a variant wherein the entire CaStGR1 gene is absent. Skim whole genome sequencing validated this apparent whole gene deletion and to obtain deletion- spanning amplicons. A molecular lesion in this candidate gene co-segregates with the seed color phenotype in an F2 population.

A representative subgroup of germplasm lines for the four StGR1 alleles were evaluated for physiological traits wherein StGR1 loss-of-function lines were similar to wild type chickpea for plant growth, development and responses to atmospheric and soil water deficit.

Marker-assisted backcrossing of CaStGR1 loss-of-function alleles has been initiated into cultivars of Ethiopia, India and Pakistan, material that will allow for rigorous examination of the effects of the staygreen gene's loss-of-function on phenotypes, physiology, and agronomic traits in common genetic backgrounds, and to advance introgression of this potentially useful trait into farmer-preferred cultivars of chickpea. Efforts to ascertain market demand for fresh market chickpea per se, and with StGR1 introgressions are being explored, as the parallel and necessary prelude to deployment and uptake of this promising genetic technology for biofortification.

The delayed de-greening of pods of the green-seeded genotypes, and the higher nutrient levels of this morpho-form suggest a novel niche market for an improved fresh market chickpea. Lengthening the shelf-life of vegetable chickpea from incorporating the ‘stay green’ trait would benefit a range of value chain participants, including small-scale farmers, local processors, and retailers that typify the fresh market chickpea production value chain, particularly in the developing world. Greater access to a fresh chickpea type would broaden options for consumers and could encourage consumption of a more sustainable plant-based protein diet.  


Developing methodology to evaluate chickpeas for resistance to root-lesion nematode Pratylenchus thornei.

Roslyn Reen, Rebecca Zwart, John Thompson Centre for Crop Health, University of Southern Queensland Toowoomba Australia. Email:Roslyn.reen@usq.edu.au

Chickpea (Cicer arietinum) is a major pulse crop grown worldwide and is susceptible to the root-lesion nematode (RLN) Pratylenchus thornei. The impact of RLN can be up to up to 20% yield loss of intolerant Australian cultivars. Globally Pratylenchus rank second to Meloidogyne as the most devastating nematodes constraining chickpea crop production. The aim of our study was to optimise methods for screening chickpea for RLN resistance. Three experiments using chickpea cultivars with a range of RLN susceptibility were conducted under controlled glasshouse conditions. Experiments assessed the effects on P. thornei reproduction by (1) slow-release granules and solution based fertilisers with and without rhizobium, (2) the effect of the fungicide seed dressing P- Pickel T, and (3) a timeline experiment to determine optimum harvest time for discriminating between resistant and susceptible cultivars. Results for the fertiliser x rhizobium experiment showed significant interactions for cultivar x fertiliser (P=0.01) and rhizobium x fertiliser (P=0.027). Overall solution based fertiliser in combination with rhizobium gave the highest nematode numbers in susceptible cultivars. The P-Pickel T treatment had no significant effect on nematode reproduction. Optimum harvest time after inoculation was 18 and 20 weeks giving the highest numbers of P. thornei for the susceptible cultivars Kyabra and Sona. However, 18 weeks gave better discrimination between susceptible cultivars and the resistant Cicer echinospermum ILWC 246, which confirms earlier research. Based on these results current experiments will implement seed dressing, a solution- based fertiliser with rhizobium and harvest at 18 weeks for assessing chickpea accessions for resistance to RLN. Two hundred new wild Cicer accessions from Turkey will be screened for RLN resistance using this methodology in order to select superior genotypes for breeding new elite resistant cultivars. 


Evaluation of Wild Relatives of Chickpea for Resistance to Pod Borer, Helicoverpa armigera

Gashaw Sefera1, 2, Hussien Mohammed2, Douglas R Cook3 and Hari C Sharma*1

1International Crops Research Institute for Semi Arid Tropics (ICRISAT), Patancheru 502314, Telangana, India. 1 Hawassa University, Hawassa, Ethiopia. 3Univerity of California, Davis, USA. *Email: h.sharma@cgiar.org

Abstract

Chickpea is an important legume crop in semi-arid tropics and warm temperate zones. Its production is constrained by different biotic and aboitic factor among which Helicoverpa armigera is the most damaging insect pest worldwide. Having limitations of resistance to this pest in cultivated chickpea, we evaluated 26 wild accessions of chickpea along with 4 cultivated checks with the objective to identify germplasms with better resistance to H. armigera resistance in the wild chickpeas using detached leaf assay and HPLC analysis to estimate the concentrations of organic acids found in chickpea leaf exudates. Based on detached leaf assay during vegetative and flowering stage, there was considerable variation among wild relatives of chickpea for damage rating, larval survival, larval weight and host suitability index. Damage rating ranged from 2.2 up to 6 and 2.2 up to 6.8 during vegetative and reproductive stages respectively. Larval survival also varied from 44 – 94% and 64 – 96% while larval weight varied 0.49 – 2.75 mg during vegetative and 0.74 – 2.39 mg during reproductive stages. The test materials also showed variation for host suitability index from 0.95 – 5.8 during both growth stages of the plant. Oxalic acid was detected in all test materials ranging from 0.95mg/g up to 19.19 mg/g on dry weight basis while malic acid and acetic acid were found only in few wild accessions. EC 868753, EC 868765, EC 868768, EC 868769 from the wild accessions and ICC 506EB from checks showed better resistance during both or one of the growth stages. The resistance expressed in 506EB is correlated to high amount of oxalic while it was other factors in the remaining wild accessions. 


Identification of DNA markers through Genome Wide Association Studies (GWAS) and in wild (Cicer reticulatum L.) and cultivated chickpea (Cicer arietinum L.)

Bahattin Tanyolac, Ege Unversity, Turkey

Abstract

Understanding genetic relationships is important in utilization of wild and cultivated chichpea (Cicer ssp.) in breeding programs, due to high content of carbohydrates and protein than other pulses. Chickpea is a good source of important vitamins and essential amino acids. Turkey is one of the important chickpea producers in the World with 535 M tons value and ranking 3rd place. We genotyped 100 wild and 100 cultivated chickpea accessions using genotypig by sequencing which can represent the general profile of high the genetic diversity in the Turkey. Nutrients are essential elements needed in small amounts for adequate human nutrition and include the elements Fe, Mg, K, Ca, Zn, P, Cu, Mn. The elements concentration in seed was determined using the method of Kacar (1972). The total Fe, Mg, K, Ca, Zn, P, Cu, Mn concentration of the prepared extract was measured by atomic absorption spectrophotometry (AAS) (Varian, SpectrAA 220/ FS, CA, USA) (Kacar 1972; Kacar and Inal 2008).

Genotyping by sequencing (GBS), a new low-cost, high-throughput sequencing technology was used to genotype chickpea accessions. A total of 312,000 SNP markers were detected in GBS analysis were sequenced on Illumina Hiseq 2000 platform. After filtering raw and redundant data, a total of 121,622 SNPs were obtained for population structure and association mapping. For determining population structure using the STRUCTURE, populations K=2 to K=10 were tested and divided into 4 populations (POPI, POPII, POPIII, POPIV. The results indicated that there were obvious genetic variations between accessions of wild and cultivated chickpea.

Genome-wide association studies (GWAS) offer high resolution through historical recombination accumulated in natural populations. GWAS analyses were performed for the elements Fe, Mg, K, Ca, Zn, P, Cu, Mn and SNPs in TASSEL 5.0.5. Three different approaches (GLM, MLM+K and MLM+K+Q) were used to control for false-positive results in association tests and compared for their capacity to fit the data. Using a GLM, we identified 7 SNPs that were significantly associated, and a MLM+K revealed 65 SNPs associated while MLM+K+Q revealed 6 SNPs associated with phenotypes. Three models detected 13 significant markers distributed on 3 of the eight. Phenotypes most associated were Mg, Fe and K with associated SNPs. Our findings could contribute to more effective screening of elite germplasm to find resistance alleles for marker-assisted selection in breeding programs. 


Genetic Resources for Resistance to Ascochyta Blight of Chickpea

M. Tekin, D. Sari, N. Cherigui, M. Talip, C. Toker*
Akdeniz University, Faculty of Agriculture, Department of Field Crops, Antalya, Turkey * E-mail
: toker@akdeniz.edu.tr

Ascochyta blight is one of the most important foliar diseases of chickpea (Cicer arietinum L.) worldwide and may cause complete yield losses under cool and humid weather conditions. The pathogen of ascochyta blight of chickpea has both an anamorph [Ascochyta rabiei (Pass.) Labr. syn. Phoma rabiei (Pass.) Labr] and a teleomorph [Didymella rabiei Kovachevski syn. Mycosphaerella rabiei Kovachevski]. The fungus shows a high pathogenic and genetic variability and a number of pathotypes of fungus were reported by molecular markers; pathotype I (less aggressive), pathotype II (aggressive), pathotype III (most aggressive) and new A. rabiei pathotype (pathotype IV), respectively. Although cultural and chemical methods is available to control with the disease, they are not effective. The use of resistant plants seems to the best way to struggle with ascochyta blight. From this point, screening of germplasm and selection of resistant plants are very important for selection of resistant resources in breeding programs. There are approximately 75 genetic materials of cultivated chickpea known to be resistant to ascochyta blight in gene banks. In addition to these, wild genetic resources including C. bijugum, C. chorassanicum, C. cuneatum, C. echinospermum, C. judaicum, C. pinnatifidum, C. reticulatum, C. yamashitae, C. canariense of chickpea have been reported for resistance to disease until now. However, it could not be benefited from wild resistance resources because of hybiridization barriers except for C. reticulatum and C. echinospermum. In this time, using of molecular markers reduces the time consumed with field screening in breeding programs. Some QTL markers (GA16, TS82, TA194, TR58, GAA47, SCY17, TA130 and TA2) known to provide resistance to the disease have been reported. This study aims to review current knowledge about ascochyta blight of chickpea and use of resistant genetic resources. 


Enhancing Food Security through Improved Productivity, Nutrition and Marketing of Chickpea in Central and Western Ethiopia

Kassahun Tesfaye (PI) 1, 2, Teklehaimanot H/Selassie 1, Fasil Aseffa 2, Paulos Asrat 3, Douglas Cook4

1Institute of Biotechnology, AAU, Addis Ababa, Ethiopia
2Department of Microbial, Cellular and Molecular Biology, AAU, Addis Ababa, Ethiopia 3 College of Development Studies, AAU, Addis Ababa, Ethiopia
4University of California, Davis, USA
(PI Email:- kassahuntesfaye@yahoocom)

Summary

Chickpea is the world’s second most widely grown pulse crop and a major source of human protein nutrition. Forty percent of Africa’s chickpea crop is grown in Ethiopia, where cultivated acreage has increased from 140,244 ha to 229,720 ha during the past decade. However, Ethiopia’s chickpea yield remains low (1.8 t/ha; CSA 2006, 2014), well below its yield potential. This yield gap derives from shortage of improved technology and limited availability of high-yielding and stress tolerant varieties. Among the constraints to chickpea production is its sensitivity to Aluminum toxicity, which is a defining feature of low pH soils that are widely distributed in Ethiopia. It is critical to identify tolerant germplasm and understand the molecular genetic basis of aluminum tolerance. Moreover, because chickpea yield depend on beneficial soil micro-organisms, especially nitrogen-fixing symbiotic bacteria and phosphate solubilizing micro-organisms (PSMs), a parallel need is to identify acid/Al- tolerant chickpea microbes.

We propose to identify stable and high yielding lines of chickpea and co-occurring symbiotic bacteria that are adapted to the current threats of crop production, particularly acid soils (aluminum toxicity). Ultimately, chickpea lines with high yield and better nutrition along with efficient PSMs inoculants will be combined with marketing, seed system and value chain strategies to increase the incomes of local farmers and to stimulate economic growth for sustainable development in the central and western highlands of Ethiopia.

During the project implementation periods of two quarters, we have organized three field germplasm collection missions and carried out one field visits to selected three districts out of six potential districts planned for the chickpea value chain project activity. Overall a total of 44 chickpea seed samples and 174 nodulated chickpea sample were collected from the central and northern highlands of Ethiopia. More than 200 chickpea germplasms and 15 released cultivars were also assembled from the Ethiopian Institute of Biodiversity (EBI) and NAR systems, respectively. Moreover, three districts have been selected for chickpea value-chain study in the central highland. The samples collected will be further processed and evaluated under lab and field conditions for detailed analysis, while in-depth survey will be conducted by the socio-economics team members in the near future. 


Assessing chickpea’s wild relatives for resistance to root-lesion nematodes

John Thompson, Roslyn Reen, Rebecca Zwart, Centre for Crop Health, University of Southern Queensland Toowoomba Australia. Email:John.Thompson@usq.edu.au

The root-lesion nematodes Pratylenchus thornei and P. neglectus are widely distributed in grain growing areas of Australia and other countries. They have a wide host range attacking both cereal and pulse crops. In the sub-tropical grain region of Eastern Australia, chickpea (Cicer arietinum) is the most important pulse crop and it is often grown in rotations with wheat, a major host for root-lesion nematodes. The benefits of a wheat-chickpea rotation such as nitrogen fixation and mutual break crops for fungal diseases are diminished in fields containing root-lesion nematodes. Similar problems occur in other grain regions of Australia and elsewhere in the world such as the Mediterranean Basin and north-west USA. Therefore developing resistant cultivars of both wheat and chickpea is now a major objective of the Grains Research Development Corporation (GRDC) and Australian plant breeding companies.

Earlier research with a limited number of accessions of the wild chickpea species Cicer reticulatum and Cicer echinospermum showed the potential for obtaining effective resistance genes to root-lesion nematodes from these species for use in chickpea breeding programs (Thompson et al. 2011). Therefore GRDC is funding our project USQ00017 ‘Assessing accessions of wild chickpeas for resistance to root-lesion nematodes’ to screen a larger number of new accessions obtained from recent collecting in Turkey. Previously we have developed glasshouse methods for assessing resistance of wheat and chickpea cultivars based on reproduction rate of the nematodes in the roots and soil of plants in glasshouse experiments. These glasshouse methods correlate well with assessments of resistance in field experiments, but permit larger numbers of genotypes to be assessed under more reproducible conditions than in the field. In the first stage of this project we have been fine-tuning the glasshouse method for determining resistance of chickpeas (results reported separately by Reen et al. at this meeting). Meanwhile the wild chickpea accessions have been grown through quarantine in Australia and seed multiplied for distribution to a number of linked projects. In the 2016 winter season we will commence assessing up to 200 accessions for resistance to P. thornei and P. neglectus and this work will be continued in the 2017 winter season. Parallel screening of these accessions will occur in a project led by Professor Halil Elekcioglu at Cukurova University in Adana, Turkey, using methods optimised there. Combined information will permit the selection of the best accessions of C. reticulatum and C. echinospermum for future germplasm development, chickpea breeding and genetic analysis. 


Effect of nitrogen availability on chickpea leaf and root morphology

Rebecca A Valls1, Emmanuel Dacosta-Calheiros1, Edward Marques, 1 Joseph Rahm1, Roxana Soler1, Oscar Blanco1, Reyaz Rouf Mir2,3, R. Varma Penmetsa2, Douglas Cook2,
Eric von Wettberg1,*

* eric.vonwettberg@fiu.edu

1 Florida International University, Biological Sciences, and, International Center for Tropical Botany, Miami, FL, USA

2 University of California at Davis, Department of Plant Pathology, Davis, CA, USA
3 Sher-E-Kashmir University of Agriculture and Technology, Division of Plant Breeding and

Genetics, Jammu, INDIA

Abstract

Chickpea (Cicer arietinum) is the second most important pulse legume crop in the world producing over 13 million tons in over 50 countries worldwide. Whether directly through consumption or by feeding livestock, chickpeas and other legumes provide 30% of our nutritional nitrogen; legumes accomplish this by creating symbiotic relationships with nitrogen fixing rhizobial bacteria within nodules found on the plant’s roots where the legume provides sugar in exchange for bioavailable nitrogen. Due to their nitrogen fixing ability, chickpeas are often grown in rotation with major agricultural crops like wheat and corn in order to reintroduce nitrogen and other nutrients into heavily depleted soils. Although it is widely observed that increasing nitrogen fertilizer applications on legumes inhibits their relationship with nitrogen-fixing rhizobia, there is little information on how these limitations affect leaf and root morphologies across wild and cultivated lines of chickpeas. Wild chickpeas grow in extremely nitrogen-limited environments, while cultivated chickpea occurs in deeper soils with more naturally available mineral nitrogen. Root and leaf morphologies hold information regarding plant development, and understanding the plasticity of chickpeas in response to nitrogen level can help to increase efficiency in their agricultural production. We are interested in seeing how various concentrations of nitrogen fertilizer affect the morphology of both root and shoot development of both wild and cultivated lines of chickpeas. We are using several computational tools to analyze multiple measurements in the roots and shoot including: internode and petiole length, frequency of branching, leaf shape and root architecture. We note several differences in root and shoot morphology between wild and cultivated chickpea. 


Exploring Genetic Diversity in Landrace Chickpea Accessions in the Secondary Centers of Diversity and in the Fertile Crescent

Kassaye N Dinegde1,2, Syed Gul Abbas Shah Sani1,7, Peter L Chang1, Noelia Carrasquila- Garcia1, Zehara Muhammad2, Yayis Rezene2, Bahattin Tanyolac3, Abdullah Kahraman4, Kassahun Tesfaye5, Muhammad Farooq Hussain Munis7 Asnake Fikre2,, R. Varma Penmetsa1, Eric JB von Wettberg6, Douglas R Cook1

1University of California at Davis, USA
2 Ethiopian Institute of Agricultural Research, Ethiopia 3 Ege University, Izmir, Turkey
4 Harran University, Izmir, Turkey
5 Addis Ababa University, Ethiopia
6 Florida International University, USA
7 Quaid-i-Azam University,Islamabad, Pakistan

Abstract: Many chickpea accessions are stored in ex situ genebanks and in situ at the farmers’ hands in the developing world. Little is known about the extent of genetic diversity contained in these landrace cultivars and its relation to advanced elite lines. On the other hand, landraces are among the basic raw materials for chickpea genetic improvement with potential to insure chickpea production against the prevailing climate change threats. Development of high throughput genotyping by sequencing methods opened the possibility for genome-wide characterization of large collections of germplasm accessions. In this study we used 26,547 SNP markers to estimate the genetic diversity and population structure in a collection of 952 chickpea accessions from Ethiopia, Pakistan and Turkey including landraces, elite materials and cultivated varieties. STRUCTURE analyses identified substantial admixture among these groups, and is being used to refine the genotype data sets for further analysis via a variety of methods (STRUCTURE, principal component analyses, AMOVA, etc). Data from these preliminary analyses will be presented.