EVALUATION OF LENTIL GERMPLASM FOR RESISTANCE TO WILT, RUST AND STRMPHYLIUM BLIGHT

Introduction

Lentils are considered to be one of the oldest food crops of mankind that  researchers have traced back to 7000-8000 BC (Hawtin, et al., 1980; Cubero, 1981; Lev- Yadun et al., 2000). Worldwide, lentil is grown on a total area of 1.8 million hectares, of which 60% is in the South Asian region which includes the lentil producing countries of Bangladesh, Burma, India, Nepal and Pakistan (Nazir et al., 1994). High protein content, starch, low level of antinutrients and ability to grow in water stress conditions are main attributes that make lentil an important crop (Anonymous, 2003). Morphological characterization is the first step in the classification and description of any crop germplasm (Ghafoor et al., 2001; Upadhyaya et al., 2001). Use of botanical descriptors for genotyping has advantages and disadvantages (Ramanatha & Riley, 1994; Watanabe et al., 1995). Among biochemical techniques, seed proteins and isozymes are practically reliable methods due to independence of environmental fluctuation (Murphy et al., 1990). Electrophoresis adds information to taxonomy and should not be dissociated from morphological, anatomical and cytological observations (Piergiovanni and Taranto, 2003; Ghafoor et al., 2002). Advancement in molecular biology has introduced DNA markers which is an attractive method for genotype identification (Samec & Nasinec, 1996; Sonnante & Pignone, 2001). Several kinds of DNA markers have been recognized and among these RAPD has been reported useful tool for evaluation of intra-specific variation (Thormann et al., 1994; Ribu & Hilu, 1996). No single method is adequate for assessing genetic variation because the different methods sample variations at different levels and differ in their power of genetic resolution as well as in the quality of information content. To enhance the scope and usefulness of lentil genetic resources preserved in the genebank, the present research was undertaken to investigate genetic diversity for botanical descriptors, proteins and DNA markers to identify landraces to enhance breeders’ efficiency for crop improvement. The results of this study are expected of practical utilization by the genebank curators and plant breeders for future germplasm management and crop improvement.

 FUSARIUM WILT

Fusarium wilt of lentil is an important disease reported in every-continent wherelentil is grown except Australia (Beniwal et al., 1993; Tosi and Cappelli, 2001).The disease may cause complete crop failure under favorable conditions for disease development, and can be the major limiting factor for lentil cultivation in certainareas (Chaudhary and Amarjit, 2002). The common name lentil wilt has beenused to describe many general wilting and dying symptoms. Hence a number ofpathogens have been reportedly associated with-lentil wilt (Khare, 1981) possiblybecause of the difficulty in species identification and confusion in the fusarium taxonomy. Strictly speaking, the causal organism of vascular wilt of lentil isFusarium oysporum Schlecht. Emend. Snyder & Hansen f. sp. lentis Vasudeva andSrinivasan. Although its sexual state has not been found, it is generally believed tobelong to the Hypocreales of Ascomycetes. In culture, the mycelium of the pathogen is hyaline, septate and much branched.Growth patterns on media vary from fluffy to appressed and vary in color from nocolor to pink. F. oxysporum f. sp. lentis produces three kinds of spores: microconidia;multi-septate macroconidia, which have a distinct foot cell and a pointed apicalcell; and chlamydospores (Khare 1980). Microconidia are ovoid or kidney-shaped,hyaline and usually one celled. Macroconidia are long with pointed apical cell andnotched basal cell, and two to seven celled. Chiamydospores are oval or spherical,one-celled, and thick walled, formed singly in macroconidia or apical or intercalaryin the hyphae.

Symptoms

Fusarium wilt usually occurs near or at reproductive stages (flowering to dod-filling) of crop growth. Symptoms include wilting of top leaves that resembI6 waterdeficiency, stunting of plants, shrinking and curling of leaves from the lower prt ofthe plants that progressively move up the stems of the infected plant. Plants finallybec6hie completely yellow and die. Rbot symptoms include reduced growth withmarked browr!i discoloration, tap root tips that are damaged and prOliferatin ofsecondary roots above the area of tap root injury. Disoloration of vascular" ascula tissue in the lower stem may not always beisible. However, in India, the disease has I. also been reported to occur at th& sedling stge. General symptoms at the'séedlingstage include seed rot and sudden drooping 'mOre like wilting and damping off (Khare"198O) . Field diagnosis should he done in coñOection with field cropping historyRecent lentil ioduction specially with a histöry Off fusarim wilt will indicate potential wilt problems Suspect stunted and wilted plants should he carefully removed fromthe soilo that the rootscan be cleckeifor reduced giowth without external fungalgrowth.t External fungal 'iñdicites the essence of the &disease such ascollar rot. Lower stems should he split to check for vascular discolor rati Althoughvascular discoloration is not always symptomatic of fusarium wilt the presence ofdiscoloration would confirm the disease. Culturing of infected plant tissue in thelaboratory should be done with caution because of the )sibl presence of othersaprophytic Fusariurn spp. that appear similar to F. oxysporum f. sp. lentis. Apathogenicity test on lentil is necessary to confirm F. oxvsporuin f. sp. lentis.

Epidemiology

Like many other formae specilaes of F. oxysporun, the pathogen has a very limitedhost range as it only infects lentil in nature. In inoculation studies, F. oxysporumf. sp. lentis was unable to infect cowpea, french-bean, bengal gram, lathyrus (Khare, 180). The disease is favourable'warm and dry conditions (Bayaa and 'Erskine, 1998) With an optimal temperature'f 22-25 °C. F. oxysporun f. sp. lentis is a soilbOrne pathogen, although seed infestation andiifectioii is common. The chlathydospores can survive in soil either in dormant formor saprophiicaIly for several years without a suitable host. A survey of soil samplesfroth Sangod Tehsilof Kota, Rajasthan, India, found that F. ox)'sporum f. sp. Ienti.s' was the most prevalent lentil pathogen (Chaudhaiy and Amarjit, 2002). Synergisticinteracti(ih between F. orvsporum f. sp. lentis and root knot nematode Meloidogynejàvanica was observed in lentil cultivars resistant or susceptible to fusarium wilt ci al.. 2001'). 'Presehcè of the nematode significantly increased wilt incidence,iussed significant reduction in shoot length, toot length and nodulation in both susceptible and resistant cultivars (De ci al., 2001).

 Control methods

The most 'econorriical hieins to control fusariuth wilt 'of lentil is through 'the use of resistant.cultivars,(Bayaa etl., 1997; Stoilova and Chavdarov, 2006). Resistantor moderately resiStant lentil cultivars (OPL 58, 'DPL 61 and DPL 62) signifi-'C'antly reduced wilt incidence and severity of root rot, and increase grain yield(Chaudhiry and Athdrjit, 2002).

LENTIL RUST

Rust, caused by Uromyces viciae-fabae (Pers.) Schroet, (Uredina/es, Pucciniaceae)is regarded as the most important foliar disease of lentil (Figure. Ib) (Erskineet al. .1994). Complete crop failures can occur due to this disease (Beniwal et al. 1993).Rust of lentil is widespread globally; but is considered to be a production problem inAlgeria, Bangladesh, Canada, Ethiopia, India, Italy, Morocco, Pakistan, Nepal, Syriaand Turkey (Erskine et al. 1994). The disease also occurs widely in South Americaincluding Argentina, Brazil, Chile, Colombia, Ecuador, and Peru (Bascur 1993). Rust is an autoecious fungus, completing its life cycle on lentil. The aecia of U. viciae-fabae are amphigenous or hyphyllous, usually in groups surroundingthe pycnia or sometimes scattered, cupulate, 0.3-0.4 mm diam. The aeciosporesare spheroidal, 18-26 p.m diam.; wall hyaline, verrucose, I p.m thick. Uredia areamphigenous and on the petioles and stems, scattered, cinnamon, 0.5—I mm diam.Uredospores are ellipsoidal or obovoidal 22-28 x l9-22 pm; wall luteous to sienna,very finely echinulate, 1 —2.5 p.m thick; pores 3-4, equatorial or occasionally scattered on Lathyrus. Telia are like the uredia but black and larger: 1-2 mm diam.Teliospores are ellipsoidal, obovoidal or cylindrical, rounded or subacute above,25-40 x l8-26m; wall chestnut, smooth, 1-2 Km thick at the sides, 5—I2 iim thickabove; pedicels sienna to luteous, up to 100 Km long. (Laundon and Waterson 1965).

Symptoms

Rust starts with the formation of yellowish-white pycnidia and aecial cups on thelower surface of leaflets and on pods, singly or in small groups in a circular form(Agrawal et al. 1993). Later, brown uredial pustules emerge on either surface ofleaflets, stem and pods (Figure 1 b). Pustules are oval to circular and up to 1 mmin diameter. They may coalesce to form larger pustules (Bayaa and Erskine 1998).The telia, which are formed late in the season, are dark brown to black, elongatedand present mainly on branches and stems. In severe infections leaves are shed andplants dry prematurely (Bakr 1993), the affected plant dries without forming anyseeds in pods or with small shriveled seeds. The plant has a dark brown to blackishappearance, visible in affected patches of the paddock or in the whole paddock iftotally infected (Beniwal et al. 1993).

 Epidemiology

The disease generally starts from low-lying patches in the paddock and radiatestowards the border (Bayaa and Erskine 1998). Lentil seed may be contaminatedwith pieces of rust-infected leaf, stem and pericarps, which can act as primaryinoculum for the recurrence of the disease in most years (Khare 1981, Agrawalet al. 1993). Rust may also perpetuate on weed hosts from where it may infect lentilcrops by windborne teliospores. High humidity, cloudy or drizzly weather withtemperatures 20 to 22°C favour disease development (Agrawal et al. 1993). Thedisease generally occurs during the flowering /early podding stage. Aeciosporesgerminate at 17-22°C and infect other plants forming either secondary aecia attemperatures of 17 —22°C or uredia at 25 °C. Uredosori develop later in the seasonand are rapidly followed by telia (Beniwal et al. 1993). After harvest, aecia anduredia present on lentil trash die out, but teliospores tolerate high temperaturesand allow the fungus to survive the summer. At lower temperatures, uredosporescould be an important means of survival (Bayaa and Erskine 1998). Uredomyceliumis highly resistant to heat and sunlight and is probably important for continueddevelopment and survival of rust in hot, dry conditions. The predominant form ofsurvival will vary with the environment and location (Bayaa and Erskine 1998).Teliospores germinate at 17-22°C without a resting period and cause new outbreaksof the disease each season. There are 70 recorded hosts of U. viciae-fabae including lentil, chickpea, fieldpea, Lathvrus spp and Vicia spp. (Parry and Freeman 2001). Degrees of hostspecialisation and pathogenic variability do exist within populations of U. viciae-fabae worldwide. Much research has been performed regarding race identification within U. viciae-fabae over many years with conflicting outcomes regarding thesuggestion of forma speciales within the species.

Control Methods

Cultural control methods currently recommended for control of U. viciae-fabaeinclude: control of volunteer plants over summer; isolation of new season cropsfrom old host crop stubbles (MacLeod 1999) and destruction of old lentil stubbles(Prasada and Verma 1948). Early studies on the control of lentil rust in Indiafound seed treated with Agrosan (phenylmercury acetate) to control seed-borneinoculum (Prasada and Verma 1948). Singh (1985) found Vigil (diclobutrazole),applied as a seed dressing prevented the appearance of U. viciae-fabae up to70 days following inoculation with uredospores; bayleton (triadimefon) preventeddisease appearance up to 40 days post inoculation and the untreated control wasseverely infected with rust 35 days after inoculation. Experiments investigating theuse of foliar fungicides for rust control by Agarwal et al. (1976) found Hexaferb(Ferric dimethyldithiocarbamate) and Dithane M-45 to give the best control of U.viciae-fabae in experimental plots at Jabalpur, India. In addition, Dithane M-45also increased plot yield by 82% and grain weight by 24% when compared to theuntreated control. The use of host plant resistance is the best means of rust control(Bayaa and Erskine 1998). Genetic differences among genotypes and sources ofresistance have been reported worldwide, with several rust resistant lines available.Resistance to rust is reported to be controlled by a single dominant gene (Sinha andYadav 1989). Studies in factors influencing the mechanism of resistance to rust inlentil (Reddy and Khare 1984) reported that resistant cultivars contained more leafsurface wax, F, K, S, Zn, Fe, Cu levels of phenols than susceptible cultivars whichhad higher levels of amino acids, N, Mn, and sugars. Structurally there were nosignificant differences found between resistant and susceptible cultivars.

STEMPHYLIUM BLIGHT

Stemphylium blight of lentil is caused by the pathogen. Stemphylium botryosurnWallr (Pleosporales, P!eosporaceae) (teleomorph: Pleospora herbarum (Fr) Rab:).The disease has been reported on lentil from Bangladesh (Bakr 1993), Canada(Morrall 2003), Egypt, Syria (Bayaa and Erskine 1998) and the USA (Wilson andBrandsberg 1965). The disease has the potential to cause yield losses of up to 62%under conducive conditions (Figure Ic) (Bakr 1993).

Conidiophores of S. botryosuin have 1-7 septate. 20-72 x 4-6 p.m, pale brownto brown, with a swollen apical sporogenous cell 7-11 p.m diam., and slightlyroughened toward the apex. They possess a single apical pore 5-8 p.m diam. Conidiaare oblong, olive to brown, ovoid to subdoliiform, occasionally constricted at 1-3transverse septa and at the 1-3 longitudinal septa if complete, 19.5 x 28.5 p.m witha single basal pore 8 p.m diam. and a roughened outer wall. Ascostromata arescattered, immersed to erumpent in the tissue of the host, 100-500 p.m in diam.Asci are 90-250 x 20-50 p.m containing eight ascospores, cylindrical to slightly club shaped. AscOspores are light to 'yellow brOwn, ellipsoid to club shaped with7 septate, slightly constricted at the three primary transverse septa, muriform and26-50 x I0-201.tm (Booth and Pirozynski 1967).

6.1. 'S3mp40ffls

Symptoms of stemphylium blight start with the appearance of small pin-headedlight brown to tan coloured spots on leaflets. Under ideal conditions the smallspots enlarge rapidly, covering the entire leaflet surface within a 2-3 day period.The infected tissue appears light cream in colour, often with angular patterns oflighter and darker areas1that spread across, or long, the entire leaflet (Morrall 2003;Figure Ic). The affected foliage and stems gradually turn dull yellow, giving ahlihted appearance to the crop (Bakr 1993). The infected leaves can be abscised  I rapidly, leaving only the terthinal leaflets on the stems. The stems bend down, dry and gradually turn ashy white, but pods remain green. White mycelial growth can sbthetimes be seen Oh the infected stems(Bkr 1993).

Epidemiology

Important sources, df S. botryosuminoculum include infected crop debris andinfected seed. Infected crop debris can be a source ofprimary inoculum in the formofair-horheascosporesor asresting mycelium, basedonthe studiesof thepthogenon other hostcrops such as alfalfa (GilchristI990) Stemphylium botrocum isknbwn to bc carried on seea (Booth and Pirozynski 1967) and Stemphylium spp hasheen'isolated oft lentil seed in Australia (Nasir and Bretag 1997) but the significanceof seed borne S botrocum inoculum on disease initiation in lentil is not clearlyhndërstobd (Mwakthhya 2066). -Bdkr\(l 993)has represented from Bangladesh that –the pathogen commeilces infection when the ambient night temperature remains above86C. Th RH canopy must also reach 94% In India Singh'and Singh (1993) found that an average meantemperature of '18 C ± 2 C andRH of 85-90% in the morning was favourablefdr disease d6v1opme'nt and spread. Most recently, inCanada Nlwakutuya (2006)'fo.indthat symptbrn development of S.hotyosu,'as optimised after 48h of leaf wetness at'temperature  above 25°C. The host range of S. hotiyosun is wide and includes a large iiUrhhér Ornamental, horticultural and field crop species. The se include lentil (Bakr 1993), lupid (Tate 1970), tomato (Bashi and Rotem 1975)spinach (Koike et al. 2001), alfalfa, clover (Smith'1940); lettuce ('Tate4970), apple,onion and gladiolus (Booth and'Pirozynski 1967).

 Control Methods

There is little published information available regarding cultural control methods forS. botryosum in lentil. Being a stubble-borne iiseasestr'átcgies suCh as destruction of old crop residues, and crop rotation would assist in. decreasing potential inoculumsources. In Bangladesh, delayed sowing was found to significantly decrease theincidence of stemphylium blight in lentil, but later sowing resulted in reduced cropyields and heavy infection by U. viciae-fahae (Bakr 1993). Foliar fungicides havebeen found to he effective in the management of stemphylium blight. In Bangladeshthe application of Royal 50 WP was found to effectively control the disease whenapplied three times at weekly intervals starting from the initiation of the disease(Bakr 1993). In other horticultural crops, such as asparagus and garlic, Stemphyliumspp. has been successfully controlled using chiorothalonil (Meyer et al. 2000),tebuconazole and procymidone (Basallote-Ureha et al. 1998). Sources of host plantresistance have been identified in screening nurseries in Bangladesh. The resistantvarieties 'Barimasur 3' and Barimasur 4' were released with resistance to S.botryosunz (Sarker et al. 1999a, b). Studies by Chowdhury et al. (1997) foundlentil cultivars with resistance to S. botryosum had a. higher number of epidermalhairs, thicker cuticle, thicker epidermal cell layer and thicker cortical layers. Inaddition, resistant lines were also, found to have fewer stomata than susceptiblecultivars.

Developing a Breeding Strategy for Stemphylium Blight Resistance in Lentil

Stemphylium blight (SB),caused by Stemphylium botryosum Wallr. is a major biotic constraint in many lentil production regions of the world, particularly the northeastern lentil growing area of South Asia. It is a major problem in Bangladesh and Nepal and has appeared in fields in North Dakota and Saskatchewan in recent years (Kumar 2007, Holzgang and Pearse, 2001). The disease causes defoliation, stem deformation and yield loss. It is increasing in importance in western Canada as the lentil crop expands into new production areas. With the increase of lentil production and deployment of resistance to ascochyta blight and anthracnose in new cultivars, SB has become a more serious problem (Vandenberg and Morrall, 2002). To date, there have only been a few investigations into the genetics of SB resistance and management of this disease. Four suitable isolates of Stemphylium botryosum (SB-16, SB-17, SB-19 and SB-BAN) were identified for indoor screening for SB resistance in University of Saskatchewan(Podder et al., ). For breeding purposes, in order to differentiate between susceptible and resistance genotypes, 2 to 4 week old plants should be inoculated. A newly developed semi-quantitative scale will be used to evaluate disease severity of lentil plants infected by SB to avoid the difficulties that were encountered using the Horsfall-Barratt scale this scale is relatively simple, taking into account lesion development as well as leaf drop. It allows large numbers of plants to be evaluated in

relatively short periods of time.

Multiple resistance by gene pyramiding

When there are different pathotypes and corresponding resistance genes, one method to breed for resistance is to combine different resistance genes into a single cultivar (gene pyramiding). By combining genes conferring resistance to different pathotypes, the cultivar can be used in more diversified environments where different pathotypes are likely to be dominant. In addition, multiple resistance genes may have additive effects. Even if they do not show additive effects, the presence of more genes implies that pathotypes have to be virulent to all the genes before a resistant cultivar will lose resistance (Crute 1988). This procedure has been suggested for breeding for durable resistance in many crops. The introduction of resistant cultivars with major resistance genes into production such as ‘Laird’ and ‘ILL 5588’ will increase the chance of the development of resistant pathotypes (Burdon 1993). It has been suggested that the large-scale cultivation of the moderately resistant cultivar ‘Laird’ is the cause of the increased aggressiveness of Canadian isolates of A. lentis (Ahmed and Morrall 1996).

Using wild relatives

Transfer the resistance genes from wild species into elite cultivars will be animportant approach in breeding for resistance. The only wild species that can be intercrossed withcultivated lentil easily is L. orientalis, the progenitor of cultivated lentil (Ladizinsky 1979, 1993).A recent report showed that viable hybrids could be obtained from the crosses between the cultivated and four wild lentil species by applying gibberellic acid (GA3) after pollination (Ahmadet al. 1995). Upon improvement, this technique may lead to an efficient method to transfer usefulresistance genes from these wild species into cultivated species. In vitro culture has been used to promote the use of wild relatives in lentil. Cohen et al. (1984) established a two-stage in vitro technique for the development of interspecific hybrid embryos. Fourteen-day-old fertilized ovules were cultured on Murashige and Skoog (MS) medium supplemented with zeatin, followed by release of the embryos from the ovular integuments. Ladizinsky et al. (1985) also obtained vegetatively normal L. culinaris × L. ervoides hybrids using embryo culture techniques. Micropropagation of the limited F1 materials has been explored as a way to enlarge the F1 population and eliminate the requirement of large-scale pollination to obtain enough hybrid material for further genetic study and breeding (Ye et al. 2000c). With the refinement of in vitro culture techniques and an artificial crossing method, it will be possible to use resistance genes from wild species by repeated backcrossing.

Molecular technology

Genes conferring resistance are conventionally introgressed into an elite background by repeated backcrossing. If, instead of tracking the gene itself, a marker tightly linked to the gene is used to trace its segregation in the selection process, then this method is referred as marker-assisted introgression. For resistance conferred by major genes, classical Mendelian linkage analysis can be used to identify linked markers. Using bulk segregation analysis. Transgenic technology provides plant breeders with new tools in breeding for resistance (Bent & Yu 1999). Genetic transformation in lentil can be achieved, as confirmed by GUS assay, but regeneration of transgenic plants was very difficult (Warkentin and McHughen 1993). However, electroporation of DNA into intact nodal meristems has resulted in the production of transgenic

plants (Chowrira et al. 1995). Oktem et al. (1999) obtained transgenic shoots from cotyledonary nodes containing GUS gene transferred by particle bombardment. Therefore, production of transgenic lentil plants and consequently the application of transgenic techniques may soon

become feasible.

Factors which need to be considered in breeding for resistance

Correlated selection response

The changes in other traits caused by selection for a trait is termed the correlated selection response (Falconer 1989). A recent study showed that the major gene for resistance in ‘ILL 5588’ and W63241 has adverse effects on seed yield/plant (Ye et al. 2001b).

1994).

Genotype × environment interaction

Genotype × environment (GE) interaction is of concern if the resultant cultivar is to be used across a large area. There are two reasons why GE interaction is more important in breeding for resistance than for other traits. First, pathogens may vary in their aggressiveness under different

environments. Furthermore, physiological races may be different across environments. Second, the growth, development and physiological status of host genotypes may change across environments. There is a paucity of information regarding the GE interaction in lentil. However, the different levels of aggressiveness among isolates from different locations and the recent identification of pathotypes suggest that GE interaction could be important.

Resistance sources are available both in cultivated and wild lentil species. Several major resistance genes have been identified, and minor genes have also been shown to play a role in resistance. Different pathotypes are present in the pathogen population and resistances conferred by major genes seem to be pathotype-specific. However, many challenges remain both for plant pathologists and for breeders. From a plant breeding viewpoint, more studies in the following areas are urgently required to sustain breeding progress.

Identification of more resistance resources

Large-scale screening of germplasm for resistance is required. What is important for the future screening of germplasm is to characterize the resistance reactions more carefully. It is necessary to evaluate the resistance several times and to test against different isolates. Three types of germplasm should be selected: (1) highly resistant with rating 1–2, (2) germplasm with slow blighting and (3) germplasm with multiple resistance.

Careful characterization of the host–pathogen interaction system

To identify and incorporate new sources of resistance genes into the breeding programme, it is necessary to have a good understanding of the pathogenic variability of A. lentis. While six different pathotypes are known, more pathotypes may be present. Molecular marker-based methods have been used successfully to provide additional information on the pathogenic variability and have the potential to fingerprint pathotypes in many host–pathogen systems. It can be expected that a better understanding of the pathogenic variability of A. lentis will soon be obtained by using these novel techniques. Armed with the understanding of pathogen populations, the host genotypes can be easily determined using different pathotypes to challenge host genotypes and, consequently, gene pyramiding becomes feasible.

Identification of molecular markers tightly linked to different resistance genes

With the development of modern molecular techniques, more and more marker systems become available and their costs will be reduced. It is possible to develop rapid and cost-effective techniques for screening large populations for markers linked to resistance genes. Once developed, the benefits of marker-assisted selection (introgression) will be available.

 Reference

Ahmad, M., A. G. Fautrier, and D. L. McNeil, 1996: Identification and genetic characterization of different resistance sources to Ascochyta blight within the genus Lens. Euphytica 97, 311—315.

Ahmed, S., and R. A. A. Morrall, 1996: Field reactions of lentil lines and cultivars to isolates of

Ascochyta fabae f. sp. lentis. Can. J. Plant Pathol. 18, 362-369.

Bavaa, B., Erskinc W and Khour, L.,Arab Journal of Plant Protection, (1986)4: 18- 119

Bent, A. F., and I. C. Yu, 1999: Applications of molecular biology to plant disease and insect

Burdon, J. J., 1993: Genetic variation in pathogen populations and its implications for adaptation to host resistance. In: T. H. Jacobs, and J. E. Parlevliet (eds), Durability of Disease Resistance. Kluwer Academic Publ., Dordrecht

Chowrira, G. M., V. Akella, and P. F. Lurquin, 1995: Electroporation-mediated gene transfer into intact nodal meristems in planta: generating transgenic plants without in vitro tissue culture. Mol. Biotechnol. 3, 17-23.

Cohen, D., G. Ladizinsky, M. Ziv, and F. J. Muehlbauer, 1984: Rescue of interspecific Lens

cross Barimasur-4 × CDC Milestone. M.S. Thesis, University of Saskatchewan, Canada.

Crute, I. R., 1988: The elucidation and exploitation of gene-for-gene recognition. Plant Pathol. 47, 107-113.

expression in lentil cotylendonary nodes using particle bombardment. Lens Newsl. 26, 3-6.

Falconer, D. S., 1989: Introduction to Quantitative Genetics. Longman, London.

Hamdi A. and AM.Hassancin, Sur of Fungal Diseases of Lentil in North Egypt. Lens NcsIcLtcr, (1996), I&2,pp. 52-53

Holzgang, G., and P Pearse. 2001. Diseases diagnosed on crop samples submitted to the Saskatchewan Agriculture and Food Crop Protection Laboratory in 2000. Can Plant Dis Surv 81:21-27.

hybrids by means of embryo culture. Plant Cell Tiss. Org. Cult. 3, 343-347.

ICARDA. Food legume Improvement Program, Anrual Report tbr 1990, Akppo, Syria. ICARDA, (1990), 129 pp.

J kharc, M. N.. Wilt ot Lentil,. Jahalpur, M.I-. India: JNKVV ,( 1980), pp. 155
 kharc, M. N.. In: Lentils, (cds. C. 4hh and G. I-{awtin). UK: ICARDA/CAB.(1981),pp. 163-172

J Mihov, M., I. Stocva and P. Ivanov, RastcnicIni nauki,(1987),24: 45-51

Kharc.M. N. S.C.AgarI and AC. Jam.. Discases of LCNII arJ thcir control. Ta.hnicul flufleim JNKVV. Jalnipur, M.P., India,(1979)

Kumar, P. 2007. Genetics of resistance to stemphylium leaf blight of lentil (Lens culinaris) in the

Ladizinsky, G., 1979: The origin of lentil and its wild genepool. Euphytica 28, 179-187.

Ladizinsky, G., 1993: Wild lentils. Crit. Rev. Plant Sci. 12, 169-184.

LattY KS., ompitionano quality ot IcntiI (1.cns ctdinaris NIcdic. A rcvw. Canadian Institute of Food Scncc and Tcchndogv,(1988),21(2) 144.160

multiplication of lentil hybrids without genetic change by culturing single node explants.

Oktem, H. A., M. Mahmoudian, F. Eyidodan, and M. Yucel, 1999: GUS gene delivery and

potential of this explant for transformation by Agrobacterium tumefaciens. Lens Newsl. 20, 26 28.

R. Podder, S. Banniza and A. Vandenberg()Developing a Breeding Strategy for Stemphylium Blight Resistance in Lentil. Department of Plant Sciences/Crop Development Centre, University of Saskatchewan, 51 Campus Drive, Saskatoon, Saskatchewan S7N 5A8

resistance. Adv. Agron. 66, 251-299.

SABRAO J. Breed. Genet. 32, 13-21.

Saxena, D.R.and Khare, M.N., Indian Phytopathology (1988), 41:69-74

 Stoákwa T. and G. Percira, Morphdogical characterization of 120 land (Lens culinans Medic.) xcessions. Lens Newslctier, (1999), 1 &2, pp. 7-9

Vandenberg, A., and R.A.A. Morrall. 2002. Pulse crop variety development strategies in Saskatchewan. Saskatchewan Pulse Growers Pulse Days 2002, Saskatoon.

Warkentin, T. D., and McHughen, 1993: Regeneration from lentil cotyledonary nodes and

Ye, G., D. L. McNeil, A. J. Conner, and G. D. Hill, 2000c: Improved protocol for the