Jalpa P. Tewari

The camalexins : new phytoalexins produced in the leaves of Camelina sativa (cruciferae)

Abstract Two new thiazoyl substituted indole phytoalexins, camalexin (5) and methoxycamalexin (8), were isolated from Camelina sativa leaves following elicitation by the fungus Alternaria brassicae.

Characterization of Plasmodiophora brassicae populations from Alberta, Canada

Clubroot, caused by Plasmodiophora brassicae, was identified in a number of canola (Brassica napus) fields in central Alberta in 2003. To characterize the virulence of the pathogen in the province, field populations from a number of locations in the Edmonton region were tested on the two most widely used sets of differential hosts, those of P.H. Williams and the European Clubroot Differential (ECD) series. Populations from British Columbia and Ontario were included for comparison. While the reaction of some hosts could be clearly defined as either resistant or susceptible, others showed intermediate disease index scores. If disease indices of 0%–49% and 50%–100% were regarded as resistant and susceptible, respectively, then populations from Alberta were classified as ECD 16/15/12 and 16/15/0 on the ECD set, or pathotypes 3 and 5, respectively, on the hosts of Williams. The population from British Columbia was classified as ECD 16/2/12 or pathotype 6, and the Ontario population was classified as ECD 16/0/14 or pathotype 6. The Alberta populations were more virulent on the B. napus hosts than those from other provinces, perhaps a reflection of their canola origin. In addition, 48 canola cultivars included in the 2004 Prairie Canola Variety Trials were screened for resistance to a local population of the pathogen, and all appeared to be highly susceptible. If clubroot were to become more widely established in western Canada, it could have a major negative impact on yields.

Resistance to Alternaria brassicae and phytoalexin-elicitation in rapeseed and other crucifers

Abstract An accession of Brassica campestris ssp. rapifera was less susceptible to Alternaria brassicae than B. campestris ssp. oleifera and B. napus ssp. oleifera . Accessions of Camelina sativa and Capsella bursa-pastoris were very resistant to A. brassicae , showing no symptoms. Production of phytoalexins of different types and amounts was found in all the above mentioned crucifers in response to A. brassicae . The differences in the susceptibility of these plants may be due, in part, to qualitative and/or quantitative differences in phytoalexin production. This appears to be the first report of elicitation of phytoalexins in crucifers upon being challenged by a fungal pathogen.

RAPD analysis of three Alternaria species pathogenic to crucifers

Genetic variation in Alternaria brassicae, A. brassicicola, and A. raphani collected from geographically diverse regions of the world was studied with RAPD and RFLP markers. Twenty 10-mer primers of arbitrary nucleotide sequences were tested for amplification of genomic DNA of A. brassicae using PCR. Of these, five primers amplified the genomic DNA from 20 A. brassicae isolates and produced reproducible RAPD profiles. UPGMA analysis of RAPD data showed that isolates collected from geographically distinct regions could be broadly classified into four groups. Intra-regional variation between isolates was less apparent. Variation was, however, higher in A. brassicicola, as based on RAPD analysis. Two isolates (from Canada and France) of A. raphani also showed variability with different RAPD profiles generated by all five primers tested. Five polymorphic, distinct RAPD products were used as hybridization probes for RFLP analysis to detect inter- and intra-specific variation. Variation among A. brassicae, A. brassicicola, and A. raphani was evident. Non-radioactive probes were also used to hybridize with Southern blots of A. brassicae, A. brassicicola, Leptosphaeria maculans, Rhynchosporium secalis and Brassica juncea for the selection of A. brassicae-specific probe(s). Probe AbP3 specifically hybridized with restriction digests of A. brassicae but not with those of A. brassicicola or other tested species. This probe should, therefore, be useful for distinguishing between two important pathogens of crucifers, i.e. A. brassicae and A. brassicicola both in culture and infected tissues.

Relationship between conidial concentration, germling growth, and phytoalexin production by Camelina sativa leaves inoculated with Alternaria brassicae

Germination and germ-tube growth of Alternaria brassicae conidia were lower with higher conidial concentration and lower on leaf surfaces of Camelina sativa than on glass slides. Leaves of C. sativa were highly resistant to A. brassicae . No disease was seen until leaves began to senesce after 6 d, and then only as localized brown flecks after inoculation with 100–10000 conidia. Camelina sativa produced a phytoalexin even when very few conidia were deposited on leaves. The phytoalexin concentration increased with increasing inoculum until leaves started to senesce. The phytoalexin slowed germination and inhibited germ-tube growth of A. brassicae conidia in vitro . The phytoalexin accumulated within leaf areas under and in conidial droplets. The rapid rate of phytoalexin accumulation shortly after inoculation could account for inhibition of fungal growth on the leaf surfaces.

Interactions of Alternaria brassicae conidia with leaf epicuticular wax of canola.

Epicuticular wax on canola leaves reduced the rate of germination and number of germ tubes produced by Alternaria brassicae conidia. The effect was more pronounced in cvs with greater amounts of wax. The wax in canola is in the form of a fluffy layer and may affect germination by impeding movement of foliar exudates to the surface.

Resistance to Rhizoctonia solani and Presence of Antimicrobial Compounds in Camelina sativa Roots

Camelina sativa (L.) Crantz was significantly more resistant to Rhizoctonia solani Kühn than [itBrassica napus L. cv Westar. Emergence of C. sativa seedlings was 22 to 33% greater than those of Westar in R. solani-infested soil. The greater resistance of C. sativa seedlings to R. solani appeared to be due to greater amounts of antimicrobial compounds present in C. sativa roots. These antimicrobial compounds inhibited the growth of both weakly virulent and virulent R. solani] isolates to the same extent. Four antimicrobial compounds were purified from C. sativa roots and their structures elucidated. Two were identified as the phytoalexins (camalexin and methoxycamalexin) previously described from C. sativa leaves. This appears to be the first report of elicitation of phytoalexins from roots of crucifers. Two preformed antimicrobial compounds were identified as methyl 1-methylindole-3-carboxylate and 10-methyl sulfinyldecylisothiocyanate.

The infection process of Alternaria cirsinoxia on Canada thistle (Cirsium arvense) and host structural defence responses

The infection process of Alternaria cirsinoxia was studied on Canada thistle (Cirsium arvense) in the controlled environment and in the field. In the controlled environment, germination of conidia began at 2 h and appressoria formation at 4 h after inoculation. Approximately 75% of appressoria formed at the anticlinal wall junctions of the epidermis. Leaf penetration occurred between 6 and 24 h, most commonly in between adjoining anticlinal walls. Penetration through stomata was rare. After penetration, large, intracellular infection hyphae formed and branched within epidermal cells, ramifying throughout the leaf tissues inter- and intracellularly by 24 h. In field conditions, infection coincided with prolonged rainfall and conidia remained viable on the leaf surface for 8–9 days (d) before causing infection on leaves. A host response occurred after penetration, involving deposition of lignin and callose in the infected epidermal and mesophyll cell walls. High levels of silicon were detected in epidermal cells directly below appressoria, often appearing to form entirely silicified infected cells which were resistant to collapse after air-drying. This study shows that A . cirsinoxia has the potential for rapid invasion of the leaf tissues of Canada thistle under good moisture conditions. The implications of the host responses, in terms of defence against infection, are discussed.

Calcium oxalate crystal formation in Rhizoctonia solani AG 2-1 culture and infected crucifer tissue: relationship between host calcium and resistance

Crystals of various forms were observed in liquid cultures of Rhizoctonia solani AG2-1 and on the surface of infected hypocotyls of canola ( Brassica napus , cv. Westar), mustard ( Sinapis alba , cv. Arda) and Camelina sativa . The crystals were identified as calcium oxalate based on solubility, staining reaction, potassium permanganate titration and energy-dispersive X-ray microanalysis. More calcium-containing crystals were formed on hypocotyls of the susceptible cv. Westar than on hypocotyls of the resistant cv. Arda and on C. sativa , and on 1-wk-old plants than on 3-wk-old plants. There was no significant difference in total calcium content in non-infected hypocotyl tissues among the three plant genera, and the calcium content did not increase with age of plant tissue. The production of oxalic acid in culture and the formation of calcium oxalate crystals in infected tissues suggested that calcium in plant cells was sequestered by oxalic acid produced by R. solani .

Abscisic acid from Alternaria brassicae

Abstract Abscisic acid has been characterized in the mycelium of the black spot fungus, Alternaria brassicae .