of coriander seed in Australia, Europe and America. The second refers to the grey angular lesions in particular, where it is thought that the pinprick indentations could be the result of programmed cell death related to a hypersensitive response (HR). Experiments are currently being carried out to investigate these two hypotheses.
The use of multi-locus sequence typing (MLST) to study genetic variation within Ralstonia solanacearum
J. Danial, G.S. Saddler & P. van de Graaf
Scottish Agricultural Science Agency, 82 Craigs Road, East Craigs, Edinburgh EH12 8NJ, UK
Multilocus sequence typing (MLST) is a nucleotide sequence based approach for the characterisation of isolates of bacteria and other microorganisms. With MLST, isolates of microorganisms are characterised using the nucleotide sequences of the internal fragments (approximately 450-500 base pair) of selected housekeeping genes. For each housekeeping gene, sequence differences between isolates are assigned as distinct alleles, and for each isolate the alleles at each of the loci define the allelic profile or sequence type. The sequence types identified for each species can be stored in a database to allow Internet access and easy identification of species and strains by other MLST users. MLST provides an accurate and sensitive molecular typing system that can be used to track different strains of important pathogens in epidemiological studies. Thus far, MLST has mainly been applied to human pathogens, such as Streptococcus pneumoniae, methicillin resistant Staphylococcus aureus and recently the yeast Candida albicans. SASA is the first to use this particular technique for the genetic characterisation of a plant pathogen, namely the bacterium Ralstonia solanacearum, the cause of potato brown rot. The aim of this study is to identify possible genetic differences within R. solanacearum biovar 2, race 3, which has not been possible with other techniques. It is hoped that any differences found can be related to differences in virulence and used to clarify other aspects of brown rot epidemiology. To create a MLST database for R. solanacearum, a number of candidate housekeeping genes were selected by mining the complete genome sequence of the strain GMI1000, a race 1 isolate from tomato. On the basis of the chosen housekeeping genes, nested primers were designed which were tested against a selection of R. solanacearum strains. After this initial screening, seven housekeeping genes involved in small molecule metabolism were selected for further testing. The external primers were used as PCR primers and the nested primers as sequencing primers. Initially, 110 strains of R. solanacearum are being sequenced, which includes different biovars from around the world. The preliminary results indicate genetic variation between the biovars but not within biovar 2.
Potential of bacteria to control Fusarium diseases of cereals
Khan, M. R., Fischer, S., Egan, D. and Doohan, F. M.
Molecular Plant Microbe Interaction Group, Department of Environmental Resource Management, Agriculture and Food Science Building, University College Dublin, Belfield, Dublin-4, Ireland.
One hundred and fifty Irish bacterial isolates were collected from Irish cereal rhizospheres and phyllospheres and in vitro dual culture and in vitro and in vivo seedling blight tests were used to determine if any such organisms showed potential for either the in vitro growth inhibition of cereal- pathogenic Fusarium species or Fusarium seedling blight disease control. In the dual culture test, ten bacterial isolates were antagonistic to Fusarium culmorum, F. graminearum and F. poae. Of these ten bacterial isolates, Pseudomonas fluorescens strain MKB 90 and strain MKB 156 and thirteen other bacteria (that did not directly inhibit the growth of Fusarium species in dual culture tests) reduced F. culmorum infection on germinating wheat (cv. GK-Othalom) seeds, as determined using an in vitro seedling blight test. Of the fifteen bacteria that inhibited Fusarium infection of germinating wheat cv. GK-Othalom, seven were non-host cultivar-specific, i.e. they inhibited Fusarium seedling blight disease to varying degrees on all wheat cultivars tested (cvs Access, Marshal, Claire, Soisson and Baldus). In vivo seedling blight tests showed that Pseudomonas sp. strain MKB 158 reduced F. culmorum infection of wheat and barley stem bases by 79% and 82%, respectively, relative to Fusarium-infected control seedlings.
ROOT ROT SYMPTOMS IN AUSTRALIAN CANOLA
S. J. Sprague1,2 B. J. Howlett2 and J. A. Kirkegaard1
1CSIRO Plant Industry, GPO Box 1600, Canberra 2601 AUSTRALIA, 2Department of Botany, The University of Melbourne, Parkville 3010 AUSTRALIA
Leptosphaeria maculans, the causal fungus of blackleg (phoma), causes stem canker and is the major pathogen of canola (B. napus) in Australia and worldwide. Although L. maculans is described as a foliar pathogen, it can also infect roots following inoculation of wounded laterals with spores. Recently in Australia, premature death of canola plants lacking visible external symptoms of blackleg or other disease have been observed. These plants often have stunted root systems and diseased tissue in the roots similar to that caused by L. maculans in the stem. In a survey of 70 commercial canola crops in south-eastern Australia, 86% of crops, some of which were in paddocks that had never grown canola previously, had plants with root rot symptoms. To determine whether infection of roots occurs through the root or via foliar infection under field conditions, plots were fumigated with methyl bromide prior to sowing. Plants in fumigated plots had similar levels of root rot at maturity as plants growing in plots that had not been fumigated, indicating that the main pathway of infection is from airborne inoculum. Leptosphaeria maculans has been isolated from all diseased root tissue and inoculation of canola plants with this pathogen has reproduced the root rot symptoms.