: There are increasing opportunities for the development of highthroughput in-vitro screens to aid the discovery of fungicides with novel modes of action. In the past, such screens were developed when biochemical targets were validated by fungicides with deÐned modes of action. However, genetic information is beginning to have a major impact both on the way in-vitro targets are selected and on the speed at which mode-of-action information is gained on current fungicides having an, as yet, undeÐned mode of action.This paper discusses issues concerning target selection and high-throughput screening, using examples taken from the current literature and from investigations at Zeneca Agrochemicals, using inhibition of fungal respiration as an example.Saccharomyces cerevisiae is discussed as model for fungicide research, both in terms of its sensitivity to known fungicides and its well deÐned molecular genetics, which makes it amenable to such techniques as gene dosage for mode of action determination.1998 Society of Chemical Industry ( Pestic. Sci., 54, 338È344 (1998)

The fluorescent ELISA technique for the identification of bacteria was compared with antibiotic resistant mutants as marker systems for use with Rhizobium trifolii in root nodules and in soil. With an effective(CP3B) and an ineffective (R4) strain as a mixed 1:1 inoculum, there was a highly significant correlation (P < 0.001) between the two techniques when the plants were grown at pH 5.5 when the majority of nodules were inhabited by the effective strain. At pH 6.5, where the ineffective strain predominated in the nodules, there was no correlation. The reason was that 85% of R4 nodules had volumes less than 0.1 mm3 with bacterial numbers obviously below the necessary threshold for detection using the serological method. Both methods were efficient at enumerating rhizobia from soils although the recovery rate from a brown earth soil was significantly higher than from a peat soil. Fluorescent ELISA was able to detect rhizobia at 8.0 times 105 cells/ml soil suspension (1 g soil to 10 ml water) in the brown earth soil and at 2.0 times 105 cells/ml in the peat soil. The results are discussed in terms of the limitations of both techniques in ecological studies.

S U M M A R YThe nodule occupancy of white clover cv. Milkanova three months after inoculation in an upland soil with a 1 : l mixture of an effective (CP3B) and an ineffective strain (R4) of R. trifolii, differed significantly depending on the lime level. The percentage of nodules occupied by the effective strain decreased from 64% to 29% with increasing lime level whilst the ineffective strain nodules increased from 4% to 24% with mixed nodule infections also increasing from 9% to 22% There was no such difference after 14 months with this cultivar nor with cv. S184 at either harvest. Inoculation with a higher ratio of the less preferred strain (R4) resulted in this strain dominating the nodules and significantly reducing the dry weight of the plants (first harvest) and clover cover of plots (second harvest). Where antibiotic resistant mutants of these strains were used, with one exception, there were no significant differences between the wild-type strains and their mutant derivatives for any of the parameters measured. In a comparison of the two marker techniques used, fluorescent ELISA was less sensitive than the antibiotic resistance technique for the identification of the occupants of small ineffective nodules. The use of both techniques in large field experiments is discussed. Clover establishment was affected by lime level during the establishment phase but the beneficial effect of lime had disappeared by the second harvest year. Experimental siteThe experiment was conducted at Pant-y-dwr, Powys (Welsh Plant Breeding Station Hill Research Station). A uniform area 50 X 30m supporting a Molinia caerulea dominant vegetation was 'flail-mown' during March 1981 and the site was fenced to exclude straying animals. The peaty grey podzolic soil (pH 3.9-4.2) was tested for indigenous rhizobia using the 'most probable number assay' (Vincent, 1970) anti by direct 'baiting' of lime amended soil (pH 6.0) using six clover varieties. In neither test were nodules formed on the clover root systems indicating the complete absence of R. trgolii in the soil. An application of N, P205, K20 9:25:25 (ICI No. 3) at a rate of 153 kg ha-l was uniformly spread over the mown area. This was followed by ground limestone (applied by hand) at the rates of 2 and 4 t ha-1 during the month of May. Paths 1 m wide were left unlimed between the 1 . 5 X 1 m plots to

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