P. G. Long

Ethylene production by Botrytis cinerea

Abstract Ethylene was produced when isolates of the postharvest pathogen Botrytis cinerea Pers.: Fr., derived from fruit of strawberry, blueberry and kiwifruit and leaves of grape and camellia, were grown on a modified Pratts medium containing 35 mM methionine in shaken or static cultures at 22 °C in the dark. Cultures grown on basal media containing glutamate or α-ketoglutarate produced no more ethylene than controls. Optimum growth occurred at pH 3.5 and 4.5 for shake and static cultures, respectively. When B. Cinerea was grown in a methionine-amended basal medium, maximum production of ethylene occurred after 3–4 days of incubation. However, maximum ethylene production per unit dry wt of mycelium (780 μl/g/h) occurred within 48 h of inoculation, after which it declined. That high ethylene production occurs with such small amounts of mycelia suggests a possible role for fungal produced ethylene in B. cinerea pathogenesis of sensitive fruit such as kiwifruit.

The combined effect of delayed application of yeast biocontrol agents and fruit curing for the inhibition of the postharvest pathogen Botrytis cinerea in kiwifruit

Abstract Application of biocontrol agents (BCAs) and use of induced host resistance for the inhibition of pathogen infection have often been examined separately. The present study focused on application of yeast BCAs alone and the interaction between kiwifruit curing and BCA application for inhibition of infection by the pathogen Botrytis cinerea. Kiwifruit pedicels were removed at the natural abscission layer and the pedicel wound was used to evaluate efficacy of up to five yeast candidates, applied at increasing delay intervals after B. cinerea challenge. Other fruit were treated with combinations and various sequences of fruit curing (incubation at 10°C) and topical yeast application. All yeast candidates conferred a significant level of biocontrol following applications made simultaneously with, or up to 96 h after B. cinerea inoculation. Biocontrol activity was further increased with an additive effect of BCA and fruit curing combined but only when BCA application was made after 96 h of fruit curing. These results suggest that a degree of protection of the kiwifruit could be achieved with the application of the yeast to the pedicel wound. If kiwifruit curing is initiated, host resistance mechanisms may not be specific to the pathogen, B. cinerea, since the effect of epiphytic microbes used for biocontrol appear to be similarly reduced by factors induced in the first 24–48 h of curing.

Thaumatin‐like protein in kiwifruit

A 21-kDa protein extracted from woody tissue underlying the pedicel of kiwifruit (Actinidia deliciosa var deliciosa [A Chev] Liang and Ferguson) was purified to homogeneity by cation exchange, gel filtration, sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) on a 15% gel and electro-transfer onto a polyvinylidene difluoride (PVDF) membrane, followed by excision of the band of interest. Amino terminal sequence analysis of this band revealed a high degree of homology with basic endochitinase (EC 3.2.1.14) in orange and thaumatin-like proteins in tobacco, barley, maize and tomato. This is the first record of a thaumatin-like protein in a subtropical berryfruit. Potential implications of this finding and the need for future research are discussed. © 1999 Society of Chemical Industry

New genetic markers for Botrytis cinerea ( Botryotinia fuckeliana )

Selenate-resistant (SelR) mutants of Botrytis cinerea were selected by plating conidia or mycelial plugs onto minimal medium amended with selenate and taurine. Ultraviolet irradiation of conidia to give 5% survival increased mutant yield twenty-fold. Mutants could be divided into three classes based on growth in the presence of selenate or chromate and on response to taurine in minimal media. Nitrate non-utilizing (Nit) mutants, generated as spontaneous sectors on minimal media amended with chlorate, behaved as nit1 mutants in growth tests (i.e. putatively defective in nitrate reductase apoenzyme). When nit1 mutants were paired on medium with nitrate as sole nitrogen source some pairings complemented, behaviour attributed to intragenic complementation. Selected crosses of SelR and nit1 mutants with wild-type strains gave 1:1 segregation of both phenotypes and no evidence of linkage to either Mbc1 (benzimidazole resistance) or Daf1 (dicarboximide resistance) markers; loose linkage was confirmed between Mbc1 and Daf1. Strains showing the SelR phenotype may result from mutations in different genes; the genotypic symbol Sel1 is allocated to one of these mutations. Four-point crosses involving the markers at Sel1, nit1, Mbc1 and Daf1 generated progeny with all 16 genotypes. Both Sel1R- and nit1 mutants were stable following subculture and retained pathogenicity in a bean seedling assay. Complementation was demonstrated between SelR and nit1 mutants selected from the same parent.

Curing of kiwifruit for control of postharvest infection by Botrytis cinerea

Abstract The effects of temperature and humidity during curing on subsequent infection levels of B. cinerea in kiwifruit were investigated in 1993 and 1994. After harvest, each fruit was inoculated with 25 000 spores in a 17 μl droplet of Tween 20. Dry conidial application was also included in 1994 experiments. The largest curing effect was obtained at 10°C. Disease incidence was highest at 0°C and the curing effect diminished at temperatures above 10°C. In 1994 a three day curing period was used and 10°C gave the lowest subsequent disease incidence. After twelve weeks of cool storage (1993) there was less disease in fruit cured at 89–95% relative humidity than at lower humidities. In 1994, comparable results were obtained. Firmness fluctuated with harvest and in general, decreased with storage, although a satisfactory firmness was maintained throughout cool storage for all treatments. For all experiments, weight loss increased with increased curing temperature or with decreased relative humidity.

Use of fungicides to assess the effects of root disease: Effects of prochloraz on red clover and microbial populations in soil and roots

Abstract Benomyl and six demethylation inhibiting (DMI) fungicides were assessed in Petri plate tests for their suitability for use in field studies on the effects of pathogenic soilborne fungi on the growth of red clover ( Trifolium pratense L.). Prochloraz was selected for glasshouse pot experiments because it had the greatest activity against seven fungi isolated from diseased red clover roots, and was the least inhibitory of the DMI fungicides to seedling growth. Pots of naturally-infested soil were given a single drench of prochlorazat the time of sowing red clover, then maintained at constant soil temperature (15, 20 or 25°C). After 4 weeks, the total populations of fungi in soil and in the roots of plants from pots treated with prochloraz at concentrations of, or exceeding. 80 mg prochloraz kg −1 dry soil (at 15°C) or 240mg kg −1 (at 20 and 25°C). were lower than those from untreated controls. However, emergence and growth of red clover was reduced, and total soil bacterial populations were increased, in soil from pots treated with high concentrations of prochloraz (730 and 2200 mg kg −1 ). Growth of red clover was also reduced in prochloraz-trcatcd soil which had been disinfcsted by microwave treatment. Provided that it is used at an appropriate rate, and that effects on target and non-target organisms are monitored, prochloraz appears useful for field studies on the effects of root discuses on growth of forage legumes.

Review of literature on camellia flower blight caused by Ciborinia camelliae

Abstract Ciborinia camelliae Kohn is the most destructive pest or disease problem of camellias (Camellia spp.) The pathogen is related to common and widespread plant pathogens in the genera Sclerotinia and Botrytinia (anamorph=Botrytis). Sclerotia form in infected petals and remain dormant in plant debris until the next season. In early spring, apothecia are produced from the sclerotia and release windborne ascospores. Infection causes the petals to turn brown and the flowers to fall prematurely. The disease has been identified in Japan (1919), the United States (1938), New Zealand (1993), and parts of Europe (1999). It has now spread over the lower North Island and upper South Island of New Zealand, with isolated outbreaks in Christchurch and Auckland. Control of this disease has proved difficult even though: (1) only camellia flowers are infected, (2) there is no secondary infection, and (3) ascospores are present for only 2–3 months each year. To date, fungicides have given less than satisfactory control of the disease and possible control measures are reviewed. Interest in potential biocontrol agents is growing but remains an unexplored alternative. Resistant varieties offer the best management option for the future.

Competition between aggressive and non-aggressive strains of Botrytis cinerea (Botryotinia fuckeliana) on French bean leaves

Competition was assessed between aggressive and non-aggressive strains of Botrytis cinerea on French bean (Phaseolus vulgaris) leaves by inoculating strains simultaneously and sequentially (6, 12 or 24 h later). The lesions produced when an aggressive strain (forming large lesions) and a non-aggressive strain (forming small, restricted lesions) were inoculated simultaneously were intermediate in size between those produced by the strains when inoculated individually. suggesting that the two strains were co-existing in the lesion. However, in sequential inoculation when the second inoculation was delayed 6 h or more, lesion sizes typical of the first inoculated strain were found indicating that the first strain to be inoculated dominated the infection site. Competition between two aggressive strains, individually identifiable by genetic markers, was found to follow a similar pattern, suggesting that the dominating effect of the first strain is a general phenomenon. Domination of the lesion site by the nonaggressive strain prevented an aggressive strain from forming spreading lesions suggesting a possible use for these strains as a biocontrol agent in some disease situations.

The potential for resistance to Botrytis cinerea by kiwifruit

Abstract Botrytis cinerea (Pers.: Fr.) is the major post-harvest pathogen of kiwifruit ( Actinidia deliciosa var. deliciosa cv. Hayward (A. Chev.) C.F. Liang et A.R. Ferguson). It causes the disease commonly known as stem-end rot or grey mould. Current control of this pathogen by pre-harvest fungicide application is inadequate due to fungicide resistance and an inability to target the infection site created at the point of harvest. Host resistance represents a more environmentally sound approach, but requires long-term research to realise its full potential. In this review, components of host resistance are described and their relevance to the kiwifruit- B. cinerea interaction is discussed to highlight those areas which merit further study.

Fungal invasion of red clover roots in a soil naturally infested with a complex of pathogens : effects of soil temperature and moisture content

Abstract A quantitative method employing tissue maceration and plating was used to determine the internal fungal flora of roots of red clover ( Trifolium pratense ) plants, 4, 8 and 12 weeks after sowing in field plots, and in pots of field soil kept at different constant temperatures (10. 15. 20 or 25°C) and moisture contents (40, 60 or 80% of soil water holding capacity) in the glasshouse. In field plots, the total number of colonies isolated per root increased at each successive harvest whereas the extent of fungal invasion (number of fungal colonies g 1− root tissue) decreased. A similar decrease in the invasion of roots of plants in pots reflected a slow rate of fungal infection and an accelerating production of lateral roots. Plant growth in pots was greatest at 25 C and 60% WHC but slowed in most treatments between 8 and 12 weeks as plants became potbound. The fungi which dominated at 4 weeks ( Verticillium dahliae and Fusarium solani ) were isolated most commonly from plants grown at 10°C. while at 12 weeks most colonies of the dominant fungi ( Cylindrocladiun scoprium, F. oxysporum, F. solari, and Trichocladium basicola ) were obtained from plants grown at 20 and 25 C. Roots from soil at high moisture content (80% WHC) yielded more fungal colonies in total than those from less moist soil at 4 and 12 weeks, however, during the first 8 weeks, F. solani was most often recovered from dry soil (40% WHC). Since the root mycoflora of field-grown and pot-grown plants was similar, choice of appropriate soil temperature and moisture conditions for glasshouse experiments would allow simulation of events in the field.