Biological control of onion white rot using Trichoderma harzianum

Abstract

The biological control of Sclerotium cepivorum, the causal agent of onion white rot, by Trichoderma harzianum and T. virens was studied in a glasshouse trial. T. Harzianum (isolates C52 and D73) and T. virens (isolate GV4) afforded early season disease control when applied to the soil as a bulk carrier (sand:bran:fungal mix, 1:1:2) at the time of planting. T. harzianum C52 gave extended control to 12 weeks after planting, but by the end of the trial, disease pressure was extreme (98%) and none of the treatments gave continuous control. Similar results were found in the field, where T. harzianum C52 was unable to control onion white rot disease under high disease pressure (87%).

The results of these disease control trials suggested that T. harzianum may be better applied as part of an integrated disease management programme rather than alone. Ecological studies were undertaken with T. harzianum C52 and S. cepivorum to optimise application strategies. T. harzianum C52 was both rhizosphere and rhizoplane competent when introduced into the soil as a seed coat (0.2 x 10¹ to 1.8 X 10² cfu/g soil) and a bulk carrier (7.5 x 10³ to 3.6 X 10⁵ cfu/g soil) in a 16 week pot trial. Under field conditions, the addition of commercial formulations of T. harzianum C52 (Trichopel and Trichodry) prepared by Agrimm Technologies Ltd. to the planting furrow enabled T. harzianum C52 to establish in the rhizosphere. Proliferation in the rhizosphere was formulation dependent with Trichopel maintaining concentrations greater than 10⁵ cfu/g soil for the majority of the 19 week trial compared with 10³ cfu/g soil for the Trichodry formulation.

A molecular marker was developed using the molecular biology technique UP-PCR. Initially, genetic variation between T. harzianum and Trichoderma isolates was studied and a band profile was identified for T. harzianum C52 that enabled this isolate to be distinguished from other Trichoderma isolates present in the soil. The combination of unique morphological characteristics, genetic fmgerprinting and selective sampling of the rhizosphere allowed T. harzianum isolates recovered at the field site to be identified with a 95% degree of certainty.

A complementary study examined the delivery of T. harzianum C52 in transplant potting mix. T. harzianum C52 successfully colonised the rhizosphere of onion transplants when Trichodry was incorporated with potting mix and populations were similar to those achieved in the field with a direct seeded crop (10³ cfu/g soil).

In vitro assays were conducted to determine possible modes of action of T. harzianum in the field. T. harzianum C52 significantly reduced the mycelial growth of S. cepivorum colonies in dual culture and prevented three of the four S. cepivorum isolates tested from producing sclerotia. Competition was determined as a likely mode of action for T. harzianum C52. Previous studies indicated that antibiosis was also a mode of action as this isolate was shown to produce the antimicrobial metabolite six-pentyl-a-pyrone (6P AP) (Dodd-Wilson, 1996; McLean, 1996). Therefore, it was postulated that T. harzianum C52 controls onion white rot by colonising the rhizosphere, out-competing S. cepivorum for nutrients and space and by producing 6PAP, to reduce S. cepivorum mycelial growth.

S. cepivorum preferential infection sites on onion roots were studied to determine if the planting furrow was optimum for delivering T. harzianum to soil for root protection. Sclerotia banded at 1, 10 and 20 cm depths in soil germinated and infected onion roots at 5, 7 and 13 weeks after planting, respectively. A second trial determined that the severity of infection varied when sclerotia were placed at the stem base, along the length of a root and at the root tip. A significantly greater number of stem base infections led to total plant collapse compared with infections occurring along the length of a root or at the root tip. As the stem base infections would contribute the most to yield losses, targeting T. harzianum to the planting furrow should protect the stem base from infection.

The fungicide sensitivity of T. harzianum C52 was studied to determine the compatibility of this fungus with fungicides routinely used on onion crops. In vitro assays determined that T. harzianum C52 mycelium was highly sensitive to half, normal and twice the recommended field rate concentrations of captan, benomyl, thiram, iprodione and carbendazim. The spore sensitivity assay showed that 100% of the T. harzianum C52 spores were prevented from germinating when exposed to mancozeb, copper oxychloride, tebuconazole, thiram and the combination of mancozeb and metalaxyl. A selection of the most commonly used fungicides were applied to T harzianum C52 infested soil. All fungicides initially reduced the T harzianum C52 concentration (2.2 x 10³ - 3.4 X 10⁴ cfu/g soil), but numbers quickly recovered within 12 days except for mancozeb formulations, where the population took longer (30 days) to recover. These results indicate that T harzianum would be compatible with most fungicides used on onion crops, except thiram and mancozeb.

The use of soil solarisation was considered as part of an integrated disease management programme to reduce sclerotial viability and, thereby, relieve disease pressure. Eight week solarisation trials in Canterbury and Blenheim reduced sclerotial viability to 31.5 and 0%, respectively, when sclerotia were contained in polyester mesh bags. The addition of a mixture of soil and sand to the bags with the sclerotia further reduced sclerotial viability to 8.7% in Canterbury. The combination of sub-lethal temperatures and lethal temperatures for short periods were responsible for the decrease in sclerotial viability. Soil temperature data from New Zealand's three main onion growing regions indicated that soil solarisation would be applicable in all regions to relieve disease pressure.