Evaluation of an antagonistic Trichoderma strain for reducing the rate of wood decomposition by the white rot fungus Phellinus noxius

Biological Control 61 (2012) 160–168

Contents lists available at SciVerse ScienceDirect

Biological Control journal homepage: www.elsevier.com/locate/ybcon

Evaluation of an antagonistic Trichoderma strain for reducing the rate of wood decomposition by the white rot fungus Phellinus noxius Francis W.M.R. Schwarze a,⇑, Frederick Jauss a, Chris Spencer b, Craig Hallam b, Mark Schubert a a

EMPA, Swiss Federal Laboratories for Materials Science and Technology, Wood Laboratory, Section Wood Protection and Biotechnology, Lerchenfeldstrasse. 5, CH-9014 St. Gallen, Switzerland b ENSPEC, Unit 2/13 Viewtech Place, Rowville, Victoria 3178, Australia

h i g h l i g h t s

g r a p h i c a l a b s t r a c t

" Antagonism of Trichoderma species against Phellinus noxius varied in the in vitro studies. " Weight losses by P. noxius were higher in angiospermous than gymnospermous wood. " Biocontrol of P. noxius depends on the specific Trichoderma strain and its host.

a r t i c l e

i n f o

Article history: Received 31 October 2011 Accepted 30 January 2012 Available online 8 February 2012 Keywords: Biological control White rot Ganoderma australe Dry weight loss Interaction tests in wood blocks

a b s t r a c t The objective of these in vitro studies was to identify a Trichoderma strain that reduces the rate of wood decomposition by the white rot fungus Phellinus noxius and Ganoderma australe. For this purpose, dual culture and interaction tests in wood blocks of three hardwoods, Delonix regia, Ficus benjamina, Jacaranda mimosifolia, and one softwood, Araucaria bidwillii, as well as investigations of fungal growth under different environmental conditions, were performed. The effect of Trichoderma ghanense, two strains of T. harzianum and T. reesei on wood colonization and decomposition by four P. noxius strains and G. australe were quantitatively analyzed by measuring the dry weight loss of wood. All Trichoderma species and wood-decay fungi showed optimum growth at a mean temperature of 25–35 °C and a high water activity (aw) of 0.998. At 35 °C and aw 0.928, no growth was recorded for any of the wood-decay fungi after 1 week, whereas most Trichoderma species were still actively growing. The different Trichoderma species all showed an antagonistic potential against P. noxius in the in vitro studies. The species of wood-decay fungi showed significant differences in their sensitivity when challenged by the volatile organic compounds (VOCs) of Trichoderma species. Reduction in the rate of wood decomposition by different Trichoderma species against all wood-decay fungi varied strongly according to the specific plant host. T. harzianum 121009 and T. atroviride 15603.1 showed the highest reduction in weight losses. P. noxius 169 strongly decomposed untreated and pretreated wood of D. regia, whereas weight losses of F. benjamina and J. mimosifolia pretreated with Trichoderma strains were significantly lower. Weight losses by G. australe were significantly reduced for A. bidwillii, D. regia and F. benjamina by all Trichoderma species, but no affect was recorded for J. mimosifolia. The in vitro studies show that only after careful monitoring (i.e. selecting the appropriate strain for the target pathogen and its niche (wood species) can Trichoderma species be used to significantly reduce the growth and rate of wood decomposition by different P. noxius strains. Ó 2012 Elsevier Inc. All rights reserved.

⇑ Corresponding author. Fax: +41 582747694. E-mail address: [email protected] (F.W.M.R. Schwarze). 1049-9644/$ - see front matter Ó 2012 Elsevier Inc. All rights reserved. doi:10.1016/j.biocontrol.2012.01.016

F.W.M.R. Schwarze et al. / Biological Control 61 (2012) 160–168

1. Introduction Species of the genus Trichoderma are ubiquitous in soils. Since Weindling (1932) recognized the antagonistic effect of Trichoderma species against plant pathogens, several have been extensively studied as biological control agents against fungal pathogens (Chet, 1990; Chet et al., 1998; Harman et al., 2004; Howell, 1998). As potential biocontrol agents, Trichoderma species have the following advantages: they grow on, but do not occur on, most organic matter, sporulate readily in culture and, in natural conditions, can act as either secondary antagonists or primary colonizers (Alabouvette et al., 2006; Highley, 1997; Holdenreider and Greig, 1998; Schoeman et al., 1999). Furthermore, some Trichoderma species survive as chlamydospores under unfavourable conditions and are fairly resistant to common fungicides and herbicides (Sariah, 2003). Much of the biological control research in the tropics has focused on the development of strains of Trichoderma species (subdivision: Ascomycota) that show antagonistic activity against fungal root fungal pathogens (Harman et al., 2004; Prasad and Naik, 2002; Raziq and Fox, 2006; Sariah, 2003; Sariah et al., 2005; Susanto et al., 2005; Widyastuti, 2006). Soepena et al. (2000) successfully formulated a biofungicide comprising Trichoderma koningii Oud. isolate Marihat (MR14) to manage basal stem rot in Elaeis guineensis (Jacq.) (oil palm) caused by Ganoderma orbiforme (Fr.) Ryvarden (=G. boninense Pat.). This pathogen is recognized as the single major disease constraint to sustainable production of oil palm throughout Asia (Ariffin et al., 2000; Durand-Gasselin et al., 2005; Flood et al., 2000; Paterson et al., 2000; Singh, 1991; Turner, 1981). In in vitro experiments, the growth of two unknown Ganoderma species, previously isolated from diseased Acacia mangium in Indonesia, were shown to be strongly inhibited by T. koningii, T. harzianum and T. reesei (Widyastuti, 2006). In field experiments carried out at different locations in France and Germany, a total of 159 angiospermous trees and 1431 wounds on six different species (Platanus  hispanica Miller ex Münchh., Acer pseudoplatanus L., Tilia platyphyllos Scop., Populus nigra L., Quercus rubra L., Robinia pseudoacacia L.) were treated with different conidial suspensions of T. atroviride strain T-15603.1 (Schubert et al., 2008a–c). T15603.1 significantly suppressed growth (82.3%) in wounds artificially inoculated with three basidiomycetes Ganoderma adspersum, Inonotus hispidus and Polyporus squamosus in comparison with growth in untreated control wounds (Schubert et al., 2008b). Although these studies show that Trichoderma species can be successfully used against wood-decay fungi, to our knowledge little research on the biological control of the basidiomycete Phellinus noxius (Corner) G.H. Cunningham exists (Bolland et al., 1988). This white-rot fungus was first described in 1932 as Fomes noxius by Corner (1932), who was investigating the cause of a brown root rot disease of trees in Singapore. It was reclassified as P. noxius by Cunningham (1965). The devastating brown root rot disease affects a wide variety of important agricultural and forest plant species (Ann et al., 2002). Many early reports of root rot caused by P. noxius were made without demonstration of pathogenicity. In 1984, Bolland was the first to fulfill Koch’s postulates for the disease on hoop pine (Araucaria cunninghamii Aiton ex D. Don.). An inoculation technique for pathogenicity tests was described by Ann et al. (2002). P. noxius is a basidiomycete with a pan-tropical/subtropical distribution, and has been found in Africa, Asia, Australia and Oceania, Central America, and the Caribbean. The pathogen has a wide host range, spanning over 100 genera of Gymnospermae and both classes (Monocotyledons and Dicotyledons) within the Angiospermae. Cross-inoculation studies have shown a lack of host specificity for various strains of P. noxius, although varying degrees of resistance are seen in different hosts (Ann et al., 1999; Chang, 1995, 2002;

161

Mohd Farid et al., 2009; Sahashi et al., 2010; Wu et al., 2011). The USDA-ARS Systematic Botany and Mycology Laboratory maintains a website with an updated list of hosts and information on geographical distribution; it currently lists 153 host species and some of the most notable include mahogany (Swietenia mahagoni King.) teak (Tectona grandis L.), rubber (Hevea brasiliensis (Willd. ex Adr de Juss.) Muell. et Arg.), oil palm (Elaeis guineensis Jacq.), tea Camellia sinensis (L.) Kuntze, coffee (Coffea canephora Pierre ex A. Froehner) and cacao (Theobroma cacao L.), as well as a variety of fruit, nut, and ornamental trees. The fungus is believed to be responsible for the death of many Hoop Pines (A. cunninghamii), and other trees throughout Australia (Bolland, 1984). The disease is highly invasive and has already killed many significant trees throughout the Greater Metropolitan area of Brisbane Queensland, including street and park trees located in the suburbs of Shorncliffe, Taringa, New Farm, Eagle Farm, West End, the city’s centre and beside the Brisbane River. As the demand for alternatives to chemical control of plant pathogens has become stronger, owing to concerns about the safety and environmental impact of chemicals, the application of Trichoderma as biological control of P. noxius shows promise. The objective of this study was to evaluate the potential of different Trichoderma species as biocontrol agents and to identify a competitive strain that can be used against P. noxius. For this purpose, a range of bioassays were conducted to evaluate the antagonistic mechanisms of different Trichoderma species against P. noxius and Ganoderma australe (Fr.) Pat. an important decay fungus on urban trees in Australia. As a benchmark, we compared the European T. atroviride strain 15603.1, which has been shown to have high biocontrol efficacy against several wood-decay fungi (Schubert et al., 2008a–c), with four Australian native Trichoderma strains (T. ghanense, T. reesei, and two strains of T. harzianum). 2. Materials and methods 2.1. Micro-morphological and molecular identification All cultures were identified microscopically (Bissett, 1984; 1991a–c, 1992; Gams and Bissett, 1998; Rifai, 1969) and additionally the internal transcribed spacer (ITS) 1-5.8S-ITS2 region of the rDNA was amplified and sequenced for each strain (Schubert, 2006). The origins of the Trichoderma strains and wood-decay basidiomycetes are provided in Table 1. All cultures were maintained on 2% malt extract agar (MEA) at 4(±1) °C. For further studies, Petri dishes with MEA were inoculated with 5 mm diameter agar plug cut from the growing edge of colonies of the strains and stored in the dark at 25 (±1) °C and 70% ambient relative humidity. The sequences were deposited in the EMBL Data Bank (Table 1). 2.2. Growth of Trichoderma species and wood-decay fungi under different conditions The effects of temperature (20 °C, 25 °C, 30 °C, 35 °C) and water activity (aw: 0.928, 0.955, 0.978, 0.998) on hyphal growth were monitored on 2% MEA. All agar plates (90 mm) were inoculated centrally with a 5-mm disc of the respective Trichoderma species and wood-decay fungi taken from the margin of growing cultures and incubated at 25(±1) °C and 70% relative humidity. For each experimental treatment (aw and temperature), 10 replicates were performed. The growth rate (mm/day) was determined by colony diameter measurements carried out along two perpendicular axes after 24 h (Schubert et al., 2009). The aw of the substrate was controlled by the addition of appropriate weights of the non-ionic solute, glycerol, prior to autoclaving (Dallyn, 1978).

162

F.W.M.R. Schwarze et al. / Biological Control 61 (2012) 160–168

Table 1 Origin of Trichodermaspeciess and wood-decay fungi used in the present study. Trichoderma T. atroviride Karsten T. ghanense Yoshim. Doi, Y. Abe and Sugiy. T. harzianum Rifai T. reesei E.G. Simmens T. harzianum

Strain No. T-15603.1 T-4510b

EMBL No. a

T-4428c T-3967d T-121009e

FR178524 FR821772 FR821773 FR821774 FR821767

Wood-decay fungi Phellinus noxius (Corner) G.H. Cunningham P. noxius P. noxius P. noxius Ganoderma australe (Fr.) Pat.

Strain No.

EMBL No.

f

FR821775

g

FR821769 FR821770 FR821771 FR821768

133

144 169h 178f 101009i

a

From EMPA culture collection, Switzerland; isolated from a basidiocarp of Armillaria mellea (Vahl) P. Kumm., Freiburg, Germany. From Commonwealth Scientific and Industrial Research Organisation (CSIRO) culture collection, North Ryde New South Wales (N.S.W.) Australia; isolated from soil, North Carolina, USA. c From CSIRO culture collection, isolated from wooden flooring of shipping container, Sydney, N.S.W. d From CSIRO culture collection; isolated from mutant of ATCC 24449, e From Mt. Coot-tha, Queensland, Australia; isolated from a basidiocarp of Laetiporus portentosus (Berk.) Rajchenb. f Provided by Dr. Geoff Pegg, Department of Employment, Economic Development and Innovation (DEEDI), Brisbane, Queensland, Australia, g From New Farm Park, Queensland Australia; isolated from basidiocarp of Ganoderma australe on Jacaranda mimosifolia D. Don, h From New Farm Park, Queensland Australia; isolated from a basidiocarp of mature Phellinus noxius on a dead Ficus species, i From City Botanic Gardens, Brisbane, Queensland, Australia; isolated from basidiocarp of Ganoderma spp. on mature Ficus microcarpa var. hillii (F.M. Bailey) Corner. b

2.3. Inhibitory effects of volatile compounds produced by Trichoderma species on wood-decay fungi The effect on wood-decay fungi of volatile organic compounds (VOCs) produced by Trichoderma strains was evaluated with the following techniques as described by Dennis and Webster (1971). Trichoderma strains were centrally inoculated onto 2% MEA by placing 5-mm discs taken from the margin of 7-day-old cultures and then incubating the plates at 25(±1) °C and 70% relative humidity for 3 weeks. MEA plates were inoculated centrally with 5-mm discs with the wood-decay fungi and then the top of each plate was replaced with the bottom of a Trichoderma-inoculated plate. Replicates without Trichoderma species were used as the control. Ten replicates were maintained for each treatment. The pairs of Petri dishes were fixed and sealed together with Para film and incubated at 25(±1) °C and 70% relative humidity. The diameter of the wood-decay fungi colonies was measured after an incubation period of 7 days and the inhibition of mycelial growth was calculated. 2.4. Dual culture and interaction tests on wood Mycoparasitism of all Trichoderma strains against the selected wood-decay fungi was assessed in dual culture according to the method of Schubert et al. (2006). The agar disc method was carried out on 2% MEA. Mycelial discs (5 mm) were removed from 1-weekold MEA cultures of each of the five wood-decay fungi and placed equidistantly at the margin of Petri dishes (90 mm) containing 2% MEA. These were incubated at 25(±1) °C and 70% relative humidity for 3–4 days. Next, discs (5 mm) were removed from the margins of actively growing 1-week-old cultures of the Trichoderma species and placed at opposite sides of the dish, and incubated in the dark at 25(±1) °C and 70% relative humidity for 4 weeks. Petri dishes without Trichoderma were used as the control. Twenty replicates were used for each experiment. Petri dishes were examined at regular intervals. The sporulation tufts and pustules of Trichoderma fungi were used as an indication of its activity (Naár and Kecskes, 1998). In order to check whether the antagonist was able to overgrow and parasitize the challenged wood-decay fungus, three agar discs (5 mm) were removed from non-sporulating regions of the mycelium of the wood decay fungus and placed on a Trichoderma-selective medium (Askew and Laing, 1993). After 7 days of incubation at room temperature, discs were observed for Trichoderma colonies. Competition (mycoparasitism rate) was assessed as follows: 0 = no overgrowth; 1 = slow overgrowth; 2 = fast overgrowth; 3 = very fast overgrowth and deadlock of the wood-decay

fungi. The ability of Trichoderma species to eliminate the wood-decay fungi (lethal effect) during the four week incubation period was evaluated by aseptically transferring 5-mm discs from test plates to MEA with 2 mL thiabendazole (2-(40-thiazolyl)- benzimidazole; Merck, Darmstadt, Germany; 0.46 mg dissolved in lactic acid). T-MEA suppresses the growth of Trichoderma species but allows growth of wood-decay fungi (Sieber, 1995). The lethal effect of Trichoderma species was expressed as a percentage of the wood-decay fungi which were eliminated. In addition, interaction tests in sap-wood blocks (Ave. 5  25  40 mm) of three hardwoods, Delonix regia (Boj. ex Hook.) Raf., Jacaranda mimosifolia D. Don, and Ficus benjamina L., and one softwood, Araucaria bidwillii (Molina) K. Koch, were performed as described by Schubert et al. (2008a). For studies of colonization behaviour, wood blocks were inoculated with a conidial suspension of Trichoderma species (colony-forming units: 105/mL + 0.2% D-glucose + 0.1% urea) and placed onto Petri dishes with 2% MEA. After the wood blocks were completely colonized by Trichoderma they were placed with their cross sections onto 2-week-old cultures of the wood-decay fungi and incubated in the dark at 25(±1) °C for 12 weeks. Untreated wood blocks served as the control. Twenty replicates were used for each experiment. Before incubation wood blocks were ovendried at 105 °C for 24 h to determine the wood dry weight. The decay tests were run for 12 weeks at 25 °C, after which the wood blocks were removed, cleaned and oven-dried for measuring dry weight losses (Schwarze and Fink, 1998). 2.5. Statistical analysis Growth data were log-transformed and data that were expressed as percentages, such as the wood weight loss, were arcsine-transformed prior to analysis (ANOVA) and back-transformed to numerical values for presentation (expressed as mean ± SE). Means were separated using Dunett’s test at significance levels of p < 0.05 and p < 0.0001. To compare the performance of the Trichoderma isolates with each other a Tukey’s HSD (Honestly Significant Difference) test (P < 0.05) was additionally performed. The statistical package used for all analyses was SPSSÒ (Version 17.0, SPSS Inc., Chicago, IL, USA). 3. Results 3.1. Growth under different conditions The influence of temperature and aw on the mean growth rates of the Trichoderma species and wood decay fungi is shown in

163

F.W.M.R. Schwarze et al. / Biological Control 61 (2012) 160–168

Tables 2 and 3, respectively. Generally, the growth rates of all Trichoderma strains increased with increasing aw (Table 2). T4510 showed the strongest growth at a mean temperature of 35 °C and at the highest aw of 0.998 (Table 2). T-3967 and T121009 showed an optimum growth at a mean temperature of 30 °C and at the highest aw of 0.998, whereas the European strains T-15603.1 and T-121009 showed optimum growth at a mean temperature of 25 °C and the highest aw of 0.998. Even at 35 °C and the lowest water activity (aw 0.928), growth was recorded for some Trichoderma species. P. noxius strains 144, 169 and 178 showed optimum growth at a mean temperature of 25–30 °C and a high aw of 0.998 (Table 3). By contrast, P. noxius 133 showed optimum growth at a mean temperature of 30 °C and moderate aw of 0.978. At the lowest aw (0.928) and at 25 °C and 30 °C, growth was only limited, and at 35 °C growth of the P. noxius strains failed completely. G. australe showed optimum growth at a mean temperature of 30 °C and high aw of 0.998, but no growth was recorded at any temperature at the lowest aw (0.928). 3.2. Evaluation of antagonistic activity in dual cultures During initial screening of the Trichoderma strains, a variety of reactions were recorded as a result of antagonism. Growth of all wood-decay basidiomycetes was inhibited by at least one of the Trichoderma strains. Contact between wood-decay fungi and Trichoderma species occurred, but the ability to overgrow and parasitize the mycelia of the wood-decay fungi was highly dependent on the antagonistic potential of each Trichoderma strain and the resistance of the challenged wood-decay fungus to antagonism (Table 4). The lethal effect of mycoparasitism by Trichoderma species was most prevalent for T. harzianum 121009. It showed a high antagonistic

Table 2 Mean growth rate (mm/day) of the Trichoderma species under different conditions. Temperature (°C)

Water activity (aw) 0.998

0.978

0.995

T. atroviride 15603.1 20 25 30 35

0.928

8.3 ± 1.3 10.6 ± 0.7 8.8 ± 0.5 0.0 ± 0.0

4.6 ± 0.3 6.1 ± 0.5 6.2 ± 0.4 0.0 ± 0.0

2.3 ± 0.2 2.9 ± 0.3 2.1 ± 0.4 0.0 ± 0.0

0.2 ± 0.1 0.4 ± 0.1 0.0 ± 0.0 0.0 ± 0.0

T. ghanense 4510 20 25 30 35

9.0 ± 1.3 11.8 ± 2.9 16.1 ± 1.6 18.1 ± 1.6

5.2 ± 0.4 7.8 ± 1.1 8.9 ± 0.5 9.1 ± 0.5

1.9 ± 0.6 3.5 ± 0.6 3.9 ± 0.4 3.5 ± 0.4

0.1 ± 0.1 0.6 ± 0.9 0.9 ± 0.2 0.3 ± 0.1

T. harzianum 4428 20 25 30 35

8.6 ± 0.7 12.0 ± 0.6 8.1 ± 0.5 0.0 ± 0.0

5.4 ± 0.7 7.6 ± 1.2 6.4 ± 0.5 0.1 ± 0.0

2.2 ± 0.3 2.9 ± 0.3 2.7 ± 0.5 0.0 ± 0.0

0.3 ± 0.1 0.7 ± 0.2 0.2 ± 0.2 0.00 ± 0.0

T. reesei 3967 20 25 30 35

10.7 ± 0.4 11.7 ± 3.1 13.7 ± 0.4 11.7 ± 0.4

3.5 ± 0.4 7.3 ± 0.4 9.7 ± 0.5 9.7 ± 0.5

1.1 ± 0.2 1.7 ± 0.3 3.1 ± 0.5 3.1 ± 0.5

0.1 ± 0.1 0.5 ± 0.2 0.3 ± 0.2 0.3 ± 0.2

T. harzianum 121009 20 25 30 35

13.0 ± 1.7 11.8 ± 1.9 14.4 ± 1.4 6.3 ± 0.8

4.5 ± 0.3 6.7 ± 0.5 8.7 ± 0.5 4.1 ± 0.7

1.5 ± 0.2 2.6 ± 0.2 3.5 ± 01.0 1.1 ± 0.2

0.1 ± 0.1 0.2 ± 0.2 0.2 ± 0.2 0.0 ± 0.0

For each experimental treatment (water activity aw and temperature), 10 replicates were performed. The growth rate (mm/day) was determined by colony diameter measurements carried out along two perpendicular axes after 24 h. The aw of the substrate was controlled by the addition of appropriate weights of the non-ionic solute, glycerol, prior to autoclaving.Data were analyzed by analysis of variance (ANOVA) to assess the effect of aw and temperature on growth rate (mm/day). ±, Standard deviation.

Table 3 Mean growth rate (mm/day)of the Phellinus noxius strains and Ganoderma australe under different conditions. Temperature (°C)

Water activity (aw) 0.998

0.978

0.995

0.928

P. noxius 133 20 25 30 35

5.5 ± 0.9 7.7 ± 1.0 7.8 ± 1.0 1.9 ± 0.8

4.2 ± 0.8 7.1 ± 1.0 9.1 ± 1.2 3.7 ± 1.0

0.5 ± 0.2 1.9 ± 0.4 2.9 ± 0.5 1.2 ± 0.3

0.0 ± 0.0 0.0 ± 0.0 0.0 ± 0.0 0.0 ± 0.0

P. noxius 144 20 25 30 35

5.0 ± 1.1 7.1 ± 1.6 7.9 ± 1.6 3.1 ± 1.6

2.9 ± 1.0 5.6 ± 1.3 7.0 ± 1.7 6.3 ± 0.7

0.5 ± 0.2 1.4 ± 0.6 2.6 ± 0.5 1.4 ± 0.3

0.0 ± 0.0 0.0 ± 0.0 0.1 ± 0.13 0.0 ± 0.0

P. noxius 169 20 25 30 35

4.8 ± 0.8 7.2 ± 0.7 9.3 ± 0.5 2.8 ± 0.7

3.7 ± 0.5 6.0 ± 0.7 7.2 ± 0.8 3.5 ± 1.0

0.4 ± 0.2 1.3 ± 0.4 2.2 ± 0.7 0.4 ± 0.3

0.0 ± 0.0 0.0 ± 0.1 0.0 ± 0.1 0.0 ± 0.0

P. noxius 178 20 25 30 35

5.4 ± 1.4 8.4 ± 1.7 9.2 ± 1.8 3.4 ± 0.7

4.1 ± 0.7 6.5 ± 0.9 7.6 ± 1.0 5.6 ± 1.0

0.5 ± 0.2 1.5 ± 0.3 2.6 ± 0.5 0.8 ± 0.3

0.0 ± 0.0 0.0 ± 0.0 0.0 ± 0.1 0.0 ± 0.0

G. australe 20 25 30 35

2.5 ± 0.5 4.1 ± 0.5 6.3 ± 0.8 3.5 ± 0.6

0.6 ± 0.2 1.0 ± 0.6 1.8 ± 0.4 1.4 ± 0.3

0.0 ± 0.0 0.0 ± 0.0 0.0 ± 0.1 0.0 ± 0.0

0.0 ± 0.0 0.0 ± 0.0 0.0 ± 0.0 0.0 ± 0.0

For each experimental treatment (water activity aw and temperature), 10 replicates were performed. The growth rate (mm/day) was determined by colony diameter measurements carried out along two perpendicular axes after 24 h. The aw of the substrate was controlled by the addition of appropriate weights of the non-ionic solute, glycerol, prior to autoclaving. Data were analyzed by analysis of variance (ANOVA) to assess the effect of aw and temperature on growth rate (mm/day). ±, Standard deviation.

potential against three P. noxius strains and G. australe. P. noxius 178 showed moderate resistance to T. harzianum 121009. Trichoderma reesei 3967 revealed the weakest effect against all wood-decay fungi. The highest resistance of P. noxius to antagonism of Trichoderma species was recorded for strain 133. The highest susceptibility of P. noxius to antagonism of Trichoderma species was recorded for strain 169 (Table 4). 3.3. Inhibitory effects of volatile compounds produced by Trichoderma species on wood-decay fungi The results revealed that after 6 days’ incubation, the VOCs produced by T. atroviride 15603.1 caused a significant (p < 0.0001) inhibition of growth of all P. noxius strains (Table 5). None of the VOCs from the Australian Trichoderma strains were effective in reducing pathogen growth except T-4428 that inhibited P. noxius 178. The weakest effect against the P. noxius strains was recorded for T. harzianum 121009. In the presence of VOCs, the growth of P. noxious strains was often enhanced. By contrast, the VOCs of T. atroviride 15603.1 and T. harzianum 121009 markedly inhibited the growth of G. australe. Among the Trichoderma species, with regard to the production and effect of VOCs, the wood-decay fungi differed significantly in their reaction. P. noxius strains 169 and 133 showed a strong sensitivity to the VOCs, whereas P. noxius 144 showed a lower sensitivity (Table 5). In summary, the effect of VOCs against P. noxious strains resulted in the inhibition of growth of approximately 19–25%, whereas growth of G. australe was inhibited by approximately 15% (Table 5). Trichoderma atroviride 15603.1 inhibited the growth of P. noxius strains and G. australe by 70–85% and 46%, respectively (Table 5).

164

F.W.M.R. Schwarze et al. / Biological Control 61 (2012) 160–168 Table 4 Classification of the degree of mycoparasitism of different Trichoderma species against different Phellinusnoxiusstrains and Ganodermaaustrale on malt extract agar. Trichoderma species

Phellinus noxius 133 Phellinus noxius 178 Phellinus noxius 144 Phellinus noxius 169 Ganoderma australe

T. atrovirde 15603.1

T. ghanense 4510

T. harzianum 4428

T. reesei 3967

T. harzianum 121009

1.1a ± 0.1 1.6 ± 0.1 2.5 ± 0.1 2.0 ± 0.4 3.0 ± 0.6

1.2 ± 1.0 1.4 ± 0.2 2.2 ± 1.2 1.5 ± 1.3 2.3 ± 1.1

2.9 ± 0.7 3.0 ± 0.1 2.9 ± 0.2 2.8 ± 0.3 3.0 ± 0.7

1.8 ± 0.8 1.1 ± 0.7 1.9 ± 0.2 1.8 ± 0.4 1.7 ± 0.2

3.0 ± 0.9 2.8 ± 0.1 3.0 ± 0.5 2.6 ± 0.6 2.9 ± 1.5

[80]b [70] [100] [100] [100]

[60] [50] [60] [60] [60]

[60] [100] [60] [100] [60]

[30] [60] [30] [60] [0]

[100] [60] [100] [100] [100]

For each experimental treatment (interactions and lethal effect) in dual cultures, 10 replicates were performed. a Following system was used to classify the rate of mycoparasitism in dual cultures: 0 = no overgrowth; 1 = slow overgrowth; 2 = fast overgrowth; 3 = very fast overgrowth and deadlock of the wood decay fungi within 4 weeks. ±, Standard deviation. b The ability of Trichoderma species to eliminate the wood-decay fungi (lethal effect in %) during the four week incubation period was evaluated by aseptically transferring 5-mm discs from test plates to MEA with 2 mL thiabendazole (2-(40-thiazolyl)- benzimidazole; dissolved in lactic acid). Table 5 Inhibition of radial growth (in %) of wood decay fungi by volatile organic compounds (VOCs) produced by Trichoderma species. Phellinus noxius strain

Control T. atrovirde15603.1 T. ghanense 4510 T. harzianum 4428 T. harzianum121009 T. reesei 3967

Ganoderma australe

133

144

169

178

9.3 ± 2.0 2.2 ± 1.4** 11.0 ± 2.4n.s. 5.3 ± 0.5n.s. 10.8 ± 2.7n.s. 5.8 ± 0.5n.s.

6.5 ± 1.3 1.9 ± 2.0** 6.3 ± 1.5n.s. 5.3 ± 0.5n.s. 6.9 ± 1.5n.s. 5.8 ± 0.5n.s.

6.9 ± 2.6 1.0 ± 1.2** 6.1 ± 2.4n.s. 4.4 ± 1.1n.s. 8.1 ± 3.4n.s. 6.28 ± 2.7n.s.

7.7 ± 2.6 1.4 ± 1.5** 8.4 ± 3.0n.s. 3.6 ± 1.1** 8.8 ± 3.4n.s. 7.1 ± 1.7n.s.

4.5 ± 1.4 2.4 ± 0.7** 5.8 ± 2.2n.s. 3.8 ± 1.8n.s. 3.4 ± 1.8n.s. 4.4 ± 1.4n.s.

For assessing the inhibition of radial growth (in %) 10 replicates were performed for each combination of wood decay fungus and Trichoderma species.*Significant influence on growth rate by VOCs. Dunnett’s test: ±, Standard deviation. * p < 0.05. ** p < 0.0001. n.s. p P 0.05.

3.4. Interaction tests in wood All wood-decay fungi had completely colonized the control wood samples, but showed distinctive differences in their potential to decompose the wood of D. regia, J. mimosifolia, F. benjamina and A. bidwillii. In untreated wood blocks of A. bidwillii the wood-decay fungi only caused negligible weight losses (Table 6). In untreated wood blocks of D. regia and F. benjamina, G. australe caused greater weight losses than all four P. noxius strains (Table 6). By contrast, in untreated wood blocks of J. mimosifolia the weight losses caused by G. australe were always lower than those of the P. noxius strains. Interactions among the wood-decay fungi, Trichoderma species and the wood substrate resulted in interesting differences in weight losses (Table 6). A significant reduction in weight losses in wood by G. australe was recorded for A. bidwillii, D. regia and F. benjamina pretreated with Trichoderma species (Table 6). By contrast, in J. mimosifolia wood the weight losses caused by G. australe were not reduced by the Trichoderma strains. In wood of D. regia and F. benjamina pretreated with T-15603.1 and T-3967, the weight losses caused by P. noxius 133 were significantly reduced (Table 6). In J. mimosifolia wood pretreated with Trichoderma species, the weight losses by P. noxious 144 and 178 were also significantly reduced (Table 6). Wood blocks of F. benjamina and D. regia pretreated with Trichoderma species showed the greatest reduction in weight loss. By contrast, wood blocks of A. bidwillii and J. mimosifolia pretreated with Trichoderma species only showed a weak reduction in weight losses. 4. Discussion 4.1. Growth under different conditions The competitiveness of Trichoderma species is based on rapid growth (i.e. a decisive feature for antagonism) (Chet, 1990; Chet

et al., 1998; Hjeljord and Tronsmo, 1998). Physical as well as chemical factors influence growth and germination, therefore knowledge of the optimal conditions for growth, as well as the influence of ecological factors on the antagonist and the target pathogen, is essential for successful application in the field (Hjeljord and Tronsmo, 1998; Kredics et al., 2000, 2003). In this study, the growth of the Trichoderma strains was affected by the environmental factors tested. Most Trichoderma strains showed optimum growth at a mean temperature of 25–30 °C and a high aw of 0.998. Even at 35 °C and the lowest aw (0.928) growth was recorded for some strains. The results have to be interpreted carefully because predicting the behaviour of Trichoderma species under specific conditions is complicated by the mutual effect of environmental parameters (Harman, 2006). Growth of all P. noxius strains at different temperatures did not differ significantly. Optimum growth for most strains was recorded at 30 °C, which is in good agreement with the experimental findings of Ann et al. (2002), who showed that the optimum growth of P. noxius is at 30 °C, and that of Albrecht and Venette (2008), who defined the optimum growth range as approximately 25– 31 °C. All of the P. noxius strains in this study were isolated from urban sites in Brisbane, Queensland, Australia. Interestingly, the optimum growth of the P. noxius strains correlates closely with a recent study on the local soil temperature. Prangnell and McGowan (2009) used a soil temperature calculation equation to calculate soil temperature at various depths in a cemetery located in Brisbane. Their study revealed that throughout the year the mean temperature at a soil depth of 1 m was approximately 28 °C. In the absence of vegetation cover, extreme temperatures of approximately 17 °C and 40 °C were measured. In soils with vegetation cover, only minor temperature fluctuations were reported (Prangnell and McGowan, 2009). The present study showed that the growth of different P. noxius strains does not appear to be strongly affected by different temperatures.

165

F.W.M.R. Schwarze et al. / Biological Control 61 (2012) 160–168 Table 6 Dry weight losses (in %) caused by Phellinus noxius and Ganoderma australein untreated controls and in wood blocks pretreated with different Trichoderma strains. P. noxius 133 Araucaria bidwillii Control T. atrovirde15603.1 T. ghanense 4510 T. harzianum 4428 T. reesei 3967 T. harzianum121009 Delonix regia Control T. atrovirde15603.1 T. ghanense 4510 T. harzianum 4428 T. reesei 3967 T. harzianum121009 Ficus benjamini Control T. atrovirde15603.1 T. ghanense 4510 T. harzianum 4428 T. reesei 3967 T. harzianum121009 Jacaranda mimosifolia Control T. atrovirde15603.1 T. ghanense 4510 T. harzianum 4428 T. reesei 3967 T. harzianum121009

P. noxius144

P. noxius169

P. noxius178

G. australe

0.94 ± 0.82 1.71 ± 3.07n.s. 0.00 ± 0.00n.s. 0.62 ± 0.83n.s. 1.80 ± 2.78n.s. 2.75±3.54n.s.

1.60 ± 0.98 0.32 ± 0.65n.s. 1.36 ± 1.34n.s. 1.69 ± 1.80n.s. 1.16 ± 0.72n.s. 1.90±2.53n.s.

1.38 ± 1.40 0.33 ± 0.67n.s. 0.00 ± 0.00* 0.05 ± 0.12* 0.00 ± 0.00* 3.04±2.79n.s.

3.43 ± 0.33 1.12 ± 0.63* 0.29 ± 0.59** 0.00 ± 0.00** 0.00±0.00** 2.99±4.10*

4.42 ± 4.38 0.55 ± 1.24* 0.06 ± 0.13** 0.00 ± 0.00** 0.00±0.00** 1.03±2.31*

34.78±17.02 13.01±5.17* 23.05±7.40n.s. 25.43±3.69n.s. 13.61±4.35* 24.05±10.42n.s.

21.34±3.36 14.65±7.94n.s. 18.07±8.24n.s. 20.82±5.76n.s. 20.15±2.04n.s. 23.62±5.71n.s.

22.03±5.44 23.95±9.29n.s. 23.45±6.37n.s. 21.44±9.18n.s. 22.20±5.35n.s. 22.83±2.71n.s.

18.43±9.74 12.31±3.66n.s. 20.97±9.82n.s. 21.07±5.23n.s. 22.61±3.73n.s. 19.96±5.61n.s.

59.89±4.19 18.97±7.60** 16.65±8.49** 18.08±6.58** 26.15±7.12** 28.59±5.99**

23.30±8.74 7.58±1.19* 7.33±2.17* 7.28±1.69* 8.49±2.75* 6.51±6.02*

22.40±4.60 6.70±2.42** 7.70±3.15** 7.59±3.00** 8.23±1.91** 16.32±11.51n.s.

9.83±3.98 0.78±1.68** 5.02±2.85n.s. 9.77±2.23n.s. 5.39±1.91n.s. 10.17±6.31n.s.

18.00±2.60 5.89±1.31** 5.02±2.85** 5.39±1.91** 8.42±3.58* 7.84±4.51**

36.91±10.02 6.76±1.92** 7.23±2.61** 7.45±3.07** 11.19±2.41** 5.70±5.24**

11.82±3.32 9.79±2.38n.s. 9.10±1.17n.s. 8.25±1.70n.s. 8.46±1.30n.s. 11.93±13.75n.s.

20.62±3.10 12.66±5.20n.s. 10.81±2.22* 10.33±3.32* 8.27±1.45* 11.86±7.88*

9.50±4.71 8.54±1.16n.s. 7.03±4.19n.s. 11.16±2.34n.s. 7.24±7.79n.s. 12.74±12.96n.s.

14.04±0.97 7.96±1.58** 9.41±1.98* 0.68±1.93* 9.58±2.36* 15.16±10.47n.s.

4.46±2.39 9.07±2.14n.s. 6.60±2.24n.s. 8.98±1.89n.s. 8.10±2.95n.s. 8.66±5.12n.s.

Untreated wood blocks served as the control. Twenty replicates were used for each experiment. Before incubation wood blocks were oven-dried at 105 °C for 24 h to determine the wood dry weight. The decay tests were run for 12 weeks at 25 °C, after which the wood blocks were removed, cleaned and oven-dried for measuring dry weight losses. Significant reduction of wood decay (wood weight loss in %). Dunnett’s test: ±, Standard deviation. * Significant p < 0.05. ** Highly significant p < 0.0001. n.s. Not significant p P 0.05.

In comparison with the P. noxius strains, growth of most Trichoderma species was more rapid at the highest and lowest water activity values (0.998 and 0.928). At the lowest aw (0.928) and at 25 °C and 30 °C, growth was minimal and at 35 °C the growth of the P. noxius strains failed completely. The European species, T. atroviride 15603.1, that was used as a ‘‘bench mark’’ failed to grow at 35 °C, which may indicate its limited adaptation to the subtropical climate of Brisbane where soil temperature can exceed 35 °C. This finding demonstrates the importance of screening Trichoderma species for the specific niche where they are envisaged to be applied (i.e. increasing target specificity). 4.2. Evaluation of antagonistic activity in dual culture The direct mycoparasitic activity of Trichoderma species is one of the major mechanisms proposed to explain their antagonistic activity against soil-borne plant-pathogenic fungi. In the dual culture tests, hyphal contact between Trichoderma species and wood-decay fungi was observed for all host–pathogen combinations. However, not all strains of Trichoderma were able to overgrow and parasitize the mycelia of the wood-decay fungi. In this study three lethal effect of mycoparasitism by Trichoderma species was most prevalent for T. harzianum 121009 which was isolated from a basidiocarp of Laetiporus portentosus (Berk.) Rajchenb. collected at Mt. Coot-tha, west of Brisbane, Queensland. It showed a high antagonistic potential against three P. noxius strains and G. australe. The European strain T-15603.1 and T4428 showed a high antagonistic potential against all wood-decay fungi, whereas the antagonistic potential of T-4510 and T-3967 was limited.

Pathogen and strain resistance to mycoparasitism by Trichoderma species is well documentated (Sivan and Chet, 1989; Cortés et al., 1998; Zeilinger et al., 1999). In vitro tests conducted by Sivan and Chet, 1989 showed that two strains of Trichoderma harzianum failed to parasitize colonies of Fusarium oxysporum f. vasinfectum (G.F. Atk.) W.C. Snyder and H.N. Hansen and Fusarium oxysporum f. melonis W.C. Snyder and H.N. Hansen. However, these strains were strongly mycoparasitic on Rhizoctonia solani J.G. Kühn and Pythium aphanidermatum (Edson) Fitzp. (Sivan and Chet, 1989). The studies suggest that proteins in the cell walls of F. oxysporum may make these walls more resistant than those of R. solani to degradation by extracellular enzymes of T. harzianum. Savoie et al. (2000) studied interactions between Lentinula edodes (Berk.) Pegler and Trichoderma spp. and observed that there were great differences between two modified substrates for each strain. Similary, Tokimoto and Komatsu (1979) observed that carbon rich medium favours L. edodes while a nitrogen rich medium favours Trichoderma and that mycoparasitism could be reduced by controlling nutritional conditions. Giovannini et al. (2004) selected resistant strains of the wood decay fungi Grifola frondosa (Dicks.) Gray and Fomitopsis pincola (Sw.) P. Karst. with a Teflon tubes confrontation method. Results showed great variations in behaviour between strains, but also for a given strain growing on different substrates, indicating the importance of trophic factors (Giovannini et al., 2004). Interestingly in their experiments with F. pinicola they demonstrated that the substrate with the lower nitrogen content was more favourable for Trichoderma (Giovannini et al., 2004). In a previous study on mycoparasitism of a range of Trichoderma species against wood decay fungi, the white rot fungus Polyporus

166

F.W.M.R. Schwarze et al. / Biological Control 61 (2012) 160–168

squamosus was not only able to circumvent parasitism but also adapted its hyphal structure, to overgrow the mycelia of the Trichoderma isolates (Schubert et al., 2008a). 4.3. Inhibitory effects of volatile compounds produced by Trichoderma species on wood-decay fungi Antibiosis was recognized and initially described by Weindling (1932) and is defined as the production of secondary metabolites that have an antimicrobial effect, even at low concentrations (Cooney et al., 1997a,b; Galindo et al., 2004; Howell, 1998; Scarselletti and Faull, 1994; Schirmböck et al., 1994; Wheatley et al., 1997). The VOCs produced by T. atroviride 15603.1 strongly inhibited growth of all P. noxius strains used in this study. Interestingly, the VOCs produced by T. harzianum 121009 and T. ghanense 4510 appeared to stimulate the growth of P. noxius strains, however the effect was not significant. The VOCs studies also indicated that the antagonistic potential of Trichoderma species against P. noxius varies from strain to strain, so for successful selection of a Trichoderma species for biological control of P. noxius these variations in strain resistance have to be considered. 4.4. Interaction tests on wood In order to assess the antagonistic potential of Trichoderma species on its natural substrate, interaction studies were performed on wood blocks. After 12 weeks’ incubation, the Trichoderma species failed to completely inhibit decomposition, as measured by dry weight losses. This may be partly explained by the degradation of readily accessible carbohydrates by Trichoderma species within parenchymal cells and pits (Kubicek-Pranz, 1998). The inoculum potential in turn is crucial for the invasiveness of pathogens (Redfern and Filip, 1991). Nevertheless, a significant reduction in dry weight loss was induced after pretreatment of the wood with a conidial suspension of Trichoderma species. Significant differences between the species and strains of Trichoderma were evident. Interestingly the antagonistic potential of different Trichoderma species against P. noxius strains varied according to the specific wood substrate. Thus, P. noxius strains showed a different degree of adaptation to the wood. Phellinus noxius 169 strongly decomposed the wood of D. regia, whereas weight losses on F. benjamina and J. mimosifolia were significantly lower. By contrast, only negligible weight losses were recorded from A. bidwillii. The variations in weight loss may be explained by the specific lignin composition of the different wood species (Schwarze, 2007). Gymnospermous wood consists almost exclusively of guaiacyl monomers, whereas angiospermous wood consists of approximately equal ratios of guaiacyl and syringyl monomers (Whetten and Sederoff, 1995). Fungal decay types fall into three categories according to their mode of degradation of the woody cell wall: brown rot, white rot and soft rot (Schwarze, 2007). White rot is subdivided into simultaneous rot and selective delignification. UItrastructural studies have revealed that P. noxius causes a simultaneous rot (Nicole et al., 1995). Many basidiomycetes cause simultaneous rot in angiosperms, but only rarely in gymnospermous wood (Schwarze, 2007). This selection may be related to the extremely resilient S3 layer of tracheids, which hampers degradation by hyphae from within the cell lumen outwards (Schwarze, 2007), and may further explain the resistance of gymnospermous wood to degradation by P. noxius. The high resistance of A. bidwillii wood to decomposition by different P. noxius strains appears to suggest an enhanced durability of softwoods to decay. However, Ann et al. (2002) report that wood of other Araucaria species (e.g. A. cunninghamii) is susceptible to decomposition by P. noxius. On the basis of the present study results, the selection of one Trichoderma strain for biological control of P. noxius is difficult.

The European strain, T. atroviride 15603.1, showed a high antagonistic potential against all wood-decay fungi and its VOCs greatly inhibited their growth. However, its failure to grow at 35 °C indicates its limited adaptation to the subtropical climate of Brisbane where temperatures can exceed 35 °C. Even at high temperatures and low aw, T-4510 was a very competitive species and mycoparasitism in wood resulted in a reduction in wood weight losses, but its VOCs actually stimulated the growth of some P. noxius strains. These results demonstrate that one and the same Trichoderma species may possess positive and negative attributes for biological control. As one Trichoderma species rarely has only positive attributes, the selection of one potential Trichoderma species is not a trivial exercise and will require further screening studies. For biological control a number of Trichoderma species with different antagonistic properties may have to be combined to successfully inhibit the growth of the target pathogen in the field. In conclusion, the present study shows that the success of biological control of P. noxius depends on the Trichoderma species, the strain of wood-decay fungus, the specific wood species and the prevailing environmental conditions (temperature/water activity). The in vitro screening of antagonistic potential used in this study allowed a systematic investigation of several Trichoderma strains, as well as specific ecological factors, and thus identification of effective strains. The study shows that Trichoderma species can be used to significantly inhibit the growth of different P. noxius strains, and have a lethal effect on some basidiomycota species in controlled growing conditions. The study also showed that Trichoderma species can be used to reduce the rate of decomposition by P. noxius and Ganoderma australe in the wood of the species studied here. However, positive results obtained from in vitro studies are only indicative, as experimental conditions do not take all ecological and endemic factors into account. For this reason, field studies are essential to test the selected competitive biocontrol agent under field conditions (Schubert et al., 2008b). Acknowledgments We thank Brisbane City Council for assisting in the preparation of wood materials and facilities, and Keith Foster for technical and project support. Finally, the financial and technical support (i.e. data collection and study design) provided by ENSPEC and Brisbane City Council is gratefully acknowledged. References Alabouvette, C., Olivain, C., Steinberg, C., 2006. Biological control of plant diseases: the European situation. European Journal of Plant Pathology 114, 329–341. Albrecht, E.M., Venette, R.C., 2008. Phellinus noxius. In: Venette, R.C. (Ed.), CAPS (Cooperative Agricultural Pest Survey). Northern Research Station, USDA Forest Service, St. Paul, MN, pp. 173–184. Ann, P.J., Tsai, J.N., Wang, I.T., Hsieh, M.J., 1999. Response of fruit trees and ornamental plants to brown root rot disease by artificial inoculation with Phellinus noxius. Plant Pathology Bulletin 8, 61–66. Ann, P.J., Chang, T.T., Ko, W.H., 2002. Phellinus noxius brown root rot of fruit and ornamental trees in Taiwan. Plant Disease 86, 820–826. Ariffin, D., Idris, A.S., Singh, G., 2000. Status of Ganoderma in oil palm. In: Flood, J., Bridge, P.D., Holderness, M. (Eds.), Ganoderma Diseases of Perennial Crops. CAB International, Oxon, UK, pp. 49–69. Askew, D.J., Laing, M.D., 1993. An adapted selective medium for the quantitative isolation of Trichoderma species. Plant Pathology 42, 686–690. Bissett, J., 1984. A revision of the genus Trichoderma. I: section Longibrachiatum sect. Nov.. Canadian Journal of Botany 62, 924–931. Bissett, J., 1991a. A revision of the genus Trichoderma. II: infrageneric classification. Canadian Journal of Botany 69, 2357–2372. Bissett, J., 1991b. A revision of the genus Trichoderma. III: section Pachybasium. Canadian Journal of Botany 69, 2372–2417. Bissett, J., 1991c. A revision of the genus Trichoderma. IV: additional notes on section Longibrachiatum. Canadian Journal of Botany 69, 2418–2420. Bissett, J., 1992. Trichoderma atroviride. Canadian Journal of Botany 70, 639–641. Bolland, L., 1984. Phellinus noxius: cause of a significant root-rot in Queensland hoop pine plantations. Australian Forestry Journal 47, 2–10.

F.W.M.R. Schwarze et al. / Biological Control 61 (2012) 160–168 Bolland, L., Tierney, J.W., Winnington-Martin, S.M., Ramsden, M., 1988. Investigations into the feasibility of biological control of Phellinus noxius and Poria vincta in Queensland hoop pine plantations. In: Morrison, D.J., Victoria, B.C. (Eds.), Seventh International Conference on Root and Butt Rots, Vernon and Victoria, British Columbia, August 9–16, 1988. Forestry Canada, pp. 72–82. Chang, T.T., 1995. Decline of nine tree species associated with brown root rot caused by Phellinus noxius in Taiwan. Plant Disease 79, 962–965. Chang, T.T., 2002. The biology, ecology and pathology of Phellinus noxius in Taiwan. Tropical Mycology 1, 87–99. Chet, J., 1990. Mycoparasitism: recognition, physiology and ecology. In: Baker, R.R., Dunn, P.E. (Eds.), New Directions in Biological Control: Alternatives for Suppressing Agricultural Pests and Diseases. Alan Liss, New York, pp. 725–733. Chet, I., Benhamou, N., Haran, S., 1998. Mycoparasitism and lytic enzymes. In: Harman, G.E., Kubicek, C.P. (Eds.), Trichoderma and Gliocladium, Enzymes, Biological Control, and Commercial Applications, vol. 2. Taylor & Francis, London, pp. 153–172. Cooney, J.M., Lauren, D.R., Perry-Meyer, L.J., 1997a. A novel tubular bioassay for measuring the production of antagonistic chemicals produced at the fungal/ pathogen interface. Letters in Applied Microbiology 24, 460–462. Cooney, J.M., Vannests, J.L., Lauren, D.R., Hi ll, R.A., 1997b. Quantitative determination of the antifungal compound 6-pentyl-a-pyrone (6PAP) using a simple plate bioassay. Letters in Applied Microbiology 24, 47–50. Corner, E.J.H., 1932. The identification of the brown-root fungus. Gardens’ Bulletin, Straits Settlements, Singapore 5, 317–350. Cortés, C., Gutiérrez, A., Olmedo, V., Inbar, J., Chet, I., Herrera-Estrella, A., 1998. The expression of genes involved in parasitism by Trichoderma harzianum is triggered by a diffusible factor. Molecular Genetics and Genomics 260, 218–225. Cunningham, G.H., 1965. Polyporaceae of New Zealand. New Zealand Department of Science and Industry Research Bulletin 164, 221–222. Dallyn, H., 1978. Effect of substrate water activity on growth of certain xerophilic fungi. Ph.D. Thesis, South Bank University, London. Dennis, C., Webster, J., 1971. Antagonistic properties of species-groups of Trichoderma. II: production of non-volatile antibiotics. Transactions of the British Mycological Society 57, 41–48. Durand-Gasselin, T., Asmady, H., Flori, A., Jacquemard, J.C., Hayun, Z., Breton, F., de Franqueville, H., 2005. Possible sources of genetic resistance in oil palm (Elaeis guineensis Jacq.) to basal stem rot caused by Ganoderma boninense: prospects for future breeding. Mycopathologia 159, 93–100. Flood, J., Husan, Y., Turner, P.D., O’Grady, E.B., 2000. The spread of Ganoderma from infective sources in the field and its implications for management of the disease in oil palm. In: Flood, J., Bridge, P.D., Holderness, M. (Eds.), Ganoderma Diseases of Perennial Crops. CAB International, Oxon, UK, pp. 101–113. Galindo, E., Flores, C., Larralde-Corona, P., Corkidi, G., Rocha-Valadez, J.A., SerranoCarreón, L., 2004. Production of 6-pentyl-a-pyrone by Trichoderma harzianum cultured in unbaffled and baffled shake flasks. Biochemical Engineering Journal 18, 1–8. Gams, W., Bissett, J., 1998. Morphology and identification of Trichoderma. In: Kubicek, C.P., Harman, G.E. (Eds.), Trichoderma and Gliocladium: Basic Biology, Taxonomy and Genetics. Taylor & Francis, London, pp. 3–34. Giovannini, I.S., Job, D., Hmamda, A., 2004. Selection of Grifola frondosa and Fomitopsis pinicola resistant strains in teflon tubes confrontation method. Mycological Progress 3, 329–336. Harman, G.E., 2006. Overview of mechanisms and uses of Trichoderma species. Phytopathology 96, 190–194. Harman, G.E., Howell, C.R., Viterbo, A., Chet, I., Lorito, M., 2004. Trichoderma species: opportunistic, avirulent plant symbionts. Nature Review Microbiology 2, 43–56. Highley, T.L., 1997. Control of wood decay by Trichoderma (Gliocladium) virens. I: antagonistic properties. Material und Organismen 31, 79–89. Hjeljord, L., Tronsmo, A., 1998. Trichoderma and Gliocladium in biological control: an overview. In: Harman, G.E., Kubicek, C.P. (Eds.), Trichoderma and Gliocladium, Enzymes, Biological Control, and Commercial Applications, vol. 2. Taylor & Francis, London, pp. 131–151. Holdenreider, O., Greig, B.J.W., 1998. Biological methods of control. In: Woodward, S., Stenlid, J., Karjalainen, R., Hüttermann, A. (Eds.), Heterobasidion annosum: Biology, Ecology, Impact and Control. CAB International, London, pp. 235–259. Howell, C.R., 1998. The role of antibiosis. In: Harman, G.E., Kubicek, C.P. (Eds.), Trichoderma and Gliocladium, Enzymes, Biological Control, and Commercial Applications, vol. 2. Taylor & Francis, London, pp. 173–184. Kredics, L., Antal, Z., Manczinger, L., 2000. Influence of water potential on growth, enzyme secretion and in vitro enzyme activities of Trichoderma harzianum at different temperatures. Current Microbiology 40, 310–414. Kredics, L., Antal, Z., Manczinger, L., Szekeres, A., Kevei, F., Nagy, E., 2003. Influence of environmental parameters on Trichoderma strains with biocontrol potential. Food Technology and Biotechnology 41, 37–42. Kubicek-Pranz, E.M., 1998. Nutrition, cellular structure and basic metabolic pathways in Trichoderma and Gliocladium. In: Harman, G.E., Kubicek, C.P. (Eds.), Trichoderma and Gliocladium: Basic Biology, Taxonomy and Genetics. Taylor & Francis, London, pp. 95–119. Mohd Farid, A., Lee, S.S., Maziah, Z., Patahayah, M., 2009. Pathogenicity of Rigidoporus microporus and Phellinus noxius against four major plantation tree species in peninsular Malaysia. Journal of Tropical Forest Science 21, 289–298. Naár, Z., Kecskes, M., 1998. Factors influencing the competitive saprophytic ability of Trichoderma species. Microbiological Research 53, 119–129. Nicole, M., Chamberland, H., Rioux, D., Xixuan, X., Blanchette, R.A., Geiger, J.P., Ouellette, G.B., 1995. Wood degradation by Phellinus noxius: uItrastructure and cytochemistry. Canadian Journal of Microbiology 41, 253–265.

167

Paterson, R.R., Holderness, M., Kelley, J., Miller, R., OGrady, E., 2000. In vitro biodegradation of oil-palm stem using macroscopic fungi from South-East Asia: a preliminary investigation. In: Flood, J., Bridge, P.D., Holderness, M. (Eds.), Ganoderma Diseases of Perennial Crops. CAB International, Oxon, UK, pp. 129– 139. Prangnell, J., McGowan, G., 2009. Soil temperature calculation for burial site analysis. Forensic Science International 191, 104–109. Prasad, M., Naik, S.T., 2002. Management of root rot and heart rot of Acacia mangium Willd. Karnataka Journal of Agricultural Sciences 15, 321–326. Raziq, F., Fox, R.T.V., 2006. The integrated control of Armillaria mellea 2. Field experiments. Biological Agriculture and Horticulture 23, 235–249. Redfern, D.B., Filip, G.M., 1991. Inoculum and infection. In: Shaw, C.G., Kile, G.A. (Eds.), Armillaria Root Disease. USDA, Washington D.C., pp. 48–61. Rifai, M.A., 1969. A revision of the genus Trichoderma. Mycological Papers 116, 1– 56. Sahashi, N., Akiba, M., Ishihara, M., Miyazaki, K., Kanzaki, N., 2010. Cross inoculation tests with Phellinus noxius isolates from nine different host plants in the Ryukyu Islands, southwestern Japan. Plant Disease 94, 358–360. Sariah, M., 2003. The potential of biological management of basal stem rot of oil palm: issues, challenges and constraints. Oil Palm Bulletin 47, 1–5. Sariah, M., Choo, C.W., Zakaria, H., Norihan, M.S., 2005. Quantification and characterisation of Trichoderma species from different ecosystems. Mycopathologia 159, 113–117. Scarselletti, R., Faull, J.L., 1994. In vitro activity of 6-pentyl-a-pyrone, a metabolite of Trichoderma harzianum, in the inhibition of Rhizoctonia solani and Fusarium oxysporum f sp. lycopersici. Mycolological Research 98, 1207–1209. Savoie, J.-M., Delpech, P., Billette, C., Mata, G., 2000. Inoculum adaptation changes the outcome of the competition between Lentinula edodes and Trichoderma spp. during shiitake cultivation on pasteurized wheat straw. In: Van Griensven, L.J.L.D. (Ed.), Science and Cultivation of Edible Fungi. Balkema, Rotterdam. Schirmböck, M., Lorito, M., Wang, Y.L., Hayes, C.K., Arisan-Atac, I., Scala, F., Harman, G.E., Kubicek, C.P., 1994. Parallel formation and synergism of hydrolytic enzymes and peptaibol antibiotics, molecular mechanisms involved in the antagonistic action of Trichoderma harzianum against phytopathogenic fungi. Applied Environmental Microbiology 60, 4364–4370. Schoeman, M.W., Webber, J.F., Dickinson, D.J., 1999. The development of ideas in biological control applied to forest products. International Biodeterioration and Biodegradation 43, 109–123. Schubert, M., 2006. In vitro und ad planta Studien zum Einsatz von TrichodermaArten für die Biologische Kontrolle Holz abbauender Pilze an Bäumen. Ph.D.Thesis, Albert-Ludwigs-Universität Freiburg im Breisgau. Schubert, M., Fink, S., Schwarze, F.W.M.R., 2008a. In vitro screening of an antagonistic Trichoderma strain against wood decay fungi. Part I. Arboricultural Journal 31, 227–248. Schubert, M., Fink, S., Schwarze, F.W.M.R., 2008b. Field experiments to evaluate the application of Trichoderma strain (T-15603.1) for biological control of wood decay fungi in trees. Part II. Arboricultural Journal 31, 249–268. Schubert, M., Fink, S., Schwarze, F.W.M.R., 2008c. Evaluation of Trichoderma spp. as a biocontrol agent against wood decay fungi in urban trees. Biological Control 45, 111–123. Schubert, M., Mourad, S., Fink, S., Schwarze, F.W.M.R., 2009. Ecophysiological responses of the biocontrol agent Trichoderma atroviride (T-15603.1) to combined environmental parameters. Biological Control 49, 84–90. Schwarze, F.W.M.R., 2007. Wood decay under the microscope. Fungal Biology Reviews 1, 133–170. Schwarze, F.W.M.R., Fink, S., 1998. Host and cell type affect the mode of degradation by Meripilus giganteus. New Phytologist 139, 721–731. Sieber, T.N., 1995. Pyrenochaete ligni-putridi sp. nov., a new coelomycete associated with butt rot of Picea abies in Switzerland. Mycological Research 99, 274– 276. Singh, G., 1991. Ganoderma: the scourge of oil palms in the Coastal Areas. The Planter 67, 421–444. Sivan, C.J., Chet, I., 1989. Degradation of fungal cell walls by lytic enzymes of Trichoderma harzianum. Journal of General Microbiology 135, 675–682. Soepena, H., Purba, R.Y., Pawirosukarto, S., 2000. A control strategy for basal stem rot (Ganoderma) on oil palm. In: Flood, J., Brignolas, F., Holderness, M. (Eds.), Ganoderma Diseases of Perennial Crops. CAB International, Oxon, UK, pp. 83– 88. Susanto, A., Sudharto, P.S., Purba, P.Y., 2005. Enhancing biological control of basal stem rot disease (Ganoderma boninense) in oil palm plantations. Mycopathologia 159, 153–157. Tokimoto, K., Komatsu, M., 1979. Effect of carbon and nitrogen sources in media on the hyphal interference between Lentinus edodes and some species of Trichoderma. Annals of the Phytopathological Society of Japan 45, 261–264. Turner, P.D., 1981. Oil Palm Diseases and Disorders. Oxford University Press, Kuala Lumpur, Malaysia. Weindling, R., 1932. Trichoderma lignorum as a parasite of other soil fungi. Phytopathology 22, 837–845. Wheatley, R., Hackett, C., Bruce, A., Kundzewicz, A., 1997. Effect of substrate composition on the production of volatile organic compounds from Trichoderma species inhibitory to wood decay fungi. International Biodeterioration and Biodegradation 39, 199–205. Whetten, R., Sederoff, R., 1995. Lignin biosynthesis. Plant Cell 7, 1001–1013. Widyastuti, S.M., 2006. The biological control of Ganoderma root rot by Trichoderma. In: Potter, K., Rimbawanto, A., Beadle, C. (Eds.), Heart Rot and Root Rot in Tropical Acacia Plantations (Proceedings of a Workshop held in Yogyakarta,

168

F.W.M.R. Schwarze et al. / Biological Control 61 (2012) 160–168

Indonesia, February 7–9, 2006). ACIAR Proceedings No. 124, Australian Centre for International Research, Canberra, pp. 67–74. Wu, J., Peng, S.L., Zhao, H.B., Tang, M.H., Li, F.R., Chen, B.M., 2011. Selection of species resistant to the wood rot fungus Phellinus noxius. European Journal Plant Pathology 130, 463–467.

Zeilinger, S., Galhaup, C., Payer, K., Woo, S.L., Mach, R.L., Fekete, C., Lorito, M., Kubicek, C.P., 1999. Chitinase gene expression during mycoparasitic interaction of Trichoderma harzianum with its host. Fungal Genetics and Biology 26, 131– 140.