Biological treatment for controlling wood deteriorating fungi

A method for retarding the growth of wood-degrading fungi by treating the wood with an effective amount of viable, nonsporulating Streptomyces rimosus SC-36 NRRL 21063.

FIELD OF THE INVENTION 
This invention is a method for the treatment of wood and wood products with 
living actinomycete cells of Streptomyces rimosus non-sporulating isolate 
SC-36, NRRL 21063, for the purpose of preventing or retarding the 
reproduction and growth of wood deteriorating fungi. The metabolites of a 
Streptomyces rimosus nonsporulating isolate, designated herein as SC-36, 
has been found to be particularly effective in preventing basidiospore and 
conidia germination of wood-attacking fungi. The effect of combining 
diluted boron with such metabolites, as well as combining the metabolites 
with ethylene glycol, have been found to have a synergistic effect in 
preventing such germination. 
BACKGROUND OF THE INVENTION 
Actinomycetes, such as Streptomyces rimosus, are well known actinomycetes 
useful for the production of antibiotics (see U.S. Pat. Nos. 2,516,080 to 
Ben A. Sobin et al and 2,963,401 to Jacob W. Davison et al). A new strain 
of such Streptomyces rimosus is disclosed by U.S. Pat. No. 3,377,242 to 
Donald Lefemmine et al. 
It has now been discovered that actinomycetes such as those described in 
the above references and extracts from such actinomycetes are particularly 
useful in treating and preventing fungal attack on wood as well as their 
antibiotic uses. Such use constitutes a significant advance in the 
treatment of wood for this purpose because there are environmental and 
health concerns about many of the synthetic chemical fungicides presently 
used to protect wood from fungal attack. 
It is accordingly the object of the present invention to provide a means 
for retarding, stopping and preventing the fungal attack of wood by the 
treatment of such wood with living actinomycete cells. 
It is also an object of the present invention to treat wood with the 
metabolites, i.e. a cell-free supernatant or a reconstituted cell-free 
compsotion, of the actinomycete to retard, stop or prevent fungal attack. 
It is also an object of the present invention to treat wood with the 
actinomycete living cells of Streptomyces rimosus non-sporulating isolate 
SC-36 for the purpose of preventing, retarding or stopping fungal attack. 
A further object of the present invention is to treat wood with a 
metabolite extract from the actinomycete Streptomyces rimosus 
non-sporulating isolate SC-36 for the purpose of preventing, retarding or 
stopping fungal attack. 
A still further object of the present invention is to treat wood with a 
combination of a metabolic extract of the actinomycete Streptomyces 
rimosus non-sporulating isolate SC-36 and diluted boron for its 
synergistic effect in preventing, retarding or stopping fungal attack. 
SUMMARY OF THE INVENTION 
It has now been discovered that living actinomycete cells prevent, retard 
or stop basidiospore and conidia germination of wood-attacking fungi. 
There has been particular success with Streptomyces rimosus nonsporulating 
isolate we have designated as SC-36 (NRRL 21063). It has also been 
discovered that the metabolites of Streptomyces rimosus nonsporulating 
isolate SC-36 are particularly effective against wood-attacking fungi and 
particularly where such metabolites are combined with diluted boron. In 
the latter instance there is a synergistic antifungal effect. 
These treatments completely prevented wood degradation and discoloration of 
sapstain fungi, mold fungi, brown-rot fungi and white-rot fungi in 
laboratory wood block tests and in pine log sections exposed in field 
trials. The combination of diluted ethylene glycol and these metabolites 
also inhibited basidiospore germination of brown-rot and white-rot fungi.

DETAILED DESCRIPTION OF THE INVENTION 
In the practice of the present invention a new strain of actinomycete 
isolate, Streptomyces rimosus nonsporulating isolate SC-36, specifically 
S. rimosus SC-36, NRRL 21063, under proper culture conditions, described 
below, produces potent antifungal metabolites and living actinomycete 
cells in complete liquid medium (CLM). S. rimosus was deposited at 
Agricultural Research Culture Collection (NRRL) International Depository 
Authority (Peoria, Ill.), at accession number 21063 on Apr. 1, 1992. The 
CLM medium may contain, per liter: D-glucose, 10 g; yeast extract, 5 g; 
lactalbumin hydrolysate, 15 g; sucrose, 2.8 g; calcium carbonate, 1 g. 
Mycelia from nutrient agar supplemented with 1% bacto peptone (NAP) slant 
are inoculated into 30 ml of CLM medium in 125-mml Erlenmeyer flasks and 
incubated at 30.degree. C. with vigorous shaking on a platform shaker at 
200 r/min for 1-2 days. The settled growth is transferred into 200-ml CLM 
medium in 500-ml Erlenmeyer flasks and allowed to grow for an additional 
2-3 days. The harvested and packed actinomycete cells are suspended in 
equal volumes of 0.5% sodium chloride solution. Treatment of wood and 
wood-products with the actinomycete cells provided for application as a 
biological control agent of wood-attacking microorganisms. 
For metabolite production, the cultural conditions recommended for the 
above consists of stock cultures of Streptomyces rimosus SC-36 maintained 
on nutrient agar supplemented with 1% peptone (NAP) slants. Cultures are 
grown in vegetative growth medium, 1x sporulation broth (SB), for 
activation and adaptation of inoculum. The sporulation broth medium 
contains per liter: 1.0 g yeast extract, 1.0 g beef extract, 2.0 g 
tryptose, 1 mg FeSO.sub.4, and 10.0 g glucose. The pH of medium is 
adjusted to 7.2 before autoclaving. For vegetative growth, mycelia of 
SC-36 from cultures grown on a NAP slant for 5 days are scraped from the 
entire surface of the slant and cultured in 50 ml of SB medium in 125-ml 
Erlenmeyer flasks and incubated at 27.degree. C. with vigorous shaking on 
a platform shaker at 200 r/min for 1-2 days. All cultures grow well in 1x 
SB medium, producing small (0.1-3 mm) pellets with average diameter of 0.2 
to 0.4 mm. Approximately 3,000 to 5,000 small pellets are formed in 50 ml 
of growth medium in a 125-ml Erlenmeyer flask. The settled pellet is 
transferred into 100 ml of SB medium in 250-ml flasks and allowed to grow 
for an additional 40 h. The settled pellet-inoculum (100%) was then 
transferred into 50 ml of SB medium at various concentrations supplemented 
with 0.05% Tween 20 in 125-ml Erlenmeyer flasks to find the production 
medium. Flasks are incubated at 30.degree. C. at 200 r/min. 
Antifungal activity is determined over time by measuring the 
clear-zone-diameter around the wells with 25 ul of metabolites in 
Aspergillus (Aspergillus niger) plate bioassay. Doubling the metabolite 
concentration increased the diameter of the clear-zone area around wells 
by 2 to 3 mm. The best results are obtained with the 1/3x concentration in 
the clear-zone area around the wells after 19 h. Therefore, 1/3x SB medium 
is used as the production medium (see FIG. 1 and Table 1). The volume of 
the culture and flask size do not influence the development of antifungal 
activity in the plate bioassay when they are varied proportionately. The 
activity is the same in 125-ml Erlenmeyer flasks with 50 ml medium and in 
1,000-ml Erlenmeyer flasks with 400 ml of the same caused by synchrony in 
culture development. The pellets are recycled continuously for several 
months. The pellets can be recycled as long as they remain uncontaminated. 
The antifungal metabolites which are found in the culture fluid, can be 
harvested every day. Production, and thereby harvest frequency, can be 
controlled by the quantity and type of nutritional supplements provided to 
the pellets. Fermentation is enhanced, scaled up, and accelerated by 
promoting secondary metabolism and inhibiting biomass production using 
large (100%) and microscopic pellet-inoculum of the new strain, 
Streptomyces rimosus (SC-36) with a new production medium (1/3x), and by 
varying culture volume and shaking speed. The cessation of pellet growth 
signals secondary metabolism. Treatment of wood and wood products with 
metabolites or the synergistic effect of diluted or undiluted metabolites 
and diluted boron also provided for application as biological control 
agents of wood-atacking fungi. The combination of diluted metabolites and 
diluted glycol ethylene inhibits basidiospore germination of wood-rotting 
fungi and therefore, prevents wood deterioration, 
After following the above procedures inhibition of radial growth of 
sapstain and mold fungi was accomplished. The antagonistic effect of 
Streptomyces rimosus (SC-36) against various sapwood-inhabiting fungi was 
tested in petri plate assays. The growth of all the sapwood-inhabiting 
fungi tested was inhibited at a considerable distance, leaving a clear 
zone of inhibition between the fungi and Streptomyces rimosus (SC-36) on 
nutrient agar (NA) plates after 20 days of incubation. Streptomyces 
rimosus (SC-36) inhibited 81% to 86% of the radial growth of the fungi in 
the petri plates (Table 2). 
The results of the interaction between wood-attacking fungi and metabolites 
in plate bioassay on 2% malt extract agar plates are given in Table 3. A 
basidiospore and conidia suspension of wood-attacking fungi (500,000 
basidiospore for wood-rotting fungi or 200,000 conidia for sapstain and 
mold fungi) was spread into the surface of 2% malt extract agar plates. 
The basidiospore and conidia germination of all wood-attacking fungi was 
inhibited by 25 ul of 1-day-old metabolites from the production medium 
(1/3x) introduced in each well. The conidia and basidiospore germination 
of wood-attacking fungi was completely inhibited, leaving a clear-zone 
around the wells. The diameter of the inhibition or clear-zone are varied 
from 18 to 38 mm per 25 ul of crude metabolites or from 891 to 3,971 
mm.sup.3 per 25 ul of metabolites. 
The antagonistic characteristics of the combination of diluted boron and 
diluted metabolites from Streptomyces rimosus SC-36 against various 
wood-attacking fungi tested in the petri plate bio-assay are shown in 
Table 4. A basidiospore or conidial suspension of wood-attacking fungi 
(500,000 basidiospore or 200,000 conidia) was spread onto the surface of 
2% malt extract agar plates. Diluted (1/4x) or undiluted (1x) metabolites 
with or without a boron-containing co-biocide, namely disodium octaborate 
tetrahydrate (Na.sub.2 B.sub.8 O.sub.13 4H.sub.2 O), was added to the 
precut wells. The boron-containing co-biocide which provided the desired 
level of effectiveness is marketed by US Borax Inc. under the registered 
tradename TIM-BOR. The clear-zone-diameter around the wells was measured 
after incubation at 27.degree. C. and 70% relative humidity for 2 to 10 
days. The growth of all the wood-attacking fungi tested was not affected 
by either TIM-BOR (4% BAE) or diluted metabolites (1/4) alone. However, 
basidiospore or conidia germination of wood-attacking fungi was inhibited 
by combination treatments with diluted (1/4) metabolites from Streptomyces 
rimosus SC-36 and TIM-BOR (4% BAE) in the plate bioassay. In fact, the 
combination of diluted metabolites and TIM-BOR produced a synergistic 
effect that increased the antifungal activity by showing clear-zone area 
around the wells. The combination resulted in a synergistic effect that 
was more effective in the clear-zone area around the wells than that of 
undiluted metabolites (Table 4). The antifungal activity increased up to 
23x by by Ceratocystis pilifera (Table 4). Doubling the metabolite 
concentration typically increased the diameter of the clear-zone area 
around the wells by 2 to 3 mm (S.D. 1.4, 1.6 respectively). 
Table 5 shows that treatment of wood blocks with living actinomycete cells 
of Streptomyces rimosus (SC-36) completely inhibited basidiospore or 
conidia germination. After actinomycete cells were removed by washing with 
a stream of water at the end of incubation, the underlying wood was free 
of fungal attack, discoloration, or deterioration. 
Table 6 shows the inhibition of basidiospore germination on blocks of 
Southern Yellow Pine (Pinus sp) inoculated with brown-rot fungi and blocks 
of sweetgum (Liquidambar styraciflua L.) inoculated with white-rot fungi. 
Blocks were treated with a combination of diluted boron and diluted 
metabolites of Streptomyces rimosus SC-36 for controlling wood-rotting 
fungi using the soil-block procedure (ASTM 1917) with one-half million 
basidiospore inoculum instead of mycelial plug. The undiluted metabolites 
(1x) and the combination of diluted boron (0.5% boric acid equivalent, 
BAE) and diluted metabolites (1/4x) of Streptomyces rimosus SC-36 on wood 
blocks completely inhibited basidiospore germination of the brown-rot 
fungi, Gloeophyllum trabeum and Antrodia carbonica, and the white-rot 
fungi, Phanerocheate chrysosporium and Trametes versicolor, and therefore 
prevented wood deterioration. Neither diluted metabolites nor diluted 
boron inhibited basidiospore germination of brown-rot and white-rot fungi 
after 12- to 16-week incubation. However, diluted metabolites prevented 
weight losses by 95 to 100% (Table 6). 
Table 7 shows inhibition of basidiospore germination on wood blocks of 
Southern Yellow Pine (Pinus sp.) inoculated with brown-rot fungi and 
blocks of sweetgum (Liquidambar styraciflua L.) inoculated with white-rot 
fungi. Blocks were treated with a combination of diluted ethylene glycol 
and diluted metabolites of Streptomyces rimosus SC-36 for controlling 
wood-rotting fungi using the soil-block procedure (ASTM 1917) with 
one-half million basidiospore inoculum. Wood blocks treated with water for 
control or diluted ethylene glycol (40%) were unable to protect the wood 
from fungal growth. However, undiluted metabolites (1x) or the mixture of 
the diluted ethylene glycol (40%) and diluted metabolites (1/4x) inhibited 
basidiospore gemination of wood-rotting fungi and therefore prevented 
weight losses in wood blocks after 12- to 16-week incubation at 27.degree. 
C. and 70% relative humidity (Table 7). The mixture of metabolites (1/4x) 
and ethylene glycol demonstrated a synergistic effect. 
Table 8 shows control of sapstain and mold fungi on wood blocks by the 
combination of diluted boron and undiluted metabolites of Streptomyces 
rimosus SC-36. Blocks of two wood species, Pinus sp. (Southern Pine) and 
Liquidambar styraciflua L. (sweetgum) were used. Blocks were presoaked to 
duplicate the moisture content of green wood and presterilized at normal 
atmospheric pressure and then were dipped for 60 seconds into each 
treatment: distilled water for control, TIM-BOR (1%, 2%, and 4% boric acid 
equivalent, BAE), metabolites (diluted, 1/4x; undiluted 1x; concentrated, 
10x), and the mixture of diluted TIM-BOR and diluted (1/4x) or undiluted 
metabolites. These were then inoculated with 400,000 to 500,000 conidia of 
sapstain or mold fungi and were incubated at 27.degree. C. and 70% 
relative humidity. The mixture of both combinations of TIM-BOR (4% BAE) 
and undiluted metabolites produced a synergistic effect that inhibited 
spore germination of sapstain and mold fungi on wood blocks as effectively 
as 10x metabolites and therefore prevented wood discoloration and 
deterioration. 
Table 9 shows control of wood-attacking fungi on green pine log sections by 
living actinomycete cells, concentrated metabolites, and the combination 
of diluted boron and unconcentrated metabolites of Streptomyces rimosus 
SC-36. The green Southern Pine sapwood log sections were dipped for 60 
seconds into each treatment. The results of the field or simulated field 
trials of living actinomycete cells or the combination of unconcentrated 
metabolites and diluted boron on pine log sections inhibited conidia and 
basidiospore germination of all wood-attacking fungi (Table 9). For the 
field trials, the treated log sections were hung on a fence for 8 weeks 
during the summer at the Vally View test site near Madison, Wis. The 
simulated field trials were undertaken using an air-permeable 
polypropylene bag containing soil from the test site. Microorganisms 
existing in the soil were restricted within the bag and the entire bag was 
incubated for 8 weeks at 27.degree. C. and 70% humidity. Green Southern 
yellow pine sapwood log sections treated with either sodium 
pentachlorophenate (0.1%) or 4% boron (boric acid equivalent, BAE, of 
TIM-BOR) were unable to protect the wood from fungal growth. However, the 
combination of unconcentrated metabolites and 4% boron resulted in a 
synergistic effect that gave greater inhibition of basidiospore and 
conidia germination of all the wood-attacking fungi than that of 
concentrated metabolites or the TIM-BOR alone (Table 9). When the treated 
green Southern pine sapwood log sections were cut just below the area 
where the living actinomycete cells had been applied, these sections were 
free of fungal attack, discoloration, or deterioration. These treatments 
did not alter the physical state of the wood. Even after actinomycete 
cells (SC-36) had dried up, the pine log sections continued to be 
protected from wood-attacking fungi, and therefore the nondecayed living 
actinomycete cells are believed to be useful for long-term protection of 
wood against sapstain, mold, and wood-rotting fungi without altering the 
ecosystem. 
Mostly purified abiotic metabolites are relatively nontoxic with an 
LD.sub.50 of 1400 mg/kg when injected intraperitoneal into 20-g mouse. 
0.09 mg abiotic metabolites inhibited conidia germination of Aspergillus 
niger by showing a 25-31 mm diameter clear-zone (2,156 mm.sup.3) around 
the wells in the plate bioassay. 
TABLE 1 
______________________________________ 
Effect of SB medium concentration on production of 
antifungal metabolites from Streptomyces rimosus SC-36 in 
Aspergillus plate bioassay 
Inhibition zone.sup.a 
SM medium.sup.b 
Diameter Area Maximum 
concentration 
(mm/25 ul) (mm.sup.2 /25 ul) 
Activity 
______________________________________ 
1x 28.5 (0.7) 638 (0.4) 4 days 
1/2x 31.0 (1.4) 755 (1.5) 3-4 days 
1/3x 34.5 (2.1) 940 (3.5) 19 hours 
1/4x 23.5 (0.7) 435 (0.4) 2 days 
______________________________________ 
.sup.a Standard deviation in parentheses. 
.sup.b SB is sporulation broth. 
TABLE 2 
______________________________________ 
Inhibition of radial growth of sapwood-inhabiting 
fungi by Streptomyces rimosus SC-36 in petri plate assay 
Inhibition of radial growth.sup.a 
Fungus (percent) 
______________________________________ 
Sapstain 
Aureobasidium pullulans 
86.0 (2.9) 
Ceratocystis coerulescens 
84.6 (2.4) 
C. minor 84.6 (0.9) 
C. pilifera 81.3 (1.8) 
Mold fungi 
Aspergillus niger 
84.5 (1.6) 
Penicillium spp. 
85.6 (11.0) 
Trichoderma spp. 
81.2 (1.6) 
______________________________________ 
.sup.a Standard deviation in parentheses. 
TABLE 3 
______________________________________ 
Inhibition of basidiospore and conidia ger- 
mination of wood-attacking fungi by metabolites from 
Streptomyces rimosus SC-36 in plate bioassay on 2 percent 
malt extract agar.sup.a 
Inhibition zone.sup.b 
Diameter Volume 
Fungus (mm/25 ul) 
(mm.sup.3 /25-ul) 
______________________________________ 
Brown-rot fungi 
Gloeophyllum trabeum 
22.0 (2.8) 
1,331 (22) 
Neolentinus lepideus 
26.5 (2.1) 
1,946 (12) 
Mold fungi 
Aspergillus niger 31.3 (1.2) 
2,694 (4) 
Penicillium spp. 19.3 (1.2) 
1,024 (4) 
Trichoderma spp. 28.0 (1.2) 
2,156 (4) 
Sapstain 
Aureobasidium pullulans 
30.0 (2.0) 
2,475 (11) 
Ceratocystis coerulescens 
38.0 (2.0) 
3,971 (11) 
C. minor 32.7 (1.2) 
2,959 (4) 
C. pilifera 18.0 (1.2) 
891 (4) 
White-rot fungi 
Phanerochaete chrysosporium 
31.0 (1.4) 
2,643 (5) 
Schizophyllum commune 
31.5 (2.1) 
2,746 (12) 
Trametes versicolor. 
31.0 (1.4) 
2,643 (5) 
______________________________________ 
.sup.a The clearzone-diameter around the wells was measured after 
incubation at 27.degree. C. and 70% relative humidity for two to ten days 
.sup.b Standard deviation in parentheses. 
TABLE 4 
______________________________________ 
Inhibition of conidia and basidiospore germination 
by metabolites of Streptomyces rimosus SC-36 with TIM-BOR 
as a co-biocide for controlling wood-attacking fungi in malt 
extract agar plate bioassay.sup.a 
Inhibition zone 
Wood-attacking TIM-BOR Metab Metab 1/4x Met 
fungi 4% BAE 1X 1/4X TIM-BOR 
______________________________________ 
Brown rot fungi 
Gleophyllum trabeum 
0 20-22 0 35-36 
Mold fungi 
Aspergillus niger 
0 25-27 0 30-32 
Penicillium spp. 
0 20-22 0 26-32 
Trichoderma spp. 
0 25-28 0 35-37 
Sapstain fungi 
Aureobasidum pullulans 
0 30-33 0 18-20 
Ceratocystis coerulescens 
0 36-39 0 36-38 
C. minor 0 24-26 0 33-35 
C. pilifera 0 10-18 0 28-30 
White rot fungi 
Phanerochaete 0 30-32 0 30-36 
chrysosporium 
Schizophyllum commune 
0 30-33 0-10 33-35 
Trametes versicolor. 
0 30-32 0 30-31 
______________________________________ 
.sup.a The clearzone-diameter around the wells was measured after 
incubation at 27.degree. C. and 70% relative humidity for two to ten days 
TABLE 5 
______________________________________ 
Inhibition of conidia and basidiospore germination on wood 
blocks treated with living actinomycete cells of Streptomyces 
rimosus SC-36 for controlling wood-rotting fungi. 
Basidiospore and 
conidia germination.sup.a 
Living actino- 
Control mycete cells 
Wood-attacking fungi 
Gum Pine Gum Pine 
______________________________________ 
Brown-rot fungi 
Antrodia carbonica 
4 4 0 0 
Gloeophyllum trabeum 
4 4 0 0 
Neolentinus lepideus 
4 4 0 0 
Postia placenta 4 4 0 0 
Mold fungi 
Aspergillus niger 
4 4 0 0 
Penicillium spp. 4 4 0 0 
Trichoderma spp. 4 4 0 0 
Stain fungi 
Aureobasidum pullulans 
4 4 0 0 
Ceratocystis coerulescens 
4 4 0 0 
C. minor 4 4 0 0 
C. pilifera 4 4 0 0 
White-rot fungi 
Phanerochaete chrysosporium 
4 4 0 0 
Schizophyllum commune 
4 4 0 0 
Trametes versicolor. 
4 4 0 0 
______________________________________ 
.sup.a The blocks were rated for fungal growth or stain after 8 to 10 
weeks exposure at 27.degree. C. and 70% relation humidity using the 
following scale: 
0 Clean: no stain or mold 
1 Minor: stain or mold covering less than 5% of upper surface. 
2 Light stain: stain or mold covering 5-20% of upper surface. 
3 Moderate stain: stain or mold covering 20-40% of upper surface. 
4 Heavy stain: stain or mold covering more than 40% of upper surface. 
TABLE 6 
__________________________________________________________________________ 
lnhibition of basidiospore germination on wood blocks of Southern Yellow 
Pine for brown-rot fungi and wood blocks of 
sweetgum (Liquidambar styraciflue L.) for white-rot fungi treated with 
the combination of diluted boron 
(Tim-Bor) and/or diluted metabolites of Streptomyces rimosus (SC-36) for 
controlling wood-rotting fungi using 
soil-block procedure (ASTM 1917). 
Basidiospore germination.sup.a and weight loss.sup.b 
Brown-rot Fungi White-rot Fungi 
Antrodia Gloeophyllum 
Phanerochaete 
Trametes 
carbonica trabeum chrysosporium 
versicolor 
Treatment Growth 
% wt loss 
Growth 
% wt loss 
Growth 
% wt loss 
Growth 
% wt 
__________________________________________________________________________ 
loss 
Control 4 1.3 (0.1) 
4 37.5 (2.8) 
4 25.9 (8.2) 
4 27.7 (8.9) 
0.25% BAE 3 1.9 (0.8) 
4 29.4 (4.7) 
4 6.3 (3.6) 
4 16.0 (7.2) 
0.5% BAE 2-3 0.6 (0.8) 
4 24.3 (4.3) 
4 4.0 (1.5) 
4 0 
1.0% BAE 0 0 4 22.1 (4.4) 
4 5.7 92.0) 
2 0 
2% BAE 0 0 2 0.2 (0.5) 
2 2.9 (2.1) 
2 0 
Metabolites (1/4x) 
3 0 3 0 4 1.3 (1.3) 
2 0 
1/4X metabolites + 0.5% BAE 
0 0 0 0 0 0 0 0 
Undiluted metabolites 
0 0 0 0 0 0 0 0 
__________________________________________________________________________ 
.sup.a The blocks were rated for the fungal growth after 12-16 weeks 
exposure at 27.degree. C. using the following scale: 
0, no growth; 1, slight growth; 2, poor growth; 3, moderate growth; 4, 
good growth based on the control. 
.sup.b Standard deviation in parenthesis (%). 
TABLE 7 
__________________________________________________________________________ 
Inhibition of basidiospore germination on wood blocks of Southern Yellow 
Pine for brown-rot fungi and wood blocks of 
sweetgum (Liquidambar styraciflua L.) for white-rot fungi treated with 
the combination of diluted ethylene 
glycol and metabolites of Streptomyces rimosus (SC-36) for controlling 
wood-rotting fungi using soil-block 
procedure (ASTM 1917). 
Basidiospore germination.sup.a and weight loss.sup.b 
Brown-rot Fungi White-rot Fungi 
Antrodia Gloeophyllum 
Phanerochaete 
Trametes 
carbonica trabeum chrysosporium 
versicolor 
Treatment Growth 
% wt loss 
Growth 
% wt loss 
Growth 
% wt loss 
Growth 
% wt 
__________________________________________________________________________ 
loss 
Control 4 1.3 (0.1) 
4 37.5 (2.8) 
4 25.9 (8.2) 
4 27.7 (8.9) 
Ethylene glycol (40%) 
4 1.9 (0.9) 
4 21.7 (4.3) 
4 28.4 (6.9) 
4 39.3 (17.7) 
Metabolites (1/4x) 
3 0 3 0 4 1.3 (1.3) 
2 0 
Metabolites (1/4x + ethylene glycol) 
0 0 0 0 0 0 0 0 
Undiluted metabolites 
0 0 0 0 0 0 0 0 
__________________________________________________________________________ 
.sup.a The blocks were rated for the fungal growth after 12-16 weeks 
exposure at 27.degree. C. using the following scale: 
0, no growth; 1, slight growth; 2, poor growth; 3, moderate growth; 4, 
good growth based on the control. 
.sup.b Standard deviation in parenthesis (%). 
TABLE 8 
__________________________________________________________________________ 
Inhibition of conidia germination on wood blocks by metabolites from 
Streptomyces rimosus SC-36 
with boron co-biocide for controlling mold fungi and sapstain. 
Degree of discoloration.sup.a 
Mold fungi Saptain 
Aspirgillus 
Penicillium 
Trichoderma 
Aureobasidium 
Ceratocystis 
Ceratocyctis 
Ceratocystis 
niger sp. sp. pullulans 
coerulescens 
minor pilifera 
Treatment 
Gum 
Pine 
Gum 
Pine 
Gum 
Pine 
Gum Pine 
Gum 
Pine 
Gum 
Pine 
Gum 
Pine 
__________________________________________________________________________ 
Control 4 4 4 4 4 4 4 4 4 4 4 4 4 4 
1% BAE 4 4 4 4 4 4 4 4 4 4 4 4 4 4 
2% BAE 4 4 4 4 4 4 4 4 4 4 4 2 4 4 
4% BAE 3 3 4 4 4 4 4 4 4 4 4 2 4 4 
Metabolites (1/4x) 
4 4 4 4 4 4 4 4 4 4 4 4 4 4 
Metabolites (1/4x) 
+ 1% BAE 4 4 4 4 4 4 4 4 4 4 4 4 4 4 
+ 2% BAE 4 4 2 2 4 4 4 4 4 4 1 1 3 1 
+ 4% BAE 2 2 0 0 0 1 4 4 0 0 0 0 1 0 
Metabolites (1x) 
4 4 3 3 4 4 1 3 3 1 1 0 1 0 
Metabolites (1x) 
+ 2% BAE 3 3 1 1 3 1 0 0 0 1 0 0 0 0 
+ 4% BAE 0 0 0 0 0 0 0 0 0 0 0 0 0 0 
Metabolites (10x) 
0 0 0 0 0 0 0 0 0 0 0 0 0 0 
__________________________________________________________________________ 
.sup.a The blocks were rated for the fungal growth or stain after 8 weeks 
exposure at 27.degree. C. using the following scale: 
0 Clean: no stain or mold. 
1 Minor: stain or mold covering less than 5% of upper surface. 
2 Light stain: stain or mold covering 5-20% of upper surface. 
3 Moderate stain: stain or mold covering 20-40% of upper surface. 
4 Heavy stain: stain or mold covering more than 40% of upper surface. 
TABLE 9 
______________________________________ 
Inhibition of wood-attacking fungi or pine log 
sections treated with living actinomycete cells, metabolites 
(1x or 10x), and metabolites from Streptomyces rimosus, 
SC-36, with diluted boron as a co-biocide (Exposed in a 
simulated field test or in field trails*) 
Rating for fungal growth.sup.a 
Treatment or stain 
______________________________________ 
Control 4 
0.1% PCP 4 
4% BAE boron 4 
metabolites (1x) 1* or 2 
metabolites (1x) + boron (4% BAE) 
0* or 0-1 
metabolites (1ox) 1* or 1-2 
Living actinomycete cells 
0 
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.sup.a The pine log sections were rated for the fungal growth or stain of 
woodattacking fungi after 8 weeks simulated field exposure at 27.degree. 
C. and 70% relative humidity or field trails* using the following scale: 
0 Clean: no stain or mold 
1 Minor: stain or mold covering less than 5% of upper surface. 
2 Light stain: stain or mold covering 5-20% of upper surface. 
3 Moderate stain: stain or mold covering 20-40% of upper surface. 
4 Heavy stain: stain or mold covering more than 40% of upper surface. 
Having thus described the invention in its preferred embodiment, it will be 
clear that modifications may be made without departing from the spirit of 
the invention. Also the language used to describe the inventive concept 
and the drawings accompanying the application to illustrate the same are 
not intended to be limiting on the invention. Rather it is intended that 
the invention be limited only by the scope of the appended claims.