S.Nami Kartal·Evren Terzi·Tsuyoshi Yoshimura

Abstract Sodium fluoride(NaF)and disodium octaborate tetrahydrate(DOT)are well-known fungicides for preventing and remediating decay in wood and wood structures,due to their excellent diffusion properties in wood.The efficacy of both compounds against drywood termites is,however,inadequately studied;there is a lack of information on the development of proper protection and control methods against drywood termites using these diffusible chemicals.In this study,the threshold values of fluoride and boron against drywood and subterranean termites and decay and mold fungi were assessed in laboratory conditions.Treated Cryptomeria japonica D.Don(sugi)sapwood specimens with four different concentrations of NaF and DOT applied by either dipping or vacuum processes were subjected to attack by drywood termites,Incisitermes minor(Hagen)for 6 weeks.Treated specimens were also tested against subterranean termites,Coptotermes formosanus Shiraki for 3 weeks.Laboratory decay and mold resistance tests were then performed to determine the efficacy of the compounds against fungal attack.Results showed 80% or greater drywood termite mortality in wood specimens treated with NaF at a retention level of 1.7 kg m?3 or more.In DOT treated groups,higher retention levels(greater than 6.8 kg m?3)were needed to reach up to 80% or more termite mortality.In general,weight losses in NaF-treated specimens were slightly lower than those in DOT-treated ones.Retention levels of around 1.2 kg m?3 for NaF and 1.6 kg m?3 for DOT were high enough to reach the JIS K 1571 threshold requirement of weight loss less than 3% in subterranean termite resistance tests.Slightly higher termite mortality values were seen in the DOT than NaF treatments at respective retention levels.In decay resistance tests,no weight losses were seen at a NaF retention level of 2.3 kg m?3;however,a retention level of nearly 4 kg m?3 for DOT was needed for complete protection against both test fungi.In most cases,lower weight losses were obtained in NaF-treated specimens when compared to DOT.In mold resistance tests,neither NaF nor DOT at even the highest retention level was able to completely inhibit mold growth on the surfaces of treated specimens.

Keywords NaF·DOT·Remedial treatments·Termites·Fungi

Introduction

In Japan,the native subterranean termites as well as both drywood and dampwood termites are among various species that can damage structural wood.Drywood termites are able to survive in wood above ground since they do not require contact with soil or moist wood.Severe damage to wooden elements in buildings from drywood termite feeding can occur in as 2 to 4 years(Lebow and Anthony 2012).Indrayani et al.(2007)and Indrayani(2007)found that the western drywood termite, Incisitermes minor(Hagen),one of the most economically important and destructive termites,causes severe damage in wood-based materials and wooden structures and mostly attacks commercial timbers prepared from oak,alder,redwood,walnut,cherry and Douglas fir,among others(Indrayani et al.2007;Indrayani 2007).The first infestation of I.minor in Japan was found in Tokyo in 1976,followed by appearances in west Chiba Prefecture.Since that time,I.minor has been spreading throughout Japan for more than 40 years,and American timbers such as Douglas fir and western hemlock as well as Japanese hinoki and sugi have been attacked(Indrayani et al.2007).Since 1976,records of I.minor in Japan have steadily increased;currently,more than half of the prefectures in Japan are considered to be infested(Yoshimura 2011).Indrayani et al.(2004,2005)stated that attacks by I.minor are most commonly observed in roofing materials in Japanese buildings.The secondmost commonly attacked parts are exterior materials,while materials beneath the floor,flooring materials,and pieces of furniture are hardly ever attacked by I.minor.

There are various control and preventing methods against termites in buildings.Preservative treatment of wood and wood-based components in buildings to prevent termite attack is one of the most common and effective methods.In recent years,the introduction of bait systems,normally used for subterranean termite control,has been used to eliminate I.minor colonies.While gel formulations against I.minor can be considered,laboratory studies have presented high deviations in termite mortalities using gel.Special attractants that spread into the whole attacked areas may be more beneficial in remedial treatments(Indrayani 2007).Remedial treatments employ non-pressure chemicals in applications and distribute the protective chemical into the various areas of a structure.Diffusible chemicals are generally selected for remedial treatments of buildings and wooden materials since these types of chemicals move slowly through water within the wood,and do not fix themselves in the wood(Lebow and Anthony 2012).

In terms of diffusible compounds such as DOT,borax,boric acid,boron has long been used in dip/diffusion processes for treatment of building frames to prevent beetle attack,while fluoride has been used to treat wooden windows and door frames(Freitag and Morrell 2005).Even though remedial treatments by borates are not registered by the Japan Termite Control Association,the use of borates for this purpose is increasing in Japan.Tascioglu et al.(2017)have stated that sodium fluoride(NaF),a colorless,odorless,non-corrosive crystalline solid,has been used effectively for long time in Europe and the USA as a registered preservative for solid wood against wood-degrading fungi and subterranean termites.Due to its high efficacy against decay fungi,NaF has become an important wood-protecting chemical in both preventive and remedial treatments since it diffuses via free water through wood.In addition,as shown by Pan et al.(2015)and Pan and Wang(2015),even low rates of fluoride treatment considerably inhibit termite feeding on wood.The termite resistance choice test of Pan et al.(2015)showed significant inhibition by NaF at the low retention rate of 0.057% ,emphasizing its anti-feedant activity against termites.

Boron-based remedial treatments may have also several advantages for protecting wood against biodeterioration.Boron has high efficacy against both decay fungi and insects and has relatively low toxicity to humans.Furthermore,borates are odorless and colorless(Terzi et al.2017). Boron retention levels in wood for disodium octaborate tetrahydrate(DOT)as determined by the BS EN 117 standard(BS EN 117 2005)are 5.60 kg m?3for protection against termites(Schoeman and Lloyd 1998;Kartal and Yoshimura 2016). The AWPA U1-12 standard demands a retention level of 4.50 kg m?3of inorganic boron compounds for sawn products used above ground and in interior construction,for either dry or humid conditions against C.formosanus termites.However,in case of other termites rather than C.formosanus,lower retention levels(2.70 kg m?3)are advised in the wooden elements above ground(AWPA U1-12 2012).Freeman et al.(2009)have reviewed that control of the drywood termite(Cryptotermes domesticus)requires a retention rate of 0.15% (m/m)boric acid equivalent(BAE)DOT in wood(Drysdale 1994).Even though borate formulations such as DOT are commonly used for drywood and subterranean termite control,they have been suggested to be insufficient for remedial control of drywood termite infestations in laboratory and field tests(Ferster et al.2001).Ohmura et al.(2011)found that the average weight loss of treated sugi sapwood specimens with DOT at 6 kg m?3BAE in drywood termite resistance tests was 2% , satisfying the threshold of the Japan Industrial Standard(JIS K 1571 2010)(≤3% weight loss).They also stated that wood elements used in interior and dry conditions require a DOT retention level of over 1.2 kg m?3BAE to be effective against drywood termites. In general, higher retention levels of wood preservatives seem to be necessary for resistance to I.minor.

The efficacy of NaF and DOT against drywood termites is poorly documented,and the development of proper protecting and control methods against drywood termites requires a much more thorough understanding of the treatment levels to provide effective chemical loading into the wood.Obtaining threshold values for the two compounds in treated wood might be also beneficial to develop better management methods against I.minor.The development of novel preventive and remedial strategies against I.minor might be important for preservation in countries where this species is widespread.In this paper,we performed laboratory tests to assess the thresholds for NaF and DOT against drywood termites as well as subterranean termites in dip- and vacuum-treated wood specimens.Treated specimens were also evaluated against decay and mold fungi.Our objective was to determine the minimum amount of chemical compound needed to arrest drywood termite attack in these chemical control methods.

Materials and methods

Wood specimens

Sugi sapwood wood specimens were used for all tests.The specimens were free of knots or any visible concentration of resins,and showed no visible evidence of infection by mold,stain,or wood-destroying fungi or attack by insects or termites.

Treatments of wood specimens

Wood specimens for pressure treatment were placed in a vacuum desiccator,where the pressure was reduced to 100 mm of mercury by a vacuum pump and held at that pressure for 30 min.Prepared aqueous solutions of NaF and DOT at 0.10,0.25,0.50,and 1.0% concentrations were then poured into the desiccator at sufficient volume such that the specimens would remain submerged after the treatment was completed.After the vacuum step,specimens were left in the solution for 30 min at atmospheric pressure.For dip treatments,the specimens were submerged into the treatment solutions for 3 min in a glass container at atmospheric pressure.Following both treatment processes,specimens were re-weighed after treatment to calculate the retention levels of the compounds.Specimens were then first air-dried overnight and then ovendried at 60°C for 72 h.All test specimens along with untreated controls were then conditioned to 12% equilibrium moisture content (EMC) in a temperature- and humidity-controlled conditioning chamber.

Drywood termite resistance tests

Incisitermes minor(Hagen)pseudergates were collected from infested timbers in Wakayama Prefecture,Japan,and used as test organisms.The termites were extracted from the timbers and kept in plastic containers with lids containing small wood particles as both food sources and harborage.The containers with the termites were kept in a dark termite culturing room of the Research Institute for Sustainable Humanosphere(RISH),Kyoto University,at(27±2)°C and＞65% relative humidity(RH)for at least 1 week before testing to ensure that only healthy termites would be used in the experiment.

Sugi sapwood wood specimens of 30 mm(radial-R)×30 mm(tangential-T)×50 mm(longitudinal-L)were used for the tests.A hole measuring 40 mm(depth)by 10 mm(diameter)was drilled into the center of each wood specimen to accommodate the termites.The specimens with holes were treated with either NaF or DOT solutions by either dipping or vacuum treatments.For the tests,twenty pseudergates of I.minor with no external evidence of wing buds or eyes were placed into the hole of a test wood specimen,and the hole was then covered with a fine mesh screen tightened by two stainless steel wires.Three replicates were employed for each treatment group.

Test units were kept in the same dark termite culturing room at(27±2)°C and 65% RH for 6 weeks.At the end of the tests,the specimens were recovered,oven-dried at 60°C for 72 h,and reweighed to measure their oven-dried weights after exposure to the termites to calculate the percentage of wood weight loss.Termite mortality was also calculated based on the differences in the initial and final numbers of live termites after the 6-week-exposure.

Subterranean termite resistance tests

Sugi sapwood specimens measuring 20 mm(R)×20 mm(T)×10 mm(L)were treated with either NaF or DOT solutions by either dipping or vacuum treatments.The specimens were exposed to the subterranean termites,C.formosanus according to the Japan Industrial Standard(JIS K 1517 2010).An acrylic cylinder(80 mm diameter,60 mm height)whose lower end was sealed with a 5-mmthick hard plaster was used as a container.A test specimen was placed at the center of the plaster bottom of the test container.A total of 150 worker termites collected from a laboratory colony of the RISH,Kyoto University,Japan were introduced into each test container together with 15 termite soldiers.The assembled containers were set on damp cotton pads to supply water to the specimens and kept at(27±2)°C and＞85% RH in a dark termite culturing room for 3 weeks.The weight losses of the specimens due to termite attack were calculated based on the differences in the initial and final oven-dry weights of the specimens after cleaning off the debris from the termite attack.Termite mortality was also calculated based on the differences in the initial and final numbers of live termites after the 3-week-exposure.

Fungal decay resistance tests

Sugi sapwood specimens measuring 20 mm(R)×20 mm(T)×10 mm(L)were treated with either NaF or DOT solutions by either dipping or vacuum treatments. A monoculture decay test was conducted with the brown-rot fungus,Fomitopsis palustris(Tyromyces palustris)(Berk.et Curt)Gilbn.&Ryv.(FFPRI 0507)(TYP)or the white-rot fungus,Trametes versicolor(Coriolus versciolor)(L.:Fr.)Pilat. (FFPRI 1030) (COV) previously grown in Petri dishes including malt-agar growth medium.After measuring oven-dry weights,wood specimens were sterilized with gaseous ethylene oxide.When the mycelium fully covered the medium in the plates,three specimens were placed onto the top of growing mycelium in each plate with a plastic mesh spacer.The plates were then incubated at(27±2)°C and 65% RH for 12 weeks.Twelve replicates were tested for each decay fungus per treatment.The extent of the fungal attack was expressed as the average weight loss(% )calculated from oven-dry weights of 12 wood specimens before and after the decay procedure.

Mold resistance tests

Sugi sapwood specimens measuring 10 mm(R)×5 mm(T)×100 mm(L)were treated with either NaF or DOT solutions by dipping only.No vacuum treatments were applied in these tests.The specimens were evaluated for resistance to mold fungi over a 4-week period according to the American Society for Testing and Materials D4445(ASTM D4445-10 2012).Three mold fungi,Aspergillus niger 2.242,Penicillium chrysogenum PH02,and Trichoderma viride ATCC 20476,were grown and maintained on 2% malt agar at(27±2)°C and 80% RH.A mixed spore suspension of the three test fungi was prepared by washing the surface of individual 2-week-old Petri plate cultures with 10-15 mL of sterile DI water.Washings were combined in a spray bottle and diluted to approximately 100 mL with DI water to yield approximately 3×107spore/mL by hemocytometer count method.The spray bottle was adjusted to deliver 1-mL inoculum per spray.Wood specimens(five specimens per treatment group in each Petri dish)were sprayed with 1 mL of mixed mold spore suspension and incubated at(27±2)°C and 80% RH for 4 weeks.Following incubation,the specimens were visually rated on a scale of 0-5 with 0 describing a specimen that is completely free of mold growth and 5 describing a specimen completely covered with mold growth(Muhcu et al.2017).

Results and discussion

Chemical retention levels,weight loss and termite mortality of wood specimens exposed to drywood termites are shown in Table 1.Figure 1 illustrates the relationship between weight losses and retention levels of NaF and DOT compounds in these tests.In addition,the relationship between termite mortality and retention levels of the compounds is given in Fig.2.In general,dipping treatments with either compound generated less termite mortality and more weight losses in the specimens due to the lower retention levels obtained in the dipping processes.When compared to DOT,NaF treatments in both dipping and vacuum processes resulted in higher termite mortality(Table 1).Termite mortalities increased as the concentrations of the both compounds increased.In general,termite mortality was in good accordance with the weight losses that occurred in the specimens;however,in NaF treatments by vacuum at 0.1 and 0.25% concentrations,higher termite mortality(85% )was seen in the specimens with higher weight loss(0.42% ).Results showed that drywood termite mortalities higher than 80% were obtained when the wood specimens were treated at a NaF retention level of 1.7 kg m?3or more.However,higher retention levels(more than 6 kg m?3)were needed to reach up to 80% or more mortality values when DOT was employed.In the dipping treatments,only the termite mortalities at 0.1% concentration levels were significantly different from the results of other treatments;however,the treatments after 0.25% treatment level showed some trend-level differences.In the dipping treatments the minimum retential levels at which no weight loss was seen was around 0.25 kg m?3for NaF and 0.55 kg m?3for DOT(Fig.1).In the vacuum treatments,only specimens treated with NaF and DOT at 0.1% concentration level showed significant differences based on the weight losses.Considering the weight losses lower than 2% ,all NaF and DOT treatments(both dipping and vacuum treatments)met the performance requirements of the Japan Industrial Standard(threshold limit value of 3% mass loss)(JIS K 1517 2010).

Table 2 shows the weight losses occurring in specimens and termite mortality values in termite resistance tests against C.formosanus.In general,slightly lower weight losses were seen in the specimens treated with NaF than with DOT.However,the compounds had a mixed effect on both termite mortality and weight losses occurring in the specimens.Similar to the drywood termite resistance tests,termite mortalities increased as the concentrations of both compounds increased,and termite mortalities were also in good accordance with the weight losses that occurred.A NaF retention level of nearly 1 kg m?3was needed to obtain weight losses less than 1% ;higher retention levels(between 3 and 4 kg m?3)for DOT resulted in similar weight losses.Figure 3 illustrates the relationship between weight losses and retention levels of NaF and DOT compounds in these tests.No significant differences among the different treatment levels were observed for weight losses in either of the treatments. When the weight losses occurred in these tests were evaluated based on the JapanIndustrial Standard threshold requirement(JIS K 1517 2010)of＜3% ,the threshold was satisfied by NaF and DOT-treated specimens at 0.5% to 1% in dipping and 0.25 to 1.0 in vacuum processes.Relationships between worker termite mortality and retention levels of the compounds are given in Fig.4.No significant differences in soldier termite mortalities were observed in the dipping and vacuum treatments among all treatment levels.When considering worker termite mortalities,there was no significant differences between the treatments at any concentration levels in vacuum treatments;however,in dipping treatments,NaF at 0.50% showed significantly greater termite mortality when compared to DOT at the same level.

Table 1 Retention levels,weight losses and termite mortality in drywood termite resistance tests

Fig.1 Relationship between weight loss and NaF and DOT retention in wood in drywood termite resistance tests(For interpretations,data were compiled from the values in both dip and vacuum treatments)

Fig.2 Relationship between termite mortality and NaF and DOT retention in drywood termite resistance tests(For interpretations,data were compiled from the values in both dip and vacuum treatments)

Table 2 Retention levels,weight loss and termite mortality in subterranean termite resistance tests

Although NaF has been used against insects and termites in wood preservation for a long time, its termiticidal effectiveness to I.minor drywood termites has been poorly studied and documented.In a previous study,the concentrations of NaF in filter paper need to be above 0.5% to kill Reticulitermes flaviceps workers in a no-choice test.Meanwhile,retention of about 0.06% NaF in wood could significantly inhibit termites feeding in a choice test(Pan et al.2015).These results showed that termite mortalities reached a maximum of 47% at the 2% NaF concentration.In our study,0.5% -1.0% solution concentrations of NaF and DOT inhibited drywood termite feeding;however,0.1% NaF in vacuum treatments(0.7 kg m?3retention level in wood)were effective against I.minor attack.Comparing termite mortalities and weight losses in termite resistance tests,NaF demonstrated anti-feedant effects against termites.This phenomenon was also seen in a study by Pan et al.(2015).Ohmura et al.(2011)found that the average mass loss of specimens treated with DOT at 6 kg m?3BAE and exposed to I.minor was 2% ,satisfying the threshold of the Japan Industrial Standard(JIS K 1517 2010).They concluded that wood material to be used indoors should be treated with boron compounds at over 1.2 kg m?3BAE to protect the wood from drywood termites. While their results suggest that higher retention levels are required for resistance to I.minor,our recent study advises lower retention levels of DOT(around 0.1-0.25% concentrations,less than a 1 kg m?3retention level)against drywood termite attack.

Fig.3 Relationship between weight loss and NaF and DOT retention in subterranean termite resistance tests(For interpretations,data were compiled from the values in both dip and vacuum treatments)

Fig.4 Relationship between worker termite mortality and NaF and DOT retention in subterranean termite resistance tests(For interpretations,data were compiled from the values in both dip and vacuum treatments)

Weight losses in specimens exposed to decay resistance tests are given in Table 3.The brown-rot fungus resulted in slightly higher weight losses in the specimens than the white-rot fungus tested.Compared to the untreated controls,both NaF and DOT treated blocks showed reduced decay even at the lowest retention levels,with weight loss values of 29% and 31% for white-and brown-rot fungi,respectively.In the dipping and vacuum treatments,NaF was more effective than DOT in reducing weight loss in the specimens exposed to the fungi.Significant differences in the weight loss specimens exposed to white rot fungus were observed only in the dipping treatments with 0.1% ,0.25% and 0.50% solution concentrations while all treatments for the brown-rot fungus were significant improvements over the controls.In the vacuum treated specimens for both fungi,only the differences for treatments at 0.1 and 0.25% levels were significant.Figure 5 illustrates the relationship between the weight losses and retention levels of NaF and DOT.A retention level of nearly 4 kg m?3for both NaF and DOT treatments was needed to achieve zero weight losses in the specimens exposed to the test fungi.Freitag and Morrell(2005)studied the threshold of boron and fluoride against Postia placenta,Gloeophyllum trabeum and Trametes versicolor fungi in Douglas fir wood.Boron and fluoride were both highly effective at extremely low levels against the decay fungi tested.For P.placenta and G.trabeum,thresholds for sapwood were between 0.40 and 0.44 kg m?3boric acid equivalent(BAE)for boron;the threshold for fluoride was 0.33 kg m?3while weight losses for T.versicolor were too low to allow for threshold determination.In a previous study by Findlay(1953),a boron threshold of 1.0 kg m?3BAE for G.trabeum was suggested.However,higher threshold values have been reported, such as 2.9 kg m?3BAE by Williams and Amburgey(1987)and 2.2 and 4.0 kg m?3BAE by Harrow(1950).In our recent study,a DOT retention level of 1.6-1.9 kg m?3in treated specimens was able to limit weight losses to around 2% or less.For NaF threshold values,Freitag and Morrell(2005)found a target retention of 0.04 kg m?3for fluoride while thresholds for G.trabeum and T.versicolor on the sapwood specimens were about0.022 kg m?3of fluoride.In this study,NaF retention levels of 0.5-1.9 kg m?3were high enough to limit weight losses to around 2% in specimens exposed to the test fungi.

Table 3 Retention levels and weight losses in fungal decay resistance tests

Fig.5 Relationship between weight loss and NaF and DOT retention in fungal decay resistance tests(COV.T.versicolor;TYP:F.palustris)(For interpretations,data were compiled from the values in both dip and vacuum treatments)

Results for mold resistance tests are given in Table 4.In mold resistance tests,lower NaF and DOT retentions(up to 0.8 kg m?3)for dipping treatments were tested.Compared to the drywood and subterranean termite and fungal resistance tests,NaF and DOT compounds were less effective in preventing mold growth.No compounds tested were able to completely inhibit surface mold growth; however,slightly lower mold scores were recorded in both NaF-and DOT-treated specimens when compared to untreated control specimens.Probably much higher retention levels of NaF and DOT compounds are needed for complete protection against mold growth.Previous studies have suggested that DOT does not provide effective protection against mold,and staining fungi and concentrations more than 1% are generally needed(Kartal et al.2005).Clausen and Yang(2007)and Barnes et al.(1989)have discussed that borates are only slightly effective at controlling moldfungi.Clausen and Yang(2003)showed that 5% DOT was unable to substantially inhibit mold fungi.Clausen and Yang(2005)estimated the minimum fungicidal concentration as 7.6% for borates.On the other hand,Klyosov(2007)stated that zinc borate at a loading level of 3% was effective against surface mold growth in composite products and only 3 and 5% of zinc borate prevented mold growth in wood plastic composites with maple flour and high-density polyethylene while a 1% loading level was not effective against molds(Laks et al.2005).Freeman et al.(2009)found that DOT provides limited protective effect against molds,and Micales-Glaeser et al.(2004)showed that around 2% BAE DOT is able to retard mold growth.Lloyd et al.1990 explained the reason borates are unable to prevent mold growth:namely,that high amounts of glucose and fructose in the wood make complexes with the boron,rendering it unavailable to complex with the polyols in the fungi under favorable(moist)conditions for fungi.

Table 4 Retention and mold scoring in mold resistance tests

Conclusion

This study evaluated the efficacy and threshold values of NaF and DOT against drywood and subterranean termites as well as decay and mold fungi.Higher I.minor mortalities were seen in NaF treatments when compared to DOT in all treatments and at levels.Considering weight losses in drywood termite tests,solution concentrations higher than 0.5% for NaF and DOT resulted in no weight losses.Similar results were obtained in the subterranean termite and fungal resistance tests;however,NaF and DOT at the tested concentration levels did not completely inhibit mold growth.

AcknowledgementsWe gratefully acknowledge the Japan Society for the Promotion of Science(JSPS)for providing the first author a research opportunity at RISH,Kyoto University,Japan under the JSPS Invitational Fellowship.