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Genetic relationships of local infestations by Anobium punctatum, Xestobium rufovillosum and their associated predator Korynetes caeruleus from buildings in North-Eastern Germany
2021 - IRG/WP 21-10982
Wood-destroying pests such as Anobium punctatum and Xestobium rufovillosum cause damage to art and cultural objects as well as to buildings. Monitoring population dynamics of pest species as well as of their naturally occurring counterparts are an essential part in the development of biological control measures as alternatives to conventional wood protection. Therefore, both the dispersal and homogeneity of pest and beneficial insect populations across multiple sites and buildings were investigated in the present study using DNA barcoding. Specifically, beetles of Anobium punctatum (de Geer 1774) (Coleoptera, Ptinidae), Xestobium rufovillosum (de Geer, 1974) (Coloeptera, Ptinidae), and Korynetes caeruleus (de Geer 1775) (Coleoptera, Cleridae) were collected from buildings at four different sites in Mecklenburg-Western Pomerania, North-Eastern Germany. DNA analysis was performed using mitochondrial cytochrome c oxidase subunit I (COI). For A. punctatum, low base pair variability was found in the gene segment studied (4-5 SNPs) within one building (Greven) and between four spatially separated sites. Conversely, in X. rufovillosum, the sequences from two sites studied were homogeneous within a site but differed between locations by nine base pair positions (SNPs). The main result of this study is that the pests A. punctatum and X. rufovillosum showed a higher variability in the investigated gene segment than the natural counterpart K. caeruleus.
C von Laar, C Baar, R Plarre, D P McMahon


What can DNA fingerprinting, aggression tests and morphometry contribute to the identification of colonies of the Formosan subterranean termite
2000 - IRG/WP 00-10371
Multilocus DNA fingerprinting, aggression tests and morphometry were compared to evaluate their potential for the identification of colonies of the Formosan subterranean termite, Coptotermes formosanus (Isoptera: Rhinotermitidae) in Hawaii. DNA fingerprinting separates the termites from all studied collection sites. Since the genetic similarity between termites from different collection sites lies in the range of the genetic background similarity in the population, collection sites in this study represent independent colonies. No significant differences could be found in the intra- and intercolonial aggression levels. While aggression tests do not support colony identification, morphometric measurements do show differentiations between colonies. However, classification of individuals to their original colony does not reach the 100% success provided by genetic analyses. No correlation between genetic similarities and aggression levels or morphometric distances could be found. This suggests that neither aggression levels nor morphometric parameters are significantly influenced by genetic factors in this species. Genetic studies appear to be the most useful approach to the identification of colonies and the analysis of small scale population structures in C. formosanus.
C Husseneder, J K Grace


Using DNA probes to characterize the metabolic pathway of pigment production in several wood-staining fungi
1996 - IRG/WP 96-10146
During shipment and storage, lumber is susceptible to sapstain, a wood discoloration caused by fungi. Currently kiln drying and chemical applications are used to control sapstain. However, the chemicals used to protect wood have a broad range of action, and so can affect other organisms. In addition, in Canada most of these chemicals are under temporary registration. Thus there is a need to develop alternative strategies for wood protection. Instead of inhibiting fungal growth one approach would be to block the production of the fungal pigment responsible for discoloration. Very little is known about pigmentation in sapstaining fungi. However, in Ophiostoma piliferum, Ophiostoma piceae and Alternaria alternata, the discoloration is due to melanin production. While fungal melanin can be synthesized by several different metabolic pathways, Alternaria alternata produces melanin via the dihydroxynapthalene (DHN) pathway. To determine whether several common sapstaining fungi also utilize this pathway, a gene from Alternaria alternata's DHN pathway was radiolabeled and used as heterologous probe to screen Southern blots from six species of staining fungi. The results suggest that most of these fungi have homologous (similar) genes for the enzymes of the DHN pathway. The genes from Ophiostoma piceae will be further characterized, to facilitate more directed development of anti stain strategies.
R Eagen, S Riecken, J Kronstad, C Breuil


Application of DNA fingerprinting methods to identify biocontrol strains of fungi imperfecti
1994 - IRG/WP 94-10068
We have analyzed a number of biocontrol strains of Trichoderma harzianum and other Trichoderma strains with the methods DNA fingerprinting and PCR fingerprinting to differentiate and identify these strain which is not possible with morphological or biochemical methods. We could differentiate even gamma-ray induced mutants from each other as well as different strains form the same and different species. The gamma-ray induced mutants are patented strains of Trichoderma harzianum and are used for biocontrol of wood-decaying fungi. The ability to differentiate the mutants is very important to identify the patent strain in nature because its "fingerprint" is unique and different of wildtypes of the same strain. Furthermore the two methods can be used to follow the path of a strain genotype in nature which could be of importance for ecological questions. We found that "DNA fingerprinting" as well as "PCR fingerprinting" are powerful methods for this task although PCR fingerprinting is faster.
A Schlick, K Kuhls, W Meyer, E Lieckfeld, T Börner, K Messner


rDNA-ITS sequence of Serpula lacrymans and other important indoor rot fungi and taxon-specific priming PCR for their detection
1999 - IRG/WP 99-10298
Taxon-specific priming polymerase chain reaction (TSPP) is a powerful molecular tool for fungal diagnosis. For its application to indoor rot fungi, the internal transcribed spacer (ITS) of the ribosomal DNA (rDNA) of the main fungal species causing wood rot in European buildings was amplified with the polymerase chain reaction (PCR). The ITS region was sequenced. The complete sequences are presented. From base sequence divergency among the fungi, species-specific oligonucleotide primers were designed for TSPP. These marker molecules were suitable for the differential diagnosis of the dry rot fungus, Serpula lacrymans, the wild merulius, S. himantioides, the oak polypore, Donkioporia expansa, the brown cellar fungus, Coniophora puteana, the broad-spored white polypore, Antrodia vaillantii, the sap polypore, Tyromyces placenta, and the yellow-red gill polypore, Gloeophyllum sepiarium. Each specific marker identified isolates of its respective target species. Cross reaction with 'foreign' fungi was the exception. Species identification from unknown field samples from rot damage in buildings is also possible, because DNA from contaminating organisms does not response to the specific primers. Our variant of the technique is fast, because no preceding fungal pure cultures, no special DNA extraction/purification, and no restriction by endonucleases are necessary.
O Schmidt, U Moreth


Aureobasidium or Hormonema? A Genetic Approach.
2004 - IRG/WP 04-10529
Aureobasidium pullulans is the main organism causing disfigurement of coatings on wood and the surface of exposed timber. This disfigurement of timber in-service is referred to as “bluestain in-service”. A. pullulans is also associated with the sapstaining of dead wood in the forest and in-service. A. pullulans is noted for its highly variable growth forms (polymorphisms). This variability presents problems when identifying environmental isolates, due to the morphological similarity of other blue stain fungi, mainly Hormonema dematioides. Molecular analysis has proven to be both accurate and reliable in distinguishing between these two morphologically similar bluestain fungi. The method was devised as part of a bigger research project designed to look in detail at the polymorphisms and physiology of Aureobasidium-type fungi. The establishment of a working library of strains and isolates was aided by many members of the International Research Group on Wood Preservation. The devised method illustrated the existing problem of distinguishing between A. pullulans and H. dematioides. The protocol has been devised to simplify what can be a long complicated procedure. By sequencing direct from amplified PCR (Polymerase Chain Reaction) products, there is no need for cloning large DNA fragments into vectors, followed by sequencing. The extraction method offered here will give DNA of sufficient purity to allow for further genetic analysis if required. This method allows consistent differentiation between isolates of A. pullulans and H. dematioides. Aureobasidium pullulans is the main organism causing disfigurement of coatings on wood and the surface of exposed timber. This disfigurement of timber in-service is referred to as “bluestain in-service”. A. pullulans is also associated with the sapstaining of dead wood in the forest and in-service. A. pullulans is noted for its highly variable growth forms (polymorphisms). This variability presents problems when identifying environmental isolates, due to the morphological similarity of other blue stain fungi, mainly Hormonema dematioides. Molecular analysis has proven to be both accurate and reliable in distinguishing between these two morphologically similar bluestain fungi. The method was devised as part of a bigger research project designed to look in detail at the polymorphisms and physiology of Aureobasidium-type fungi. The establishment of a working library of strains and isolates was aided by many members of the International Research Group on Wood Preservation. The devised method illustrated the existing problem of distinguishing between A. pullulans and H. dematioides. The protocol has been devised to simplify what can be a long complicated procedure. By sequencing direct from amplified PCR (Polymerase Chain Reaction) products, there is no need for cloning large DNA fragments into vectors, followed by sequencing. The extraction method offered here will give DNA of sufficient purity to allow for further genetic analysis if required. This method allows consistent differentiation between isolates of A. pullulans and H. dematioides.
M J Ray, D J Dickinson, M Buck


Development and Implementation of a DNA – RFLP Database for Wood Decay and Wood Associated Fungi
2004 - IRG/WP 04-10527
We are developing Restriction Fragment Length Polymorphism (RFLP) and Internal Transcribed Spacer (ITS) sequence databases for wood decay basidiomycetes and other fungi associated with wood. These databases currently house information for 39 fungal species consisting of 9 brown-rot basidiomycetes, 12 white rot basidiomycetes, 1 soft rot, 1 stain fungi, and 16 molds or other ascomycetes or imperfect fungi. We plan to add 6 brown rot, 14 white rot and 8 other species that we have in culture by summer 2004. Using the RFLP database, we were able to identify wood decay basidiomycetes that were isolated from a local forest and that could not be distinguished based on morphology. In addition, RFLP data confirmed identifications of several other wood associated fungi. One of our ultimate goals is to establish a web-based database of wood basidiomycetes and wood-associated fungi emphasizing ITS sequence and RFLP pattern data plus morphological characteristics in one searchable system. These databases will be able to provide sensitive and reliable identifications of the wood decay community and important wood decay fungal species.
S V Diehl, T C McElroy, M L Prewitt


Computerised data acquisition and barcode technology - Examples of its application and use in wood preservation research
1992 - IRG/WP 92-2404
Experimental samples in wood preservation research are usually individually labelled with either some form of durable marking pen or more commonly with a stamped metal or plastic tag. The label must remain intact and legible for the duration of the experiment which, in the case of a field trial, can be many years. Preservative performance data have traditionally been recorded manually on a data sheet. Since the advent of the personal computer there has been a progressive switch towards the new technology for manipulation of the recorded data. Unfortunately most information is laboriously entered into the computer by hand. This is both inefficient and prone to human error. In recent years barcode technology and portable field computers have progressed to the point where they are relatively inexpensive and simple to use. This paper describes one application of this technology to wood preservation research. The relative advantages of the new technology are discussed.
K J Archer


A comparison of fatty acid and molecular profiles for identification of wood colonizing basidiomycota
2003 - IRG/WP 03-20278
Two methods that are currently being employed to detect and identify wood decay fungi are Fatty Acid Methyl Ester (FAME) analysis and Restriction Fragment Length Polymorphism (RFLP) analysis. A FAME library and RFLP library for 9 species and up to 10 strains within each species have been developed. The profiles generated by these methods have been compared for each species and strain to test for accurate species identification and consistency within species strains. Single FAME libraries were created for Trametes versicolor and T. pubescens, indicating that all isolates of these species clustered as a single species. More than one FAME library was created for P. chrysosporium, P. sordida, Gloeophyllum trabeum, G. sepiarium, G. striatum and T. hirsuta, indicating that not all isolates of these species clustered as a single species. Blind samples could not distinguish between isolates of T. pubescens and T. hirsuta nor G. sepiarium and G. striatum. Restriction enzyme digestion with AluI, TaqI, RsaI, HaeIII, DraI, and Hinf reliably resolved species; however, no single restriction enzyme profile was sufficient to resolve all of the species groups. The species groups could be resolved with the combined information from Alu I and Taq I restriction profiles, while the addition of Hinf and /or Hae III restriction profiles improved resolution within species and between species. The addition of all other restriction profiles added more but not significantly more resolution within and between species. The ultimate goal of this work is to develop a sensitive and reliable assay to survey the wood decay community and accurately detect important wood decay fungal species.
T C McElroy, L Prewitt, S V Diehl


Detection of the dry rot fungus Serpula lacrymans by amplified ribosomal DNA restriction analysis
1998 - IRG/WP 98-10245
Isolates of the dry rot fungus Serpula lacrymans and the wild merulius S. himantioides were investigated by amplified ribosomal DNA restriction analyses (ARDRA) of the internal transcribed spacers (ITS). The technique uses the polymerase chain reaction (PCR) to amplify the variable region of the ITS between the conserved 18S and 28S genes of the nuclear ribosomal DNA (rDNA). The ITS region of all isolates of Serpula lacrymans and S. himantioides was successfully amplified. The length of the amplified product was about 630 bp. Subsequent double digest of the ITS with the restriction endonucleases Haelli and Taql separated the closely related fungi by species-specific fragments. Thus, ARDRA-ITS proved to be suited for the detection of the dry rot fungus.
O Schmidt, U Moreth


Detecting fungal DNA in treated and non-treated wood
2007 - IRG/WP 07-10621
Isolating fungi from wood has long involved culturing on selective media followed by identification using various keys. This process can be cumbersome, costly, and, most importantly, not always capable of detecting all of the fungi present. The recent development of molecular methods for isolation and identification of fungi has created tremendous opportunities for expanding our knowledge of the microbial ecology of wood. Among the issues with these methods is the ability of some wood extracts to inhibit the amplification of DNA by the polymerase chain reaction (PCR), but it is unknown whether the resins in wood-based composites and wood preservatives are similarly inhibitory. A variety of wood species and composites were inoculated with wood decay fungi and the resulting fungal DNA was extracted from the decayed wood. It was not always possible to successfully extract DNA. In order to determine if the failure was caused by interferences or with a lack of fungal DNA present, known quantities of purified fungal DNA were added to sawdust of various species with or without preservatives during the extraction procedure and then recovered. Fungal DNA was recovered from nearly all samples including red oak, OSB, plywood, wood plastic composites as well as wood treated with pentachlorophenol, CCA, ACZA and other copper based preservatives.
C Freitag, M Freitag, J Morrell


A step towards a better understanding of fungal colonization of modified wood - QRT-PCR studies
2008 - IRG/WP 08-10653
The area of wood protection is in a period of change. New wood protection systems have been developed while their mode of action remains insufficiently understood. The development of molecular methods provides potential tools to investigate the interaction between modified wood and decay fungi. One small step to tackle some of the unsolved questions about the mode of action of modified wood is taken in this study. A specific and quantitative real-time PCR (QRT-PCR) assay was now established for identifying and quantifying early stages of fungal colonisation in modified wood and for profiling growth dynamics of the white-rot fungus Trametes versicolor through different stages of decay. QRT-PCR of colonisation of three different wood modification systems (acetylation, furfurylation, thermal modification), two reference treatments (Cu-HDO, CCA) and Scots pine sapwood as control was performed. Incubation time was 2, 4, 6, 8 and 10 weeks. While the fungal colonisation in untreated control samples showed a continuous increase during the experimental period, the amount of fungal DNA in modified wood had an initial peek after two weeks, followed by a gradual decline. The furfurylated samples had lower fungal colonisation than all the other treatments except for Cu-HDO. In the reference samples of copper nearly no fungal colonisation was found at all during the study period of 10 weeks. In the CCA samples there was an initial colonization peek at 2 weeks, but for the remaining part of the experimental period the colonization level remained very low.
G Alfredsen, A Pilgård, A Hietala


Analysis on dynamics of basidiomycetes in decayed wood by a method using non-specific amplification of DNA
2009 - IRG/WP 09-20426
A combination of non-specific amplification of DNA by Phi29 DNA polymerase and specific amplification of the internal transcribed spacer (ITS) region of ribosomal DNA (rDNA) by polymerase chain reaction (PCR) was applied to elucidate the ratio of basidiomycete species inhabiting decayed wood from an outdoor bench. After more than 8 hours incubation with Phi29 DNA polymerase, concentration of DNA were reached to 50 ng/μl, which was approximately 1,000 times higher than that of DNA in the original sample. It was revealed that the increase of PCR cycles biased basidiomycete species from original ratio, whereas longer incubation time of the non-specific amplification less affected on it, suggesting the advantage of amplification by Phi29 DNA polymerase to analysis on dynamics of basidiomycetes in decayed wood.
T Wada, K Igarashi, M Samejima


Detection of Wood Destroying Fungi Using DNA Microarray Technology
2010 - IRG/WP 10-20435
Wood decay fungi of the phylum Basidiomycota cause serious damage to wooden constructions and building elements. The elimination and the appraisement of fungal decay require an assured species identification. Conventional fungal diagnostics are mainly based on morphological characteristics by macro- and microscopy analysis. For some years, standardised and highly sensitive molecular methods focussing on DNA analysis are increasingly used. However, these methods still have some disadvantages in routine diagnostics, like long operating time and infeasibility of analysing mixed samples, where several fungi occur simultaneously in one sample. To overcome these drawbacks, a DNA microarray for the confident identification of the 27 most important wood destroying basidiomycetes was developed that is already used in the laboratory routine. First of all, species-specific DNA probes were generated on the basis of validated rDNA ITS sequences from 121 fungal isolates. The specificity of these DNA probes was assured using the polymerase chain reaction (PCR) and microarray detection technology. Afterwards, the microarray detection procedure that is based on the arrayed ligation reaction (ALR) was optimized and validated. For a comprehensive field testing and validation of the newly developed detection technique, 36 samples from decayed timber have been collected. The test specimens were analysed by microarray analysis as well as by conventional methods like microscopy analysis and ITS sequencing determination in parallel. Thereby, at 33 samples the fungus species could be addressed directly by microarray analysis. In the case of three samples, the concentration and the quality of the DNA preparations were insufficient for a distinct diagnosis. In these samples the conventional morphological identification as well as the sequencing determination failed, too. The comparison methods provided a higher level of failure, in which 22 % (ITS sequencing), respectively 33 % (conventional diagnostics) of the samples could not be identified to the species level. The field test showed that the microarray based diagnostic system ensures a fast and reliable identification of wood decay fungi. Since January 2010, the Mycotype® BasidioQS Microarray Detection Kit is available as the first commercial product. This DNA microarray was developed in a research cooperation between the Institute of Wood Technology, Dresden (IHD) and the Biotype Diagnostic GmbH, Dresden.
K Jacobs, N Rangno, W Scheiding, B Weiss, D Müller, C Hiller, W Brabetz


DNA-based tools for rapidly detecting, quantifying and monitoring ophiostomatoid fungi on beetles, in trees and wood products
2010 - IRG/WP 10-20450
Approximately half of the trees harvested for commercial purposes are lost because of native or introduced insects or insect-vectored microorganisms. Ophiostomatoid fungi, which are well adapted to dissemination by insects, include ~140 species of saprobes and pathogens. They are present worldwide, have high economical impact and many are subject to quarantine regulation. Thus, it is necessary to quickly and efficiently characterize which fungi are associated with beetles, host trees, and wood products, and to establish the relative degree of damage or quarantine risk associated with these pests. During the past decade, molecular biology approaches have provided diverse DNA-based diagnostic tools that have gradually made their way into forestry and forest products industries. Such tools are typically sensitive, able to detect pests without the need to grow the microorganisms and able to provide quick answers. The new tools are rapidly becoming the new standard for detecting, monitoring and quantifying pests worldwide. In this paper we will review the molecular techniques available, as well as some of our own results, discussing issues like single and multiple gene amplification, sequencing and phylogeny, genetic markers to differentiate species or individual in a species, quantitative PCR and meta-genomic sequencing.
L Khadempour, Young Woon Lim, S Massoumi Alamouti, C Breuil


The effects of acetylation level on the growth of Postia placenta
2011 - IRG/WP 11-10751
To understand the defence mechanisms utilized by decay fungi when exposed to different wood protection systems the study of gene expression can give us some answers. When the DNA sequences are known, primers can be designed to detect transcripts of genes with gene products related to basic cellular processes and hyphal growth. The characteristic gene products induced in different fungi by different wood protection systems can be identified. Studies on the expression of fungal genes will give us a better understanding of the fungal degradation of wood and we can optimize wood protection systems. Hence, no single technique will give us the answer to all questions about the decay of wood we need to gather small pieces of the puzzle using different approaches. The aim of the present study was to investigate the effects of acetylation level on the growth of Postia placenta with regard to amount of total DNA and gene expression targeting 7 different genes. This paper presents preliminary results after 4 weeks of incubation. The results presented in this paper are parts of a larger project which reaches over a period of 36 weeks with sampling times after 12, 20, 28 and 36 weeks. We found no mass loss in the acetylated samples after 4 weeks of incubation in a modified soil-block test. The presence of P. placenta DNA and the absence of mass loss could indicate on an inability of the mycelia to establish a wood exploitation phase. Two genes related to carbohydrate metabolism were expressed in a higher amount in P. placenta during growth on untreated wood than during growth on acetylated wood. However, for a third gene, also related to carbohydrate metabolism, the relationship was the opposite. Two genes related to oxidative metabolism were expressed in a higher amount in P. placenta during growth on acetylated wood than during growth on untreated wood and another two genes related to oxidative metabolism showed inconsistent results.
A Pilgård, G Alfredsen, C G Fossdal, C J Long II


Variation in two Postia placenta strains, MAD-698-R and FPRL 280 – mass loss, DNA content and gene
2012 - IRG/WP 12-10781
Brown-rot fungi such as Postia placenta are common inhabitants of forest ecosystems and brown rot fungi are also largely responsible for the destructive decay of wooden structures. The aim of this study was to compare two commonly used strains of Postia placenta – MAD-698-R and FPRL 280. Scots pine sapwood samples were exposed for two and eight weeks to both fungal strains. The following was investigated: mass loss, fungal gDNA content and gene expression. A significant difference was found in mass loss after eight weeks between the P. placenta strains MAD-698-R and FPRL 280. MAD-698-R gave higher mass loss than FPRL 280. However, MAD-698-R seems to have a slightly slower growth rate than FPRL 280, reflected in lower gDNA content after two weeks. After eight weeks of exposure the gDNA content dropped and no significant difference was found between MAD-698-R and FPRL 280. We observed differences in mass loss, colonization-rate and gene expression between the two Postia strains. Results suggest significant differences in the regulation of key lignocellulose degrading enzymes between MAD-698-R and FPRL 280.
N Thaler, G Alfredsen, C G Fossdal


Analyses of premature failure of utility poles
2012 - IRG/WP 12-40584
In this study a total number of 18 utility poles of Scots pine (Pinus sylvestris) impregnated with a copper-chromium containing preservative were investigated. They were part of different lower voltage transmission lines in the western part of Germany and failed before predicted minimum service life. All poles in this study were less than 15 years in use. The type of decay and fungi were evaluated. Furthermore the copper content of undecayed areas of the same poles was analysed. The poles were strongly decayed in the sapwood area as well as in sap- and heartwood areas. The major part of the pole sections were infested by brown rot particularly by the copper tolerant fungi Antrodia spp.. The species of fungi were determined by molecular diagnostics. The analyses of Cu content of undecayed areas of poles after utilization showed a wide range (0.7- 4.7 kg/m³). The Cu content before utilization was not known.
S Bollmus, N Rangno, H Militz, A Gellerich


The effects of acetylation level on the growth of Postia placenta over 36 weeks
2012 - IRG/WP 12-40589
Genomic sequencing gives us a tool to systematically and rapidly discover novel genes, how their products function in the cell, and explore their interactions. When the DNA sequences are known, primers can be designed to detect transcripts of genes with gene products related to basic cellular processes and hyphal growth. The characteristic gene products induced in different fungi by different wood protection systems during decay can be identified. This knowledge will give us a better understanding of the fungal degradation of wood and we can optimize wood protection systems. Hence, no single technique will give us the answer to all questions about the decay of wood we need to gather small pieces of the puzzle using different approaches. The aim of the present study was to investigate the effects of acetylation level on the growth of Postia placenta with regard to amount of total DNA and gene expression targeting six different genes. This paper presents preliminary results after 36 weeks of incubation. We found no mass loss in the acetylated samples treated to a high treatment level after 36 weeks of incubation in a modified monoculture soil-block test. The presence of P. placenta DNA and the absence of mass loss could indicate on an inability of the mycelia to establish a wood exploitation phase. The results also showed that P. placenta increased the expression of AlO (involved in production of H2O2), cytochrome P450 (related to breakdown of toxic compounds), and QRD (involved in generating biodegradative hydroxyl radicals via redox cycling) along the incubation time, growing on acetylated wood treated to a high treatment level.
A Pilgård, G Alfredsen, C G Fossdal, C J Long II


Biodegration of treated wood waste by native fungal communities of tropical soil in French Guiana
2012 - IRG/WP 12-50285
Woods have been protected with fungicides for a long time, and the effects of these fungicides on soil after being leached into the ground have turned out to be a true environmental issue. It is in this perspective that we are proposing to study fungal communities of these contaminated woods in a purpose of bioremediation. Most of precedent studies have focused on ability of some Basidiomycetes and white rot fungi to degrade these biocide products. Treated and reference (non-treated) woods samples have been incubated in containers of forest soil in Guyana. The first two samplings of these woods and soils have been realized five months apart. A crop and molecular study allowed us to isolate and identify forty strains of Ascomycetes able to develop on wood and resist xenobiotics. Until now, no Ascomycete was known to resist xenobiotics. Furthermore, a study of fungal communities of the woods and soil were done by D-HPLC and SSCP, and then analyzed by ACP. According to these analyses, biocides are leached in the soil and have an impact on these fungal communities, which are different depending on time of sampling and the way wood is processed.
A Zaremski, L Gastonguay, C Zaremski, F Chaffannel, J Beauchêne, G LeFloch


Characterization of test fields
2013 - IRG/WP 13-20508
Test field characteristics and impact of test fields on wood degradation is important when testing wood protection. The current EN 252 standard has no requirement for knowledge of decay hazards, but most commonly a test field is known as a “brown”,- “white”,- or “soft rot” field. To understand which decay hazard wood preservatives are tested against, each test field should be characterized. Our purpose here is to outline a number of different approaches that we have been using to characterize four test fields belonging to the Department of Forest Products, SLU, Sweden. For characterization we isolated fungi from EN 252 Scots pine stakes by using different media and identified the isolated fungi by DNA sequencing. DNA profiles were established for Scots pine mini- stakes as well as microscopy characterization of stake cross sections for decay type and degree of attack. MOE and mass loss was measured for the mini-stakes at one test field. For all fields, the pick test of EN 252 stakes were performed according to the European Standard EN 252. A summary of problems and possibilities of the techniques are presented. Our results show that by combining different approaches like molecular techniques with microscopy and/or strength tests, it is possible to obtain good characterization of test fields, the inherent fungi, and their decay capacity cannot be described by using only one approach.
U Råberg, N Terziev, G Daniel


Extraction and analysis of DNA from green and seasoned timber as basic methods for determination of wood species and origin
2013 - IRG/WP 13-20523
Against the background of the European timber trade regulation EUTR, commenced to law by March 2013, the determination of wood species and tracing of its origin is getting a great importance. A promising approach for establishing fast and reliable tracking systems for wood products is DNA analysis. A critical point is the extraction of analysable DNA from the wood and its lignified cell walls. Thus, aim of the work was the analysis of DNA content in different areas of the cross section of green and dry, well-seasoned wood from pine, spruce and beech, using quantitative, real-time PCR analysis. DNA from green timber was successfully extracted and amplified with primer set ycf3hm at all three wood species and zones of cross-section. Whereas the results with pine and spruce confirmed the expectation, that the DNA content decreases from cambium to pith, this was not observed with beech. DNA detection with 20 year old well-seasoned wood was successful only with beech. Further systematic studies are necessary to get better information about the influence of wood processing and ageing on DNA quantity and quality.
K Jacobs, H Mende, W Scheiding


Molecular characterization and biodiversity of wood-decaying fungi in French Guiana
2014 - IRG/WP 14-10825
Fungi from tropical regions are currently under-represented in the classification system. Indeed, difficult access to tropical forests and irregular occurrence carpophores make it complicated to study fungus species in such environments, unlike in European zones where fungal diversity and taxonomy are better known. The purpose of this work was to enhance classification by integrating new data that would bring out the importance of certain traits of these fungi, and provide a clearer understanding of how the biodiversity of fungi from the forest ecosystems of French Guiana is organized, particularly those causing wood decay through white rot, brown rot or soft rot. In our study, we chose to work in the zone comprising the internal transcribed spacers ITS1 and ITS2, which are relatively variable, and the 5.8 S small ribosomal subunit, which is not highly variable. The primers ITS 1(5’-TCCGTAGGTGAACCTGCGC-3’) and ITS 4 (5’-TCCTCCGCTTATTGATATGC-3’), specific to fungi, were chosen for this taxonomic analysis of the studied species. This study was carried out on 101 fungus fruiting bodies at the Paracou forest site in French Guiana. Of those 101 fungi, 72 were identified by BLASTn. Four species were Ascomycetes of the genus Muscodor and Xylaria. The other 68 species, all in the class of the Basidiomycetes, were divided into the following orders: 31 Agaricales, 1 Atheliales, 2 Boletales, 1 Gomphales, 12 Polyporales, 1 Trechisporales and 1 Tremellales. There was also an indeterminate taxon very similar to the lichens. Within the order Polyporales, the main genera were found, such as Antrodiella, Coriolopsis, Fomitopsis, Ganoderma, Lentinus, Pycnoporus, Steccherinum, Trametes, Fomitoporia. All these fungi have the particularity of causing wood decay.
A Zaremski, L Gastonguay, C Zaremski, J Beauchene


Microbial Community Analysis of Naturally Durable Wood in an Above Ground Field Test
2014 - IRG/WP 14-10826
This paper presents preliminary results of an above ground field test wherein eight naturally durable wood species were exposed concurrently at two sites in North America. Surface samples were taken at regular intervals from non-durable controls and compared to their more durable counterparts. Terminal Restriction Fragment Length Polymorphism was performed to characterize the microbial (bacteria, fungi, and basidiomycetes) communities present. Differences were noted among wood species and seasonal shifts in microbial diversity were noted at both sites. Attempts to correlate diversity indices with decay ratings were unsuccessful, but differences in species richness were noted for several of the naturally durable species. Western red cedar had significantly fewer bacterial species compared to other wood species. Fungal and basidiomycete species richness differed due to site and fungal species richness increased with increased exposure. Clustering of fungal and basidiomycete communities suggests seasonal patterns of colonization at both sites, but was more defined in the more southern site; Saucier, MS (MS). Future analyses will focus on comparison of years to model successional patterns of bacteria, fungi, and basidiomycetes.
G T Kirker, S V Diehl, P K Lebow


Communities of mold fungi in moisture damaged building materials
2014 - IRG/WP 14-20542
The critical conditions needed for the development of mould and decay fungi have been modelled for different building materials. However, current knowledge of indoor microbes growing on building materials relies on culture-based methods and more advanced molecular biological techniques should be employed to study the complex microbial communities in building materials. In this paper molecular biological techniques were optimized and used to study microbial diversity in building materials exposed to different moisture conditions. Different naturally contaminated and inoculated building materials were exposed to different humidity conditions (relative humidity 90% and 98%) in laboratory-scale experiment. The DNA extraction method was optimized to different building materials and microbial communities were studied by fungal ITS region targeted PCR-DGGE and sequencing. Fungal communities differed between building materials and humidity conditions. In RH 90% the majority of the sequences obtained belonged to genus Aspergillus. As expected, in RH 98% the fungal community was more diverse containing e.g. genera Penicillium, Aspergillus and Oidiodendron. The fungal diversity was highest in wood-based building materials.
E Sohlberg, H Viitanen


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