**1. Introduction**

Wood is among the most durable cellulosic materials, but it can be degraded by a number of biotic and abiotic agents. These agents often act simultaneously making it difficult to completely separate causal agents. This review will concentrate on the role of fungi in degradation but will also discuss other agents as they relate to the overall process. Our discussion will focus on those fungi capable of degrading the primary cell wall polymers of cellulose, hemicellulose and lignin, but will also touch on the many mold and stain fungi that not only cause aesthetic concerns and disfigure coatings but can digest the stored compounds in the parenchyma cells and the pit membranes while promoting more limited damage to the structural elements of wood.

Wood poses a major challenge to organisms seeking to extract the energy from its polymeric structure. While the stored compounds in parenchyma cells are digestible by many organisms, accessing the more complex polymers is a key challenge. The chemistry and arrangement of the cellulose and lignin polymers in the wood cell wall sharply reduces the number of agents capable of causing damage. Many fungi are cellulolytic but are unable to unlock the chemistry of the lignin polymer that both enrobes and is interspersed with the cellulosic components of the lignocellulosic matrix. Only those fungi that have developed strategies to surmount the recalcitrance of lignin are able to fully extract the embodied energy of the lignocellulose cell wall.

#### **2. Wood as a Polymeric Material**

Cellulose represents 40–44% of most wood and endows wood with many of its unique material properties [1]. At the nanoscale, cellulose is arranged in discrete units known as elementary fibrils, with the inner core of those fibrils being a tightly packed crystalline material and the outer layers being more loosely packed amorphous cellulose. The crystalline nature of the inner core of the elementary cellulose fibrils gives wood its exceptional tensile strength—comparable to steel and aramid (Kevlar) at the nanoscale [2]. Crystallinity also renders the inner portion of the cellulose elementary fibril more resistant to degradation than the other more amorphous carbohydrate polymers also present in the wood cell wall.

Lignin represents 18 to 35% of the wood cell wall and is a heteropolymer consisting of repeating units of phenyl propane with a diverse array of bonds between three variations of the monomer form. This diversity in bonding pattern makes lignin extremely resistant to degradation and few microorganisms other than select wood degrading fungi have been able to unravel this system. Furthermore, the higher levels of lignin in wood compared to other plant materials, and manner in which lignin is intimately integrated with the holocellulose components, are major reasons for wood's resistance to degradation.

Hemicellulose is a branching heteropolymer of pentose and hexose sugar monomers representing 15–32% of the wood cell wall. Hemicellulose is a more amorphous polymer compared to cellulose, and it is considered to be the link between cellulose and lignin that allows the three polymers to behave as an integrated matrix. Hemicellulose is also more susceptible to degradation and the many forms of the hemicellulose polymer can be degraded by excessive heat such as that occurring in thermal modification processes used in industrial wood protection [3,4], or by relatively dilute acids, as was observed with significant mechanical property losses following treatment by some early-industrial acidic fire-retardant and preservative formulations [5]. These forms of degradation are discussed more completely in the section on Abiotic Degradation below. Important relative to this chapter are several types of fungi that attack hemicellulose in early stages of decay leading to mechanical property loss [6]. Although hemicellulose is an integral part of the wood structure, it is often viewed as the weak link in the armor of wood as a material, since its degradation profoundly affects the ability of lignocellulose to behave as a rigid structural system. Hemicellulose is often the first major component of wood to be attacked and deconstructed by fungi [7,8].

In addition to the cell wall polymers, other components including proteins, fatty acids, pectin, starch and other polysaccharides and sugars are also present at levels ranging from 2 to 15% (Average ~6%) of the wood mass [1]. While these materials can have dramatic effects on resistance to microbial attack, this review will concentrate on the roles of the three primary cell wall polymers in relation to fungal attack.

#### **3. The Importance of the Wood–Water Relationship**

Water is an important constituent of wood materials and biomass in the natural environment and can affect wood properties both directly and indirectly. The uptake of water is an integral factor associated with wood's ability to be attacked by fungi and to decay, but moisture also impacts wood by moving into the cellular structure to associate with the cellulose, causing dimensional changes. Even though water is held in wood primarily by hydrogen bonding at the molecular level, the collective force of the hydrogen bonds formed is sufficient to swell the wood to the extent that, at a practical level, is capable of splitting rocks, tearing thin metal plates or forcing structural walls apart. Shrinkage and swelling of wood with changes of humidity in the environment creates substantial challenges for coatings. Polymeric coatings typically are unable to prevent moisture vapor from diffusing in and out of wood as humidity changes. This results in repeated swelling and shrinkage of wood, particularly in exterior environments. Few polymeric coatings can withstand the recurring cycles of expansion and contraction that wood surfaces undergo with significant humidity changes.

The stress on wood coatings due to moisture changes, combined with other factors such as mechanical action and abrasion, creates an environment where coatings are subject to rupture at the microscopic level. Wood surfaces are subjected to a continual dusting of fungal propagules that, once moisture levels and environmental conditions are correct, can germinate and exploit the micro-cracks that can develop in coatings. This will be addressed in the section on future research needs for bio-based coatings.

#### **4. Abiotic Degradation**

Non-living agents can cause different types of wood degradation that are often confused with fungal degradation. For this reason, it is important to review some of the most important abiotic (non-biological) degradation agents which are often confused with fungal attack of wood.

Wood weathering is the most common type of non-biological degradation occurring in exterior-use wood [9]. The low energy levels in sunlight can be quite disruptive to the polymeric components of wood at the molecular level. During UV (sunlight) exposure, the primary energetic components which can attack lignocellulose are reactive oxygen species (ROS or oxygen radicals) that randomly attack the cell wall polymers, especially lignin. This energy transforms lignin into a variety of radical forms that transfer the energy to hemicellulose and cellulose which are then readily depolymerized [10,11]. This process begins almost immediately after timber is exposed to sunlight, but the effect on the timber is shallow until the outer layers of UV-degraded wood are sloughed off. Weathering related to UV exposure is typically slow but can have substantial effects on adhesion of coatings as well as declines in appearance leading to premature replacement. For example, a preweathering exterior exposure of Western redcedar siding in direct sunlight for 1, 2, 4, 8 or 16 weeks prior to being painted reduced paint adhesion by up to 50% [12,13]. Increased preweathered was associated with shorter paint service life: paint on panels preweathered for 16 weeks began to flake and peel after 4 to 5 years; preweathering for 8, 4, 2 or 1 weeks resulted in paint failures after 7 to 8 years, 9 to 10 years, 10 to 11 years or 13 years, respectively. Coatings on non-preweathered controls were still in almost perfect condition after 17 years of exposure.

Another common abiotic source of wood degradation that can be confused with fungal attack is heat- or thermal degradation. Wood has a well-known propensity to burn, but it also experiences more subtle changes upon extended exposure to temperatures between 65 ◦C and the ignition point [5,14]. Heating affects each polymer differently, with hemicelluloses being the most susceptible, followed by cellulose and finally, lignin [15]. Thermal-related modification can have negative effects on wood, particular if the wood has been modified with other treatments such as some types of fire-retardant treatment. In the 1980s, most of the fire retardants used in North America were acidic. During that period, issues gradually emerged where fire-retardant treated wood in structures began experiencing extensive degradation [5,16]. Subsequent investigations showed that the acidic formulations tended to react with, and degrade, wood maintained at temperatures typically found in attics and enclosed roof spaces during the summer. Further study revealed that hemicelluloses were the first polymers affected, again illustrating the critical role played by these polymers in lignocellulose performance [14]. Disruption of the biological degradation process by thermally modifying hemicellulose has been studied since the 1930s [17]. This research was initially directed at modifying colors to render lighter, less valuable woods darker, but it was also shown to affect wood/moisture relationships. The process was never fully exploited, but was revisited in the 1980s as European researchers sought non-biocidal methods for improving timber durability [4]. Extensive subsequent research has shown that thermal modification alters the hemicelluloses, thereby reducing moisture uptake which in turn reduces, but does not completely eliminate, the risk of decay.

#### **5. Biotic Deterioration—Wood Decay and Requirements for Fungal Attack**

Wood decay is largely caused by fungi that fall into categories depending on the appearance of the degraded wood which is, in turn, related to polymeric materials that are degraded. Brown rot decay is an informal name for the most common type of decay occurring in timber products. Fungi that cause brown rot depolymerize cellulose and hemicellulose (holocellulose) for digestion, while lignin is also depolymerized and modified before being rapidly repolymerized. The general categories of white rot fungi and soft rot fungi are the other major types of decay, and these are covered later in this review, as these fungal decays can be quite important in certain environments. Fungi are Eukaryotic organisms that are in the same Domain in the Tree of Life as plants and animals [18]. Species within other Domains in the tree of life comprising the Bacteria and Archaea can also live in wood. Some species of Bacteria have been shown to cause limited wood deterioration over long periods of time (several centuries), resulting in mechanical property loss in wood. Because of their minor importance in deterioration of structures, these microorganisms will not be considered further in this chapter, although we recognize that they are almost always present and have been suggested to play supporting roles in the degradation process including pre-conditioning of wood, and extractives detoxification. Bacteria are also active in long-term degradation of submerged wooden foundation piling, which typically occurs over many centuries [19]. Insects and some types of marine boring animals also can cause significant biodeterioration of wood under some circumstances, but deterioration by these animals is reviewed elsewhere [20].

For all fungi, spores (microscopic seeds) or other small fragments of the fungi must be produced and be transported either in the air, water or on other organisms (such as insects) to other pieces of wood where a new fungal colonization can initiate. Several requirements must be met for colonization to occur. In addition to the wood substrate itself, these include:

#### *5.1. Water and Air*

Typically, wood must be at or near the point where the wood cell wall is saturated with "bound" water, known as the fiber saturation point (FSP) for the fungal spores or fragments to germinate and initiate new fungal colonies. There is considerable debate about the minimum moisture level required for fungi to colonize and decay wood exposed out of soil contact [21]; however, because the filamentous strands of fungi (hyphae-see Fungal Physiology/Anatomy below) are required to be surrounded by a watery extracellular polysaccharide matrix (ECM) when wood is being attacked, some level of free moisture in the wood cell lumen is required to support fungal growth. In most cases, fungal decay can begin at approximately 30% moisture content (oven dry basis), reaching an optimum between 40 and 80%, then declining with increasing moisture levels above 100% as cell lumens begin filling with water and oxygen becomes limiting. Water is often the most important limiting factor in decay, and some paradigm-shifting current literature [22] focuses on the critical moisture content that allows fungal metabolites responsible for depolymerization of the wood polymers to diffuse within the wood cell wall. However, fungi require liquid water to initiate the secretion of metabolites required for decay, even in the absence of wood or other suitable substrates. Because the biosynthesis of these metabolites is a prerequisite for their diffusion within the cell wall, it is important for wood to first attain a moisture content where fungi can synthesize metabolites as well as one that supports robust ECM production surrounding and attaching the fungal cells to the wood cell wall to allow compounds to diffuse between fungus and the wood cell wall. In this regard, appropriate moisture conditions for fungal activity are required even before diffusional aspects of metabolites within the wood can be considered, and inhibition of fungal ECM production represents a fruitful area for future research in controlling fungal growth.

Wood in contact with the ground is frequently above the FSP and, in addition to having a fully saturated cell wall, contains liquid water in the lumens of the fibers. For fungal growth to occur in wood, the moisture content of wood must also not be too high to preclude adequate oxygen levels. Although fungi do not require as much oxygen as humans, they are aerobic organisms and wood decay fungi typically will not grow on wood that is completely saturated or submerged. Oxygen is rarely limiting for fungal attack, although complete saturation of logs by "ponding" (submersion in natural or artificial bodies of water) has been used to limit decay for periods of several months prior

to processing in mills. In some cases, logs that sank during freshwater storage have been retrieved decades and sometimes even centuries later and processed with little deterioration noted. Many of these logs remain on the bottom of fresh water bodies and are in pristine condition because of the lack of oxygen, and also because relatively cold temperatures limited anaerobic bacterial deterioration.

#### *5.2. Temperature*

Temperature is critical for most physiologic reactions. Fungi may continue to grow as temperatures decline to levels near freezing [23], but reaction rates involved in both fungal metabolism and in the chemistry/biochemistry of wood depolymerization decline, and the decay process ultimately stops at freezing temperatures. Metabolic reactions increase with increasing temperature with most fungi having growth optima between 24 and 32 ◦C [20]. This is well within the temperature range inside most inhabited structures but specialized fungi can grow well outside this range. As a result, temperature is generally not a limiting factor in decay. For most decay fungi, fungal metabolism becomes more constrained as temperatures exceed 39–40 ◦C; however, some thermophilic fungi survive and have been observed to be active at temperatures exceeding 50 ◦C in specialized environments such as pulp chip piles [24,25]. Exposure to temperatures about 56 ◦C results in permanent denaturing of proteins and DNA, effectively killing most non-thermophilic organisms.

#### *5.3. Other Essential Components for Fungal Growth and Decay Potential*

As noted above, a primary requirement for decay is a nutrient source which is typically the timber itself. This wood or surrounding supporting materials/soils must also contain various micronutrients and microelements, including nitrogen, that are essential to fungal growth and the decay process. There are vast volumes written about the relationships between decay and wood properties, e.g., [26]. Relatively few fungi have evolved the ability to degrade and utilize the three primary cell wall polymers, and the mechanisms by which they accomplish this task will be the subject of the remainder of this paper.

#### **6. The Decay Environment**

Many microorganisms are ubiquitous and will colonize almost any substrate meeting the basic requirements for growth of oxygen, food, moisture & temperature. However, the nature of the wood substrate affects the types of wood degrading fungi that attack it, the colonization patterns, and the rates at which wood is utilized. For this discussion, we will ignore aquatic environments and concentrate on the terrestrial decay process. Decay can be categorized as either in ground or above ground, recognizing that these are arbitrary terms that overlap at their margins. In-ground or ground-contact decay is typically more rapid, reflecting the presence of adequate moisture along with an abundance of organisms with differing wood-attacking or wood-inhabiting capacities that are always present and in competition with each other. The above-ground environment is more challenging with fewer organisms present, and a greater potential for drying that can slow the decay process. There is also more limited potential for the transfer of moisture and exogenous nutrient resources into the wood and into the fungal thallus. These differences result in dramatic differences in the rates of degradation as well as substantial changes in the organisms involved.

### **7. Basics of Fungal Anatomy**/**Physiology and Evolution**

#### *7.1. Biology and Evolution*

Within the Eukarya, the Kingdom of Fungi are classified into Subkingdoms and further into Orders/Divisions. The subkingdom of Dikarya, which are fungi that largely produce filamentous cells known as hyphae, are of particular interest with regard to coatings [27], and are described in more detail below. Two Orders are encompassed by Dikarya, the Basidiomycota and the Ascomycota, and both can play significant roles in wood and/or coating degradation. Ongoing research suggests

that both Basidiomycota and Ascomycota evolved from earlier progenitor fungi between 400 and 500 million years ago [28–31], with species of white rot fungi that had the ability to decay lignocellulose evolving only perhaps 280 million years ago within a Class of fungi known as the Agaricomycetes [31]. Present day Agaricomycetes include many well-known mushroom-producing species, but many of the Agaricomycetes are rather nondescript in their macroscopic features, yet they have the ability to aggressively attack wood and disrupt any overlying coating materials.

It is interesting to note that plants initially evolved the ability to produce the cellulosic components of their cell walls long before they evolved the ability to produce lignin, which only later started to appear as a cell wall component in some types of woody plants [32]. Lignin "stiffened" the cellulose into a rigid polymer-matrix composite, allowing the precursors of modern-day trees to grow much taller than the grasses and sedges which existed prior to the Devonian period. The first lignin-producing plants evolved about 420 million years ago [33], long before fungi evolved enzymes and non-enzymatic systems with the capacity to depolymerize and further degrade lignin. Precursors to white- and brown rot fungi evolved lignin-degrading enzymes only about 295 million years ago [32], meaning that woody biomass was unable to be efficiently degraded by fungi for approximately 125 million years, and it has been proposed that inability to decay wood over this period of time led to the formation of coal seams in many areas of the world. However, some geology experts [33] examining fossilized woody tissue from the Carboniferous period dispute this premise and report that fungi were present in the fossilized wood cells showing "evidence of decay that is pervasive", suggesting that the fungi of that period were still somehow capable of decaying the woody tissues. However, the authors do not show convincing evidence of fungal attack of wood cell walls, and the images of fossilized fungal hyphae they present for interpretation are larger in diameter, and more consistent with non-decay Ascomycota species rather than Basidiomycota decay fungi. This suggests that there is still much more to be learned about the how wood decay fungi arose on earth, and their impacts, including the impact they have in decaying current day wood products and disrupting the coatings on those products.

#### *7.2. Growth and Infection of Wood*

Wood decay fungi primarily initiate as fungal spores or mycelial fragments. If conditions are favorable, spores germinate to produce fine hair-like structures known as fungal hyphae, which are elongated cells of the fungus which grow end-to-end. Hyphal fragments landing on wood can also in many cases initiate growth leading to broader colonization of the wood. Single-celled fungal growth is not common in wood degrading fungi except during spore formation. As the fungal hyphae grow along the surface of materials, some species can form a mat consisting of multiple layers of interwoven hyphae known as mycelium or a mycelial mat. The tips of the fungal hyphae initially seek out relatively simple pathways through the microstructure of wood, exploiting interconnecting cell wall pits (interconnecting channels between wood cells) to extend from one fiber to the next, and ramifying through the wood in this manner. During this initial growth phase, all wood-inhabiting fungi seek out stored products in the parenchyma as a ready nutrient source for energy for the fungus, and also to build up fungal biomass within or on the surface of the wood structure. This initial stage of growth of decay and some stain fungi results in activation of biochemical "machinery" within the hyphae resulting in secretion of a diverse suite of extracellular metabolites and enzymes that can depolymerize and digest select polymeric components of wood. One of the most common ways to measure wood degradation in laboratory studies is to monitor mass loss as decay progresses and wood cell wall components are converted to CO2 by the fungus. Although mass loss is not always an adequate indicator of structural strength loss, it is a straight-forward means for assessing some types of fungal degradation. Fungi have diverse suites of metabolites and enzymes that effect deterioration, and the discussion below provides information on how these fungi can be differentiated and how they may degrade wood in different ways.

Decay types: brown rot white rot, soft rot and their effects on wood chemistry: The fungi that degrade the cell wall polymers have long been segregated on the basis of the appearance of the damaged wood into brown, white and soft rot fungi. These separations are arbitrary and we now recognize that the decay types are more of a continuum [34], but the categories are helpful in generally classifying these fungal degradative agents, and for discussion.
