There are multiple options for dealing with forest insect pests and pathogens, but feasibility and success vary widely. To assess the unique challenges and opportunities that a biotech tree may present as a tool for addressing forest health, it is important to understand the other options available. Given the spatial and temporal nature of forest health threats from insect pests and pathogens, it is also likely that a combination of approaches might be needed to ensure proper management of an infestation. The most cost-effective approach for protecting forest health from nonnative insect pests and pathogens (Finnoff et al., 2007) is to prevent introduction, followed by early eradication after arrival (Liebhold et al., 2016). Once established, the impact and cost of dealing with the infestation rapidly increase (Roy et al., 2014). Given that human mobility and trade volumes—major drivers of pest introductions—are likely to continue to rise, the enforcement and enhancement of preventive measures will become even more critical (Lovett et al., 2016). Even where prevention or eradication has been successful, forests will remain vulnerable to repeated introductions of the same nonnative insect pests and pathogens over time.
Once established and spreading in forests, whether pests are native or nonnative, multiple management options may exist. Management can focus on trying to minimize the damage and mortality to the forests (including the large, old-growth trees), on actively preparing to regenerate or restore a species, or on both strategies. If the impacts are not severe enough to alter the species’ ecological footprint or management actions appear unrealistic or undesirable, managers may decide that taking no action is the best alternative. However, if the decision is to take action, the focus turns to early detection and response, containment, and long-term management to restrict further expansion and impact (Liebhold et al., 2017). Management options include biological control and integrated pest management, and various forms of site management (e.g., pesticide use, containment, fire, thinning) (Liebhold et al., 2017). When outbreaks of insect pests and diseases affect only one or a few tree species, the larger impact of such pests is directly related to the dominance of the host species. Thus, maintaining high levels of diversity may be an effective management approach to minimize impact. In low-diversity forests, other approaches may be more important.
In many of the most extreme cases, because of the high susceptibility of native tree species to some nonnative insects and pathogens with substantial dispersal potential, it will not be possible to
prevent extremely high mortality in the affected tree species. Once an insect or pathogen is established, there are a number of key management tools that might be considered to retain the presence of the tree species in North American forests into the future. These include (1) the enhancement of genetic resistance, (2) the development and use of biocontrol agents, (3) the development and use of chemical control methods, and (4) management practices to prevent or decrease the infestation. The enhancement and use of genetic resistance can proceed through the development and deployment of selective-resistance breeding within either the native species or from closely related nonnative species or the development and deployment of resistance using biotechnology.
The effectiveness of these varied approaches to prevent and manage insect pests and pathogens varies across systems and infestations (Lovett et al., 2016). The time line for use of these tools in management activities for forest trees and forest health will depend on a number of factors, but the biology of the species involved (both tree and insect or pathogen) and the environments in which the tree species exist will have a major influence. Insecticides and fungicides are often used in attempts to preserve existing forest stands or to protect large individual trees in urban settings. They are usually expensive, may have negative environmental impacts, and in some cases provide only a stopgap measure to give time to consider or develop other alternatives or the hope that future environmental conditions become less conducive for the damaging insect or pathogen. The same can often be said of many biological control agents. This chapter provides an overview of the different approaches and the approximate time required for implementation. The case study species are featured to illustrate the differences between species and considerations of the merits of different approaches.
Preventing the introduction of insect pests and pathogens yields the largest ecological and economic benefits (e.g., Mack et al., 2000; GAO, 2015; see Figure 3-1). International trade agreements include clauses aimed at reducing these introductions (Burgiel et al., 2006). These are being implemented by the Animal and Plant Health Inspection Service (APHIS) of the U.S. Department of Agriculture (USDA) and include quarantine, interception (e.g., inspection, decontamination), and pre-border treatments (e.g., fumigation, immersion, spraying, irradiation, extreme temperatures) at the point of origin and during shipment (Haack et al., 2014; Leung et al., 2014).
The International Standards for Phytosanitary Measures Protocol 15 (ISPM-15) were developed under the International Plant Protection Convention to reduce the movement of wood-boring insects in pallets and other wooden shipping materials (Haack et al., 2014). Wood borers are among the most serious of insect pest invaders: 58 species of wood borers became established in the United States between 1909 and 2008 (Leung et al., 2014). The approved methods for wood treatment in the United States include heat treatment (conventional and dielectric) and fumigation with methyl bromide. An economic analysis by Leung et al. (2014) concluded that implementation of ISPM-15, though expensive and not fully effective, would save the United States more than $11 billion by 2050 in avoided impacts. These pre-border efforts are often coupled with the post-border protection efforts of inspection, quarantine, and treatment of imported materials that facilitate interception of insect pests and pathogens prior to their potential escape.
Early detection and response programs are essential to prevent the spread of introduced insect pests and pathogens, although these practices may not be effective against microscopic species (i.e., most pathogens) (Liebhold et al., 2016). In addition, public awareness through educational programs may be instrumental in minimizing the entry of harmful organisms and their early detection.
Surveillance methods to facilitate early detection can include deployment of pheromone and other traps, monitoring of sentinel trees or vulnerable sites, and solicitation of reported sightings
(Kalaris et al., 2014). Spatial modeling of locations of highest risk of invasion (Venette et al., 2010) can guide deployment of early detection efforts. Liebhold et al. (2016) reviewed both the uses of and methods for surveillance, ranging from baseline early detection to infestation delimitation, to verify the success of an eradication effort.
Eradication (see Figure 3-1) is dependent on effective early detection efforts because eradication is more successful when introduced populations cover small areas (Liebhold et al., 2016). Success is also dependent on the detectability of the insect pest involved and whether species-specific control tools are available (Tobin et al., 2014). Chemical traps, mating disruption (e.g., releasing sterile insects), and insecticide fumigations can be used to eradicate small populations of insect pests. For example, pheromone traps have been deployed at the advancing front of the introduced gypsy moth (Lymantria dispar) for early detection of spreading populations that can then be treated (Sharov et al., 2002). For pathogens, mechanical removal of the infected host may be the only viable practice, given that detection of the pathogen may not be feasible before infestation. Overall, eradication can either remove or contain the threat or delay the spread of the insect pest or pathogen while more effective management methods are developed (Liebhold et al., 2017).
Although the ability to eradicate pests has improved over time (Liebhold et al., 2016), many attempts have been unsuccessful. For example, eradication of white pine blister rust (Cronartium ribicola) to protect species of five-needle pine was a multimillion-dollar effort extending over more than 50 years in the 20th century. The principal approach was through removal of Ribes species (e.g., currants and gooseberries), the alternative host for the pathogen to complete its life cycle.
However, this effort is regarded as a failure in the western United States (Maloy, 1997). As of 2018, it is generally acknowledged that white pine blister rust will have a permanent presence in North America. Emerald ash borer (EAB), first detected in Michigan in 2002, was found in Maryland and Ohio in 2005, indicating that eradication efforts were not effective. Attempts at eradicating EAB were unsuccessful in part because of unintentional long-distance dispersal in nursery stock, movement via infested firewood and vehicles, the long-distance dispersal ability of the insect, the difficulty in detecting the early stages of infestation, the absence of a long-range sex or aggregation pheromone, and the lack of a suitable attractant for mating disruption (Mercader et al., 2011, 2016; McCullough and Mercader, 2012). In other instances, eradication efforts have been constrained by negative public reactions to the methods used, such as removal of potential host trees, release of irradiated insects, or broad spraying of a pesticide (Liebhold et al., 2016). Further review of this literature can be found in Liebhold et al. (2016).
When eradication of a nonnative is not possible or the spread of a native or established nonnative pest is inevitable, a variety of management options may be pursued. One option is to take no action. Although the option to take no action often is the de facto outcome because the discovery of a new introduction or recognition of increasing impacts of a species already present lags the infestation of hosts (Liebhold et al., 2017), the committee defines “no action” as a deliberate management decision that is weighed against other options. Options to minimize the effects of insect pests and pathogens include (1) biological control, (2) site management practices (including applying the types of chemical, host removal, and sterile insect techniques also used for eradication), and (3) enhancement of genetic resistance through selective breeding, hybridization, or biotechnology. These options may be implemented independently or in combination.
The decision to take no action may result from a determination that the insect pest or pathogen is unlikely to have significant (further) impacts on individual species or forest health, a lack of resources, or an inability to identify an effective action to take. This last reason would likely inspire further research if significant impacts are anticipated. The decision to take no action recognizes that vulnerable tree species may decline or be lost entirely, with potential cascading impacts on other species and ecosystem services. The ecological effects will depend on the role of the tree species in the environment, whether replacement species fill similar niches, and whether replacement species are themselves later subject to pest outbreaks. The no-action decision may be made at the time that the host tree is threatened or when restoration (e.g., via breeding or breeding in combination with a biotechnology approach) is considered.
Biological Control and Integrated Pest Management
Biological control is the intentional introduction or application of populations of natural enemies or competitors to control insect pest species (Kenis et al., 2017). Two types of biological control have been effective against introduced insects. The insect pest itself can be manipulated to reduce population growth (e.g., release of sterile males to suppress population growth by competing with fertile males). Alternatively, if the lack of natural enemies (i.e., enemy release) in the new range is the major driver of the outbreak, specialist natural enemies can sometimes be identified in the indigenous range of the introduced insect pest species and released into the area of invasion (Liebhold et al., 2017).
Biological control can be non-self-sustaining, such as when large numbers of sterile males of the insect pests are released, inundating the population sufficiently to dominate breeding, thus reducing the growth of a pest population. In this case, the released organisms are not self-sustaining in the environment, so this approach requires release whenever population control is necessary.
Self-sustaining biological control methods include introduction or augmentation of natural enemies that reproduce and are maintained in that location without successive applications. For North American trees, these include control of species such as the winter moth (Operophtera brumata) and the larch case bearer (Coleophora laricella) with parasitoid insects introduced from overseas or from another region of the North American continent (Wainhouse, 2005; Kenis et al., 2017). Management practices that favor native predators of the insect pests (conservation biological control), such as providing shelter and alternative food sources for those species, are also common strategies to minimize the likelihood of damage from insect pests (Tscharntke et al., 2007). Overall, a review of biological control in the United States concluded that establishment of biological control agents targeting insect pests of trees has been more successful than those targeting pests of herbaceous species (Kenis et al., 2017); still, the success of these programs can be quite variable (see Box 3-1). Both Liebhold et al. (2017) and Kenis et al. (2017) provide reviews of biological control efforts to reduce the impacts of insect pests on trees.
Hypovirulence is a biological control strategy for mitigating or suppressing the effects of some fungal pathogens. Some viruses can infect pathogenic fungi, reducing their ability to infect, colonize, kill, and reproduce on susceptible hosts (Boland, 2004). If these viruses are spread in the area infested with a problematic pathogen, in some instances they may reduce the virulence of the pathogen of interest. Success with this strategy has been demonstrated in Europe on strains of Dutch elm disease (Boland, 2004) and chestnut blight (Grente and Sauret, 1969; Grente and Berthelay-Sauret, 1978; see Box 3-1).
Site Management Practices
Whether to contain a pest from spreading or as a strategy for long-term management, there are a number of site management practices that create conditions unconducive to a pest outbreak. Use of chemicals, such as pesticides and fungicides, are common practices in managed forests. However, reliance on chemical controls is generally not a long-term solution because of the long-recognized potential consequences of widespread pesticide use. These consequences may include evolution of resistance in the pest, nontarget impacts, substantial expense associated with repeated treatments, and public opposition to widespread use of potential toxins (Mack et al., 2000; Gould et al., 2018).
As in eradication efforts, quarantines may be put in place to prevent the movement of contaminated wood, and infested or infected trees may be removed. For example, when the Asian longhorned beetle (Anoplophora glabripennis) was discovered infesting multiple tree species in urban parks and suburban neighborhoods (Haack et al., 2010) in New York in 1996 and in Chicago in 1998, survey, chipping and burning of infested trees, and quarantined movement of potential host species (including nursery stock) were all rapidly implemented. Uninfested host trees within specific distances from infested trees were treated with insecticide. However, despite successful eradication in several sites, repeated introductions of the beetle means that it remains a threat to U.S. forests (Haack et al., 2010). With regard to EAB, tree removal, quarantine, and insecticides have not been effective (see Box 3-1).
Another site management practice to minimize the conditions that favor the onset of an outbreak is thinning. Silvicultural thinning of managed stands is often conducted to improve growth; this practice also promotes individual vigor, increasing tree defenses against insect pests and pathogens (Gottschalk, 1993; Maher et al., 2018). For example, mountain pine beetle (Dendroctonus pon
derosae) is one of the native insect pest species predicted to expand its distributional range under climate change (Raffa et al., 2013). Outbreaks of this species are driven by drought and even-aged stands of mature trees. In this situation, thinning stands and removing infested trees may reduce the beetle population below the outbreak threshold. Thinning of the understory also removes fuel for wildfire, reducing the probability of tree mortality.
Pruning, rather than thinning, can also be an option in silvicultural stands. For example, branch pruning of white pines can reduce the impact of white pine blister rust (Ostry et al., 2010; Schwandt et al., 2010). This approach is feasible only where white pine species are in silvicultural management. Pruning focuses on protecting existing trees and their genetic diversity but does not increase the genetic resistance of future progeny.
Maintaining diverse forests and planting mixed stands where the site naturally supported multiple species is another management tool to promote ecosystem resistance to insect pests and pathogens. More diverse forests are subject to lower levels of herbivory by insects than are more homogeneous forests, and this effect increases with the taxonomic distances among trees and with the proportion of unaffected species (Jactel and Brockerhoff, 2007). This phenomenon is due to the
In the case of nonnative pathogens or insects, a greater understanding of controls on pest population dynamics in the home range of the pathogen or insect would provide a valuable perspective on what type of resistance might have a high likelihood of success. For example, in Asia, what resistance is present in the native white pine species, where presumably the pines and the white pine blister rust fungus have coevolved? It would be helpful to know whether similar resistance exists (even at low frequency) in North American white pines. In addition, in some areas of Asia, white pine blister rust has become problematic (La, 2009; Zhang et al., 2010). Knowing the cause of that change—perhaps due to new land management practices, evolution of greater virulence in the rust, or changing climate—would also be useful information. Further study from a global perspective to understand coevolved systems and how they can be disrupted will help design strategies to restore species (including how to use any biotech option) and help refine models to increase understanding of the potential long-term efficacy of resistance and its impacts over the landscape.
Another way to introduce genetic resistance into a susceptible tree species is to hybridize the susceptible species with a related resistant tree species. After hybridization, the offspring are backcrossed with different trees of the susceptible tree species to maintain genetic diversity. In theory, repeated backcrossing will result in resistant trees with genomes that are almost entirely consistent with those of the susceptible parent species, with the exception of the regions containing alleles that confer resistance to the insect pest or pathogen (Woodcock et al., 2017). In practice, this result is rarely achieved without intensive monitoring with DNA markers and large backcross population sizes. Backcross breeding is most effective for introgression of resistance when resistance is due to one or two dominant factors. Even in the simple case of only one or two factors, if one or both of these factors are recessive, the breeding strategy must include selfing or intercrossing alternating with backcrossing. Backcross breeding strategies may be greatly accelerated using marker-aided selection or genomic selection, particularly when the goal is to capture multiple resistance factors from the non-
native species. A good example of a backcross breeding program is the one undertaken to introduce blight resistance into the American chestnut from the Chinese chestnut (see Box 3-2).
Using Biotechnology to Enhance Resistance
Another way to generate resistant trees is through the use of biotechnology. Biotechnological research to introduce or modify traits in trees has been explored in a wide range of economically and ecologically important tree species throughout the world. Appendix C contains reports on biotech tree species at all stages of research and development from 1987 through 2018. Often the reports cover establishment of initial proof-of-concept transformation and regeneration systems, which demonstrate bacterial resistance genes from donor species. In species where this system is robust, the appendix includes reports demonstrating the incorporation of genes conferring various traits in the target species. These traits include insect and fungal resistance, early flowering, phytoremediation, tolerance to metal toxicity, herbicide tolerance, improvement of wood quality, changes to lignin content, and tolerance to drought, frost, and salt. The primary approach used has been transgenesis via Agrobacterium tumefaciens; however, the table includes reports describing biotechnological approaches such as RNA interference (RNAi) and clustered regularly interspaced short palindromic repeat (CRISPR) (see Box 3-3). For a more comprehensive treatment of the developments in the field of tree biotechnology, see Chang et al. (2018).
The first application for a field test of a biotech tree in the United States was submitted to USDA in 1989 for a poplar modified for glyphosate tolerance (Fillatti et al., 1987). As of 2018, about 700 permits have been issued by USDA-APHIS. However, only two tree species modified using biotechnology had been grown outside of field trials in the United States in that time.1 The first tree species to reach this stage was papaya (Carica papaya). Varieties with resistance to papaya ring spot virus incorporated via transgenesis have been grown in Hawaiian orchards since 1998. The second was apple (Malus × domestica) in which RNAi has been used to suppress the expression of polyphenol oxidase genes, resulting in fruit flesh that does not brown when peeled or cut. Packages of sliced nonbrowning apples became available to U.S. consumers in 2017.
To use biotechnology to confer resistance to a forest health threat, the first step would be to identify the gene that would be targeted for modification, introduction, or silencing. If a gene target is not already in hand, then a gene discovery process would be required. This step has traditionally been hindered in trees due to the characteristics that make them difficult as experimental organisms: large size, long generation time, potentially weak correlations between seedling and adult phenotypes, and (in the case of conifers) immense genomes. Purification of high-quality genomic DNA from forest trees requires modest modifications of standard procedures used for extracting DNA from plants due to high phenolic content, large fragile cells, or highly lignified tissue, such as stems or inner bark. The sheer size of many tree genomes presents a less tractable problem. Conifers have some of the largest genomes ever sequenced. The pine genomes can exceed 20,000 megabase pair (Mbp) per haploid genome (Zimin et al., 2014); sugar pine (Pinus lambertiana) has a genome size of 31,000 Mbp (Stevens et al., 2016). In comparison, Arabidopsis has only 135 Mbp and rice has 420 Mbp per haploid genome. Huge genome sizes and high content of repetitive DNA present difficulties in DNA sequencing and in genome assembly after sequencing.
Another problem results from the sequence diversity of forest tree genomes. Forest trees have high levels of heterozygosity due to their large population sizes and outcrossing breeding systems (Williams et al., 1999; Remington and O’Malley, 2000). There are no inbred lines of forest trees and few, if any, haploid individuals. Consequently, sequencing a tree genome is made more difficult
1 A variety of plum has been modified via Agrobacterium-mediated transgenesis to have resistance to plum pox virus, and this variety has met U.S. government regulatory requirements. However, it is not grown commercially as of 2018.
because, even for an individual tree, there are two diverse haploid genomes that are being sequenced at the same time, creating a challenge for sequence assembly. In sequencing and assembling the loblolly pine (Pinus taeda) genome, direct sequencing of DNA from haploid megagametophyte seed storage tissue avoided this difficulty (Zimin et al., 2014). Unfortunately, the seed storage tissues of angiosperm (broad-leaved) forest trees (such as ash) are diploid or even polyploid, and so this strategy is not available.
Genome sequences can provide substantial insights into organismal evolution, but their applicability in biotechnological approaches requires functional characterization of the components of the genome, including transcribed sequences (genes) and regulatory elements. One of the most significant methods for learning about the function of tree genomes is comparative sequence analysis, which reveals homologous sequences in different genomes that, in turn, implies similar functions. Proof of function of homologs requires further testing, such as evidence of transcription and translation, purification of a functional product, or genetic complementation. Here again, forest trees are at a severe disadvantage. Many advances in functional genomics have come from work on the genetic model plant Arabidopsis thaliana and herbaceous crops such as maize. Many tree species have structural and developmental differences that either required adaptation of methodology or precluded application altogether. Furthermore, many characteristics common to trees (e.g., dormancy, wood formation, and obligate outcrossing) have few or no analogs in annual crop species.
Recent technological developments have mitigated some of these shortcomings such that dramatic progress has been made in understanding the organization, structure, and function of tree genomes, thereby facilitating potential biotech modifications aimed at pressing problems in forest health. These developments include large-scale quantitative trait locus (QTL) or genome-wide association studies coupled with in-depth analyses of transcriptional and metabolic responses to insect or pathogen attack. Additionally, genome editing can produce a “meiotic drive” (a kind of gene drive) function that converts a heterozygous individual to a homozygous one at one or more loci of interest, eliminating a generation of selfing to produce homozygous trees in one generation (see Box 3-4).
The second phase of deployment in using biotechnology to modify phenotypes is production of trees containing the desired gene sequence. Transgenesis and genome editing require a transformation and tissue culture protocol in which the desired modification can be introduced into a single cell (usually in callus culture), and whole plants are generated from the transformed cell by regeneration of roots and shoots from disorganized callus tissue (organogenesis) (Birch, 1997). Many species of trees remain recalcitrant to the process of cell culture and regeneration. Even when regeneration is possible, the regeneration of a plant from a single cell may not produce an individual that has the desired genetic change in every cell. In well-studied plant species that are amenable to this process, embryos can be produced through somatic embryogenesis, a process where the manipulated cell or cells originate from a totipotent embryo and then are induced to make more embryos (Hakman and Von Arnold, 1985; Suprasanna et al., 2005). Regeneration can be stepwise and sequential, where shoots are induced first and rooting is induced subsequently through organogenesis. This process is complex and must be customized not only at the species level but also for individual cultivars within a species (Busov et al., 2005). The third phase is field testing. The case study species demonstrate varying degrees of progress with regard to the application of biotechnology to mitigating forest health threats (see Box 3-5).
The speed and cost of approaches to mitigate threats to forest health can vary widely. They are variable not only by approach (e.g., biological control, breeding, or biotechnology) but also by the state of knowledge about the target tree and the target pest (and its potential predator).
Speed and Cost of Biological Control
The speed and cost of biological control efforts vary depending on the biology of the target pest and the availability of biocontrol agents. A best-case scenario might be represented by the ash whitefly (Siphoninus phillyreae Haliday), an exotic insect from Eurasia and Africa that caused extensive defoliation of urban ornamental trees in California between 1988 and 1991 (Pickett et al., 1996). A wasp parasitoid, Encarsia inaron Walker, was imported from Italy and Israel, reared and tested under confined conditions, and then released into 43 counties in California by 1992. This resulted in nearly total control of the ash whitefly within the first year, with populations in Riverside, California, being reduced by a factor of 10,000 (Bellows et al., 1992). The total investment for this program was estimated at $1.2 million, and the cost savings were estimated to be between $220 million and $300 million, based primarily on the retail cost of removing and replacing urban trees (Pickett et al., 1996). This case was simpler than average for several reasons. First, the insect was restricted to California, and so federal regulations about interstate movement were not a factor, and only one state regulatory agency and USDA were involved. Second, the parasitoid was readily available and highly specific, so the risk assessment was simplified. A more typical biological control effort could be expected to cost between $2 million and $5 million and take 5–10 years to complete (Dr. Mark Hoddle, University of California, Riverside, personal communication, August 20, 2018). Other biocontrol efforts may not be successful, as has been the case for EAB (see Box 3-1). Additionally, the introduction of nonnative predators to control nonnative pests can often take years, first to identify the appropriate predators and second to obtain regulatory approval for their use (Rose, 2018).
Speed of Selective Resistance Breeding
The time line for selective resistance breeding in forest trees is dependent on several factors. In the best-case scenario for selective breeding, the infrastructure for a breeding program already exists, tree breeding expertise exists, and the biology of the tree species and the insect or pathogen is known. As discussed above (see section “Breeding to Enhance Resistance”), the first steps in selective breeding are to determine whether there is genetic resistance within the affected species, the frequency of resistance over its range, and type and degree of resistance available (i.e., is it immediately usable or will breeding be required?).
Determining the frequency and distribution of resistance, where it exists, may take several years or decades. Seed collection and testing of seedlings from hundreds or thousands of parent trees may be necessary. Those steps must then be followed with the development of breeding or orchard populations that have useful types and degrees of genetic resistance and sufficient genetic diversity to use for restoration (e.g., Sniezko et al., 2012; Dudley et al., 2017).
Two of the fastest evolving selective-breeding programs in forest trees have been development of resistance to the soilborne pathogen Phytophthora lateralis in Port-Orford-cedar (Chamaecyparis lawsoniana) on the West Coast (Sniezko et al., 2012) and development of resistance to the fungal disease Fusarium oxysporum in the Hawaiian koa tree (Acacia koa; Dudley et al., 2017). However, these two operational resistance programs only progressed rapidly once the basic data about resistance had been collected. In Port-Orford-cedar, the initial assessment erroneously concluded
that there was no resistance (Hansen et al., 1989). In the koa, there was initial uncertainty as to the causative agent of mortality. The operational program for resistance in Port-Orford-cedar started in 1996 (after a significant research period), and the first orchard seed was released by 2003. The operational koa wilt resistance program started in 2003; the first orchard seed was not available until more than a decade later. Even though programs for these two species were producing resistant seed as of 2018, the work was not complete. Seed was available for only some breeding zones, and the number of resistant parent trees in some orchards is too low to ensure that genetic diversity is preserved.
By contrast, the program to develop white pine blister rust resistance in sugar pine has taken longer to develop because of lower degrees of resistance and the longer time to reproductive maturity in sugar pine. The sugar pine resistance breeding program has continued for 50 years, with slow but steady progress (Sniezko et al., 2000; Kegley and Sniezko, 2004; McDonald et al., 2004; Kinloch et al., 2008, 2012).
The case of whitebark pine with resistance to white pine blister rust demonstrates the variation in the time it takes from resistant parent identification to deployment of resistant seedlings based on geography and parent genetics (Sniezko et al., 2007, 2011). Six restoration plantings were established in Crater Lake National Park from 2009 to 2016 (see Figure 3-7), using seedlings from some of the most resistant parents from the park. Seedling testing of progeny of Crater Lake parent trees was started in 2004, making the time from first testing to the first restoration planting only 5 years. However, seed orchards are also planned for Oregon and Washington seed zones, and in this case it may take from 10 to 20 years before resistant seed from these orchards is available. Thus, the overall time to begin restoration efforts using selective resistance breeding with whitebark pine can vary from as little as 5 years to several decades or more. The highly sporadic nature of good cone crops can also slow progress in resistance testing of candidate trees. Even in the case of Crater Lake, additional parent trees are being evaluated to increase the genetic base of seedlots used for restoration. With enough funding, a good cone crop, and a good seed collection effort, hundreds or even thousands of whitebark pine parent trees could be evaluated in a short time.
The backcross breeding program for the American chestnut, which occurred over six generations, took 35 years, a relatively short amount of time for several generations of tree breeding (see Figure 3-2). However, the program did not include the establishment of resistant seed orchards, the incorporation of genetic diversity in resistant seeds, or the development of resistant populations of chestnut for different geographic areas.
Relative Speed and Cost of Biotechnological Approaches in Trees
One of the commonly cited advantages of biotechnological approaches to create resistant genotypes of trees is the speed with which they can be deployed. Selective breeding and backcrossing are slow processes in some tree species (such as sugar pine) because the low initial degree of resistance and the long juvenile periods of most trees translate to long generation times and therefore very slow breeding cycles (Harfouche et al., 2012; Isik et al., 2015). Furthermore, there is often a poor correlation between traits measured in juvenile trees compared to those in mature trees, which necessitates expensive field testing over multiple years for each breeding cohort (White et al., 2007). Consequently, most forest breeding programs have only progressed through a few of generations, leading to modest genetic gains compared to annual commodity crops such as maize and wheat (Isik et al., 2015). Biotechnology has been promoted as a means to accelerate the domestication of forest trees by shortening the breeding cycle (e.g., through early flowering; Martín-Trillo and Martínez-Zapater, 2002; Flachowsky et al., 2009), using marker-aided selection (Harfouche et al., 2012; Isik et al., 2015), or bypassing breeding entirely by manipulating DNA (Merkle and Dean, 2000; Harfouche et al., 2011).
The actual speed of biotechnological approaches depends on a number of practical factors that can potentially limit implementation. In the area of marker-aided selection, it is becoming increasingly clear that the efficacy of this approach is limited by the complex genetic architecture of quantitative traits. Genome-wide association studies have clearly demonstrated that complex traits are polygenic, that is, controlled by hundreds or even thousands of loci, each of which has small genetic effects, and complex epistatic interactions (Boyle et al., 2017). This complexity means that alleles that control traits in one population are often not effective at predicting phenotypes in an unrelated population, thus requiring expensive and time-consuming marker discovery and model training in each subpopulation (Resende et al., 2012). Although this problem should be diminished for some disease resistance traits with simpler genomic architecture (e.g., for cases of major gene resistance), these types of resistance are expected to be less durable than quantitative resistance based on multiple unlinked loci (McDonald and Linde, 2002), so applications are limited.
The situation could be different when using biotechnology to make individual genetic modifications to produce dramatic phenotypic changes. For example, introduction of the crystalline endotoxins derived from Bacillus thuringiensis (Bt) can confer complete resistance to feeding by Lepidoptera (moths and butterflies), even in host species that are normally highly susceptible to such damage (Shelton et al., 2002). A wide variety of Bt toxins are already available (de Maagd et al., 2003), and existing toxins can be modified using mutagenesis to enhance efficacy against a particular insect once introduced into the plant via transgenesis (de Maagd et al., 1999). Similar examples exist for pathogen resistance as well, such as the introduction of the oxalate oxidase gene (see Box 3-5), which shows broad efficacy against fungal pathogens such as the Cryphonectria blight in American chestnut (Zhang et al., 2013) and Septoria leaf spots in Populus (Liang et al., 2001). Nevertheless, the relative speed with which biotechnology solutions can be implemented depends on a number of factors, and significant impediments at each stage of development could substantially slow the process.
As reviewed above (see section “Using Biotechnology to Enhance Resistance”), the first step to make use of biotechnology to introduce genetic resistance is gene identification. Although the gene discovery process has become remarkably more efficient with advances in genome sequencing technology, this step could take a number of years. For example, in the case of EAB, there was no genome or transcriptome sequence available for the insect or host in the early stages of the epidemic, and it is time-consuming and difficult to measure ash tree resistance, which requires infestation of trees that are 2–3 years old (Koch et al., 2015).
The second step is producing trees with the desired gene sequence. Under the best-case scenario, represented by the model transformation clones in Populus, it takes 4–8 months to produce transformed plants that could be transplanted to pots (Busov et al., 2010). At least another several months would then be required to vegetatively propagate enough material for a field trial. In other species for which transformation systems are not readily available, the process of regenerating a plant from somatic embryos to a seedling growing under ordinary conditions in a greenhouse can take more than 1 year. If the introduced gene is present in the germ line, progeny of the transformed plant will also have the inserted gene. In many tree species it can take 5–10 years or more until flowers are produced and the gene can be passed on to the next generation.
The third step is field testing. The length of this phase would depend on the growth rates of the trees and the life history of the insect pest or pathogen. In some cases, resistance is best evaluated in adult trees, which requires many years. For example, both Septoria and Cryphonectria cankers take years to develop, though effective in vitro assays have been developed for both diseases (LeBoldus et al., 2010; Newhouse et al., 2014b). Furthermore, multiple field trials over a large geographic area are desirable, especially in cases where significant genotype × environment interactions occur for host susceptibility. Given the expense and difficulty of performing field trials, testing would typically begin on a limited basis to demonstrate efficacy before scaling up to larger and more widespread trials. This slow rollout would add years to the process. As a result, the full process would take more than a decade.
Estimating costs of the application of biotechnology for forest health is difficult because it requires estimation of processes and products that have not yet been developed or information that is proprietary and not available. The cost of gene identification or developing a new DNA transfer system is undefined because in some cases the efforts could be unsuccessful; therefore, the project could have high costs with no results. Producing trees with the desired gene sequence through clonal propagation can be expensive.
However, when it comes to a comparison of costs between breeding and biotechnology, the costs may be similar or at least similarly variable. With regard to the identification of a trait of interest, for the biotech tree the expense is in gene discovery and integrating the desired change into the tree’s DNA; for breeding, the costs are related to screening and testing to find resistance. For both approaches, the costs can vary widely depending on the biology of the tree and the pest, the state of knowledge about the tree and pest biology, and the robustness of the biotech or breeding program associated with the species of interest. When it comes to the next step of clonal propagation, the costs are likely to be similar between the two approaches.
The major difference in costs between the two would be those associated with any regulatory approval that a biotech tree may need to obtain; a selectively bred or hybrid tree does not have to go through a regulatory process in the United States.
Management to mitigate damage to forests from insects or pathogens takes significant time and resources. With regard to nonnative insects and pathogens, the first line of defense is preventing their introduction. When introduced pests have become established or native pests are expanding
their range or increasing in virulence, there are a number of management options that may be employed, including taking no action. Chemical or biological control can control pests in some cases, but these approaches are often not acceptable to the public, effective, or timely. The development and actual deployment of genetic resistance, whether via breeding or biotechnology, will usually take decades from the initial research phase to even the beginning of the restoration plantings. However, given the repeated introduction of nonnative pests and the likelihood of continued abiotic stress from climate change, incorporating genetic resistance may be the effective strategy for the long term.
Conclusion: Substantial literature supports the need for sustained investment in prevention and eradication as the most cost-effective and lowest impact approaches for managing introduction of nonnative insect pests and pathogens.
Economic analysis has found that the United States could save billions of dollars in avoided impacts from nonnative pests by increasing its efforts to prevent the entry of nonnative pests. Inspection, quarantine, and treatment of imported materials can facilitate the interception of insect pests and pathogens prior to their potential escape and establishment.
Recommendation: Investment in effective prevention and eradication approaches should be the first line of defense against nonnative species in efforts to maintain forest health.
Conclusion: Any single management practice alone is not likely to be effective at combatting major pest outbreaks.
Site management practices—such as pesticide use, thinning, reintroduction of fire, and removal of infested trees—can minimize conditions that favor a pest outbreak. Biological control agents can suppress insect pest populations or mitigate the effects of a fungal pathogen. However, experience with the American chestnut, whitebark pine, ash, and poplar indicates that these practices will be insufficient to curtail the loss of affected tree species.
Recommendation: Management for forest health should make use of multiple practices in combination to combat threats to forest health.
Conclusion: A variety of biotech and nonbiotech approaches have been and will be developed to address insect pest and pathogen threats. The time line for use of these tools in management activities for forest trees and forest health will depend on a number of factors, but the biology of the species involved (both tree and insect or pathogen) and the environments in which the tree species exist will have a major influence on effective mitigation.
The time line for using approaches to mitigate forest health such as biological control, breeding, or biotechnology vary by the state of knowledge about the target tree and the target pest. The availability of natural enemies, the size of the tree genome, and the environment will also affect the deployment of mitigation tools.
Conclusion: Many tree species have some degree of resistance to particular native and nonnative pests that may be harnessed to combat infestations and epidemics.
It is often possible to find resistance to damaging insects and diseases in the field and use it to develop resistant trees for restoration planting. However, this outcome depends on the resources to
find resistant trees and established breeding programs to develop resistant seedlings. This strategy has been successfully deployed for blister rust resistance in whitebark pine and, at the time of the committee’s report, were also in use for ash against EAB. For whitebark pine, there are still no programs for drought tolerance or resistance to the mountain pine beetle.
Recommendation: Entities concerned about forest health should devote resources to identifying resistant trees within a population that have survived a pest outbreak. Research to understand the role of resistance in coevolved systems from the perspective of a global host–pest system, where the nonnative pathogen or insect originate, would help guide efforts in North America.
Conclusion: Using biotechnology to introduce resistance to threats in forest trees has been hampered by the complexity of tree genomes, the genetic diversity in tree populations, and the lack of knowledge about genetic mechanisms that underlie important traits. However, recent technological developments have improved functional genomic tools, facilitating the potential for biotechnology to help address forest health problems.
At the time the committee was writing its report, there was insufficient knowledge about the fundamental mechanisms involved in resistance to pests to efficiently identify genomic means to mitigate pest damage. Most tree genomes had not been sequenced, and there were still many unknowns about the underlying nature of resistance, including its heritability and on whether it will be durable. Investigations in trees species are needed to uncover all forms of resistance, not just those due to easily discernible single major genes. Likewise, in using biotechnology, greater efforts are needed to understand what types of resistance or combinations of resistance are likely to be durable.
Recommendation: More research should be conducted on the fundamental mechanisms involved in trees’ resistance to pests and adaptation to diverse environments under a changing climate.
Conclusion: The time it takes to identify resistance in an affected population, breed resistant seedlings, and plant resistant seedlings in the field can vary from a few years to multiple decades, depending on the species. Incorporating resistance via biotechnology into a tree species is also a lengthy process, the duration of which varies by species.
The amount of natural genetic resistance in a population can vary by species or by the geographic distribution of a species, and the reproductive cycle of the tree will affect how quickly resistant offspring can be generated. When introducing resistance via biotechnology, the number of the genes involved in the expression of resistance will affect how long it takes to identify and incorporate resistance in biotech trees.
Recommendation: Sufficient investment of time and resources should be made to successfully identify or introduce resistance into tree species threatened by insects and pathogens.
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