Darling 58

This happened at the very beginning of the program, likely in 2016. It is a “switched at birth” scenario, where pollen which was thought to have been gathered from a Darling 58 (D58) tree was actually gathered from a Darling 54 (D54) tree. Pollen from those two originating D54 first-generation trees were the basis for all research moving forward on what has since been assumed to be Darling 58.

In December 2023, TACF made the difficult decision to discontinue support for the development of the Darling line of transgenic trees (D54 & D58). This decision was based primarily on performance issues observed in scientific trials and the potential social ramifications of releasing an ineffective transgenic tree.

• Performance issues
  • Extreme variability in blight resistance, leading to the eventual shut-down of resistance over several generations
  • Growth penalties: upwards of 25% reduced growth in trees carrying the OxO gene (D54) compared to non-D54 trees
  • Higher mortality rates in trees carrying the OxO gene (D54) than those without, in greenhouse and field trials
  • Lethality in the homozygous state: offspring that inherit the OxO gene from both parents rarely survive
  • Genetic deficiencies: the insertion of the OxO gene in D54 caused a deletion of the SAL1 gene and may cause performance issues
• Social ramifications
  • Reduced trust in the use of biotechnology in forest health due to poor performance and lack of longer-term testing

Yes, TACF still supports transgenics as a restoration tool. The technology itself remains safe and represents one of several branches of research TACF is investigating for American chestnut restoration. While TACF believes the Darling line to be ineffective, the trees are not unsafe for the surrounding ecology. Our concerns are that we do not want the underlying genetic issues to be passed on to future generations of American chestnut restoration populations.

We also do not want a defective product released into the public domain. During this deregulation process, we have seen what appears to be an increase in overall public acceptance of these technologies. To bolster that trust, our organization, its scientists, and its advisors, therefore, must sometimes make difficult decisions which will ensure that the safest and most effective product will be what is deployed for restoration.

The first reason not to start over with the “real” Darling 58 is that Darling 58 also uses the 35S constitutive promoter which appears to result in variety of metabolic costs to the tree and causes some or most of the performance issues we are seeing. Secondly, there are only a handful, and perhaps even only one, D58 tree in existence. These trees are at either the T0 or T1 stage (the original event or first generation). If work is to start over at those early diversification stages, it makes the most sense to focus on lines that are likely to provide better competitive performance, for example, with wound-inducible promoters.

One of the greatest achievements of The American Chestnut Foundation (TACF) is the establishment of a broadly diverse, sexually-reproducing population of American chestnuts containing thousands of trees and representing hundreds of breeding lines.

Please continue to plant and care for these trees! Our ongoing efforts are dedicated to achieving enhanced disease-resistant American chestnuts. These wild trees and germplasm conservation orchards (GCOs) will continue to play a pivotal role in testing and research, as well as the ultimate reintroduction of a robust and diverse population back into the native range. Along with their potential outreach benefits, these plantings are vital for preserving genetic diversity, which is essential for the long-term survival and adaptation of the American chestnut. For more detailed information on our current scientific strategies, please refer to the Science Strategy page.

TACF remains open to transparent collaboration with partners who employ safe and effective scientific methods for American chestnut restoration, including ESF.

TACF is privileged to work with many partners throughout the native range of the American chestnut, all of whom inform the direction of our scientific inquiry. As TACF strives to fulfill our mission to restore the iconic American chestnut to its native range, it is imperative to include data and input from many credible sources, from the work of private landowners to university-level, ongoing projects.

No. Testing has demonstrated that the Darling prototype trees do not present plant pest risks different from native chestnuts. While Darling trees would not adversely affect the natural environment, TACF has concluded that these trees are unsuitable for restoration due to their performance issues (decreased growth and fitness, increased mortality, and variable resistance). Premature distribution of this or other inferior varieties may also unfairly skew public perception against the use of biotechnology solutions to save threatened forest tree species. TACF is confident that our path will eventually yield safe and effective restoration trees.

The regulatory review is focused exclusively on safety and not efficacy (i.e. how well the tree would perform in restoration populations). The studies performed on environmental safety, as well as data regarding American chestnut spread and silviculture, continue to suggest that the researched Darling line will not constitute a plant pest risk.

However, it is the producer who is in charge of ensuring that the product is effective for its intended purpose, not the regulatory agencies. This means that research into the ability of Darling 58 to survive blight and effectively compete and reproduce in a forest setting has been ongoing throughout the regulatory review. The accumulation of data concerning performance is a long-term process and can continue long after a product is deemed environmentally safe.

At TACF, we want to ensure that the product (trees destined for restoration efforts) are effective before they “hit the market”. We value greatly the trust granted to us by supporters which has allowed us to devote 40 years to the process, and which will take many more decades for successful restoration to be realized.

The process of science leads to learning. One of the major lessons learned from this process is the need for field test data and results, not just lab and greenhouse data, available before planning deployment and release of a given product.

Tree life spans and cycles necessitate longer-term timelines than those of most crop plants. Despite a perception that modern transgenic and other genetic engineering technologies are a “quick fix”, their application and testing remain subject to “tree time”.

TACF has two tools that allow us to track and implement quality control on the trees we produce and plant. The first is called the Germplasm Agreement (GPA). The GPA is a type of material transfer agreement (MTA) that is signed between TACF and someone using material from our research programs. The GPA ensures that the cooperator understands that, because the germplasm is still in a research trial, the material is not to be used for commercial purposes, and cannot be distributed beyond land which that person owns. Second, the GPA has provisions on guarantees. These research materials cannot be guaranteed for performance, and we reserve the right to recall the materials should they be found unsuitable for restoration.

The second tool we have is a customized, online database called dentataBase. This tool allows us to track the progress of our breeding and research programs and the locations of trees we have distributed as part of our research programming.

TACF is committed to ensuring any product it distributes and deploys is both safe and effective for long-term restoration goals.

Yes, TACF has provided information to regulatory agencies regarding its concerns with Darling’s performance, and the organization’s new stance that these trees should remain under permitted research status.

In the immediate future, American chestnut restoration will involve forest plantings of trees that incorporate various approaches to disease resistance (backcross, transgenic, gene-editing, etc.) in scientifically selected sites, and testing various treatments to overcome obstacles to growth and survival.  These plantings will require collaboration with diverse partners who are prepared to take chestnut restoration through the 21st century.

In the longer term, American chestnut restoration will involve monitoring forest plantings for flowering, reproduction, regeneration, recruitment, and population dynamics over one to two decades. It will also include monitoring sustained impacts on other plants, animals, and microbes, on soil health, and on other important ecosystem processes. We must continue to partner with diverse stakeholders who are committed to the long haul and to incremental improvement through the rest of this century and beyond.

American chestnut restoration is a much broader and more complex endeavor than developing blight-resistant varieties of trees and mass-producing them.

Successful reintroduction of self-sustaining and growing populations of our target species requires a whole-ecosystem approach. This means not only tree improvement for disease resistance and competitive ability, but also identifying and targeting optimal habitats and improving establishment techniques that protect from animal predation, novel pests and diseases, climate change, habitat degradation, and anything else that is likely to stand in the way of a seed becoming a seed-bearing tree.

The forests of eastern North America have changed drastically in many ways and will continue to change rapidly through this century and beyond. We must discover the role that disease-resistant American chestnuts will play in these new forests through a combination of predictive models and long-term in-forest performance trials.

Restoration is also a socio-ecological exercise, not solely biological or technical. Just as forests have and continue to change, so will the relevance of the American chestnut tree to current and future generations of people who will carry this work forward.

As in all scientific endeavors, TACF has gained much from the experience with the Darling lines. Enthusiasm for this project has expanded an already large network of engaged people with a passion for the species, within our chapters and across the range. Their efforts have conserved dozens of diverse, wild American chestnut populations, and public support for integrated solutions to forest health issues is at an all-time high. Our diversified portfolio of research initiatives and the partners implementing them ensures that other disease-resistant prototypes – those created from breeding and biotechnology methods that are already in the research pipeline – can be comprehensively vetted and released in a reasonable time frame.

The following achievements gained through the process of studying the Darling line will continue to have a positive impact on our research and restoration programming.

• Creation of additional, better-performing transgenic lines that can be used for research and deployment in the next few years (see What’s Next section above)
• Ongoing expansion of research and implementation partnerships
• Field tests with large sample sizes, wide geographies, and 12-month+ direct inoculations of research trees are essential to gathering appropriate performance indicators
• Increased acceptance of transgenic and biotechnologies for forest health applications
• Greater understanding of the regulatory process and its application for forest health concerns
• Expanded and ongoing conservation of wild-type American chestnut germplasm
• Deployment & improvement of high-light methods for “speed breeding”
• Increased standardization of processes between TACF and external research locations which minimize confounding variables

Your donation and membership was, and continue to be, vital for our capacity to research transgenic technologies. That funding has allowed us to comprehensively review the field performance of advanced-generation Darling lines, the analysis of which has led to this unfortunate, but science-based conclusion. The beauty of science is that we learn and then apply those lessons to future work.

TACF is grateful for the support of its thousands of members, donors, and volunteers. We have all embarked on this journey with an understanding that working with trees requires timespans that will surpass our lifetimes.

As outlined in our “What’s Next” section above, TACF follows a multi-pronged approach to American chestnut restoration and pursues a range of viable options simultaneously. With the disappointing realization that Darling 58 is not an effective pathway, we must now focus our efforts on other research initiatives that show promise.

Your membership and donation dollars are more important than ever in allowing TACF and its research collaborators to develop safe and effective methods toward restoration of the American chestnut. We hope you will continue to support the exciting new and ongoing research that TACF and its partners have in the pipeline. 

Distribution is dependent upon deregulation and then whether SUNY-ESF still chooses to distribute Darling 58/54 material at that time. TACF does not support further development or deregulation of the Darling line of American chestnuts because, in our opinion, they have unacceptable performance and underlying genetic issues that make them unsuitable for release and restoration.

The Darling tree is still pending approval by regulatory agencies. There is currently no information about whether trees from the Darling line will be distributed if they reach deregulated status.

TACF’s position is that the Darling line is ineffective as a restoration tree, and may compromise future generations of disease-resistant American chestnut populations. We are researching other transgenic lines that show promise and, as always, we encourage you to plant wild-type American chestnuts, assist with the planting and research efforts of our Chapters as a volunteer, and become a member of The American Chestnut Foundation to have priority access to the most advanced populations of disease-resistant material.

Due to performance concerns, likely caused by a constitutive promoter used to express the OxO gene in the Darling transgenic American chestnut, TACF is no longer planning to incorporate any Darling trees in our species restoration efforts.

The Darling tree also has yet to be deregulated (a step that TACF no longer endorses) and the original D58 event is at a very early stage of production. If we have to start at such an early stage of research and diversification of a transgenic product, we believe that our efforts would be much more efficiently placed on events with different genetic constructs. TACF looks forward to focusing on current and new alternative approaches.