The Animal Protection Program of USDA CSREES National Research Initiative (NRI) announces a funding opportunity as part of their competitive grants program. A research priority of the program, listed under subsection 2 on Animal Well-Being Assessment and Improvement, includes a request to “assess the behavior and well-being of genetically modified food animals enhancing animal well-being throughout the food production cycle.” This emphasis will provide information about how animals of agricultural importance in the U.S. interact with the production environment and respond to animal management practices. The program invites fundamental and mission-linked research proposals, as well as integrated proposals.

CONFERENCE: BIOSAFETY CONSIDERATIONS IN THE USE OF GENETICALLY MODIFIED ORGANISMS FOR MANAGEMENT OF ANIMAL POPULATIONS

Modern biotechnology can provide innovative approaches to management of animal populations, but what about potential risks? This online conference will facilitate exchange of views on a range of important issues such as the use of GM microorganisms, nematodes and insects for biological control (including those used to cause sterility in pest species for conservation and/or commercial purposes); the use of GM viruses to protect mammal populations against disease; and the use of GM insects for reducing disease transmission rates.

A long prevailing dogma has been that plants only use inorganic nitrogen sources such as ammonium and nitrate for their nutrition. As early as the middle of the previous century, plant scientists showed, however, uptake of organic nitrogen sources by plants. Despite this, only in the last 10 years or so have organic nitrogen and preferably amino acids been recognized as important nitrogen forms for plant nutrition. Several studies have shown that amino acids are of importance for plant nutrition in natural habitats, especially in alpine, arctic, and boreal regions where mineralization of organic matter and production of inorganic N is slow. Other studies suggest the capacity for root absorption of different amino acids is ubiquitous to plants. Furthermore, studies of different amino acid transporters have indicated that several of these have a very broad substrate range, including also D-enantiomers of amino acids. We have discovered, while conducting research in the fields just described, important differences between optical isomers of amino acids. The nutritional value of individual amino acids was tested on sterile-grown Arabidopsis thaliana. In these tests we could show that several L-amino acids promoted growth while none of the D-enantiomers did so. We also found that some D-amino acids were very toxic to plants (e.g., D-alanine and D-serine), while others had slight negative effects, and still others had no effect at all (e.g., valine and isoleucine).

D-amino acids and their metabolism
All protein amino acids except glycine are chiral molecules and have at least two optical isomers. Optical isomers are nonsuperimposable mirror images having identical physical and chemical properties, such as melting and boiling points, and chemical reactivity against molecules that are not optical isomer themselves.

The metabolism of amino acids is described in detail for a large number of different organisms. Studies of amino acid metabolism are in many cases narrowed down to only L-amino acids, although many organisms metabolize the less abundant optical isomer, D-amino acids. One of the most studied metabolic routes involving D-amino acids is in bacteria, where the peptidoglycan layer of the bacterial cell wall contains amino acids of both optical isomers. Peptidoglycan consists of peptides with both L/D and D/D amino acids. Furthermore, D-amino acid metabolism is also described in many other organisms, such as humans, mammals, birds, insects, fish, algae, and yeast, but very little is known about D-amino acid metabolism in plants.

One of the best known enzymatic pathways for metabolism of D-amino acid is via oxidative deamination by D-amino acid oxidase (encoded by the DAO1 gene). D-amino acid oxidase is one of the most studied enzymes, discovered more than 60 years ago, and is a model flavoprotein for which the structure and catalytic mechanism have been described in great detail. Enzymes such as the D-amino acid oxidase, which is almost ubiquitous to organisms of many kingdoms, is missing in all plants investigated hitherto.

During the 1970s, a fair amount of research was carried out within the field of D-amino acid metabolism in plants. The majority of that research used pea seedlings or other leguminous plants as study objects. Radiotracer studies with D-amino acids showed that plant metabolism of these compounds preferably leads to the formation of N-malonyl and N-acetyl derivatives. Such conjugation is believed to be a way to inactivate potentially toxic compounds. The fact that plants cannot use the optical isomer of protein amino acids as nitrogen sources, and that they are conjugated into what is thought to be dead end products, suggests that plants treat D-amino acids as intrusive compounds. Thus, plants metabolize D-amino acids differently than other organisms.

D-amino acids in biotechnology
The potential for using D-amino acids and their metabolism in biotechnology is suggested from what is described (above). Because plants lack the metabolic pathway common to many other organisms, they are sensitive to several D-amino acids and cannot use such compounds as N sources. We hypothesized that this sensitivity could be alleviated through expression of a D-amino acid metabolizing enzyme in a plant. Furthermore, we hypothesized that introducing this metabolism into a plant would also enable it to use D-amino acids as N sources. A candidate for such an experiment was the dao1 gene. Hence, our first test was to see if the toxicity exerted by D-serine and D-alanine could be relaxed through expression of dao1 and if D-amino acids promoted growth of these plants. These experiments gave very promising results and suggested to us that dao1 could function as a selectable marker.

One of the unique features of the D-amino acid oxidase selectable marker system is, however, that it enables both positive and negative selection, simply by choosing different selective media. This feature of dao1 was discovered when we screened for effects of a range of D-amino acids on wild type and on dao1-expressing plants and found that some D-forms that were indifferent for wild type plants had a strong negative effect on the transgenic plants. Selecting for the transgenic plants/tissues/cells occurs through dao1 alleviation of the toxicity exerted by D-alanine and D-serine, for example. D-alanine is decomposed by D-amino acid oxidase into ammonium, hydrogen peroxide, and pyruvate while D-serine forms hydroxyl-pyruvate instead of pyruvate when metabolized by the enzyme. Selecting against dao1-carrying plants, tissues, or cells is possible through conversion of non-toxic D-amino acids (e.g., D-isoleucine, D-valine) into toxic products (3-methyl-2-oxo pentanoic acid and 3-methyl-2-oxo butanoic acid, respectively) by the action of D-amino acid oxidase. In Figure 1, the effect of D-alanine and D-isoleucine on dao1-expressing and wild type plants is shown.

Figure 1. Arabidopsis thaliana wild type plants (left half of the plate) and plants expressing dao1 (right half of the plate) sown on media containing D-isoleucine (upper half of the plate) and D-alanine (lower half of the plate). Wild type plants are killed by D-alanine but not by D-isoleucine. Transgenic plants are not harmed by D-alanine but suffer when exposed to D-isoleucine.

The use of selectable markers is currently debated. There are arguments raised against the use of antibiotic resistance markers due to the risk that it might compromise the therapeutic value of antibiotics. Whether this risk is real or not is still an open question; in any case, it is still a matter of public concern and it is likely that these markers will be phased out, at least in commercial crops, in the near future. Obviously, the best way to treat the problem of selectable markers is to delete it. One advantage with the D-amino acid oxidase marker is that it can be used for both positive and negative selection. Thus, in combination with existing techniques for marker removal, dao1 may find its primary function in the production of marker free transgenic plants. Preliminary studies on a range of crop plants show that all species tested react similarly as Arabidopsis upon exposure to D-amino acids. This suggests that dao1 may work as a selectable marker in commercially important crops as well as in the model plant, and future research will show if, and to what extent, dao1 may replace selectable markers used today.

Reference
Erikson O, Hertzberg M, & Näsholm T. (2004) A conditional marker gene allowing both positive and negative selection in plants. Nature Biotechnology, 22: 455-458.

T7 RNA POLYMERASE-BASED OVEREXPRESSION OF FOREIGN GENES IN PLANTS
Vanga Siva Reddy

Potential of genetic engineering in plant biotechnology
Genetic engineering allows the introduction and expression of selective gene(s) derived from a wide range of organisms into plants. These genes may cause heritable changes in the plant’s ability to resist pathogens, sustain growth during adverse agro-climatic conditions, and improve shelf life of plant produce. In addition, plants are a possible source of recombinant proteins useful in industry and human health. In order to use a plant-based expression system as an alternative to microbial expression systems, further improvements in foreign gene expression are needed. Although significant increases in transgene expression can be achieved through promoter optimization, protein targeting, and codon optimization, the inability to achieve a high level of foreign gene expression in a desired tissue has limited its widespread application.

An alternate approach to overproducing foreign proteins in plants is through chloroplast genetic engineering. However, chloroplast transformation is achieved routinely thus far only in tobacco. Moreover, certain substances such as “edible-vaccines” and provitamin A are more desirable when produced in edible plant tissues (e.g., tomato fruit, potato tuber, and rice and wheat grains, etc.), which are mostly devoid of chloroplasts. Also, posttranslational modifications such as glycosylation of recombinant proteins, essential in some cases for their biological function, may be a limiting factor when expressed in chloroplasts.

The robust T7-expression system
The bacteriophage T7 RNA polymerase (T7 RNAP) is a single subunit DNA-dependent RNA polymerase that exhibits very high specificity despite its rather short length (23 bp). The T7 RNAP-based transcription is both rapid and processive, requiring no additional host factors for promoter recognition. At present, it is the most commonly used transcription system for overproducing recombinant proteins of commercial and academic interest in E. coli, a prokaryotic organism having single circular DNA as its genome. On the other hand, eukaryotic organisms such as plants have genomes that are very large, and the DNA is tightly packed into highly condensed chromosomes. Therefore, it was unknown whether T7 RNAP would be able to recognize its promoter and transcribe a randomly integrated foreign gene into plant chromosomes.

As reported in a recent publication¹, we demonstrated the potential of the T7 RNAP-based expression system in plants to overproduce foreign proteins in desired tissues constitutively and upon induction. To examine the species range, the T7 expression system was tested in tobacco, a dicot plant used as a model for transformation, and in rice, a monocot cereal crop cultivated all over the world.

Unlike the conventional approach in which foreign gene expression is dependent on transcription by host RNA polymerase, in the “T7 system,” foreign gene expression is solely dependent on transcription by the introduced T7 RNAP, and in turn, the expression of T7 RNAP is dependent on host RNAP. In the T7 system, foreign gene expression is modulated further by controlling the expression of T7 RNAP using various tissue-specific promoters or chemically inducible mechanisms. This was achieved by using a modified T7 RNAP coding region fused with SV40 nuclear localization signal² to target the polymerase to the nucleus. In this study, the uidA (GUS) reporter gene was cloned in the same construct and placed under T7 promoter and termination signal sequences. For a better comparison of expression levels, stably transformed tobacco plants were grown with the GUS gene placed directly under six promoters.

Tissue specific expression
To test tissue specificity of transgene expression under the T7 system, expression of T7 RNA polymerase was regulated through one of the six previously characterized tissue-specific plant promoters. Three promoters were tested: 1) the promoter for the small subunit of ribulose-bisphosphate carboxylase (rbcS-3A) that expresses in chloroplast-containing tissues such as leaves; 2) the kin1, and cor6.6 promoters that express in leaf tissue, but more prominently in guard cells of stomata; and 3) the phenylalanine ammonia-lyase (pal1) and pal1∆ (a deletion version) promoters that express differentially in different tissues but more prominently in conducting tissues.

Results from all six promoters demonstrated that recombinant protein can be expressed at several fold (3 – 10 times) higher levels as compared to transgene expressed under these promoters directly. The transcript initiation analysis showed that T7 RNAP recognizes its short (17bp) but highly specific promoter when integrated randomly into plant genome. Primer extension studies showed that GUS transcripts initiated from the same nucleotide, ‘G’, are specific for the T7 promoter. Similar to tobacco, GUS activity under the T7-system was high in rice plants when compared to GUS expressed directly under the CaMV 35S promoter. Again, transcript analysis revealed that the increased activity of GUS was due to an increase in uidA transcription by the T7 RNAP.

Current transformation methods for plants introduce foreign genes randomly into genomes. Consequently, large variations are observed in the level of transgene expression among independently transformed plants, depending on the site of their integration and the number of events integrated into the genome. This necessitates the production and screening of large numbers of transgenic plants to identify ones having the desired level of transgene expression, which is a laborious and time-consuming process, especially in the crop plants recalcitrant for regeneration and transformation procedures. At present, this is a major hurdle in several crop species, such as legumes, fruit trees, vegetable crops, and in certain cereals. Detailed expression analysis involving large numbers of transgenic plants from each group of plants transformed with different promoter-constructs revealed that variations in GUS expression was low among independently transformed plants under the T7 system, as opposed to large variations observed for GUS expression when expressed directly under any of the promoters tested. This capability of the T7 RNAP system to affect a near-uniform expression level in plants will be extremely useful for minimizing the number of transgenic plants required for screening.

Inducible expression
T7 RNAP-based recombinant protein production in E. coli is highly amenable to induction studies. To test the possibility of engineering a similar system capable of inducing the expression of recombinant protein in plants, a previously well-characterized tetracycline (Tc) inducible expression system was used³. Induction kinetics showed that GUS expression is completely repressed when the same plants also express a repressor protein (tetR) that binds to the modified 35S CaMV promoter driving the T7 RNAP. However, such a repression can easily be de-repressed with Tc treatment. The expression of GUS after Tc treatment was correlated with the co-transcription of both T7 RNAP and uidA (GUS) genes, demonstrating the inducible expression of GUS under the T7-system.

Conclusions and future perspectives
In principle, these studies demonstrate that the T7 system can be useful in both dicot and monocot plants, and the expression levels achieved are quite significant when compared to transgene expression observed directly under any of the plant promoters tested. The tissue specificity was maintained for the transgene under all promoters tested. Under the T7-system, transgene expression can be regulated through inducible mechanisms. Most importantly, there is a near uniformity in transgene expression among transgenic lines derived from independent transformation events. Such widely applicable, regulated, and tissue specific high-level expression makes the T7-system an effective tool for genetically manipulating various agronomic traits, for over-producing proteins, and for aiding in understanding gene functions. Expressing a large number of foreign proteins in a wide range of plant species is required to authenticate the final utility of the T7 RNAP-based expression system in plants.

References
1. Nguyen HT, Leelavathi S, & Reddy VS. (2004) Bacteriophage T7 RNA polymerase directed inducible and tissue specific overexpression of foreign genes in transgenic plants. Plant Biotechnol. J. 2: 301-310.

2. Dunn JJ et al. (1988) Targeting bacteriophage T7 RNA polymerase to the mammalian cell nucleus. Gene 68: 259-266.

3. Gatz C, Frohberg C, & Wendenburg R. (1992) Stringent repression and homogeneous de-repression by tetracycline of a modified CaMV 35S promoter in intact transgenic tobacco plants. Plant J. 2: 397-404.

HOS9 MEDIATED COLD ACCLIMATION OUTSIDE THE CBF REGULON
Ray A. Bressan, Jianhua Zhu, Paul M. Hasegawa

The genetic basis for plant cold tolerance, that is, which genes are needed to convey cold tolerance to plants, has been a long standing goal of plant scientists. Two general molecular genetic approaches toward achieving this goal have been implemented in the past decade or two using the model plant Arabidopsis. First, directly screening for plants with altered freezing tolerance has yielded several mutants such as the sfr mutants which are sensitive to freezing¹ and the eskimo mutant² that displays increased freezing tolerance. So far, few of the genes responsible for these altered freezing phenotypes have been identified. Only recently has one of the SFR mutant genes, sfr2, been cloned³. Another approach utilizing the identification of genes that have altered expression after exposure to cold temperatures such as the kin (cold induced), lti (low temperature induced) and cor (cold regulated) genes has resulted in the identification and cloning of more genes that are required for cold tolerance.

After the identification of the cold/desiccation induction recognition sequence motifs called CRT/DRE (C repeat/dehydration responsive element) in the promoters of cor target gene⁴, Stockinger et al.⁵ identified the CBF1 (cor binding factor) gene that encodes an APETELA2/ethylene response element class transcription factor that regulates the expression of cor target genes by binding to the CRT/DRE promoter region. Later, using yeast one-hybrid screening, other members of the CBF transcription factor family were identified and called DRE binding (DREB) proteins⁶.

Overexpression of a number of the CBF transcription factor genes in Arabidopsis has resulted in increased stress tolerance⁷. A modification of this approach toward identifying cold stress tolerance genes has utilized the CRT/DRE-containing promoter of the cor gene RD29A fused to the LUCIFERASE marker gene⁸. Genes that affect cold tolerance, such as HOS1, LOS2, and HOS2⁹, have been identified by using this luciferase marker strategy. Some of the genes that affect acclimation to cold appear to function as regulators of transcription. HOS1 is a negative regulator of genes that are targeted by CBF⁹ and appears to act through a transcription factor called ICE which controls CBF gene expression by binding to the CBF promoter¹⁰. The central role of the CBF family of genes in cold tolerance has been highlighted by the absence of any known genes (identified sequences) that affect cold tolerance without affecting expression of the CBF gene family. Because the CBF genes are not expressed before cold treatment⁷, the identification of mutants in genes such as ESKIMO that directly increase cold tolerance without cold treatment and subsequent induction of expression of CBF genes, and the known occurrence of cold-induced target genes outside of the CBF regulon¹¹, have indicated that there should exist cold tolerance signal pathway(s) independent of the CBF mediated regulon. Using the RD29A::LUCIFERASE screening strategy, Zhu et al.¹¹ have recently reported the existence of such a pathway that is mediated by the HOS9 gene product.

HOS9 encodes a putative homeodomain family transcription factor. Zhu et al. showed that the protein encoded by HOS9 when fused to GFP protein is localized to the nucleus. Using microarray analysis they also demonstrated that the HOS9 gene controls the expression of about 175 gene targets that appear not to be in the CBF regulon. Also, 41 of the genes targeted by HOS9 were reported to be cold-induced. Thus, HOS9 appears to encode a nuclear factor that acts, at least in part, separately from the CBF family and its target genes to mediate cold-induced cold acclimation in Arabidopsis. The hos9 mutation led to the alteration of other characteristics including trichome development, growth rate, and flowering time. Interestingly, both growth rate and flowering time are also altered in CBF-overexpressing transgenic plants⁷.

Although it is known that CBF affects growth through a gibberillic acid-mediated mechanism, the mechanism by which HOS9 controls growth is unknown. Thus it appears that both CBF-mediated and HOS9-mediated pathways control cold acclimation, growth rate and flowering time. Growth rate, and flowering time are phenological traits that are associated with stress avoidance mechanisms¹². The consistent alteration in growth rate and flowering time in mutants with altered stress tolerance indicates that these are likely to be important traits that also affect cold acclimation and tolerance and need to be coordinated by cross-talking signal pathways. Like CBF genes, HOS9 has low expression in flower tissues that are particularly vulnerable to cold temperature, especially in valuable fruit crops such as citrus and stonefruit species.

The strategy of screening mutants in a genetic background containing a cold-inducible promoter (R929A or CBF) fused to the luciferase marker has been remarkably successful in finding genes that control cold acclimation and should result in the discovery of other components of the cold acclimation signal pathway system.

References

1. Warren G, McKown R, Martin AL, Teutonico V (1996) Isolation of mutations affecting the development of freezing tolerance in Arabidopsis thaliana (L.) Heynh. Plant Physiol 111:1011-1019

2. Xin Z, Browse J (1998) eskimo1 mutants of Arabidopsis are constitutively freezing-tolerant. Proc Natl Acad Sci USA 95: 7799-7804

3. Thorlby G, Fourrier N, Warren G (2004) The SENSITIVE TO FREEZING2 gene, required for freezing tolerance in Arabidopsis thaliana, encodes a β-glucosidase. Plant Cell 16: 2192-2203

4. Yamaguchi-Shinozaki K, Shinozaki K (1994) A novel cis-acting element in an Arabidopsis gene is involved in responsiveness to drought, low-temperature, or high-salt stress. Plant Cell 6: 251-264

5. Stockinger EJ, Gilmour SJ, Thomashow MF (1997) Arabidopsis thaliana CBF1 encodes an AP2 domain-containing transcription activator that binds to the C-repeat/DRE, a cis-acting DNA regulatory element that stimulates transcription in response to low temperature and water deficit. Proc Natl Acad Sci USA 94: 1035-1040

6. Liu Q et al. (1998) Two transcription factors, DREB1 and DREB2, with an EREBP/AP2 DNA binding domain, separate two cellular signal transduction pathways in drought- and low temperature-responsive gene expression, respectively, in Arabidopsis. Plant Cell 10: 1391-1406

7. Thomashow MF (2001) So what's new in the field of plant cold acclimation? Lots! Plant Physiol 125: 89-93

8. Ishitani M et al. (1998) HOS1, a genetic locus involved in cold-responsive gene expression in Arabidopsis. Plant Cell 10: 1151-1161

9. Viswanathan C, Zhu J-K (2002) Molecular genetic analysis of cold-regulated gene transcription. Phil Trans R Soc Lond B 357: 877-886

10. Chinnusamy V et al. (2003) ICE1: a regulator of cold-induced transcriptome and freezing tolerance in Arabidopsis. Genes & Dev 17: 1043-1054

11. Zhu J t al. (2004) An Arabidopsis homeodomain transcription factor gene, HOS9, mediates cold tolerance through a CBF-independent pathway. Proc Natl Acad Sci (USA) 101: 9873-9878

12. Maggio A, Joly RJ, Hasegawa PM, Bressan RA (2003) Can the quest for drought tolerant crops avoid Arabidopsis any longer? In: (Goyal SS, Sharma SK, Rains DW, eds) Journal of Crop Production, The Haworth Press, Inc. Vol 7, No. 1/2, pp. 99-129

For over a decade, researchers have devised methods for producing genetically modified (GM) plants that synthesize therapeutically active proteins and industrial chemicals, a technology known as plant molecular farming or biopharming. Hailing the technology as the next wave in biotech, supporters claim that the biopharm industry may be worth $100 billion by 2015. This enthusiasm stems from the conviction that biopharming can provide a means to manufacture therapeutic compounds and chemicals at lower cost and in greater amounts than conventional techniques. The past half year brought signs that justify such optimism.

Accelerating advances
In June, scientists at the National Institute of Public Health in Tokyo announced their success in creating GM tobacco plant cells designed to synthesize a human recombinant monoclonal antibody that binds with hepatitis B virus. The plant-made antibodies stimulated cytotoxicity in a manner similar to anti-hepatitis B virus human antibodies used in the clinic. The National Institute’s Dr. Akira Yano told Reuters Health that, while technical problems in antibody production should be resolved in a few years, the greater challenge lies in public acceptance of plant-derived therapeutics.

The European Union has shown its interest in plant-made pharmaceuticals by investing 12 million euros for the first five years of the Pharma-Planta Consortium. Officially launched on July 12, the consortium comprises academic laboratories and industrial partners from 11 European countries and South Africa. The group aims to perfect techniques for the production of antibodies and vaccines to prevent or treat human diseases. They intend to implement the entire process of developing a plant-made therapeutic: from the design of recombinant genes to clinical trials by 2009. Pharma-Planta’s candidate plants include tobacco, maize, potatoes, and tomatoes, with a preference for plants that can express desired protein products in high quantities in easily harvestable seeds. Their first product will probably be an antibody that can be used to block HIV transmission, followed by a post-bite vaccine for rabies.

In the United States, ProdiGene Inc. (College Station, TX) announced the availability of TrypZean™, the first commercial plant-produced recombinant bovine-sequence trypsin. The company says that the enzyme, synthesized in maize, has essentially the same properties as bovine trypsin, which is used to manufacture insulin and vaccines, and to enhance wound healing. ProdiGene also produces AproliZean™, a recombinant bovine aprotinin expressed in maize. Aprotinin—an inhibitor of trypsin, plasmin and other serine proteases—can be used to reduce bleeding and provides a component in fibrin sealants. During the summer, ProdiGene requested permission from the U.S. Department of Agriculture to begin large-scale cultivation of GM corn in an area southwest of San Antonio, Texas. These fields would generate the company’s trypsin and aprotinin products.

California almost became the first state to host production-sized fields of a drug-producing food crop. Ventria Bioscience (Sacramento, CA) had applied for the go-ahead to cultivate GM rice that synthesizes the antimicrobial pharmaceutical proteins, lactoferrin and lysozyme. Although the California Rice Commission favored Ventria’s plans for a 120-acre development, the California Department of Food and Agriculture rejected the application, because the company lacked approval from federal agencies. A few months later, reports surfaced that Ventria has decided to scale back plans for production of pharmaceutical rice in California. The company intends to plant a test crop of lactoferrin-producing GM barley in Iowa, a state that does not have an established barley industry.

While many biopharming products require purification before use, edible vaccines do not. Synthesized in food crops, these vaccines employ plant components as delivery vehicles. ProdiGene recently announced a study of its edible vaccine targeted against transmissible gastroenteritis virus. The company engineered transgenic corn to produce recombinant vaccine antigens in the seed. When porcine patients ate ground corn feed that contained the vaccine antigens, they displayed an increased lactogenic immunity. Dow AgroSciences (Indianapolis, IN) shares an interest in vaccines aimed at the animal health industry. The company is building a plant-based vaccine manufacturing facility in Lincoln, NE, and plans to launch the first plant-made vaccine for the poultry industry in 2006.

Pete Siggelko, vice president of Dow AgroSciences, recently expressed confidence about biopharming during a Congressional hearing. The manufacture of “antibodies, vaccines, industrial products and pharmaceuticals is no longer a pipe dream,” he said. “It is a reality.” Not everyone wishes to see the promise of biopharming realized, however.

Arresting advances
In June, the Center for Science in the Public Interest published a report about the acceleration of biopharming’s progress. The Center observed that the USDA received 16 new applications for biopharming permits from May 2003 to April 2004, with half submitted during the last three months. The organization intended to warn about biopharming’s progress, not to praise it.

The Center’s report emphasizes that many of the USDA applicants want to plant a GM crop in a state that harbors significant commercial production of a conventional counterpart for human and animal consumption. This perceived threat against established agriculture sparked protests against GM corn in Texas and GM rice in California.
ProdiGene’s plans for large-scale production of its trypsin- and aprotinin-producing transgenic corn in Texas drew a protest from the Grocery Manufacturers of America. In a letter to the USDA, the group opined that the government provides inadequate oversight of crops engineered for pharmaceutical and industrial purposes and insisted that the FDA should evaluate the safety of the crops before they are approved for cultivation.

At the same time, consumer and environmental organizations requested California state agencies to investigate potential hazards posed by Ventria’s plan to produce pharmaceutical drugs from GM rice. The Friends of the Earth, Center for Food Safety, Consumers Union, and Environment California delivered a 22-page report detailing their concerns that the “pharmaceutical traits” of Ventria’s GM rice could pass to conventional rice through transport in the guts of birds, flooding, or pollen dispersal by bees or high winds. They also listed potential environmental impacts, such as the creation of hardier weeds and disruption of soil ecology. The groups urged California authorities to impose a moratorium on such plant-produced drugs until state agencies have conducted an independent review.

The report brought a swift rebuttal from representatives of the International Academy of Life Sciences. In a letter to California health, agricultural and environmental agencies, Drs. Hilmar Stolte (Hannover Medical School, Germany) and Robert Rich (University of Illinois, Urbana-Champaign) countered that the report does not present an objective or accurate perspective of the risks. Characterizing the document as a “laundry list of potential problems,” they accused the report’s authors of intentionally creating confusion by presenting hazards as if they were risks. “Just because a hazard can happen,” they wrote, “does not necessarily mean it will, or that it is even likely to happen.” Drs. Stolte and Rich concluded that the claimed health and environmental risks of the GM rice are negligible.

Using a different tactic to terminate Ventria’s plans, the SLO GE Free group promotes Measure Q-04, an initiative on the November ballot that would prohibit the propagation of GM organisms in San Luis Obispo County. The county Farm Bureau, the San Luis Obispo Chamber of Commerce, and academics at California Polytechnic State University oppose the measure, which they say is too broad and would chill the county’s small biotechnology industry. The notion of forbidding the growth of any GM organism is not novel. California’s Mendocino and Trinity counties have such bans in place, while Humboldt, Marin and Butte counties have measures on their November ballots that would establish similar prohibitions.

Hawai’i hosted yet another tactic for discouraging biopharm technology. In August, Judge David A. Ezra made history by ordering the USDA to identify the Hawaiian locations of four companies’ open-air test sites for food crops engineered to produce industrial chemicals and drugs. The order ensued from a lawsuit filed by Earthjustice in the Honolulu federal district court on behalf of the Center for Food Safety. The Department of Agriculture, State of Hawai’i, had denied the Center’s earlier request for this data on the basis that the records contain confidential business information protected from disclosure under federal law. According to Ezra’s order, the locations of the test fields will be revealed to Earthjustice and its client, but they must keep the information confidential for at least 90 days. During this time, the defendants can try to persuade the judge that public disclosure will result in a specific harm, such as the destruction of the fields. If the judge is not convinced, the information will become publicly available.

Representatives on both sides of the case say that this would be the first time in the United States that locations of biopharm tests would be revealed to an outside party. The decision could encourage similar disclosures in other states. The American Farm Bureau Federation’s public policy specialist, Michelle Gorman, said that her organization is concerned about the effect of the ruling on farmers. “Disclosure of the test site locations has no role in protecting public health or the environment,” she said. “To the contrary, releasing the location of the test sites could leave them vulnerable to vandalism.” The destruction of a biopharm test field not only affects farmers. Michael Rodemeyer, executive director for the Pew Initiative on Food and Biotechnology, warns that vandalism may disperse genetically engineered crops, creating the harm feared by biopharming’s adversaries.

A combined symposium and discussion session entitled, “Biodiversity and Biotechnology: Understanding the Potential Conservation Risks and Benefits of Genetic Engineering,” was held at the 2004 Society for Conservation Biology (SCB) Annual Meeting in New York City on July 31 and August 1. The SCB is an international organization of conservation professionals who are dedicated to promoting the “scientific study of the phenomena that affect the maintenance, loss, and restoration of biological diversity”¹.

The symposium and discussion were organized by Kelly Paulson and Erika Rivers, graduate students at the University of Minnesota’s (Twin Cities) Conservation Biology Program, and Dr. Emily Pullins, former Biotechnology Governance Program Manager at the Institute for Social, Economic, and Ecological Sustainability (ISEES) at the University of Minnesota. The organizers linked the symposium with a discussion session to meet two core needs of the SCB members: 1) to educate conservation practitioners about the potential benefits and risks of genetic engineering to biodiversity, and 2) to initiate a dialogue within the SCB about potential actions that the Society might take to address the myriad issues associated with biotechnology and biodiversity conservation. Prior to this event, the SCB journal had only considered the potential of genetic engineering for conservation purposes in one paper².

It appears from the attendance at the event that conservation biologists were eager to hear more about the potential—and risks—of biotechnology for conservation of biodiversity. Both sessions were well attended, with eleven symposium papers drawing audiences of 35–150 conferees, and the discussion session engaging approximately 50 conferees in a conversation about the potential actions the SCB might take to address the roles (positive and negative) of biotechnology in the conservation of biodiversity. Topics covered in both of the sessions included using genetic engineering for conservation purposes, the role of risk assessment, and how SCB and its members might become more involved in the issue of genetic engineering.

Of great interest to symposium participants was the potential use of genetically engineered organisms to further a wide variety of conservation goals—from controlling exotic species, to increasing productivity of already-converted agroforestry lands, to the conservation benefits of alleviating human health problems. For example, symposium speaker, Dr. Ron Thresher of the Commonwealth Scientific & Industrial Research Organisation (CSIRO), discussed the possibility of using genetically engineered carp to control destructive non-native carp populations in Australian waterways. In a similar vein, CSIRO’s Dr. Tony Peacock outlined the potential of a genetically engineered immunocontraceptive virus as a means of controlling Australia’s notorious exotic rabbits. In the realm of silviculture, Dr. Steve Strauss, of Oregon State University, discussed the potential of genetically engineering trees for more efficient conversion to wood products, implying that less land would be needed for agroforestry operations. The potential of genetic engineering to alleviate human health problems by genetically engineering mosquitoes resistant to malaria was introduced by Harvard University’s Dr. Andrew Spielman, along with guidelines for implementing such a technology.

Concerns more familiar to Conservation Biologists were represented by speakers such as Dr. LaReesa Wolfenbarger of the University of Nebraska at Omaha, who joined Dr. Karen Oberhauser of the University of Minnesota – Twin Cities in elucidating the potential of agricultural biotechnology to harm non-target organisms. Drs. Anne Kapuscinski and David Andow, both from the University of Minnesota, introduced the intricacies of risk assessment as applied to genetically engineered fish and the potential for development of pest resistance to genetically engineered crops, respectively. To sate the political appetites of the Society’s multidisciplined audience, presentations by Drs. Susan Haseltine (U.S. Geological Survey) and Ryan Hill (Secretariat of the Convention on Biological Diversity) outlined ways in which SCB members could direct their resources to help such organizations craft scientifically grounded policies vis-à-vis genetic engineering.

Regardless of whether a genetically engineered organism is intended for conservation, human health, or consumer purposes, it is customary to assess the ecological risks posed by this technology by some means. The intricacies of risk assessment captured the interdisciplinary nature of the Society’s membership: the science, policy, social consequences, limitations, and even philosophical underpinnings of risk assessment were discussed at length. Dr. David Ehrenfeld (Rutgers University), a co-founder of the SCB and its academic journal, raised many concerns about the utility of risk assessment, the degree of certainty we can expect from such assessments, as well as the failures of many previous “techno-fixes” employed in the name of conservation biology. Many discussion participants expressed a desire for a more proactive and holistic risk assessment process, with a transparent structure that allowed for more public and scientific input. One participant concluded the event with the interesting observation that by becoming more involved in defining scientific and ethical guidelines for adequate risk assessment, and by engaging in risk assessment policy formation (such as the opportunities for input presented by the Convention on Biological Diversity), Conservation Biologists might be in a better position to advocate for the development and evaluation of biotechnologies that serve the public good.

THE PEW INITIATIVE ON FOOD AND BIOTECHNOLOGY (PIFB) POSTS ARCHIVED WEBCASTS

Biotech Bugs Conference
On September 20-21, 2004, in Washington, DC, a workshop was convened to look at the science and public policy surrounding the release of GM insects. The Pew Initiative on Food and Biotechnology (PIFB) hosted this two-day multidisciplinary workshop on “Biotech Bugs: A look at the science and public policy surrounding the release of GM insects.” The workshop provided an exploration of the potential benefits and risks of genetically engineered insects and the public policy and ethical implications of releasing them. Representatives of government, academia, consumer and environmental groups, and policy leaders attended this important event.