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Despite benefits, commercialization of transgenic horticultural lags

David Clark Harry Klee Abhaya Dandekar able in the United States: , sweet commercialized. Disease resistance, par- ▼ corn, squash and a carnation. Except ticularly to viruses, can be developed for transgenic papaya, which accounts using , and potato and for approximately 50% of the Hawaiian papaya engineered for virus (HASS 2001), the fraction of the to- resistance have been commercialized, The acreage of agronomic crops tal horticultural commodities represent- but many potential applications are cur- (soybean, cotton, corn and canola) ed by transgenic varieties is miniscule. rently underutilized. Improving traits developed using recombinant DNA The absence of significant com- that directly benefit consumers, such as technology has expanded dramati- mercialization of transgenic varieties nutritional or aesthetic quality, is also cally since their introduction in 1996, in is not due to lack of technically feasible now in many horti- while the commercialization of bio- potential products or value (Dandekar cultural crops, but only a few products and Gutterson 2000; see sidebar, page have reached the market. tech horticultural crops (vegetables, 94). The basic techniques of molecular , nuts and ornamentals) has Approved traits and technologies have become routine, and con- languished. This is not due to a lack siderable research is being conducted on The major technologies that have of both current and potential traits horticultural crops. For example, her- been approved and widely adopted that could be utilized in horticultural bicide resistance has been transferred by the industry focus on input traits, crops, as ongoing research is identify- into bentgrass and bluegrass to make or those affecting production of the ing a diverse array of applications. control in municipal and highly crop rather than the qualities of the fi- However, commercialization is stalled managed turf environments such as nal product. Although most approved golf courses more efficient. However, genes confer insect resistance and herbi- by market reluctance to accept bio- they have not been commercialized. cide tolerance, a range of genetic traits tech products, particularly in the ab- Similarly, some horticultural crops, in- has been approved by the U.S. regula- sence of clear benefits to consumers. cluding lettuce and , have been tory system (table 1). High regulatory costs and restricted engineered with resistance Insect resistance. Insect resistance access to intellectual property create and tested in field trials but remain un- has been engineered primarily by using additional hurdles for specialty crops. These challenges are causing the hor- ticultural industry to forego a number Florigene of current benefits. New products with clear advantages for producers, marketers and consumers may be required before the potential of bio- technology can be realized.

2003, nearly 106 million acres of transgenic or genetically engineeredIN (GE) crops was planted in the United States, part of 167 million acres of such crops grown worldwide (James 2003). Despite the fact that the first commercialized transgenic food crop was the Flavr Savr tomato, four agronomic crops (corn, soy, cotton and canola) account for virtually all of the current acreage. Last year, only four horticultural crops developed using re- Florigene, of Melbourne, Australia, markets transgenic carnations engineered for blue-violet combinant DNA technology were avail- color under the variety name ‘Moonshadow’.

http://CaliforniaAgriculture.ucop.edu • APRIL-JUNE 2004 89 TABLE 1. Approved transgenic traits for U.S. crops

Trait Examples Herbicide tolerance Bromoxynil, glufosinate, , sulfonylurea Flavr Savr tomato. Transgenic horti- Insect resistance Bt kurstaki, Bt tenebrionis cultural crops providing direct benefits Virus resistance Papaya ringspot virus, cucumber mosaic virus, zucchini to the consumer have also been devel- yellow mosaic virus, watermelon mosaic virus, oped. Calgene’s Flavr Savr tomato potato roll virus, potato virus Y silenced the gene encoding polygalac- Male sterility Barnase/barstar turonase, an enzyme implicated in Modified ripening ACC synthase, ACC deaminase, SAM hydrolase, softening. The expectation was that the polygalacturonase tomato would soften and spoil more Modified oils High lauric, myristic, oleic acids slowly and could be picked at a later stage of maturity. This later harvest, in principle, would permit greater devel- opment of flavor compounds and bet- two classes of bacterial genes derived Virus resistance. The use of viral ter taste. This product, first marketed from Bacillus thuringiensis (Bt) ssp. coat protein genes to confer resistance in 1994, was a success with consumers kurstaki and ssp. tenebrionis (de Maagd has been approved for several virus- but failed economically for a variety of et al. 2003). These Bt genes control a crop combinations (table 1). The most reasons (Martineau 2001). This same broad spectrum of lepidopteran and commercially successful has been pa- gene was also used in a tomato variety coleopteran insects, respectively. The payas engineered for resistance to the processed for paste and marketed by genes have been approved for use in papaya ringspot virus. This product Zeneca in the United Kingdom. The major row crops (feed corn and cotton) has revived the Hawaiian papaya in- trait reduced processing costs and and some horticultural crops (sweet dustry, which was devastated by the vi- consumers accepted the clearly labeled corn and potato). Potato and sweet rus in the 1990s (see sidebar, page 92). product, until the European uproar corn varieties engineered for resistance Small acreages of transgenic squash over biotech foods forced it off the su- to Colorado potato beetle and corn resistant to mosaic viruses are also permarket shelves. earworm, respectively, were in com- grown. Virus-resistant potato varieties U.S. regulatory agencies also ap- mercial production for several years were formerly commercialized but are proved several other delayed-ripening and were technically and agronomi- not currently being marketed. Newer tomato varieties based on strategies tar- cally successful, allowing significant technologies, such as RNA interfer- geted to block the ethylene biosynthetic reductions in insecticide use (Shelton ence or RNA silencing (Waterhouse et pathway (ACC deaminase and anti- et al. 2002). However, the transgenic al. 2001), offer promise for developing sense/cosuppressed ACC synthase) that potato varieties were withdrawn from resistance to other damaging diseases, is essential for ripening. None of these the market after major processors and such as those caused by geminiviruses products is currently marketed, despite distributors chose not to purchase and (Gilbertson et al. 1998). their technical feasibility and potential market them. Bt sweet corn, while still consumer benefits. Rather, they were available, is not widely grown for the preempted by a nonbiotech approach same reason (Cornell Cooperative Ex- utilizing the naturally occurring rin tension 2003). mutant of tomato that delays fruit rip- Herbicide tolerance. Several ening. Heterozygous produce herbicide-tolerance genes are registered Courtesy of USDA-ARS fruits that ripen at a significantly slower for use. The most widely commercial- rate than normal fruits. A nontransgenic ized gene is a bacterial enzyme confer- approach achieved essentially the same ring tolerance to glyphosate, the active objective, and aggressive breeding and ingredient of Roundup herbicide. marketing of the long-shelf-life rin hy- Transgenes conferring tolerance to bro- moxynil (Buctril), glufosinate (Liberty) and sulfonylurea (Glean) are also approved for use in a wide variety of crops. In addition, crops tolerant to imidizolinone (Clearfield) and sulfony- Wilson/USDA-ARS Doug lurea herbicides have been developed through nontransgenic methods based on natural or induced mutations. How- ever, no horticultural crops engineered Left, scientists are investigating ways to prevent Pierce’s disease in grapes by genetically en- for herbicide resistance have been gineering genes from a naturally resistant variety, Muscadinia rotundifolia, into susceptible commercialized, although several have varieties. In a peer-reviewed study, a parasitoid that controls the diamondback moth, been developed and tested. right, a canola pest, was not affected by Bt canola.

90 CALIFORNIA , VOLUME 58, NUMBER 2 brid tomatoes made the transgenic ap- potential mar- proach redundant. ket (Gillis 2000). Lengthening time. Clearly, growers,

Similarly, virtually all bananas mar- distributors and site Web Courtesy of UC Biotech keted in the United States are naturally consumers must deficient in their ability to initiate eth- all see biotech ylene synthesis, allowing them to be crops as in their shipped green and ripened by exposure best interests for to ethylene gas prior to sale. However, commercialization where natural mutants are not available, to be successful. the general approach of manipulating Products offering ethylene synthesis has great potential compelling value for application in other climacteric fruits could alter the (whose ripening is mediated by ethyl- economic forces in- Biotechnology is contributing to the development of sensitive diagnostic ene), particularly tropical fruits with fluencing producer techniques. Above, samples are placed in wells and subjected to a short postharvest lives such as , choices and could color-based detection system; yellow shows evidence of disease. papaya and banana. A higher quality create consumer fruit bringing tangible value to the con- demand to pull such products through provide broader spectrum disease sumer could improve the market accep- the marketplace. control and reduced dependence upon tance of biotech crops. Loss of . One factor that chemical pesticides (Lincoln et al. 2002). Despite their early introduction and may significantly alter grower economics Resistance to viral diseases would be initial market success, and in contrast to is the loss of currently registered pesti- valuable in many horticultural crops, as what many consumers may believe, the cides due to environmental and health there are few other options for control, only biotech horticultural commodities concerns. For example, methyl bromide and methods for engineering virus re- currently marketed in the United States is scheduled to be withdrawn from use sistance are well established. are Hawaiian , a small amount in the United States in 2005 because it fruit, nuts and grapes. Research of squash and sweet corn, and a few contributes to depletion of the ozone is well under-way to build a robust plat- carnation varieties. The commonly cited layer. Methyl bromide is widely used in of technologies to utilize estimate that as much as 70% of food horticultural crops to control soilborne in the discovery of useful traits for products in U.S. supermarkets contain diseases and and to fumigate (Dandekar et al. 2002). Transformation ingredients from GE crops is not attrib- harvested crops to eliminate insects. The technology has been developed and utable to fruits and vegetables, but rath- majority of this use is in horticultural trait evaluation is under way on apple, er to the widespread use of corn, canola crops (primarily , tomatoes, almond, , citrus, walnut, pear, and soybean oil, soybean protein, corn peppers, ornamentals and nurseries, and plum, grapevine and persimmon. Good and related products in virtually tree crops) with California and Florida progress has been made in developing all processed foods. together accounting for 80% of the 35 resistance to codling moth and fireblight million pounds applied each year for in apple, plum pox virus in plum/ Opportunities for biotechnology preplant fumigation (Economic Research Prunus, crown gall and codling moth in As global acreage of biotech agro- Service 2000). Many genes are available walnut, citrus tristeza virus (CTV) in cit- nomic crops sustains its eighth consecu- that potentially could be used to enable rus and Pierce’s disease in grapevine. tive year of double-digit growth rates alternative weed-control strategies. Engineering of resistance to cod- (increasing 15% from 2002 to 2003) Horticultural crops are also limited ling moth in apple to reduce the use of (James 2003), it is paradoxical that the in the numbers of herbicides registered chemical pesticides has advanced to the trend in horticultural crops is exactly for use. Loss of registration for a few point of commercial interest in product the opposite, particularly as many of key chemicals could markedly limit development. Work is also under way to the approved genes fit naturally with grower options, making crop resistance develop productivity and quality traits the needs of horticultural crops. Fear of to broad-spectrum herbicides more criti- such as modified metabolism and consumer rejection on the part of both cal. Resistance to fungal and bacterial ripening in apple and regulation of self- producers and marketers of horticul- diseases would also be desirable, as in incompatibility in almond and other tural products is a major impediment to some areas extensive use of pesticides is Prunus species. Some deployment strat- wider utilization of biotechnology, even currently undertaken for their control. egies for transgenic trees are also being though many consumer polls do not As for herbicides, it is also difficult to developed, such as the use of transgenic find a majority negative opinion about it maintain registrations for minor crops trees as “trap crops” to control insects (see page 99). Many food companies are grown on smaller acreages, which are in conventional and the use of unwilling to risk the consequences of primarily horticultural. Biotech strate- transgenic rootstocks to control diseases alienating even a small fraction of their gies are being developed that could and pests in nontransgenic scion variet-

http://CaliforniaAgriculture.ucop.edu • APRIL-JUNE 2004 91 Virus-resistant transgenic papaya helps save Hawaiian industry

Dennis Gonsalves

he pivotal year in the history of gated and established Hawaii’s papaya industry was in a field trial in T1992. In May 1992, papaya ringspot vi- Waimanalo on Oahu rus (PRSV) was discovered in Puna on in April 1992. By De- Hawaii Island, where 95% of Hawaii’s cember 1992, it was

papaya was being grown. Just one evident that line 55-1 Manshardt Courtesy of R. month earlier, a small field trial to test was resistant under the resistance of a transgenic papaya field conditions. line had been started on Oahu Island, From the 1992 field where papaya production had previ- trial, two cultivars ously been devastated by PRSV. The were developed and timely commercialization of PRSV- designated ‘SunUp’ resistant transgenic papaya trees has and ‘Rainbow’. ‘Sun- revived Hawaii’s papaya industry and Up’ is homozygous provides an example of the challenges for the coat protein and opportunities for horticultural bio- gene while ‘Rainbow’ Two varieties of papaya resistant to papaya ringspot virus technology. is an Fl of have been developed using biotechnology: SunUp, left, and Rainbow, right. They have performed well for Hawai- In 1945, D.D. Jensen made the first ‘SunUp’ and the non- ian growers, even under prolonged and heavy disease report in Hawaii of PRSV, a potyvirus transgenic ‘Kapoho’. pressure. that is transmitted nonpersistently by Unfortunately, by Oc- aphids (Gonsalves 1998). PRSV was tober 1994, PRSV had spread through- tion had dropped to only 26 million first discovered on Oahu and caused out much of Puna, causing HDOA to pounds as PRSV spread throughout such severe damage that the papaya abandon rouging efforts to slow the the region. Since then, the transgenic industry was relocated to Puna in the spread of PRSV. The was on to varieties have enabled farmers to late 1950s and early 1960s. The papaya move the transgenic papaya line to reclaim infected areas and in 2001, industry expanded and prospered in commercialization. A 1995 field trial in Puna produced 40 million pounds of Puna conclusively showed papaya. The resistance has held up The experience in Hawaii shows that ‘SunUp’ and ‘Rainbow’ remarkably well and remains stable were resistant under pro- after 5 years of extensive plantings. that transgenic virus resistance is longed and heavy disease Hawaii also exports papaya to an excellent approach for controlling pressure. Canada and Japan. The transgenic viral diseases in horticultural crops. The U.S. Department of papaya was recently deregulated in Agriculture’s Animal Plant Canada, which is a relatively small Health Inspection Service market for Hawaii. The main chal- Puna, primarily because PRSV was (APHIS) deregulated transgenic line lenge is deregulation of transgenic absent. However, by the 1970s PRSV 55-1 in November 1996, and the U.S. En- papaya in Japan, where Hawaii sells was found only about 19 miles away vironmental Protection Agency deregu- about 30% of its papaya. Presently, in Hilo, and the Hawaii Department of lated it in August 1997. The consultation nontransgenic papaya must also be Agriculture (HDOA) took rouging (the process with the U.S. Food and Drug produced in Hawaii to satisfy the removal of infected trees) and quar- Administration was completed in Sep- Japanese market, but this is increas- antine actions to prevent its spread to tember 1997. Licenses to commercialize ingly difficult due to the disease Puna. In 1986, efforts were initiated to the transgenic papaya were obtained by pressure. Exporters face added ex- develop a virus-resistant transgenic the Papaya Administrative Committee penses to guard against the acciden- papaya by transforming commercial in Hawaii by April 1998. A celebration tal shipment of transgenic papaya lines of Hawaiian papaya with the coat was held to mark the debut of the trans- to Japan. In December 2000, Japan’s protein gene of PRSV from Hawaii. genic papaya on May 1, 6 years after Ministry of Agriculture, By 1991, the team of Maureen Fitch, PRSV was discovered in Puna and after and Fisheries approved line 55-1, Jerry Slightom, Richard Manshardt the first field trial of line 55-1 was initi- and the Ministry of Health, Labor and Dennis Gonsalves identified a ated. The transgenic fruit is currently and Welfare is reviewing a recently transgenic line (55-1) that showed sold throughout the United States. submitted petition for deregulation. resistance under inocula- In 1992, Puna produced 53 million Anticipated approval of transgenic tions. These plants were micropa- pounds of papaya, but by 1998 produc- papaya in Japan will allow Hawai-

92 CALIFORNIA AGRICULTURE,AGRICULTURE, VOLUMEVOLUME 5858,, NUMBERNUMBER 22 Simply the diversity of crops utilized in horticulture slows the adoption of new technologies. For example, as many as 60 distinct cultivars of iceberg lettuce alone may be grown throughout the year.

ies (see sidebar, page 96). The latter ap- over years of traditional hybridization proach avoids the task of transforming and selection, or creating new fragrances D. Gonsalves and S. Ferreira Gonsalves and S. D. many varieties of a particular tree crop in plants, has considerable potential and and in the future may be used to regu- appeal. Research on genes controlling the late quality and productivity traits. different biochemical pathways for vari- Nutrients, consumer qualities. Al- ous floral fragrances is being conducted though more difficult technically and on wild plants and on crops such as snap- therefore not close to market, there dragon, petunia and rose (Vainstein et al. are many potential opportunities for 2001). enhancing the nutritional value or con- Plant size. Currently, growth- sumer appeal of horticultural products regulating chemicals are applied to through biotechnology. In addition to ornamental plants to inhibit gibberel- modification of ripening, projects to lic acid (GA) synthesis and reduce plant increase the content of vitamins, miner- height during crop production. Many als or nutraceuticals in horticultural newly introduced ornamental species are products are in progress (Grusak and receiving particular attention via conven- Papaya ringspot virus causes small, darkened Della Penna 1999). The development of tional breeding for dwarf plants because rings on the surface of fruit, as well as foliar Golden Rice with enhanced beta- their natural habits do not fit into market- damage. carotene (pro-vitamin A) in the grain (Ye ing systems requiring compact plants. et al. 2000) demonstrated the potential The manipulation of GA metabolism via for biotechnology to increase nutritional biotechnology has the potential to pro- ian growers to expand their transgen- value. Whether such products will have duce ornamental and flowering plants ic papaya markets and will eliminate sufficient consumer appeal in fully de- with reduced-height phenotypes (Clark et excessive costs associated with segre- veloped markets to drive their commer- al. 2003). The development of grass- gating trans-genic and nontransgenic cialization remains to be seen. es that require significantly less frequent fruits. mowing is another obvious application. , ornamental plants The experience in Hawaii shows Early experiments suggest that expression that transgenic virus resistance is an Since floricultural and ornamental of genes controlling height can be applied excellent approach for controlling vi- plants are grown for aesthetic or other to many plant species. ral diseases in horticultural crops. This nonedible purposes, there may be less Leaf life. Engineering of plants to industry was fortunate to have a po- potential for public concern about GE delay leaf senescence (yellowing) is also tential product already under devel- varieties than there has been with bio- being pursued in ornamental crops. For opment when PRSV was discovered in tech food crops. years, ornamental breeders have selected the main growing area of Puna. There color. Several ornamental new cultivars of plants with more attrac- are many reports that virus-resistant plants, including carnation, rose and tive “stay green” phenotypes. Cytokinins transgenic plants are being developed gerbera, have been engineered for mod- are plant hormones well known to delay in diverse crops, but few have been ified flower color. Research has focused the loss of in ; using commercialized. The papaya story on the manipulation of either anthocya- biotechnology, targeted expression of shows that this approach can provide nins (red and blue colors) or carotenoids genes involved in cytokinin synthesis is a stable and safe option for virus pro- (yellow and orange colors), with the in- now possible. When a gene promoting cy- tection that can be essential for the tent of creating a wider range of flower tokinin biosynthesis is inserted into plants success of specific horticultural crops. colors than occurs naturally, as well as in conjunction with a regulator (promot- to produce natural dyes for industrial er) that turns the gene on only when the purposes (Lu et al. 2003). Florigene is leaf starts to senesce, leaf life is extended selling Transgenic Moon in transgenic plants exposed to drought, series carnations engineered for dark vi- and pathogen stress (Gan and D. Gonsalves is Director, Pacific Basin olet-purple color around the world. The Amasino 1995; Clark et al. 2004). Agricultural Research Center, USDA- varieties are developed in Australia and Ethylene sensitivity. As in fruit ripen- ARS, Hilo, Hawaii. are produced primarily in South ing, manipulation of ethylene synthesis Reference America for marketing in the United or sensitivity has applications in the orna- Gonsalves D. 1998. Control of papaya States and Japan. mental plant industry. Ethylene accelerates ringspot virus in papaya: A case study. Floral scent. Putting the scent back floral and foliar senescence, and chemical Annu Rev Phytopathol 36:415–37. into flowers that have “lost” this trait methods have been developed to miti-

http://CaliforniaAgriculture.ucop.edu • APRIL-JUNE 2004 93 Biotechnology expands pest-management options for horticulture

Leonard Gianessi

ruit and vegetable crops are un- is planted on a small Scott Bauer/USDA-ARS der number of acres and constantF pressure from pests such has reduced use of in- as weeds, viruses, fungi, bacteria, secticide sprays. insects and . If not con- Withdrawn variet- trolled, many of these pests substan- ies. Two transgenic tially lower yields. Successful agri- horticultural variet- cultural production has depended on ies were available for the use of pesticides for 100 years, a short time in the and, yet, losses still occur due to cer- United States but were tain pests that are poorly controlled. withdrawn due to Some crops incur high costs for hir- marketing concerns. ing laborers to weeds because Insect- and virus-resis- Plums resistant to the plum pox virus have been developed by there are no effective herbicides. In tant New Leaf potatoes scientists with the U.S. Department of Agriculture but are not yet addition, new pests routinely arrive were planted on 4% available to growers. for which effective controls have not of the nation’s acreage yet been developed. in 1999 and were credited with reduc- $30 million and replace the use of 4.2 mil- Agricultural researchers continu- ing insecticide use. If the transgenic lion pounds of fumigants. ously seek out new methods to control varieties had not been withdrawn due UC researchers have tested herbicide- pests, including biological agents, to processor resistance they could have tolerant lettuce that could reduce her- new chemicals and plant resistance been planted extensively in the North- bicide use by 140,000 pounds a year. through classical breeding. Biotech- west, reducing insecticide use by 1.4 Herbicide-tolerant strawberries could nology also offers a solution in some million pounds. save Eastern growers several hundred situations where tra- In 1999, the U.S. En- dollars per acre in weed-control costs. ditional methods are If plum pox virus vironmental Protection -resistant pineapple is being ineffective or costly. reaches California, Agency (EPA) granted developed at the University of Hawaii Numerous researchers the transgenic plum Wisconsin sweet-corn to replace 1.4 million pounds of fumi- around the world are growers emergency per- gants. Insect-resistant broccoli developed investigating biotech- could help prevent mission to spray herbi- at Cornell University could improve nological solutions to losses to the state’s cide-tolerant varieties (see yields in years of heavy insect pressure. pest problems of horti- multibillion dollar sidebar, page 110). The Virus-resistant raspberries developed by cultural crops. In 2002, transgenic varieties were U.S. Department of Agriculture (USDA) the National Center for stone-fruit industry. not widely planted due to researchers in the Northwest could help Food and Agricultural marketing concerns and combat bushy dwarf virus, which is pres- Policy released a study of current and growers have not reapplied for the use ent in 80% of Northwest plantings. And potential biotechnological approaches despite continued production losses. transgenic apples resistant to fire blight to pest management in a wide array of Crops currently being tested. Nu- bacteria have been developed and tested crops (Gianessi et al. 2002). merous fruits and vegetables have been at Cornell University; the transgenic va- Current plantings. The study transformed through genetic engineer- rieties would replace the use of antibiot- identified three varieties of trans- ing and are being tested for their poten- ics, which are used to kill the bacteria on genic fruits and vegetables that are tial role in improving pest management. 25% of U.S. apple acreage. currently planted on small acreages For example, University of Florida Emerging pests. Several research in the United States: virus-resistant researchers are testing virus-resistant to- programs are focused on biotechnologi- squash is grown on 5,000 acres in matoes as a substitute for the extensive cal approaches to control emerging pest the Southeast, to prevent late-season insecticide spraying currently utilized to problems. Plum pox virus was detected losses to mosaic viruses; virus- control insects vectoring geminiviruses. in the United States for the first time in resistant papaya is widely planted in In California, herbicide-tolerant process- Pennsylvania, where efforts are under way Hawaii (2,000 acres)(see sidebar, page ing tomatoes have been tested and have to eradicate it by destroying infected trees. 92); and insect-resistant sweet corn the potential to reduce grower costs by USDA researchers have developed a virus-

94 CALIFORNIA AGRICULTURE,AGRICULTURE, VOLUMEVOLUME 5858,, NUMBERNUMBER 22 Monsanto

resistant plum that is being tested in Europe. If plum pox virus reaches Cali- fornia, the trans-genic plum could help prevent losses to the state’s multibillion dollar stone-fruit industry. Pierce’s disease threatens Cali- fornia , and insecticide spraying has occurred to control the Cotton has been genetically engineered to express a protein from a naturally occurring bac- disease carrier, the glassy-winged terium, Bacillus thuringiensis, which is toxic to insect pests such as bollworm and budworm. sharpshooter. A researcher at the This cotton is widely planted in California and elsewhere in the United States. University of Florida (a state where Pierce’s disease has been a problem gate its effects (Sisler and Serek 2003). valuable applications. However, several for 80 years) has transformed grape Ethylene sensitivity can be reduced in fundamental issues inherent to horti- by inserting an antibacterial floriculture crops through applications of cultural crops create significant hurdles protein from another species into the the ethylene antagonist silver thiosulfate (see sidebar, page 84). grape genome. As a result, the trans- (STS), but unfavorable environmental Biological diversity. Simply the di- formed grape plant can destroy the aspects such as metal contamination of versity of crops utilized in horticulture bacteria without the need for insecti- groundwater restrict its commercial use. slows the adoption of new technologies. cide sprays targeting the carrier. Another compound, 1-methylcyclopro- For any given crop, there may be sev- Tristeza virus has killed 45 mil- pene, also blocks the ethylene receptor eral different species and dozens of cul- lion citrus trees in Latin America and protein and makes plant tissues insensi- tivars that are currently marketed, and threatens the Texas citrus industry. tive to ethylene, delaying ripening or the turnover of new cultivars from year Researchers at Texas A&M University senescence. Although this compound to year is tremendous. For example, as have developed and are field testing is effective in many crops, its action de- many as 60 distinct cultivars of iceberg virus-resistant trees. creases with time after treatment as the lettuce alone may be grown throughout Bacterial canker is present in tissues synthesize new ethylene receptor the year as production locations shift Florida citrus orchards, and the state proteins during postharvest transit. By seasonally. Add to this the dozens of ad- is trying to eradicate the disease by expressing a mutant form of the ethylene ditional varieties for romaine, leafy, red destroying infected trees, including receptor protein or by blocking expres- and other specialty types, and it is evi- millions of and backyard sion of components of the ethylene- dent that introducing a new biotech trait citrus trees. A University of Florida signaling pathway, petunia plants with for lettuce requires developing not just researcher has developed and is test- longer lasting floral displays have been one but many new varieties. In perenni- ing a canker-resistant citrus tree. produced (Wilkinson et al. 1997). Unfor- als such as trees and , on the other tunately, negative side effects, such as hand, the choice of a variety is a long- higher susceptibility to fungal pathogens term commitment, making growers cau- and decreased rooting of vegetative cut- tious in selecting novel varieties. L. Gianessi is Director, Crop Protection tings, have limited the commercial use Market acceptance. Currently, the Research Institute, CropLife Founda- of these technologies. The key to effec- largest impediment to adoption of at tion, Washington, D.C. The foundation tive manipulation of ethylene sensitivity least some biotech horticultural prod- is an independent, nonprofit research will be the use of promoters limiting ucts is the lack of market acceptance. organization. transgene expression to the target tissue, Biotech products having documented leading ultimately to plants that have agronomic, economic and environmen- Reference longer lasting flowers with no negative tal advantages have been removed from Gianessi LP, Silvers CS, Sankula S, Carpen- side effects. the market due to the concerns of pro- ter JE. 2002. Plant Biotechnology: Current cessors and distributors about potential and Potential Impact for Improving Pest Hurdles to commercialization Management in U.S. Agriculture; An Analy- consumer rejection. sis of 40 Case Studies. National Center for The lag in commercialization of Intellectual property. Large corpo- Food and Agricultural Policy, Washington, transgenic horticultural crops clearly rations focused on the major agronom- DC. www.ncfap.org/40CaseStudies.htm. is not due to a lack of useful genes or ic crops own the majority of patents on

http://CaliforniaAgriculture.ucop.edu • APRIL-JUNE 2004 95 Transgenic trap crops and rootstocks show potential

John Driver Javier Castillón Abhaya Dandekar

iotechnology may offer unique opportunities for pest control in perennialB tree and crops (Dandekar et al. 2002). Trap crops — plants that an insect pest prefers to the commercial crop — have been tested in a number of agricultural settings, but in most cases have not achieved control levels high enough to completely replace chemical pesticides. Insects are attracted to the trap plant, but they multiply there and Left, apple engineered to silence bacterial genes are resistant to crown gall for- can spread to the adjacent crop. A vari- mation. Right, control (nontransgenic) roots infected with the same bacterial strain show extensive gall proliferation. ant on this concept is to incorporate ex- pression of the Bacillus thuringiensis (Bt) insecticidal protein into the trap plant. press the Bt protein directly (Dandekar bacterium result in the formation of a When the insect feeds on the transgenic et al. 2002), an attractive feature of this gall, an unorganized mass of plant cells trap plant, it dies and the insect popula- scheme is that the walnuts themselves that results from overproduction of two tion is reduced, thereby protecting the are not transgenic and the method plant hormones. The bacterium has the nearby commercial crop. could be used to protect existing or- natural ability to transfer some of its Dry Creek Laboratories of Hugh- chards by interplanting the Bt-express- genes into the host plant’s genome fol- son, Calif., demonstrated this concept ing apple or crabapple trees. Broader lowing infection. The transferred genes with codling moth (Cydia pomonella), application of this approach could code for three specific enzymes. When a major pest of apples, pears, walnuts result in more effective trap crops for a the plant expresses these genes, the and other fruits. The female moth lays number of annual and perennial crops. enzymes synthesize the two hormones eggs on the leaves or fruit, which then Unfortunately, Dry Creek Laboratories that induce the plant to form the tumor, hatch into larvae that burrow into the is unable to move forward at this time or gall, on which the bacteria live. Even- fruits, making them unmarketable. with commercialization of the Bt apple tually, the galls can girdle the stems and sprays and pheromone dis- plants due to the costs associated with reduce the vigor of the tree or vine. A biotechnology When the insect feeds on the transgenic trap plant, it dies and the insect tool called “gene si- population is reduced, thereby protecting the nearby commercial crop. lencing” has been used to generate resistance ruption are generally used to control the regulatory process required for bio- to crown gall. This method involves this pest. However, the female moth tech crops (see page 106). transforming plants with DNA that, prefers to lay its eggs on apple trees. Another opportunity for biotechnol- when expressed, produces signals Under license from Monsanto, Dry ogy in perennial crops that are normally that block the expression of any genes Creek Laboratories developed apple grafted is to engineer only the rootstock with the same sequence as the inserted trees capable of expressing a Bt pro- for desirable traits. Commercial tree cul- DNA. Plants transformed with these tein that was toxic to the codling moth tivars grafted onto transgenic rootstocks interfering versions of the three en- larvae, with the intention of using could benefit from increased rootstock zyme genes would be primed to block these plants as trap crops in and adja- productivity or disease resistance while the function of the corresponding cent to walnut orchards. producing nontransgenic and bacterial genes in infected plants. This A 90-acre field trial was established fruit. For example, such applications would prevent the formation of the in 1997, and in the 4 subsequent years in grapes could offer new solutions to damaging galls without even needing worm damage to the walnuts was Pierce’s disease or Phylloxera by to kill the bacterium itself. The feasibil- almost completely controlled without traditional varieties onto resistant trans- ity of this approach was demonstrated pesticide applications, equivalent to genic rootstocks. The feasibility of this in tomato and Arabidopsis plants (Esco- that in the plots sprayed three times approach was recently demonstrated for bar et al. 2001). Furthermore, both wal- per season with pesticides. While wal- resistance to crown gall disease (Agrobac- nut (see photo; Escobar et al. 2002) and nuts have also been transformed to ex- terium tumefaciens). Infections by the apple (see photo; J. Driver et al., un-

96 CALIFORNIA AGRICULTURE,AGRICULTURE, VOLUMEVOLUME 5858,, NUMBERNUMBER 22 the genes and enabling technologies registration of that herbicide for mil- (such as transformation protocols and lions of acres of field crops. promoters) required for genetically Regulatory requirements. Extensive engineering plants. They are generally safety testing is required for regula- not interested in the smaller horticul- tory approval (deregulation) of biotech tural markets, and may not want to li- crops beyond what is required for va- cense their technologies, depending on rieties bred using traditional methods the impact it could have on their other (see page 106). If the trait has already approved crops (see page 120). been approved in other crops, the costs Post-commercialization. Post- are lower as prior data can be used to commercialization stewardship is also support an application. However, for an increasingly important consider- novel traits likely to be of interest for Crown gall formation was suppressed in ation to technology owners in deciding horticultural crops, the costs could be walnut plants engineered to turn off spe- whether to license their intellectual millions of dollars. For example, by cific bacterial genes. (A) The control property. In Bt crops, for example, some estimates it will cost $20 million exhibits a large, undifferentiated tumor at insect-resistance management pro- to achieve deregulation of Golden Rice 5 weeks after inoculation with a virulent A. tumefaciens strain, while (B) a shoot en- grams must be developed and moni- for humanitarian purposes in six devel- gineered for resistance exhibits no tumor. tored after commercialization. Identity oping countries (I. Potrykus, UC Davis Source: Escobar et al. 2002. preservation programs and segregation seminar, Jan. 22, 2003). Since each trans- of products in the distribution channels genic event (each insertion of a gene in a published results) plants resistant may be required when marketing in genome) must be separately tested and to crown gall have been produced. locations where they are not approved. approved, it is not feasible to transform As most crown gall infections occur Herbicide applications to diverse horti- multiple varieties with a given trait to in the rootstock, nontransgenic sci- cultural crops have the potential to in- amortize the research and technology ons grafted on resistant transgenic crease the Average Daily Intake (ADI) investment across a given crop. Instead, rootstocks would be protected from over the maximum permitted level for a single insertion event is approved for the disease. Rootstock engineering the pesticide active ingredient. (ADI is commercialization and then must be holds great promise for the im- the total residues of a pesticide that a transferred via standard backcrossing to provement of tree and vine crops consumer can be exposed to, consider- other varieties. This is highly inefficient by preserving the horticultural ing all sources; the government sets and often makes it difficult to regain characteristics of existing varieties limits for each pesticide.) An agro- the unique properties of all the diverse used as scions while incorporating chemical company will not approve the varieties. Public-private partnerships beneficial traits into the rootstocks. use of its herbicide-resistance trait in a are one way to reduce the costs of com- small acreage crop if it endangers the mercialization (see page 116). The IR-4

J. Driver is former President and Syngenta J. Castillón is Director of Research, Dry Creek Laboratories, Hughson, Calif.; and A. Dandekar is Professor, Depart- ment of , UC Davis. References Dandekar AM, Fisk HJ, McGranahan GH, et al. 2002. Different genes for dif- ferent folks in tree crops: What works and what does not. Hort Sci 37:281–6. Escobar MA, Civerolo EL, Summerfelt KR, Dandekar AM. 2001. RNAi-mediated oncogene silencing confers resistance to crown gall tumorigenesis. Proc Natl Acad Sci USA 98:13437–42. Escobar MA, Leslie CA, McGranahan GH, Dandekar AM. 2002. Silencing crown gall disease in walnut (Juglans regia L.). Plant Sci 163:591–7. “Golden Rice” has been genetically engineered to produce beta-carotene, the precursor to vitamin A. However, for a variety of reasons it is not yet available to farmers in developing countries, where vitamin A deficiencies are common.

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