U.S. Department of Agriculture Agricultural Research Service July 2012

Studying the Syndrome of COLONY COLLAPSE

page 4

Agricultural Research Service‡6ROYLQJ3UREOHPVIRUWKH*URZLQJ:RUOG ostharvest control of insect pests The dried fruits and tree nuts industry tion until they drown, and that means that is an integral part of maintaining also requires insect pest control measures. Asian citrus psyllids will very likely not Psafe, high-quality, abundant produce Much of the U.S. production of dried fruits be present in commercially packed fruit. domestically and for export. Insect pests and tree nuts occurs in the Central Valley This research will help maintain access can travel with fresh produce from the of California. To provide consumers with of California citrus to Australia, a market field and into processing plants and storage quality produce, processors must control valued at $60 million annually. facilities. Agricultural Research Service postharvest insect pests. Scientists in Parli- Also on the world stage, tephritid fruit scientists have found many innovativeand er, California, are developing nonchemi- flies are serious economic pests worldwide. environmentally friendly methods to help cal treatments to solve this issue, using The larval stages feed within host fruits, meet this challenge. temperature extremes or vacuum, as well making infestation difficult to detect. Fruit ARS laboratories in Manhattan, Kansas, as insect parasitoids as control agents. The imported into theUnited States is currently and Gainesville, Florida, in collaboration target pests for most of this work include checked for infestation by cutting open with others, are using unique ways to both field pests of postharvest significance a small sample of fruit and looking for detect and disinfest stored-grain facilities and stored-product pests. Several methods fly larvae. This is a time-consuming and of insect pests like the red flour beetle, a are being tested, including cold storage for potentially limited way to detect the insect, major pest of the flour-milling industry. spotted wing Drosophila on grapes, radio so more sensitive screening methods are Effective pest detection and monitoring frequency treatments for cowpea weevil in needed. USDA-ARS scientists in Miami, are critical to pest management because dried pulses, and low-temperature vacuum Florida, in conjunction with USDA’s they provide necessary information for treatments for codling moth in fresh fruits. Animal and Plant Health Inspection timing and targeting the application of Another insect, the Indianmeal moth, is Service, also in Miami, conducted research control measures. globally the most important stored-product to evaluate gas chromatography (GC) as Researchers in Gainesville have devel- moth pest. ARS scientists in Parlier, along a detection method. Grapefruits infested oped anew trap for monitoring thepresence with collaborators from the University with larvae of the Caribbean fruit fly were of red flour beetles. The trap uses ultraviolet of Michigan, demonstrated that mating examined to determine whether infested light, a chemical attractant, and a physical disruption controls Indianmeal moth in fruit emitted chemicals distinct from those configuration that guides beetles into a pit dried beans in central California. Mating of healthy fruit. GC analysis indicated that where they become trapped. This monitor- disruption is safer and less disruptive than there were volatile chemicals indicative ing tool has promise for use in flour mills fumigants and aerosol sprays, but it is less of citrus fruit injury and others associated and other food-processing plants. widely used for control of Indianmeal with larval infestation. These “signature In Manhattan, Kansas, ARS scientists moth. This demonstration of successful chemicals” were also detectable with a are trying to slow the growth of red flour control of the Indianmeal moth will encour- portable, ultra-fast GC analyzer. This beetle populations by altering its hormonal age adoption of mating disruption, thereby is just one example of studies that hold system. Treatment with growth regulators protecting food in distribution channels the potential in development of a rapid can kill immature insects, and survivors while reducing use of insecticides and screening protocol for detection of infested may have reduced fitness. When larvae improving worker safety. fruit at U.S. ports of entry. were exposed to growth regulators, males The possible presence of the Asian The article on oxygenated phosphine were less likely to survive to the adult citrus psyllid in loads of citrus arriving fumigation, on page 10 of this issue, is stage than females. Adult males that sur- in Australia threatens California’s ability another example of ARS research on ways vived larval exposure to growth regulators to export citrus into that market. Parlier to make U.S. produce as freeofinsect pests produced fewer offspring than unexposed researchers found that Asian citrus psyl- as possible for the consumer. males. Results show that the nonlethal lids are completely washed from fruits that are submerged, flooded, or sprayed effects of growth regulators potentially Daniel Strickman increase the impact of insecticide treat- at high temperatures using soak tanks and wash lines consistent with commercial ARS National Program Co-Leader ments on population growth rates, thus Methyl Bromide Alternatives practices in California. Nearly 99 percent making growth regulators more effectivein Beltsville, Maryland pest management than initially estimated. of the insects remain trapped by the solu-

2 Agricultural Research O July 2012 July 2012 Vol. 60, No. 6 ISSN 0002-161X Solving Problems for the Growing World

(D2549-1) Agricultural Research is published 10 times a year by the Agricultural Research Service, U.S. Department The root-knot nema- of Agriculture (USDA). The Secretary of Agriculture tode can penetrate the has determined that this periodical is necessary in the transaction of public business required by law. roots of some crops Tom Vilsack, Secretary and rob plant nutrients. U.S. Department of Agriculture ARS scientists and Catherine E. Woteki, Under Secretary colleagues have Research, Education, and Economics released a resistant Edward B. Knipling, Administrator Agricultural Research Service cotton line to help Sandy Miller Hays, Director breeders fight the pest. Information Staff Story begins on Editor: Robert Sowers (301) 504-1651 Associate Editor: Sue Kendall (301) 504-1623 page 16. Art Director: BA Allen (301) 504-1669 Photo Editor: Tara Weaver-Missick (301) 504-1663 Staff Photographers: Peggy Greb (301) 504-1620 Photo by WILLIAM WERGIN and RICHARD SAYRE. Colorized by STEPHEN AUSMUS. Stephen Ausmus (301) 504-1607 Most information in this magazine is public property and may be reprinted without permission (except where copyright is noted). Non -copyrighted articles and high-  Colony Collapse Disorder: An Incomplete Puzzle resolution digital photos are available at ars.usda.gov/ar. Paid subscriptions are available from the U.S. Cracking Down on Poultry Disease with Egg Yolk Government Printing Office (Superintendent of Documents). See back cover for ordering infor mation. Oxygenated Phosphine Fumigation for Pest Complimentary 1-year subscriptions are available directly from ARS to public libraries, schools, USDA employees, Control on Harvested Fruits and Vegetables and the news media. Call (301) 504-1638 or e-mail [email protected].  Getting Your Kids To Eat More Vegetables: Scientists This magazine may report research involving pesticides. It does not contain recommendations for their use, Scrutinize “Parenting Practices” nor does it imply that uses discussed herein have been registered. All uses of pesticides must be registered by  Beneficial Yeast: A New Weapon for Keeping Tree appropriate state and/or federal agencies before they can be recommended. Nuts Safe to Eat Reference to any commercial product or service is made with the understanding that no discri mination is intended  The Search for Nematode-Resistant Cotton and no endorsement by USDA is implied. The USDA prohibits discrimination in all its programs  No-Till Crops Can Improve Air Quality and activities on the basis of race, color, national origin, age, disability, and where applicable, sex, marital in the Pacific Northwest status, familial status, parental status, religion, sexual orientation, genetic information, political beliefs,  Searching for Genes To Protect reprisal, or because all or part of an individual’s income is derived from any public assistance program. (Not Soybeans From Flooding and Diseases allprohibited bases apply to all programs.) Persons with disabilities who require alternative means for  Locations Featured in This Magazine Issue communication of program information (Braille, large print, audiotape, etc.) should contact USDA’s TARGET Center at (202) 720-2600 (voice and TDD). To file a complaint of discrimination, write to USDA, Director, Cover: A honey bee being inoculated with 1RVHPD to determine bee infection rates and Office of Civil Rights, 1400 Independence Avenue, S.W., immune responses. ARS scientists and others have been working for years to try to Washington, D.C. 20250-9410, or call (800) 795-3272 solve the puzzling honey bee syndrome known as “colony collapse disorder.” The story (voice) or (202) 720-6382 (TDD). USDA is an equal that begins on page 4 examines some of the potential causes and rules out others. opportunity provider and employer. Photo by Stephen Ausmus. (D2484-7)

Agricultural Research O July 2012 3 Colony Collapse Disorder An Incomplete Puzzle

hen it comes to solving For the last 3 years, self-reported the puzzling syndrome surveys of beekeepers have found Wknown as “colony collapse that winter colony losses have aver- disorder” (CCD), which has been at- aged about 30 percent, with around tacking honey bee colonies since 2006, losses of 30 one-third of those losses ascribed to the best that can be said is that there is to 90 percent of the CCD, according to Jeff Pettis, research good news and bad news. The good news hives in their apiaries with no apparent leader of the Bee Research Laboratory in is that the rate of honey bee losses seems cause. The defining characteristic of CCD Beltsville, Maryland, who heads up the to have leveled off rather than continuing is the disappearance of most, if not all, of Agricultural Research Service’s CCD to increase. The bad news is that the cause the adult honey bees in a colony, leaving research effort. This compares to colony or causes of CCD remain unclear. behind honey and brood but no dead bee losses that were averaging 15-20 percent In the United States, the problem sur- bodies. This definition has recently been before CCD. faced in October 2006, when an increasing revised to include low levels of Varroamite “The faint good news in the survey number of beekeepers began reporting and other pathogens, such as Nosema, as numbers is that the CCD problem does probable contributing factors.

Entomologist Jeff Pettis inspects honey bee combs at Beltsville, Maryland, for disease. Honey bees are disappearing at an alarming rate. ARS researchers have been working diligently to solve the mysterious syndrome known as “colony collapse disorder.”

STEPHEN AUSMUS (D2489-12)

4 Agricultural Research O July 2012 STEPHEN AUSMUS (D2482-20) not seem to be getting worse,”Pettis stressors. The researchers have been says. “But—and this is a big ‘but’— analyzing samples from healthy and 33-percent losses each year are prob- CCD-struck colonies and applying a ably not economically sustainable for variety of stressors from the four groups commercial beekeeping operations.” to colonies in hopes of provoking a While many possible causes for CCD colony response that duplicates CCD. have been proposed, reported, and dis- “While a number of potential causes cussed—both in the scientific literature have been championed by a variety of and popular media—no cause has been researchers and interest groups, none proven. (See sidebar, page 7.) of the causes has stood up to detailed “We know more now than we did scrutiny. Every time someone has a few years ago, but CCD has really proclaimed a potential smoking gun, been a 1,000-piece jigsaw puzzle, and further investigation has not been able the best I can say is that a lot of pieces to make the leap from correlation to have been turned over. The problem is cause-and-effect for one reason or that they have almost all been blue-sky another. Other times, there hasn’t pieces—frame but no center picture,” even been a scientific correlation,” Pettis explains. Pettis says. The beelab’s scientists have been ARS’s research work, along with looking for the cause or causes of that of university and other scientists, CCD within four broad categories: “pretty well supports the idea that CCD is caused by multiple factors—possibly pathogens; parasites, such as Varroa In Beltsville, Maryland, ARS entomologist Jay Evans mites or Nosema; environmental inspects a comb of honey bees for signs of mites and working individually, but more likely stressors, such as pesticides or lack brood disease. in combination,”Pettis adds. “But we of nectar diversity; and management still can’t say whether it’s the same set

Entomologist Jay Evans (left) and postdoctoral research associate Ryan Schwarz use a microscope to look at spores of the honey bee fungal parasite 1RVHPDFHUDQDH which can replicate in cells lining the honey bee gut.

STEPHEN AUSMUS (D2483-5)

Agricultural Research O July 2012 5 STEPHEN AUSMUS (D2487-3)

ARS scientists are studying the transfer of food or other fluids among members of a bee community through mouth-to-mouth feeding as a possible facilitator of colony collapse disorder. of factors in every CCD incident or—if it exposure to either pesticides or disease, Finding Varroa mites to be a good is the same factors—that they are occur- respectively, there was no significant predictor of winter declines in this study ring in the same sequence in every case.” difference between CCD and non-CCD was not really surprising, according to Pathogens continue to stand out as one colonies. Evans, though other studies in the United of the important puzzle pieces, according The team did find considerable differ- States have not found as high a correlation to ARS entomologist Jay Evans, also with ences between CCD hives on the west coast between CCD and Varroa mites. the Bee Research Laboratory. He was part and the east coast. “Finding Kashmir bee “Even if Varroa mites themselves do of a team that used genetic analyses to virus in a hive was the best predictor of not directly cause CCD, we know they can look for correlations between bee health CCD in the western United States, while transmit multiple viruses to honey bees,” and bee pathogens and activity levels of deformed wing virus, an unrelated RNA Evans says, “and higher total pathogen honey bee genes. virus, was a better predictor in the East,” levels (rather than infection by any specific Two picornalike viruses—acute bee Evans says. pathogen) have been our best correlation paralysis virus and Kashmir bee virus— Evans and Bee Research Laboratory with CCD so far. Butshould the pathogens along with deformed wing virus, black colleague Judy Chen were also part of an be considered primary casual agents— queen cell virus, and two species of No- international team that closely followed multiple pathogens specifically causing sema, were found to be more abundant in 29 European honey bee colonies, carefully CCD—or are they indirectly involved CCD hives. Infection by multiple picorna- monitoring for pathogens, parasites, and because they weaken bees, making these like viruses could result in honey bees hav- bee proteins. This study found that four bees more vulnerable to something else ing reduced abilities to synthesize certain factors appeared to be the best predictors we don’t know yet?” proteins, the lack of which would leave the so far of winter honey bee loss: presence Pesticides as Cause? bees more vulnerable to additional stresses of the microsporidian Nosema ceranae; Pesticides—individually and in like pesticides, nutrition problems, or other levels of the protein vitellogenin, which general—have been repeatedly nominated pathogens—which sounds like a possible strongly reflects the bee’s protein status and as a cause of CCD, often without direct root cause of CCD, Evans points out. plays an immune system role; Varroa mite scientific data to support the idea. In When the researchers looked at the infestation; and the presence of deformed a pesticide survey conducted by U.S. bees’ turning on of detoxification and im- wing virus, a virus often associated with Department of Agriculture and university mune genes, which would have reflected Varroa mites. scientists that analyzed wax, pollen, and

6 Agricultural Research O July 2012 Many Suspects, But No Culprit

Transportation stresses from migra- apiaries of beekeepers who do not has been a subject of interest in sci- tory beekeeping feed HFCS. Others have suggested entific journals and the popular media Pollination-service beekeepers stack a possible connection with HFCS since the syndrome first appeared in colonies on tractor-trailers and trans- produced from genetically modified 2006. Despite numerous and differ- port them thousands of miles during corn, combining two popular villains. ing claims, nothing has actually been the growing season. For honey bees, But the simple management change shown to be the cause of the problem. orientation to their hive is vital, and of not feeding any HFCS does not being regularly relocated must be Pathogens stop CCD. stressful. Additionally, moving hives One or more pathogens remain scien- Global climate change tists’ most likely choice as the cause around the country may spread dis- Weather changes, such as unusu- or partial cause. But various viruses eases and pathogens as honey bees ally warm winters, earlier springs, and bacteria have had higher correla- intermingle in the fields. It is possible drought, and flooding, can lead to tions with CCD in different parts of the that such stresses play into CCD, but changes in flowering times. Plants United States and in different coun- there is no scientific evidence of it at may blossom early, limiting nectar tries. Before any pathogen can be this time. and pollen supplies. But bees used legitimately accepted as the cause, Monoculture for pollination contracts are moved science must demonstrate that when Wild honey bees forage on a wide to fields to coincide with flowering it is introduced into a healthy colony, variety of nectar sources. Honey bees of crops. Still, some believe global CCD results. used for commercial pollination are warming is to blame, if only in part, mostly limited to one crop at a time, Parasites for CCD. and it is possible that they may suf- A parasite is the other perennial sus- Ozone pect, either by itself or in combination fer nutritional deficiencies that stress The level of the air pollutant ozone with one or more pathogens. 1RVHPD their immune systems. has been steadily dropping since and 9DUURD mites remain high on the Genetically modified crops the early 1990s. Since CCD did not probable-cause list. Genetically modified (GM) crops, appear until 2006, the timing doesn’t most commonly %W corn, have been New pests or diseases match for ozone to be related. offered up as the cause of CCD. But Some believe that a previously un- Cell phones and cell phone towers discovered or unidentified pest or there is no correlation between where The idea of cell phones causing CCD pathogen is involved in CCD. But GM crops are planted and the pattern began with the misinterpretation of claims that such an agent has been of CCD incidents. Also, GM crops a study in which a cordless home identified have not held up scientifi- have been widely planted since the phone, not a cell phone, was shown cally so far. late 1990s, but CCD did not appear to have some impact on honey bee until 2006. In addition, CCD has been navigation. The study author has Pesticides reported in countries that do not al- There are many classes of pesticides repeatedly stated that the phone he low GM crops to be planted, such as tested is nothing like a cell phone and to which honey bees can become Switzerland. German researchers exposed. Among those that have has nothing to do with CCD. But the have noted in one study a possible idea remains popular. One of the most been stamped with a “CCD cause” correlation between exposure to %W label are the neonicotinoids, like recent “proofs,” (published in &XUUHQW pollen and compromised immunity 6FLHQFH in 2010) claimed evidence imidacloprid and clothianidin. One to 1RVHPD. issue with making that link is the suggesting “that colony collapse does lack of amatching pattern between High-fructose corn syrup occur as a result of exposure to cell neonicotinoid residues in colonies Some researchers have attributed phone radiations”while also reporting and CCD outbreaks. France, which CCD to the practice of feeding high- that the impact of cell phones in both banned imidacloprid in 1999, and fructose corn syrup (HFCS) to supple- of the test hives resulted in more bees Germany, which along with France ment bee colonies. But there are staying in the hive longer—the exact banned clothianidin in 2008, still have many reports of CCD occurring in the opposite of the definition of CCD.* CCD problems.

Agricultural Research O July 2012 7 bee samples for the presence of 121 of crops and ornamentals from many dif- To better account for such sublethal different pesticides or their metabolites, ferent insects. The dosages used in the impacts, Pettis is working as part of an the most commonly found pesticides study were intentionally well below the international group of scientists and were fluvalinate and coumaphos. While levels that have been documented to kill regulators to help the U.S. Environmental about 60 percent of the 259 wax and 350 honey bees after short-term exposure and Protection Agency (EPA) develop pollen samples did show the presence of reflected levels that have been measured recommendations for pesticide-testing at least one systemic pesticide, almost all in the environment. guidelines that factor sublethal effects into were found at levels well below what is After the third generation, newly test protocols. ARS scientists have already considered lethal to honey bees. emerged adult bees from these colonies developed a sublethal-impact cage assay There was no overall pattern of exposure were exposed to spores of N. apis and that has been provided to EPA. among the samples for a specific pesticide N. ceranae, gut parasites that have been Could imidacloprid and Nosema to- or class of pesticides. The study did not a growing problem for U.S. beekeepers gether be the cause of CCD, as some claim? look specifically at the pesticides as they since the 1990s. “This study did not look for nor establish might be related to CCD, but if a specific There was up to a fourfold increase in the any connection between either imidaclo- class of pesticides were involved, a pattern levels of Nosema in honey bees from the prid or Nosemaand CCD,”Pettis explains. of residues should have been discernable, imidacloprid-exposed colonies, regardless “But the effect of the combination of explains Pettis, who co-led the study. of whether 5 or 20 ppb were fed. imidacloprid and Nosema demonstrates Not all pesticide impact is about directly “While these increased Nosema lev- that there are many complex interactions killing honey bees, however. Sublethal els were found in individual bees, there between stress factors that need to be con- doses of the pesticide imidacloprid—one was no measurable impact at the colony sidered in looking for a cause of CCD and of the neonicotinoid group of pesticides— level,”Pettis says. “Imidacloprid was high honey bee mortality in general.”—By were found to make honey bees more chosen for this study because of its J. Kim Kaplan, ARS. susceptible to the gut parasite Nosema, widespread use and beekeepers’ con- This research is part of Crop Production, according to a study by Pettis and Uni- cerns about it. But it was only found in an ARS national program (#305) described versity of Maryland researchers Dennis 3 percent of the pollen samples at www.nps.ars.usda.gov. vanEngelsdorp, Josephine Johnson, and checked in the pesticide-survey study, To reach scientists mentioned in this Galen Dively. usually at very low levels, and no article, contact Kim Kaplan, USDA-ARS The researchers fed three generations connection with CCD has ever Information Staff, 5601 Sunnyside Ave., of honey bee colonies either 5 or 20 been made scientifically,” Beltsville, MD 20705-5128; (301) 504- parts per billion (ppb) of imidacloprid, he adds. 1637, [email protected]. which is used to protect a wide variety *

A honey bee, with pollen attached to its hind leg, pollinating a watermelon flower.

STEPHEN AUSMUS (D2368-1)

8 Agricultural Research O July 2012 t’s not mother’s milk, but egg yolk may be the closest remedy for boosting Ithe immune system of newly hatched chickens against infectious diseases such as coccidiosis. Amajor diseaseof chickens, coccidiosis is caused by intestinal parasites—single- celled protozoans in the genus Eimeria. Disease-affected birds are unable to absorb feed or gain weight, costing thepoultry industry more than $600 million annually in the United States and $3 billion worldwide. Scientists at the Agricultural Research Service Henry A. Wallace Beltsville [Maryland] Agricultural Research Center (BARC) and collaborators from different universities and the Mexican company ARS molecular IASA (Investigacíon Aplicada, S.A.) have biologist Sung Hyen Lee (left) and visiting developed a novel, antibiotic-free method scientist Seung Ik that uses hyperimmune egg yolk antibodies Jang prepare live to control intestinal poultry diseases. coccidia to test a “Coccidiosis is associated with other chick’s immunity. The chick should be pathogens, such as the one that causes immune because it necrotic enteritis—a prevalent gut disease previously consumed of poultry,” says avian immunologist Hyun hyperimmune egg Lillehoj, who works in BARC’s Animal yolk antibodies from Parasitic Diseases Laboratory. “By control- immune chickens.

ling one, you’re also reducing the impact D2494-2) of the other.” B ( G RE Good management practices and live vac- cinations reduce the spread of coccidiosis, P EGGY but alternative strategies are needed to help the disease. Egg yolk is spray dried, mixed significant increase in body weight gain, controldrug-resistantstrains and to enhance with feed, and given to chicks that have reduced fecal Eimeria shedding, and fewer organic farming for the poultry industry. no immune protection right after hatching. gut lesions compared to control birds fed a Generally, a host can develop two types Lillehoj teamed up with ARSvisiting nonsupplemented diet. of immunity—active and passive—to resist scientist Sung Hyen Lee from the Rural De- “It’s very simple technology, and it infection. Passive immunity allowsimmune velopment Administration in South Korea, works,”Lillehoj says. molecules that are already formed to be IASA scientist Eduardo Lucio, and other transferred from the hen, viathe yolk, to the Based on these results, one company has researchers to conduct different experiments chick. Active immunity relies on vaccines developed a commercial product that can to demonstrate the efficacy of inducing pas- to build immunity in the birds. be fed to chickens to control coccidiosis. sive immunity against coccidiosis. Similartechnology maybe usedin thefuture “When chicks hatch, they have no im- One-day-old broiler chickens were con- to guard against other devastating poultry munity to this pathogen. But if we give tinuously fed a standard diet containing a diseases.—By Sandra Avant, ARS. preformed immune proteins to 1-day-old commercially available egg yolk powder progeny, they are ready to fight infection,” This research is part of Animal Health, prepared from hens hyperimmunized with she says. “It’s similar to how immunity is an ARS national program (#103) described multiple species of Eimeria. They were passed to newborns through milk.” at www.nps.ars.usda.gov. then given a challenge infection with live Hyun Lillehoj is in theUSDA-ARS Ani- The method involvesextracting antibodies coccidia. Body weight gain between days from yolks of eggs from pathogen-free birds mal Parasitic Diseases Laboratory, 10300 0 and 10 and fecal shedding between days Baltimore Ave., Bldg. 1043, BARC-East, that have been hyperimmunized, meaning 5 and 10 postinfection were analyzed. they possess greater-than-normal immunity Beltsville, MD 20705-2350; (301) 504- Chickens given 0.5 percent or less of the 6170, [email protected]. due to an abundance of antibodies against hyperimmune egg yolk antibodies had a *

Agricultural Research O July 2012 9 hipments of U.S. fresh fruits and Entomologist Yong-Biao Liu, at the U.S. enhances phosphine toxicity against in- vegetables can face obstacles in Agricultural Research Station in Salinas, sects. Liu calls the treatment “oxygenated Sexport to overseas markets if they California, is working on low-temperature phosphine fumigation.” harbor unwanted pests. There are few fumigation with phosphine as an alterna- In a series of experiments, Liu tested workable options to kill the pests, and tive to methyl bromide for control of pests phosphine fumigation under high levels of methyl bromide fumigation is the most on harvested fresh fruits and vegetables. oxygen against four insects and at different common one. But methyl bromide destroys Phosphine has been used for more than life stages: western flower thrips adults and atmospheric ozone, and its production 80 years as a fumigant to control stored- larvae, leafminer pupae, grape mealybug is being phased out globally. Therefore, product pests. It acts slowly against insects. eggs, and Indianmeal moth eggs and pupae. methyl bromide fumigation is unsustain- Many insects, especially at egg and pupal Liu says that the four species represent a able—as well as expensive. In addition, stages, are very tolerant of phosphine, and range of insect types and life stages for methyl bromide fumigation injures some it may take more than 10 days of fumiga- which quarantine treatments are needed. fresh produce, such as lettuce. Scientists tion treatment to control them. In working “Itis important to test the fumigation are searching for alternative ways to solve on fumigation with pure phosphine at on eggs and pupae because, in general, postharvest pest problems on exported low temperatures, Liu found that oxygen eggs and pupae are more tolerant of fresh products. Directly below: Lettuce aphids, 1DVRQRYLDULELVQLJUL infesting a lettuce At the Crop Improvement and Protection Research Unit in Salinas, leaf. Bottom: Close-up of alettuce aphid. This pest (about 1-3 millimeters California, entomologist Yong-Biao Liu injects fumigant samples into a long) appeared in California’s Salinas Valley in 1998 and is now found in gas chromatograph to determine phosphine concentrations. all lettuce-production areas of that state and Arizona.

YONG-BIAO LIU (D2534-1)

TIFFANY MASUDA (D2532-1) STEPHEN AUSMUS (D689-7)

10 Agricultural Research O July 2012 phosphine than larvae and adults,” says phosphine fumigation treatments for phosphinefumigation makes thetreatment Liu. “If we can control insects at the egg pest control. The magnitude of toxicity less phytotoxic than regular phosphine and immaturestages, wecan besureadults enhancement will likely vary depending fumigation. will also be controlled. These insects eat, on the insect species and their life stage.” Fumigating under high oxygen is also and thus damage, produce while they are In a follow-up study for control of lettuce safer to conduct than regular phosphine fu- in the larval stage, but all stages require aphid on lettuce, Liu demonstrated that migation. Used alone, phosphine will burn quarantine treatment.” oxygenated phosphine fumigation is not in the air if its concentration exceeds 1.8 The small-scale study demonstrated only more effective against the pest but also percent. Past research showed that oxygen that in 5-hour fumigations with 1,000 less toxic to plants than regular phosphine will suppress phosphine ignition and thus ppm (parts per million) phosphine at 5˚C, fumigation. At a low temperature of 3°C, reduce risk of fire. The shorter treatment control of western flower thrips on lettuce regular phosphine fumigation took 3 days time and lower phosphine concentration increased from 80 percent to 98 percent at more than 2,000 ppm concentration to used in oxygenated phosphine fumigation when oxygen was increased from 21 per- control lettuce aphid, and the treatment also reduce fire risk. Liu says that this is cent to 40 percent. When the oxygen level resulted in significantly higher percent- particularly true for fresh products as they was increased to 80 percent, mortality of ages of both romaine and head lettuce are fumigated under low temperature and western flower thrips reached 99 percent. having injuries than the 2-day oxygen- high humidity conditions. Western flower thrips are a common pest ated phosphine fumigation at 1,000 ppm Even though oxygenated phosphine of fruits and vegetables in the United States concentration under 60 percent oxygen, fumigation is more complex and expen- and are often found on fresh products ex- which also controlled lettuce aphid. Liu sive than regular phosphine fumigation, ported to Taiwan, where it is a quarantined says that the shorter treatment combined Liu is optimistic that its benefits in terms pest. Currently, fresh fruits and vegetables with lower phosphinelevels in oxygenated of reduced treatment time and enhanced exported to Taiwan arefumigated ability to control tolerant pests with methyl bromide to control An insulated, covered pallet used in fumigation trials to control will more than compensate for western flower thrips. western flower thrips on head lettuce. The trials were conducted to the added costs and complexity. test phosphine fumigation under high levels of oxygen against insects Liu expects that the oxygenated Leafminer pupae control was at different life stages. also improved when oxygen was phosphine fumigation he devel- introduced: At normal oxygen oped will have an impact on the levels, more than 70 percent sur- phosphine fumigation industry, vived, but only 16 percent were because many insects that cannot still alive at 40 percent oxygen, be controlled by regular phos- and just 1 percent survived at 80 phine fumigation in a reasonable percent oxygen. This fumigation time frame can now be controlled treatment was conducted over effectively with the new method. 24 hours and contained 500 ppm The research studies described of phosphine at 5˚C. Complete in this story were published control of leafminer pupae was between 2008 and 2012 in achieved in a 24-hour fumigation the Journal of Asia-Pacific with 1,000 ppm of phosphine at Entomology and the Journal of 5˚C under 60 percent oxygen. Economic Entomology.—By “Increased oxygen levels also Sharon Durham, ARS. resulted in significantly lower This research is part of Crop survival rates of Indianmeal moth Protection and Quarantine (#304) pupae in response to 24-hour fu- and Methyl Bromide Alterna- migationswith 500 and 1,000 ppm tives (#308), two ARS national phosphine at 10˚C, and complete programs described at www.nps. control was achieved at 1,000 ppm ars.usda.gov. phosphine at 60 percent oxygen,” Yong-Biao Liu is in the USDA- says Liu. ARS Crop Improvement and “Oxygen enhanced the phos- Protection Unit, U.S. Agricultural phine toxicity significantly for Research Station, 1636 E. Alisal all life stages tested. This has St., Salinas, CA 93905; (831) not been reported previously, and 755-2825, y ongbiao.liu@ars. the findings have potential to be usda.gov. used to develop more effective *

YONG-BIAO LIU (D2533-1)

Agricultural Research O July 2012 11 Getting Your Kids To Eat More Ve

(D2538-1) Maybe you’re one of those Service’s Children’s Nutrition Research Center (CNRC) in Houston, Texas, and a lucky parents whose kids professor of pediatrics at Baylor College already love vegetables. of Medicine, also in Houston. The college operates the nutrition center in cooperation If you’re not, you’re among legions of with ARS. moms, dads, grandparents, and others who “Kiddio: Food Fight!”—the lively, know that some of the youngsters in their upbeat video game that Baranowski’s lives should eat more of these good-for-you team is creating, will offer users a series foods, but don’t. of short, interactive episodes that they can Help may be on the way. play on their smart phone. The engaging, And it’s coming from a perhaps unlikely fast-paced game features “Kiddio,”an source: Your smart phone. appealing preschooler who doesn’t like vegetables. “We’re creating a fun, science-based video gamethatgives parents of preschool- Each episode will give users several ers a quick, easy way to learn some of the choices of what to do to improve the balky best approaches for getting their kids to eat youngster’s eating behaviors. Importantly, more veggies,”says Tom Baranowski, a parents can customize the game so that psychologist at the Agricultural Research Kiddio’s temperament matches that of their child. “That way, whatparents learn can help them reshape their own child’s eating hab- its,”says Baranowski. “We want the game to “Kiddio,”an appealing character who doesn’t like vegetables, stars in a fun, science-based video game that helps be relevant to the real- parents learn some of the best approaches for getting their preschool kids to eat more veggies. Kiddio’ whether to take a bite, or say something like “Yuk!” are based on what researchers have learned about kids’ world food-choice is- to parental tactics. Images courtesy of ARCHIMAGE, Inc. sues of their household.” In the course of each Some of these options, says Baranowski, episode, parents will be “create effective, ‘teachable moments,’ such able to select—with a as when the parent says, ‘That’s a really tasty quick touch on thesmart- veggie.’Other options may express a perhaps- phone screen—multiple ineffective, ‘firm discipline’ approach in which options for influencing the parent tells Kiddio, ‘You will taste it before Kiddio. For example, you leave the table!’ after deciding whether “Each of the options is based on a parenting to offer Kiddio a serv- practice that we’ve studied in our research. And ing of broccoli, carrots, Kiddio’s responses to these options—whether to corn, or peas, players next take a bite or to say something like ‘Yuk!’—are select what to say to him based on what we’ve learned so far about kids’ Parents who want their kids to eat more fru to increase the chances reactions to these parental tactics.” ma its and vegetables y involve the youngsters in helping to that he will at least taste supermarket produce sect select items from the By working their way through the various ion. the veggie. options, “parents can learn which tactics USDA-FNS (D2535-1)

12 Agricultural Research O July 2012 eVegetables

(D2538-2) The video-game project, funded by ARS American Journal of Preventive Medicine and a grant from the National Institute of and the Journal of Diabetes Science and Child Health and Human Development, Technology. will draw upon five studies that the Houston Survey Reveals Some Parenting scientists have conducted over the past Practices decade. These investigations, involving Among the studies that are helping shape thousands of parents, kids, and nutrition- the new “Kiddio”series is an investigation related professionals, are examples of that drew upon the real-life experiences of “behavioral nutrition,” a comparatively more than 700 Alabama and Texas parents new scientific discipline that has roots in and their preschool-aged children. “Kid- both psychology and nutrition. dio”collaborator Teresia O’Connor, M.D., The field is “all about exploring—and an assistant professor of pediatrics at the explaining—the internal and external CNRC and at Baylor College of Medicine, factors that influence our food choices,” led this study, analyzing—from adiffer- says Baranowski. His work, and that of ent perspective—data collected as part of his Houston coinvestigators, has helped an earlier, larger investigation headed by make the CNRC an international leader CNRC colleague Theresa Nicklas. in behavioral-nutrition research geared to Unlike some previous studies, this understanding—and helping solve—the one didn’t focus on just one category of most urgent nutrition-related problems of America’s children and adolescents. How do veggies fit “Kiddio,”an appealing character who doesn’t like vegetables, stars in a fun, science-based video game that helps into this picture? parents learn some of the best approaches for getting their preschool kids to eat more veggies. Kiddio’s responses— are based on what researchers have learned about kids’ reactions Increased vegetable ARCHIMAGE, Inc. consumption helps kids get the recommended succeed,”says Baranowski. “The point is to amounts of several give them a safe, low-risk, nonthreatening way vitamins and minerals to sharpen their parenting skills and to boost and is thought by some their confidence in their decisions. experts to help reduce “We plan to make the episodes increasingly the risk of chronic dis- difficult, so players won’t become bored or eases such as diabetes, complacent. We hope parents will want to play heart disease, and some each episode several times, and that they’ll cancers. learn something new every time.” Peer-reviewed articles Baranowski says that by limiting each by Baranowski and col- episode to just a few minutes, the team will leagues about the use of “make it convenient for on-the-go parents to video games to improve home environment where kids are play and learn in spare moments, such as when kids’ eating habits have Proactive actions, such as creating a see a parent fruits and vegetables and to likely to see and be served effective they’re waiting for their kids at the dentist or been published in the ables, are believed to be more enjoying eating fruits and veget at soccer practice.” these healthful foods. ways to get children to eat USDA-FNS (D2537-1)

Agricultural Research O July 2012 13 parenting practices. Instead, O’Connor’s are related to children’s vegetable and “In general, those surveyed agreed that team looked at an array of categories and fruit intake,” O’Connor notes. She plans it’s more helpful for parents to be proac- at combinations of specific tactics from to use this research as the starting point tive than reactive in getting children to eat within each category. for a longer study. “We looked at one fruits and vegetables. Proactive actions, “Parents don’t do just one thing when time period—essentially, 3 daysin the such as creating a home environment in trying to influence their child’s eating lives of our volunteers. Now we want to which kids are likely to see and be served behaviors,” saysO’Connor. “Rather, look at how parenting practices influence fruits and vegetables, to see their parent they do a combination of things. So, we children’s intake of vegetables and fruits enjoying eating fruits and vegetables, attempted to investigate this by looking at over a longer period of time, such as 1 or and to have the chance to help a parent data pertaining to five different types of 2 years.” select and prepare fruits or veggies, were behaviors that parents in our study reported The findings were documented in a believed to be more effective techniques, using when trying to get a child to eat a 2009 issue of Public Health Nutrition, a in the long term, for getting children to veggie or a fruit. peer-reviewed journal. eat these foods.” “These categories were: ‘teachable mo- Lessons From Home and Abroad: Pros On the other hand, being reactive by ments,’such as telling your son or daughter Share Their Insights pressuring, scolding, or punishing the to try a couple of bites of a vegetable or Other parent-and-kid-focused research child who’s not eating fruit or vegetables a fruit, but that he or she doesn’t have to led by O’Connor has yielded a globe- was believed to be ineffective—or even eat all of it; ‘practical methods,’ such as spanning glimpse of parenting practices counterproductive—in the long run.” adding something to make a veggie or fruit pertaining to fruits and veggies. Her Hous- According to O’Connor, these consen- taste better to the child; ‘firm discipline,’ sus opinions “can be useful for parents likepreventing your child fromhaving USDA-FNS (D2536-1) who are trying to find new ways to sweets if he or she doesn’t eat the veg- encourage their child to eat more fruits gie or fruit; ‘restriction of junk foods,’ and vegetables, and also for public such as not keeping any junk foods in health and healthcare specialists who the house; and ‘enhanced availability are developing strategies to promote and accessibility,’ such as keeping a increased fruit and vegetable intake container of ready-to-eat carrots on a among young children. lower shelf of your fridge that your “At Houston, we’re using what we preschooler can easily reach. learned from this study, and others, “We then grouped parents into to develop food-based strategies for three clusters according to their use doctors and other clinicians to use as of tactics that are within these general a first-line treatment of obesity among categories of practices,” O’Connor their younger patients.” says. “On average, no matter what O’Connor and colleagues reported group their parents were in, kids ate their findings in a peer-reviewed ar- less than the recommended number of ticle published in 2010 in the Journal daily servings of veggies and fruits. of the American Dietetic Association. But children of the parents who used Many U.S. kids eat more servings of fruits than “Today, most kids in this country eat less of the reactive ‘firm discipline’ vegetables, but most eat less of each than they should. less than the recommended amounts tactics and showed a preference for of veggies and fruits,” O’Connor the proactive ‘teachable moments’and ton team, and severaluniversity re search ers says. “We hope that findings from ‘enhanced availability and accessibility’ based in the United States and abroad, de- our studies will help change this for the approaches ate slightly more veggies and signed, conducted, and analyzed results of better.”—By Marcia Wood, ARS. fruits than children whose parents were an Internet survey that tapped the expertise This research supports the USDA pri- in the other two groups. The finding was of nearly 900 doctors, nurse practitioners, ority of improving children’s health and statistically significant.” registered dietitians, and other healthcare nutrition and is part of Human Nutrition, specialists, mostly in Australia, Chile, Using combinations of proactive prac- an ARS national program (#107) described Mexico, Spain, and the United States. tices “appears to be more effective than at www.nps.ars.usda.gov. using combinations of other parenting Survey participants were asked to rate Tom Baranowski and Teresia M. tactics,” she says. “So, we now want to the long-term effectiveness of nearly O’Connor are with the USDA-ARS determine which specific combinations 40 different parenting practices. “The Children’s Nutrition Research Center, give the best results.” people who took part in this survey have 1100 Bates St., Houston, TX 77030; firsthand experience counseling parents This study was “one of the first to look (713) 798-6767 [Baranowski], (713) 798- about their preschoolers’ eating habits,” at how parents use combinations of parent- 6782 [O’Connor], [email protected], says O’Connor. ing practices and how these combinations [email protected].*

14 Agricultural Research O July 2012 STEPHEN AUSMUS (D2505-5)

omorrow’s orchards of almonds, pistachios, and walnuts might be Tsprayed with fine mists of a ben- eficial yeast, Pichia anomala. Studies led by Agricultural Research Service plant physiologist Sui-Sheng T. (Sylvia) Hua have shown that this yeast can undermine a troublesome mold, Aspergillus flavus. Themold is of concernbecause itproduces aflatoxin, a natural carcinogen. Federal food safety standards and quality-control procedures at U.S. pack- inghouses help ensure that these crunchy, healthful tree nuts remain safe to eat. Nonetheless, growers and processors have a continuing interest in new, environmen- tally friendly ways to combat the mold. Hua is one of several scientists at ARS’s Western Regional Research Center in Al- bany, California, who are investigating new strategies for thwarting A. flavus. The idea of developing a practical, af- fordable way for growers to use a yeast to fight a mold isn’t new. But Hua’s tree- nut-focused investigations of P. anomala may be among the most extensive of their kind to date. Her research has included exploring the $ 1HZ:HDSRQIRU.HHSLQJ7UHH1XWV6DIH7R(DW yeast’s talents as a biocontrol candidate in a series of laboratory tests at Albany and in a field test at a California pistachio orchard. generating ATP (adenosine triphosphate) be involved in the cell wall and cell mem- The orchard study, documented in a patent system, vital for the mold’s survival. The brane damage observed in the fluorescence issued to Hua in 2009, indicated that the findings also suggest that the yeast dam- assays.”—By Marcia Wood, ARS. yeast was responsible for a 96-percent aged mold cell walls and cell membranes. This research supports the USDA pri- reduction in the number of mold spores. Walls and membranes perform the essential ority of ensuring food safety and is part For ongoing laboratory research, Hua role of protecting cell contents. of Food Safety, an ARS national program has selected, refined, and applied several The team used a different analytical (#108) described at www.nps.ars.usda.gov. analytical procedures to discover precisely procedure—quantitative reverse tran- Sui-Sheng T. (Sylvia) Hua, Maria T. how the yeast disables the mold. “If we scriptase PCR (polymerase chain reaction) Brandl, and Bradley J. Hernlem are at the understand the underlying mechanisms,” assays—to analyze the activity of certain USDA-ARS Western Regional Rese arch she says, “wemay be able to use that knowl- P. anomala genes in the presence of the Center, 800 Buchanan St., Albany, CA edge to increase the yeast’s effectiveness.” mold. Preliminary findings, which Hua 94710; (510) 559-5905 [Hua], (510) 559- In a collaborative experiment with Al- reported at the annual national meeting 5885 [Brandl], (510) 559-5937 [Hern- bany coinvestigators Bradley J. Hernlem, of the American Society for Microbiology lem]; [email protected], maria. a chemical engineer; and Maria T. Brandl, in 2010, suggest that exposing the yeast [email protected], bradley.hernlem@ a microbiologist, the mold was exposed to the mold may have triggered the yeast ars.usda.gov. to the yeast and later to several different to turn on genes that code for production * compounds that fluoresce red or green of two enzymes—PaEXG1 and PaEXG2. “These enzymes are capable of degrad- Above: Plant physiologist Sylvia Hua (right) and when evidence of specific changes in the technician Siov Sarreal display petri dishes show- mold’s cells is detected. ing the mold’s cell walls and causing dam- ing the effectiveness of a biocontrol yeast against Results of these assays, documented in age to membranes,”Hua notes. $VSHUJLOOXVÀDYXV On the left, a mutant $ÀDYXV Though further studies are needed, Hua turns the agar orange, signifying aflatoxin produc- a peer-reviewed article in the scientific tion. On the right, when the same $ÀDYXV was journal Mycopathologia, suggest that the says these early, PCR-based findings point inoculated between two streaks of yeast, growth yeast interfered with the mold’s energy- to “gene-controlled mechanisms that may was inhibited and no aflatoxin was produced.

Agricultural Research O July 2012 15 The Search for Nematode-Resistant Cotton

gricultural Research Service sci- Plant breeders have struggled to develop Nichols, seniordirectorfor Cotton Incorpo- entists in Georgia and Mississippi resistant lines in part because cotton has a rated, which is funding much of the work. Aare helping cotton growers deal diverse and complicated genome—some The research has taken on a sense of with the double-barreled threat posed by plants have two sets of chromosomes urgency because a pesticide widely used to two nematode species that lurk in their and some have four—making it difficult control nematodes in cotton fields, Temik, fields. The root-knot nematode (Meloido- to cross “wild” resistant germplasm with is in short supply and is scheduled to be gyne incognita) thrives in the sandy soils commercial cultivars and come up with a discontinued in the years ahead because of throughout much of the southern United hybrid that will produce seed. Develop- health and environmental concerns. The States and can cause crop losses of up to ing lines resistant to root-knot nematode phase-out of the pesticide, also known as 10 percentworldwide. Thereniformnema- has been particularly challenging because “aldicarb,”is “prodding everyone working tode (Rotylenchulus reniformis) is limited resistance is a multi-gene trait, and that in this area to step lively,”Nichols says. to warmer regionsof the Cotton Belt, but its makes developing a resistant cultivar time Eliminating undesirable traits in cotton range is expanding. It causes an estimated consuming and extremely expensive. is a team effort in which researchers $130 million in losses each year to the ARS efforts have attracted supportfrom U.S. cotton industry. In some areas, crop cotton growers looking for environmen- Plant pathologist Sally Stetina (left) and losses caused by the reniform nematode tally friendly ways to repel soil pests. technician Kristi Jordan examine cotton roots are as high as 75 percent, depending on “Our best hope for future management with a microscope to determine the level of weather conditions. Losses are greatest of nematodes is to achieve through plant infection by reniform nematode. By comparing under drought stress that typically occurs infection levels in resistant test lines to those in breeding much of what we are now doing susceptible controls, they can identify lines with from midsummer to early fall. with chemical treatments,” says Robert the most resistance.

STEPHEN AUSMUS (D2513-2)

16 Agricultural Research O July 2012 STEPHEN AUSMUS (D2511-15) Geneticist John Erpelding cross-pollinates tool for breeders and provides a source of *RVV\SLXP cotton flowers to develop new resistance to root-knot nematode, along populations. with yields higher than and quality superior to a breeding line released in 1989 and still used in many field trials as a research Davis and colleagues at the University standard. Davis released the new line in of Georgia have released a root-knot-nem- a recent report in the Journal of Plant atode-resistant line for breeders to work Registrations. with, and they are hunting for additional genetic markers that will open pathways “What makes this release significant is toward development of commercially vi- that it has extremely good fiber quality, able resistant lines. it resists the root-knot nematode, and it can grow all over the southeastern United Davis has been focused on combating States,”Davis says. nematode resistance for years. In 2006 he and Peng Chee, his University of Georgia Developing Cotton That Resists Both partner, published a paper that identified Nematodes areas of the cotton genome where root-knot At the Crop Genetics Research Unit resistance genes are likely to reside. They in Stoneville, Mississippi, ARS plant pa- have since refined the search by mapping thologist Sally Stetina and plant geneticist portions of the chromosome where the John Erpelding are conducting a program resistancegenes arelocated and identifying to insert genes for reniform nematode “flanking markers” that lie on either side resistance into cultivated upland cotton essentially “pass the baton” to plant of the genes themselves. These results, varieties. Those resistance genes will breeders to develop commercial varieties. published in Theoretical Applied Genetics, come from several distant relatives: G. Scientists use molecular tools to link will be critical in the search for the specific aridum, G. arboreum, G. herbaceum, and nematode resistance with certain patterns genes that confer resistance to nematodes. G. barbadense. in the plant’s DNA, and those patterns Their new line is the result of sev- But crossing cultivated cotton with its are referred to as “markers.”Researchers eral years of field trials where researchers distant cousins isn’t easy, mainly because provide new cotton lines with those evaluated crosses among cotton plants, of chromosomal incompatibilities. markers to plant breeders and they use some raised in fields inoculated with the “Upland cotton is tetraploid—meaning them to screen for resistance based on nematode and others raised in fields free it hasfour sets of chromosomes—and most the markers, crossing plants that have of it. The new line is susceptible to the of the related species with reniform nema- them with adapted commercial lines. This reniform nematode and is not intended as tode resistance are diploid, having two process eventually leads to lines with both a commercial cultivar. Butit is an excellent sets of chromosomes,” explains Stetina. resistanceand the desirabletraits inherited from commercial varieties. Agronomist Jack McCarty (left) and geneticist Johnie Jenkins study one of the cotton lines that resist ARS researchers are making it easier for root-knot nematode. In ongoing studies, this resistant line is being crossed with other cotton plants to transfer resistance. breeders to develop commercially accept- able materials by transferring resistance genes from wild plants into cotton cultivars and releasing the resulting lines as breeding tools. They are also developing molecu- lar markers to speed up identification of key nematode-resistance genes. Much of the research is focused on upland cotton (Gossypium hirsutum), which is native to Mexico and Central America and is one of two principal types of cotton, making up more than 95 percent of U.S. production. “Finding genetic markers is critical if we want cotton breeders and private compa- nies to get involved and begin developing commercialvarieties with nematoderesis- tance,” says Richard Davis, an ARS plant pathologistattheCoastalPlain Experiment Station in Tifton, Georgia.

RUSS HAYES (D2518-1)

Agricultural Research O July 2012 17 “When you cross these directly, you get a Stetina and Erpelding have teamed with search in root-knot nematode resistance in triploid hybrid, a plant with three sets of MSU researchers Peggy Thaxton and Ted cotton,”says Jenkins.Raymond Shepherd, chromosomes that is sterile; it will not set Wallace to develop cotton varieties with a retired ARS scientist, was instrumental seed, and the resistance you moved in will resistance to the two nematode species by in using root-knot nematoderesistance in a never be passed to the next generation.” using marker-assisted selection. Offspring line of wild cotton from Mexico to develop The researchers’ solution was to create from crosses are first selected based on the resistant germplasm, he says. an intermediary cotton strain, known as presence of markers for resistance. Plants Jenkins and his colleagues found pat- a “bridging line,”using a series of com- that are found to have multiple sources terns of DNA associated with root-knot plicated procedures, including embryo of resistance are directly challenged with nematode resistance and key genetic under- rescue and chromosome doubling. Its the nematodes to confirm the resistance. pinnings that confer resistance to reniform express purpose is to serve as a bridge Advanced lines of upland cotton that resist nematode. The markers theydeveloped for between species so that genes for reniform one or both of the nematode pests may be resistance to root-knot nematode in upland nematode resistance can be passed from ready for release in 2 to 4 years. cotton—found on chromosomes 11 and cotton’s distant relatives into cultivated A Pest for the Past 100 Years 14—should be useful in selecting plants varieties or germplasm lines used to breed At the ARS Genetics and Precision Agri- with resistance. They also found that re- them. However, says Stetina, “When you cultural Research Unit in Mississippi State, sistance to reniform nematode in a wild G. bring in resistance from the related species, Mississippi, geneticist Johnie Jenkins and barbadense line is governed by more than you can introduce undesirable traits such his colleagues have also made significant one gene, and they have identified markers as smaller bolls, limited flower- linked to these genes on chromo- ing, poor fiber quality, and poor (D2549-1) somes 21 and 18. They published performance under typical U.S. separate articles on the root-knot crop conditions. That’s why ad- nematode work and the reniform ditional crosses with adapted lines nematode work in Theoretical that have desirable agronomic and Applied Genetics. traits are critical to getting the Commercial breeders had right combination of resistance steered away from efforts to and crop performance.” breed root-knot resistance into Markers provide an important upland cotton lines over the years tool to track resistance over because it was governed by more multiple generations of crossing than one gene and seemed so to ensure successful transfer, costly and time-consuming, says Erpelding adds. ARS agronomist Jack McCarty. Erpelding and Stetina aim to But the research contributions develop markers associated with from Jenkins and his colleagues reniform nematode resistance in may change that due to the use of G. arboreum and G. herbaceum marker-assisted selection. and make themavailable to breed- “This research has sparked ers. Markers are already available interest from some plant breeding for reniform nematode resistance companies in trying to develop from G. longicalyx, G. aridum, high levels of resistance to root- and G. barbadense sources. knot and reniform nematode in These were developed by teams upland cotton,” he says.—By of researchers from ARS, Texas Dennis O’Brien, Jan Suszkiw, A&M University, Mississippi and Sharon Durham, ARS. State University (MSU), Cotton A juvenile root-knot nematode, 0HORLGRJ\QHLQFRJQLWD penetrates a tomato root. Once inside, the juvenile, which also attacks cotton This research is part of Plant Incorporated, and Monsanto roots, causes a gall to form and robs the plant of nutrients. Photo by Diseases (#303) and Plant Ge- Company, Stetina says. William Wergin and Richard Sayre. Colorized by Stephen Ausmus. netic Resources, Genomics, and Depending on the field in Genetic Improvement(#301), two which it is grown, cotton can be strides in coming up with nematode- ARS national programs described at www. attacked by many different nematodes, so resistant cotton lines. nps.ars.usda.gov. varieties with resistance to two or more Root-knot nematode has been recog- To reach scientists mentioned in this nematode species can be beneficial. In nized as a cotton pestforthe past 100years, article, contact Dennis O’Brien, USDA- Mississippi, reniform nematode and root- according to Jenkins. “Since the 1930s, ARS Information Staff, 5601 Sunnyside knot nematode are the biggest challenges scientists have been looking for resistance Ave., Beltsville, MD 20705-5129; (301) to profitable cotton production. to nematodes. Inthe 1960s, ARS started re- 504-1624, [email protected].*

18 Agricultural Research O July 2012 No-Till Crops Can Improve Air Quality

in the Pacific Northwest “One major windstorm can gener- ate enough airborne dust to exceed air quality standards for PM10,”Sharratt winter wheat. Soils are most exposed or more than 100 years, farmers says. “But annual no-till cereal crops after sowing, and sowing schedules in the Pacific Northwest interior appear to be a viable strategy that coincide with the seasons when high Fhave favored winter wheat/sum- farmers can use to control erosion winds prevail in the Pacific Northwest. mer fallow production systems. Since and meet air quality regulations in the rain typically falls during themild The scientists found that in the Pacific Northwest.” The next challenge winters, this schedule gives the growing spring, soils in spring barley and spring is finding ways to make annual no-till crops the water they need when they wheat rotations were wetter than soils crop systems as profitable as the cur- need it. But this rotation can require up in traditional winter wheat systems. In rent winter wheat/fallowsystem.—By to eight tillage passes during the fallow late summer, the no-till spring barley Ann Perry, ARS. season to control weeds and conserve rotation also had more standing stubble soil water. Tillage also creates a dry, than the other two rotations. Stubble This research is part of Climate loose bed of fine soil particles that is helps keep soil on the ground and out Change, Soils, and Emissions(#212), easily carried away by the strong sum- of the air. an ARS national program described mer winds. Results from the study highlighted at www.nps.ars.usda.gov. “In the inland Pacific Northwest, other soil-quality payoffs from using To reach the scientists mentioned when atmospheric levels of PM10— spring wheat/spring barley rotations. in this story, contact Ann Perry, particulate matter that is 10 microns or Soils had larger and more continuous USDA-ARS Information Staff, 5601 lessindiameter—exceed federal limits, pore space, higher water-infiltration Sunnyside Ave., Beltsville, MD 20705- it’s usually because of erosion from rates, higher saturated hydraulic con- 5129; (301) 504-1628, ann.perry@ farm lands,”says Agricultural Research ductivity, and higher drainage rates. ars.usda.gov.* Service scientist Brenton Sharratt. He Given these findings, the team con- Near Ritzville, Washington, is the research leader in the ARS Land cluded that annual no-till spring cereal Management and Water Conservation scientists use a crops could significantly improve water wind tunnel to Research Unit in Pullman, Washington. infiltration and retention and help retain measure wind “Since the U.S. Environmental Protec- crop surface residue in the late summer. erosion of soil particles from a tion Agency regulates air quality, farm- Farmers could benefit from improved ers in this area are looking for ways to recently planted soil quality—and a reduced risk of wheat field. reduce erosion from their fields and wind erosion. assist communities in complying with regulations.” To help address these concerns, Sharratt, ARS agronomist Frank Young, and Washington State University re- search associate Gary Feng conducted an 11-year study that evaluated whether no-till spring cereal rotations could help mitigate wind erosion. The systems they studied included thetypicalwinter wheat/summer fallow rotation, a no-till spring barley/spring wheat rotation, and a no-till spring wheat/chemical fallow rotation. During the study, they tracked several soil characteristics, including aggregation, moisture, roughness, crusting, and crop residue cover. Soil properties were measured twice during the study: once after sowing spring wheat and once after sowing

BRENTON SHARRATT (D2539-1)

Agricultural Research O July 2012 19 PEGGY GREB (D2524-1) ;MIZKPQVO NWZ /MVM[

Tara VanToai, retired ARS plant physiologist, and Thomas Doohan, a student at Ohio State University, collect soybean plants and root samples to analyze them for response to flooding stress.

oybean varieties that grow in rice pad- VanToai worked with plant pathologist The team works on multiple fronts, in- dies in Southeast Asia could provide Anne Dorrance and soybean breeders cluding molecular plant breeding, with the ;the United States with much-needed Grover Shannon and Henry Nguyen in the help of DNA markers, genetic transforma- genes for developing soybeans tolerant to search for genes that protect against both tion, and soilmanagement—allin an effort flooding—as well as to root rot and other flooding and plant diseases. Dorrance is to protect soybeans growing on wet soils. plant diseases found in waterlogged soils. at Ohio State University’s Ohio Agricul- Flooding—Vietnam to Ohio Tara VanToai pursued the genes for these tural Research and Development Center Growing up in the Mekong Delta of traits. She recently retired from the Agri- in Wooster; Shannon and Nguyen are at Vietnam, and then working in Ohio, Van- cultural Research Service’s Soil Drainage the University of Missouri, in Portageville Toai has experienced firsthand the harm Research Unit in Columbus, Ohio. and Columbia, respectively. flooding does to crop yields. VanToai has

20 Agricultural Research O July 2012 PEGGY GREB (D2529-1) been studying flood tolerance in soybeans Australia, respectively. for more than two decades, in greenhouse, The initial experiments lab, and growth-chamber conditions as well were done in outdoor as in experimental and farm fields in Ohio “screenhouses,”which and in Missouri. She has also studied and are greenhouses with collected soybean lines in Vietnam and screens instead of China, including backyard soybean plants glass. VanToai found that survived China’s 1991 flood-of-the- that the screenhouses, century. The series of research studies is while not complete- beingfunded by ARS, the U.S. Department ly duplicating field of Agriculture’s Foreign Agricultural Ser- results, allowed for vice, the United Soybean Board, and the accurate predictions North Central Soybean Research Program. of flood tolerance in She has also collaborated with scientists soybeans. from Brazil, China, France, and Hungary, Flood-tolerance is as well as Vietnam. defined as the ability U.S. Soybeans Need New Genes of a plant to survive 10 VanToai and colleagues need to incor- days of steady flooding porate genes from plants native to other during the plant’s criti- countries to supplement the narrow genetic cal flowering stage. Technician G. John Lazur (formerly with ARS) scans soy- bean root with winRHIZO software to digitize root growth. base of U.S. soybeans and to improve toler- For the screenhouse ance to wet soil and associated diseases. tests, the plants were The varieties of soybeans used by most grown in pots. When each plant was in channels that enable roots of plants—rice, American farmers are damaged by even full bloom, it was placed for 2 weeks in a for example—to grow under water. These short periods in waterlogged soil. Yield bucket of water so that the water level was open channels allow flooded roots to losses as high as 25 percent are estimated 2 inches above the soil surface. snorkel air from the above-water parts of in the Mississippi Delta region, Asia, and In later field tests, the plants were sub- the plants. ARS scientists and university other regions of the world where soybean jected for 2 weeks to water 4 to 6 inches breeders haveintroduced aerenchymainto crops are rotated with paddy rice. The above the soil surface. wheat and corn experimentally. losses are from injuries due to flooding The screenhouse tests killed up to 100 VanToai found that soybean plants that and flood irrigation. percent of susceptible varieties. The test survive flooding use some of the same Tests of 21 soybean varieties in flooded accurately identified the top three flood- mechanisms as rice growing in paddies. experimental fields at Can Tho, Vietnam, tolerant lines, later borne out by the The two standbys that rice roots rely on revealed three lines of soybean—VND2, flooded-field tests. for its ability to grow in water are adventi- Nam Vang, and ATF15-1—with superior Soybean Secrets tious roots and aerenchyma. Adventitious flood tolerance. VanToai has also analyzed flood-tolerant roots grow out of the plant’s stem near the VanToai and the team did this research eastern gamagrass in the United States soil surface. They work in tandem with with Tran Thi Cuc Hoa and Nguyen Thi for its secrets, one of which turned out aerenchyma to provide even more oxygen Ngoc Hue—both with the Mekong Delta to be aerenchyma. This is tissue with air to flooded roots. Rice Research Institute, where the experi- ments were done. Soybean Plants with Superior Flood Tolerance Plants from the three flood-tolerant lines Ohio State University grew tallest and had the biggest seeds and scientist Liming Chen examines flooded the highest yields. soybean plants. The 21 lines tested included plants na- tive to Vietnam and Cambodia and those developed through selection by farmers and gardeners. They also included lines developed by modern breeding practices and imported from Australia, China, Japan, and Taiwan. Nam Vang is native to Cambodia, while VND2 and ATF15-1 are from China and ANN HOUSER (D2531-1)

Agricultural Research O July 2012 21 PEGGY GREB (D2527-1) PEGGY GREB (D2528-1) Far left: Soybean plants after 2 weeks of flooding stress. Left: Soybean plants that were not flooded.

ceptible U.S. variety S99-2281 lost more than 80 percent of its yield. The scientists crossed PI 408105A with S99-2281 to study and transfer the resistance genes. They evaluated 200 lines from these crosses in fields in Ohio and Missouri for 2 years and identified several lines with a consistent tolerance to flooding. DNA markers were found that could effectively identify plants that carried the resistance genes. Clear genetic differences were found between the roots of flood- tolerant and flood-susceptible plants. Because the parents PI 408105A and Genes for Flood Survival lines. Some soybean lines developed S99-2281 also show differences in resis- The scientists are looking for genes that adventitious roots as early as 2 days after tance to soybean root and stem rot diseases, trigger thedevelopmentof thosetwo stand- being flooded. They discovered that a Dorrance used the same 200 lines of the bys, as well as other flood-tolerant aids. flood-tolerant South Korean variety, PI population to look for genes for root rot They found that flood-tolerant soybean 408105A, showed only a 30-percent yield resistance. lines develop more adventitious roots and reduction after 10 days of flooding during Are Flood- and Root Rot-Resistance more aerenchyma than flood-susceptible the critical flowering stage, while the sus- Genes the Same? “Now we’re wondering whether root rot CHRISTINE STONE D521-1 was contributing to the yield reductions Resistance to Soybean Rust Asian soybean rust, we saw in susceptible plants,” VanToai a major disease of soybeans worldwide, was found in says. “Did the tolerant plants do so much soybean fields in South America in 2000. It was first better because they were tolerant not only of flooding but also of root rot? Are some found in the continental United States 4 years later—most likely of the same genes involved?” VanToai its spores delivered by winds of an unusually busy hurricane season. wonders. From 2006 to 2009, ARS plant pathologists Tara VanToai (retired), She and colleagues have mapped the at Columbus, Ohio, and Glen Hartman, at Urbana, Illinois, and other genes and found that there is an overlap colleagues at the University of Illinois at Urbana and the University of of genes for resistance to flooding and Missouri at Columbia collaborated with scientists at the Plant Protection wet-soil-borne diseases, which indicates Research Institute and the Thai Nguyen Education University in Hanoi, some genes are involved in both, while Vietnam,to test 65 soybean varieties for resistance to soybean rust. These others are not. varieties included rust-resistant accessions from the USDA Soybean Several of the plant lines VanToai and Germplasm Collection identified in earlier research, resistant varieties colleagues have developed over the years from the Vietnam collection, and susceptible checks. In all, she and col- are included in the ARS Soybean Germ- leagues found that the Vietnamese soybean variety DT 2000 showed plasm Collection in Urbana, Illinois.—By the strongest resistance in all five experiments. Response of varieties Don Comis, formerly with ARS. with known resistance genes varied from experiment to experiment. This research is part of Water Avail- ability and Watershed Management (#211), Asian soybean rust is constantly forming new strains. “This means that Plant Genetic Resources, Genomics, and we need to develop standardized ‘tester’ strains so that we can compare Genetic Improvement (#301), and Plant results across studies,”VanToai says. Biological and Molecular Processes The next step will be to identify and map the resistance genes present (#302), three ARS national programs de- in the resistant variety found in the Vietnam study and determine their scribed at www.nps.ars.usda.gov. effectiveness against specific soybean rust strains. To reach scientists mentioned in this VanToai says that the rust-resistant soybean varieties from Vietnam may article, contact Robert Sowers, USDA- have genes that could be helpful wherever rust strains have a similar ARS Information Staff, 5601 Sunnyside Ave., Beltsville, MD 20705-5129; (301) virulence to those in Vietnam.—By Don Comis, formerly with ARS. 504-1651, [email protected].*

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