College of Natural Resources, UC Berkeley

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October 31, 2003

Microbial Infection Can Significantly Shorten Shelf Life of Eggs

by Sarah Yang

Berkeley - Research led by University of California, Berkeley, biologists has implicated microbial infection as a culprit for why some birds start incubating eggs in a nest before the clutch is complete, resulting in eggs hatching at different times and putting younger chicks at a significant disadvantage that often leads to death.

Incubation seems to protect eggs from bacterial and fungal infection, according to the study, to be published in the Nov. 22 issue of the Proceedings of the Royal Society B.

"We found that freshly laid eggs that aren't incubated have a shorter shelf life," said Steven Beissinger, principal investigator of the study and professor and chair of the Department of Environmental Science, Policy and Management at UC Berkeley's College of Natural Resources. "Without incubation, eggs are more vulnerable to infection from micro-organisms. This paper is the first to present evidence that microbes may be connected to egg viability and incubation patterns in wild birds."

Beissinger explained that birds generally produce no more than one egg per day, so there may be a one-to-two-week lag between the laying of the first and last eggs in a nest. Ornithologists have puzzled over why many birds start to incubate their eggs before all are laid, which causes them to hatch at different times. The pattern leads to a high mortality rate for the clutch's younger hatchlings because they cannot compete with their bigger siblings that got a head start in life.

"The dominant theory is that birds intentionally do this because they would not be able to adequately feed all the young if they hatched at once," said Beissinger. "By staggering the hatching order, they can control the food demand from the number of young they have to care for at any one time."
However, a number of recent studies, including one conducted by Beissinger, have caused some to question that theory. By moving eggs that were laid around the same time into one clutch - a procedure they nicknamed "egg bingo" - researchers found that bird parents were able to successfully raise more young if they hatched together rather than weeks apart.

So, the hunt began for an alternate explanation of birds' hatching patterns. Beissinger worked with Mark Cook, a UC Berkeley post-doctoral researcher in ecosystem sciences and lead author of the paper; Gary Toranzos and Roberto Rodriguez from the University of Puerto Rico's Department of Biology; and Wayne Arendt from the USDA Forest Service International Institute of Tropical Forestry.

The researchers conducted the study at two research sites in Puerto Rico. One site was at the top of the Luquillo Experimental Forest, a cool, very humid cloud forest with an average daily temperature of about 70 degrees Fahrenheit. The other site was a drier lowland forest at the base of the mountain with daily temperatures averaging 81 degrees.

The researchers used freshly laid, free range chicken eggs provided by a local supplier. They sampled the shell surface of 164 eggs for baseline bacteria levels, then randomly assigned the eggs to one of the two sites and exposed them for periods of one, three, five or seven days. After the exposure time, the eggs' yolk and albumen, or egg white, were separately tested for the presence of microbes.

They found that microbes were rapidly able to enter the egg through its pores and overwhelm the embryo within three to five days. The longer the exposure time, the greater the rate of infection for eggs at both sites. Microbes appeared on the inner membrane after just one day of exposure. Microbes reached the egg white after three days, and the yolk after five days.

Eggs in the cloud forest were more susceptible to infection. Sixty percent of the eggs were infected by microbes after five days in the cloud forest compared with 25 percent of eggs infected in the lowland forest.

"It's damper in the cloud forest, which makes it easier for the microbes to get through the egg shell," said Cook. "Water tends to help bacteria and fungi penetrate the pores of the shell. Also, once fungi take hold on the shell, it quickly starts deteriorating the thin protective cuticle that covers the shell."

The next step was to determine whether microbial infection could impact hatching success. The researchers began another round of experiments with 152 freshly laid chicken eggs. They randomly assigned the eggs to either the cloud forest or lowland forest sites, and exposed them for five days. In addition, half the eggs were wiped twice daily with alcohol to kill germs on the shell surface. After five days, the eggs were then incubated to test hatching success.

Several dozen baby chickens later, the researchers found that of the eggs in the cloud forest, 76.5 percent of the cleaned eggs hatched compared with 22.7 percent of the eggs that had been left uncleaned. In the lowland forest, only 40 percent of the eggs hatched regardless of whether they had been cleaned or not, suggesting that temperature played an important role along with microbes in controlling embryo viability.

Incubation warms eggs up to about 95 F, the optimal temperature for development. When temperatures drop to about 77 F or below, cell division stops, but the embryo can remain alive in a state of limbo.

But with temperatures in the danger zone of 77 to 95 F, tissues are more likely to develop incorrectly, leading to higher embryo mortality.
Eggs in the cloud forest that were cleaned had the highest rate of viability because they were protected from both microbial infection and dangerous temperatures.

"For the cleaned eggs that didn't hatch in the lowland forest, we're primarily seeing the impact of temperature," said Cook. "The two factors - microbes and temperature - can act independently or together to reduce the shelf life of the non-incubated eggs."

The results also shed light on why some birds partially incubate their eggs. That is, they may only incubate their eggs for two or three hours in a day.
"Some eggs are filled in the albumen with anti-microbial agents," said Beissinger. "For those agents to react with the enzymes in the albumen and work well, they need to be warmed. Warming the eggs up a bit may serve to clean out the bacteria and fungi that may have gotten in through the shell."

"Our research shows the tradeoff that birds are making in their incubating behavior," said Beissinger. "Birds may be beginning incubation early in part to maintain the viability of the embryos inside the eggs, even though they are risking a disadvantage to their latter-hatching offspring."

This research was supported by the National Science Foundation and the USDA Forest Service.

October 30, 2003

Scientists Accurately Map Plant Genome, Which Could Lead to New Generation of Hybrid Crops

by Sarah Yang

Berkeley - In a study led by researchers at the University of California, Berkeley, and the Salk Institute in La Jolla, Calif., scientists have accurately mapped the genes of the common mustard weed, Arabidopsis. The achievement may lead to the next generation of genetically modified crops that can grow faster, produce more food and resist disease.

The study, which appears in the Oct. 31 issue of Science, reveals the existence of nearly 6,000 genes, about one-third of the genes that exist in Arabidopsis. Knowing these genes and how they work can allow researchers - in a short period of time - to use them to change the characteristics of other plants.

"Arabidopsis has all the genes a plant needs," said Joe Ecker, Salk professor of plant biology. "All flowering plants are closely related, and so the genes that encode various traits are also shared. It's possible, then, to take a gene for flowering from Arabidopsis and insert it into rice or poplar, and have that gene function."

Ecker and Athanasios Theologis, adjunct professor at UC Berkeley's College of Natural Resources and senior scientist at the Plant Gene Expression Center, are the principal investigators on the project, which includes a team of 72 scientists from nine institutions in the United States and Japan. The Plant Gene Expression Center is a collaboration between UC Berkeley's Department of Plant and Microbial Biology and the USDA's Agricultural Research Service.

The findings revealed some shortcomings of computer-based gene prediction programs, including those that have been used to sequence the human genome and the Arabidopsis plant - the plant biologists' equivalent of the fruit fly for genetics research.

The researchers point out that computer algorithms can't always distinguish whether a piece of code corresponds to a single gene or to two overlapping genes. And while the programs have become increasingly accurate in recent years, the researchers added, computer programs may still put genes' parts in the wrong places, find genes that aren't really there, or miss genes altogether. What researchers say they often get from an initial sequence of a genome is a "best-estimate" lineup of transcription units.

To get the real picture of what's there and what's not, researchers say they need empirical, experimental verification.

The research team placed the entire Arabidopsis genome, consisting of about 25,000 suspected genes, on a series of six gene chips, and then analyzed the chips for any protein-making activity, the primary function of genes. They isolated one-third of the plant's genes, which will be publicly available for researchers to fix errors in the current blueprint of the genome. In addition to finding shortcomings in the much-heralded, computerized methods of sequencing a genome, they discovered about 3,300 functioning genes for the first time.

"By putting the entire genome on the gene chips, we could find that what the computers predicted as genes were wrong about a third of the time," said Ecker. "But we also found other genes we had not seen before. Genetically, plants are much simpler than animals, so this information can be used almost immediately to improve crop yields and disease resistance."

"We eventually want to be able to understand the function of all the proteins within an organism," said Theologis. "If you know the correct gene structure, you can clone DNA to express and study proteins. This type of research eventually will lead to advances in proteomics."

Many of the researchers on this study were part of the team that sequenced the genome of Arabidopsis nearly three years ago. The initial genome work and the current research are funded by the National Science Foundation (NSF), which established a project to identify an entire plant genome by 2010.

"The technology used in this research will be able to reveal the dark matter in a genome," said Theologis. "We will be able to identify never-before-seen RNA in regions that were once thought to contain no genes. Researchers could also use this method to get a more definitive answer to how many genes are in the human genome."

"Finding the genes that lurk in the DNA sequence sounds like an easy problem, but in fact is tremendously challenging," said Robert Last, program director of the NSF's plant genome research program. "Completion of the DNA sequence of a genome such as Arabidopsis is an important milestone towards understanding the function of every gene in the plant, and discovering the genes that can positively influence the productivity, nutritional and medical value of the plant to human beings."

October 28, 2003

Federal, Academic Scientists Partner to Streamline Environmental Research

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by Kathryn Stelljes

Berkeley — A new era of scientific collaboration for the benefit of the environment begins today (Tuesday, Oct. 28) at the University of California, Berkeley, as the Californian Cooperative Ecosystem Studies Unit (CESU) holds its first planning meeting to set goals for the next five years.
This unit and others like it nationwide establish a streamlined method for matching the research and technical assistance needs and funding of federal agencies with the scientific interests and expertise of university faculty and graduate students. The Californian CESU, hosted by the College of Natural Resources at UC Berkeley, brings together nine University of California campuses, three California State University campuses, and six federal agencies.
While other CESUs have been established for a dozen other geographic regions nationwide, the Californian has several unique traits.
“The Californian CESU provides the largest scope of collaboration among educational institutions,” said Steven Beissinger, chair of the College of Natural Resources’ Department of Environmental Studies, Policy and Management. Beissinger and professor Craig Moritz, director of UC Berkeley’s Museum of Vertebrate Zoology, proposed its establishment, hosted by UC Berkeley.
Other leaders of the UC Berkeley team include Barbara Allen-Diaz, executive associate dean of the College of Natural Resources, and James Shevock, a research coordinator with the National Park Service’s Pacific West region.
“The potential benefit of this arrangement to the environment is huge. California has a tremendous amount of public land managed by the federal government, along with the greatest number of endangered species on the mainland and a population that includes one of every nine people in the U.S.,” said Beissinger. “The CESU provides a way to help maintain biodiversity and protect ecosystems by providing expertise to the land managers who need scientific information.”
“The great thing about this CESU is the breadth of expertise available. This allows us to broaden our research perspectives beyond the natural and physical sciences,” said Shevock. “That’s important for the National Park Service, which manages a wide array of historical, cultural, archeological, and natural places.”
Shevock’s role will be to market the research and technical assistance needs of the National Park Service and match those needs with the expertise found at the CESU universities. He will reside on the Berkeley campus beginning January 1.
Unlike typical federal agreements, research under CESUs is collaborative in design.
“The CESU is a genuine two-way relationship. Agency and university researchers will plan projects together, and share data and expertise, rather than a contractual relationship where a researcher completes a study and may never see how it is used,” said Moritz. “As a researcher, the collaboration is much more satisfying.”
Another benefit is bringing together agencies that may be facing similar problems. “The larger-scale problems are the ones that will really stimulate the interest of faculty,” said Moritz.
The collaborative nature also allows the project to evolve as new information becomes available.
This arrangement provides “a great avenue for graduate students to do research and receive funding,” said Shevock. “It would be much harder for a student to obtain a federal contract. And we’re also hoping that some of these graduate students will consider careers in resource management and become federal employees.” And, belonging to the Californian CESU has wider benefits.
“Once your university is a member of a CESU, any other CESU institution can tap your expertise,” said Beissinger.
The Californian CESU was officially established this summer, but meets as a unit for the first time today. Cooperating institutions are the University of California’s Berkeley, Davis, Irvine, Los Angeles, Merced, Riverside, San Diego, Santa Barbara, and Santa Cruz campuses; California State University at Fresno and Los Angeles; San Francisco State University; the Bureau of Land Management; the Bureau of Reclamation; the U.S. Geological Survey; the National Park Service; the USDA Forest Service; and the National Aeronautics and Space Administration.
Although the initial CESU agreement lasts five years, Shevock believes it will be successful and that the CESU will be renewed into the future. “Environmental issues aren’t going away, and having access to the best research available is an ongoing need of the federal government,” he said. “We’re in this partnership for the long haul.”

October 12, 2003

Don Dahlsten Memorial

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Public Memorial Planned for Don Dahlsten

The Don Dahlsten memorial service is scheduled for Sunday, October 12, 2003, at noon on the east lawn of Giannini Hall on the UC Berkeley campus. A Giant sequoia will be planted in his honor following the ceremony on the northwest lawn of Giannini Hall facing Tolman Hall.

RSVP is requested to memorial@nature.berkeley.edu.

On-street parking is free and generally available on Sundays in Berkeley. In addition, fee parking is available in many campus parking lots. If you need special accommodations, please contact Kathryn Moriarty Baldwin at (510) 643-6641.

Donations can be made in Dahlsten's memory to fund outreach programs benefiting K-12 students. Checks can be sent to the Donald Dahlsten Outreach Fund, c/o the College of Natural Resources, University of California, Berkeley, 101 Giannini Hall, #3100, Berkeley, CA 94720-3100.

For a story about Don’s career, visit

http://www.cnr.berkeley.edu/site/news_item_detail.php?id=58

October 2, 2003

New Treatment for Sudden Oak Death Approved

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by Sarah Yang

BERKELEY – State regulators yesterday (Wednesday. Oct, 1) opened the door for a treatment for Sudden Oak Death to be legally applied by licensed professionals to oaks and tanoaks. A University of California, Berkeley, researcher has proven the treatment effective in preventing or slowing down infection, and he developed an innovative technique to make it significantly easier to administer.

The Department of Pesticide Regulation approved a special registration for this treatment on a fast-tracked, special needs basis. It is the first and only treatment approved by the state for use against a pathogen that has killed tens of thousands of coastal oak trees from California's Big Sur to the border of Oregon.

"The likely areas where the treatment would be used include mostly private-owned land, but trees around high-use facilities in public parks may also be potential candidates," said the UC Berkeley researcher, Matteo Garbelotto, who is an adjunct assistant professor of ecosystem sciences and cooperative extension specialist at UC Berkeley's College of Natural Resources. "It's really meant for individual oaks or tanoaks that are at high risk for infection, such as those in the vicinity of infected California bay laurel trees."

Garbelotto explained that in the wild, California bay laurel trees are considered the most important vector for spreading Sudden Oak Death. Spores congregate on the leaves of those trees, where they can easily become airborne.

Garbelotto and David Rizzo, associate professor of plant pathology at UC Davis, first identified Phytophthora ramorum three years ago as the fungus-like pathogen responsible for Sudden Oak Death. Since the disease was first observed in Marin County in 1994, it has spread to 12 California counties, including Alameda and Contra Costa counties, and it has been confirmed in at least 25 different plant species, including California bay laurels and rhododendrons. It has also been found in nurseries in Washington state and British Columbia, Canada.

"It hasn't been long since the pathogen was first isolated," said Garbelotto, who presented results of his research at a recent meeting of the American Phytopathological Society. "The development of a treatment has been incredibly rapid."

The approved treatment is a phosphite compound sold under the brand name Agri-Fos, and its effectiveness has been proven for oaks and tanoaks. Agri-Fos is a fungicide that has been effective for other Phytophthora species, but Garbelotto was the first to test it on P. ramorum. Agrichem, the Australian based company that sells Agri-Fos, will make the treatment available to trained professionals licensed by the state to apply pesticides.

arbelotto emphasized that the treatment is not a cure-all, and said there is no evidence to show that it would be recommended on a widespread basis. The treatment has not been tested on other plant and tree species susceptible to Sudden Oak Death.

The confirmation last year that redwood and Douglas fir - two of the state's most highly prized trees - were susceptible to the pathogen sparked concerns of the pathogen's ability to do more damage. But it is the state's oak trees that have suffered the pathogen's most dramatic impact, exhibiting oozing lesions and cankers as they die.

Garbelotto has conducted more than 30 independent trials of treatment protocols. Steven Tjosvold, a cooperative extension specialist with Santa Cruz County, helped Garbelotto with field experiments.

To test whether the treatment worked on diseased trees, Garbelotto infected potted oaks and tanoaks with P. ramorum and then waited several days to several weeks before injecting the phosphite compound. The injections slowed down the growth of cankers on trees that were treated.

Not surprisingly, he found that the effectiveness of the treatment declines steadily the longer the tree has been infected. "Ideally, this would be used as soon as symptoms are noted," said Garbelotto. "The treatment is not recommended if the symptoms have been around for a year or more."

He noted that the treatment does not kill the pathogen, but that it stops its growth if used in the early stages of infection.
He also found that injecting oaks and tanoaks with the chemical first and then inoculating them with the pathogen effectively prevented infection. When used as a preventative agent, the treatment completely protected smaller trees, and it reduced the canker size by half on adult trees.

Through his experiments, Garbelotto also developed an innovative way of increasing the effectiveness of the phosphite compound by combining it with an organosilicate carrier, named Pentra-Bark, that helps trees gradually absorb the treatment over time. Instead of injecting the compound, Garbelotto sprayed it directly onto the bark of the trees.

"This method of delivering the treatment to the tree is completely new," said Garbelotto. "It's a discovery that can revolutionize the way trees are treated."

The organosilicate used in this treatment had not been proven successful in carrying chemicals through the bark until Garbelotto paired it with the phosphite compound. The matching works well because the molecular structure of the phosphite is compatible with that of the organosilicate, he said.

"By combining the phosphite compound with organosilicate, the treatment can be sprayed onto the bark instead of injected," said Garbelotto. "The application is so easy compared to injection, which is complicated and requires a lot of skill to do right. With injection, it is hard to know if you are getting the compound into the tree's vascular system, so if you don't know what you're doing, a lot of the compound can get lost."

In addition, applying the treatment to the bark makes the treatment available to the tree for more than six months. "Once applied, it doesn't rub off," said Garbelotto. "It's comparable to a time-release drug patch for humans."

However, the topical application has thus far only been shown effective in oak trees. The compound has not been successfully absorbed through the bark of tanoak trees.

Garbelotto noted that phosphites have been used for more than 10 years and are considered a low-toxicity chemical. In addition, the methods of application - both injection and bark spray - are targeted so that the chemical is not dispersed into surrounding areas.

Nevertheless, he emphasized the need for training before the treatment is used for Sudden Oak Death. Training sessions for professional arborists will be offered through UC Cooperative Extension on Monday, Oct. 20, in Marin County, and Tuesday, Oct. 21, in Santa Cruz County. Details on the classes will be available online within the next week at http://www.suddenoakdeath.org/.

"Until now, many oak trees in northern California faced a potential death sentence from Sudden Oak Death," said Assemblywoman Patricia Wiggins (D-Santa Rosa), who was instrumental in expediting the approval of the phosphite-based treatment. "Dr. Garbelotto and his associates moved quickly, professionally and with great care to find a treatment, for which we are very grateful."

Garbelotto's research was supported by the Gordon and Betty Moore Foundation and the USDA Forest Service Pacific Southwest Region. The Valley Crest Tree Company provided hundreds of potted trees used for the research.

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Microbial Infection Can Significantly Shorten Shelf Life of Eggs
Scientists Accurately Map Plant Genome, Which Could Lead to New Generation of Hybrid Crops
Federal, Academic Scientists Partner to Streamline Environmental Research
Don Dahlsten Memorial
New Treatment for Sudden Oak Death Approved

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