FEATURES


Catching the Bad Guy

DNA sequencing establishes the origin of a destructive bacterium
By Rocky Womack


Genetic sequencing has helped law enforcement catch the bad guys and cleared some of the good guys. Now a select number of university professors and their research teams have used genetic sequencing to identify a bacterium that is harming the kiwifruit. First, a little bit about the destruction so far.


Boris Vinatzer, an associate professor with Virginia Tech, says that by sequencing DNA, researchers can trace the origin of a bacterium that has caused severe losses to the kiwifruit industry.
Photo courtesy of Virginia Tech.

The bacterium, Pseudomonas syringae pathovar actinidiae (PSA), was discovered in the 1980s in Japan and China. In 2008, PSA caused disease in Italy, and it was found in New Zealand in 2010, when the U.S. banned all imports of kiwifruit plant material and pollen so it wouldn't infect crops here.

Since that time, PSA has threatened the world's kiwifruit, destroying orchards in Europe, New Zealand and South America. The destruction overseas has caused hundreds of millions of dollars in economic losses.

Because of the economic losses in 2012, the New Zealand government declared PSA an adverse event and pledged to fund recovery for the country's affected kiwifruit growers operating on the northern part of the island, according to the New Zealand Kiwifruit Growers, Inc. (NZKGI). That funding would assist in:


Photo court esy of Alvimann/morg uefile.
Because the disease is a weather event, the assistance is available for 12 months for the Rural Assistance Payment and 18 months for the welfare and other support from the date of the declaration, according to NZKGI. After that time, the government will review the time limits.

"Since PSA arrived in New Zealand two years ago, the kiwifruit industry has really banded together to support those affected by PSA. As a result, many initiatives are already in place to support the kiwifruit growing community," says Ian Greaves, a representative of Kiwifruit Vine Health, Inc., an organization established to minimize the impact of PSA on the New Zealand kiwifruit industry. "This includes counseling, pastoral care, well-being meetings and printed literature. However, up until now this has been funded from within the industry, and it has been a real challenge for everyone involved. Therefore, this important commitment from the government is very much a welcome relief."

Luckily, PSA has not entered the kiwifruit growing region of California - and if it does, perhaps the U.S. government will support the industry with its own funding.

There's a real possibility of the disease entering the U.S. "The risk is clearly there," says Boris Vinatzer, an associate professor in the department of plant pathology, physiology and weed science at Virginia Tech's College of Agriculture and Life Sciences in Blacksburg, Va. "It may have been imported already, but maybe it did not cause any disease yet," he continues. "It is also possible that the climate in California is not very conducive to the disease. It is a good thing that importation of all plant material had been stopped in 2010 after the disease broke out in New Zealand."

If PSA does enter the U.S., growers may feel better about it because of the research being done to slow down the pathogen's spread. Extension agents and researchers may be able to accurately diagnose it early enough to keep it in check.


When a plant is infected, PSA can cause red or white bacterial slime to ooze out from the branches and stems.
Photos courtesy of Giorgio Balestra, Tuscia University, Italy.

Tracing the bacterium

With the huge economic losses and the threat of the disease entering the U.S., university researchers believed it was vitally important to detect the bacterium. "Only by detecting the bacterium can we accurately diagnose plants and possibly eliminate them before the bacterium spreads to more plants," Vinatzer says. "Based on the bacterium's DNA sequence, we can also develop tests to test plant material or pollen that is being imported from other countries. However, because of this disease, the U.S. has stopped all import of kiwifruit plant material."

With the disease detected, Giorgio Balestra, a senior researcher in the department of agriculture, forestry, nature and energy at the Tuscia University in Italy, and his team "cultured bacteria from the plant, compared the bacteria to the ones from Japan and found that it was very similar. It was the same pathogen, but a different variant of it," Vinatzer explains.

Vinatzer and Balestra and their teams sequenced the entire deoxyribonucleic acid (DNA) of PSA bacteria from kiwifruit trees in China, Italy and Portugal. In addition, they analyzed bacteria from New Zealand, where the kiwifruit is almost a $1 billion industry. They found that DNA sequencing technology helped trace the origin of the pathogen back to China. In May 2012, they published their findings in a paper for the scientific journal PLOS ONE.

Vinatzer and Balestra were the lead authors of the paper; co-authors were Angelo Mazzaglia of Tuscia University and David Studholme with the University of Exeter in the United Kingdom. Others involved included David Guttman, professor with the University of Toronto in Canada; Nalvo Almeida at the Federal University of Mato Grosso do Sul in Brazil; Tokia Goodman, an undergraduate student at Virginia Tech; and Rongman Cai, a graduate student at Virginia Tech. Vinatzer's research was funded by the National Science Foundation. Balestra's research was funded by the Italian Ministry of Agricultural, Food and Forestry Policies.

"It causes very severe symptoms," Vinatzer says of PSA. "The whole kiwifruit plant can wilt, so it is relatively easy to detect. In Italy, it practically killed all yellow pulp kiwifruit trees. The green ones are a little more tolerant, but get sick also."

Leaf spots are seen first, and then branches begin to wilt. In addition, PSA causes a red or white bacterial slime to ooze out of the plant's stems and branches. Eventually the whole plant may die.


Pseudomonas syringae pv. actinidiae (PSA) can cause kiwifruit leaves and branches to wilt.

To find the root of the disease, researchers examined how the bacteria evolved from the same ancestor. They compared the DNA from the different bacteria to each other and found that the bacteria from China, Europe and New Zealand were almost identical. However, one small difference in one region of the DNA linked the New Zealand outbreak to the Chinese bacteria. Vinatzer, Balestra and their colleagues believe the bacterium was imported from China to Italy and from China to New Zealand, independently.

"The first step in stopping the spread of aggressive bacteria like PSA is knowing where they come from and how they have spread," Balestra explains. "Now that we have sequenced the DNA and found its likely origin, we can start to figure out ways to stop it and similar bacteria from doing so much damage in the future."

Vinatzer says the origin of the bacterium is likely China, but he believes more research needs to be done to confirm it. "We compared the DNA sequence of bacteria from China with those from Europe and New Zealand, and the bacteria in China were found to be more similar to the inferred ancestor of the bacteria in New Zealand and Europe," he says.

Besides having direct practical applications, the study will also lead to new insights into the adaptation of bacterial plant pathogens to crops. Vinatzer says this research "was the first step in identifying the source of the outbreak and how it spread from there. Reconstructing how it spread can help improve the measures in place to block the spread of other plant diseases and/or the further spread of this pathogen to the U.S."

Rocky Womack has written about agriculture and business for more than 25 years and currently serves as a contributing writer and correspondent for agriculture and business magazines, domestically and internationally.