by Sarah Yang
BERKELEY - During World War II, soldiers from the Fifth U.S. Army set up camp at an exclusive hunting estate in Italy, regrouping between military drives north against German troops and fascist leader Benito Mussolini. Sixty years later, forest pathologists are pointing to huge gaps of dead trees in the estate as the visible reminders of that brief stay.
In a new study published in the April issue of Mycological Research, researchers at the University of California, Berkeley, and in Italy, have unlocked the mystery of how the destructive Heterobasidion annosum pathogen could have spread to the Presidential Estate of Castelporziano, which has been sealed off from the public for centuries.
They were able to trace the origins of the pathogen back to eastern North America, where U.S. troops departed for Europe during World War II. The researchers say the pathogen likely hitched a ride in transport crates, pallets or other military equipment made from untreated lumber from infected trees. It took decades for the pathogen to establish itself, but since symptoms were first noticed in the 1980s, the root fungus has wiped out large swaths of stone pine trees in the Castelporziano estate less than 15 miles southwest of Rome.
"The spread of exotic plant and tree diseases is not new, but this is the first evidence of a pathogen being introduced into a different continent through military activity," said Matteo Garbelotto, an adjunct assistant professor of ecosystem sciences at UC Berkeley's College of Natural Resources and co-author of the study.
"Quarantines and regulations already exist to guard against the introduction of pests and pathogens from lumber brought in through commercial and other sources, but there is no equivalent standard for lumber brought in by the military," said Garbelotto, who is also a UC cooperative extension specialist in forest pathology. "This study suggests that when planning military operations abroad, there is a need, and a responsibility, to check for potential micro-organisms that could be introduced to foreign lands, and to take measures to prevent them from spreading."
Control of the nearly 15,000-acre estate has passed through the hands of various leaders in Italy's history. For centuries it was used as a hunting ground favored by the popes before becoming the exclusive property of the king by 1870. After World War II, when Italy voted to dissolve the monarchy to become a republic, the estate was turned over to the president, but public access remained restricted.
"The estate is of naturalistic importance because it is a pristine example of a Mediterranean forest," said senior author Paolo Gonthier, assistant professor at the University of Torino's Department of Exploitation and Protection of the Agricultural and Forestry Resources. "Except for a few eucalyptus trees, the estate contains exclusively native Italian flora."
In the mid-1980s, estate staff started noticing symptoms of infection in the stone pines. They did not know what was causing the die-off at the time, but the disease spread quickly, killing several hundred trees throughout the forest.
"There is a gap in the forest of about 100 meters from where trees were cut down because they were dead from infection by the fungus," said Gonthier. "There are now just a few isolated stone pine trees that remain in the gap."
In an effort to find the culprit, the investigators combined cutting-edge DNA analysis an extensive search through historical documents and interviews with staff members at the preserve.
Gonthier and his colleagues collected samples of the pathogen from fertile fruiting bodies, allowing researchers to obtain pure cultures for analysis.
But when the researchers analyzed the genetic material, they unexpectedly found a telltale insertion of a piece of DNA in the mitochondria known to be present in North America, but not in Europe.
Researchers in Garbelotto's lab had just finished constructing a genetic database for H. annosum, enabling them to narrow down the region where the samples taken from Castelporziano originated. They sequenced four different genes, all of which matched the DNA of North American species. Two of the four matched eastern North American species, and one matched a species in the Southeast region of the United States.
Further genetic analysis revealed that the samples they collected from the woodlands each had different genotypes, indicating that the population was several generations removed from the "founding" pathogens that originally settled the estate.
"At this point, the H. annosum population at Castelporziano is indistinguishable from a natural population in the eastern United States, which indicates that it has become well-established," said Garbelotto. "DNA analysis also indicates that the Castelporziano population is diversified and is made up of many different individuals, as it is normally seen in the wild in the United States. This process takes decades, so we know the infection was not a recent occurrence."
What the researchers did not know at the time, however, was how the non-European pathogen could have ended up in an Italian estate that saw very few visitors, both foreign and domestic. They ruled out the introduction of an infected plant since exotic species had not been planted in the forest, except for a small group of eucalyptus trees that are far from the areas of infection.
"The only other possible mode of transport is through untreated lumber," said Garbelotto. "This pathogen is not that easy to move. It doesn't move through soil, and its airborne spores are short-lived. And until now, the only documented transport of this pathogen was on the scale of a few kilometers, certainly not thousands of miles."
The connection to military activity came when researchers learned from interviews with estate staff that the U.S. Army had set up an encampment on the grounds during World War II. A search through accounts of military history confirmed that, for at least one month in the summer of 1944, the 85th Infantry Division, part of the Fifth U.S. Army, stayed on the grounds shortly after capturing Rome.
"There is little doubt in my mind that the source of the pathogen was the U.S. Army," said Garbelotto. "Everything matches - the timeline and location of infection, plus the region of origin of the fungus."
Garbelotto said the findings support a common suspicion among plant pathologists in Europe that certain diseases in that region are linked to U.S. Army bases. "The sources of tree diseases that have afflicted Europe this past century, including Chestnut blight, often seem to be near U.S. military bases, but they had no way of proving the link," said Garbelotto. "This study is as close to a link as we've gotten."
He said the lesson in this is that when there is an organized mass movement of people and equipment through unregulated military channels, the introduction of microbes is a real risk.
"This study suggests that self-imposed regulations may be a useful implementation by the military," said Garbelotto. "The effects of the introduction of exotic microbes may not be evident for decades, and by then eradication of the exotic organism may be impossible."
The researchers are not sure how they can protect the remaining stone pine trees in Castelporziano, but they are watching to see if the pathogen manages to spread beyond the estate. While the pathogen infects a number of plants, it does the most damage to pine, juniper and cedar trees.
"Right now we know that this pathogen is virulent on Italian stone pines, but we don't yet know if it has affected other pines or broad-leaf species in Europe," said Gonthier. "When tourists visit the region, they expect to see these stone pines. It's the landmark tree for the Mediterranean coast. We need to ensure that this exotic pathogen does not spread to regions outside the estate."
The other authors of this study are Rachel Warner of the UC Berkeley Department of Environmental Science, Policy and Management; and in Italy, Giovanni Nicolotti of the University of Torino Department of Exploitation and Protection of the Agricultural and Forestry Resources, and Angelo Mazzaglia of the University of Studies of Tuscia Department of Plant Protection.
This research was conducted with the cooperation of the Technical and Scientific Commission of the Presidential Estate of Castelporziano.
NOTE: To reach Matteo Garbelotto, call (510) 643-4282 or e-mail firstname.lastname@example.org.