Farm Progress

PCR test remains industry-standard for determining disease"Clustering" theory developed by USDA researcherCalifornia officials watching several "hot spots" for disease

tfitchette, Associate Editor

January 27, 2016

10 Min Read
<p>From left, citrus industry experts: David Bartels, entomologist, USDA APHIS; Mike Irey, director of research and business development, Southern Garden Citrus, Florida; Neil McRoberts, associate professor, UC Davis; and, Carolyn Slupsky, professor, UC Davis, spoke&nbsp; on various issues centering on Huanglongbing at a summit in Visalia, Calif.</p>

For California citrus growers, trying to stay ahead of an insidious disease that is difficult to detect, easily spread and has a record of destruction in other states is likely their only short-term hope.

Meanwhile, time is not their friend.

A researcher with the U.S. Department of Agriculture in Texas says the same ominous trend witnessed in Texas ahead of its Huanglongbing (HLB) explosion is now being observed in California.

David Bartels is an entomologist with the U.S. Department of Agriculture in Edinburg, Texas. Among his studies is the Asian citrus psyllid (ACP), the only known insect to vector HLB. Bartels believes that California could be on the brink of an HLB explosion similar to what is currently being seen in Texas.

At a meeting with citrus growers and industry officials in Visalia in early December, Bartels placed an emphasis on qualifying statements, staying away from solid predictions of an upcoming HLB outbreak in California.

According to Bartels, Texas saw a rapid rise of HLB-positive psyllids over period of two years leading up to nearly 2,000 HLB discoveries in the Rio Grande Valley.

“You have to make a lot of assumptions there, but if you want to make the assumptions that the conditions are the same you might see this in California in the near future,” he said.


Prior to discovering HLB in Texas Bartels said officials there were sampling psyllids and plant material as fast as they could. This was on the heels of Florida’s HLB outbreak and expansion of the disease throughout that state’s entire citrus belt.

In Sept. 2008 Texas officials began surveying a large area covering about 17,000 sites. Inspections happened about twice a year, he said.

From these they would collect over 10,000 psyllids a year that were tested for the Candidatus Liberibacter asiaticus (CLA) bacterium responsible for causing HLB. Industry-standard polymerase chain reaction (PCR) tests were used to determine whether the psyllids had the bacteria.

The PCR test is a diagnostic exam that amplifies a short segment of DNA, which can be sequenced and determined to have the CLA bacterium. The figure generated from this test is called the “cycle threshold,” or cT score.

Bartels explains that high cT values (larger numbers) have fewer copies of DNA in them than lower values. For instance, a cT value of 38 will have one CLA bacterium in it while a cT value of 17 has 3.2 million bacteria.

For regulatory purposes, a cT value of 40 or higher is considered negative for the bacteria that cause HLB while a score of 32 or lower is positive.

It’s those scores between 32 and 40 that present questions and concerns for Bartels as DNA copies of the bacteria are present in scores below 38, but not in high enough numbers to make a regulatory determination of HLB.

Bartels’ clustering theory came into play when he discovered data in Texas that showed clusters of cT scores in the high 30s from captured psyllids just before HLB was discovered in the same general area. By studying the data collected in Texas, he says there appears to be a correlation between cT scores in this “grey zone” and later discoveries of HLB in plant material.

The same thing is now being seen in California, particularly in the Los Angeles basin, which is what led to HLB discoveries in the San Gabriel Valley last summer.

San Gabriel

Part of the California Department of Food and Agriculture’s ACP/HLB protocols is to collect live psyllids for testing. Last summer those tests revealed psyllids with cT scores of 37 in the San Gabriel area, according to Bartels. That opened the door to closer inspections of plant materials and the ultimate discovery of almost a dozen HLB-positive trees in the city of San Gabriel.

California’s first HLB-positive tree was discovered in nearby Hacienda Heights in 2012 in generally the same manner: positive psyllids were discovered which led officials to a single tree that had the disease.

Bartels suspects there may be more HLB trees in the region as CDFA samples of psyllids from the area are showing cT scores in the high 30s – not enough to declare a regulatory negative result, but not yet positive either.

At a citrus grower meeting earlier in the summer, citrus program manager for the CDFA, Victoria Hornbaker, said PCR tests of psyllids captured in the San Joaquin Valley community of Farmersville revealed a cT score of 38.6.

For officials, the needle-in-the-haystack search for HLB-positive trees in southern California is daunting. There can be multiple citrus trees in nearly every yard in the Los Angeles region and visible symptoms of the disease can be difficult to spot or sometimes confused with nutritional deficiencies in a tree.

While small and misshapen fruit can be obvious to spot, it can take over a year for trees to show those kinds of symptoms. Before then visible symptoms can be limited to single branches as the disease travels in the tree’s phloem.

Bartels says the work is complicated because psyllids travel and trees do not. In Texas his discovery of questionable psyllids tended not to be exactly over the HLB finds, which led him to believe that prevailing winds in the area could have blown infected psyllids around.

High wind conditions at certain times of the year in the Los Angeles basin are suspected of relocating psyllids throughout the region.

Message to growers

The same citrus meeting in which Bartels spoke included Mike Irey, director of research and business development with Southern Garden Citrus in Florida.

During his presentation Irey painted the HLB picture in Florida for growers in the audience, telling them that nothing short of aggressive sampling should be tolerated in California.

Irey spoke briefly to the topic of early detection technologies. Currently Florida citrus growers are being inundated by marketers trying to sell new technologies and nutritional supplements that range from the practical to the ridiculous.

California Citrus Mutual is warning its members of these efforts, cautioning growers not to assume every new product or technology will help them.

Texas is currently on the path of repeating the Florida experience, Irey said.

Florida citrus production is down from a high of 150 million boxes to about 74 million boxes, Irey says. Growers there are no longer removing HLB-infected trees, opting instead to blend fruit when making juice.

Irey’s doom-and-gloom message to growers was aimed at encouraging them to survey their groves aggressively for the psyllid and for signs of HLB. According to Bartels, Texas research reveals young groves to be more susceptible to HLB as they tend to be more attractive to the psyllid because of their almost-constant new growth.

Irey and Bartels recommend regular surveys of trees on the outer boundaries of groves. Young groves that flush almost constantly need to be inspected throughout the block while mature groves with more closed canopies and growth that is closer together can have inspections limited to rows on the exterior of the grove and along wide alleys.

Irey cautions California growers to not assume that current psyllid management area protocols will solve the problem. They may slow the spread of HLB, but they won’t stop it, he said.

He also warned California growers that once HLB takes hold in commercial citrus expect production costs to possibly double as quality and production decline.

“Don’t think it’s going to be easy or that you can manage your way out of these HLB losses,” he said.

Ivey shared the following opinion based on experience as to what California growers can do in the near term:

  • Stay in eradication mode for the ACP as long as possible;

  • Reduce the inoculum early. Pull trees and continue with aggressive treatment options; and,

  • Sample your groves – the more the better.

He further recommends growers not wait for new early HLB-detection methods or better control methods for the ACP. Use the current tools available and use them all, he says.

Moreover, Ivey recommends focusing on tree health and minimizing tree stress, which he said in his experienced helped. While HLB cannot be cured through good tree health or nutritional supplements, it can possibly stretch the time an infected tree produces marketable fruit.

Early detection

Carolyn Slupsky, a University of California, Davis professor working on early detection methods for HLB, told the audience that it could be five to 15 years before growers have useful early-detection methods for HLB.

By early-detection, scientists mean the ability to find questionable trees that may not yet have visible and outward signs of HLB, or will not yet test positive for HLB using the industry-standard PCR test.

During her discussion she did highlight research using dogs to sniff groves for volatiles that may be present within HLB-infected trees. Of the different ideas out there, using dogs seems to be the closest early-warning technology that could become available to citrus growers in the near future.

One of the struggles she points to with HLB is how the bacterium can be unevenly spread throughout a tree. Testing the wrong part of the tree can lead to a false negative, even with the current PCR method.

Neil McRoberts, an associate professor of plant pathology at UC Davis says that as early-detection methods are developed these will not be regulatory in nature, but will be tools growers can employ to make management decisions up to, and including the removal of suspect trees.

California currently has the authority to pull citrus trees, including those from commercial groves, that test positive for HLB.

Moreover, McRoberts points to already-known symptoms for HLB and good research growers can lean on to make management decisions today to protect their groves.

“We’re not talking about regulatory decision-making,” he said. “We’re talking about private decision making in your own groves.”

McRoberts says some areas in the San Joaquin Valley already have measurable risk factors, which is where he recommends growers begin looking for the disease.

Earlier this year McRoberts sampled grower and PCA opinions at various meetings in California. By asking a series of questions at citrus grower meetings in Palm Desert, Ventura and Exeter, McRoberts was able to gauge the opinions of growers on issues including how important they thought HLB was as an issue and their thoughts on it being found in their region.

For all the news about ACP/HLB, McRoberts found that growers attending the Palm Desert gathering thought it was more likely HLB would be found in their region than did growers in Ventura and Exeter. For all the media attention on the issue, he said growers in southern California stood up and took notice of the disease when it first appeared in California while growers in Ventura and the San Joaquin Valley remained a bit more skeptical that the disease would be found in their region.

He warns that not adhering to quarantine regulations “is an effective way to take the disease and put it where it wasn’t once located.”

He also looked at data of how the ACP spread in California, particularly in the San Joaquin Valley and Bay Area and found that it was along major highway corridors where the pest was being relocated.

McRoberts urged growers to participate with voluntary psyllid management area protocols as a tool to control ACP populations.

Moreover, he cautions against divisive attitudes where large grower operations ignore the needs of smaller operations.

“We saw this in Florida … the big guys cannot afford to let the little guys sink,” he said.

What happened there, and can be seen in southern California, is the abandoning of citrus groves by growers. Just as bad are the city parks in southern California with citrus that are untreated and become host to numerous citrus pests, including the ACP. This leaves weakened trees with the HLB inoculum there to be spread by psyllids.

About the Author(s)


Associate Editor, Western Farm Press

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