Antarctic fungi found to be effective against citrus canker
Brazilian researchers have identified activity against Xanthomonas citri in 29 fungi isolated from samples collected in Antarctica; one of the compounds inhibited reproduction of the bacterium by up to 98 percent
A research team at the São Paulo State University's Bioscience Institute (IB-UNESP) in Rio Claro, Brazil, has identified 29 fungi with proven action against Xanthomonas citri, a bacterium responsible for citrus canker, an endemic disease in all citrus-producing countries. The origin of the fungi is surprising. They were isolated from samples of soil and marine sediment collected in Antarctica.
"These fungi live in isolated conditions and proliferate under inhospitable conditions including low temperatures and high levels of ultraviolet radiation," says Daiane Cristina Sass, a Professor at UNESP who heads a project engaged in a search for microorganisms that produce compounds with antibacterial action for use in agriculture, with support from the São Paulo Research Foundation - FAPESP.
"How have they adapted to survive in an environment so hostile to life? We wanted to see if they produced molecules with unique structures that protected them from infections and might therefore be capable of antibacterial action." Sass wrote an article published in Letters in Applied Microbiology - jointly with IB-UNESP colleagues Lara Durães Sette and Henrique Ferreira, among others - which shows some of the research's results.
More efficiency on fighting citrus canker
Although the bacterium can be combated in several ways, none is sufficient to eradicate the disease. Therefore, new chemical or biological methods of protecting citrus groves have to be pursued.
The disease is controlled directly by growers. The recommended measures include spraying trees with copper-based products and replacing infected trees with healthy new plantings derived from more resistant varieties. Control of the citrus leaf miner (Phyllocnistis citrella) is also advisable. The wounds made by larvae of this moth in feeding on the plant exacerbate citrus canker by serving as an entry point for X. citri.
"The main method for combating citrus canker consists of spraying trees with copper compounds. The downside is that when even small amounts are used for a long period, copper accumulates in the fruit, soil and water, eventually contaminating the entire environment. For this reason, we're looking for new compounds that are less aggressive to the environment and also less harmful to humans," Sass explained.
Collection and isolation of the Antarctic fungi
On the extent of the Sass-headed project and its research on biotechnology, the team came up with the idea of investigating the collection of fungi curated by Professor Sette, which resulted from Antarctic summer expeditions to the South Shetland Islands in 2013 and 2015 as part of Project Microsfera, conducted under the aegis of the Brazilian Antarctic Program (PROANTAR) and sponsored by the National Council for Scientific & Technological Development (CNPq).
Sette leads the project "Marine and Antarctic mycology: diversity and environmental application", also supported by FAPESP.
Sette isolated 33 filamentous fungi from samples collected in soil under rotten wood on Deception Island and 53 filamentous fungi from marine sediments at a depth of 20 meters in Admiralty Bay, King George Island. All fungal strains are kept at UNESP's Microbial Resource Center (CRM).
The FAPESP-funded research found that 29 of the 86 Antarctic fungi they isolated (19 of marine origin and ten terrestrial) contained compounds with proven action against X. citri.
Isolating the compounds produced by the fungi and verifying their antibacterial activity involved several stages. The process began with isolation of the fungi, which were then grown for several days in culture dishes with nutrients.
The fungi were cultured in liquid medium and shaken for 20 days at 15 °C. The solid biomass was separated from the liquid portion, and both parts were submitted to processing with solvents to obtain intracellular and extracellular extracts.
The researchers obtained 158 extracts. Each extract was diluted at several concentrations (2.10 mg/ml-0.02 mg/ml) and tested against X. citri. In the case of the soil fungi, most of the extracts with antibacterial action were intracellular in origin, while for the marine fungi, only the extracellular extracts hindered the bacterium's growth.
"We wanted to determine the lowest concentration of each extract that inhibited growth in 90% of cases," Sass said.
Some (12) of the extracts affected bacterial growth at lower concentrations than the highest tested, and ten of these inhibited growth in more than 90% of cases at concentrations of 1.5 mg/ml-1.0 mg/ml.
"At maximum concentration, one extract inhibited growth by up to 98%, and another inhibited it by about 80% at 0.52 mg/ml," Sass said. "It's important to note that we're talking about extracts [which contain varying amounts of molecules]. If an extract contains only one compound that's responsible for this bioactivity, the compound may display good antibacterial activity at much lower concentrations."
Twenty of the isolated fungi with action against X. citri belonged to the genus Pseudogymnoascus and were extracted from terrestrial and marine samples. Next came Penicillium (five), followed by Cadophora (two), Paraconiothyrium (one) and Toxicocladosporium (one), all extracted from marine sediments.
Having identified the extracts with action against X. citri, the researchers are now working to find out which chemical compounds give them this antibacterial capability.
"We expect to identify and purify some of these bioactive compounds, as well as to complete toxicology testing on them, within 18 months or less," Sass said.
The researchers plan to patent the compounds they identify. They also hope to persuade pesticide manufacturers to develop commercial products for combating citrus canker based on these compounds.
Twenty of the isolated fungi with action against X. citri belonged to the genus Pseudogymnoascus and were extracted from terrestrial and marine samples. Next came Penicillium (five), followed by Cadophora (two), Paraconiothyrium (one) and Toxicocladosporium (one), all extracted from marine sediments./h1>
- Author: Guy B Kyser
A recent request from the San Diego area has prompted the reposting of this blog by Guy Kyser, UC Davis Plant Sciences Specialist
A neighbor asked me to identify a robust perennial that keeps coming up in his garden. It had long, tropical-looking leaves and floppy racemes with small white flowers. This was a new one for me. Turned out it was common pokeweed (Phytolacca americana), a native of eastern North America. In the south some people eat it (poke salad), and a few southerners probably brought it west as a garden vegetable. But the whole plant is toxic if improperly prepared, so it's the fugu of weeds.
A couple of weeks later my daughter brought home a stalk of purple berries and asked if she could eat them. “No,” I said, “they contain numerous saponins and oxalates.” I began to wonder if there's more pokeweed around than I realized.
Then Gillies Robertson of Yolo RCD sent photos of a purple-berried plant found along a slough near Grimes. Common pokeweed again.
Pokeweed is in the Phytolaccaceae. This weed can grow to 10 feet tall. It dies back in winter then reemerges from the ground in spring, growing from a fat fleshy storage root. The leaves are large, 3 inches to a foot long and 1 to 5 inches wide, often with reddish stalks and lower veins. From August to October, pokeweed produces racemes of white flowers followed by reddish-purple berries. In its natural state, all parts of the plant, especially the root, are toxic to humans. Birds can eat the berries but sometimes act funny afterwards.
This plant can be found in most of the contiguous states. In drier regions, it prefers gardens and irrigated areas. Southerners with pokeweed experience suggest controlling it by digging up as much of the taproot as possible and/or by cutting off the stalks and painting the stubs with concentrated glyphosate (e.g. Roundup). Either way, treatments will probably have to be repeated until the plant's storage reserves are worn down. And it's a good idea to deal with pokeweed before it produces berries and seeds.
Since this is the first year I've seen it, and since I suddenly ran into it in three locations within a few weeks, I'm guessing that the common pokeweed population is expanding. This plant seems robust enough to cause some trouble if it becomes established in natural riparian areas.
- Author: Sara Garcia Figuera, Jennifer Reed and Brianna McGuire
Western Plant Protection Network at UC Davis
Early detection technologies (EDTs) are tests that indicate the presence of disease before signs or symptoms of the disease can be seen. In the same way that a doc-tor measures a patient's blood pressure to look for heart problems, a grower might use a trained “sniffer” dog to detect changes in a tree that looks healthy but has huanglongbing (HLB) disease. By using the EDT, the grower is able to uncover HLB earlier, and can decide on an early, cost-saving course of action.
In the case of HLB, there are many EDTs under development. Some of them look for patterns in the microorganisms that live on the citrus leaves (Leveau snapshot); some look for patterns in the chemicals that are produced by the tree in response to HLB (Pourreza, Davis and Slupsky); and others look for the molecules that the bacterium injects in the tree to cause disease (Ma). A description of some of these EDTs can be found on the Science for Citrus Health website.
Why do we need EDTs for HLB?
To understand why EDTs are needed and what their potential value is, it is necessary to understand the difference between the incubation period for a disease and the latent period. The incubation period is the time between exposure to the pathogen and the appearance of symptoms. The latent period is the time between exposure and the newly-infected host becoming infectious. Huanglongbing (HLB) has a long incubation period and a very short latent period, which means that a tree can be dis-eased for a long time without showing any visible symptoms, while being infectious for a large fraction of that time. Even if a tree does not seem diseased, it can serve as a home for the bacterium (Candidatus Liberibacter asiaticus, CLas) that causes HLB. If a psyllid feeds on the infected tissue of a tree (with or without symptoms), CLas that is present in the leaf tissue can be picked up by the insect and transmitted to other trees when the psyllid moves on to feed. Information from an EDT can help a grower detect the disease in a tree a long time before it would be detected by eye. This cuts down the time psyllids are able to feed on it and transmit the disease, slowing the spread of HLB to neighboring trees.
Why is it important to remove infected trees as early as possible?
If a tree that tests positive for CLas is not treated or removed, the bacterium will spread throughout the tree. Over time, an increasing proportion of the tree's tissues will become infected, increasing the chances that a psyllid will become infected upon feeding, and subsequently spread the infection to healthy neighboring trees. If the infected tree is removed, there is no opportunity for psyllids to feed on the infected tissue and spread the disease. Once CLas is detected, tree removal is the only surefire way to prevent the spread of the infection, and it is extremely time-sensitive. The sooner an infected tree is removed, the lower the chances that psyllids will get infected. The savings associated with early infected tree removal will be proportional to the amount of surrounding trees that would have been infected with CLas due to that tree, and the number of months that it would be left on the ground.
Who is working on the project?
Several research teams in different universities and research stations, supported by a variety of funding organizations, have been working on the development of a variety of EDTs. These EDTs, designed under laboratory and greenhouse conditions, are being validated under field conditions in Texas and Florida. In California, where HLB has not been detected in citrus orchards, samples of different citrus varieties have been collected from healthy trees and trees affected by other diseases from all over the state. These samples are being used to calibrate the EDTs, and to test if they can distinguish between healthy and HLB-diseased trees, and between HLB-diseased trees and trees affected by other common citrus diseases. Dr. Neil McRoberts and his team at UC Davis are evaluating the data from these experiments and providing support to the EDT researchers.
What are the challenges and opportunities?
Currently, regulations require HLB infected trees to be removed if a certain amount of CLas DNA is detected in leaf samples through polymerase chain reaction (PCR). However, CLas is unevenly distributed in the sap of citrus trees, and the leaf samples collected might not be PCR-positive even though the bacterium is already present elsewhere in the tree. EDTs offer the possibility to detect infected trees before they are PCR-positive, so they could be removed earlier in the HLB epidemic. Therefore, the value of EDTs relies on the voluntary removal of EDT-positive trees before the law requires them to be removed.
No EDT gives perfect diagnostic results. Sometimes healthy trees will produce EDT scores that look like diseased trees (so-called “false positives”). Removing such trees will result in an immediate financial loss. However, because the economic damage caused by leaving an infected tree in place is much bigger than the value of a healthy tree, using an EDT to guide decisions has the potential to result in a long-term economic benefit to individual growers and communities, by reducing the spread of HLB. Losing a few healthy trees along the way is the unavoidable cost of stopping the disease from spreading. Like-wise, some trees will seem healthy based on EDT scores but might end up showing symptoms (“false negatives”). The proportion of true positives, false positives, true negatives and false negatives represents the accuracy of a diagnostic test. Dr. McRoberts' team is analyzing the accuracy of the EDTs, and preliminary results suggest that the best performing EDTs could be correctly determining the status of the trees 95% of the time.
The results of this analysis could be used to foster the adoption of EDTs among the citrus grower community, promoting the idea that the sooner infected trees are detected and removed, the smaller impact HLB will have on California's citrus production. Unless there is sufficient cooperation in integrated management of HLB by removing infected trees as early as possible, controlling the ACP on an area-wide scale, and using certified plant material, the California citrus industry is likely to suffer un-sustainable economic losses to HLB.
So, this weekend we had some hot weather and the damage from that heat is apparent in all kinds of plants. Sycamores, cottonwoods and willow in the Santa Clara River bottom look torched. Redwoods in the landscape look like a new disease has hit them.
Even old coast live oak in Ojai have been toasted. Orchards have been hit also with been hit without exception. This has been a widespread weather phenomenon like a major freeze. And the trees should be treated as if they have been freeze damaged.
So, what to do with the avocados and citrus that have been hit? Well, if it's just a slight toasting, nothing. They will grow out of it. It's a setback. The growing points, the terminal buds, have been damaged and in the case of avocados those may not flower next spring. If the damage is not extensive, the whole canopy has not been damaged, then flowering should be sufficient for a good crop next year. If the whole canopy has been hit, it's likely that flowering will be minimal next year.
If the trees have lost significant portions of the canopy, though, the heat damage is not the problem, it's the sunburn damage that is going to happen that is the problem. It's the loss of the leaves that transpire and cool the tree that lead to this kind of damage that can kill small trees and lead to significant branch loss in older trees.
The leaves act like the radiator in a car. They move water through the tree and that water movement carries off the heat that accumulates in the branches and stems. When water flow stops, the bark heats up and tissue is damaged. The worst-case scenario occurs when a “renovated” tree that has been brought down to 6 feet in January and since then there has been new growth all over the tree. The heat fries that new growth and now the whole tree structure is exposed to sunburn damage.
The branches exposed to the sun need to be protected with whitewash. The whitewash needs to be WHITE, not grey. It needs to be able to reflect the sun and prevent the surface from heating. The tops of branches and the west and south sides need to be the most protected, so it often involved hand work. And it needs to be done soon after the canopy loss. That wood heats up fast and damage occurs soon after it heats up.
So what else needs to be done? No canopy, no water loss, so it's necessary to manage the water differently. With no leaves, there is no water moving from soil through the tree, so it just sits there, and the ground stays wet. Perfect conditions for root rot.
Growers who were watering their trees knowing that a heat spell was coming, did the right thing. It probably reduced the severity of the damage, but even growers who had water on before the heat and it was running during the heat have had damage. With canopy damage and loss, applied water needs to be restricted to just enough to get tree recovery without creating a wet, soggy condition. And with tree recovery, it's going to need a continually changing irrigation schedule as new growth occurs.
So now more than ever, water to the tree's growing needs. And the normal fertilizer program needs to be adjusted. There's probably sufficient nutrients in the soil from prior fertilization that nothing new needs to be applied.
And don't' prune the trees. Leave the hanging leaves there. They will help protect the tree from sunburn, but the extent of the damage is not clear. Let the tree push new growth and that will tell you sometime in the future 3-6 months, even a year from this event, when to do significant pruning.
Phlood, Phyre, Phrost, Fytophthora and Phahrenheit continue to plague our industry. It seems like we are always coping with some natural and some unnatural issues affecting agriculture. Oh, yeah and pH.
Photo: Heat singed new avocado growth.