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Protecting passion fruit

New early diagnostic kit for bacteria that devastates orchards

CARLA MUNHOZDNA sequence of the pathogen used to detect Xanthomonas in passion fruitCARLA MUNHOZ

An edible fruit used in juices, jams, syrups and cosmetics, passion fruit does it all, even it’s rind is used to make flour. Brazil is its largest producer and consumer of passion fruit in the world, having produced more than 700,000 tons in 2009, but the industry has seen significant losses in orchards due to a disease that is difficult to control and widespread in its occurrence – passion fruit blight damages the plants leaves and renders the fruit unfit for consumption. The bacterium Xanthomonas axonopodis is responsible for this disease that attacks passion fruit plants and causes producers to lose 20% to 30% in yield.

“Since there is no efficient means of chemical control and eradication of the disease, when it spreads widely enough, whole orchards have to be eliminated, “says Professor Maria Lúcia Carneiro Vieira of the University of São Paulo (USP). She and her doctoral student, Carla de Freitas Munhoz, developed an early diagnosis kit that indicates the presence of the bacteria when it still in the early stages of infecting a passion fruit plant, making it more possible to prevent the spread of the pathogen and slow its progression. With funding from FAPESP, the work included the participation of researchers from the Agrony Institute of Campinas (IAC) and the State University of Londrina (UEL). The initial study investigated the genetic structure of populations of the pathogen in Brazil. For this purpose, bacteria were collected in orchards in the Southeast, South and Central Regions of the country.

“From this we collection we obtained 87 bacterial isolates,” says Maria Lucia from the Department of Genetics at Luiz de Queiroz College of Agriculture in Piracicaba, São Paulo State. Each isolate corresponds to different line of the disease and that has not yet been studied genetically. An analysis of genetic variability was conducted to assess whether there were identifiable differences between the bacteria samples, according to their region of origin. “There are differences, yes. They are associated with the region where they were collected,” she says. Samples collected in São Paulo, for example, are genetically more similar to samples from Minas Gerais than to those collected in Paraná. With this information, a comparative study was possible. “We did an analysis of the genome of the Xanthomonas axonopodis affecting passion fruit and compared it to isolates of different types (or partoviruses) of the same bacteria that attack other plant species such as grapes, garlic, manioc and beans,” she says. “We discovered the presence of a nucleotide base that distinguishes isolates of the bacterium that attack passion fruit from all of all others.” Encouraged by this finding, the researchers were able to develop a molecular method based on the polymerase chain reaction or PCR – a technique used to make multiple copies of a specific DNA fragment – to develop a diagnostic kit for detecting the presence of Xanthomonas even before a plant begins to show symptoms of disease.

“A small piece of a leaf is enough to make the diagnosis, and the test can be applied in plant nurseries and orchards,” says Maria Lucia. You don’t need to take the samples back to a lab in order to run the diagnostic test. It is sufficient to simply mail samples in for analysis. The cost of testing each sample will be around R$3.00. A very small investment compared to the losses suffered when an entire plantation must be eliminated. “Apparently there are no orchards resistant to this bacterium, which attacks both the tart and sweet varieties of passion fruit,” says the researcher, who has been studying the problem using different molecular genetics techniques since the 90’s. The rapid spread of the disease makes it extremely difficult to control, particularly since the bacteria can be spread by the wind, acquired from contaminated farm tools and new seedlings introduced to an orchard. In citrus trees, like oranges, Xanthomonas causes a sort of cancer in the fruit. In passion fruit, it makes the skin of the fruit oily and dark yellow in color, tending towards brown. The fruit eventually rots while it is still on the vine. “When the disease reaches the fruit, the vine has already been contaminated.” Tart passion fruit represents about 97% of the total planted area and volume sold throughout the country. The Northeast Region of Brazil accounts for 73% of all national production of the fruit, according to data from 2009 compiled by the Municipal Agricultural Production at the Brazilian Institute of Geography and Statistics (IBGE). The main buyers of fresh fruits are Argentina and Uruguay, while the fruit juice concentrate is preserved and sold to many European countries.

The growing of passion fruit occurs mainly on small family farms. Because of this, the idea of the researchers is to offer the diagnostic kit to juice manufacturers or plant nurseries. In this way, infected seedlings can be eradicated and the small producers, who supply the fruit to the juice processing industry, will ultimately benefit from the reduction in losses. Large nurseries also will benefit from the practical application of such test results because they are the main suppliers of seedlings for the whole country. “Upon buying a new stock of seedlings, the bacteria may be present and go undetected by just a visual examination,” says Maria Lucia.

The Project
Study of the molecular diversity of Xanthomonas axonopodis pv. passiflorae using AFLP and REP-PCR markers and its diagnosistic applications (nº 2008/58494-0); Modality RegularAid Research Project; Coordinator Maria Lucia Carneiro Vieira – USP; Investment
R$ 126,439.46 (FAPESP)

Scientific article
MUNHOZ, CF et al. Genetic diversity and PCR-based method for Xanthomonas axonopodis detection in passion fruit. Phytopathology. V.101, n. 4, p. 416-24. April 2011.