3.7.59
FIVE-YEAR FIELD AND LABORATORY STUDY SHOWS ABSENCE OF CITRUS CANKER IN NORTH-WESTERN ARGENTINA

NV RAMALLO1, CJ RAMALLO1, LD PLOPER1, ML VERA2, V GONZALEZ1, G MUSLERA3, ML FONALLERAS2 and HM FIGUEROA4

1EEAOC, (4101) Las Talitas, Tucuman, Argentina; 2SENASA, Buenos Aires, Argentina; 3AFINOA, Buenos Aires, Argentina; 4SEAG, Tucuman, Argentina

Background and objectives
Citrus canker, caused by Xanthomonas axonopodis pv. citri, poses serious problems to citrus growers in northeastern Argentina (Provinces of Corrientes, Entre Rios, and Misiones). The disease severely affects the commercial value of the citrus crops in this region because it reduces the quality of fresh fruit for export. To date, the disease has still not been detected in the citrus growing areas of northwestern Argentina (NWA) (Provinces of TucumAn, Catamarca, Salta, and Jujuy). Lemon (Citrus limon) production is a very important activity in this region, especially in the Province of Tucuman, which produces, in 27,000 ha, 80% of the lemons of the country and was the second leading world exporter during 1997. In 1990, researchers sent by the European Union conducted a first survey for citrus canker in NWA, using visual inspection, indirect immunofluorescence, isolation in semi-selective media, and pathogenicity tests [1]. This survey revealed the absence of the disease and the causal agent in NWA. Due to the importance of citrus production for the economy of NWA, an extensive effort is currently being devoted to programs aimed at maintaining this status by preventing the introduction of citrus canker in the region. Two coordinated actions have been undertaken: a) check points at all sites of entry into the region to restrict the introduction of propagating materials, fruits, and used fruit containers (since 1991); and b) periodical area surveys to verify the absence of the disease by visual examination and laboratory analyses (since 1993). These actions are possible through the joint efforts of farmers, consultants, and officers of provincial institutions and the federal plant protection service (SENASA). The objectives of this study were to analyze the results of the first five years of the periodical survey for citrus canker in the Tucumn/Catamarca subregion and determine the status of the disease in NWA.

Materials and methods
The citrus subregion in Tucuman and Catamarea was divided into 5 areas, and samples were collected from each area following a systematic sampling approach. From 1993 through 1995, surveys were conducted Wce a year, during spring (October-December) and summerlautumn (February-April), following growth flushes. In 1996 and 1997, samples were collected only in December. A total of 5,786 samples have been processed during the 1993-1997 period. In the laboratory, leaves were blended using distilled water, and the extracts were analyzed by indirect immunofluorescence. In addition, aliquots of the leaf extracts were streaked onto a semi-selective medium. Pathogenicity tests, when necessary, were performed by applying droplets of the extracts to needle-puncture wounds on the lower surface of detached young leaves of grapefruit (Citrus paradisi) and lemon [2]. A similar procedure was carried out in the subregion of Salta and Jujuy.

Results and conclusions
Results of laboratory analyses were all negative for the presence of X. axonopodis pv. citriin Tucuman and Catamarca, substantiating the results of visual inspections of citrus groves in the region, which had revealed the absence of canker symptoms. Identical results were obtained for the Salta and Jujuy samples. The indirect immunfluorescence procedure, approved by the EPPO, U.S.A., and Chile for area surveys, proved to be very efficient for examining large numbers of samples. This technique has been shown to be more sensitive than ELISA for detecting bacterial pathogens in plant parts. Furthermore, this technique allows the viewing of the bacterial cells, therefore decreasing the chance of detecting faise positives. The specificity of these surveys was increased when the indirect immunfluorescence procedure was coupled with the isolation and pathogenicity tests. The results from this five-year study confirm that NWA is a citrus canker free area. Efforts will continue throughout this region to maintain such status and international recognition.

References
1. OEPP/Smith IM, ed., 1990. Bulletin OEPP/EPPO Bulletin 20, 263-272.
2. Randhawa PS, Civerolo EL, 1985. Phytopathology 75, 1060-1063.