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Distribution of Citrus Tristeza Virus in Grapefruit and Sweet Orange in Florida and South Africa*, ** R

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Distribution of Citrus Tristeza Virus in Grapefruit and Sweet Orange in Florida and South Africa*, ** R Distribution of Citrus Tristeza Virus in Grapefruit and Sweet Orange in Florida and South Africa*, ** R. F. Lee, S. M. Garnsey, L. J. Marais, J. N. Moll, and C. 0. Youtsey ABSTRACT. The distribution of citrus tristeza virus (CTV) in field trees was determined by enzyme-linked immunosorbent assay on individual flushes collected from citrus trees in Florida and South Africa. In Florida, most CTV isolates were unevenly distributed in grapefruit trees, especially in late summer. However, one mild isolate with some cross-protecting ability (T26) was consistently distributed throughout the trees regardless of season. Citrus tristeza virus was more evenly distri- buted in sweet orange trees in Florida, but occasionally very young flush tissue was found CTV-free. In South Africa, CTV was found evenly distributed throughout both grapefruit and sweet orange trees. South African isolates were highly invasive in recently inoculated plants. The even distribution of CTV within trees is a trait which may be important for effective cross protection by mild CTV strains. Index words. ELISA, cross protection, stem pitting. Citrus tristeza virus (CTV), a of effective cross protection. Selection member of the closterovirus group, is of cross-protecting CTV strains has the most economically important cit- thus far been an empirical process. rus virus worldwide (1). There are The distribution of the mild strain of many strains of CTV which differ in virus within the plant host may play their biological activity (1, 3). Quick an important role in its ability to cross decline of trees on sour orange protect the host against subsequent rootstock induced by CTV has killed challenge by severe isolates (9). millions of trees worldwide, but this We used the enzyme-linked im- aspect of tristeza can be controlled by munosorbent assay (ELISA) to study the use of CTV-tolerant rootstocks. the distribution of CTV within Strains of CTV which cause severe grapefruit and sweet orange trees in stem pitting on scions cause losses Florida where severe stem-pitting even in trees on CTV-tolerant strains of CTV are not yet present rootstocks. For these severe stem pit- and in trees in South Africa where se- ting strains of CTV, the use of mild vere stem pitting strains are endemic. strains for cross protection is proba- Results are presented in this paper bly the most effective control strat- and discussed in relation to cross pro- egy (5, 7). With mild strain cross pro- tection success. tection, a plant previously infected with a mild virus strain will show in- METHODS AND MATERIALS terference or delay in the expression Virus isolates. For some experi- of severe symptoms after being chal- ments, Marsh grapefruit trees on lenge-inoculated with a severe strain sour orange rootstock were inocu- of the same virus. Severe stem-pit- lated with the previously charac- ting strains of CTV have been con- terized CTV isolates T3, T4, or T26 trolled successfully by use of mild (11). These trees were in an experi- strain cross protection in Brazil (5) mental field plot in central Florida. and in South Africa (7). Isolate T3 causes decline of sweet While mild CTV strains are com- orange on sour orange rootstock and mon, few of these strains are capable causes seedling yellows (SY). Isolate *Florida Agricultural Experiment Station Journal Series No. 7952. **Mention of a trademark, warranty, proprietary product, or vendor does not constitute a guaran- tee by the U.S. Department of Agriculture and does not imply its approval to the exclusion of other products or vendors that may also be suitable. Tenth IOCV Conference T4 causes strong stunting, stem pit- tember, and February. Similar re- ting and vein clearing in Mexican petitive assays were made from four lime, but no decline of sweet orange sweet orange trees. Some trees in- on sour orange and no SY. Isolates fected with naturally-occurring CTV T26, T32, and T55 produce mild strains were sampled only one time symptoms and little stunting on Mex- by collecting the indicated number of ican lime, and cause no decline of flushes from throughout the tree sweet orange on sour orange, or SY. canopy. Root sprouts were forced by Isolate Ti33 is an uncharacterized exposing and cutting some of the CTV field strain which produces mod- roots in an 80-year-old grove of Marsh erate vein clearing and stunting on grapefruit on rough lemon rootstock Mexican lime. located in central Florida. About 8 The Nartia mild, Bolton severe, weeks later, new flush tissue was col- and Nkwalini mild CTV isolates from lected from the forced root sprouts South African have been previously and also from new flushes on the described (6, 10). The CSFRI CTV grapefruit scions. strains from South Africa were natur- Tissue sources-South Africa. ally-occurring field selections which Twenty-five flushes were collected did not cause decline of sweet orange from individual trees from ll-year-old on sour orange rootstock. Isolate Nartia grapefruit in the Nkwalini GFSS-1 is a severe strain of CTV Valley, Natal Province, and from 10- which severely stunts grapefruit, and year-old Rose grapefruit propagated GFMS-10 produces moderate stem from the Bolton budwood source. pitting and mild stunting on grape- Samples were also collected from 5- fruit (12). year-old seedling Marsh grapefruit Sample collection and enzyme trees on an experimental plot near linked immunosorbent assay Malelane, Transvaal Province. (ELISA). Samples of young flush Twenty-five samples were collected which had the basal leaves at or near from each of five trees infected with full expansion were collected from three different CTV isolates in the field trees and stored in plastic bags latter location. at 4 C until processed for ELISA, A greenhouse experiment was set usually within 1-7 days. A 0.25-gm up in South Africa to determine the aliquot of chopped bark tissue was uniformity of CTV infection within homogenized in 5.0 ml of 0.05 M Tris- the inoculated plants 4 weeks after HCl buffer, pH 8.0. The double anti- graft inoculation. Two large Marsh body sandwich ELISA procedure (2) grapefruit seedlings and two Mexican was used with antisera prepared lime plants were inoculated using leaf against unfixed CTV (4). These anti- pieces infected with Florida isolates sera have reacted to all CTV isolates T55 and T32 and South African iso- tested. Healthy tissue gave OD405 lates Nartia, GFMS-10 and GFSS-1. readings of 0.02-0.03, and values After 4 weeks, the young growth twice this were considered as the from four different branches of each threshold for a CTV-positive reac- plant was collected for assay. tion. Most OD values were 0.4 or greater; only rarely were OD values RESULTS less than 0.4 in the samples declared CTV positive. Distribution of CTV within Tissue sources-Florida. Thirty grapefruit trees in Florida. In a pre- individual branches were labeled on liminary test, 30 flushes were col- Marsh grapefruit trees systemically lected from each of two 12-year-old infected with CTV isolates T3, T4, Marsh grapefruit trees on rough T26, and Ti33. Samples of new flush lemon rootstock during the spring and tissue were collected from the same were assayed by ELISA. Only eight individual branches in July, Sep- of the flushes from one tree and 12 Tristexa and Related Diseases from the other reacted positively for In another study, 22 Pineapple CTV. In a test of the scions and the sweet orange trees on sour orange rootsprouts from an 80-year-old grove rootstock were sampled at three in- of Marsh grapefruit on rough lemon tervals. The first collection was Janu- rootstock, CTV was present in both ary 1985, after a severe freeze had de- the scion and rootsprouts of 21 trees, foliated the trees and the new flush in only the scion of seven trees, and was about 2-4 cm long. Only eight of in only the rootsprout of six trees. It the 22 trees were found CTV positive was not detected in either scion or using a sampling of 25 flushes from rootstock of nine trees. each tree. Subsequent collections of The apparent erratic distribution five flushes from each tree were made of CTV within grapefruit trees was in April 1985 and August 1986, and examined more closely by individually all flushes from all trees tested posi- labeling 30 separate branches on four tively for CTV infection. 6-year-old Marsh grapefruit trees, Distribution of CTV in grape- each inoculated with a different CTV fruit and sweet orange in South Af- strain. Young flush tissue was col- rica. Several grapefruit and sweet lected for ELISA from each tagged orange trees were tested for the pres- branch in summer (July), late summer ence of CTV by collecting 25 flushes (September), and during spring flush from around the canopy of each tree. (February) (table 1). Isolate T26 was A list of the trees tested and the sev- well distributed throughout the tree erity of the CTV symptoms is given with 90% of the branches positive on in table 3. All individual flushes from all three sampling dates. Isolate Ti33 all trees tested in South Africa as- was very unevenly distributed sayed positively for CTV. throughout the tree. Only 53% of the The movement of Florida and branches indexed CTV positive in South African CTV isolates into July, while 37% were positive in Sep- freshly inoculated plants was deter- tember, and 83% were positive in mined by testing the new flush tissue February. Only 13% of the tagged from plants 4 weeks after graft inocu- branches were CTV positive on all lation. All South African isolates were three sampling dates, whereas 10% of detected in all flushes from grapefruit the branches were CTV negative on and Mexican lime seedlings.
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