Detection of a Tristeza-Seedling Yellows Strain in Spain

In late 1984, some growers were found to be using budwood of a very early maturing satsuma mandarin of uncertain origin, for topworking citrus orchards near Valencia. Indexing of this material showed that it was infected with a very severe strain of citrus tristeza virus (CTV). It induced severe stunting, stem pitting, vein clearing and vein corking in Mexican lime and C. macrophylla; conspicuous stunting and seedling-yellows on Eureka lemon and Duncan grapefruit and stem pitting on sweet orange, grapefruit arid rough lemon. None of the common CTV strains previously studied in Spain induces these severe symptoms. There is some evidence indicating that the original satsuma was illegally introduced from Japan. A program has been established for the eradication of this CTV strain

The Citrus Quarantine Station in Spain

A Citrus Quarantine Station has been recently established in Spain. It is based in a tissue culture procedure developed for safe budwood introduction, consisting of the following steps: a) preliminary budwood inspection; b) budwood disinfestation; c) budwood culture in vitro at 32 C for flushing; d) disinfestation of flushes obtained; e) isolation of 0.1-0.2 mm long shoot tips; f) grafting shoot tips in vitro; g) transplanting micrografted plants to a quarantine greenhouse; h) indexing of micrografted plants; i) release of budwood. Any abnormal or contaminated tissue or plant found during the process and all introduced plant material except the shoot tips are destroyed. Pests, fungi, bacteria and even virus and virus-like diseases introduced with the original material can be easily eliminated by this procedure. The estimated time from budwood introduction until budwood release is 24-30 months, including 18-24 months for xyloporosis indexing

Persistent Transmission of Citrus Vein Enation Virus by Aphis gossypii and Myzus persicae

Citrus vein enation virus (CVEV) is widely distributed in Spain and circumstantial evidence suggests a high rate of natural spread. No experimental transmission was achievedin extensive trials with several aphid species using a 2-day acquisition period and a 2-day inoculation period. In further experiments, using a 5-7 day acquisition period and a 9-16 day inoculation period, a 95% transmission efficiency was obtained with Aphis gossypii and 10% with Myzus persicae. Furthermore, a 10% transmission efficiency was achieved with viruliferous A. gossypii maintained on healthy plants for a period of up to 14 days after acquisition then given an 11-day inoculation access feed. These results show that CVEV is persistently transmitted by A. gossypii in Spain, thus explaining the high rate of natural spread in the field

Virus Content and Growth Patterns of Callus Cultured in vitro from Healthy and Virus-Infected Citrus Species

Stem segments from healthy and virus-infected Pineapple sweet orange, Mexican lime and Etrog citron were cultured on callus-inducing medium. Explants infected with citrus infectious variegation virus (CIVV), psorosis, vein enation and cachexia produced significantly less primary callus than healthy controls, whereas citrus tristeza virus (CTV) did not affect callus production. The growth of secondary callus during subculturing was similar in healthy and infected tissues. Indexing of callus indicated that the number of calli infected with CTV and vein enation decreased drastically with subculturing, whereas the number of calli infected with CIVV and psorosis did not decrease after five subcultures

The Citrus Variety Improvement Program in Spain in the Period 1975-2001

The Citrus Variety Improvement Program in Spain (CVIPS) started in 1975. It has the following objectives: a) to recover pathogen-free plants of local cultivars by shoot-tip grafting in vitro (STG); b) to import foreign genotypes through a STG based quarantine procedure; c) to maintain healthy genotypes in a Germplasm Bank; and d) to release healthy budwood to citrus nurseries through a certification program. Plants recovered by STG are biologically indexed by inoculation to the following indicator plants: Mexican lime, Pineapple sweet orange, Dweet tangor, Citrus excelsa, Etrog citron, and Parson’s Special mandarin. In addition, they are indexed by sPAGE or imprint-hybridization for viroids, by RT-PCR for Citrus leaf blotch virus, by dsRNA analysis for viruses that produce dsRNA during their replication cycle, and by tissue print-ELISA for Citrus tristeza virus. Only healthy genotypes are included in the Germplasm Bank, which has a field collection used for research and horticultural evaluation, a cryo-stored collection for longterm maintenance, and a screen-house collection that is used to release budwood to nurseries. It contains a total of 428 genotypes, 237 selected in Spain and 191 imported from other countries, representing 43 Citrus species and 32 species from 17 Citrus-related genera. Release of healthy budwood from this program to nurseries started in 1979. At that time, there were only 10 registered nurseries, but in the last few years the number has now increased to 39. For commercial propagation all nurseries are using budwood from the Germplasm Bank. Since the beginning of the program, about 85 million certified nursery trees from this origin have been produced. This represents more than 70% of the Spanish citrus industry. The CVIPS has had a very high impact on the citrus industry. Virus and virus-like diseases do not currently induce any significant damage in the new plantings, and a wide selection of healthy material from the best varieties is available for growers