Psorosis, sometimes also associated with ringspot symptoms, is a widespread and damaging disease of citrus in many parts of the world including South America and the Mediterranean basin. We describe the application of RT‐PCR and DAS‐ELISA diagnostics to an isolate of citrus ringspot virus (CtRSV‐4) and other virus isolates associated with this disease. Fragments of cDNA from bottom‐component RNA of CtRSV‐4 were cloned and sequenced, and PCR primers were designed, 5′ACAATAAGCAAGACAAC upstream, and 5′CCATGTCACTTCTATTC downstream. RT‐PCR experiments using these primers allowed detection of CtRSV‐4 in infected citrus leaves down to a tissue dilution of 1/12 800 representing 2 μg of tissue, and less sensitive detection of the related citrus psorosis‐associated virus (CPsAV90‐1‐1) and four other psorosis isolates from Argentina and the USA. In addition, CtRSV‐4 particles were partially purified from local lesions in Chenopodium quinoa, and the preparations used to raise a rabbit antiserum. The antiserum was absorbed with extracts of healthy C. quinoa leaves, and a DAS‐ELISA kit was prepared and tested for detection of CtRSV‐4, CPsAV90–1‐1, and other psorosis isolates from Argentina, the USA, Italy and Spain. The ELISA detected CtRSV‐4 down to a tissue dilution of 1/1600, and most other psorosis isolates down to dilutions of 1/200–1/800. Three of a total of 20 heterologous isolates were consistently negative. Comparison of the PCR and ELISA results suggests that both methods can be used for detection of a range of psorosis isolates, but that variation of the viruses in the field might cause problems for any one diagnostic test.

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Somatic embryogenesis was used to eliminate Citrus psorosis virus (CPsV) from three citrus species (common mandarin, sweet orange and Dweet tangor), all of which regenerated somatic embryos with different embryogenic potential from stigma and style explants. CPsV was detected by double antibody sandwich-indirect-enzyme-linked immunosorbent assay (DASI-ELISA) in explants and embryogenic callus, but was not detected in any of the plants obtained from somatic embryos, even 24 months after regeneration. Loss of juvenile characters (disappearance of thorns) was observed in the first year of growth and was retained in plants propagated by grafting from thornless stems. Somatic embryogenesis appears to be a very promising technique for the production of healthy citrus stocks.

Citrus tristeza virus (CTV) represents one of the major threats to citrus production worldwide. In the East Adriatic region, CTV symptoms are mostly absent due to traditional citrus grafting on trifoliate orange (Poncirus trifoliata), a CTV-tolerant rootstock. Therefore, the virus has been continuously spreading by the propagation of infected material. The genetic variability of CTV was studied on nineteen citrus samples, collected from orchards in the coastal region of Croatia, Montenegro and Albania, that previously tested positive by ELISA and immunocapture RT-PCR. Single-strand conformation polymorphism of the amplified coat protein gene demonstrated the presence of different CTV variants in each amplicon, while sequence analysis of cloned CP gene variants confirmed their clustering into six out of the seven phylogenetic groups so far delineated. Four of these groups include sequences of severe quick decline, seedling yellows and stem-pitting (SP) isolates, thought to be found only rarely in the Mediterranean region. Regardless of the lack of symptoms in the field, CTV isolates from the East Adriatic displayed high genetic variability and pathogenic potential, additionally confirmed by biological characterisation. The high percentage of mixed infections suggest the potential for further diversification and a greater risk of severe variants spreading into new areas.

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