Imported tomato fruits infected with Tomato yellow leaf curl virus (TYLCV) were identified on the market in northern Europe using paper-based FTA Classic Cards (Whatman), polymerase chain reaction (PCR) and partial DNA sequence analysis. Trade tomatoes originating from southern Europe, Africa and the Middle East were sampled in Estonia and Sweden, and tested for infection with begomoviruses. Out of 100 batches analysed with five fruits sampled in each batch (58 batches from Estonia and 42 from Sweden), 20 batches were positive (16 from Estonia and four from Sweden). Rolling circle amplification (RCA) and full-length genome sequence analysis of one isolate collected in Estonia and one isolate in Sweden, revealed highest nucleotide sequence identity at 99% to TYLCV-IL for the Estonian isolate and at 97% to TYLCV-Mld for the Swedish isolate. In this study, TYLCV was identified for the first time in imported tomato fruits on the market in northern Europe. FTA cards proved to be an effective means to collect, extract and store begomovirus DNA from tomato fruits and the subsequent molecular analysis.

There is a risk that Tomato yellow leaf curl virus (TYLCV) and its vector, whiteflies of the Bemisia tabaci species complex, will become established in greenhouses in temperate regions of the world, including northern Europe. In this study, TYLCV isolated from imported tomato fruit in Estonia (originating from Spain) was shown to be able to infect plants of tomato and Nicotiana benthamiana using Agrobacterium-mediated inoculation with an infectious clone as well as using biolistic delivery of products from rolling circle amplification (RCA). A 1.8-mer genomic construct of TYLCV was engineered and efficiently agroinfiltrated into plants of tomato and N. benthamiana, and induced symptoms characteristic of natural infection. With Agrobacterium-mediated inoculation, the infection efficiency was 100% for both tomato and N. benthamiana, whereas biolistic inoculation using RCA products resulted in efficiencies of 57% and 36%, respectively. Particle bombardment with monomeric linear genome failed to produce any infection in tomato or N. benthamiana. The genome of TYLCV amplified from tomato fruit was infectious confirming that tomato fruit may serve as a source of virus inoculum. This is the first report of agroinfiltration and particle bombardment assay using TYLCV DNA derived from infected tomato fruit tissue.

Fruit constitutes a strong sink organ and thus accumulates infecting viruses, but there is limited information about the infection process of viruses in fruit. Tomato yellow leaf curl virus (TYLCV, genus Begomovirus, family Geminiviridae) is one of the most important viruses affecting the production of tomato fruit. Using real-time quantitative PCR, TYLCV was shown to accumulate with increasing titres in early developing tomato fruit tissues from anthesis until 21 days post-anthesis. In situ hybridization demonstrated that TYLCV DNA and transcripts of the coat protein gene localized specifically to the phloem tissue of young fruit as well as sepals and petals. Embryos of developing seeds were also found to be infected. Expression of a host histone H4 gene was used as a marker for S-phase and the gene was also found to be expressed in phloem cells of young tomato fruit. The results indicate that TYLCV is transported to developing tomato fruit, where the virus titre gradually increases because of movement and probably also due to virus replication. In this study, the accumulation and localization of TYLCV in early developing tomato fruit are monitored for the first time.