Abstract

Potato (Solanum tuberosum L.) is one of the most important crops grown in Norway, and virus-free plants are required for commercial potato production and for preservation of potato germplasm. The present study evaluates three in vitro therapies – meristem culture, cryotherapy, and chemotherapy combined with thermotherapy – to eliminate viruses from eight historically valuable potato cultivars belonging to the Norwegian potato germplasm. Potato virus Y, potato virus M, potato virus X and potato virus S were present in eight selected old potato cultivars due to long-term conservation in open field. Double-antibody sandwich enzyme-linked immunological assay (DAS-ELISA) and biological indicators were the standard tests used to confirm virus infection in our study. Six virus-free plants from four potato cultivars were obtained after meristem culture, and no virus-free potato cultivars were obtained after cryotherapy. Virus-free frequency for eight different potato cultivars after combining chemotherapy with thermotherapy varied from 36.4% to 100%, with single virus elimination rates of between 74.2% and 92.9%. Chemotherapy compared with thermotherapy was the most effective of the three in vitro therapies used in this study. Highly sensitive small RNA high-throughput sequencing (HTS) was used to evaluate the virus status of potato virus-free materials after virus eradication, and no virus was found, which was consistent with the results of DAS-ELISA and biological indicators. Small RNA HTS has been reported for the first time to evaluate the virus status after virus elimination and to control virus-free potato nuclear stocks.

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Abstract

Virus diseases have been a great threat to production of economically important crops. In practice, the use of virus-free planting material is an effective strategy to control viral diseases. Cryotherapy, developed based on cryopreservation, is a novel plant biotechnology tool for virus eradication. Comparing to the traditional meristem culture for virus elimination, cryotherapy resulted in high efficiency of pathogen eradication. In general, cryotherapy includes seven major steps: (1) introduction of infected plant materials into in vitro cultures, (2) shoot tip excision, (3) tolerance induction of explants to dehydration and subsequent freezing in liquid nitrogen (LN), (4) a short-time treatment of explants in LN, (5) warming and post-culture for regeneration, (6) re-establishment of regenerated plants in greenhouse conditions, and (7) virus indexing.

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Abstract

The presence of viral diseases poses a significant challenge to the high-quality, efficient, and sustainable production of apples. Virus eradication and the use of virus-free plants are currently the most crucial method for preventing viral diseases. Among the viruses affecting apples, apple stem grooving virus (ASGV) and apple chlorotic leaf spot virus (ACLSV) present particular challenges in efficient eradication from apples. This study investigated the effects of exogenous salicylic acid (SA) treatment on the efficient eradication of ASGV and ACLSV from apple cultivar ‘Yanfu 8’. Shoots were excised from in vitro 4-week-old stock and cultured in shoot proliferation medium supplemented with 10 μM SA combining thermotherapy with shoot tip culture or cryotherapy for ASGV and ACLSV eradication. The results showed that including of 10 μM SA in thermotherapy significantly reduced the concentrations of ASGV and ACLSV by 33% and 14% in shoots compared to thermotherapy without SA. SA treatment also improved the shoot tips survival and regrowth after combining 2 or 4 weeks of thermotherapy followed by shoot tip culture or shoot tip cryotherapy, while maintaining the higher (75–100%) of virus eradication efficiencies. Therefore, the application of SA in combination with thermotherapy followed or not by cryotherapy proves to be a promising approach for enhancing the efficiency of virus eradication in apple.