In this study, melatonin (MEL)‐mediated plant resistance to tobacco mosaic virus (TMV) was examined to study local infection in Nicotiana glutinosa and systemic infection in Solanum lycopersicum. Exogenous application of 100 µm MEL increased anti‐virus infection activity to 37.4% in virus‐infected N. glutinosa plants. The same treatment significantly reduced relative levels of virus RNA analysed by qRT‐PCR and virus titres measured by dot‐ELISA, and increased the relative expression levels of the PR1 and PR5 genes analysed by qRT‐PCR, in virus‐infected S. lycopersicum. MEL treatment induced considerable accumulations of salicylic acid (SA) and nitric oxide (NO) but did not significantly affect production of hydrogen peroxide (H2O2) in the virus‐infected S. lycopersicum plants. Transgenic nahG N. tabacum was used to determine whether MEL‐induced TMV resistance was dependent on the SA pathway. The results showed that the relative RNA level of the TMV analysed by qRT‐PCR and virus titres analysed by dot‐ELISA were not reduced by the MEL treatment in the nahG transgenic N. tabacum seedlings treated twice with 100 µm MEL. The increased relative expression levels of PR1 and PR5 were greatly reduced when cPTIO, an NO scavenger, was included in the MEL treatment. A working model of MEL‐mediated plant resistance to TMV is proposed. MEL‐mediated plant resistance to viruses provides a new avenue to control plant viral diseases.

Production and maintenance of virus‐free planting materials is pivotal for the control of viral diseases. The present study attempted to test exogenous application of melatonin for eradication of apple stem grooving virus (ASGV) from virus‐infected in vitro shoots of apple cultivar Gala. Exogenous application of 15 μm melatonin to the shoot proliferation medium significantly increased the number of shoots and shoot length. The level of endogenous indole‐3‐acetic acid (IAA) was the highest in the shoots proliferating on the shoot proliferation medium containing 15 μm melatonin. Shoot regrowth levels were significantly higher in shoot tips of the virus‐infected shoots cultured for 4 weeks on this medium than the control. In addition, culture of shoot tips of the virus‐infected in vitro shoots proliferated for 4 weeks on this medium resulted in 95% of shoots being virus‐free, while no virus‐free shoots were obtained in shoot tips of the virus‐infected shoots cultured without melatonin. Analyses by microtissue direct RT‐PCR and RT‐qPCR showed that ASGV concentration decreased in shoot tips of the virus‐infected shoots proliferating on the medium containing 15 μm melatonin for 4 weeks. Virus localization showed that exogenous application of melatonin enlarged the virus‐free area in the virus‐infected shoot tips. These data provide explanations as to why exogenous application of melatonin can efficiently eradicate ASGV. Exogenous application of melatonin provides an alternative means for plant virus eradication and has the potential to produce virus‐free plants.

Here, we report efficient eradication of Jujube witches' broom phytoplasma (Candidatus Phytoplasma ziziphi) from Chinese jujube (Ziziphus jujuba) by cryopreservation. Shoot tips (1.0 mm in size) with 5-6 leaf primordia (LPs) excised from diseased in vitro stock shoots were subject to droplet-vitrification cryopreservation. Shoot tips following cryopreservation were post-cultured on a recovery medium for survival. Plantlet regeneration was obtained by micrografting of surviving shoot tips upon in vitro rootstocks. With this protocol, 85% of shoot tips survived following cryopreservation, among which 75% regenerated into whole plantlets and all of them were free of phytoplasma, regardless of the sizes used for cryopreservation. Ultrastructural studies demonstrated that phytoplasma was absent in the apical dome, and leaf primordia (LPs) 1 and 2, while abundance of phytoplasma was present in the lower parts of shoot tips, leaf primordium 3 and older tissues. Histological observations showed that much more damage was found in cells located in the lower part of apical dome, leaf primordium 3 and older tissues than in those at the upper part of apical dome and in the LPs 1 and 2. These cells were most likely to survive and regenerate into phytoplasma-free plantlets following cryopreservation and micrografting. Ploidy levels analyzed by flow cytometry (FCM) were maintained in plantlets regenerated from cryopreservation followed by micrografting. Results reported here would provide technical support for production of phytoplasma-free plants and for long-term storage of germplasm of Chinese jujube.

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