Carlos Marcelino Rodriguez Lopez

bioRxiv2016

Botrytis cinerea loss and restoration of virulence during in vitro culture follows flux in global DNA methylation

O_LIPathogenic fungi can lose virulence after protracted periods of culture but little is known of the underlying mechanisms. Here we present the first single-base resolution methylome for the plant pathogen B. cinerea and identify differentially methylated genes/genomic regions associated with virulence erosion.\nC_LIO_LICultures were maintained for eight months with subcultures and virulence testing every month. Methylation-sensitive amplified polymorphisms were performed at monthly intervals to characterise global changes to the pathogens genome during culture and also on DNA from mycelium inoculated onto Arabidopsis thaliana after eight months in culture. Characterisation of culture-induced epialleles was assessed by whole-genome re-sequencing and whole-genome bisulfite sequencing.\nC_LIO_LIVirulence declined with time in culture and recovered after inoculation on A. thaliana. Variation detected by methylation-sensitive amplified polymorphisms followed virulence changes during culture. Whole-genome (bisulfite) sequencing showed marked changes on global and local methylation during culture but no significant genetic changes.\nC_LIO_LIWe imply that virulence is a non-essential plastic character that is at least partly modified by changing levels of DNA methylation during culture. We hypothesise that changing DNA methylation during culture may be responsible for the high virulence/low virulence transition in B. cinerea and speculate that this may offer fresh opportunities to control pathogen virulence.\nC_LI

Genomics↗

bioRxiv2016

Effect of cryopreservation and post-cryopreservation somatic embryogenesis on the epigenetic fidelity of cocoa (Theobroma cacao L.)

While cocoa plants regenerated from cryopreserved somatic embryos can demonstrate high levels of phenotypic variability, little is known about the sources of the observed variability. Previous studies have shown that the encapsulation-dehydration cryopreservation methodology imposes no significant extra mutational load since embryos carrying high levels of genetic variability are selected against during protracted culture. Also, the use of secondary rather than primary somatic embryos has been shown to further reduce the incidence of genetic somaclonal variation. Here, the effect of in vitro conservation, cryopreservation and post-cryopreservation generation of somatic embryos on the appearance of epigenetic somaclonal variation were comparatively assessed. To achieve this we compared the epigenetic profiles, generated using Methylation Sensitive Amplified Polymorphisms, of leaves collected from the ortet tree and from cocoa somatic embryos derived from three in vitro conditions: somatic embryos, somatic embryos cryopreserved in liquid nitrogen and somatic embryos generated from cryoproserved somatic embryos. Somatic embryos accumulated epigenetic changes but these were less extensive than in those regenerated after storage in LN. Furthermore, the passage of cryopreserved embryos through another embryogenic stage led to further increase in variation. Interestingly, this detected variability appears to be in some measure reversible. The outcome of this study indicates that the cryopreservation induced phenotypic variability could be, at least partially, due to DNA methylation changes.\n\nKey messagePhenotypic variability observed in cryostored cocoa somatic-embryos is epigenetic in nature. This variability is partially reversible, not stochastic in nature but a directed response to the in-vitro culture and cryopreservation.

Plant Biology↗