The foodborne pathogen Escherichia coli O157:H7 is increasingly associated with fresh produce (fruit and vegetables). Bacterial colonization of fresh produce plants can occur to high levels on the external tissue but bacteria have also been detected within plant tissue. However, questions remain about the extent of internalization, its molecular basis, and internal location of the bacteria. We have determined the extent of internalization of E. coli O157:H7 in live spinach and lettuce plants and used high-resolution microscopy to examine colony formation in roots and pathways to internalization. E. coli O157:H7 was found within internal tissue of both produce species. Colonization occurred within the apoplast between plant cells. Furthermore, colonies were detected inside the cell wall of epidermal and cortical cells of spinach and Nicotiana benthamiana roots. Internal colonization of epidermal cells resembled that of the phytopathogen Pectobacterium atrosepticum on potato. In contrast, only sporadic cells of the laboratory strain of E. coli K-12 were found on spinach, with no internal bacteria evident. The data extend previous findings that internal colonization of plants appears to be limited to a specific group of plant-interacting bacteria, including E. coli O157:H7, and demonstrates its ability to invade the cells of living plants.

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Engineering resistance genes to gain effector recognition is emerging as an important step in attaining broad, durable resistance. We engineered potato resistance gene R3a to gain recognition of the virulent AVR3aEM effector form of Phytophthora infestans. Random mutagenesis, gene shuffling and site-directed mutagenesis of R3a were conducted to produce R3a* variants with gain of recognition towards AVR3aEM. Programmed cell death following gain of recognition was enhanced in iterative rounds of artificial evolution and neared levels observed for recognition of AVR3aKI by R3a. We demonstrated that R3a*-mediated recognition responses, like for R3a, are dependent on SGT1 and HSP90. In addition, this gain of response is associated with re-localisation of R3a* variants from the cytoplasm to late endosomes when co-expressed with either AVR3aKI or AVR3aEM a mechanism that was previously only seen for R3a upon co-infiltration with AVR3aKI. Similarly, AVR3aEM specifically re-localised to the same vesicles upon recognition by R3a* variants, but not with R3a. R3a and R3a* provide resistance to P. infestans isolates expressing AVR3aKI but not those homozygous for AVR3aEM.

In plants infected with Potato leafroll virus (PLRV), or other luteoviruses, infection is very largely

The whole RNA genomes of six European Potato virus Y (PVY) isolates, representing isolates of the PVY O , PVY N and PVY NTN subgroups, were sequenced and the reactions of a range of tobacco and potato cultivars after the inoculation were determined. One isolate induced symptoms of veinal necrosis when inoculated to tobacco, but reacted with PVY O -specific antibodies. The genome sequence of this isolate showed that it had a recombinant genome comprising large sections of sequence that are characteristic of N and O sub-group isolates. Another isolate caused mosaic symptoms when inoculated to tobacco, and reacted with PVY N -specific antibodies. The nucleotide sequence of its genome was very closely related to the genome sequence published for two potato-infecting PVY isolates from North America. These three isolates are the members of interesting group NA (North American). They have highly similar genome sequences throughout their length, but biologically represent isolates from the PVY O , PVY N and PVY NTN subgroups. The occurrence of these biologically diverse isolates with their highly homologous genomes brings into question the exact nature of gene structure in relation to function.Resumen Se secuenciaron los genomas completos de ARN de seis aislamientos europeos del virus Y de la papa (PVY) representantes de aislamientos de los subgrupos de PVY O , PVY N y PVY NTN y se determinaron las reacciones de una serie de cultivares de tabaco y papa después de las inoculaciones. Un aislamiento indujo síntomas de necrosis de las nervaduras cuando fue inoculado en tabaco pero reaccionó con los anticuerpos específicos de PVY O . La secuencia del genoma de este aislamiento mostró que tenía un genoma recombinante con grandes secciones de secuencias que son características de aislamientos de los subgrupos N y O. Otro aislamiento causó síntomas de mosaico cuando se inoculó a tabaco y reaccionó con anticuerpos específicos de PVY N . La secuencia de nucleótidos de su genoma fue relacionada muy cercanamente con la secuencia publicada de dos aislamientos de PVY de Norteamérica que infectan papa. Estos tres aislamientos son miembros del grupo NA (Norte Americano). Ellos tienen una secuencia de genoma altamente similar, pero biológicamente representan aislamientos de los subgrupos PVY O , PVY N y PVY NTN . La presencia de estos aislamientos biológica-mente diferentes con sus genomas altamente homólogos, pone en cuestión la naturaleza exacta del gen con relación a su función.