L. Otten

Genome Sequences of Three Agrobacterium Biovars Help Elucidate the Evolution of Multichromosome Genomes in Bacteria

The family Rhizobiaceae contains plant-associated bacteria with critical roles in ecology and agriculture. Within this family, many Rhizobium and Sinorhizobium strains are nitrogen-fixing plant mutualists, while many strains designated as Agrobacterium are plant pathogens. These contrasting lifestyles are primarily dependent on the transmissible plasmids each strain harbors. Members of the Rhizobiaceae also have diverse genome architectures that include single chromosomes, multiple chromosomes, and plasmids of various sizes. Agrobacterium strains have been divided into three biovars, based on physiological and biochemical properties. The genome of a biovar I strain, A. tumefaciens C58, has been previously sequenced. In this study, the genomes of the biovar II strain A. radiobacter K84, a commercially available biological control strain that inhibits certain pathogenic agrobacteria, and the biovar III strain A. vitis S4, a narrow-host-range strain that infects grapes and invokes a hypersensitive response on nonhost plants, were fully sequenced and annotated. Comparison with other sequenced members of the Alphaproteobacteria provides new data on the evolution of multipartite bacterial genomes. Primary chromosomes show extensive conservation of both gene content and order. In contrast, secondary chromosomes share smaller percentages of genes, and conserved gene order is restricted to short blocks. We propose that secondary chromosomes originated from an ancestral plasmid to which genes have been transferred from a progenitor primary chromosome. Similar patterns are observed in select Beta- and Gammaproteobacteria species. Together, these results define the evolution of chromosome architecture and gene content among the Rhizobiaceae and support a generalized mechanism for second-chromosome formation among bacteria.

Opines in crown gall tumours induced by biotype 3 isolates of Agrobacterium tumefaciens.

Pathogenic properties and opine markers of biotype 3 (grapevine) isolates of Agrobacterium tumefaciens (Smith and Townsend) Conn were compared with those of biotypes 1 and 2. By contrast with biotypes 1 and 2, grapevine isolates induced only small tumours on Kalanchoe daigremontiana Hamet and Perrier stems but produced large organogenic galls on decapitated shoot tips of this plant species. Grapevine agrobacteria were classified according to their opine catabolic properties and the type of opine they induced in Kalanchoe tubiflora Hamet and grapevine ( Vitis vinifera L.) tumours. Crown gall tissues obtained after infection with octopine-utilizing grapevine isolates contained octopine in very large amounts compared to biotype 1 tumours. A similar quantitative difference was not observed when nopaline production of tumours incited by biotypes 1 and 2 and by grapevine isolates was examined. Tumours formed by a third class of grapevine isolates synthesized a novel opine, for which we propose the trivial name vitopine. Octopine, nopaline, agropine and vitopine were also detected in healthy tissues of plants infected with the appropriate Agrobacterium isolates, indicating that these opines can selectively promote the colonization of the whole plant by agrobacteria. Biotype 3 octopine isolates induced octopine synthesis at the infection sites of a crown gall-resistant grapevine hybrid, showing that pathogenic agrobacteria can transform plants without tumour formation.

Tartrate Utilization Genes Promote Growth of Agrobacterium spp. on Grapevine

Crown gall on grapevine is mainly caused by Agrobacterium vitis, which metabolizes tartrate. Competition experiments between a tartrate-utilizing strain and its non-utilizing derivative showed that tartrate utilization confers a selective advantage on grapevine.

Further evidence that the vitopine-type pTi's of Agrobacterium vitis Represent a novel group of Ti plasmids

To study the incompatibility properties of the vitopine Ti plasmids of Agrobacterium vitis, pPM739 containing the cloned ori region of pTiS4 was introduced by triparental mating into seven Agrobacterium tumefaciens strains carrying incRh-1, incRh-2, and incAg-1 plasmids and into eight A. vitis strains. All strains containing pPM739 retained their original plasmids and virulence or the ability to grow on tartrate, except for the three vitopine strains S4, Sz1, and NW11. Furthermore, pTiS4 was stably maintained in S4 cells following introduction of the ori/inc clones of the incRh-1, incRh-2, and incRh-3 plasmids. These results show that vitopine Ti plasmids represent a novel incompatibility group for which we propose the name incRh-4.

Major differences between the rrnA operons of two strains of Agrobacterium vitis.

Abstract The sequence of the rrnA operon and its flanking regions was determined for the Agrobacterium vitis type strain NCPPB3554. Compared to the earlier obtained rrnA sequence of A. vitis strain S4, several important differences were noted: the sequences diverged at the 5′-flanking region, within the 16S–23S intergenic region, and within the 23S rRNA sequence. The B8 stem-loop structure at the 5′-end of the 23S rRNA of strain NCPPB3554 was 142 nt shorter than that of strain S4. These findings have important consequences for the use of ribosomal RNA gene sequences in phylogenetic comparisons.