Genome Analyses of an Aggressive and Invasive Lineage of the Irish Potato Famine Pathogen

Pest and pathogen losses jeopardise global food security and ever since the 19th century Irish famine, potato late blight has exemplified this threat. The causal oomycete pathogen, Phytophthora infestans, undergoes major population shifts in agricultural systems via the successive emergence and migration of asexual lineages. The phenotypic and genotypic bases of these selective sweeps are largely unknown but management strategies need to adapt to reflect the changing pathogen population. Here, we used molecular markers to document the emergence of a lineage, termed 13_A2, in the European P. infestans population, and its rapid displacement of other lineages to exceed 75% of the pathogen population across Great Britain in less than three years. We show that isolates of the 13_A2 lineage are among the most aggressive on cultivated potatoes, outcompete other aggressive lineages in the field, and overcome previously effective forms of plant host resistance. Genome analyses of a 13_A2 isolate revealed extensive genetic and expression polymorphisms particularly in effector genes. Copy number variations, gene gains and losses, amino-acid replacements and changes in expression patterns of disease effector genes within the 13_A2 isolate likely contribute to enhanced virulence and aggressiveness to drive this population displacement. Importantly, 13_A2 isolates carry intact and in planta induced Avrblb1, Avrblb2 and Avrvnt1 effector genes that trigger resistance in potato lines carrying the corresponding R immune receptor genes Rpi-blb1, Rpi-blb2, and Rpi-vnt1.1. These findings point towards a strategy for deploying genetic resistance to mitigate the impact of the 13_A2 lineage and illustrate how pathogen population monitoring, combined with genome analysis, informs the management of devastating disease epidemic

Oxygen metabolism in plantlbacteria interactions: characterization of the oxygen uptake response of plant suspension cells

In recent years the accumulation of reactive oxygen species (ROS) has been studied in plant cell suspension systems treated with bacterial pathogens. However, the associated utilization of molecular oxygen has not been well characterized. Using a multi-electrode oxygen analyser, the rates of oxygen consumption by tobacco cells during bacterial interactions were monitored. Heat-killed (HK) bacteria, which initiate an immediate ROS response in plant cells, were used as an elicitor to avoid complications of oxygen consumption by viable bacteria. An increase in oxygen uptake by the tobacco cells occurred within 4 min after addition of HK-bacteria and lasted for about 10 min, returning to a steady state at approximately twice the initial basal rate. The initial burst in oxygen uptake coincided with production of H202. Calculation of the total oxygen consumption by the plant cells indicated that less than 5 % of the increased oxygen uptake was utilized in ROS production. Use of respiratory inhibitors indicated that respiration, especially the cytochrome pathway, played a significant role in this response. Results from the use of K-252, a protein kinase inhibitor, and DPI, an inhibitor of membrane bound NADPH oxidases, indicated that triggering of the oxygen uptake response may involve protein phosphorylation and is at least partially activated by the membrane bound NADPH oxidase activity. The involvement of mitochondrial respiration in the oxygen uptake response described here indicates that early events in plant recognition of pathogens involves more of the cellular machinery than previously hypothesized

Oxidative metabolism in plant/bacteria interactions: characterization of a unique oxygen uptake response of potato suspension cells

Plant suspension cells have been shown to respond to bacteria or microbial elicitors by producing active oxygen as well as increasing oxygen uptake. Here we characterize a unique two stage oxygen uptake response of potato suspension cells to heat-killed bacteria. Stage 1 occurred within minutes after the addition of heat-killed bacteria; the potato suspension cells responded with a rapid increase in oxygen uptake and reached a steady state approximately 50 % greater than the initial basal rate. Stage 2 began 20-30 min after this new steady state was achieved and was characterized by a slow increase in the oxygen uptake rate over the remaining 90 min period. Calculation of the total oxygen consumption by the plant cells indicated that only a small fi-action of the increased oxygen uptake was due to the concomitant production of reactive oxygen species. The protein kinase inhibitor, K-252, inhibited the oxygen uptake response by 80-90 %, suggesting the involvement of protein phosphorylation in the oxygen uptake response. The alternate oxidase inhibitor, SHAM, inhibited the elicited oxygen uptake response by about 25 % while a combination of SHAM and KCN almost completely blocked respiration as well as the elicited response. The data indicate that mitochondrial respiration and, in particular, the alternate oxidase, play a significant role in the elicited oxygen uptake response of potato cells

Oxidative metabolism in plant/bacteria interactions: characterization of the oxygen uptake response of bacteria

An increase in oxygen uptake has been previously described in plant cell suspensions treated with bacteria or bacterial elicitors. These studies, regarding oxygen uptake, have all been undertaken from the perspective of the host plant cell reacting to the invading pathogen. In contrast, here we describe and characterize an increase in oxygen uptake by bacterial cells in response to plant suspensions or autoclaved plant cell filtrates. Autoclaved plant cell filtrates stimulated bacterial oxygen uptake by as much as sevenfold within a few minutes after addition. This oxygen uptake response was proportional to both the concentration of the plant cell filtrate and the concentration of the bacteria. KCN inhibited the bacterial response, suggesting that bacterial respiration may be involved. Unlike the plant oxygen uptake response to bacteria, there was no concurrent H202 accumulation and the NADPH oxidase inhibitor, DPI, had no effect on the bacterial response. Streptomycin, an inhibitor of bacterial protein synthesis, inhibited the bacterial oxygen uptake response to the plant cell filtrate. K-252, a protein kinase inhibitor that strongly inhibits the plant oxygen uptake response to bacteria, had no effect upon the bacterial oxygen uptake response. When potato/bacterial cell suspensions were pretreated with either streptomycin or K-252, the combined plant/bacterial oxygen uptake response was inhibited by 15 or 70 %, respectively. This indicates that as much as 15-30 % of the increased oxygen consumption during plant suspension cell/bacteria interactions may be attributable to bacteria, which comprise less than 1 % of the total cell mass

Apoplastic redox metabolism: Synergistic phenolic oxidation and a novel oxidative burst

The plant apoplast is an important mediator of communication between the cell cytoplasm and its surroundings. Plant cell suspensions offer a convenient model system to gain insight into apoplastic physiology. Here, we describe a novel phenomenon that took place when two naturally occurring phenolics were added together to either soybean or tobacco cell suspensions. Acetosyringone (AS) and/or hydroxyacetophenone (HAP), phenolics found in the extracellular/apoplast of tobacco cells, were added to soybean or tobacco cell suspensions undergoing an oxidative burst. Individually, AS appeared to be utilized as a typical peroxidase substrate to scavenge hydrogen peroxide, while HAP was utilized at a much lower rate. However, when added together the rate of utilization of both phenolics increased and surprisingly resulted in the production of hydrogen peroxide. We have further characterized this novel phenomenon in suspension cells. This study demonstrates that certain phenolics in plants can cause co-oxidation which, as in animals, could alter the structure and bioactivity of surrounding phenolics

Induction of redox sensitive extracellular phenolics during plant–bacterial interactions

This study focuses on the transient and complex nature of phenolics that accumulate in the extracellular environment of plant suspension cells during the first few hours of the interaction between these plant cells and bacterial pathogens. Using suspension cells of Nicotiana tabacum we identified four acetophenones and four hydroxycinnamic acid amides that accumulate in this extracellular environment. Treatment of the suspension cells with isolates of the plant pathogen Pseudomonas syringae or heat-killed bacteria increased elicitation of extracellular phenolics and changed the composition of the compounds that accumulated. These phenolics were sensitive to oxidative stress; when suspension cells were treated with bacterial strains or elicitors that triggered an oxidative burst, these phenolics were oxidized and depleted for the duration of the burst. The qualitative and quantitative makeup of phenolics produced by N. tabacum suspensions was also affected by plant cell age and density. To our knowledge, this is the first study that closely follows the kinetics of individual extracellular phenolic compounds and the concurrent oxidative stress during the first few hours of a plant–bacterial interaction