Genetic Change Within Populations of Phytophthora infestans in the United States and Canada During 1994 to 1996: Role of Migration and Recombination

ABSTRACT Dramatic changes occurred within populations of Phytophthora infestans in the United States and Canada from 1994 through 1996. Occurrence of the US-8 genotype, detected rarely during 1992 and 1993, increased rapidly and predominated in most regions during 1994 through 1996. US-7, which infected both potato and tomato and made up almost 50% of the sample during 1993, was detected only rarely among 330 isolates from the United States analyzed during 1994. It was not detected at all in more limited samples from 1996. Thus, ability to infect both potato and tomato apparently did not increase the fitness of this genotype relative to US-8, as predicted previously. US-1, the previously dominant genotype throughout the United States and Canada, made up 8% or less of the samples analyzed during 1994 through 1996. A few additional genotypes were detected, which could indicate the beginnings of sexual reproduction of P. infestans within the United States and Canada. However, clonal reproduction still predominated in all locations sampled; opportunities for sexual reproduction probably were limited, because the A1 and A2 mating types usually were separated geographically. The high sensitivity of the US-1 genotype to the fungicide metalaxyl also could have reduced opportunities for contact between the mating types in fields where this compound was applied. The previous correlation between metalaxyl sensitivity and genotype was confirmed and extended to a new genotype, US-17: all US-1 isolates tested were sensitive; all isolates of the US-7, US-8, and US-17 genotypes tested to date have been resistant. Isolates of P. capsici and P. erythroseptica, two other species often found on tomato and potato, could be easily distinguished from each other and from P. infestans using a simple allozyme assay for the enzyme glucose-6-phosphate isomerase. This technique could be useful for rapid identification of species, in addition to genotype of P. infestans. It generally was not possible to predict which genotypes would be present in a location from 1 year to the next. Long-distance movement of US-8 in seed tubers was documented, and this was probably the primary means for the rapid spread of this genotype from 1993 through 1996.     

rep-PCR-Mediated Genomic Fingerprinting: A Rapid and Effective Method to Identify Clavibacter michiganensis

ABSTRACT The genomic DNA fingerprinting technique known as repetitive-sequence-based polymerase chain reaction (rep-PCR) was evaluated as a tool to differentiate subspecies of Clavibacter michiganensis, with special emphasis on C. michiganensis subsp. michiganensis, the pathogen responsible for bacterial canker of tomato. DNA primers (REP, ERIC, and BOX), corresponding to conserved repetitive element motifs in the genomes of diverse bacterial species, were used to generate genomic fingerprints of C. michiganensis subsp. michiganensis, C. michiganensis subsp. sepedonicus, C. michiganensis subsp. nebraskensis, C. michiganensis subsp. tessellarius, and C. michiganensis subsp. insidiosum. The rep-PCR-generated patterns of DNA fragments observed after agarose gel electrophoresis support the current division of C. michiganensis into five subspecies. In addition, the rep-PCR fingerprints identified at least four types (A, B, C, and D) within C. michiganensis subsp. michiganensis based on limited DNA polymorphisms; the ability to differentiate individual strains may be of potential use in studies on the epidemiology and host-pathogen interactions of this organism. In addition, we have recovered from diseased tomato plants a relatively large number of naturally occurring avirulent C. michiganensis subsp. michiganensis strains with rep-PCR fingerprints identical to those of virulent C. michiganensis subsp. michiganensis strains.     

Bracken effects on inorganic nitrogen leaching from an upland podzol

Leaching of inorganic N species to stream waters from upland areas of the UK is increasing, reflecting the increases in atmospheric deposition of nitrogen species due to increased levels of vehicular emissions. Bracken cover in UK uplands is also increasing overall ( Taylor, 1986 ), and the architectural nature of bracken has been shown to both increase deposition of atmospheric constituents to soils and change their chemical and physical nature. We have tested the nutrient status of upland podzols from the Lake District, Cumbria, UK, from under both moorland grass and bracken. The results show reduced levels of base cations and nitrate throughout the profile and decreases in the organic matter content of surface horizons as a consequence of bracken encroachment. We also report increased ammonium concentrations in lower soil horizons under bracken, possibly leading to increased leaching to stream waters. These results suggest either increased leaching of soil nutrients due to bracken encroachment or increased storage of nutrients within the bracken rhizome system. Results also suggest changes in either microbial activity or the microbial community of podzols, possibly due to the allelopathic nature of bracken. This may have caused changes in the soil nutrient dynamics, possibly accounting for some of the changes seen. We hypothesize that increased bracken cover within the British uplands has the potential to increase leaching of inorganic nitrogen to upland streams.     

Development of a pheromone trap monitoring system for orange wheat blossom midge, Sitodiplosis mosellana, in the UK

Field-trapping experiments with synthetic 2,7-nonadiyl dibutyrate, the female-produced sex pheromone of the orange wheat blossom midge, Sitodiplosis mosellana (Géhin), demonstrated that pheromone traps were highly attractive to males and caught very few non-target organisms. Different formulations of pheromone were tested to identify the optimum release rate and dispenser type for use in pheromone traps in the UK. Key findings were that racemic pheromone was as effective as enantiomerically pure (2S,7R)-2,7-nonadiyl dibutyrate, that release rates higher than 0.5 microg day(-1) were not necessary and that the optimal formulation was a 1 mg pheromone loading in a rubber septum. Pheromone traps gave a reliable indication of peak midge emergence, onset of flight and abundance of midges throughout the season. A strong correlation between maximum trap catch and crop infestation levels was obtained.     

Eliminating pond differences with cross-over designs

Large variability between ponds in aquaculture experiments often leads to imprecise estimates of treatment effects. One solution would be to increase the number of experimental ponds but the availability of ponds is usually limited. A cross-over trial makes comparisons directly within the same pond, instead of across ponds, thus taking account of individual pond effects and potentially removing much of the variability. A cross-over trial was initiated to test the feasibility of using such designs in pond aquaculture research. When compared to a completely randomized design, the cross-over trial was more efficient for comparing the fish growth for each of the two treatments, and, on average, a considerable reduction in the standard error of the treatment estimate was achieved. Thus, under certain circumstances, the cross-over design has great potential to improve the precision of pond aquaculture experimentation.     

Non-intertidal habitat use by shorebirds: a reflection of inadequate intertidal resources?

Many species of shorebird typically forage almost exclusively on intertidal habitats. When such strongly maritime species choose to forage on non-intertidal habitats, it may either be a response to deteriorating intertidal conditions or to the discovery of more profitable resources in non-intertidal areas. Methods which allow distinction between these two will clearly be important for identifying problems in intertidal habitats. Since January 1998, turnstone (Arenaria interpres) on the Wash estuary, eastern England have been foraging on the docksides of Port Sutton Bridge (mainly on spilt wheat and fishmeal), fields and river edges, resulting in concern that intertidal food supplies were no longer sufficient to support the population. We quantified the distribution and behaviour of turnstone within the Wash in relation to season, tidal state and weather and used a depletion model to predict the number of turnstone that could be supported by the port, under a range of resource densities and environmental conditions. Numbers of turnstone on non-intertidal habitats increased over the winter and use of the port was significantly greater around high tide and on colder days. The depletion model showed that under virtually all conditions, the port could support a much greater proportion of the turnstone population than current peak numbers. The use of non-intertidal habitats therefore suggests that the preferred intertidal food supplies are not currently capable of supporting the turnstone population throughout the winter. Habitat switches such as this can potentially be important advance warnings of ecological changes for species, which have not yet led to reductions in population size.     

Silencing of host basal defense response-related gene expression increases susceptibility of Nicotiana benthamiana to Clavibacter michiganensis subsp. michiganensis

Clavibacter michiganensis subsp. michiganensis is an actinomycete, causing bacterial wilt and canker disease of tomato (Solanum lycopersicum). We used virus-induced gene silencing (VIGS) to identify genes playing a role in host basal defense response to C. michiganensis subsp. michiganensis infection using Nicotiana benthamiana as a model plant. A preliminary VIGS screen comprising 160 genes from tomato known to be involved in defense-related signaling identified a set of 14 genes whose suppression led to altered host-pathogen interactions. Expression of each of these genes and three additional targets was then suppressed in larger-scale VIGS experiments and the effect of silencing on development of wilt disease symptoms and bacterial growth during an N. benthamiana-C. michiganensis subsp. michiganensis compatible interaction was determined. Disease susceptibility and in planta bacterial population size were enhanced by silencing genes encoding N. benthamiana homologs of ubiquitin activating enzyme, snakin-2, extensin-like protein, divinyl ether synthase, 3-hydroxy-3-methylglutaryl-coenzyme A reductase 2, and Pto-like kinase. The identification of genes having a role in the host basal defense-response to C. michiganensis subsp. michiganensis advances our understanding of the plant responses activated by C. michiganensis subsp. michiganensis and raises possibilities for devising novel and effective molecular strategies to control bacterial canker and wilt in tomato.