An Ephemeral Sexual Population of <i>Phytophthora infestans</i> in the Northeastern United States and Canada

<div><p><i>Phytophthora infestans</i>, the causal agent of late blight disease, has been reported in North America since the mid-nineteenth century. In the United States the lack of or very limited sexual reproduction has resulted in largely clonal populations of <i>P. infestans</i>. In 2010 and 2011, but not in 2012 or 2013, 20 rare and diverse genotypes of <i>P. infestans</i> were detected in a region that centered around central New York State. The ratio of A1 to A2 mating types among these genotypes was close to the 50∶50 ratio expected for sexual recombination. These genotypes were diverse at the <i>glucose-6-phosphate isomerase</i> locus, differed in their microsatellite profiles, showed different banding patterns in a restriction fragment length polymorphism assay using a moderately repetitive and highly polymorphic probe (RG57), were polymorphic for four different nuclear genes and differed in their sensitivity to the systemic fungicide mefenoxam. The null hypothesis of linkage equilibrium was not rejected, which suggests the population could be sexual. These new genotypes were monomorphic in their mitochondrial haplotype that was the same as US-22. Through parentage exclusion testing using microsatellite data and sequences of four nuclear genes, recent dominant lineages US-8, US-11, US-23, and US-24 were excluded as possible parents for these genotypes. Further analyses indicated that US-22 could not be eliminated as a possible parent for 14 of the 20 genotypes. We conclude that US-22 could be a parent of some, but not all, of the new genotypes found in 2010 and 2011. There were at least two other parents for this population and the genotypic characteristics of the other parents were identified.</p></div

Discriminant Analysis of Principal Components (DAPC) using 12 microsatellite loci.

<p>This scatterplot shows the first two principal components of the DAPC of <i>Phytophthora infestans</i> genotypes found in the United States. Groups are shown by different colors and inertia ellipses, while dots represent individual strains. Cluster 1 includes lineages US-6, US-7, US-11, US-12, and US-16; Cluster 2 includes isolates in lineage US-23; Cluster 3 includes lineages GDT-02, GDT-07, GDT-13, GDT-18, and GDT-20; Cluster 4 includes lineages GDT-03, GDT-04, GDT-08, GDT-08.1, GDT-14 and GDT-15; Cluster 5 includes lineages US-8, US-14, US-20, and US-24; Cluster 6 includes lineages GDT-05, GDT-06, GDT-09, GDT-10, GDT-11, GDT-12, GDT-16, GDT-17, and GDT-19; Cluster 7 includes lineages US-1; and Cluster 8 included lineages US-17, US-19, US-21 and GDT-01.</p

Spatial occurrence of the NYS-2010/11 population of <i>Phytophthora infestans</i> detected in western New York State.

<p>Genotypes that are underlined are those detected in 2010 all other genotypes were detected in 2011. Genotypes shown in bold are those that were found in several counties (GDT-01, GDT-04, and GDT-08). In New York State we detected only A1 individuals in six counties, and only A2 individuals in another three counties. However, both A1 and A2 individuals were reported from yet two other counties. Because of our limited sample size, we cannot conclude with certainty that both mating types were not present in counties where only a single mating type was detected.</p

<i>P. infestans</i> population displacement in Great Britain by the <i>13_A2</i> genotype.

<p>(A) Frequency of multilocus genotypes (MLGs) over the course of 11 years from more than 4000 potato blight outbreaks. The number of isolates fingerprinted each year and dominant MLGs of each mating type are indicated. Isolates of MLGs that occurred at a very low frequency in a single year are grouped under the category termed <i>‘misc’</i>. The shading between the bars indicates the proportion of A1 and A2 mating type isolates. (B) Minimum Spanning Trees based on the alleles at 11 SSR loci indicating the relatedness of the main MLGs and decrease in population diversity between the periods 2003–5 and 2008. The numerous short branches from the <i>13_A2</i> MLG node reflect the high mutation rate in some SSR markers that results in intra-MLG diversity (<i>n</i> is the number of isolates from which the trees are derived). (C) Spatial pattern of spread two dominant MLGs across Great Britain (GB) from 2006–2008 (the numbers of isolates are indicated on each pie chart).</p

Summary of nonsynonymous and synonymous single nucleotide polymorphisms (SNPs) in coding genes (CDSs) of <i>P. infestans</i> 06_3928A compared to T30-4 reference genome strain.

*<p>count of SNPs causing loss of stop codons were omitted;</p>†<p>dN/dS rates were calculated using Yang method reported in Yang and Nielsen <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1002940#ppat.1002940-Yang2" target="_blank">[56]</a>.</p><p>Nonsynonymous and synonymous SNPs were calculated for all genes, core orthologs and RXLRs. Core orthologs as genes showing orthologous sequences 1∶1∶1 in <i>P. infestans: P. ramorum: P. sojae</i> genomes respectively <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1002940#ppat.1002940-Haas1" target="_blank">[23]</a>.</p

Gene expression polymorphisms correlate with extended biotrophy in <i>P. infestans 13_A2</i> isolate 06_3928A.

<p>(A) Number of genes (left) and RXLR effector genes (right) that are induced during potato infection in <i>P. infestans</i> T30-4, 06_3928A and NL07434 strains. Only a small subset of genes is consistently induced in the three strains. (B) Average gene expression pattern during potato infection for all genes (left) and RXLR effector genes (right) induced in all three <i>P. infestans</i> strains analyzed. A consistent divergence is observed at 3 days post inoculation (dpi) when gene induction is maintained in the 06_3928A isolate only. (C) Number of all (left) and RXLR effector genes (right) induced at various time points during potato infection in each of the three <i>P. infestans</i> strains. Compared to other strains, 06_3928A shows the highest number of genes that are induced both at 2 and 3 dpi. (D) Variation in the size of the biotrophic area (infected living host tissue) in lesions induced by three <i>P. infestans</i> strains during potato infection. Error bars are s.e.m. over 28 measurements at 2, 3 and 4 dpi. Representative pictures illustrate the sizes of the necrotrophic (infected dead host tissue, centre of the lesion) and biotrophic (periphery of the lesion, lighter grey ring) growth (mm) with the respective <i>P. infestans</i> strain in color (blue for 06_3928A, red for T30-4 and orange for NL07434 strain) from lesions at 3 dpi.</p

Field aggressiveness of five <i>P. infestans</i> genotypes estimated using mark-and-recapture methods during a controlled blight epidemic.

<p>The proportion of each multilocus genotypes (MLG) recovered amongst 716 foliar blight lesions over a 21 day epidemic initiated with 5 <i>P. infestans</i> isolates of known MLG presented by (A), sampling date and (B), Cultivar. Note; in each case the Y-axis scale is set from 0.7 to 1.0 to more clearly reveal the proportion of the non-<i>13_A2</i> MLGs. The isolates of ‘Other’ MLGs were of non-introduced clonal MLGs that migrated into the trial.</p

<i>13_A2</i> genotype is among the most aggressive <i>P. infestans</i> genotypes on potato.

<p>Aggressiveness of 26 <i>P. infestans</i> isolates grouped into 10 multilocus genotypes (MLGs) on leaves of five potato cultivars (A to E) estimated using mean latent period (x-axis) and mean lesion size at 6 days post inoculation (dpi) (y-axis). Measurements made at 13°C and 18°C are indicated with empty and filled symbols, respectively, and the three most aggressive MLGs colour-coded. (F) The sum of the ranked positions of each MLG according to lesion size at 13°C and 18°C indicates that <i>6_A1</i> and <i>13_A2</i> isolates more often had the largest lesions (particularly at 13°C). The standard errors (s.e.) and degrees of freedom (d.f.) for cultivar by MLG comparisons of latent period and lesion size at 13°C and 18°C (charts A to E) are shown in the lower corner of the figure.</p

Distribution of polymorphism in genes of <i>P. infestans 13_A2</i> isolate 06_3928A.

<p>Y-axis indicates the frequency of synonymous substitutions (dS) in (A) and nonsynonymous substitutions (dN) in (C) given for RXLR genes, other (non-RXLR) genes and core ortholog genes <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1002940#ppat.1002940-Haas1" target="_blank">[23]</a>. Y-axis shows the rates of synonymous substitutions (dS) in (B) and nonsynonymous substitutions (dN) in (D) given for RXLR genes, other (non-RXLR) genes and core ortholog genes. Box and whisker plots in (B and D) show median, first and third quartile, and first values beyond 1.5 times the interquatile range.</p

Invariant avirulence genes in <i>P. infestans</i> 06_3928A identify efficient plant resistance genes.

<p>(A) Expression patterns of three avirulence genes in <i>P. infestans</i> T30-4 (red) and 06_3928A strains (blue) during potato infection. In addition to sequence conservation, these genes also show conserved gene induction patterns in 06_3928A. (B) The corresponding potato resistance genes confer resistance to <i>P. infestans</i> T30-4 and 06_3928A strains.</p