Diversity of chosen effectors in samples of Polish and Norwegian populations of Phytophthora infestans
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1 Diversity of chosen effectors in samples of Polish and Norwegian populations of Phytophthora infestans Emil Stefańczyk 1, Marta Brylińska 1, May Bente Brurberg 2, Ragnhild Naerstad 2, Abdelhameed Elameen 2, Sylwester Sobkowiak 1, Jadwiga Śliwka 1 1 IHAR-PIB, Młochów, Poland 2 The Norwegian Institute of Bioeconomy Research, Ås, Norway
![potato cultivation 2 15 Norway yield [t] 0.36 mln yield [t] 7.](/docs-images/74/70330843/images/2-0.jpg "7 mln area [ha] 12 000 area [ha] 280 000 climate moderate - subarctical climate moderate Poland cultivars Asterix, Saturna,")
2 potato cultivation 2 15 Norway yield [t] 0.36 mln yield [t] 7.7 mln area [ha] area [ha] climate moderate - subarctical climate moderate Poland cultivars Asterix, Saturna, Mandel, Folva, Lady Claire cultivars Vineta, Denar, Lord, Tajfun, Satina
![potato cultivation 2 15 Norway yield [t] 0.36 mln yield [t] 7.](/docs-images/74/70330843/images/3-0.jpg "7 mln Poland area [ha] 12 000 area [ha] 280 000 climate moderate - subarctical climate moderate")
3 potato cultivation 2 15 Norway yield [t] 0.36 mln yield [t] 7.7 mln Poland area [ha] area [ha] climate moderate - subarctical climate moderate cultivars Asterix, Saturna, Mandel, Folva, Lady Claire cultivars Vineta, Denar, Lord, Tajfun, Satina P. infestans population structure influenced by: cultivation system chemical protection crop rotation potato cultivars seed tubers origin
4 P. infestans structure in Norway Genotype: Poland no dominant clonal lineages high genetic diversity occurence of sexual reproduction source:
5 aim of the study forecasting the late blight resistance genes durability by analyzing the diversity of effector genes Avr-vnt1 recognized by the Rpi-phu1 (Rpi-vnt1.1) gene AvrSmira1 recognized by the Rpi-Smira1 gene
6 material 5 15
7 material reference isolates 4 from Scotland 1 from Ecuador Lees et al., 2006
8 methods primers design Primer3, NetPrimer Avr-vnt1 AvrSmira1 2. effectors sequencing (external companies) 3. sequences processing Chromas Lite 2, NovoSNP, PHASE 2.1 SeaView4, MEGA6 4. data analyses effectors SSR product size (bp) gene size (bp) DNASP 5.10, TCS 1.21, Arlequin 3.5, LOSITAN, omegamap Gene Mapper 4.0, XLSTAT, GenAlex 6.5
9 Avr-vnt1 results 7 15 one polymorphic site all (42) Norwegian 41 Polish pos. 321 aa PRO 3rd position in a codon synonymous mutation nine polymorphic sites found in 8 Polish isolates 3 synonymous 6 nonsynonymous pos aa VAL ILE THR ASN MET LEU ASP ALA PRO ASP ASN LEU PHE ASP TYR
10 Avr-vnt1 results 7 15 one polymorphic site all (42) Norwegian 41 Polish pos. 321 aa PRO 3rd position in a codon synonymous mutation nine polymorphic sites found in 8 Polish isolates 3 synonymous 6 nonsynonymous pos aa VAL ILE THR ASN MET LEU ASP ALA PRO ASP ASN LEU PHE ASP TYR 8 of 9 polymorphic sites behind the RXLR-DEER motif
11 Avr-vnt1 results 8 15 N=192 three Avr-vnt1 alleles hypothetical missing allele * V (n) nonsynonymous mutation number of sequences representing given allele
12 Avr-vnt1 results 8 15 N=192 three Avr-vnt1 alleles EC1 neither Polish isolate, regardless of allele possesed, was virulent to plants with Rpi-phu1 gene hypothetical missing allele * V (n) nonsynonymous mutation number of sequences representing given allele
13 AvrSmira1 results polymorphic sites 7 synonymous 13 nonsynonymous pos aa ALA PRO ASN ARG GLN LYS MET ARG LYS LEU MET LEU GLN ARG ALA ARG GLN GLN LEU GLN LYS PHE LEU THR LYS PRO GLU LYS ARG HIS ARG ASP ASN
14 AvrSmira1 results polymorphic sites 7 synonymous 13 nonsynonymous pos aa ALA PRO ASN ARG GLN LYS MET ARG LYS LEU MET LEU GLN ARG ALA ARG GLN GLN LEU GLN LYS PHE LEU THR LYS PRO GLU LYS ARG HIS ARG ASP ASN N- RXLR-DEER C-terminal 18 of 20 polymorphic sites behind the RXLR-DEER motif
15 AvrSmira1 results N= AvrSmira1 alleles nine alleles unique for Polish isolates one allele unique for Norwegian isolates
16 AvrSmira1 results N= AvrSmira1 alleles nine alleles unique for Polish isolates one allele unique for Norwegian isolates 29 of 48 Polish isolates virulent to plants with Rpi-Smira1 absence of AvrSmira1 variants, carrying the M 156 and R 170 amino acids among the P. infestans isolates avirulent to Rpi-Smira1 plants virulent Rietman et al., 2012 M 156 R 170 L 156 R 170 M 156 Q 170 L 156 Q 170 avirulent
17 test for selection omegamap effector sequences divided according to their origin and tested separately Avr-vnt1 no positively selected sites among sequences from both sample sets AvrSmira1 the same potentially selected sites among sequences from both sample sets amino acids , 156, 170, 178, , 199, 205, 209, 221
18 test for selection omegamap effector sequences divided according to their origin and tested separately Avr-vnt1 no positively selected sites among sequences from both sample sets AvrSmira1 the same potentially selected sites among sequences from both sample sets amino acids , 156, 170, 178, , 199, 205, 209, 221 Rietman et al statistically significant statistically insignificant positive selection 128, 130, 156, , 180, 199, 209
19 Ri-1B1 P-3S P-3B1 P-5S MP1710 M-1B3 HS-3B2 HS-3S HS-4S3 HS-3B3 HS-3B1 HS-1B1 HS-2B1 HS-4S2a HS-4S2b Ri-1B3 V-3B EC L-1B6 L-1B7 L-1B2 L-1B3 MP1699 A-3B1 A-2B3 A-1S A-2B1 A-6B3 A-6B1 A-5B3 A-5B2 A-4B2 A-4B3 H-1B4 P-4S T-2B3 V-5B3 P-1B1 P-2B1 K-4B1 K-5B3 K-1B3 K-5B2 S-1B1 S-2S2 MP1681 MP1689 MP1687 MP1652 MP1653 MP1713 MP1685 MP1709 MP1708 MP1755 MP1804 MP1705 MP1794 MP1787 MP1680 MP1698 MP1728 MP1765 MP1703 MP1706 MP1781 MP1783 MP1776 MP1784 MP1753 MP1796 MP1721 MP1774 MP1772 MP1768 MP1764 MP1712 MP1711 MP1707 MP1692 MP1702 MP1695 MP1782 MP1704 MP1700 MP1688 MP1769 MP1792 MP1778 MP1727 MP1683 MP1725 0,08 0,18 0, UPGMA (Jaccard s similarity coefficient) 12 SSR loci 96 P. infestans isolates 66 genotypes (7 clusters) 0,38 0,48 0,58 0,68 0,78 0,88 0,98
20 Ri-1B1 P-3S P-3B1 P-5S MP1710 M-1B3 HS-3B2 HS-3S HS-4S3 HS-3B3 HS-3B1 HS-1B1 HS-2B1 HS-4S2a HS-4S2b Ri-1B3 V-3B EC L-1B6 L-1B7 L-1B2 L-1B3 MP1699 A-3B1 A-2B3 A-1S A-2B1 A-6B3 A-6B1 A-5B3 A-5B2 A-4B2 A-4B3 H-1B4 P-4S T-2B3 V-5B3 P-1B1 P-2B1 K-4B1 K-5B3 K-1B3 K-5B2 S-1B1 S-2S2 MP1681 MP1689 MP1687 MP1652 MP1653 MP1713 MP1685 MP1709 MP1708 MP1755 MP1804 MP1705 MP1794 MP1787 MP1680 MP1698 MP1728 MP1765 MP1703 MP1706 MP1781 MP1783 MP1776 MP1784 MP1753 MP1796 MP1721 MP1774 MP1772 MP1768 MP1764 MP1712 MP1711 MP1707 MP1692 MP1702 MP1695 MP1782 MP1704 MP1700 MP1688 MP1769 MP1792 MP1778 MP1727 MP1683 MP1725 0,08 0,18 0, UPGMA (Jaccard s similarity coefficient) 12 SSR loci 96 P. infestans isolates 66 genotypes (7 clusters) 0,38 0,48 0,58 0,68 0,78 0,88 0,98
21 SSR genotype / effectors alleles alleles combinations origin SSR genotype number of isolates Avr-vnt1 AvrSmira Norway Poland
22 SSR genotype / effectors alleles alleles combinations origin SSR genotype number of isolates Avr-vnt1 AvrSmira Norway Poland
23 conclusions Avr-vnt1 high conservation polymorphisms unrelated to virulence no selection pressure AvrSmira1 highly diversified (most polymorphic sites located in C-terminal region) positions 156 and 170 may be related to virulence not the only mechanism of avoiding recognition (presence of virulent isolates without the M 156 R 170 polymorphism) selection pressure acting on amino acid positions eight positions consistent with previous research (Rietman et al., 2012) isolates belonging to the same SSR genotype contain different alleles of the Avr-vnt1 (1 Polish SSR genotype) and AvrSmira1 (1 Norwegian, 4 Polish) two/multi-compartment genome
24 Diversity of chosen effectors in samples of Polish and Norwegian populations of Phytophthora infestans Emil Stefańczyk THANKS FOR YOUR ATTENTION
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