Моnilinia fructicola a potential threat for sweet and sour cherry production in Serbia

Transcription

1 Моnilinia fructicola a potential threat for sweet and sour cherry production in Serbia Brankica Tanović and Jovana Hrustić Institute of Pesticides and Environmental Protection Belgrade, Serbia brankica.tanovic@pesting.org.rs

2 Stone fruit production 30% fresh or frozen 70% processed

3 Stone fruit production in the world Million tons Sour cherry Višnja Sweet Trešnja cherry Apricot Kajsija Plum Šljiva Breskva Peach godina godina *FAOSTAT, 2011 *FAOSTAT, Million tons

4 Stone fruit production in Serbia Pome fruits 27% Berry fruits 11% Stone fruits 61% Sour cherry Višnja Sweet cherry Trešnja Apricot Kajsija Nuts 2% Plum Šljiva Peach Breskva

5 Disease symptoms Stigmina carpophila Glomerella cingulata Blumeriella jaapii, Glomerella cingulata, Monilinia spp., Stigmina carpophila, Apiognomonia erythrostoma, Armillaria mellea, Pythium spp., Rhizopus stolonifer, Chondrostereum purpureum, Tranzschelia pruni-spinosae, Verticillium spp. Apiognomonia erythrostoma Blumeriella jaapii Monilinia laxa

6 Monilinia spp.

7 Monilinia spp. causal agents of blossom and twig blight and fruit rot of stone fruits Worldwide distribution in all major fruit growing areas Cause one of the most important pre- and postharvest fungal diseases of stone fruits Three species of this genus are economically important

8 Monilinia laxa economically the most important blossom blight twig and branch blight fruit rot stone fruits pome fruits apricot cherry peach sour cherry plum Monilinia fructigena pome fruits fruit rot plum stone fruits blossom, twig and branch blight Monilinia fructicola the most destructive species of the genus Monilinia fruit rot including latent infection peach and nectarine

9 Distribution of Monilinia spp. in the world M. M. fructigena fructicola laxa M. laxa M. fructigena M. fructicola

10 Distribution of Monilinia fructicola in Europe detected not known absence confirmed

11 Separation of species of the genus Monilinia 1 Symptoms Morphological and cultural characteristics 2 3 Molecular identification

12 Monilinia spp. control Breeding for resistance, Cultural practice, Sanitation, Biological control, Chemical control. Fungicides registered: azoxistrobin, bitertalon, ciram, dichlofluanid, dikoran, dithianon, fenhexamid, fludioxonil, fluopyram, fluoroimid, imibenconazole, captan, mepanipyrim, oksokonazol fumarate, propioneb, triadimefon, triflumizole, thiram, triforine

13 Tasks To establish presence and distribution of Monilinia spp. on stone fruits in Serbia To complete identification procedure to the species level To determine fitness components of the isolates of all found species of the genus Monilinia in order to assess the risk of spreading of quarantine pathogen M. fructicola in stone fruit orchards: growth rate at different temperatures temperature range of the growth growth rate at different media ph range of the growth sporulation virulence sensitivity to fungicides To analyze potential ways of introduction and spreading of M. fructicola

14 Sampling hosts 119 locations 15 regions 362 samples

15 Samples

16 Isolation and pathogenicity test Isolation Origin of the isolates Number of isolates mature fruit 232 mummified fruit 45 twigs 11 total 288 Artificial inoculation wounded detached fruits unwounded detached fruits

17 Pathogenic characteristics 1 Identification 2 Morphological, ecological and cultural characteristics Sequencing of the ITS rdna region 3

18 Morphological characteristics M a c r o s c o p i c Key for identification of Monilinia spp.: colony colour colony margin rosetting sporulation presence of sporulation in concentric rings presence of black arcs quantitative growth rate M i c r o s c o p i c shape and size of the conidia germ tube length and morphology

19 I group 257 isolates II group 8 isolates III group 23 isolates Colony characteristics - light to dark gray - lobed margins - absent sporulation - absent concentric rings of spores - creamy to yellow - entire margins - rare sporulation - concentric rings of spores - black arcs - hazel zonate - even margins - abundant sporulation - concentric rings of spores

20 I group II group III group Conidia characteristics μm x μm μm x μm μm x μm 40 µm 197 µm 365 µm

21 I group 257 isolates II group 8 isolates III group 23 isolates Population structure Identification

22 2011 one isolate M. fructicola on green market nectarine 2012 (217 samples) two isolates M. fructicola on green market no isolates in the field nectarine isolates M. fructicola in three independent orchards nectarine, peach, apricot, plum

23 Tasks To establish presence and distribution of Monilinia spp. on stone fruits in Serbia To complete identification procedure to the species level. To determine fitness components of the isolates of all found species of the genus Monilinia in order to assess the risk of spreading of quarantine pathogen M. fructicola in stone fruit orchards: growth rate at different temperatures temperature range of the growth growth rate at different media ph range of the growth sporulation virulence sensitivity to fungicides To analyze potential ways of introduction and spreading of M. fructicola

24 Growth rate of the isolates Monilinia spp. 16 isolates PDA medium Seven days at 23 o C ANOVA, software Statistika M. laxa M. fructigena M. fructicola

25 M. laxa M. fructigena M. fructicola Growth rate at different temperatures 16 isolates PDA o C Seven days ANOVA, software Statistika Temperatures

26 M. laxa M. fructigena M. fructicola Growth rate at extreme temperatures 16 isolates PDA Temperatures 0-4 o C Temperatures o C Seven days ANOVA, software Statistika Growth rate (mm/7days) Temperatures 0 C 1 C 2 C 3 C 4 C TPGR ŠMLEs ŠPSV TPBN ŠMLE VGRSD BPSV BMLEs TPAL VGRSE VPBG TPGO ŠPPR NPGM NPUD1 NPUD2 45 Growth rate (mm/7days) C 32 C 33 C 34 C 35 C TPGR ŠMLEs ŠPSV TPBN ŠMLE VGRSD BPSV BMLEs TPAL VGRSE VPBG TPGO ŠPPR NPGM NPUD1 NPUD2

27 Growth rate on different media Potato-dextrose agar PDA Water agar - WA Czapek agar CzA Malt agar - MA Vegetable-8 juice agar - V8A 12 isolates Seven days ANOVA, software Statistika Media M. laxa M. fructigena M. fructicola

28 Growth rate on different ph 12 isolates PDA ph 2 -ph 12 Seven days at 24 C ANOVA, software Statistika M. laxa M. fructigena M. fructicola

29 Sporulation M. laxa M. fructigena M. fructicola 11,2-15,4 μm x 8,2-11,1 μm 19,2-23,5 μm x 11,9-13,5 μm 16,0-16,9 μm x 9,9-10,7 μm 40 µm 197 µm 365 µm Conidia production 0.29 x 10 4 /cm x 10 4 /cm x 10 4 /cm 2

30 Sporulation Control Wounded mature plum fruits Artificial inoculation Isolates M. laxa M. fructigena M. fructicola Mildora Stenley Čačanska rodna Čačanska lepotica

31 Sporulation Wounded immature plum fruits Control Artificial inoculation Isolates M. laxa M. fructigena M. fructicola Mildora Stenley M. laxa Čačanska rodna Čačanska lepotica

32 Virulence test Nine isolates: four isolates M. laxa four isolates M. fructigena one isolate M. fructicola Artificial inoculation of wounded mature stone fruits: peach, apricot, nectarine, plum, sweet cherry, sour cherry Total: 15 fruits/species/isolate Incubation in moisture chamber at 24 o C ANOVA, software Statistika

33 Virulence 60 lesion expansion rate (mm/3 days) Nectarine Peach Plum Apricot TPGR ŠPSV VGRSE BMLEs TPGO ŠPPR VPBG BŠPBA NPGM M. laxa M. fructigena M. fructicola

34 Virulence lesion expansion rate (mm/2 days) BMLEs VPBG NPGM 0 Sweet cherry Sour cherry M. laxa M. fructigena M. fructicola

35 Sensitivity of Monilinia spp. isolates to fungicides from different chemical groups 12 isolates 7 fungicides Probit analysis Parameters: EC-50 (mg/l) b (slope of regression line) Product Company Active substance Chemical group A.S. content Concentration range (mg/l) Bravo 720 SC Syngenta Chlorothalonil Chloronitrile 720 g/l 2; 1; 0,5; 0,25; 0,125 i 0,06 Kidan EC Bayer 10; 5; 2,5; 1,25; 0,6; 0,5; 0,3; Iprodione Dicarboximides 255 g/kg CropScience 0,25; 0,125; 0,06 i 0,03 Spartak 450-EC Sinochem 0,05; 0,025; 0,0125; 0,006 i Prochloraz Imidazoles 500 g/kg Ningbo 0,003 Quadris Syngenta Azoxystrobin Strobilurins 250 g/kg Akord Galenika Fitofarmacija Tebuconazole Triazole 250 g/kg Cantus BASF Boscalid Carboxamide 500 g/kg Luna Privilege Bayer CropScience Fluopyram Pyridinyl-ethylbenzamides 500 g/l 100; 25; 10; 5; 1; 0,1; 0,01; 0,001 0,5; 0,25; 0,125; 0,06; 0,05; 0,03; 0,025; 0,0125 i 0, ; 1500; 1000; 750; 500; 0,1; 0,05; 0,025; 0,0125 i 0, ; 500; 250; 125; 62,5; 2; 1; 0,5; 0,25; 0,125 i 0,06

36 Sensitivity of Monilinia spp. isolates to fungicides from different chemical groups M. laxa M. fructigena M. fructicola Active substance EC 50 (mg/l) Chlorothalonil Iprodione Prochloraz Azoxystrobin Tebuconazole Boscalid Fluopyram M. laxa M. fructigena M. fructicola M. laxa M. fructigena M. fructicola Monilinia fructicola: Sensitivity of NPGM isolate on iprodione

37 Tasks To establish presence and distribution of Monilinia spp. on stone fruits in Serbia To complete identification procedure to the species level. To determine fitness components of the isolates of all found species of the genus Monilinia in order to assess the risk of spreading of quarantine pathogen M. fructicola in stone fruit orchards: growth rate at different temperatures temperature range of the growth growth rate at different media ph range of the growth sporulation virulence sensitivity to fungicides 4 To analyze potential ways of introduction and spreading of M. fructicola

38 Tasks 3 To determine fitness components of the isolates of all found species of the genus Monilinia in order to assess the risk of spreading of quarantine pathogen M. fructicola in stone fruit orchards: growth rate at different temperatures temperature range of the growth growth rate at different media ph range of the growth sporulation virulence sensitivity to fungicides 4 To analyze potential ways of introduction and spreading of M. fructicola

39 SSR marker analysis of M. fructicola isolates from Serbia Three isolates M. fructicola originating from Serbia Multiplex PCR kit Termocycler (SensoQuest) Protocol: 95 o C / 15 min 94 o C / 40 s 50 o C / 90 s 72 o C / 90 s 60 o C / 30 min 35 cycles ABI PRISM 3100 DNK capillary sequencer (Applied Biosystems)

40 SSR marker analysis of M. fructicola isolates Allele (in bp) composition of M. fructicola haplotypes found in Serbian samples Isolate CHMFc5 CHMFc1 CHMFc4 CHML5 CHMFc12 NPUD1 86 bp 173 bp 216 bp 220 bp 160 bp US50 NPUD2 86 bp 173 bp 216 bp 220 bp 160 bp US50 NPGM 86 bp 173 bp 223 bp 220 bp 160 bp ITA01 Haplotype according to Jansch et al. (2012)

41 plums peach/nectarine apricots sour cherries Vojvodina Production area (ha) Belgrade Šumadija and Western Serbia 0 Belgrade Vojvodina Sumadija and Western Serbia Southern and Eastern Serbia Kosovo and Metohija Kosovo and Metohija Southern and Eastern Serbia

42 Projects: III COST FA 1104