Characterization of Diaporthe ampelina isolates and their Sensitivity to Hot-Water Treatments and Fungicides in in vitro
Abstract
Diaporthe ampelina (=Phomopsis viticola) is one of the most important pathogens causing both cane/leaf spot and wood canker diseases in grape growing countries in the world. In this research, morphological, molecular and pathogenic characterization of 23 D. ampelina isolates were studied and their sensitivity was tested against hot-water treatments and some of the fungicides used in vineyards. Morphologically, the isolates were grouped according to “W type” and “G type” colony appearance and microscopic features. In molecular characterization, beta-tubulin, calmodulin and translation elongation factor (tef1-α) gene regions were amplified with PCR. The nucleotide sequences were analyzed using NCBI-BLAST search and recorded in GenBank, through which species identity was also confirmed. Mycelial viability was tested against hot-water treatments (46 – 50°C for 30 and 45 min) in centrifuge tubes and thermal inactivation point was determined. It was also tested against some of the fungicides (azoxystrobin, boscalid, cyprodinil, tebuconazole, azoxystrobin + cyproconazole + tebuconazole, cyprodinil + fludioxonil, azoxystrobin + tebuconazole and fludioxonil) in vitro and EC50 values were calculated. The morphological and molecular study results showed that all the isolates were D. ampelina and they were pathogenic on wood tissues of vines. Thermal inactivation of “W type” isolates was ensured at 48°C-30 min hot-water treatments. Although this treatment also reduced colony growth of “G type” isolates, it did not inhibit it completely and 48°C-45 min treatment was needed to reach full eradication. Considering fungicide sensitivity, fludioxonil or tebuconazole containing fungicides were the most effective in suppressing the mycelial growth of the fungus. However, azoxystrobin, boscalid, cyprodinil could not perform a strong inhibition when compared to fludioxonil and tebuconazole.
Supporting Institution
Çukurova Üniv. BAP Birimi Koordinatörlüğü
Project Number
FYL-2017-8158
Thanks
This study was supported by the Cukurova University Scientific Research Projects Department, in Project FYL-2017-8158, and was a MSc Thesis Project. The authors thank The Rectorate of Cukurova University.
References
- Akgül DS, Savaş NG, Teker T, Keykubat B, Mayorquin JS, Eskalen A 2015. Fungal Trunk Pathogens of Sultana Seedless Vineyards in Aegean Region of Turkey. Phytopathol Mediterr 54 (2):380-393.
- Akgül DS, Ahioğlu M 2019. Fungal pathogens associated with young grapevine decline in the Southern Turkey vineyards. 42. World Congress of Vine & Wine, 15-19 July 2019, Geneva, Switzerland, 15 (01027):1-6.
- Baumgartner K, Fujiyoshi PT, Travadon R, Castlebury LA, Wilcox WF, Rolshausen PE 2013. Characterization of Species of Diaporthe from Wood Cankers of Grape in Eastern North American vineyards. Plant Dis 97: 912-920.
- Carbone L, Kohn LM 1995. A method for Designing Primer Sets for Speciation Studies in Filamentous Ascomycetes. Mycologia 91: 553–556.
- Caudwell A, Larrue J, Boudon-Padieu E, McLean GD 1997. Flavescence Dorèe Elimination from Dormant Wood of Grapevines by Hot-Water Treatment. Aust J Grape Wine R 3: 21–25.
- Crous PW, Swart L, Coertze S 2001. The Effect of Hot-Water Treatment on Fungi Occurring in Apparently Healthy Grapevine Cuttings. Phytopathol Mediterr 40: 464–466.
- Dissanayake AJ, Liu M, Zhang W 2015. Morphological and Molecular Characterization of Diaporthe Species Associated with Grapevine Trunk Disease in China. Fungal Biol 119: 283–294.
- Elena G, Di Bella V, Armengol J, Luque J 2015. Viability of Botryosphaeriaceae Species Pathogenic to Grapevine After Hot Water Treatment. Phytopathol Mediterr 54: 325–334.
- Erincik O, Madden LV, Ferree DC, Ellis MA 2001. Effect of Growth Stage on Susceptibility of Grape Berry and Rachis Tissues to Infection by Phomopsis viticola. Plant Dis 85:517-520.
- Gessler NN, Egorova AS, Belozerskaia TA 2014. Melanin Pigments of Fungi Under Extreme Environmental Conditions (review). Prikl Biokhim Mikrobiol 50: 125–134.
- Glass NL, Donaldson GC 1995. Development of Primer Sets Designed for Use with the PCR to Amplify Conserved Genes from the Filamentous Ascomycetes. Appl Environ Microbiol 61: 1323–1330.
- Gomes RR, Glienke C, Videira SIR, Lombard R, Groenewald JZ, Crous PW 2013. Diaporthe: A Genus of Endophytic, Saprobic and Plant Pathogenic Fungi. Persoonia 31: 1–41.
- Gomez KA, Gomez AA 1984. Statistical Procedures for Agricultural Research. 2nd ed. Wiley, New York, USA. Görür V, Akgül DS 2019. Fungicide Suspensions Combined with Hot-Water Treatments Affect Endogenous Neofusicoccum parvum Infections and Endophytic Fungi in Dormant Grapevine Canes. Phytopathol Mediterr 58(3): 559-571.
- Gramaje D, Garcia-Jimenez J, Armengol J 2008. Sensitivity of Petri disease Pathogens to Hot-Water Treatments in vitro. Ann Appl Biol 153: 95-103.
- Gramaje D, Aroca A, Raposo R, Garcia-Jimenez J, Armengol J 2009. Evaluation of fungicides to control Petri disease pathogens in the grapevine propagation process. Crop Prot 28: 1091-1097.
- Guarnaccia V, Groenewald JZ, Woodhall J, Armengol J, Cinelli T 2018. Diaporthe Diversity and Pathogenicity Revealed from A Broad Survey of Grapevine Diseases in Europe. Persoonia 40: 135–153.
- Howard RJ, Valent B 1996. Breaking and Entering: Host Penetration by the Fungal Rice Blast Pathogen Magnoporthe Grisea. Annu Rev. Microbiol 50: 491-512.
- Kanematsu S, Kobayashi T, Kudo A, Ohtsu Y 1999. Conidial Morphology, Pathogenicity and Culture Characteristics of Phomopsis Isolates from Peach, Japanese Pear and Apple in Japan. Ann Phytopathol Soc Jpn 65: 264–273.
- Kim DS, Park SH, Kwon SB, Joo YH, Youn SW, Sohn UD, Park KC 2003. Temperature Regulates Melanin Synthesis in Melanocytes. Arch Pharm Res 26:840–845.
- Mostert L., Denman S, Crous PW (2000) In vitro Screening of Fungicides Against Phomopsis viticola and Diaporthe perjuncta. S. Afr. J. Enol Vitic. 21 (2): 62-65.
- Mostert L, Crous PW, Kang JC, Phillips AJL 2001. Species of Phomopsis and a Libertella sp. Occurring on Grapevines with Specific Reference to South Africa: Morphological, Cultural, Molecular and Pathological Characterization. Mycologia 93: 146-167.
- O’Donnell K, Cigelnik E, Nirenberg HI 1998. Molecular Systematics and Phylogeography of the Gibberella Fujikuroi Species Complex. Mycologia 90(3): 465-493.
- Rawnsley B, Wicks TJ, Scott ES, Stummer BE 2004. Diaporthe perjuncta Does Not Cause Phomopsis Cane and Leaf Spot Disease of Grapevine in Australia. Plant Dis 88: 1005–1010.
- Rehnstrom AL, Free SJ 1996. The Isolation and Characterization of Melanin Deficient Mutants of Monilinia fructicola. Physiol Mol Plant P 49: 321–330.
- Rego C, Nascimento T, Cabral A, Silva MJ, Oliviera H 2009. Control of Grapevine Wood Fungi in Commercial Nurseries. Phytopathol. Mediterr 48: 128-135.
- Rooney SN, Gubler WD 2001. Effect of Hot Water Treatments on Eradication of Phaeomoniella chlamydospora and Phaeoacremonium inflatipes from Dormant Grapevine Wood. Phytopathol Mediterr 40: 467–472.
- Schilder AMC, Erincik O, Castlebury L, Rossman A, Ellis MA 2005. Characterization of Phomopsis spp. Infecting Grapevines in the Great Lakes Region of North America. Plant Dis 89: 755–762.
- Sosnowski MR, Loschiavo AP, Wicks TJ, Scott ES 2013. Evaluating Treatments and Spray Application for the Protection of Grapevine Pruning Wounds from Infection by Eutypa lata. Plant Dis. 97: 1599-1604.
- Úrbez-Torres JR, Peduto F, Smith RJ, Gubler WD 2013. Phomopsis dieback: A Grapevine Trunk Disease Caused by Phomopsis viticola in California. Plant Dis. 97: 1571-1579.
- Waite H, Armengol J, Billones-Baaijens R, Gramaje D, Halleen F 2018. A Protocol for the Management of Grapevine Rootstock Mother Vines to Reduce Latent Infections by Grapevine Trunk Pathogens in Cuttings. Phytopathol Mediterr 57: 384−398.
Diaporthe ampelina İzolatlarının Karakterizasyonu ve in vitro’da Bazı Fungisitlere ve Sıcak Su Uygulamalarına Duyarlılıkları
Abstract
Diaporthe ampelina (=Phomopsis viticola) dünyadaki üzüm yetiştirilen ülkelerde hem sürgün/yaprak lekesi hem de odun kangrenine neden olan en önemli patojenlerden biridir. Bu araştırmada, 23 D. ampelina izolatının morfolojik, moleküler ve patojenik karakterizasyonu çalışılmış ve bunların bağcılıkta kullanılan bazı fungisitlere ve sıcak su uygulamalarına karşı duyarlılığı test edilmiştir. Morfolojik olarak izolatlar, “W” tipi ya da “G” tipi koloni görünümleri ve mikroskobik özelliklerine göre gruplandırılmıştır. Moleküler karakterizasyonda beta-tubulin, calmodulin ve translation elongation factor (tef1-α) gen bölgeleri PCR ile çoğaltılmıştır. Nükleotid dizileri NCBI-BLAST yazılımı kullanarak analiz edilmiş, gen bankasına kaydedilmiş ve tür tanısı doğrulanmıştır. Miseliyal canlılık sıcak su uygulamalarına karşı (46–50°C’de 30-45 dk.) santrifüj tüplerinde test edilmiş ve termal inaktivasyon noktası saptanmıştır. Bu aynı zamanda in vitro’da bazı fungisitlere karşı (azoxystrobin, boscalid, cyprodinil, tebuconazole, azoxystrobin + cyproconazole + tebuconazole, cyprodinil + fludioxonil, azoxystrobin + tebuconazole and fludioxonil) test edilmiş ve EC50 değerleri hesaplanmıştır. Morfolojik ve moleküler çalışma sonuçları tüm izolatların D. ampelina olduğunu ve bunların asma odunsu dokularında patojen olduklarını göstermiştir. “W” tipi izolatların termal inaktivasyonu 48°C-30 dakikalık sıcak su uygulamasıyla sağlanmıştır. Bu uygulama “G” tipi izolatların koloni büyümesini azaltmasına rağmen tamamen inhibe etmemiş, tam eradikasyona ulaşmak için 48°C-45 dakikalık uygulamaya gerek duyulmuştur. Fungisit duyarlılığı ele alındığında, fludioxonil ve tebuconazole içeren fungisitler fungusun miseliyal büyümesini baskılamada en etkili olmuşlardır. Ancak azoxystrobin, boscalid ve cyprodinil, fludioxonil ve tebuconazole ile kıyaslandığında güçlü bir inhibisyon sergileyememiştir.
Project Number
FYL-2017-8158
References
- Akgül DS, Savaş NG, Teker T, Keykubat B, Mayorquin JS, Eskalen A 2015. Fungal Trunk Pathogens of Sultana Seedless Vineyards in Aegean Region of Turkey. Phytopathol Mediterr 54 (2):380-393.
- Akgül DS, Ahioğlu M 2019. Fungal pathogens associated with young grapevine decline in the Southern Turkey vineyards. 42. World Congress of Vine & Wine, 15-19 July 2019, Geneva, Switzerland, 15 (01027):1-6.
- Baumgartner K, Fujiyoshi PT, Travadon R, Castlebury LA, Wilcox WF, Rolshausen PE 2013. Characterization of Species of Diaporthe from Wood Cankers of Grape in Eastern North American vineyards. Plant Dis 97: 912-920.
- Carbone L, Kohn LM 1995. A method for Designing Primer Sets for Speciation Studies in Filamentous Ascomycetes. Mycologia 91: 553–556.
- Caudwell A, Larrue J, Boudon-Padieu E, McLean GD 1997. Flavescence Dorèe Elimination from Dormant Wood of Grapevines by Hot-Water Treatment. Aust J Grape Wine R 3: 21–25.
- Crous PW, Swart L, Coertze S 2001. The Effect of Hot-Water Treatment on Fungi Occurring in Apparently Healthy Grapevine Cuttings. Phytopathol Mediterr 40: 464–466.
- Dissanayake AJ, Liu M, Zhang W 2015. Morphological and Molecular Characterization of Diaporthe Species Associated with Grapevine Trunk Disease in China. Fungal Biol 119: 283–294.
- Elena G, Di Bella V, Armengol J, Luque J 2015. Viability of Botryosphaeriaceae Species Pathogenic to Grapevine After Hot Water Treatment. Phytopathol Mediterr 54: 325–334.
- Erincik O, Madden LV, Ferree DC, Ellis MA 2001. Effect of Growth Stage on Susceptibility of Grape Berry and Rachis Tissues to Infection by Phomopsis viticola. Plant Dis 85:517-520.
- Gessler NN, Egorova AS, Belozerskaia TA 2014. Melanin Pigments of Fungi Under Extreme Environmental Conditions (review). Prikl Biokhim Mikrobiol 50: 125–134.
- Glass NL, Donaldson GC 1995. Development of Primer Sets Designed for Use with the PCR to Amplify Conserved Genes from the Filamentous Ascomycetes. Appl Environ Microbiol 61: 1323–1330.
- Gomes RR, Glienke C, Videira SIR, Lombard R, Groenewald JZ, Crous PW 2013. Diaporthe: A Genus of Endophytic, Saprobic and Plant Pathogenic Fungi. Persoonia 31: 1–41.
- Gomez KA, Gomez AA 1984. Statistical Procedures for Agricultural Research. 2nd ed. Wiley, New York, USA. Görür V, Akgül DS 2019. Fungicide Suspensions Combined with Hot-Water Treatments Affect Endogenous Neofusicoccum parvum Infections and Endophytic Fungi in Dormant Grapevine Canes. Phytopathol Mediterr 58(3): 559-571.
- Gramaje D, Garcia-Jimenez J, Armengol J 2008. Sensitivity of Petri disease Pathogens to Hot-Water Treatments in vitro. Ann Appl Biol 153: 95-103.
- Gramaje D, Aroca A, Raposo R, Garcia-Jimenez J, Armengol J 2009. Evaluation of fungicides to control Petri disease pathogens in the grapevine propagation process. Crop Prot 28: 1091-1097.
- Guarnaccia V, Groenewald JZ, Woodhall J, Armengol J, Cinelli T 2018. Diaporthe Diversity and Pathogenicity Revealed from A Broad Survey of Grapevine Diseases in Europe. Persoonia 40: 135–153.
- Howard RJ, Valent B 1996. Breaking and Entering: Host Penetration by the Fungal Rice Blast Pathogen Magnoporthe Grisea. Annu Rev. Microbiol 50: 491-512.
- Kanematsu S, Kobayashi T, Kudo A, Ohtsu Y 1999. Conidial Morphology, Pathogenicity and Culture Characteristics of Phomopsis Isolates from Peach, Japanese Pear and Apple in Japan. Ann Phytopathol Soc Jpn 65: 264–273.
- Kim DS, Park SH, Kwon SB, Joo YH, Youn SW, Sohn UD, Park KC 2003. Temperature Regulates Melanin Synthesis in Melanocytes. Arch Pharm Res 26:840–845.
- Mostert L., Denman S, Crous PW (2000) In vitro Screening of Fungicides Against Phomopsis viticola and Diaporthe perjuncta. S. Afr. J. Enol Vitic. 21 (2): 62-65.
- Mostert L, Crous PW, Kang JC, Phillips AJL 2001. Species of Phomopsis and a Libertella sp. Occurring on Grapevines with Specific Reference to South Africa: Morphological, Cultural, Molecular and Pathological Characterization. Mycologia 93: 146-167.
- O’Donnell K, Cigelnik E, Nirenberg HI 1998. Molecular Systematics and Phylogeography of the Gibberella Fujikuroi Species Complex. Mycologia 90(3): 465-493.
- Rawnsley B, Wicks TJ, Scott ES, Stummer BE 2004. Diaporthe perjuncta Does Not Cause Phomopsis Cane and Leaf Spot Disease of Grapevine in Australia. Plant Dis 88: 1005–1010.
- Rehnstrom AL, Free SJ 1996. The Isolation and Characterization of Melanin Deficient Mutants of Monilinia fructicola. Physiol Mol Plant P 49: 321–330.
- Rego C, Nascimento T, Cabral A, Silva MJ, Oliviera H 2009. Control of Grapevine Wood Fungi in Commercial Nurseries. Phytopathol. Mediterr 48: 128-135.
- Rooney SN, Gubler WD 2001. Effect of Hot Water Treatments on Eradication of Phaeomoniella chlamydospora and Phaeoacremonium inflatipes from Dormant Grapevine Wood. Phytopathol Mediterr 40: 467–472.
- Schilder AMC, Erincik O, Castlebury L, Rossman A, Ellis MA 2005. Characterization of Phomopsis spp. Infecting Grapevines in the Great Lakes Region of North America. Plant Dis 89: 755–762.
- Sosnowski MR, Loschiavo AP, Wicks TJ, Scott ES 2013. Evaluating Treatments and Spray Application for the Protection of Grapevine Pruning Wounds from Infection by Eutypa lata. Plant Dis. 97: 1599-1604.
- Úrbez-Torres JR, Peduto F, Smith RJ, Gubler WD 2013. Phomopsis dieback: A Grapevine Trunk Disease Caused by Phomopsis viticola in California. Plant Dis. 97: 1571-1579.
- Waite H, Armengol J, Billones-Baaijens R, Gramaje D, Halleen F 2018. A Protocol for the Management of Grapevine Rootstock Mother Vines to Reduce Latent Infections by Grapevine Trunk Pathogens in Cuttings. Phytopathol Mediterr 57: 384−398.
Details
| Primary Language | English |
|---|---|
| Subjects | Agricultural, Veterinary and Food Sciences |
| Journal Section | RESEARCH ARTICLE |
| Authors | |
| Project Number | FYL-2017-8158 |
| Publication Date | December 30, 2022 |
| Submission Date | November 7, 2021 |
| Acceptance Date | December 19, 2021 |
| Published in Issue | Year 2022Volume: 25 Issue: 6 |
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