University of Agriculture in Krakow, Cracow, Poland FUNGI INHABITING THE RHIZOSPHERE OF PERSIAN CYCLAMEN AND THEIR IMPACT ON POTENTIAL PATHOGENS OF THE PLANT H. Kurzawińska Abstract The diseased cyclamen plants were populated by a number of fungal species, predominantly by: Fusarium oxysporum, Cylindrocarpon destructans and Alternaria alternata. All the mentioned species were pathogenic to cyclamen roots but to different extent. The community of the cyclamen rhizosphere fungi supported the growth of the pathogens under investigation. Key words: Persian cyclamen, rhizosphere, Alternaria alternata, Cylindrocarpon destructans, Fusarium oxysporum, biotic relations Introduction Persian cyclamen (Cyclamen persicum)belongs to most commonly grown pot plants. The cultivars of longer pedicels are also grown for cut flowers (Startek and Mynett 2003). The most common pathogenic fungi occurring on this plant include: Fusarium oxysporum f. sp. cyclaminis, causing leaf wilting and bulb rot, Cylindrocarpon destructans and Rhizoctonia solani, both responsible for root rot, and Botrytis cinerea causing grey mould (Gabarkiewicz et al. 1995, Orlikowski 2000). The accumulation of pathogenic fungi that infest the plant through the root system can lead to infection and development of the disease process. Among saprotrophic microorganisms there may be also species antagonistic to pathogenic fungi, able to reduce its aggressiveness (Kurzawińska 2006, Kurzawińska and Duda 2005, Mańka 1990). The understanding of the structure and phytopathological function of fungal community can bring information, very important from the environmental point of view, on plant disease condition and ability to control such diseases. Phytopathologia 57: 5 10 The Polish Phytopathological Society, Poznań 2010 ISSN 2081-1756
6 H. Kurzawińska The aim of this study was to identify fungi populating the infested plants and to recognize fungal species composition in the rhizosphere of Persian cyclamen and its effect on the growth of Alternaria alternata, Cylindrocarpon destructans and Fusarium oxysporum. Material and methods The study was carried out on diseased Persian cyclamen plants showing symptoms of wilting, from a private horticultural farm located in Southern Poland (Małopolska province). Isolation of microorganisms from diseased cyclamen bulbs and roots and further culturing, was previously described in detail by the author (Kurzawińska 2006). The isolated fungi were identified according to identification keys and monographs by Booth (1966), Domsch et al. (1980), Nelson et al. (1983) and Ramirez (1982). The isolates selected for further analysis were tested for their pathogenicity to cyclamen. The following three fungal species, densely populating the cyclamen plants, were selected: Alternaria alternata, Cylindrocarpon destructans and Fusarium oxysporum. The tests were carried out under laboratory conditions. Healthy roots of Persian cyclamen were surface disinfected with 70% ethyl alcohol and rinsed in sterile distilled water. After drying in filter paper, the roots were placed in cuvettes lined with wet filter paper. Agar disks (3 mm diam.)with mycelium taken from 14-day-old cultures growing on potato dextrose agar (PDA)at 22 23 o C were placed on each root. The cuvettes were covered tightly with a film and incubated at 22 23 C. The experiment was performed in the randomized block design with four repetitions, each having five roots. The degree of pathogenicity was expressed by the surface area of necrosis caused by fungi under consideration and measured after 14 days of incubation. The results were analyzed statistically by analysis of variance. The significance of differences between combinations was estimated with Duncan s test at = 0.05. Fungi populating the rhizosphere of Persian cyclamen were isolated with the method described by Mańka and Mańka (1993). To recognize biotic functions of the fungal community populating the cyclamen rhizosphere to A. alternata, C. destructans and F. oxysporum, the most commonly isolated saprotrophic species (85% of total isolates)were tested. The tests were carried out with the Mańka biotic series method (1974). Results From diseased bulbs and roots of Persian cyclamen 83 fungal colonies, representing 10 genera and 13 species, and 10 bacterial colonies were isolated (Table 1). The following fungi were most frequent: F. oxysporum (20.5%), C. destructans (13.3%)and A. alternata (12.1%). Also C. radicicola, F. avenaceum (10.8% each)and
Fungi inhabiting the rhizosphere of Persian cyclamen... 7 Table 1 Microorganisms isolated from diseased tubers and roots of Cyclamen persicum Fungal species Number of isolates Percentage Fusarium oxysporum 17 20.5 Cylindrocarpon destructans 11 13.3 Alternaria alternata 10 12.1 Botrytis cinerea 8 9.6 Cylindrocarpon radicicola 9 10.8 Fusarium avenaceum 9 10.8 Penicillium verrucosum var. corymbiferum 5 6.0 Rhizoctonia solani 4 4.8 Penicillium expansum 2 2.4 Phytophthora cactorum 2 2.4 Thielaviopsis basicola 2 2.4 Trichoderma viride 2 2.4 Verticillium albo-atrum 2 2.4 Total 83 100.0 Bacteria 10 Botrytis cinerea (9.6%)were quite frequently found. The remaining less numerous species included: Penicillium verrucosum var. corymbiferum (6.0%), Rhizoctonia solani (4.8%), Verticillium albo-atrum, P. expansum, Phytophthora cactorum, Thielaviopsis basicola and Trichoderma viride 2.4% each (Table 1). In the pathogenicity tests the fungal species under consideration caused necrotic changes in the inoculated cyclamen roots. Fusarium oxysporum displayed the highest pathogenic activity, as compared to those of other species (Fig. 1). Both F. oxysporum and C. destructans as well as A. alternata were isolated from the diseased spots, thus confirming their pathogenicity. A total of 178 fungal colonies were isolated from the cyclamen rhizosphere. The most frequently occurring fungi belonged to the following genera: Penicillium, Cylindrocarpon, Trichoderma, Fusarium and Alternaria (Table 2). The rhizosphere fungal community of Persian cyclamen displayed no ability to suppress the growth of the pathogens under investigation. This resulted in negative summary biotic effects on F. oxysporum ( 191), on C. destructans Fig. 1. Pathogenicity of the three pathogens to Cyclamen persicum roots; means followed by different letters are significantly different at p = 0.05 according to Duncan s multiple range test
8 H. Kurzawińska Table 2 Effect of fungal communities isolated from cyclamen rhizosphere on the growth of three pathogens Biotic effect on Fungal species Frequency A. alternata C. destructans F. oxysporum IBE GBE IBE GBE IBE GBE Penicillium expansum 21 2 42 2 42 2 42 Penicillium verrucosum var. 19 4 76 4 76 4 76 corymbiferum Alternaria alternata 16 0 0 2 32 2 32 Penicillium chrysogenum 16 2 32 2 32 4 64 Cylindrocarpon radicicola 14 0 0 0 0 0 0 Fusarium oxysporum 13 +2 +26 0 0 0 0 Penicillium brevi-compactum 11 4 44 4 44 4 44 Cylindrocarpon destructans 8 0 0 0 0 0 0 Phoma eupyrena 7 4 28 4 28 4 28 Fusarium avenaceum 6 +1 +6 0 0 0 0 Trichoderma viride 6 +8 +48 +8 +48 +8 +48 Verticillium albo-atrum 6 4 24 4 24 4 24 Botrytis cinerea 5 +5 +25 +2 +10 +2 +10 Cladosporium cladosporioides 5 3 15 2 10 2 10 Trichoderma harzianum 5 +8 +40 +8 +40 +8 +40 Trichoderma polysporum 5 +8 +40 +7 +35 +7 +35 Acremonium butyri 4 4 16 4 16 4 16 Penicillium citrinum 4 4 16 4 16 4 16 Trichoderma koningii 4 +8 +32 +7 +28 +7 +28 Rhizoctonia solani 3 +2 +6 0 0 0 0 Total 178 Summary biotic effect 70 159 191 IBE individual biotic effect, GBE general biotic effect. ( 159)and A. alternata ( 70; Table 2). The degree of pathogens growth promotion varied. The suppressive effect on pathogens growth depended mainly on the share of fungi of antagonistic abilities. The strongest antagonists to the pathogens under consideration included Trichoderma spp. (Table 2). Discussion Among fungal colonies isolated from diseased bulbs and roots of Persian cyclamen, the genus Fusarium, including F. oxysporum and F. avenaceum predominated. According to Orlikowski (2000), vascular fusariosis of cyclamen caused by F.
Fungi inhabiting the rhizosphere of Persian cyclamen... 9 oxysporum f. sp. cyclaminis is one of the most serious threats to cultivation of the plant. The fungus causes also bulb rot. Fusarium spp. may survive in the substrate for dozens of years without the presence of host plant. They may be easily spread on seedlings or seeds with no visible disease symptoms (Orlikowski and Skrzypczak 1998). Fusarium oxysporum is destructive to vascular bundles. Fungi of the genus Cylindrocarpon were represented by C. destructans and C. radicicola. According to Gabarkiewicz et al. (1995), C. destructans (along with R. solani) causes root rot in cyclamen. The examined community of cyclamen rhizosphere fungi had no suppressing effect on the pathogens under investigation. The values of summary biotic effects were negative. The impact of this community was considerably less supporting on A. alternata than on C. destructans and F. oxysporum. The biotic activity characteristics of its components point to the fact that too small and insufficient amount of antagonistic fungi can not prevent vascular fusariosis and root rot in cyclamen. Positive individual biotic effects on the pathogens under consideration were shown by fungi belonging to the genus Trichoderma. Antagonistic impact of these fungi on A. alternata, C. destructans and F. oxysporum was earlier confirmed by Kurzawińska and Duda (2005) and Mańka et al. (2001). Streszczenie GRZYBY ZASIEDLAJĄCE RYZOSFERĘ CYKLAMENA PERSKIEGO I ICH ODDZIAŁYWANIE NA POTENCJALNE PATOGENY TEJ ROŚLINY Określono skład gatunkowy grzybów zasiedlających chore rośliny i ryzosferę cyklamena, frekwencję gatunków, chorobotwórczość niektórych izolatów w stosunku do korzeni cyklamena oraz stosunki biotyczne między zbiorowiskiem grzybów ryzosferowych a Alternaria alternata, Cylindrocarpon destructans i Fusarium oxysporum. Chore rośliny cyklamena były zasiedlone głównie przez F. oxysporum, C. destructans i A. alternata. Gatunki te były chorobotwórcze dla korzeni cyklamena. Stwierdzono również, że zbiorowisko grzybów zasiedlających ryzosferę cyklamena wpływało sprzyjająco na wzrost F. oxysporum, C. destructans i A. alternata. Literature Booth C., 1966: The genus Cylindrocarpon. Mycol. Pap. 104. Domsch K.H., Gams W., Anderson T.H., 1980: Compendium of soil fungi. Academic Press, London. Gabarkiewicz R., Łabanowski G., Orlikowski L., Saniewska A., Skrzypczak Cz., Soika G., 1995: Ochrona ozdobnych roślin cebulowych. Plantpress, Kraków. Kurzawińska H., 2006: An interaction of potato crop soil fungi population on fungi responsible for tuber superficial diseases. J. Plant Prot. Res. 46, 4: 339 346. Kurzawińska H., Duda J., 2005: Wpływ zbiorowisk grzybów ryzosferowych dwuletnich sadzonek stewarcji pseudokameliowatej (Stewartia pseudokamellia Max.)na wzrost Fusarium oxysporum Schlecht. i Rhizoctonia solani Kühn. Progr. Plant Prot. / Post. Ochr. Rośl. 45, 2: 834 837.
10 H. Kurzawińska Mańka K., 1974: Zbiorowiska grzybów jako kryterium oceny wpływu środowiska na choroby roślin. Zesz. Probl. Post. Nauk Roln. 160: 9 23. Mańka K., 1990: Saprofityczna mikoflora środowiska glebowego a zdrowotność roślin. Phytopathol. Pol. 11 (Materiały z sympozjum odbytego w dniach 12 14 września w Szczecinie na temat Niepatogeniczna mikoflora w patologii roślin ): 122 134. Mańka K., Mańka M., 1993: O metodzie izolacji z ryzosfery drzew leśnych. In: Materiały z IV Konferencji Sekcji Biologicznych Metod Ochrony Roślin przed Chorobami PTFit, Skierniewice 22 23 kwietnia 1993. Polskie Towarzystwo Fitopatologiczne, Instytut Sadownictwa i Kwiaciarstwa, Skierniewice: 3 7. Mańka K., Mańka M., Stępniewska S., Kacprzak M., 2001: Damping-off of Scots pine (Pinus sylvestris) seedlings in Wronczyn forest nursery versus soil fungi community. Phytopathol. Pol. 22: 163 170. Nelson P.E., Tousson T.A., Marasas W.F.O., 1983: Fusarium species. An illustrated manual for identification. The Pennsylvania State University Press, University Park. Orlikowski L., 2000: Fuzarioza cyklamena szkodliwość i możliwości ograniczenia występowania. Ochr. Rośl. 1: 40. Orlikowski L.B., Skrzypczak Cz., 1998: Rola niektórych mikroorganizmów odglebowych jako czynników ograniczających rozwój fuzariozy naczyniowej. Zesz. Probl. Post. Nauk Roln. 461: 331 338. Ramirez C., 1982: Manual and atlas of the Penicillia. Elsevier, Amsterdam. Startek L., Mynett K., 2003: Rośliny ozdobne. Hortpress, Warszawa. Author s address: Prof. Dr. hab. Halina Kurzawińska, University of Agriculture in Krakow, Al. 29 Listopada 54, 31-425 Kraków, Poland, e-mail: h.kurzaw@ogr.ur.krakow.pl Accepted for publication: 25.07.2010