Fungi and fungi-like Oomycetes isolated from affected leaves of rhododendron MARIA KOWALIK Department of Plant Protection, University of Agriculture in Kraków Al. 29. Listopada 54, PL-31-425 Kraków, mkowalik@ogr.ar.krakow.pl Kowalik M.: Fungi and fungi-like Oomycetes isolated from affected leaves of rhododendron. Acta Mycol. 43 (1): 21–27, 2008. The aim of the work is to identify fungi and fungi-like Oomycetes occurring on affected leaves of rhododendron Rhododendron L. Mycological analyses were carried out on 200 leaves collected from green areas of Kraków from May till September 2005. Isolated fungi- like Oomycetes belonged to 67 taxa. The most frequently found fungi included: Alternaria alternata, Aspergillus niger, Botrytis cinerea, Coelophoma empetri, Nigrospora sphaerica, Pestalotia sydowiana, Phialophora cyclaminis, Phomopsis archeri, Septoria azalea and Sordaria fimicola. Among fungi-like organisms Phytophthora cinnamomi and P. citricola were isolated. Key words: rhododendron, leaves, isolation, fungi-like Oomycetes INTRODUCTION Rhododendrons growing in urban green areas are susceptible to various patho- genes. Infected plants develop discolouration, brown spots and necroses, affecting their aesthetic value. The symptoms of infection, observable from spring to autumn, increase when the plants are in bloom resulting in dieback and leaf drop. The dam- age is caused by fungi-like Oomycetes of genera Pythium and Phytophthora and fungi of genera: Botrytis, Cercospora, Colletotrichum, Cylindrocladium, Exobasidium, Mi- crosphaera, Ovulinia, Pestalotia, Phomopsis, Phyllosticta, Pycnostysanus, Ramularia, Rhizoctonia and Septoria (Farr et al. ����� �er�er� ��a��a ����� �erre� ����� �r������ �er�er� ��a��a ����� �erre� ����� �r- liko��ki ����� Łaba�o��ki et al. 2001; Kita, Mazurek 2003; Kowalik et al. 2006; Kowalik, Muras 2007). The aim of this work was to identify the organisms occurring on affected rhodo- dendron leaves in green areas of Kraków. ACTA MYCOLOGICA Vol. 43 (1): 21–27 2008 22 M. Kowalik MATERIAL AND METHODS Leaves of rhododendron with the symptoms of discoloration, brown spots or ne-rhododendron with the symptoms of discoloration, brown spots or ne- croses were collected from the plants in the Botanical Garden of the Jagiellonian University, the Zoological Garden and town lawns in Kraków. 200 leaves from 50 plants were collected for mycological analysis. The numbers of leaves taken from the top part (T), central part (C) and lower part (L) of the plant were approximately equal. The leaf samples were disinfected in 70% ethyl alcohol and 5 parts of each leaf from the area between ill and healthy tissue were put on Petri dishes with 2% PDA medium. The colonies of fungi and fungi-like Oomycetes �ere ide�tified u�i�g variou� media for ide�tificatio�: Guba (����)� Dom�ch et al. (����)� Sutto� (����)� Ho (1981), Ellis and Ellis (1987). RESULTS From the collected material 721 colonies of fungi, 19 colonies of fungi-like Oomycetes and 150 colonies of bacteria (not being the subject of this paper) were isolated. The isolated fungi and Oomycetes belonged to 67 taxa within 37 genera. Among the dominat fungi the following constitued more than 66 per cent of the total: Alternaria alternata, Aspergillus niger, Botrytis cinerea, Cladosporium macrocar- pum, Coelophoma empetri, Humicola grisea v. grisea, Nigrospora sphaerica, Pestalotia sydowiana, Phialophora cyclaminis, Ph. richardsiae, Phoma fimeti, Ph. macrostoma, Phomopsis archeri, Pycnostysanus azaleae, Septoria azalea and Sordaria fimicola. Less frequent were: Acremonium furcatum, Cladosporium sphaerospermum, Colletotri- chum gloeosporioides, Cylindrocladium scoparium, Epicoccum purpurascens, Exoba- sidium vaccinii, Humicola fuscoatra v. fuscoatra, Phoma chrysanthemicola and Ph. putaminum. Fungi belonging to 20 taxa occurred once or twice. Fungi-like Oomycetes were represented by Phytophthora cinnamomi and Ph. citricola (Tab. 1). Among the species isolated from the leaves situated in the central part of the plants, the following were predominant: A. alternata, B. cinerea, C. empetri, C. gloeosporioides, E. purpurascens, N. sphaerica, P. sydowiana, S. fimicola and fungi belonging to genera: Aspergillus, Chaetomium, Cladosporium, Humicola and Penicillium. From leaves situated in the lower parts of the plant, the following were isolated: Cylindrocarpon destructans, C. scoparium, Ph. archeri, Rhizoctonia solani, S. fimicola, Trichothecium roseum and fungi of genera: Acremonium, Fusarium, Mortierella and Phialophora. From the lower parts also Ph. cinnamomi and Ph. citricola were iso- lated. From the leaves situated at the top part of the plant, numerous colonies of A. alternata, C. empetri, S. azalea, S. fimicola were isolated. Less frequent were: E. vaccinii, Mucor hiemalis, P. sydowiana, Pycnostysanus azaleae and fungi of genera Trichoderma and Phoma. The frequency of species occurrence of fungi and fungi-like Oomycetes on leaf- blades was differentiated. The following species occurred at the highest frequen- cy: A. alternata, S. fimicola, P. sydowiana, S. azalea and C. empetri. They were iso- lated (respectively) from 27, 11, 9, 8 and 7 leaves. Less frequently (on 5-6 leaves) occurred: Aspergillus niger, B. cinerea, Cladosporium macrocarpum, C. gloeosporioides, Fungi and fungi-like 23 Ta b l e 1 Fungi and fungi-like Oomycetes isolated from affected leaves of rhododendron Species Fre- quency of occur- rence Percent- age of occur- rence The number of leaves the fun- gus spe- cies was isolated from Localiza- tion of leaves* Acremonium butyri (van Beyma) W. Gams 6 0.81 2 L Acremonium furcatum F. et V. Moreau ex W. Gams 8 1.08 2 L Acremonium fusidioides (Nicot) W. Gams 1 0.14 1 C Acremonium kiliense Grütz 4 0.54 1 L Alternaria alternata (Fr.) Keissler 131 17.70 27 TCL Arthrinium pheosphermum (Corda) M. B. Ellis 2 0.27 1 C Aspergillus niger van Tiegh. 21 2.84 6 C Aspergillus wentii Wehmer 4 0.54 1 C Botrytis cinerea Pers. ex Nocca et Balb. 21 2.84 6 C Chaetomium globosum Kunze ex Steud. 3 0.41 1 C Chaetomium indicum Corda 2 0.27 1 C Chrysosporium asperatum Carm. 6 0.81 2 C Chrysosporium merdarium (Link ex Grev.) Carm. 1 0.14 1 L Cladosporium cladosporioides (Fres.) de Vries 6 0.81 2 C Cladosporium macrocarpum Preuss 19 2.57 6 C Cladosporium sphaerospermum Penz. 10 1.35 2 C Coelophoma empetri (Rostr.) Petrak 25 3.38 7 TC Colletotrichum gloeosporioides (Penz.) Sacc. 14 1.89 5 C Coniothyrium minitans W. A. Campbell 6 0.81 2 C Cylindrocarpon destructans (Zins.) Scholten 6 0.81 2 L Cylindrocarpon tenue Bugn. 2 0.27 1 C Cylindrocladium scoparium Morg. 8 1.08 2 L Doratomyces stemonitis (Pers. ex Steud.) Morton 2 0.27 1 T Epicoccum purpurascens Ehrenb. ex Schlecht. 12 1.62 4 C Exobasidium vaccinii (Fuckel) Woron. 12 1.62 3 T Fusarium culmorum (W. G. Sm.) Sacc. 5 0.68 1 L Fusarium graminearum Schwabe 2 0.27 1 L Fusarium stilboides Wollenw. 2 0.27 1 L Gloeosporium rhododendri Briosi et Cav. 2 0.27 1 C Humicola fuscoatra Traaen var. fuscoatra 8 1.08 4 C Humicola grisea Traaen var. grisea 17 2.30 5 C Mammaria echinobotryoides Ces. 7 0.95 2 C Morenoina rhododendri J. P. Ellis 2 0.27 1 C Mortierella alpina Peyronel 2 0.27 1 L Mortierella horticola Linnem. 1 0.14 1 L Mortierella isabellina Oudem. 1 0.14 1 L Mortierella parvispora Linnem. 4 0.54 1 L Mucor hiemalis Wehmer f. hiemalis 4 0.54 1 T Myrothecium cinctum (Corda) Sacc. 1 0.14 1 C Nectria inventa Pethybr. 2 0.27 1 C Nigrospora sphaerica (Sacc.) Mason 21 2.84 5 C Penicillium expansum Link ex Gray 7 0.95 2 C Pestalotia sydowiana Bres. 40 5.41 9 TC Phialophora cyclaminis van Beyma 23 3.11 5 L Phialophora richardsiae (Nannf.) Conant 16 2.16 4 L Phoma chrysanthemicola Hollòs 11 1.49 3 T Phoma epicoccina Punithalingam, Tulloch et Leach 7 0.95 2 T Phoma eupyrena Sacc. 6 0.81 2 T 24 M. Kowalik Humicola grisea v. grisea, N. sphaerica, Ph. cyclaminis, Ph. fimeti, Ph. macrostoma, Ph. archeri and P. azaleae. On 3-4 leaves occurred E. purpurascens, E. vaccinii, H. fuscoatra v. fuscoatra, Ph. richardsiae, Ph. chrysanthemicola and Ph. cinnamomi. The remaining were isolated form 1-2 leaves (Tab. 1). Rhododendron symptoms caused by fungi and fungi-like Oomycetes were var- ied. From chlorosis affected leaves with discoloration changes alongside the main veins, C. destructans, C. tenue, C. scoparium, and also three species of Fusarium were isolated. Fungus R. solani, causing the rhizoctoniosis in rhododendron, was isolated from the leaves with irregular brown-grey spots, surrounded by a large yellow area. B. cinerea, the cause of Botrytis petal blight, was isolated from rapidly dying leaves, with the symptoms of necrosis, such as large brown spots covered with dense grey mould. Fungus E. vaccinii, was isolated from deformed leaves with small reddish and brownish galls, typical of azalea leaf a�d flo�er gall. The cause of large oval necrotic �pot� �ith dark li�i�g� a�d vi�ible pic�idia �a� ide�tified a� Ph. archeri. Other pathogens occurring in phyllosphere, such as C. gloeosporioides (causing rhododendron anthracnose), Gloeosporium rhododendri, Morenoina rhododendri, C. empetri and S. azalea (the reason for leaf scorch) caused round, oval or irregular necrotic spots of different shapes. A sign of C. gloeosporioides infection was grey spots, with visible rings and picnidia. In case of S. azalea, small oval, grey-brown spots with darker lining were seen. Necrotic spots caused by Ph. cyclaminis and P. richardsiae were small irregular with uneven edges. P. sydowiana was isolated from large light brown necrotic spots appearing on the edges and the tops of the leaves. Leaf petioles and blades, with brown discolouration and necroses forming a cline �hape� �ere the reflectio� of Ph. cinnamomi and Ph. citricola presence. These patho- gens were also isolated from fast expanding brown oval spots along the leaf margin. Phoma fimeti Brun. 18 2.43 6 T Phoma herbarum Westd. 3 0.41 1 T Phoma leveillei Boerema et Bollen 1 0.14 1 T Phoma macrostoma Mont. 17 2.30 6 T Phoma medicaginis Malbr. et Roum. 5 0.68 1 T Phoma pomorum Thüm. 5 0.68 1 T Phoma putaminum Speg. 9 1.22 2 T Phomopsis archeri Sutton 23 3.11 6 L Phytophthora cinnamomi Rands 12 1.62 3 L Phytophthora citricola Sawada 7 0.95 2 L Pseudeurotium zonatum van Beyma 2 0.27 1 C Pycnostysanus azaleae (Peck) E. Mason 16 2.16 5 T Rhizoctonia solani Kühn 2 0.27 1 L Septoria azalea Voglino 33 4.46 8 T Sordaria fimicola (Rob.) Ces. et de Not. 53 7.16 11 TCL Trichoderma harzianum Rifai 2 0.27 1 T Trichoderma pseudokoningii Rifai 2 0.27 1 T Trichoderma viride Pers. ex Gray 4 0.54 1 T Trichothecium roseum (Pers.) Link ex Gray 3 0.41 1 L Total 740 100.00 - - Abbreviations: * – localization of leaves: T – top part of the plant; C – central part of the plant; L – lower part of the plant. Tab. 1 cont. Fungi and fungi-like 25 DISCUSSION Different discolorations, brown spots and necroses, visible on rhododendron leave� i� gree� area� of �rakó�� �ere cau�ed by ma�y orga�i�m� cla��ified a� Fungi and �omycete�. Accordi�g to Farr et al. (����)� �er�er a�d ��a��a (����)� �erre� et al. (����)� Łaba�o��ki et al. (����) rhodode�dro� leave� ca� be i�fected by fungi and fungi-like organisms of different genera including: Botrytis, Cercospora, Colletotrichum, Cylindrocladium, Exobasidium, Gloeosporium, Macrophoma, Microsphaera, Monochaetia, Pestalotia, Pestalotiopsis, Phomopsis, Phyllosticta, Phytophthora, Septoria, Rhizoctonia and others. While Kita and Mazurek (2003) isolated fungi of genera: Alternaria, Botrytis, Cladosporium, Epicoccum, Fusarium, Humicola, Mucor, Penicillium, Pestalotia, Phoma, Rhizopus, Sclerotinia and Trichode- rma from the phyllosphere of rhododendron and azalea growing in the botanical garde� a�d arboretum. Thi� re�earch �ork co�firm� that most of fungi and fungi- like Oomycetes isolated from the affected leaves of rhododendron belong to the mentioned above taxa. The determination of organisms inhabiting affected rhododendron leaves enabled to show the relation between pathogens and pathologic symptoms. The isolation of fungi such as: C. destructans, C. tenue, C. scoparium in the leaves with the discoloura- tio� alo�g�ide the mai� vei� i�dicate� that the root �y�tem of the pla�t� �a� fir�ty at- tacked� cau�i�g root rot� a� me�tio�ed by �er�er (���5) a�d Łabanowski et al. (2001). Also in the work by Kowalik and Muras (2007) fungi C. destructans, C. scoparium and numerous species of Fusarium were numerously isolated from dropped rhodo- dendron leaves. Fungus R. solani was the reason for rhododendron rhizoctoniosis, B. cinerea caused grey mould, C. gloeosporioides – anthracnose, and S. azalea – rho- dode�dro� leaf �corch (Łabanowski et al. 2001), while fungus E. vaccinii spending �i�ter i� flo�er a�d leaf bud� of rhodode�dro��� i� �pri�g cau�ed the di�ea�e called azalea leaf a�d flo�er gall (Orlikowski 1999). The occurrence in phyllosphere of pathogens such as C. gloeosporioides, G. rhododendri, M. rhododendri, C. empetri and S. azalea (numerously isolated in this reasearch work) and leaf symptoms caused by them were documented in the papers by Werner and ��a��a (����)� Łaba�o��ki et al. (2001), Kita and Mazurek (2003). In this work P. sydowiana was frequently isolated. Łaba�o��ki et al. (2001) do not attribute necroses to this species. The role of P. sydowiana in causing necroses of heathers in nurseries and in permanent sites was described by Kowalik and Sa-heathers in nurseries and in permanent sites was described by Kowalik and Sa- gan (2005) as well as Kowalik and Wandzel (2005), while Kita and Mazurek (2003), in phyllosphere of azaleas and rhododendrons found quite numerously occurring: Pestalotia fibricola, P. rhododendrii and P. truncata. In the works by Kowalik and Muras (2007) fungus P. sydowiana together with A. alternata, occurred causing spots and necroses of rhododendron leaves and intensifying the process of leaf drop were isolated at large frequency. The description of phytophtorosis symptoms on leaves and petioles of rhodo- de�dro�� a�d their effect� i� rhodode�dro� pla�ti�g� are co�firmed i� the �ork� by Łaba�o��ki et al. (����)� �erre� (����)� �er�er (���5)� �o�alik et al. (����). The occurrence of Ph. citricola in rhododendron nurseries and susceptibility of many rhododendron taxa to this pathogen is described by Orlikowski and Szkuta (2003), 26 M. Kowalik while Orlikowski and Szkuta (2002) highlight the role Ph. cinnamomi in causing phy- tophtorosis on container-grown ericaceous plants in ornamental nurseries. While analysing the plant material, it was found out that, in addition to the above mentioned causes of discolorations, brown spots and necroses, the secondar- ily i�vadi�g fu�gi o� pathological leave� �ere ide�tified. They �ere the follo�i�g: A. alternata, S. fimicola, N. sphaerica, A. niger and the species of genera: Acremo- nium, Cladosporium, Humicola, Mortierella, Mucor, Penicillium, Phoma and Tri- choderma. Accordi�g to �er�er a�d ��a��a (����) the�e fu�gi ca� i�te��ify patho- genic processes. Inhabiting small necrotic areas in a short time they rapidly inten-in a short time they rapidly inten- sify the process of necrosis. A large participation of fungi from genera: Alternaria, Cladosporium, Epicoccum and Phoma in the phyllosphere of rhododendrons is men- tioned by Kita and Mazurek (2003). Their work indicates that these saprotrophs dominate in the structure of the populations of isolated fungi. The obtained results co�firm co��iderable participation of A. alternata, E. purpurascens, C. macrocarpum, C. sphaerospermum and many species of Phoma in the populations of fungi isolated from necrotic tissues of rhododendrons. It can be concluded that a large biodiversity of fungi and fungi-like Oomycetes isolated from the affected rhododendron leaves in the green areas of Kraków caused different pathologic symptoms and decreased decorative value of the plants. CONCLUSIONS The most numerous fungi occurring in the rhododendron plantings in the green 1. areas of Kraków were: Alternaria alternata, Aspergillus niger, Botrytis cinerea, Coelophoma empetri, Nigrospora sphaerica, Pestalotia sydowiana, Phialophora cyclaminis, Phomopsis archeri, Septoria azalea and Sordaria fimicola Among the fungi-like Oomycetes2. Phytophthora cinnamomi and Ph. citricola were isolated from the affected rhododendron leaves. The causes of discolorations, brown spots and necroses of the rhododen-, brown spots and necroses of the rhododen-spots and necroses of the rhododen- and necroses of the rhododen-and necroses of the rhododen-3. dron leaves were the pathogens belonging to genera: Botrytis, Colletotrichum, Cylindrocarpon, Cylindrocladium, Exobasidium, Fusarium, Pestalotia, Phomopsis, Phytophthora, Rhizoctonia and Septoria. Commonly occurring saprotrophs of genera: 4. Acremonium, Alternaria, Cladosporium, Coelophoma, Epicoccum, Humicola, Nigrospora, Phoma, Sordaria a�d other�� i�te��ified the proce�� of leaf �ecro�i�. On the rhododendron plants the symptoms of phytophthorosis, grey mould, 5. rhizoctoniosis, anthracnose, leaf scorch, azalea leaf a�d flo�er gall were found. The effect was leaf drop and, in consequence decreased decorative value of the plants. REFERENCES Domsch K. H., Gams W., Anderson T. H. 1980. Compendium of soil fungi. Acad. Press. London, New York, Toronto, Sydney, San Francisco. 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Rhododen- dron und immergrüne Laubgehölze. Jahrbuch der Deutschen Rhododendron Gesselschaft 1999: 99–113. Grzyby i organizmy grzybopodobne Oomycetes wyizolowane z porażo�ych li�ci róża�ecz�ika S t r e s z c z e n i e Celem pracy była ide�tyfikacja grzybó� i orga�izmó� grzybopodob�ych �omycete� �y- �tępujących �a porażo�ych li�ciach róża�ecz�ika Rhododendron L. Materiał do badań �ta- �o�iło ��� li�ci (z obja�ami przebar�ień� plami�to�ci i �ekroz) zebra�ych z 5� krze�ó� �a terenie Ogrodu Botanicznego UJ, Ogrodu Zoologicznego oraz na skwerkach miejskich Kra- ko�a. �yodręb�io�o grzyby i orga�izmy grzybopodob�e �omycete� (� liczbie 74� kolo�ii)� �ależące do �7 tak�o�ó�. Na porażo�ych li�ciach �ajlicz�iej �y�tępo�ały grzyby: Alternaria alternata, Aspergillus niger, Botrytis cinerea, Coelophoma empetri, Nigrospora sphaerica, Pestalo- tia sydowiana, Phialophora cyclaminis, Phomopsis archeri, Septoria azalea i Sordaria fimicola. Spo�ród orga�izmó� grzybopodob�ych� z porażo�ych li�ci róża�ecz�ika �yizolo�a�o Phy- tophthora cinnamomi i Ph. citricola. Spra�cami przebar�ień� plami�to�ci i �ekroz �a li�ciach róża�ecz�ika były patoge�y z rodzajó�: Botrytis, Colletotrichum, Cylindrocarpon, Cylindro- cladium, Exobasidium, Fusarium, Pestalotia, Phomopsis, Phytophthora, Rhizoctonia i Septoria. Licz�ie �y�tępujące �aprotrofy z rodzajó�: Acremonium, Alternaria, Cladosporium, Coelopho- ma, Epicoccum, Humicola, Nigrospora, Phoma, Sordaria i i��e i�te��yfiko�ały proce� �ekro- tyzacji li�ci. Na róża�ecz�ikach rozpoz�a�o obja�y fytoftorozy� �zarej ple��i� rizokto�iozy� a�trak�ozy� �eptoriozy� po�łocz�ika azalio�ego� � �y�iku których dochodziło do zamiera�ia i opada�ia li�ci� a tym �amym ob�iże�ia �aloró� dekoracyj�ych ro�li�. 2014-01-01T11:47:00+0100 Polish Botanical Society