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Fungi Associated with a Decline of Pinus Nigra in Urban Greenery Original Paper

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Fungi Associated with a Decline of Pinus Nigra in Urban Greenery Original Paper Acta fytotechn. zootechn., 18, 2015(2): 36–43 Helena Ivanová: Fungi associated with a decline of Pinus nigra in urban greenery Original Paper Fungi associated with a decline of Pinus nigra in urban greenery Helena Ivanová Institute of Forest Ecology of the Slovak Academy of Sciences, Zvolen, Slovak Republic During the years 2013–2014, complex mycoflora of Austrian pine trees (Pinus nigra Arnold) was monitored within a survey on health state of trees growing in urban environment. Nine species of microscopic fungi were isolated and identified from samples collected from different part of Pinus nigra needles during the study period. This study reports the occurrence of the fungi Asterosporium asterospermum, Beltrania rhombica, Camarosporium pini, Coleosporium sp., Cyclaneusma niveum, Lophodermium seditiosum, Nigrospora sp., Phomopsis sp., Trichothecium roseum of P. nigra. In Austrian pine the greatest damage has been caused by the fungi Lophodermium seditiosum and Cyclaneusma niveum. Damage caused by rust fungus (Coleosporium sp.) and by fungi Asterosporium asterospermum and Beltrania rhombica occurs less frequently. Keywords: Austrian pine, disease symptoms, health state, parasitic microscopic fungi 1. Introduction by the low temperatures in winter period and by the In recent years, among urban trees Austrian pine also drought from the spring to summer season (Grove, 1922; called European black pine (Pinus nigra Arnold) belongs James, 1984; Manoharachary et al., 2003; Karadžić and to the particularly affected tree species. At the same Milijašević, 2008). time with regard to the expected global climate change, The aim of this study was to research distribution, this change may cause the pathogens to adapt to the disease symptoms, some important characteristics in previously resistant species of woody plants (Kolářík et pure culture and distinctive morphological features of al., 2005). Austrian pine is a bold-textured and urban- the most important parasitic fungi in Austrian pine. The tolerant pine having a broad-pyramidal growth habit presence of these fungi in host tissue of symptomatic trees with ascending branches and showy spring candles and has been described using classical phytopathological often developing a flat-topped crown and ornamental approaches and microscopical identification based on bark with age. This evergreen tree in pine family is used morphological keys. either as a single specimen or in group or mass plantings as visual screens or windbreaks (Sinclair et al., 1987). 2. Material and methods Numerous fungal diseases affect pine trees, including From spring to autumn 2013–2014 needles of Pinus Austrian pine. Some of them are managed easily by nigra with blight symptoms were collected at several applying chemical substances or biological means, the locations from plants growing in private gardens and other spread rapidly through pine populations, as there public greenery of the town of Nitra (Nitra-Zobor, Nitra- have been no known means for controlling the fungal Chrenová, Nitra-Sihoť, Nitra-Kynek). Altogether 20 trees spread yet (Sinclair et al., 1987). Among pathogenic fungi, were studied. The age of evaluated trees was between pathogens identified from the dead pine tissues include 35–40 years. Samples were taken from some sections of also Asterosporium asterospermum (Pers.) S. Hughes, trees with damaged needles from every locality. Every Beltrania rhombica Penz., Camarosporium pini (Westend.) sample was cultivated on 30 Petri dishes (PD) with 3% Sacc., Coleosporium sp., Cyclaneusma niveum (Persoon ex PDA (potato-dextrose agar – complete name of medium, Fr.) DiCosmo, Peredo & Minter, Lophodermium seditiosum when first use) medium. The samples of material have Minter, Staley & Millar, Nigrospora sp., Phomopsis sp. and been deposed at the Institute of Forest Ecology of Trichothecium roseum (Pers.) Link. the Slovak Academy of Sciences, Branch for Woody The occurrence of causal pathogens causes withering Plants Biology in Nitra. Classical phytopathological of branches and tree thinning in the lower parts of the approaches were used to isolate and obtain pure hyphal tree crown as a beginning of woody plants weakening cultures. Fungi were isolated from the needles first *Correspodence: Helena Ivanová, Institute of Forest Ecology of the Slovak Academy of Sciences Zvolen, Branch for Woody Plants Biology, Akademická 2, 949 01 Nitra, Slovak Republic, [email protected] © Slovak University of Agriculture in Nitra Faculty of Agrobiology and Food Resources, 2015 http://www.fapz.uniag.sk/ – 36 – DOI: 10.15414/afz.2015.18.02.36–43 Acta fytotechn. zootechn., 18, 2015(2): 36–43 Helena Ivanová: Fungi associated with a decline of Pinus nigra in urban greenery immersed for one minute into 70% alcohol and then flattened acervuli with a wide opening. Conidiomata for 15 minutes into sodium hypochlorite (1% available were pycnidial with a wide ostiole of more than 100 µm chlorine). After that, the needles were washed twice in diameter. Conidial septation was euseptate. Central cell sterilized distilled water and cut to fragments of 2–5 mm was indistinct. Conidia stellate, brown, composed of which were placed on the nutritive medium, potato- 4 equally developed arms, 30.5–55 (–61) µm between the dextrose agar (PDA). Petri dishes with the medium widest points, arms 17–28 (–30) × 8–9.5 (–11) µm (Tanaka and host fragments were incubated at 24±1 °C and et al., 2010). 45% humidity in dark in a versatile environmental test The hyphomycetous fungus Beltrania rhombica Penz. chamber MLR-351H (Sanyo). Pure fungal cultures were which occurs on dead leaves of many trees and shrubs obtained using multiple purifications. The identification caused foliar damage and reduced the ornamental was performed using morphological keys according value of host trees (Manoharachary et al., 2003). Fungus to Minter et al. (1978), Woodward (2001), Tanaka et al. was dominant between endophytic fungi on Pinus (2010), Luo et al. (2010) or morphological studies in thunbergii (Rambelli et al., 2008; Kim et al., 2012). On Botella et al. (2010). All the isolates were examined from PDA produced gray, later dark red-brown, floccose, time to time and the rate of colony development was effuse colonies. Conidiophores simple, or sometimes observed, as well as the fructifications in the culture, branched at the base, gently flexuous, clear or light etc. The fungi were identified based on the sporulation brown, cylindrical, smooth, septate. Conidiogenous in the culture, the appearance of hyphae, rate of growth, cells integrated, terminal, polyblastic, sympodial, etc. Visual characteristics of symptomatic needles were denticulate, sub-clavate; separating cells oval, swollen, examined with a stereomicroscope SZ51 (Olympus) 9.5 × 8 µm. Conidia solitary, biconic, appendiculate- and fungal structures observations were accomplished spicate, 0-septate, smooth, dark reddish-brown, with with a microscope BX41 (Olympus) under a 400× hyaline transverse band in the widest part of the magnification. conidium, 21–25 × 9–12 µm (Rambelli et al., 2008) or conidia unequally biconic, unicellular, dark brown with 3. Results and discussion a pale brown band, 26–31 × 8.5–12 µm (Mulas et al., Asterosporium asterospermum (Pers.) S. Hughes, syn. 1993). In our experiments from Pinus nigra needles this Stilbospora asterosperma Pers. is fungus associated with fungus formed on PDA medium red-brown mycelium. a severe needle blight of ponderosa and lodgepole pines Conidia were light brown, biconic, 20–28 × 10–14 µm, (Hunt, 1985) or scots pine (Kowalski and Kehr, 1992). In solitary with double external wall and dark brown our experiments the fungus was isolated from needles transverse septum in widest part of the cell (Figures of Austrian pine. Conidia stellate, brown composed with 1b, c). Comparable results achieved Shi et al. (2012) three arms were 15–17–25 × 10 µm, with two septa by the study of popular ornamental plant Tibouchina (Figure 1a). Number and length of arms are important semidecanta Cogn. which is damaged by the fungus distinquising criterion (Prášil and Réblová, 1995). Fruting Beltrania rhombica. The fungus formed on the leaves bodies 1.5 mm in diameter were subepidermal at first, round spot with a brown center surrounded by a reddish splitting irregularly to expose the dark spore mass later. brown border and on PDA medium gray, floccose colonies Conidiogenous cells cylindrical, hyaline. Mature conidia with light brown, cylindrical, simple or sometimes massive, dark brown, smooth in pulvinate acervuli on branched at the base conidiophores 105–202 × 3–5 µm twigs. Conidia appear as star shape that consisting of in size and unequally biconic, unicellular, dark brown 4 arms at 90° to each other. The length of conidia was with a pale brown or subhyaline band just above the 20–55 µm from tip to tip of the branches which were widest part conidia 26–31 × 8.5–12 µm in size. 10–25 µm thick at the base. Conidia are formed solitary Fungus Camarosporium pini (Westend.) Sacc., syn. at the apex of coniodiophores (Boujari et al., 2012). Hendersonia pini Westend., causes severe infection According to Kobayashi and Kubano (1986) acervuli that can result in significant growth reduction. The 800–930 µm in diam. occurred on dead beech bark are fungus parasitizes the needles of Pinus nigra weakened black, scattered, at first immersed within the epidermal by the low temperatures in winter period and by the layer, then erumpent. Conidiophores are slender, simple, drought from the spring to summer season. On the dry hyalin to pale brown, 121.5–80 µm long. Dark brown, needles little black spots which represent pycnidia of terminal, holoblastic conidia are smooth and consisting 180–300 µm in diameter arranged linearly and in parallel of 4 arms at 90° in angle to each other, connected with to venation were observed. Pycnidia were immersed conidiophore at the center cell, 30–50 µm long from in the bark, at first scattered, than arranged in line, up the tip of one arm to that of the opposite one: arms to 700 µm diameter, subglobose, brown, perforating 15–25 × 7.5 × 12.5 µm with 3–4 septa.
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