Cent. Eur. J. Biol. • 8(2) • 2013 • 195-200 DOI: 10.2478/s11535-013-0122-4
Central European Journal of Biology
Coniochaeta prunicola - first record for Slovakia and Europe
Research Article
Helena Ivanová1,*, Slávka Bernadovičová2
1Institute of Forest Ecology SAS Zvolen, Branch for Woody Plants Biology, Sk-949 01 Nitra, Slovakia
2Ministry of Agriculture and Rural Development of the Slovak Republic, Sk-812 66 Bratislava, Slovakia
Received 05 March 2012; Accepted 21 May 2012
Abstract: This study reports the first record of Coniochaeta on Laurocerasus officinalis Roem. from the Nitra district. This is the first record of Coniochaeta for Slovakia and also for Europe. The fungus Coniochaeta prunicola Damm & Crous (Coniochaetales, Sordariomycetes, Ascomycota) was isolated from damaged leaves and twigs of host trees. Morphological analyses demonstrate that Coniochaeta prunicola and Coniochaeta velutina are distinct species.
Keywords: Ascomycota • Laurocerasus officinalis • Morphological characteristic ©VersitaSp.zo.o.
with or without setae, and dark brown, discoid, nearly 1. Introduction globose or ellipsoidal ascospores [5,6]. The genus Coniochaeta (anamorph: Lecythophora) The Coniochaeta (Sacc.) Cooke genus is recognised including ascomycetous fungi are known pathogens of as a large and highly diversified ascomycetous genus woody plants, but some species can also cause human with non-stromatic, globose or subglobose short-necked infections. Coniochaeta contains more than 80 species perithecia with broad ostiole [6]. Species of the genus occurring mostly on wood and bark, leaves and leaf litter Coniochaeta and their Lecythophora anamorphs occur of different trees, in dung of various animals, and in soil on various substrates and media: in plants (wood, bark, and water. leaves leaf litter), animal faeces, soil and in a trongly Different genera of the family Coniochaetaceae acidic water with high heavy metal concentrations [7-10]. were identified based on the differences in their Some Coniochaeta species have significant biochemical ascomata (ostiolate in Coniochaeta, and non-ostiolate in properties. Species of Coniochaeta were isolated from Coniochaetidium and Ephemeroascus) [1,2]. Ornaments various body parts of the representative genus Prunus. in ascospore walls are another useful criterion, and the C. ligniaria (Grev.) Massee was isolated from decaying presence of pitted ascospores indicates Poroconiochaeta bark of Prunus avium L. in the Netherlands (CBS [3]. Phialidic, verticillate conidiogenous cells enable 178.75). [11] reported several species on fruit trees in differentiation Ephemeroascus from Coniochaetidium Moldavia: on dry twigs of apricot and cherry C. ambigua [2]. The most distinctive morphological features are (Sacc.) Cooke, on twigs of cherry and plum was C. calva germ-slits in ascospores, differentiating these fungi from Tode, on dry twigs and wood of plum trees C. ligniaria Sordariaceae and its phialidic anamorphs belonging to (Grev.) Massee. C. velutina (Fuckel) Munk was isolated Lecythophora [4]. Coniochaetaceae are characterized from Prunus sp., C. africana Damm & Crous, sp. nov. by dark brown to black ascocarps, ostiolate peridia from wood of Prunus salicina Lindl., C. prunicola Damm
* E-mail: [email protected] 195 Coniochaeta prunicola - first record for Slovakia and Europe
& Crous. from wood of Prunus armeniaca L. (syn. and ascospores). The identification was performed Armeniaca vulgaris Lam.), Prunus salicina Lind. and according to morphological keys [10,14-17] and other Prunus sp. [12]. reference guides [4-6,12]. During an investigation on mycoflora of cherry laurel trees growing in urbanized area, besides the fungi of the classes Hyphomycetes and Coelomycetes isolated 3. Results and Discussion from affected cherry laurels [13], the ascomycetous fungus Coniochaeta prunicola (Coniochaetaceae, The Lecythophora-like fungi isolated from Prunus Coniochaetales) that affects leaves and twigs of the host wood were classified in 13 species representing three trees was noticed. This is the first record of this fungus phylogenetically distinct genera: Collophora gen. as a pathogen of L. officinalis and also the first record of nov., Coniochaeta (anamorph: Lecythophora) and its occurrence in Slovakia and also in Europe. Although Phaeomoniella. Two species of Coniochaeta proved the incidence of disease is sporadic, the infected trees distinct from the known species, and are newly described. showed relatively severe damage. One is Coniochaeta africana Damm & Crous, sp. nov., The recently-noticed new disease of cherry laurel named after the continent of its origin (Africa) and the trees becomes an especially relevant issue. The aim of other is Coniochaeta prunicola Damm & Crous, named our study was to isolate Coniochaeta species as a causal after the host from which it was isolated, Prunus species. factor involved in health state decline of Laurocerasus The isolates from Prunus sp. which were identified officinalis and to present morphological description with as Coniochaeta included, besides C. prunicola and distinctive features. C. africana, also C. velutina (Fuckel) Munk. [12] (Table 1). The fungus isolated from cherry laurel branches with symptoms of necrosis and leaf blight displayed 2. Experimental Procedures morphological characteristics and differences pointing at Coniochaeta prunicola. The issue was studied on samples of leaves and twigs of Coniochaeta prunicola Damm & Crous (Figure 1). Laurocerasus officinalis showing blight symptoms. The Taxonomy: Anatomical and morphological samples were gathered from plants growing in private description. The ascomata were perithecial, gardens and in public greenery of the town Nitra, during subglobose to pyriform, with central ostiole, solitary, spring – autumn 2009 and 2010 and in spring 2011. The 119-159×162-221 µm in size (Figure 1a,b). The collected material was deposed at the Institute of Forest ascomata necks were 38-42 (60) µm long (Figure 1b). Ecology of the Slovak Academy of Sciences, Branch for The peridium walls were thick, multiple-layered. The Woody Plant Biology in Nitra. outer layers were composed of brown, thick-walled For isolation and obtaining pure cultures we used angular cells (Figure 1c) with relatively scarce setae. The classical phytopathological approaches. Leaf and twig setae were 3-4.5 µm and 35-51 µm in size (sometimes parts separated from the diseased plants were surface- shorter), brown or hyaline, smooth walled, straight or sterilized by immersion in a sodium hypochlorite bent, with globose or subglobose apices (Figure 1d,e). solution (1% available chlorine) for 20 minutes, rinsed The literature gives evidence that although the asci and twice or three times with sterile distilled water, and dried ascospores are indicative, the setae remain the definitive carefully with filter paper. Then the plant samples were characteristic for classification of the most Coniochaeta cut into 3–5 mm fragments, placed in Petri dishes with species. The literature gives evidence, that although the a 3% potato-dextrose agar (PDA), cultivated at 24±1°C asci and ascospores are indicative, the setae, in most and 45% air humidity in dark conditions in a versatile cases with brown to black rigid hairs, straight or bent, environmental test chamber MLR-351H (Sanyo) and unbranched with a sharp apex scattered over perithecial isolated on 3% PDA medium. Pure fungal cultures walls or concentrated in their upper parts remain the were obtained by multiple purifications. The obtained definitive characteristic for classification of the most isolates were transferred on 3% PDA medium to induce Coniochaeta species [5]. Some of species have been sporulation. The fungal structures were examined with referred as lacking setae [17]. The paper [12] informs a clinical microscope BX41 (Olympus) under a 400× that fungus C. prunicola formed perithecial, solitary, and 1000× magnification. superficial on pine needles, immersed or superficial on The isolated fungi were identified by microscopic PDA, subglobose to pyriform ascomata, with a central analyses based on the morphological characteristics ostiole, 200-250 µm in diameter, neck 50-60 µm long; of the fruiting bodies (perithecia), spore bearing peridium pseudoparenchymatous, 20-25 µm (5-8 layers), organs (asci), and reproduction organs (conidia outer wall consists of dark brown textura angularis,
196 H. Ivanová, S.Bernadovičová
Authors Examined material [12]
Causal agent C. prunicola C. prunicola
Armeniaca vulgaris Host/ Laurocerasus officinalis Prunus salicina Plant part twigs, leaves wood perithecial, solitary, subglobose to pyriform perithecial, solitary, subglobose to pyriform with a central ostiole, Ascomata 162-221×119-159 µm, neck 38-42 µm 200-250 µm diam., setose, neck 50-60 µm
hyaline or brown setae, smooth walled brown or hyaline, straight, cylindrical, tapering to a round tip, smooth- Setae 3-4.5×35-51 µm walled or granulate, 2.5-3.5 µm wide, 80 µm long
Paraphyses hyaline, septate, 3-4×74-78 µm hyaline, septate, 2-3×60-100 µm
cylindrical, unitunicate with obtuse end unitunicate, cylindrical, apedicillate Asci 8 ascospores/ascus, 68-81×8-10 µm 8 ascospores/ascus 63-73×8-10 µm
uniseriate, 1-celled, brown, smooth-walled with uniseriate, 1-celled, brown, smooth-walled, broadly ellipsoidal in top Ascospores granular content 9(10-)13×(5-)6-7(-8) µm view and reniform from the side, granular content, germ slit, longitudinal germ slit 7×6 µm (7.5-)8.5-10(-11)×(5-)6-7.5(-8)×(3-)4-5 µm
Guttules absent absent
Hyphae hyaline, 2-3 µm wide hyaline, 1-4 µm wide
hyaline, 1-celled, smooth-walled, cylindrical to hyaline, 1-celled, smooth-walled, mainly allantoid, cylindrical to ovoid, Conidia ovoid, sometimes allantoid, (2-)3-4(-7) × 1-2 µm (2.5-)3.5-6(-8)×1-2(-3) µm
pale buff to white, flat with sparse aerial flat with sparse aerial mycelium, pale saffron, pale buff to white on Colonies mycelium on PDA PDA, 28 mm in diam. in 2 weeks
Chlamydospores lacking lacking
Table 1. Comparison of biometric characteristics and morphological features of different species in genus Coniochaeta on Prunus sp. reported by [12] and examined material from Slovakia. setose. Setae brown (or hyaline), straight, cylindrical, ornamented or broadly umbonate ascospores, or by tapering to a round tip, smooth-walled or granulate, 2-3.5 lacking Lecytophthora anamorphs. Ascospore sizes by µm wide, up to 80 µm long. most of the Coniochaeta species varied: Coniochaeta Asci were 4-5 µm long, fasciculate, unitunicate, leucoplaca (Berk. & Ravenel) isolated from twigs of cylindric, with a truncate apex and small apical rings, Pistacia vera L. [10,20] have ascospore size 7-10×5- growing from the bottom of the perithecium between 9×4-8 µm, but Coniolariella ershadii (Zare, Asgari & W. less numerous paraphyses which were hyaline, septate, Gams) Zare, Asgari & W. Gams (basionym Coniochaeta 74-78×3-4 µm in size (Figure 1g). There were formed ershadii Zare, Asgari & W. Gams) 16×18×9.5-10 µm. unitunicate cylindrical asci, 68-81×8-10 µm with eight Coniolariella gamsii (Asgari & Zare) Dania García, ascospores/ascus (Figure 1f-h). Ascospores were Stchigel & Guarro (basionym Coniochaeta gamsii brown, one-celled, ellipsoidal (Figure 1i), smooth- Asgari & Zare) isolated from leaves of Hordeum vulgare walled, without ornamented walls and without guttules, L. [20,21] has size 16-19×6-11 µm and Coniochaeta 9(10-)13×(5-)6-7(-8) µm in size, with granular contents ligniaria (Grev.) Massee has size 9-20×8-15×4-8 µm (Figure 1j). The mature broadly almond-shaped [5]. Ascospore size of Coniochaeta rhapalochaeta ascospores were ellipsoidal with a longitudinal germ slit sp. nov. (Romero & Carmarán) isolated from wood of 6-7 µm long (Figure 1k). Bulnesia retama (Gillies ex Hook. & Arn.) Griseb. has The key provided in [10] suggests identifying our 10-14×7.5-9×5-6 µm [17]. results as Coniochaeta velutina, but cultures of this Isolates were obtained from the leaf and twig tissues species turn dark and ascospores in comparison of showing rusty to brown colour blight symptoms. Colonies Coniochaeta prunicola are smaller and have guttules. on PDA low growing, first white, than becoming pale buff Review of the literature [18] and [12] shows that had not turned as dark as Coniochaeta velutina cultures this fact is in accordance with our results. The other [4,12]. Conidia in the culture media were abundant. species [19] differed from C. prunicola by having Perithecia produced abundantly on PDA maturing in
197 Coniochaeta prunicola - first record for Slovakia and Europe
Figure 1. Coniochaeta prunicola on Laurocerasus officinalis. a-k Teleomorph a ascocarps; b ascocarps in dehiscence; c peridium; d, e peridial setae; f rosettes of asci; g asci with paraphyses; h. 8-spored ascus; i ascospores; j granular content of ascospores; k ascospore germ slit; l-r Anamorph l colony on PDA after 24 days; m hyphal coil; n-q conidiogenous cells; r conidia Scale bars: a, c-e, g-r =20 µm; b, f =50 µm
198 H. Ivanová, S.Bernadovičová
4-5 weeks solitary, black, subglobose to pyriform. The and not regular allantoid conidia. The facts are in good same was observed in our study with isolates of accord with our observations. fungus C. prunicola from cherry laurel. Our colonies In the investigated samples, the fungus C. prunicola placed on PDA in dark conditions at 24°C were was relatively rare. An important fact is that C. prunicola slow-growing, with aerial, pale saffron, later tanned has been identified as a new pathogenic fungus causing mycelium (Figure 1l). Vegetative hyphae were hyaline, blight symptoms of L. officinalis in Slovakia and also 2-3 µm wide, lacking chlamydospores. Conidiophores in Europe. At the same time, the present report is the formed directly on hyphae, mostly reduced to first from Slovakia on the genus Coniochaeta. Further conidiogenous cells (Figure 1n-q). Phialides were studies are required to assess the pathogenicity and either short cylindrical or ampulliform. Collarettes were relevance of Coniochaeta infection in connection with usually inconspicuous. Conidia (Figure 1r), size of cherry laurel damage. (2-)3-4(-7)×1-2 µm formed on hyphal coils (Figure 1m) are hyaline, one-celled, cylindrical, mostly allantoid. The anamorph stage of C. velutina known from Acknowledgements various trees and shrubs of Lecythophora genus, had varying size of conidia obtained from pure This study was conducted thanks to financial support cultures: 3-6×2-4 µm [22], 2.5-3.5×1.5-2 µm [23], of the project No. 2/0149/10 of scientific grant agency 2-4×1-2.5 µm [24], and 3-8 µm long [25]. According to of the Ministry of Education of the Slovak Republic [12], the anamorph of Coniochaeta prunicola is similar and Slovak Academy of Sciences VEGA and the to the anamorph of Coniochaeta velutina, but the latter grant of Slovak Research and Development Agency has shorter colarettes, not beyond 1 µm, and wider No. APVV-0421-07.
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