Nova Biotechnol Chim (2021) 20(1): e760 DOI: 10.36547/nbc.760 1 Nova Biotechnologica et Chimica Antagonistic effectiveness of Macromycetes against Candida albicans strains and Issatchenkia orientalis Tetiana A. Krupodorova 1 , Victor Yu. Barshteyn 1,, Olena V. Pokas 2 1 Institute of Food Biotechnology and Genomics of the National Academy of Sciences of Ukraine, Osipovskogo Str. 2a, Kyiv 04123, Ukraine 2 Institute of Epidemiology and Infectious Diseases of the National Academy of Medical Sciences of Ukraine, M. Amosova Srt. 5, Kyiv 03038, Ukraine  Corresponding author: barmash14@gmail.com Article info Article history: Received: 18 th November 2020 Accepted: 21 st April 2021 Keywords: Dual-culture plate assay Macromycetes Saccharomycetaceae Species interactions Abstract The trend to search novel natural antifungal compounds has recently been increasing. Interspecific interactions between 30 Macromycetes species and fungal pathogens (Issatchenkia orientalis, Candida albicans strains) have been evaluated using dual culture plate assay. Interaction reactions between studied fungi were different: deadlock after mycelia contact or at a distance, overgrowth without initial deadlock, partial or complete replacement after initial deadlock with contact. Domination of replacement (78.7 %) of pathogenic fungi by Macromycetes was established. Complete replacement was almost twice more frequent (30.7 %) than partial replacement (19.3 %). These results clearly indicate effectiveness of Macromycetes against pathogenic fungi via contact antagonism. Strong antagonistic activity with high antagonistic index (AI) was established by xylotrophic species Ganoderma applanatum (AI = 22.5), Lentinula edodes (AI = 21.0), Flammulina velutipes, Irpiciporus litschaueri and Pleurotus ostreatus (AI = 20.5), leaf litter decay fungus Crinipellis schevczenkoi (AI = 21), and soil saprotroph Lyophyllum shimeji (AI = 21).  University of SS. Cyril and Methodius in Trnava Introduction One of the major and global health care threat in dermatology, gynecology, and pediatrics continue to represent fungal infections. The most common fungal pathogen is Candida albicans that is the predominant cause of candidiasis. Such fungal infections can be subdivided into major groups: cutaneous (skin and its appendages), mucosal (oropharyngeal, esophageal, and vulvovaginal) and systemic (bloodstream infections, i.e., candidemia and other forms of invasive candidiasis) (Papon et al. 2013). AIDS and cancer patients as well as in transplant individuals are at high risk of candidiasis due to the immunosuppression. In addition, frequently isolated non-albicans Candida species like Issatchenkia orientalis (formerly Candida krusei) have shown gradual emergence as a cause of invasive candidiasis (Samaranayake and Samaranayake 1994; Samaranayake et al. 2014) and also onychomycosis in patients with chronic mucocutaneous infections (Hossain and Ghannoum 2001). Also of concern is the continued emergence of new facts of resistance to the existing commercial drugs. mailto:barmash14@gmail.com Nova Biotechnol Chim (2021) 20(1): e760 2 At the same time, poor effect of some drugs and high possibility of candidiasis recurrence, as well as the toxicity of drugs of modern drug therapy, based on ketoconazole, emphasize the search for new effective antifungal agents with low bayside effect. It is worth considering that interspecific fungal antagonism can be a powerful emerging tool in the development of mycoproducts (or biotech- products) against pathogenic fungi. The encouraging results of studies of the antifungal activity of Macromycetes against most pathogenic fungi have contributed to the growing interest in this problem. Numerous experiments have shown inhibition of the growth of Candida sp. under the influence of various species of Basidiomycetes: Lentinula edodes (Hirasawa et al. 1999; Hearst et al. 2009), Agrocybe perfecta, Climacodon pulcherrimus, Oudemansiella canarii, Pycnoporus sanguineus (Rosa et al. 2003), Oudemansiella spp. (Vahidi and Namjoyan 2004), Laetiporus sulphureus (Turkoglu et al. 2007; Petrović et al. 2014), Coprinus cinereus (Ndyetabura et al. 2010), Agaricus brunnescens, Lactarius vellereus (current name: Lactifluus vellereus), Terfezia boudieri (Dogǎn et al. 2013), Amanita muscaria, A. phalloides, Lactarius spp., L. densifolius, L. gymnocarpoides (current name: Lactifluus gymnocarpoides), Russula kivuensis (Chelela et al. 2014), Cantharellus cibarius (Popova 2015), Agaricus bisporus, Trametes gibbosa (Waithaka et al. 2017), Morchella esculenta, Verpa bohemica (Shameem et al. 2017), Entoloma speculum (Kodiyalmath and Krishnappa 2018), Pleurotus ostreatus (Gutef et al. 2020). These examples show clearly the significant antifungal potential of fungi and testify the relevance and prospects of such investigations. Altogether, the knowledge and understanding of different mechanisms of biological, chemical, physical action, directly caused by the species interaction of fungi, reveals alternative ways of their use as natural biotechnological tools for ecological bioremediation, biologically control the pathogens of agricultural crops or forests and as biological helpers in healing serious diseases (Petre 2015). The aim of our study was to investigate the interspecific interactions between Macromycetes species and Saccharomycetaceae fungi in dual- culture. Experimental Fungal species Thirty macrofungi from different ecophysiological (wood decaying, saprotrophic, entomophilic, and leaf litter decaying) and taxonomic groups were screened (Table 1). All the mushroom species used in this study were kindly supplied by the Mushroom Culture Collection (IBK) of the M.G. Kholodny Institute of Botany of the National Academy of Sciences of Ukraine (Bisko et al. 2016). Stock cultures were maintained on beer- wort agar-agar slants at 4 ºC. The Issatchenkia orientalis Kudryavtsev 301, Candida albicans (C.P. Robin) Berkhout 17/138 and clinical strains of C. albicans 311, 315, 319 were from the Culture Collection of Microorganisms of the Institute of Epidemiology and Infectious Diseases of the National Academy of Medical Sciences of Ukraine. Strains C. albicans 311, 315, 319 were isolated from people with different infections with undergone long-term antibiotics therapy and show resistance to antibiotic drugs (mainly to streptomycin, penicillin, oxacillin, ampicillin, some of them to amoxicillin and others). Stock cultures were maintained on potato- dextrose-agar (PDA) slants at 4 ºC. All fungi were transferred from stored cultures to PDA Petri dishes and cultured at 26 ± 1 ºC to obtain mycelial colonies. Mycelial plugs 8-mm were cut from at the mycelial active growth stage using a sterile borer and used as inoculum in dual culture. Dual-culture experiments Macromycetes were screened for their ability to suppress the mycelial growth of Saccharomyceta- ceae fungi in vitro dual-culture plate assays on PDA by the method described by Badalyan et al. (2002; 2004). The antagonistic ability of each fungal organism was determined using a rating scale for the 3 main types of reactions (A, B, C) and 4 sub-types (CA1, CB1, CA2, and CB2). Types A and B were deadlock (mutual inhibition, in which Nova Biotechnol Chim (2021) 20(1): e760 3 neither organism was able to overgrow the other) at mycelial contact (A), or at distance (B); type C replacement, overgrowth without initial deadlock. The intermediate sub-types scored were: CA1 partial and CA2 complete replacement after initial deadlock with mycelial contact; CB1 partial, and CB2 complete replacement after initial deadlock at a distance. The following score was assigned to each type or sub-type of reaction: A = 1; B = 2; C = 3; CA1 = 3.5; CB1 = 4; CA2 = 4.5; CB2 = 5. The antagonism index (AI) was then calculated for each fungal species using the following formula (Eq. 1): AI=A(nx1)+B(nx2)+C(nx3)+CA1(nx3.5)+CB1(nx4) +CA2(nx4.5)+CB2(nx5) (1) where n = frequency of each type or sub-type of reaction (Badalyan et al. 2002; 2004). Saccharomycetaceae together with the macromycetes were incubated in the thermostat at 26 ºC. The growth of fungi was monitored daily for a month. The morphological changes (coloration of mycelia and agar, visualization of the demarcation line, producing of exudate drops, formation of dense zones of mycelium, fruiting body primordia etc.) in interacted colonies were noted. The type of interaction was recorded on the 30 th day of the experiment. All experiments were independently conducted in three replicates. Results and Discussion The interaction between fungi is quite broadly, engaging and important field of study. Of particular interest is the cause-effect relationship of biological and biochemical bases for immune and defense reactions of competing species. Different concepts have been used to characterize the Macromycetes interaction with other fungi in co-culture (Bruce and Highley 1991; Badalyan et al. 2002; 2004; Barinova et al. 2008; Owaid 2017; Pasaylyuk 2017). To our opinion, the determination of antagonistic ability according to the method of Badalyan et al. (2002; 2004) using a rating scale and calculation of an antagonistic index is more appropriated for the screening and evaluation of the antifungal potential of Macromycetes in solid medium. The interactions between all tested fungi were established. The results obtained at the process of growth in dual cultures are shown in Table 1 and some examples are illustrated in Fig. 1. Performed screening allowed easy to determine the most active fungus species with high antagonistic index (AI) from different ecological groups: xylotrophic species G. applanatum (AI = 22.5), L. edodes (AI = 21.0), F. velutipes, I. litschaueri and P. ostreatus (AI = 20.5), leaf litter decay fungus C. schevczenkoi (AI = 21), and soil saprotroph L. shimeji (AI = 21). Previous experiments also showed that xylotrophic fungi P. ostreatus, F. velutipes, Ganoderma sp. had the strongest competitive effect against mycoparasitic fungi (Clonostachys rosea, Trichoderma harzianum, T. pseudokoningii, and T. viride) (Badalyan et al. 2004). Moreover, it was found that P. ostreatus possessed strongest antagonistic activity against cereal pathogenic fungi (Gaeumannomyces graminis var. tritici (current name: Gaeumannomy- ces tritici), Bipolaris sorokiniana, Fusarium culmorum, Rhizoctonia cerealis (current name: Ceratobasidium cereale) (Badalyan et al. 2002). Studied by us fungi produced a different variety of interaction reactions: deadlock after mycelial contact or at a distance, overgrowth without initial deadlock, partial or complete replacement after initial deadlock with contact (Fig. 2). However, domination of replacement (78.7 %) of pathogenic fungi by Macromycetes was established. Furthermore, complete replacement was almost twice more frequent (30.7 %) than partial replacement (19.3 %). These results clearly indicate effectiveness of Macromycetes against pathogenic fungi via contact antagonism. This tendency is in line with observation of antagonistic activity of 17 species xylotrophic Basidiomycotina against pathogenic fungi (Bipolaris sorokiniana, Fusarium culmorum, Gaeumannomyces graminis var tritici, Rhizoctonia cerealis) (Badalyan et al. 2002). Probably different mechanisms affect species interactions. Generally, any type of antagonism (at mycelial or hyphal level) or parasitism is caused by production of different compounds such as ferments, toxins, antimycotic substances that are capable of volatility as well as able to the diffusion (Woodward and Boddy 2008). It is important to Nova Biotechnol Chim (2021) 20(1): e760 2 remark that response of species interaction in certain model experimental conditions can change Table 1. The interactions of studied fungi and their antagonism index (AI). Fungi species Strain I. orientalis 301 C. albicans strains AI 17/ 138 311 315 319 Auriporia aurea (Peck) Ryvarden 5048 C C CA1 C A 13.5 Coprinus comatus (O.F. Müll.) Pers. 137 CA2 CA2 CA1* CA1 CA1* 19.5 Cordyceps militaris (L.) Fr. 1862 CA1 CA2 CA1* CA1 CA1* 18.5 Crinipellis schevczenkoi Bukhalo 31 C CA2 CA2 CA2 CA2 21.0 Cyclocybe aegerita (V. Brig.) Vizzini 1853 C C CA1 C C 15.5 Flammulina velutipes (Curtis) Singer 1878 CA2 CA2 CA2* CA1 CA1* 20.5 Fomes fomentarius (L.) Fr. 355 CA2 CA2 CA2 CA2 A* 19.0 Fomitopsis betulina (Bull.) B.K. Cui, M.L. Han &Y.C. Dai 327 CA2 C CA2 CA2 A* 1 7.5 Fomitopsis pinicola (Sw.) P. Karst. 1523 C C C* C C 15.0 Ganoderma applanatum (Pers.) Pat. 1701 CA2 CA2 CA2* CA2 CA2 22.5 Ganoderma lucidum (Curtis) P. Karst. 1900 C C C* C C 15.0 Grifola frondosa (Dicks.) Gray 976 A B CA2* A CA2 13.0 Hericium erinaceus (Bull.) Pers. 970 A A B* A CA1 8.5 Hypsizygus marmoreus (Peck) H.E. Bigelow 2006 CA1 A A* CA1 CA1* 12.5 Inonotus obliquus (Fr.) Pilát. 1877 C C CA1 C A 13.5 Irpiciporus litschaueri (Lohwag) Zmitr. 5312 CA2 CA2 CA1 CA1 CA2 20.5 Laetiporus sulphureus (Bull.) Murrill 352 C C C* C C* 15.0 Lentinula edodes (Berk.) Pegler 502 C CA2 CA2* CA2 CA2* 21.0 Lepista luscina (Fr.) Singer 64 A A A* A A* 5.0 Lyophyllum shimeji (Kawam.) Hongo 1662 CA2 CA2 C* CA2 CA2 21.0 Morchella esculenta (L.) Pers. 1953 A B A A* A 6.0 Ophiocordyceps sinensis (Berk.) G.H. Sung, J.M. Sung, Hywel-Jones & Spatafora 1928 CA1 CA1 CA2 CA1 CA1 18.5 Oxyporus obducens (Pers.) Donk 5085 C C C C C 15.0 Phellinus igniarius (L.) Quél. 1578 A CA2 CA1 CA1 A 13.5 Pleurotus djamor (Rumph. ex Fr.) Boedijn 1526 B CA2 CA1* CA2 CA2 19.0 Pleurotus eryngii (DC.) Quél. 2015 CA2 CA2 CA1* C CA1 19.0 Pleurotus ostreatus (Jacq.) P. Kumm. 551 CA2 CA2 CA1* CA2 CA1* 20.5 Schizophyllum commune Fr. 1768 C CA2 C A C* 14.5 Trametes versicolor (L.) Lloyd 353 C C C C C 15.0 Xanthoporia radiata (Sowerby) Ţura, Zmitr., Wasser, Raats & Nevo 2454 A A A* A* A* 5.0 Legend: A – deadlock after mycelial contact, B – deadlock at a distance, C – overgrowth without initial deadlock, CA1 – partial replacement after initial deadlock with contact, CA2 – complete replacement after initial deadlock with contact, * – the presence of polymorphism of C. albicans. in natural habitat due to the effecting any antagonistic or parasitism mechanisms that provide full functioning of ecosystem (Bruce and Highley 2001). Generally, C. albicans 17/138 and I. orientalis were more sensitive to all investigated fungi than C. albicans clinical isolates. Current studies may provide the basis to determine the correlation between the interaction of Macromycetes and the degree of Candida strains virulence. During the experiments the polymorphism (formation pseudohyphae or/and hyphal form) of C. albicans clinical isolates has been established (Fig. 3): C. albicans 311 in co-cultivation with 17 fungi (entomophilous C. militaris, soil saprotrophic like C. comatus, L. luscina, L. shimeji and xylotrophic F. velutipes, F. pinicola, G. applanatum, G. lucidum, G. frondosa, H. marmoreus, H. erinaceus, L. sulphureus, L. edodes, P. djamor, P. eryngii, P. ostreatus, X. radiate), C. albicans 315 ̶ with 2 fungi (soil saprotroph M. esculentaand xylotroph X. radiata), C. albicans 319 ̶ with 12 fungi (entomophilous C. militaris, soil saprotrophic C. comatus, L. luscina, and xylotrophic F. velutipes, 4 Nova Biotechnol Chim (2021) 20(1): e760 5 F. betulina, F. fomentarius, H. marmoreus, L. sulphureus, L. edodes, P. ostreatus, S. commune, X. radiate). Thus, all three clinical isolates of C. albicans Fig. 1. Illustration of competitive interactions between mycelia of studied fungi: A – deadlock at mycelial contact of M. esculenta and C. albicans 315; B – deadlock at distance, interaction of P. djamor and I. orientalis 301; C – overgrowth without initial deadlock, interaction of F. betulina and C. albicans 17/138; D – partial replacement after initial deadlock with contact of P. eryngii and C. albicans 319; E, F – complete replacement after initial deadlock with contact of I. litschaueri and C. albicans 17/138, front and reverse sides of Petri dishes, respectively. Note: right - pathogenic fungus, left - macromycete fungus. showed polymorphism in the case of co-cultivation only with X. radiata. Two clinical isolates C. albicans 311 and 319 showed this feature with growth in dual culture with C. militaris, C. comatus, L. luscina, F. velutipes, H. marmoreus, L. sulphureus, L. edodes, P. ostreatus, X. radiata. It is A B C D E F Fig. 2. Frequency of type (A – deadlock after mycelial contact, B – deadlock at a distance, C – overgrowth without initial deadlock) and subtype (CA1 – partial replacement or CA2 – complete replacement after initial deadlock with contact) interactions between colonies of studied fungi, expressed as percentage of the total number (450) of pairings tested. Nova Biotechnol Chim (2021) 20(1): e760 6 known that the basic function of hypha formation of opportunistic pathogens like Candida spp. is to invade the substrate they are adhered to. However, probably, in this case, this may be a response to a change the conditions in medium, it can become more extreme in particular, a sharp change in pH as Fig. 3. Illustration of polymorphism of C. albicans clinical isolates (right) in co-cultivation with macromycetes (left): H. erinaceus and C. albicans 311(A), X. radiata and C. albicans 315 (B), F. fomentarius and C. albicans 319 (C). a reply of the synthesis of metabolites by antagonist. The competition or antagonism experienced during co-cultivation led to a change in the morphological characteristics of fungal colonies (Fig. 4). These changes can be mediated by the up-regulation of genes involved in antagonism (Iakovlev et al. 2004). The formation of a mycelial ridge of G. frondosa upon contact with a colony of I. orientalis has been established (Fig. 4A). During deadlock reaction localized aerial mycelia of F. betulina was produced at the interface between confronted fungi (Fig. 4B). The presence of central stria was observed in case of co-cultivation G. applanatum with I. orientalis (Fig. 4C). After 20 days of growth, observed the appearance of fungal metabolites on F. fomentarius colony in the form of exudate of dark brown drops (Fig. 4D) and on P. ostreatus and L. shimeji colonies in the form of shining small golden drops of liquid (Fig. 4E, 4F). The production of analogical golden secondary metabolites was also found on edges of colonies L. edodes in dual culture with Verticillium sp. and Pythim sp. (Owaid 2017). Fungus F. fomentarius is distinguished by its ability to form primordia in culture. However, it should be noted that this process took place more intensively in the area of C. albicans location (Fig. 4D). Fruit-body primordia were also reported only in pairings of Lentinus tigrinus with Bipolaris sorokiniana (Badalyan et al. 2002). Interaction of C. comatus with C. albicans 311 resulted in the formation of brown barrages in the antithetic zone line of interaction with the pathogen, сolony morphology and color changes in the mycelia and reversume of xylotrophic fungus colony (Fig. 4G, 4H, 4J, 4K). Similar finding with brown barrages has already been reported as a resistance of L. edodes strain UFLA-LE5 to Trichoderma sp. (Santos 2019). Changes in colony color and morphology were also observed in co- cultivation of C. comatus with C. albicans 319 (Fig. 4G, 4I, 4J, 4L). It is probably that diffusion of metabolites of Candida albicans strains stimulated laccase activity (oxidation of polyphenols) of C. comatus and the producing of brown pigments in C. comatus hyphae by activating some general cellular mechanisms of stress resistance. It should be noted that the presence of laccase was previously established by us in 21 fungi species (Krupodorova et al. 2014), but only co-cultivation of C. comatus and C. albicans strains 311, 319 allowed us to reveal it. Thus, co-cultivation in our case can be considered as a biotechnological tool that allows increasing the production of C. comatus laccase. Obtained from C. comatus mycelia (strain JT-01) laccase was shown previously as antipathogenic protein that possessed antiproliferative and HIV-1 reverse transcriptase inhibitory activities (Zhao et al. 2014). The enhanced activity of lignin-modifying enzymes (particular laccase, manganese peroxidase, phenol- B C A Nova Biotechnol Chim (2021) 20(1): e760 7 Fig. 4. Illustration the changes in the morphological characteristics of fungal colonies: the formation of a mycelial ridge of G. frondosa upon contact with a colony of I. orientalis 301 (A); localized aerial mycelia of F. betulina at the interface between confronted fungi C. albicans 319 (B); the presence of central stria in case of co-cultivation G. applanatum with I. orientalis 301 (C); primordia formation of F. fomentarius in dual culture with C. albicans 17/138 (D); producing in form shining small golden drops of liquid on colonies of P. ostreatus by co-cultivation C. albicans 311 (E) and of L. shimeji by co-cultivation with C. albicans 311 (F); formation of brown barrages in the antithetic zone line of interaction with the pathogen, сolony morphology and color changes in the mycelia of C. comatus (H) and reversume of xylotrophic fungus colony (K) in dual culture with C. albicans 311; changes in front colony colour and morphology (I) and reversume (L) of C. comatus in co- cultivation with C. albicans 319. G and J - front and reversume of C. comatus colonies in mono culture, respectively (control). Note: right – pathogenic fungus, left – macromycete fungus. A B C D E F H I K L J G Nova Biotechnol Chim (2021) 20(1): e760 8 oxidase and peroxidase activities) in fungal co- culture and the important role of these enzymes in the antagonistic interaction between species pointed out by different researchers (Li 1981; 1983; Bruce and Highley 1991; Nia 1992; Freitag and Morrel 1992; Rayner et al. 1994; Savoie et al. 1998; Hatvani et al. 2002; Badalyan et al. 2002; 2004; Uzar et al. 2017). In addition, dark zones can be due to melanin production. It can function as a physical barrier, preventing access by cell wall- degrading enzymes of other organisms (Bull 1970), and also it can be an important part of the protective response against mycelial invasion that allows to adapt to environmental stress or to attack of antagonists (Lee et al. 2008). Generally, exudates producing as well as color changes should be considered a consequence of the fact, that сo-cultivation lead to a significantly enhanced production of constitutively present compounds and/or to an accumulation of cryptic compounds that are not detected in axenic cultures of the producing strain (Marmann et al. 2014) and there are could be responsible for the antifungal or/and fungistatic activity. Conclusion The study touched on the antagonistic activity is one of the integral stages into the understanding fungal-fungal competition, physiological as well as biochemical processes of their interaction. Presented research may provide valuable insights into interspecific fungal interactions. Co-cultivation led to a change in the morphological characteristics of some Macromycete fungal colonies such as the formation of a mycelial ridge, aerial mycelia, barrages, drops of exudate, changed the colony color, and polymorphism of C. albicans clinical isolates. Obtained results indicate the considerable antifungal potential of studied 30 Macromycetes against Issatchenkia orientalis and Candida albicans strains. Studied fungi produced a different variety of interaction reactions: A - deadlock after mycelial contact (18.7 %), B - deadlock at a distance (2.6 %), C - overgrowth without initial deadlock (28.7 %), and subtypes CA1 - partial replacement (19.3 %) or CA2 - complete replacement after initial deadlock with contact (30.7 %). Domination of replacement (78.7 %) of pathogenic fungi by Macromycetes was established. The complete replacement was almost twice more frequent (30.7 %) than partial replacement (19.3 %). These results clearly indicate the effectiveness of Macromycetes against pathogenic fungi via contact antagonism. Performed screening allows to determining the most active mushrooms with high antagonistic index (AI): xylotrophic species G. applanatum (AI = 22.5), L. edodes (AI = 21.0), F. velutipes, I. litschaueri and P. ostreatus (AI = 20.5), leaf litter decay fungus C. schevczenkoi (AI = 21), and soil saprotroph L. shimeji (AI = 21). These fungi are promising species for further studies to investigate and isolate potential antifungal metabolites with strong action. Acknowledgement We thank to Prof. Nina A. Bisko (M.G. Kholodny Institute of Botany of the National Academy of Sciences of Ukraine) for providing of fungi from the Culture Collection of Mushrooms (IBK). This research was funded by the National Academy of Sciences of Ukraine (State Registration Number 0118U003812) for the Institute of Food Biotechnology and Genomics of the National Academy of Sciences of Ukraine and by the National Academy of Medical Sciences of Ukraine (State Registration Number 0119U002041) for Institute of Epidemiology and Infectious Diseases of the National Academy of Medical Sciences of Ukraine. Conflict of Interest The authors declare that they have no conflict of interest. References Badalyan SM, Innocenti G, Garibyan NG (2002) Antagonistic activity of xylotrophic mushrooms against pathogenic fungi of cereals in dual culture. Phytopathol. Mediterr. 41: 220-225. Badalyan SM, Innocenti G, Garibyan NG (2004) Interactions between xylotrophic mushrooms and mycoparasitic fungi in dual-culture experiments. Phytopathol. Mediterr. 43: 44-48. 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