Biotechnology of morel mushrooms: successful fruiting body formation and development in a soilless system
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- Masaphy, S. Biotechnol Lett (2010) 32: 1523. doi:10.1007/s10529-010-0328-3
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Morchella spp. ascocarps (morels) are some of the world’s most sought-after mushrooms. Successful cultivation of morels is still a rare and difficult task despite over 100 years of effort. Here we provide the first report of successful Morchellarufobrunnea fruiting body initiation and development in laboratory-scale experiments. Mushroom initials appeared 2 to 4 weeks after first watering of pre-grown sclerotia incubated at 16 to 22°C and 90% humidity. Mature fruiting bodies reached 7 to 15 cm in length and were obtained after the five morphological developmental stages of this Morchella species: sclerotium formation, scelerotium germination, asexual spore formation, formation of initial knots and development of the fruiting body.
KeywordsFruiting bodyMorchella rufobrunneaMorelMorphological developmentMushroom cultivationSoilless culture
Morels (Morchella spp.) are naturally growing and commercially important edible mushrooms with a delicate taste and a unique appearance that make them one of the world’s most sought-after mushrooms (Carluccio 1989). In the western world, morels are considered a gourmet food, resulting in their increased harvesting from the wild. In nature, morels have a short season, usually on the order of a few weeks, mainly in spring, and fresh mushrooms can only be found in the markets for a few weeks per season. This, and the increasing number of reports on accumulation of metal in ascocarps that are picked from natural habitats (Isildak et al. 2004; Shavit 2008), have resulted in the need to develop a biotechnological process to cultivate morels under controlled conditions.
Although such efforts began over 100 years ago (Roze 1882), it was only in 1982 that the first indoor cultivation of morels was reported (Ower 1982), with Morchella esculenta being the first Morchella species, followed by others, reported as culturable by the same author (Ower et al. 1986). However, growth of morel ascocarps under controlled conditions and repeating Ower’s success have proven to be difficult tasks. The failure to cultivate morels has been suggested to be partly related to insufficient knowledge of the factors controlling fruiting body initiation and development, but also to insufficient knowledge of which species are suitable for cultivation, the latter being an important aim.
In the past, morels were divided phenotypically into two groups, the Black and Yellow morels (Boudier 1897), and were studied as such for their indoor cultivation options (Ower et al. 1986). Recently, a new group of morels has been characterized: the red-brown blushing morel represented by the species Morchella rufobrunnea. It differs morphologically from the known Yellow and Black morel species (Kuo 2008; Masaphy et al. 2009; Pilz et al. 2007) in having a conical head with pale tan pits and reddish brown ridges when mature. This species has been found in different sites throughout the world, and confirmed as genetically distinct from the Yellow and Black morels (Guzman and Tapia 1998; Kuo 2008; Masaphy et al. 2009; Pilz et al. 2007). The various habitats in which it has been discovered—disrupted soil in a healthy forest, the forest after a fire, and road beds—imply its saprophytic ecotype.
Here we provide a first report on the successful initiation and development of fruiting bodies of M. rufobrunnea in a soilless controlled process which has been conducted at MIGAL’s mushroom experimental cultivation growth facility since 2002. The developmental changes in the fungus’s morphology during the cultivation process are also reported.
Materials and methods
The M. rufobrunnea (MIGAL strain MS3-730) described in this report has been reported previously (Masaphy 2005).The isolate has been preserved in the author’s collection at MIGAL Institute (Kiryat Shmona, Israel).
Growth conditions were as described in an earlier work (Masaphy 2005). For the sclerotial production phase of the process, the fungal mycelium was inoculated into a sterile (1 h at 120°C) of potting soil buffered with limestone. The inoculated medium was placed over a layer of nutritionally rich medium based on wheat grains, and incubated for 2–3 weeks at 18–25°C. The sclerotium-containing layer was kept at 4°C until its use for the second phase of the process, in which sclerotia were subjected to continuous watering for 5–24 h. The induced sclerotia were then incubated at 16–22°C for 2–4 weeks for carpogenic initiation and fruiting body development. The different morphological developmental stages were followed during the growing process and photographed.
Scanning electron microscopy
Conidial production was studied by scanning electron microscopy (SEM) according to Masaphy (2005). A sample of the conidial layer was transferred to a solution of 4% (v/v) glutaraldehyde in 0.1 M phosphate buffer (pH 7.2) for 1 h. The sample was dehydrated in a graded series of ethanol washes (10, 25, 50, 70 and 100% ethanol for 60, 15, 15, 60, 60, 15 min, respectively). After dehydration, the samples were critical-point-dried through liquid CO2 adhered to stubs, coated with a gold–palladium layer, and observed under a scanning electron microscope (SEM) (JEOL JSM 35C, Japan).
Results and discussion
In nature, morel fruiting body formation is associated with a broad range of environmental stress conditions, some of which have yet to be defined. This conclusion is based on the high diversity of habitats in which morels are found (Pilz et al. 2007), and the high number of Morchella species reported (Gessner 1995).
Here we provide the first report of successful M. rufobrunnea fruiting body production under controlled conditions, and M. rufobrunnea’s entire life cycle in the experimental controlled cultivation system is described. Some of these developmental stages have been previously described for other reported cultured Morchella species (Ower 1982; Ower et al. 1986). However, M. rufobrunnea was not mentioned in the list of cultivable species in those reports. The results may indicate identical life-cycle stages in different species of Morchella, within the groups of Black, Yellow or Red-Brown morels. Successful fruiting body initiation of M. rufobrunnea in soilless culture confirms that it is a saprophyte, and the search for other saprophytic species that can produce fruiting bodies under controlled conditions is therefore warranted. However, this goal will be easier to achieve once the species confusion surrounding the genus Morchella is resolved.
The authors thank Prof. Dan Levanon, Dr. Doron Goldberg and Dr. Ofer Danai for their help. This work was supported by the Office of the Chief Scientist, Ministry of Agriculture, Israel.