著者

大賀 祥治

出版者

九州大学

雑誌

九州大学農学部演習林報告 (ISSN:04530284)

巻号頁・発行日

vol.61, pp.1-90, 1989-11

被引用文献数

3

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シイタケ菌とヒポクレア菌などの害菌の間には拮抗現象が期待できることを見出した.この場合,特にシイタケ菌の蔓延率が重要な因子となり,蔓延が進み培地表面を完全に覆った状態であれば,ヒポクレア菌の侵入に対し,十分抵抗性を示す結果を得た.また,他の因子として,培養温度,培地含水率,pH,ヒポクレア菌の胞子濃度等も重要な因子であった.これらの結果から,より速くシイタケ菌糸を蔓延させ高い菌糸密度を保つことが害菌対策上,極めて重要であることが明らかになった.シイタケ菌の初期蔓延を活発にし,培地をより早く完熟化すること,また害菌の胞子濃度が低い環境を選定することがそれぞれ重要事項であることが示唆された.そこで,シイタケ菌糸を積極的に蔓延させる手段として,手近に得られる蔬菜類を中心とした天然物の熱水抽出物,パルプ廃液成分,クロレラ熱水抽出物等を添加物として取り上げ,まず室内試験を中心として検討した.シイタケ菌糸の蔓延を著しく促進する作用を示すものが見出された.これらは,寒天,木粉,小径木等培地形態の違いにより,異なった挙動を示した.寒天培地ではパルプ廃液成分,クロレラ熱水抽出物,木粉培地ではクロレラ粉末,ネギ煎汁,小径木ではニンジン煎汁,ミカン煎汁がそれぞれ有効であった.そして,大型の鋸屑培養,すなわち菌床栽培の手法で検討を行った.添加物としてネギ煎汁を投与することで,菌糸の蔓延に対し促進あるいは刺激作用を加えたものは完熟菌床に至る期間が短く,しかも,後続する子実体発生に際しても良好な効果を持続し,発生量の増加が期待できることが分った.次に,添加物投与による効果をほだ木でのシイタケ栽培に適用することを考えた.ところが,ほだ木内におけるシイタケ菌の蔓延度,特に菌糸密度の高さを示す熟度を正確に知る適当な方法がなく,室内試験で明らかにした添加物の影響についての判定が困難であった.シイタケ栽培では,熟度を正確に把握することが子実体発生のための必須条件となっているにもかかわらず,判定法がこれまでに確立されていなかった.そこで,菌糸蔓延度の判定法を確立することが添加物投与の検討を行ううえで必要事項と思われた.呈色反応,炭酸ガス放出盛量,培地表面の白色度を測定することが有効な方法であることを認めた.特に,pH指示薬であるブロモフェノールブルーを噴霧した際の呈色反応により,簡易にしかも,正確にほだ木の菌体量を把握できることを明らかにした.5年間のほだ木栽培での結果,添加物投与の効果が野外試験においても認められた.添加物投与により菌糸の初期蔓延が活発になったほだ木では,ほだ木内により速くシイタケ菌糸が蔓延し,完熟ほだ木になるまでの期間が短縮された.その結果として,害菌による被害率が低くなり,さらに後続する子実体発生量が多くなった。効果はミズナラ,コナラ,クヌギ,アラカシ,ツブラジイ,マテバシイおよびタブノキの7樹種のうち,ほとんどのもので確認された。子実体発生試験はミズナラほだ木で行ったが分散分析の結果,有意差が認められたものが多く,しかも子実体の形質が優れ,特に菌傘の厚さが増すという結果が得られた.添加物の投与時期は種駒接種と同時に行うのが,最も効果が期待できることを明らかにした.菌糸が蔓延して完熟ほだ木になった時点では添加物投与を行っても効果はみられず,この段階では保水力の高い物質を加えることにより子実体発生量が増加した.また,添加物を授与する手段として,脱脂綿に含浸させて種駒接横穴に挿入し,その上から直接種駒を打ち込む方法(脱脂綿挿入法)を新しく考案した。添加物としてはネギ煎汁が最も優れ,穫駒接種時の投与によりほだ木内への菌糸蔓延率が高まり,さらに子実体発生数が明らかに多くなり,きわだった効果をみせた。これまでの検討でシイタケ栽培における添加物として最も効果が高いネギ煎汁中に存在する生育活性化物質の単離,同定を行った。シイタケ歯の生育に対し,煎汁の固形分濃度が大きな因子となり,また,その作用は培地形態により異なる結果が得られた.ゲルろ過法により活性物質を分画でき,活性の主体は核酸関連物質であることが明らかになり,種々の分析でこれらは3'-AMP,アデノシンおよびアデニンであることが判明した。種々の核酸関連物質のうち,アデノシン,アデニン等がシイタケ菌の生育に対し,極めて高い促進効果を示すことを明らかにした.また,ネギ煎汁にはニンジン煎汁,モヤシ煎汁に比べ,これらの核酸成分の含有量が高いことが分った.さらに,ネギ煎汁に含有されるシスチンおよびシステイン等の含硫アミノ酸が活性を示し,アデノシンならびにアデニンと相乗効果を示した.これらの結果からネギ煎汁中のシイタケ薗生育活性化物質の主体はアデノシン,アデニン等の核酸関連物質とシスチン,システイン等の含硫アミノ酸であると考えられた.As a result of tests on the administration of additives in the production of the shiitake mushroom (Lentinus edodes (Berk.) Sing.), the following was made clear. 1. Antagonism of Shiitake Fungi with Hupocrea spp. Fungi An antagonistic phenomenon was discovered between shiitake fungi and Hypocrea spp., an injurious fungi. Namely, results showed if shiitake fungi did not completely cover the surface of a medium, such as an agar medium, Hypocrea spp, invaded the shiitake fungi. On the other hand, if the surface of the medium was covered throughout, the shiitake fungi showed sufficient resistance against Hypocrea, and shiitake fungi increased its biomass. Further, other important factors such as; culture temperature, the water content of a medium, pH, the amount of spores of Hypocrea spp. fungi or the like were also found. Accordingly, as a measure to protect shiitake fungi from injurious fungi, it is extremely important to grow shiitake fungi rapidly to keep a high mycelial biomass in the medium. Results indicate that initial activity of shiitake fungi is an important factor in protecting from an injurious fungi (Chapter I). 2. Effect of Additives in Indoor Tests As a means for promoting the growth of shiitake mycelium, several additives were tested, that is; hot water extracts of natural substances centering around vegetables; liquor-components from sulfite pulping (lignin carbohydrate complex sulfonate) ; hot water extract of chlorella; or the like. They were chosen to perform experiments in a laboratory, at first. The liquor components from sulfite pulping and chlorella extract were effective in the agar medium. Chlorella and welsh onion extracts were effective in the wood flour medium. Lastly, carrot and orange peel extracts were effective with the small log. Following tests for selecting proper additives, a sawdust substrate cultivation test was conducted. To promote the growth of mycelium, the administration of the welsh onion extract was applied as an additive, resulting in a shorter spawning period. Furthermore, successive growth of mycelium was positively affected in succeeding generations, resulting in an increase in the production of fruit bodies (Chapter II). 3. Determination of Mycelial Biomass An idea was pondered as to whether the administration of additives would promote the production of shiitake mushroom on natural bed log cultivations. There was no proper means, however, for accurately determining the mycelial biomass of shiitake fungi in a bed log. Therefore, it was difficult to judge the effect of additives discovered in indoor tests. In the production of shiitake mushroom, though the accurate determination of the mycelial biomass is an essential matter for the formation of a fruit body, a weighing method has not been established up to date. Therefore, the establishment of a method for judging the mycelial biomass was considered to be a necessary matter in order to perform an investigation on the administration of additives. Some methods were effective, that is; measuring color reaction; the evolution quantity of carbon dioxide; and, the surface whiteness of a medium. Especially, it was shown that the mycelial biomass in bed log could be simply and accurately determined by color reaction generated when bromophenol blue, a pH indicator, was sprayed (Chapter Ⅲ). 4. Effect of Additives in Field Tests The amount of fruit bodies harvested from natural bed logs was analyzed for five years. The effect on the administration of additives was confirmed in field tests. In bed logs having greater mycelial biomass, by means of the administration of additives, the spawn run time was shortened. As a result, the damage rate caused by Hypocrea spp. decreased and, further, in succeeding generations, the amount of fruit bodies was increased. This effect was cofirmed in almost all seven species of trees, that is; Quercus mongolica ; Q. serrata ; Q. acutissima ; Q. glauca ; Castanopsis cuspidata ; Pasania edulis ; and Persea thunbergii, Tests of fruit body production were conducted using a Q. mongolica bed log and, using a variance analysis, a significant difference was confirmed in many cases and, moreover, results showed that the size of fruit bodies was excellent and the pileus thickness was greatly increased. By performing the administration of additives simultaneously with the inoculation of a spawn plug, the maximum effect of generating fruit bodies will be achieved. On the other hand, if the additives are administered at a time when the mycelium covers the bed log completely, instead of administering them simultaneously with the inoculation of a spawn plug, no positive effects were confirmed. By adding a substance having a high water retentivity when the mycelium covers the bed log completely, it was found though, that the amount of fruit bodies was increased. Further, as a means for administering the additives, a mothod for inserting absorbent cotton, impregnated with additives, in a spawn plug inoculating hole and directly inoculating the spawn plug, was newly devised. As for the additives, the hot water extract of a welsh onion was most efficient and growth into the bed log was enhanced by the administration of the extract at the inoculation time of the spawn plug and, further, the amount of fruit body production was clearly increased and a marked effect was developed (Chapter Ⅳ). 5. Growth Activating Substances to Shiitake Fungi in Hot Water Extract of Welsh Onion On the basis of the foregoing tests, the growth activating substance present in the hot water extract of the welsh onion had the highest effect as an additive in the cultivation of shiitake. The concentration of extract becomes an important factor with respect to the growth of shiitake mycelium and, further, according to different medium, several results were obtained. Active substances could be fractionated by gel-filtration, it was mainly based on nucleic acid related substances. These substances were adenosine-3'-triphosphate, adenosine and adenine by various analytical methods, that is; colorimetric analysis; ultraviolet spectrum; thin-layer chromatography; high-performance liquid chromatography; and, gas chromatography-mass spectrometry. Adenosine, adenine or the like among various nucleic acid related substances showed extremely high growth promoting effects with respect to shiitake mycelium. Further, it was found out that the content of these nucleic acid components was high in the welsh onion extract as compared with extracts of carrot or bean sprout. Furthermore, sulfur-containing amino acids such as cystine, cysteine or analogues contained in the welsh onion extract showed activity and imparted synergistic effects to adenosine and adenine. From the results, the growth activating substances of shiitake mycelium in the welsh onion extract were based on nucleic acid related substances such as adenosine, adenine or the like and sulfur-containing amino acids such as cystine, cysteine or the like (Chapter V).