Production of Mushroom Mycelium

Harry Humfeld.

Growing mushrooms has become an important industry in the United States. Of the estimated annual production of 62 million pounds, 20 million reach the consumer as fresh mushrooms, 18 million in cans, and 24 million as flavoring in soups.

Consumption might be even greater if production costs could be reduced. With the trend towards packaging prepared, ready-to-serve food products, the use of mushroom flavor in a number of such preparations doubtlessly will increase. The problem is to incorporate mushroom flavor without markedly increasing the cost of the product.

The mushrooms in the retail markets are really the fruits of a plant that grows as a mat of fine strands or threads on the organic material in the soil or in a mushroom bed. When conditions of temperature, moisture, and food supply are right, such a mat, or mycelium, periodically forms mushroom fruits near the surface of the soil. These are the familiar buttons and caps. Soon after they push up through the bed they are ready for harvest.

Although people over the world eat mushrooms of several species, in the United States only one species Agaricus campestris is grown commercially.

In 1947, I discovered that the vegetative, or underground, part (mycelium) of that species will grow under conditions of submerged propagation.

The technique offers the possibility of greatly cutting the cost of a mushroom-flavored food material, although the physical form and texture usually associated with mushrooms must be sacrificed.

The principles of the method are based on the fact that when a liquid that contains the required chemicals and food supply is continuously stirred vigorously at the right temperature and mixed thoroughly with a constant supply of air, many micro-organisms reproduce in it with great rapidity. The method is used commercially in the production of baker's yeast and feed yeast. More recently it has been applied to the large-scale production of antibiotics, such as penicillin and streptomycin, as well as to the manufacture of citric and gluconic acids.

The mycelium proved to be not too particular about the medium on which it will grow, but it does have certain requirements. It seems now that a good commercial product can be grown on any medium that contains a suitable sugar and other essential nutrients, that does not contain an ingredient inhibitory or toxic to the growth of the mycelium, and that does not impart a characteristic flavor-of its own during its use as a medium.

To OBTAIN PURE-CULTURE mushroom mycelium, we either germinated mushroom spores or made tissue cultures of various parts of the mushroom. Then we grew the mycelium on an agar medium in order to maintain the pure cultures. The cultures may be inoculated into sterilized compost, grain, or tobacco stems, and allowed to grow to provide the material known in the trade as mushroom spawn. The spawn, when planted in mushroom beds, produces the mushrooms sold commercially.

For the submerged propagation of the mycelium, the procedure now in use consists in transferring some of the mycelium grown on agar in a test tube to 50 milliliters of sterile liquid medium in a 250-milliliter Erlenmeyer (conical) flask. The flasks are shaken on a machine in a room at 77 F. until a heavy growth of mycelium has developed. The time usually required is 5 to 6 days. This inoculum is then transferred to 2 liters of medium in fermentors made from Fernbach (spherical) flasks and incubated to maximum growth. This culture suspension is used as inoculum in the larger fermentors with a culture-medium capacity of 16 to 20 liters.

It has been necessary to consider variations in so-called strains of Agaricus campestris. Plant species, like animal species, have familylike variations. Therefore I have isolated and tested more than 40 cultures of strains of Agaricus campestris.

Among them I have found three that are well able to adapt themselves to growth in an agitated, aerated liquid medium. Each has characteristic mushroom flavor. Each can be distinguished from the others by a characteristic difference in flavor. Two of the strains were isolated from mushrooms of the white variety and one from the cream or brown variety of Agaricus campestris. All have certain characteristics in common. They grow more rapidly than the other isolations. The hyphae, or mycelial threads, are more slender. They produce an abundance of so-called secondary spores in the liquid medium. The secondary spores were first described by Albert Kligman in 1932 at Pennsylvania State College. He found them in old cultures on Solidified (agar) medium. From observations up to the present time, it seems likely that the ability to produce the secondary spores may account for the ability of the strains to adapt themselves to submerged growth in a liquid medium.

The process also has been carried successfully through a semipilot-plant stage. That is, we have grown all three strains in 40-gallon batches in our pilot-plant fermentor. Each batch has produced 35 to 38 pounds of mycelial cake with good mushroom flavor.

In both the Fernbach and the larger fermentors, a rate of air flow of 1 liter of air a minute per liter of culture medium gives rapid growth. Lower rates give slower growth. Higher rates do not materially increase the growth rate.

We harvest when good growth and good mushroom flavor have been produced in the large fermentors by taking out half to three-fourths of the culture and adding an equal volume of fresh sterile culture medium. In this manner a number of consecutive harvests are obtained without contamination of the culture medium.

Immediately after harvesting, we separate the mycelium from the culture liquid by centrifuging. Then we re-suspend the mycelium cake in water and again centrifuge.

We pack the mycelium into suitable cans and seal and sterilize the filled cans at 15 pounds of steam pressure for 20 minutes. In some cases we have frozen it and stored it in a freezer until used. Drying the mycelium, either by lyophilizing (drying under vacuum from the frozen state) or on a drum drier, has been tried, with some loss of flavor in both cases.

We have obtained good yields on media composed of juices pressed from various fruit and vegetable wastes, such as asparagus butts and pear waste. Our more recent investigations show that a desirable product is obtainable from media made from pear waste, a rice-bran extract, or beet molasses, and also from a synthetic medium containing dextrose and inorganic salts. Media containing asparagus juice or alfalfa juice seemed to impart the flavor of those constituents and were deemed less desirable.

Basic studies in progress in 1949 demonstrated that the essential nutrient requirements for the growth of the mycelium of A. campestris are comparatively simple. Sources of nitrogen utilized include ammonia, urea, peptone, monosodium glutamate, a mixture of amino acids, and probably a number of other nitrogen compounds. Most of the sugars tested, which included hexoses, pentoses, and disaccharides, give good growth. The polysaccharides (soluble starch and dextrin) also are suitable sources. However, a form of soluble cellulose (sodium carboxymethyl cellulose) cannot be fermented. Apparently a wide range of carbohydrates can serve as sources of energy for growth of mycelium of A. campestris.