Comparative Study of Pleurotus Djamor

Cultivation on Sustainable Waste Substrates

Obuasi Boulware, Jonathan Carr, Julie Deslauriers, Shannon Foley, and Victoria Kreinbrink
BSC 4861L, Department of Biology, University of Central Florida
Orlando, FL 32816, USA

Abstract

Fungi are diverse and essential to the health of many ecosystems. They break down
organic material from waste and in turn are able to provide other organisms with essential
nutrients. Mushrooms are critical to the health of Earth because they recycle tremendous
amounts of waste. This experiment tested the productivity of 4 different recycled substrates and
their ability to produce the largest final mass of Pleurotus Djamor. Oak mulch, palm mulch,
coffee grounds, and a mix of all three were tested. The levels of mycelium growth were measured
rather than mushroom mass due to time constraints. The mixed substrate was predicted to have
the best results; however, coffee surpassed it and had the overall fastest development times. The
mixed substrate had the second quickest colonization, the palm mulch came in third, and oak
mulch mycelium growth was the least, never surpassing 20%. Although coffee had the quickest
colonization rates, the experimenters determined the mixed substrate to be the most sustainable
because of coffee’s vulnerability to environmental factors. Even though this is a small-scale
experiment, certain outcomes may lead to future large-scale changes, thus reducing the amount
of waste in public spaces by a noticeable amount.

Introduction
1,543 pounds of trash each year. An average
For hundreds of years, scholars have college student (21 years old) has already
been debating the carrying capacity of earth produced 16.2 tons of trash over their
in relations to the logistic growth model. lifetime. At this rate, if an American lives
Carrying capacity is the maximum for 80 years, they produce roughly 62 tons
population size that an environment can of trash. Putting it in a different perspective,
support (Campbell et al. 2011). With over an 80-year-old person produces a quantity of
seven billion people on Earth, when will waste that is equivalent in mass to about
Earth reach its maximum carrying capacity approximately 31 elephants. If we compare
and what will cause the human population this trend to the entire planet, Earth’s 7
decrease? Could it be that human’s leading billion inhabitants produce 5 billion tons of
predator is waste itself? trash each year.

According to a recent BBC This waste comes from the Linear

publication on global waste production, the Model of Production, a very common model
average American produces over 700 kg of used in industrialized countries. There is a
waste per year (BBC, 2013). That means long history of linear models of economic
each person in the United States produces activity, tracing back to Soviet economists
material balance tables (Blume). These

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models fluctuate between general and partial of utmost importance in order for a system
equilibrium and have no recycling pattern to to maintain itself (Bernard, 2013). This
balance the system (Blume). More experiment was designed to maximize
production means more disposal and in this “profit” through using free, recycled
linear model, there is no use for the waste. materials as growing mediums for
As seen in Figure 1, businessmen go home mushrooms that can be sold. Through using
happy with their paycheck, but will they be a variety of natural, readily available
happy when the Earth’s resources are resources, this experiment explores the most
barren? effective substrate for mushroom

Figure 1: Diagram From Story of Stuff by

Annie Leonard (Leonard, 2007). cultivation. This experiment also focuses on
“planet”, as the mushrooms will potentially
Where does all this waste go? If degrade their substrates sufficiently enough
there were no decomposers, this waste to be used as compost, which in turn can be
would suffocate the planet and massively used to grow other plants. “People” were
pollute living conditions. It is easy to say also focused on through the use of purely
that without decomposers, trash really would waste products as the substrates. Even
be a human ‘predator’ and the carrying though this is a small-scale experiment,
capacity of Earth would have been reached certain outcomes may lead to future large-
hundreds of years ago. However, scale changes, thus reducing the amount of
decomposers are essential organisms that waste in public spaces by a noticeable
degrade organic materials into a form that amount. These three pillars of sustainability:
can be easily utilized by other organisms. people, planet, and profit are all benefitted
Decomposers in nature often form a by the closed loop system created by
sustainable cycle of death, growth, and birth. decomposers (Figure 2, pg. 3).
When biotic organisms die, decomposers
grow from the remnants and break down the Edible mushrooms, particularly fast
larger molecules into a form more readily growing ones, such as Pleurotus spp. can
for plant uptake (Stamets, 2000). successfully form a closed loop system. The
first part of the system is to provide an
Sustainability is the key focus of this edible product (mushrooms) by using waste
experiment. The three pillars of byproducts (i.e., coffee grounds, tree
sustainability, people, planet, and profit, are trimmings, palm fronds, etc.) as the growing

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medium. Once the mushrooms have been making products ranging from bread to
sold, the spent substrate can in turn be antibiotics” (Campbell, 2008). Also, human
composted and eventually converted into nutritional needs are met by the fruiting
fertilizer to add nutrients back into the soil. body of the mushroom. Commercial
The fertilizer encourages other plant growth mushroom producers could benefit greatly
and eventually cycles around to produce and increase their profit by creating their
more mushrooms (Rinker, 2002). If not used “product” from free, recycled substrates.
for compost, the spent substrates can be While humans have already found a wide
utilized to produce animal feed or other range of uses for the products and services
crops and purify water and soil (Rinker, produced by these organisms, there is still
2002). room for further incorporation into societies’

Figure 2: Sustainable Cycle of Pleurotus Djamor

everyday means of operation.
According to an article published in
Conservation Biology, “Ecological Decomposers can be separated into
Sustainability as a Conservation Concept”, the 5 phyla of fungi, or Eumycota (Campbell
sustainability is achieved by meeting human et al. 2011). Fungi are diverse and essential
needs without compromising the health of to the health of many ecosystems. They
an ecosystem (Callicott, 1997). Mushrooms break down organic material from useless
exceed these criteria for sustainability by waste, and provide other organisms with
benefitting the ecosystem with a wide array essential nutrients. The 5 phyla of fungi
of ecosystem services. “Humans benefit include: chytrids, zygomycetes,
from fungi’s services to agriculture and glomeromycetes, ascomycetes (sac), and
forestry as well as their essential role in

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basidiomycetes (Campbell et al. 2011). shops supplied the coffee. All three
Basidiomycetes, or mushrooms, are the most substrates in the experiment are readily
commonly known phyla of fungi and will be available, inexpensive, and have few
the main focus of this experiment. alternative uses, which make them ideal
substrates for this experiment.
Similar to plants, basidiomycetes
disperse their genetic material via aerial or Additional components that are
water dispersal in seed-like capsules known typically used to increase mycelia growth
as spores. The visible part of a mushroom is and mushroom production such as rice bran,
actually an entire fruiting body, woven with calcium carbonate, and cottonseed oil will
reproductive hyphae in thread-like structures not be used in this experiment (Stamets,
(Campbell, 2008). The fungi produce a 2000). This is to allow direct observation of
fruiting body above ground in order to each individual substrate’s ability to provide
maximize the surface area in which the adequate conditions for mycelia growth and
spores will be exposed too. After the spores mushroom production. For example, coffee
are released, they spread out in search of already has high nitrogen content, and
nutrients and a stable place for survival adding an additional source of nitrogen is
(Campbell, 2008). Substrate is focused on in unnecessary. Also, adding supplements to
this experiment because ideal substrate and the substrate increases production costs and
climatic conditions are essential for maximal creates a need for stricter sanitation
mycelia growth and mushroom production. (Stamets, 2000).

Substrates chosen for this study were The species, Pleurotus Djamor, was
recycled materials that could all be found chosen for this experiment for many
within the UCF campus. Coffee was chosen reasons. Pleurotus cultivation popularity has
because it has high nitrogen content and has recently increased due to its desired taste as
been used many times as additives to well as numerous nutritional and medicinal
traditional substrates. Oak mulch was benefits. In 2008, oyster mushrooms rose to
chosen due to the observation that the third most commercially produced
mushrooms often typically grow on trees, mushroom in the world (Dundar, Yildiz,
such as oak, in nature. Lastly, palm mulch 2008). P. Djamor prefers tropical and
was chosen because palm trees are an subtropical regions and is known for its
abundant natural resource in Florida. As “speed to fruiting, adaptive ability to
well, many previous studies have been flourish on a wide variety of base materials,
found to test palm fronds as a substrate with and high temperature tolerance” (Stamets,
success (Kalita & Mazumder, 2001). A 2000). Also, P. Djamor grows on a wider
blend of all three substrates will also tested range of forest and agricultural waste
be as a fourth trial. Hypothetically, products than any other species within the
Pleurotus Djamor mushrooms should basidiomycota phyla. In past research, P.
produce at the highest yield on the blend of Djamor has been cultivate on most all
coffee grounds, oak mulch, and palm fronds hardwoods, on wood by-products (sawdust,
because each substrate will contribute to paper, pulp sludge), all the cereal straws,
maximize the nutrient content. corn and corn cobs, sugarcane bagasse,
coffee residues (coffee grounds, hulls, stalks
The UCF Arboretum supplied the and leaves) banana fronds, cottonseed hulls,
oak mulch and palm mulch. Campus coffee agave waste, soy pulp, and numerous other

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materials containing lignin and cellulose largest final mass of Pleurotus Djamor. A
(Stamets, 2000). total of six trials were performed for each
substrate, creating an overall test of 24
Further research within this genus substrates (Table 1).
has been stressed due to the species’ A 25 lb. bag of Pleurotus Djamor
adaptability. The potential to utilize Sawdust Spawn was ordered from an online
otherwise considered waste products has distributor (www.everythingmushrooms.com).
been studied as a means of reducing hunger The recycled substrates were collected
in developing nations (Anoliefo et al., directly from the University of Central
1999). Other research has been performed Florida campus to portray the accessibility
about the bioremediation abilities of P. and sustainability of the chosen substrates.
Djamor. When grown on materials The UCF Arboretum provided the oak
containing significant levels of mercury they mulch and palm mulch. A local campus
have been found to accumulate these heavy coffee shop provided the coffee grounds.
metals from the ground. Growing
mushrooms in such conditions may not be
useful for human consumption but can play
valuable role in the bioremediation of
contaminated ecosystems (Bressa et. al,
1988). In a study done by Paul Stamets,
oyster mushrooms were used to breakdown
residual oil in soil in a lot near a high traffic
area resulting in significantly less oil in the
soil and mushrooms free from petroleum
residues (Stamets, 1999). Pleurotus spp.
have also been shown to stun and digest
nematodes for access to necessary nitrogen,
which can be a nuisance in agriculture and
water quality (Thorn and Barron, 1984).

Methods

Oak Palm
Coffee Blend
Mulch Mulch
Trial 1 x3 x3 x3 x3
Trial 2 x3 x3 x3 x3
Table 1: Testing the substrates consisted of 3
replicates of each substrate, for a total of 24
jars. Overall, there were 6 jars of coffee ground Figure 3: Above- Substrates were submerged in
substrate, 6 jars of oak mulch substrate, 6 jars water in order to reach 100% moisture content.
of palm debris substrate, and 6 jars with a Below- To reach 55-65% moisture percentage,
proportional mixture of all three substrates. substrates with excess moisture were baked at
350°F for 15 min intervals.
This experiment was designed to test
The growing mediums (coffee, oak
the productivity of 4 different recycled
mulch, and palm mulch) were collected in
substrates and their ability to produce the

5
separate five-gallon buckets and submerged was too high, the substrate was baked at 350
in water. The substrates were soaked for degrees for15 minute intervals.
about three hours to ensure 100% moisture
content. The wet substrates were then laid The substrates were then placed in
out on tarps to sundry for 3 hours. baking trays, covered, and heated to 160° F
for one hour by oven. Each substrate was
Initial moisture content is of utmost then placed into an individual autoclaved
important for proper mycelium growth. wide mouth glass quart jars. Pleurotus
Many hobbyists use a method called the Djamor spawn was layered within the
palm test to determine proper moisture. The substrate to ensure thorough colonization In
palm test is determined by squeezing a order to minimize risk of contamination; this
handful of substrate. If the substrate is step was completed under a laminar flow
squeezed and it drips, the moisture content is hood. The jars were then capped, labeled
too high. Optimal moisture content is accordingly, and placed in a dark incubation
achieved when wet substrate is squeezed and room at ideally 75-85° F.
no water drips out (Stamets, 2000). There is
another method called the oven method
(described below). This method is typically
preferred by scientists. In this experiment,
the oven method was used to get more
consistent moisture content in the substrates.

Initial Moisture Content Per Substrate

Coffee Palm Oak
Trial 1 62.70% 58.70% 62.43%
Trial 2 61.60% 57.67% 61.79% Figure 4: Capped jars were placed in a dark
incubation room at 75-85° F.
Table 2: The initial moisture percentage for
both trials. Moisture percentage was Over the colonization period, the
determined by subtracting the dry mass from amount of mycelium growth in each jar was
the wet mass, dividing the total quantity by the observed every two days. The data was
dry mass, and multiplying by 100. recorded based on visual observation of the
quickness and quality of mycelium growth
The mass of one cup of each per substrate. Due to time constraints of this
substrate was found using a gram scale. The class, the data cut off point was collected
dry substrate mass was subtracted alongside before full mycelium colonization.
the experiment by placing a cup of each Therefore, results were recorded in
substrate in the oven at 350°F for 1 hour. mycelium growth. However, the experiment
The dry and wet masses were used to find continued and just a few days after the data
the percent moisture content, ideally aiming cut off point, the jars were fully colonized
for 55% to 65%. Proper moisture is essential and ready to fruit.
for mycelium colonization, and 55% to 65%
moisture content is the optimal condition At around 100% colonization (about
(Stamets, 2000). If the percentage was too 24 days), the jars were uncapped and placed
low, water was added, and if the percentage in a terrarium containing 100% humidity for

6
fruiting. The jars were wrapped in aluminum prediction that the mixed substrate would
foil to encourage vertical growth. have the greatest success in growing
mushrooms.

Figure 5: Jars covered in aluminum foil in

terrarium.

Results
The results of this experiment were
measured as a percentage of visible
mycelium growth rather than mass of
fruiting bodies due to time constraints. As
visible in Figure 6, coffee had the overall
fastest colonization time within this
experiment. Coffee was followed by the
mixed substrate, which had the second
fastest colonization time, and palm mulch
finished third. Oak mulch had barely any
colonization remaining less than 20%
throughout the experiment.
Figure 6. Above-The overall growth rates
Additionally, the second run of the averaged among the two trials. Below- Trial 1
is represented solid lines and Trial 2 with
experiment had faster development times dashed lines.
than the previous run. This held true for all
four substrates being tested. The mixed
Discussion
substrate’s second trial results developed at
the same pace as the first trial of coffee, only
being surpassed at the end. The second trial Contrary to the original hypothesis
of palm mulch surpassed the growth the first that the mixture of palm mulch, oak mulch,
mixed trial at the end of collection as well and coffee grounds would colonize at the
(Figure 6). fastest rate, the coffee grounds used as a
substrate gave the quickest colonization
The findings that coffee produces the times. The mixed substrate had the second
highest yield, contradicts the original quickest colonization, the palm mulch came

7
in a close third, and oak mulch mycelium Due to time constraints of the
growth never surpassed 20% growth. As a project, the experimental data was collected
result of this, it is believed that by oak before fruiting bodies emerged. Therefore,
mulch being in the mixture, it may have instead of determining the efficiency
decreased the nutritional value of it enough substrates by comparing mushroom mass,
to put it at a disadvantage when compared to the mycelium growth was analyzed. As a
coffee alone, which is extremely nutrient result of this, there may be some bias in the
rich. If not the nutritional value, then the mycelium growth results reported as they
high density of oak mulch pieces may have were based on outward visual appearance,
been the culprit for the slow rate of and inner growth in the jars could not be
mycelium growth. While mushrooms are properly accounted for. This was discovered
one of the few things able to break down after the appearance of mushrooms in all of
lignin, a very complex polymer found in the jars. These results were unexpected,
wood, it can cause growth to be a much especially in the case of oak mulch, where
slower process as the nutrients are not as barely any mycelium growth had been
easily accessible as in other substrates, ie. observed. It is believed that even though the
coffee (Buswell & Odier, 1987). mycelium growth may not have been
visible, it was still occurring in the middle of
Although coffee had the quickest the substrate, where it was not visible. It is
colonization rates, it was later determined possible this is due to the density differences
that the mixed substrate is the most among the substrates, something like coffee
sustainable because of coffee’s vulnerability grounds having a very low density and being
to environmental factors. Despite proper easy to maneuver through, compared to
pasteurization techniques, the nutrient chunks of oak mulch being hard to break
richness of coffee attracted other types of apart.
fungi and some jars were contaminated. This
was visible through a green mold growing
throughout 3 of the jars. Contamination such
as this would be detrimental to mushroom
growers. In Pennsylvania in the 1990s, the
mushroom industry experienced losses of
between 30-100% as a result of this mold
(Tisdale, 2004). Coffee was also determined
to be less stable than the other substrates. At
one point late in the experiment, the
humidifier stopped for a brief period of time
(approximately 5 hours), which caused the
coffee substrate pull away from the sides of Figure 7. Mushrooms growing on the coffee
the jar and mushrooms to slightly shrivel, substrate.
whereas the other substrates remained
stable. Coffee’s inability to maintain Mushrooms started to appear only a
moisture content would be unfavorable in few days after the experimental cut off
larger outdoor experiments that could point, and shocking observations were
potentially be faced with periods of drought reported. Even the oak mulch, which
and flood. visually had the least mycelium growth
produced mushrooms. A future experiment
analyzing the mass of fruiting bodies will

8
provide more information about the Future Work
effectiveness of growing on each substrate.
The findings of this experiment have
As far as measuring the percentage led to further questions being asked. After
of substrate converted to compost, discovering that coffee rather than mixed
mushrooms grow as they decompose more grew the fastest, possibly being due to high
and absorb more from their substrate. As a lignin content/high density or low nutritional
result of this, it is reasonable that the larger value of oak mulch in the mix, the
the mushrooms are, the more the soil has experiment was rethought out. The
been broken down into compost. Based off experiment should be repeated in the future
of these results, both coffee and the mixed with a mix of coffee and palm alone.
substrates were composted the most, as they Predicted results from this experiment are
produced the largest mushrooms, shown in Figure 8.
presumably from degrading the soil at a
higher rate. If removal of waste is the main
focus of future growers, then mixed
substrate would be the best option as it
allows for conversion of three types of waste
into compost rather than just one.

This can cycle back into the concept

of sustainability. By using the spent
substrates, they can then be composted and
eventually converted into compost to add
nutrients back into the soil. Coffee grounds
are especially a readily available source of
“waste”, which is highly beneficial to the
process and will allow for a continuous
supply of substrate to produce at high yields
for the community. Figure 8. Previous experiment results along
with predicted results for a palm and coffee
mixture (dashed line).
Some may see placing all jars in the
same terrarium as another possible bias in In theory, the combination of palm
this experiment; however, even if spores and coffee without oak will be the best
transfer substrates during the cultivation combination of the three. This is because the
period, the spores will not have enough time palm much should decrease the risk of
to develop into fruiting bodies and will contamination as it will just slightly lower
therefore not affect the final mass, creating the richness of the coffee. Additionally, the
no experimental bias. Also, there is no need oak mulch will not be an obstacle for
to worry about cross contamination of mycelia growth. As well, by using a
species since all of the spawn used across combination of two of the three rather than
the various substrates will be one consistent one alone, more waste will be able to be
species, Pleurotus Djamor. composted.

9
 An additional issue in this September 26, 2013, from
experiment was finding the exact moisture http://www.bbc.co.uk/schools/gcsebitesize
content of the substrates in the beginning. /geography/wasting_resources/waste_poll
Eventually, the oven method was decided ution_rev1.shtml
on, but even though this was the most
precise method found, it was still not exact. Blume, L. (n.d.). Linear models of
While pasteurizing the substrates in the production. Informally published
oven, some moisture tended to escape manuscript, Cornell University & The
through the lids of the containers, so the Santa Fe Institute & IHS, Ithaca, NY,
final exact moisture content was most likely http://elaine.ihs.ac.at/~blume/LinearModel
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As a result, a further experiment is Bressa, G., L. Cima & P. Costa. 1988.

planned in which autoclavable bags will be Bioaccumulation of Hg in the mushroom
filled with dried substrates (0% moisture Pleurotus ostreatus. Ecotoxicology and
content) to start. The amount will be Environmental Safety Oct. 16(2), 85-89
weighed and 65% water will be added to the
bags. The bags will then be sealed shut and Buswell, J.A. & Odier, O. 1987. Lignin
autoclaved. This process should biodegradation. Critical Review of
theoretically cause the water within the bags Biotechnology 6, 1-60.
to steam the substrate throughout, thus
creating a completely uniform moisture Callicott J (1997). Conservation Biology.
content within the bags. The substrates Ecological Sustainability as a
should then give more consistent results. Conservation Concept. 11(1), 32-40.

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