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DELIVERABLE REPORT
Grant Agreement number: 289843
Project acronym: GRATITUDE
Project title: Gains from Losses of Root and Tuber Crops
Funding Scheme: Seventh Framework Programme
Date of latest version of Annex I against
which the assessment will be made:
2011-11-15
Deliverable Number: D 4.1
Deliverable Title: Vegetative growth and mushroom production
on cassava waste
Deliverable leader, organisation, tel and
email:
Anton Sonnenberg, Plant Research
International, Wageningen UR. +31 317
481336. [email protected]
Contributing authors to this deliverable: Johan Baars, Marcel Visser
Name, title and organisation of the
scientific representative of the project's
coordinator:
Dirk Pottier, Scientific Officer, European
Commission - DG for Research and Innovation,
E3: COV2 07/157, B-1049 Brussels, Belgium,
Tel: +32 229 67209;
Project website address: www.fp7-gratitude.eu
ii
Table of Contents
Table of Contents ............................................................................................................................. ii
1. Summary ......................................................................................................................................... I
Introduction .................................................................................................................................................. I
2. Key Findings ................................................................................................................................. II
3. Deliverable Objectives .............................................................................................................. II
4. Background ................................................................................................................................... II
5. Methodology ................................................................................................................................. II
Preparation of substrates for testing vegetative growth ...................................................................... II
6. Results ............................................................................................................................................ V
6.1 Tests on vegetative growth of fungal species/strains on cassava waste. ....................... V
6.2 Effects of fungal growth on digestibility by rumen microflora ......................................... IX
6.3 Mushroom production of selected strains on cassava waste ............................................ XI
7. Conclusions ............................................................................................................................... XIII
8. References ................................................................................................................................. XIV
Annes 1 Pictures of mushroom production ....................................................................... XIV
Annex 2 Statistical analysis of yield data ............................................................................ XIV
I
1. Summary
Varieties of three different species of edible fungi have been grown vegetatively on cassava
peels, cassava sticks and a mixture of the two (1 : 1 (w/w). Growth has been estimated by
measuring linear growth rate in tubes containing the different substrates. Very accurate
growth rate could be estimated in this way. The order of growth rate on different substrates
was stalks > a 1:1 mixture of stalks & peels > peels. The order of growth rate for different
species was Pleurotus ostreatus > Pleurotus pulmonarius > Pleurotus eryngii > Lentinula
edodes. Variation between strains within one species was limited. After full colonization,
each strain-substrate combination was further incubated for an extra 2, 4 and 6 weeks.
These samples have been analysed for digestibility by ruminants in a model system showing
that especially cassava sticks (stalks) can be made digestible almost comparable to peels
after pretreatment with fungi of edible mushrooms.
Mushroom production of cassava waste (peels, sticks and 1:1 mixtures of peels and sticks)
were used to produce mushrooms of grey oyster and lung oyster mushrooms. Yield up to
15% biological efficiency (fresh weight mushrooms/fresh weight substrate) were achieved
indicating that waste of cassava only is in principle suitable for the production of edible
mushrooms. Addition of nitrogen rich waste (such as rice or wheat bran) will certainly
increase yields to a level comparable to substrates now used (mainly saw dust)..
Introduction
For a viable business on mushroom production on cassava waste materials (peels and
sticks), fungal strains producing edible fungi must first grow well vegetatively on waste
materials. Subsequently, species/strains that show a good colonization of waste can be
tested for the production of edible mushrooms. Species should be selected that are
commercially available now and thus known to consumers and also easy to cultivate. The
first part of the report deals with the selection of the best mushroom species/strains-
substrate combination for small scale testing of mushroom formation on cassava waste. For
this, 30 mushroom strains representing 4 species have been tested for vegetative growth on
cassava peels, cassava sticks and a mixture of the two (1:1). Four strains from FIIRO Nigeria
were included. A selection of strains were incubation for an additional period (2 and 4
weeks) to see if this increases digestibility and makes cassava waste a better substrate for
animal feed.
A selection of 4 strains, representing 2 species, showing a good colonization of different
waste materials were subsequently selected for mushroom cultivation on a small scale to
evaluate the potentials for mushroom production. Strains were selected that were shown to
produce mushrooms on an elevated temperature (>22 oC), since cooling capacity is limited
especially in African countries and would make cultivation on household level and small
enterprises impossible.
II
2. Key Findings
Mycelia of different varieties of the common oyster mushroom, brown oyster mushroom,
king oyster mushroom and shiitake were grown cassava peels, cassava stalks or a 1:1
mixture of both waste products. In general all species and varieties did grow on all waste
types but growth rate differs per species and waste type, i.e. for waste type: stalks> mixture
stalks/peels and for species: Pleurotus ostreatus > Pleurotus pulmonarius > Pleurotus eryngii
> Lentinula edodes.
Pretreatment of especially cassava sticks with fungi increases the digestibility for animals
considerably as tested in a rumen model. Substrates based on cassava waste only can be
used in principle to produce oyster mushrooms with yield up to 150 grams per kilogram
substrate (biological efficiency of 15%). This is high considering the very low nitrogen
content of the cassava waste based substrates. It is expected that addition of nitrogen rich
waste (like rice and wheat bran) can increase yield top levels comparable to saw dust based
substrates used at this moment.
3. Deliverable Objectives
To develop technologies for making food products from waste which is either in the form of
food products, animal feeds or mushrooms made from composting waste.
4. Background
Fungi are specialist in mineralization of a broad range of waste materials, especially the
most abundant waste on earth, i.e. lignocellulose. A large variety of waste steams can thus
be used to produce new food as edible mushrooms. To make cassava production a more
profitable crop it is, therefore, obvious to test also if waste products generated by cassava
production can be used to generate new food products such as edible mushrooms. When
good starting materials is available (strains and inoculum, the latter also known as spawn),
mushrooms is a low tech business and can be done on a small and large scale. Mushroom
production has thus potentials to for developing countries to generate food on waste and
additional income for small enterprises.
5. Methodology
Preparation of substrates for testing vegetative growth
III
During the first half year of 2013 preliminary work was done on optimization of substrate
composition. For this we used peels (milled and sieved, ca. 2 mm sieve, i.e. MESH 60 or 80),
stalks (milled and sieved, ca. 2 mm sieve, MESH 60 or 80) and a mixture of milled peels and
milled stalks (on a dry weight basis 50/50, w/w). For proper wetting, substrates were
submerged overnight in water. After this, excess water was removed from the substrates by
pressing as much water as possible from the substrate. The wetted substrate was used to fill
glass tubes to a height of 12 cm. Substrate was filled into the tubes in small portions. After
each portion, the substrate was “packed” by inserting a metal rod into the tube until it
rested on the substrate. The metal rod fitted neatly into the tubes and by its weight
produced an evenly packed substrate throughout the full height of the tube (i.e. 12 cm).
After this the tubes were capped and sterilised (1 hr. 121 oC). The three different substrates
were tested for their suitability for colonization by 30 strains listed in Table 1. Strains were
first grown on malt extract agar. After a few days of growth (colony diameter about 4-5 cm),
5 mm (Ø) plugs were taken from the edge of the growing colony. In total 5 of these plugs
MES-number Strain name Origin Remarks
Lentinula edodes
MES 02089 4B Su Xiang China Tested previously 1
MES 02008 sh 02/07 China Tested previously 2
MES 02110 le 03/13 Unknown Tested previously, commercial strain? 3
MES 02052 sh 02/02 China Tested previously 4
MES 02090 S600 Unknown Tested previously, commercial strain? 5
MES 00001 donko type 3715 Unknown Commercial strain 6
MES 00052 Lentinula edodes -2 Indonesia 7
MES 00061 Lentinula edodes -3 Indonesia 8
MES 02015 sh 03/01 Indonesia 9
MES 02016 sh 03/02 Indonesia 10
Pleurotus ostreatus
MES 14029 Florida Nigeria 11
MES 01997 pc 8-2-02 p2 Brasilia 12
MES 01998 pp 8-3-02 Brasilia 13
Pleurotus pulmonarius
MES 14030 Lagos Strain Nigeria 14
MES 14031 Ibadan Strain Nigeria 15
MES 11698 Jamur Amis - 3 Indonesia 16
MES 03448 pl 00/11 Unknown Commercial strain 17
MES 03444 G21 Unknown Commercial strain 18
Pleurotus eryngii
MES 12060 Le Lion PE Unknown Commercial strain 19
MES 03467 Sylvan 3065 Unknown Commercial strain 20
MES 02073 Mycelia 2600 Unknown Commercial strain 21
MES 03757 Pl02/08 China 22
MES 02001 Pl03/01 China 23
MES 12920 AL10 Iran 24
MES 12936 F21 Iran 25
MES 12961 M07 Kourdistan 26
MES 12970 M16 Kourdistan 27
MES 12929 D29 Iran 28
MES 12939 G02 Iran 29
MES 12978 S07 Kourdistan 30
Table 1. Species/strains used to test vegetative growth on cassava waste.
IV
were used to inoculate a single glass tube. After inoculation the strains were incubated at
24oC and checked at 3-5 day intervals for linear growth. Growth fronts were marked on 2
perpendicular axes on the side of the tube. For each strain linear growth was tested in 3
tubes (so called “race-tubes”). At the end of the experiment, the tubes were used for
determination of rumen digestibility. For this one tube has been frozen en freeze-dried after
2 weeks of colonisation, the second tube after 4 weeks and the last tube after 6 weeks.
Digestibility has been assessed after the method described by Cone et al. (1996). In short:
Rumen fluid was collected in the morning from two non-lactating Holstein–Friesian cows fed
1 kg of concentrate daily and grass silage ad libitum. Samples were incubated in triplicate
with rumen fluid for 72 h to determine gas production characteristics of the control and the
fermented substrates.
Mushroom cultivation trials on cassava waste
Peels and woody parts (sticks) of commercial cassava cultivation were send by FRI (Accra).
The woody parts comprised of sticks with a diameter of about 2-3 cm. The peels were sent
as dried material. The cassava waste materials were used to prepare 5 different substrates
(Table 2), three of which consisted solely of cassava waste material. Two consisted of
cassava waste materials mixed with saw dust (oak). As a control we included commercially
prepared wheat straw based substrate that is used by Dutch oyster mushroom growers.
Table 2. Substrate composition for small trials of mushroom production
Cassava stalks Cassava peels Saw dust CaCO3
Substrate 1 25% 75% 0% 1%
Substrate 2 50% 50% 0% 1%
Substrate 3 75% 25% 0% 1%
Substrate 4 25% 25% 50% 1%
Substrate 5 12.5% 12.5% 75% 1%
Substrate 6 Commercially available wheat straw based substrate (used by Dutch oyster
mushroom growers.
For the experiment 4 strains of Oyster mushroom were selected; Pleurotus pulmonarius
(lung oyster) strains MES03444 (commercial strains, originating from Slovakia) and
MES03448 (commercial strain, country of origin unknown) and Pleurotus ostreatus (grey
oyster) strains MES01997 and MES01998 (originating from Brazil). These strains were
chosen since they fructify at a higher temperature than strains used by European growers
(cultivation temperatures between 16 and 18 oC). Cassava peels are rich in starchy
components. In mushroom cultivation the presence of starchy components in substrates
poses a risk for contamination of the substrate with green moulds such a Trichoderma and
Aspergillus species. To reduce this risk, we subjected substrates 1 till 5 to fermentation at
elevated temperatures before inoculating the mushroom strains into the substrate.
Fermentation was performed in aerated boxes in an acclimatised room at an air
temperature of 43-45oC for a period of about 11 days. If substrates get warmed up to this
V
temperature, usually a thermophilic population starts to grow which heats up the substrate
to even higher temperatures. In our experiment the substrates only slightly elevated
temperatures were recorded. This indicates that the thermophilic population had difficulty
to grow. Likely, the low nitrogen content of this type of waste materials are the main cause
of low activities. Nevertheless, an extended period of elevated temperature will probably
kill many contaminating spores of fungal moulds. In households and small enterprises this
fermentation step can be done by “composting” waste on piles (or dikes) and leave it for 1
to 2 weeks with mixing the material every 2 to 4 days.
After fermentation, substrates were cooled to 24oC and inoculated with spawn of the Oyster
mushroom strains in a clean room. Strains were grown in plastic bags containing 2 kg of
inoculated substrate. After a 14 day period of spawn-run at 24oC, substrates were fully
colonised by the fungi. No infections with green moulds occurred. Mushroom production
was initiated by venting the room and lowering air temperature to 20oC. First mushrooms
started to appear 7 days after the onset of venting. Mushrooms were sampled for analysis
of dry weight, heavy metal content, presence of food pathogenic bacteria, mycotoxins,
pesticide residues and cyanide. These analyses are currently being performed.
6. Results
6.1 Tests on vegetative growth of fungal species/strains on cassava
waste.
The data are not complete since some strains need a very long time to colonize the
substrate and some still have not reached full colonisation after 44 days of colonization
time. These strains were excluded. Those strains that had reached full colonization of the
substrates are reported here and from these a selection was made for mushrooms growing
and upgrading waste into animal food. Colonization rate of substrate by a fungal strain is
well reflected by the linear growth rate of the strain on that substrate (figure 1).
Figure 2 shows the growth curve of L. edodes strain MES00052 on a mixture of cassava peels
and cassava stalks. In the period from 0 to 6 days the growth rate is slowly increasing. This
could be the adaptation phase in which the fungus needs to switch from feeding on malt
extract agar (in the inoculum points) to the plant material. However, this “lag” period can
also be due to a relatively poor contact between the agar plugs used for inoculation and the
Figure 1. Glass tubes filled with moistured and sterilized cassava waste and inoculated at the top with spawn of a fungal strain. The position of the growth front was measured every 3 days to estimate the colonization rate of each strain on different type of cassava wastes.
VI
rather uneven layer of plant material. After 6 days a long period with a nearly constant
growth rate can be seen. At the end of the incubation period, the growth rate seems to
diminish as the mycelium reaches the bottom of the tube. This may look like a depletion of
nutrients. However, it is more likely that during the long period of incubation some water
may have collected on the bottom of the tube. During the wetting of the substrate, excess
water (i.e. not taken up by the substrate and present in between the substrate particles) has
been pressed from the substrate as good as possible. However, small amounts of this excess
water may still have been present in the substrate when filling the tubes. After a long period
this water may have accumulated on the bottom of the tube. Such “free” water in the
substrate is known to retard fungal growth.
As the growth rates at the beginning and the end of the incubation period may not reflect
the true potential of the fungal strains, we have used only the linear part of the growth
curve for calculation of the growth rate (like is shown in Figure 3).
Results on growth of L. edodes strains on cassava based substrates are shown in Figure 4.
As can be seen for 7 out of 10 strains grow on all cassava waste types and the best growth is
shown on a 1:1 mixture of peels and sticks. Growth rate on a mixture varies from 2.6 to 3.5
mm/day at 24 oC.
Results on growth of P. ostreatus strains on cassava based substrates are shown in Figure 5.
The best growth of this species was on cassava sticks and growth rate varied between 6.5
and 7.1 mm/day. Growth rates on a mixture of peels and sticks was ca. 6.6 mm/day,
considerably faster than the L. edodes strains. For each combination of strain and substrate,
three replicates are present in the experiment. However, currently we do not have the data
for all replicates.
Figure 2. Growth curve of L. edodes strain MES00052 on a mixture of cassava peels and stalks.
Figure 3. Part of the growth curve used
for calculation of the linear growth rate.
VII
Results on growth of P. pulmonarius strains on cassava based substrates are shown in Figure
6. For this species growth is slowest on cassava peels, intermediate on a mixture of peels
and stalks and highest on cassava stalks, similar to growth rates of P. ostreatus. Growth of P.
eryngii strains on cassava based substrates are shown in Figure 7. Growth on cassava peels
was poor and only for 4 strains growth was recorded. Also this species showed a better
growth on sticks than on a mixture of sticks with peels.
Statistical analyses of the measurements are given in appendix I
Figure 5. Linear growth rate of P. ostreatus strains on cassava based substrates. As these are merely preliminary results, we can only show growth rate on a substrate consisting of a mix of cassava peels and stalks and a substrate based on only cassava stalks. Results on cassava peels only are not available yet. MES 14029 is a FIIRO strain.
0
1
2
3
4
5
6
7
8
MES 01997 MES 01998 MES 14029
P. ostreatus
Max
imu
m li
ne
ar g
row
th r
ate
(m
m/d
ay)
Cassava peels Cassava peels with 50% (w/w) stalks Cassava stalks
Substrate
Species Strain
Average of Linear growth rate (mm/day)
Figure 4. Linear growth rate of L. edodes strains on cassava based substrates. Growth on cassava sticks was very slow and discarded. Growth of strains that were very slow on peels and mixture of peels and sticks were also excluded.
0
0.5
1
1.5
2
2.5
3
3.5
MES 00001 MES 00052 MES 00061 MES 02008 MES 02015 MES 02016 MES 02052 MES 02089 MES 02090 MES 02110
L. edodes
Linear growth rate of Lentinula edodes (shiitake) on different cassava waste products
Cassava peels Cassava peels with 50% (w/w) stalks Cassava stalks
Substrate
Species Strain
Average of Linear growth rate (mm/day)
VIII
Figure 6. Linear growth rate of P. pulmonarius strains on cassava based substrates. MES 14030 is a FIIRO strain.
0
1
2
3
4
5
6
7
8
MES 03444 MES 03448 MES 11698 MES 14030 MES 14031
P. pulmonarius
Max
imu
m li
ne
ar g
row
th r
ate
(m
m/d
ay)
Cassava peels Cassava peels with 50% (w/w) stalks Cassava stalks
Substrate
Species Strain
Average of Linear growth rate (mm/day)
Figure 7. Linear growth rate of P. eryngii strains on cassava based substrates. As
these are merely preliminary results, we can only show growth rate on substrates consisting of a mix of cassava peels and stalks and a substrate based on cassava stalks only. Results on cassava peels only are largely not available yet.
0
1
2
3
4
5
6
MES 02001 MES 02073 MES 03467 MES 03757 MES 12060 MES 12920 MES 12929 MES 12936 MES 12939 MES 12961 MES 12970 MES 12978
P. eryngii
Max
imu
m li
ne
ar g
row
th r
ate
(m
m/d
ay)
Cassava peels Cassava peels with 50% (w/w) stalks Cassava stalks
Substrate
Species Strain
Average of Linear growth rate (mm/day)
IX
6.2 Effects of fungal growth on digestibility by rumen microflora
After the completion of the vegetative growth tests, a selection of the tubes were used to
test the digestibility by rumen microflora as described in Cone et al., 1996. Gas production
was compared to non-inoculated (autoclaved) cassava waste. No effects was expected for
cassava peels since this material contains much starch. This starch will be consumed by
fungi before they will degrade the more fibrous material. The digestibility was, therefore,
tested with strains of one species only, i.e. P. pulmonarius. The digestibility of untreated
peels was considerably since the presence of starch (figure 8). As expected, the gas
production and thus the digestibility decreased after incubation with P. pulmonarius strains.
For the mixture of peels and sticks (1:1) an increase in gas production was seen with some
strains, especially those from L. edodes. The highest effect, however, was seen with the
cassava sticks. Sticks produce only minor amounts of gas in the rumen model as expected.
After especially treatment with strains from P. eryngii the amount of gas increased
considerably, up to levels close to 65% of peels showing potentials of these fungi to improve
digestibility of woody materials.
0.0
50.0
100.0
150.0
200.0
250.0
300.0
0 0 0 2 2 4 6 6
MES 11698 MES 03444 MES 11698 MES 11698 MES 03444
Uninoculated autoclaved control P. pulmonarius P. pulmonarius P. pulmonarius P. pulmonarius P. pulmonarius
Cassava Peels
Figure 8. Gas production (ml/gram organic matter) in the rumen model by cassava peels with or
without pretreatment with strains of P. pulmonarius. The digits under each bar indicate the
number of weeks of fungal pretreatment.
Figure 9. Gas production in a rumen model of different cassava waste products after fungal
pretreatment. The first panel shows the gas production by peels after treatment with P.
ostreatus. Gas production decreases compared to untreated samples probably due to
consumption of starch by the fungus. The second graph shows the gas production of a mixture
of cassava peels and stalks (1:1) after treatment with 2 Pleurotus species. Numbers under the
bars indicate the number of weeks of incubation. The third graph in figure 9 shows the gas
production of stalks after petreatment with the 3 Pleurotus species. Here, a clear increase of gas
production is seen compared to the untreated samples.
XI
6.3 Mushroom production of selected strains on cassava waste
Two P. ostreatus and 2 P. pulmonarius strains were used to test mushroom yield on cassava
waste basted substrates. Mushrooms were harvested during a period of ca. one month (2
flushes for each strain). The production of the strain used by Dutch growers (SPOPPO) on
commercial substrate (wheat straw) did produced earlier (6 days after climate change to
induce fruiting) than all other treatments. The best production was seen by the commercial
strain used by Dutch growers on commercially available substrate (wheat straw). This is not
unexpected since the strain/substrate has been developed especially for cultivation in Dutch
systems (similar to what we have used here). Also, Dutch substrate is supplemented with
nitrogen and the cassava waste based substrates has been used either as they are or
supplemented with oak tree saw dust (low in nitrogen). Despite the absence of supplements
it is remarkable that yields on cassava waste only are reasonably good, varying from 6% up
to more than 15% (figure 8). The highest yield is seen on a 50:50 mixture of cassava peels
and sticks with a Brazilian P. ostreatus strain. If yields are compared for cassava based
substrates only (with or without oak saw dust) and averaged for all strains, a mixture of
peels and stalks (1:1) shows the best production (figure 9). It is surprising that the addition
of saw dust had no positive effect on the production. This might be either due to the low
quality of the saw dust or the inability of the fungi to utilize saw dust well without enough
nitrogen available.
Figure 8. Yield of oyster mushroom strains of different substrates expressed as Biological
Efficiency (fresh weight mushrooms/fresh weight substrate). The commercial wheat straw
substrate (substrate 6) has the highest yields and is supplemented with nitrogen, contrary to the
other cassava waste based substrates. Nevertheless, yields up to 15% can be reached.
XII
Substrate bags were weighted at the beginning and at the end of the cultivation. In this way
yields of mushrooms can be compared with the decrease in weight of the substrate. A very
clear correlation was seen (figure 10) indicating the good quality of the data.
0
50
100
150
200
250
300
Substrate 1 Substrate 2 Substrate 3 Substrate 4 Substrate 5
Gra
mm
es
mu
shro
om
s p
er
2 k
g su
bst
rate
Total yield in 2 flushe
substraat
Average of Totale opbrengst 2 vluchten (g)
Figure 9. Yield (gram mushrooms per bag=2 kg substrate) for substrates based on
cassava waste only. The mixture of peels and stalks gave the best production.
Figure 10. Correlation between mushroom yield and decrease of
substrate weight. A clear correlation is shown indicating that the
measurements were sound.
y = 0.0014x + 0.0218R² = 0.8329
0
0.2
0.4
0.6
0.8
1
1.2
0 100 200 300 400 500 600 700 800De
cre
ase
su
bst
rate
we
igh
t (k
g)
Yield mushrooms (grammes)
Correlation between yield and decrease substrate weight
XIII
7. Conclusions
The main conclusions of this research are:
At least 4 different fungal species used to produce edible fungi grow well
vegetatively on cassava waste (peels, stalks and a mixture of both (1:1)). Comparison
of growth rate for the 4 species tested
o L. edodes: Mixture of peels & stalks> peels or stalks alone
o P. ostreatus: Stalks> mixture> peels
o P. pulmonarius: Stalks> mixture> peels
o P. eryngii: Stalks> mixture> peels
Pretreatment of especially stalks with P. eryngii increases the digestibility for rumen
flora considerably.
The production of grey oyster mushrooms (P. ostreatus) on substrate consisting of
cassava waste only is good (up to 15% fresh weight mushrooms/fresh weight
substrate).
Experiments are started now to test yields on cassava waste supplemented with
nitrogen rich waste. We expects that addition (like bran) can increase yields up to
levels of commercial production.
XIV
8. References
Cone, J.W., van Gelder, A.H., Visscher, G.J.W., Oudshoorn, L., 1996. Influence of rumen fluid
and substrate concentration on fermentation kinetics measured with a fully automated time
related gas production apparatus. Anim. Feed Sci. Technol. 61, 113–128.
Aro S O, Aletor V A, Tewe O O and Agbede J O 2010: Nutritional potentials of cassava tuber
wastes: A case study of a cassava starch processing factory in south-western Nigeria.
Livestock Research for Rural Development. Volume 22, Article #213. Retrieved December 6,
2013, from http://www.lrrd.org/lrrd22/11/aro22213.htm
Annes 1 Pictures of mushroom production
Annex 2 Statistical analysis of yield data
Program used; Genstat, 16th edition.
Raw data
Strain Substrate Tray Yield (g) Strain Substrate Tray Yield (g)
MES 01997 Substrate 1 25 247 MES 03444 Substrate 1 6 193
MES 01997 Substrate 1 36 284 MES 03444 Substrate 1 40 90
XV
MES 01997 Substrate 1 73 268 MES 03444 Substrate 1 56 140
MES 01997 Substrate 1 78 261 MES 03444 Substrate 1 82 290
MES 01997 Substrate 2 18 336 MES 03444 Substrate 2 9 152
MES 01997 Substrate 2 47 341 MES 03444 Substrate 2 28 70
MES 01997 Substrate 2 67 308 MES 03444 Substrate 2 59 140
MES 01997 Substrate 2 87 254 MES 03444 Substrate 2 88 168
MES 01997 Substrate 3 24 313 MES 03444 Substrate 3 16 150
MES 01997 Substrate 3 38 321 MES 03444 Substrate 3 48 150
MES 01997 Substrate 3 75 324 MES 03444 Substrate 3 74 130
MES 01997 Substrate 3 99 282 MES 03444 Substrate 3 93 115
MES 01997 Substrate 4 1 268 MES 03444 Substrate 4 5 114
MES 01997 Substrate 4 31 307 MES 03444 Substrate 4 30 145
MES 01997 Substrate 4 60 282 MES 03444 Substrate 4 52 118
MES 01997 Substrate 4 97 271 MES 03444 Substrate 4 98 111
MES 01997 Substrate 5 14 300 MES 03444 Substrate 5 2 93
MES 01997 Substrate 5 43 254 MES 03444 Substrate 5 27 146
MES 01997 Substrate 5 66 272 MES 03444 Substrate 5 61 0
MES 01997 Substrate 5 85 172 MES 03444 Substrate 5 79 59
MES 01997 Substrate 6 19 481 MES 03444 Substrate 6 8 441
MES 01997 Substrate 6 44 676 MES 03444 Substrate 6 41 390
MES 01997 Substrate 6 71 589 MES 03444 Substrate 6 65 414
MES 01997 Substrate 6 77 542 MES 03444 Substrate 6 94 384
MES 01998 Substrate 1 22 273 MES 03448 Substrate 1 11 298
MES 01998 Substrate 1 34 212 MES 03448 Substrate 1 29 242
MES 01998 Substrate 1 58 162 MES 03448 Substrate 1 63 330
XVI
MES 01998 Substrate 1 89 185 MES 03448 Substrate 1 83 187
MES 01998 Substrate 2 15 253 MES 03448 Substrate 2 12 333
MES 01998 Substrate 2 33 209 MES 03448 Substrate 2 49 305
MES 01998 Substrate 2 54 190 MES 03448 Substrate 2 51 319
MES 01998 Substrate 2 81 166 MES 03448 Substrate 2 91 334
MES 01998 Substrate 3 17 227 MES 03448 Substrate 3 13 309
MES 01998 Substrate 3 50 163 MES 03448 Substrate 3 39 306
MES 01998 Substrate 3 53 39 MES 03448 Substrate 3 55 185
MES 01998 Substrate 3 86 171 MES 03448 Substrate 3 100 231
MES 01998 Substrate 4 7 147 MES 03448 Substrate 4 3 235
MES 01998 Substrate 4 32 232 MES 03448 Substrate 4 35 231
MES 01998 Substrate 4 69 183 MES 03448 Substrate 4 62 254
MES 01998 Substrate 4 84 95 MES 03448 Substrate 4 92 214
MES 01998 Substrate 5 4 135 MES 03448 Substrate 5 21 180
MES 01998 Substrate 5 46 82 MES 03448 Substrate 5 45 177
MES 01998 Substrate 5 72 170 MES 03448 Substrate 5 68 170
MES 01998 Substrate 5 96 139 MES 03448 Substrate 5 90 176
MES 01998 Substrate 6 23 493 MES 03448 Substrate 6 10 700
MES 01998 Substrate 6 37 561 MES 03448 Substrate 6 42 636
MES 01998 Substrate 6 64 493 MES 03448 Substrate 6 57 644
MES 01998 Substrate 6 80 371 MES 03448 Substrate 6 95 471
Analysis of variance.
Variate: Yield_g
Source of d.f. s.s. m.s. v.r. F pr.
XVII
variation
Tray stratum
Strain 3 390814 130271 47.98 <.001
Substrate 5 1342896 268579 98.92 <.001
Strain.Substrate 15 43355 2890 1.06 0.404
Residual 72 195492 2715
Total 95 1972558
Information summary
All terms orthogonal, none aliased.
Message: the following units have large residuals.
Tray 82 111.8 s.e. 45.1
Tray 95 -141.8 s.e. 45.1
Tables of means
Variate: Yield_g
Grand mean 260.1
Strain MES 01997 MES 01998 MES 03444 MES 03448
331.4 223.0 175.1 311.1
Substrate Substrate
1
Substrate
2
Substrate
3
Substrate
4
Substrate
5
Substrate
6
228.9 242.4 213.5 200.4 157.8 517.9
Substrate
Strain Substrate Substrate Substrate Substrate Substrate Substrate
XVIII
1 2 3 4 5 6
MES
01997
265.0 309.7 310.0 282.0 249.5 572.0
MES
01998
208.0 204.5 150.0 164.2 131.5 479.5
MES
03444
178.2 132.5 136.2 122.0 74.5 407.2
MES
03448
264.2 322.8 257.8 233.5 175.8 612.8
Standard errors of differences of means
Table Strain Substrate Strain x Substrate
rep 24 16 4
d.f. 72 72 72
s.e.d. 15.04 18.42 36.85
Least significant differences of means (5% level)
Table Strain Substrate Strain x Substrate
rep 24 16 4
d.f. 72 72 72
l.s.d. 29.99 36.73 73.45
Strain Substrate
Average yield (g)/2 kg of
substrate
(wet weight/wet weight)
l.s.d. is 73.45 at P=0.05
XIX
MES 03444 Substrate 5 75 a
MES 03444 Substrate 4 122 a b
MES 01998 Substrate 5 132 a b c
MES 03444 Substrate 2 133 a b c
MES 03444 Substrate 3 136 a b c d
MES 01998 Substrate 3 150
b c d e
MES 01998 Substrate 4 164
b c d e f
MES 03448 Substrate 5 176
b c d e f
MES 03444 Substrate 1 178
b c d e f g
MES 01998 Substrate 2 205
c d e f g h
MES 01998 Substrate 1 208
e f g h
MES 03448 Substrate 4 234
f g h i
MES 01997 Substrate 5 250
g h i j
MES 03448 Substrate 3 258
h i j
MES 03448 Substrate 1 264
h i j
MES 01997 Substrate 1 265
h i j
MES 01997 Substrate 4 282
i j
MES 01997 Substrate 2 310
j
MES 01997 Substrate 3 310
j
MES 03448 Substrate 2 323
j
MES 03444 Substrate 6 407
k
MES 01998 Substrate 6 480
k
MES 01997 Substrate 6 572
l
MES 03448 Substrate 6 613
l
Values sharing the same letter show not statistically significant difference at p=0.05