*2.5. Application of Post-Growth Treatments*

Grown materials received were either dried, baked, compacted then dried, or compacted then baked. Drying myceliated materials dehydrates them to make the fungus dormant and stop its growth. If the water content of the materials is increased after the drying process, the fungus is usually able to resume growth [5]. Myceliated materials were dried in an oven for a total of 12 h at 40 ◦C. Due to the restrictions of the COVID-19 regulations, they were dried in 2 sessions of 6 h each and kept in the oven between these sessions.

The baking process is supposed to kill the fungus. In comparison with the drying process, if the water content of the material is increased after the baking process, no fungal growth should be observed. They were placed in an oven for 6 h at 100 ◦C to bake.

Half of the specimens were compacted to half of their thickness with an industrial hydraulic press (20 TON shop press from Central Machinery, Moses Lake, WA, USA) between metal plates. Further details on the application of post-growth treatments are explained in Appendix G.

## *2.6. Mechanical Testing Procedure*

### 2.6.1. Compression Testing

Due to the lack of testing standards to assess the uniaxial compressive properties of myceliated materials, various ASTM standards specific to soil (i.e., ASTM D2166/D2166M-13), wood-based panels (i.e., ASTM D3501 and ASTM D1037), and thermal insulations (i.e., ASTM C165-07) have been used by research teams [8,9,11,42,55–57]. Elsacker et al., 2019, followed ASTM D3501, whereas their use of cylindrical specimens followed ASTM D2166/D2166M-13 [9]. This latter standard specifies that the specimen should have a height:diameter ratio ranging from 2:1 to 2.5:1. In their study, Elsacker et al. opted for a 0.5:1 height:diameter ratio with specimens measuring 3.75 cm in height (or 10 cm in height for pre-compressed specimens) for 7.5 cm in diameter [9]. Yang et al., 2017, followed ASTM D2166/D2166M-13, while growing their specimens to a height of 6 cm and a diameter of 5 cm (i.e., 1.2:1 height:diameter ratio) [8]. To facilitate data comparison and reduce the effect emerging from corners in rectangular specimens, ASTM D2166/D2166M-13 was used [55]. Our cylindrical specimens were grown to attain a 2:1 height:diameter ratio after post-growth treatments, as specified by the standard. However, due to the large amount of contamination, the height had to be reduced to 11 cm before the application of post-growth treatments. Therefore, specimens were tested with a height of 11 cm for the uncompacted specimens and 5.5 cm for the compacted specimens. Since the diameter could not be reduced easily, the diameters remained the same (i.e., 7 cm), resulting in a 1.57:1 height:diameter ratio.

The uniaxial compressive tests were performed at room temperature at the University of Akron's Olson mechanical testing facility. The testing room had a temperature of 20.06 ± 0.39 ◦C and a relative humidity of 62.6 ± 1.36%. Due to the high humidity, the specimens were stored in the Biodesign lab, which had a temperature of 25.53 ± 0.20 ◦C and a relative humidity of 46.14 ± 0.64%. Previous research projects also performed mechanical tests on myceliated materials under ambient conditions: 23 ◦C [7], and 25◦C for a 50% relative humidity [9]. A 5567 Instron with a loading cell of 10 kN was used for the compressive tests. Before starting the test, the loading cell was lowered onto the specimens until a load of 1 N was reached. ASTM 2166-13 requires a loading rate that will produce a strain of 0.5 to 2%/min. Yang et al., 2017 used a 2%/min loading rate and our team decided to follow the same rate [8]. The standard also specifies that the specimen should fail within 15 min, but our specimens did not fail. Elsacker et al., 2019 stopped their compressive tests when the strain produced a height deformation between 70 to 80% strain [9]. In this regard, our tests were stopped when the specimen experienced 75% deformation (i.e., strain of 0.75) or the maximum load of 10 kN was reached. The specimens' dimensions (i.e., height, diameter, and weight) were measured according to the ASTM C303-10 to determine specimens' density before the first test [58].

Since the mycelium-based specimens did not fail under the compressive load, the team decided to reload the specimens grown from *Pleurotus ostreatus* to study the hysteresis and dilatation of mycelium-based materials. Therefore, the specimens were first loaded up to a 75% height deformation (i.e., strain of 0.75) or a load of 10 kN. Once one of these limits was reached, the load was released. Specimens were left to dilate for 10 min. After 9 min of being released, the specimens' height and diameter were measured. After 10 min of being released, the specimen was loaded a second time with an updated loading rate. The same formula was used to calculate the rate of testing (i.e., 2%/min). The height of the specimen changed between the start of each test. Thus, the rate was recalculated with the updated height for the second test. The height of all specimens was taken before and after both tests, 1 h after the end of the second test, and after 1 week of testing to study the dilatation over time. The width of the specimens was also recorded but was not useable due to the pieces falling off from the sides. The weight of the specimens was not recorded after the beginning of the test as specimens broke into multiple pieces every time they were handled.

Different methods can be used to measure Young's modulus. The slope of the linear portion of the stress–strain curve is commonly used to calculate Young's modulus. Our materials did not fail, and the stress–strain curves did not possess linear portions. Elsacker et al., 2019, seemed to have used the entire stress–strain curve to calculate Youngs' modulus [9]. Since our specimens did not fail, the resistance to the load started an exponential trend towards the middle of the test (i.e., densification). Therefore, extracting Young's modulus from the entire deformation of the specimen seems to be incorrect. Two studies obtained Young's moduli of myceliated specimens at 20% height deformation (i.e., strain of 0.20) [10,11], so this technique was chosen. Furthermore, the top and bottom surfaces of the myceliated specimens were not entirely flat, but highly textured. Therefore, the data collected at the beginning of the tests (up to around a strain of 0.05) only showed the compressive resistance of parts of the specimens. Calculating Young's modulus at a strain of 0.2 (i.e., 20% height deformation) allowed the removal of all this variability in the data. To facilitate comparison between studies, Young's modulus was still calculated with three different techniques used in the literature. First, it was calculated from the slope of the stress–strain curve between strain values of 0.19 and 0.20. This technique shows Young's modulus at a strain of 0.20. Second, it was calculated from the slope from the start of the test up to a strain of 0.20. Third, it was calculated from the stress–strain slope of the entire specimen deformation. Additional information about the calculation of Young's modulus is presented in Appendix H.

### 2.6.2. Three-Point Bending Testing

The bending testing followed the procedure from ASTM D1037-12: Standard Test Methods for Evaluating Properties of Wood-base Fiber and Particle Panel Materials, Section 9. Static Bending [54]. Since each mold contained six specimens grown as one entity, each specimen was cut to the desired size (i.e., 76 mm) prior to testing.

The bending tests were performed with the same Instron that was used for compression tests. A 5567 Instron with a loading cell of 100 N was used for the bending tests. A specific testing apparatus was built according to the ASTM standard. Rounded supports had a span of 266 mm, which is equal to 24 times the average nominal thickness of the panels. Before starting the test, the upper point was lowered onto the specimens until a load of 0.01 N was reached. ASTM D1037-12 requires a uniform loading rate to achieve an outer fiber strain rate of 0.005 mm/mm/min. Since each specimen had a slightly different thickness, each specimen was tested with a different loading rate. As a reference, the average loading rate of all specimens tested in bending for the first batch was 5.043 mm/min. Tests were stopped once the specimen failed, or the load applied reached less than 20% of the maximal load achieved during the test.

The specimens' dimensions were measured according to the ASTM C303-10 to determine specimens' density before testing [58]. Some specimens broke asymmetrically (i.e., they did not fracture at the location of the upper point pushing down onto the specimen or the middle of the specimen) and/or the fracture was not parallel to the apparatus supports. After the test, the fracture location and angle were measured to study asymmetric failure (Figure A1). Additional information about the bending test data analysis is provided in Appendix H.

### **3. Results**
