GrowCast: Sustainable, Stronger, and Affordable Patient-Specific 3D-Printed Mycelium Casts for Orthopedic Care

1. Comments to Author:

This is a very interesting manuscript. I enjoyed reading it. Your study demonstrates the application of mycelium in a custom 3D-printed cast to enhance the mechanical properties and biocompatibility of an orthopedic cast. The hypothesis is well thought out, and the experiments performed support your hypothesis. To improve your manuscript, I suggest making the following changes to the introduction and adding clarifications to the methods sections, which will help others replicate your experiment and achieve repeatable results. The introduction is one of the most important sections, as it sets the tone for the 'why,' i.e., why you are trying to solve a problem. The manuscript could also benefit from additional images, such as a comparison of the gypsum cast with the 3D-printed cast, an image of the cast during testing, and so on. There are some holes in the methods section that can cause confusion to the reader. I have suggested some recommended changes that might help you improve the article.

1. Summary:

This manuscript presents the authors' hypothesis and demonstrates that a mycelium-infused 3D printed cast outperforms a traditional gypsum cast, achieving a significantly higher compressive force. The authors 3D-scanned a forearm and then created a custom Voronoi lattice pattern using 3D CAD software. The authors then packed the cultivated mycelium into the 3D-printed cast and tested its mechanical properties, comparing them to those of a typical gypsum cast. The authors demonstrated that there is a significant increase in the compressive as well as stiffness of the mycelium-infused 3D printed cast.

1. Scientific Changes Recommended:

Introduction –

· L71: Please check the work by D’Amado et al. It can be added as another citation to support your statement (https://doi.org/10.1016/j.stlm.2024.100146). 

 · L72: You do a good job of explaining some advantages of mycelium. Suggest adding some references or work done by others here, explaining how they have incorporated mycelium into their work, and briefly explaining their observation, i.e., whether it helped or not. This will help inform the reader of the current trends.

 · L78: Could use some references here.

 Results –

· L97: Please add an image of the compression test setup. Check similar work by D’Amado et al., for reference (https://doi.org/10.1016/j.stlm.2024.100146).  Please note how they clearly describe their test setup with a test article inside the machine. This is crucial to understanding how you tested your cast and will be helpful for others who want to reproduce your results.

 · L106: Please clarify how the displacement was monitored. Did you use an extensometer or a strain gauge?

 · L108-L109: You specify that 3 different sizes were fabricated to evaluate the scaling effects, but you did not individually specify the average compressive forces of specific sizes. Only one average is mentioned. Is this the average compressive force of all 3 sizes combined or of a specific size? Could you please clarify whether you noticed a significant difference in compressive force for different sizes? If you did, then please report the averages of each size. If you do see a difference, then consider normalizing the data. That can help clean the data.

 Discussion –

· L126-L129: Can you please expand on this, i.e., why does mycelium infusion help improve the mechanical properties? Have similar observations been made by others? The reasoning here is based on experimental observation but lacks a thorough discussion. This is where digging a bit deeper into the WHY is important. It will be helpful to state how mycelium helps. This can be done by doing a brief literature review here. Please check these references as examples (https://prezi.com/p/b5bzqdlivbp3/mycoplaster-orthopedic-casts-with-biomimetic-mycelium/; https://doi.org/10.1016/j.matdes.2023.112592).

 Materials and Methods –

· L157: Why did you choose a Voronoi pattern? What was the reason behind it? Did you try modeling or testing any other lattice structures? Please justify this. One way to do this is, for example, comparing the work done by others, examining different lattice shapes, and highlighting the advantages of the Voronoi pattern.

 · L161: Just to clarify, all three sizes were fabricated on different machines? Why was this done? Although probably not an issue for your experiment, but just for your reference, there are noticeable differences between machines to machine even when printing using the same material. As each machine and manufacturer has their own parameter set, they differ enough to cause irregularities.

 · L161: Please specify the direction of print for all, i.e., whether all were printed in the same orientation or different orientations?

 · L179: Can you please specify what the loading direction was with respect to the printing direction? What I mean is, was the compressive test performed parallel or perpendicular to the material extruded direction? Please check the resource mentioned here (https://akademiabaru.com/submit/index.php/mjcsm/article/view/4528/3363). As you can see from the references, the direction of the printing has a significant impact on the results. Please clarify this in the manuscript. If possible, it might be worthwhile to test in different printing orientations to compare printing orientation and load-bearing properties.

1. Presentation Changes Recommended:

Summary –

· L5-7: Think about the sequence of events and reorganize it. i.e, first is scan, then design, followed by printing, then testing

 Introduction –

· L22: Please reword this sentence. One suggestion is to add ‘and was previously used in..’. This will help the sentence read better.

· L28-L34: You should think about moving this paragraph towards the end of the introduction section, i.e., right before you start with your results. This will help improve the structure. Also, it will help you set the tone. Think about it this way, first you explain WHAT the issue is and WHY it is an issue, and towards the end you are touching upon HOW, i.e., how you propose to solve this problem. This structure will help you with a broad 'what' statement and then narrow it down using 'why', with 'how' being the final tip of the pyramid.

· L53: Instead of ‘mesh cast has large openings’, you should think about rewording it to something like ‘mech can have large openings. As you rightly pointed out in L51, a 3D printed cast can be customized to suit the patient's specific needs. Based on the requirement, you may or may not have large openings. Instead, you can have a bunch of smaller openings that can still allow sufficient air circulation.

· L55: You should expand on abbreviations, such as what PLA and ABS stand for. This will help the reader understand what PLA stands for. Once you expand on the abbreviation here, you can use it everywhere else.

· L56: You mention PLA and biodegradable filaments separately, while under Materials for 3D printed cast you mention PLA as biodegradable. One suggestion can be to reword the sentence to read something like ‘Materials like Acrylonitrile butadiene styrene (ABS) and biodegradable polymers such as Polylactic Acid (PLA) allow for….’

· L59: I think this line is not needed, as you are comparing a 3D printed cast with casts made from gypsum. These gypsum casts are not injection molded. This sentence is unnecessary and may confuse the reader regarding the paper's scope.

· L67: Please expand upon PCL and PHA

· L85: Can you expand on some long-term cost benefits? For example, a rough cost estimate of the number of parts vs the break-even point? It is not necessary, but it will help strengthen this section.

Results –

· L108: Please specify compressive force instead of just force

· L108-L109: Please reference the figures

· L116: What were they? Please specify here

Discussion –

· N/A

Materials and Methods –

· L157: Please specify other types of lattice structures that are available. For example, beam lattice, gyroid, honeycomb, and Voronoi (https://www.ntop.com/resources/blog/guide-to-lattice-structures-in-additive-manufacturing/).

· L176: Please mention the entire abbreviation of RH as relative humidity.

Acknowledgement-

· L191: While not necessary, thanking and acknowledging Dr. Bhamla and his lab for printing the transparent prototype will be nice and  kind gesture

1. Figure Changes Recommended:

Results –

· L103: Please add an image of the quarter, half, and full-scale models’ side by side next to each other and compare that to the gypsum cast. Check similar work by D’Amado et al., for reference (https://doi.org/10.1016/j.stlm.2024.100146).

Materials and Methods -

· L162-L164: Please add a picture of the quarter, half, and full-scale model for reference and compare it to the cast model for ease of understanding. Check similar work by D’Amado et al., for reference (https://doi.org/10.1016/j.stlm.2024.100146).

· L174: A picture of pre- and post-mycelium application into the 3D printed cast. It would be nice to include this here; alternatively, you can refer to the image here if you plan to display the post mycelium application elsewhere. This will help the reader visualize how the mycelium was bonded with the cast.

Figures and Tables -

· Please merge and center the cells for the heading to make it clear