Last week Professor Hibbett, and two undergraduate research assistants (myself and Sam Kovaka), visited the site. It was a beautiful sunny day! We met Gene Bernat there, and he gave us a tour of the greenhouse. He updated us on the status of the project. We have a lot to do! Soon we will inoculate the mycelium beds in the greenhouse. We found mushrooms growing at the banks of the canal where the previous wood chips were laid out. We also spotted some turtles swimming in the canal! – Vanessa June 15, 2013 Plans After our meeting at Clark; Gene, Jim Rice, Brian Seitzman, myself and David set goals and plans for the project. Our biggest challenge was that of deciding on a name. An important goal for the project was to establish it as a teaching platform for education and outreach. We heard about Jim’s passive sampler study and his plans to start this summer. Jim told all of us about the time frame he had planned for the testing, deployment of 10-12 samplers in July, and the analysis in August. Brian also told us about his project,using the site as a business model for economic sustainability. He wishes to see the project as a path to generate income as well and integration into the local economy. Inoculating June 25, 2013 David and I moved forward with our plans for the site to reinoculate the mycoreactors. Our first step was to culture the sample we got from Paul Stamets through Gene. I made some agar plates and cultured the three species (Irpex, T.V.G, and Pleurotus). We made some media out of rye grain in small mason jars and milk jugs. We autoclaved them and then I inoculated three jars with different volumes of water (20 ml, 30 ml, 40 ml) and 1 milk jug for each species. We inoculated one extra jug for pleurotus so that we could grow some oyster mushrooms for food on coffee grounds from Acoustic Java (a local coffee shop near campus). The mycelium has been growing in the plates, jars, and jugs. Everything is going well! – Vanessa

More Plans 06/27/2013 Today Professor Hibbett and I developed more concrete plans for the Fisherville mycoreactors. We shook the mycelium jugs and jars (this was after a week) some of them, mostly the Irpex, had really taken over the rye grain and shaking them loose was hard. We realized there was a lot that needed to be done, quickly!

• subculturing the pleurotus Professor Hibbett brought on to new plates
• Autoclaving bags on Monday with rye grain and coffee grounds (from Acoustic Java).
• Inoculation on Tuesday

We also realized the water volume in some of the jars was too dry so we need to increase it. – Vanessa Subculturing and Increasing Water Volumes 06/28/2013 Today was the start of our new plans! I autoclaved some water and then added it to the jars of Pleurotus and T. R.G. I added 10ml to each jar. I did this under hood using sterile technique. I also subcultured the Whole Foods Pleurotus onto 4 new MEA plates. – Vanessa

Some Inoculation 07/01/2013 Today I filled 8 bags with rye grain and 5 with coffee grounds. For the rye the ratio was 1L of rye and 900 ml of water per bag. For the coffee the ratio was 2L of coffee grounds and 250ml of water per bag. One bag had 1L and no water added, and another bag had 2L and no water added either. I autoclaved these bags and left them under the hood to cool overnight. – Vanessa The Big Inoculation Day 07/02/2013 Today we inoculated 20 bags of rye with the Irpex, Pleurotus, and Trametes culture we had growing in jugs and jars. The ratio remained 1L of rye to 900 ml of water. It was a very big day because I could only autoclave 12 bags at a time, before they were ready (sterile and cool enough) to inoculate. I used the bag sealer that Gene sent us to seal them. unfortunately 2 bags ripped in the autoclave and I had to throw them out. Nevertheless, the day was a success! -Vanessa The Bags 07/17/2013 After being away for some time I checked on the bags and they’re doing great! It was really impressive to see the mycelium really take over the rye in the bags. One of the bags got some bacteria in it so we need to throw it out. Unfortunately the Pleurotus in the coffee grounds didn’t do as well as the rye.

– Vanessa Subculturing 07/17/2013 We received a package from Fungi Perfecti with Stropharia. I subcultured it onto new MEA plates. I also did a tissue explant onto plates with another Pleurotus strain Professor Hibbett brought. -Vanessa Inoculating 07/22/2013 Today I inoculated 9 bags of rye, each with 1L of rye and 900 ml of water, with the Pleurotus from Fungi Perfecti that we had in a jug. The Stropharia culture on the plates is growing!

-Vanessa  More Inoculating 07/28/2013 Today I inoculated six jars of rye (30g of rye and 40 ml of water) with the previous cultures of pleurotus ostreatus strain. – Vanessa Passive Samplers 07/31/2013 Jim Rice sent us information on what he did in the canal with the passive samplers. Here is a brief description he wrote for us.

08/02/2013 Today I inoculated 8 bags with the new stropharia culture we got from Fungi Perfecti. I also Inoculated 12 bags with the Pleurotus Eryngii cultures.

-Vanessa The Big Day 08/07/2013 Today was the big day! Professor Hibbett and I went to the Fisherville site. We took the 23 bags of mycelium inoculated with Irpex, Trametes, and Pleurotus. We joined Gene, his son Nick, and Max from John Todd Ecological Designs. First we cleaned the 23 bins with water. We added about 20 lbs of hard wood pellets (an oak and maple blend). Next we added about four gallons of water to each bin and let the pellets absorbs the water. It was amazing to see the pellets grow with water! We labeled the bins with a labeler that Nick had with the names of each of the fungi. We then gloved up and inoculated the bins with the bags of mycelium. We had to be careful and break apart the mycelium on the rye. It was a really interesting texture: slimy and but very cohesive. We spread out the mycelium in the wood chips as thoroughly as we could.We then carried the bins inside the trailer with air-conditioning and covered them up. The bins are now ready to be placed in the eco-machine.