Progress 09/01/24 to 08/31/25

Outputs
Target Audience:Students, farmers, researchers in mushroom production, organic agricultural farmers, and people in the animal and poultry science field are targeted currently. Next year, we will present the results from these studies to agriculture and poultry farmers during Agriculture Field Days at the respective universities, as well as scientists and researchers at academic conferences. Changes/Problems:-No problems or changes are necessary. None. What opportunities for training and professional development has the project provided?As part of this project, a dedicated research assistant professor has been recruited at the UH to oversee research activities and mentor student participants. Additionally, one graduate student has been appointed at each partnering institution--UH, PVAMU, and UGA--to carry out research tasks related to mushroom stem processing and poultry nutrition studies. Several undergraduate students at UH have also received hands-on training in mushroom collection, handling, and processing techniques, contributing to various stages of the project such as cleaning, drying, and powder preparation. At PVAMU, two postdoctoral associates have been actively engaged in both laboratory and field-based research. Their responsibilities have included washing, cleaning, and processing mushroom stems, as well as conducting poultry trials involving layer chicks and broilers, in alignment with the project's objectives. Ongoing research at PVAMU focuses on evaluating the inclusion of BMS powder in layer diets. This work involves collaborative efforts among graduate students, postdoctoral researchers, and faculty and provides valuable data on the performance, health, and environmental impact of using BMS as a partial replacement for soybean meal. Through this multi-institutional effort, the project continues to promote experiential learning, research capacity building, and workforce development in the areas of sustainable agriculture and animal nutrition. How have the results been disseminated to communities of interest?To date, the project has resulted in one published review article titled "Harnessing Mushrooms for Poultry Nutrition," which highlightsthe potential of mushroom byproducts as sustainable feed ingredients. In addition, two research manuscripts based on the use of brown and white mushroom stems in layer chick diets have been accepted/are in press at Poultry Science and Frontiers in Veterinary Science, respectively. Dissemination efforts have also included student-led presentations at national scientific meetings. A master's student from PVAMU delivered an oral presentation on BMS powder supplementation in layer chick diets at the American Society of Animal Science (ASAS) 2025 Southern Section Joint Meeting, held in Arlington, Texas, on April 6, 2025. At the same event, an undergraduate student from PVAMU presented findings from the evaluation of white mushroom stems as a sustainable alternative to soybean meal in poultry nutrition during the Undergraduate Student Oral Competition. Additionally, a hands-on workshop was organized at the Poultry Center at PVAMU to train farmers on mushroom cultivation, stem collection and preparation, and their incorporation into poultry feed. These outreach and dissemination activities have strengthened academic engagement, promoted farmer education, and increased awareness of sustainable alternatives in poultry production systems. What do you plan to do during the next reporting period to accomplish the goals?We plan to finalize the design and begin testing of the complete mushroom stem processing system, which includes the cutting machine, root detection algorithm, and integration of robotic components. Alongside this, we will complete the techno-economic analysis of the processing workflow, optimizing key parameters to improve operational efficiency, reduce production costs, and ensure consistent product quality for potential commercial-scale implementation. In collaboration with the PVAMU team, we will continue formulating poultry feed containing DMS and conducting feeding trials to generate robust performance and health data. We will also work closely with the UGA team to analyze the results from the broiler and challenge studies, making any necessary adjustments to DMS processing or feed formulations based on those findings. Additionally, we aim to complete and submit a technical manuscript detailing the compositional and nutritional analysis of DMS for publication in a peer-reviewed journal. Efforts are also underway to explore partnerships with commercial mushroom producers to scale up DMS production and expand its reach as a sustainable feed ingredient. To support community engagement and education, we plan to organize additional farmer training sessions through extension agents at PVAMU and UGA. These sessions will focus on sustainable mushroom cultivation and the use of mushroom stems in poultry feed, while also providing students with valuable opportunities to interact with industry professionals and gain practical agricultural knowledge. Moreover, we are actively exploring opportunities for students to participate in farm visits and workshops, further enriching their hands-on experience in the agriculture and animal science sectors.

Impacts
What was accomplished under these goals? For major goal 1: Develop a process to produce dried mushroom stems (DMS) as a feed ingredient. Milestone 1.1: Develop a process to collect and prepare mushroom stems for further processing. We have developed and refined a process for upcycling mushroom stems, focusing on efficiently separating the stem from the root for further use as poultry feed. The initial step involves cleaning the mushrooms and removing stems from the roots. In our first prototype, we used a bowl feeder to load mushroom legs into a designated zone. A camera, paired with an image processing algorithm, helped locate each mushroom, allowing a linear actuator to position it under a guillotine-style cutter that separated the leg into stem and root. These components were then collected in separate containers. While the design met the technical goals, its processing speed was too low for commercial use, especially considering the large quantity of stems needed for feed trials and future scaling. To enhance productivity, we explored commercially available equipment. We acquired a small vegetable cleaning machine through vendor negotiations, which significantly reduced the time and labor needed for cleaning--previously a manual step. Additionally, we tested a commercial mushroom leg cutting machine with a rotary conveyor and automated loading system. Drawing inspiration from these trials, we began developing a redesigned mushroom leg cutting machine with improved functionality and productivity. The major components include a full mushroom loader, conveyor belt, precision cutting blades, and a robotic system featuring either a 6DOF robotic arm or dual arms. The operation proceeds as follows: a robotic arm or human operator loads either a full mushroom or leg onto the system. If a full mushroom is used, the cap is removed and collected, and the leg proceeds along the conveyor. As the leg moves, a camera monitors its orientation, and a robot positions it accurately for cutting using an image processing algorithm. At the end of the belt, the cutting blade separates the leg into stem and root, which are sorted into different containers. Several algorithms support the machine's function, including those for mushroom identification, cap handling, cutting line detection, and precise actuation. We submitted the design for the newly developed mushroom leg cutting machine to the UH Intellectual Property (IP) Office in June 2025 for consideration as a provisional patent. Milestone 1.2 Develop methods to freeze-dry and pulverize DMS into a fine powder for use in chicken feed formulation. We have successfully produced and stockpiled substantial quantities of DMS powder to support a wide range of animal feeding trials and research activities. To date, a total of 66 kg of dried mushroom powder--comprising both brown and white varieties--has been processed and prepared over the course of the project. Since the yield of dried powder is approximately 10% of the fresh weight, this production corresponds to the processing of over 660 kg of fresh mushroom stems sourced from farms, underscoring the significant scale and operational efficiency of our upcycling process. Milestone 1.3 Determine the chemical composition and nutritional value of DMS. Comprehensive analyses have been performed on DMS samples to evaluate their nutritional and safety profiles. These include proximate analysis (measuring moisture, protein, carbohydrates, fiber, lipids, and ash), amino acid profiling, vitamin content analysis, and heavy metal screening. For major goals 2 and 3: We conducted 3controlled feeding trials and initiated a 4thstudy at PVAMU. Experiment 5 was conducted at UGA. Experiment 1 focused on assessing the impact of brown mushroom stems (BMS) powder on the performance and health of layer chicks. A total of 160 three-week-old Lohmann LSL Lite pullets were used in this five-week trial. Following a four-day adaptation period on a control diet, birds were randomly assigned to one of four treatment groups receiving 0% (control), 2%, 4%, or 6% BMS in place of soybean meal. Results showed no significant (P > 0.05) differences in body weight, weight gain, feed intake, or feed conversion ratio across groups, indicating that BMS can replace soybean mealwithout compromising growth performance. Experiment 2 evaluated the inclusion of white mushroom stem (WMS) powder in layer chick diets. The same experimental design used in Experiment 1 was followed, involving 160 Lohmann LSL Lite chicks divided into four groups (0%, 2%, 4%, and 6% WMS). Statistical analysis revealed no significant (P > 0.05) differences in body weight, weight gain, FCR, or internal organ weight across the groups. Blood biochemical markers also remain unchanged, demonstrating the safety of WMS as a feed ingredient. Interestingly, gas emissions from excreta showed both linear and quadratic (P < 0.05) responses with increasing WMS levels, mirroring results observed in the BMS trial and pointing to a reduced environmental footprint. These findings suggest that WMS, like BMS, can be used to partially replace soybean mealwithout compromising bird health or productivity while offering environmental benefits. Further research is recommended to validate these findings in commercial settings. Experiment 3 focused on broiler chickens and investigated the effects of BMS inclusion on growth performance, blood chemistry, carcass traits, meat quality, and gas emissions. Data collection and analysis are underway to determine whether the findings from layer chick studies also apply to meat-producing birds. Experiment 4 is currently being conducted on layer hens to assess the impact of BMS on egg production performance and egg quality at PVAMU. Experiment 5 was conducted on broilers challenged with Eimeria spp. at UGA. A total of 240 one-day-old Cobb500 broilers were allocated with a 2 x 3 factorial design (challenge vs. non-challenge; 0, 3, 6% WMS). Each treatment group had 5 replicated cages with 8 birds per cage. On day 7 (dpi 0: days post inoculation 0), birds in the challenged groups were orally inoculated with 1 mL of a sporulated oocyst solution containing a mixed species of Eimeria acervulina (50,000 oocysts/mL), E. maxima (10,000 oocysts/mL), and E. tenella (10,000 oocysts/mL), while those in the non-challenged groups were inoculated with 1 mL of PBS. For growth performance, there were no significant interactions between the Eimeria challenge and the WMS level. The challenged groups had significantly lower body weight gain and feed intake but a higher feed conversion ratio compared to the non-challenged groups during the acute challenge period (dpi 0-6; day 7-13) (P<0.05). In summary, Eimeria challenges significantly reduced growth performance and gut health in broilers, while 6% WMS may be too high to show any beneficial effects under Eimeria challenge conditions. For major goal 4 Students from UH have played an active role in mushroom stem processing studies, participating in various stages such as sample collection, cleaning, freeze-drying, data collection, and analysis. Their involvement extends to literature reviews and assisting in the optimization of processing methods. Engineering students at UH have also contributed by helping to design and develop a machine for cutting, cleaning, and washing mushroom stems, enhancing the technical capacity of the project. In collaboration with researchers and students at PVAMU, additional efforts were made to prepare samples--through washing, drying, and grinding--for poultry feeding trials. DMS samples, derived from both white and brown mushrooms, were also sent to the UGA for rooster assay and challenge studies, where students participated in conducting animal trials to evaluate the nutritional quality of the feed materials. As part of outreach activities, a dedicated session was held at PVAMU during the "Ag Day on the Hill" Family Field Day on April 12, 2025.

Publications

• Type: Peer Reviewed Journal Articles Status: Published Year Published: 2025 Citation: Md Salahuddin, Venkatesh Balan, Kayla G. Stamps, Ahmed A. A. Abdel-Wareth, Maedeh Mohammadi, Woo Kyun Kim, Weihang Zhu, and Jayant Lohakare. (2025). Harnessing Mushrooms for Poultry Nutrition: Boosting Health, Immunity, and Productivity. Poultry Science, 105223. https://doi.org/10.1016/j.psj.2025.105223
• Type: Conference Papers and Presentations Status: Published Year Published: 2025 Citation: Kayla G. Stamps, Ahmed A.A. Abdel-Wareth, Md Salahuddin, Venkatesh Balan, Maedeh Mohammadi, Woo Kyun Kim, Weihang Zhu, Ahmed A. Ahmed, Cassandra D Gray, Adrian M.W. Avi�a, Taylor R. Carter, Trahmilla Carr, Amri Williams, Adebowale Bakare and Jayant Lohakare. 2025. Exploring brown mushroom stem as a sustainable soybean meal alternative in layer chicks diet. Abstract accepted and presented for Graduate Student Oral competition for American Society of Animal Science, ASAS 2025 Southern Section Joint Meeting, April 06, Arlington, Texas.
• Type: Conference Papers and Presentations Status: Published Year Published: 2025 Citation: Taylor Rounds-Carter, Kayla G. Stamps, Ahmed A.A. Abdel-Wareth, Md Salahuddin, Venkatesh Balan, Maedeh Mohammadi, Woo Kyun Kim, Weihang Zhu, Ahmed A. Ahmed, Cassandra D Gray, Adrian M.W. Avi�a, Trahmilla Carr, Amri Williams, Adebowale Bakare and Jayant Lohakare. 2025. Evaluating white mushroom stem as a sustainable alternative to soybean meal in poultry nutrition. Abstract accepted and presented for Undergraduate Student Oral competition for American Society of Animal Science, ASAS 2025 Southern Section Joint Meeting, April 06, Arlington, Texas.
• Type: Conference Papers and Presentations Status: Published Year Published: 2025 Citation: Ahmed A.A. Abdel-Wareth, Kayla G. Stamps, Md Salahuddin, Ahmed A. Ahmed, Venkatesh Balan, Maedeh Mohammadi, Woo Kyun Kim, Weihang Zhu, Cassandra D Gray, Adrian M.W. Avi�a, Taylor R. Carter, Trahmilla Carr, Amri Williams, Adebowale Bakare and Jayant Lohakare. 2025. Assessing Brown and White Mushroom Stem Powders as Sustainable Substitutes for Soybean Meal in Poultry Nutrition. Poultry Science Association, July 2025 PSA Annual Meeting Raleigh, NC. Accepted.
• Type: Peer Reviewed Journal Articles Status: Accepted Year Published: 2025 Citation: Kayla G. Stamps, Ahmed A.A. Abdel-Wareth, Md Salahuddin, Venkatesh Balan, Maedeh Mohammadi, Woo Kyun Kim, Weihang Zhu, Ahmed A. Ahmed, Cassandra D Gray, Adrian M.W. Avi�a, Taylor R. Carter, Trahmilla Carr, Amri Williams, Adebowale Bakare and Jayant Lohakare. 2025. Utilizing Brown Mushroom Stem Waste as an Eco-Friendly Alternative to Soybean Meal in Layer Chick Nutrition. Frontiers in Veterinary Sciences. In press.
• Type: Peer Reviewed Journal Articles Status: Awaiting Publication Year Published: 2025 Citation: Kayla G. Stamps, Ahmed A.A. Abdel-Wareth, Md Salahuddin, Venkatesh Balan, Maedeh Mohammadi, Woo Kyun Kim, Weihang Zhu, Ahmed A. Ahmed, Taylor Rounds-Carter, Cassandra D Gray, Adrian M.W. Avi�a, Trahmilla Carr, Amri Williams, Adebowale Bakare and Jayant Lohakare. 2025. Utilizing white mushroom stem powder as a sustainable substitute for soybean meal in layer chick diets. Poultry Science. Awaiting Publication. Final proof before publication submitted. doi: https://doi.org/10.1016/j.psj.2025.105571

Progress 09/01/23 to 08/31/24

Outputs
Target Audience:Students, farmers, and researchers in mushroom production, organic agricultural farmers, and animal and poultry science-related people are targeted at this time. Next year, we will present the results from these studies to agriculture and poultry farmers during Agriculture Field Days at the respective universities, as well as scientists and researchers at academic conferences. Changes/Problems:No problems or changes are necessary. None. What opportunities for training and professional development has the project provided?One research assistant professor at UH, and one graduate student each at UH, PVAMU, and UGA are recruited for this project, and several UG students have learned mushroom collection, handling, and processing. In addition,one postdoctoral associate at PVAMU was involved in washing, cleaning, andprocessingmushroom stems and will conduct research on poultry as planned in this project. How have the results been disseminated to communities of interest?At this time, no data has been disseminated.However, one MS student presented an oral presentation on the mushroom supplementation in chickens to other animal science students, scientists, and researchers at the Association of Research Directors (ARD, 2024) Symposium in Nashville, TN on April 07, 2024. Two workshops were conducted (one at Poultry Center, PVAMU and one at Tyler, Texas) for farmers to learn how to grow mushrooms,collect and prepare mushroom stems, and use them in poultry feed. What do you plan to do during the next reporting period to accomplish the goals?1. Begin formulating poultry feed incorporating DMS, considering the nutritional composition of the samples to ensure balanced nutrition. 2. Fine-tune and finalize the design and begin the development of the cleaning and sanitizing machine. 3. Use DMS powder in the poultry studies at Prairie View A&M University and the University of Georgia for feeding trials. 4. Continue exploring partnerships with mushroom companies to scale-up the production of DMS and make it more widely available to organic poultry producers. 5. Plans are underway to organize training sessions for local farmers using extension agents on these projects from PVAMU and UGA, focusing on sustainable mushroom cultivation practices and the potential use of mushroom stems in poultry feed. These sessions will provide students with opportunities to engage with industry professionals and gain practical knowledge of agricultural practices. 6. We are exploring possibilities for students to participate in farm visits and workshops, allowing them to gain hands-on experience in the agriculture and animal industry sectors. We plan to work with all team members on this project in terms of interpreting the analytical results generated and preparing manuscripts for publication.A review article on mushroom feeding in chickens is in the developing stage, and we hope to get it published before the next reporting period.

Impacts
What was accomplished under these goals? For major goal 1: Develop a process to produce dried mushroom stems (DMS) as a feed ingredient. Progress: Milestone 1.1 Develop a process to collect and prepare mushroom stems for further processing. The process of upcycling mushroom stems is developed effectively, involving cleaning the mushrooms and cutting the stems from the roots. The process begins by feeding the mushroom stems into a vibrational bowl feeder, which directs them to a custom-designed cutting machine that meets specific requirements. Following the cutting stage, the mushrooms are washed to ensure cleanliness. While commercially available vibrational bowl feeders and washing machines are utilized, the cutting machine is tailored to precise specifications for optimal performance in the upcycling process. The mushroom root trimming machine is designed to trim the roots of mushrooms. The stems are trimmed from the root and can be utilized as chicken feed. The mushroom stems are placed into a vibrational bowl feeder, which aligns and feeds them into the cutting machine developed at University of Houston (UH). The main function of the bowl feeder is to ensure that the mushroom is delivered into the cutting machine in a single line. After exiting the bowl feeder, a linear actuator was employed to move each mushroom forward through the stem cutting machine. Computer vision algorithms will determine whether the mushroom needs adjustment to reorient the stems. After alignment and rotation, if necessary, the blade cuts the mushroom stem, extracting the usable portion for chicken feed while discarding the waste root. This separation is facilitated by swinging arms that turn and place the mushrooms into their respective sorted piles. Milestone 1.2 Develop methods to freeze dry and pulverize DMS into a fine powder for use in chicken feed formulation. We have successfully doubled our freeze-drying capacity by purchasing freeze dryers (two at UH and (1 at PVAMU). This enhancement will allow us to process larger quantities of mushroom stems, supporting the scaling up of our DMS production. We visited mushroom farms to become familiar with the mushroom cultivation process and to educate workers on properly harvesting button mushrooms and caps. The students gained valuable hands-on experience and insights into the mushroom industry. Two sets of samples were prepared to investigate the effect of different cleaning methods on the mushroom stems' composition: Method 1: The freshly harvested mushroom stems without the dirt were cleaned using soft nylon brushes to remove surface dirt and freeze dried. Method 2: The harvested mushroom stems with the dirt were washed with water to produce dirt-free stems. This method was used to check how many soluble minerals, vitamins, and other compounds get lost during the water washing process. For each cleaning method, both brown and white varieties of button mushrooms were used. The cleaned and washed mushroom stem samples were freeze-dried until the moisture content was reduced to below 5 wt.%, to enhance the shelf-life of mushroom stem. Due to the low moisture content of the freeze-dried mushroom stems, a single milling method was sufficient to pulverize the stems into a fine powder in a few minutes, minimizing energy consumption. Dried mushroom stems were then finely ground into powder to be infused into chicken feed. Milestone 1.3 Determine the chemical composition and nutritional value of DMS. 1. Amino Acid Profile Analysis: We have completed a comprehensive amino acid profile analysis for both brown and white mushroom stems, comparing brush-cleaned and water-washed samples. All the essential amino acids required by poultry are present in all the samples. White mushroom stems generally showed higher amino acid content compared to brown mushroom stems. Water washing appeared to have a more significant impact on the amino acid content of white mushroom stems compared to brown mushroom stems. 2.Vitamin Analysis: We conducted a detailed vitamin analysis, focusing on B vitamins, vitamin E family, vitamin D family, and vitamin A family in DMS. Notable results include: Vitamin B2 (riboflavin) is present in relatively high concentrations (16.3 to 19.5 mg/kg) across all mushroom stem samples, with levels significantly higher than those found in common chicken feed ingredients such as corn, wheat, and soybean meal. Vitamin D2 levels were significantly higher than Vitamin D3 levels in all samples, potentially offering a valuable plant-based source of this essential nutrient for organic poultry diets. Interestingly, water-washed samples showed higher levels of vitamins across several families, including both water-soluble (B vitamins) and fat-soluble vitamins (E, D, and A), compared to brush-cleaned samples. 3. Heavy Metal Analysis: We performed heavy metal analysis to ensure the safety of DMS as a feed ingredient. Arsenic, cadmium, lead, and mercury were detected in low concentrations across all samples, ranging from 0.1-0.255 ppm, 0.02-0.021 ppm, 0.1 ppm, and 0.01 ppm respectively. These levels are well below the maximum allowable limits for animal feed (2 ppm for arsenic, 1 ppm for cadmium, 5 ppm for lead, and 0.1 ppm for mercury), indicating that the mushroom stems are safe for use in poultry feed. Interestingly, water-washed samples showed slightly higher levels of some heavy metals, particularly arsenic in white mushroom stems (0.125 ppm in brush-cleaned vs. 0.255 ppm in water-washed). Despite these variations, all values remained well within safe limits. For major goal 2, we are collecting enough dried mushroom stems for poultry studies at Prairie View A&M University. Once we have the required amount of DMS material, we will conduct studies on broilers and layers to measure the effects on performance and product quality. For major goal 3, we have sent brown and white dried mushroom stemproduct for rooster assay studies at the University of Georgia at Athens to evaluate the nutritional value of the DMS. After the rooster assay studies, we will send more DMS materials for chicken health and challenge studies at UGA. For major goal 4, Students from the University of Houston have been actively involved in sample preparation and data collection for mushroom stem processing studies. This includes participating in collecting, cleaning, and freeze-drying procedures. While the detailed analyses (amino acid, vitamin, and heavy metal studies) were conducted by external laboratories, students gained valuable experience in sample preparation, data interpretation, and understanding the importance of these analyses in feed development. Students are continuously involved in various aspects of the research project, including literature reviews, data analysis, and assisting in the optimization of processing methods. Along with the students and researchers at the University of Houston, researchers and students at Prairie View A&M University were involved in collection, washing, cleaning, freeze drying, and preparing the samples for poultry studies. The students from the Department of Engineering at UH were actively involved in designing and developing a machine for cutting, cleaning, and washing mushroom stems. DMS samples from brown and white mushrooms are sent to UGA to conduct rooster assay studies. Students are being involved in these animal studies too.

Publications

• Type: Conference Papers and Presentations Status: Published Year Published: 2024 Citation: K. G. Stamps*, A. A. A. Abdel-Wareth, and J. Lohakare. 2024. The Use of Mushrooms in Poultry Nutrition: Current and Future Prospects. Association of Research Directors Symposium. Nashville, TN. April 06-09, 2024. Presented orally by Ms. Stamps. USDA-NIFA-OREI support was acknowledged.