Enhancing biogas production from co-digestion of organic wastes mixed with cattle manure using rumen fluid and Saccharomyces cerevisiae isolate MUTJ0F

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Biogas is a cost-effective, ecologically friendly, sustainable energy source that is also a reliable method to handle food waste. Methods This study used rumen fluid and S. cerevisiae isolate MUTJ0F to maximize the production of biogas from household consumption of three substrates such as cattle manure, cafeteria leftover foods, and fruit and vegetable wastes. Plastic 0.6 L digesters operating at a mesophilic temperature (30°C) were used to produce biogas from the mixed waste. Biogas production was recorded after the 60-day retention period. Standard methods were used to figure out parameters like pH, total solids, organic carbon volatile solids, and moisture content of the raw mixed wastes. This study was carried out experimentally using factorial patterns and a completely randomized 6 × 2 design. The water displacement method was used to estimate the average amount of biogas generated by a 0.6 L digester. Result The corresponding VS/TS ratios for cattle manure, fruit and vegetable waste, and leftover cafeteria food waste were 91.2%, 80.2%, and 78.3%. More biogas was produced using rumen fluid and the S. cerevisiae isolate MUTJ0F than from mixed wastes without inoculums. The highest volume of biogas (6900.3 ml) was generated by mixing 10 ml of S. cerevisiae isolate MUTJ0F with the rumen fluids (100 ml). The digester that included 100 ml of rumen fluid and S. cerevisiae was the optimum performance for biogas production. Conclusion This result suggests that using rumen fluid and the S. cerevisiae isolate MUTJ0F as inoculums in a biogas digestor can enhance the production of biogas. 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F1000Research 2025, 14 :199 ( https://doi.org/10.12688/f1000research.161561.1 ) NOTE: If applicable, it is important to ensure the information in square brackets after the title is included in all citations of this article. Close Copy Citation Details Export Export Citation Sciwheel EndNote Ref. Manager Bibtex ProCite Sente EXPORT Select a format first Track Share ▬ ✚ Research Article Enhancing biogas production from co-digestion of organic wastes mixed with cattle manure using rumen fluid and Saccharomyces cerevisiae isolate MUTJ0F [version 1; peer review: 1 not approved] Mulugeta Fentahun https://orcid.org/0009-0007-3025-7156 1 , Birhanu Kahsay 2 Mulugeta Fentahun https://orcid.org/0009-0007-3025-7156 1 , Birhanu Kahsay 2 PUBLISHED 13 Feb 2025 Author details Author details 1 College of Natural and Computational Sciences, Department of Biology, Debre Markos University, Debre Markos, Amhara, 269, Ethiopia 2 Institute of Biotechnology, Mekelle University, Mek'ele, Tigray, Ethiopia Mulugeta Fentahun Roles: Conceptualization, Data Curation, Formal Analysis, Funding Acquisition, Investigation, Project Administration, Resources, Software, Supervision, Validation, Visualization, Writing – Original Draft Preparation, Writing – Review & Editing Birhanu Kahsay Roles: Conceptualization, Data Curation, Investigation, Methodology, Resources, Software, Validation, Writing – Review & Editing OPEN PEER REVIEW DETAILS REVIEWER STATUS This article is included in the Energy gateway. Abstract Background Biogas production from different types of biodegradable wastes used as an alternative for fossil fuels for energy consumption and biodegradable waste management. Biogas is a cost-effective, ecologically friendly, sustainable energy source that is also a reliable method to handle food waste. Methods This study used rumen fluid and S. cerevisiae isolate MUTJ0F to maximize the production of biogas from household consumption of three substrates such as cattle manure, cafeteria leftover foods, and fruit and vegetable wastes. Plastic 0.6 L digesters operating at a mesophilic temperature (30°C) were used to produce biogas from the mixed waste. Biogas production was recorded after the 60-day retention period. Standard methods were used to figure out parameters like pH, total solids, organic carbon volatile solids, and moisture content of the raw mixed wastes. This study was carried out experimentally using factorial patterns and a completely randomized 6 × 2 design. The water displacement method was used to estimate the average amount of biogas generated by a 0.6 L digester. Result The corresponding VS/TS ratios for cattle manure, fruit and vegetable waste, and leftover cafeteria food waste were 91.2%, 80.2%, and 78.3%. More biogas was produced using rumen fluid and the S. cerevisiae isolate MUTJ0F than from mixed wastes without inoculums. The highest volume of biogas (6900.3 ml) was generated by mixing 10 ml of S. cerevisiae isolate MUTJ0F with the rumen fluids (100 ml). The digester that included 100 ml of rumen fluid and S. cerevisiae was the optimum performance for biogas production. Conclusion This result suggests that using rumen fluid and the S. cerevisiae isolate MUTJ0F as inoculums in a biogas digestor can enhance the production of biogas. READ ALL READ LESS Keywords Anaerobic digestion, Biogas production, Inoculum, Rumen fluid Corresponding Author(s) Mulugeta Fentahun ( [email protected] ) Close Corresponding author: Mulugeta Fentahun Competing interests: No competing interests were disclosed. Grant information: This research was supported by Debre Markos University. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Copyright: © 2025 Fentahun M and Kahsay B. This is an open access article distributed under the terms of the Creative Commons Attribution License , which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. How to cite: Fentahun M and Kahsay B. Enhancing biogas production from co-digestion of organic wastes mixed with cattle manure using rumen fluid and Saccharomyces cerevisiae isolate MUTJ0F [version 1; peer review: 1 not approved] . F1000Research 2025, 14 :199 ( https://doi.org/10.12688/f1000research.161561.1 ) First published: 13 Feb 2025, 14 :199 ( https://doi.org/10.12688/f1000research.161561.1 ) Latest published: 18 Apr 2026, 14 :199 ( https://doi.org/10.12688/f1000research.161561.3 )  There is a newer version of this article available. Suppress this message for one day. 1. Introduction The sources of renewable energy have gained attention due to growing concern about the negative environmental effects of fossil fuels and the depletion of global petroleum reserves ( Akinbami et al ., 2001 ; Imri and Valeria, 2007 ). Long-term possible strategies for sustainable growth should be address the world’s current environmental issues and potential fossil fuel crisis ( Farid, 2022 ). Under this situation one of the most promising and feasible options appears to be renewable energy ( Alemayehu, 2014 ). Biogas remains in use worldwide as a source of renewable energy, less expensive, environmentally friendly, clean and readily available ( Abdulkareem, 2005 ; Arthur and Brew-Hammond, 2010 ; Alemayehu, 2014 ). Biogas may be generated from bio-waste or biomass and used for cooking, heating, lighting and absorption refrigeration, compressing gas for storage, use in vehicles, and generating electricity ( Harris, 2008 ; Corral et al. , 2008 , Nasir et al ., 2012 ). Biogas is generated when bacteria, and microorganisms break down bio-waste or biomass without oxygen ( Membere et al ., 2012 ). An innovative solution to increase anaerobic waste digestion yields are co-digestion due to inexpensive, simple technology, enhance the rate anaerobic digestion process by creating a better nutrient balance from the materials mixed to feed the digester, provide positive synergism for bacterial growth, and increased in biogas production ( Sosnowski et al ., 2003 ; Mata-Alvarez et al ., 2000 ; Mshandete et al ., 2004 ; Leta et al ., 2015 ). Several studies have used co-digestion to increase the rate at which organic matter is converted biologically in the biogas system to improving biogas performance ( Huang et al ., 2016 ; Alemayehu, 2014 ; Mata-Alvarez et al ., 2014 ; Abbas et al ., 2021 ). Three useful bacteria groups break down complex organic materials such as fermentative, acidogenic, and methanogenic microbes interdependently during anaerobic digestion. These microorganisms are in responsible of the four-stage process that transforms complex organics into biogas. The hydrolysis stage comes first, followed by the acidogenesis stage, the acetogenesis stage, and the methanogenesis stage ( Jia et al ., 2020 ). One of the waste products from slaughterhouses that is regularly dumped into drainage systems is rumen fluids ( Zhang et al ., 2016 ). A diverse range of fungus, bacteria, protozoa, and archaea inhabit the rumen, an anaerobic microbial habitat ( Sylvester et al ., 2004 ; Sonakya et al ., 2003 ). The rumen contains a variety of microorganisms, including cellulolitic and methanogenic bacteria ( Lopes et al ., 2004 ; Yue and Yu, 2009 ). Rumen could be helpful as an activator in the anaerobic fermentation process that produces biogas. This fermentation process resembles the biogas digester process ( Achmad et al. , 2011 ). Saccharomyces cerevisiae is an anaerobic microorganism that can increase fibre degradation, stimulate cellulolytic bacterial, fungal growth, and increase pH in digesters through organic acid production ( Lynd et al ., 2002 ; Achmad et al. , 2011 ). Debre Markos University one of the Ethiopia’s federal universities and currently there are above 20,000 resident students in the main campus during regular academic year and summer time. For this reason, these students have their meal in the university. We have an enormous amount organic waste and leftover food, which might all be utilized as inputs for the generation of anaerobic biogas. The primary ingredients of the meals provided to students at Debre Markos University include bread, injera, spaghetti, rice, meat, and various stews and sauces. The majority of the food leftovers were disposed of near the fence in the back of the male student home, which produced an unpleasant odor and suitable environment for the growth of harmful bacteria. However, various juice house wastes are among the municipal wastes that are becoming difficult to manage, this kind of waste is typically dumped in landfills along with other household wastes, which greatly pollutes the environment in Debre Markos town. Finding alternate ways to handle those organic wastes and convert them into a source of energy is therefore necessary. Therefore, by co-digestion of these organic wastes with rumen fluids and Saccharomyces cerevisiae is an excellent opportunity to produce biogas, which would be a useful way to manage those wastes and a potential source of energy for different homes enterprises. Even though, there is information on production biogas from students cafeteria wastes and juice house wastes ( Alemayehu, 2014 ; Abayneh et al ., 2014 ; Hammad et al ., 2018 ; Earnest and Singh, 2013 ) but there is limited report on method for increasing biogas generation particularly the one that using rumen fluids and Saccharomyces cerevisiae. The aim of this study was designed to know the effect of rumen fluids and S. cerevisiae isolate MUTJ0F as fermentation activator on the amount of biogas generated from food waste co-digestion in anaerobic conditions and cattle manure and also to optimize the volume of rumen fluids and S. cerevisiae isolate MUTJ0F for biogas production . Therefore, this study may lead to better management of animal dung and other solid wastes, lower ground water contamination, better health and reduced respiratory infections, improved air quality, and less deforestation and consequent soil erosion. 2. Methods 2.1 The study area The research was carried out in microbiology laboratory of department of biology, Debre Markos University, East Gojjam, Ethiopia. The university is found in Debre Markos town. Debre Markos is located at latitude and longitude of 10°20’N 37°43’E/10.330°N 37.717°E, elevation of 2,446 meters above sea level. It is 300 km away from Addis Abeba, capital city of Ethiopia and 265 km from Bahir Dar regional state of Amhara. In Debre Markos, there are 107,684 residents, comprising 49,893 men and 57,791 women ( Aynalem et al ., 2014 ). The average annual rainfall is 380 mm, while the lowest and maximum temperatures are 150°C and 220°C, respectively. 2.2 Yeast strain used in experiments Test of strain Saccharomyces cerevisiae isolate MUTJ0F (OR209280.1) with accession numbers from the National Center for Biotechnology Information was acquired from previously isolated traditional fermented alcoholic beverage (Tej) in Ethiopia. These strains of Saccharomyces cerevisiae isolate MUTJ0F (OR209280.1) were obtained in our stock cultures from the previous study ( Fentahun and Andualem, 2024 ). 2.3 Substrate collection and preparation The study was aimed at evaluating the effect of ruminant fluids and Saccharomyces cerevisiae isolate MUTJ0F (OR209280.1) on biogas generation from various mixed organic wastes under co-digestion with cattle manure. Various wastes from fruits and vegetables were gathered from the town fruit houses, leftovers food from Debre Markos University student cafeteria, fresh cattle manure (CM) from Monkorer Agroindustry Enterprise and rumen fluids (RL) from nearby slaughterhouse was taken and used as activator for biogas production. Waste from fruits and vegetables was gathered from Debre Markos town’s juice shops. Unwanted which are non-digestible materials was carefully separated from the substrate. Food scraps from leftovers were gathered from cafeteria of students found in the Main Campus of Debre Markos University daily for a week. Indigestible wastes, such bones, were carefully removed from the substrate from the gathered meal. The mixture of substrates includes peels of bread, injera, spaghetti, papaya, mango, banana, and avocado were used in this research work. To improve and maintain anaerobic digestions process, the organic wastes were manually chopped to a size of 1-4 mm ( Leta et al ., 2015 ). The CM were separated and allowed to dry for two days in direct sunlight on a plastic tray then it was shredded to an average particle size of 2 mm and kept in a refrigerator at 4°C ( Tamrat et al ., 2013 ). After measuring the samples of total solids (TS), the de-sized cattle manure and food waste were mixed separately with distilled water in 1:5 (solid waste: distilled water) volume ratio, in order to maintain the total solid in the digester between 8 to 15%, which is the optimum value for wet anaerobic digestion ( Ituen et al ., 2007 ). 2.4 Preparation of inoculums After filtering the rumen fluids, the filtrate was stored in a refrigerator until used. Then, different amount of the filtrate was added into each digester to start up the reaction ( Aurora, 1983 ; Genet et al ., 2018 ). The S. cerevisiae isolate MUTJ0F inoculum was prepared in Yeast Extract Peptone Dextrose Broth (YEPD Broth) (Sigma-Aldrich (Oxoid Limited, USA) medium containing (g/l): yeast extract 10, peptone 20, and dextrose 20. The media was sterilized at 121°C for 15 min in an autoclave. A loop full of a chosen 48 hrs old culture was inoculated into a 250 ml flask with 100 ml of the medium, and it was then shaken at room temperature at 25°C on a rotary shaker (SHKA4450-1CE) (121rpm) for 72 hrs. The inoculums were specific to each digester. 2.5 Analysis of the physic-chemical properties of the substrate Total solids (TS), volatile solids (VS), moisture content, organic carbon, and pH were measured in each sample of biodegradable cattle manure, cafeteria leftovers, and fruit and vegetable wastes using the standard methods ( APHA, 1999 ). 2.5.1 Total solids (TS) An evaporating dish (crucible) was cleaned and dried for one hour at 105°C, cooled in desiccators and weighed immediately before use 5 gram of cattle manure, fruit and vegetable waste. Five gram of sample was weighed separately applying for an standard analytical balance (LX200ABL) and placed on a pre-dried and weighed evaporating dish. Then the dish (crucible) was placed inside an oven (Contherm 260M) maintained at 105°C. The dish (crucible) was allowed to stay in the oven (Contherm 260M) for 24 hrs and then removed and allowed to cool in desiccators and weighed. Using the formula stated in APHA (1999) , the percentage of the TS was determined as follows. % TS = mDS mFS × 100 Where, % TS = percentage of total solid mDS = mass of dry sample mFS = mass of fresh sample. 2.5.2 Volatile solids (VS) The total solid was ignited at 550°C in a muffle furnace (BiBBY, Stuart) for 3 hrs to determine the volatile and fixed solids of the sample. Then volatile solid content in the sample was determined using the formula: APHA (1999) . % VS = mDS − m ( ash ) mDS × 100 Where, mDS= mass of dry samples whereas m(ash) = mass of ash 2.5.3 Organic carbon (C) According to Haug (1993) , using data from volatile solids and an empirical equation, the organic carbon was calculated, and the organic carbon content of the sample was calculated by taking into account the volatile solids content, which was expressed as a percentage: % carbon = % VS 1.8 2.5.4 Moisture content determination Oven-dying method was used to figure out the amount of moisture as the percentage of wet (initial) weight of the material lost through heating. To achieve this, 10 g of sample was dried in an oven (Contherm 260M) at 105°C for 24 hrs and weighed. Moisture content was then calculated using the formula ( Elias et al ., 2010 ): % MC = W − D W × 100 where W is the sample’s initial weight (g), D is the sample’s weight during a 24 hrs drying period at 105°C, and M is the moisture content. 2.5.5 pH determination A digital pH meter was applied for measurement the pH value (Hanna ECI pH meter, Hanna scientific, USA). The pH meter was calibrated using pH 4.0 and 7.0 buffers prior to measuring the samples of pH, resulting in a measurement that was within the proper pH range ( Arogo et al., 2009 ). 2.6 Experimental set up This study was consisted of anaerobic digestion of substrate in 12 treatments. The 12 treatment types used for anaerobic co-digestion digestion was cattle manure (CM), fruit and vegetable wastes (FVW) and leftover foods (LF) in mixtures. The first factor was dosage of rumen liquid i.e. 0 ml/100 gram, 25 ml/100 gram, 50 ml/100 gram, 75 ml/100 gram, and 100 ml/100 gram, while the second factor was dosage of S. cerevisiae isolate MUTJ0F i.e. 5 ml/100 gram, and 10 ml/100 gram of the mixed organic waste. Three replicates were used for the treatments. The study was conducted at the room temperature (30°C). Each digester water content was calculated in accordance with the suggestion of ( Ituen et al. , 2007 ). Feed stocks was mixed with distilled water to get about 8% of TS suspension. The following formula was then used to determine how much water needed to be added: 8 % = mFTS A + B where A is the mass of the freshly added sample, B is the amount of water and inoculums to be added in order to achieve an 8% TS suspension in the digester, and mFTS is the mass of the fixed total solid. 2.7 Construction of the anaerobic digesters for batch system Anaerobic digesters (plastic bottle) were constructed for bench-scale experiments with which biogas was produced out of the degradation of substrates in 0.6 L digester. The three plastic bottles were set up so that the substrate was in the first bottle, the acidified brine solution was in the center, and the last bottle was collecting the brine solution that was expelled from the second container. All the three containers were interconnected with a plastic tube having a diameter of 1 cm. The lids of all digester were sealed tightly using super glue in order to control the entry of oxygen and loss of biogas. 2.8 Biogas measurements An acidified brine solution was produced by adding NaCl to water until a solution that was supersaturated formed. The brine solution was then acidified by adding two to three drops of sulfuric acid by the method of ( Elijah et al ., 2009 ). Finally, this formed solution was contained in the second chamber. The biogas was moved to the second chamber while it was being produced in the fermentation chamber. A pressure buildup served as the catalyst for the solution displacement because the biogas is insoluble in it. The amount of gas collected was equal to the amount of water that was pushed from the cylinder. By looking at the cylinder’s graduation, the displacement of water was measured. According to Budiyono et al . (2010) , the “liquid displacement method” was used to measure the amount of biogas produced. 2.9 Burning test The measurement cylinder was filled with gas after collecting. The lit match sticks were positioned close to the cylinder mouth. The gas formed in the digester that contains methane was burnable. If no flame forms the gas produced in the digester was non-burnable. 2.10 Data analysis Version 23.0 of SPSS (IBM SPSSInc., Chicago, IL, SPSS (RRID:SCR_002865), https://www.ibm.com/support/pages/downloading-ibm-spss-statistics-23 ) was used to analyze the data. The mean and standard deviations of the triplicates analysis were calculated using analysis of variance (ANOVA). 3. Results and Discussion 3.1 Analysis of the physic-chemical properties and composition of the mixed wastes The physical-chemical properties of different mixed wastes, including their moisture contents, pH, organic carbon, total solids, and volatile solids, are shown in Table 1 . Pre-digestion of three mixed wastes, including cattle manure, cafeteria leftovers, and vegetable and fruit wastes were varied in the amount of the composition, which was due to the variability in the composition of the samples of the different substrates. Each digester mixed wastes had between 6.8 and 7.6 in pH. High moisture content percentage of mixed wastes were ranged from 62.5 to 77.6, which makes anaerobic digestion facilitates ( Fernández et al ., 2008 ). The mixed wastes physicochemical characteristics were showed a low percentage of volatile solids relative to total solids. The VS of cafeteria leftover food waste was greater (30.8%) than that of cattle manure and fruit and vegetable wastes, suggesting a comparatively higher energy content that is advantageous for biogas production. The VS/TS ratios for cafeteria leftover food waste and cattle manure were 78.3% and 91.2%, respectively. For effective biogas generation, the amount of biodegradable organic matter should be between 70 and 95 percent of the dry matter content ( Steffen et al ., 1998 ; Buffiere et al ., 2006 ). Table 1. Physico chemical properties of mixed wastes (mean ± SD). No Component Percent by weight (% w/w) Total solid (TS) Volatile solids (VS) Organic carbon (C) Moisture content (MC) pH VS/TS (%) 1 Cafeteria leftover food 22.5 ± 0.15 39.4 ± 0.06 30.8 ± 0.12 41.6 ± 0.23 62.5 ± 0.21 7.6 ± 0.21 78.3 ± 0.36 2 Fruit waste and Vegetable 17.3 ± 0.1 31.5 ± 0.3 25.2 ± 0.06 34.4 ± 0.15 66.3 ± 0.06 6.8 ± 0.06 80.2 ± 0.87 3 Cattle manure waste 47.8 ± 0.12 21.3 ± 0.15 19.4 ± 0.1 25.3 ± 0.1 77.6 ± 0.12 6.8 ± 0.06 91.2 ± 1 Total 100 - - - - - - All of the substrates were considered acceptable for anaerobic digestion because all the mixed wastes had the highest ratio of VS to TS in Table 1 . Cattle dung is used to speed up the production of biogas by promoting bacterial growth in the digester. Co-digestion has a positive synergistic effect by neutralising pH, increasing buffering capacity, reducing the effects of harmful compounds, and supplying more balanced nutrients like vitamins, trace metals, and other substances required for microbial growth ( Fang, 2010 ; Aragaw et al., 2013 ; Jianzheng et al ., 2011 ). 3.2 The effect of rumen liquid and S. cerevisiae isolate MUTJ0F on biogas production The amount of biogas generated from biogas production using rumen fluids and S. cerevisiae isolate MUTJ0F as a fermentation activator is presented in Table 2 . The production of biogas at the mesophilic temperature (30°C) using rumen fluid and the S. cerevisiae isolate MUTJ0F was more intense than that without inoculums. Rumen fluid and S. cerevisiae isolate MUTJ0F inoculums increased production biogas more than by four times in compare to a mixed substrate without inoculums. This implies that the high anaerobic bacterial concentration in rumen fluid efficiently breaks down organic substrate from a mixed waste. The findings are consistent with the work of other researchers ( Tamirat, 2012 ; Sakar et al., 2008 ; Yitayal, 2011 ; Forster-Carneiro et al ., 2008 ; Abdullahi et al ., 2011 ). Table 2. The effect biogas production on 12 treatments used in anaerobic digestion of mixed wastes combinations with rumen fluid and S. cerevisiae isolate MUTJ0F doses (ml). Treatments Mixed Wastes (MW) 100 g Rumen fluid S. cerevisiae isolate MUTJ0F Methane (mean ± SD) T1 MW 1 0 ml RF 5 ml 1600 ± 0.49 ml T2 MW 2 0 ml RF 10 ml 1600 ± 0.62 ml T3 MW 3 25 ml RF 5 ml 2100.7 ± 0.67 ml T4 MW 4 25 ml RF 10 ml 2300.3 ± 0.17 ml T5 MW 5 50 ml RF 5 ml 3400.4 ± 0.49 ml T6 MW 6 50 ml RF 10 ml 3800.6 ± 0.56 ml T7 MW 7 75 ml RF 5 ml 6000.4 ± 0.17 ml T8 MW 8 75 ml RF 10 ml 6500.5 ± 0.25 ml T9 MW 9 100 ml RF 5 ml 6800.4 ± 0.12 ml T10 MW 10 100 ml RF 10 ml 6900.3 ± 0.26 ml T11 MW 11 150 ml RF 5 ml 6300.57 ± 0.47 ml T12 MW 12 150 ml RF 10 ml 6350.13 ± 0.06 ml T control MW 0 0 ml RF 0 ml 1500.2 ± 0.1 ml According to the results, the maximum amount of biogas (6900.3 ml) was produced by combining 100 ml of rumen fluids with 10 ml of S. cerevisiae isolate MUTJ0F/100 gram mixed waste, followed by 100 ml of rumen fluids and 5 ml of S. cerevisiae isolate MUTJ0F/100 gram mixed waste. As the digester rumen content varies, the results also showed that the amount of biogas generated increases when the dose of S. cerevisiae isolate MUTJ0F is increased from 5 ml to 10 ml. The addition of the S. cerevisiae isolate MUTJ0F culture to the digester enhanced the number of ruminal bacteria and their activity while also improving the digestibility of dry matter, crude protein, and hemicelluloses ( Lynd et al ., 2002 ; Wiedmeier et al ., 1987 ; Wilson, 2011 ). According to the findings, 100 ml of rumen fluid mixed with 10 ml of S. cerevisiae isolate MUTJ0F was the optimum quantity to provide the best performance of biogas production. 3.3 pH variations in digestor T10 during fermentation The pH value was checked every 10 days to examine the effect of change during digestion on bacterial activity ( Figure 1 ). The pH value drops rapidly, reached 4.3 on day forty. After that, the pH value then increased to 6.4 over the period of the following sixty days. This low pH value was permitted very little methanogenic bacterial activity and the acid-formers might yet be able to proliferate and generate large amounts of volatile acids, which lowers the pH of the digester’s contents ( Joyce et al ., 2018 ; Li et al ., 2018 ; Stabnikova et al. , 2005 ; Lin et al. , 2011 ). On the other hand, the pH starts to rise on day forty of fermentation. This could be because proteins break down to release ammonia, which causes alkalinity ( Gerardi, 2003 ). This also brings the pH closer to neutral, which makes it easier for methanogenic microbes to multiply and produce methane. Methanogenic bacteria the optimum pH values between 6.8 and 7.2 ( Anunputtikul and Rodtong, 2004 ; Budiyono et al. , 2010 ). Figure 1. T10 pH variation during the digestion process. 3.4 Combustibility of biogas A Bunsen burner attached to the digester gas outlet was utilized to evaluate the biogas combustibility after the 60 th day of the digestion period in a 0.6 L digester. The flammable gas was observed at the burner’s mouth confirming presence of methane in the biogas ( Figure 2 ). Figure 2. Performance of biogas production after 60 days observation. A. Mix of wastes which was subjected to digesters; B. 0.6 L plastic bottle digesters; and C . Combustibility of Biogas. 4. Conclusion S. cerevisiae isolate MUTJ0F and Rumen fluid seeded to a biodigester significantly impacted the total amount of biogas produced. Rumen fluid and S. cerevisiae isolate MUTJ0F inoculums enhanced production of biogas by more than four times in comparison to a mixed substrate without inoculums. The optimal results for biogas production were achieved with a 100 ml rumen fluid. A mixture of 100 ml of rumen fluids and 10 ml of S. cerevisiae isolate MUTJ0F/100 grams of mixed waste produced the highest volume of biogas (6900.3 ml). Biodegradation, microbial strength, and biogas generation can all be enhanced by using animal rumen fluid and the S. cerevisiae isolate MUTJ0F as inoculums in biogas digesters. Author contribution statement Conceptualization, M.F. and B.K; Methodology, B.K.; Data analysis, M.F.; Investigation, M.F. and B.K.; Resources, M.F. and B.K.; Writing – original draft, M.F.; Writing – review and editing, M.F. and B.K.; Visualization, M.F.; Funding acquisition, M.F. All authors have read and agreed to the published the manuscript. Consent for publication Not applicable. Ethical approval Not applicable. Data availability statement Underlying data Figshare: The effect biogas production on 12 treatments used in anaerobic digestion of mixed wastes combinations with rumen fluid and S. cerevisiae isolate MUTJ0F doses (ml) of data analysis. https://doi.org/10.6084/m9.figshare.28284806 ( Fentahun and Kashay, 2025a ). This project contains the following underlying data: - Raw data. 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Publisher Full Text Comments on this article Comments (0) Version 3 VERSION 3 PUBLISHED 13 Feb 2025 ADD YOUR COMMENT Comment Author details Author details 1 College of Natural and Computational Sciences, Department of Biology, Debre Markos University, Debre Markos, Amhara, 269, Ethiopia 2 Institute of Biotechnology, Mekelle University, Mek'ele, Tigray, Ethiopia Mulugeta Fentahun Roles: Conceptualization, Data Curation, Formal Analysis, Funding Acquisition, Investigation, Project Administration, Resources, Software, Supervision, Validation, Visualization, Writing – Original Draft Preparation, Writing – Review & Editing Birhanu Kahsay Roles: Conceptualization, Data Curation, Investigation, Methodology, Resources, Software, Validation, Writing – Review & Editing Competing interests No competing interests were disclosed. Grant information This research was supported by Debre Markos University. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Article Versions (3) version 3 Revised Published: 18 Apr 2026, 14:199 https://doi.org/10.12688/f1000research.161561.3 version 2 Revised Published: 19 Dec 2025, 14:199 https://doi.org/10.12688/f1000research.161561.2 version 1 Published: 13 Feb 2025, 14:199 https://doi.org/10.12688/f1000research.161561.1 Copyright © 2025 Fentahun M and Kahsay B. This is an open access article distributed under the terms of the Creative Commons Attribution License , which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Download Export To Sciwheel Bibtex EndNote ProCite Ref. 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Key to Reviewer Statuses VIEW HIDE Approved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested Approved with reservations A number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit. Not approved Fundamental flaws in the paper seriously undermine the findings and conclusions Version 1 VERSION 1 PUBLISHED 13 Feb 2025 Views 0 Cite How to cite this report: Emmanuel JK. Reviewer Report For: Enhancing biogas production from co-digestion of organic wastes mixed with cattle manure using rumen fluid and Saccharomyces cerevisiae isolate MUTJ0F [version 1; peer review: 1 not approved] . F1000Research 2025, 14 :199 ( https://doi.org/10.5256/f1000research.177603.r401840 ) The direct URL for this report is: https://f1000research.com/articles/14-199/v1#referee-response-401840 NOTE: it is important to ensure the information in square brackets after the title is included in this citation. Close Copy Citation Details Reviewer Report 19 Aug 2025 Jovine Kamuhabwa Emmanuel , Chemistry, University of Dar es Salaam, Dar es Salaam, Dar es Salaam, Tanzania Not Approved VIEWS 0 https://doi.org/10.5256/f1000research.177603.r401840 Title The title captures the content of the current work. Abstract The abstract has no information on the study, methodology used and key findings ... Continue reading READ ALL Title The title captures the content of the current work. Abstract The abstract has no information on the study, methodology used and key findings and recommendation based on findings. Authors must re-write this to reflect the current work. Other comments are found in the main document. Introduction The section has not been written properly and not easy to follow and understand. Authors must write this in a scholarly manner to reflect an article for publication. The novelty is not well feature thus, authors must reveal the novelty of the work. Other comments are found in the main document. Materials and methods Proper details and step after step must be provided for readers to follow. Authors need to take time and re-do it properly. Other comments are found in the main document. Results and discussion Results are not well organized and discussed with previous work. Authors need to re-think on how they can organize these data for publication. Other comments are found in the main document. Conclusions and recommendations This section is not well written thus, must be re-written for clarity. Only key findings and recommendation must be presented and not otherwise. Other comments are found in the main document. General comment The manuscript must benefit from a serious English Language editing before being indexed. The manuscript must be enriched with up-to-date citations which are available in the literature. Other comments are found in the main document. Is the work clearly and accurately presented and does it cite the current literature? No Is the study design appropriate and is the work technically sound? Yes Are sufficient details of methods and analysis provided to allow replication by others? Partly If applicable, is the statistical analysis and its interpretation appropriate? Not applicable Are all the source data underlying the results available to ensure full reproducibility? Partly Are the conclusions drawn adequately supported by the results? Partly Competing Interests: No competing interests were disclosed. Reviewer Expertise: Catalysis, materials science and bioenergy I confirm that I have read this submission and believe that I have an appropriate level of expertise to state that I do not consider it to be of an acceptable scientific standard, for reasons outlined above. Close READ LESS CITE CITE HOW TO CITE THIS REPORT Emmanuel JK. Reviewer Report For: Enhancing biogas production from co-digestion of organic wastes mixed with cattle manure using rumen fluid and Saccharomyces cerevisiae isolate MUTJ0F [version 1; peer review: 1 not approved] . F1000Research 2025, 14 :199 ( https://doi.org/10.5256/f1000research.177603.r401840 ) The direct URL for this report is: https://f1000research.com/articles/14-199/v1#referee-response-401840 NOTE: it is important to ensure the information in square brackets after the title is included in all citations of this article. COPY CITATION DETAILS Report a concern Author Response 29 Dec 2025 Mulugeta Fentahun , College of Natural and Computational Sciences, Department of Biology, Debre Markos University, Debre Markos, 269, Ethiopia 29 Dec 2025 Author Response Mulugeta Fentahun Dr. Jovine Kamuhabwa Emmanuel thank you very much for your insightful comments and the time you dedicated to review. We greatly appreciate your feedback and suggestions. We ... Continue reading Mulugeta Fentahun Dr. Jovine Kamuhabwa Emmanuel thank you very much for your insightful comments and the time you dedicated to review. We greatly appreciate your feedback and suggestions. We provide a detailed response to each comment, along with the corresponding changes made in the revised manuscript below: Title: Comment : The title captures the content of the current work. Response : Thank you for your positive feedback. Abstract Revision: Comment : The abstract has no information on the study, methodology used, and key findings and recommendations based on the findings. Authors must re-write this to reflect the current work. Response : Thank you for your insightful comment. We have thoroughly revised the abstract part in response to your suggestions (see the revised manuscript). Introduction Revision: Comment : The section has not been written properly and is not easy to follow and understand. Authors must write this in a scholarly manner to reflect an article for publication. The novelty is not well featured; thus, authors must reveal the novelty of the work. Response : We sincerely thank the reviewer for your constructive feedback. We have thoroughly revised the introduction section to improve clarity, coherence, and scholarly manner. Additionally, we have clearly highlighted the novelty of the study by emphasizing its unique contributions in the revised manuscript (refer to the revised manuscript). Methodology Revision: Comment : Proper details and step after step must be provided for readers to follow. Authors need to take time and re-do it properly. Response: Thank you for this valuable observation. We have substantially revised the materials and methods section to improve clarity, completeness, and reproducibility. The updated version now provides detailed, step-by-step procedures for the experimental work, enabling readers to follow the methodology accurately and enhancing the scientific rigor of the manuscript (see the revised manuscript). Results and Discussion Revision: Comment : Results are not well organized and discussed with previous work. Authors need to re-think on how they can organize these data for publication. Response: We sincerely thank the reviewer for your constructive feedback. We have carefully revised the results and discussion section to enhance clarity and coherence. The revised version enriched with up-to-date citations (recent literature) and emphasizes the key findings. Please refer to the updated section in the revised manuscript. Conclusions and recommendations Revision: Comment : This section is not well written thus, must be re-written for clarity. Only key findings and recommendation must be presented and not otherwise . Response: Thank you for your insightful comment. We have revised the conclusions section to clearly highlight the study’s key findings and recommendation. Please see the updated section in the revised manuscript. General comment: Comment : The manuscript must benefit from a serious English Language editing before being indexed. Response: We sincerely thank the reviewer for the valuable suggestion regarding the language quality of our manuscript. We have thoroughly revised the manuscript to improve English clarity, grammar, and readability. Comment : The manuscript must be enriched with up-to-date citations which are available in the literature. Response: Thank you for your comments. We have updated the manuscript by incorporating recent literature, as suggested (see the revised manuscript). Thank you once again for your valuable feedback. Your comments have significantly contributed to improving the quality of this paper. Best regards, Authors Mulugeta Fentahun Dr. Jovine Kamuhabwa Emmanuel thank you very much for your insightful comments and the time you dedicated to review. We greatly appreciate your feedback and suggestions. We provide a detailed response to each comment, along with the corresponding changes made in the revised manuscript below: Title: Comment : The title captures the content of the current work. Response : Thank you for your positive feedback. Abstract Revision: Comment : The abstract has no information on the study, methodology used, and key findings and recommendations based on the findings. Authors must re-write this to reflect the current work. Response : Thank you for your insightful comment. We have thoroughly revised the abstract part in response to your suggestions (see the revised manuscript). Introduction Revision: Comment : The section has not been written properly and is not easy to follow and understand. Authors must write this in a scholarly manner to reflect an article for publication. The novelty is not well featured; thus, authors must reveal the novelty of the work. Response : We sincerely thank the reviewer for your constructive feedback. We have thoroughly revised the introduction section to improve clarity, coherence, and scholarly manner. Additionally, we have clearly highlighted the novelty of the study by emphasizing its unique contributions in the revised manuscript (refer to the revised manuscript). Methodology Revision: Comment : Proper details and step after step must be provided for readers to follow. Authors need to take time and re-do it properly. Response: Thank you for this valuable observation. We have substantially revised the materials and methods section to improve clarity, completeness, and reproducibility. The updated version now provides detailed, step-by-step procedures for the experimental work, enabling readers to follow the methodology accurately and enhancing the scientific rigor of the manuscript (see the revised manuscript). Results and Discussion Revision: Comment : Results are not well organized and discussed with previous work. Authors need to re-think on how they can organize these data for publication. Response: We sincerely thank the reviewer for your constructive feedback. We have carefully revised the results and discussion section to enhance clarity and coherence. The revised version enriched with up-to-date citations (recent literature) and emphasizes the key findings. Please refer to the updated section in the revised manuscript. Conclusions and recommendations Revision: Comment : This section is not well written thus, must be re-written for clarity. Only key findings and recommendation must be presented and not otherwise . Response: Thank you for your insightful comment. We have revised the conclusions section to clearly highlight the study’s key findings and recommendation. Please see the updated section in the revised manuscript. General comment: Comment : The manuscript must benefit from a serious English Language editing before being indexed. Response: We sincerely thank the reviewer for the valuable suggestion regarding the language quality of our manuscript. We have thoroughly revised the manuscript to improve English clarity, grammar, and readability. Comment : The manuscript must be enriched with up-to-date citations which are available in the literature. Response: Thank you for your comments. We have updated the manuscript by incorporating recent literature, as suggested (see the revised manuscript). Thank you once again for your valuable feedback. Your comments have significantly contributed to improving the quality of this paper. Best regards, Authors Competing Interests: No competing interests were disclosed. Close Report a concern Respond or Comment COMMENTS ON THIS REPORT Author Response 29 Dec 2025 Mulugeta Fentahun , College of Natural and Computational Sciences, Department of Biology, Debre Markos University, Debre Markos, 269, Ethiopia 29 Dec 2025 Author Response Mulugeta Fentahun Dr. Jovine Kamuhabwa Emmanuel thank you very much for your insightful comments and the time you dedicated to review. We greatly appreciate your feedback and suggestions. We ... Continue reading Mulugeta Fentahun Dr. Jovine Kamuhabwa Emmanuel thank you very much for your insightful comments and the time you dedicated to review. We greatly appreciate your feedback and suggestions. We provide a detailed response to each comment, along with the corresponding changes made in the revised manuscript below: Title: Comment : The title captures the content of the current work. Response : Thank you for your positive feedback. Abstract Revision: Comment : The abstract has no information on the study, methodology used, and key findings and recommendations based on the findings. Authors must re-write this to reflect the current work. Response : Thank you for your insightful comment. We have thoroughly revised the abstract part in response to your suggestions (see the revised manuscript). Introduction Revision: Comment : The section has not been written properly and is not easy to follow and understand. Authors must write this in a scholarly manner to reflect an article for publication. The novelty is not well featured; thus, authors must reveal the novelty of the work. Response : We sincerely thank the reviewer for your constructive feedback. We have thoroughly revised the introduction section to improve clarity, coherence, and scholarly manner. Additionally, we have clearly highlighted the novelty of the study by emphasizing its unique contributions in the revised manuscript (refer to the revised manuscript). Methodology Revision: Comment : Proper details and step after step must be provided for readers to follow. Authors need to take time and re-do it properly. Response: Thank you for this valuable observation. We have substantially revised the materials and methods section to improve clarity, completeness, and reproducibility. The updated version now provides detailed, step-by-step procedures for the experimental work, enabling readers to follow the methodology accurately and enhancing the scientific rigor of the manuscript (see the revised manuscript). Results and Discussion Revision: Comment : Results are not well organized and discussed with previous work. Authors need to re-think on how they can organize these data for publication. Response: We sincerely thank the reviewer for your constructive feedback. We have carefully revised the results and discussion section to enhance clarity and coherence. The revised version enriched with up-to-date citations (recent literature) and emphasizes the key findings. Please refer to the updated section in the revised manuscript. Conclusions and recommendations Revision: Comment : This section is not well written thus, must be re-written for clarity. Only key findings and recommendation must be presented and not otherwise . Response: Thank you for your insightful comment. We have revised the conclusions section to clearly highlight the study’s key findings and recommendation. Please see the updated section in the revised manuscript. General comment: Comment : The manuscript must benefit from a serious English Language editing before being indexed. Response: We sincerely thank the reviewer for the valuable suggestion regarding the language quality of our manuscript. We have thoroughly revised the manuscript to improve English clarity, grammar, and readability. Comment : The manuscript must be enriched with up-to-date citations which are available in the literature. Response: Thank you for your comments. We have updated the manuscript by incorporating recent literature, as suggested (see the revised manuscript). Thank you once again for your valuable feedback. Your comments have significantly contributed to improving the quality of this paper. Best regards, Authors Mulugeta Fentahun Dr. Jovine Kamuhabwa Emmanuel thank you very much for your insightful comments and the time you dedicated to review. We greatly appreciate your feedback and suggestions. We provide a detailed response to each comment, along with the corresponding changes made in the revised manuscript below: Title: Comment : The title captures the content of the current work. Response : Thank you for your positive feedback. Abstract Revision: Comment : The abstract has no information on the study, methodology used, and key findings and recommendations based on the findings. Authors must re-write this to reflect the current work. Response : Thank you for your insightful comment. We have thoroughly revised the abstract part in response to your suggestions (see the revised manuscript). Introduction Revision: Comment : The section has not been written properly and is not easy to follow and understand. Authors must write this in a scholarly manner to reflect an article for publication. The novelty is not well featured; thus, authors must reveal the novelty of the work. Response : We sincerely thank the reviewer for your constructive feedback. We have thoroughly revised the introduction section to improve clarity, coherence, and scholarly manner. Additionally, we have clearly highlighted the novelty of the study by emphasizing its unique contributions in the revised manuscript (refer to the revised manuscript). Methodology Revision: Comment : Proper details and step after step must be provided for readers to follow. Authors need to take time and re-do it properly. Response: Thank you for this valuable observation. We have substantially revised the materials and methods section to improve clarity, completeness, and reproducibility. The updated version now provides detailed, step-by-step procedures for the experimental work, enabling readers to follow the methodology accurately and enhancing the scientific rigor of the manuscript (see the revised manuscript). Results and Discussion Revision: Comment : Results are not well organized and discussed with previous work. Authors need to re-think on how they can organize these data for publication. Response: We sincerely thank the reviewer for your constructive feedback. We have carefully revised the results and discussion section to enhance clarity and coherence. The revised version enriched with up-to-date citations (recent literature) and emphasizes the key findings. Please refer to the updated section in the revised manuscript. Conclusions and recommendations Revision: Comment : This section is not well written thus, must be re-written for clarity. Only key findings and recommendation must be presented and not otherwise . Response: Thank you for your insightful comment. We have revised the conclusions section to clearly highlight the study’s key findings and recommendation. Please see the updated section in the revised manuscript. General comment: Comment : The manuscript must benefit from a serious English Language editing before being indexed. Response: We sincerely thank the reviewer for the valuable suggestion regarding the language quality of our manuscript. We have thoroughly revised the manuscript to improve English clarity, grammar, and readability. Comment : The manuscript must be enriched with up-to-date citations which are available in the literature. Response: Thank you for your comments. We have updated the manuscript by incorporating recent literature, as suggested (see the revised manuscript). Thank you once again for your valuable feedback. Your comments have significantly contributed to improving the quality of this paper. Best regards, Authors Competing Interests: No competing interests were disclosed. Close Report a concern COMMENT ON THIS REPORT Comments on this article Comments (0) Version 3 VERSION 3 PUBLISHED 13 Feb 2025 ADD YOUR COMMENT Comment keyboard_arrow_left keyboard_arrow_right Open Peer Review Reviewer Status info_outline Alongside their report, reviewers assign a status to the article: Approved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested Approved with reservations A number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit. Not approved Fundamental flaws in the paper seriously undermine the findings and conclusions Reviewer Reports Invited Reviewers 1 2 3 Version 3 (revision) 18 Apr 26 read Version 2 (revision) 19 Dec 25 read read Version 1 13 Feb 25 read Jovine Kamuhabwa Emmanuel , University of Dar es Salaam, Dar es Salaam, Tanzania Dominic Yellezuome , University of Mines and Technology, Tarkwa, Ghana Josef Marousek , Technical University of Košice, Košice–Sever, Slovakia Comments on this article All Comments (0) Add a comment Sign up for content alerts Sign Up You are now signed up to receive this alert Browse by related subjects keyboard_arrow_left Back to all reports Reviewer Report 0 Views copyright © 2026 Yellezuome D. This is an open access peer review report distributed under the terms of the Creative Commons Attribution License , which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. 29 Apr 2026 | for Version 3 Dominic Yellezuome , University of Mines and Technology, Tarkwa, Western Region, Ghana 0 Views copyright © 2026 Yellezuome D. This is an open access peer review report distributed under the terms of the Creative Commons Attribution License , which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. format_quote Cite this report speaker_notes Responses (0) Approved With Reservations info_outline Alongside their report, reviewers assign a status to the article: Approved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested Approved with reservations A number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit. Not approved Fundamental flaws in the paper seriously undermine the findings and conclusions The manuscript requires a thorough revision before it can be considered for acceptance. The final sentence in the Methods section of the abstract should either be removed or integrated with the preceding sentence for better flow and clarity. Several statements are supported by multiple references (three or more) without clearly justifying the relevance of each, and some cited works appear outdated. For example, the last sentence of the first paragraph of the Introduction. The manuscript does not clearly articulate its scientific novelty in comparison to existing studies. This should be strengthened. A proper reference should be provided for the rainfall and temperature data reported for Debre Markos. In Subsection 2.5.1, the formula does not clearly indicate how the empty crucible is accounted for. Additionally, the explanation of variables should be formatted horizontally for consistency with the rest of the manuscript. The authors indicated that subsection 2.5.4 has been deleted; however, this subsection still remains, which describes the same procedure as subsection 2.5.1. In Section 2.6, “The 12 treatment types used for anaerobic co-digestion were cattle manure, fruit and vegetable waste, and cafeteria leftover food in mixtures of 1:1 ratio.” How do the three substrates have a mixing ratio of 1:1? How is the information provided in Section 2.3 “After measuring the total solids (TS) of the samples, the de-sized cattle manure and food waste were mixed separately with distilled water in a 1:5 ratio (w/v) to maintain the total solid in the digester between 8 and 15%, which is the optimum value for wet anaerobic digestion (Ituen et al., 2007).” and Section 2.6 “Feed stock was mixed with distilled water to obtain approximately 8% of TS suspension.” different. Section 2.7 “Anaerobic digesters (plastic bottles) were constructed for bench-scale experiments, in which biogas was produced from the degradation of substrates in a 0.6 L digester with a working volume of 450 mL.” Why use a different unit? The retention time (duration) of anaerobic digestion is not specified in Section 2.7, despite being mentioned in the authors’ response. This information should be clearly included. The control test was not described in the methodology section. Figure 1 should be referenced in Section 2.7. Additionally, the experimental setup shown does not indicate that the digesters were maintained in an oven, which should be clarified. It is also recommended that Section 2.7 be presented before Section 2.6 for better logical flow. Referencing Figure 1 again in Section 3.2 is unnecessary and can be removed. Biogas production should be reported in terms of specific yield (e.g., mL/gVS or mL/gTS) to allow comparison with existing literature. The total solids or volatile solids before digestion are used for this assessment. The response to the comment regarding the exclusion of rumen fluid (Table 2) is insufficient. The rationale for evaluating only the Saccharomyces cerevisiae isolate (MUTJ0F) instead of including rumen fluid should be clearly justified. In Table 2, how relevant is the column containing MW 0 – MW 12 ? Although the authors state that pH was monitored for 80 days to assess post-digestion stabilization, this was not included in the study objectives nor discussed in the Results and Discussion section. This inconsistency should be addressed. What is the difference between Figure 1b and Figure 3a? Some responses to reviewer comments lack clarity, making it difficult to understand the revisions made (e.g., Comments 31, 32 and 33). These responses should be improved for transparency. Competing Interests No competing interests were disclosed. Reviewer Expertise Anaerobic digestion, pyrolysis and gasification I confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard, however I have significant reservations, as outlined above. reply Respond to this report Responses (0) Yellezuome D. Peer Review Report For: Enhancing biogas production from co-digestion of organic wastes mixed with cattle manure using rumen fluid and Saccharomyces cerevisiae isolate MUTJ0F [version 1; peer review: 1 not approved] . F1000Research 2025, 14 :199 ( https://doi.org/10.5256/f1000research.197955.r476295) NOTE: it is important to ensure the information in square brackets after the title is included in this citation. The direct URL for this report is: https://f1000research.com/articles/14-199/v3#referee-response-476295 keyboard_arrow_left Back to all reports Reviewer Report 0 Views copyright © 2026 Marousek J. This is an open access peer review report distributed under the terms of the Creative Commons Attribution License , which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. 13 Jan 2026 | for Version 2 Josef Marousek , Technical University of Košice, Košice–Sever, Slovakia 0 Views copyright © 2026 Marousek J. This is an open access peer review report distributed under the terms of the Creative Commons Attribution License , which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. format_quote Cite this report speaker_notes Responses (1) Not Approved info_outline Alongside their report, reviewers assign a status to the article: Approved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested Approved with reservations A number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit. Not approved Fundamental flaws in the paper seriously undermine the findings and conclusions The urgency and significance of the research hypothesis needs to be better justified and placed into [economic/industrial/environmental] context. It is appropriate to adapt he methodology to the current state of knowledge. The applicability needs to be strenghtened [cost breakdown and corresponding financial analyses are hard to find]. It is advisable to improve the conclusions in terms of synthesis and fertilization. Is the work clearly and accurately presented and does it cite the current literature? No Is the study design appropriate and is the work technically sound? Partly Are sufficient details of methods and analysis provided to allow replication by others? Partly If applicable, is the statistical analysis and its interpretation appropriate? Partly Are all the source data underlying the results available to ensure full reproducibility? No Are the conclusions drawn adequately supported by the results? No Competing Interests No competing interests were disclosed. Reviewer Expertise phytotechnology, commercialization I confirm that I have read this submission and believe that I have an appropriate level of expertise to state that I do not consider it to be of an acceptable scientific standard, for reasons outlined above. reply Respond to this report Responses (1) Author Response 18 Apr 2026 Mulugeta Fentahun, College of Natural and Computational Sciences, Department of Biology, Debre Markos University, Debre Markos, 269, Ethiopia Mulugeta Fentahun Professor Josef Marousek thank you very much for your insightful comments and the time you dedicated to review our article. I greatly appreciate your feedback and suggestions. Please find my responses to your comments below: Comment1 : The urgency and significance of the research hypothesis needs to be better justified and placed into [economic/industrial/environmental] context. Response : Thank you for this valuable comment. We have revised the manuscript to better clarify the urgency and significance of the research hypothesis within economic, industrial, and environmental contexts. Refer to the introduction part of the updated manuscript. Comment2 : It is appropriate to adapt the methodology to the current state of knowledge Response: Thank you for this helpful comment. We have revised the methodology to align with the current state of knowledge by updating the experimental method based on recent literature. Refer to the method section of the updated manuscript. Comment3 : The applicability needs to be strengthened [cost breakdown and corresponding financial analyses are hard to find]. Response: Thank you for this important comment. We have strengthened the applicability of the study by considering the economic feasibility and practical potential of the study. Comment4 : It is advisable to improve the conclusions in terms of synthesis and fertilization. Response: Thank you for this valuable suggestion. We have revised the conclusion section (refer to the revised manuscript). Thank you once again for your valuable feedback. Your comments have significantly contributed to improving the quality of this paper. Best regards, Authors View more View less Competing Interests No competing interests were disclosed. reply Respond Report a concern Marousek J. Peer Review Report For: Enhancing biogas production from co-digestion of organic wastes mixed with cattle manure using rumen fluid and Saccharomyces cerevisiae isolate MUTJ0F [version 1; peer review: 1 not approved] . F1000Research 2025, 14 :199 ( https://doi.org/10.5256/f1000research.193404.r449755) NOTE: it is important to ensure the information in square brackets after the title is included in this citation. The direct URL for this report is: https://f1000research.com/articles/14-199/v2#referee-response-449755 keyboard_arrow_left Back to all reports Reviewer Report 0 Views copyright © 2026 Yellezuome D. This is an open access peer review report distributed under the terms of the Creative Commons Attribution License , which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. 09 Jan 2026 | for Version 2 Dominic Yellezuome , University of Mines and Technology, Tarkwa, Western Region, Ghana 0 Views copyright © 2026 Yellezuome D. This is an open access peer review report distributed under the terms of the Creative Commons Attribution License , which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. format_quote Cite this report speaker_notes Responses (1) Approved With Reservations info_outline Alongside their report, reviewers assign a status to the article: Approved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested Approved with reservations A number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit. Not approved Fundamental flaws in the paper seriously undermine the findings and conclusions This manuscript aimed at enhancing biogas production from a co-digestion of various organic wastes using rumen fluid and Saccharomyces cerevisiae isolate MUTJ0F as activators. The manuscript requires a careful major revision before it could possibly be accepted. There is too much information in the methods paragraph in the Abstract Section, in which some of it can not be found in the Methods Section, such as “This study was conducted experimentally using factorial patterns and a completely randomized 6 × 2 design.” And some of them are not important to be in the Abstract Section. Such as “Standard techniques were used to measure the physicochemical characteristics of the mixed waste, such as pH, total solids, volatile solids, organic carbon, and moisture. This study was conducted experimentally using factorial patterns and a completely randomized 6 × 2 design. The water displacement method was used to estimate the average amount of biogas generated by a 0.6 L digester.” These sentences are similar; hence should be merged. “Biogas production from different types of biodegradable waste is an alternative to fossil fuels for energy consumption and biodegradable waste management. Biogas is a cost-effective, environmentally friendly, and sustainable energy source that provides a reliable method for handling food waste.” “An innovative solution to increase anaerobic waste digestion yields is codigestion, which is an inexpensive and simple technology that enhances the anaerobic digestion rate by creating a better nutrient balance from the materials mixed to feed the digester, provides positive synergism for bacterial growth, and increases biogas production (Sosnowski et al., 2003; Mata-Alvarez et al., 2000; Mshandete et al., 2004; Leta et al., 2015). Several studies have used co-digestion to increase the rate at which organic matter is converted biologically in the biogas system to improve biogas performance (Huang et al., 2016; Alemayehu, 2014; Mata-Alvarez et al., 2014; Abbas et al., 2021).” “A diverse range of fungi, bacteria, protozoa, and archaea inhabit the rumen, an anaerobic microbial habitat (Sylvester et al., 2004; Sonakya et al., 2003). The rumen contains a variety of microorganisms, including cellulolytic and methanogenic bacteria (Lopes et al., 2004; Yue and Yu, 2009).” “Institutions such as universities, agro-industries, and municipalities that produce large amounts of food and fruit-vegetable waste should implement co-digestion systems using these inoculants to improve waste management, lower environmental pollution, and produce renewable energy. Additionally, by offering a cleaner, sustainable alternative for household and industrial energy, expanding this strategy could improve public health, reduce reliance on firewood, and mitigate deforestation.” Authors cited lists of 3 or more references without proper justification of the relevance of each one. Hence, avoid citing more than two references and outdated references. “Although there is information on biogas production from organic waste or single inoculums (Alemayehu, 2014; Abayneh et al., 2014; Hammad et al., 2018; Earnest and Singh, 2013) but there is limited reports on methods for increasing biogas generation, particularly those that use rumen fluid and S. cerevisiae.” This paper is not clear in the description of the scientific novelty in comparison to related previously published and referenced in your work below: Achmad KT, Gani SA, Sunarto SI, et al.: The use of rumen liquid and Saccharomyces cerevisiae as activators in biogas production from fresh market garbage. Lucrări Ştiinţifice. 2011; 56. Aragaw T, Mebeaselassie A, Amare G: Co-digestion of cattle manure with organic kitchen waste to increase biogas production using rumen fluid as inoculums. International Journal of the Physical Sciences. 2013; 8: 443–450. Tamirat A, Mebeaselassie A, Amare G: Co-digestion of cattle manure with organic kitchen waste to increase biogas production using rumen fluid as inoculums. International Journal of Physical Sciences. 2013; 8: 443–450. Tamrat A: Effects of Co-Digestion of Cattle Manure with Organic Kitchen Waste Using Rumen Fluid as Inoculums on the Rate and Amount of Biogas Yield. Haramaya university; 2012. Master’s thesis Yue ZB, Yu HQ: Anaerobic batch degradation of cattail by rumen cultures. International Journal of Environmental Pollution. 2009; 38: 299–308. Zhang H, Zhang P, Ye J, et al.: Improvement of methane production from rice straw with rumen fluid pretreatment: A feasibility study. International Biodeterioration & Biodegradation. 2016; 113: 9–16. “…there are 107,684 residents, comprising 49,893 men and 57,791 women (Aynalem et al., 2014).” These statistics are outdated. “This study aimed to evaluate the effects of ruminant fluids and S. cerevisiae isolate MUTJ0F (OR209280.1) on biogas generation from various mixed organic wastes co-digested with cattle manure. Various wastes from fruits and vegetables were gathered from the town fruit houses, leftover food from the Debre Markos University student cafeteria, fresh cattle manure (CM) from Monkorer Agroindustry Enterprise, and rumen fluids (RL) from a nearby slaughterhouse and used as activators for biogas production.” These sentences contains repititions in the next paragraph, hence should be avoided. “… the de-sized cattle manure and food waste were mixed separately with distilled water in a 1:5 (solid waste: distilled water) volume ratio …” What is the ratio between cafeteria leftovers, cattle manure, and fruit and vegetable waste? How was the solid waste measured in volume? What is the size of the filter used in filtering the rumen fluid? Some of the citations are not necessary, such as “Then, different amounts of the filtrate were added to each digester to initiate the reaction (Aurora, 1983; Genet et al., 2018).” “After calibration, the electrode was submerged in the substrate samples, rinsed with distilled water, and the pH values were recorded (Arogo et al., 2009).” In subsection 2.5.1, how was the measured empty crucible used in the formula? In subsection 2.5.2, the authors indicated the measurement of fixed solids, but it was not presented in Table 1. Authors should be consistent with the way they explain parameters in an equation, whether horizontal or vertical. The procedure for moisture content determination is the same as that of total solids, so you can indicate the formula of MC there and delete subsection 2.5.4 “After calibration, the electrode was submerged in the substrate samples, rinsed with distilled water, and the pH values were recorded (Arogo et al., 2009).” Are the authors sure this was the sequence used in measuring the pH value? “The first factor was the dosage of rumen liquid i.e. 0 ml/100 g, 25 ml/100 g, 50 ml/100 g, 75 ml/100 g, and 100 ml/100 g), …” Why is the 150 ml rumen fluid not stated here but in Table 2? “The study was conducted at room temperature (30°C).” How did the authors maintain this temperature in the room? “Feed stock was mixed with distilled water to obtain approximately 8% of TS suspension.” Is this information similar to the “1:5 (solid waste: distilled water) volume ratio” indicated in Section 2.3? In section 2.6, the retention time or duration of the anaerobic digestion was not mentioned. Section 2.7 should come before Section 2.6. Also, the authors need to provide an image of the three plastic bottles set up for anaerobic digestion, since it was constructed for this study. What is the working volume of the digester? Sections 2.8 and 2.7 can be merged. Was the biogas measured daily or at what interval? In Section 3.1, what does the use of “mixed wastes” mean? For example: “The pH of the mixed waste ranged from 6.8 to 7.6, …” Does it mean the mixture of the various feedstock or the individual feedstock? “The VS/TS ratios of the cafeteria food waste and cattle manure were 78.3% and 91.2%, respectively.” What about fruit and vegetable waste? “Cattle dung is used to accelerate biogas production by promoting bacterial growth in the digester.” Provide a reference. “Co-digestion has a positive synergistic effect by neutralizing pH, increasing buffering capacity, reducing the effects of harmful compounds, and supplying more balanced nutrients, such as vitamins, trace metals, and other substances required for microbial growth (Fang, 2010; Aragaw et al., 2013; Jianzheng et al., 2011).” The relevance of this information in the Results and Discussion Section is not clear since it has already been stated in the Introduction Section with different references cited. “The 12 methane production treatments varied significantly, reflecting variations in the mixed-waste composition and inoculum dosage.” Are the authors sure that the variations in the mixed-waste composition may have caused the significant difference in biogas production? “This implies that the high anaerobic bacterial concentration in rumen fluid efficiently breaks down organic substrates from mixed waste.” Is it only the rumen fluid that increased biogas production? “According to the results, the maximum amount of biogas (6900.3 ml) …” Normally, biogas production is evaluated using methane yield (mL/gVS or mL/gTS) to enable comparison with other published studies. In Table 2, why was only rumen fluid not investigated? The heading “Methane (mean ± SD)”, did the authors measure methane or biogas? In Figure 1, please plot pH on the y-axis and days on the x-axis. Why plot only T10? It is better to plot all the treatments. Why was the pH measured up to 80 days, but the anaerobic digestion was observed for 60 days, as indicated in Figure 2, the Abstract and Section 3.4? Figure 2A and B should come before Figure 1. These sentences could have been verified by plotting the daily biogas production. “This low pH value was permitted very little methanogenic bacterial activity …” “This also brings the pH closer to neutral, which makes it easier for methanogenic microbes to multiply and produce methane. Methanogenic bacteria the optimum pH values between 6.8 and 7.2 (Anunputtikul and Rodtong, 2004; Budiyono et al., 2010).” The authors should clarify which equipment was used for the combustion test, either “a lit matchstick” as indicated in Section 2.9 or “a Bunsen burner” as indicated in Section 3.4. Also, Figure 2C was not well captured to help identify which equipment was used. In section 3.4, it is better use “Figure 2C” rather than “Figure 2”. “Biodegradation, microbial strength, and biogas generation can all be enhanced by using …” How were biodegradation and microbial strength measured in this study? Future perspectives or studies could be added to the Conclusion section. The language must be checked thoroughly. Some statements are not clear or too long and should be revised. For examples: “However, various juice house wastes are among the municipal wastes that are becoming difficult to manage.” Also, indicate why they are difficult to manage. “… food waste co-digestion under anaerobic conditions and cattle manure, …” “The strain Saccharomyces cerevisiae isolate MUTJ0F (OR209280.1) with accession number was used in this study.” “A mixture of substrates, including peels of bread, injera, spaghetti, papaya, mango, banana, and avocado was used in this study.” Which substrates? “The CM was separated …” Should it be “separated” or “spread”? “The inoculum was specific to each digester.” “The crucibles were dried, cooled to room temperature in a desiccator, and weighed again.” Should it be “dried” or “removed”? “According to Haug (1993), using data from volatile solids and an empirical equation, the organic carbon was calculated, and the organic carbon content of the sample was calculated by taking into account the volatile solids content, which was expressed as a percentage:” “The first factor was the dosage of rumen liquid i.e. 0 ml/100 g, 25 ml/100 g, 50 ml/100 g, 75 ml/100 g, and 100 ml/100 g), while the second factor was the dosage of S. cerevisiae isolate MUTJ0F i.e. 5 ml/100 g, and 10 ml/100 g) of the mixed organic waste.” What does the “factor” stands for? “… until a solution that was supersaturated formed.” “After the biogas was collected, a measurement cylinder containing the gas was carefully placed.” “… varied in the amount of composition owing to the variability in the composition of the samples of the different substrates.” “Among with significant potential for biogas production (Panahi et al., 2022; Zeng et al., 2022). All the substrates, cafeteria leftover food had the highest organic carbon content (41.6%), …” “This makes it a high-energy feedstock substrates were considered acceptable for anaerobic digestion because all the mixed wastes had the highest ratio of VS to TS in Table 1.” “This low pH value was permitted very little methanogenic bacterial activity …” Avoid stating the mesophilic condition twice in the abstract section. "physic-chemical properties" = "physicochemical properties" Use SI units. For example “hrs” = “h” These sentences should be merged to improve clarity. “The brine solution was then acidified by adding two to three drops of sulfuric acid by the method of (Elijah et al., 2009). Finally, this formed solution was contained in the second chamber.” “These values indicated that there were sufficient biodegradable volatile solids in each substrate to enable effective anaerobic digestion (Li et al., 2013; Pagliacci et al., 2016; Kawai et al., 2014). The amount of biodegradable organic matter in the dry matter content should be between 70 and 95% for efficient biogas production (Wu et al., 2021; Buffiere et al., 2006).” “This increase is due to ammonification processes …” Since this sentence was not verified by measurement, it would be better to express it as “This increase may be due to ammonification processes …” Is the work clearly and accurately presented and does it cite the current literature? Partly Is the study design appropriate and is the work technically sound? Partly Are sufficient details of methods and analysis provided to allow replication by others? Partly If applicable, is the statistical analysis and its interpretation appropriate? Not applicable Are all the source data underlying the results available to ensure full reproducibility? Partly Are the conclusions drawn adequately supported by the results? Partly Competing Interests No competing interests were disclosed. Reviewer Expertise Anaerobic digestion, pyrolysis and gasification I confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard, however I have significant reservations, as outlined above. reply Respond to this report Responses (1) Author Response 18 Apr 2026 Mulugeta Fentahun, College of Natural and Computational Sciences, Department of Biology, Debre Markos University, Debre Markos, 269, Ethiopia Mulugeta Fentahun Thanks to Dr. Dominic Yellezuome for your constructive comments. We conducted a number of changes. I greatly appreciate your feedback and suggestions. Please find my responses to your comments below: Comment 1 : There is too much information in the methods paragraph in the Abstract Section, in which some of it can not be found in the Methods Section. Some of them are not important to be in the Abstract Section. Response: Thank you for this helpful comment. We have revised the abstract according to your feedback. Refer to the abstract part in the updated manuscript Comment 2 : These sentences are similar; hence should be merged. Response : Thank you for this observation. We have revised the manuscript by merging similar sentences to improve readability and eliminate redundancy (Refer to the revised manuscript). Comment 3 : Authors cited lists of 3 or more references without proper justification of the relevance of each one. Hence, avoid citing more than two references and outdated references (Refer to the revised manuscript). Response: Thank you for this constructive comment. We have revised the manuscript by limiting citations and avoiding outdated references (Refer to the revised manuscript). Comment 4 : Although there is information on biogas production from organic waste or single inoculums (Alemayehu, 2014; Abayneh et al., 2014; Hammad et al., 2018; Earnest and Singh, 2013) but there is limited reports on methods for increasing biogas generation, particularly those that use rumen fluid and S. cerevisiae.” This paper is not clear in the description of the scientific novelty in comparison to related previously published and referenced in your work below: Response: Thank you for this important comment. We have revised the manuscript based on your comments (Refer to the revised manuscript). Comment 5 : There are 107,684 residents, comprising 49,893 men and 57,791 women (Aynalem et al., 2014).” These statistics are outdated. Response: Thank you for pointing this out. We have updated the demographic information using the most recent and reliable data sources to ensure accuracy and relevance in the manuscript. Comment 6 : This study aimed to evaluate the effects of ruminant fluids and S. cerevisiae isolate MUTJ0F (OR209280.1) on biogas generation from various mixed organic wastes co-digested with cattle manure. Various wastes from fruits and vegetables were gathered from the town fruit houses, leftover food from the Debre Markos University student cafeteria, fresh cattle manure (CM) from Monkorer Agroindustry Enterprise, and rumen fluids (RL) from a nearby slaughterhouse and used as activators for biogas production.” These sentences contains repititions in the next paragraph, hence should be avoided. Response: Thank you for this comment. We have revised the manuscript to remove the repetitive sentences . Comment 7 : The de-sized cattle manure and food waste were mixed separately with distilled water in a 1:5 (solid waste: distilled water) volume ratio …” What is the ratio between cafeteria leftovers, cattle manure, and fruit and vegetable waste? How was the solid waste measured in volume?. Response: Thank you for this insightful comment. We have revised the manuscript to clarify the mixing ratios and measurement procedures (Refer to the revised manuscript). Comment 8 : What is the size of the filter used in filtering the rumen fluid? Response: Thank you for this comment. We have revised the manuscript based on your comments (Refer to the revised manuscript). Comment 9 : Some of the citations are not necessary, such as ○ “Then, different amounts of the filtrate were added to each digester to initiate the reaction (Aurora, 1983; Genet et al., 2018).” “After calibration, the electrode was submerged in the substrate samples, rinsed with distilled water, and the pH values were recorded (Arogo et al., 2009). Response: Thank you for this comment. We have revised the manuscript by removing unnecessary citations. Comment 10 : In subsection 2.5.1, how was the measured empty crucible used in the formula? Response: Thank you for this comment. We have revised and clarified the method in subsection 2.5.1 based on your feedback (Refer to the updated manuscript). Comment 11 : In subsection 2.5.2, the authors indicated the measurement of fixed solids, but it was not presented in Table 1. Response: Thank you for this comment. We have revised the manuscript by include the measured fixed solids in Table 1 or clarified in the manuscript. Comment 12 : Authors should be consistent with the way they explain parameters in an equation, whether horizontal or vertical. Response: Thank you for this comment. We have revised the manuscript to present all equation parameters consistently in a horizontal format (Refer to the updated manuscript). Comment 13 : The procedure for moisture content determination is the same as that of total solids, so you can indicate the formula of MC there and delete subsection 2.5.4. Response: Thank you for this suggestion. We have removed subsection 2.5.4 and included the formula for moisture content within the total solids section, streamlining the methods and avoiding redundancy. Comment 14 : After calibration, the electrode was submerged in the substrate samples, rinsed with distilled water, and the pH values were recorded (Arogo et al., 2009).” Are the authors sure this was the sequence used in measuring the pH value? Response: Thank you for this comment. We have revised the correct sequence and refer to the updated the manuscript. Comment 15 : The first factor was the dosage of rumen liquid i.e. 0 ml/100 g, 25 ml/100 g, 50 ml/100 g, 75 ml/100 g, and 100 ml/100 g), …” Why is the 150 ml rumen fluid not stated here but in Table 2? Response: Thank you for this comment. We have revised the manuscript to include the 150 ml/100 g rumen fluid level in the text for consistency with Table 2. Comment 16 : The study was conducted at room temperature (30°C).” How did the authors maintain this temperature in the room? Response: Thank you for this comment. We have clarified in the manuscript that the study was conducted under mesophilic conditions at 38 °C using an oven, and the text has been corrected to fix the typographical error. Comment 17 : Feed stock was mixed with distilled water to obtain approximately 8% of TS suspension.” Is this information similar to the “1:5 (solid waste: distilled water) volume ratio” indicated in section 2.3? Response: Thank you for this comment. We have clarified in the manuscript that the 8% TS suspension is distinct from the 1:5 ratio described in section 2.3. Comment 18 : In section 2.6, the retention time or duration of the anaerobic digestion was not mentioned. Response: Thank you for this comment. We have revised section 2.7 based on your comment. Comment 19 : Section 2.7 should come before Section 2.6. Also, the authors need to provide an image of the three plastic bottles set up for anaerobic digestion, since it was constructed for this study. What is the working volume of the digester? Response: Thank you for this comment. We have revised the manuscript and we have added an image of the three plastic bottle digesters used in this study and the working volume of digester. Comment 20 : Sections 2.8 and 2.7 can be merged. Was the biogas measured daily or at what interval? Response: Thank you for this comment. We have merged sections 2.7 and 2.8 for clarity and streamlined the methods. We also clarified that biogas production was measured at regular daily intervals throughout the experiment. Comment 21 : In section 3.1, what does the use of “mixed wastes” mean? For example: “The pH of the mixed waste ranged from 6.8 to 7.6, …” Does it mean the mixture of the various feedstock or the individual feedstock? Response: Thank you for this comment. We have clarified in section 3.1 based on your comments. Refer to the results and discussion parts in the updated manuscript . Comment 22 : The VS/TS ratios of the cafeteria food waste and cattle manure were 78.3% and 91.2%, respectively.” What about fruit and vegetable waste? Response: Thank you for this comment. We have revised the manuscript based on your comments. Comment 23 : Cattle dung is used to accelerate biogas production by promoting bacterial growth in the digester.” Provide a reference. Response: Thank you for this comment. We have added a relevant reference. Refer to the updated manuscript. Comment 24 : Co-digestion has a positive synergistic effect by neutralizing pH, increasing buffering capacity, reducing the effects of harmful compounds, and supplying more balanced nutrients, such as vitamins, trace metals, and other substances required for microbial growth (Fang, 2010; Aragaw et al., 2013; Jianzheng et al., 2011).” The relevance of this information in the Results and Discussion Section is not clear since it has already been stated in the Introduction Section with different references cited. Response: Thank you for this comment. We have revised the results and discussion section and remove the redundant statement. Comment 25 : The 12 methane production treatments varied significantly, reflecting variations in the mixed waste composition and inoculum dosage.” Are the authors sure that the variations in the mixed-waste composition may have caused the significant difference in biogas production? Response: Thank you for this comment. We have revised the manuscript based on your comments. Comment 26 : This implies that the high anaerobic bacterial concentration in rumen fluid efficiently breaks down organic substrates from mixed waste.” Is it only the rumen fluid that increased biogas production? Response: Thank you for this comment. We have revised the manuscript based on your comments. Comment 27 : According to the results, the maximum amount of biogas (6900.3 ml) …” Normally, biogas production is evaluated using methane yield (mL/gVS or mL/gTS) to enable comparison with other published studies. Response: Thank you for this comment. We clarify that our results were reported based on the total biogas volume (mL), and we did not measure total solids or volatile solids after digestion (methane yield (mL/gVS or mL/gTS) in this experiment. Comment 28 : In Table 2, why was only rumen fluid not investigated? The heading “Methane (mean ± SD)”, did the authors measure methane or biogas? Response: Thank you for this comment. We have clarified that Table 2 presents biogas production and not methane specifically; the heading has been corrected accordingly. Additionally, rumen fluid alone was not included as a treatment because the study focused on co-digestion with mixed wastes to evaluate the synergistic effect of dual inoculants. Comment 29 : In Figure 1, please plot pH on the y-axis and days on the x-axis. Why plot only T10? It is better to plot all the treatments. Why was the pH measured up to 80 days, but the anaerobic digestion was observed for 60 days, as indicated in Figure 2, the Abstract and Section 3.4? Response: Thank you for this comment. We have revised Figure 1 to plot pH on the y-axis against days on the x-axis and included all treatments for clarity. Additionally, we clarified that pH measurements were continued up to 80 days to monitor post-digestion stabilization, even though the main anaerobic digestion period lasted 60 days. Comment 30 : Figure 2A and B should come before Figure 1. Response: Thank you for this comment. We have revised the manuscript so that Figures 2A and 2B now appear before Figure 1. Comment 31 : These sentences could have been verified by plotting the daily biogas production. Response: Thank you for this comment. We have revised the sentences in the updated manuscript. Comment 32 : The authors should clarify which equipment was used for the combustion test, either “a lit matchstick” as indicated in Section 2.9 or “a Bunsen burner” as indicated in Section 3.4. Also, Figure 2C was not well captured to help identify which equipment was used. In section 3.4, it is better use “Figure 2C” rather than “Figure 2” Response: Thank you for this comment. We have revised the manuscript in the method section and the results and discussion sections. Comment 33 : Biodegradation, microbial strength, and biogas generation can all be enhanced by using” How were biodegradation and microbial strength measured in this study Response: Thank you for this comment. We have revised the manuscript based on your comments. Comment 34 : Future perspectives or studies could be added to the Conclusion section. Response: Thank you for this comment. We have revised the conclusion section to include future perspectives. Comment 35 : The language must be checked thoroughly. Response: Thank you for this comment. We have carefully reviewed and revised the manuscript to improve language, grammar, and clarity throughout the text. Thank you once again for your valuable feedback. Your comments have significantly contributed to improving the quality of this paper. Best regards, Authors View more View less Competing Interests No competing interests were disclosed. reply Respond Report a concern Yellezuome D. Peer Review Report For: Enhancing biogas production from co-digestion of organic wastes mixed with cattle manure using rumen fluid and Saccharomyces cerevisiae isolate MUTJ0F [version 1; peer review: 1 not approved] . F1000Research 2025, 14 :199 ( https://doi.org/10.5256/f1000research.193404.r445154) NOTE: it is important to ensure the information in square brackets after the title is included in this citation. The direct URL for this report is: https://f1000research.com/articles/14-199/v2#referee-response-445154 keyboard_arrow_left Back to all reports Reviewer Report 0 Views copyright © 2025 Emmanuel J. This is an open access peer review report distributed under the terms of the Creative Commons Attribution License , which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. 19 Aug 2025 | for Version 1 Jovine Kamuhabwa Emmanuel , Chemistry, University of Dar es Salaam, Dar es Salaam, Dar es Salaam, Tanzania 0 Views copyright © 2025 Emmanuel J. This is an open access peer review report distributed under the terms of the Creative Commons Attribution License , which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. format_quote Cite this report speaker_notes Responses (1) Not Approved info_outline Alongside their report, reviewers assign a status to the article: Approved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested Approved with reservations A number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit. Not approved Fundamental flaws in the paper seriously undermine the findings and conclusions Title The title captures the content of the current work. Abstract The abstract has no information on the study, methodology used and key findings and recommendation based on findings. Authors must re-write this to reflect the current work. Other comments are found in the main document. Introduction The section has not been written properly and not easy to follow and understand. Authors must write this in a scholarly manner to reflect an article for publication. The novelty is not well feature thus, authors must reveal the novelty of the work. Other comments are found in the main document. Materials and methods Proper details and step after step must be provided for readers to follow. Authors need to take time and re-do it properly. Other comments are found in the main document. Results and discussion Results are not well organized and discussed with previous work. Authors need to re-think on how they can organize these data for publication. Other comments are found in the main document. Conclusions and recommendations This section is not well written thus, must be re-written for clarity. Only key findings and recommendation must be presented and not otherwise. Other comments are found in the main document. General comment The manuscript must benefit from a serious English Language editing before being indexed. The manuscript must be enriched with up-to-date citations which are available in the literature. Other comments are found in the main document. Is the work clearly and accurately presented and does it cite the current literature? No Is the study design appropriate and is the work technically sound? Yes Are sufficient details of methods and analysis provided to allow replication by others? Partly If applicable, is the statistical analysis and its interpretation appropriate? Not applicable Are all the source data underlying the results available to ensure full reproducibility? Partly Are the conclusions drawn adequately supported by the results? Partly Competing Interests No competing interests were disclosed. Reviewer Expertise Catalysis, materials science and bioenergy I confirm that I have read this submission and believe that I have an appropriate level of expertise to state that I do not consider it to be of an acceptable scientific standard, for reasons outlined above. reply Respond to this report Responses (1) Author Response 29 Dec 2025 Mulugeta Fentahun, College of Natural and Computational Sciences, Department of Biology, Debre Markos University, Debre Markos, 269, Ethiopia Mulugeta Fentahun Dr. Jovine Kamuhabwa Emmanuel thank you very much for your insightful comments and the time you dedicated to review. We greatly appreciate your feedback and suggestions. We provide a detailed response to each comment, along with the corresponding changes made in the revised manuscript below: Title: Comment : The title captures the content of the current work. Response : Thank you for your positive feedback. Abstract Revision: Comment : The abstract has no information on the study, methodology used, and key findings and recommendations based on the findings. Authors must re-write this to reflect the current work. Response : Thank you for your insightful comment. We have thoroughly revised the abstract part in response to your suggestions (see the revised manuscript). Introduction Revision: Comment : The section has not been written properly and is not easy to follow and understand. Authors must write this in a scholarly manner to reflect an article for publication. The novelty is not well featured; thus, authors must reveal the novelty of the work. Response : We sincerely thank the reviewer for your constructive feedback. We have thoroughly revised the introduction section to improve clarity, coherence, and scholarly manner. Additionally, we have clearly highlighted the novelty of the study by emphasizing its unique contributions in the revised manuscript (refer to the revised manuscript). Methodology Revision: Comment : Proper details and step after step must be provided for readers to follow. Authors need to take time and re-do it properly. Response: Thank you for this valuable observation. We have substantially revised the materials and methods section to improve clarity, completeness, and reproducibility. The updated version now provides detailed, step-by-step procedures for the experimental work, enabling readers to follow the methodology accurately and enhancing the scientific rigor of the manuscript (see the revised manuscript). Results and Discussion Revision: Comment : Results are not well organized and discussed with previous work. Authors need to re-think on how they can organize these data for publication. Response: We sincerely thank the reviewer for your constructive feedback. We have carefully revised the results and discussion section to enhance clarity and coherence. The revised version enriched with up-to-date citations (recent literature) and emphasizes the key findings. Please refer to the updated section in the revised manuscript. Conclusions and recommendations Revision: Comment : This section is not well written thus, must be re-written for clarity. Only key findings and recommendation must be presented and not otherwise . Response: Thank you for your insightful comment. We have revised the conclusions section to clearly highlight the study’s key findings and recommendation. Please see the updated section in the revised manuscript. General comment: Comment : The manuscript must benefit from a serious English Language editing before being indexed. Response: We sincerely thank the reviewer for the valuable suggestion regarding the language quality of our manuscript. We have thoroughly revised the manuscript to improve English clarity, grammar, and readability. Comment : The manuscript must be enriched with up-to-date citations which are available in the literature. Response: Thank you for your comments. We have updated the manuscript by incorporating recent literature, as suggested (see the revised manuscript). Thank you once again for your valuable feedback. Your comments have significantly contributed to improving the quality of this paper. Best regards, Authors View more View less Competing Interests No competing interests were disclosed. reply Respond Report a concern Emmanuel JK. Peer Review Report For: Enhancing biogas production from co-digestion of organic wastes mixed with cattle manure using rumen fluid and Saccharomyces cerevisiae isolate MUTJ0F [version 1; peer review: 1 not approved] . F1000Research 2025, 14 :199 ( https://doi.org/10.5256/f1000research.177603.r401840) NOTE: it is important to ensure the information in square brackets after the title is included in this citation. The direct URL for this report is: https://f1000research.com/articles/14-199/v1#referee-response-401840 Alongside their report, reviewers assign a status to the article: Approved - the paper is scientifically sound in its current form and only minor, if any, improvements are suggested Approved with reservations - A number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit. 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