Effect of biological treatment of Pig dung using Pseudomonas fluorescens on biogas production

Introduction

Pigs play a crucial role as versatile livestock, providing an affordable source of protein and enhancing livelihoods in developing nations such as India. Since the 1970s, the global pig population has doubled, with forecasts suggesting a further increase of 25% by 2030 (Hu et al., 2021) [4] . However, swine farming generates significant waste; in India, the pig population produces approximately 10.1 million tons of excreta annually (Kaur et al., 2017) [5] . Importantly, pig manure is richer in nutrients compared to cow dung (Makara et al., 2019) [7] . Despite its potential as an organic fertilizer and renewable energy source, social taboos and poor management practices hinder the effective utilization of pig waste, leading to environmental pollution and health risks (Ramírez et al., 2020) [12] . Utilizing pig manure for biogas production can alleviate some of these challenges, helping to reduce reliance on nonrenewable resources (Hamelinck et al., 2005) [3] while simultaneously addressing issues such as environmental degradation and energy shortages. Therefore, this research aims to investigate the impact of biologically treating pig dung with Pseudomonas fluorescens on biogas production.

Materials and Methods

Work plan

In this study, we employed fresh pig dung sourced from uniform age groups of animals as the substrate. To optimize biogas production, the dung was analysed for proximate principles as detailed in Table .1 and standardized to contain 10% dry matter before its introduction into the digester to enhance the efficiency and yield of biogas generation. A slurry with a dry matter content of 10% was prepared by adding the appropriate amount of water to the substrate. To this mixture, inoculum of Pseudomonas fluorescens was introduced at concentrations of 3% and 6% as detailed in Table 2. The treatment groups were designated as P0 (control), P6 (3% inoculum), and P6 (6% inoculum). The prepared syringes containing 20 ml of this slurry were then incubated in a water bath at 30 °C, the optimal temperature for the growth of Pseudomonas fluorescens. Biogas production was observed over a 50-day period, with three replicates for each treatment group. Every 10 days, six syringes from each group were retrieved for fiber analysis and bacterial quantification using qPCR techniques.

Statistical analysis

Data analysis was conducted using SPSS software, version 16.0. A one-way ANOVA was performed to evaluate differences between treatment groups, with significance determined at the 5% level as per Snedecor and Cochran (1994). The Duncan Multiple Range Test (DMRT) was used for further comparison of significant differences. Biogas output increased over time for all groups, with P6 consistently producing the highest amounts. In the 0-10 days interval, the values ranged from 38.68 to 47.81 ml/gm VS, with no significant difference among treatments (P = 0.6867). From day 11 onwards, P6 showed significantly higher production (151.54, 221.39, 277.40, 322.06) compared to P0 and P3, which had similar values within each interval. The differences among treatments were statistically significant (p < 0.05) from day 11 onwards and persisted till day 50. Over all Cumulative Biogas production increased gradually over time for all treatments, with 6% inoculum (P6) generating 12.51% and 3% inoculum (P3) generating 11.93% more compared to control.

Results and Discussion

Discussion

The impact of biological treatment with Pseudomonas fluorescens on cumulative biogas production from pig dung was evident throughout the 50-day experimental period. During the initial phase (days 0-10), cumulative gas yields did not show significant differences (p > 0.05) among the control, 3%, and 6% culture-treated groups, which likely reflects a short adaptation (lag) phase for the methanogenic bacteria, resulting in a slow start to hydrolytic processes. However, starting from days 11-20, the 6% treatment group (P6) demonstrated significantly higher cumulative gas production compared to the control (p < 0.05), and this trend continued through day 50, with the 6% treatment consistently outperforming both the control and the 3% treatment groups. These results suggest that a 6% inoculum level of P. fluorescens provides a reproducible and dose-dependent increase in biogas yield, whereas the 3% level failed to achieve a substantial improvement. The enhancement observed at the 6% level may be attributed to improved hydrolysis of readily degradable organic compounds, such as simple sugars, soluble carbohydrates, proteins, and lipids, alongside partial lignocellulose degradation. Pseudomonas fluorescens is known to produce DyP-type peroxidases (Rahmanpour and Bugg, 2014) [11] , and some strains of Pseudomonas exhibit endo-β-1,4-glucanase and exo-β-1,4-glucanase activities (Pathak et al., 2016) [9] , which are extracellular enzymes with potential lignin-degrading and cellulose-acting capacities. Thus, the enzyme secretion by P. fluorescens likely contributes to the degradation of the lignincarbohydrate complex, enhancing the accessibility of cellulose and hemicellulose. The lack of significant enhancement at the 3% inoculum level suggests that a critical threshold of bacterial biomass or enzyme activity was not reached, limiting substrate modification and biogas yield. Similar findings were noted by Bruni et al. (2010) [1] , who reported that suboptimal dosing of microbial cultures had negligible effects on methane production. Similar findings have been reported by Mustafa et al. (2016) [8] , who observed a 120% increase in biogas yield from unprocessed rice straw, and Fugol et al. (2023) [2] , who noted a 62% increase in biogas production from maize silage treated with Pseudomonas spp. In other studies, Phutela et al. (2011) [10] found a 53.8% increase in biogas yield compared to controls, while Tuesorn et al. (2013) [13] reported a 55% increase. Kaur et al. (2014) [6] also demonstrated a 30.7% rise in biogas yield compared to untreated paddy straw in their biological treatment experiments with various substrates.

Conclusion

The present study suggests that inoculation of sheep dung slurry with Pseudomonas fluorescens showed significant enhancement in biogas yield at 6% inoculum compared to 3% and control.

Acknowledgement

AcknowledgementWe are grateful to our Dean, Veterinary college Bengaluru, and KVAFSU, Bidar, Karnataka for providing necessary facilities and funding to carry out this research work.

References

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