Reduced Cytochrome c: Exploring Its Promoting Function in Synergistic Denitrification by Pseudomonas sp. JM-7 and Thiobacillus denitrificans

Excessive accumulation of nitrate in natural ecosystems can lead to eutrophication of water bodies, resulting in environmental impacts such as algal blooms and red tides that pose serious threats to environmental and human health. Autotrophic denitrification is a resource-efficient biological nitrogen removal technology, but the slow growth and low electron transfer efficiency of autotrophic denitrifiers limit their practical application. In this study, the heterotrophic denitrifying bacterium Pseudomonas sp. JM-7 (P. JM-7) was used in mixed culture with the autotrophic denitrifying Thiobacillus denitrificans ATCC 25259 to form a heterotrophic-autotrophic synergistic denitrification process (mixPT) to enhance electron transfer and improve the denitrification efficiency. Our results for an initial nitrate concentration of 430 mg/L showed that P. JM-7 reduced approximately 210 mg/L nitrate (48.8 %) in 90 h, with 1 g/L yeast extract powder (YEP) as the electron donor, while Thiobacillus denitrificans barely removed nitrate. In contrast, the mixPT system reduced approximately 360 mg/L nitrate (83.3 %), nearly 1.7 times greater than P. JM-7 alone. The mixPT culture also exhibited excellent denitrification in low carbon waters (10 mg/L), and removed 40 mg/L nitrate completely within 120 h. Cytochrome c produced by Pseudomonas sp. JM-7, which is an excellent electron transfer mediator for denitrification by Thiobacillus denitrificans, played a crucial role in the heterotrophic-autotrophic syntrophic denitrification. The adsorption effect of Pseudomonas sp. JM-7 shortened the contact distance between the autotrophic denitrifying bacteria and cytochrome c, which made it easy for cytochrome c to transfer the electrons to the autotrophic bacteria directly, thereby reducing the loss of electrons in the process of transfer, and improved the electron utilization efficiency. These findings have demonstrated the potential applications of mixPT system in controlling eutrophic waters and propose cytochrome c of Pseudomonas sp. JM-7 could serve as a novel and high-quality electron transfer mediator. Furthermore, this study have overcome the restriction of low denitrification efficiency exhibited by heterotrophic denitrifying bacteria Pseudomonas sp. JM-7 in low-carbon environments and expanded the utilisation of Pseudomonas sp. JM-7 in low carbon environments.