Nitrogen fertiliser is manufactured using the industrial Haber–Bosch process, although it is extremely energy-consuming. One sustainable alternative technology is the electrochemical promotion of biological nitrogen fixation (BNF). This study reports the promotion of BNF activity of anaerobic microbial consortia by humin, a solid-phase humic substance, at any pH, functioning as an extracellular electron mediator, to levels of 5.7–11.8 times under nitrogen-deficient conditions. This was evidenced by increased acetylene reduction activity and total nitrogen content of the consortia. Various humins from different origins promoted anaerobic BNF activity, although the degree of promotion differed. The promotion effected by humin differed from the effects of chemical reducing agents and the effects of supplemental micronutrients and vitamins. The promotion of anaerobic BNF activity by only reduced humin without any other electron donor suggested that humin did not serve as organic carbon source but as extracellular electron mediator, for electron donation to the nitrogen-fixing microorganisms. The next generation sequencing (NGS) of partial 16S rRNA genes showed the predominance of Clostridiales (Firmicutes) in the consortia. These findings suggest the effectiveness of humin as a solid-phase extracellular electron mediator for the promotion of anaerobic BNF activity, potentially to serve for the basis for a sustainable technology.

The effect of pesticides on cellulose degradation in a soil was estimated by incubation experiments under upland conditions, transitional flooded conditions from aerobic to anaerobic, and fully anaerobic flooded conditions. Pesticides examined were trichlamide [(RS)-N-(1-butoxy-2,2,2-trichIoroethyl)salicyl-amide], chlorothalonil (TPN, 2,4,5,6-tetrachloro-1,3-isophthalonitrile), quin-tozene (PCNB, pentachloronitrobenzene), and hymexazol (3-hydroxy-5-meth-ylisoxazol) as fungicides, paraquat (1,1′-dimethyl-4,4′-bipyridium dichloride), thiobencarb (benthiocarb, S-p-chlorobenzyldiethyl thiocarbamate), propanil (DCPA, 3′,4′-dichloropropionanilide), and butachlor [2-chloro-2′,6′-diethyI-N-(butoxymethyl)acetoanilide] as herbicides, and diazinon (2-isopropyl-4-niethyl-pyrimidyl-6-diethylthiophosphate) as insecticide. Trichlamide inhibited completely the cellulose degradation under the two flooded conditions, while the inhibition under upland conditions was weak. The inhibition by chlorothalonil was strong under all conditions, while quintozene and hymexazol did not display any inhibitory effect. There was a difference in the effect of fungicides between upland and flooded conditions, but not between the two flooded conditions. The inhibitory effect of the herbicides and insecticide was not observed under upland and transitional flooded conditions.

Anoxic aquifers suffer from energy limitations due to the unavailability of organic substrates, as dictated by hydrogen (H₂) for various electron-accepting processes. This deficiency often results in the accumulation of persistent organic pollutants, where bioremediation using organic compounds often leads to secondary contamination. This study involves the reductive dechlorination of pentachlorophenol (PCP) by dechlorinators that do not use H₂ directly, but rather through a reduced state of humin—a solid-phase humic substance—as the extracellular electron donor, which requires an organic donor such as formate, lactate, etc. This shortcoming was addressed by the development of an anaerobic mixed culture that was capable of reductively dechlorinating PCP using humin under autotrophic conditions induced by homoacetogenesis. Here, H₂ was used for carbon-dioxide fixation to acetate; the acetate produced was used for the reduction of humin; and consequently used for dechlorination through reduced humin. The 16SrRNA gene sequencing analysis showed Dehalobacter and Dehalobacterium as the possible dechlorinators, while Clostridium and Oxobacter were identified as the homoacetogens. Thus, this work contributes to the development of an anaerobic consortium that balanced H₂ dependency, where efficiency of humin reduction extends the applicability of anaerobic microbial remediation in aquifers through autotrophy, syntrophy, and reductive dechlorination.