Lack of inhibitory effects of 1-Octyne and PTIO on ammonia oxidizers, nitrite oxidizers, and nitrate formation in acidic paddy soils

The feasibility of employing double inhibitors consisting of PTIO and 1-octyne with the 15 N pool dilution technique to distinguish comammox process from archaeal and bacterial nitrification remains to be confirmed. As such, we carried out a nitrification inhibition experiment on acidic paddy soils at a pH of 5.0, 5.3, and 5.4 to investigate the effects of 1-octyne and PTIO on nitrate (NO3-) formation, ammonia (NH3)-oxidizing archaea (AOA) and bacteria (AOB), comammox Nitrospira, and nitrite (NO2-)-oxidizing bacteria (NOB) under urea fertilized- and unfertilized-conditions. We observed that 1-Octyne stimulated the growth of comammox Nitrospira clades A and B, and Nitrobacter-like-like NOB and Nitrospira-like-like NOB without inhibiting AOB growth. This observation indicates that 1-Octyne has the potential to promote comammox process and NO2- oxidization, and its usage may lead to an underestimation of AOB activity. Unexpectedly, we found that PTIO was incapable of causing a rapid inhibition of AOA growth and NO3- formation. In the short term, PTIO was usually not detrimental to the growth of comammox Nitrospira clades A and B, AOB, and Nitrobacter-like-like NOB. Instead, increased growth of AOB and Nitrospira-like-like NOB were observed following its application. Likely due to increased bacterial nitrification and NO2- oxidation and the reaction of PTIO with nitric oxide (NO) produced by non-nitrification processes, increased NO3- formation with PTIO was observable in acidic soils with varying N statuses. These findings highlight the need to develop novel and more efficient inhibition strategies to actualize rate distinction among AOA, AOB, comammox Nitrospira in acidic soils.