Effects Of Different Carbon And Nitrogen Ratio Of Substrate Additionson Soil Ammonia Oxidizers And Their Underlying Mechanisms

Nitrification is a key process in the nitrogen(N)cycle that critically modulates plant N utilization,ecosystem N retention and environmental effects of reactive N.Nitrification includes two steps,with ammonium being oxidized to nitrite and nitrite to nitrate.Microbial ammonia oxidation,the first and rate-limiting step of the nitrification process,is primarily controlled by ammonia oxidizing archaea(AOA)and ammonia oxidizing bacteria(AOB).Soil factors,particularly soil pH and ammonium(NH4+),have been proposed as primary drivers that control AOA and AOB abundances.However,field results have been highly variable,and there is still lack of a unifying framework describing the patterns of global AOA and AOB distribution.In this study,I first performed a meta-analysis to explore global patterns of AOA and AOB abundance and identify the potential drivers that dominate these patterns.The results obtained showed that soil carbon to nitrogen(C:N)ratio explained the most variance of AOA and AOB distribution.Then,a microcosm experiment was designed to assess how different C:N ratios of substrate affect AOA and AOB in two agricultural soils with long-term contrasting fertilization histories.In addition,three independent but complementary experiments were conducted to further investigate how biotic interactions control AOA and AOB abundances.The main results are listed below.1.A database was constructed through an extensive searchof published data from AOA and AOB related studies(Web of Science and Google Scholar before 2016)to examine the global distribution of AOA and AOB and the major drivers.Results from the meta-analysis showed that the mean AOA abundance was almost one magnitude higher than AOB across all ecosystems.While ammonium concentrations and soil C:N ratios were negatively related with AOA and AOB abundances,soil pH was positively related to them.AOA abundance nonlinearly increased with increasing total organic C,whereas no relationship between AOB and total organic C was detected.Results from the SEM analysis showed that soil C:N ratios explained the most of variance in AOA and AOB abundances,and the soil pH exhibited the secondary effect.Based on these results,a conceptual model was proposed in which biotic interactions among plants,heterotrophic microbes and ammonia oxidizers for substrate N predominantly control AOA and AOB abundances.In this model,the relative N availability rather than the absolute N content was most related to the abundance of AOA and AOB.2.To examine how different C:N ratios of substrates affect AOA and AOB,an incubation study was carried out to quantify the effects of differing carbon(cellulose)and nitrogen(ammonium sulfate)additions on AOA and AOB abundances and their community structure.Soil samples were collected from Qiyang Red Soil Experimental Station of Chinese Academy of Sciences,Qiyang,Hunan Province under two fertilization regimes,one with organic manure(OM)and another with mineral fertilizer(NPK).The results showed that the C:N ratio had significant effects on AOA and AOB abundances.Although the AOA and AOB abundances in the control with no substrate additions remained relatively stable over the three-month incubation period,they rapidly decreased with the increasing of C:N ratios in both soils.Both AOA and AOB decreased more significantly at high than low N addition in the presence of cellulose.The results of 454-pyrosequencing analysis,targeting amoA gene,revealed that that C:N ratio had significant impacts on the composition of the AOB but not AOA community.3.To further assess the impact of microbial interactions on AOA and AOB dynamics,different competition intensities were created through adding different amounts and ratios of organic C(cellulose)and mineral N[(NH4)2SO4].The experiment used the four soil samples from three different regions,including a pine forest soil(PINE),a vegetable field(VF),two fertilized soils that either received chemical fertilizers(NPK)or organic manures(OM)from Qiyang Red Soil Experimental Station of Chinese Academy of Sciences.Rapid decreases in AOA and AOB abundances concurred with high total microbial biomass and activities(respiration),suggesting intensive competition between ammonia oxidizers and other microbes.4.Since we observed extensive growth of Trichoderma spp.in our cellulose-amended soils,we suspect that these organisms may play a dominant role in suppressing AOA and AOB.Trichoderma aureoviride strain M-12,as a model microorganism,was used to evaluate the competitive effect on AOA and AOB.Soil samples were collected from Jiangxi red soil experimental station with two treatments,including organic manure(OM)and mineral fertilizer(NPK).The soil respiration rates were clearly separated by the addition of carbon and nitrogen(carbon and nitrogen>Carbon>Nitrogen).The highest abundance of Trichoderma occurred in the inoculated soil,and carbon addition alone promoted the growth of indigenous Trichoderma in the NPK soil,but had no effect in the OM soil.The inoculation of Trichoderma,especially in combination with the addition of carbon and nitrogen sources,reduced the abundance of AOA and AOB,indicating that Trichoderma spp.may interfere the nitrification process by inhibiting AOA and AOB.5.To further determine whether inhibitive interactions occur with metabolic compounds,we tested the effect of water extracts of soils amended with high ratios of cellulose and ammonium sulfate on the abundances of AOA and AOB.The results showed that extracts of the C:N ratio of 50 and 100 treatments significantly reduced AOA and AOB populations,but this effect abated over time.The metabolic compound addition of Trichoderma also inhibited the growth of AOA and AOB.Together,these results suggest that microbes also inhibit AOA and AOB through their secondary metabolites.In conclusion,above results provide a framework highlighting the dynamic,interactive controls of soil abiotic(carbon and nitrogen)and biotic factors over AOA and AOB.And C:N ratios function as a critical mediator.In the end,microbial interactions dominate the population density of AOA and AOB.