| The carbon-to-nitrogen ratio(C/N)in biofloc systems is the core control factor.By adding a carbon source,the activity and populations of the heterotrophic bacterial community can be stimulated and enhanced favourably.Also,ammonia nitrogen can be absorbed and converted into bacterial protein,thereby removing the ammonia nitrogen in the water.In production practice,the focus is often on adjusting the C/N in the rearing water to an appropriate range,but the actual ammonia nitrogen control does not achieve the ideal ammonia nitrogen removal rate as well as the ammonia nitrogen conversion rate,which may cause the death of the aquaculture animal or the delay of production time.Studies have shown that heterotrophic bacteria can only play the role of ammonia nitrogen conversion when a certain population of bacterial communities are established and the community is stablized.Therefore,when the ammonia nitrogen conversion effect is not appropriate or within the expected range,it may be due to the insufficient concentration of the substrate available for heterotrophic bacteria,which result inadequate establishment of the heterotrophic bacteria community in the biofloc system.At the same time,in articles published by different scholars,it is found that the"N"in C/N refers to different"N"elements,among which the common ones are total ammonia nitrogen(TAN)and dissolved inorganic nitrogen(DIN),and total nitrogen(TN).These are often used in determining the amount of carbon source to be added,and researchers believe that the organic matter content in the environment is positively correlated with the number of heterotrophic microorganisms.In addition,biological flocs can not only play a role in water quality treatment,but the content of crude protein in biological flocs is also considerable,which is a potential protein feed;bioflocs serve as a carrier for many microbial probiotics in aquaculture water,which is useful for breeding.Disease prevention of the subject can also play a role.This experiment mainly focused on the influence of C/N ratio on ammonia nitrogen conversion rate,microbial community structure,and functional marker genes related to nitrogen conversion enzymes.And just as important,different types of carbon and nitrogen sources are used to inoculate and enhance bioflocs under different carbon source addition strategies,and compare the ammonia nitrogen conversion effect of the flocs,the nutritional components of the flocs,and the influence on the number of vibrio and the number of heterotrophic bacteria.The functional development and utilization of flocculation provide a theoretical basis.The experiment is mainly divided into the following three parts:1.Effects of different concentrations of carbon and nitrogen on ammonia nitrogen conversion rate and microbial community structure of bioflocsThis aspect of the study investigated the effects of different concentrations of carbon and nitrogen sources on the water quality treatment capacity and the composition of the flocculent bacteria under the same carbon-to-nitrogen ratio of bioflocs.The experiment maintained the C/N ratio of the three groups at 20 and the concentration gradient of carbon and nitrogen sources for the groups were respectively set as:group A=200 mg/L:10 mg/L,group B=100 mg/L:5 mg/L,group C=50 mg/L:2.5 mg/L.Also,the influence of ammonia nitrogen conversion rate,bacterial community structure,biofloc composition and nitrification and denitrification function marker genes in the water were analyzed.The results showed that the removal rates of TAN in groups A,B and C were 1.89±0.02 mg TAN/g TSS·h,1.63±0.03mg TAN/g TSS·h and 1.32±0.02 mg TAN/g TSS·h,respectively.There was significant differences in the removal rate(P<0.05)recorded in this study.Within the concentration gradient range of carbon and nitrogen sources designed in the experiment,the ability of flocs to treat ammonia and nitrogen increases significantly with the increase of carbon and nitrogen concentration.Additionally,the crude protein content of the flocs in group A(29.22±0.20%)was significantly lower than that of group B(30.56±0.32%)and group C(30.83±0.20%),respectively.Further,the crude fat content in the groups were ranked as follows;group A(6.65±2.12%)>group B(5.06±0.81%)>group C(4.42±2.66%),but there is no significant difference among the groups.In essence,the microbial community structure found in this study included the following phyla;Proteobacteria,Bacteroides,Actinomycetes,Verrucomicrobia,and Planktomycetes.The dominant bacterial groups described at the genus level found in this study were;Gemmobacter.,Hydrogenophaga,and Microbacterium.With the increase of the concentration of carbon and nitrogen sources,Shannon index showed that the diversity of the bacterial community structure decreased;the number of heterotrophic bacteria increased significantly,and the total number of heterotrophic bacteria in group A was significantly higher than that in groups B and C.Groups A and B whch were characterized with higher concentrations of carbon and nitrogen sources had relatively low gene copy numbers for nitrification,but a relatively high level of gene copy numbers for denitrification.2.Effects of carbon-to-nitrogen ratio and carbon source addition to different expressions of nitrogen concentrations in the rearing water on the conversion rate of ammonia nitrogen and nutrient components of bioflocsThis experiment studied the assimilation process of bioflocs,based on the different expressions of the concentration of nitrogen including;"TN","DIN",and"TAN"which referred to the"N"in C/N,respectively.At C/N=20,we explored the rapid effect of ammonia nitrogen on bioflocs.The effect on transformation ability,nutrient composition,expression of the the functional genes of nitrification and denitrification and the total counts of heterotrophic bacteria.It was found that during the rapid conversion of ammonia nitrogen,the removal rate of TAN was 2.11±0.05mg TAN/g TSS·h in group A,2.00±0.02 mg TAN/g TSS·h in group B and 2.09±0.02mg TAN/g TSS·h in group C,respectively.Ammonia nitrogen conversion rate was rate as follows;group A>group C>group B,which revealed a significant difference in conversion rate between group A and group B(P<0.05)while there was no significant difference between group C and groups A and B(P>0.05).At the end of the experiment,each group recorded crude fat content(7.10%～8.98%),crude protein content(25.92%～26.86%),VSS content(76.76%～82.08%),and crude ash content(19.94%～23.24%)of the floc components within the recommeded ranges.Compared to the 4.12%crude fat content,31.06%crude protein content,and 65.56%VSS content of the inoculated flocs before the experiment,the crude ash content(34.44%)of the inoculated flocs was significant decreased.After the experiment,the relative content of amino acids in each group only maintained the level of inoculated floccules before the experiment with cystine,and the relative content of other amino acids decreased significantly.The reason remains to be pursued further.The relative content of the eight fatty acids decreased compared with the relative content of the inoculated bioflocs before the experiment.Stearyl carbonic acid(C18:0),octadecadienoic acid(C18:2n6t)and eicosatetraenoic acid(C20:4n6)),tetradecanoic acid(C14:0),hexadecenoic acid(C16:1n7)and octadecenoic acid(C18:1n9c,C18:1n9t)are all improved compared with the floccules before the experiment.The copy numbers of the functional marker genes(amo A,nxr B,nar G,nap A,nir S,nos Z)related to nitrification and denitrification enzymes were increased significantly compared with the copy numbers in the water before the experiment.Due to the addition of carbon sources,the total number of heterotrophic bacteria in each group increased to varying degrees at the end of the experiment,and the total number of heterotrophic bacteria in group B(2.13×10~7CFU/m L)was the highest.3.Effects of different carbon sources and the different sources of nitrogen on bioflocs inoculation and formationThis experiment studied different carbon and nitrogen sources and different carbon source addition strategies(one-time addition and 48 hours equally divided addition)to increase the biofloc.The carbon and nitrogen sources of groups A and B were feed and glucose monohydrate.The carbon and nitrogen sources of group C and D were ammonium chloride and dextrose monohydrate,the nitrogen sources of group E was the feed,the nitrogen sources of group F was ammonium chloride,and the carbon sources of groups E and F are polyhydroxybutyrate(PHB).In terms of carbon source addition strategy,groups B and D were added equally 4 times within 48 hours.The effects of total suspended solids(TSS)production in each group,including the respective crude protein,crude fat,amino acids,fatty acids and other nutritional components in the bioflocs,as well as the total number of heterotrophic bacteria and vibrio in the water were also evealuated.It was found that,by comparing the nitrification and assimilation of the bioflocs after the flocs increased in concentration the nitrification function was enhanced when it was exposed to a salinity shock at 10.The results showed that adding carbon source at one time and adding carbon sources in batches during the enhancement of the flocculation process only affects the peak of nitrite nitrogen at the expansion stage.Essentially,by adding the carbon source group(groups A and C)to the batches at one time,the addition of carbon source groups(groups B and D)recorded a high nitrous nitrogen peak and the different carbon and nitrogen sources have a significant impact on the amount of bioflocs.The addition of carbon sources in feed(groups A and B)generateed a TSS values which were significantly higher than the others.It was also observed that in the PHB group which was with no carbon source released(group E),in the case of only feed,the final TSS was still higher than the ammonium chloride and glucose group.In terms of biofloc components,the different nitrogen sources and carbon sources affected the nutritional components of flocs.The results show that compared with the feed group,ammonium chloride as the nitrogen source(C,D)improved the crude protein,17 amino acids,crude fat and fatty acid contents to a certain extent.Carbon source and nitrogen sources was also found to have a significant impact on the total number of heterotrophic bacteria.The total number of heterotrophic bacteria in the feed group(groups A,B,E)was higher than that of the ammonium chloride group(group C,D,F).Addirionally,the number of bacteria in the E group was significantly higher than the other groups and the nitrification performance of the bioflocs when treated with 10 salinity after formation,the nitrification process of the bioflocs in most groups was inhibited.In each group of assimilation performance tests,the two performances did not change due to the ongoing nitrification process.Due to the high carbon to nitrogen ratio and the availability of carbon and nitrogen sources,it was concluded that there was a process of dissimilatory nitrate reduction to ammonium ongoing in the system. |
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