Toxicity And Metabolic Effects Of Nickel On C.testosteroni ZG2 And Bacillus.D2

Heavy metal pollution is the environmental pollution caused by heavy metals or their compounds.Because heavy metals have the characteristics of enrichment and difficult degradation,the pollution problem of heavy metals has attracted worldwide attention.Nickel is a microelement needed for the growth and metabolism of organisms,but it will be toxic if the concentration exceeds a certain level.Nickel toxicity can cause oxidative stress and DNA damage.Due to the increasingly serious nickel pollution in the environment,the exposure of bacteria and other microorganisms to nickel stress has been a common problem.In order to avoid nickel toxicity,bacteria must enhance their tolerance to environmental pressure and regulate the concentration of nickel in cells to maintain the stability of the internal environment,so that the growth and metabolism can be carried out normally.Therefore,the study of the mechanism of bacterial tolerance to nickel is helpful to understand the response of bacteria to environmental pressure conditions,and can provide a theoretical basis for the study of microbial treatment of nickel pollution and environmental remediation.In this study,C.testosteroni ZG2 and Bacillus.D2 were used as the research objects.The growth curves of the two bacteria were measured by turbidimetric method under different concentrations of nickel to evaluate the effects of nickel on the growth of the two bacteria.The removal efficiency of nickel by bacteria was studied by flame atomic absorption spectrometry.The morphological changes of the bacteria under nickel stress were observed by scanning electronic microscopy(SEM).In addition,fourier transform infrared spectroscopy(FTIR)analysis and lactate dehydrogenase(LDH)assay were carried out to evaluate the damage characteristics of membrane caused by nickel toxicity to the two bacteria.Finally,LC-MS and GC-MS metabonomics were used to analyze the different metabolites and metabolic pathways to explore the possible mechanism of nickel tolerance of the two bacteria.The main results showed that nickel inhibited the growth of ZG2 and D2,resulting in the lag of logarithmic growth period of ZG2 and the significant decrease of biomass of D2.Under nickel stress for 12 h,the highest antibacterial rates of ZG2 and D2 were93.03% and 94.14%,respectively(P < 0.01).The nickel removal efficiency of ZG2 strain was only 14.96% within 48 h,while related studies showed that the nickel removal efficiency of D2 strain was 16.82% at 10°C(P < 0.05).Nickel toxicity also caused changes in the morphology of strains ZG2 and D2,and the results of FTIR and LDH tests also confirmed that the cell membranes of the two strains were damaged to different degrees.Finally,through metabolomic analysis,238 different metabolites were identified in strain ZG2,and the metabolites with very significant differences included amino acids,sugars and other substances.The contents of amino acids such as isoleucine,proline and 5-phosphoric acid-D-ribulose were significantly upregulated.Adenine,uracil,thiamine and acetyl-acetyl-coenzyme A were significantly down-regulated(P < 0.01).Differential metabolites were mainly concentrated in multiple metabolic pathways such as amino acid metabolism,cofactor synthesis,ABC transporter synthesis,fructose and mannitose metabolism(P < 0.05).A total of 415 different metabolites were identified in strain D2,among which the most significant metabolites were amino acids.The contents of various amino acids including lysine were significantly down-regulated,while the contents of ergothionein and EDTA were significantly up-regulated(P < 0.01),which had certain effects on the metabolism of amino acids such as cysteine and methionine,the synthesis of ABC transporter,pyruvate metabolism and other metabolic pathways(P < 0.05).Finally,the addition of exogenous amino acids under the condition of nickel stress was confirmed to restore the growth of strain D2.The above results indicate that strain ZG2 has an energy-saving life mode,which can reduce the damage caused by redox reaction,adjust metabolism to resist osmotic stress and form EPS to effectively isolate nickel ions.The strain D2 has a strong antioxidant capacity,and can use metal chelation to alleviate the toxicity of nickel,so as to improve energy metabolism and sugar metabolism to maintain its own survival.Metabolomics analysis enables us to understand the changes of bacterial metabolites and metabolic pathways under nickel stress,which provides new clues and ideas for understanding the reaction mechanism of microorganisms under nickel stress,and also provides theoretical basis for the study of microbial bioremediation.