Studies On Mutation Breeding Of Acidithiobacillus Ferrooxidans By Heavy Ion Beam Irradiation And Its Molecular Mechanism

As the decline of gold deposits,it is expected that refractory gold ores will become the main resource for gold production.Biooxidation,an eco-friendly and low-cost method compared with traditional methods,has been successfully applied in the field of refractory gold ores pretreatment.However,biooxidation of refractory gold ores has not been widely commercialized because of the low leaching efficiency which is the result of the long growth cycle and slow oxidation activity of the bacteria Acidithiobacillus ferrooxidans.Therefore,breeding new bacterial strains with higher oxidation efficiency is crucial to industrial application.In this study,carbon ion beam irradiation and X-ray irradiation were applied to a novel strain of Acidithiobacillus ferrooxidans,YS-1,to improve its ferrous iron oxidation capacity.In addition,an expression model of specific genes involved in ferrous iron oxidation in the m RNA level was investigated to analyze the underlying cause of high ferrous iron oxidation capacity.1.Isolation and identification of original Acidithiobacillus ferrooxidans strainA novel strain YS-1 was isolated,and the morphological characteristics and molecular identification suggested that it belonged to Acidithiobacillus ferrooxidans species.The cells of strain YS-1 were motile,rod-shaped,and the length and diameter were 1.2-1.5 ?m and 0.4-0.7?m,respectively.The strain YS-1 was a chemoautotrophic microorganism which can obtain energy by oxixizing ferrous iron and sulfur or sulfur compounds.The optimal growth condition of YS-1 was: p H 2.0,shaking speed 150 rpm,inoculation quantity 10% and temperature 35?.Compared with Acidithiobacillus ferrooxidans ATCC 23270 grows at 30°C,YS-1 has a better tolerance of temperature of 35°C.It is suggested that YS-1 may have a promising prospect for biooxidation industrial application.2.Mutant breeding of strain YS-1 by carbon ion beam and X-ray irradiationTo evaluate the effects of carbon ion beam and X-ray irradiation on cell viability,YS-1 cells which treated with different doses of irradiation were assessed in terms of viable cell number.The results revealed that cell survive rate was declined with increasing doses of carbon ion irradiation.The cell mortality reached 95% at 320 Gy.As for X-ray irradiation,Cell mortality reached 82.78% at 360 Gy.After irradiation,two mutants were screened.These two strains could grow under the condition of 39°C or 90mmol/L arsenic respectively.For carbon ion irradiation,the highest oxidation ability strain was obtained from the 320Gy-treated group(named SJ-1).Compared with the oxidization process of 60 h by the original strain,SJ-1 could finish the process within 48 h,the other mutant(JW-1)with high ferrous iron oxidation activity was obtained from 360 Gy X-ray irradiation,and the ferrous iron oxidation process was 10 h earlier than the original strain.Both of strain SJ-1 and JW-1 were good genetic stabilities.3.Biooxidation and cyanide leaching of refractory gold oresTo compare the biooxidation capacity for refractory gold minerals among the original strain YS-1 and irradiation-treated samples,the leaching rates of total iron were investigated.The highest leaching rate of iron was found in the culture of mutant SJ-1,which reached up to 73.15% after 28 days of incubation,and the leaching rate was 28.12% higher than strain YS-1.The strain JW-1 made the leaching rate of total iron reached to 66.19%,which was 21.16% higher than the original strain.The effect of biooxidation pretreatment on the gold recovery of refractory gold mineral was evaluated by cyanide leaching.A remarkable 43.87% increase in the extraction of gold occurred when the mineral had been subjected to biooxidation by the mutant SJ-1 before cyanidation,and the gold recovery of the ore biooxidized by the strain JW-1 reached to 89.89%,which was 42.03% higher than that of direct cyanidation.4.Transcription levels of genes involved in ferrous iron oxidationTo explore the molecular mechanism of carbon ion beam and X-ray irradiation-induced oxidation capacity changes on Acidithiobacillus ferrooxidans strains,the transcriptional profiles of genes involved in ferrous iron oxidation in mutant SJ-1,JW-1 and strain YS-1 were detected during the lag,logarithmic,stationary and aging phases.The results showed that the transcription levels of genes varied substantially between strains,and the changes generally included a considerable increase in transcription levels within the mutants SJ-1 and JW-1.During growth of the mutant SJ-1 and JW-1,transcription levels of almost all genes investigated were higher than their basal levels in strain YS-1,and the changes of gene expression levels in strain SJ-1 were much more significant than them in JW-1.The expression profile analysis of genes involved in ferrous iron oxidation indicated that upregulated expression of these genes might be closely related to higher oxidation capacity.