| Objective: With the development of human productivity,the pollution of metal-like arsenic in the environment is becoming more and more serious.Arsenic,as a non-essential element,can enter the human body from the environment through the food chain,and then cause damage to life and health.In view of the environmental,health and social problems caused by metal-like arsenic pollution,remediation technology of heavy metal pollution has attracted much attention.Among them,plant-microbial remediation technology is an efficient,economic and safe means of heavy metal contaminated soil remediation.Ectomycorrhizal fungi are the main microbial members,and it is of great significance to study the response mechanism of ectomycorrhizal fungi in the remediation of heavy metal contaminated soil.Methods: An arsenic-tolerant fungus W9 was isolated from several fungal fruiterbodies collected from arsenic-contaminated areas and identified as Suillus Luteus.Under five different arsenic concentrations(0,50,100,250 and 500 mg/L),the physiological and biochemical characteristics of W9 strains under arsenic stress were investigated by measuring biomass,micromorphology,distribution of arsenic at subcellular level,and secretion of organic acids.Then,transcriptome analysis was performed on W9 strain under different arsenic concentrations to elucidate the molecular mechanism of potential arsenic tolerance by exploring the differential gene expression and gene enrichment characteristics of W9 strain under different arsenic concentrations.Results:(1)The results of arsenic tolerance experiment showed that strain W9 could grow under 5 arsenic concentrations(0,50,100,250,500 mg/L)in both solid and liquid culture.There was no significant difference in colony diameter between 0 mg/L group and 50mg/L group under solid culture.With the increase of arsenic stress concentration,the colony diameter decreased.The results of liquid culture showed that there was no significant difference in the biomass of the four groups under arsenic stress concentration of 0,50,100 and 250mg/L,but the biomass of the four groups was significantly different from that of the 500 mg/L group.(2)The results of morphology showed that the mycelia of the strain treated with arsenic became dense,and the pores became smaller and deformed.Arsenic could be detected on the surface of the bacteria under50 and 250 mg/L arsenic stress,and the element proportion increased with the increase of arsenic stress concentration.(3)The results of W9 adsorption experiment showed that when the concentration was 100 mg/L,the arsenic adsorption capacity of cell wall was the highest,accounting for 3%,and when the concentration was 50mg/L,the arsenic adsorption capacity was 2%.When the concentration reached 100 mg/L,the highest concentration of intracellular arsenic was144.26 μg/g,and when the concentration reached 500 mg/L,the biomass decreased significantly.(4)Succinic acid secretion showed that when arsenic stress concentration reached 500mg/L,succinic acid content increased 194%compared with 250 mg/L treatment group.(5)Transcriptome analysis showed that the number of differential gene expression in M group(100 mg/L)was the highest,followed by H group(500 mg/L)and L group(50 mg/L).(6)GO enrichment in the L group is mainly enriched in the plasma membrane,the periphery of the cell and ion transporters,etc.;in the M group,it is mainly enriched in cell division and protein metabolism,etc.;H is mainly enriched in protein misfolding reaction and cell stimulation reaction,etc.In addition,organic acid synthesis and metabolism related pathways were enriched.(7)KEGG enrichment results showed that the L group was mainly carbon source metabolism.In addition to metabolism-related pathways and genetic information processing,differences existed in group M.Group H showed differences in cellular processes compared to the other two groups.Among them,the GSH pathway showed differential enrichment.(8)Among the results of differentially expressed genes,gene CY34DRAFT＿807068 showed significant expression in L,M and H treatment groups,and expression multiple was positively correlated with concentration.Fungal hydrophobic protein-related genes were differentially expressed in both L and M groups at over 10 times.Differential expression of methyltransferase domain related gene CY34DRAFT＿807069 was observed in L group and M group,and the expression multiple was 5.88 and 11.56,respectively.Differential expression of cytochrome P450(CYP 450)gene showed differential expression in all the three treatment groupsConclusion:(1)The purified exomycorrhizal fungus S.luteus W9 was collected and isolated in the mine area and showed good arsenic tolerance.Under arsenic stress,W9 strains can improve their own arsenic tolerance by cell wall adsorption and intracellular arsenic absorption.Mass secretion of low molecular weight organic acid succinate may be a response strategy for strains under high concentrations of arsenic stress.(2)Strains have different coping strategies under different concentrations of arsenic stress.It depends on extracellular as well as plasma membrane-related pathways at low concentrations,and more on intracellular-related metabolism at high concentrations.Arsenite efflux may be the main mechanism of arsenic tolerance in W9.Glutathione metabolism,intracellular detoxification,as well as methyltransferase metabolism may play an important role in arsenic tolerance in W9.19 figures,3 tables,79 references... |
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