Screening Of Citrinin-degrading Microorganisms And The Study Of The Possible Degradation Mechanisms

Citrinin(CIT)is a nephrotoxicity fungal metabolite produced by several fungi of the genera Penicillium,Aspergillus and Monascuc.It is now know that the mechanism of toxicity is CIT that causing intracellular redox system and mitochondrial membrane permeability dysfunction.Moreover,the CIT pollution area is worldwide with many foods and feeds.So CIT was regularly synergy with other mycotoxins,such as patulin,ochratoxin.Therefore,CIT is a high risk of mycotoxin which seriously harming human health.The key of improving food security is to find ways to control the citrinin and reduce the risk of human exposure.Taking C.podzolicus Y3,high efficiency citrinin-degradation yeast as the research object,this paper has studied the degradation effect and the molecular mechanism in the process of CIT-degradation.The main research results are as follows:(1)Seven strains of yeasts were compared to find that Y3 had high efficiency of degrading CIT.Y3 was identified as Cryptococcus podzolicus with the 5.8S rDNA-ITS sequences sequenced and the morphology identified.The safety test results showed that this yeast was nontoxic and could be used for controlling of CIT in food.(2)Different factors influencing the degradation efficiency of CIT by C.podzolicus Y3 were analyzed,including different initial concentration of CIT,different initial concentration of C.podzolicus Y3,culture conditions,temperature,pH value,etc.The results showed that The initial concentration of C.podzolicus Y3 was larger,the efficiency of degradation was higher;The best temperature condition of C.podzolicus Y3 degrading CIT was 28?C and the high pH could discourage the CIT-degradation;High concentration of CIT could motivate C.podzolicus Y3 to degrade CIT;CIT could be degraded by C.podzolicus Y3 on NYDB but not on PDA medium.(3)Cell wall,extracellular metabolites and cell body of C.podzolicus Y3 was analyzed under laboratory conditions.The results showed that the cell wall and extracellular metabolites of live C.podzolicus Y3 and dead C.podzolicus Y3 had no ability of CIT-degrading.Cell body of C.podzolicus had no ability of CIT-degrading too.(4)The proteins expression was compared between two different culture conditions of C.podzolicus Y3 by using proteomics.The two conditions were NYDB containing 20 mg/L CIT and not.The comparison showed that there were differential expression of proteins with glycosyl transferase family 2,malate dehydrogenase,NAD-dependent,superoxide dismutase [Cu-Zn],cysteine peroxiredoxin,double-strand break repair Rad50 ATPase,cytochrome c and so on.(5)The differentially expression genes of C.podzolicus Y3 at two different culture conditions of NYDB containing CIT and not were analyzed by transcription technique.The RT-qPCR tested nine related genes.The results showed that: Total 14551 genes were identified and 1208 had different expression(|log2(FoldChange)|>2)including increasing gene number of 551,43.05% of all differential expression proteins,and down gene number of 657,56.95% of all differential expression proteins.Six RT-qPCR tested genes with Flavin-binding monooxygenase,Alcohol dehydrogenase,FAD dependent oxidoreductase,Glutathione S-transferase,Acetyltransferase and beta-D-glucuronidase were increasing genes,which were related with degradation of CIT.Multidrug resistance regulator 1 and Peroxisomal membrane were increase genes,while DNA polymerase family A was down gene,which were all related with the damage of CIT.