Mechanism Research On Graphene And Tetracycline Influence On Bacteria Community And Resistome In SBR

Graphene is a newly developed material which have been synthesized and studied prevalently in electric products these years.Graphene has broad spectrum antibacterial activity.And it is a promising material in antibacterial application.like antifouling bacteria on membrane surface,eliminating multidrug-resistance-bacteria in environment.However,there are limited research reporting graphene' influence on bacteria community and antibiotic resistance genes?ARGs?.Meanwhile,tetracycline is a broad spectrum antibiotics which could be detected in water environment globally.And tetracycline could change the bacteria community and antbiotic resistance genes dramatically.Our study will discuss graphene'influence on bacteria community and resistome by adding graphene into activated sludge in sequencing batch reactor?SBR?.Meanwhile,we will take tetracycline,a broadly used antibiotics,as a reference.SBRs have been assigned into 4 different groups.1,Control group;2,Graphene group?100mg/L?;3,Tetracycline group?48mg/L?;4,Combine group?100mg/L graphene and 48mg/L tetracycline?.Chemical oxygen demand?COD?and ammonia removal efficiency decreased from 99.3%,97.7%in control group to 92.3%and 92.7%in tetracycline group respectively.And relative abundance of Nitrospira decreased from 0.6%in control group to 0.23%in tetracycline group.The nitrification rate was inhibited consequently.While graphene did not show significant influence on SBR removal efficiency.Illumina Miseq amplicon sequencing platform was used to sequence 16S rRNA in activated sludge and analyze the bacterial community.Compared to tetracycline,graphene shown limited ability in changing bacterial community.In tetracycline group,tetracycline resistant bacteria,like Lactococcus,Parabacteroides became the abundant bacteria.In contrast,a multidrug resistant bacteria Tolumonas became the dominant bacteria in combined group.33 genra relative abundance are higher than 0.2%.31 of them are gram-negative bacteria while 2 of them are gram-positive bacteria.Total abundance of gram-positive bacteria decreased in all three groups,which indicate that gram-positive bacteria are more sensitive to tetracycline and graphene.Metagenome of activated sludge were sequenced by Illmina Hiseq 2000.Blast the sequence to antibiotic resistance gene database to annotate ARGs.Graphene would not increase total abundance of ARGs in metagenome.In tetracycline group,ARGs abundance was 5 times higher than control group and the resistome shown specific tetracycline resistance.However,in graphene and tetracycline combining group,the resistome shown multidrug resistance.Based on co-occurrence relationship between ARGs and abundant genera and mobile gene elements?MGEs?we could found:1.ARGs sharing same function?efflux pump?shown strong co-occurrence.2.ARGs hold significant co-occurrence relationship with their hosts?P<0.01,R>0.8?.There was strong corelationship between bacteria community and ARGs.3.ARGs abundance and MGEs abundance shown significant linear correlation?R>0.95?.MGEs,including plasmids pO111₁ and pGMI1000MP,played important role in ARGs distribution,Graphene shown moderately change in bacterial community when compared to tetracycline.And it would not increase total abundance of ARGs.But graphene would increase abundance of multidrug resistant bacteria and multidrug resistance genes.Which maybe result from that multidrug resistance bacteria will hold efflux pump protein on their membrane,and make the bacteria tolerant to graphene.