Study On Typical Phage Isolation From Wastewater Systems And Its Effect On The Transmission Of Antibiotic Resistance Genes

In recent years,many studies have focused on the threats of environment and human health posed by antibiotic resistance.The essential nature of resistance occurrence is due to the presence of resistance genes on the bacterial genome.Wastewater treatment plants are hotspots for the spread of resistance genes.Resistance genes can be transmitted among bacteria through horizontal gene transfer.Phages are abundant and diverse in wastewater treatment plants,but their impact on antibiotic resistance gene transmission is still unclear.In this paper,we first investigated the distribution characteristics of viral communities and resistance genes in wastewater by macrovirology,explored the co-occurrence relationship between phages and resistance genes,and further revealed the distribution pattern and transmission risk of viruscarried resistance genes in different processes.Moreover,the lytic phage MG1 with drug-resistant bacteria as hosts was isolated from activated sludge,and the effects of phage MG1 on the growth process of host bacteria and on the ability of different host bacteria to accept resistance genes were investigated.The main conclusions from this paper are as follows.(1)Thirty-three families of viruses were detected in the influent,effluent of biological treatment units and effluent of deep treatment units from the wastewater plant,while the percentage of phages reached 28.89% with the highest relative abundance of the long-tailed phage family(6.90%).A total of 81 resistance gene subtypes were detected in the viruses with the highest percentage of fluoroquinolone antibiotic resistance genes(33.57%).There was a strong correlation between 35 viruses and 13 resistance genes at the genus level,and it is likely that these viruses carry some of the resistance genes and are likely to transmit resistance genes under certain circumstances.(2)A phage MG1 was successfully isolated and purified from the sewage plant secondary sludge mix of E.coli MG 1655 with methicillin resistance.The optimal infection complex of phage MG1 was 0.001.And the sequence identity of this phage was 96.43% with the complete genome of E.coli phage Lilleput(MK-629525.1),which belongs to the microphage family.The phage was highly active from 0 to 40°C and at p H=5 to 8 environment.The phage MG1 inhibited the growth of the host bacteria most significantly at the optimal infection complex,while the maximum specific growth rate of the host bacteria was the highest(0.0084 min-1).(3)When the donor bacteria were host bacteria and the recipient bacteria were non-host bacteria,the phage was able to reduce the transfer rate by affecting the population size of the host bacteria(0.14% to 0.10%).When the recipient bacterium was a sludge mix,the phage may also infest the recipient bacterium,and the addition of the phage reduced the transfer rate(2.11% to 0.59%).When the recipient bacterium was also the host bacterium,the phage preferentially infected the sensitive recipient bacterium,resulting in a larger proportion of donor bacteria and thus facilitating transfer(0.02% to 0.15%).The phage is able to control the population size of resistant host bacteria to control the transmission of resistance genes.