A Study Of The Inhibition Of Microcystis Aeruginosa By A Comamonadaceae Bacteria

Water bloom or algal bloom is a harmful ecological phenomenon in which some phytoplankton species accumulate rapidly and produce toxins and odorous substances.Water bloom may cause adverse effects to the aquatic ecosystem,environmental safety and human health.Due to the increase of human social activities and the dramatic changes in the global climate,water blooms are widespread worldwide.Water blooms occur frequently in many lakes and reservoir areas in China in recent years.The dominant phytoplankton are often cyanobacteria such as Microcystis,Anabaena,Aphanizomenon,and Oscillatoria et al,and Microcystis aeruginosa is one of the main dominant algae species.The outbreak of water bloom is the result from the combined effects of water nutrition level,climatic conditions and biological factors.In recent years,many studies have shown that algae or cyanobacteria can interact with other microorganisms in either beneficial or harmful formats,which may affect greatly their growth and morphological development.These interactions are believed to play an important role in the outbreak and regression of water blooms.Therefore,studying the interaction between phytoplankton and other microorganisms is critical for understanding the complex interspecies relationship of microorganisms in aquatic conditions and the control of water blooms.In our previous study,we isolated a strain WR11,belonging to the family Comamondaceae,from the water bloom sample of the Nan-Lake in Wuhan.WR11 has a strong inhibitory effect on Microcystis aeruginosa,and may play an important role in the outbreak and regression of water bloom,while the mechanism is not known.In this study,we combined microbial techniques and multi-omics methods to explore the mechanism of the inhibition of M.aeruginosa by WR11.The main results of this study are as follows:(1)Inhibition of M.aeruginosa by WR11 and its regulation by environmental condition.Under the full day light condition,WR11 showed a strong algicidal effect against M.aeruginosa at the beginning.In the co-culture,the cells of M.aeruginosa are ruptured and the cellular contents are released,thereby causing the death of M.aeruginosa.Increasing the ratio of bacterial inoculation enhanced the algicidal effect.But in the later stage of cultivation,M.aeruginosa resumed growth and maintained a good growth state for a long time in the presence of WR11.However,when WR11 and the Microcystis aeruginosa were co-cultured under a short day light condition,the bacteria can continuously maintain the lytic effect.These results suggested that the light conditions have a regulatory effect on the interaction between algicidal bacteria and the M.aeruginosa.(2)Phenotype study of the algicidal dysfunction mutant of WR11.We isolated a spontaneous mutant,M20,which lost the algicidal effect on M.aeruginosa.While both the supernatant and pellet fraction of WR11 culture showed a strong algicidal effect,those of M20 culture had no obvious phenotype.However,the cellular contents released from lysed M20 still showed the algicidal effect.These results implied that the bacteria may secrete algicidal substances to the outside of the cell.The supernatants of the two strains were compared with HPLC scanning and LC-MS.The results showed an absorption peak at 430 nm in the supernatant of WR11 has disappeared in the supernatant of M20.The algicidal ability and molecular Structure of this compound need to be further determined.(3)Genomics study of WR11 and M20.The genome sequences of WR11 and M20 were determined by the third-generation Sequencing technology.The 16S rRNA gene sequence of WR11 has a homology of 97.04%with the Paucibacter genus.Comparative genonmics analysis revealed there are multiple point mutations or deletion mutations in the mutant genome,which provided the targets for subsequent search for the key mutant genes.(4)Proteomics study of the interaction between algicidal bacteria and M.aeruginosa.We set up the following five experimental groups,single culture of M.aeruginosa,co-culture of WR11 and M.aeruginosa,co-culture of M20 and M.aeruginosa,and single cultures of WR11 and M20,for the proteomics assays using a high resolution liquid chromatography-mass spectrometer.The results showed that the algicidal bacteria WR11 significantly affected the membrane protein synthesis and photosystem assembly of M.aeruginosa in the co-cultures.The photosynthetic system of Microcystis aeruginosa was influenced by WR11,resulting into an inhibition effect on the growth.By comparing the expression of WR11 and M20 proteome,it was found that the expression of certain outer membrane proteins,ABC transporters and type II secretion system proteins were significantly down-regulated in M20,which might be important for algicidal ability.In summary,we used the tools of microbiology,genomics and proteomics to study the mechanism of inhibition of M.aeruginosa by a Comamonadaceae Bacteria,and explored the influence of light condition on this interaction.The results will provide a useful basis for further in-depth study of the bacterial algicidal mechanism,and help to understand the complex relationships between microbes across the water bloom event.