1. Identification of Cyanophage Ma-LBP and Infection of the Cyanobacterium M. aeruginosa from an Australian Subtropical Lake by the Virus ( Stephen & Peter, 2004) Prof.: Dr. Hul Seingheng Subject :General Microbiology Group 2 : DYSI Nora EK Pichmony Hak Rany HE Ravy Horn chanrithy Academic year 2010-2011 Institute of Technology of Cambodia Department of Chemical and food technology 1

2. Key terms Cyanophage : ◦ phage that infects to cyanobacteria ◦ Virus specific to cyanobacteria 2 Infection: Lysis : dissolution or destruction of cells, such as blood cells or bacteria, as by the action of a specific lysin that disrupts the cell membrane.

3. Why was this research conducted? Toxic cyanobacteria are common seasonal inhabitants of subtropical lakes in Queenland, Australia. ( late autumn and early spring) 3 It could be because of :  Natural control by cyanophage  Environmental factors : influence the ability of virus to infect, lyse the host.

4. Objectives of research to determine whether Cyanophage influence on the abundance of M. aeruginosa blooms. Identify the Cyanophage Ma-LBP. 4

5. Material and Methods Materials Sampling site : Lake Baroon, South EastQueenland, Australia 5

6. Materials Field sample : - collected in March, April, June and July 2001 - from intake tower, north and south swimming pool( at the surface, at 3, 6 and 9m depths) Water samples mixed before further subsampling Host growth medium : B-12 medium for isolated and grow M.aeroginosa. 6 Material and Methods

7. Methods 7 Material and Methods (con’t) Isolating M.aeroginosa from Lake Baroon Calculate the growth and replication Collecting viruses from Lake Baroon Collect the virus from supernatant M.aeroginosa growth and generation time assaysDetermining viral numbers

8. Stained cyanophage used to assess infectionCyanophage replication rate and lysis assay Add the natural viral population to the cultured host population ( 6 days) Determining cyanophage burst size Quantified the cyanophage released Isolation of M.aeruginosa phage for TEMData analysis 8 Material and Methods (con’t)

9. Results M.aeruginosa growth rate 9

10. Lysis assays 10 Results  The more VLPs that were present at the start of the incubation, the more and faster the host population decreased.

11. Results Observing viral infection and lysis 11 A.Cell wall rupture B.Cytoplasm leakage C. Whole cells Viral burst size : - 28 cyanophage per cell -each cycle of replication cycle 11.2h

12. TEM studied to characterize the cyanophage 12 Results T7-like morphology with short geometries tail : family Podoviridae

13. Discussion Important finding: the quantitive demonstration of cyanophage that are infective for M.aeruginosa TEM of Cyanophage Ma-LBP is same as TEM picture of Podoviridae 13

14. Discussion Infection, lysis and immunity ◦ Host decrease 95% within 6 days when Cyanophage Ma-LBP in original sample=0.23% from the sampling 14 No bloom can form, if nothing interfere with the Cyanophage ability

15. Discussion However….. After the lysis assays ◦ host cells (after 95% reduction). Then we add Cyanophage again. ◦ This acclimated host cells attained natural population densities in 3 weeks with a resistance to the Cyanophage Ma-LBP. 15 The host may have simply become resistant

16. Conclusion Control of bloom formation ◦ Cyanophage presence possibly suppressed M.aeruginosa abundance. ◦ Cyanobaterial blooms may results from condition in the lake that both favor host growth and prevent viral infection and lysis. 16

17. Recommendation Research aims ◦ to understand physical and chemical factors that might control toxic cyanobaterial blooms 17 consider the factors might interfere with the binding, infection, and lysis of the host’s cyanophage. Should also..

18. Reference Tucker, S. & Pollard, P. (2005). Applied and Environmental Microbiology: Identification of Cyanophage Ma-LBP and Infection of the Cyanobacterium Microcystis aeruginosa from an Australian Subtropical Lake by the Virus.71:2. pp 629-635. Doi: 10.1128/AEM.71.2.629-635.2005 18

19. Thank you for attention! Welcome for questions and comments!