Nijoy John Postdoctoral Researcher Jex Lab, Walter and Eliza Hall Institute of Medical Research Water Research Centre, UNSW

Population genetics and toxigenicity of Victorian sites of a recent (2016) cyanobacterial bloom in the Murray River

Nijoy John Postdoctoral researcher Jex lab, Walter and Eliza Hall Institute of Medical Research Water Research Centre, UNSW

Microscopy data

• Predominant species: Chrysosporum ovalisporum • Subdominant species: Planktolyngbya, Chroococcales, Cyanogranis sp., Aphanocapsa sp. • Small quantities : Raphidiopsis raciborskii, Microcystis sp., Dolichospermum sp. • Total BGA count: more than drinking and recreational water guideline

• Cylindrospermopsin producer • Generally found is tropical Queensland • Aphanizomenonaceae family • Nitrogen fixing cyanobacteria

C. ovalisporum (Crawford et al., 2017) Multiplex-Tandem PCR (MT-PCR) WaterRA project 1043

• Diagnostic specificity and sensitivity: 100% and 97.7% respectively • Simultaneous amplification of 6 gene targets MT-PCR data −34 Prevalence

31% Toxin 39% −35 CYN MC

SXT

30% Latitude −36

Abundance (>1000 gene copies/mL)

7% Downstream Upstream −37 31% 142144 146 Longitude 62%

● Ouyen Pental Island● Barmah Cobram● Murray U/S 290S28 Wemen● Taylors (Gunbower) Picola● MV channel Barnes ● Piangil Leitchville● Nathalia Yarrawonga Nyah● Torrumbarry Numurkah● Lake Mulwala ● Lower Murray Murray (Echuca)● Torgannah Mays Bridge Murray (Wahgunyah) Anatoxin producers in the Murray River

• Nested PCR using previously established primers • Anatoxin = 59/82 • Confirmation by ELISA 16S rRNA short read amplicon sequencing

Greengenes database SILVA database

Acknowledgements: Jacob Munro; Louise Baker, Katharina Stracke, Stephen Wilcox Long read sequencing of Phycocyanin gene

Aphanizomenon flos-aquae TR183 (Sivonen et al., 2002)

Acknowledgements: Jacob Munro; Sarah Charnaud, Stephen Wilcox Can we detect C. ovalisporum if present ?? Multiple toxin producers in the Murray River Sequencing vs MT-PCR

−34

Toxin −35 CYN MC

SXT Latitude −36

Downstream Upstream −37 142144 146 Longitude

Microscopy Illumina 16S rRNA PacBio PC-IGS

C. ovalisporum D. planctonicum NIVA-CYA 659 Aphanizomenon flos-aquae TR183 (Crawford et al., 2017) (Ballot et al., 2010) (Sivonen et al., 2002) Opportunities to the water sector o MT-PCR: insight into toxigenic and T/O potential of cyanobacterial blooms o Highly sensitive and specific technique: rapid assessment, early warning tool o Molecular approaches: significant understanding about bloom complexity in a shorter period

of time o Study interactions between cyanobacteria and other microbial populations o Molecular techniques and physicochemical data: causes and potential impacts of blooms. Acknowledgements Supervisors PHI Division • Assoc. Prof. Aaron R Jex • Dr. Brendan Ansell • Jacob Munro • Dr. Nick Crosbie, Melbourne Water • Dr. Sarah Charnaud • Dr. Judy Blackbeard, Melbourne Water • Natalie Senzo WEHI Genomics Lab Jex Lab, WEHI • Dr. Louise Baker • Dr. Stephen Wilcox • Dr. Samantha Emery Water Research Australia • Dr. Hayley Joseph • Carolyn Bellamy • Katharina Stracke Mentor • Balu Balan • Dr. Daniel Hoefel (SA Water) • Swapnil Tichkule • Dr. Najoua Lalaoui (WEHI) • Rikvin Singh • Amrita Vijay Industry Partners • Pradip Roy • Goulburn Murray Water • Goulburn Valley Water The University of Melbourne • Grampians Wimmera • Dr. Anson V. Koehler • North East Water • Coliban Water