Status and next steps The Australian Marine Microbial Biodiversity Initiative: Establishing a continental scale, long term oceanography observatory Brown, M.V., Van de Kamp, J., Ostrowski, M., Seymour, J.R., Bissett, A., Goossen, K., Ingleton, T., Messer, L., Laverock., B, Siboni., N, Jeffries T., Bibiloni Isaakson, J., Nelson, T., Coman, F., Davies, C., Frampton D., Robert S., Holmes, B., Abell, G., Craw, P., Kahlke T., Sow, S.L-S., Mcallister, K., Windsor, J., Skuza, M., Crossing, R., Patten, N., Malthouse, P., Paulsen, I., Steinberg, C., Richardson, A., VanRuth, P., Koval, J., Fuhrman, J.A., Fitzgerald, A., Moltmann, T., Bodrossy, L. Levente Bodrossy

AMMBI – Status and next steps BPA Coastal/estuarine opportunities Autonomous sampling devices Next steps

AMMBI – Status and next steps BPA Coastal/estuarine opportunities Autonomous sampling devices Next steps

Multihead peristaltic pump What’s involved 2L seawater Sterivex filter -80 °C Multihead peristaltic pump $ 16 $ 5000 $ 60 $180 - $800 500-3000 ng DNA Genomic analyses Aliquot 1 – for first genomic analyses Aliquots 2-6 – freeze dried, at -80C, for a range of projects Aliquot 7 – QIAsafe 96 well plate, -80C, long term storage

Community composition for: Bacteria Archaea Rottnest Island Since July 2015 6 depths 12x per year Port Hacking Since July 2010 Maria Island Since July 2012 Darwin Since July 2015 3 depths 4x per year* 2L water Community composition for: Bacteria Archaea Eukaryotes (~phytoplankton) Functional composition (recent addition) Sequenced until May/June 2016 Yongala Since July 2015 4 depths 12x per year North Stradbroke Island Since July 2012 6 depths 12x per year Kangaroo Island Since July 2015 6 depths 6x per year

7 Stations, 48/12 months, 5 voyages Bacteria, rchaea and eukaryotes > 30 000 OTUs AMMBI represents the first methodologically standardized, continental scale, microbial ocean-observing network. The data provide a critical baseline against which to measure, and from which to predict, changes in microbial assemblages under future scenarios.

Martin Ostrowski, Macquarie University

Martin Ostrowski, Macquarie University

Conditionally abundant bacteria (<0.01% overall, but >10% in at least 1 sample) MAI PHB NSI 30% 10% 2% Conditionally abundant ones Kirianne’s poster!

Conditionally abundant bacteria (<0.01% overall, but >10% in at least 1 sample) MAI PHB NSI 10% 0.1% 0% Conditionally abundant ones Kirianne’s poster!

Bacteria-phytoplankton interactions Darwin (12.4° S) Port Hacking (34.1° S) Maria Island (42.6° S) Yongala (19.3° S) North Stradbroke Island (27.4° S) Rottnest Island (34.8° S) Kangaroo Island (34.8° S) SAR11Oligo 12567 SAR11Oligo 12503 Mamiellaceae Chloroplast OTU Mark Brown, UNSW

Bacteria-phytoplankton interactions Port Hacking (34.1° S) Maria Island (42.6° S) SAR11Oligo 12567 SAR11Oligo 12503 Mamiellaceae Chloroplast OTU Δ: SAR11Oligo 12567 – SAR11Oligo 12503 Mamiellaceae Chloroplast OTU Mark Brown, UNSW

AMMBI – Status and next steps BPA Coastal/estuarine opportunities Autonomous sampling devices Next steps

BPA Marine Microbial project – pelagic component Rottnest Island Yongala Kangaroo Island Darwin North Stradbroke Island Port Hacking Maria Island Inv2014-e03 CSC_Trip5913 SAIMOS2014_RC48/51 SS2013_t03+t07 SS2010_c09 Pelagic component = AMMBI (IMOS) samples + oceanic voyages extending spatial coverage 2 years, 7 stations + voyage samples, ~1000 samples Amplicon sequencing, ~1000 Bacterial 16S Archaeal 16S Eukaryote 18S Shotgun metagenome sequencing, ~500 Genome sequencing of bacterial isolates, ~20 Transcriptome sequencing of eukaryote isolates, ~10 Total sequencing costs ~$400k

Australian Microbiome Database

AMMBI – Status and next steps BPA Coastal/estuarine opportunities Autonomous sampling devices Next steps

Genomics tools for estuarine status assessment 10 g sediment contains tens of millions of microbes and micro-invertebrates – thousands of different species These organisms live in intimate relationship with the environment and are sensitive indicators of its status (of many different aspects of it) Using genomics we can tell their stories Read their community structure as a fingerprint In order to be able to read this fingerprint we need to establish a baseline database Australia-wide baseline project: Microbial community structure fingerprint vs. estuarine status around 10 g of sediment that can tell the story of thousands of bacteria, phytoplankton and microinvertebrates (amongst others) living in it. these are living in an intimate relationship with the environment and their community structure is a fingerprint of the status and health of the environment. we are not yet able to read this fingerprint in detail, to a large extent because we don't have the baseline dataset to interpret it. to this end, we are working on an Australia-wide baseline of estuarine genomics – BGC – environmental status dataset.

Fragmented efforts Report Report Survey Management decision Publication Publication Management decision Report Report Publication PhD thesis a second one that visually tells the story of the fragmented efforts into estuarine observations and how these could be glued together by support for sequencing (BPA likely to provide it), for bioinformatics (BPA/CSIRO) and for BGC (missing). maybe add modelling plans there, leading to improved advice to estuarine management. Management decision = Agencies, GMOs, research groups, etc. running separate projects, collecting samples, running various analyses (chemical, physical, sometimes genomics observations). In vastly different ways to allow for direct comparison, minimizing any chance for leverage.

Fragmented efforts – can we leverage? Chemistry Sequencing Bio-informatics a second one that visually tells the story of the fragmented efforts into estuarine observations and how these could be glued together by support for sequencing (BPA likely to provide it), for bioinformatics (BPA/CSIRO) and for BGC (missing). maybe add modelling plans there, leading to improved advice to estuarine management. Publications, PhD theses Reports, surveys Best practice, cost efficiency Continent-wide baseline database, leading to: Better understanding of estuarine health and status Modeling of estuarine BGC processes, leading to: Improved support for estuarine management decisions

CSIRO Environomics Future Science Platform Environmental genomics of coastal environments – proposal 2 years initially; 4 FTE; BPA support Relationship between microbial community phenotype and process rates  incorporating microbial data into process models Biodiversity assessment and baseline data establishment at a range of coastal/estuarine sites within Australia (seek partnerships) Define indicators of ecosystem health within these environments Apply modelling (at 1 or 2 sites?)

AMMBI – Status and next steps BPA Coastal/estuarine opportunities Autonomous sampling devices Next steps

SAFA Sample Filtration & Archival unit Function: In-situ microbial sampling and preservation for subsequent laboratory analysis. Collects up to 24 samples. Cost: ~$25-30,000 Status: Second generation; field test successful (30 m) First customers will take delivery in March 2017

STAN Standalone unit for Low cost, high flexibility monitoring Function: Collection, filtration and preservation of a single 2L water sample for lab based analysis. Can be triggered manually, by time or remotely Very low cost allows high spatial and temporal coverage Sampling location flexibility. Cost: ~$200 per unit Status: First field tests done – leak at 30 m, being fixed

MOBI Microbial Oceanography Biosensing Instrument Function : Near real-time, in-situ microbial sampling and analysis. ddPCR with telemetry Cost: ~30k for core instrument Status: Prototype development. Assay validation

AMMBI – Status and next steps BPA Coastal/estuarine opportunities Autonomous sampling devices Next steps

Next steps Merge with other Australian environmental microbiome sequencing projects (AMD) Enter data into Atlas of Living Australia database (-> release data, initiate broad community involvement) Extend into coastal and estuarine environments Quantification – B16S, A16S, E18S qPCRs? Flow cytometry including SYBR Green? Remotely deployable samplers – event based sampling and higher frequency sampling Linking genomics observations of phytoplankton to microscopy and satellite observations HABs: understanding events and underlying microbial community dynamics leading to blooms Phytoplankton productivity – cyanobacterial community dynamics; biogeochemical cycling supporting phytoplankton

Thank you © Malou Zuidema