Romagnan, J-B., Elineau A., Picheral, M., Stemmann, L. Observatoire Oceanologique de Villefranche sur mer, LOV, UMR 7093

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1 Development of new tools and strategies for the monitoring of bottle and net collected plankton. A system based on image acquisition and semi-automatic analysis. Romagnan, J-B., Elineau A., Picheral, M., Stemmann, L. Observatoire Oceanologique de Villefranche sur mer, LOV, UMR 7093 contact: Observatoire Oceanologique de Villefranche sur mer, LOV, UMR

2 Framework WP 10 : Improved existing & emerging technologies! Improvement of coastal observatories! Development of a new set of biological parameters! Better precision of existing parameters! Automation of acquisition " high frequency Task 10.1: Development of new tools and strategies for the monitoring of key biological compartments and processes! Development of a new imaging software Deliverable 10.2: Set of software!zooprocess: Zooscan " FlowCAM contact: Observatoire Oceanologique de Villefranche sur mer, LOV, UMR

3 Why Imaging? Ecological indices for zooplankton Zooscan Taxonomy " abundance & biodiversity indices Size " Biomass & Size structure contact: Observatoire Oceanologique de Villefranche sur mer, LOV, UMR

4 Sensors & Softwares Zooscan! mesozooplankton FlowCAM! Microplankton Zooprocess Freeware open source ImageJ Macro Langage Robust Metadata Archiving Easy outputs :.txt files & vignettes.jpg Easy to handle analysis process totally under control Visual SpreadSheet Proprietary software = closed source Upgrade = ~5000 $ No Metadata Archiving outputs :.lst files & collages Not so easy to handle analysis process totally opaque Development of a zooprocess for flowcam 4

5 Background for development Zooscan Zooprocess : image analysis PkID & Tanagra: automatic 5 identification

6 Systems Single image Zooscan Light 10² objects 2 GB / Sample Sensor µm FlowCAM Multiple images = movie 1-5 objects / image Light Sensor objects total µm 6 Up to 30 GB / Sample

7 Steps of development Zooprocess 7.00 released : Compatible with Windows Xp and Windows 7 Wednesday 24 November 2010! Update Zooprocess for up-to-date Windows OS = enhanced computing power Zooprocess 7.06 and Zooscan V3 coming soon! Thursday 28 July 2011! ZooScan resolution upgraded to 4800 dpi : pixel size = 5 µm! Zooprocess updated to handle 4800 dpi images Zooprocess 7.11 released : NOW compatible with UVP5 and FLOWCAM :USER/ADVANCED mode, new metadata filling method, Stop button and more... Monday 30 July 2012! Zooprocess adapted to handle and analyse multiple images samples : FLOWCAM & UVP!! Zooprocess 7.12 released : Scanning operator now filled with fraction metadata Wednesday 5 September 2012! Metadata management and archiving made more robust & new tools for metadata QC Zooprocess 7.14 released : improvement of UVP5 and GENERIC tools Tuesday 2 April 2013! Zooprocess usable with ANY KIND OF IMAGING DEVICE!! Current test: inverted microscopy 7

8 Zooprocess 7.15 (latest) Launch ImageJ Start interface Launch Zooprocess Project architecture = ZooScan Project name = flowcam_* 8 Background operations Process config files Metadata file Vignettes operations & output files Raw images / films stats Work in progress : step by step archiving of process & log

9 Zooprocess 7.15 (latest) Main menu 4 sensors options 9

10 Zooprocess 7.15 (latest) Image analysis settings Main menu 10

11 Zooprocess 7.15 (latest) Metadata form associated to each sample Main menu 11

12 Outputs Set of raw images for each sample in the project " archive Associated Metadata 2.pid file log of image processing (sensor config + zooprocess config) metadata associated to sample sample flowcam run info (from flowcam) features data for each object imaged To be used for semi automatic identification Optionnal : non - identified vignettes of each object imaged 12

13 Local JERICO Image analysis Network Marc Picheral 2013 Server Computing 13

14 Local JERICO Image analysis Network Imaging Enginner Server Computing 14

15 Application : Sampling 0 m Colonne d eau intégrée 75 m Homogenous and quantitative Sampling Seasonnal time series over 6 order of magnitude in size 15

16 Application: image analysis Size Continuum & taxonomy Zooprocess 7.xx & PkID 1.3 nano 2 categories micro 47 categories meso- macro 38 categories 87 plankton categories + 11 non plankton categories Semi automaticcally identified with PkID millions objects analysed, predicted, and sorted 16

17 Hétérotrophic Bacteria Prochlorococcus Synechococcus Pico-Eukaryotes Nanoplankton Silicoflagellates Other Small Protists Diatoms Dinoflagellates Ciliates Rhizaria Small Omnivorous Zoo Copepods Other Crustaceans Chaetognaths Filtering Gélatinous Carnivorous Gélatinous Ptéropods Application : Multiparametric ecological series 18 Ecological groups time series 17

18 Application : extended size spectrum Multi instrumental size spectrum partial spectrum Micro/flowcam Pico Nano Micro Meso Macro Total spectrum Partial size spectra series 18

19 Conclusions Image analysis software : Zooprocess! Enhanced for zooscan and zooplankton analysis! Adapted specifically to FlowCAM and Microplankton analysis! Adapted to ANY KIND OF IMAGING DEVICE! Inverted microscopy tested! UVP tested Holistic sampling strategy developped to get appropriate samples to test Holistic analysis of plankton dynamics based on zooprocess and combining 3 imaging instruments : JB Romagnan PhD thesis (09/2013) 19

20 Conclusions Image analysis software : Zooprocess! Enhanced for zooscan and zooplankton analysis! Adapted specifically to FlowCAM and Microplankton analysis! Adapted to ANY KIND OF IMAGING DEVICE! Inverted microscopy tested! UVP tested Holistic sampling strategy developped to get appropriate samples to test Holistic analysis of plankton dynamics based on zooprocess and combining 3 imaging instruments : Romagnan et al., 2014, Plos One Lots of ongoing collaborations around imaging and the zooprocess!! 20

21 Nanoplankton (inverted microscopy)

22 Radiolarians

23 You are here Thanks for your attention! 23

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