Marine fish eggs and larvae from the

East coast of South Africa Dirk Steinke, Allan Connell, Tyler Zemlak, Paul Hebert

History

Allan Connell

1985 - a major effluent pipeline was about to begin

discharging industrial effluent over a shallow

continental shelf area in Richards Bay

1985 - surface plankton samples were collected, over

several years, to assess the diversity of fish species

spawning in the area, and the intensity and seasonality

of spawning.

1986 – a second study in Park Rynie was started in

order to collect alive specimens.

1987 – cataloguing of eggs and hatched larvae started

2004 – DNA Barcoding was added to the procedure

(including sampling of adults for reference library)

History

Work flow

• By collecting both offshore (5km) and inshore (0.5km) a reasonable

assessment of location of spawning was obtained for all the common

eggs in the study area.

• A simple “key” based on the physical features of pelagic fish eggs,

was used to separate eggs into basic groups.

• eggs were hatched and both eggs and larvae were photographed.

• once larvae had fully pigmented eyes, they

were anaesthetised with MS222, prior to

fixing in 98% alcohol for DNA Barcoding.

• other larvae were reared to the point

where fin counts and juvenile features

aided in identification.

Work flow

• DNA extraction was done using standard protocols at the CCDB.

• A reduced elution volume was used.

• PCR used Fish Cocktail (Ivanova 2007).

• Sequences were queried against BOLD using its Identification

Engine (only 100% were considered).

• reared larvae of the same batch were fixed in formalin and serve as

„para-vouchers‟

Today

• some 2100 larvae have been barcoded since 2005

• the local adult reference library (assembled in parallel) contains

some 900 species

• some 1500 species of marine fishes from South Africa are barcoded

• 9000 fish species have been barcoded world-wide

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Are we done?

Phylo

genetic D

ive

rsity (

PD

)

# barcodes

• PD calculated using Conserve based on

NJ trees generated in MEGA 4.0

• Sample size progressively increased by 10

random sequences

Results

• 1638 specimens (78%) could be identified using BOLD

• they represent 280 known species

• 10 of those are new records for South Africa

• the remaining 22% could not be matched to any barcode sequence

on BOLD or GenBank.

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% Offshore

% Inshore

Monodactylus falciformis Cubiceps pauciradiatus Pomadasys olivaceus

Some observations

Mean monthly eggs per sample, averaged over 24 years

Trends

Trends

• large rainfall causing mud to be washed out from rivers

• the high nutrient load of such a deluge caused massive increase in egg

numbers

• three most prolific pelagic egg spawners: Sardinops sagax, Etrumeus

teres, and Scomber japonicus

www.fisheggsandlarvae.com

Acknowledgements:

Erin Corstorphine

Tyler Zemlak

Philip Heemstra

Biodiversity Institute of

Ontario

Thank you!