This interactive shows how Cawthron scientists found out what killed the dogs on Auckland beaches.

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Rights: 2012. University of Waikato. All Rights Reserved. Published 28 June 2016 Referencing Hub media

Interact with some of the processes that Cawthron scientists used to discover what killed the dogs on Auckland beaches in 2009. Collect samples, analyse them and identify a toxin in one of the samples.

You will need the Adobe Flash Player to view this.

Transcript

Screen 1

Introduction

Can you help to solve this mystery?

Several dogs have died on a beach. The owners said the dogs vomited and acted strangely before they died. It seems to indicate some kind of poisoning. The vet noticed that the stomach contents of some of the dogs contained seawater, cooked bone fragments, sea creatures and rubbish.

Scientists have already tested most of the items on the beach – marked with an x. The results for all of them showed negative for levels of toxins high enough to have killed the dogs. There are five items left. Drag and drop them into your bucket for testing in the lab.

Screen 2

In the Lab

You have taken your items to the lab and homogenised some of each one in a blender. You added some other ingredients to further break down and separate the particles. You are testing for toxins in the samples.

Drag the samples into the liquid chromatography-mass spectrometer (LC-MS) to find out if they contain a toxin and, if so, what the molecular mass is for that toxin (to compare to a known toxin).

Check the toxin level and then check out the LD50 to find out how poisonous it is. If the micrograms (µg/kg) for the LD50 are low and the level of toxin is high, look out!

Liquid chromatography-mass spectrometer

Video transcript: Paul McNabb

An LC-MS is really two instruments. It’s an LC – so liquid chromatograph – and this separates compounds based on their chemical properties. Toxins that are a different polarity will emerge from an LC system at different times. As they emerge from the LC system, they are volatilised, so they’re put from a liquid form into a gaseous form, and then they’re analysed within a mass spectrometer as charged particles. Certain chemicals will emerge from the LC, and then those chemicals give very distinctive signals in the mass spectrometer, and that signal is based on the size or the mass of the molecule.

So the mass spectrometer will separate compounds based on their mass. The mass spectrometer that we have will see things from a mass of around about 10 or 20 through to 3 or 4 thousand daltons. Daltons are a unit of atomic mass.

Results

Chicken bones – no toxin present
Sea slug – extreme level of toxin present, molecular mass 319.28 Da
Crab – no toxin present 
Dead mussel – low level of toxin present, molecular mass 805 Da
Rotting fish – no toxin present

Screen 3

You have identified two items containing toxins – the sea slug and the mussel. The toxin in the mussel is at a low level but the toxin in the sea slug appears to be at an extreme level.

To identify the sea slug toxin, match the molecular mass with one of the five toxins below. 
Sea slug molecular mass = 319.28 Da

Toxins

Brevetoxin 
C52H74NO17SNa
Molecular mass = 1039 Da
LD50 = 200 µg/kg

Saxitoxin
C10H15N7O4
Molecular mass = 294 Da
LD50 = 10 µg/kg

Tetrodotoxin 
C11H17N3O8
Molecular mass = 319.28 Da
LD50 = 10 µg/kg

Okadaic acid
C44H68O13
Molecular mass = 805 Da
LD50 = 200 µg/kg

Domoic acid
C15H21NO6
Molecular mass = 311.14 Da
LD50 = 3600 µg/kg

Results: Tetrodoxin

Well done! This toxin not only has an extreme level but has an LD50 of 10 – very toxic!!
Great detective work. You have identified tetrodotoxin as the toxin found in the grey side-gilled sea slug (Pleurobranchaea maculata). This slug was indeed the culprit that killed the dogs.

Video transcript: Dr Susie Wood

The grey side-gilled sea slug or Pleurobranchaea maculata, they’re found all around New Zealand, Australia and South-East Asia. The slugs live for about a year. We find them usually at about 6–10 metres depth. Typically, we find them on rocky reefs. They’ll eat pretty much anything and they’ll also eat themselves, and that’s possibly one reason that we see such a huge variation in the levels in TTX between slugs. We like to say that the sea slug is New Zealand’s most toxic animal – we know that there’s enough TTX in one slug to kill four adult humans.

Acknowledgement: 
Jarrod Walker