Taking a sample is just the beginning and preserving and processing specimens requires more than just the e-word.
Having completed eight dives, at least sixteen shore excursions, one nightlighting session, six trips to the fish markets, several roadside purchases and a surprise swim up to a fishing boat, the scientists have well and truly justified that purchase of 90 litres of ethanol on Day 1.
On each trip the preserving began in the field and continued back at our accommodation. The specimens will soon be shipped to the Museum where a thorough analysis can be performed, possibly including SEM photography and DNA sequencing. After that, identifications can be confirmed if necessary and scientific papers can be published.
So how did the scientists start to process their samples here in Timor? I spoke with about half the team to find out (because talking to all of them would've made this post longer than a Dead Sea scroll).
Lauren studies amphipods, an order of crustaceans, and was picking samples from dive sites (and one nightlighting session) with these small (usually less then 10mm) creatures in mind. She uses the freshwater dip method which means placing her substrate samples in a bucket of tap water as it encourages saltwater animals to release from their holds.
Lauren then elutriated (swirled) the bucket, causing the heavier sediment to fall to the bottom and the lighter amphipods to rise to the top. The swirling water was slowly tipped out into a sieve. What was filtered out was then placed in a tray and examined for animals, which were picked up with forceps or pipettes and placed into jars of ethanol. After that, the habitat samples that remained in the bucket were also placed in the tray and similarly examined for fauna.
We had a team of five fish scientists (dubbed ‘the fishos’) on this trip who worked together to process their samples and make strange, bawdy jokes whenever possible. While they did collect samples at the fish markets, the vast majority of their specimens were taken on dives and kept in plastic bags of seawater (in eskies) until they returned to camp.
Hijacking the dinner table for hours at a time, their processing often looked and sounded like question time at Parliament, but on closer examination was actually a highly organised and efficient affair. Their processing started with placing the fishes into trays and tubs of ice, dividing the day’s catch roughly by type.
From there the team began to identify the fishes, with each member focussing on those species they specialise in: Barry covered the wrasses, for example, and Jeff the cardinal fishes. They would often check with each other, however, and consult the stack of reference books they brought with them.
Once a fish was named it would go to Mark for photographing, sometimes having their fins pinned out if they had distinctive colours. Sally was the chief scribe during all of this, recording names, sizes and number of specimens, as well as writing small labels for Mark to photograph with the fishes.
If a fish was identified as one that hadn't been previously collected on the trip, then it would go to Amanda who would extract a small piece of muscle tissue with a scalpel and place it into a vial of DMSO solution (which is conducive to DNA analysis). The rest of the fish would be placed in formalin.
Penny came on this trip to collect crustaceans and fishes and was involved in the processing of invertebrate samples. Like Lauren, Penny did most of her sorting in a tray, but her samples remained in saltwater until they were picked out and placed in jars of ethanol.
From experience she was able to remove large pieces of reef rock from the tray that were unlikely to contain animals (those that shake ‘clean’ in the water for example) and make use of chisels to break down the likely ones (those with cracks and crevices).
Nerida and Greg frequently collected sediment from the ocean floor, elutriating at the back of the boat and transferring the filtered portion to plastic bags. Back at their makeshift lab they would examine this remainder in dishes under the microscope, looking for sea slugs and similar animals not much bigger than the grains of sediment they move between.
The process involved multiple dishes and frequent changing and cleaning of the seawater to make it easier to observe these microscopic animals. At the end of the process they would have a collection of specimens and would decide then which ones to process further. These would be photographed while still alive and depending on their size, have either a subsample taken for DNA analysis (and placed in a vial of ethanol) or be placed whole in ethanol or formalin.
Rosemary searched for tiny sea snails in mangrove and intertidal zones in Timor, collecting samples of mud, leaf matter and debris which she would ‘coarsely wash’ in the field using a very fine sieve. Keeping these samples cool in a bucket, she would return to the lab and use a spoon to scoop this ‘washed off’ matter into a petri dish.
The contents of the dish were swirled around so that it would settle into one layer, making it easier to see crawling animals or shapes that she recognises. She placed her specimens in ethanol for photographing and DNA sequencing back at the Museum.
Mandy surveyed the dive sites, markets and random fishing boats for cephalopods (octopuses, squid and cuttlefish). Back at the hotel, she would photograph each specimen and take two tissue samples: one muscle and one from the gills.
These samples would be placed in separate vials of ethanol. Later, the ethanol will be poured off and the samples added to our frozen tissue collection at the Museum and made available for DNA analysis.
The remainder of the specimens she acquired were fixed in formalin, with the beaks and radulas of the squid and the cuttlebones of the cuttlefish being detached beforehand (yes, squid have beaks). These will eventually be transferred to 70% ethanol, given a number in our database and added to our collection for long term storage.