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Rapid Testing of Resistance of Timber to Biodegradation by Marine Wood-Boring Crustaceans
Chapters
Summary January 29th, 2022
This protocol presents a method for assessing the feeding rate of the wood-boring crustacean, Limnoria, by measuring faecal pellet production. This method is designed for use in non-specialist labs and has potential for incorporation into standard testing protocols, to evaluate enhanced wood durability under marine conditions.
Transcript
Wood is an excellent material for construction, widely used for marine structures, such as piers, jetties, wharves and coastal defenses, such as groynes. So really excellent for these structures because of the strength, the impact resistance, the ability of people to shape, repair, and modify structures, all of those things are good. However, there's a major problem that results in damage that must run into the billions of dollars per year in terms of damage to marine structures.
This damage is caused either by ship worms, which tunnel through the word and create quite large tunnels and as we will see in this particular presentation, a small crustacean. So here's a tube with some of them in it just about visible, the size of a small ant, but they occur in large numbers. They erode away the surface of wood and, eventually, cause collapse.
What are we going to do about that problem? Well, the traditional approach is effective. It employs broad spectrum biocides, but as their name implies, these biocides are chemicals which do not distinguish between the different sorts of organisms.
So a marine structure may be emitting chemicals that are damaging to the wider environment. With these concerns in mind, legislation has shifted what is permitted for marine use in North America, in the European waters, and in Australian waters. There's a major shift which requires innovation where the methods of protecting the wood are directed specifically at the borders and are not things which are going to cause problems in the broader environment.
So we are setting out in this presentation to demonstrate how we test wood protection methods against the gribble, the tiny crustacean, and what we're presenting is a rapid response test. The traditional methods of testing wood materials for use in the sea require a five-year testing period and companies need to be much more nimble than that allows them to be. So we are looking and have developed a rapid method of evaluating preservation methods.
This allows us to interact with the engineers producing the new methods, allows them to modify and us to test again. So here in this presentation, we'll be looking at a rapid testing method for evaluating ways of protecting wooden structures from the wood boring crustacean, the gribble. This is a bucket laboratory assessment to test for resistance of timber to biodegradation by marine wood borers.
We are using the standard method to assess the feeding rate of a wood boring crustacean, the gribble, by measuring its fecal pellet production as well as assessing its vitality and mortality. After any treatment processes are complete, cut dry wood into test sticks to size two millimeters by four millimeters by 20 millimeters. Add dry sticks to a constant weight under laboratory conditions.
At least five rep occurs of each wood being tested should be used. Post wood preparation. Place sticks under mesh in a food safe plastic container inside the vacuum desiccator and replace the lid, ensuring that there is a tight seal facilitated by a coating of vacuum grease.
Attach three-way valve between the tubing connecting the desiccator and pump with the third tube leading to open air. Ensure that the three-way valve is closed off to air and run the pump to achieve a vacuum of between 0.75 to minus one bar within the vacuum desiccator and hold this vacuum for 45 minutes to one hour. Submerge the open end of the third tube into a container of seawater.
Switch the pump off and close the valve leading to the pump and slowly open the valve until seawater is drawn by the vacuum into the desiccator. Allow the water to flow until it fills the plastic container above the level of the mesh. Then withdraw the tube from the seawater in the container, allowing air to enter until the desiccator attends to atmospheric pressure.
Keep the sticks submerged under the mesh until they sink to the bottom of the plastic container. Submerge seawater saturated test sticks in sea water contained in 50 millimeter Falcon tubes. Replace water regularly for a period of 20 days.
Extract individual specimens of gribble from an infested woodblock. Use a pair of fine forceps and a thin paintbrush. Carefully peel back any wood that is covering the gribble burrow with the forceps.
Once the gribble have been exposed, use a paintbrush to gently pick out individuals from underneath and deposit them in a Petri dish filled with seawater. Check the gribble under a microscope to identify the species and ensure that no damage was caused while extracting. Any females brooding eggs should be discarded as gravid females have a reduced feeding capacity.
Limnoria quadripunctata can be identified under a stereo microscope by the four distinct tubercules arranged in a square pattern on the animal's pleotelson in addition to an on the fifth In well plates with wells of diameter 20 millimeters, place one test stick in five millimeters of unfiltered seawater between 32 and 35 PSU per well. Place treatments or species of woods randomly throughout the well plate. Add one Gribble per well.
Twice per week, remove the test stick and each gribble, one per well from the plate and place into a freshly prepared well plate containing five millimeters of sea water per well. Use a paintbrush to gently brush off any fecal pellets from the stick before transferring and retain the fecal pallets within the original well. Plates can be kept in constant dark conditions as photo period does not have an effect on gribble feeding rate.
For changing the well plate and collecting the fecal pellets, vitality of individual gribble should be assessed. If a gribble has died, this cause a vitality of one. A score of two is given to gribble that are no longer on the wood and lethargic or slowly moving.
Three is given to gribble that are actively swimming or moving, but not on the wood. Gribble that are crawling on the wood's surface are given a vitality score of four. Finally, the highest score of five is given to gribble who have created burrows within the wood.
Use a fine paint brush to separate any clumps so the individual pellets are visible and brush any pellets away from the very edges of the well. Take a detailed photograph under a stereo microscope at one times magnification and upload to a computer. Ensure the pellets are in focus in the background as uniform with no shadows or light reflection on the surface of the water as this can interfere with imaging.
Upload a stack of images by dragging and dropping or by selecting File, Import, Image Sequence, and Browse. Do not change any parameters and then select OK.Next, use the circle tool to select the bottom section of a well containing the fecal pellets. Remove the well edges and select Edit, Clear Outside.
Make the image binary by selecting Process, Make Binary. Calibrate by selecting Analyze, Set Scale, and choose the number of pixels per millimeter for your image. Count the pellets and select Analyze, Analyze Particle in the box next to the size unit squared.
Then select a lower threshold that is the same as the smallest size of a pellet using the unit scale set earlier. Invert show dropdown box, select Outlines, and then tick Summarize and press OK.Convert pellet counts to pellets per day, which give an indirect measure of feeding rate. Discard data from any molting individuals on days that molting occurred.
Our representative results look to the species beech, Scots pine, turpentine, Ekki, and sweet chestnut on the feeding rate and vitality of gribbles. Daily fecal pellet production was calculated and averaged among eight replicates. Counts from individuals that were molting or had previously died were not included in the averages.
The two control species beech and Scots pine saw the highest fecal pellet production while the hardwood Ekkis were below it. The highest vitality of five, shown in dark blue, is seen only on beech and Scots pine wood. Mortality represented by black, a vitality of one, was highest on sweet chestnut.
The majority of remaining living individuals stayed a vitality of four in sweet chestnut, Ekki, and turpentine. The benefits of laboratory trials is that we can rapidly assess really promising modified woods for their durability against marine wood boring organisms and biodegradation in the ocean. I have some woods right here that has been really heavily degraded by these wood boring organisms.
If you imagine, this was quite a decent sized piece of wood and it has been really, really eaten and chewed up. We can see how damaging these organisms are. By testing woods in our laboratory, it is cheaper, quicker, and more efficient than by simply going straight to trials in the ocean and we can actually get results really, really quickly.
We can start to see whether or not the organisms can destroy and eat and survive on the wood and then we can start to pick really, really promising, modified wood that can then progress towards more expensive marine trials.
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