Aquacultural Determination of the Ecophysiological Effects of Microplastic Consumption and Retention in Marine Fish

Marine plastic pollution is ranked as one of the greatest threats to marine life (1, 16) and this plastic pollution consists of both macroplastic (> 5 mm) and microplastic (< 5 mm) particles (9).

Whilst primary microplastics are those which were manufactured to be < 5 mm in size, secondary microplastics occur as a result of

the disintegration of larger plastic items (2).

Microplastic Retention Time Experiments

Fish specimens were starved for 48 h before being force-fed a known quantity of

food. They were then held individually in small tanks and observed periodically

until the presence of faeces was detected in order to determine the natural gut

retention times of various fish species.

In order to determine the gut retention times of specific microplastics in the

various fish, known amounts of selected shapes and sizes of UV fluorescent

microplastics were administered to the fish via force-feeding. The fish were then

maintained individually in small tanks and observed periodically under UV light to

determine the presence of microplastics in the faeces. The faeces were retained

and analysed using a novel separation technique in order to quantify microplastic

gut retention time.

MICROPLASTIC SETTLEMENT TIMES

Samples containing known amounts of microplastics of various combinations of

size/structure suspended in water with varying salinities were analysed using a

Fluorometer to observe settlement times.

Baseline readings of the Samples were taken after complete settlement (centrifuging)

and the samples were then re-suspended and fluorometer readings were taken at 1 min

intervals until the baseline fluorescence was reached. The settlement times were then

calculated accordingly

AIMS OF THIS STUDY1- Determine the effects of plastic type, size, shape, surface-area fouling and solvent salinity on the settlement times of various microplastic particles.

2- To investigate the gut retention times of various microplastics in a variety of ecologically important fish species (Mullet- M. cephalus L., Spotted Grunter- Pomadasys commersonni Laćpède and Glassies -Ambassis Ambassis).

3- Assess the effects of prolonged microplastic consumption on the overall physiology (length of fish, weight of fish, girthof fish or Gonado/Hepato-somatic indices) as well as gut histology (microvilli damage, changes in microvilli length, changes in microvilli thickness and changes in the number of gut mucus cells) in M. cephalus L.

4- Determine whether microplastics < 100 µm are able to pas through the gut lining and become assimilated into somatic tissue.

Ingestion by Marine Organisms

Microplastic particles are prone to ingestion by a variety of organisms due to their presence in both the sediment and the water column (3, 6).

Microplastics have been found in the guts of both pelagic and demersal fish species (10) yet little is known about the retention time of these plastics

within the fish (1).

The retention of microplastic particles may have several effects on digestive processes. They could potentially block or abrade the digestive tract, as

well as hinder the assimilation of food items. However, the impacts of microplastic pollution on marine organisms are not well established (10, 15).

Observational studies of natural systems are unable to accurately correlate the extent of microplastic consumption with any

physiological effects on marine organisms due to a high degree of natural variability within these systems (8, 10). Such

aspects of the eco-physiology of marine organisms may therefore be better examined in aquacultural systems (5, 10).

Microplastics and Persistent Organic Pollutants (POPs)

Ingested microplastics may serve as a delivery mechanism for POPs which have a high affinity for the hydrophobic

surfaces of the microplastic particles (12).

Filter-feeding organisms which ingest microplastics in large quantities may

facilitate and enhance the bio-accumulation of POPs throughout the food

web (5).

An important factor of this is the retention time of microplastics in an organism's gut. To date there are no studies that

investigate the gut evacuation times of microplastics in filter-feeding fish.

● Settlement time and gut retention time experiments(Figures 1 & 2) indicate that several important links exist between the size/structure of microplastic particles and their bio-availability, their distribution and their retention time in fish. Smaller microplastic particles such as microfibres and microbeads (< 1 mm) are more likely to remain bio-available to surface/ pelagic filter-feeders for longer periods than larger microplastics (2 – 5 mm).

● The presence of food items with microplastics increases the chance of retention during feeding. Microplastics such as microfibres which remain in suspension in the photic zone for longer periods are more likely to undergo bio-fouling (11), increasing their chances of being retained by filter-feeding fish.

● During these extended periods of suspension microplastic particles are increasingly subjected to the sorption of POPs and other toxins from the water (12, 14).

● Larger microplastic particles are more likely to end up in the benthic environment which may have implications on the ecology of organisms which exist there (13).

● Larger microplastic particles have implications on the physiology of the fish as they diminish the stomach's capacity for food items for periods up to three times greater than the natural gut retention time. They may also be more likely to block the gut as a result of their size (17).

● Ingested microbeads (> 2 mm) exhibit relatively long gut retention time of up to 39.6 hours. Therefore, even though they have a relatively small surface area for the sorption of POPs, they are retained within a fish's gut for a longer period.

● With regard to the ecological impacts of microplastics, the use of aquaculture enables us to perform ingestion/retention experiments with a greater degree of accuracy and to observe trends that would otherwise not be visible to us in natural systems.

Preliminary Results and Findings

PROLONGED EXPOSURE

Fish specimens will be maintained in one of the following exposure treatments for a period of three months.

1- Control: 0 mg 2- Low Conc: 10 mg/dm3 High Conc: 150 mg/dm3

Monthly - 5 fish of each species taken from each treatment for for the following analyses

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Matthew. Coote, Dr. Deborah Robertson-Andersson, G.K Moodley

University of KwaZulu–Natal – Westville Campus, College of Agriculture, Engineering and ScienceSchool of Life Sciences, Private Bag X 54001, Durban, 4000, South Africa

GUT TRANSLOCATION

Fish specimens will be isolated, tagged and force-fed a known amount of microplastics

(< 100 µm) every 48 hours for a period of 10-12 days, whereafter the specimens will

be dissected and the various tissues digested using NaOH, filtered through GFF filters

and analysed under a microscope using UV light in order to determine

presence/absence of microplastics.

Additionally, Histological samples of the gut-lining cells will be taken and observed for

the presence of microplastics assimilated by the gut-lining via possible amoeboid

uptake.

Materials and Methods

A common example of Secondary

Microplastics- the microfibres produced by

synthetic textiles during the washing process

Microfibers enter the marine environment

through waste-water sources that are only

filtered for larger particles (6, 7, 10).

THE MICROPLASTIC PROBLEM

LITERATURE CITED

1) Andrady, A. L. 2011. Microplastics in the marine environment. Marine Pollution Bulletin. 62 (8). 1596–16052) Barnes, K.A., Galgani, F., Thompson, R.C. and Barlaz, M. 2009. Accumulation and fragmentation of plastic debris in global environments. Philosophical Transactions of the Royal Society Biological Sciences. 364. 1985-1998.3) Boerger, C.M., Lattin, G., Moore, S.I. and Moore, C.J. 2010. Plastic ingestion by planktivorous fishes in the North Pacific Central Gyre. Marine Pollution Bulletin. 60. 2275-2278.4) Bowmer, T., Kershaw, P., 2010. Proceedings of the GESAMP International Workshop on Micro-plastic Particles as a Vector in Transporting Persistent, Bio-accumulating and Toxic Substances in the Oceans June 2010. UNESCO-IOC, Paris 5) Clements, K. D., Raubenheimer, D., & Choat, J. H. 2009. Nutritional ecology of marine herbivorous fishes: ten years on. Functional Ecology. 23 – 1. 79-92.6) Cole, M., Lindeque, P., Halsband, C. and Galloway, T.S. 2011. Microplastics as contaminants in the marine environment: A review. Marine Pollution Bulletin. 62. 2588-2597.7) Fendall, L.S. and Sewell, M.A. 2009. Contributing to marine pollution by washing your face: Microplastics in facial cleaners. Marine Pollution Bulletin. 58. 1225-1228.8) Foekema, E. M., De Gruijter, C., Mergia, M. T., van Franeker, J. A., Murk, A. J. & Koelmans, A. A. 2013. Plastic in North sea fish. Environmental science & technology. 47 – 15. 8818-8824.9) Hidalgo-Ruz, V., Gutow, L., Thompson, R.C. and Thiel, M. 2012. Microplastics in the Marine Environment: A Review of the Methods Used for Identification and Quantification. Environmental Science and Technology. 46. 3060-3075.10) Lusher, A.L., McHugh, M. and Thompson, R.C. 2012. Occurrence of microplastic in the gastrointestinal tract of pelagic and demersal fish from the English Channel. Marine Pollution. 1-6.11) Lobelle, D. & Cunliffe, M. 2011. Early microbial biofilm formation on marine plastic debris. Marine Pollution Bulletin. 62-1. 197-200.12) Mato Y. 2001. "Plastic resin pellets as a transport medium for toxic chemicals in the marine environment", Environmental Science & Technology 35(2), pp. 318–324 13) Moore, C. J. 2008. Synthetic polymers in the marine environment: a rapidly increasing, long-term threat. Environmental Research. 108-2. 131-139. 14) Teuten, E. L., Rowland, S. J., Galloway, T. S. & Thompson, R. C. 2007. Potential for plastics to transport hydrophobic contaminants. Environmental science & technology. 41-22. 7759-7764.15) Thompson, R.C., Olsen, Y., Mitchell, R.P., Davis, A., Rowland, S.J., John, A.W.G., McGonigle, D., Russell, A.E., 2004. Lost at sea: where is all the plastic? Science 304, 838.16) Todd, P.A., Ong, X. and Chou, L.M. 2010. Impacts of pollution on marine life in Southeast Asia. Journal of Biodiversity and Conservation. 19. 1063-1082. 17) Wright, S. L., Thompson, R. C. & Galloway, T. S. 2013. The physical impacts of microplastics on marine organisms: A review. Environmental Pollution. 178. 483-492.

Figure 2. Gut Retention Times of Various Microplastics in mullet (M. cephalus L)

Figure 1. Comparative Settlement Times of Various Shapes/Sizes of Microplastic

A- Force-feeding apparatus, B- Gut dissection (M. cephalus L), C- Micrograph of UV fluorescent microplastics

A B C

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