Percentage of Primnoella chilensis polyps activity states under sediment exposure of 15 days, observation period 21 h, Control, Punta Gruesa@en

To determine behavioural stress, polyp activity states of Primnoella chilensis colonies were observed for 21 hours after eleven days under turbid seawater exposure in an experimental setup (Treatment: equals sedimentation rate of 450 mm/y; control group: ambient seawater). Image analysis was conducted by drawing a segmented selection line along the centre of the colonies' internal axis using the image-processing program ImageJ. This line was moved to the side by 175 % of the internal axis' width measured in pixels and 250 % respectively, resulting in two lines that were named selection A and B. These distances were chosen because fully expanded polyps showed a horizontal spread of about two times the internal axis' diameter and fully closed polyps less than one time respectively. For both lines, a plot profile was created showing grey values of all pixels under the segmented selection line. Lighter pixels of the extended polyps in comparison to the black background resulted in countable peaks. Therefore, the plot was divided by the highest and lowest grey values and bisected horizontally using the vector graphics editor Adobe Illustrator. The number of peaks in the upper part of the divided plot-image was then counted in ImageJ using the function 'analyze particles'. Lastly, the number of polyps of a category was calculated: category 0 (closed): = total polyps observed minus counted peaks for selection line A; category 1 (semi-open): = counted peaks for selection line A minus counted peaks for selection line B; category 2: = counted peaks for selection B.