Now that I officially have all the oysters sampled accounted for, staged, and recorded I split up my results by population as suggested by Laura. I went through and reviewed all my oysters I had previously labeled as HPM or HPF and, based on the literature, (Oates) decided if they were developing hermaphroditic oysters or or actually oysters going towards being male or female (sperm or eggs were degrading). That gave me only three categories to work with and much more visible results. She was right! There do seem to be more differences between populations than between treatments overall. It also seems like overwintering temperature has a bigger impact than OA treatment does, but that is just at first glance. I’m not sure how to pull any significant results from my figures at this point….my statistic skills need some brushing up on.
Check out these results and see if anything should potentially be combined in different ways. The full updated excel document is posted under Histology staging results.
I first made pivot tables to get all the total distributions of sex within each population and treatment:
Then I organized it all in a new spreadsheet so that figures can be easily generated in the future using any combinations of these results:
Then here are some figures I made that show the comparison between overwintering temperature results! Hood Canal oysters show a significant different between the two temperature treatments, South Sound oysters are pretty similar within the population across temperature treatments, and then the Fidalgo Bay oysters are interesting in that they have the exact same results, but there are no females…
I also started play with OA comparison figures. I could do it in two ways: Compare, for example, both the treatments that were overwintered at 10 degrees but had different OA treatments OR I could compare oysters that had the same OA treatment but were overwintered at different temperatures.
For now I am going to work on making a google doc to begin typing up my methods.
1) Finished staging all of the pictures I took of Laura’s slides and inputed the information into the details section of all of the “low” images on the google drive
2) Identified the tissues on the cassettes that Sam is going to be using based on Laura’s original pictures and her histology key just as I did when I was imaging them. These labeled slides can be lined up with the cassettes to identify which oysters are which!
I took pictures of all the histology slides and cassettes, so I will work on going through and labeling all of Laura’s slides properly to make a usable new key.
3) TO DO LIST FOR THE NEXT WEEK OR SO:
-Stage all of Grace’s post-overwintering histology images and add that information to her google drive.
-Create an excel document with all the oysters from both the post-temperature and post-OA treatments, their, and their maturity stage. This can be used to sort the images and compare treatments/locations.
-Create some example figures for how this information can be sorted in different ways.
-Original cassette orientation/slide matching to create new histology sample key for the post-OA slides.
This To-Do list should allow me to start making observations across treatments!
6/30/17 Finished taking pictures of all of Laura’s OA only slides. I just finished labeling them and they are all uploaded to the google drive. The next step is to figure out how to score them! I’m going to read the papers Grace and Laura have sent me about gonad tissue and rating it today.
Here is a compiled list of all the oysters that I either am not sure whether or not I found gonad tissue (labeled: ?) or oysters that I did not image at all (labeled: no) because I could find nothing that resembled gonad tissue:
HL-6_21: no (only 5 oysters on slide and 6 on key)
Took pictures of slides 17-21 today. Took a really long time to figure out which oysters were which because 19 was another split slides, and almost all the oysters were rearranged. The ink and Laura’s pictures online continue to be helpful though!
Oly 17 slide: no issues
Oly 18 slide: no issues
Oly 19 slide: Split slide
Oly 20 slide: SN-6_31 and NF-6_31 have no images as I could not find any gonad tissue
Oly 21 slide: no issues
Here is the link to the google drive where I have been uploading all the images!
Took pictures of slides 12-16 today. Ran into a few problems as one of the cassettes (14) isn’t photographed or just wasn’t included in the file I’ve been looking at on Laura’s GitHub. This ALSO happens to be a slide that was split into two slides and contained oysters from two different treatments. So there really is no way to tell which oysters are which on slide 14.
Summary of issues/notes on the slides I worked on today:
Oly 12 slide: There are 6 oyster locations listed on the histology key, but only 5 oysters on the slide/cassette. All the of the supposed 6 oysters were HL-6, so I am not too concerned.
Oly 13 slide: HL-10_18, not sure if I captured gonad tissues
Oly 14 slide: Split slides. 3 smaller oysters on the first slide and two larger oysters on the second. From Laura’s key it looks as if the largest oyster was supposed to be NF-10_26 (third numerically on slide 14), but it was on the second slide. I called the inked oyster and middle oyster on slide one HL-10_20 and HL-10_21, the super large oyster on the second slide NF-10_26, the third oyster on the first slide I called NF-10_25, and the smaller oyster on slide two I called NF-10_27. Unless the original picture can be found though this is just my best guess.
Oly 15 slide: no problems
Oly 16 slide: no problems
I will go back through once I’m done and accumulate a list of the oysters I was unsure about or couldn’t find gonad tissue for. I should be done by the end of this week.
****note: 7/7/17 Found picture of slide 14, fixed image titles on google drive and all tissues have been identified
Went in to UW to continue imaging Laura’s slides. Imaged slides 3-10. Here are the samples that I either couldn’t find gonad tissue in or I wasn’t quite sure about:
Oly 3 slide: HL-6_13 (not sure if I captured gonad tissue), HL-6_14 (none, no picture)
Oly 6 slide: NF-6_17 (none, no picture), NF-6_19 (not sure if I captured gonad tissue)
Oly 7 slide: NF-6_18 (not sure if I captured gonad tissue)
Oly 8 slide: SN-6_18 (not sure if I captured gonad tissue)
I will go through all of my noted questionable slides when I’m done.
Grace showed me how to image histology slides today in the same way as she did the post-overwintering samples. I will image all of the slides at low (4x) and high (40x) magnification with the high magnification looking at the gonads of the oyster and the low magnification being the zoomed out image of the same place. Some of the oysters were split between slides or moved around after Laura sent them in, but between her histology key and her images of what the oysters looked like in the cassettes I’ve been able to figure out which oysters are which.
The pictures are labeled in the same way as Laura’s histology key (location-overwintering temp-sample #) and then labeled as either high or low (magnification). I was unable to find any gonad tissue in a couple of the smaller oysters and I will have to go back and figure out a way to make a note of that. The two I was unable to find gonad tissue in thus far were HL-10_14 and HL-6_11.
Here is the URL for the drive where I will be uploading all my images:
Went into UW today and checked all the silos for larvae. Two out of the 13 individual oysters had spawned with one having thousands of larvae and one having only a few. Also possible presence ciliates, but there were larvae there!
Here are my counts:
In addition, I talked with Steven and Grace about how to image all of Laura’s histology slides from after the OA treatment. From there I will work to go through them all and look at gonad maturity between treatments and after just the overwintering as well as after the OA treatments.