Progress Reports: Ocean acidification and emerging diseases in the Pacific Northwest | Roberts Lab
Wild-collected adult C. gigas were strip spawned and gametes
(separate eggs and sperm) were pooled. Fertilizations occurred
in water equilibrated to two different pCO2: 380 ppm (current
levels) and 840 ppm (near end-of-century estimation).
<<WOW!!!>>
Fertilization times were staggered and there were 3 fertilization
times per treatment with 3 replicates per time (9 replicates per
treatment). Developing embryos were sampled at times
post-fertilization that correspond to important stages in
development: 1 h, 2h, 5h, 17h, and 24h. We collected data on the
proportion of larvae that had reached cleavage, hatching, etc.
for the different time points. We found that the larvae
developing in the 840 ppm treatment were slower to develop than
larvae at 380 ppm.
In collaboration with the Northwest Fisheries Science Center
(NOAA, NWFSC) we reared larvae until 48 hours post-fertilization
at 4 different pCO2 treatments:
280 ppm (pre-industrial levels),
380 ppm (current day), 750 ppm (projected mid-century), 2000 ppm
(pessimistic end-of-century). <<MORE POISON Dr. Frankenstein>>
There were 6 replicates per treatment.
As described above, adult wild-collected oysters were strip spawned and larvae were monitored in the different
treatments for the ability to reach developmental milestones,
percent fertilization, morphological abnormalities, and swimming
activity. At 24 hours post-fertilization larvae were also
assessed for percent calcification using polarized light. Also
at 24 hours post-fertilization we took samples of larvae from 3
larval chambers per treatment to store for gene expression
analysis.
We found that C. gigas larvae in this experiment had
the greatest success of calcification at 380 ppm and the poorest
at 2000 ppm. Calcification at 280 and 750 ppm decreased when
compared to 380 ppm, but the majority of the larvae were fully
calcified.
<<Lemme repeat a part of that>>
Calcification at
280 and 750 ppm decreased when
compared to 380 ppm, but the majority of the larvae were fully
calcified.
<<They couldn't hack the pre-industrial level of CO2>>
The greatest proportion of larvae with normal
morphology was observed at 380 ppm and the greatest incidence of
abnormal morphology was observed at 2000 ppm. Gene expression
analysis of hsp70 revealed elevated expression at 750 ppm when
compared with 380 ppm, but decreased expression at 2000 ppm
compared to 750 ppm. One intepretation of this is that the
larval stress response reaches a threshold in its ability to
respond to this stressor in water that in greater that 750 ppm
pC02.
Disease susceptibility of Pacific oyster larvae at elevated ��atm
CO2 levels- Disease challenges were performed exposing C. gigas
larvae to a combination of elevated ��atm CO2 and Vt.
These
experiments examined C. gigas larval susceptibility to vibriosis
caused by Vt at three ��atm CO2 levels (ambient (~380), 750 and
2000 ppm ��atm CO2). Specialty gas mixtures of air and CO2 (2000
and 750 ppm ��atm CO2) (Praxair, Inc.) were used to produce
elevated ��atm CO2 conditions.
No significant differences in larval susceptibility was detected
at elevated either elevated ��atm CO2 when compared to ambient
levels.
Although LD50 values are lower for prodissoconch I larvae
at 2000 ��atm CO2, no significant differences were detected when
compared to ambient or 750 ��atm CO2 (Table 5). Figures 3a and 3b
illustrate proportions of larval survival at both larval stages
over 72 hours of exposure.
Again, no significant differences in
larval survival was seen at elevated ��atm CO2.
Further statistical analyses are still underway to further validate these
findings.
<<I BET -- NOAA must be fuming>>
Oyster Larvae Growth and Calcification at Three Different pCO2-
Experimental conditions were maintained using a flow-through
seawater system in Friday Harbor, Washington, USA.
Three
experimental treatments were chosen to correspond with dissolved
CO2 levels of 400, 700 or 1000 ppm in the atmosphere. These
levels correspond to near current ambient oceanic conditions,
projections for mid-century pCO2, and end-of-century,
respectively (IPCC 2007).
Larval size (shell height and hinge length) was similar across
experimental treatments after 24 hours, however by day 3 larvae
grew significantly larger (height and length) in the Ambient and
MidCO2 compared to the HighCO2 treatment. Between days 1 and 3
larvae increased in size under Ambient conditions (shell height,
P < 1e-7) and MidCO2 conditions (shell height, P < 1e-7; Figure
4). Developmental rate did not vary across treatments.
As part of second experiment, C. gigas juveniles were exposed to
6 different levels of pCO2 (the same as above) for 1 month. At
the end of the month, oysters were sampled for transcriptomics,
proteomics, and histology. A subset of the oysters were subjected
to mechanical stress and similarly sampled. The goal of the
mechanical stress was to simulate a general secondary stressor to
see if OA affects response to other stress. Another group of
oysters from each treatment was subjected to one of three
temperatures representing thermal stress: 44°C (lethal
temperature, or LT), 43°C (LT-1°C), or 42°C (LT-2°C). Mortality
was monitored over a week and any surviving oysters were sampled
at the end of the week.
There was slightly more shell growth at ambient conditions
compared to all elevated pCO2 conditions, although there was no
overall consistent trend in the effect of pCO2 on shell growth.
We also accomplished preliminary proteomics on one sample to
ensure that we will be able to sequence and identify proteins.
The sequencing was successful and we were able to annotate almost
300 peptides.