DISCLAIMER
All views and opinions expressed on this page are those of the individual and not necessarily those of the University of Southampton.
Plymouth Field Course 2019
Group 7
Flow cytometry
The flow cytometry instrument was used to quantify the numbers of numbers of cells within the samples taken offshore and image the large cells that might be of interest. 5 samples were taken offshore on Callista for flow cytometry analysis; one sample was taken at Station 18 at 24.3m and four samples at Station 23 at 39.4, 26.2, 9.9 and 3.2m depth in order to produce a profile through the water column. 50ml of sample water from the Niskin bottle was added to glutaraldehyde to preserve it. The samples were run through the machine in the lab in order to get cell counts and quantify the red fluorescence (chlorophyll) present.
The graph shows that microplankton are making a much smaller contribution to total cell counts and total chlorophyll than nanoplankton, which are numerically dominant. Both the red fluorescence and cell counts peak at the sample taken at 26.2m depth, suggesting the presence of a deep chlorophyll maximum between 10 and 40m.
DAPI (4',6-diamino-2-phenylindole dihydrochloride)
Purpose:
DAPI testing was also carried out to determine the abundance of microbes in water column samples. Multiple DAPI samples taken from a single CTD measurement and total cell counts were compared to photosynthetic cell counts to make deductions about the position and abundance of chlorophyll within the water column.
Method:
A CTD was deployed on the 5th of July 2019 and at 13:03:07 UTC from the position N50ᵒ03.227’ W4ᵒ23.224’, near the E1 station. Four Niskin bottles were deployed at different depths to collect samples that were then readied for the lab. Here the sample was filtered and the filter used placed under a slide. The slide was then taken to the MBA to be analysed under the Leica SP8 confocal microscope.
DAPI is a blue fluorescent cell stain that binds to AT regions of DNA, staining every microbial cell fluorescing bright blue when exposed to 405nm of light. This can be compared to the photosynthetic cells that will be excited by within the region of 650-675nm and show up as red. Upon counting each individual microbe for both wavelengths the percentage of microbes in the sample that have photosynthetic properties can be found.
Using these counts and the knowledge of how large the filter paper and volume of water filter mean cells per ml were calculated; this number and multiplied by 12.5 fg (average microbe size) yielded a biomass estimate.
A fourth sample was collected from a depth of 3.2 metres, however this was not analysed under the light microscope due to time pressures and the conclusion being drawn that this sample was closest to the surface and therefore of the least importance for microbial abundance
Only one DAPI sample was successfully analysed as there were issues with the DAPI staining. This may be due to formaldehyde being used instead of para-formaldehyde meaning that the DAPI smudged easily on the slides making it impossible to count individual microbes for all samples apart from bottle 12. The percentage of microbial biomass that are chlorophyll for this bottle can then be calculated. This percentage comes to 29% but is obsolete without anything to compare it to.
Looking at just the chlorophyll biomasses, it’s apparent that the greatest number of microbes were found at 26.2m. This is to be expected, as we inferred on the boat that the deep chlorophyll maximum is found at this depth within the water column due to the high levels of PAR irradiance recorded. This is concordant with all other data points; biomass is much lower for the chlorophyll at 39.4m. The sample at 9.9m gives a value that lies closer to sample 12 than the deeper sample. This would be expected before the deep chlorophyll maximum.
Zooplankton
Introduction
Zooplankton samples were collected at all Stations but Stations 21 and 24, at which only CTD profiles were taken; these samples were then analysed, identified and counted in the lab on 06/07/2019.
Method
Depth of zooplankton net samples were first decided based on ADCP backscatter data and features observed on the CTD profile.
A 200µm net with a 50cm diameter opening and a 1L bottle was then deployed by the deck crew in order to collect the plankton. The net was closed manually at desired depths by deploying a weight along the line which, once reached the net, fired the closing mechanism. The first time the net was deployed, the closing mechanism failed due to an entanglement with the rope. Once retrieved to deck the net was washed down with an inline seawater pump fed hose, to wash plankton into the sample bottle. The 1L sample then had 100% formalin added to it and was stored in a cool box to preserve the sample.
Lab processing
Samples collected during the offshore boat work were then analysed within the lab on 09/07/2019. 10ml from each bottle was divided into 2x5ml counting chambers under a light microscope. Numbers of zooplankton were counted and consequently scaled up to give the full m3 concentrations.
Results
Zooplankton abundances generally correlated with expectations based on ADCP backscatter and CTD profiles. At all stations, dinoflagellates and copepods were the dominant zooplankton collected, though proportions of these groups differed between sites. Lower abundances of other taxa were also observed throughout, such as Cladocera, Polychaeta and Appendicularia. Stations further onshore with less developed stratification were dominated by dinoflagellates; Stations 22 and 23 offshore with more pronounced stratification exhibited higher abundances of copepods. Between 42m and 38m depth at Station 22, where high backscatter but high transmissivity was observed, zooplankton were expected; copepod abundances were estimated to be 26865 per m3. Abundances remained high at this station at different sample depths, with dinoflagellate abundance also being high between 24m and 20m around the DCM. Similarly, copepods dominated zooplankton abundance at depth at Station 23 (Figure 2b-c), though with decreasing depth dinoflagellate abundance increased dramatically, and just above the chlorophyll maximum, dinoflagellate abundance reached 29030 per m3 as they became the most abundant taxon. Figure 2a presents the depth profile at this Station, with Figures 2b-d presenting proportions of taxa at different depths.
Phytoplankton
Station: C18
At the depth of 9m, low species diversity (3 species) and low abundance were observed probably due to low nutrient concentrations.
Deep chlorophyll maximum was observed at depth of 24m. Where there is an increase in species diversity (5 species) and in abundance in relation with the shallower depth sample.
Station: C19
At the depth of 6.5m, very low species diversity (1 species) and very low abundance were observed probably due to low nutrient concentrations.
And at the depth of 26m, deep chlorophyll maximum was observed. Where there is a big increase in species diversity (7 species) and in abundance in relation with the shallower depth sample.
Station: C20
At the depth of 2.8m, low species diversity (4 species) and low abundance were observed probably due to low nutrient concentrations.
A small chlorophyll maximum was found at a depth of 19m. However, low species diversity (4 species) and very low abundance were observed.
Finally, at depth 44m, surprisingly there is a substantial increase in species diversity (8 species) and a small increase in abundance.
Station: C22
At depth 4.6m a coccolithophore bloom probably exists. As both ADCP had scatter in top 10m, where Coccolithophores are expected, and low transmissivity was found in the region. Also, chalky colour was profound in the area suggesting that coccolithophores exist. Furthermore, the low abundance of other phytoplankton species in this station, may suggest nutrient depletion from a coccolithophore bloom. However, it is not possible to analyse the data until late September, when the university of Southampton opens, as there is no access to SEM in Plymouth Marine Station.
The deep chlorophyll maximum was found at a depth of 24m. Where low species diversity (3 species) was found but high abundance of the Chaetoceros family was observed.
Finally, at depth 39m low abundance and low species diversity (4 species) were observed due to small irradience.
Station: C23
At Depth 3.2m, a coccolithophore bloom probably exists. As both ADCP had scatter in top 10m, where Coccolithophores are expected, and low transmissivity was found in the region. Also, chalky colour was profound in the area suggesting that coccolithophores exist. Furthermore, the low abundance of other phytoplankton species in this station, may suggest nutrient depletion from a coccolithophore bloom. However, it is not possible to analyse the data until late September, when the university of Southampton opens, as there is no access to SEM in Plymouth Marine Station.
At the depth of 10m low nutrients were found, and low species diversity (3 species) and moderate abundance.
Deep chlorophyll maximum was found at 26m, where low species diversity was found but high abundance of the Chaetoceros family was observed.
Finally, at depth 39m very low abundance and species diversity were found due to small irradiance.
Figure 1: Graph depicting Red Fluorescence (mg/l^3) and cell counts of nano and micro plankton and total values per 100 ml in relation with depth (m). depicting Red Fluorescence (mg/l^3) and cell counts of nano and micro plankton and total values per 100 ml in relation with depth (m).
Table 1: Table displaying bottle numbers, Dapi/chlorophyll, depths sampled (m) and biomass.
Figure 2: a) Graph of salinity, temperature and fluorescence vertical profiles of station
b) Pie chart visualizing zooplankton (and dinoflagellates) species composition of station C23 at depth: 18-16 meters. Dinoflagellates: 29030; Copepoda: 9167; Chaetognatha: 3056
d) Pie chart visualizing zooplankton (and dinoflagellates) species composition of station C23 at depth: 35-30 meters. C23. Copepoda: 5297; Copepoda nauplii: 713; Dinoflagellates: 713
c) Pie chart visualizing zooplankton (and dinoflagellates) species composition of station C23 at depth: 28-24 meters. Copepoda: 14388; Dinoflagellates: 10695
b) Pie chart visualizing phytoplankton species composition of station C18 at the depth of 24 meters (Ceratium lineatum 1 cells/ml, Guinardia flaccida 1 cells/ml, Rhizosolenia imbricata 2 cells/ml, Ceratium furca 9 cells/ml, Lauderia annulata 4 cells/ml).
Figure 3: a) Pie chart visualizing phytoplankton species composition of station C18 at the depth of 9.2 meters (Guinardia flaccida 1 cells/ml, Nitzchia spp. 2 cells/ml, Guinardia deliculata 2 cells/ml).
Figure 4: a) Pie chart visualizing phytoplankton species composition of station C19 at the depth of 6.5 meters (Ceratium furca 1 cells/ml).
b) Pie chart visualizing phytoplankton species composition of station C19 at the depth of 26 meters (Diploneis 1 cells/ml, Rhizosolenia 1 cells/ml, Coscinodiscus 1 cells/ml, intinnids 5 Mesodinium 1 cells/ml, Euglena 1 cells/ml, Proboscia 2 cells/ml).
Figure 5: a) Pie chart visualizing phytoplankton species composition of station C20 at the depth of 2.8 meters (Ceratium lineatum 8 cells/ml, Thalassiosira 5 cells/ml, Skeletonemia 14 cells/ml, Leptocylindrus danicus 1 cells/ml).
b) Pie chart visualizing phytoplankton species composition of station C20 at the depth of 19 meters (Mesodinium 4 cells/ml, Guinardia 3 cells/ml, Ceratium lineatum 1 cells/ml, Prorocentrum 2 cells/ml).
c) Pie chart visualizing phytoplankton species composition of station C20 at the depth of 44 meters (Coscinodiscus 2 cells/ml, Euglena 1 cells/ml, Protoperidinium 2 cells/ml, Mesodinium 1 cells/ml, Phaeocystis colony 1 cells/ml, Ceratium fusus 2 cells/ml, Pseudonitzschia 4 cells/ml, Dinoflagellate 2 cells/ml).
Figure 6: a) Vertical profile graph of Temperature, Salinity and Fluorescence at station C22.
c) Phytoplankton species composition of station C23 at the depth of 26 meters (Chaetoceros 524 cells/ml, Ceratium lineatum 1 cells/ml, Rhizosolenia alata 1 cells/ml, Guinardia flaccida 1 cells/ml).
d) Phytoplankton species composition of station C23 at the depth of 39 meters (Cylindrotheca closterium 1 cells/ml).
b) Phytoplankton species composition of station C23 at the depth of 9.9 meters (Leptocylindrus danicus 1 cells/ml, Chaetoceros 160 cells/ml, Mesodinium rubrum 2 cells/ml).
Figure 7: a) Pie chart visualizing phytoplankton species composition of station C23 at the depth of 3.2 meters (Dinoflagellate 4 cells/ml, Striatella 2 cells/ml, Chaetoceros 200 cells/ml).
b) Pie chart visualizing phytoplankton species composition of station C22 at the depth of 4.6 meters (Cylindrotheca closterium 1 cells/ml).
c) Pie chart visualizing phytoplankton species composition of station C22 at the depth of 24 meters (Pleurosigma 5 cells/ml, Chaetoceros spp 341 cells/ml, Rhizosolenia imbricate 41 cells/ml).
d) Pie chart visualizing phytoplankton species composition of station C22 at the depth of 39 meters (Lauderia annulata 13 cells/ml, Leptocylindrus danicus 5 cells/ml, Chaetoceros sp 37 cells/ml, Mesodinium rubrum 1 cells/ml).