Fast repetition rate fluorometer (FRRF) study of sea ice algae in low iron conditions. Algae were grown in an ice tank and the measurements were made at the end with a Chelsea Insrtuments FRRF. Materials and Methods (see the download document for original formatting and formulas) 1. Ice tank incubation The polar pennate diatom Fragilariopsis cylindrus, isolated from Antarctic pack ice in 2015 (Davis station, East Antarctica) was incubated in a purpose designed ice tank (Island Research, Tasmania). The ice tank, which was contructed of titanium to minimise dissolved Fe, was placed into a freezer (–20 degrees C), and the ice thickness and temperature gradient controlled by interaction between a basal heater and the adjustable ambient freezer temperature (see Kennedy et al., 2012). This enabled an ice thickness of approximately 5.5 cm to be maintained during the experiment. The diatom F. cylindrus was incubated in Aquil media (Price et al. 1989) buffered with ethylenediaminetetraacetic acid (EDTA) at 150 micro mol photons m−2 s−1 (PAR), a salinity of 35, and a Fe concentration of 400 nM, where the concentration of total inorganic forms of Fe (Fe') was 1.54 nM, this being continuously supplied to the medium and the exact value calculated using the software Visual MINTEQ, ver. 3.1 (https://vminteq.lwr.kth.se). Before a freezing cycle started, the seawater temperature was maintained at 2.5 degrees C, and a sample was obtained to assess the original physiological state of the algae (Day−5, hereafter). After obtaining the sample, the seawater temperature was set to −1.8 degrees C to initiate ice formation in the ice tank. Once ice had formed at Day−2, the under-ice seawater was partially replaced with ultrapure water to reduce the salinity down to 35, because the salinity had increased (to approximately 38) as a result of brine rejection from the ice. After a 2-day acclimation to the new salinity, ice samples were obtained every 5 days for 20 days (i.e., Days 0, 5, 10, 15, and 20). To minimize the heterogeneity among ice cores, ice samples were randomly collected from the tank chamber with a trace metal-free hand drill (2 cm in diameter) from randomly annotated grids on the ice surface, following normal random sampling numbers generated by the software R (https://www.r-project.org/). To assess the effects of melting and high light exposure, the ice was melted at 2.5 degrees C for 2 days. After the ice had completely melted, the seawater was exposed to a high light level, which was adjusted to represent the likely summer light intensity at the surface in ice-edge regions (800 micro mol photons m−2 s−1; MODIS Aqua), Seawater samples were obtained both after the melting and light exposure events (Melt and Light, respectively, hereafter). Fast repetition rate (FRR) fluorometry To monitor the photophysiology of F. cylindrus during the freezing and melting processes, variable chlorophyll a fluorescence (ChlF) measurements were conducted using a bench-top Fast Repetition Rate fluorometer (FRRf) (FastOcean Act2Run Systems, Chelsea Technologies) with Act2Run software (Chelsea Technologies). Ice samples were directly thawed at 2 degrees C in the dark for 30 min, and the slushily melted ice samples were placed in a quartz tube and their flouresence (ChlF) was measured. A single turnover protocol was applied for the ChlF measurements; 100 flashlets with 1 micro second duration at a wavelength 450 nm and 2 micro second intervals for excitation of reaction centres of photosystem II (PSII, hereafter), and 20 flahlets with 1 μs duration and 100 micro second intervals for relaxation. Eighteen light steps were applied to generate a rapid light curve (RLC) from 0 to 1800 μmol photons m−2 s−1, taking less than 5 min to complete one RLC. At each light step (~15 s), at least five induction and relaxation curves were averaged to obtain ChlF yields, described in Table, after calibrating the ChlF yields with filtered seawater. According to the models proposed

From the abstract of one of the papers: Oxygen microelectrodes were used to measure the photosynthetic rates of Antarctic fast ice algal mats. Using the oxygen flux across the diffusive boundary layer below the fast ice at Davis, a productivity range of 0-1.78mg C per square metre per hour was measured. This is at the lower end of fast ice productivity estimates and suggests that conventional carbon 14 techniques may overestimate sea ice algal mat productivity. Photosynthetic capacity (P max) approached 0.05 mg per C.(mg chlorophyl a) per hr. Onset of photosynthesis saturation, E k, was found at about 14 micromol photons per square metre per second. The irradiance of photoinhibition onset, E inh, was about 20 micromol photons per square metre per second and the irradiance at the compensation point, E c, was 4 micromol photons per square metre per second.

Metadata record for data from ASAC Project 102 See the link below for public details on this project. From the abstracts of some of the referenced papers: Six species of marine microalgae, namely Phaeodactylum tricornutum Bohlin, Dunaliella tertiolecta Butcher, Isochrysis galbana Parke, Porphyridium purpureum (Bory) Ross, Chroomonas sp., and Oscillatoria woronichinii Anis., have been examined with respect to their gas exchange characteristics and the inorganic carbon species taken up by the cells from the bulk medium. All species showed a high affinity, in photosynthesis, for inorganic carbon and low CO2 compensation concentrations. Such data are suggestive of operation of a 'CO2-concentrating mechanism' in these microalgae. Direct measurements of internal organic carbon pools in four of the species studied confirm this (O. woronichinii and Chroomonas were not tested). By comparison of achieved photosynthetic rates with calculated rates of CO2 supply from the dehydration of bicarbonate, it was shown that Phaeodactylum, Porphyridium and Dunaliella could utilise the bicarbonate present in the medium. Data for the other species were inconclusive although the pH dependence of K 1/2CO2 for photosynthesis by Oscillatoria indicated that this species too could utilise bicarbonate. Such observations could, however, not be used as evidence that, at least in the eucaryotic algae examined, bicarbonate was the inorganic carbon species crossing the plasmalemma as Phaeodactylum, Porphyridium and Dunaliella, and Isochrysis all showed the presence of carbonic anhydrase activity in intact cells as well as in crude extracts. 'External' carbonic anhydrase activity represented from 1/4 to 1/2 of the total activity in the cells of these algae. It is concluded that, as a consequence of a CO2-concentrating mechanism, photorespiration was suppressed in the marine microalgae examined although the data obtained did not allow any firm conclusions to be drawn regarding the species of inorganic carbon transported into the cell. Analysis of the age composition of a given species within a community is fundamental to any study of population dynamics and to the subsequent analyses of community interactions such as competition, succession and productivity. A problem exists in that calendar age often provides little information on the role played by any given individual plant within a population. For many populations the most useful definition of population structure is obtained from an analysis of both the functional age and the vitality of the component plants. Data from such studies on populations of marine macroalgae are lacking mainly because of the lack of suitable methods. This paper provides a review of the methods which have ben applied to such analyses in both terrestrial and marine communities, discusses these methods in the context of marine algae and presents the results of a case study on the analysis of population structure in the large brown alga Durvillaea potatorum. Evidence is presented for the occurrence of sexual reproduction including plasmogamy and meiosis, events previously undescribed in the life history of Ascoseira mirabilis. Ascoseira is monoecious. Gametangia are formed in chains within conceptacles. Synaptonemal complexes, structures concerned with chromosome pairing in meiosis, have been observed in the nucleus of gametangial initials. Mature male and female gametes have the same size and appearance, and resemble typical brown algal zoids. Sexual interaction begins after the female gamete settles down, and both zygotes and unfused gametes develop into sporophytes. It is concluded that Ascoseira has the same basic pattern of life history that characterises the order Fucales, and it is argued that this is probably the result of convergent evolution rather than being indicative of close phylogenetic relationship. Life histories are of central importance in understanding evolution and phylogeny of brown algae. Like other hereditary

The ASPeCt - Bio dataset is a compilation of currently available sea ice chlorophyll a (chl-a) data from pack ice (i.e., excluding fast ice) cores collected during 32 cruises to the Southern Ocean sea ice zone from 1983 to 2008 (Table S1). Data come from peer-reviewed publications, cruise reports, data repositories and direct contributions by field-research teams. During all cruises the chl-a concentration (in micrograms per litre) was measured from melted ice core sections, using standard procedures, e.g., by melting the ice at less than 5 degrees C in the dark; filtering samples onto glassfibre filters; and fluorometric analysis according to standard protocols [Holm-Hansen et al., 1965; Evans et al., 1987]. Ice samples were melted either directly or in filtered sea water, which does not yield significant differences in chl-a concentration [Dieckmann et al., 1998]. The dataset consists of 1300 geo-referenced ice cores, consisting of 8247 individual ice core sections, and including 990 vertical profiles with a minimum of three sections. An updated dataset was provided in 2017-12-15, which included a compilation Net CDF file.

The productivity of Antarctic waters may be controlled by the amount of iron. Experiments have shown that this is probably the case for phytoplankton but as yet we do not know if iron limits the growth of sea ice algae. This study will assess whether iron limits sea ice algae production and will conduct experiments to work out how these algae use iron. Measurements have been made to determine whether sea ice algae are limited by Fe. Sea ice samples were taken and this spreadsheet refers to those ice cores Columns A-G are self explanatory Column G is the depth in the ice core from the bottom Column H is the chlorophyll concentration in mg Chl m-2 Column I is the phaeophytin concentration in mg m-2 J is the total amount of protein in the sample ng m-2 K is the total amount of the protein flavodoxin ng m-2 L is the total amount of ferrodoxin ng m-2 These last two enable the Fe limitation status to calculated (not completed).

Field-based sampling: As part of Australian Antarctic Science project # 4298, a total number of 44 sea ice sites were sampled for bio-optical measurements along 4 transects on land-fast sea ice off Davis Station (Antarctica) during November – December 2015. Measurements included simultaneous hyperspectral down-welling (ice surface) irradiance (triplicate) and under-ice radiance (triplicate) measurements (320 – 900 nm, 3.3 nm resolution) with a TriOS ACC and Trios ARC radiometer, respectively. The radiance measurements were conducted with the TriOS ARC radiometer mounted onto an L-shaped arm (for deployment details see Melbourne-Thomas et al. 2015). Subsequently, snow thickness was measured with a ruler and an ice core was collected directly above the radiometer location. Sea-ice freeboard (tape measure) and ice thickness (ice core length) were also recorded. Ice cores (9 cm internal diameter) were cut into sections, and these were melted in the dark at +4 degrees C, filtered onto GFF filters and then used to measure ice algal pigment content (using High Performance Liquid Chromatography (HPLC) and spectral ice algal absorption coefficients (ap, ad, aph) for entire vertical profiles or for the lower-most 0.1 m of ice cores. The location of the sampling grid had its origin (x=0, y=0) at GPS position: -68.568904, 77.945439. Transects (128m – 512 m in length) started at x=60, x=70, x=80 and x=90 m and were sampled at y-positions of 0m, 0.5m, 1m, 2m, 4m, 8m, 16m, 32m, 64m, 128m, (256m, and 512m) on 19/11/2015, 23/11/2015, 29/11/2015 and 02/12/2015, respectively. Analysis of ice algal chlorophyll a concentration: For pigment analysis, 0.25 to 1.0 litres of melted ice core subsamples were passed through 25 mm diameter glass-fiber (Whatman GF/F) filters. The filters were then frozen and stored below −80 degrees C prior to analysis using HPLC. Samples were extracted over 15 to 18 hours in acetone before analysis by HPLC using a modified C8 column and binary gradient system with an elevated column temperature [Van Heukelem and Thomas, 2001]. Pigments were identified by retention time and absorption spectra from a photo-diode array (PDA) detector, and concentrations were determined from commercial and international standards (Sigma; DHI, Denmark). Analysis of particulate (algal and non-algal) absorption: The optical density (OD) spectra of the particulate material on these filters (see section above) were measured over the 350 to 750 nm spectral range in 0.9 nm increments, using a Cintra 404 UV/VIS dual-beam spectrophotometer equipped with an integrating sphere. The pigments on the sample filter were then extracted using the method of Kishino et al. [1985]'s method to determine the OD of the non-algal particles in a second scan. The OD due to ice algae was then obtained by calculating the difference between the optical density of the total particulate and non-algal fractions. The OD measurements were converted to absorption spectra using blank filter measurements, and by first normalizing the scans to zero at 750 nm and then correcting for the path length amplification using the coefficients of Mitchell [1990]. A detailed description of the method is given in Clementson et al. [2001], and followed SeaWiFS protocols [Muller et al., 2003]. An exponential function was fitted to all spectra of non-algal particulate material: ad(λ) = ad(350 nm) exp[−S(λ − 350 nm)] + b, (1) where ad(λ) is the residual absorption coefficient over the wavelength (λ) range 350 to 750 nm of the particles after methanol extraction, also referred to as absorption of detritus [m−1] although this may include absorption of non-extractable pigments and heterotrophic protists. A non-linear least-squares technique was used to fit Equation 1 to the untransformed data, where S and b are empirically-determined constants. The inclusion of an offset b allows for any baseline correction. In some samples, pigment extraction was incomplete, leaving small residual peaks in

Chlorophyll data was used to measure growth rates of sea ice algae in CO2 incubations. Sea ice brine microalgae was collected from sackholes. Replicate samples were incubated in ambient air (~0.04% CO2), 0.1% CO2, 1.0% CO2 and 2.0% CO2 concentrations. Three incubation experiments were carried out at SIPEX stations 4 (expt 1) 7 (expt 3) and 8 (expt 4). Growth rate calcualtions followed a standard exponential growth model, i.e Bf = Bi x e(rt) Where Bf equals final biomass, Bi equals initial biomass, r = growth rate and t = time (in days).

The distribution and abundance of ice-associated copepods in the fast ice of the Australian Antarctic Territory were investigated over a distance of approximately 650 km between October and December 1995. The six sites where collections were made were: offshore from Mawson station, Larsemann Hills (including Nella Bay), Rauer Islands (ice edge near Filla Is), O'Gorman Rocks and Bluff Island near Davis Station, and Murphy Rocks in the northern Vestfold Hills. Ice cores were obtained using SIPRE ice augers. Five to ten cores were collected along transects several km in length. Thickness of sea ice and snow cover were measured at each sampling site. Chlorophyll a concentrations were determined for each core. Copepods were isolated from the melted core water and identified and counted. Zooplankton tows were also made at each site where cores were collected. Nine species of copepods were identified from the cores. However, of these, only three were recorded regularly: Paralabidocera antarctica, Drescheriella glacialis and Stephos longipes. The abundance of copepods ranged between 0 and 147/L. The highest densities were recorded at the Larsemann Hills and the lowest at Murphy Rocks. Within the cores, the highest abundances were found in the bottom 10 cm of ice, irrespective of the species. Chlorophyll a concentrations ranged between 0.9 and 373 mg/m3. Data available: excel files containing sampling dates, sampling sites and abundances (number per L) of three dominant sea ice copepods, Paralabidocera antarctica, Drescheriella glacialis, Stephos longipes. Data are presented for developmental stages (nauplii, copepodites and adults) where available. Totals are also provided. Vertical distribution in some cores is also provided. Chlorophyll a concentrations (ug/L) provided for most sites. Detailed information about each of the spreadsheets is provided below: The chlorophyll spreadsheet shows chlorophyll concentrations for 5 sites in the AAT. The column headings are: core - reference number of the core collected subsection - depth in the core in cm volume - vol of melted core water volume added - 1 L of filtered seawater for melting % original - amount of total that core water represents (i.e. minus the 1L added) aliquot - volume subsampled for chlorophyll analysis acetone - amount added (mL) for extraction 750, 664, 647, 630 - wavelengths where absorbance was measured chloro a - amount of chlorophyll a in the sample ug/L - chloro a expressed as a concentration The spatial spreadsheet shows species abundances of three copepods at 4 sites N1 to NVI - nauplius stage 1 to 6 of a species CI to CVI - copepodite stage 1 to 6 of a species F or M - female or male of copepodite stage 5 or 6 1,1 etc - cores 1 and 2 from site 1 within a major location (e.g. 2 cores close together in the Larsemann Hills) The temporal spreadsheet shows abundances over time at 2 sites (O'Gorman Rocks, Bluff Is) near Davis and two species (Paralabidocera antarctica and Drescheriella glacialis) on several sampling dates N1 to N3 - total nauplii in each of three cores (i.e. not separated into stages as above) C1 to C3 - total copepodites A1 to A3 - total adults Then at the bottom are the means of each three cores.