Feature ArticlesExpanding Applications For Variable Fluorescence Fluorimeters
By Justin Dunning
Chelsea Technologies Group Ltd.
West Molesey, England
Since the introduction of commercially available variable fluorescence fluorimeters in the 1990s, these tools have been applied to a growing number of applications in which information on phytoplankton physiological state within ocean waters is required. Data from such fluorimeters had originally been driven by research requirements, and the parameters generated have taken time to reach maturity. There is now a general acceptance of these fluorimeters’ value for inclusion within operational monitoring programs.
The latest generation of variable fluorescence fluorimeter from London, England-based Chelsea Technologies Group (CTG), the FASTtracka II, has seen a significant increase in applications over the last two years. These range from traditional oceanographic research—for example, in characterizing productivity within the world’s oceans—to monitoring water quality in coastal systems. With increased functionality and adaptability, detailed analysis of phytoplankton within the laboratory (whether internally cultured or sampled from natural systems) can now be achieved.
The variable fluorescence of chlorophyll a allows interrogation of oxygenic photosynthesis, particularly the functioning of photosystem II (PSII). A number of parameters are obtained, including, among others, variable fluorescence/maximum fluorescence value (Fv/Fm), which provides an estimate of PSII efficiency. Two common methods are used, one being fast repetition rate fluorescence (FRRF), which provides a single turnover (ST) protocol, and pulse amplitude modulation, which provides a multiple turnover (MT) protocol. The advantage of the ST protocol is that the measurement is made very quickly, and it allows analysis of moving samples. ST protocol also benefits from providing additional kinetic data relating to PSII flux. The original FASTtracka was restricted to measurements using the ST protocol. The FASTtracka II measures in both ST and MT protocols.
Success of the original FASTtracka fluorimeter could be attributed to the fact that it was unique as an in-situ device providing FRRF protocols. Two fluorescence chambers (one for ambient readings and one for dark adaptation) were provided in the one instrument, yet many users were only utilizing one. This prompted CTG to divide these chambers for the successor instrument to form the basis of a flexible deployment instrumentation package.
Since the FASTtracka II instrument was released in 2007, CTG has continued to develop tools and accessories with a focus on widening its applications.
The first and possibly most significant development was a user-friendly software interface that could provide robust, real-time-processed FASTtracka II data as well as a utility to process and manipulate recorded data. The FASTtracka II graphical user interface (GUI), FASTpro, offers flexibility in the comparison of data sets and the selection of data for processing.
A key request from the FASTtracka user community was the capability to take FASTtracka II readings under controlled light conditions. This prompted the development of the FASTact system, which is provided as an add-on to the current FASTtracka II instrument. FASTact provides a sample chamber that incorporates white light-emitting diodes, a water jacket (for temperature control) and a pump. FASTact allows users to conduct rapid light curves (RLCs) on low-volume samples and includes such features as a sample mix function and automated sample change. FASTpro gives users complete control of RLC settings and allows such features as dark adaptation between selected steps.
In-situ measurements of both dark-adapted and ambient readings can still be obtained simultaneously (as with FASTtracka) by deploying two FASTtracka II instruments together. If rapid profiling or towing operations provide insufficient time for samples to dark adapt, water samples may be taken and analyzed with the FASTact system for accurate readings.
Optional depth and photosynthetically active radiation sensors are available that interface directly to the FASTtracka II, and data from these sensors are recorded and displayed by the FASTpro software. These and other accessories allow the user to configure the system for a range of applications.
Fish Survival Assessments
The University of British Columbia has been taking FASTtracka II measurements as part of a large field study in a fjord on the central coast of the province.
Research associate Dr. Brian Hunt explained that the tool is being used to measure primary production as part of a larger program, the Rivers Inlet Ecosystem Study, which is investigating the ecosystem’s seasonal cycle, interannual variation, response to environmental conditions and, ultimately, drivers of sockeye salmon population fluctuations. Water samples are collected at discrete depths and brought to the surface, where controlled FASTtracka II runs are conducted under conditions of unlimited light and complete darkness.
These measurements are being conducted on regular two-week intervals at three separate sites. It is planned that in-situ measurements will be made at these sites this year. Ultimately, data from the FASTtracka II will feed into a trophic model describing the functioning of the ecosystem’s food web.
The Royal Netherlands Institute for Sea Research will be using FASTtracka II as part of a coastal monitoring project, IN-PLACE, in the western Wadden Sea. This program is financed by the Coast and Sea Program of the Netherlands Organization for Scientific Research.
Dr. Jacco Kromkamp of The Netherlands Institute of Ecology reports that the FASTtracka II data will be used to measure primary production online using an automated system installed on a continuous monitoring platform. Nutrients and water currents will also be recorded. FASTtracka II data will be calibrated against standard carbon-14 measurements. The same instrumentation will also provide cruise data about spatial patterns in nutrients; conductivity, temperature and depth; and primary production.
Previous work with the FASTact system has included analysis of samples taken at five stations in the Wadden Sea, each over a period of 13 hours on two-hour intervals, to assess changes across the tidal cycle. High-quality RLC data from low phytoplankton concentrations were obtained.
“The setup used allows the frequent measurements of photosynthesis parameters, which are otherwise difficult to obtain,” Kromkamp said.
Iron Fertilization Experiments
The National Institute of Oceanology (NIO), India, deployed FASTtracka II during the Lohafex experiment in early 2009 in the Southern Ocean. This Indo-German cruise on the RV Polarstern was part of the Council of Scientific and Industrial Research (India) network program titled “Atmosphere Carbon Di-oxide Sequestration Through Fertilisa-tion of High-Nutrient Low-Chlorophyll Oceanic Region With Iron.” The FASTtracka II was sent down to 200 meters on a cable winch system. It measured phytoplankton photosynthetic efficiency inside and outside of the iron-enriched patch over a 41-day period.
Dr. Mangesh Gauns of the NIO reported that the data were collected during the expedition using two systems, one for profiling and one for underway measurements. This application builds on previous FASTtracka deployments during iron fertilization experiments, such as the Southern Ocean Iron Enrichment Experiment, also conducted in the Southern Ocean in 1999, and the earlier IronEx program in the Equatorial Pacific.
Monitoring Algal Blooms
The University of Copenhagen and Aarhus University in Denmark deployed FASTtracka II during a project called “The North Atlantic Bloom Experiment 2008” under a National Science Foundation-funded ocean carbon and biogeochemistry program.
The FASTtracka II was used to examine processes related to the spring bloom (its development and culmination) during April and May of 2008 in the waters southeast of Iceland.
Data from the FASTtracka II was also used to provide calibration material for a number of instrumented floats and gliders that were deployed during the project.
“Thanks to the Fv/Fm data, we could determine that some part of the spring bloom sank out of surface waters in spite of the fact that the phytoplankton were, with respect to their photosynthetic physiology, in good condition,” professor Katherine Richardson said.
The data noted considerable differences in Fv/Fm in surface waters over the study area.
Data from the FASTtracka II have allowed the group the opportunity to analyze the state of the phytoplankton bloom in real time.
Water Contamination Detection
The FASTtracka II was installed in a flow-through configuration at water intakes to water processing plants, including the Washington Suburban Sanitary Commission’s facility at Rocky Gorge, Maryland; Veolia Water’s (Paris, France) plant in London; and United Utilities’ (Warrington, England) facility in Lancashire, England. Trials have been conducted with these units to generate “real-life” fluorescence data for use in the development of a fully automated red, amber and green (RAG) system for the detection of contamination events.
The RAG detection system is fully integrated into the FASTtracka II FASTpro GUI. A significant number of potential toxicants have been tested with the FASTtracka II, and it has been demonstrated within laboratory conditions that many of these toxicants attack PSII in different ways, resulting in changes to different FASTtracka II parameters. During the course of these trials, the RAG system was developed to produce robust detection, reporting negligible false positives.
In addition, a new flow-through manifold has been developed, triggered by early issues with silt affecting FASTtracka II readings.
The new manifold has been designed to provide rapid, near-laminar flow across the surface of the front lens, which minimizes the opportunity for silt to accumulate. Results from this design have shown little to no silt accumulating on the lens or inner chamber surfaces.
Success of the system was confirmed in late 2009, when the system installed on the site in West London singularly detected a contamination event over a 12-minute period that was later confirmed and identified as triclosan at levels of 22.6 micrograms per liter.
Laboratory Culture Monitoring
The National Oceanography Centre is utilizing a FASTtracka II on a number of programs funded by the National Environmental Research Council, among others. In addition to in-situ deployments, such as within the SeaSoar towed undulating vehicle, they regularly utilize the FASTtracka II to investigate the physiological response of a diverse range of phytoplankton types to variable environmental growth conditions in the laboratory.
Similarly, the University of Essex is taking FASTtracka II measurements from laboratory cultures and making use of the FASTact system to generate RLCs on a number of these cultures. Their work includes comparison of measured gross oxygen evolution with gross photosynthesis calculated from fluorescence on individual culture species.
Justin Dunning has been sales manager at Chelsea Technologies Group (CTG) since 1996, selling CTG’s range of environmental sensors within the ocean, coastal and inland water sectors as well as the industrial processing sector. Before this, he was responsible for the marine systems group within CTG’s research and development department.