The Dark Decay Assay (DDA)
for aquatic plankton ecosystems
by Dr J Floor Anthoni
First published: January 2005; On web: 10 Aug 2005.

Since the discovery of the missing ecological factor in the sea, the planktonic decomposers, and the formulation of the Plankton Balance hypothesis, the search continued to measure and quantify the two most important components of planktonic ecosystems, the phytoplankton and the decomposers. This led to the discovery of a new procedure that enables anyone to measure the most important plankton characteristics using a simple pH meter. The DDA has opened a new window of interest on aquatic ecosystems, discovering new environmental laws in the process. This document describes the ideas behind it, methods used and results obtained. From necessity, the examples used are all from New Zealand. 

Important notice! The information provided here is NOT in the 'Public Domain' and remains the intellectual property of Dr J Floor Anthoni. By using the described methods, you may infringe copyrights unless being registered. Please read the conditions of use and how to register (register.htm). Also please note that the documents provided here are 'living' documents, being updated from time to time.

An introduction into the history of the discovery of the missing ecological factor, the Plankton Balance hypothesis and the Dark Decay Assay. Principles behind the method, results and more.   (16 pages, this document)
Rather than measuring physical quantities, the DDA measures the interaction between the most important components of plankton ecosystems. It measures the health of the aquatic environment. As a result, it overturns much of our thinking. (on this page)
With the DDA several important ecological discoveries were made. (on this page)
The Dark Decay Assay method has many applications, summarised here (on this page)
A summary of new and old words used in the DDA chapters (on this page)
Further reading (on this page)
Recent changes and additions (on this page)
DDA for dummies
This chapter begins from the beginning, assuming you know little or nothing at all about ecosystems, the sea and degradation. Also for people who have little time, read this first. (10 pages)
Complete instructions on how to do the DDA method, equipment, limitations, pitfalls, calculations and more. A practical manual. (8 pages)
A full description of the equipment required and how to make or adapt your own (3 pages)
Many tests have been performed to improve the accuracy of the DDA and to understand how it works. Using simple tests, the DDA has been verified but more needs to be done. A history of how the DDA developed. Fascinating and revealing. (18 pages)
What is pH and how is it metered? How does a pH meter work?  How best maintained? (3 pages)
Read the conditions of use and register now with easy incremental payments through PayPal. (3 pages)
Frequently Asked Questions about the DDA method. (2 pages)
Scientific publications that either support or refute our findings. Most interesting reading. (2 pages, growing)
A list of people, institutions, businesses and community groups who wish to be networked. (1 pages)
marine studies
Studies made with the DDA in the marine environment, supported by further experimentation. 
Note! for best printing results, narrow the page margins to 0.3" and 0.2" and top/bottom to 0.4"
  • Mapping the sea (1): mapping biodensity & RoA around Northland, New Zealand, Feb-May 2005. (10 pages)
  • _
  • freshwater studies
    Studies made with the DDA in the fresh water environment. Because freshwater plankton ecosystems are more stable and less complex than their marine counterparts, new insight is gained.
  • fresh01.htm: Seasonal studies of Rotorua, Waikato and Northland freshwater lakes (10 pages, growing)
  • _
  • aquariums
    The marine aquariums at the Seafriends field centre in Leigh are based on a self-circulating ecosystem. As a result, the degradation happening in the sea has been experienced there too. With the DDA technique these aquariums are now being prograded and this reverse process gives new insight about degradation and indicator species. Interesting and challenging. ( in progress)
  • aquarium description (aqua1.htm): a description of the Seafriends ecosystem aquariums, how they work, how they are managed and how they are prograded. (10 pages)
  • aquarium species (aqua2.htm): an annotated list of species and how well they fare now compared to before. (9 pages)
  • progradation timeline (aqua3.htm): an annotated timeline of events and observations, with photos. (in progress)
  • _
  • further research
    The (simple and exploratory) research done so far can only open a view to further possibilities, many of which require extensive instrumentation and scientists in many different scientific disciplines. Here is a wish list of what could be done next. It also gives you an insight into the limitations encountered so far. (postponed)
    related chapters
    Supporting documents: documents found elsewhere on this web site.
  • principles of conservation: resource management, biodiversity, conservation principles, marine conservation, marine reserves and much more (a very large section and important reading).
  • principles of degradation: the coastal seas are degrading precipitously. Why is this happening, and why is it happening so fast? Why also in New Zealand and why now? What are the principles behind it? (27 pages)
  • the Plankton Balance Hypothesis: the most important ecological factor affecting marine life has been overlooked. Plankton does not only feed but it also kills by means of its decomposers which pose a health threat to all marine organisms. (16 pages)
  • examples of degradation: a section with photographic examples of what degradation looks like (16 pages, growing)

    For comments and suggestions, please e-mail me, Dr Floor Anthoni.
    Reader please be aware that the information provided in this section is entirely new work that has not been confirmed yet by conventional science. It is therefore not suitable yet for teaching. Readtips for printing.
    -- seafriends home -- DDA index -- plankton index -- decay index --
    Rev 20050201, 20050630,20050810,20050830,20051008,20080703,

    The Dark Decay Assay method may well be the culmination of a personal quest to understand and mitigate the problems in our aquatic environments. It began in 1990 when I decided to devote the remainder of my life towards saving our seas.  But this cannot be done without understanding precisely what is wrong with it and how degradation works. One cannot prescribe an effective cure without thoroughly examining the patient. My dogged pursuit led to several discoveries, all documented on the Seafriends web site. But the discovery of the missing ecofactor, the planktonic decomposers, followed by a simple method to measure them, may well rank amongst the most important discoveries in recent times, changing the way we think about the sea and hopefully giving us the insight to reverse the declining situation.

    For over 40 years now I have been keenly observing the marine environment while taking underwater movies and later, still pictures. These gave me a kind of registration from which I learnt more as I viewed them again. It allowed me to discover the ill effects of degradation at an early stage (1987), which urged me to devote my life to this matter in 1990. Because degradation has far-reaching ill effects on nearly all species, I had to rely on observation alone to further my understanding. This led to so many paradoxes that I concluded that an important ecological factor (like sunlight, wave action, nutrient level, etc.) had been overlooked.
    The plankton ecosystemIt didn't take long to discover that this had to be the planktonic decomposers consisting of bacteria, fungi and viruses. These living organisms are so small that they occupy the pico- and femto-plankton,  invisible to the optical microscope. What people had overlooked is that the food chain is not all the service that plankton delivers, but it also contains potent decomposers that recirculate the nutrients from wastes and dead bodies. Their task is to break dead biomatter apart, thereby consuming the last of its solar energy, as nutrients contain no such energy. Enthusiastically these decomposers will also attack living organisms such that living in the sea depends on a precarious balance - the Plankton Balance that feeds but also kills. Even those organisms not depending on plankton for food, still need to breathe it, and even marine mammals who do not breathe it, are bound to be affected.
    The diagram shows the plankton ecosystem. Phytoplankton (producers, green) convert sunlight into biomatter using nutrients dissolved in the water. The phytoplankton is eaten (grazed) by the zooplankton which is in turn eaten by larger predators, which forms the food chain in its many stages, not shown here. All dead biomatter is recycled by the planktonic decomposers (brown) while larger parts sink to the sea soil where they are decomposed by the benthic (bottom) decomposers. Both kinds of decomposers return dissolved nutrients to the water, thereby closing the cycle. The planktonic decomposers play a large role because of the high turnover rates in the first stages of the food chain.

    The organisms most affected by the decomposers are those taking part in the plankton, particularly the producers or phytoplankton. It was then that I realised that a kind of arms race between the two main guilds, the producers and the decomposers can have a selective influence on both. Such an arms race can be initiated by overfeeding (eutrophication) as is now common in all coastal seas. As the decomposers (the attackers) become more aggressive, the producers (phyto plankton, the defenders) become more defensive as the hardy species replace the sensitive ones. This in turn selects for even more aggressive attackers, and so on. As the phytoplankton becomes more hardy, it also becomes less digestible as at the same time the decomposers become more deadly. Thus the Plankton Balance within the plankton can shift profoundly from powering the food chain (powerplankton) to powering the decomposers while becoming a threat to all other marine organisms (killerplankton), as if the sea were eating itself! This agrees with the environmental degradation symptoms I have been observing.
    I postulated that the health of the sea, particularly where eutrophication reigns, depends almost entirely on the decomposers (while not ignoring sedimentation), and that a method for measuring these was needed.

    Decomposition connects all trophic levels in an ecosystem - Floor Anthoni

    simplified plankton ecosystemMeasuring the decomposers
    By simplifying the planktonic ecosystem to the diagram shown here, it was realised that only a pH meter could measure the biomatter (or energy) flows, as the densities of phytoplankton are very low indeed. The producers (green arrow) photosynthesise (put together by means of energy from light) chains of living matter containing many hydrogen bonds, and therefore scavenge hydrogen ions from the water: the pH increases as hydrogen ions become scarcer. The foodchain (orange arrow) consumes the living matter and oxygenates it: the pH decreases somewhat. Finally the decomposers break the remaining hydrogen bonds, thereby returning hydrogen ions to the water: the pH decreases as hydrogen ions become more numerous.
    Simplistically thought, a high pH is healthy whereas a low pH is not. However, this is not quite borne out by measurements as the system is more complicated.
    Note the orange side loop, thought necessary in case phytoplankton respires at night, is of minor concern. But the brown side loop, which feeds phytoplankton direct to the decomposers, can become rather influential.

    DDA: the breakthroughBirth of the Dark Decay Assay
    Taking the pH of sea water is disappointing as the resulting values do not correspond with the quality of the water such as turbidity, colour and smell. In the map for instance, measurements were done in the Manukau Harbour (New Zealand) where most of Auckland's treated sewage is discharged. As heavy nutrient loads overfertilise the water, the phytoplankton blooms at maximum density, which decreases towards the harbour mouth where the water is visibly much clearer, but this is not borne out by the pH of the water (see inset map).
    It then occurred to me that the producers should be disabled and 'fed' to the decomposers and the process followed in a sealed container to keep all reagents inside. By placing the test vials in the dark, the decomposers would immediately cease their activity and eventually die, to be decomposed. In the process H+ions (hydrogen ions) are released and these can be measured accurately. The graph shows how in curve A the plankton decomposes very rapidly (beginning slope) whereas its total biomass (end of decomposition) is no different from curve B. At the very mouth of the harbour, point D the decomposers are less active as also biomass density (biodensity) is less. A high tide rock pool (E) contains water with similar density as A and B but beginning with a very high pH. It was later discovered that long-lived algae excel in scavenging hydrogen ions, thereby pushing the pH up high.
    It was surprising that the curves showed so much latent or chronic decomposition, even when the phytoplankton should still be alive, later recognised as a sure sign of sick plankton.
    Note that we assumed that the released hydrogen ions are a proxy for (representative of) biomass (measured as hions), something that needs further confirmation. Note also the logarithmic nature of the pH scale where decomposed biomass increases steeply down the graph as a step from 8 to 7 represents a tenfold increase. 

    discovery of powerplanktonPowerplankton
    While using the DDA to get a feel for what it is capable of, also a coastal area was sampled, which usually has clear water, near Cape Brett, New Zealand. Travelling over a patch of water with a different smell, a sample was taken (R) which subsequently showed to consist of a peculiar plankton assemblage. For about one day and a half, no latent or chronic decomposition was measured, followed by decomposition occurring so fast that most of it was missed. As the curves are not caused by artefacts, it must be assumed that this 'suicide' is a special adaptation of the plankton assemblage, perhaps to retain nutrients in the photic (light-) zone. But it is also bound to make this type of plankton better digestible, as no pickings remain for the decomposers (proved by the flat curve at the beginning). This powerplankton thus transfers the solar energy almost completely into the food chain without threatening health. It could well be that this powerplankton powered the once bountiful coastal fisheries. Unfortunately it has now become rare, reason perhaps for collapsing fisheries world wide. It is possible that the disappearance of the powerplankton is the first sign of degradation, happening at around 12-15m visibility. One day perhaps, satellites may recognise this kind of plankton.

    Improving the method
    Once the DDA had shown its promise, we concentrated on making it more accurate, as it can easily suffer from loss of hydrogen ions during the short periods of measurement. We also explored temperature stabilisation because the Rate of Attack (the beginning slope) depends on the biological activity of microorganisms, and thus on temperature. Eventually we settled for an accelerated temperature of 27ºC as the ideal incubation temperature. We also minimised the number of measurements, such that the nature of the decay curve is preserved with minimal loss of hydrogen ions to the air. With these improvements in place, measurements could now be compared from place to place and time to time, as also the whole DDA incubation is completed within two weeks.
    DDA exampleHere is an example of a 250km round-trip in the outer Hauraki Gulf, New Zealand. The distance between points I and M is about 40 nautical miles (75 km). M is located at the territorial boundary which also marks the edge of the continental shelf. The graph shows measured pH vertically and days incubation at the standard temperature of 27ºC. The red rectangle is a movable window of 48 hours to measure the 48 hour Rate of Attack from the drawn curves. Note that decomposed biomass (H+ion proxy or hions) increases downward with decreasing pH. Note how the initial pH always climbs towards the end of the day, due to producer activity opposing that of the decomposers. Note that the total decomposed biomass at day 13 shows only small differences between sites. Highest density (lowest final pH) is D inside a small harbour whereas lowest density (highest final pH) is M. Although M has a noticeably smaller biomass, it also has the lowest Rate of Attack (5) in 48 hours. Conversely, D has the highest RoA (15). These measurements were taken at the end of summer (16 April) when the waters clear up as their nutrients have been consumed and blue water mixes with the coastal water. Even so, it was an unusually murky year with water clarity ranging from 4m (D) to 29m (P). All sites suffered from chronic decay. These results have also been mapped in the coastal map below.

    Northland NZ, RoA + densityMapping the health of the sea
    The map shows the northern part of the North Island (New Zealand), with the city of Auckland straddling its narrowest part. Here the sewage of 1 million inhabitants finds its way to the sea. On the West Coast, the Waikato River flows into the sea, infused with fertiliser and excrement from stock and people from a large area of land to the south of this map.
    Not surprisingly, the (green) values for biomass density are very high here and with these the bacterial activity or chronic Rate of Attack (RoA). Inside the Manukau Harbour where the main sewage works are located, the RoA may reach values of 69, which are not conducive to rich life (even oysters and cockles die), but even at its mouth very high values are found (51), just tolerated by mussels. Notice that RoAs should be interpreted with care because of the possible arms race between decomposers and producers. A low RoA on hardy phytoplankters may mean a high RoA on more sensitive species. As a guideline, consider RoA<20 as healthy, 20-40 as sick and >40 as killer plankton.
    Following the West Coast northward, one comes across a patch of low RoA which shows how a tongue of blue water from the north lays over the more aggressive green-brown water from the south.
    Whereas the west coast is characterised by dirty waters, the east coast is usually clear, with 10-20m viz. However, surprises abound as RoA stays high in many places. Furthest out lies the Poor Knights Islands marine reserve where very high biomass densities and Rates of Attack (45/303) were found. Closer investigation showed a concentration of biomass near the surface, with very high bacterial activity. It explained why we saw much degradation happening in the shallows, whereas such shallows are usually exempted along the coast, due to the intensity of the waves that cleanse it, and more thorough mixing of the water.
    decomposers increase suddenlyIn order to better understand the data in the above map, memorise the diagram shown here, plotting phytoplankton density (green) and decomposer density (orange) on a linear scale (but logarithmic visibility scale). It shows how the phytoplankton reaches a maximum between 4-5m viz. It also shows how the decomposers can increase suddenly between 11-9m viz. The resulting rate of attack (RoA), however, is more erratic but follows an average according to the red curve.
    To focus your thoughts while browsing the map, imagine a hion density of less than 60 as healthy, 60-90 as sick and above 90 as killerplankton, even though this has not been proved conclusively. Note that biodensity and RoA must be considered together.
    Remember also that the map is but a snapshot in time, taken in the autumn period between March and May 2005 when the water becomes clear (and healthier?). Note also that the new technique of alcohol enhancement will find different biodensity values as it also decomposes the slush in the sea.
    Note how the inner Hauraki Gulf misses its RoA values (red) because measurements were done at an unknown ambient temperature. However the green values for biomass density are accurate.
    The data for maps like these can now be collected by conscientious amateurs equipped with a small and low-cost portable laboratory costing no more than $500. Their results can be poured into a national database with Internet access such that a finger can be kept on the pulse of the sea. Local care groups now have a tool to monitor the health of their aquatic ecosystems, a tool that also allows them to monitor the results of their actions and the origins of eutrophication.

    relationship density - visibilityBiodensity and visibility
    Wherever possible, we also measured the clarity of the water by a modified Secchi disc method as this provides another means of verifying the data. In theory the data points for hion biomass should lie along two straight lines marking the reciprocal relationships between visibility and phytoplankton biomass. Vertically the hion density calculated by subtracting the initial pH value from the final pH value (by proper anti-log conversion) and then plotting it again logarithmically. So it resembles the logarithmic pH scale very much.
    The right-hand line marks the nutrient-limited region where visibility increases proportionally as density decreases. The lef-hand line marks the light-limited region where the sunlight can maintain proportionally less biodensity if the visibility decreases (but depth also plays a role). Note in this respect that the DDA does not measure nutrient concentrations, but only their effects on planktonic life. Note also that the graph maps plankton densities rather than the total amount of plankton found in the sea. Ironically, as the water becomes clearer, it enables the sunlight to penetrate deeper too, such that the total plankton biomass over all depth strata remains almost constant between 5m and 30m visibility where it is limited by nutrients.

    Hion density vs visibility
    Unfortunately, the DDA method cannot distinguish between decomposable biomass originating from various sources, such as phytoplankton, sewage, dissolved organic matter (DOM) and zooplankton which tend to draw the data points down and to the left of the green boomerang where they were expected. We recently discovered an invisible thick layer of high density in clear water where this was not expected and named it the plankton graveyard, possibly an indicator of food chain failure when unconsumed biomass heaps up. 
    The visibility measurement on the other hand, cannot distinguish between sediment, small bubbles and phytoplankton, thereby drawing the data points to the left of the green boomerang.
    The combined effects are shown in the diagram, from original data. We are still in the process of mapping results as these arrive, in order to further our understanding. Also the recent technique of alcohol enhancement will have a major influence on the data points to such extent that two separate mapa may result.

    Along the vertical axis, which is a logarithmic scale, also the actual values for hion density have been marked in green. The colour of the datapoints corresponds to the batches obtained from the coloured regions shown on the inset map. The datapoints marking the plankton graveyard lie in the grey shaded region. They have all been confirmed by sampling at greater depths (10, 20, 40m) where hion densities are normal.
    Of main concern is the orange region between the plankton limits and the maximum densities measured. In this region the biomass of the decomposers can increase suddenly between 11m and 9m viz. Unexpectedly decomposers can attain densities exceeding that of the phytoplankton, suggesting that most degradation in the sea happens between 5m and 9m viz, which agrees with our observations! Paradoxically, a constant ingress of mud (in the light-limited region) may limit the action of the decomposers because there is less light to produce the phytoplankton that feeds them!
    It would be interesting to know whether the graph shown above is valid for all the world's seas. A new map is being made with biodensities derived from alcohol-enhanced decomposition.
    Decomposition deficit
    In the data shown above, it was thought that DDA decomposition had expired after settling (9-13 days). However, failed tests for linearity showed that decay continued rapidly after adding an energy food source (fuel) like agar, jelly, sugar or alcohol. It was then realised that the first and second laws of thermodynamics insist that the energy embedded in organic molecules cannot be enough for decomposing them. At some stage a small amount of additional fuel is needed to cover thermodynamic conversion losses. 
    Drastic influence of alcohol on decompositionAfter some experimentation, we saw that pure alcohol was the most suitable substance, as it is also the most basic of fuels, influences pH only little and is easy to keep and to administer. By adding two drops of 20% alcohol at an anaerobic stage (after day 5), decomposition resumes very rapidly, resulting in hion biodensities of two to ten times larger. Some samples (Murrays Bay, French Bay, Seafriends aquariums) even began to stink as pH dropped below 6.4!
    The graph shown here demonstrates alcohol's drastic effect on decomposition, as well as the ability to keep samples in one litre 'aquariums'. These aquariums were sealed and kept in a window facing away from the sun for over one month. The aquariums were then scraped with a clean dishcleaner brush to resuspend all attached algae and bacteria. As one can see, the DDA proceeds in much the same manner, with very similar rates of attack and would have ended as the original set had 2 drops of alcohol not been added after day five. The curve of Leigh Harbour (B, brown) shows how it bottomed out at day five, but under alcohol enhancement continues to reveal a biodensity of 438 instead of 78 hion!

    The discovery of a thermodynamic limitation preventing complete decomposition may have profound ecological consequences on our understanding of ecosystems. It explains why plants provide fuels like sugars and alcohol from fruits to the soil, such that decomposition indeed returns the desired nutrients and minerals. In the sea it may well be the jelly component of the plankton providing the same service, or the slime produced by plants. There may be other providers like perhaps cyanobacteria using sunlight to provide for the missing fuel.

    The oak tree and the aphids
    In our garden stands an oak tree which rains sticky sugars down onto the decking and chairs. It is a nuisance caused by thousands of aphids sucking its sap and wasting sugars. I often wondered why the oak tree had no defences against these little sapsuckers. Now I know that the rain of sugar is needed for the soil decomposers to complete the decomposition of fallen leaves from the previous season for the oak tree to get the nutrients and minerals it craves for. The oak tree does not make sweet fruits, you see?

    The first laws of thermodynamics
    1. Energy can neither be created nor destroyed. Thus all forms of energy ultimately convert to heat.
    2. All physical processes lead to a decrease in the availability of the energy involved. This defines that all energy conversions such as photosynthesis and decay are accompanied by intrinsic losses. Some of these losses are due to the energy being used by organisms for the sake of living. Other losses occur from the thermodynamic conversions of one energy (a hydrogen bond, e.g.) to another. The second law in essence prohibits a perpetuum mobilae, a device running on no energy.


    Freshwater explorations
    Maximum biodensity and natural pHLakes and rivers seem outside the interest of the marine biologist but for us they became very interesting as their planktonic ecosystems are simple and stable and cannot flow away on tidal currents. Because lakes are more tranquil, their clarity is more meaningful as a measure of phytoplankton density. They also have a large range of parameters of which intrinsic acidity or natural pH as we call it, is the most important.
    Natural pH is measured by eliminating all forms of life, either by complete decomposition or by other means. Each lake has its own natural pH and when their biodensities are plotted, a straight linear relationship is found as shown on the graph. Even the sea fits into this relationship (Leigh 250-330; Murrays Bay 400-600 due to ingress of raw sewage).

    The lake explorations gave us the insight to link all water bodies (fresh and salt) in the world by means of a single equation. We discovered that the maximum density and productivity of life depend on the availability of hydrogen ions. Thus hydrogen ions are an important limiting factor of any aquatic ecosystem, according to the maximum biodensity formula discovered:

    maximum biodensity = ALOG( 1.55 - natural pH  )   hion, where the factor 1.55 needs further confirmation.
    The simpler freshwater ecosystems also showed that bacterial activity increases suddenly between 100 and 200 hion biodensity, very similar to the marine situation. Eutrophication begins here. Some eutrophied lakes have huge biodensities (Blue Lake =  2000) and corresponding bacterial biodensities and activity. Blue Lake's clarity (4m viz) defines its producer biodensity at around 250 hion, leaving 1750 hion in decomposer biodensity, or 7 times more. We discovered that the producers are able to feed this large amount of bacteria because they are extremely productive, also following the maximum biodensity formula.

    Read more about all this in method.htm and fresh01.htm.

    plankton with symbiontic decomposersThe slush hypothesis or symbiotic decomposer hypothesis
    It is undeniable that the (un)availability of hydrogen ions is an important (but so far overlooked) limiting factor. An 'acidic' lake like Blue Lake  (natural pH=7.19) does not only have more biodensity than the sea according to the maximum biodensity formula (8-10 times), but also a correspondingly larger productivity to keep up with its high decomposition rate. The sea's pH between 8.0 and 8.3 is so high that it limits plant growth.

    It is also undeniable that full decomposition in the sea does not take place unless an additional bacterial fuel is provided. So the sea is apparently awash in semi-decomposed organic matter (slush, as in semi-molten snow) waiting to be decomposed further. This could have interesting ecological consequences. Plants for instance may well live in a symbiotic relationship with friendly bacteria inside the slime that they produce. As the slime feeds these bacteria, it also enables them to fully decompose the slush, providing not only the craved-for nutrients and minerals, but also for scarce hydrogen ions. The slime on plants should thus be relatively acidic (pH 7 instead of 8), which needs to be tested. One could think of seaweeds living inside a thin cocoon of slime and bacteria that offer a more productive environment. In the scarce environment of salt water, this could well be nature's solution to higher plant productivity. However, it is also a risky way of life, reason perhaps why seaweeds are so sensitive to degradation, a paradox because degradation is accompanied by higher free nutrient levels.

    This idea carries further to the powerplankton, which could well consist of larger phytoplankters (like diatoms) living in a symbiotic relationship with friendly bacteria and benefiting from them in a similar way. It makes these phytoplankters much more productive, while it also provides for the bacteria to rapidly decompose the host once it dies. The reason we are not measuring either productivity or decay is that this happens in a short cycle on the skin of the phytoplankter. As slush is decomposed, its products (hydrogen ions + carbondioxide + nutrients) are immediately converted into plant matter with no measurable change in pH in the water outside. This also suppresses surrounding bacterial activity as the phytoplankters act like a bacterial substrate. As the producers' productivity increases, the planktonic (free-floating) decomposers' activity decreases, which makes the plankton nutritious and safe to live in. The producers' increased productivity is evident from a 0.10 higher initial pH.

    coral polyps by dayThe slush hypothesis also insists that corals which are known as mixotrophs (mixed-feeders) for the plant cells in their tissues, cannot work without symbiotic decomposers (bacteria) on their skins. The DDA method found high concentrations of a mysterious substance in the sea (slush), particularly in the blue desert seas with very little nutrients and chlorophyll. Mixotrophs like corals are designed to harvest this slush which has up to 8 times more biomass than the phytoplankton. This is possible only with symbiotic decomposers on their skins. It may also explain why these creatures are sensitive to changes in their environment like eutrophication and higher temperatures. Thus corals are animal + plants + bacteria. The slush hypothesis predicts that many animals like these remain to be discovered, even in the zooplankton.
    The photograph shows a close-up of the coral polyps of a tropical coral. Their brown colour is caused by brown symbiotic algae in their tissues. The polyps have lost their tentacles for catching zooplankton because their main food now comes direct from sunlight. Protected by their limestone skeletons, these polyps dare to expose themselves by day at risk of being scraped by fish like parrotfish.

    One may wonder where most of the slush comes from. The second law of thermodynamics insists that the bottom decomposers deeper than the photic zone will not be able to fully decompose wastes and dead bodies. Thus the deep sea decomposers must continuously provide slush that eventually finds its way to the surface. How this happens and how old most of the slush is, remains a mystery.

    "Worldwide, mixotrophy is very widespread in the top layer of oceanic water (up to 90% of organisms can be mixotrophic) and in the layer beneath mixotrophs would account for the greatest part of the organisms present there. It is a phenomena that has been very important in evolution and is very important for the continuing of life in the oceans, with their rapidly changing environmental conditions and highly variable food and anorganic nutrients concentrations and light intensities. Symbiosis between a heterotrophic multicellular organism and phototrophic unicellulars can also be seen as a form of mixotrophy. When these forms are also taken into account, mixotrophy may well be the most abundant strategy in oceanic lifeforms."
    Ecophysiology of mixotrophs, Erwin Schoonhoven, January 19, 2000

    The importance of first the discovery of the planktonic decomposers as the missing ecofactor, then the formulation of the Plankton Balance hypothesis and finally a method to measure the health of the sea, cannot be overstated. It required someone to think outside the square, someone who is not part of the scientific community, as it is not being taught at universities, does not appear in any text book or scientific publication and has simply been overlooked completely. The DDA and the thinking leading up to it has given mankind new eyes through which he can see new things, and old things in a new light.
    The Dark Decay Assay is now the first test to consider for anyone working with aquatic ecosystems. It is cheap and easy but unfortunately takes 1-2 weeks of incubation, preferably at a standard temperature. This brings the laboratory into the hands of conscientious amateurs who are willing to do something to save our seas. Einstein observed that the problems of today cannot be solved with the consciousness that created them. I hope that the DDA will bring the new consciousness that enables us to turn the tide in the predicament of the seas, lakes and waterways, as it also empowers many to take part. For me this is a very exciting aspect.

    The problems that exist in the world today cannot be solved by the level of thinking that created them.
    - Albert Einstein

    The advantages of the DDA are:

    The DDA brings a new way of thinking:
    The importance of something new can be measured by how much it destroys of what is old.
    - Floor Anthoni, 2005

    Whenever a new measuring technique is invented, new discoveries are bound to follow. This chapter summarises them.
    Science requires only one investigator ... with results that are verifiable
    by reference to the real world. - Michael Crichton, 2003

    The way science really works is that new methods of acquiring information are created, and data is collected.
    The resulting data is always surprising and never entirely predicted.
    This type of science has been almost totally destroyed by the corruptions which have overtaken science.
    - Gary Novak, Independent Scientist

    International Census of Marine Microbes (ICoMM) (April 2010)
    The International Census of Marine Microbes (ICoMM) found an astonishing number of new microbe species in the sea, estimating their variety at close to one billion species, 20,000 of which have so far been catalogued. "The marine microbes in fact constitute somewhere between 50 to 90 percent of all ocean biomass, and by volume weigh the equivalent of 240 billion African elephants." 
    The average weight of an African Elephant is 4.5 ton, which makes the estimated marine microbial biomass 1080 Gt - many times the combined weight of all other species together.

    The DDA in its generality has many applications where bacterial activity and biomatter are involved, particularly in aquatic ecosystems. Here is a list of useful applications.

    As happens when a new field of scientific investigation is opened, new words are invented to describe new items and processes. Here is an alphabetical list of  new and old terms used, and their meanings.

    aerobic: (Gk: aer= air; bios= life; life growing in air) aerobe= an organism growing in the presence of air or needing air for growth. aerobic= growing in contact with air and thus a supply of oxygen and escape of carbondioxide.
    alcohol enhancement: often the DDA does not complete due to energy conversion losses, such that a small quantity of additional energy is needed to complete the decay. Alcohol was preferred over other energy foods like sugar and agar because it is the most basic of energy foods that only bacteria can make use of, almost immediately. It is also easily obtained, calibrated and kept and it does not affect the measurement. Not all samples need it but on some it makes a 300% difference! One drop of alcohol at 20% is added for every 10ml of sample liquid on day 3-5 of incubation, when decomposition slows down.
    anabolic: (Gk: ana= up; ballo= throw; upbuilding) the synthesis of complex molecules from simple ones, together with the storage of energy. Photosynthesis is an anabolic activity.
    anaerobic: (see aerobic) growing in the absence of or contact with air, resulting in a lack of oxygen. Animals and plants cannot live anaerobically but some bacteria can. Anaerobic decomposition produces different intermediate molecules like hydrogen sulphide which gives a putrid smell.
    assay: (Fr: essay= try, test) the determination of the content or strength of a sample.
    assemblage: (L: ad= to; simul= together; Fr: assembler= get together) a collection of species within an ecosystem. The assemblages of species and their numbers can change rapidly within plankton ecosystems but together they form a functional whole.
    autotroph: (Gk: auto- self; trephos= feed, nourish; self-feeding) a plant; an organism feeding from sunlight.
    biomass: (bio + mass) the total weight of organisms in a given area or volume, often expressed in kg dry matter. Note that the DDA cannot distinguish between living and dead organic matter and we use the words biomass and biodensity to include both.
    biodensity: (bio + density) a new word to describe the biomass per litre as is necessary for planktonic ecosystems. It is expressed as dry matter per litre of liquid, or as the weight of carbon per litre.
    buffer: (a thing that cushions) Chemical: a substance that maintains the hydrogen ion concentration of a solution when an acid or alkali is added or when polluted somewhat.
    calomel: (Gk: calo= beautiful; melas= black) a mercury compound, medically used as cathartic (purgative). Mercurous chloride HgCl2.
    calomel electrode: a conductor made with mercurous chloride, through which electricity enters or leaves a liquid or gas.
    catabolic: (Gk: cata= down; ballo= throw; breaking down) the breaking down of complex molecules into simpler ones with release of energy. Decomposition and decay are catabolic activities.
    chronic: (Gk: chronos= time) persisting for a long time or continuously.
    chronic decay: the decomposition (decay) found even when organisms like phytoplankton are still alive. The decomposition of organisms that died earlier on. Chronic decay indicates a continuous dying of organisms, a sign of ill health.
    DDA: Dark Decay Assay, a new term coined for the measuring technique that places a sample of natural water containing life forms, in a dark sealed container to achieve that plants die from lack of light while decomposers decompose these. The resulting increase in hydrogen ions, measured with a pH meter, is a proxy for the biomass (biodensity) decomposed and the activity of the decomposers. The term dark decay is also used for the loss of electronic charge in charge-coupled devices (CCDs) used for photographic imaging.
    decay: (L: de-= down, completely; cadere=fall; to fall down) to rot, decompose.
    decomposers: (L: de-=down, completely; com-= with, altogether; ponere= to put; to decay or rot) decomposers are an important guild of lifeforms that are able to break down the complex molecules of life, including woody substances. They consist mainly of animals with appendages to physically tear matter, fungi and single-celled bacteria, and viruses. Planktonic decomposers must from necessity be very small, consisting mainly of single-celled fungi, bacteria and viruses.
    degrade: (L: de-= down; gradus= step; step down) to reduce to a simpler molecular structure, converting energy to a less convertible form. To reduce to a lower rank or degree. This word is now extensively used to describe the deterioration in the environment.
    density: the degree of compactness of a substance. The density of seawater means its specific weight which increases due to saltiness. The biodensity of seawater is used to describe its density in biomass. Density is expressed as kg per litre. The density of water is 1.0.
    DOM: dissolved organic matter, a catch-all term for unknown organic substances in the water. See also slush.
    ecofactor: (Gk: oikos= house, environment; environmental factor) ecological factor or environmental factor or limiting factor or life-determining factor. The limiting factors on land are all well known. Of these moisture and temperature are the most decisive since life depends so much on these. But in the sea these two are of less consequence. The planktonic decomposers have completely been overlooked as an important ecofactor. The availability of hydrogen ions has also been overlooked as an important ecofactor.
    ecosystem: a biological community of interacting organisms and their physical environment. An ecosystem is not a complete ecosystem if nutrients are not used by plants and regenerated by decomposers. The cycles of energy and nutrients are closed.
    eutrophication: (Gk: eu-= well; trephos= feed, nourish; overfed, overnourished) an aquatic environment rich in nutrients and therefore upsetting the natural balance. It may result in obscuring light from deeper plants, excessive rot which uses up all oxygen, and aggressive decomposers that attack all life forms.
    expired: (L: ex= out of; spirare= to breathe) to come to an end. This word is used to describe natural water which through full decomposition in darkness has lost all decomposable biomass and is then allowed to equilibrate (ventilate) with the atmosphere. It is natural water that has lost all its solar energy. The resulting pH is called natural pH.
    femtoplankton: (Danish: femten, femto= 15) 0.02-0.2 µm (viruses)
    fuel: (L: focus= hearth; Old French: fouaille) a material used as a source of heat or power. It describes food high in energy like carbohydrates but low in proteins, like sugars and fats. Alcohol can be used as fuel for yeasts which convert it to vinegar in the presence of oxygen, but only bacteria are able to use alcohol as fuel in the absence of oxygen.
    glass electrode: the glass bulb electrode used for measuring the pH of liquids.
    graveyard: (plankton graveyard) a descriptive word for the dense plankton biomatter aggregating near the surface in layers up to 5 metre deep. See also slush.
    healthy plankton: a name used to describe a plankton assemblage high in food value but low in bacterial threat to health. It is used qualitatively but may later be defined quantitively.
    hion: a new unit, introduced here as a proxy for biodensity. One hion is the amount of biomass per litre decomposed in darkness, measured in pH units. It is calculated from the initial and final pH values as: antilog( - final pH) - antilog( - initial pH) in parts per billion (1E-9). The number of hydrogen ions at a pH of 9.00 corresponds to one hion. The hion unit allows for comparison without knowing the actual densities as dry organic matter per litre.
    hydrogen ion: a free hydrogen atom with the charge of one unit. H+
    hydronium ion: the same as a hydrogen ion, but describing it better as still bound to a water molecule: H2O.H+ or H3O+
    hydroxyl ion: the OH- ion, counterpart of the hydrogen ion. Bound together, they form water HOH. Some of the water is dissociated into (relatively) free ions H+.OH-
    incubator: a box or bath that keeps a constant temperature for hatching eggs or nursing premature babies, or for growing micro-organisms. Since the DDA technique relies on the activity of micro-organisms, it is sensitive to temperature. The incubator used with the DDA enables one to compare results wherever and whenever.
    killerplankton: a plankton assemblage very high in chronic decay, showing a steep drop in its DDA curve and high biodensity. This kind of assemblage transfers the solar energy into the bacteria rather than the food chain. Its high bacterial activity threatens all aquatic life forms.
    latent: (L: latent= hidden) concealed, dormant. Existing but not well developed. Ever-present but not developed.
    lysis: (Gk: lysis= loosening) the disintegration of a cell.
    mesoplankton: (Gk: mesos= middle) 0.2-20 mm (copepods)
    microplankton: (Gk: micros= small)  20-200 µm (diatoms, dinoflagellates)
    mixotroph: (L:miscere= to mix; mixtus= mixed; Gk:trephos= to feed, nourish) an organism that feeds in mixed ways, referring to animals with plant cells in their tissues like corals and some anemones. The new slush hypothesis insists that these organisms can exist only if they have symbiotic bacteria as well. Thus corals are animal + plant + bacteria. Mixotroph species are also bound to be discovered in the zooplankton. The unicellular mixotrophic microbes are capable of feeding from both sunlight (phototroph) and something else (phagotroph), and these are found amongst many single-celled aquatic organisms (flagellates [dinoflagellates, prymnesiophytes, chrysophytes, cryptophytes], ciliates, sarcodines,  and radiolarians) as well as in sponges, corals, rotifers, and even in higher plants
    mol, mole, molar: (from molecule) the SI unit of amount of substance equal to the quantity containing as many elementary units as there are atoms in 12 gram of carbon-12. Thus one mole of water contains two hydrogen atoms of weight 1 and one oxygen atom of weight 16, totalling 18 gram.
    nanoplankton: (Gk: nanos= dwarf) planktonic microbial organisms between 2 and 20 µm in size. (diatoms, dinoflagellates, coccolithophorids)
    natural pH: the intrinsic pH of natural water without any influences from biotic (life) factors. It is water that through full decomposition in darkness followed by full exposure to the atmoshpere, has lost all its solar energy and thereby all possible influences that could have altered its pH.
    pH: potential of hydrogen, meaning the electrical potential caused by hydrogen ions. It is about 60mV per pH unit. A logarithm of the reciprocal of the hydrogen ion concentration in moles per litre of a solution, giving a measure of its acidity or alkalinity.
    phagotroph: (Gk: phagos= eat; trephos= nourish) an organism that eats something to nourish itself: grazers, predators, scavengers and detritus-feeders but is mainly used for small planktonic microbes.
    photic: (Gk: phos, photos= light) relating to light. In aquatic systems the photic zone refers to the depth to which light still causes photosynthesis. Light is easily diminished by dense plankton and mud.
    photoheterotroph: same as mixotroph.
    photosynthesis: the process by which the energy of sunlight is used to synthesise carbohydrates and other molecules of life from carbon dioxide and water and other elements.
    photototroph: (Gk: photos= light; trephos= feed, nourish; light-feeding) a plant; an organism feeding from sunlight.
    phyto-: (Gk: phyton= plant) relating to plants. Phytoplankton is the assemblage of plant plankton.
    picoplankton: (Spanish: pico= a little bit) planktonic organisms in the range of 0.2-2 µm, mostly bacteria. (cyanobacteria, other bacteria, Prochlorococcus, and Synechococcus). The open oceans contain mostly picoplankton to 200m depth. Marine pelagic bacteria are small: 0.03-0.4 µm.
    phagotroph: (Gk: phagos=food; trephos= feed, nourish; a something eater) to distinguish plants (autotrophs or phototrophs) from all others (phagotrophs) like grazers and predators. But bacteria living from Dissolved Organic Carbon (DOC or slush) are called saprotrophs. Phagotrophs have no photosynthesising pigments.
    plankter: a single plankton organism. Hence zooplankter and phytoplankter.
    plankton: (Gk: planktos= wandering; plazomai= to wander) organisms spending some part of their life cycle suspended in the water. It includes jellyfish, larvae of non-planktonic adults, microscopic animals living entirely planktonic, single-celled plants of a large range of sizes and single-celled decomposers including bacteria and viruses. Plankton is conveniently divided into groups depending on size:  Megaplankton 20-200 cm, Macroplankton 2-20 cm, Mesoplankton 0.2 -20 mm, Microplankton 20-200 µm, Nanoplankton 2-20 µm, Picoplankton 0.2-2 µm (mostly bacteria), Femtoplankton smaller than 0.2 µm (marine viruses). Note in this respect that the wavelength of blue-green light is 0.5µm.
    plankton balance: a new term coined by us drawing attention to the dual nature of plankton: on the one hand it feeds with the plant matter it synthesises, whereas on the other hand it kills by means of its decomposers. The plankton balance within the plankton can change rapidly from being nutritious and healthy (power plankton, healthy plankton) to being a threat to all life, without providing digestible food (sick plankton, killer plankton).
    powerplankton: a plankton assemblage very low in chronic decay, showing a flat shoulder in its DDA curve, lasting for over 24 hours, followed by very rapid decay. It is postulated that this kind of assemblage provides the most food value for the least threat by committing suicide (lysis). The loss of this kind of plankton could have serious consequences for fisheries.
    producer: a photosynthesiser or plant.
    prograde: the opposite of degrade, a new word denoting improvement in the environment.
    proxy: (from old English procuracy= the taking care of; L: procurare= to take care of, to buy for someone else) a person authorised to act as a substitute. In the case of the DDA method, the measured change in hydrogen ions is a substitute for density of biomass in the sample. The proposed unit for this is the hion.
    RDOC: Recalcitrant or Refractory (stubborn) Dissolved Organic Carbon, the name used in some scientific articles for a fraction of DOC that is resistant to bacterial decomposition. It is identical to our term slush.
    RoA: Rate of Attack. The rate at which decomposers decompose biomass, particularly at the very beginning of the DDA test, when phytoplankton in the sample is not deemed to have died yet. The Rate of Attack measures decomposed biomass in hions after a set time, like 48 hours. For convenience we use RoA48 and RoA24.
    saprotroph: (Gk:sapros= rot; trephos= feed, nourish) a decomposer living from rot but more specifically, an organism living from DOC (Dissolved Organic Carbon) like slush. These are obviously bacteria.
    scavenge: (from Flemish: scauwen, schouwen= to show) to search for and collect discarded items (from a beach or dump). It is used to describe the process of obtaining 'free' but infrequent items, like carbon dioxide from air (one in 3000 molecules) and hydrogen or other ions from water.
    Secchi: Fr. Pietro Angelo Secchi, an Italian astrophysicist, was requested to measure transparency in the Mediterranean Sea by Commander Cialdi, head of the Papal Navy. Secchi was the scientific advisor to the Pope. Secchi used some white discs to measure the clarity of water in the Mediterranean in April of l865. Various sizes of discs have been used since that time, but the most frequently used disc is an 8 inch (20cm) diameter metal disc painted in alternate black and white quadrants.
    The Secchi disc is used to measure how deep one can see into the water. It is lowered into the water by unwinding the waterproof tape to which it is attached and until the observer loses sight of it. The disc is then raised until it reappears. The depth of the water where the disc vanishes and reappears is the Secchi disc reading. The depth level reading on the tape at the surface level is recorded.
    sick plankton: a new name for a type of plankton assemblage with high bacterial activity (and threat to life) but low food value. At present used qualitatively.
    slush: (comes originally from sludge; watery mud or thawing snow, indicating its half-accomplished state) a new word to describe the incompletely decomposed biomass in the sea which waits for an additional bacterial fuel to complete the decomposition process that finally releases nutrients. The term Dissolved Organic Matter (DOM) covers all forms of biomatter, including slush.
    symbiont: (Gk: syn-= with/together; bios= life; symbio= living together) two organisms of different type living together for mutual benefit.
    thermodynamics: (Gk: therme= heat; thermos= hot; dynamis= power) the science of the relationships between heat and other forms of energy. dynamics= any branch of science in which forces or changes are considered.
    vial: (Gk: phiale= a broad, flat vessel) a small glass container, especially for holding liquid medicines. A test tube.
    visibility: the range of vision as possible with given conditions of light and atmosphere. In this chapter, visibility means the distance one can see objects under water. It is measured by looking at a white object to see its shape disappear. See also Secchi above.
    viz: divers' short word for visibility.

    Books and articles relevant to the Dark Decay Assay.
    [let me know]  Presentation of The Plankton Balance Hypothesis to the New Zealand Marine Sciences Conference 2003 in Auckland. (p29 of 171pp, June 2004) Presentation of The Dark Decay Assay (DDA), a new technique for measuring the health of plankton to the NZMSS conference 2005 in Wellington. (p23 of 271pp, Aug 2006)

    Changes and additions
    Check this chapter for the latest changes and additions. This research has been curtailed due to lack of funds.

    yyyymmdd - description
    20080703 - new page added about scientific publications that support or refute our discoveries.
    20060712 - DMS and cloudformation diagram added to dda for dummies.
    20051008 - DDA for dummies added (10p) and corrections made to various chapters.
    20050830 - Changes made to the wordings in various chapters
    20050826 - Presented to the NZ Marine Sciences conference 'human impacts on the marine environment' in Wellington.
    20050803 - Published on the Web
    20050510 - A beginning made writing the DDA chapters.
    20050109 - Discovery of the Dark Decay Assay method
    20030902 - The Plankton Balance hypothesis presented to the NZ Marine Sciences Conference in Auckland, NZ.
    20030617 - Discovery of the missing ecological factor in aquatic ecosystems, the planktonic decomposers.