J.D. van der Toorn (1987)
A biological approach to water purification: I. Theoretical aspects
From: Aquatic Mammals 13(3): 83-92

Table of contents

Water composition
The biology of water treatment
Organisms in a trickling filter
Organic matter and its conversion
Nitrification and denitrification
Foam fractionation


The most common disinfecting agents are chlorine and ozone. Especially for aquaria UV-irradiation is also often employed to disinfect the water (Spotte, 1979b). However its use for marine mammal pools is very limited, due to its very localized and not very strong effect (Spotte and Buck, 1981).

On the other hand, the need for powerful disinfection in a biologically sea water recirculation system may not be very high, since bloom of micro-organisms in such systems is very rare (Wickins and Helm, 1981). Aside from extensive grazing of ciliates on bacteria in a biological filter, there are substances excreted by diatoms that inhibit the growth of Staphylococcus aureus (Aubert and Pesando, 1969) and also certain lipids, that occur in sea water can inhibit the growth of bacteria (Sieburth, 1971).

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The biology of water treatment

(the following is based mainly on Mudrack and Kunst (1986). For more information on this subject the reader is referred to this book.)

The biological treatment of water is based on the natural self-purification capacity of natural waters. When looking at the organisms responsible for this, we can distinguish 2 different groups: surface attached organisms and suspended (free floating) organisms. This is reflected in 2 different possibilities for water treatment. The first type is a so-called fixed bed bio-reactor, usually referred to as a trickling filter. The aim of the construction of such a filter is to get an internal surface area, that is as big as possible, combined with an external volume that is as small as possible. To this end such a filter is filled up with irregular shaped bodies. In waste water treatment very often stones are used as filling material. Small scale trickling filters have also been successfully used for aquarium water treatment (Wolff, 1981), often with poly-ethylene filling bodies, which are, because of their shape, sometimes referred to as "hedgehogs".

Also slow sand filters can act as fixed-bed bio-reactors (Wickins and Helm, 1981). On the top a thick biologically active mud layer will form, that will have to be scraped off every now and then (slow sand filters cannot be backwashed). Due to their size, these filters are not very practical for use in dolphinaria. Rapid and high rate sand filters do not sustain a stable microbial population for 2 reasons. First of all the water flow through the filter bed is usually too fast for the micro-organisms to properly break down dissolved substances. And when the filter is backwashed (nearly) all the micro-organisms are washed out and a new population has to be established (Kinne, 1976). It is therefore better to separate biological and mechanical filtration (Wickins and Helm, 1981). In the other process use is made of the suspended organisms. In order for this to work, it is essential that the organisms make suspended colonies, or flocs. These flocs are kept in suspension by constant aeration of the water. In a second stage these flocs are allowed to settle and the clear water is removed while the settled flocs, usually referred to as sludge, are recycled. This is called the activated sludge process. This process is probably less suitable for dolphinarium or aquarium water purification for 2 reasons. First of all it needs a lot of space and second it works best at relatively high loads (at least, activated sludge reactors are currently used only for the treatment of waters with high organic loads) (Kinne, 1976). It is quite possible that in an aquarium or dolphinarium type situation the amount of sludge formed will be too low for proper operation of the system. In the following, only the trickling filter process will therefore be considered.

In a trickling filter, water is divided over the top and is allowed to flow slowly over the filter bed. In the course of that flow organic matter, both dissolved and suspended, is attacked by micro-organisms. Because some reactions proceed faster than others you can see a stratification in the filter. In the first part of the filter (the top) mainly carbon decomposition will take place. The depth of that carbon decomposition zone depends on the load: if the water contains much organic carbon the zone will be very deep. In heavily loaded filters carbon decomposition may be the only process taking place. In the lower part of the trickling filter nitrification can occur. It will take rather long before a population of nitrifying bacteria has been established, since they grow slowly. It will take several weeks before a filter is properly conditioned (Spotte (1979b), Kinne (1976)).

If a trickling filter is just put into operation the first organisms that will grow in it will be bacteria and fungi. These will coat the filling material: a bio-film is formed. On this film the secondary colonizers will start growing. To this group belong among others protozoa and nematodes. The growth of organisms in and on the bio-film will increase its thickness. If a film becomes too thick it will start sloughing. This sloughed film will appear in the water as flocs. These flocs settle easily, so if water from the trickling filter is led through a settling area first, nearly all organic matter can be removed.

In short what is happening in a trickling filter is the following:

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Organisms in a trickling filter

The organisms that can be found in a trickling filter do not differ much from the ones found in activated sludge. No differences have been found between the bacterial flora of activated sludge and that of trickling filters (Lin, 1984). In activated sludge most are attached to suspended flocs, whereas in a trickling filter they will be attached to the filter bed. Apart from heterotrophic and nitrifying bacteria, they following organisms take part in the purification process (see Mudrack and Kunst (1986), Tri (1975), Fair et al (1968)):

Figure 1: Some organisms, commonly encountered in trickling filters and activated sludge (Original magnification 400x)

a) A zooflagellate, Bodo sp.

b) A nematode

c) A stalked ciliate: Vorticella sp.

d) A free swimming ciliate: Euplotes sp.

e) A heliozoan amoeba, possibly Actinophrys sp.

Other organisms that are occasionally encountered include crustaceans, mites and insect larvae. The so-called sewage fungus appears only in highly loaded or over loaded systems in significant amounts. This is in fact not a fungus but a filamentous bacteria species from the genus Sphaerotilus.

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Organic matter and its conversion

Most of the organic matter that is put into an aquatic system as food, will at one time or another end up in the water as metabolic end products through urine and faeces. Most of it will still be in organic form and can be converted by micro -organisms. These micro-organisms will break down the organic matter to inorganic forms. This process is called mineralization.

Large molecules will first be broken down to smaller ones by bacterial exo-enzymes (enzymes that bacteria excrete) (Tri, 1975). The most important organisms involved in these conversions are heterotrophic bacteria. It is extremely difficult to identify bacterial genera, let alone species, from water purification systems, but it is obvious that especially the genera Flavobacterium and Pseudomonas are important (Lin, 1984). These bacteria are also very common in seawater (Rheinheimer, 1980). Vibrio species are common in coastal environments and are also commonly present in marine mammal pools. Marine mammals are apparently carriers of these bacteria, usually without any adverse effects (Buck and Spotte, 1986). Most of the heterotrophic bacteria are proteolytic, which means that they can break down proteins. In this process, ammonia is released into the water. Pseudomonas species play a key role in proteolysis. Most important in the breakdown of fats are Pseudomonas and Vibrio, which are also responsible, together with Flavobacterium, for most of the conversion of carbohydrates (Rheinheimer, 1980). The resulting smaller molecules can be absorbed and will be further mineralized, thereby providing the organism with energy, or will be converted into macromolecules the organism needs. In the mineralization process 3 nutrient elements are of interest: carbon, nitrogen and phosphorus.

The carbon in organic matter is mostly converted into carbon dioxide and will leave the water as carbon dioxide gas. It can in water form carbonic acid as thus play a role in the buffering system (see under that heading for more details).

The phosphorus is released into the water as (reactive) phosphate. This is not converted any further. Some of it can be taken up by certain algae, but in aquarium systems, most of it will either precipitate with calcium or magnesium, or will be bound to organic matter that can be removed for instance by foam fractionation. Phosphates that have been complexed with calcium can be re-released into the water by bacteria from the genera Pseudomonas, Aeromonas and Escherichia (Rheinheimer, 1980).

The fate of nitrogen is somewhat more complex. Some algae can convert urea, the main nitrogen source in mammalian urine, directly into useful products and this may also be true for certain amino acids. But in most cases nitrogen in organic matter is mineralised to ammonia (the amount of nitrogen present as ammonia is usually referred to as NH4-N). Ammonia is toxic to most organisms in high concentrations. It can however be converted to nitrate in a process called nitrification. Nitrate is not known to have toxic properties even at high concentrations.

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Nitrification and denitrification

Nitrification, the conversion of ammonia to nitrate, is a process that takes place under aerobic conditions. This is done by bacteria. The first step in the process is the conversion of ammonia, NH4+ to nitrite, NO2-. According to Kinne (1976), there are 5 species of bacteria that can do this conversion in sea water: Nitrosomonas, Nitrosocystis (2 species), Nitrosospira and Nitrosolobus. Spotte (1979b) mentions Nitrosomonas as being the most important ammonia oxidizer. Helder and de Vries (1983) tested the growth on Nitrosomonas at different salinities (from 0 to 3.5%) and found that this species grew equally well at all salinities. They noted that it needed some time to adapt to a different salinity, but once adapted, grew well.

The next step in the nitrification process is the conversion of nitrite, NO2-, to nitrate, NO3-. There are 4 species of nitrite oxidizers: Nitrobacter (2 species), Nitrococcus and Nitrospina (Kinne, 1976). Helder and de Vries (1983) found that Nitrobacter sp. adapted fast to different salinities and grew equally well over the whole range from 0 to 3.5%. Under aerobic conditions, nitrate is the end stage of nitrogen metabolism.

For the removal of nitrate there are 2 possible pathways. There are several large algae, among others Ulva sp. (sea lettuce), that can use nitrate nitrogen for the formation of organic nitrogen compounds, provided there is plenty of light available (Wickins and Helm, 1981). If there is not enough light, they will start metabolizing organic nitrogen and in that way increase the nitrate concentration.

Under anaerobic conditions, several bacteria species can use nitrate in respiration instead of oxygen. Among these, bacteria from the Pseudomonas group are predominant (Rheinheimer, 1980).In the process elementary nitrogen, N2, is formed, which will escape as a gas. Since the energy gain from nitrate-induced respiration is about 10% less than that of oxygen-induced respiration, the latter will have preference if oxygen is present (Mudrack and Kunst, 1986).

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