Aquaculture 270 (2007) 1 – 14

www.elsevier.com/locate/aqua-online

Review article
Nitrogen removal techniques in aquaculture
for a sustainable production
Roselien Crab a,b , Yoram Avnimelech c , Tom Defoirdt a,b ,
Peter Bossier b , Willy Verstraete a,⁎
a
Laboratory of Microbial Ecology and Technology (LabMET), Ghent University, Coupure Links 653, 9000 Gent, Belgium
b
Laboratory of Aquaculture and Artemia Reference Center, Ghent University, Rozier 44, 9000 Gent, Belgium
c
Faculty of Agricultural Engineering, Technion, Israel Institute of Technology, Haifa 32000, Israel
Received 16 February 2007; received in revised form 2 May 2007; accepted 5 May 2007

Abstract

As the aquaculture industry intensively develops, its environmental impact increases. Discharges from aquaculture deteriorate
the receiving environment and the need for fishmeal and fish oil for fish feed production increases. Rotating biological contactors,
trickling filters, bead filters and fluidized sand biofilters are conventionally used in intensive aquaculture systems to remove
nitrogen from culture water. Besides these conventional water treatment systems, there are other possible modi operandi to recycle
aquaculture water and simultaneously produce fish feed. These double-purpose techniques are the periphyton treatment technique,
which is applicable to extensive systems, and the proteinaceous bio-flocs technology, which can be used in extensive as well as in
intensive systems. In addition to maintenance of good water quality, both techniques provide an inexpensive feed source and a
higher efficiency of nutrient conversion of feed. The bio-flocs technology has the advantage over the other techniques that it is
relatively inexpensive; this makes it an economically viable approach for sustainable aquaculture.
© 2007 Elsevier B.V. All rights reserved.

Keywords: Water quality; Sustainability; Biofilters; Periphyton; Bio-flocs technology; C/N ratio

Contents

1. Overview of problem. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2
2. N removal outside the culture unit . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3
2.1. Earthen treatment ponds or reservoirs. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3
2.2. Biofiltration . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4
3. N removal within the culture unit . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7
3.1. The periphyton treatment technique. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7
3.2. Bio-flocs technology . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8

⁎ Corresponding author. Tel.: +32 9 264 59 76; fax: +32 9 264 62 48.
E-mail address: [email protected] (W. Verstraete).
URL: http://labmet.ugent.be (W. Verstraete).

0044-8486/$ - see front matter © 2007 Elsevier B.V. All rights reserved.
doi:10.1016/j.aquaculture.2007.05.006
2 R. Crab et al. / Aquaculture 270 (2007) 1–14

4. Conclusions and future perspectives . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11

Acknowledgements . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11

1. Overview of problem waste than live human biomass. The reason is that the
scope of digestion in fish is limited; a relatively large
Aquaculture is a rapidly growing food producing fraction of feed remains undigested and is excreted
sector. The sector has grown at an average rate of 8.9% (Amirkolaie, 2005). The feeding habit of fish is reflected
per year since 1970, compared to only 1.2% for capture in the digestive anatomy. The gut length of fish is short
fisheries and 2.8% for terrestrial farmed meat-production and the ratio of gut length to body length is small
systems over the same period (FAO, 2004). In contrast to (Hertrampf and Piedad-Pascual, 2000). For instance, the
aquaculture, capture fisheries landings as a whole is intestine of carp is 2.0–2.5 times longer than the body,
stagnant. Although catch rates for some species did not while that of cattle and sheep is respectively 20 and 30
decline during the 1990s, most ocean fisheries stocks are times longer. The human intestine is about 3 to 4 times
now recognized as fully or over fished. The worldwide longer than the body. Consequently, in fish, the chyme
decline of ocean fisheries stocks and the further expansion stays in the gut only for a short time. For this reason, fish
of the human population are an incentive for the further feed must have a high digestibility. Typically, fish body
growth of aquaculture. Despite the growth of the sector, contains 65 to 75% protein (Hertrampf and Piedad-
aquaculture production still needs to increase 5-fold in the Pascual, 2000). In addition, fish use proteins for energy
next 2 decades in order to satisfy the minimum protein production to a large extent, unlike terrestrial animals that
requirement for human nutrition (FAO, 2004). use mostly carbohydrates and lipids (Hepher, 1988). Fish
The intensive development of the aquaculture protein requirement, therefore, is about two to three times
industry has been accompanied by an increase in higher than that of mammals. Ammonium is one of the
environmental impacts. The production process gener- end products of protein metabolism (Walsh and Wright,
ates substantial amounts of polluted effluent, containing 1995). All these factors contribute to the high nitrogen
uneaten feed and feces (Read and Fernandes, 2003). residues in aquaculture water (Fig. 1). In water, NH3
Discharges from aquaculture into the aquatic environ- (ammonia) and NH4+ (ammonium) are in equilibrium
ment contain nutrients, various organic and inorganic depending on the pH and the temperature (Timmons et al.,
compounds such as ammonium, phosphorus, dissolved 2002). The sum of the two forms is called total
organic carbon and organic matter (Piedrahita, 2003; ammonium nitrogen (TAN). Although both NH3 and
Sugiura et al., 2006). The high levels of nutrients cause NH4+ may be toxic to fish, unionized ammonia is the more
environmental deterioration of the receiving water toxic form attributable to the fact that it is uncharged and
bodies. In addition, the drained water may increase the lipid soluble and consequently traverses biological
occurrence of pathogenic microorganisms and introduce membranes more readily than the charged and hydrated
invading pathogen species (Thompson et al., 2002). NH4+ ions (Körner et al., 2001). Ammonia-N is toxic to
To produce 1 kg live weight fish one needs 1–3 kg dry commercially cultured fish at concentrations above
weight feed (assuming a food conversion ratio about 1–3) 1.5 mg N/l. In most cases, the acceptable level of union-
(Naylor et al., 2000). About 36% of the feed is excreted as ized ammonia in aquaculture systems is only 0.025 mg
a form of organic waste (Brune et al., 2003). Around 75% N/l (Neori et al., 2004; Chen et al., 2006). However, the
of the feed N and P are unutilized and remain as waste in toxicity threshold depends strongly on the species, size,
the water (Piedrahita, 2003; Gutierrez-Wing and Malone, fine solids, refractory organics, surface-active com-
2006). An intensive aquaculture system, which contains pounds, metals, and nitrate (Colt, 2006).
3 ton tilapia, can be compared on a biomass basis to a In addition to the generation of large amounts of waste,
human community with 50 inhabitants (Helfman et al., the use of fishmeal and fish oil as prime constituents of
1997). This intensive aquaculture system can also be feed is another non-sustainable practice in aquaculture.
compared on grounds of waste generation to a community Approximately one-third of the global fishmeal produc-
of around 240 inhabitants (Aziz and Tebbutt, 1980; tion is converted to aquaculture feeds (Delgado et al.,
Flemish government, 2005). It can thus be concluded that 2003). The proportion of fishmeal supplies used for fish
live fish biomass generates approximately 5 times more production increased from 10% in 1988 to 17% in 1994
 R. Crab et al. / Aquaculture 270 (2007) 1–14 3

Fig. 1. Nitrogen cycle in aquaculture ponds with a long hydraulic residence time. The N-input considered is formulated feed. A part of the feed
remains unconsumed in the system (Franco-Nava et al., 2004b). The consumed feed is partially converted into fish biomass and partially excreted as
ammonium or egested as feces (Jiménez-Montealegre et al., 2002). The uneaten feed and feces contribute to the organic mater load of the system. The
microbial decomposition of organic matter in the system leads to increased levels of TAN and nitrite, both harmful to fish even at low concentrations
(Meade, 1985; Jiménez-Montealegre et al., 2002; Torres-Beristain et al., 2006). The TAN present in the system may be transformed into nitrite, nitrate
and gaseous nitrogen. The formation of nitrogen gas is considered negligible in aquaculture ponds (El Samra and Olàh, 1979). The bacteria present in
the water and sediment carry out these nitrogen transformations by nitrification and denitrification. Both TAN and nitrate can be assimilated by the
phytoplankton, present in the water column. The phytoplankton can be consumed by the cultured organism (Turker et al., 2003). The consumption of
phytoplankton by fish is minimal in this network. In stagnant water ponds TAN tends to accumulate within the system due to insufficient nitrification
activity (Grommen et al., 2002).

and 33% in 1997 (Naylor et al., 2000). Hence, aquaculture nitrification tanks as also used in domestic wastewater
is a possible panacea, but also a promoter of the collapse treatment plants. It should be noted that the real nitrogen
of fisheries stocks worldwide. The ratio of wild fish:fed removal processes are those that involve the release of
farmed fish (both live weight base) is about 1.41:1 for fixed nitrogen back to the atmosphere (van Rijn et al.,
tilapia and 5.16:1 for marine finfish, (Naylor et al., 2000). 2006). However, these are not discussed here.
Purchase of commercially prepared feed for fish culture
comprises 50% or more in the production costs; this is 2.1. Earthen treatment ponds or reservoirs
primarily due to the cost of the protein component
(Bender et al., 2004). On average some 25% of the This treatment procedure consists of the direct linkage
nutrient input of these feed sources is converted into of, and water recirculation between the intensive produc-
harvestable products (Avnimelech and Lacher, 1979; tion ponds and treatment ponds. The effluent water of the
Boyd, 1985; Muthuwani and Lin, 1996; Avnimelech and production pond is retained in a basin for several hours to
Ritvo, 2003). To make further sustainable increase of days to allow natural physical, chemical, and biological
aquaculture production possible, the search for inexpen- processes to improve its quality for reuse (Diab et al., 1992;
sive protein sources and a higher efficiency of nutrient Hargreaves, 2006). Important practical parameters in this
conversion of feed is needed. system are the hydraulic retention times of the intensive
fish culture unit and the treatment pond, homogeneous
2. N removal outside the culture unit mixing of the treatment pond, and the periodic aeration of
the pond sediment by drainage. The use of treatment ponds
The most common water purification treatments in encounters problems due to algal collapse and anaerobiosis
aquaculture systems can be subdivided in different types of the sediment (van Rijn, 1996). The main disadvantage is
of water treatments: 1) earthen treatment ponds or the unstable purification resulting from unpredictable
reservoirs, and 2) a combination of solids removal and fluctuations of phytoplankton biomass and speciation in
4 R. Crab et al. / Aquaculture 270 (2007) 1–14

the treatment pond (Hargreaves, 2006). An important operations apply physical forces to remove contaminants.
advantage is that the microalgae grown in the treatment Solid removal is accomplished by sedimentation (settle-
pond can be used to produce a second crop, such as bivalve able solids) or mechanical filtration (suspended and fine
seed or Artemia, which can be sold to generate income solids) (van Rijn, 1996). Two commonly used types of
(Wang, 2003). mechanical filtration in aquaculture include screen fil-
A possible system configuration comprises of a fish tration and expendable granular media filtration (Twar-
farm with nutrient assimilation by molluscs and seaweed owska et al., 1997; Franco-Nava et al., 2004a). For fine
(Fig. 2). Here, nutrients released in the culture system can solids removal, foam fractionation – a process also
be converted into plant or other biomass, which can easily referred to as air stripping or protein skimming – is often
be removed and may often be a valuable by-product. The employed (Timmons, 1984; Hussenot, 2003). Chemical
nutrient-assimilating photoautotrophic plants can be used unit processes used for aquaculture wastewater treatment
to turn nutrient-rich effluents into profitable resources are customarily used in conjunction with physical unit
(Neori et al., 2004). Biofiltration by plants generates in the operations and biological processes. The inherent disad-
culture system a mini-ecosystem, in which, if properly vantage of most chemical unit processes is that they are
balanced, plant autotrophy counters fish (or shrimp) and additive processes; the chemicals tend to stay for a major
microbial heterotrophy, not only regarding nutrients but part in the water. This is a significant factor if the waste-
also with respect to oxygen, pH and CO2 (Neori et al., water is to be reused. The main chemical unit process used
2004). As a result, plant biofiltration diminishes the net in aquaculture is disinfection by means of ozonation
environmental impact of aquaculture production systems. (Summerfelt, 2003). Disinfection by UV irradiation is
Today's integrated intensive aquaculture approaches, considered as a credible alternative to chemical disinfec-
developed from traditional extensive polyculture, inte- tion, because of the absence of toxic by-products which
grate the culture of fish or shrimp with vegetables, are usually generated and identified during chemical
microalgae, shellfish and/or seaweed (Neori et al., 2004). disinfection (Hassen et al., 2000). These techniques avoid
By dividing the production process into stages, we can the addition of chemical substances that are hazardous to
increase the constancy of the biomass in the system and the cultured organism. Biological processes are the most
improve the utilization efficiency of the physical facility important ones with respect to aquaculture wastewater
(Wang, 2003). treatment and the major biological process is nitrification.
Nitrification is carried out in a variety of systems, which
2.2. Biofiltration can be grouped into 2 general types: emerged (rotating
biological contactors, trickling filters) and submerged
The treatment methods that are applied to treat (e.g. fluidized bed filters, bead filters) fixed film filters
aquaculture wastewater are broadly classifiable into phy- (van Rijn, 1996; Ling and Chen, 2005; Malone and
sical, chemical and biological processes. Physical unit Pfeiffer, 2006). Biological filters are used for freshwater

Fig. 2. Integrated farming: nitrogen and phosphorus budget (after Kautsky, 2004).
 R. Crab et al. / Aquaculture 270 (2007) 1–14 5

and marine operations (Hovanec and DeLong, 1996; tions. Rotating biological contactor technology is based
Gutierrez-Wing and Malone, 2006; Malone and Pfeiffer, on the rotation of a submerged substrate, which is made of
2006). This paper reviews recirculating systems on high-density polystyrene or polyvinyl chloride, attached
biofiltration technologies for freshwater systems. to a shaft (Tawfik et al., 2004; Park et al., 2005; Brazil,
Nitrification in the bacterial film of the biofilter is 2006). Nitrifying bacteria grow on the media and because
affected by a variety of parameters such as substrate and of the rotation they alternately contacting nitrogen rich
dissolved oxygen concentrations, organic matter, temper- water and air. As the rotating biological contactor rotates,
ature, pH, alkalinity, salinity and turbulence level (Satoh it exchanges carbon dioxide, generated by the bacteria,
et al., 2000; Chen et al., 2006). Nitrifying bacteria are with oxygen from the air. In general, rotating biological
sensitive organisms and are extremely susceptible to a contactor systems are divided into a series of independent
wide variety of inhibitors such as high concentrations of stages or compartments (Lavens and Sorgeloos, 1984;
ammonia and nitrous acid, low dissolved oxygen levels Brazil, 2006). Compartmentalization creates a plug-flow
(b 1 mg/l) and pH outside the optimal range (7.5–8.6) pattern, increasing overall removal efficiency. It also pro-
(Masser et al., 1999; Villaverde et al., 2000; Ling and motes a variety of conditions where different organisms
Chen, 2005). Nitrification, and especially the second step can flourish to varying degrees. As the water flows
(NO2 → NO3), is very sensitive to even traces of sulphides through the compartments, each subsequent stage
(Joye and Hollibaugh, 1995). Sulphides are present in receives influent with a lower organic content than the
sediments and in sludges accumulated in intensive previous stage; the system thus enhances organic removal
aquaculture systems. For higher C/N ratios, the hetero- (UN, 2003; Watten and Sibrell, 2006). Complimentary,
trophic bacteria out-compete nitrifiers for available the rotating biological contactor has low head require-
oxygen and space in the biofilters (Michaud et al., ments to move water through the vessel. This advantage
2006). Hence, nitrification necessitates a low C/N ratio. implies passive aeration and carbon dioxide removal, and
Fig. 3 illustrates the N cycle in aquaculture systems low chance of clogging (Brazil, 2006).
equipped with an external biofilter. Miller and Libey (1985) demonstrated that a rotating
Rotating biological contactors have been used in the biological contactor provided better TAN areal removal
treatment of domestic wastewater for decades and are now rates, in the range of 0.19–0.79 g TAN/m2 day, than a
widely used as nitrifying filters in aquaculture applica- packed tower or fluidized bed reactor (0.24 g TAN/m2

Fig. 3. Nitrogen cycle in aquaculture systems equipped with an external biofilter. The nitrogen cycle is similar to that of in water ponds with a long
hydraulic residence time, but now the water rich in TAN is sent to an external biofilter. In this biofilter, nitrification is enhanced and through nitrite,
nitrate is formed out of TAN. Nitrate is less toxic to fish than is TAN or nitrite (Meade, 1985; Lyssenko and Wheaton, 2006). Although such a system
avoids TAN to accumulate, nitrate build up may take place. The biofilter creates space and optimal conditions for nitrifying bacteria to grow.
6 R. Crab et al. / Aquaculture 270 (2007) 1–14

day), when treating the same fish culture water at Downflow microbead filters are combinations of
comparable hydraulic loadings. Brazil (2006) described trickling filters and granular type biological filters
the performance and operation of a rotating biological (Timmons et al., 2006a). The use of floating media in
contactor in a tilapia recirculating aquaculture system. downflow configurations has the advantage of being
The system obtained an average TAN areal removal rate capable of using smaller media and the associated higher
of about 0.42 g/m2 day. Increasing influent dissolved specific surface areas. As the recirculating water passes
organic carbon levels decreased ammonia removal ef- through the packed bed, suspended solids are captured
ficiency. However, there was no detectable relationship and biofiltration processes are active (Malone and
between the feed loading rate and ammonia oxidation Beecher, 2000). The configuration offers the added ad-
performance. In addition to organic loading, mass and vantage of using high hydraulic loadings without the need
hydraulic loading, rotational speed and staging affected for sophisticated mechanical structures in the reactor to
the ammonia oxidation performance. retain the media within the reactor vessel (Greiner and
Trickling filters consist of a fixed medium bed Timmons, 1998). The medium consists of polystyrene
through which aquaculture wastewater flows down- beads that are 1–3 mm in diameter and have a porosity of
wards over a thin aerobic biofilm (Eding et al., 2006). 36–40% (Timmons et al., 2006a). Depending on these
As it trickles down, the water is continuously oxygen- features the specific surface area ranges from 1150 to
ated, while the carbon dioxide is degassed and removed 3936 m2/m3 (Greiner and Timmons, 1998; Malone and
by the ventilated air. Trickling medium has a specific Beecher, 2000; Timmons et al., 2006a).
surface area ranging from 100 to 1000 m2/m3. Finturf Greiner and Timmons (1998) observed TAN areal
artificial grass (284 m2/m3), Kaldnes rings (500 m2/m3), removal rates of about 0.45–0.60 g/m2 day. A study
Norton rings (220 m2/m3) and Leca or light weight clay using a commercial microbead filter system reported
aggregate (500–1000 m2/m3) are some of the most fre- an average TAN areal removal rate of 0.30 g/m2 day
quently used media (Greiner and Timmons, 1998; (Timmons et al., 2006a).
Lekang and Kleppe, 2000; Timmons et al., 2006a). Fluidized sand biofilters have been widely adopted in
The organic material present in the wastewater is adsorbed recirculating systems that must reliably maintain ex-
on the biological slime layer and degraded by aerobic cellent water quality (Summerfelt, 2006). Filter sand has
microorganisms. a high specific surface area, i.e. 4000–20000 m2/m3 and
Kamstra et al. (1998) reported TAN areal removal has a moderate cost (Summerfelt, 2006). A disadvantage
rates between 0.24 and 0.55 g TAN/m2 day for a com- of the FBS is that they do not aerate, as do trickling
mercial-scale trickling filter. For three different applied filters (Summerfelt, 2006). Therefore, additional aera-
filter medium types in commercial farms and a range of tion is needed. These filters also must operate within a
hydraulic surface loading conditions, the highest ob- narrow water flow range in order to maintain proper bed
served TAN areal removal rate for a trickling filter was expansion (Summerfelt, 2006).
1.1 g TAN/m2 day, with an average TAN areal removal Miller and Libey (1985) demonstrated that the TAN
rate of about 0.16 g TAN/m2 day (Schnel et al., 2002; removal rate of a fluidized bed reactor was around
Eding et al., 2006). Lyssenko and Wheaton (2006) re- 0.24 g N/m2 day. Timmons and Summerfelt (1998)
ported TAN areal removal rates of 0.64 g TAN/m2 day. found similar rates in their research.
In the same study they found similar TAN areal removal Table 1 gives an overview of the average TAN areal
rates for a submerged expandable upflow sand filter. removal rate and the cost per kg of fish produced per

Table 1
General overview of the average TAN areal removal rate for frequently used biofilters in aquaculture systems
Biofilter type Average TAN areal removal rate Cost a References
2
(g TAN/m day) (Euro/kg yr)
Rotating biological contactor 0.19–0.79 1.143 Miller and Libey, 1985; Brazil, 2006
Trickling filter 0.24–0.64 1.036 Kamstra et al., 1998; Schnel et al., 2002; Eding et al.,
2006; Lyssenko and Wheaton, 2006
Bead filter 0.30–0.60 0.503 Greiner and Timmons, 1998; Timmons et al., 2006a
Fluidized sand biofilter 0.24 0.198 Miller and Libey, 1985; Timmons and Summerfelt, 1998
Also the costs for various biofilter choices based upon their capitalization cost to support a 454 ton per year tilapia farm are summarized.
a
Data from Timmons et al. (2006b).
 R. Crab et al. / Aquaculture 270 (2007) 1–14 7

year for each biofilter type. Rotating biological con- greaves, 2006). An important advantage is that microbial
tactors have the highest TAN areal removal rate, bio-flocs and periphyton can be consumed and used as a
followed by bead biofilters and trickling filters, and source of feed by the cultivated organisms (Burford et al.,
fluidized sand biofilters. Although rotating biological 2003, 2004; Hari et al., 2004; Azim and Wahab, 2005;
contactors show good performance concerning TAN Keshavanath and Gangadhar, 2005). As explained in the
removal rate, they are together with trickling filters more following paragraphs, both approaches are possible
expensive then the other biofilter types discussed. solutions for water quality problems, and can decrease
Fluidized sand biofilters and bead biofilters are the the use of fish oil and fishmeal utilization in aquaculture.
least expensive options for water treatment when the
cost per kg of fish produced per year is considered. 3.1. The periphyton treatment technique

3. N removal within the culture unit The periphyton community consists of attached
aquatic biota on submerged matrices. It harbours algae,
The three nitrogen conversion pathways naturally bacteria, fungi, protozoa, zooplankton and other inverte-
present for the removal of ammonia–nitrogen in brates (Azim et al., 2005). As with phytoplankton,
aquaculture systems are photoautotrophic removal by periphyton can be found in almost every type of water
algae, autotrophic bacterial conversion of ammonia– body from small ponds to large oceans and in trophic
nitrogen to nitrate–nitrogen, and heterotrophic bacterial conditions that range from the most oligotrophic to the
conversion of ammonia–nitrogen directly to microbial most eutrophic (Azim and Asaeda, 2005). Given adequate
biomass (Ebeling et al., 2006). light, up to about 0.5 m depth in the water, high rates of
Developing and controlling dense heterotrophic mi- photosynthesis and autotrophic production can be
crobial flocs in the water column or attached micro- achieved (Craggs et al., 1996; Vermaat, 2005). Values
organisms called periphyton can accelerate the biological for periphyton productivity are typically in the range of
removal of organic and inorganic wastes in ponds 1–3 g C/m2 substrate day or 2–6 g dry matter/m2 day
(Avnimelech, 2005; Azim et al., 2003a,c). These (Azim et al., 2005). Periphyton entraps organic detritus,
processes are integral parts of the culture unit (Har- removes nutrients from the water column and helps

Fig. 4. Nitrogen cycle in extensive aquaculture ponds with substrates for periphyton growth. The nitrogen cycle is similar to that of in water ponds
with a long hydraulic residence time, but now TAN concentration does not build up in the water column, neither is the nitrate concentration. The
periphyton community takes up both TAN and nitrate and edible biomass is formed. The cultured fish can graze on the periphyton community and
hence nitrogen, originating from wasted feed and excretion by fish, is redirected towards the cultured organism and therefore, this technique enhances
the overall efficiency of nutrient conversion of feed.
8 R. Crab et al. / Aquaculture 270 (2007) 1–14

control the dissolved oxygen concentration and the pH of is around 4 g dry matter/m2 day and the protein content of
the surrounding water (Azim et al., 2002; Dodds, 2003; periphyton is around 25% of the dry matter (Azim et al.,
Bender et al., 2004). 2002, 2005). This corresponds to a particular feed
Supplying substrates improves the nitrogen-related quantity that can diminish the overall feed cost.
processes developing in the water column and the nitro- Besides the large area needed, the problem within this
gen flow is mainly linked to autotrophic and heterotrophic system is that the process is completely dependent on the
activity that takes place in the periphyton (Fig. 4) availability of sunlight (Azim and Asaeda, 2005). On
(Milstein, 2005). The beneficial influence of periphyton cloudy days or on days with insufficient sunlight, the
on the water quality in different aquaculture systems has maximum nitrogen uptake rate will not be reached.
been investigated, as well as the impact of grazing by fish Another problem is the laborious task to harvest the
on periphyton communities (Huchette et al., 2000; Azim periphyton. One can conclude that application of the
et al., 2001, 2002, 2003a,b,c, 2004). Not all fish are able to periphyton treatment technique in the intensive aquacul-
graze on periphyton; morphological and physiological ture sector is not feasible. Nevertheless, the technique of
adaptations to periphyton grazing are required (Azim using this natural feed may be significant, particularly in
et al., 2005). Although direct experimental evidence is smaller, extensive-level aquaculture systems in develop-
scarce, the aquaculture fish species that can effectively ing countries. The addition of the ‘periphyton loop’ in
utilize the periphyton assemblage are probably more aquaculture ponds can be accomplished by adding static
numerous than those that are exclusively phytoplankti- substrates to the pond (Azim et al., 2005), such as poles
vorous (van Dam and Verdegem, 2005). Besides spe- horizontally planted in the ponds. Substrates used are
cialist (macro)herbivores, more general detritus and bamboo, hizol and kanchi (Azim et al., 2002, 2003c).
benthos feeders can also thrive on periphyton (van Dam Since periphyton can be easily cultured in modified fish-
et al., 2002). ponds and demands little management, the benefits may
Periphyton has an average C/N ratio of 10 (Azim and be substantial.
Asaeda, 2005). Its assimilation capacity is around 0.2 g N/
m2 day. From this it is clear that one needs a large surface, 3.2. Bio-flocs technology
which allows periphyton growth, to treat intensive
aquaculture wastewater without compromising the water Suspended growth in ponds consists of phytoplankton,
quality. Besides N removal, biomass is formed. The yield bacteria, aggregates of living and dead particulate organic

Fig. 5. Amorphous aggregate, which consists of phytoplankton, bacteria, aggregates of living and dead particulate organic matter, and grazers (The
flocs were examined with light microscopy, and digital images were captured with a 1-CCD camera).
 R. Crab et al. / Aquaculture 270 (2007) 1–14 9

matter, and grazers of the bacteria (Fig. 5) (Hargreaves, niques are available, such as diffuser aeration, mechanical
2006). If carbon and nitrogen are well balanced in the aeration and packed column aeration. For turbulent fluids,
solution, ammonium in addition to organic nitrogenous the mean shear rate G is determined from the power input
waste will be converted into bacterial biomass (Schneider to the fluid per unit volume of the fluid. In intensive
et al., 2005). By adding carbohydrates to the pond, aquaculture systems the average power input to the fluid is
bacterial growth is stimulated and nitrogen uptake around 1–10 W/m3 = 101–102 cm2/s3 or 10 b G b 100 s− 1
through the production of microbial proteins takes place (Boyd, 1998; McGraw et al., 2001; Schuur, 2003). At
(Avnimelech, 1999). This promoted nitrogen uptake by these moderate mixing rates, cells growing in permeable
bacterial growth decreases the ammonium concentration aggregates can profit from advective flow and grow better
more rapidly than nitrification (Hargreaves, 2006). Immo- than single dispersed cells (γ N 1). One can calculate that
bilization of ammonium by heterotrophic bacteria occurs the relative growth rate of aggregated cells in this energy
much more rapidly because the growth rate and microbial regime is greater than the growth rate of free cells (Logan
biomass yield per unit substrate of heterotrophs are a and Hunt, 1987, 1988). When more intense aeration is
factor 10 higher than that of nitrifying bacteria (Har- applied, the advantage of growing in flocs disappears and
greaves, 2006). The microbial biomass yield per unit cells growing solely show higher growth.
substrate of heterotrophic bacteria is about 0.5 g biomass We can conclude that the biological flocs can be
C/g substrate C used (Eding et al., 2006). considered as a kind of fast growing microbial mixed
In natural environments, microorganisms tend to form culture in which the ‘waste’-nitrogen is recycled to
amorphous aggregates. The settling velocity of these flocs young cells, which subsequently are grazed by the fish
appears not to relate to the square of the size, as expected (Fig. 7). Uptake of the bio-flocs by fish depends most
from Stokes' law (Logan and Hunt, 1987, 1988). If an probably on the fish species and feeding traits, fish size,
aggregate is highly porous, fluid streamlines will pene- floc size and floc density (Avnimelech, 2007). With
trate the aggregate resulting in advective flow through it. respect to feeding, this technique operates at “neutral
This will improve the supply of nutrients to the cells cost”, because it upgrades starch to protein. Moreover,
present in the aggregate and will decrease the settling one does not need to invest in an external water treatment
velocity of the flocs in the pond. system. This method is applicable to extensive as well as
Using the relative uptake factor γ, defined as growth intensive aquaculture systems. In addition, the heterotro-
rate of aggregated cells/growth rate of free cells, one can phic microbial biomass is suspected to have a controlling
make a comparison of the substrate uptake by aggregated effect on pathogenic bacteria (Michaud et al., 2006).
versus dispersed cells. Fig. 6 depicts the relative uptake Preliminary results at our laboratories have shown the
predictions for microbial cells in permeable flocs (Logan presence of poly-β-hydroxybutyrate in bio-flocs. PHB-
and Hunt, 1988). The power input to the fluid originates accumulating bacteria may abate pathogenic bacteria in
from the aeration of the ponds. Different aeration tech- aquaculture (Defoirdt et al., 2007; Halet et al., 2007).

Fig. 6. Relative growth prediction for microbial cells in permeable flocs (after Logan and Hunt, 1988).
10 R. Crab et al. / Aquaculture 270 (2007) 1–14

Fig. 7. Nitrogen cycle in bio-floc ponds. The nitrogen cycle is similar to that of in water ponds with periphyton. In contrast to the periphyton system, this
system is also applicable to intensive systems. The added carbon source, together with the waste nitrogen, is converted into microbial bio-flocs, which in
turn can be eaten by the cultured organism. This technique provides an inexpensive protein source with a higher efficiency of nutrient conversion of feed.

In what follows, some examples are discussed con- synthesis by increasing the C/N ratio of the feed and by
cerning performance of the bio-flocs technology in prac- further increasing the C/N ratio through carbohydrate
tice in freshwater systems. addition to the ponds. The added carbohydrate facilitat-
Avnimelech (1999) pointed out the use of the C/N ratio ed increased heterotrophic growth thereby augmenting
as a control element in aquaculture systems. Nitrogen shrimp production. The levels of inorganic nitrogen
control was induced by adding carbohydrates to the water, species in the water column were lower due to uptake by
and through the subsequent uptake of nitrogen by heterotrophic bacteria, making farming more sustain-
heterotrophic bacteria. This resulted in the synthesis able. The TAN levels in the water column in the study
of microbial proteins that can be eaten by the cultured fish were 0.01 mg/L, which is low compared to levels
species. Experiments with sediment suspension amended reported in other studies (0.5–3.0 mg/L) (Hopkins et al.,
with about 10 mg N/L ammonium and glucose at a 1993). Consumption of microbial flocs increased
concentration 20 times higher than that of the TAN nitrogen retention from added feed by 13% (Hari
showed that almost all the added ammonium disappeared et al., 2004).
over a period of about 2 h. Avnimelech et al. (1994) found Burford et al. (2004) promoted the growth of the
that protein utilization by fish in intensive bio-floc natural microbiota in ponds by routine addition of grain
systems is almost twice as high as the protein utilization feed (18–22% protein) and molasses as carbon sources.
in conventionally fed intensive aquaculture ponds, due to Fishmeal-based feeds and ammonia from shrimp
a recycling of the excreted nitrogen into utilizable excretion were used as nitrogen sources. The study
microbial protein. Protein recovery by tilapia rose from supports the theory that natural biota can provide a
23% in the control to 43% in the floc treatment. It was nitrogen source for shrimp, and that flocculated particles
concluded from this study that the price of feed for fish are likely to be a significant proportion of this.
production using sorghum supplemented granules (pellets Hari et al. (2006) reported that carbohydrate addition
containing only 20% protein and sorghum as a carbona- in combination with a decreased dietary protein level
ceous substrate) is just about 50% of the conventional cost improved the sustainability of shrimp farming in exten-
when 30% protein pellets were used. sive shrimp culture systems through 1) increased nitro-
The bio-flocs technology is also applicable to saline gen retention in harvested shrimp biomass, 2) reduced
systems, as discussed below. demand for feed protein, 3) reduced concentrations of
Hari et al. (2004) facilitated the development of potentially toxic TAN and NO2–N in the system, and 4)
heterotrophic bacteria and the related in situ protein reduced water based nitrogen discharge to the
 R. Crab et al. / Aquaculture 270 (2007) 1–14 11

environment. If carbohydrate was added to the water Avnimelech, Y., 2005. Tilapia harvest microbial flocs in active
column to enhance heterotrophic bacterial protein suspension research pond. Glob. Aquac. Advocate. October 2005.
Avnimelech, Y., 2007. Feeding with microbial flocs by tilapia in minimal
production, the protein level in the diet could be discharge bio-flocs technology ponds. Aquaculture 264, 140–147.
reduced from 40% to 25%, without compromising Avnimelech, Y., Lacher, M., 1979. A tentative nutrient balance for
shrimp production. intensive fish ponds. Bamidgeh 31, 3–8.
Avnimelech, Y., Ritvo, G., 2003. Shrimp and fish pond soils: processes
and management. Aquaculture 220, 549–567.
4. Conclusions and future perspectives
Avnimelech, Y., Kochva, M., Diab, S., 1994. Development of controlled
intensive aquaculture systems with a limited water exchange and
Possible effluent treatment technologies in aquaculture adjusted carbon to nitrogen ratio. Bamidgeh 46, 119–131.
are diverse. The challenge to the designers of aquaculture Azim, M.E., Asaeda, T., 2005. Periphyton structure, diversity
systems is to develop systems that maximize production and colonization. In: Azim, M.E., Verdegem, M.C.J., van Dam,
capacity per cost unit of capital invested. To do so, com- A.A., Beveridge, M.C.M. (Eds.), Periphyton — Ecology,
Exploitation and Management. CABI Publishing, Wallingford,
ponents used in recirculating systems need to be designed pp. 15–33.
and developed to reduce the cost of the unit while main- Azim, M.E., Wahab, M.A., 2005. Periphyton-based pond polyculture. In:
taining reliability. The bio-flocs technology, the periphyton Azim, M.E., Verdegem, M.C.J., van Dam, A.A., Beveridge, M.C.M.
treatment technique, integrated treatment ponds, fluidized (Eds.), Periphyton — Ecology, Exploitation and Management. CABI
sand biofilters, bead filters, trickling filters and the rotating Publishing, Wallingford, pp. 207–222.
Azim, M.E., Wahab, M.A., van Dam, A.A., Beveridge, M.C.M.,
biological contactors can be considered as good effluent Verdegem, M.C.J., 2001. The potential of periphyton-based culture
treatment technologies. The bio-flocs technology provides of two Indian major carps, rohu Labeo rohita (Hamilton) and gonia
a sustainable method to maintain water quality in Labeo gonius (Linnaeus). Aquac. Res. 32, 209–216.
aquaculture systems and moreover concurrently fish feed Azim, M.E., Wahab, M.A., Verdegem, M.C.J., van Dam, A.A., van
is produced. Since the purchase of commercially prepared Rooij, J.M., Beveridge, M.C.M., 2002. The effects of artificial
substrates on freshwater pond productivity and water quality and
feed in fish culture has a share of 50% or more in the the implications for periphyton-based aquaculture. Aquat. Living
production costs, an effluent treatment technique that Resour. 15, 231–241.
maintains water quality and simultaneously produces in Azim, M.E., Milstein, A., Wahab, M.A., Verdegem, M.C.J., 2003a.
situ fish feed has a large asset over other techniques. Periphyton–water quality relationships in fertilized fishponds with
Additional research in this field concerning management artificial substrates. Aquaculture 228, 169–187.
Azim, M.E., Verdegem, M.C.J., Mantingh, I., van Dam, A.A., Beveridge,
of the floc production, the floc dynamics in intensive M.C.M., 2003b. Ingestion and utilization of periphyton grown on
aquaculture systems, the nutritional value of flocs and the artificial substrates by Nile tilapia, Oreochromis niloticus L. Aquac.
health effects of flocs is needed, more specifically the Res. 34, 85–92.
effect on growth and survival of the cultured organisms. Azim, M.E., Verdegem, M.C.J., Singh, M., van Dam, A.A., Beveridge,
M.C.M., 2003c. The effects of periphyton substrate and fish
Also microbiological aspects need further investigation,
stocking density on water quality, phytoplankton, periphyton and
particularly the microbiological characterization of the fish growth. Aquac. Res. 34, 685–695.
flocs, possible manipulation of the microbial community Azim, M.E., Wahab, M.A., Biswas, P.K., Asaeda, T., Fujino, T.,
and presence of pre/probiotic organisms in the microbial Verdegem, M.C.J., 2004. The effect of periphyton substrate density
community of the flocs are challenging fields of interest. on production in freshwater polyculture ponds. Aquaculture 232,
441–453.
Azim, M.E., Beveridge, M.C.M., van Dam, A.A., Verdegem, M.C.J.,
Acknowledgements 2005. Periphyton and aquatic production: an introduction. In: Azim,
M.E., Verdegem, M.C.J., van Dam, A.A., Beveridge, M.C.M. (Eds.),
This work was supported by the “Instituut voor de Periphyton — Ecology, Exploitation and Management. CABI
aanmoediging van Innovatie door Wetenschap en Publishing, Wallingford, pp. 1–13.
Technologie in Vlaanderen” (IWT grant no. 51256). A Aziz, J.A., Tebbutt, T.H.Y., 1980. Significance of COD, BOD and
TOC correlations in kinetic models of biological oxidation. Water
special thank to the reviewers (ir. Han Vervaeren and ir. Res. 14, 319–324.
Peter Deschryver) for critically reading the manuscript. Bender, J., Lee, R., Sheppard, M., Brinkley, K., Philips, P., Yeboah, Y.,
Wah, R.C., 2004. A waste effluent treatment system based on
microbial mats for black sea bass Centropristis striata recycled-
References water mariculture. Aquac. Eng. 31, 73–82.
Boyd, C.E., 1985. Chemical budgets for channel catfish ponds. Trans.
Amirkolaie, A.K., 2005. Dietary carbohydrate and faecal waste in the Am. Fish. Soc. 114, 291–298.
Nile tilapia (Oreochromis niloticus L.). PhD dissertation. Univer- Boyd, C.E., 1998. Pond water aeration systems. Aquac. Eng. 18, 9–40.
sity Wageningen, Wageningen, The Netherlands. Brazil, B.L., 2006. Performance and operation of a rotating biological
Avnimelech, Y., 1999. Carbon/nitrogen ratio as a control element in contactor in a tilapia recirculating aquaculture system. Aquac. Eng.
aquaculture systems. Aquaculture 176, 227–235. 34, 261–274.
12 R. Crab et al. / Aquaculture 270 (2007) 1–14

Brune, D.E., Schwartz, G., Eversole, A.G., Collier, J.A., Schwedler, T.E., Halet, D., Defoirdt, T., van Damme, P., Vervaeren, H., Forrez, I., Van de
2003. Intensification of pond aquaculture and high rate photosyn- Wiele, T., Boon, N., Sorgeloos, P., Bossier, P., Verstraete, W., 2007.
thetic systems. Aquac. Eng. 28, 65–86. Poly-β-hydroxybutyrate-accumulating bacteria protect gnotobiotic
Burford, M.A., Thompson, P.J., McIntosh, R.P., Bauman, R.H., Pearson, Artemia franciscana from pathogenic Vibrio campbellii. FEMS
D.C., 2003. Nutrient and microbial dynamics in high-intensity, zero- Microbiol. Ecol. 60, 363–369.
exchange shrimp ponds in Belize. Aquaculture 219, 393–411. Hargreaves, J.A., 2006. Photosynthetic suspended-growth systems in
Burford, M.A., Thompson, P.J., McIntosh, R.P., Bauman, R.H., aquaculture. Aquac. Eng. 34, 344–363.
Pearson, D.C., 2004. The contribution of flocculated material to Hari, B., Madhusoodana, K., Varghese, J.T., Schrama, J.W., Verdegem,
shrimp (Litopenaeus vannamei) nutrition in a high-intensity, zero- M.C.J., 2004. Effects of carbohydrate addition on production in
exchange system. Aquaculture 232, 525–537. extensive shrimp culture systems. Aquaculture 241, 179–194.
Chen, S., Ling, J., Blancheton, J.P., 2006. Nitrification kinetics of biofilm Hari, B., Kurup, B.M., Varghese, J.T., Schrama, J.W., Verdegem, M.C.J.,
as affected by water quality factors. Aquac. Eng. 34, 179–197. 2006. The effect of carbohydrate addition on water quality and the
Colt, J., 2006. Water quality requirements for reuse systems. Aquac. nitrogen budget in extensive shrimp culture systems. Aquaculture
Eng. 34, 143–156. 252, 248–263.
Craggs, R.J., Adey, W.H., Jessup, B.K., Oswald, W.J., 1996. A controlled Hassen, A., Mahrouk, M., Ouzari, H., Cherif, M., Boudabous, A.,
stream mesocosm for tertiary treatment of sewage. Ecol. Eng. 6, Damelincourt, J.J., 2000. UV disinfection of treated wastewater in
149–169. a large-scale pilot plant and inactivation of selected bacteria in a
Defoirdt, T., Halet, D., Sorgeloos, Vervaeren, H., Boon, N., Van de Wiele, laboratory UV device. Bioresour. Technol. 74, 141–150.
T., Sorgeloos, P., Bossier, P., Verstraete, W., 2007. The bacterial Helfman, G.S., Collette, B.B., Facey, D.E., 1997. The Diversity of
storage compound poly-β-hydroxybutyrate protects Artemia fran- Fishes. Blackwell publishing, Malden, USA. 1006 pp.
ciscana from pathogenic Vibrio campbellii. Environ. Microbiol. 9, Hepher, B., 1988. Nutrition of Pond Fishes. Cambridge Univ. Press,
445–452. Cambridge, NY, USA. 388 pp.
Delgado, C.L., Wada, N., Rosegrant, M.W., Meijer, S., Mahfuzuddin, A., Hertrampf, J.W., Piedad-Pascual, F., 2000. Handbook on Ingredients
2003. Outlook for fish to 2020 — meeting global demand. A 2020 for Aquaculture Feeds. Kluwer Academic Publishers, Dordrecht,
Vision for Food, Agriculture, and the Environment Initiative. The Netherlands. 624 pp.
International Food Policy Research Institute, Washington, D.C., U.S.A. Hopkins, J.S., Hamilton, R.D.L., Sandifers, P.A., Browdy, C.L.,
Diab, S., Cochaba, M., Mires, D., Avnimelech, Y., 1992. Combined Stokes, A.D., 1993. Effect of water exchange rate on production,
intensive–extensive (CIE) pond system, A: inorganic nitrogen trans- water quality, effluent characteristics and nitrogen budgets of
formations. Aquaculture 101, 33–39. intensive shrimp ponds. J. World Aquac. Soc. 24, 304–320.
Dodds, W.K., 2003. The role of periphyton in phosphorus retention in Hovanec, T.A., DeLong, E.F., 1996. Comparative analysis of
shallow freshwater aquatic systems. J. Phycol. 39, 840–849. nitrifying bacteria associated with freshwater and marine aquaria.
Ebeling, J.M., Timmons, M.B., Bisogni, J.J., 2006. Engineering analysis Appl. Environ. Microbiol. 62, 2888–2896.
of the stoichiometry of photoautotrophic, autotrophic and heterotro- Huchette, S.M.H., Beveridge, M.C.M., Baird, D.J., Ireland, M., 2000.
phic removal of ammonia–nitrogen in aquaculture systems. Aqua- The impacts of grazing by tilapias (Oreochromis niloticus L.) on
culture 257, 346–358. periphyton communities growing on artificial substrate in cages.
Eding, E.H., Kamstra, A., Verreth, J.A.J., Huisman, E.A., Klapwijk, Aquaculture 186, 45–60.
A., 2006. Design and operation of nitrifying trickling filters in Hussenot, J.M.E., 2003. Emerging effluent management strategies in
recirculating aquaculture: a review. Aquac. Eng. 34, 234–260. marine fish-culture farms located in European coastal wetlands.
El Samra, M.I., Olàh, J., 1979. Significance of nitrogen fixation in fish Aquaculture 226, 113–128.
ponds. Aquaculture 18, 367–372. Jiménez-Montealegre, R., Verdegem, M.C.J., van Dam, A., Verreth, J.A.J.,
FAO, 2004. State of World Fisheries and Aquaculture. FAO, Rome. 2002. Conceptualization and validation of a dynamic model for the
Flemish government, 2005. Afvalwaterproblematiek op melkveebedrij- simulation of nitrogen transformations and fluxes in fish ponds.
ven. Flemish Government, Department Agriculture and Fisheries, Ecol. Model. 147, 123–152.
Department Sustainable Agricultural Development, Belgium. 61 pp. Joye, S.B., Hollibaugh, J.T., 1995. Influence of sulphide inhibition of
Franco-Nava, M.A., Blancheton, J.P., Deviller, G., Charrier, A., Le-Gall, nitrification on nitrogen regeneration in sediments. Science 270,
J.Y., 2004a. Effect of fish size and hydraulic regime on particulate 623–625.
organic matter dynamics in a recirculating aquaculture system: Kamstra, A., van der Heul, J.W., Nijhof, M., 1998. Performance and
elemental carbon and nitrogen approach. Aquaculture 239, 179–198. optimisation of trickling filters on eel farms. Aquac. Eng. 17,
Franco-Nava, M.A., Blancheton, J.P., Deviller, G., Charrier, A., Le- 175–192.
Gall, J.Y., 2004b. Particulate matter dynamics and transformations Kautsky, N., 2004. Aquaculture and the Environment — Integrated
in a recirculating aquaculture system: application of stable isotope Aquaculture. Course, University Ghent, Belgium.
tracers in seabass rearing. Aquac. Eng. 31, 135–155. Keshavanath, P., Gangadhar, B., 2005. Research on periphyton-based
Greiner, A.D., Timmons, M.B., 1998. Evaluation of the nitrification aquaculture in India. In: Azim, M.E., Verdegem, M.C.J., van Dam,
rates of microbead and trickling filters in an intensive recirculating A.A., Beveridge, M.C.M. (Eds.), Periphyton — Ecology,
tilapia production facility. Aquac. Eng. 18, 189–200. Exploitation and Management. CABI Publishing, Wallingford,
Grommen, R., Van Hauteghem, I., Van Wambeke, M., Verstraete, W., pp. 223–236.
2002. An improved nitrifying enrichment to remove ammonium Körner, S., Das, S.K., Veenstra, S., Vermaat, J.E., 2001. The effect of
and nitrite from freshwater aquaria systems. Aquaculture 211, pH variation at the ammonium/ammonia equilibrium in wastewa-
115–124. ter and its toxicity to Lemna gibba. Aquat. Bot. 71, 71–78.
Gutierrez-Wing, M.T., Malone, R.F., 2006. Biological filters in Lavens, P., Sorgeloos, P., 1984. Controlled production of Artemia
aquaculture: trends and research directions for freshwater and cysts under standard conditions in a recirculating culture system.
marine applications. Aquac. Eng. 34, 163–171. Aquac. Eng. 3, 221–235.
 R. Crab et al. / Aquaculture 270 (2007) 1–14 13

Lekang, O.-I., Kleppe, H., 2000. Efficiency of nitrification in trickling Schnel, N., Barak, Y., Ezer, T., Dafni, Z., van Rijn, J., 2002. Design and
filters using different filter media. Aquac. Eng. 21, 181–199. performance of zero-discharge tilapia recirculating system. Aquac.
Ling, J., Chen, S., 2005. Impact of organic carbon on nitrification Eng. 26, 191–203.
performance of different biofilters. Aquac. Eng. 33, 150–162. Schuur, A.M., 2003. Evaluation of biosecurity applications for intensive
Logan, B.E., Hunt, J.R., 1987. Advantages to microbes of growth in shrimp farming. Aquac. Eng. 28, 3–20.
permeable aggregates in marine systems. Limnol. Oceanogr. 32, Sugiura, S.H., Marchant, D.D., Wigins, T., Ferraris, R.P., 2006. Effluent
1034–1048. profile of commercially used low-phosphorus fish feeds. Environ.
Logan, B.E., Hunt, J.R., 1988. Bioflocculation as a microbial response Pollut. 140, 95–101.
to substrate limitations. Biotechnol. Bioeng. 31, 91–101. Summerfelt, S.T., 2003. Ozonation and UV irradiation — an introduction
Lyssenko, C., Wheaton, F., 2006. Impact of positive ramp short-term and examples of current applications. Aquac. Eng. 28, 21–36.
operating disturbances on ammonia removal by trickling and sub- Summerfelt, S.T., 2006. Design and management of conventional
merged-upflow biofilters for intensive recirculating aquaculture. fluidized-sand biofilters. Aquac. Eng. 34, 275–302.
Aquac. Eng. 35, 26–37. Tawfik, A., Klapwijk, B., Van Buuren, J., El-Gohary, F., Lettinga, G.,
Malone, R.F., Beecher, L.E., 2000. Use of floating bead filters to re- 2004. Physico-chemical factors affecting the E. coli removal in a
condition recirculating waters in warmwater aquaculture production rotating biological contactor (RBC) treating UASB effluent. Water
systems. Aquac. Eng. 22, 57–73. Res. 38, 1081–1088.
Malone, R.F., Pfeiffer, T.J., 2006. Rating fixed film nitrifying biofilters used Thompson, F.L., Abreu, P.C., Wasielesky, W., 2002. Importance of bio-
in recirculating aquaculture systems. Aquac. Eng. 34, 389–402. film for water quality and nourishment in intensive shrimp culture.
Masser, M.P., Rakocy, J., Losordo, T.M., 1999. Recirculating aqua- Aquaculture 203, 263–278.
culture tank production systems — management of recirculating Timmons, M.B., 1984. Use of foam fractionation in aquaculture. Dev.
systems. SRAC Publication, vol. 452. Aquac. Fish. Sci. 27, 247–279.
McGraw, W., Teichert-Coddington, D.R., Rouse, D.B., Boyd, C.E., Timmons, M.B., Summerfelt, S.T., 1998. Application of fluidized sand
2001. Higher minimum dissolved oxygen concentrations increase bed biofilters to aquaculture. In: Libey, G.S., Timmons, M.B.
penaeid shrimp yields in earthen ponds. Aquaculture 199, 311–321. (Eds.), Recent Advances in Aquaculture Engineering; Proceedings
Meade, J.W., 1985. Allowable ammonia for fish culture. Prog. Fish-Cult. of the Second International Conference on Recirculating Aqua-
47, 135–145. culture. Virginia Polytechnic Institute and State University,
Michaud, L., Blancheton, J.P., Bruni, V., Piedrahita, R., 2006. Effect of Roanoke, VA, pp. 342–354.
particulate organic carbon on heterotrophic bacterial populations and Timmons, M.B., Ebeling, J.M., Wheaton, F.W., Summerfelt, S.T.,
nitrification efficiency in biological filters. Aquac. Eng. 34, 224–233. Vinci, B.J., 2002. Recirculating aquaculture systems, 2nd edition.
Miller, G.E., Libey, G.S., 1985. Evaluation of three biological filters suit- NRAC Publication, vol. 01-002.
able for aquaculture applications. J. World Maric. Soc. 16, 158–168. Timmons, M.B., Holder, J.L., Ebeling, J.M., 2006a. Application of
Milstein, A., 2005. Effect of periphyton on water quality. In: Azim, M.E., microbead biological filters. Aquac. Eng. 34, 332–343.
Verdegem, M.C.J., van Dam, A.A., Beveridge, M.C.M. (Eds.), Timmons, N., Timmons, M.B., Ebeling, J.M., 2006b. Recirculating
Periphyton — Ecology, Exploitation and Management. CABI Aquaculture System (RAS) technologies — Part 2. Aquac. Mag.
Publishing, Wallingford, pp. 179–190. 32–39 (September/October 2006).
Muthuwani, V., Lin, C.K., 1996. Water quality and nutrient budget in Torres-Beristain, B., Verdegem, M., Kerepeczki, E., Verreth, J., 2006.
intensive shrimp culture ponds. Book of Abstracts, The 1996 Annual Decomposition of high protein aquaculture feed under variable
Meeting of the World Aquaculture Society, p. 270. January 29 – oxic conditions. Water Res. 40, 1341–1350.
February 2, Bangkok Thailand. Turker, H., Eversole, A.G., Brune, D., 2003. Comparative Nile tilapia
Naylor, R.L., Goldburg, R.J., Primavera, J.H., Kautsky, N., Beveridge, and silver carp filtration rates of Partitioned Aquaculture System
M.C.M., Clay, J., Folke, C., Lubchenco, J., Mooney, H., Troell, phytoplankton. Aquaculture 220, 449–457.
M., 2000. Effect of aquaculture on world fish supplies. Nature 405, Twarowska, J.G., Westerman, P.W., Losordo, T.M., 1997. Water
1017–1024. treatment and waste characterization evaluation of an intensive
Neori, A., Chopin, T., Troell, M., Buschmann, A.H., Kraemer, G.P., recirculating fish production system. Aquac. Eng. 16, 133–147.
Halling, C., Shpigel, M., Yarish, C., 2004. Integrated aquaculture: UN, 2003. Wastewater treatment technologies: a general review.
rationale, evolution and state of the art emphasizing seaweed Economic and Social Commission for Western Asia. United
biofiltration in modern mariculture. Aquaculture 231, 361–391. Nations, New York. 122 pp.
Park, D., Lee, D.S., Young, J.Y., Park, J.M., 2005. Comparison of van Dam, A.A., Verdegem, M.C.J., 2005. Periphyton: systems ecology
different bioreactor systems for indirect H2S removal using iron- perspective. In: Azim, M.E., Verdegem, M.C.J., van Dam, A.A.,
oxidizing bacteria. Process Biochem. 40, 1461–1467. Beveridge, M.C.M. (Eds.), Periphyton — Ecology, Exploitation
Piedrahita, R.H., 2003. Reducing the potential environmental impact and Management. CABI Publishing, Wallingford, pp. 91–111.
of tank aquaculture effluents through intensification and recircu- van Dam, A.A., Beveridge, M.C.M., Azim, M.E., Verdegem, M.C.J.,
lation. Aquaculture 226, 35–44. 2002. The potential of fish production based on periphyton. Rev.
Read, P., Fernandes, T., 2003. Management of environmental impacts Fish Biol. Fish. 12, 1–31.
of marine aquaculture in Europe. Aquaculture 226, 139–163. van Rijn, J., 1996. The potential for integrated biological treatment systems
Satoh, H., Okabe, N., Watanabe, Y., 2000. Significance of substrate C/N in recirculating fish culture — a review. Aquaculture 139, 181–201.
ratio on structure and activity of nitrifying biofilms determined by in van Rijn, J., Tal, Y., Schreier, H.J., 2006. Denitrification in recirculating
situ hybridization and the use of microelectrodes. Water Sci. Technol. systems: theory and applications. Aquac. Eng. 34, 364–376.
41, 317–321. Vermaat, J.E., 2005. Periphyton dynamics and influencing factors. In:
Schneider, O., Sereti, V., Eding, E.H., Verreth, J.A.J., 2005. Analysis of Azim, M.E., Verdegem, M.C.J., van Dam, A.A., Beveridge, M.C.M.
nutrient flows in integrated intensive aquaculture systems. Aquac. (Eds.), Periphyton — Ecology, Exploitation and Management.
Eng. 32, 379–401. CABI Publishing, Wallingford, pp. 35–49.
14 R. Crab et al. / Aquaculture 270 (2007) 1–14

Villaverde, S., Fdz-Polanco, F., García, P.A., 2000. Nitrifying biofilm Wang, J.-K., 2003. Conceptual design of a microalgae-based
acclimation to free ammonia in submerged biofilters, Start-up in- recirculating oyster and shrimp system. Aquac. Eng. 28, 37–46.
fluence. Water Res. 34, 602–610. Watten, B.J., Sibrell, P.L., 2006. Comparative performance of fixed-
Walsh, P.J., Wright, P.A., 1995. Nitrogen Metabolism and Excretion. film biological filters: application of reactor theory. Aquac. Eng.
CRC Press, Florida, USA. 352 pp. 34, 198–213.