An Experimental Comparison of Sandbed and Plenum-Based Systems: Part 2: Live Animal Experiments

Last month we presented the
results from our experiments comparing deep sandbed and
plenum-based aquaria under controlled conditions in the
laboratory as Part 1 of this series.
Although those results are very useful in understanding the real
effects of the different aquarium designs, few people are really
interested in keeping a tank without any live animals. This
month, we will continue our experimental comparison of sandbed
and plenum-based systems under more realistic conditions. This
second experiment uses live rock, fish and invertebrates along
with the full complement of natural sandbed infauna found in
similarly-sized sediments on Hawaiian reefs to evaluate the
relative nutrient processing capacity of sandbeds of various
depths and grain sizes with and without a plenum beneath them. In
this article we will explain our live animal experiments, and
present the results of this experiment to scientifically examine
the relative contribution of: (1) a plenum void-space (sandbed
with or without plenum); (2) the sediment depth of the bed (2.5
versus 9.0 cm); and (3) the mean particle size of
sediments in the bed (2.0 versus 0.2 mm mean particle
diameter) to their nutrient processing capacity and performance
as a lone filtration method for recirculating aquaria.

We presented the background for this study in Part 1 of this
series, and will not repeat it here. If you have not already done
so, please read Part 1 for the
relevant introduction to this work.

Experimental Methods & Materials:

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a) Aquarium set up and nutrient dosing

As described in the previous article, we set up a factorial
design experiment with three replicate nano-tanks (27 cm long x
17 cm wide x 30 cm high) for each factor: with or without plenum,
deep or shallow, and coarse or fine sediments for a total of 24
experimental aquaria (Fig. 1a,b). Unlike the previous experiment,
the live animal trials were run under natural conditions which
were quite variable through time. Aquaria were maintained
outdoors in a shade enclosure to protect the aquaria from inputs
of rain and direct sunlight, but both temperature and light
fluctuated at natural rates. During the experiment, the maximum
air temperature recorded was 33ºC (~91ºF) and the minimum air
temperature recorded was 19ºC (~66ºF); aquarium temperatures
varied less than these extremes, and ranged from 22 and 30ºC (~72
to 86ºF).

Figure 1a

fig1a.jpg

Schematic of aquarium design experiment to
compare directly the effects of the presence or absence of a
plenum, the depth of the sediment bed, and the mean particle
size of sediments in recirculating aquarium systems.

Sediments from the previous experiment were removed from each
tank and combined together along with an equivalent volume of
natural sediments of equivalent size collected from the lagoon at
Coconut Island (Hawaii Institute of Marine Biology, Kaneohe, HI).
These sediments, along with the natural infauna community, were
mixed thoroughly by hand and then redistributed among the aquaria
for each treatment. As with the previous experiment, the deep
sandbed treatments contained 9.0 L (~2.4 gallons) of wet sediment
to provide a constant depth of roughly 9.0 cm (~3.6″).
Shallow sandbed treatments contained 2.5 L (~0.7 gallons) of wet
sediment to provide a constant depth of roughly 2.5 cm
(~1″). Florida crushed coral gravel (#0, mostly oblong,
averaging ~2x4mm, with a mean particle diameter ~2.0 mm) from the
first experiment was mixed with similarly sized particles
collected from around the reefs in Kaneohe Bay, Hawaii. Southdown
Tropical Play Sand (mean particle diameter ~0.2 mm) from the
first experiment was mixed with similarly sized sediments
collected from the lagoon at Coconut Island in Kaneohe Bay,
Hawaii. Sediments of each type were thoroughly mixed before being
redistributed among the treatment aquaria (Figure 1b).

Figure 1b

fig1b.jpg

Photographs of the aquaria used in this
experiment.

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Treatments were assigned to the same aquaria as in the first
trial, and were set up in the same way as outlined in Part 1.
After sediments were distributed among aquaria, the tanks were
allowed to settle for 1 week without any nutrient additions prior
to the introduction of live animals. Water was circulated using a
CAP-180 powerhead as described previously for the dosing
experiments. Water parameters were tested (see below) for each
aquarium at the end of this week to determine the starting
conditions for each trial aquarium.

After the 1 week stabilization period, we added 1 kg of
“live rock” (consisting of 1-3 pieces of natural coral
rubble collected from the nearby reef), a single Hawaiian
sharpnose puffer (Canthigaster jactator), a small rock
urchin (Echinometra oblongata), ten left-handed hermits
(Calcinus laevimanus), and 10 snails (5
Littorina sp. and 5 Nerita sp.) to each
aquarium. Although this sounds like only a light stocking level,
it is important to keep in mind that the nano-tanks used for this
experiment were only 3 gallons. Also, the deep sediment trials
were half filled with sand leaving only half the aquarium volume
for water and animals. If we were to scale this stocking level up
to a 50 gallon tank, we’d have 50 lbs of live rock, 90g of
fish (roughly equivalent to 8 or 9 adult yellow tangs), 16
golf-ball sized urchins, 220 hermit crabs, 220 snails and all the
natural infauna associated with a natural coral reef environment.
Clearly when you think of the stocking level on that scale, each
tank contained a decent bioload relative to a well-stocked reef
tank.

We prepared homogenized squid (Loligo sp.) pellets
for food as described in Pawlik et al. (1995). Fish were fed
ad libitum each week day (unfed on weekends) until they
did not ingest the final pellet offered to them. The final
uneaten pellet was left in the aquarium to provide food for
scavengers in the tank. The number of pellets fed to each tank
therefore differed from day-to-day and tank-to-tank. The final
number of pellets fed to each tank was different at the end of
the experiment. However, over the course of the entire
experiment, there were no significant differences in the number
of squid pellets fed per treatment. Any deaths of animals in the
aquaria were recorded at each testing period, and replacement
animals were added as necessary to maintain a constant bioload of
the same number of live animals in each treatment throughout the
experimental period.

The experiment ran for 118 days after the addition of live
animals without any water changes. Again, the salinity of each
aquarium was adjusted to ~53 mS every other day as outlined in
Part 1.

b) Aquarium water testing

All tanks were initially filled from a large holding tank of
well-mixed natural seawater taken from the seawater system at the
Hawaii Institute of Marine Biology. A single 50ml sample of this
water was collected and frozen at -80°C until water analyses were
completed at the end of the experiment. Likewise, a single 50ml
sample from each aquarium was collected at the end of the
experiment and also frozen at -80ºC. At the completion of both
portions of the experiment (this experiment and the laboratory
dosing experiment outlined in Part 1 of this series), all water
samples were transported frozen to the University of Hawaii at
Manoa and water nutrient concentrations were determined using
colorimetric methods on a Technicon AutoAnalyzer as outlined in
Laws et al. (1999).

Each experimental aquarium was also tested twice or three
times per week for salinity, pH, ammonia, nitrite, nitrate,
oxygen, phosphate, calcium, alkalinity, and organics using
standard aquarium testing equipment from an online aquarium
supplier. Salinity was determined using an electronic PinPoint
salinity meter (calibrated to 53.0 mS using IAPSO seawater) and
pH was measured with the electronic PinPoint pH probe (after
2-point calibration to 7.0 and 10.0). All other water parameters
were measured using standard Salifert aquarium test kits as
outlined in Part 1.

Figure 2

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fig2.jpg

Plot of the readings obtained from the
Salifert aquarium test kits used to record daily nutrient
levels presented, and the readings provided from the Water
Chemistry Analysis Lab at the University of Hawaii at Manoa
using a Technicon AutoAnalyzer. For direct comparison, the
readings obtained from the Salifert test kits (mg/L) are
converted into the same concentration units as those obtained
from the Technicon AutoAnalyzer (µM).

c) Statistical analyses

If you are not familiar with statistics at all, you will
probably want to skip this section. It won’t stop you from
being able to read the rest of the article, but we present the
details here for those readers who want to know how the analyses
were done.

We performed all statistical tests using JMPin ver. 4.0.2
Academic Version (SAS Institute Inc.). We performed a bivariate
linear fit of aquarium test kit readings (response variable) to
the AutoAnalyzer readings (factor), and tested the significance
of the Lack of Fit test as implemented in JMPin. All other
statistical analyses were carried out using an Analysis of
Variance as implemented in JMPin. We first confirmed conformity
to assumptions of normality using Shapiro-Wilks, and homogeneity
of variance using Bartletts test (alpha = 0.01) before performing
the ANOVA analysis. The full ANOVA model used as the presence or
absence of a plenum, the mean particle size of sediments, the
depth of the bed and interactions among them as fixed effects;
the salinity, pH, ammonia, nitrite, nitrate, oxygen, phosphate,
alkalinity, and calcium were measured as response variables.
Significant differences among treatment pairs (plenum
vs. none; fine vs. coarse particles; deep
vs. shallow sediments) were determined for each response
variable through effect tests as implemented in JMPin. Data were
plotted using PSI Plot ver, 7.01 (Poly Software International,
Inc.).

Our Experimental Results

a) Live animal aquarium experiments

Time-series of ammonia, nitrite and nitrate concentrations in
aquaria showed little difference among treatments (Figs. 3-5). As
with the dosing experiments presented in Part 1, the time-series
of pH, salinity, ammonia, nitrite and nitrate concentrations in
aquaria showed no significant differences among treatments (data
not shown). Analyses of variance for each water parameter
revealed no significant differences among the final salinity,
ammonia, nitrite, oxygen, or organic concentrations, nor were
there any significant interactions among experimental treatments
for any of these water parameters (data not shown). There were
significant differences among treatments for the remaining water
parameters and the variances are uniformly larger among
treatments including live animals than in the dosing experiments
(i.e., there is much more variation from tank-to-tank when live
animals are included in the aquarium).

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Figure 3

fig3.jpg

Time series plot of the mean ammonia
concentration (mg / L) in experimental aquaria. Note that the
ammonia concentrations throughout the entire experiment are
lower than those observed in the dosing experiments reported in
Part 1.

Figure 4

fig4.jpg

Time series plot of the mean nitrite
concentration (mg / L) in experimental aquaria. Again, readers
should note that the nitrite concentrations throughout the
experiment are lower than those observed in the dosing
experiment presented in Part 1.

Figure 5

fig5.jpg

Time series plot of the mean nitrate
concentration (mg / L) in experimental aquaria. It is
noteworthy that peak nitrate levels in the live animal
experiments are equal to those observed in the dosing
experiments presented in Part 1, but this may be a limitation
of the Salifert test kits which have a maximum reading of 100
mg / L. Overall, the concentrations of nitrate recorded
throughout the experiment are significantly lower than those
reported for the dosing experiments (similar to the results for
ammonia and nitrite).

By the end of the experiment, pH was significantly higher in
aquaria with fine (8.22 ± 0.02 SE) than coarse (8.10 ± 0.02 SE)
sediments (df = 1, F = 7.68, p = 0.01). For nitrate, the overall
analysis of variance was not significant (df = 7, F = 1.25, p =
0.34). However, there was a significant particle size by depth
interaction effect (df = 1, F = 6.48, p = 0.02), in which deep,
coarse (27.41 mg / L ± 6.89 SE) and shallow, fine (20.42 mg / L ±
6.89 SE) sediments have the highest average final nitrate
concentration, while shallow, coarse (12.08 mg / L ± 6.89 SE) and
deep, fine (0.67 mg / L ± 6.89 SE) sediments consistently had the
lowest final nitrate concentrations. Phosphate ended up
significantly higher in aquaria with coarse (0.35 ppm ± 0.02 SE)
than fine (0.02 ppm ± 0.02 SE) sediments (df = 1, F = 119.69, p
< 0.01). Phosphate was also significantly higher among deep
(0.22 mg / L ± 0.02 SE) than among shallow (0.15 mg / L ± 0.02
SE) sediment treatments (df = 1, F = 5.70, p = 0.03), although
this comparison becomes non-significant after Bonferroni
correction. Alkalinity was significantly higher in tanks with
fine (1.97 meq / L ± 0.06 SE) than with coarse (1.69 meq / L ±
0.06 SE) sediments (df = 1, F = 12.03, p < 0.01). Finally,
calcium concentrations were significantly higher in tanks with
fine (340.42 mg / L ± 2.89 SE) than with coarse (327.92 mg / L ±
2.89 SE) sediments (df = 1, F = 9.35, p < 0.01).

No other source variables or interaction terms were
significant, however all but one of the water parameters tested
(see below) showed similar trends to the dosing experiments
presented in Part 1. Overall most comparisons that were
significant in dosing experiments presented in Part 1 were also
close to significant (0.1 < p > 0.05) despite the higher
variability in live animal trials presented here. This similarity
shows that the addition of live animals to the experiment had
little overall effect on the results. Given that similarity, and
the increased variability with the inclusion of live animals, an
increased sample size for the live animal experiments would
almost certainly have shown identical trends among the two
experiments. In fact, the only parameter that showed an opposite
effects between the dosing and live animal experiments was
alkalinity in the presence or absence of a plenum (Fig. 6).

Figure 6

fig6.jpg

Comparison of final nutrient concentrations
in experimental aquaria with and without plenums. Bars
represent the mean concentration among tanks with (blue
cross-hatch) and without (red cross-hatch) a plenum beneath the
sediments. Error bars are standard errors among replicates, and
parameters that show a significant difference between a DSB and
plenum design are flagged with an asterisk. Salinity is
measured in mS, alkalinity in meq, and organics are presented
as a relative colorimetric measure. Nitrate, calcium, oxygen,
ammonia, phosphate and nitrite are all presented in mg / L.
Treatments which differ significantly between the live animal
and laboratory dosing experiments are highlighted with a bar
above them.

b) Aquarium water testing

Comparisons of the nutrient concentration in initial and final
water samples determined by AutoAnalyzer to the results obtained
from the Salifert aquarium test kits were sufficiently
well-correlated (r2 = 0.75, F = 10.19, P <
0.01) to use the aquarium test kit values as a relative
measure
of aquarium nutrients throughout the experiment.
This correlation (also presented in Part 1) is a direct
comparison of the readings from the aquarium test kits (in mg/L)
with the concentration readings from the AutoAnalyzer (in µM).
However, that is not actually a fair comparison. If we convert
the results from the aquarium test kits to µM concentrations (for
a direct comparison of the accuracy of the test, rather than the
relative magnitude as reported by the correlation above), the
correlation is actually better (r2 = 0.83, F = 5.41,
P < 0.01, Fig. 2). Although there is a decent
correlation between the values obtained with an AutoAnalyzer and
the Salifert aquarium test kit, the overall Salifert test kit
readings (143.5 ± 25.4) were significantly higher on average than
were the AutoAnalyzer (84.7 ± 15.9) readings (df = 1, F = 356.0,
p < 0.01). Although this is encouraging because it leans
toward the conservative (high) side for the test results, it is
important to note that the difference between readings could be
more than double or less than half the true value (Fig. 2
above).

So, as long as you use the test kits as a ballpark figure for
the concentration of nutrients in the aquarium, this kit performs
quite reliably (we did not test any other kits, and cannot
comment on their accuracy). However, there is considerable
variability among readings given by the Salifert kit, and if you
are interested in a highly accurate reading of nutrients in the
aquarium, I suggest you read Dana Riddle’s article on
Water Testing Devices for Advanced and
Professional Aquarists
in the Advanced Aquarist
archives.

Figure 7

fig7.jpg

Comparison of final nutrient concentrations
in experimental aquaria with deep (9.0cm) and shallow (2.5cm)
sediments. Bars represent the mean concentration among tanks
with deep (blue) and shallow (red) sediments. Error bars are
standard errors among replicates, and none of the parameters
show a significant difference between deep and shallow
sediments. Salinity is measured in mS, alkalinity in meq, and
organics are presented as a relative colorimetric measure.
Nitrate, calcium, oxygen, ammonia, phosphate and nitrite are
all presented in mg / L. Treatments which differ significantly
between the live animal and laboratory dosing experiments are
highlighted with a bar above them.

Figure 8

fig8.jpg

Comparison of final nutrient concentrations
in experimental aquaria with coarse (2.0mm mean diameter) and
fine (0.2mm mean diameter) particles. Bars represent the mean
concentration among tanks with coarse (purple) and fine (green)
sediments. Error bars are standard errors among replicates, and
parameters that show a significant difference between particle
sizes are flagged with an asterisk. Salinity is measured in mS,
alkalinity in meq, and organics are presented as a relative
colorimetric measure. Nitrate, calcium, oxygen, ammonia,
phosphate and nitrite are all presented in mg / L. Treatments
which differ significantly between the live animal and
laboratory dosing experiments are highlighted with a bar above
them.

c) Death rates among live animal treatments

We also kept track of all animal deaths in the live animal
experiments. Each animal in the experiment was treated as
equivalent, and the total number of individuals that had to be
replaced throughout the experiment was compared. We compared the
mean death rate for animals: 1) among experimental treatments
(Fig. 9), and 2) by similarity to popular aquarium designs (Fig.
10).

Figure 9

fig9.jpg

Comparison of mean death rate of all animals
(fish, snails, urchins & hermits combined) among
experimental treatments. Treatments that do not differ
significantly from one another are labeled with the same letter
above each bar (only shallow sediment trials are significantly
higher than deep sediment trials).

Figure 10

fig10.jpg

Comparison of mean death rate of all animals
(fish, snails, urchin & hermits combined) among
experimental treatments most closely resembling the setup of
popular aquarium design methods. The treatments are
abbreviations of the plenum, sandbed depth and sediment grain
size in that order. NP denotes “no plenum” while P
denotes a “plenum” is present. D stands for

“deep” beds (9.0 cm) whereas S stands for
“shallow” (3.0 cm) beds, and C denotes
“coarse” sediments (2.0 mm) while F denotes
“fine” (0.2 mm) sediment beds. It is critical to note
that we did not actually test any of the aquarium designs named
in this figure – we are simply indicating which design most
closely matches one of the experimental treatments we used in
our experiment for readers to compare. Typical plenum designs
would be halfway between PDC and PSC in depth, and
“Jaubert” is therefore labeled twice on the figure,
because the expected mortality rate would be halfway between
these extremes.

For the among treatment comparison, the overall analysis of
variance was not significant (df = 7, F = 0.88, p > 0.5).
However, there were nearly twice as many animal deaths overall in
shallow as in deep sediment tanks (Fig. 9). On average 2.91 ±
0.46 animals had to be replaced in the shallow sediment
treatments, whereas only 1.47 ± 0.46 animals had to be replaced
in the deep sediment trials (df = 1, F = 5.23, p < 0.05). No
other treatment or interaction term significantly affected the
death rate in our experiment.

If we examine the overall death rates among all depth,
particle size and aquarium design combinations, we find there are
considerable differences in the mean death rates among some of
the treatments. If we examine the aquarium design in our
experiment that is most similar to some of the common aquarium
designs, we can see some interesting differences (Fig. 10).
Although we never intended to set out to test all possible
aquarium setup designs, the factorial approach we have taken
gives us experimental tanks that are close in design to many of
the popular aquarium set ups. This allows us to compare the
relative performance of shallow coarse sediments (such as used in
the Berlin aquarium design) to shallow fine sediments (such as
used in the Miracle Mud aquarium design). Clearly these specific
designs have other components (such as a skimmer or Miracle Mud)
that would be required to directly compare these designs to those
of a deep sandbed or plenum design. However, it also seems
reasonable to us that a skimmer could be added to any of these
tanks and therefore show a different effect, so the relative
comparison of the sediments themselves (as used in the most
popular aquarium setups currently) is worth some attention. We
have labeled the sediment treatment that is closest to each
popular aquarium design in Figure 10 for comparison. The Jaubert
plenum design uses an intermediate depth sediment bed, so we have
labeled both the deep and shallow plenum design with
“Jaubert.” Given our results, we would expect the true
Jaubert plenum design to fall roughly halfway between these two
extremes.

Overall, deep coarse sediments without a plenum had the lowest
death rate in the experiment (1.0 ± 0.58 animals replaced), while
shallow coarse sediments above a plenum had the highest overall
death rate (4.0 ± 1.53) in the experiment. Shallow coarse
sediments without a plenum (2.67 ± 0.88) and shallow fine
sediments with (2.67 ± 0.33) or without (2.33 ± 1.20) a plenum
were all on the higher end of the death rates recording in the
experiment as well (Fig. 10). These higher mortality sediment
combinations are closest in design to those of the Berlin (NPSC)
and Miracle Mud (NPSF) systems (Figure 10).

Discussion & Conclusions

As we explained in Part 1, public aquaria and hobbyists at
home have long used recirculating systems based on some form of
sediment filtration to aid in the processing of nitrogenous
wastes produced by tank inhabitants (reviewed by Delbeek, Sprung,
1994a; b; Carlson, 1999; Toonen, 2000a; b; Borneman, Lowrie,
2001; Delbeek, Sprung, In press). The design of these sediment

filtration units for recirculating systems to culture coral reef
organisms fall largely into only a few major types: Berlin,
Miracle Mud, plenum and sandbed-based systems. However, these
systems can also be viewed as a simple continuum from virtually
no sediment with complete reliance on live rock and
protein-skimming in Berlin systems, to extreme amounts of
sediment and no skimmer with some deep sandbed systems. Despite
the diversity of opinions on the value of these different
designs, the relative utility of each of these types, and the
most effective means to design them are still a subject of
considerable controversy (reviewed by Toonen, 2000a; b).

There have been some studies to compare the relative
performance of a given design (e.g., Auger, 1999; Hovanec, 2003),
however to date these studies have all been unreplicated and only
show results based on comparisons from a single aquarium of each
design. In Part 1, we showed how even in a replicated experiment
set up in a laboratory without any live animals and with ammonia
dosed in to simulate an identical bioload among tanks, there is
far too much variability to draw any conclusions
based on a single tank. We reiterate the point made by Terry
Siegel in his editorial last
month
: we need experimental evidence to draw any intelligent
conclusions about the relative advantages of any particular
aquarium design or additive.

The varied opinions and continuous debate of this subject were
what led us to begin this experiment a couple of years ago, and
here we finally present the experimental data that compare
directly a variety of recirculating nano-reef aquarium designs.
We performed a controlled and replicated factorial design
experiment to determine the relative effect of the presence or
absence of a plenum, the depth of the sediments and the size of
the particles in the sandbed on the concentration of nutrients in
the aquarium. Put simply, our experiment shows that the presence
of a plenum has no measurable benefits over simply depositing the
same sediments directly on the bottom of the aquarium (at least
for nano-tanks over the time scales that we tested).

With a single exception, the results of the live animal
experiments were similar to those of the animal-free dosing
experiments (Figs. 7-9). Only alkalinity showed the opposite
pattern of significance in the presence or absence of a plenum
among the dosing and live animal experiments (Fig. 7). Although
final concentrations of nitrate and calcium did not vary among
plenum, sediment depth or particle size treatments within either
the dosing or the live animal experiments, both differed
significantly between the two experiments. Nitrate concentrations
of experimental aquaria in the live animal experiments (15.15 ±
17.51) were significantly lower than those of the dosing
experiments (62.76 ± 14.47) (df = 1, F = 150.33, p < 0.01).
Likewise, final calcium concentrations of experimental aquaria
with live animals (334.17 ± 11.81) were significantly lower than
those in the aquarium dosing experiments (446.67 ± 37.15) (df =
1, F = 199.95, p < 0.01).

We cannot exclude the possibility that the presence of live
animals in the aquarium may alter the buffering capacity or the
rate of denitrification. However, the most likely explanation for
reduced final calcium concentrations is uptake by organisms in
the trial aquaria which could not happen in the dosing
experiments presented in Part 1. The same could be said for the
nitrate concentration, but there are at least three additional
potential explanations for the differences between the live
animal and dosing trials in the final nitrate concentration.
First, the presence of the coral rubble (‘live rock’) in
the live animal trials could well have increased the biological
filtration capacity, and could account for the reduced final
nitrate concentrations. Second, the waste introduced to the
aquarium by the live animals is likely much lower than 0.5mg
NH4+ / L / day. Based on a rough
calculation of size-specific nitrogenous waste production from
Qian and colleagues (2001), we estimate that the rate of ammonium
production in the live animal trials was probably between 0.05
and 0.08 NH4+ / L / day. Finally, the live
animal trials were conducted outside beneath a shade enclosure,
and the presence of algae in these treatments could easily
account for significant nitrate uptake relative to the aquarium
dosing treatments. Further experimentation would be required to
address the specific cause of the reduced nitrate concentrations
in the live animal trials.

To our surprise, the majority of the nutrient processing
capacity appears to be explained quite simply by microbial
processes. These experiments show no evidence that the presence
or absence of live animals and sediment infauna have a measurable
effect on the nutrient processing capacity of sediments (Figs.
7-9) – at least in nano-tanks on the time scales covered by this
experiment. However, the question of how these results scale up
to larger aquaria, and the role of sediment infauna in the
long-term stability of closed systems certainly remains a subject
for future studies. We cannot address these questions with our
data, and hope that someone will follow up on this study to
specifically research the question of time and scale in these
systems.

Perhaps the most perplexing result from this experiment is the
significant interaction of sediment particle size and depth in
the aquaria. The simple prediction based on sandbed depth would
be that deeper and finer sediments should always have reduced
oxygen penetration and therefore increased nitrate processing
capacity (Toonen, 2000a; b; Shimek, 2001; Delbeek, Sprung, In
press). Therefore, it is hard to explain why deep, coarse (27.41
mg / L ± 6.89 SE) and shallow, fine (20.42 mg / L ± 6.89 SE)
sediments have the highest average final nitrate concentration,
while shallow, coarse (12.08 mg / L ± 6.89 SE) and deep, fine
(0.67 mg / L ± 6.89 SE) sediments consistently had the lowest
final nitrate concentrations. Nitrate reduction in deep, fine
sediments is easily explained by reduced oxygen penetration to
the sediments. However, the increased final nitrate
concentrations in aquaria with deep, coarse and shallow, fine
sediments relative to the shallow, coarse treatment is harder to
understand. Additional research will be required to explain the
source of denitrification in shallow, coarse sediments and
account for this unexpected result. During his MACNA XVI
presentation, Julian Sprung discussed his research into the
physical effects of water motion on the biological filtration
capacity of sediment beds in aquaria. The basic conclusion from
that work (covered in more detail in Delbeek, Sprung, In press)
is that the location and volume of rock as well as the surface
shape of the sand or gravel (e.g., mounds, sloped, or flat) can
dramatically affect the efficiency of water flow, oxygen
diffusion and nutrient processing in the sandbed. The results we
present here likewise argue that there are complex interactions
between sandbed depth, particle size and flow that are sometimes
counter-intuitive. Obviously, additional research along these
lines may prove very fruitful to our ultimate understanding of
biological filtration in recirculating aquaria.

Overall, both the results of the dosing and live animal
experiments suggest that there is no measurable difference
between most of these common sediment filtration designs for
maintaining suitable water parameters. There were no significant
differences among depth, particle size or plenum treatments for
the processing of ammonia or nitrite in recirculating aquarium
systems. Deep, fine sediments had the lowest average final
concentration of nitrate in these trials, but these values were
not significantly less than the average final concentration of
nitrate in shallow, coarse sediment treatments. Also, contrary to
our expectations, the presence or absence of live animals and
sandbed infauna made no significant difference to the nutrient
concentrations across the time periods tested here. So what does
explain the differences among aquaria in these experiments? Well,
it turns out that the best predictor of aquarium nutrient levels
is quite simply the bioload and any animal deaths in the tanks.
Aquaria that had low (even undetectable) levels of ammonia,
nitrite and nitrate would suddenly show a substantial peak in
nitrogenous wastes following the death of an animal in the
aquarium (Fig. 11). Our results suggest that stocking level of
the aquarium, and any animal deaths, have a much greater effect
on the overall water quality than the specific design of the
aquarium set-up you chose to follow.

Figure 11

fig11.jpg

Time series of nitrate in an experimental
aquarium prior to and after the death of a fish in the
aquarium. The arrow on the plot marks the point at which the
fish died, and the ammonia, nitrite and nitrate level all show
a significant increase immediately after that event (we present
only one plot as a representative data set). Overall, bioload
and animal deaths in the aquarium appear to be the best
predictors of water quality in these experiments.

Ultimately, however, we suspect that most aquarists are less
concerned about the exact concentrations of any of these water
parameters, and are instead acutely concerned with whether or not
animals survive in their aquaria. Our experiment showed that
sandbed systems had a slightly lower mortality rate than systems
based on a plenum; likewise, mortality in tanks with coarse
sediments was very slightly lower than those based on fine
sediment, but neither effect was significant (Fig. 9). The only
significant effect was that death rate in shallow sediments was
significantly higher than (almost twice) that of tanks with deep
sediments (Fig. 9), and the highest death rate of all was
observed in aquaria with shallow, coarse sediments over a plenum
(Fig. 10). Insofar as the sediments themselves are an important
component of the aquarium design, these results can be used to
infer the relative efficiency of a variety of aquarium designs
with different depths and sizes of sediment included within
them.

In conclusion, regardless of whether we look at the laboratory
dosing or the live animal experiments, our results show no
evidence for any of the espoused benefits of a plenum. To the
contrary, these experiments suggest that any benefits seen are a
direct consequence of the sediments themselves rather than the
void space beneath it. However, at least over the time scales we
could test, our experiments also showed little support for the
espoused benefits of a natural community of infauna in the
sediments. Overall, there was only one qualitatively different
trend observed between the laboratory dosing and live animal
trials, and that was alkalinity rather than any of the
nitrogenous waste products (Figs. 6-8). This study is the first
experimental comparison of aquarium designs to compare their
relative performance. We cannot address any long-term effects on
aquarium maintenance or survivorship in this experiment, and we
hope that others will follow up on our work to address this
issue. This sort of experimental data is necessary to evaluate
alternative aquarium designs or additives objectively, and
anecdotal evidence or unreplicated studies should always be
viewed with skepticism.

Overall Summary:

  • Our experiment shows no evidence for any of the espoused
    benefits of a plenum (reviewed by Goemans 1999) either with or
    without live animals in the design. Instead our results suggest
    that any benefits seen are a direct consequence of the presence
    of the sediments themselves rather than the void space beneath
    it.
  • Each sediment-based aquarium design appeared capable of
    handling nutrient inputs up to 0.5 mg / L / day of
    NH4+ – which is equivalent to a
    well-stocked reef aquarium. At this input level, final
    concentrations of ammonia, nitrite and nitrate did not differ
    significantly among aquaria 1) with or without plenums, 2)
    containing deep (9.0 cm) or shallow (2.5cm) sediments, or 3)
    containing coarse (2.0mm) or fine (0.2mm) mean particle sizes.
    Bioload and animal deaths in the aquarium show a much greater
    effect on the water quality than does the specific design for
    the tank.
  • The greatest differences among experimental treatments were
    observed as decreased buffering capacity, and higher final
    phosphate concentration of aquaria with coarse sediments
    relative to those with fine sediments. However, the chemical
    composition of the gravel may be responsible for this effect,
    and we have not tested other gravel types of similar size. We
    recommend that aquarists test any new gravel for dissolution
    before adding a lot of it to their aquarium.
  • We show that there can be extreme variation among identical
    tanks, even without any live animals included as outlined in
    Part 1. Given the added variability as soon as live animals are
    included into the mix, our results highlight the problem with
    drawing any conclusions based on a single aquarium – no matter
    how beautiful it may be. The results from any study lacking
    proper replication and controls should be viewed with
    suspicion. We argue that anecdotal evidence is simply
    presentation of an opinion in cases such as this, and more than
    5 years of heated debate on the merits of DSB vs. plenum
    systems has resulted from the staunch defense of opinions
    without data.
  • We show that even high-quality aquarium tests provide only
    a ballpark estimate of the actual concentration of nutrients in
    the aquarium. However the readings from the Salifert test kits
    were sufficiently correlated with the true nutrient values to
    be reliable for comparisons among aquaria.
  • Overall death rates were roughly twice as high in aquaria
    with shallow sediments as in deep sediment treatments. The
    highest overall death rates were seen in aquaria with shallow
    coarse sediments over a plenum, and the lowest death rates
    occurred in aquaria with a sandbed composed of deep coarse
    sediments. The treatments that were closest to the design
    aquarists employ for deep sandbed, Miracle Mud and Jaubert
    plenum aquaria had intermediate death rates. The shallow coarse
    sediment design that is closest to that used in Berlin systems
    had one of the highest death rates, and the deep coarse
    sediment design for which there is currently no accepted name
    had the lowest overall mortality (Fig. 10). We did not test
    bare bottom tanks, but the data clearly suggest that the
    shallower the sediment, the higher the mortality rate, and you
    can’t get much shallower than a bare bottom tank!
  • Experimental results were surprisingly similar between the
    aquarium dosing and live animal experiments. Contrary to our
    expectations, the presence of live animals and sediment infauna
    does not have any measurable effect on final nutrient
    concentrations in our experimental aquaria.

Acknowledgements

This research was funded in part by a Program Development
Award to RJT from Hawaii Sea Grant. Additional funding came via
donations from Reed Mariculture, Catalina Aquarium and my very
understanding wife, Carol Fong. Water testing was performed by
Saipologa Toala and Houston Lomae as part of a Pacific Islander
Undergraduate Mentorship in Environmental Biology (UMEB)
internship, and we greatly appreciate their diligence and hard
work assisting with this project. We thank Ross Shaw for taking
photographs of our aquarium setups. This manuscript was improved
by discussion and comments from Eric Borneman, Anthony Calfo,
Charles Delbeek, Tom Frakes, Richard Harker, Tim Hovanec, Larry
Jackson, Julian Sprung and the many other excellent aquarists at
the XVI Marine Aquarium Conference of North America.

References

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