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International Journal of Aquaculture 2012, Vol.2, No.6, 29-39
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35
environmentally friendly and biosecure practice for
the production of food size
L. vannamei.
Pre-conditioned
water with an established heterotrophically dominated
community of microorganisms may serve to provide a
more balanced system for the control of harmful
nutrients such as NO
2
-N and NH
3
-N and help alleviate
the risk of disease in these systems. This water may be
an alternative to commercial probiotic supplementation
and an alternative to individual culture treatments. The
solid control methods used in this study behaved
similarly unlike in other studies and this may be an
effect of pre-conditioned culture water. Flow
cytometry served as a cost-efficient high-throughput
method for monitoring general community structure
of microorganisms within a RAS. Due to the inherent
limitations of flow cytometry for community biology
we would recommend that future studies be performed
in parallel with molecular techniques for a more
comprehensive higher resolution investigation of the
changes in bacterial communities.
4 Materials and Methods
4.1 Experimental design
A 108
-
d study was conducted in four 40 m
3
(25.4 ×
2.7 m with 68.5 m
2
bottom area) raceways lined with
EPDM (ethylene propylene diene monomer, Firestone
Specialty Products, Indianapolis, IN, USA). All
raceways were constructed with a longitudinal
partition centered over a PVC pipe (5.1 cm long)
containing spray nozzles. Six 0.91 m long air diffusers
(1.9 cm OD, Aero-Tube
TM
, Tekni-plex Aeration,
Austin, TX, USA) and six banks, each constructed
with three 5.1 cm airlift pumps, were positioned at
equal distances on both sides of the partition on each
of the raceways used in this study. A 2 hp centrifugal
pump and a 5.1 cm Venturi injector (model 2081
-
A,
Mazzei Injector Corporation, Bakersfield, CA, USA)
served to maintain dissolved oxygen concentrations
consisting of atmospheric air or a mixture of 100%
bottled oxygen and air. Water for this experiment was
obtained from a preceding 62-d nursery study. Four
raceways were used in which two were equipped with
a commercial foam fractionator (VL65, Aquatic Eco
System, Apopka, FL, USA) and the remaining two
furnished with a 8.6 m
3
cylindroconical settling tank
(4.5 m
3
working water volume). All four raceways
were stocked (450 shrimp/m
3
) with juveniles (0.99 g ±
0.17 g) of the Pacific White Shrimp,
Litopenaeus
vannamei
, cultured at high density for 62-d in the
same tanks. Shrimp were fed seven days a week
specially formulated commercial 35% crude protein
(CP) feed for intensive systems operated with limited
discharge (Hyper-Intensive 35, Zeigler Bros., Gardners,
PA, USA). Until Day 18, feed was offered at four
equal portions during the day. From Day 19 onward,
two-thirds ration partitioned into four equal portions
were fed during the day with the remainder fed
throughout the night using belt feeders (spring loaded,
12
-
hour clock 4.5 kg capacity, Zeigler Bros.,
Gardners, PA, USA). After stocking our tanks in the
first week, shrimp feed was mixed with food using in
the nursery trial (Fry #4, 30% CP, Rangen Inc., Buhl,
ID). Each day rations were calibrated on a FCR of
1:1.4, approximating growth of 1.4 g/wk and
assuming a mortality rate of
≤
0.5%/wk. Settling
tanks (ST) and foam fractionators (FF) were used 23 days
after stocking. Both the ST and FF were used
intermittently targeting concentrations between 400 to
500 mg/L of culture water total suspended solids (TSS)
and settleable solids (SS) between 10~14 mL/L. Water
flow into the ST varied between 2 and 6 L/min to
provide between 4.6 and 13.9 tank turnovers/day. Both
the FF and the ST were operated via a side loop,
which received the water from the pump installed in
each RW. Raceways were maintained with zero water
exchange throughout the study. Municipal chlorinated
freshwater was added to compensate for water loss
due to evaporation and operation of the FF.
4.2 Multiparameter monitoring and statistical analyses
Water temperature, salinity, dissolved oxygen, and
pH was simultaneously monitored twice daily using
an YSI 650 Series multi-probe (YSI Inc., Yellow
Springs, OH, USA). Alkalinity (Method # 2320 B,
APHA, 1995), and SS (Imhoff Cone Method # 2540
F, APHA, 1995) were monitored every two to three
days. Turbidity (Spectronic 21, Milton Roy Co,
Ivyland, PA), TSS (Method # 2540 D, APHA, 1995),
volatile suspended solids (VSS) (Method # 2540 E,
APHA, 1995), and five-day carbonaceous biochemical
oxygen demand (cBOD
5
) (Method #5210 B, APHA,
1995) were monitored weekly. A Autoanalyzer (dual
channel FIAlab-2600, FIAlab Instruments, Inc.,
Bellevue, WA, USA) was used to monitor weekly
total ammonia-nitrogen (TAN) (salicylate method),
nitrite-nitrogen (NO
2
-N), nitrate-nitrogen (NO
3
-N)
(nitrate/nitrite method), and phosphate (water based
samples, orthophosphate method). Sodium bicarbonate