This article was downloaded by: [Anselmo MIranda]

On: 04 January 2015, At: 15:20

Publisher: Taylor & Francis
Informa Ltd Registered in England and Wales Registered Number: 1072954 Registered office: Mortimer House,
37-41 Mortimer Street, London W1T 3JH, UK

North American Journal of Aquaculture

Publication details, including instructions for authors and subscription information:
http://www.tandfonline.com/loi/unaj20

Proximate Composition of Bioflocs in Culture Systems

Containing Hybrid Red Tilapia Fed Diets with Varying
Levels of Vegetable Meal Inclusion
a

Jos Antonio Lpez-Elas , Anglica Moreno-Arias , Anselmo Miranda-Baeza , Luis Rafael

a

Martnez-Crdova , Martha Elisa Rivas-Vega & Enrique Mrquez-Ros

a

Departamento de Investigaciones Cientficas y Tecnolgicas, Universidad de Sonora,

Boulevard Colosio s/n, Edificio 7J, Hermosillo, Sonora 83000, Mexico
b

Laboratorio de Tecnologas de Cultivo de Organismos Acuticos, Universidad Estatal de

Sonora, Carretera Huatabampo Km 5, Navojoa, Sonora 85800, Mexico
c

Click for updates

Departamento de Investigacin y Posgrado en Alimentos, Universidad de Sonora, Boulevard

Colosio s/n, Edificio 7F, Hermosillo, Sonora 83000, Mexico
Published online: 31 Dec 2015.

To cite this article: Jos Antonio Lpez-Elas, Anglica Moreno-Arias, Anselmo Miranda-Baeza, Luis Rafael Martnez-Crdova,
Martha Elisa Rivas-Vega & Enrique Mrquez-Ros (2015) Proximate Composition of Bioflocs in Culture Systems Containing
Hybrid Red Tilapia Fed Diets with Varying Levels of Vegetable Meal Inclusion, North American Journal of Aquaculture, 77:1,
102-109, DOI: 10.1080/15222055.2014.963767
To link to this article: http://dx.doi.org/10.1080/15222055.2014.963767

PLEASE SCROLL DOWN FOR ARTICLE

Taylor & Francis makes every effort to ensure the accuracy of all the information (the Content) contained
in the publications on our platform. However, Taylor & Francis, our agents, and our licensors make no
representations or warranties whatsoever as to the accuracy, completeness, or suitability for any purpose of the
Content. Any opinions and views expressed in this publication are the opinions and views of the authors, and
are not the views of or endorsed by Taylor & Francis. The accuracy of the Content should not be relied upon and
should be independently verified with primary sources of information. Taylor and Francis shall not be liable for
any losses, actions, claims, proceedings, demands, costs, expenses, damages, and other liabilities whatsoever
or howsoever caused arising directly or indirectly in connection with, in relation to or arising out of the use of
the Content.
This article may be used for research, teaching, and private study purposes. Any substantial or systematic
reproduction, redistribution, reselling, loan, sub-licensing, systematic supply, or distribution in any
form to anyone is expressly forbidden. Terms & Conditions of access and use can be found at http://
www.tandfonline.com/page/terms-and-conditions

North American Journal of Aquaculture 77:102109, 2015


C American Fisheries Society 2015
ISSN: 1522-2055 print / 1548-8454 online
DOI: 10.1080/15222055.2014.963767

ARTICLE

Proximate Composition of Bioflocs in Culture Systems

Containing Hybrid Red Tilapia Fed Diets with Varying Levels
of Vegetable Meal Inclusion
Jose Antonio Lopez-Elas and Angelica Moreno-Arias
Departamento de Investigaciones Cientficas y Tecnologicas, Universidad de Sonora, Boulevard Colosio
s/n, Edificio 7J, Hermosillo, Sonora 83000, Mexico

Downloaded by [Anselmo MIranda] at 15:20 04 January 2015

Anselmo Miranda-Baeza*
Laboratorio de Tecnologas de Cultivo de Organismos Acuaticos, Universidad Estatal de Sonora,
Carretera Huatabampo Km 5, Navojoa, Sonora 85800, Mexico

Luis Rafael Martnez-Cordova

Departamento de Investigaciones Cientficas y Tecnologicas, Universidad de Sonora, Boulevard Colosio
s/n, Edificio 7J, Hermosillo, Sonora 83000, Mexico

Martha Elisa Rivas-Vega

Laboratorio de Tecnologas de Cultivo de Organismos Acuaticos, Universidad Estatal de Sonora,
Carretera Huatabampo Km 5, Navojoa, Sonora 85800, Mexico

Enrique Marquez-Ros
Departamento de Investigacion y Posgrado en Alimentos, Universidad de Sonora,
Boulevard Colosio s/n, Edificio 7F, Hermosillo, Sonora 83000, Mexico

Abstract
Biofloc culture systems, which are based on the development of microorganisms that recycle inorganic nutrients and
organic matter, may contribute to the nutrition of some farmed species. Juvenile red tilapia (Nile Tilapia Oreochromis
niloticus Mozambique Tilapia O. mossambicus) cultured in saltwater were fed pelleted diets in which 0, 33, 67, or
100% of the fish meal was substituted with a vegetable meal mix (corn, wheat, and sorghum meals). The proximate
composition of the biofloc produced in the culture systems was evaluated. Four experimental diets and one control
diet (isocaloric and isoproteic) were randomly assigned to 15 experimental tanks. Samples of biofloc were periodically
collected to measure the total suspended solids, organic matter, and ash content and to determine the protein, lipid,
and carbohydrate contents. At the end of the study, variables describing red tilapia production were determined. The
biofloc volume, total suspended solids, ash, and organic matter showed significant differences among treatments, but
carbohydrate (33.039.0%), lipid (2.63.5%), and protein (23.725.4%) levels were similar. No significant differences
were observed in red tilapia survival, final biomass, or feed conversion ratio. We conclude that the substitution of fish
meal with vegetable meal in the pelleted feed had no adverse effect on the production response of saltwater-cultured
red tilapia.

*Corresponding author: anselmo.miranda@ues.mx

Received June 25, 2014; accepted September 5, 2014

102

Downloaded by [Anselmo MIranda] at 15:20 04 January 2015

DIET EFFECT ON BIOFLOC PROXIMATE COMPOSITION

Biofloc is a term used to designate the aggregation of

particles in a colloidal dispersion. These aggregates contain
heterotrophic and nitrifying bacteria, microalgae, fungi, and
detritus as well as flagellates, ciliates, rotifers, and nematodes
(Buford et al. 2004). Tilapias can be successfully farmed at
high concentrations of suspended solids; thus, tilapia species
are good candidates for culture in biofloc systems (Emerenciano
et al. 2013) since the suspended organic matter, detritus, and unconsumed food can provide an important nutritional source. The
retention of residues in a culture system and their conversion to
biofloc can be achieved by constant aeration and water column
agitation to keep nutrients available (Crab et al. 2012); it is also
necessary to add carbon sources as a substrate for bacteria that
consume organic matter (Azim et al. 2008).
The microbiota associated with bioflocs is able to remove
toxic compounds and recycle nutrients (Audelo-Naranjo et al.
2012). Bioflocs are also a source of live feed for farmed organisms, allowing for the partial substitution of formulated feeds
(Tacon et al. 2002; De Schryver et al. 2008) while also reducing
production costs and pollution. The proximate composition of
the biofloc may vary according to several factors, such as culture
density, aeration intensity, dissolved oxygen, carbon source and
availability, temperature, salinity, and pH as well as the type of
microbial community that has developed (Avnimelech 2007; De
Schryver et al. 2008; Martnez-Cordova et al. 2014).
Most of the previous biofloc experiments have focused on (1)
calculating the effects of bioflocs on water quality characteristics
(e.g., pH, dissolved oxygen, and total ammonia nitrogen [TAN])
and (2) the production response of cultured organisms (e.g.,
weight gain, feed conversion ratio [FCR], total biomass, and
survival). However, there is little available information about
the biochemical composition of biofloc and how it varies in
relation to the type of diet provided or the age of the culture.
In recent years, the increased demand for and scarcity of
fish meal have caused increases in the price of formulated feed,
thereby affecting the economic feasibility of aquaculture and
necessitating studies for the development and evaluation of alternative diets. Some studies have suggested biofloc technology (BFT) as an alternative to complement the nutritional requirements of shrimps (pink shrimp Farfantepenaeus paulensis,
pinkspot shrimp F. brasiliensis, and Pacific white shrimp Litopenaeus vannamei) and Nile Tilapia Oreochromis niloticus (Azim
and Little 2008; Kuhn and Boardman 2008; Ballester et al. 2010;
Emerenciano et al. 2011). The benefits of biofloc-based cultures
are related to the better use of nutrients, diminished production
of wastewater, reduction in farm surface area, decreases in environmental impacts, and reduction in dependence on fish meal
(Moreira de Souza et al. 2013).
Azim and Little (2008) studied the quality of biofloc that developed in a culture system wherein two formulated feeds with
different protein content (24% and 35%) were administered to
Nile Tilapia; their results indicated that lipid and protein composition and energy content did not show significant differences
among treatments. Similar results were reported by Megahed

103

(2010) in comparing the quality of biofloc developed during the

culture of green tiger prawns Penaeus semisulcatus that were fed
diets with different levels of crude protein (31.15, 21.6, 18.45,
and 16.25%); the protein (19.9%) and lipid (11.8%) content of
the biofloc did not differ among treatments.
The feasibility of culturing tilapia in saltwater represents
an opportunity for farming these fishes in enclosure facilities
using seawater (Miranda-Baeza et al. 2010). Advancement in
the knowledge of tilapia nutrition, including the use of BFT,
may contribute to the sustainability of tilapia culture.
Based on the above information, we sought to evaluate two
hypotheses: (1) in a BFT system, the unconsumed food, remains of dead organisms, and metabolites can be used by
microorganisms to produce biomass of high nutritional quality; and (2) the origin of the protein (animal or vegetable) in
the pelleted feed does not significantly affect biofloc composition. We focused on evaluating the variation in biofloc proximate composition as a function of the fish meal substitution
level in the diet and the age of the tilapia culture in a BFT
system.

METHODS
Experimental culture.Juvenile red tilapia (Nile Tilapia
Mozambique Tilapia O. mossambicus hybrids; mean individual
weight SD = 16.2 1.0 g) were stocked in fifteen 200-L, indoor tanks at a density of 2.0 kg/m3 (1215 fish/tank depending
on the individual weight). The experimental fish were maintained under constant aeration supplied by an electric blower
(0.333 hp).
One control diet and four experimental diets (isocaloric, isolipidic, and isoproteic) were evaluated in triplicate over a 42-d
period. In the formulation and preparation of the experimental
diets, fish meal was included at four different levels: 0% (T0),
10% (T10), 20% (T20), and 30% (T30). To obtain the desired
protein level (35%), a mixture of vegetable meals (corn, wheat,
and sorghum) was included at different proportions for each
diet (100, 67, 33, or 0% of the fish meal was substituted, respectively; Table 1). The control diet consisted of a commercial feed
formulated for tilapia, and the crude protein content and crude
fat content were similar to those of the experimental diets (35%
and 8%, respectively). In total, 1,200 g of feed were administered to each tank, with a feeding rate equal to 3% of the tilapia
biomass.
Fifteen days prior to the stocking of red tilapia, the indoor
culture tanks (0.6 m wide, 1.00 m deep) were filled with 200 L
(0.71-m depth) of filtered seawater (35) and then were inoculated with mature, microbe-dominated biofloc at 1%. The protocol for biofloc formation and maturation consisted of adding organic matter (commercial pulverized pellet at 0.025 gL1d1;
500-m diameter, 35% crude protein) and a carbon source
(unrefined granulated sugar) to maintain a C:N ratio of 20:1.
Temperature was not controlled; rather, it was completely dependent on the local temperature variation (2428 C during the


IAS ET AL.
LOPEZ-EL

104

TABLE 1. Formulation and composition of experimental diets fed to red

tilapia that were cultured in saltwater systems with biofloc (T0 = 0% fish meal;
T10 = 10% fish meal; T20 = 20% fish meal; T30 = 30% fish meal).

Diet

Downloaded by [Anselmo MIranda] at 15:20 04 January 2015

Ingredient or
component
Ingredients
(g/kg of diet)
Soybean
concentrate
Fish meal
Cornmeal
Wheat meal
Sorghum meal
Fish oil
Soybean lecithin
Vitamin premix
Mineral premix
Sodium phosphate
Choline chloride
Vitamin C
Butylated
hydroxytoluene
Proximate
composition (%
dry weight basis)
Crude protein
Crude fat
Crude fiber
Ash

T30

T20

T10

T0

200.0

373.9

549.8

725.0

300.0
376.1
50.0
10.3
32.5
10.0
8.0
5.0
5.0
2.0
1.0
0.1

200.0
150.0
150.0
51.5
43.5
10.0
8.0
5.0
5.0
2.0
1.0
0.1

100.0
114.5
150.0
10.0
44.6
10.0
8.0
5.0
5.0
2.0
1.0
0.1

0.0
150.0
50.0
5.0
38.9
10.0
8.0
5.0
5.0
2.0
1.0
0.1

35
7
3.128
2.31

35
8
3.526
2.497

35
8
4.157
2.768

35
7.196
4.996
3.16

experiment). There was no water exchange during the experiment. Freshwater was added only to compensate for losses from
evaporation. Oxygen was maintained at high levels (46 mg/L).
Dissolved oxygen and temperature were measured twice per
day by using a multiparameter YSI 550A meter. The pH was
measured daily with a Denver Instruments UP-10 pH meter, and
salinity was determined every 3 d with a refractometer (Aquatic
Eco-Systems).
During the first 10 d of culture, the carbon : nitrogen (C:N)
ratio was maintained at approximately 20:1 by estimating the
carbon and nitrogen added in the feed and sugar. For the remainder of the trial, the C:N ratio was reduced to 12:1.
Total ammonia nitrogen and nitrite-nitrogen measurement.
Samples (250 mL) of culture water were collected every 2 d and
filtered with glass-microfiber filters (Whatman GF/C, 47-mm
diameter). Total ammonia nitrogen was then determined by the
salicylate method (Hach 8155) and nitrite-nitrogen (NO2 -N)
was determined by the diazotization method (Hach 8507) in
accordance with the manufacturers protocols (Hach Co. 2005).

Biofloc volume estimation.Samples of culture water were

placed in 1-L Imhoff cones and were left for 45 min to allow
sedimentation. Suspended solids on the walls were removed
carefully with a glass stirrer, and the samples were allowed to
settle for an additional 15 min. Biofloc volume (mL/L) was then
recorded.
Biofloc biomass evaluation.Samples (500 mL) of culture
water were taken from each tank on days 0, 7, 21, 35, and 42 of
the experiment; the samples were stored in plastic bottles and
were kept frozen at 80 C until analysis. Water samples were
filtered through glass microfiber filters (Whatman GF/C), and
the filters were dried in a convection oven (70 C for 72 h) to
calculate the total suspended solids (TSS) content. The biofloc
biomass was calculated based on the difference in filter weight
before and after filtration and drying. The ash content was evaluated by incinerating the sample in a muffle furnace at 450 C
for 4 h. The organic matter in the biofloc was estimated by calculating the difference in weight between the TSS and the ash
content.
Biofloc proximate composition.To evaluate the total protein content of the biofloc, the filtered samples were treated
with a sodium hydroxide solution (NaOH, 0.1 N) for extraction of protein according to Lowrys method (Lowry et al.
1951, as modified by Lopez-Elas et al. 1995). Total lipid content of the biofloc was determined by extraction with a mixture of chloroformmethanolwater (5:2.5:1) following Pandes
method (Pande et al. 1963, as modified by Lopez-Elas et al.
1995). Total carbohydrate was evaluated by calculating the difference between the weight of organic matter and the sum of
total protein and lipid.
Production response of red tilapia.The production response of red tilapia in the treatments was evaluated based on
biometric indices (individual weight and biomass), survival, and
FCR.
Statistical analysis.Biofloc development was depicted
graphically with mean volume, total particulate matter, and
proximate composition. Repeated-measures ANOVA was used
to compare biofloc volume, total particulate matter, organic matter, and ash as well as water quality variables among treatments
for each sampling day. A chi-square test of independence was
used to verify whether biofloc composition (carbohydrate, lipid,
protein, and ash percentages) significantly differed among treatments (Sokal and Rohlf 1981). Survival (arcsine transformed),
mean final weight, final biomass, and FCR of red tilapia were
analyzed using one-way ANOVA. Tukeys honestly significant
difference test (Zar 1996) was used to compare and rank treatment means. Statistical analyses were performed in Statistica
version 7.0 for Windows (Statsoft, Inc.).
RESULTS
Water Quality Variables
The pH averaged between 7.68 and 7.72 at the beginning of
the experiment and averaged 7.657.75 at the end of the study,

105

DIET EFFECT ON BIOFLOC PROXIMATE COMPOSITION

TABLE 2. Global mean ( SD) volume, total suspended solids (TSS), ash content, and organic matter content of biofloc in treatments with different levels of
fish meal inclusion in pelleted diets for red tilapia (C = control with 30% fish meal; see Table 1 for definition of experimental diets T0T30). Within a row, means
with different letters are significantly different (P < 0.05).

Variable

Downloaded by [Anselmo MIranda] at 15:20 04 January 2015

Volume (mL/L)
TSS (mg/L)
Ash (mg/L)
Organic matter (mg/L)

T0
9.22
537.3
213.0
324.2

2.81 x
108.1 y
50.1 y
63.0 y

T10
8.67
351.3
116.9
234.4

3.74 yx
57.7 z
31.1 z
38.3 z

without significant differences among treatments (P > 0.05).

Total ammonia nitrogen concentration was low in all treatments,
and there were no significant differences (P > 0.05). A slightly
higher mean TAN of 0.63 mg/L was recorded for the control,
while T30 exhibited a lower value of 0.40 mg/L. The NO2 N concentration began to increase on day 6, and maximum
levels of 34 mg/L were observed on days 912. During the
entire experiment, the mean NO2 -N values ranged from 0.82
to 0.85 mg/L and did not significantly differ among treatments
(P > 0.05).
Biofloc Volume
Mean biofloc volume showed significant differences among
treatments (P < 0.05); mean volume was 9.22 mL/L for T0,
8.67 mL/L for T10, 6.78 mL/L for T20, and 4.22 mL/L for
T30, whereas the mean for the control was 5.56 mL/L (Table 2).
From the beginning to the end of the study, the highest biofloc
volumes (mL/L) were recorded for treatments with a lower fish
meal content. In general, the biofloc volume increased with

T20
6.78
402.3
156.1
246.1

5.26 zy
151.2 z
102.17 zy
59.6 z

T30
4.22
404.0
167.0
237.0

2.63 z
120.4 z
63.9 zy
59.6 z

C
5.56
354.6
132.3
222.3

2.18 zy
88.9 z
60.6 z
36.4 z

the age of the culture, but the rate of increase differed among
treatments (Figure 1). On day 42 (the end of the culture period),
the highest biofloc volume (19 mL/L) was observed for T0,
while the lowest volume was observed for T30 (6.5 mL/L).
Biofloc Biomass
Mean TSS in the biofloc ranged from 351.3 mg/L in T10 to
537.3 mg/L in T0 (Table 2). The TSS concentration for T0 was
significantly higher than values observed for the other treatments
(P < 0.05). For T10, T20, T30, and the control, TSS values were
stable through time (Figure 2A).
On day 0 (i.e., 1 d before the fish were stocked), lower
TSS concentrations within the range of 200300 mg/L were
recorded, and there were no significant differences among treatments. On day 7 (the second sampling day), an increase of
approximately 100 mg/L was observed in all treatments. From
day 21 to day 42, T0 consistently presented the highest TSS
concentrations (Figure 2A).

FIGURE 1. Mean ( + SD) biofloc volume (mL/L) obtained in the treatments with different levels of fish meal inclusion in pelleted diets for red tilapia (C =
control with 30% fish meal; experimental diets: 0 = 0% fish meal; 10 = 10% fish meal; 20 = 20% fish meal; 30 = 30% fish meal).

Downloaded by [Anselmo MIranda] at 15:20 04 January 2015

106

IAS ET AL.
LOPEZ-EL

FIGURE 2. Changes in mean ( SD) values of (A) total suspended solids,

(B) ash content, and (C) organic matter content of the biofloc that developed
in red tilapia culture with different levels of fish meal inclusion in the pelleted
diets (see Figure 1 for definition of treatments).

FIGURE 3. Changes in mean ( SD) values of (A) total protein, (B) total
carbohydrate, and (C) total lipid of the biofloc developed in red tilapia culture
with different levels of fish meal inclusion in the pelleted diets (see Figure 1 for
definition of treatments).

Ash content in the biofloc of T10, T20, T30, and the control was maintained at stable levels throughout the experiment; no significant changes related to culture age were observed (Figure 2B). The global mean ash content of the biofloc
ranged from 116.9 to 213.0 mg/L, with significant differences
(Table 2).
Mean organic matter content in the biofloc varied from 222.3
to 324.2 mg/L, and significant differences among treatments
were identified (P < 0.05; Table 2). From day 7 to the end of
the experiment, organic matter showed low variation in T10,
T20, T30, and the control, whereas it increased over the culture
period in T0 (Figure 2C).

Biofloc Proximate Composition

The protein concentration in the biofloc ranged from 46 to
70 mg/L on day 0 and increased until day 21, when the highest
means were recorded; a general decrease was observed thereafter (Figure 3A). At the end of the culture period (day 42),
T0 had the highest protein concentration (132 mg/L). Considering the whole experiment, the mean protein content in the
biofloc varied from 23.7% to 25.4% and showed no significant
differences among treatments (P > 0.05; Table 3).
On day 0, the carbohydrate concentration in the biofloc
was similar among treatments, with means ranging from 70
to 80 mg/L. Carbohydrate values reached 120150 mg/L on day

107

DIET EFFECT ON BIOFLOC PROXIMATE COMPOSITION

TABLE 3. Proximate composition (global mean SD) of the biofloc (dry basis) developed with different levels of fish meal inclusion in pelleted diets for red
tilapia (C = control with 30% fish meal; see Table 1 for definition of experimental diets T0T30).

Variable

Downloaded by [Anselmo MIranda] at 15:20 04 January 2015

Ash (%)
Carbohydrate (%)
Protein (%)
Lipid (%)

T0
39.3
33.0
24.5
2.6

T10
3.6
3.9
2.3
0.3

33.0
39.0
24.1
3.5

T20
5.4
3.7
5.3
0.6

7 and increased continuously over the remainder of the culture

period (Figure 3B). Considering the entire experimental period, the carbohydrate content of the biofloc varied from 32.2%
to 39.0%, without significant differences among treatments
(P > 0.05; Table 3).
The lipid content did not show any clear trend during the
culture period (Figure 3C). Mean lipid content ranged from 2.6%
to 3.5% and did not significantly differ among the treatments
(P > 0.05; Table 3).
Production Response of Red Tilapia
The survival of red tilapia did not present significant amongtreatment differences (P > 0.05); nevertheless, survival tended
to be greater when fish meal inclusion in the pelleted diet was
lower (Table 4). Final individual weight was significantly different among treatments (P < 0.05): the lowest weight (45.0 g) was
observed for T0, while the highest weight (64.1 g) was recorded
for T20. Final fish biomass varied from 5.8 to 7.1 kg/m3, and
FCR ranged from 1.2 to 1.7; in both cases, no significant differences were observed among the treatments (P > 0.05; Table 4).
DISCUSSION
Ammonia is the main metabolic waste of fish and is found in
two different forms in the water column: un-ionized ammonia
(NH3 ) is the form most toxic to fish, whereas ionized ammonia
(NH4 + ) is less toxic (El-Sayed 2006). The form of ammonia
present is closely correlated with pH; at high pH, un-ionized
ammonia is dominant. Timmons et al. (2002) indicated that TAN
levels less than 3.0 mg/L are safe for warmwater fish farming
if the pH is between 6.5 and 8.5. In our study, pH ranged from
7.53 to 7.92, and under these conditions the nontoxic form of

35.8
35.5
23.7
2.8

T30
10.1
4.7
5.5
0.7

40.4
32.2
25.4
3.2

C
4.9
8.9
9.3
0.8

35.9
36.5
23.9
2.9

8.3
5.2
5.3
0.5

ammonia was dominant; additionally, the TAN concentration

was maintained under 0.7 mg/L due to the prior maturation of the
biofloc. The C:N ratios used in our experiment (20:1 and 12:1)
contributed to maintaining a high concentration of heterotrophic
bacteria, which have been demonstrated to remove ammonia and
organic matter from the water column (Avnimelech 2007).
Nitrite-nitrogen is an intermediate compound in the nitrification process; the increase in NO2 -N during the first days was
a normal process attributable to the low rate at which nitriteoxidizing bacteria (Timmons et al. 2002) increased in the biofloc
aggregates. The peak in NO2 -N and the subsequent decrease to
low concentrations indicated the occurrence of an efficient nitrification process in the biofloc culture, which has been reported
previously by several authors (Hari et al. 2006; Avnimelech
2009; Emerenciano et al. 2012).
As the amount of vegetable meal in the diet increased, a
higher biofloc volume was observed; this effect could have been
induced by the lower digestibility of vegetable meal, which creates a higher amount of flocculated material (mainly fiber) that
can be used as substrate by the bacteria and other microorganisms. Similarly, Soltan et al. (2008) found that when fish meal
substitution by a mix of vegetable meals in a tilapia diet exceeded 45%, the apparent digestibility decreased from 81% to
73%. In the present study, the increase in biofloc volume from
the beginning to the end of the trial was associated with the fish
meal content in the diet, since T0 showed the highest increase
(from an initial volume of 3 mL/L to a final volume of 19 mL/L),
while T30 recorded the lowest increase (from an initial volume
of 1.3 mL/L to a final volume of 5.6 mL/L).
The biofloc biomass recorded in this study accords with
values reported by Scopel et al. (2011), who found that the
TSS level in a Pacific white shrimp culture was greater when

TABLE 4. Mean ( SD) survival, final individual weight, initial and final biomass, and feed conversion ratio (FCR) of cultured red tilapia that received diets
with different levels of fish meal inclusion (C = control with 30% fish meal; see Table 1 for definition of experimental diets T0T30). Within a row, means with
different letters are significantly different (P < 0.05).

Variable
Survival (%)
Final individual weight (g)
Initial biomass (kg/m3)
Final biomass (kg/m3)
FCR

T0

T10

T20

T30

100 0
45.0 1.5 z
2.0
6.0 0.7
1.5 0.3

100 0
53.5 5.6 zy
2.0
7.1 0.1
1.2 0.0

91 16
64.1 1.9 y
2.0
6.8 1.4
1.4 0.5

83 10
57.4 18.7 zy
2.0
5.8 1.0
1.7 0.5

90 16
53.1 5.1 zy
2.0
6.3 0.5
1.5 0.2

Downloaded by [Anselmo MIranda] at 15:20 04 January 2015

108

IAS ET AL.
LOPEZ-EL

vegetable meal diets were administered than when fish meal diets were used. Scopel et al. (2011) suggested that the result was
due to the lower digestibility of the vegetable ingredients.
In a study of Nile Tilapia cultures, Azim and Little (2008)
compared experimental diets containing 35% and 24% protein
and observed TSS values of 597 and 560 mg/L, respectively,
similar to TSS levels found in the present study. Avnimelech
(2007) reported mean TSS values of 460643 mg/L, close to
values we found for T0. In contrast, Ray et al. (2010) observed
no difference in TSS production when Pacific white shrimp were
given fish meal diets versus vegetable meal diets.
Increases in TSS commonly affect the respiration of cultured organisms (gill obstruction) and dissolved oxygen diffusion into the water column (Crab et al. 2012; Moreira de Souza
et al. 2013). Nevertheless, the TSS values reported here (400
500 mg/L) are within the recommended range (200500 mg/L)
for a good productive response in biofloc cultures (Avnimelech
2009).
The mean ash values of the biofloc in all treatments (33
40%) were similar to previous findings (39.241.1%: Ballester
et al. 2010; 39.8%: Becerra-Dorame et al. 2012). Maica et al.
(2011) reported that the ash content in biofloc tended to increase
with increasing salinity: mean ash content was 22.12, 26.73,
and 42.19% at salinities of 2, 4, and 25, respectively. In the
present study, salinity was maintained at a constant level of
35, similar to the experiments performed by Ballester et al.
(2010) and Becerra-Dorame et al. (2012).
The organic matter content of biofloc ranged between 59%
and 66% and therefore was very close to the values of 5779%
reported by Maica et al. (2011).
With regard to the proximate composition of the biofloc, the
total protein content we found was similar to values reported by
Azim and Little (2008; 2831%), Ekasari et al. (2010; 30.4%),
Becerra-Dorame et al. (2012; 11.517.5%), and Emerenciano
et al. (2012; 37.938.4%). The results we recorded for carbohydrate content were close to previously published values of
35.4% (Becerra-Dorame et al. 2012) and 29.134.9% (Emerenciano et al. 2012). At the end of the culture period (i.e., day 42),
we observed that the protein content was decreasing, whereas
the carbohydrate content was still increasing; this result could
be due to fiber accumulation in the culture tanks. Azim and
Little (2008), Ekasari et al. (2010), and Becerra-Dorame et al.
(2012) reported that the mean lipid content in the biofloc was
3.163.2%, 69%, and 6.5%, respectively, similar to the lipid
content values observed during the present trial.
The production response of red tilapia was independent of
dietary treatment. Survival rate, final biomass, and FCR were
not affected by substituting fish meal with vegetable meal in
the diet. These results concur with those reported by Scopel
et al. (2011) after substituting fish meal with vegetable meal in
experimental diets for Pacific white shrimp cultured in biofloc
systems; those authors found no significant difference in mean
final biomass, FCR, or survival among the various diet treatment
groups. Similar results were reported by Azim and Little (2008),

who found no significant difference in mean survival, weight

gain, or FCR for Nile Tilapia that were fed diets with two
protein contents during culture in biofloc systems.
In our study, the TSS, ash content, and volume of the biofloc
were greater in diet treatments with a lower fish meal inclusion,
but we observed no negative effect on the final biomass or FCR
of red tilapia. In BFT systems, the TSS concentration is affected
by fish consumption; tilapia are able to filter particulate matter,
and in the absence of formulated feed they could use the biofloc
as a unique nutritional source. Avnimelech (2007) reported a
decrease in biofloc volume (from 40 mL/L to 20 mL/L) during
the first 6 d of Nile Tilapia culture when the fish were deprived
of exogenous feed.
In biofloc cultures, the microbial communities improve the
water quality and recycle the organic matter (Buford et al. 2004);
bacteria are consumed by other microorganisms and generate
high concentrations of live biomass. More specific studies have
revealed that biofloccules used as feed in aquaculture have considerable amounts of essential amino acids and essential fatty
acids, and the vitamin profiles are usually adequate for fish and
crustaceans (Martnez-Cordova et al. 2014). In BFT systems, the
cultured organisms have a high-quality food source that is permanently available (Browdy et al. 2001; Samocha et al. 2007)
and complements their nutrition. Recent evidence indicates that
microbial flocs and biofilms act as probiotics (Becerra-Dorame
et al. 2012; Martnez-Cordova et al. 2014); therefore, the nutritional condition of farmed organisms is a consequence not
only of the feed consumed but also of the presence of bacteria
and their exogenous enzymes, which contribute to the digestive physiology of the cultured species (Ziaei-Nejad et al. 2006;
Zhou et al. 2009). In the present study, the bacterial community
of the biofloc may have improved the assimilation of vegetable
meal that was included in the experimental diets.
Based on the proximate composition of biofloc developed
using a pelleted diet with a low fish meal content, we conclude
that the biofloc is a good, nutritious source of protein, lipid,
and carbohydrate for cultured tilapias. These findings suggest
that tilapia producers who use biofloc systems can employ diets
with a low fish meal content to reduce production costs as well
as promote an environmentally friendly method of aquaculture.
Future research on BFT should include evaluations of lower
fish meal inclusion in the diets, the amino acid and fatty acid
composition of the biofloc, and the effects of biofloc-associated
microorganisms on the digestive activity of tilapias.

ACKNOWLEDGMENTS
This study was partially financed by the Sonora State
University project entitled Evaluation of vegetable ingredients to grow tilapia in seawater with biofloc (Project C-PII
/11/26ESU0057P/01). Angelica Moreno-Arias was supported
by a scholarship from the Mexican National Council of Science and Technology as part of a masters degree. We thank

DIET EFFECT ON BIOFLOC PROXIMATE COMPOSITION

the University of Sonora for the support provided during this

research.

Downloaded by [Anselmo MIranda] at 15:20 04 January 2015

REFERENCES
Audelo-Naranjo, J. M., L. R. Martnez-Cordova, S. Gomez-Jimenez, and D.
Voltolina. 2012. Intensive culture of Litopenaeus vannamei without water
exchange and with an artificial substrate. Hidrobiologica 22:17.
Avnimelech, Y. 2007. Feeding with microbial flocs by tilapia in minimal discharge biofloc technology ponds. Aquaculture 264:140147.
Avnimelech, Y. 2009. Biofloc technology. World Aquaculture Society, Baton
Rouge, Louisiana.
Azim, M. E., and D. C. Little. 2008. The biofloc technology (BFT) in indoor
tanks: water quality, biofloc composition, and growth and welfare of Nile
Tilapia (Oreochromis niloticus). Aquaculture 283:2935.
Azim, M. E., D. C. Little, and J. E. Bron. 2008. Microbial protein production in
activated suspension tanks manipulating C:N ratio in feed and the implications
for fish culture. Bioresource Technology 2008:35903599.
Ballester, E. L. C., P. C. Abreu, R. O. Cavalli, M. Emerenciano, L. Abreu, and
W. Wasielesky. 2010. Effect of practical diets with different protein levels
on the performance of Farfantepenaeus paulensis juveniles nursed in a zero
exchange suspended microbial floc intensive system. Aquaculture Nutrition
16:163172.
Becerra-Dorame, M. J., M. Martnez-Porchas, L. R. Martnez-Cordova, M. E.
Rivas-Vega, J. A. Lopez-Elas, and M. A. Porchas-Cornejo. 2012. Production response and digestive enzymatic activity of the Pacific white shrimp
Litopenaeus vannamei (Boone, 1931) intensively pregrown in microbial heterotrophic and autotrophic-based systems. Scientific World Journal 2012:16.
Browdy, C. L., D. Bratvold, A. D. Stokes, and R. P. McIntosh. 2001. Perspectives
on the application of closed shrimp culture systems. Pages 2034 in C. L.
Browdy and D. E. Jory, editors. The new wave: proceedings of the special
session on sustainable shrimp culture, aquaculture 2001. World Aquaculture
Society, Baton Rouge, Louisiana.
Buford, M. A., P. J. Thompson, R. P. McIntosh, R. H. Bauman, and D. C. Pearson. 2004. The contribution of flocculated material to shrimp (Litopenaeus
vannamei) nutrition in a high-intensity, zero-exchange system. Aquaculture
232:525538.
Crab, R., T. Defoirdt, P. Bossier, and W. Verstraete. 2012. Biofloc technology
in aquaculture: beneficial effects and future challenges. Aquaculture 356
357:351356.
De Schryver, P., R. Crab, T. Defoirdt, N. Boon, and W. Verstraete. 2008. The
basics of bio-floc: the added value of aquaculture. Aquaculture 277:125
137.
Ekasari, J., R. Crab, and W. Verstraete. 2010. Primary nutritional content of bioflocs cultured with different organic carbon sources and salinity. HAYATI
Journal of Biosciences 17:125130.
El-Sayed, A. M. 2006. Tilapia culture. CAB International, Wallingford, UK.
Emerenciano, M., E. L. C. Ballester, R. O. Cavalli, and W. Wasielesky. 2011.
Effect of biofloc technology (BFT) on the early postlarval stage of pink
shrimp Farfantepenaeus paulensis: growth performance, floc composition
and salinity stress tolerance. Aquaculture International 19:891901.
Emerenciano, M., E. L. C. Ballester, R. O. Cavalli, and W. Wasielesky. 2012.
Biofloc technology application as a food source in a limited water exchange nursery system for pink shrimp Farfantepenaeus brasiliensis (Latreille, 1817). Aquaculture Research 43:447457.
Emerenciano, M., G. Gaxiola, and G. Cuzon. 2013. Biofloc technology (BFT):
a review for aquaculture application and animal food industry. Pages 301
328 in D. Matovik, editor. Biomass now: cultivation and utilization. InTech,
Rijeka, Croatia.
Hach. 2005. Procedures manual, DR5000 spectrophotometer. Hach, Loveland,
Colorado.
Hari, B., B. Madhusoodana Kurup, J. T. Varghese, J. W. Schrama, M. C. J.
Verdegem. 2006. The effect of carbohydrate addition on water quality and the
nitrogen budget in extensive shrimp culture systems. Aquaculture 252:248
263.

109

Kuhn, D. D., and G. D. Boardman. 2008. Use of microbial flocs generated from
tilapia effluent as nutritional supplement for shrimp, Litopenaeus vannamei, in
recirculating aquaculture systems. Journal of the World Aquaculture Society
39:7282.
Lopez-Elas, J. A., M. del, C. Baez-Duenas, and N. Huerta-Aldaz. 1995. Manual
de tecnicas analticas aplicadas al cultivo de microalgas. [Manual of analytical
techniques applied to microalgae culture.] Departamento de Investigaciones
Cientficas y Tecnologicas, Universidad de Sonora, Mexico.
Lowry, O. H., N. J. Rosenbrough, A. L. Farr, and R. J. Randall. 1951. Protein
measurement with the Folin phenol reagent. Journal of Biological Chemistry
193:265275.
Maica, P. F., M. R. Borba, and W. Wasielesky. 2011. Effect of low salinity
on microbial floc composition and performance of Litopenaeus vannamei
(Boone) juveniles reared in a zero-water-exchange super-intensive system.
Aquaculture Research 43:361370.
Martnez-Cordova, L. R., M. Emerenciano, A. Miranda-Baeza, and M.
Martnez-Porchas. 2014. Microbial-based systems for aquaculture of fish
and shrimp: an updated review. Reviews in Aquaculture 6:118.
Megahed, M. E. 2010. The effect of microbial biofloc on water quality, survival
and growth of the green tiger shrimp (Penaeus semisulcatus) fed with different
crude protein levels. Journal of the Arabian Aquaculture Society 5:119142.
Miranda-Baeza, A., J. Lizarraga-Armenta, M. E. Rivas-Vega, J. A. LopezElas, and M. Nieves-Soto. 2010. Integrated culture of tilapia (Oreochromis
mossambicus Oreochromis niloticus) and Pacific oyster (Crassostrea gigas) in a recirculation system. Journal of the World Aquaculture Society
41:763771.
Moreira de Souza, D., L. Godoy, W. Wasielesky, and E. L. C. Ballester. 2013.
Contribucion de los microorganismos en ambientes acuaticos y en sistemas
de cultivo. [Contribution of the microorganisms in aquatic environments
and in culture systems.] Pages 124 in L. R. Martnez-Cordova and M.
Martnez-Porchas, editors. Alimento natural en acuacultura. [Natural food in
aquaculture.] AGT Editor SA, Mexico, City.
Pande, S. V., R. P. Khan, and T. A. Venkitasubramanian. 1963. Microdetermination of lipids and serum total fatty acid. Analytical Biochemistry 6:415423.
Ray, A. J., B. L. Lewis, C. L. Browdy, and J. W. Leffler. 2010. Suspended
solids removal to improve shrimp (Litopenaeus vannamei) production and an
evaluation of a plant-based feed in minimal-exchange, super-intensive culture
systems. Aquaculture 299:8998.
Samocha, T. M., S. Patnaik, M. Speed, A. M. Ali, J. M. Burger, R. V. Almeida,
Z. Ayub, M. Harisanto, A. Horowitz, and D. L. Brock. 2007. Use of molasses
as carbon source in limited discharge nursery and grow-out systems for
Litopenaeus vannamei. Aquaculture Engineering 36:184191.
Scopel, B. R., R. Schveitzer, W. Q. Seiffert, V. Pierri, R. F. Arantes, F. N. Vieira,
and L. A. Vinatea. 2011. Substitution of fish meal in diets for marine shrimp
grown in a biofloc system. Pesquisa Agropecuaria Brasileira 46:928934.
Sokal, R. R., and F. J. Rohlf. 1981. Biometry, 2nd edition. Freeman, New York.
Soltan, M. A., M. A. Hanafy, and M. I. A. Wafa. 2008. Effect of replacing
fish meal by a mixture of different plant protein sources in Nile Tilapia
(Oreochromis niloticus L.) diets. Global Veterinaria 2:157164.
Tacon, A. G. J., J. Cody, L. Conquest, S. Divakaran, I. P. Forster, and O. Decamp.
2002. Effect of culture system on the nutrition and growth performance
of Pacific white shrimp Litopenaeus vannamei (Boone) fed different diets.
Aquaculture Nutrition 8:121137.
Timmons, M. B., J. M. Ebeling, F. W. Wheaton, S. T. Summerfelt, and B.
J. Vinci. 2002. Recirculating aquaculture systems. Cayuga Aqua Ventures,
Ithaca, New York.
Zar, J. 1996. Biostatistical analysis. Prentice Hall, Englewood Cliffs, New Jersey.
Zhou, X. X., Y. B. Wang, and W. F. Li. 2009. Effect of probiotic on larvae
shrimp (Penaeus vannamei) based on water quality, survival rate and digestive
enzyme activities. Aquaculture 287:349353.
Ziaei-Nejad, S., M. H. Rezaei, G. A. Takami, D. L. Lovett, A. R. Mirvaghefi,
and M. Shakouri. 2006. The effect of Bacillus spp. bacteria used as probiotics
on digestive enzyme activity, survival and growth in the Indian white shrimp
Fenneropenaeus indicus. Aquaculture 252:516524.