Aquaculture Reports 26 (2022) 101300

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Aquaculture Reports
journal homepage: www.elsevier.com/locate/aqrep

Microorganisms in biofloc aquaculture system

Mohammad Hossein Khanjani a, *, Alireza Mohammadi b,
Maurício Gustavo Coelho Emerenciano c
a
Department of Fisheries Sciences and Engineering, Faculty of Natural Resources, University of Jiroft, Jiroft, Kerman, Islamic Republic of Iran
b
Department of Environmental Sciences and Engineering, Faculty of Natural Resources, University of Jiroft, Jiroft, Kerman, Islamic Republic of Iran
c
Commonwealth Scientific and Industrial Research Organization (CSIRO), Livestock & Aquaculture Program, CSIRO Agriculture and Food, Bribie Island Research
Centre, Woorim, Australia

A R T I C L E I N F O A B S T R A C T

Keywords: Microorganisms (MOs) are an important member of any aquaculture system. The MOs in the BFT (Biofloc
Aquaculture Technology) system are also crucial, diverse and therefore have different roles. This study reviews the impor­
Biofloc tance of biofloc organisms (BFOs) in BFT, the factors affecting their population compositions, the impact of BFOs
Microorganism
on water quality, and applications as food source for cultivated aquatic species. Common MOs in the BFT system
Water quality
Feed
often include photoautotrophic (e.g. microalgae), chemoautotrophic (e.g. nitrifying bacteria), and heterotrophic
organisms, including fungi, ciliates, protozoans, and zooplankton (e.g. rotifers, copepods, and nematodes).
Various factors such as salinity, type of carbon source, carbon to nitrogen ratio, aeration, light, stocking density,
and total suspended solids affect the quality, density, and diversity of BFOs. Various MOs show different func­
tional characteristics and perform three main functions: (i) help to improve the water quality by removing
inorganic nitrogen compounds (bioaccumulation, bioassimilation, nitrification, and denitrification); (ii) act as a
supplementary food source, and (iii) create probiotic properties; key roles for any aquatic farming system.

1. Introduction
The aquaculture industry is one of the fastest growing sectors in the
food industry, supplying more that 50 % of the global demand for fish in
the face of dwindling natural stocks due to overexploitation (Naylor
et al., 2021; FAO, 2018). Among the innovations in the field of aqua­
culture, biofloc technology (BFT) has attracted great attention due to its
ability to foster the goals of sustainable aquaculture (Khanjani et al.,
2022a). If implemented correctly, BFT capitalizes on the existing or­
ganisms in water to significantly reduce water use and environmental
impact and increase biosafety at farms (Avnimelech, 2012). Biofloc or­
ganisms (BFOs) play three main roles in BFT: (i) maintaining water
quality by consuming nitrogenous compounds; (ii) providing a source of
food for the cultivated species (Emerenciano et al., 2012; Wang et al.,
2016), and (iii) pathogen competition and create probiotic properties
(Ferreira et al., 2015) often by heterotrophic organisms (e.g. Bacillus spp
bacteria).
BFOs help with maintaining water quality by reducing harmful
nitrogenous compounds such as ammonia and nitrite by metabolizing
fecal matter and unconsumed food (Suita et al., 2015; Wang et al.,
2016). The growth of BFOs is associated with control of TAN in the
environment (Khanjani and Sharifinia, 2020). Bioflocs (BFs) contains
organisms such as bacteria, microalgae, yeast, rotifers, ciliates, pro­
tozoans, nematodes, copepods and crustaceans (Monroy-Dosta et al.,
2013; Collazos-Lasso and Arias-Castellanos, 2015; Khanjani et al.,
2022a, 2022b). Greater diversity of BFOs can improve flocs’ minerali­
zation of waste (Martínez-Córdova et al., 2015), protein utilization
(Avnimelech, 2007), and pathogen control (Emerenciano et al., 2013a).
The exact make-up of the floc community in a BFT system depends on
several factors e.g., the type of external carbon, environmental condi­
tions such as salinity, light, oxygen levels, and solids load, the ratio of
carbon to nitrogen (C/N ratio), and the cultivated species (Ray et al.,
2010; Maicá et al., 2012; Meenakshisundaram et al., 2021; Khanjani and
Sharifinia, 2021; Krummenauer et al., 2021; Khanjani et al., 2022c). In
addition, the dynamics in BFT system are determined by the types of
ecological relationships (e.g., commensalism, competition, and preda­
tion) between BFOs in a trophic network (Collazos-Lasso and
Arias-Castellanos, 2015; Pacheco-Vega et al., 2018). Regarding types of
carbon, different sources promoted different BFOs community compo­
sition in BFT for Pelteobagrus vachelli cultivation (Deng et al., 2018), and

* Corresponding author.
E-mail addresses: m.h.khanjani@ujiroft.ac.ir, m.h.khanjani@gmail.com (M.H. Khanjani).

https://doi.org/10.1016/j.aqrep.2022.101300
Received 23 May 2022; Received in revised form 15 July 2022; Accepted 9 August 2022
Available online 16 August 2022
2352-5134/© 2022 The Authors. Published by Elsevier B.V. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).
M.H. Khanjani et al.
different C/N ratios influenced BFOs composition in L. vannamei
biofloc-based culture (Panigrahi et al., 2018).
The presence of BFOs in aquaculture, especially ciliates and nema­
todes, can make proteins, lipids, and vitamins available to the farmed
species, thus promoting growth and reducing the need for these com­
pounds in commercial feed (Focken et al., 2006; Loureiro et al., 2012;
Becerril-Cortés et al., 2017; Khanjani and Sharifinia, 2020). Moreover,
the identification of BFOs composition can help fine-tune the applica­
tions of BFT and achieve increased production and reduced environ­
mental footprint (Emerenciano et al., 2017; Kumar et al., 2021; Abakari
et al., 2022). The proper understanding of types, abundance and di­
versity of species, as well as their roles in the ecological dynamics (e.g.,
bacteria as decomposers, phytoplankton as primary producers, etc.)
have been identified as crucial for a proper system management in BFT,
proving yield consistency and predictability (Emerenciano et al.,
2022b). In this sense, the present manuscript aims to provide funda­
mental knowledge on BFOs and has been reviewed and organized with
the following topics:
2. BFT system
BFT systems are a recent development in aquaculture that eliminate
or minimize water exchange while maintaining high stocking density. In
such systems, addition of supplementary sources of carbon once C/N
ratio reaches 15:1, induces the organisms that are already present in
water to use the accumulated nitrogenous waste (in the form of un­
consumed feed or excrement), which in turn accelerated their growth
and leads to the formation of agglomerations known as flocs (Khanjani
and Sharifinia, 2022a; Khanjani et al., 2022a). These communities
contain different species which interact with other organisms in the
community and the environment by performing different functions
(Díaz and Wacher, 2003; Cienfuegos-Martínez et al., 2020). Through
BFT, such microscopic communities have been applied in farming
crustaceans and fish as feed and environmental ameliorants (Khanjani
et al., 2020b, 2021a; Emerenciano et al., 2022a). BFT systems are
capable of achieving resource optimization, endogenous nutrient recy­
cling, and food generation (in the form of biofloc). These microbial
aggregates, which might float freely in water or attach to surfaces
(Martínez-Córdova et al., 2015), colonize the culture environment or the
digestive tract of other organisms (Panigrahi et al., 2019a) and confer
several advantages in BFT systems, including improving the activity of
digestive enzymes (Khanjani and Sharifinia, 2021; Khanjani et al.,
2021c), fostering the health of the cultivated organisms and the systems
as a whole (Cardona et al., 2015; Kumar et al., 2020), and mitigating the
outbreak of diseases through competition with pathogens or reducing
their virulence (Hostins et al., 2019).
Therefore, it is vital that the BFOs community develops into the
‘ideal’ conditions and community (Martínez-Córdova et al., 2015). In
such systems, bacteria are often the foundation of the BFOs community
and play a major role in determining the health of the system. In
particular, heterotrophic forming bacteria perform an essential function
by making use of dissolved organic matter, and then being consumed by
protozoans, which are themselves consumed by organisms that occupy
higher levels in the food chain (Nevejan et al., 2018). However, other
communities such as chemoautotrophic and photoautotrophic also play
a key role in BFT (Emerenciano et al., 2017).
3. Microorganisms in BFT
Microorganisms (MOs) are an essential part of any organism life and
ecosystem (López and Zaballos, 2005). Many microbial groups are found
in aquaculture systems, and can have positive effects such as removal of
contaminants, serving as food for other organisms, and recycling of
organic matter, or negative effect such as pathogenicity under adverse
environmental conditions (Karigar and Rao, 2011; Abakari et al., 2022).
Therefore, it is crucial to identify the environmental and biological
2
Aquaculture Reports 26 (2022) 101300
conditions which influence MOs functions (Monroy-Dosta et al., 2015).
Some studies have focused on biofloc community structure and
composition in shrimp cultivation (Ju et al., 2008; Ballester et al., 2010;
Anand et al., 2014; Ekasari et al., 2014; Kumar et al., 2021), and have
placed the MOs in five overlapping categories including floc-forming
organisms, saprophytes, nitrifying bacteria, algae grazers, and patho­
genic bacteria (e.g., Vibrio spp). For a BFT system to function properly
and achieve its goals, species in all these categories must be in suspen­
sion (water column) and perform their functions and maintain their
interactions with other species (Kumar et al., 2021). Floc-forming MOs
secret extracellular polymeric substances which are sticky and can
contribute to the floc formation (Medina and Neis, 2007; Manan et al.,
2017).
Ju et al. (2008) studied the MOs community in Pacific white shrimp
(L. vannamei) cultivation and found in this particular condition that the
microbial biomass contains 24.6 % phytoplankton (mainly diatom spe­
cies such as Thalassiosira, Chaetoceros, and Navicula), 3 % bacteria, a
small fraction of protozoans (98 % flagellates, 1.5 % rotifers, and 0.5%
amoebae), 33.2 % dead material, and 39.25 % ash. Although hetero­
trophic bacteria are considered the dominant species among BFOs, other
key species are also found such as fungi, algae (dinoflagellates and di­
atoms), flagellates, rotifers, and ciliates (Ju et al., 2008; Kim et al., 2014;
Kasan et al., 2017; Tepaamorndech et al., 2020). Monroy-Dosta et al.
(2013) conducted research on a macrocosm-microcosm system associ­
ated with a tilapia culture. The authors identified seven genera of ciliates
and two were described: Vorticella and Epystilis. Moreover, rotifers from
the Philodina, Lecane, and Keratella genera were also detected (Mon­
roy-Dosta et al., 2013). Table 1 shows the MOs commonly present in BFT
systems for the cultivation of different farmed species. Also, rearing
conditions such as (study period, stocking density, cultivation tank
volume, carbon source used, carbon-nitrogen ratio, dissolved oxygen
(DO), pH, total suspended solids (TSS), light, and salinity) are provided.
Fig. 1 shows tanks used during the rearing Nile tilapia in a biofloc sys­
tem, commercial biofloc-based shrimp farming, as well as some of key
MOs species found in BFT.
3.1. Bacterial and associated communities
Bacteria play an important role in BFT system by transforming
organic material, removing toxic compounds and contaminants, as well
as functioning as food source for larger cultivated organisms (De
Schryver et al., 2008; Kumar et al., 2021). In-Kwon (2012) reports that
more than 2000 species of bacteria can grow well in BFT system. The
main groups of bacteria identified in BFT system are presented in
Table 1, in which nitrifying and heterotrophic bacteria are considered
the most important groups (Emerenciano et al., 2022a).
Heterotrophic bacteria (e.g., Bacillus species) play the main role in
BFT systems (Kumar et al., 2021). Ammonia levels, C/N ratio, DO,
temperature, alkalinity, salinity, light, and pH impact the composition of
these bacteria in BFT systems (Ebeling et al., 2006; Nootong et al., 2011;
Avnimelech, 2012). In BFT aquaculture, heterotrophic bacteria can be
stimulated by manipulating the C/N ratio (increasing C/N ratio to more
than 10), limiting water exchange, removing solids, and increasing
aeration and circulation within cultivation tanks. The growth rate of
heterotrophic bacteria is ten times greater than nitrifying bacteria (Hari
et al., 2004; Hargreaves, 2006). Although heterotrophic bacteria may
make smaller contributions to nitrification and denitrification, they are
involved in the digestion of excreta, unconsumed feed, and dead organic
matter, and sequester nitrogenous ammonia in harmless microbial ag­
gregates or masses (Ebeling et al., 2006).
BFT systems promote the proliferation of both autotrophic and het­
erotrophic MOs (Manan et al., 2017; Pacheco-Vega et al., 2018; Kumar
et al., 2021) although beneficial heterotrophic MOs are more abundant
in BFs (e.g., Bacillus, Acinetobacter, Sphingomonas, Pseudomonas, Rho­
dopseudomonas, Micrococcus, Nitrosomonas, Nitrospira, Nitrobacter, Cel­
lulomonas, and yeast species). These organisms can contribute to
 M.H. Khanjani et al.
Table 1
Cultivated species, rearing conditions, reported microorganisms and main groups of identified bacteria in the BFT system.
Cultivated Rearing conditions Reported microorganisms and main groups of Refs.
species identified bacteria
SP IBW CTV SD CSU C:N ratio L DO (mg/ pH S (PPT)
(days) (g) L)

O. niloticus 84 80–120 0.25 m3 12 kg/m3 Wheat flour – – 6 6.7 – Three groups of organisms were identified: Azim and Little
Protozoa, Rotifera and Oligochaeta. Among (2008)
protozoans, three genera, (Paramecium,
Tetrahymena and Petalomonas) dominated. Four
genera of rotifers were identified, (Lecane,
Trichocerca, Polyarthra and Asplanchna). Only
Tubifex was found in the group Oligochaeta.
M. rosenbergii 120 5.023 40 m2 2 Tapioca starch 10:01 – – – – About 40 genera of algae belonging to Asaduzzaman et al.
prawns/ 15:01 Bacillariophyceae (10 genera), Chlorophyceae (2010)
m2 20:01 (21), Cyanophyceae (7) and Euglenophyceae
(2) and 6 genera of attached zooplankton
belonging to Rotifer (5) and Crustacea (1) were
identified as periphyton communities in
substrate-added ponds. Among autotrophic
periphyton communities, Synedra, Tabellaria,
Navicula, Fragilaria, Cyclotella and Diatoma
(Bacillariophyceae), Chlorella, Sphaerocystes,
Palmella, Pediastrum, Microspora, Oocystis,
Ulothrix and Scenedesmus (Chlorophyceae),
Microcystis, Anabaena, Aphanizomenon,
Aphanocapsa and Gomphosphaeria
(Cyanophyceae), Euglena and Phacus
(Euglenophyceae), and among zoobenthic
3

periphyton crustacean nauplii, and Asplanchna,

Brachionus and Filinia (Rotifera) were the
dominating genera.
Marsupenaeus 106 0.09 30 m2 175 PLs/ Sucrose – – 4–6.2 7.8–8.4 22.7–23.4 Bacillus sp. (27.71 %), Proteobacterium (13.24 Zhao et al. (2012)
japonicus m2 %), Actinobacterium (8.16 %), Roseobacter sp
(1.53 %), Cytophaga sp. (0.97 %) were
detected in the BFT system.
L. vannamei 30 0.006 0.005 m3 50 PLs/L Sugarcane – – 4.6–7.26 7.9–8.2 32 Flagellates, Ciliates, Rotifers and Nematodes Loureiro et al. (2012)
molasses
Farfantepenaeus 210 7.4 20 m3 38 Sugarcane 20:01 Low (until 6.1–6.2 8.3–8.4 38.4–39.3 Microorganism assessment suggested a higher Emerenciano et al.
duorarum shrimps/ molasses and 80 %) presence of filamentous cyanobacteria followed (2013b)
m2 wheat bran by protozoa, nematodes and copepods.
L. vannamei 21 1 1.2 m3 500 g/m3 Corn meal 31.5:1 – 3.7–4 7.5–7.8 35.4 Vibrionaceae, followed by Enterobacteriaceae Luis-Villaseñor et al.
were the dominant families in the BFT and in (2016)
the control cultures, and the remaining strains
pertained to Alteromonadaceae and
Micrococcaceae in the BFT, and
Pseudoalteromonadaceae, Rhodobacteriaceae

Aquaculture Reports 26 (2022) 101300

and Alteromonadaceae in the control cultures.
L. vannamei 42 0.34 – 500 Molasses 12:01 Natural – – – In the biofloc tanks, relative abundance of Gálvez et al. (2015)
shrimps/ phytoplankton, including: Cyanobacteria
m3 (77.91 %), Chlorophyta (20.01 %),
Heterokontophyta (1.99 %), Euglenophyta
(0.14 %) and Dinophyta (0.02 %). Zooplankton,
including: Rotifers (43.10 %) Copepoda (33.21
%), Protozoa (13.52 %) and Cladocera (10.16
%).
L. vannamei 60 0.15 0.35 m3 7 7.6–7.8 25 Rajkumar et al. (2016)
(continued on next page)
 M.H. Khanjani et al.
Table 1 (continued )
Cultivated Rearing conditions Reported microorganisms and main groups of Refs.
species identified bacteria
SP IBW CTV SD CSU C:N ratio L DO (mg/ pH S (PPT)
(days) (g) L)

130 Molasses, 12 hL:12 The group of organisms was identified as

shrimps/ tapioca flour hD tintinnids, ciliates, copepods, cyanobacteria and
m2 and wheat flour nematodes.
O. niloticus 70 4.2 1 m3 0.252 g/L Molasses, 20:01 – 2.9–4.07 8.26–8.46 – Three bacterial groups were identified, Gutiérrez et al. (2016)
including:
mixture of pathogen (Aeromonas hydrophila, Enterobacter
molasses + rice sakazakii, Vibrio fluvialis, Pasteurella
powder pneumotropica, Brucella spp., Moraxella spp.
Citrobacter freundii, Erwinia spp.), degradative
(Pseudomonas cepacia, Pseudomonas fluorescens,
Enterobacter cloacae, Actinobacter baumannii,
Sphingomonas paucimobilis, Ochrobactrum
anthropi, Burkholderia cepacia, Prolinoborus
fasciculus), and probiotics (Bacillus simplex,
Lactococcus lactis, Cedecea lapagei, Rhodotorula
spp., Saccharomyces spp., Pseudomonas
oryzihabitans, Bacillus cereus, Lactobacillus spp.,)
Litopenaeus 35 4.45 0.25 m3 900 g/m2 Sugar cane 20:01 Low (until – – – Major Phyla abundance: Proteobacteria, Cardona et al. (2016)
stylirostris molasses 70 %) Bacteroidetes, Cyanobacteria, Euryarchaeota,
Actinobacteria. In BFT water, Leucothrix (20.1
%), Rhodobacteraceae (16.4 %), Stramenopiles
(8.0 %), Oceanospirillaceae (5.5 %) and
Saprospiraceae (4.7 %) were the most
dominant.
4

Penaeus 105 – 8 m3 – Molasses 10:01 – 6.67 7.36 33.66 Abundances and high varieties of Manan et al. (2017)
vannamei phytoplankton, zooplankton, protozoa,
nematodes and algae.
Heterotrophic bacteria identified from
Pseudomonas and Aeromonas family
L. vannamei 44 0.42 28 m3 400 PLs/ Molasses 15:01 – 5.06–5.33 7.4 39–39.5 Different concentrations of Vibrio spp (0–17.3 × Pacheco-Vega et al.
m3 103 CFU mL− 1) and Bacillus spp bacteria were (2018)
reported.
Pelteobagrus 42 38.1 0.3 m3 – Tapioca starch, 20:01 – 5.6–6.3 6.6–6.7 – Identified 13 bacterial phyla. Deng et al. (2018)
vachelli plant cellulose Abundant: Proteobacteria (53.4–59.5 %),
and the mixture Alphaproteobacteria and Betaproteobacteria,
of tapioca starch Deltaproteobacteria and Gammaproteobacteria.
and plant Nitrosomonas was the most abundant genus in
cellulose all treatments.
L. vannamei 35 1 0.5 m3 150/m3 Molasses 05:01 – – 7.87–8.15 30.1–30.77 In C:N5 treated water, the Vibrio of 79 % and 37 Panigrahi et al. (2018)
10:01 % were most dominant operational taxonomic
15:01 unit respectively. In C:N10, Thauera (62 %) was
20:01 most represented taxa. In C:N15, Attheyaceae

Aquaculture Reports 26 (2022) 101300

(56 %) and Peridiniaceae (30 %) were the most
dominant phyla. In C:N20, the Psychrobacter
(26 %), Proteobacteria (25 %) and Peridiniaceae
(20 %) phyla were found to be dominated
phylum.
L. vannamei 60 – Tanks 400, 600 Brown sugar 15:01 12 hL:12 – – – Abundance of identified bacterial phyla: Deng et al. (2019)
with 5 m and 800/ hD Proteobacteria, Acidobacteria,
length, 3 m2 Bacteroidetes,and
m width,
(continued on next page)
 M.H. Khanjani et al.
Table 1 (continued )
Cultivated Rearing conditions Reported microorganisms and main groups of Refs.
species identified bacteria
SP IBW CTV SD CSU C:N ratio L DO (mg/ pH S (PPT)
(days) (g) L)

1.2 m Planctomycetes. Also at genus taxonomy level

depth. abundant: Paracoccus, Nocardioides,
Rhodopirellula, and Blastopirellula.
– 31 – 0.01 m3 – – 10:01 – 6 – – The predominant phylum were Proteobacteria, Gou et al. (2019)
15:01 Bacteroidetes and Verrucomicrobia, but
20:01 Saccharibacteria occupied a dominant position
25:01:00 in the late period of the experiment. Pathogens,
such as Aeromonas, Acidovorax, Flavobacterium,
and Malikia were significantly decreased after
high C/N ratio.
Prochilodus 120 1.6 6 m3 5, 10, and Molasses 20:01 Low (until 6.8–6.9 8 – Five planktonic groups (microalgae, rotifers, Ayazo-Genes et al.
magdalenae 20 fish/ 60 %) cladocerans, copepods, and protists with ciliates (2019)
m3 predominating) were identified, also identify
ten bacterial strains: Escherichia coli,
Enterobacter sp., Klebsiella sp., Salmonella sp.
(Enterobacteriaceae), Bacillus subtilis, Bacillus
sp., Lactobacillus sp., Pseudomonas sp.
(Vibrionaceae), Micrococcus sp., and
Staphylococcus sp. (Coccus Gram+).
O. niloticus 175 79.28 – 100 fish/ Molasses 12.5:1 – 5.4–5.8 6.75–7.45 – Microalgae, ciliates, flagellates, amebas, Gallardo-Collí et al.
m3 rotifers, nematodes and annelids were observed. (2019)
L. vannamei 49 2.68 0.3 m3 0.71 g/L Molasses 12.36 12 hL:12 5.7–8.2 7.3–8.0 32 Bacteroidetes and Proteobacteria dominated in Jiang et al. (2020)
hD and rearing tanks (for more than 74 %). Also had
24 hD lower Sphingobacteriia and higher
5

Deltaproteobacteria levels than the systems

with no added carbon source
L. vannamei 49 2.68 0.3 m3 0.71 g/L No molasses 7.37 12 hL:12 5.7–8.2 7.3–8.0 32 Bacteroidetes and Proteobacteria dominated in Jiang et al. (2020)
hD and rearing tanks
24 hD
Piaractus 91 5.4 0.5 m3 20 and 40 Brown sugar 15:01 – 07-Aug 7.3 – Dominant groups of microorganisms include Sandoval-Vargas et al.
brachypomus fish/m3 ciliates, rhizopods, rotifers, and chlorophytes. (2020)
Scortum barcoo 95 347.7 – 24.54 kg/ – – – 7.18 7.75 – Chloroflexi, Bacteroidetes, Proteobacteria, Liu et al. (2020)
m3 Chlorobi, and Acidobacteria were the dominant
bacterial phyla.
L. vannamei 70 1.92 30 m3 300 Molasses 12:01 500–900 4.5–6.2 7.0–7.8 26–28 Dominant genus, including: Nitrosopumilus, Xu et al. (2020)
shrimps/ lux Nitrosopelagicus, Nitrosomonas (96.8 % in BFT),
m3 Nitrobacter, Planctomyces.
O. niloticus 30 1.7 0.16 m3 1 fish/L Molasses, 15:01 12 hL:12 5.43–6.47 7.28–7.45 8.5 Single-cell algae, protozoa, rotifers, and various Khanjani et al.
starch, corn hD types of ciliates and flagellates. (2021a)
flour, barley
flour
Tilapia, GIFT 180 5.1 300 m2 15 fish/ 10:01 5.7–5.75 7.5–7.56 Fresh Proteobacteria and Caldilinea aerophila were the Meenakshisundaram

Aquaculture Reports 26 (2022) 101300

– –
m3 water most abundant genus identified, also bacteria et al. (2021)
was the most abundant at the kingdom level
(70.8 %) followed by Eukarya (5.08 %), Archaea
(0.62 %), Viruses (0.16 %) and Unclassified
(23.35 %)
M. rosenbergii 42 0.0219 0.1 m3 1 prawn/ Wheat bran, rice 15:01 – 6.45–6.57 7.36–7.64 5.6–5.7 Total bacteria and 8 genera of zooplankton were Hosain et al. (2021)
L flour, molasses, recorded, of which 4 genera belonged to ciliates
maize starch, (Euplotes, Paramecium, Vorticella, Acineta), 3
palm kernel rotifers (Brachionus, Lecane, Colurella) and one
expeller nematode (Rhabditis)
(continued on next page)
 M.H. Khanjani et al.
Table 1 (continued )
Cultivated Rearing conditions Reported microorganisms and main groups of Refs.
species identified bacteria
SP IBW CTV SD CSU C:N ratio L DO (mg/ pH S (PPT)
(days) (g) L)

L. vannamei 70 – 35 m3 500 Molasses 15:01 – – – 28 There was a greater diversity of microorganisms Krummenauer et al.
shrimps/ (protozoa, microalgae and bacteria) and (2021)
m2 abundance (flagellate, ciliates, ameba, diatoms,
oocysts, planktonema, bacillus and free
filamentous algae) in the BFT systems with the
air injector.
Penaeus indicus 45 1 0.5 m3 0.2 g/ L – – – 5.9–6.7 7.5–7.9 33–35 Major Phyla abundance: Proteobacteria, Panigrahi et al. (2021)
Firmicutes, Bacteroidetes, Planctomycetes,
Cyanobacteria, Actinobacteria
L. vannamei 28 3.2–3.4 – – Corn starch 20:01 12 hL:12 6 – 25 In the Prokaryotic communities, the most Tinh et al. (2021)
hD abundant microbial phyla was Proteobacteria
(35 %), Planctomycetote (15 %), Bacteroidota
(15 %), Cyanobacteria (12 %) and Chloroflexi
(10 %). while the most abundant genus was
Phaeodactylibacter (4 %), Muricauda (2 %),
Planctomicrobium (2 %), Pseudoalteromonas (1
%) and Ruegeria (1 %). In the eukaryotic
communities, Chlorophyta phylum (45 % of the
relative abundance), followed by Rotifera (5 %),
Perkinsidae (4 %), Nematoda (2 %) and
Dinoflagellates (1.2 %). Among the Chlorophyta
phylum, Nannochloropsis (20 % of the relative
abundance) was the most abundant genera.
P. vannamei 42 1.4 28 m3 350 Molasses 15:01 12 hL:12 5.5–6.3 7.3–8 36–38 microorganisms associated with biofloc were Jiménez-Ordaz et al.
6

shrimp/ hD ciliates, rotifers, and nematodes. Also identified (2021)

m3 bacteria
of the genus Bacillus and Vibrio.
M. rosenbergii 32 – 0.25 m3 60 Refined sugar 17.5:1 1000, – 7.96–8.14 12 The community of phytoplankton and Tao et al. (2021)
prawns/L 10,000, zooplankton among (all treatments) was mainly
20,000, recorded with Bacillariophyta, Euglenophyta,
and Rotifera, Dinophyta, Chlorophyta and Protozoa.
30, 000
lux
L. vannamei 91 – 30 m3 580 Molasses 15:01 12 hL:12 5.7–5.87 7.4–7.5 29.7–30.1 A total of 32 bacterial phyla were detected in Xu et al. (2021)
shrimps/ hD the biofloc samples, including Proteobacteria,
m3 Bacteroidetes, Planctomycetes, Chloroflexi,
Actinobacteria, Latescibacteria,
Gemmatimonadetes, Patescibacteria,
Acidobacteria, Verrucomicrobia and
Cyanobacteria. A total of 262 bacterial genera
were shared in the biofloc treatments. five
genera of nitrifying bacteria, such as

Aquaculture Reports 26 (2022) 101300

Nitrosomonas, Nitrosococcaceae_Unclassified,
Nitrobacter, Nitrococcus and Nitrospina, were
detected. Also At genus level,
Phaeodactylibacter, Microbacterium and SM1A02
were identified as biomarkers in indoor culture
while Marivita was identified as biomarkers in
outdoor culture.
O. niloticus 49 0.58 0.03 m3 0.638 g/L – – – – – – A total of 487 genera was identified. The four Deng et al. (2021)
core genera (95 %), namely Cetobacterium,
(continued on next page)
 M.H. Khanjani et al.
Table 1 (continued )
Cultivated Rearing conditions Reported microorganisms and main groups of Refs.
species identified bacteria
SP IBW CTV SD CSU C:N ratio L DO (mg/ pH S (PPT)
(days) (g) L)

Halomonas, Mycobacterium and an unidentified

genus within the family Peptostreptococcaceae.
L. vannamei 56 4.12 0.4 m3 200 Sucrose – – 5.99–6.2 7.81–7.91 33–33.84 All of the bacterial samples were dominated by Chen et al. (2021)
shrimps/ Proteobacteria with the proportion of
m3 38.00–60.31 %, followed by Bacteroidota
(15.49–27.89 %), Actinobacteriota
(8.20–13.72 %), Cyanobacteria (1.69–8.09 %),
Firmicutes (2.72–6.04 %), Chloroflexi
(0.89–3.37 %).
A total of 272–375 genera were detected. The
dominant genera included Maribacter, Ruegeria,
norank_f_ Rhodobacteraceae and
norank_f_norank_o_Chloroplast
O. niloticus 141 – 1330 m2 – – – – – – – Five genera identified of Nitrosospira, Fan et al. (2022)
Nitrosococcus, Nitrosomonas,
Proteobacteria_unclassified, and
Nitrosomonadaceae_unclassified.
Penaeus 60 0.11 0.1 m3 400 Tapioca flour, 15:01 – – 7.5–7.7 34.2–34.6 Abundance of phytoplankton and zooplankton Chakrapani et al.
vannamei shrimps/ rice bran, wheat included Chlorophytes (41–48 %), Cyanophyta, (2022)
m3 flour, rice flour Bacillariophyta, Ciliophora, Rotifera, Nematoda,
7

proteobacteria, Charophyta and Ochcrophyta

were reported.
L. vannamei 28 2.5 5 m3 4000 Glucose 20:01 – 6.8 7.4–8 5 The bacterial community was dominated by Huang et al. (2022)
shrimps/ phyla of Proteobacteria (45.8 %), Bacteroidetes
m3 (21.1 %), Planctomycetes (13.5 %), Chlamydiae
(10.3 %) and Firmicutes (6.8 %).
P. vannamei 56 2.68 30 m3 300 Molasses 8:1, 12:1 – – – – Main bacterial genus in water biofloc: Xu et al. (2022)
shrimps/ and 16:1 Tropicibacter, Saprospiraceae_uncultured,
m3 Planctomyces, Rhodobacteraceae_uncultured, Pir4
lineage, Ardenticatenia_norank, Microbacterium
and Candidatus Alysiosphaera. main bacterial
genus in shrimp gut: Vibrio, Spongiimonas and
Pseudoalteromonas
O. niloticus 70 5.1 0.1 m3 1.275 g/L Starch 10:1–20:1 Natural 6 8.3–8.5 – Nine zooplankton species was recorded. Aboseif et al. (2022)
Four species of rotifer were observed, Anuropsis
fissa, Lecane bulla, L. closterocerca, and Philodina
sp., Protozoa were represented by three species
Arcella sp., Centropyxis sp. and Vorticella sp.,

Aquaculture Reports 26 (2022) 101300

Also, high amounts of (aerobic plate, total
diazotrophs, spore-forming, and lactic acid
bacteria) were observed.

Abbreviations: IBW: initial body weight, SP: study period, CTV: cultivation tank volume, SD: stocking density, CSU: Carbon source used, C:N: Carbon to nitrogen, L: light, DO: dissolved oxygen, S: Salinity, hL: hours light,
hD: hours dark. GIFT: genetically improved farmed tilapia. PLs: postlarvae.
M.H. Khanjani et al. Aquaculture Reports 26 (2022) 101300

Fig. 1. a) Tanks used during the rearing Nile tilapia in a biofloc system (Khanjani et al., 2021a); b) commercial biofloc-based shrimp farming (Emerenciano et al.,
2021), c) Biofloc produced in rearing tanks; d) Rotifer, e) Nematod, f) Flagellates, j) Ciliates, (Khanjani et al., 2021a), h) Copepod (Emerenciano et al., 2013a), i)
Stenostomum, j) Euplotes (Sandoval-Vargas et al., 2020).

bioremediation and improve water quality, growth, performance, and
health of the cultured organism (Monroy-Dosta et al., 2013; Das and
Dash, 2014; Adel et al., 2017; Kumar et al., 2021).
Chemoautotrophs affect water quality through nitrification. In the
process of nitrification, first ammonia is converted to nitrite by bacteria
in genera Nitrosovibrio, Nitrosolobus, Nitrosomonas, Nitrosospira, and
Nitrosococcus (Ebeling et al., 2006; Ray, 2012). In the next step, nitrite is
oxidized to nitrates by bacteria from genera Nitrobacter, Nitrospira,
Nitrospina and Nitrococcus (Ray, 2012). Ammonia and nitrite concen­
tration, DO, temperature, alkalinity, salinity, pH, substrate, and TSS
impact nitrification (Ebeling et al., 2006; Ray et al., 2010; Abakari et al.,
2021b). Overall, the ability of autotrophs to convert inorganic com­
pounds to organic compounds makes them suitable agents in bioreme­
diation (Musyoka, 2016).

8
M.H. Khanjani et al. Aquaculture Reports 26 (2022) 101300

Monroy-Dosta et al. (2013) identified and estimated the number of identified. Additional classification showed that the Proteobacteria and
bacteria in a BFT system for tilapia cultivation. The results showed that Caldilinea aerophila were the most abundant bacterial taxa. Moreover,
the prevalence of species changes over the 14-week cultivation period; Fang et al. (2019) found Proteobacteria, Actinobacteria, Bacteroidetes,
in the first weeks, pathogenic species (Vibrio spp, Aeromonas spp) Chloroflexi, Verrucomicrobia, Planctomycetes and Cyanobacteria in pond
showed higher numbers but as time progressed, the density of hetero­ (conventional) freshwater crab farming. In BFT, these groups were
trophic bacteria increased. The microbial community included hetero­ recognized as key actors involved on water quality maintenance,
trophic species such as Sphingomonas paucimobilis, Pseudomonas luteola, bioremediation, nitrogen assimilation, and pathogen control (Meenak­
Pseudomonas mendocina, Bacillus sp, Micrococcus sp, Nitrospira sp, shisundaram et al., 2021).
Nitrobacter sp, and Rhodotorula sp. (a yeast). The authors expressed that Overall, these studies mentioned above indicate that bacteria are the
these MOs are suitable for maintaining water quality and physiological dominant MOs in BFT system, highlighting groups such as Lactobacillus,
health of the cultivated species. Bacillus, and Vibrio as well as other bacterial groups such as Halomonas,
In a BFT system for Pacific white shrimp cultivation, several groups Providencia, Nitratireductor, Pseudoalteromonas (Ebeling et al., 2006;
of bacteria were found, but the major species included Vibrio spp, Vibrio Avnimelech, 2012; Chen et al., 2021; Fan et al., 2022; Kumar et al.,
rotiferianus, Photobacterium spp, Proteus mirabilis, and Marinobacter 2021). Table 2 presents the main groups of bacteria (photoautotrophic,
goseongensis, among which Vibrio rotiferianus was the most abundant chemoautotrophic, and heterotrophic), its roles and impacts on water
(Luis-Villaseñor et al., 2016). Using 16S rRNA sequencing, Cardona et al. quality. Last but not least, photoautotrophic bacteria (e.g. cyanobacteria
(2016) analyzed the bacteria community found in BFT for cultivation of or blue-green algae) also play a key role in BFT (Emerenciano et al.,
L. stylirostris, and found that species from three phyla (Proteobacteria, 2017; Miranda-Baeza et al., 2017). The roles and functions of this
Bacteroidetes, and Cyanobacteria) were the most abundant. In the photosynthetic group will be highlighted in Section 3.2.
beginning of biofloc formation, Proteobacteria was the most abundant
phylum (60 %). These bacteria are widely distributed in seawater and 3.2. Microalgae
play an important role in nutrient recycling and mineralization of
organic matter (Kerster et al., 2006). Bacteroidetes was the second most For the purpose of this review, microalgae (phytoplankton) will be
prevalent phylum (21.9 %). These bacteria are the dominant component divided into eukaryote (e.g. diatoms, chlorophytes, etc.) and prokaryote
of heterotrophic marine bacterioplankton and are found as macroscopic (e.g. cyanobacteria) groups. These organisms play an important role in
colonies attached to organic particles (Woebken et al., 2007). Cyano­ BFT system by using nitrogenous compounds to produce proteins and
bacteria were the third most common phylum of bacterial species sugars, as well as providing oxygen when light is available (Hargreaves,
(8.5 %). Overall, these three phyla made up 90% of active bacteria in 2006; Ebeling et al., 2006; Becerril-Cortés et al., 2017; Khoa et al.,
BFT in such condition. 2020). They are also consumed by zooplankton and thus transfer nu­
A comparison of bacterial communities in BFT versus ‘water ex­ trients to higher trophic levels (Brown et al., 1997; Ju et al., 2008).
change’ system by Cardona et al. (2016) found that Leucothrix (20.1 %), Microalgae are also a good source of nutrients for aquatic organisms. For
Rhodobacteraceae (16.4 %), Stramenopiles (8 %), Oceanospirillaceae instance, eukaryote (e.g. diatoms, Chlorella sp.) are good sources of
(5.5 %), and Saprospiraceae (4.7 %) were the dominant taxonomic units essential amino acids and highly unsaturated fatty acids for shrimp
in BFT. In contrast, Cryomorpgaceae (24.6 %), Pelagibacteraceae (Brito et al., 2016). In biofloc systems, diatom inoculation (culture
(10.1 %), Stramenopiles (8.4 %), Glaciecola (5.6 %), Colwelliaceae water) improved performance (e.g. growth and FCR) and enhanced the
(4.9 %), were the most abundant under ‘water exchange’ condition. fatty acid content of L. vannamei postlarvae (Godoy et al., 2012; Marinho
These differences can be caused by fluctuations in water physicochem­ et al., 2017; Abreu et al., 2019). In commercial-scale, Emerenciano et al.
ical and biological parameters such as chlorophyll a levels, nitrate (2022c) found that promotion of diatoms by routine sodium metasilicate
concentrations, and total ammonia nitrogen (TAN) in BFT, while these supplementation significantly enhanced the growth performance and
parameters remain more constant in the ‘water exchange’ system, economics of an indoor biofloc-based L. vannamei cultivation in Mekong
affecting the bacterial abundances (Cardona et al., 2016). Delta, Vietnam. In laboratory-scale, Molina-Cárdenas and
Maya et al. (2016) studied the bacteria associated with BFT in Sánchez-Saavedra (2017) observed an inhibitory effect of diatom spe­
cultivation of Puntius conchonius. In the initial weeks, Vibrio and Aero­ cies against pathogens (e.g. vibrio), presumably through the production
monas species were dominant but their numbers dropped by the 12th of antibacterial compounds.
week. In the final two weeks, the abundance of B. subtilis and Rhodo­ On the other hand, high concentration of prokaryote (e.g.
torula sp, which show probiotic properties, increased. The authors state
that this particular BFT condition could be used as a model of ecological Table 2
succession. The effect of the presence of different bacteria on water quality parameters in the
Using 16S rRNA amplicon sequencing and shotgun metagenomic BFT system (Ebeling et al., 2006; Avnimelech, 2012; Chen et al., 2021; Fan et al.,
analysis, Tepaamorndech et al. (2020) described the bacterial commu­ 2022).
nities in BFT for L. vannamei. They found that Vibrio sp. constituted 90 % Parameters Photoautotroph Chemoautotrophic Heterotrophic
of the bacterial community. Bacillus, Lactobacillus, Pseudoalteromonas,
NH3 Decrease Increase then Decrease
Clostridium, Shewanella, Acinetobacter, Photobacterium, Alteromonas, decrease
Marinifilum, and Pseudomonas species were also identified. Wei et al. NO2 Effectless Increase then Affectless
(2020) employed high-throughput sequencing for the 16S rRNA gene to decrease
identify bacteria in BFT. The results indicated that Flavobacteriaceae (e. NO3 Decrease Increase Affectless
O2 Increase Decrease Decrease
g., Marivita, Ruegeria, and Maribacter) and Rhodobacteraceae were the
CO2 In the presence of light Increase Increase
main bacterial taxa. Interestingly, Islam et al. (2021) assessed the (decrease), darkness
structure of the microbial community in BFT for farming Macrobrachium (increase)
rosenbergii. They found that at a C/N ratio of 15–20:1, Lactobacillus was pH In the presence of light Decrease Decrease
the most abundant, followed by Enterococcus sp. Meenakshisundaram (increase), darkness
(decrease)
et al. (2021) evaluated the microbial community in BFT during grow-out Total Effectless Increase Increase
phase in the cultivation of genetically modified tilapia. Metagenomic nitrogen
profile analysis using Illumina Nextseq500 shotgun sequencing revealed Growth rate Fast Slowly Very Fast
that the community included 70.80 % bacteria, 5.08 % eukarya, 0.62 % Dominant Photosynthesis, Nitrification, Assimilation
process Assimilation Removal Conversion
archaea, 0.16 % virus, while the rest of the organisms could not be

9
M.H. Khanjani et al.
cyanobacteria) can adversely impact the water quality and performance
of the cultivated organisms through the production of toxins and
blooming (Emerenciano et al., 2017). Cyanobacteria also contains ca­
rotenoids and unsaturated fatty acids (Alsenani et al., 2020). However,
in BFT system which rely on cyano-photoautotrophs, environmental
variables such as toxins, DO, CO2, pH, and TAN can fluctuate consid­
erably, which may impact the growth and survival of the cultivated
species (Emerenciano et al., 2022a). Miranda-Baeza et al. (2017) found
that the type and amount of cyanobacteria can directly impact the water
quality (pH, chlorophyll a, TSS and nitrate) and tilapia performance in
BFT farming condition. The authors observed poor final weight, final
biomass, FCR, SGR and survival in higher Oscillatoria sp. concentrations.
The relationship between algal and bacterial communities in BFT
systems can be mutualistic or antagonistic (i.e., inhibitory or stimula­
tory). For instance, some bacteria can contribute to the growth of algae,
and the extracellular substances secreted by diatoms can be used by
bacteria as an energy source (Fukami et al., 1997; Bruckner et al., 2008).
Such interactions, and other similar relationships, can contribute to the
control of harmful bacteria in BFT systems (Fuentes et al., 2016).
Furthermore, algae can produce antibiotics that directly affect the
structure of the microbial community (Fuentes et al., 2016; Khoa et al.,
2020).
The following genera and species of algae have been reported from
BFT systems: Scenedesmus quadricauda, Coelastrum, Pediastrum dúplex,
Cyclotella, Navicula, Synedra, Fragilaria, Fragilaria, Orthoseira, Rhabdo­
nema, Ulothrix, Skeletonema, Cylindrotheca, Hemiaulus, Phymatodocis,
Ulothrix, Cyanobacteria, Chlorophyta, Bacillariophyta, Euglenophyta and
Dinophyta (Monroy-Dosta et al., 2013; Brito et al., 2013;
Ferreira-Marinho et al., 2014; Castro et al., 2014). Manan et al. (2017)
found that algae from Chlorophyceae (Green algae), Bacillariophyceae
(Diatoms), and Cyanophyceae (Blue green algae) were the most common
species found in the initial stages of BFT aquaculture, in addition to
species from Dinophyceae (Dinoflagellates) in some cases. Gálvez et al.
(2015) and Schrader et al. (2011) found similar results; the most
abundant algae were Cyanobacteria (75 %), followed by Chlorophyta,
Heterokontophyta, Euglenophyta, and Dinophyta.
3.3. Fungi
Given fungi are heterotrophs, they are in competition with bacteria
for resources such as hexose sugars (fructose and glucose) in BFT sys­
tems (Hargreaves, 2006; Emerenciano et al., 2017; Abakari et al., 2022).
The yeast from the genus Rhodotorula were found in large numbers in
tilapia reared in BFT throughout 14 weeks of cultivation period, likely
acting as natural probiotic and recycling the organic material from the
environment (Monroy-Dosta et al., 2013).
Microfungi can be considered as one of the most diverse eukaryotic
organisms that play an important role in the decomposition of organic
material (Black and Black, 2008). The presence of the filamentous
microfungi in biofloc will strengthen and contribute to forming larger
and stronger biofloc particles (More et al., 2010). Some species of
microfungi such as Aspergillus sp. and Penicillium sp. were demonstrate to
be bioflocculant producing micro-organisms with high flocculation ac­
tivity (Liu and Cheng, 2010; Cheng and Hu, 2010). Moreover, interac­
tion and association of microfungi and other micro-organism within the
biofloc can stimulate the growth and survival of shrimp as well as in­
crease their resistance to disease (De Schryver et al., 2008; Kasan et al.,
2018). Nine species of microfungi from biofloc samples of L. vannamei
culture pond known as Aspergillus versicolor, A. niger, A. tamarii, A. fla­
vipes, A. aculeatus, Penicillium citrinum, P. griseofulvum, Trichoderma
virens, and Pestalotiopsis microspora were successfully recognized
through 18s rDNA sequences analysis (Kasan et al., 2018). The existence
of microfungi species such as Aspergillus sp. and Penicillium sp. in biofloc
are perfect candidates in forming biofloc that can decrease water
pollution towards circularity and sustainable aquaculture industry
(Kasan et al., 2018). In a study conducted by Tabarrok et al. (2021),
10
Aquaculture Reports 26 (2022) 101300
fungal metagenome diversity of recirculating aquaculture (RAS) and
biofloc rearing systems of common carp (Cyprinus carpio) by using ITS 2
were investigated. The authors identified as the most dominant species
Homophron spadiceum, an agaric fungus in the family Psathyrellaceae.
Certainly, more research is need in the fungi space in BFT to better
understand their relationship with other BFOs and its impact on animal
performance, water quality and nutritional value of bioflocs.
3.4. Zooplankton
Various species of zooplankton (rotifers, protozoans, crustaceans,
and nematodes) naturally occur in BFT systems and function in nutrient
recycling, maintenance of water quality, and nutrition of the cultivated
species (Emerenciano et al., 2013a; Khanjani et al., 2021a). Consump­
tion of BFOs by the cultivated species can improve growth rate, decrease
FCR, and provide a more balanced diet (Hargreaves, 2013). Rotifers can
contain 54–60 % raw protein; the same figure for cladocerans and co­
pepods is 50–68 % and 70–71 %, respectively (Ray et al., 2010). The
lipid content of these three groups is respectively 3.9–13.2 %, 1–2.9 %,
and up to 2.6 % (Ray et al., 2010). Nematodes can also be good sources
of nutrients; nematode dry matter is 40 % protein and 20 % fat, with the
remaining 40 % being composed of nitrogen-free extract and other
macronutrients (Figueroa, 2009). Ciliates feed on algae, bacteria, and
fungi and are in turn consumed by metazoans (Nagano and Decamp,
2004). Ciliates are a rich source of free amino acids, and similar to fla­
gellates, can provide other organisms with polyunsaturated fatty acids
and sterols (polyunsaturated fatty acids) and sterols for the shrimp
(Decamp et al., 2001).
Protozoa are an important component of BFOs as they can colonize
organic particles and consume bacteria (De Schryver et al., 2008; Fer­
reira et al., 2015). Ciliate densities in BFT systems for the cultivation of
shrimp can reach 39–169 per mL (Ballester et al., 2010). Maicá et al.
(2012) report ciliate densities of 164, 64 and 29 ciliates per mL under 2,
4 and 25 g/L salinity, respectively. These observations indicate that
organisms other than bacteria also inhabit BFOs. Hargreaves (2013)
reports the presence of other organisms such as nematodes, gastrotrichs,
euplotes, vorticella, ciliates, rotifers and copepods in BFOs, which acts as
links in the food chain between algae and bacteria, and organisms higher
in the food chain.
In a study by Krummenauer et al. (2021), MOs identified in BFT
systems for shrimp cultivation included coccoids, free filamentous and
attached filamentous bacteria, Vibrio and Bacillus species, and groups of
flagellates, ciliates, diatoms, oocystes, ameba, planktonema, rotifers and
nematodes. Gálvez et al. (2015) studied the zooplankton composition in
an integrated BFT system for cultivation of L. vannamei and Gracilaria
birdiae (a species of red algae) during a 42-day period of cultivation.
BFOs in this system included 43.10 % rotifers, 33.21 % copepods,
10.16 % cladocerans, and 13.52 % protozoans. The literature suggests
that rotifers play an important role in meeting the nutritional needs of
cultivated species (Asaduzzaman et al., 2010; Khanjani et al., 2021a;
Krummenauer et al., 2021). Their small size (0.06–1.00 mm) makes
them suitable food for very young larvae (Kyewalyanga et al., 2004).
Therefore, it is essential for the survival of many fish larvae that enough
rotifers be available in the environment at the right time.
Nematodes have also been reported to occur in BFT systems (Emer­
enciano et al., 2013b; Manan et al., 2017; Gutiérrez et al., 2016). These
species can be a rich source of live food in BFT systems (Jiménez-Ordaz,
Silva et al., 2021, 2021). Nutritional analysis has shown that nematodes
contain 48.3 % protein, 17.3 % lipid, and 31.3 % carbohydrates in dry
weight; it should be noted, however, that these values are affected by
environmental conditions (Biedenbach et al., 1989; Focken et al., 2006).
In future, isolation and manipulation of certain BFOs could enhance the
animal performance in BFT, especially larvae/postlarvae stage, boosting
growth and reducing the cultivation period. Therefore, more research is
needed in this field.
M.H. Khanjani et al.
4. Factors affecting BFOs composition in the BFT system
BFOs community rapidly responds to changes in environmental pa­
rameters. Although these changes can be hard to observe, they can
manifest as activation or deactivation of certain pathways and changes
in the communities’ composition, structure, and functionality (Bentzon
et al., 2016). Environmental changes, including biotic factors such as
predation and abiotic factors such as salinity, can affect BFOs’ produc­
tion, biodegradation activities, and abundance (Avnimelech, 2007). The
proliferation of BFOs can also be influenced by the type of cultivated
species, environmental conditions, food availability, and source of car­
bon (Monroy-Dosta et al., 2013).
4.1. Salinity
Salinity can affect the density of heterotrophic bacteria and other
BFOs (Khanjani et al., 2020a, 2020b; Maicá et al., 2012). In a study by
Maicá et al. (2012), chlorophytes were the dominant species of algae at
low salinities while diatoms were dominant at the highest levels of
salinity. Ju et al. (2008) reported similar results, with chlorophytes
being more abundant at 5 g/L of salinity and diatoms being abundant at
32 g/L. Chlorophytes have lower nutritional value than diatoms due to
having lower levels of essential polyunsaturated fatty acids and ineffi­
cient assimilation by zooplankton and cultivated animals; in contrast,
diatoms have lower fiber content are therefore more digestible (Brown
et al., 1997). This might explain decreased performance by the cultures
species as salinity decreased and diatoms became less abundant.
Pedros-Alio et al. (2000) pointed the salinity as the most important
factor controlling the density of ciliates, and state that increases in
salinity may decrease ciliate abundance. A similar effect was confirmed
by Decamp et al. (2003). Maicá et al. (2012) found greater numbers of
diatoms and flagellates at higher salinities, and chlorophytes and ciliates
at lower salinities in biofloc shrimp cultivation. Khanjani et al. (2020a)
studied the impact of different salinity levels and food sources on the
density of heterotrophic bacteria in biofloc cultivation of Pacific white
shrimp and found an increase in bacterial density with increased salinity
in the 10–32 g/L range.
4.2. Carbon source
Different sources of carbon affect bacterial abundances in BFT sys­
tems (Viau et al., 2013; Gutiérrez et al., 2016; Deng et al., 2018;
Khanjani et al., 2021b; Hosain et al., 2021; Chakrapani et al., 2022). As
carbohydrates become more available, heterotrophic bacteria dominate
the community of BFOs (Abakari et al., 2021a; Panigrahi et al., 2019b;
Hosain et al., 2021; Chakrapani et al., 2022). Deng et al. (2018) studied
the effects of different sources of carbon on BFT microbiota through the
addition of tapioca starch, cellulose, and their combination in cultiva­
tion of herbivorous Carp. The results of mass sequencing showed that
Proteobacteria, and Bacteroidetes were the most abundant phyla regard­
less of the source of carbon. It should be noted that these two phyla are
highly abundant in most aquatic environments and aquaculture systems
(Guo et al., 2011; Wei et al., 2016; Zhou et al., 2017).
Khanjani et al. (2021a) found that simple sources of carbon (starch,
molasses) led to higher densities of heterotrophic bacteria compared to
complex sources (barley, flour, corn). However, maintaining the balance
between BFOs densities and the right levels of dissolved oxygen (DO)
requires constant supervision (Emerenciano et al., 2012). For example
Pérez-Fuentes et al. (2016) report a sharp reduction in DO levels (from
3.2 to 1–1.5 mg L− 1) when more 0.12 g/L of molasses was added to the
system. Although such a change might not be lethal, it could have
negative effects on the bacterial community and overall shrimp-fish
performance in BFT.
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Aquaculture Reports 26 (2022) 101300
4.3. Aeration
Aeration is among the factors that affect the density and diversity of
BFOs communities in aquaculture systems (Krummenauer et al., 2021).
Adequate levels of DO increase the activity of heterotrophic and
chemoautotrophic bacteria and enables them to digest the available
sources of carbon (e.g. organic and inorganic, respectively) and assim­
ilate/transform nitrogen (Hargreaves, 2006). Crab et al. (2012) suggest
that DO must be higher than 5 mg L− 1 for BFOs and the cultivated
species to be able to survive. Another goal of aeration is to keep BFOs
suspended and prevent the particles from settling to the bottom of the
medium, creating anaerobic zones. Aeration equipment usually include
paddlewheels, diffusers couple with blowers, nozzles coupled with
high-pressure pumps, nanobubbles devices, among others, (Hargreaves,
2006; Luo et al., 2017; Emerenciano et al., 2021), as well as hybrid
systems (e.g. paddlewheels + diffusers or nozzles + diffusers;
Piñeros-Roldan et al., 2020). Regardless the aeration layout, these are
adjusted to supply concentrations > 5 mg L− 1 (Hargreaves, 2013;
Piñeros-Roldan et al., 2020). In semi-biofloc photoautotrophic-based,
(also known as ‘green-biofloc’; Emerenciano et al., 2022a), oxygen can
be naturally supplemented by phytoplankton, but this source of oxygen
is variable and may impact pH and toxicity of N-compounds (Har­
greaves, 2006). In a study by Krummenauer et al. (2021) the effects of
three aeration systems (air hoses, injector nozzle, air hoses/injector
nozzle) on the development of BFOs communities in shrimp cultivation
were studied. They found that aeration with air injector nozzles led to a
greater diversity of protozoa, microalgae, and bacteria and higher
abundance of flagellate, ciliates, ameba, diatoms, oocysts, planktonema,
Bacillus species and free filamentous algae.
4.4. Light
Light is an important parameter is aquaculture and affects the per­
formance and structure of the BFOs community (Baloi et al., 2013;
Fleckenstein et al., 2019; Jiang et al., 2020; Khanjani and Sharifinia,
2021; Tao et al., 2021). Phytoplankton abundances are affected by light
conditions, which leads to variation in the nutritional value of BFs
(Samocha, 2019). Such factors are significant in aquaculture since
cultivated species can feed on algae, which enhances their growth (Kent
et al., 2011). Studies have shown that outdoor cultivation has a more
positive impact on L. vannamei and photosynthetic organisms compared
to indoor cultivation (Neal et al., 2010; Coyle et al., 2011; Fleckenstein
et al., 2019). Type/source of light (e.g. LED) can also impact the bac­
terial community in L. vannamei BFT culture. Reis et al. (2022) observed
that green-LED enabled lower nitrite concentration compared to red,
blue, yellow and white LED.
In a research conducted by Jiang et al. (2020) the effect of carbon
sources (with addition molasses and without addition molasses) and
light (natural photoperiod and 24 h dark) on the microbial community
structure in biofloc-based L. vannamei culture systems, was examined.
The authors observed that, in all the groups, Bacteroidetes and Proteo­
bacteria accounted for more than 74 % of the entire community. The
groups with light had higher Chloroplast levels than the groups without
light, while the systems with added carbon sources had lower Sphingo­
bacteriia and higher Deltaproteobacteria levels than the systems with no
added carbon source at the last sampling. Also, in their study, beneficial
bacteria, such as the genus Paracoccus, were recognized in the BFT
systems with light, but some harmful bacteria, such as the genus Leu­
cothrix, were detected in the BFT systems without light.
Members of Paracoccus were reported to exhibit probiotic effects and
were used to promote growth and enhance intestinal innate immunity
for sea cucumber (Yang et al., 2015). Moreover, Paracoccus sp. was also
identified as an efficient denitrifier (Schreier et al., 2010; Chakravarthy
et al., 2011), which has the potential to improve water quality. Light can
also restrict the growth and activity of heterotrophic microorganisms
and nitrifier communities (Dauda, 2020; Khanjani and Sharifinia, 2021),
M.H. Khanjani et al.
and affect pH, temperature, and DO in systems that are exposed to
sunlight (Hargreaves, 2006). The efficiency of the BFT system seems to
be more effective in the presence of minimal light. Therefore, the
knowledge regarding the impact of different light intensity, types,
wavelength and photoperiod regimes in BFT systems is still scarce and
more research is needed in this area.
4.5. Total suspended solids
Suspended solids are a component of the organic matter decompo­
sition pathways in BFT systems and can directly impact the water quality
(Vinatea et al., 2010; Copetti et al., 2021), as well as diversity and
abundance of BFOs (Ray et al., 2010). Some studies demonstrated that
removal of suspended solids from water affected some MOs communities
reducing the abundance of nematodes, rotifers, cyanobacteria, and
bacteria, while chlorophytes, diatoms, and dinoflagellates were not
significantly impacted (Ray et al., 2010; Dauda, 2020; Copetti et al.,
2021). These observations highlight the importance of management
decisions and its impact on the microbial community in BFT (Ray et al.,
2010). Increases in TSS often trigger increases in microbial biomass
since MOs in BFT utilize the sources of carbon and nitrogen made
available through fertilizers and animal waste. In L. vannamei cultiva­
tion, the level of settleable solids should be kept at 10–14 mL L− 1, TSS at
250–350 mg L− 1, and for turbidity at 75–200 NTU (Samocha and
Prangnell, 2019). TSS can also be managed through integrated
multi-trophic aquaculture (IMTA) since this method combines several
species which can utilize the suspended and dissolved residue within the
system (Legarda et al., 2019; Poli et al., 2019; Khanjani et al., 2022d);
for instance, inorganic extractive species (e.g., algae) and/or organic
extractive species (e.g., mollusks; echinoderms, polychaetes) can be
integrated into the production of fish and shrimp (Granada et al., 2016;
David et al., 2017; Poli et al., 2019), combined or separately, to manage
excess of nutrients and organic matter.
4.6. C/N ratio
The C/N ratio is another factor affecting the density of heterotrophic
bacteria and other MOs in BFT systems (Asaduzzaman et al., 2010;
Panigrahi et al., 2018; Gou et al., 2019; Minabi et al., 2020; Khanjani
and Sharifinia, 2022a; Xu et al., 2022). C/N ration is often manipulated
during the biofloc development to manage TAN. Avnimelech (1999)
developed the commonly used C/N ratio in BFT systems based 40–60 %
conversion efficiency of nitrogen and stated that C/N > 10 is appro­
priate for assimilating TAN in BFT, and that to remove 1 g of nitrogen,
20 g of carbon is needed.
Studies have shown that the C/N ratio affects BFOs, ammonia
pathways, growth, and welfare of cultivated organisms in aquaculture
(Panigrahi et al., 2018). Abu Bakar et al. (2015) reported that a C/N
ratio of 15 is ideal for effective nutrient removal. Panigrahi et al. (2020)
showed a decrease in the abundance of Vibrio species and an increase in
the expression of immune genes in L. vannamei with increased C/N ra­
tios. Khanjani and Sharifinia (2022a) found that increasing C/N ratio
from 10 to 22 increased the biomass of heterotrophic bacteria. The
addition of carbonaceous organic matter to BFT system also stimulated
the development of heterotrophic bacteria (Khanjani et al., 2020a,
2020b), supressing algae blooms. The subsequent ‘crash’ of the algal
population degrades the rearing system, as large quantities of dead algal
cells and organic matter accumulate at the pond’s bottom, which pro­
motes the spread of pathogenic bacteria such as Vibrio spp. (Samocha,
2019). Guo et al. (2020) investigated the effects of different C/N ratios
on the intestinal microbiome and metabolome of L. vannamei and biofloc
formation through addition of sucrose to the systems. Increased C/N
lowered the α- diversity of the intestinal microbiome and changed the
abundance of different bacteria. Actinobacteria, Rhodobacteraceae and
Alteromonadaceae saw an increase in their numbers while Cyanobacteria,
Mycoplasmataceae and Vibrio declined. The results indicate that higher
12
Aquaculture Reports 26 (2022) 101300
C/N ratios promoted the growth of beneficial bacteria, which then
produced bioactive metabolites, suppressing pathogenic species.
In a research conducted by Xu et al. (2022) showed that the
manipulation of input C/N ratio (8:1, 12:1 and 16:1) could affect the
bacterial community of both water biofloc and shrimp gut in the envi­
ronment of a commercial L. vannamei BFT system. According to the
authors, the differences in water quality and nitrogen dynamics prob­
ably resulted from the responses of the bacterial community in water
bioflocs to different C/N ratios.
4.7. Stocking density
Stocking density is one of the factors that affect the microbial pop­
ulation in the BFT system (Deng et al., 2019; Ayazo-Genes et al., 2019;
Sandoval-Vargas et al., 2020; Fan et al., 2022), mainly due to its relation
with feeding rates and consequently nutrient inputs. At high stocking
density, the excess of suspended flocs or settled organic matter (sludge)
can be removed e.g. through settling chambers (Ray et al., 2010) and
siphoning procedures (Emerenciano et al., 2022a), impacting the floc
microbial community (Ayazo-Genes et al., 2019; Sandoval-Vargas et al.,
2020).
In the study conducted by Deng et al. (2019), the abundance of
bacterial communities at the genus level was examined in different
densities of shrimp stocking. In the water column, the highest abun­
dance of Paracoccus (5.46 %) and Nocardioides (3.81 %) at a density of
800 shrimp m− 2, the highest abundance of Rhodopirellula (0.67 %) and
Blastopirellula (0.52 %) was observed at a density of 400 shrimp m− 2.
The water microbial community is clustered into groups according to
different stocking density (Deng et al., 2019; Sandoval-Vargas et al.,
2020), implying that density can strongly shift the microbial community
in BFT and potentially change its functionality. Such changes may be
related to the different ratios of nutrient inputs in the culture system,
impacting the water abiotic environment (Deng et al., 2019). In the gut
of L. vannamei, Paracoccus (47.7 %) was a dominating genus identified at
a density of 600 shrimp m− 2 (Deng et al., 2019). The high stocking
density (800 shrimp m− 2) changed the gut microbiota by decreasing the
relative abundance of Paracoccus (20.44 %) and increasing Nocardioides
(25.64 %), which implies that the potential probiotic effect of biofloc
may decrease at high stocking density.
Members of Nocardioides were reported to produce spores (Prauser,
1976) and antibiotics (Lee et al., 2014), which enable this genus to
proliferate in shrimp gut and compete against beneficial genus of Par­
acoccus, and finally may negatively influence shrimp performance.
Nutrient availability plays an important role in the biodiversity of mi­
croorganisms (Mitri et al., 2016). The highest water and gut bacterial
diversity were identified in a high stocking density system (Deng et al.,
2019). However, the nutrient dynamics in BFT is more complex,
including residual feed, shrimp faces, exuviae, dead microbes, and
additional carbon sources. In a high stocking density biofloc system, the
accumulation of minerals and other compounds may cause more mi­
crobial reactions that increased the bacterial diversity (Deng et al.,
2019; Sandoval-Vargas et al., 2020; Fan et al., 2022). At high stocking
density, higher flocs density could help shrimp particles’ capture and
ingestion. However, Wasielesky et al. (2006) demonstrated that higher
flocs concentration didn’t impact the shrimp performance, but feed
protein content. More studies are needed to clarify the impact of
stocking density and concentration of solids on floc composition and
nutritional value, as well shrimp gut and water microbiota.
5. Application of BFOs for water quality improvement and as
feed
Optimal water quality is a key requirement for the economic and
environmental sustainability of aquatic farming, especially in intensive
conditions (Emerenciano et al., 2017). The BFT system is a complex
trophic system, where MOs (e.g photoautotrophs, chemoautotrophs and
M.H. Khanjani et al.
heterotrophs) and the processes involved (e.g. photosynthesis, nitrifi­
cation, assimilation) play a key role on water quality improvement and
maintenance (Table 2). Heterotrophic bacteria is considered the ‘core’ of
BFT, controlling algae blooms and dictating system health. As per the
‘microbial loop concept’, heterotrophs are situated at the bottom of the
food chain by utilizing dissolved organic matter, re-incorporating up to
50 % of the carbon released by microalgae, accelerating mineralization
and making the nutrients (e.g. carbon) available to higher trophic level
MOs (Azam et al., 1983), The bacteria are then consumed by protozoans,
which in turn are consumed by larger organisms in the BFOs food chain
(Nevejan et al., 2018). This recycling process is especially important in
BFT with high loads of nutrient inputs and reduced water exchange
(Samocha et al., 2007; Asaduzzaman et al., 2010; Emerenciano et al.,
2022a).
BFOs can also be effective in the control of pathogens. Unlike
traditional approaches to pathogen control, BFT does not create resis­
tance in the pathogenic species since its effects arise from competition
between beneficial organisms and pathogenic organisms. Biofloc has
probiotic properties by providing a suitable substrate for potential
probiotic bacteria (Cienfuegos-Martínez et al., 2020). The probiotic
property of BFOs increases growth, digestion, metabolism, and disease
resistance in the cultivated animals (Daniel and Nageswari, 2017; Hos­
tins et al., 2019). In a stable/mature environment with favorable water
quality conditions, probiotic bacteria likely dominate over other groups
(Monroy-Dosta et al., 2015) by secreting a broad array of exoenzymes
and polymers, reducing pathogenic load in the water and guts of farmed
aquatic species in BFT (Cienfuegos-Martínez et al., 2020, 2022). The
‘natural probiotics’ in BFT can significantly decrease water treatment
costs (up to 30 %) since BFT systems require much less water exchange
(0.5–1 % per day), (Crab et al., 2009). In a commercial-scale L. vanna­
mei culture, Emerenciano et al. (2022b) demonstrated that
chemoautotrophic-based biofloc with specific protocols for pathogenic
vibrio control resulted in higher survival, superior shrimp quality and
economics metrics.
From a nutritional point of view, a range of advantages of BFT have
been reported including: (i) reduced dietary protein requirement; (ii)
reduced feed conversion ratio; and (iii) a greater scope for protein-based
ingredients to replace usual high-cost feed ingredients, e.g. fishmeal and
soybean meal (Emerenciano et al., 2022a).
BFOs combined with commercial feeds can meet the dietary de­
mands of cultured species and enhance their growth performance,
supplementing a variety of key nutrients, e.g. unsaturated fatty acids,
minerals, and amino acids (Izquierdo et al., 2006; Ju et al., 2008;
Bakhshi et al., 2018; Khanjani et al., 2022a). Many BFT studies with
supplementation of external sources of carbon, reported the important
role BFOs played in the provision of essential nutrients (Wei et al., 2016;
Deng et al., 2018; Khanjani and Sharifinia, 2022b). However, biofloc
particles differ significantly in total protein (12–49 %), lipid
(0.5–12.5 %), and ash (13–46 %) levels (Bakhshi et al., 2018; Khanjani
et al., 2022a). Also, it has been documented that BFs are rich in bioactive
compounds such as carotenoids, chlorophylls, phytosterols, and bro­
mophenols (Ju et al., 2008; Zhao et al., 2016).
Mahanand et al. (2013) reported that the total lipid content in bio­
floc was 4.22 %, of which 21.11 % and 43.1 % were polyunsaturated
and saturated fatty acids, respectively. In Moreno-Arias et al. (2018)
study, the values of α-linoleic acid (18:2n-6, ALA), linolenic acid
(18:3n-3, LNA), eicosapentaenoic acid (20:5n-3, EPA), and docosahex­
aenoic acid (22:6n-3, DHA) in biofloc biomass were determined
12.06–15.33, 0.45–0.65, 14.70–16.44, and 10.72–13.36 (g/100 g lipid),
respectively. In the same study, total levels of essential amino acids and
non-essential amino acids were reported to be 11.69–24.49 and
13.21–24.61 (g/100 g dry sample), respectively. Amounts of calcium
(12.8–29.65 g/kg), phosphorus (11.25–22.6 g/kg), sodium
(2.41–24.4 g/kg), potassium (1.68–13.7 g/kg), and magnesium
(3.28–18.1 g/kg) in BFs has been reported in various studies (Kuhn
et al., 2016; Mabroke et al., 2019).
13
Aquaculture Reports 26 (2022) 101300
The application of BFOs biomass as complementary food source has
been shown to be effective at improving growth rate, survival, and food
conversion rate. In a study on Nile tilapia (Oreochromis niloticus) Azim
and Little (2008) report an increase of 45 % in production in the BFT
system compared with the conventional system. Burford et al. (2004)
report that 29 % of the food consumed by L. vannamei can be supplied by
biofloc. They also found that biofloc cultivation could eliminate the need
for marine protein in shrimp diet, with no negative effects on growth.
Hargreaves (2013) observed a 20–30 % increase in growth of shrimp in
a BFT system. The rate of protein and lipid assimilation was also higher
in a BFT system utilizing waste material as natural feed for the cultured
species, and the food conversion ration also showed improvements
(Zhao et al., 2012; Khanjani et al., 2022a; Khanjani and Sharifinia,
2022b).
According to Effendy et al. (2016) the growth of Fenneropenaeus
indicus exhibited a significant increase in a treatment with 35 % protein
feed in a biofloc setup compared with the clear water treatment. Other
studies have investigated the effects of decreasing protein content on
growth of shrimp in BFT systems. Ballester et al. (2010) found that a
10 % reduction in feed is feasible if Farfantepenaeus paulensis postlarvae
are reared in a BFT system. Nunes et al. (2010) report that there was no
difference between shrimp fed with 25 % crude protein in a BFT system
and those fed with 37 % crude protein in clear-water intensive culture.
In tilapia culture, Durigon et al. (2020) in brackish conditions, and
Sgnaulin et al. (2020) in freshwater conditions, reported significant
digestible protein reduction in BFT culture essays.
Consumption of BFOs reduces food conversion rate and consequently
costs in feed. BFOs are also able to rapidly assimilate nitrogen from
excreta and uneaten food into compounds which can be consumed by
shrimp. Manan et al. (2017) found that heterotrophic bacteria from
Aeromonas (Aeromonas salmonicida and Aeromonas hydrophila) and
Pseudomonas family (Pseudomonas aeruginosa) consumed the settled
material at the bottom of L. vannamei cultivation tanks and subsequently
produced flocs with high protein contents which were used by the
shrimp (Manan et al., 2017). The studies mentioned above highlight the
properties of limited-exchange BFT systems in water quality and nutri­
tion perspectives. BFOs can simultaneously recycle toxic N-compounds
and limit the need for (i) protein and (ii) expensive ingredients e.g.
fishmeal in aquafeeds. Thus, BFT provides a powerful tool for achieving
the environmental, social, and economic goals of aquaculture without
sacrificing growth (Crab et al., 2012; Ahmad et al., 2017; Rajeev et al.,
2021; Khanjani et al., 2022a).
6. Conclusion
BFT aquaculture is considered a step towards sustainable aquacul­
ture by increasing production, supplementing the animals’ diets,
contributing to bioremediation and biodegradation to maintain water
quality, and reducing the need for water exchange. The current review
highlighted the BFOs roles in BFT aquaculture; for instance (i) sup­
porting water quality, (ii) acting as a nutritional component and (iii)
pathogen competition/natural probiotic effect in aquaculture. In addi­
tion, higher immune response against pathogens are among the benefits
of BFOs. Steps forward should be taken to balance and improve these
roles in a BFT system. In addition, many factors that impact BFOs were
discussed in this review. The proper BFT management will boost pro­
biotic BFOs (e.g. Bacillus sp) and limit harmful bacteria in BFT. A better
understanding of this approach as a new strategy for controlling path­
ogenic bacteria in aquaculture needs further investigation. Studies on
microbial community dynamics are very important to increase knowl­
edge in this field for researchers, farm managers and cultivators. The
study of BFOs gives a broad view on the abundance, diversity and
ecology of microorganisms and makes it possible to accurately under­
stand the functions of microorganisms as a biotechnology in
aquaculture.
M.H. Khanjani et al. Aquaculture Reports 26 (2022) 101300

CRediT authorship contribution statement Ballester, E.L.C., Abreu, P.C., Cavalli, R.O., Emerenciano, M., Abreu, L., Wasielesky, W.,
2010. Effect of practical diets with different protein levels on the performance of
Farfantepenaeus paulensis juveniles nursed in a zero exchange suspended microbial
Conception and design of study: Mohammad Hossein Khanjani, flocs intensive system. Aquac. Nutr. 16, 163–172.
Alireza Mohammadi. Drafting the manuscript: Mohammad Hossein Baloi, M., Arantes, R., Schveitzer, R., Magnotti, C., Vinatea, L., 2013. Performance of
Khanjani, Alireza Mohammadi. Revising the manuscript critically for Pacific white shrimp Litopenaeus vannamei raised in biofloc systems with varying
levels of light exposure. Aquac. Eng. 52, 39–44.
important intellectual content: Mohammad Hossein Khanjani, Alireza Becerril-Cortés, D., Monroy-Dosta, M.C., Emerenciano, M.G.C., Castro-Mejía, G.,
Mohammadi, Maurício Gustavo Coelho Emerenciano. Revising the Cienfuegos- Martínez, K., Lara-Andrade, R., 2017. Nutritional importance for
manuscript after reviewers evaluation: Mohammad Hossein Khan­ aquaculture and ecological function of microorganisms that make up biofloc, a
review. Int. J. Aquat. Sci. 8 (2), 69–77.
jani, Alireza Mohammadi, Maurício Gustavo Coelho Emerenciano. Bentzon, T.M., Sonnenschein, E.C., Gram, L., 2016. Monitoring and managing microbes
in aquaculturetowards a sustainable industry. Microb. Biotechnol. 9 (5), 576–584.
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Declaration of Competing Interest Panagrellus redivivus as an Artemia replacement in a larval penaeid diet. Journal of
the World Aquaculture Society 20, 61–71.
Black, J.G., Black, L.J., 2008. Microbiology: Principles and Exploration. John Wiley &
The authors declare that they have no known competing financial Sons (Asia) Pty. Ltd., USA.
interests or personal relationships that could have appeared to influence Brito, L.O., Arana, L.A.V., Soares, R.B., Severi, W., Miranda, R.H., da Silva, S.M.,
the work reported in this paper. Gálvez, A.O., 2013. Water quality, phytoplankton composition and growth of
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