Review Article

Probiotics in aquaculture
Priyadarshini Pandiyan
a,
*, Deivasigamani Balaraman
b
, Rajasekar Thirunavukkarasu
a
,
Edward Gnana Jothi George
a
, Kumaran Subaramaniyan
a
, Sakthivel Manikkam
a
,
Balamurugan Sadayappan
a
a
Ph.D Research Scholar, CAS in Marine Biology, Annamalai University, Parangipettai 608502, Tamil Nadu, India
b
Assistant Professor, CAS in Marine Biology, Annamalai University, Parangipettai 608502, Tamil Nadu, India
a r t i c l e i n f o
Article history:
Received 10 January 2013
Accepted 8 March 2013
Keywords:
Probiotic
Aquaculture
Lactic acid bacteria
Bacillus sp
a b s t r a c t
Aquaculture is the worlds fastest growing food production sector. However, sh culture is
currently suffering from serious losses due to infectious diseases. The use of antimicrobial
drugs, pesticides and disinfectant in aquaculture disease prevention and growth promo-
tion has led to the evolution of resistant strains of bacteria. Thus, the research into the use
of probiotics for aquaculture is increasing with the demand for environment e friendly
sustainable aquaculture. The benets of such supplements include improved feed value,
enzymatic contribution to digestion, inhibition of pathogenic microorganisms, anti-
mutagenic and anti-carcinogenic activity, and increased immune response. These pro-
biotics are harmless bacteria that help the well being of the host animal and contribute,
directly or indirectly to protect the host animal against harmful bacterial pathogens. The
use of probiotics in aquaculture has just begun, due to the fact that gastrointestinal
microbiota of aquatic organisms has been poorly characterized, and their effects are not
studied extensively. This review summarizes and evaluates brief knowledge about the
probiotic organism, the action of probiotic in sh culture and the safety evaluation of
probiotics in aquaculture.
Copyright 2013, JPR Solutions; Published by Reed Elsevier India Pvt. Ltd. All rights
reserved.
1. Introduction
Today, aquaculture is the fastest growing food-producing
sector in the world, with an average annual growth rate of
8.9% since 1970, compared to only 1.2% for capture sheries
and 2.8% for terrestrial farmed meat production systems over
the same period.
1
World aquaculture has grown tremen-
dously during the last fty years from a production of less
than a million tonne in the early 1950s to 59.4 million tonnes
by 2004. This level of production had a value of US$70.3 billion.
The diseases and deterioration of environmental conditions
often occur and result in serious economic losses.
2
During the last decades, antibiotics used as traditional
strategy for sh diseases management and also for the
improvement of growth and efciency of feed conversion.
However, the development and spread of antimicrobial resis-
tant pathogens were well documented.
3,4
There is a risk asso-
ciated with the transmission of resistant bacteria from
aquaculture environments to humans, and risk associated
with the introduction in the human environment of non-
pathogenic bacteria, containing antimicrobial resistance
genes, and the subsequent transfer of such genes to human
pathogens.
5
Considering these factors, there has been height-
ened research in developing new dietary supplementation
* Corresponding author. Tel.: 91 9524149006.
E-mail address: priyadarshinibio87@gmail.com (P. Pandiyan).
Available online at www.sciencedirect.com
j ournal homepage: www. el sevi er. com/ l ocat e/ di t
d r ug i nv e nt i o n t od a y 5 ( 2 0 1 3 ) 5 5 e5 9
0975-7619/$ e see front matter Copyright 2013, JPR Solutions; Published by Reed Elsevier India Pvt. Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.dit.2013.03.003
strategies in which various health and growth promoting
compounds as probiotics, prebiotics, synbiotics, phytobiotics
and other functional dietary supplements have been evalu-
ated.
6
In this context, microbial intervention can play a vital
role in aquaculture production, and effective probiotic treat-
ments may provide broad spectrum and greater nonspecic
disease protection.
7,8
This review summarizes and evaluates
the broader knowledge about the probiotics, selection of pro-
bionts, commonly used probiotic organism, their mode of ac-
tion and safety regulation of probiotics in aquaculture.
2. Denition of probiotics
The word probiotic was introduced by Parker, 1974.
9
Ac-
cording to his original denition, probiotics are organisms
and substances which contribute to intestinal microbial bal-
ance. Fuller, 1989
10
revised the denition as live microbial
feed supplement which benecially affects the host animal by
improving its intestinal microbial balance. Therefore, several
terms such as friendly, benecial, or healthy bacteria are
also commonly used to describe probiotics. Although appli-
cation of probiotics in aquaculture seems to be relatively
recent,
11
the interest in such environment friendly treatments
is increasing rapidly. Moriarty, 1998
12
proposed to extend the
denition of probiotics in aquaculture to microbial water
additives. A growing number of studies have dealt explicitly
with probiotics, and it is nowpossible to survey its state of the
art, from the empirical use to the scientic approach.
13,14
3. Selection of probiotics
Selection of probiotic bacteria has usually been an empirical
process based on limited scientic evidence. Many of the
failures in probiotic research can be attributed to the selection
of inappropriate microorganisms. Selection steps have been
dened, but they need to be adapted for different host species
and environments. It is essential to understand the mecha-
nisms of probiotic action and to dene selection criteria for
potential probiotics.
15
General selection criteria are mainly
determined by bio safety considerations,
a. Methods of production and processing.
b. Method of administration of the probiotic and
c. The location in the body where the microorganisms are
expected to be active.
15
Three general modes of probiotics actions have been classi-
ed and presented by Oelschlaeger, 2010
16
as follow: (1) Pro-
biotics might be able to modulate the hosts gut defenses
includingtheinnateaswell astheacquiredimmunesystemand
this mode of action is most likely important for the prevention
and therapy of infectious diseases but also for the treatment of
inammationof thedigestivetract or parts thereof. (2) Probiotics
canalsohaveadirect effect onother organisms, commensal and
or pathogenic ones andthis principle is inmany cases is of great
importance in the prevention, treatment and restoration of the
microbial equilibriuminthegut. (3) Finally, probioticeffectsmay
be based on actions affecting microbial products, host products
andfoodingredients andsuchactions mayresult ininactivation
of toxins and detoxication of host and food components in the
gut. According to above summary, all three modes of probiotics
actions are all likelihood associated with gut and/or gut
microbiota. Therefore, it hasbecomeapparent that weareinfact
dealing with another organ, the so called microbiotic canal
with the increased knowledge of the specic activity of the gut
microbiota.
17
4. Probiotic organism
Today probiotics are quite commonplace in health promoting
functional foods for humans, as well as therapeutic, pro-
phylactic and growth supplements in animal production and
human health.
18e20
Typically, the lactic acid bacteria (LAB)
have been widely used and researched for human and
terrestrial animal purposes, and LAB are also known to be
present in the intestine of healthy sh.
21,22
Interest in LAB
stems from the fact that they are natural residents of the
human GIT with the ability to tolerate the acidic and bile
environment of the intestinal tract. LAB also function to
convert lactose into lactic acid, thereby reducing the pH in the
GIT and naturally preventing the colonization by many bac-
teria,
23
The most widely researched and used lactic acid bac-
teria are the Lactobacilli and Bidobacteria.
20,24,25
Other commonly studied probiotics include the spore form-
ingBacillus sp. andyeasts. Bacillussp. havebeenshowntopossess
adhesion abilities, produce bacteriocins (antimicrobial peptides)
and provide immunostimulation.
26e29
Gram-positive obligate or
facultative anaerobes are dominant in the gastrointestinal
microbiotaof manandterrestrial farmanimals.
30
Most probionts
belong to dominant or sub-dominant genera among these
microbiota, e.g., Bidobacterium, Lactobacillus, Streptococcus.
30
Gram-negative facultative anaerobes prevail in the digestive
tract of sh and shellsh, though symbiotic anaerobes may be
dominant inthe posterior intestine of some herbivorous tropical
sh.
31
Vibrio and Pseudomonas are the most common genera in
crustaceans,
32
marine sh and bivalves.
33,34
Aeromonas, Plesio-
monas and Enterobacteriaceae are dominant in freshwater sh.
33
Bacillus spp. hold added interest in probiotics as they can be
kept in the spore form and therefore stored indenitely on the
shelf.
35
The list of microorganism authorized as probiotics in
feeding stuffs under Council Directive 70/524/EEC are given in
Table 1. In addition, other probiotics are commercialized on the
market that has been notied, but that do not appear in the last
authorized list of feed additives published by the Commission.
5. Mechanisms of action
Different modes of action or properties are desire on the po-
tential probiotic like antagonismto pathogens
36,37
abilityof cells
to produce metabolites (like vitamins) and enzymes,
38
coloni-
zation or adhesion properties
39
enhance the immune system.
40
5.1. Competitive exclusion of pathogenic bacteria
Competitive exclusion is a phenomenon whereby an estab-
lished microora prevents or reduces the colonization of a
d r ug i nv e nt i o n t o d a y 5 ( 2 0 1 3 ) 5 5 e5 9 56
competing bacterial challenge for the same location on the
intestine. The aim of probiotic products designed under
competitive exclusion is to obtain: stable, agreeable and
controlled microbiota in cultures based on the following;
competition for attachment sites on the mucosa, competition
for nutrients and production of inhibitory substances by the
microora which prevents replication and/destroys the chal-
lenging bacteria and hence reduce colonization.
12
Different
strategies are displayed in the adhesion of microorganism to
those attachment sites as passive forces, electrostatic in-
teractions, hydrophobic, steric forces, lipoteichoic acids, ad-
hesions and specic structures of adhesion.
41
Adhesion and
colonization of the mucosal surfaces are possible protective
mechanisms against pathogens through competition for
binding sites and nutrients.
42
5.2. Production of inhibitory compounds
Bacterial antagonism is a common phenomenon in nature;
therefore, microbial interactions play a major role in the equi-
librium between competing benecial and potentially patho-
genic microorganisms.
43
Antagonistic compounds are dened
as chemical substances produced by microorganisms (in this
case bacteria) that are toxic (bactericidal) or inhibitory (bacte-
riostatic) toward other microorganisms. The presence of bacte-
ria producing antibacterial compounds in the intestine of the
host, on its surface, or in its culture water is thought to prevent
proliferation of pathogenic bacteria and even eliminate these.
The structure of the antibacterial compound is often not eluci-
dated and their mode of action has not been reported. Further-
more none of these reports demonstrate that the antibacterial
compound is produced in vivo. This will be of signicant
importance, if production of these compounds and its mode of
action are understood. If the production of antibacterial com-
poundis the onlymode of action, it is possiblethat thepathogen
eventually will develop resistance toward the compound. This
will result inanineffectivetreatment. Theriskof thepathogento
develop resistance against the active compound has to be eval-
uated, to assure a stable effect of the probiotic bacterium.
5.3. Enhancement of the immune response against
pathogenic microorganisms
The immune systems of sh and higher vertebrates are
similar and both have two integral components: 1) the innate,
natural or nonspecic defense system formed by a series of
cellular and humoral components, and 2) the adaptive, ac-
quired or specic immune system characterized by the hu-
moral immune response through the production of antibodies
and by the cellular immune response which is mediated by T-
lymphocytes, capable of reacting specically with antigens.
The normal microbiota in the GI ecosystem inuences the
innate immune system, which is of vital importance for the
disease resistance of sh and is divided into physical barriers,
humoral and cellular components. Innate humoral parame-
ters include antimicrobial peptides, lysozyme, complement
components, transferring, pentraxins, lectins, antiproteases
and natural antibodies, whereas nonspecic cytotoxic cells
and phagocytes constitute innate cellular immune effectors.
Cytokines are an integral component of the adaptive and
innate immune response, particularly IL-1b, interferon, tumor
necrosis factor-a, transforming growth factor-b and several
cehmokines regulate innate immunity.
44
The nonspecic
immune system can be stimulated by probiotics. It has been
demonstrated that oral administration of Clostridium butyr-
icum bacteria to rainbow trout enhanced the resistance of sh
to vibriosis, by increasing the phagocytic activity of leuco-
cytes. Rengpipat et al, 2000
7
mentioned that the use of Bacillus
sp. (strain S11) provided disease protection by activating both
cellular and humoral immune defenses in tiger shrimp
(Penaeus monodon). Balcazar, 2003
45
demonstrated that the
administration of a mixture of bacterial strains (Bacillus and
Vibrio sp.) positively inuenced the growth and survival of
juveniles of white shrimp and presented a protective effect
against the immune system, by increasing phagocytosis and
antibacterial activity.
5.4. Antiviral effects
Some bacteria used as candidate probiotics have antiviral ef-
fects. Although the exact mechanism by which these bacteria
exerts its antiviral effects is not known, laboratory tests in-
dicates that the inactivation of viruses can occur by chemical
and biological substances, such as extracts from marine algae
and extracellular agents of bacteria. It has been reported that
strains of Pseudomonas sp., Vibrio sp., Aeromonas sp., and
groups of coryneforms isolated from salmonid hatcheries,
showed antiviral activity against infectious hematopoietic
necrosis virus (IHNV) with more than 50% plaque reduction.
46
Girones et al, 1989
47
reported that a marine bacterium,
tentatively classied in the genus Moraxella, showed antiviral
activity against poliovirus. Direkbusarakim et al, 1998
48
iso-
lated two strains of Vibrio spp. from a black tiger shrimp
hatchery. These isolates displayed antiviral activities against
IHNV and Oncorhynchus masou virus (OMV), with percentages
of plaque reduction between 62 and 99%, respectively.
6. Safety regulation
The safety prole of a potential probiotic strain is of critical
importance in the selection process. This testing should
include the determination of strain resistance to a wide vari-
ety of common classes of antibiotics such as tetracyclines,
quinolones and macrolides and subsequent conrmation of
Table 1 e List of microorganism authorized as probiotics
in feeding stuffs under Council Directive 70/524/EEC.
S. no. Probiotic organism
1. Bacillus cereus var. toyoi
2. Bacillus licheniformis
3. Bacillus subtilis
4. Enterococcus faecium
5. Lactobacillus casei
6. Lactobacillus farciminis
7. Lactobacillus plantarum
8. Lactobacillus rhamnosus
9. Pediococcus acidilactici
10. Saccharomyces cerevisiae
11. Streptococcus infantarius
d r ug i nv e nt i o n t od a y 5 ( 2 0 1 3 ) 5 5 e5 9 57
non-transmission of drug resistance genes or virulence plas-
mids.
49
Evaluation should also take the end-product formu-
lation into consideration because this can induce adverse
effects in some subjects or negate the positive effects alto-
gether. A better understanding of the potential mechanisms
whereby probiotic organisms might cause adverse effects will
help to develop effective assays that predict which strains
might not be suitable for use in probiotic products. Further-
more, modern molecular techniques should be applied to
ensure that the species of probiotics used in aquaculture are
correctly identied, for quality assurance as well as safety.
7. Discussion
The application of probiotics in aquaculture shows promise,
but needs considerable efforts of research. However, a num-
ber of probiotic products have been thoroughly researched,
and evidenced their efcacy a possible use on aquaculture.
Benecial bacterial preparations that are species-specic
probiotics have become more widely available to the aqua-
culture community. These preparations show specic bene-
cial effect as disease prevention and offer a natural element
to obtain a stable healthy gut environment and immune sys-
tem. The establishing of strong disease prevention program,
including probiotic and good management practice can be
benecial to raise aquatic organism production.
8. Conclusion
The application of probiotics in aquaculture shows promise,
but needs considerable efforts of research. It is essential to
understand the mechanisms of action in order to dene se-
lection criteria for potential probiotics. Therefore, more in-
formation on the host/microbe interactions in vivo, and
development of monitoring tools (e.g. molecular biology) are
still needed for better understanding of the composition and
functions of the indigenous microbiota, as well as of microbial
cultures of probiotics. The use of probiotics is an important
management tool, but its efciency depends on understand-
ing the nature of competition between species or strains.
Conicts of interest
All authors have none to declare.
Acknowledgments
Authors are grateful to Rajiv Gandhi National Fellowship (F1-
17.1/2011-12/RGNF-SC-TAM-1686/(SA-III Website)) University
Grant Commission, Government of India, New Delhi for the
nancial support and sincere thanks and gratitude to Prof. Dr.
T. Balasubramanian, Dean and Director, CAS in Marine
Biology, Faculty of Marine Sciences, Annamalai University,
Parangipettai for the necessary facilities provided.
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