AND ITS APPLICATIONS IN AQUACULTURE AND FISHERIES
Trivesh S.Mayekar*, Amod A. Salgaonkar,
J.M.Koli, Pravin R. Patil, Ajit Chaudhari, Nilesh Pawar, Suhas Kamble,Abhay Giri,
Girija G.Phadke, Pankaj Kapse
Research Scholar,Central Institute of Fisheries Education,
Seven Bunglow, Versova, Mumbai-400061,India.
author E-mail: firstname.lastname@example.org
Biotechnology provides powerful tools for the
sustainable development of aquaculture, fisheries, as well as the food
industry. Increased public demand for
seafood and decreasing natural marine habitats have encouraged scientists to
study ways that biotechnology can increase the production of marine food
products, and making aquaculture as a growing field of animal research.
Biotechnology allows scientists to identify and combine traits in fish and
shellfish to increase productivity and improve quality. Scientists are
investigating genes that will increase production of natural fish growth
factors as well as the natural defense compounds marine organisms use to fight
microbial infections. Modern biotechnology
is already making important contributions and poses significant challenges to
aquaculture and fisheries development. It perceives that modern biotechnologies
should be used as adjuncts to and not as substitutes for conventional technologies in solving
problems, and that their application should be need-driven rather than
The use of modern biotechnology to
enhance production of aquatic species holds great potential not only to meet
demand but also to improve aquaculture. Genetic modification and biotechnology
also holds tremendous potential to improve the quality and quantity of fish
reared in aquaculture. There is a growing demand
for aquaculture; biotechnology can help to meet this demand. As with all biotech-enhanced foods, aquaculture will be
strictly regulated before approved for market. Biotech aquaculture also offers
environmental benefits. When appropriately integrated with other technologies
for the production of food, agricultural products and services, biotechnology
can be of significant assistance in meeting the needs of an expanding and
increasingly urbanized population in the next millennium. Successful
development and application of biotechnology are possible only when a broad
research and knowledge base in the biology, variation, breeding, agronomy,
physiology, pathology, biochemistry and genetics of the manipulated organism
exists. Benefits offered by the new technologies cannot be fulfilled without a
continued commitment to basic research. Biotechnological programmes must be fully integrated
into a research background and cannot be taken out of context if they are to
fisheries and aquaculture is an important sector of food production, providing
nutritional security to the food basket, contributing to the agricultural
exports and engaging about fourteen million people in different activities.
With diverse resources ranging from deep seas to lakes in the mountains and
more than 10% of the global biodiversity in terms of fish and shellfish
species, the country has shown continuous and sustained increments in fish
production since independence. Constituting about 4.4% of the global fish
production, the sector contributes to 1.1% of the GDP and 4.7% of the
agricultural GDP. The total fish production of 6.57 million metric tonnes
presently has nearly 55% contribution from the inland sector and nearly the
same from culture fisheries. Fish and fish products have presently emerged as
the largest group in agricultural exports of India.( Marine products export
review-MPEDA.,April 2006-March2007).The potential area of biotechnology in
aquaculture include the use of synthetic hormones in induced breeding,
transgenic fish ,gene banking , uniparental and polyploidy population and
in fish breeding
releasing hormone (GnRH) is now the best available biotechnological tool for
the induced breeding of fish. GnRH is the key regulator and central initiator
of reproductive cascade in all vertebrates (Bhattacharya et al.,2002).It is a
decapeptide and was first isolated
from pig and ship hypothalami with the ability to induce pituitary release of
luteinising hormone (LH) and
follicle stimulating hormone (FSH) (Schally et al.,1973).Since then only one
form of GnRH has been identified in most placental mammals including human
beings as the sole neuropeptide causing the release of LH and FSH. However ,in
non mammalian species (except guinea pig) twelve GnRH variants have now been
structurally elucidated ,among them seven or eight different forms have been
isolated from fish species.(Halder et al.,1991;Sherwood et al.,1993;King and
Miller,1995;Jimenez-Linan et al.,1997).The most recent GnRH purified and
characterized was by Carolsfeld et al.(2000) and Robinson et
al.(2000).Depending on the structural variant and their biological activities,
number of chemical analogues have seen prepared and one of them is salmon GnRH
analogue profusely used now in fish breeding and marked commercially under the
name of “Ovaprim” throughout the world .The induced breeding of fish is now
successfully achieved by development of GnRH technology.
Transgenesis or transgenics may be defined as the introduction of
exogenous gene / DNA into host genome resulting in its stable maintenance,
transmission and expression. The technology offers an excellent opportunity for
modifying or improving the genetic traits of commercially important fishers,
mollusks and crustaceans for aquaculture. The idea of producting transgenic
animals became popular when Palmitter et al. (1982) first produced transgenic
mouse by introducing metallothionein human growth hormone fusion gene (mT-hGH)
into mouse egg, resulting in dramatic increase in growth. This triggered a
series of attemptson gene transfer in economically important animals including
The first transgenic fish was produced Zhu et al. (1985) in China, who
claimed the transient expression n putative transgenics, although they gave no
molecular evidence for the integration of the transgene. The technique has now
seen successfully applied to a number of fish species. Dramatic growth
enhancement has been shown using this technique especially in salmonids (Devlin
et al., 1994). Some studies have revealed enhancement of growth in adult salmon
to an average of 3 – 5 times the size of non – transgenic controls, with some
individuals, especially during the first few months of growth, reaching as much
as 10 – 30 times the size of the controls (Devlin et al., 1994; Hew et al.,
1995).The introduction of transgenic technique has simultaneously put more
emphasis on the need for production of sterile progeny in order to minimize the
risk of transgenic stocks mixing in the wild populations. The technical
development has expanded the possibilities for producing either sterile fish or
those whose reproductive activity can be specifically turned on or off using
inducible promoters. This would clearly be of considerable value allowing both
optimal growth and controlled reproduction of the transgenic stocks while
ensuring that any escaped fish would be unable to breed. An increased
resistance of fish to cold temperatures has been another subject of research in
fish transgenics for the past several years (Fletcher et al., 2001). Coldwater
temperatures pose a considerable stressor to many fish and few are able to
survive water temperatures much below 0-1oC. this is often a major
problem in aquaculture in cold climates. Interestingly, some marine teleosts
have high levels (10 – 25 mg/ml) of serum antifreeze proteins (AFP) or
glycoproteins (AFGP) which effectively reduce the freezing temperature by
preventing ice-crystal growth. The isolation, characterization and regulation
of these antifreeze proteins particularly of the inter flounder Pleuronectas
americanus has been the subject of research for a considerable period in
Canada. Consequently, the gene encoding the liver AFP from winter flounder was
successfully introduced into the genome of Atlantic salmon where it became
integrated into the germ line and then passed onto the off – spring F3 where it
was expressed specifically in the liver (Hew et al., 1995).The introduction of
AFPs to gold fish also increased their cold tolerance, to temperatures at which
all the control fish died (12 h at 0o C; Wang et al., 1995). Similarly, injection or
oral administration of AFP to juvenile milkfish or tilapia led to an increase
in resistance to a 26 to 13o C. drop in temperature (Wu et al., 1998). The
development of stocks harbouring this gene would be a major benefit in
commercial aquaculture in counties where winter temperatures often border the
physiological limits of these species.
The most promising
tool for the future of transgenic fish production is undoubtedly in the development of the embryonic stem cell
(ESC) technology. There cells are undifferentiated and remain totipotent so
they can be manipulated in vitro and subsequently reintroduce into early
embryos where they can contribute to the germ line of the host. This would facilitate
the genes to be stably introduced or deleted (Melamed et al., 2002).Although
significant progress has been made in several laboratories around the world,
there are numerous problems to be resolved before the successful
commercialization of the transgenic brood stock for aquaculture. To realize the
full potential of the transgenic fish technology in aquaculture, several
important scientific break – through are required. There include (i) more
efficient technologies for mass gene transfer (ii) targeted gene transfer
technologies such as embryonic stem cell gene transfer (iii) suitable promoters
to direct the expression of transgenes at optimal levels during the desired
developmental stages (iv) identified genes of desireable traits for aquaculture
and other applications (v) informations on the physiological, nutritional,
immunological and environmental factors that maximize the performance of the
transgenics of the transgenics and (vi) safety and environmental impacts of
sex manipulation techniques to induce polyploidy (triploidy and tetraploidy)
and uniparental chromosome inheritance (gynogenesis and androgenesis) have been
applied extensively in cultured fish species (Pandian and Koteeswaran, 1998;
Lakra and Das, 1998). These techniques are important in the improvement of fish
breeding as they provide a rapid approach for gonadal sterilization, sex
control improvement of hybrid viability and clonation. Most vertebrates are
diploid meaning that they possess two complete chromosome sets in their somatic
cells. Polyploidy individuals possess on or more additional chromosome sets,
bringing the total to three in triploids, four in tetraploids and so on.
Induced triploidy is widely accepted as the most effective method for producing
sterile fish for aquaculture and fisheries management. The methods used to
induce triploids and other types of chromosome set manipulations in fishes and
the applications of these biotechnologies to aquaculture and fisheries
management are well described (Purdom, 1983; Chourrout, 1987; Thorgaard, 1983;
Pandian and Koteeswan, 1998). Tetraploid breeding lines are of potential
benefit to aquaculture, by providing a convenient way to produce large numbers
of sterile triploid fish through simple interploidy crosses between
tetraploids and diploids
(Chourrout et al., 1986; Guo et al., 1996). Although tetraploidy has been
induced in many finfish species, the viability of tetraploids was low in most
instances (Rothbard et al., 1997).
teleosts, technique for inducing sterility include exogenous hormone treatment
(Hunter and Donaldson, 1983) and triploidy induction (Thorgaard, 1983). The use
of hormone treatements, however could be limited by governmental regulation and
a lack of consumer acceptance of hormone treated fish products. Triploidy can
be induced by exposing eggs to physical or chemical treatment shortly after
fertilization to inhibit extrusion of the second polar body (For reviews see
Purdom, 1983; Thorgaard, 1983 and Ihssen et al., 1990) triploid fish are
expected to be sterile because of the failure of homologous chromosomes to
synapse correctly during the first meiotic division. Methods of triploidy
induction in clued exposing fertilized eggs to temperature shock (hot or cold),
hydrostatic pressure shock or chemicals such an colchicines, cytochalasin-B or
nitrous oxide. Triploid can also be produced by crossing teraploids and
diploids. Tetraploid induction involves fertilizing eggs with normal sperm and
exposing the diploid sygote for physical or chemical treatment to suppress the
first mitotic division. Gynogenesis is the process of animal development with
exclusive maternal inheritance. The production of gynogenetic individuals is of
particular interest to fish breeders because a high level of inbreeding can be
induced in single generation. Gynogenesis may also be used to produce all –
female populations in species with female homogamety and to reveal the sex
determination mechanisms in fish. It is convenient to use all female
gynogenetic progenies (instead of normal bisexual progenies) for sex inversion
experiments. Methodologies combining use of induced gynogenesis with hormonal
sex inversion have been developed for several aquaculture species (Gomelsky et
al., 2000). Androgenesis is the process by which would have commercial
application in aquaculture. It can also be used in generating homozygous lines
of fish and in the recovery of lost genotypes from the crypreserved sperms.
Androgenetic individuals have been produced in a few species of cyprinids,
cichlids and salmonids (Bongers et al., 1994).
Biotechnology and fish health management
Disease problem area major constraint for development of aquaculture.
biotechnological tools such as molecular diagnostic methods, use of vaccines
and immunostimulants are gaining popularity for improving the disease
resistance in fish and shellish species world over for viral diseases,
avoidance of the pathogen in very important.in this context there is a need to
rapid method for detection of the pathogen. Biotechnological tools such as gene
probes and polymerase chain reaction (PCR) are showing great potential in this
area. Gene probes and PCR based diagnostic methods have developed for a number
of pathogens affecting fish and shrimp ( karunasagar ,1999). In case of finfish aquaculture, number of vaccine
against bacteria and viruses have been developed. Some of these have been
conventional vaccines consisting of killed microorgansism but new generation of
vaccine consisting of protein subunit vaccine genetically engineered organism
and DNA vaccine are currently under development.
In the vertebrate system, immunization against disease is a common
strategy. However the immune system of shrimp is rather poorly developed,
biotechnological tools are helpful for development of molecule, which can
stimulate this immune system of shrimp. Recent studies have shown that the non
specific defense system can be stimulated using, microbial product such as
lipopolysacharides, peptidoglycans or glucans (itami et al 1998). Among the
immunostimulants known to be effective in fish glucan and levamisole enhance
phagocytic activities and specific antibody responses (Sakai, 1999).
Cryopreservation of gametes or gene banking
Cryopreservation is a technique, which involve long-term preservation
and storage of biological material at a very low temperature usually at -196 C ,the temperature of liquid
nitrogen. It is based on the principle that very low temperature tranquilize or
immobilize the physiological and biochemical activities of cell, thereby making
it possible to keep them viable for very long period.
The technology of cryopreservation of fish spermatozoa (milt) has been adopted for animal husbandary
. The first success in preserving fish sperm at low temperature was reported by
Blaxter (1953) who fertilizes Herring (Clupea herengus ) eggs with frozen thawed semen .The spermatozoa of
almost all cultivable fish species has now been cryopreserved (Lakra 1993) .
Cryopreservation overcomes problems of male maturing before female, allow
selective breeding and stock improvement and enables the conservation (Harvey
,1996)One of the emerging requirement for that can be used by breeders for
evolving new strains. Most of the
plant varieties that has been produced are based on the gene bank collections.
Aquatic gene bank however suffers from the fact that at present it is possible
to cryopreserve only the male gametes of finfishes and there in no viable technique for finfish eggs and embryos.
However , the recent report on the freezing of shrimp embryos. However , the
recent report on the freezing of shrimps embryos by subramoniam and newton
(1993) and Diwan and kandaswami (1997) look promising. Therefore, it is
essential that gene banking of cultivated and cultivable aquatic species be
Biotechnological research and development are growing at a very fast
rate. The biotechnology has assumed greatest importance in recent years in the
development of fisheries, agriculture and human health. The science of
biotechnology has endowed us with new tools and tremendous power to create
novel genes and genotypes of plants, animals and fish. The application of biotechnology
in the fisheries sector is a relatively recent practice. Neverthless,it is a
promising area to enhance fish production. The increased application of
biotechnological tools can certainly revolutionise our fish farming besides its
role in biodiversity conservation.
The paper briefly reports the current progress and thrust areas in the
transgenesis, chromosome engineering, use of synthetic hormones in fish
breeding, biotechnology in health management and gene banking.
Baras,E.,C.Prignon,Gohoungo,G.and Melard, C.,
2000.Phenotypic sex differentiations of blue tilapia under constant and fluctuating thermal
regimes and its adaptive and evolutionary implications.J. Fish Biol.
Baroiller,J.F.,Guiguen,Y. and Fostier ,
A.,1999. Endocrine and
environmental aspects of sex differentiation in fish.Cellular and molecular
Battacharya s.,Dasgupta,S, Datta,M ,and Basu, D,
2002 Biotechnology input in fish breeding .Indian journal of
Billington N, and Hebert, P.D. N,
1991.Mitochondrial DNA diversity in fishes and it implication for
Blaxter,J ,H. S, 1953 Sperm storage and cross
fertilization of spring and autumn spawning herring .Nature ,London,172:1189-1190.
Bonger,A.B.J, In”t Veld EPC, Abo-Hashema,K,
Bremmer,I.M, Eding,E.H,KOmen J and Richter, C.J.J, 1994. Androgenesis in common
carp (cyprinus carpio ) using
UV Irradiation in a synthetic ovarian fluid and heat shock, aquaculture.122:199-132.
Carolsfeld J, Powell, J.k, Park,M Fisher. W, H
Craig A.G, Chang, J.P. River, J,E and Sherwood N.M, 2000/ Primary structure and
function of three gonadotropin releasing hormone. Endocrinology, 141: 505-512.
Chourrout, D, 1987. Genetic manipulation in
fish:review of method. In k Tiews (ed), selection, hybridization and genetic
Engineering in aquaculture Bordeaux
Heenemann , Berlin, 2:111-126 .
Chourrout, D, Chevassus, B, Krieg F. Happe, G. and
Renard P, 1986. Producation of second generation triploids and tetraploid
rainbow trout by mating tetraploid males and
diplod female . potentials of tetraploid fish. Theror. Appl.gene ,
Diwan, A.D.and Kandasami, D, 1997. Freezing of
viable embryo and larvae of marine
shrimp P.Semisulcatus de haan. Aquaculture research.28:947-950.
Fiecther, G.L, Hew, C.L,Davies, P.L,2001.
Antifreeze protein of teleost fishes Anuu,Rev . Physiol, 63:359-390.
Gilberg,A ,Mikkelsen H,Sandaker,E, Ringo, E, 1997.
Probiotic effect of lactic acid bacteria in the feed on growth and survival of
fry of Atlantic cod. Hydrobiologia, 352(1-3):279-285.
Gomelsky Boris, Mims, S.D and Onders Richard J.
2000. Induced gynogenesis in Black Crappie. North American journal of
Griffith, R, Orr. J. Adam.A and Barber, I, 2000.
DNA sex identification in the three spined stickleblack. J, fish Biol, 57:
Halder ,S Sen , Bhattachery, S, Ray A.K, Ghosh, Aand jhingran, A,G, 1991. Induced spawing of
IMC and maturation of a perch and a catfish by murrel gonadotropin releasing hormone
, pimozide and calcium aquaculture, 97:373-382.
Davies,P.L.,1995.Transgenic salmon:tailoring the genome for food
production.J.Fish Biol .,47:1-9.
Karunasagar,I. and Karunasagar,I.,1999.Diagnosis
treatment and prevention of microbial diseases of fish and
Lakra,W.S. and Das ,P.,1998.Genetic engineering in
aquaculture ,Indian .J.Anim.Sci.,68(8);873-879.
export review-MPEDA.,April 2006-March2007.
Pandian,T.J.and S,G.Sheela,1995.Hormonal induction
of sex reversal aquaculture,138:1-22.
Sakai,M.,1999.Current research status of fish
immunostimulants. Aquaculture ,172:63-92.
Subramonium,T.and Newton ,S.,1993.Cryopreservation
of penaeid prawn embryos.Current science,65(2):176-177.
Zhu,Z.,G,Li,L.Je and Chen ,S., 1985.Novel gene
transfer into the fertilized egg of goldfish(Carassiius auratus ),Z.Angrw.Ichthyol,1:32-34.