This document discusses different types of vaccines that can be developed for aquaculture, including live attenuated, inactivated, subunit, DNA, and RNA vaccines. It notes the advantages and disadvantages of each type. First generation vaccines include live attenuated and inactivated whole pathogens, while second generation are subunit vaccines containing purified proteins. DNA and RNA vaccines are third generation vaccines that induce immune responses by encoding pathogen antigens intracellularly. The document also discusses novel vaccine types such as biofilm and egg yolk vaccines produced from chickens.

1. Dr. T. Citarasu
Associate Professor
Tele-fax: + 91-4652-253078
Mobile: +91-9994273822
Email: citarasu@gmail.com
citarasu@msuniv.ac.in
Website: www.msuniv.ac.in
www.msuniv.irins.org/profile/174278
CENTRE FOR MARINE SCIENCE AND TECHNOLOGY
MANONMANIAM SUNDARANAR UNIVERSITY, TIRUNELVELI

2. AQUACULTURE VACCINE
DEVELOPMENT

3. Chemotherapy and antibiotics are giving
positive effects for treating disease, there
are some drawbacks, such as residual
problem, resistance strain development,
bio magnification and host immune
system suppression etc.
Considering the potential threats of
diseases on one side and environmental
issues on the other side, attention should
be focused to developing alternative
practices for solving problems such as
vaccine and immunostimulants.
Development of immunostimulation and
vaccines with advanced protocols are
effective to boost the immune system
against bacterial, viral and parasitic
diseases.
NEED OF VACCINES

4. THE CRUCIAL TRAITS OF AQUACULTURE VACCINES
Induce long-lasting protective immune
memory and preventative immunity
Elicit humoral and cell - mediated
immunological responses in the host
B cells store the memory and trigger the
immunity when subsequent exposure of
pathogens
A vaccination is deemed safe if it
immunizes in a manner similar to a
natural infection but does not negative
side effects, and the fish vaccine must be
risk-free for the final consumer
Must be good protection, stress free and
time efficient with mass immunization
Equally effective in multiple fish species
and cost-effectiveness make profit

5. GENERATION OF VACCINES

6. FIRST GENERATION OF VACCINES
First-generation vaccinations are forms of
microbial pathogens like live, weakened,
and killed bacteria and viruses.
They do not trigger cell-mediated immune
responses; only humoral immune
responses are elicited.
Live-attenuated vaccines contain live
microorganisms that have been
"weakened" or rendered incapable of
causing disease but some occasions they
revert their virulence
Self-replicating qualities of live vaccines
they do not require booster dose
Genetic attenuation by mutation (deletion,
disruption of the metabolic pathway or
virulence gene); chemical (formaldehyde)
& physical (heat and UV radiation)

7. Contains defined protein antigens
or recombinant protein
components capable of inducing
humoral and cell mediated immune
responses, particularly T helper
antibody responses
Antigens are crude proteins
extracted and purified from
pathogens and also specific
immunogenic proteins can be
expressed and purified using
different recombinant expression
systems
Antigenic components are unable
to proliferate in the host and no
threat of pathogenicity to the host
SECOND GENERATION OF VACCINES

8. Examples of bacterial extra cellular
toxins like haemolysin, bacterial
outer membrane protein (OMP),
viral envelop protein etc.
They are incredibly safe method of
vaccination and are less side
effects.
They have the ability to elicit an
immune response and mount
immunological response and
memory.
Multiple doses of the subunit
vaccinations may be necessary,
and immunological adjuvant is
required for effective generation of
antibodies

9. THIRD GENERATION OF VACCINE
A short, circular piece of bacterial
DNA (plasmid) that has been
genetically modified to create one or
two specific proteins (antigens) from
a microorganism makes up DNA
vaccines, which are third-generation
vaccinations.
Host cells receive the vaccine DNA
and the "inner machinery" of the host
cells "reads" the DNA to produce
corresponding target proteins.
The host cells process them and
expose them on their surface, the
immune system is notified and
causes a variety of humoral and cell-
mediated powerful and long-lasting
immune responses to be triggered

10. CONVENTIONAL VACCINES AND THEIR DRAWBACKS
Unfortunately, they have several
drawbacks, such as a week or
shorter immunity, reversal of
virulence, ineffectiveness, heat
liability, etc.
The risk of new strains emerging
from recombination of various
strains in live attenuated vaccines
exists.
In order to retain an acceptable
degree of protective immunity for a
longer period of time and enhance
their efficiency, inactivated
vaccines require multiple booster
doses, proper adjuvant, etc.

11. Typically, inactivated vaccines are ineffective against
viral infections and intracellular microorganisms
Physical or chemical treatments that result in
denaturation of the proteins or damage to the nucleic
acids in inactivated and killed vaccines
Also toxoid vaccines mixed with immune adjuvant are
effective but high level bio safety is required

12. TYPES OF VACCINES

13. NON EXPRESSED VACCINES
Subunit Vaccines
The short, specialized proteins of a
pathogen that is non-infectious
because they cannot reproduce in
the host and promote strong
immunological response
By directly extracting the native
protein from pathogen cultures,
subunit vaccines can be made.
By include more than one protein
in a subunit vaccine; it is possible
to establish immunity against more
than one strain or serotype of a
bacterial or viral disease.

14. S/A-layer proteins, extracellular products
(ECP), outer membrane proteins (OMP),
synthetic peptides, & lipopolysachharides
(LPS) are common examples of subunit
proteins.
When administered by injection along
with an immunoadjuvant, they are more
efficient.
They have some drawbacks, such as the
need for immunoadjuvants and the need
for precise understanding of the epitope
structure of the protective antigens of the
pathogen for the vaccine.
Non expressed subunit vaccines have the
drawback of having a low level of
immunogenicity, requiring more doses
and immuno-adjuvants

15. BIOFILM VACCINE
Cells embedded in a matrix create
biofilms; structures that help
bacteria adapt to various micro
niches and endure harsh
environmental circumstances.
Bacterial cells within the biofilm and
those located in its matrix, which
includes polysaccharides,
extracellular proteins, and DNA, are
key sources of biofilm antigens.
The extracellular matrix, also known
as extracellular polymeric
substances, or EPS, which is made
up of proteins, lipids,
exopolysaccharides, and
extracellular DNA (eDNA), surrounds
the biofilm bacterium.

16. Biofilm antigens obtained from several bacterial pathogens
were highly influenced to boost the immune system in fish
species
Oral biofilm vaccines have been confirmed to have
protective effects for many fish, such as, Lates calcarifer,
Labeo rohita , Oreochromis sp., Cyprinus carpio, and Mugil
cephalus
The TLR 3, IL-1, and IL-8 genes were up-regulated in the
vaccinated fish 28 days after immunization, which increased
survival and immunological parameter

17. EGG YOLK EDIBLE VACCINE IgY
When chickens are faced with a foreign
virus or bacterium, they produce
antibodies to fight the invader. They pass
that immunity on to their offspring, and
antibodies wind up in the eggs.
1. Chickens are more apt than mammals
to make high-avidity antibodies
2. A single chicken can produce an
enormous amount of antibody, up to
3 grams of IgY per month, which is
10-20 times the amount of a rabbit
3. Chickens produce antibody much
quicker-high-titre antibody is
available from eggs as early as day 25
4. Storing of eggs (antibodies) is very
easy

18. APPLICATIONS AND ADVANTAGES
Research , diagnostics & therapy
High specificity to target antigens
Less cross reactivity compared with mammalian antibody
Potential therapeutic applications, such as passive
immunization
Effective against drug-resistant bacteria.
It is cheaper to feed and house chickens than rabbits

19. GENETICALLY ALTERED VACCINES
By deleting, disrupting, or altering the
metabolic pathway or virulence genes
that causes attenuation in pathogens
used as live-attenuated vaccinations.
This modification aids in turning a
pathogen into non-pathogenic and
produces a host's protective immune
response.
Attenuated live vaccines against
Streptococcus iniae, Vibrio
anguillarum, Flavobacterium
columnare, Edwardsiella ictaluri, A.
hydrophila and E. tarda.
Effective, but due to some of the
safety concerns, mostly not accepted
for aquaculture operations.

20. VECTORS EXPRESSED VACCINES
DNA VACCINE
The ability of DNA vaccines to trigger
both cellular and humoral immune
responses that provide meaningful
protection against fish diseases is
one of its numerous advantages over
conventional immunization
techniques.
Immunization with a circular piece of
DNA that code for an antigen
Plasmids consist of strong viral
promoter (SV-40/CMV ) to drive the in
vivo transcription and translation of
gene interest.

21. Intron A may sometimes be included to improve mRNA
stability and hence increase protein expression.
Plasmids also include a strong polyadenylation/transcriptional
termination signal, such as bovine growth hormone or rabbit
beta-globulin polyadenylation sequences.
Multi-cistronic vectors are sometimes constructed to express
more than one immunogen, or to express an immunogen and
an immunostimulatory protein

22. Recombinant plasmids enter to the host cell
Gene of interest is transcribed by the RNA polymerase II & synthesis
of messenger RNA (mRNA)
Translated into the corresponding protein in the cytoplasm of the host
cells
T- Cell as well as B-Cell immunity will developed & produced antibody
against the proteins/ keep the mammary
When pathogen (antigen) enters to the host cell, the T-Cells as well as
B cells recognize and killed.
How DNA vaccine plasmids Stimulate immune responses?

23. More effective at inducing antigen-specific immune responses,
especially cellular responses.
Compared to conventional vaccines, they are much cheaper,
safer, and the mRNA vaccine helped to synthesis of proteins.
When cells are done producing a protein, they swiftly degrade
the mRNA.
Vaccination of mRNA doesn't enter the nucleus and doesn't
change genome
RNA VACCINES

24. Types including
Non-replicating mRNA, which is the simplest technology; the
pathogen's mRNA strand is packaged with additional RNA strands
that ensure it will be copied once the vaccine is inside a cell.
In vivo self-replicating mRNA, which is the vaccine's mRNA strand.
This means that more antigen may be produced from a lower dose
of vaccination, ensuring a stronger immune response
Dendritic cells that may transfer antigens on their cell surfaces to
other types of immune cells in order to assist trigger an immune
response are known as in vitro dendritic cell non-replicating mRNA
vaccines.

25. RNA INTERFERNECE (RNAi)
“A process in which the introduction of double-stranded
RNA into a cell inhibits the expression of genes”
Shrimp also use RNA interference as a defense mechanism, when a
virus infects a shrimp, its RNA is recognized and targeted by the
shrimp's RNAi machinery, inhibiting the virus's ability to replicate

26. Post Translational Gene Silencing
(PTGS) Pathway
ds RNA in the cytoplasm triggers the multi
domain ribonuclease II enzyme DICER which
cleaves the ds RNA in to si RNA which are 21
to 23 nucleotide fragments
These si RNA is recognized by the RNA
induced Silencing complex (RISC), a multi
enzyme unit that brings about separation of
two si RNA stands.
The antisense Si RNA stand remains bound to
RISC while the sense strand is released.
Finally the antisense and RISC complex bind
the target mRNA allowing the nuclease activity
and degraded the target gene

27. RECOMBINANT SUBUNIT VACCINE
Targeted proteins of a microbial
pathogen expressed through
various expression system to
produce recombinant proteins.
that.
They have lack the capacity to
multiply in the host, making them
non-infectious
Need several immunoadjuvant
booster shots to guarantee long-
lasting protective immunity.
Induce immunity against more
than one strain or serotype of a
bacterial or viral disease by
including multiple proteins in a
subunit vaccination.

28. Virus-like particles (VLPs), which are subunit vaccine
components, are created when viral capsid proteins self-
assemble into particles that resemble the virus's natural
structure.
VLPs lack genomic content, unlike the genuine viral
particles, ruling out the possibility of reversion mutations
or pathogenic infection.
VLPs—which cannot replicate in the recipient—can
augment both innate and adaptive immune responses.

29. REVERSE VACCINE
The immunogenic sequences of
infectious diseases are predicted
using a bioinformatics approach
in reverse volcanology used to
develop vaccines.
The whole antigenic repertoire of
the infectious organism's genome
and the identification of many
prospective protective targets are
identified for potential vaccines
This method enables the
identification of previously
unknown and/or rare proteins.
OMP, cytoplasmic membrane, cell
wall, periplasmic, and ECP of the
proteins are relevant for vaccine
candidates

30. Bioinformatics Databases
Vaxign analysis and PSORTb
V.3.0.2 tuned to examine
transmembrane helices, adhesion
potential, and subcellular
localization.
BepiPred linear epitope prediction
tools predict B-cell epitopes.
Artificial neural network (ANN):
peptide binding ability to MHC
molecules
VirusPloc predicts the sub cellular
localization of viral proteins
Raptorx server produces the most
accurate structure of the protein
and its activities.

31. EXPRESSION METHODS FOR VACCINE DEVELOPMENT
Bacterial Expression System
Easiest and high levels of expression
for producing recombinant proteins.
E. coli is one of the dominant strains
for and Bacillus brevis, Vibrio
cholerae and Salmonella typhimurium
ofenly expression of antigens.
Clone the desired genes in to specific
protein expression vector such as
pET32a and transform to bacterial
expression host such as BL21DE3
To create the desired protein, the
transformants are multiplied, induced
with the proper inducers, they
expressed recombinant proteins and
then purified.

32. Bacterial expression systems have several benefits, multiply rapidly,
fast production of proteins with up to 10% of mass, short generation
times and low cost
Few drawbacks as including the lack of post-translational
modification due to lack of nucleus, Golgi complex and Endoplasmic
reticulam improper protein folding, biologically inactive & endotoxin
contamination from host

33. Yeast is the most economical
eukaryotic expression system for
large scale production of both
endogenous and heterologous
proteins.
Advantages are high cell densities,
ease of growth and manipulation,
proteolysis, well-adapted for
secreting proteins, and post-
translational modifications such as
glycosylations.
Non-pathogenic nature,
comprehensive genome sequence,
well-established genetics, and
intrinsic natural adjuvant, best
model system for creating
vaccines
YEAST EXPRESSION SYSTEM

34. Galactose induction system which serves as transcriptional
induction system in and potent promoters GAL1, GAL10, JUB1,
SNR52, MET17, TDH3, TPI1, ENO1, and PDC1 are responsible
for high level expression of foreign genes in S. cerevisiae.
Immunogenic genes may clone to the yeast expression vectors
like pESC-URA and transform to competent Yeast cells with
heat shock of 42º C and plated
There is no need of purification, largely scalable, easily
deliverable for oral vaccine and low cost etc.

35. Saccharomyces cerevisiae, Pichia pastoris, Hansenula
polymorpha, Yarrowia lipolytica, Arxula adeninivorans,
Kluyveromyces lactis, and Schizosaccharomyces pombe are
excellent strains
The extra benefits of yeast cell walls acting as immune
stimulators make them stronger candidates for vaccines.
Because yeast cells contain polysaccharides such as
mannan, beta-1, and 3-D-glucan (BGs), it was thought that
yeast cells were immunogenic

36. BACULOVIRUS EXPRESSION SYSTEM
Production of recombinant proteins in
insect cells and larvae using
recombinant baculovirus with a
double-stranded circular DNA genome.
The Autographa californica multiple
nucleopolyhedrovirus (AcMNPV) is the
most used baculovirus species for
biotechnological uses.
The large size of its genome gives the
baculovirus ample capacity to
accommodate large amounts of
foreign DNA, including multiple genes.
Inserting foreign genes downstream of
the polyhedron promoter will result in
high level expression upon infection of
permissive cells.

37. The polyhedron protein is replaced by the foreign gene of interest
and integrated into the viral genome by homologous recombination.
The Spodoptera frugiperda (Sf-9 and Sf-21) cell lines are capable of
producing the recombinant virus easily, and after infection, the
replication cycle is accompanied by the production of the foreign
protein by polyhedron or p10 promoters.

38. Glycosylation, fatty acid acylation, phosphorylation, and
proteolytic processing are among the post-translational
alterations that insect cells infected with baculovirus are
capable of better quality vaccines.
Other advantages including improved solubility, proper folding
with biological active, higher yields and 90 % efficiency

39. MAMMALIAN EXPRESSION SYSTEM
Plasmid-based expression such as
adenovirus, vaccinia, retroviral, and
baculovirus vectors, that transfer
the gene into mammalian cells.
Viral-mediated transduction
especially baculo-viruses
technology is an effective method
for achieving protein expression in
mammalian cells that produce
milligrams amounts of protein.
Benefits: Wide range of post-
transnationally modified proteins
with a higher degree of flexibility,
correct protein folding, genuine
glycosylation, carbohydrate
trimming & proper proteolytic
processing

40. Other alterations may involve carboxylation of glutamic acid
residues, hydroxylation of aspartic acid and asparagine
residues, sulfation of tyrosine residues, phosphorylation of
proteins via cell receptor-protein interaction, fatty acid
acylation and correct assembly of multimeric proteins.
Several proteins such as Red seabream iridovirus's (RSIV)
transmembrane domain (pCMV-569) and structural proteins
(glycoprotein) of spring viraemia of carp virus (SVCV) were
successfully expressed through mammalian cell lines.

41. MICRO ALGAL EXPRESSION
Chloroplast expression system is a
promising method of producing oral
vaccinations.
Chlamydomonas reinhardtii,
Dunaliella salina, cyanobacteria, and
other microalgae have been treated to
express antigen genes in chloroplasts.
Introducing desired gens into nuclear
or chloroplast genome through
transformation using electroporation,
glass beads micro-particle
bombardment, or agrobacterium-
mediated DNA transfer.
The foreign gene is integrated in to
the chloroplast genome by
homologous recombination

42. Antigens expressed in the chloroplast or anchored to the
surface of plasma membrane
Benefits: No purification, expensive fermentation, cold storage,
simplicity, safety etc
Additionally, they have health-promoting advantages by high
protein, fatty acids, pigments provide immunogenicity to hosts
fish.
Transgenic Anabaena sp. PCC 7120, which expressed WSSV
VP28 fed L. vannamei PL had higher resistance against WSSV.

43. Adjuvant has been used to increase the
potency and durability of immune
responses to specific antigens, vaccine
efficacy & reduce the booster doses
Types
Complete Freund's adjuvant (CFA):
Contains killed and dried
Mycobacterium tuberculosis bacteria
includes an oil-based emulsion. Highly
effective to enhancing the immune
response but some demerits such as
inflammation and granuloma formation
Incomplete Freund's Adjuvant (IFA):
Consists of the oil-based emulsion &
less inflammation used for booster
immunizations for CFA

44. Aluminum hydroxide & aluminum phosphate, are oldest adjuvants
help enhance the prolonged immune response which slowly
releases the antigen
Oil-in-water emulsions like MF59 and AS03 helps to activate
immune cells.
Liposomes are lipid vesicles that can encapsulate antigens
enhance the presentation of the antigen to the immune system
Monophosphoryl Lipid-A (MPLA), a bacterial lipopolysaccharide
(LPS) helps to activates Toll-like receptor 4 (TLR4) on antigen-
presenting cells
Saponins from plants can enhance antigen uptake and
presentation

45. Cytokines, such as interleukin-12 (IL-12), enhance the
immune response.
Virosomes are lipid vesicles containing viral proteins used to
deliver antigens
Virus-Like Particles (VLPs) are non-infectious particles can
serve as carriers for antigens and stimulate a strong immune
response.
Nucleic acid adjuvants, CpG oligodeoxynucleotides (CpG
ODNs) and double-stranded RNA (dsRNA) activate Toll-like
receptors (TLRs) and other pattern recognition receptors

46. IMMUNE SYSTEM AND INFLENCE OF VACCINES
FISH IMMUNE SYSTEM (INNATE)
Fish possesses innate and adaptive
immunity for their defense system.
Innate immune system is lack of
immunological memory and has the
physical barriers such as scales, skin,
and mucus that provide protection
against entry of pathogens
Innate immune system initiation by the
interaction between pathogens and
pattern recognition receptors (PRRs)
Toll-like receptors (TLRs) are trans-
membrane proteins receptors detecting /
interact the ligand molecules of
pathogens that culminate innate
immunity

47. Cytotoxic cells (NCC), NK-like cells
and granulocytes like neutrophils
involved in the innate immunity
Secretion of antimicrobial peptides
(AMPs) against pathogenic
interruption leading to promotion of
mast cell degranulation,
enhancement of phagocytosis and
decreasing fibrinolysis.
In cellular level of innate immunity,
secondary responses were observed
by repeated exposure to the same
pathogen.
Mononuclear phagocytic system
have monocytes/macrophages
helped to eliminate soluble foreign
waste products

48. Neutrophils, mast and rodlet cells
have the ability of degranulation
against bacterial, viral, protozoan
and fungal pathogens
ADAPTIVE IMMUNE SYSTEM
Delayed but strong and long lasting
for successful vaccination.
Production of antibodies, B and T
cells recognizing and responding to
specific pathogens, production of
cytokines that help coordinate their
immune response
Major Histocompatibility Complex
(MHC), involved in presenting
antigens to T cells, production of
interferons inhibit viral replication

49. Two major groups of lymphocytes, B cells that mediate antibody
(humoral) responses and T cells that mediate cell-mediated immune
responses.
In antibody responses, B cells are activated to secrete antibodies,
which are soluble forms and circulate in the bloodstream and
permeate the other body fluids, bind specifically to the foreign
antigen
Binding of antibody inactivates viruses and microbial toxins by
blocking their ability to bind to receptors on host cells and
destructing the invading pathogens.

50. Cell-mediated immunity, antigen-specific T cells are activated
that react directly against a foreign antigen
Unlike B cells, T cells can only recognize antigen that has been
processed and presented by antigen-presenting cells via their
MHC proteins.
T cells can kill, virus-infected host cells that have viral
antigens on their surface, thereby eliminating the infected cells
before the virus has had a chance to replicate.

51. SHRIMP IMMUNE SYSTEM
Unlike fishes, shrimp have
primitive type of innate immune
system and have lack of adaptive
immune system.
It includes physical barriers like
the exoskeleton that provides
protection against external threats.
They have humoral and cell
mediated immunity that help
recognize and destroy pathogens.
Coagulation is preventing
hemolymph loss by clotting
protein will trap microorganisms
during coagulation and this
reaction is catalysed by a
Transglutamase released from the
hemocytes.

52. Hemocytes play a crucial role for
phagocytosis by engulfing and digesting
pathogens, encapsulation and the
production of antimicrobial peptides.
Prophenoloxidase (ProPO) encounters a
pathogen, the ProPO system is activated,
leading to the production of melanin and the
formation of melanized nodules, which help
encapsulate and neutralize the invading
microorganisms.
They also stimulate several cellular defence
reactions, including phagocytosis, nodule
formation, encapsulation, and haemocyte
locomotion.
Activated haemocytes also produce extra
bactericidal substances, such as superoxide
anion (O2-), lysozyme that may increase
disease resistance and activation of Toll
Like Receptors (TLR).

53. Pattern Recognition Receptors
(PRRs) have the ability to the
interaction between host and
pathogen could be stimulate
immunity
Toll-like receptors (TLRs), has
the ability to recognize a wide
range of pathogen-associated
molecules patterns (PAMPs).
After recognizing PAMPs, TLRs
genes will trigger the innate
immunity of the shrimp by
initiating the signalling of TLRs
pathway (TLR-MYD88-TNRF)
followed by the AMP and proPO
cascade.

54. RNA interference as a defense
mechanism, inhibiting the virus's
ability to replicate.
Short immune memory, lack of true
immunoglobulin, but a short
memory of antigen as the immune
response might involve an induced
secretion of neutralizing substances
or defence proteins against
pathogenic interruption.
Quasi-immune response has been
reported against WSSV in
Marsupenaeus japonicas, resistance
against WSSV appeared 3 weeks
after the virus exposure and was
maintained for about 1 month, which
was endorsed by the appearance of
virus neutralizing factor(s) in the
haemolymph of immunized shrimp.

55. Shell Fishes
Innate immune system
No clear responses
No antibody production
Pathogen persists
Infectious to others
Humoral immunity
AMP, PO & Coagulation
Cellular immunity
Encap., nodule & pago.
Adaptive immune system
Specific responses
Antibody produced
Pathogen eliminated
May or may not
Humoral immunity
B cells
Cellular immunity
T cells
Fin Fishes
IMMUNE SYSTEM OF SHRIMP VS FISH

56. DELIVERY METHODS
Delivery method depends on various factors, including the fish
species, size, specific disease & age
Injection: Intra-peritoneal injection into the peritoneal cavity;
Intramuscular injection into the muscle tissue used for larger fish
species.
Oral vaccines, it may be inactivated bacterial/viral cells or bacterial
transformants with recombinants coated with feed pellets with oil
& feed them
Bio-encapsulation: Inactivated cells and recombinant bacterial
transformants encapsulated with Artemia or rotifer and feeding to
fish larvae

57. Immersion bath: Fish immersed in a solution containing the
vaccine for a specific period especially for smaller fish and
stress-free
Immersion vaccination: Fish/shrimp are coated with a vaccine
suspension for a brief period, skin & gills absorb the vaccine
Spray vaccines: Vaccine solution is sprayed onto the fish's
skin or nostrils, helps to absorbing the vaccines, for skin &
gills infection.
Recombinant or transgenic microalgae with expressing viral
and bacterial proteins are directly feeding to the fish and
shrimp larvae.

58. NANO DELIVERY
Facilitate controlled release, target-
based drug delivery, improve
biocompatibility and
biodegradability.
Proteins, hydrophobic and
hydrophilic biological
macromolecules, peptides, DNA
vaccines and other substances are
all delivered via nanoparticles.
This delivery improve adjuvant or
antigen uptake by APCs, resulting
greater immune responses.
Peptides, cell lysates, DNA
vaccines, and lipo-peptides have
been conjugated with Poly (Lactic –
co-Glycolic Acid) nanoparticles.

59. Natural polymers: Created through physical and chemical
processes, are extremely biocompatible for drug delivery.
Chitosan, starch, alginate, cellulose, hyaluronic acid, chondroitin
sulphate, and other extensively used natural polymer ingredients.
Chitoson have several advantages like biodegradability, excellent
biocompatibility, the ability to be functionalized, antibacterial
capabilities, and natural processability.
Tripolyphosphate (TPP), a cross-linking agent, has been added to
the aqueous phase containing chitosan in a method for creating
chitosan nanogels, absorbed into fish cells also useful to siRNA
delivery

60. CONCLUSIONS & FUTURE DIRECTIONS

61. Utilizing the advanced biotechnological tools and expression
system resulted efficient recombinant vaccine may produce
and this will solve almost disease outbreaks.
Advantages: Proper post translational modification and protein
folding that gives strong and effective immunity
Edible vaccine like IgY has been numerous benefits, including
lower cost, greater convenience, and less invasiveness

62. Transgenic micro algae has been expressing immunogenic
proteins are highly valuable and effective against bacterial and
viral pathogens especially to the Fish/shrimp young ones
Nano-based delivery is powerful tools have so many
advantages including controlled release of antigens and long
lasting immunity

63. THANK YOU