113
Fish Vaccines
Alexandra Adams1 and Rohana Subasinghe2
1
Institute of Aquaculture, University of Stirling, Stirling, Scotland
2
FUTUREFISH, Rajagiriya, Sri Lanka
9.1 ­Introduction
In 2014, the contribution of aquaculture to supply food
for human consumption overtook that for wild‐caught
fish for the first time (FAO 2016). China has played a
major role in this growth, as it represents more than 73%
of world aquaculture production today (FAO 2018).
Aquaculture currently contributes approximately 80 mil-
lion tonnes of aquatic animals (including marine and
freshwater finfish, crustaceans, and shellfish) with a
value of US$ 232 billion. With an annual percent rate of
growth of 5.8% since 2001, aquaculture still represents the
fastest growing animal production sector in the world
(FAO 2018). Twenty‐seven finfish species make up 90% of
global aquatic animal production, with Atlantic salmon
(Salmo salar) being the number one fish species in terms
of economic value and in the top 10 species in terms of
volume (FAO 2018).
Fish diseases are considered to be a major constraint to
aquaculture globally, with all finfish aquaculture sectors
affected to some extent by infectious disease (Rodger 2016).
It has been estimated that 10% of all cultured aquatic ani-
mals are lost because of infectious diseases alone, amount-
ing to >10 billion US$ in losses annually on a global scale
(Evensen 2016).
Although many bacterial diseases are now effectively con-
trolled by the use of vaccines, viral diseases still present sig-
nificant infectious disease challenges for salmonid and
marine finfish, and there are only a limited number of effec-
tive vaccines commercially available for these (Rodger 2016).
Bacterial pathogens still present some major challenges for
rainbow trout, carp, tilapia, and catfish. In addition, these
pose problems for “cleanerfish,” i.e. ballan wrasse (Labrus
bergylta) and lumpsucker (Cyclopterus lumpus), which are
currently being used in the biological control of sea lice
Veterinary Vaccines: Principles and Applications, First Edition. Edited by Samia Metwally, Gerrit Viljoen, and Ahmed El Idrissi.
© 2021 John Wiley & Sons Ltd. Published 2021 by John Wiley & Sons Ltd.
(Lepeophtheirus salmonis) in Atlantic salmon aquaculture.
Ectoparasites (including sea lice and Paramoeba perurans
which causes amoebic gill disease, AGD) currently pose the
most significant disease threat for the Atlantic salmon
industry and there are no commercial vaccines available at
present for these, nor for fungi or fungi‐like organisms. The
more common “water molds” in fish, such as Saprolegnia
and Aphanomyces, are not in fact true fungi but oomycetes
and are now considered to be opportunistic facultative para-
sites, e.g. Saprolegnia parasitica and Aphanamyces invadans.
The former, fungal‐like oomycete causes significant eco-
nomic losses to salmonid aquaculture (both eggs and fish)
while the latter causes epizootic ulcerative syndrome (EUS)
in many freshwater and brackish species in the Asia‐Pacific
region and Australia.
Vaccines are recognized as important tools for the pre-
vention and control of diseases in fish. The number of fish
vaccines commercially available has grown in recent years
but there are still numerous diseases where no vaccines are
available or cases where existing vaccines do not perform
well. Atlantic salmon and rainbow trout aquaculture in the
UK, Norway, and USA expanded in the 1980s with a con-
current increase in disease. This led to the use of large
amounts of antibiotics and consequently concerns grew
with regard to antibiotic resistance. This stimulated the
development of vaccines against bacterial pathogens and
led to the first commercially available fish vaccines against
vibriosis and enteric redmouth (ERM), followed by furun-
culosis vaccines. Commercial vaccines for fish have
expanded from two in the 1980s to 24 currently, with one
vaccine also available for lobsters (Assefa and Abunna
2018), with many of these now being multivalent. For
example, heptavalent vaccines exist for use in Atlantic
salmon and the use of antibiotics has been reduced by over
99%. In comparison, although carp and tilapia are
114 Veterinary Vaccines: Principles and Applications

well‐established cultured species, there are few vaccines Table 9.1 Advantages and disadvantages of different methods
available for them. of fish vaccine delivery.

Delivery
method Advantages Disadvantages
9.2 ­Current Fish Vaccines
Injection Exact dose known, Fish need to be caught and
Commercial fish vaccines are available for a wide range of good immune anesthetized, cannot use
fish species (reviewed by Evensen 2016; Assefa and Abunna response, adjuvants with small fry
available
2018), including Atlantic salmon, rainbow trout, sea bass
(Dicentrarchus labrax) and sea bream (Sparus aurata), Dip Exact dose known. Do not always get a good
immersion Used with small fry, immune response (depends
tilapia (Oreochromis niloticus/mossambicus), amberjack little handling on vaccine), cannot be used
(Seriola dumerili), and yellowtail (Seriola quinqueradiata) needed – net the fish with larger fish. No
in Japan, catfish (Ictalurus punctatus), and Vietnamese cat- and dip in vaccine for adjuvants available,
fish (Pangasionodon hypophthalmus). The majority of 30 s therefore protection
short‐lived
commercial fish vaccines are formalin killed whole cell
vaccines although live attenuated vaccines are licensed in Bath Exact dose known. Do not always get a good
immersion Used with larger fish. immune response, fish
the USA for use in catfish (Klesius and Pridgeon 2014). The need to be closely watched
Can also be useful
latter are not currently allowed to be used in Europe. In and water aerated. No
with small fry if dip
addition, a DNA vaccine against infectious hematopoietic vaccination is too adjuvants available,
necrosis (IHN) is licensed in Canada for use in Atlantic stressful, e.g. with therefore protection
ballan wrasse short‐lived
salmon (Alonso and Leong 2013). There is also currently
one commercial subunit vaccine (peptide; VP2) used in Oral Vaccine given with Do not always get a good
Norway (against infectious pancreatic necrosis virus, feed so no fish immune response;
handling normally used as booster
IPNV) and one recombinant vaccine against infectious vaccination and vaccine
salmon anemia virus (ISAV) in Chile. needs to be protected.
In the European Union (EU), the cost of fish vaccine pro- Vaccine dose per fish not
duction is high as there are stringent requirements for vac- known
cine manufacturers, although vaccines for use in small
markets may take advantage of the Minor Use Minor
Species Limited Market (MUMS) where the regulatory vaccination. This is either performed by hand, normally by
requirements are reduced (Cowan et al. 2016). an injection team on site (Figure 9.1), or automated
There are a number of important considerations for the machines are now available and used widely in some coun-
use of vaccines in fish, including fish species, status of the tries. Vaccination by injection can potentially cause stress
immune system, production cycle and life history, when a but no mortality is usually associated with the vaccination
disease occurs, farming technology (handling, mechaniza- process per se, although some weak fish may die due to the
tion, etc.), environment (e.g. temperature, salinity), stress handling process. Vaccine (usually 0.1–0.2 mL) is injected
factors, nutrition, and cost benefits. Guidelines on the use in the abdominal area of each anesthetized fish (>50 g),
of fish vaccines are provided by the Responsible Use of although microdoses of vaccines (0.025 or 0.05 mL) are also
Medicines in Agriculture Alliance (RUMA 2006). now being used. Fish are held ventral side up for vaccina-
tion and the needle is inserted into the peritoneal cavity
using an automated injection gun (Figure 9.2). A team of
9.3 ­Methods of Administration four people can vaccinate approximately 5000 salmon per
hour. Fish are often graded at the same time.
Fish vaccines are administered by injection, immersion, or Vaccination by injection provides a long duration of pro-
orally. Each of these methods has advantages and draw- tection (>12 months) and multiple antigens can be com-
backs (Table 9.1). bined in a single administration. In addition, each fish
receives the vaccine at the correct dose; 10 000 fish (>25 g)
can be vaccinated per liter of vaccine by IP injection.
9.3.1 Injection Vaccination
Injections are in general superior to any other vaccine appli-
The majority of commercial vaccines are administered to cation method, but they normally can only be applied to
fish by injection (normally intraperitoneal injection, IP). fish of 10 g or more (usually larger), although vaccination
This requires catching and anesthetizing the fish prior to machines are being developed for smaller fish. One major
 Fish Vaccines 115

Figure 9.1 Vaccination of sedated fish is performed by hand, normally by an injection team on site. Source: Photograph courtesy of
Pharmaq.

Figure 9.2 Fish are held ventral side up for vaccination and the needle is inserted into the peritoneal cavity using an automated
injection gun. Source: Photograph courtesy of Pharmaq.

advantage of injection vaccination is that adjuvants can be
included and there are a good range of adjuvants commer-
cially available. Retention of antigens at the injection site is
believed to be a prerequisite for long‐term protection in fish,
also known as the depot effect (Evensen et al. 2005).
9.3.2 Immersion Vaccination
There are two application methods for immersion vaccina-
tion: dip and bath. ERM and Vibrio vaccines are routinely
administered to rainbow trout by immersion.
Dip vaccination is more widely used and involves
immersing small fish for a short time (30 seconds) in a
highly concentrated vaccine solution (one part vaccine to
nine parts water). Large numbers of fish can be vaccinated
using this method (up to 100 kg of fish per liter of vaccine)
and it is widely used for vaccination of fry from 1 to 5 g.
This method of vaccination is effective and provides rela-
tively good protection. It is, however, limited in that there
is a short duration of immunity (approximately 3 months)
and a booster vaccination is required when the threat of
disease persists. This method is impractical for larger fish
116 Veterinary Vaccines: Principles and Applications
due to cost‐effectiveness and the stress of vaccination. In
addition, in fish smaller than 1 g, the immune system may
still be immature and therefore the vaccine efficacy may be
reduced.
Bath vaccination is used for larger fish and they are
exposed for a longer period, usually one to several hours, in
a lower concentration of vaccine (normally 1/100). Large
groups of fish are cut off from the rest in a cage and a low
dose of diluted anesthetic is added. Air or oxygen needs to
be continuously pumped in to avoid anoxia.
Following immersion vaccination, suspended antigens are
adsorbed by the skin and gills. Specialized cells, such as
antibody‐secreting cells, in the skin and gill epithelium are
activated and protect the fish when they are exposed to the
live pathogen at a later stage. Other cells in the epithelium of
skin and gills, such as antigen‐presenting cells (mac-
rophages), also absorb vaccine antigens and transport them
to specialized tissues where the systemic immune response
builds up.
9.3.3 Oral Vaccination
This is the most suitable method for mass vaccination but
the amount eaten by individual fish is uncertain and poor
potency can be a problem due to antigen destruction in the
stomach. Thus, the vaccine needs to be protected in some
way. Vaccine can be mixed with the feed, coated on top of
the feed (top dressed), or bio‐encapsulated. Stability in the
feed as well as stability due to destruction in the stomach
can both be an issue. Most vaccines are either incorporated
in an “antigen‐protecting vehicle” (Yersinia ruckeri, Vibrio
anguillarum, and IPNV vaccines, MSD‐Animal Health) or
in a patented MicroMatrix™ delivery system (Piscirickettsia
salmonis, ISAV, and IPNV, Centrovet) (Embregts and
Forlenza 2016). When antigens are to be incorporated in
feed, the heat sensitivity of the antigen has to be taken into
consideration. Potency can be affected due to the low pH of
the stomach. When vaccines are used as top dressing in
feed, a coating agent is usually applied, to prevent either
leaching of the antigen from the pellets or breakdown of
the antigen in the acidic environment of the stomach.
There are few oral vaccines on the market, and currently
these are used as booster vaccines. Administration meth-
ods for oral vaccines still require optimization.
9.4 ­Nonfish Vaccines
Crustaceans (shrimp) are a very important species group in
aquaculture with regard to value and volume. Although no
commercial vaccines are available for aquatic animals
other than fish (with the exception of the lobster gaffkemia
vaccine), there has been some research effort on the devel-
opment of shrimp vaccines. The mode of action of these is
not fully known and it is thought that they could simply be
stimulating the shrimp rather than vaccinating them as
such (Musthaq and Kwang 2014). Further research to
determine which alternative protection mechanisms can
be found in crustaceans (especially shrimp) should be pur-
sued. Development of specific pathogen‐free (SPF) and
specific pathogen‐resistant (SPR) stocks are currently pro-
viding some hope for control of viral pathogens, but com-
plete protection in open systems is currently not possible.
9.5 ­Immune Response to Vaccines
Fish differ from mammals in that they lack bone marrow
and lymph nodes (Press and Evensen 1999). The major lym-
phoid tissues in teleost fish are the (head) kidney, thymus,
spleen, and mucosa‐associated lymphoid tissues (Press and
Evensen 1999), including the gills (Haugarvoll et al. 2008),
skin (Xu et al. 2013), and nostrils (Tacchi et al. 2014).
The fish immune system comprises both innate and
adaptive immune responses, as in all vertebrates, with the
latter playing the key role in providing protection following
vaccination. A detailed review of the adaptive immune sys-
tem in teleost fish and how this responds to vaccination has
been recently published by Secombes and Belmonte (2016).
The adaptive immune response is mediated by T and B
lymphocytes, with T cells produced in the thymus and
migrating to other tissue sites to induce responses. The B
cells are produced at different sites in different vertebrate
groups; in teleost fish this is mainly the kidney (akin to the
bone marrow in mammals). Other differences between
mammals and fish include immunoglobulin (Ig)
classes – fish have no IgG, but instead have mainly IgM,
IgD, and IgT. Significant advances have been made in
understanding the fish immune system, with detailed
knowledge of many of the cytokines involved in its regula-
tion and assays developed to measure immune responses to
vaccination and infection. This has assisted in the develop-
ment of new vaccines.
9.6 ­Future Prospects
and Challenges for Fish Vaccines
Fish vaccines have been very successful in reducing the use
of antibiotics in the salmonid industry but additional vac-
cines are required for other fish species. Although the
number of fish vaccines commercially available has grown
in recent years, there are still numerous diseases where no

vaccines are available or cases where existing vaccines do
not perform well. Fish vaccine development is a very active
research area (Evensen 2016). It is possible to measure with
precision the responses elicited by vaccination (Secombes
and Belmonte 2016), thus assisting the development of
new vaccines for the future. The most crucial step in devel-
oping an effective vaccine is identification of “potentially”
protective antigens and confirming their protective
response in the host species by efficacy testing. The most
effective approach taken depends on the type of pathogen
and the final end‐use envisaged for the vaccine (e.g. cost,
fish species, immersion versus injection).
Fish vaccines have in general become much more sophis-
ticated in recent years. Technologies such as recombinant
and DNA vaccines are powerful tools for vaccine develop-
ment as these enable the separation of potential protective
antigens from suppressive ones. These are being developed
because the simpler approach of using inactivated whole
cell vaccines did not succeed for many important diseases,
and attempts at developing attenuated vaccines in general
have not been encouraged from a safety point of view.
Recently, the use of DNA vaccines has been authorized in
Europe, representing a major step forward. In addition,
there is much current research focused on the develop-
ment of mucosal vaccines (immersion and oral) and novel
vaccine strategies following the discovery of IgT as a
mucosal antibody in fish (Zhang et al. 2010, 2011).
Discovery and optimization of the use of novel adjuvants is
another area where improvements can be made. A variety
­References
Alonso, M. and Leong, J.A. (2013). Licensed DNA vaccines
against infectious hematopoietic necrosis virus (IHNV).
Recent Pat. DNA Gene Seq. 7 (1): 62–65.
Assefa, A. and Abunna, F. (2018, 2018). Review article:
maintenance of fish health in aquaculture: review of
epidemiological approaches for prevention and control of
infectious disease of fish. Vet. Med. Int.: 5432497.
Cowan, G., Smith, P., and Christofilogiannis, P. (2016). Fish
vaccines: the regulatory process and requirements from the
laboratory bench to a final commercial product, including
field trials. In: Fish Vaccines (ed. A. Adams), 105–118.
Basel: Springer.
Embregts, C. and Forlenza, M. (2016). Oral vaccination of
fish: lessons from humans and veterinary species. Dev.
Comp. Immunol. 64: 118–137.
Evensen, O. (2016). Development of fish vaccines: focusing
on methods. In: Fish Vaccines (ed. A. Adams), 53–74. Basel:
Springer.
Fish Vaccines 117
of adjuvants exist for use by injection but none have so far
been effective by immersion. In addition, the injectable
adjuvants still cause concern with regard to fish welfare
due to adhesions forming between organs in the fish, a
long period after vaccination.
9.7 ­Summary
In conclusion, although many fish vaccines are commer-
cially available and are effective, improvements are still
required for some of the traditional inactivated vaccines
with regard to vaccine efficacy (identification and optimi-
zation of antigen components), improved adjuvants, and
oral administration. Development of successful vaccines
against intracellular bacterial pathogens and viruses may
require the use of live attenuated vaccines and application
as oral vaccines, although there are safety concerns with
the use of live vaccines in the aquatic environment.
Vaccines against parasites and fungi‐like organisms are
also in development and these may need to rely on recom-
binant or DNA vaccine technology. There is also still a
requirement for basic information on pathogenesis,
immune response, and identification of potentially protec-
tive antigens for parasites and fungi. Autogenous vaccines
are currently used in aquaculture, for example to prevent
rainbow trout fry syndrome, and these are now also being
developed against parasite diseases in Atlantic salmon
(e.g. AGD).
Evensen, O., Brudeseth, B., and Mutoloki, S. (2005). The
vaccine formulation and its role in inflammatory processes
in fish – effects and adverse effects. Dev. Biol. (Basel) 121:
117–125.
FAO (2016). The State of World Fisheries and Aquaculture.
Contributing to Food Security and Nutrition for All. Rome:
FAO. www.fao.org/3/a‐i5555e.pdf
FAO (2018). The State of World Fisheries and Aquaculture
2016. Meeting the Sustainable Development Goals. Rome:
FAO. www.fao.org/3/i9540en/I9540EN.pdf
Haugarvoll, E., Bjerkas, I., Nowak, B.F. et al. (2008).
Identification and characterization of a novel
intraepithelial lymphoid tissue in the gills of Atlantic
salmon. J. Anat. 213: 202–209.
Klesius, P.H. and Pridgeon, J.W. (2014). Vaccination against
enteric septicemia of catfish. In: Fish Vaccination, 1e (eds.
R. Gudding, A. Lillehaug and O. Evensen), 25. Chichester:
Wiley Blackwell.
118 Veterinary Vaccines: Principles and Applications
Musthaq, S.K. and Kwang, J. (2014). Evolution of specific
immunity in shrimp – a vaccination perspective against
white spot syndrome virus. Dev. Comp. Immunol. 46:
279–290.
Press, C.M. and Evensen, O. (1999). The morphology of the
immune system in teleost fishes. Fish Shellfish Immunol. 9
(4): 309–318.
Rodger, H.D. (2016). Fish disease causing economic impact in
global aquaculture. In: Fish Vaccines (ed. A. Adams), 1–34.
Basel: Springer.
RUMA (2006). Responsible Use of Vaccines and Vaccination in
Fish Production, 1–23. Leominster: RUMA. www.ruma.org.
uk/fish/responsible‐use‐vaccines‐vaccination‐fish‐
production.
Secombes, C.J. and Belmonte, R. (2016). Overview of the fish
adaptive immune system in fish. In: Fish Vaccines (ed.
A. Adams), 35–52. Basel: Springer.
Tacchi, L., Musharrafieh, R., Larragoite, E.T. et al. (2014).
Nasal immunity is an ancient arm of the mucosal immune
system of vertebrates. Nat. Commun. 5: 5205.
Xu, Z., Parra, D., Gomez, D. et al. (2013). Teleost skin, an
ancient mucosal surface that elicits gut‐like immune
responses. Proc. Natl. Acad. Sci. U.S.A. 110: 13097–13102.
Zhang, Y.A., Salinas, I., Li, J. et al. (2010). IgT, a primitive
immunoglobulin class specialized in mucosal immunity.
Nat. Immunol. 11 (9): 827–835.
Zhang, Y.A., Salinas, I., and Oriol Sunyer, J. (2011). Recent
findings on the structure and function of teleost IgT. Fish
Shellfish Immunol. 31 (5): 627–634.