A. P. Desbois
4.2 VACCINES
50 Fish Vaccines
Concepts and Types of Vaccines 51
to the evolutionary system. There are more similarities than differences although fish immune sys- tems are primitive compared to mammals. Furthermore, there are more than 25,000 species of fish living in habitats ranging from muddy fresh water to the ocean, from polar regions to the tropics, from dark high-pressure depths to the bright low-pressure surface areas. Therefore, their response to vaccines and combating mechanisms against pathogens differ for different fish species. The study of both pathogen and vaccine-induced immunity is limited due to the lack of detailed knowledge of the immune systems in different fish species (4).
4.2.1.1 Fish Immunity
Innate immunity is more vital in fish than mammals against invading pathogens because the adap- tive immunity of fish is not as diversified; there are only three isotypes of immunoglobulins and some fish lack few major histocompatibility complexes (MHC) (12).
More than 550 million years ago, the adaptive immune system emerged along with the evolution of jawless vertebrates. Only variable lymphocyte receptors are used to recognize antigens in jaw- less vertebrates due to lack of T- and B-cells, while in jawed vertebrates, immunoglobulins (Ig) are involved in the recognition of pathogens in the process of immune exclusion (13). Immunoglobulin T is one of the key molecules of jawed vertebrate’s adaptive immunity (14).
To date, five immunoglobulin isotypes have been characterized in mammals, namely, Ig M, IgD, IgG, IgE, and IgA, whereas in fishes only three isotypes were well characterized, namely, IgM, IgD, and IgT or IgZ (15). IgT plays important roles in mucosal immunity in the gut (16), skin, and gills (17,18) of fish (19). Nayak and Nakanishi (2013) demonstrated the direct antibacterial activity of CD8+ and CD4+ T-cells and surface IgM+ cells in fish (20). The interbranchial lymphoid tissue (ILT) of Atlantic salmon originates from an embryological location that in higher vertebrates acted as origin for the development of primary and secondary lymphoid tissues (21). The skin-associated lymphoid tissue (SALT) is a mucosal lymphoid tissue in fish that is different from that of mammals that is the outermost cell layer alive and actively dividing. The SALT protection is efficient due to the high concentration of immunoglobulin M (22).
The mucosal surface of the fish digestive tract is covered with immunoglobulins, complement factors, antimicrobial peptides, and other surface defensins beneath which several types of immune cells are found in fish gut, which provides innate immunity to fish (23). The CD4 cell surface marker plays an important role in distinguishing between T-helper cell and cytotoxic T-cell (24).
Oral vaccines induce fish immune system selectively by activating CCL25/CCR9 ligand/receptor system in innate immunity (25,26). In fish, central and peripheral immune tolerance is important in the gills, where the intimate contact between gill tissue and the aqueous environment would other- wise lead to continual immune stimulation by innocuous antigens (27).
4.2.1.1.1 Fish Immune Organs
The organization of immune organs in fish is slightly different from higher vertebrates. The main difference is that fishes lack bone marrow and lymph nodes and the primary lymphoid organs are the thymus, the head kidney, or pronephros for teleosts, and the leydig and epigonal organs for chon- drichthyes, while secondary lymphoid organs are the spleen, the kidney, and mucosa-associated lymphoid tissue (MALT) present in peripheral immune tissues (28).
Bony fish (teleosts) have a complete set of lymphoid organs except for bone marrow, and they can mount both innate and adaptive immune responses. Their lymphoid organs are very different from those in mammals. For example, their thymus may involute in response to hormones or season. Age involution is inconsistent, and a thymus may be found in old fish. Fish kidneys differentiate into two sections. The opisthonephros or posterior kidney is similar to the mammalian kidney.
In contrast, the pronephros or anterior kidney is a lymphoid organ containing antibody-forming cells and phagocytes. Its function is analogous to mammalian bone marrow and lymph nodes. Fish have a spleen with a structure and function similar to that in mammals. Aggregates of lympho- cytes are prominent in the fish intestinal tract. Fish also have clusters of macrophages containing
52 Fish Vaccines melanin and hemosiderin. These melanomacrophage centers are found in the spleen, liver, and kidney. Antigens may persist in these centers for long periods, and they appear to be precursors of the germinal centers found in more evolved vertebrates. Teleosts also have dendritic-like cells that can present antigens to T-cells.
Fish lymphocytes resemble those of mammals. B-cells can be found in the thymus, anterior kid- ney, spleen, leydig organ, and blood, and their surface immunoglobulins act as antigen receptors.
These B-cells can mature into plasma cells. Unlike mammalian B-cells, however, teleost B-cells can phagocytose particles, generate phagolysosomes, and kill ingested microbes. Both helper and cytotoxic T-cells can be detected in fish (6).
4.2.1.1.2 Fish Immune Response
Immune cells recognize most pathogens and altered/infected cells through expressed pathogen- associated molecular patterns (PAMPS) or danger-associated molecular patterns (DAMPS), through nonself or MHC class I molecules and by presenting foreign nonself peptides of intracellular or extracellular origin. In order to destroy pathogens directly or by inhibiting their ability to replicate, specialized immune cells of the innate and adaptive responses were produced during evolution.
The first line of defense is represented by the evolutionarily ancient macrophages and natural killer (NK) cells. These innate mechanisms are well developed in bony fish.
Adaptive cell-mediated cytotoxicity (CMC) requires key molecules expressed on cytotoxic T lymphocytes (CTLs) and target cells. CTLs kill host cells and attract intracellular pathogens by binding of their T-cell receptor (TCR) and its co-receptor CD8 to a complex of MHC class I and bound peptide molecule on the host infected cell. Alternatively, extracellular antigens are taken up by professional antigen-presenting cells such as macrophages, dendritic cells, and B-cells to process those antigens and present the resulting peptides in association with MHC class II to CD4+ T-helper cells. Then, T-helper cells through activation of other immune cells eliminate the extracel- lular pathogens (29).
4.2.1.2 Vaccine Efficacy
Many factors affect the efficacy of vaccines. The factors include husbandry nature, handling method, fish age, smoltification, dominance hierarchies, pheromones, temperature, diet (vitamins and minerals), pollutants (metals and pesticides), seasonal variations, vaccine dose, vaccine route, nature of adjuvant usage, immunostimulants used, and antibiotics. Utmost care should be taken to achieve optimum levels of protective immunity when administering vaccines to minimize the effects of stressors (30).
4.2.1.3 Adverse Effects
Fish vaccines are no exception to the rule that adverse effects following vaccination are common problems. Vaccine producers considered adverse effects as a serious problem in recent years. The nature of delivery systems for oral or immersion administration would be an improvement to fish vaccination in the near future (4). Local injection site reactions remain an issue in aquaculture.
Intraperitoneal inoculation may lead to local or diffuse peritonitis with adhesions. Multiple granu- lomas may also develop. The fish produce multiple autoantibodies and develop immune-complex glomerulonephritis, liver thrombosis, and spinal deformities. Both polyclonal IgM and antibodies to salmon red blood cells are elevated in vaccinated fish (6).
4.2.2 HIstoryof VaccInes
The history of fish vaccinology is a documentation of how the immune system of fish can be stimu- lated by vaccines to prevent accidental effects of pathogenic microorganisms (31). The first attempts to immunize fish started date back to the 1930s, but the economic incentive to commercialize vac- cines was not until the mid- to late 1970s (30). Even though the fish vaccination process has a long
Concepts and Types of Vaccines 53
history, only in the past few decades it has become a well-established protective measure against bacterial infections (32).
The first commercially available bacterial vaccines were against enteric red mouth disease and vibriosis, introduced in the USA in the late 1970s. Fish immersion vaccines were found to be effec- tive against vibriosis in the USA. However, a new disease, furunculosis, appeared and injectable vaccines containing adjuvants were developed in the early 1990s.
The first viral vaccine was produced by Bioveta Company for fish in 1982. The vaccine was against a carp rhabdovirus, and was based on two inactivated strains of rhabdovirus emulsified in oil and administered by injection. Most available virus vaccines are based on inactivated virus or recombinant subunit proteins for aquaculture.
With the exception of the introduction of a recombinant virus vaccine in 1995, vaccination strat- egies in the fish farming industry have been more or less unchanged over the last 20 years. In the present scenario, trial and error has been the important strategy for the designing of fish vaccines.
Vaccination against the most serious bacterial diseases has been quite successful for the large-scale commercial farmed fish varieties with a few exceptions. The usage of modern sophisticated techniques is applied as a new trend for the development of new vaccines. The development of fish genomic stud- ies and their information can be incorporated for the invention of new vaccines. Usage of DNA vac- cines are safer than live attenuated vaccines but the legal issues related with the usage of DNA and genetically modified organism vaccines limited their availability. If only the method of administra- tion, efficacy, and cost of production were considered, live attenuated vaccine would be chosen as an optimal type of vaccine. But a consumer safety measure prevents the usage of live attenuated vaccine type because of the reversal of infectivity features (4). To date, there are many vaccines used to control infections such as killed vaccines, live attenuated vaccines, subunit vaccines, and DNA vaccines (33).
The field of fish vaccinology has shown an amazing development recently. The comprehensive scientific research and valuable practical experience are responsible for the production of first gen- eration fish vaccines, which will immensely contribute to social, environmental, and economical sustainability in aquaculture worldwide.