An immunostimulant is a naturally occurring substance that simulates the immune system of host against pathogens (Barman and Nen 2013). Immunostimulants can be divided into several groups depending on their sources: bacterial, algae-derived, animal-derived, nutritional factors, and hormones/cytokines (Sakai 1999). Recently, polysaccharides from seaweeds have also been tested as immunostimulants for shrimps (Declarador et al. 2014) (Fig. 6). They are broad range in efficacy and depict multitude of functions (Fig. 7). The multifarious antivirals available from variety of life forms are now discussed below.
Plants serve as repository of secure and economic chemical compounds that depict various properties such as growth promoters, immunostimulants, and antimicrobials (Citarasu et al. 2002). These have an upper hand because they consist of compounds that are non-toxic, biodegradable, and biocompatible (Citarasu et al. 2006). Underneath effort was done to provide a detailed insight into various forms of immunostimulants and chemicals derived from plants that have been reported as antiviral against WSSV.
Antiviral activity of genipin
Genipin (GN) is a bioactive compound obtained from the fruits of Gardenia jasminoides (Huang et al. 2019). GN is an aglycone obtained from an iridoid glycoside called geniposide. It is a natural cross-linker that has low toxicity and excellent biocompatibility. Therefore, it is used for synthesizing diverse biological polymers for drug delivery purpose (Sung et al. 2001; Mahgoub et al. 2017). It depicts anti-inflammatory, neuroprotective, antidiabetic, antiproliferative, antioxidative, and antiviral activities (Wang et al. 2017). Lately, the antiviral activity of GN against WSSV has been illustrated in crayfish Procambarus clarkii and in shrimp Litopenaeus vannamei. The infection caused by WSSV was impeded to the maximum level when the crayfish and shrimps were treated with 50 mg/kg of GN for a day. A detailed insight into the mechanism showed that GN deteriorates gene expression of signal transducer and activator of transcription (STAT), which in turn obstructs transcription of WSSV immediate-early gene, thus finally retarding WSSV replication. Moreover, it also caused the inhibition of Bax inhibitor-1 gene expression, which contributed further to inhibition of WSSV infectivity. Hence, GN is presumed to be an effective remedy to occlude WSSV infection (Huang et al. 2019).
Immunostimulant from various herbs
Oral administration of immunostimulants like lipopolysaccharides, etc. has shown increase in defensive effectiveness in WSD (Citarasu et al. 2003). Recently, a variety of herbal extracts have demonstrated their ability to augment immunity in shrimp against yellow head virus YHV and WSSV (Citarasu et al. 2003). Immunostimulants from 5 diverse herbal medicinal plants such as Eclipta alba, Aegle marmelos, Tinospora cordifolia, Picrorhiza kurooa, and Cyanodon dactylon depicted significant reduction in the viral load with increase in survival rate (74%) of shrimps (Citarasu 2010). Five herbs such as Adathoda vasica, Agathi grandiflora, Leucas aspera, Psoralea corylifolia, and Quercus infectoria have been used to screen the antiviral and immunostimulant activity against WSSV using different organic polar and non-polar solvents. Ethyl acetate and methanolic extracts of A. grandiflora depicted strong antiviral and immunostimulant activities. Compounds such as 1,2-benzenedicarboxylic acid, diisooctyl ester in the A. grandiflora extract–enriched diets helped to control the WSSV in shrimps (Bindhu et al. 2014). The shrimp species which were not subjected to any immunostimulant completely died in 3 days while the one who were inoculated with A. grandiflora extract–enriched diets showed reduction in the cumulative mortality of shrimps up to 60–80% (Bindhu et al. 2014).
Antimicrobials from Argemone mexicana
A. mexicana is a species of poppy that has medicinal properties and used by people in parts of Mexico, western US, and India. A. mexicana is employed to cure numerous illnesses including tumors, lumps, skin diseases, inflammations, painful joints, leprosy, malaria, and microbial infections. Such ailments can be treated due to diversified chemical constituents of plant that includes various alkaloids, mexicanol, mexicanic acid, argemonic acid, and phenolics (Palanikumar et al. 2018). Its seeds contain argemone oil that is composed of alkaloids such as sanguinarine, dihydrosanguinarine, dehydrocorydalmine, jatrorrhizine, columbamine, and oxyberberine (Gunstone et al. 1977). Variety of other alkaloids like cheilanthifoline, berberine, cryptopine, muramine, protopine, sanguinarine, stylopine, and thalifoline have also been described in the recent years (Singh et al. 2010). Methanolic and ethanolic extracts of the plant have potent astringent, antioxidant hepatoprotective activities, and antimicrobial properties (Dash and Murthy 2011).
A. mexicana roots possess antioxidant activity and therefore can be used as restorative agent in pacifying oxidative stress–associated degenerative diseases (Perumal et al. 2010). The methanolic extract accelerates the wound healing in wounded rats because of the phytochemicals like triterpenoids, alkaloids, flavonoids, and tannins. The proPO system then stimulates the production of antioxidant enzymes such as reactive oxygen intermediates (ROI) and reactive nitrogen intermediates (RNI), antimicrobial peptides, and lysozymes that eventually lead to phagocytosis. The fraction of shrimp group kept in control conditions succumbed 100% death in just 4 days, while the shrimp group subjected to A. mexicana extracts depicted the survival up to 79%.
Antiviral activity of bis(2-methylheptyl)phthalate from Pongamia pinnata leaves
Pongamia glabra vent (family Leguminosae) is an aboriginal ethnomedical plant found throughout India, tropical Asia, and Northern Australia. It is a moderate sized, expeditiously growing, and resilient plant that thrives under wide range of agroclimatic conditions and commonly occurs around roadsides, river banks, tidal forests, and even coastal areas. Its dried leaves when stored in the stocked grains serve as insect repellent. Its leaves decrease nematode infections tremendously when used as part of manure. The leaves of the plant are popularly known for its digestive, laxative properties, curing piles, lacerations, and extra soreness too. The juice of the leaves can be consumed to reduce flatulence, dyspepsia, diarrhea, cough, leprosy, and gonorrhea. The bark and roots of the plant are bitter in nature and therefore have anthelmintic properties (Al Muqarrabun et al. 2013). Feeding the purified ethanolic extract of P. pinnata leaves orally to WSSV-infected P. monodon increased the survival chances of the shrimps. Bis(2-methylheptyl)phthalate was responsible for the antiviral nature of P. pinnata. When WSSV-infected shrimp were fed with 200 g and 300 g extracts of P. pinnata, their survival rates improved by 40% and 80%, respectively (Rameshthangam and Ramasamy 2007).
Immunomodulatory effect of polysaccharide gel extracted from Durio zibethinus
D. zibethinus is the most prevalent tree species in the genus Durio most commonly found in South Asia especially Thailand. Its fruit is brown-green in coloration and exhibits presence to massive spiny thorns. The durian-rind waste is economically viable and produces polysaccharides with pharmaceutical and therapeutic uses (Pongsamart et al. 2005). The polysaccharide gel (PG) of durian rinds has been extracted, refined, and characterized (Hokputsa et al. 2004), and is now known to be a water-soluble pectic polysaccharide. The structure was deciphered and detail introspection revealed that the structure contains polygalacturonic acid with side chain made up of neutral sugars such as rhamnose, arabinose, glucose, galactose, and fructose. Regarding the bioactivity of the PG, it was discovered that it is antibacterial and also possesses wound-healing activities and immunomodulating properties (Chansiripornchai and Pongsamart 2008). Hence, PG extract from Durio zibethinus was tested as an immunostimulant to check the immunocompetency in shrimp against WSSV.
Shrimps that were given 1 and 2% of PG as diet for a period of 12 weeks demonstrated accelerated proPO activity as well as elevated values of THC (P < 0.05) when compared to the control group that was fed the basal diet without PG. Such results clearly suggested that the shrimp population in which PG-supplemented diet was orally administration emerged to be resistant to WSSV. Hence, the observed disease resistance in shrimps against WSSV may be attributed to the aggravated immune response of shrimps. Shrimps fed with 2% PG-supplemented diet indicate disease resistance of 36% (at day 4) and 100% (at day 6) (Pholdaeng and Pongsamart 2010).
Antiviral extract from Cyanodon dactylon
In another study, antiviral plant extract derived from Cyanodon dactylon was injected both by in vitro as well as in vivo process. In in vitro method, the extract was injected intramuscularly while in in vivo process, it was fed orally at concentration of 2 mg per animal. When C. dactylon plant extract was inoculated in WSSV-infected shrimps, then proPO, O2−, and NO got considerably high (P < 0.05) in values. Such evidence strongly suggests that inoculation of C. dactylon plant extract both by in vivo and in vitro procedures into shrimps enhances immunity of the shrimps. Additionally, the plant extract is very easily accessible and can serve as an excellent pre-emptive agent against WSSV disease in shrimps (Balasubramanian et al. 2007).
Phytochemicals as immunostimulants
An assortment of heterocyclic synthetic compounds like benzisoxazoles and piperidines has been found to play significant roles against microbial pathogens (Rajashekar Reddy et al. 2016). Furthermore, in silico studies provide a deeper knowledge for developing novel inhibitor against WSSV. The computational studies divulge that phytocompounds 2H-1-benzopyran-6-ol, 3,4-dihydro-2,5,7,8-tetramethyl-2-(4,8,12-trimethyltridecyl)-acetate, and 1,4-benzenediamine, N,N0-diphenyl identified from Phyllanthus amarus can serve as candidate molecules for WSD management (Dinesh et al. 2017). Another phytochemical known as quercetin has been predicted to be the suitable lead as it interacts with PmRab7 (one of the most crucial shrimp receptor proteins). Hence, use of this compound during maintenance of shrimp population in ponds, tanks, or natural reservoirs might diminish the interactions between Peneaus monodon PmRab7 and the viral proteins (Joseph et al. 2017).
A polyphenolic extract of red raspberry (Rubus idaeus) has shown to cluster viral particles cohesively to form non-infectious aggregations (Fukuchi et al. 1989). The polyphenols have further been reported to get associated with the virus particles directly thereby inhibiting the viral absorption into the host. Moreover, numerous polyphenolic compounds have been found to decrease association between human immunodeficiency virus (RNA virus) and influenza virus (RNA virus) with the host cells. Recently, 2,4-di-tert-butylphenol extracted from Emilia sonchifolia (a medicinal herb) has been recognized to have anti-WSSV and anti-8YHV (Maikaeo et al. 2015) properties and, therefore, it has been suggested to be fed to shrimp in order to boost resilience against virus-induced disease (Citarasu et al. 2006).
Anti-inflammatory activities of coumarins, which are naturally occurring pharmacologically active chemicals, are infinitely strong and efficient (Liu et al. 2020). This structure is made up of a benzo-pyranolactone moiety. Coumarins are divided into four categories: (i) simple coumarins, (ii) furanocoumarins, (iii) pyranocoumarins, and (iv) pyrone-substituted coumarins. They have a wide range of non-covalent interactions that allow them to bind to a variety of active sites. Some of the diversified functions in which coumarins majorly contribute are as follows: antioxidants (Minhas et al. 2017), antibacterials (Dastan et al. 2016), anti-inflammatory (Sahu et al. 2017), anticancer (Zhang et al. 2019), and antiviral (Kostova et al. 2006). Anti-diabetic drugs, HIV, cancer, and degenerative disorders may all benefit from the use of hydroxycoumarin derivatives in animals (Shen et al. 2010). There is a lot of potential for coumarins to be employed as antiviral medications in aquaculture because of their widespread usage as phytochemicals. Carp virus (SVCV) expression in host cells may be inhibited by 7-[6-(2-methylimidazole)-hexyloxy] coumarin (a coumarin derivative), which has been used as a potent antiviral medication (Chen et al. 2018). Epithelial papulosum cyprinid cells may be successfully treated with imidazole-coumarin compounds with four-carbon linker lengths (Liu et al. 2020). A coumarin derivative, chromene-3-carbonitrile widely known as C3007 used in Litopenaeus vannamei larvae, was tested for its anti-WSSV function. More than half of WSSV-infected larvae were able to survive for longer after taking C3007. Continued immersion in C3007 resulted in a 20% decrease in mortality after a 24-h WSSV infection. Hence, coumarin and its derivatives can be a viable treatment and prevention option for farmed shrimp WSSV infection.
Antivirals from bacteria
Other alternatives used as protection against WSD were oral inoculation of baculovirus and Bacillus subtilis spores (Syed Musthaq et al. 2009; Nguyen et al. 2014). RNA interference (RNAi) was also used to restrain the expression of viral proteins for inducing immunity in the host against WSD. It is elusive that M. japonicus has a potential to generate small interfering RNA (siRNA), which targets vp28 (vp28-siRNA) in reaction to WSSV infectivity. This further indicates that RNAi has a shielding effect for the host (Huang and Zhang 2013). In another contemporary work, when vp28-siRNAs encapsulated with β-1,3-d-glucan were injected along with WSSV in M. japonicas (Zhu and Zhang 2012), there occurred tremendous inhibition of WSSV replication.
The studies made till date laid a lot of importance on increasing the immune competency of the host. It can be done by catering the host with agents containing pathogen-associated molecular patterns (PAMPs). Studies have illustrated that feeding P. japonicus with peptidoglycans over a time span has escalated the phagocytic activity of host granulocytes therefore significantly decreasing mortality of P. japonicus upon WSSV exposure (Itami et al. 1998). Similarly, injecting β-glucan in shrimp prior to WSSV infection activated prophenoloxidase system and consequently caused reduction of morbidity in shrimps up to 25–50% when compared to control system (Song et al. 1994). They boost innate immunological responses such as phagocytic activation, neutrophil activation, alternative complement system stimulation, and enhanced lysozyme activity (Kyu et al. 2014).
Vibrio is a rod-shaped gram-negative bacteria and has been documented to reduce mortality in shrimps (Itami et al. 1989). The non-specific immune cells of shrimps such as phagocytic hemocytes are activated (Sakai 1999) by use of Vibrio. The bacterium Vibrio harveyi was reported to defend P. monodon against WSSV infection. Another important bacterial component is lipopolysaccharide (LPS). LPS are the major component of the outer membrane of gram-negative bacteria that can stimulate immune responses in different hosts. They are constituted by lipid and a polysaccharide that are joined by a covalent bond. The lipid in conjunction with polysaccharide is considered to be responsible for the biological activities of LPS (Rietschel et al. 1993). The immunostimulatory activity of LPS has been depicted in shrimps. Shrimps fed with LPS-containing diet depicted higher survival rates in comparison to the one fed with normal diet. Receptors such as anti-lipopolysaccharide factor 3 (ALF3), C-lectin, and mucin-like peritrophin located on shrimp digestive tracts were induced on using LPS-containing diet. Thus, LPS-supplemented diet was considered to be an efficient immunostimulant for Peneaus monodon (Rungrassamee et al. 2013). Another form of immunostimulants can be live bacteria. When shrimp feed is supplemented with probiotic bacteria, the cellular and humoral components of the innate immune system of shrimps get stimulated. Immunostimulation of Penaeus monodon by Bacillus S11 bacteria boosted phagocytic activity. Lactobacillus plantarum treatment increased phenoloxidase and superoxide dismutase activity, resulting in improved Vibrio alginolyticus clearance efficiency in Litopenaeus vannamei (Chiu et al. 2007).
Antivirals from algae
Seaweeds are multicellular algae that have been conceded to be a significant resource to develop resistance against pathogens for aquaculture because of prevalence of bioactive compounds, such as laminarin, fucoidan, carrageenan, and alginate that act as immunostimulants for the aquatic species. Furthermore, brown seaweeds have lately been reported as a source of polysaccharides and exhibit property of therapeutic agents and antibiotics for aquatic species. Fucoidan containing extracts from Sargassum spp. and from Cladosiphon okamuranus seaweed provide some resistance to WSD in Penaeus monodon and Marsuspenaeus japonicus (Immanuel et al. 2012), respectively. Recent studies have revealed that fucoidan plays an imperative role in the defensive mechanism of shrimp through mediating the cellular and the humoral responses in order to confront with the virus. It hampers the absorption of viral particles and facilitates the genesis of virus-induced syncytium. Feeding fucoidan orally is presumed to have defensive effects through inhibition of viral replication directly as well as triggering the innate immune defense responses.
It is seen that sodium alginate obtained from brown algae Undaria pinnatifida and Macrocystis pyrifera has potential to aggravate the nonspecific defense response of common carp Cyprinus carpio to Edwardsiella tarda (Fujiki et al. 1994). In similar manner, alginate obtained from brown algae U. pinnatifida and Lessonia nigrecans boosts Litopenaeus vannamei resistance to V. alginolyticus. Another study showed that sodium alginate availed from brown seaweed Sargassum wightii and primed in form of powder/beads was used to supplement Artemia nauplii. Such alginate-enriched nauplii when catered to Penaeus monodon could potentially impede succession of WSSV disease. When mortality between alginate-fed shrimp groups and control groups was compared, it was detected that mortality increases in alginate-fed groups by 26.5 to 58.4%.
More recently, a screening study was performed to check efficiency of Cereops tagal mangrove extracts against WSSV activity in P. monodon (Sudheer et al. 2011), and Litopenaeus vannamei immersed in the seawater containing Sargassum hemiphyllum var. chinensis powder. A significant increase in the defensive mechanism of such P. monodon (Sudheer et al. 2011) and Litopenaeus vannamei was depicted with increased protection against Vibrio alginolyticus and WSSV (Huynh et al. 2011).
Antivirals from fungi
A marine yeast Candida aquaetextoris S527 was used as diet for P. monodon with varying proportions, viz., single dose every day, once in 3 days, once in 7 days, and once in 10 days followed by challenge with WSSV. Following induction of this feed, the immune parameters such as total hemocyte count, pro-phenoloxidase, nitroblue tetrazolium reduction, and acid phosphatase activity were tested on the shrimps. The expression profiling of immune-associated proteins and pathways such as antimicrobial peptide (AMP) genes, alpha-2-macroglobulin (a-2-M) genes, prophenol oxidase (proPO) genes, and WSSV genes was further analyzed. The investigation established that when the yeast feed was inoculated one time every 7 days, then it worked as a superior immunostimulant against WSSV infection (Babu et al. 2013).
There are a wide range of components in yeast that are of nutritional benefit to humans and other animals, including proteins, carbohydrates, lipids, vitamins, and minerals. Yeast extracts are made up of water-soluble components of yeast cells, such as amino acids, peptides, carbohydrates, and salts, which are their primary constituents. Because of their nutritional properties, nitrogen components and vitamins are of the utmost economic significance. In recent years, yeasts have been shown to exhibit antioxidant and immunostimulating properties. Bakery and brewer’s yeast produce naturally occurring polysaccharides containing glucose as a structural component, connected by β-glycocidic linkages (Saccharomyces genus). Saccharomyces cerevisiae, the yeast used in the brewing process, contains a variety of compounds, including the sugars glucans and nucleic acids as well as the mannan-oligosaccharides that have been shown to improve immune responses and growth in a variety of fish species (Ortuño et al. 2002). Brewer’s yeast β-glucan (BYG) has been shown to have enhanced phenoloxidase activity, and oral treatment of BYG for 3 days to black tiger shrimp (Suphantharika et al. 2003) boosted their ability to kill Vibrio harveyi (Thanardkit et al. 2002). Pacific Food supplemented with an inactive yeast cell wall had no effect on the shrimp’s weight, survival rate, or growth rate; yet they exhibited stronger immunological measures (total hemocyte count, clearance of microorganisms) than shrimp given a standard diet (Chotikachinda et al. 2008).
Antivirals from animals
In some studies, animals have also been utilized to extract antivirals in order to cure WSSV in shrimps. In a recent study, cathelicidin 5, a type of peptide was obtained from Alligator sinensis and used as antimicrobial peptide against WSSV in caridean shrimp Exopalaemon modestus. Shrimp medicated with cathelicidin 5 and injected with WSSV revealed an appreciably lower mortality rate, lower viral VP28 amplification rate, and improved antioxidant enzyme activity and immune-related gene expression in shrimp Exopalaemon modestus (Xie et al. 2019).
Furthermore, the compound 3-(1-chloropiperidin-4-yl)-6-fluoro benzisoxazole 2 has been identified as a suitable antiviral curative agent in freshwater crabs Paratelphusa hydrodomous against WSD (Rajashekar Reddy et al. 2016). The same compound was tested against WSSV in shrimps by employing in silico methodologies such as molecular docking and molecular dynamics. The computational analysis revealed that the inhibitor binds to the polar amino acids that line the pore region of the envelope proteins. The delineation of the inhibitor molecule within the binding pocket is much plausible due to the low binding energy against VP26 and VP28. This makes the inhibitor a preferred antiviral lead against WSSV (Rajashekar Reddy et al. 2016).
Similarly, chitosan has been proposed to be a good immunostimulant. Chitin is a polysaccharide prevalent in insects, crustacean exoskeletons, and fungal cell walls, and is one of the most ubiquitous polysaccharides in nature (Esteban et al. 2005). Chitosan is derived from the alkaline deacetylation of crustacean chitin. Chitin and chitosan have been shown to protect fish and shrimp from bacterial infection when given as an injection, immersion, or dietary supplement (Siwicki et al. 1994).
Immunostimulants from synthetic sources
Currently, nanotechnology is being used to study the biological systems in much intricate detail at the atomic level (Parveen et al. 2012; Esmaeillou et al. 2017). Silver nanoparticles (AgNPs) are presently serving as very promising nanomaterials because they render comprehensive antiviral property (Galdiero et al. 2011; Bello-Bello et al. 2018; Chris et al. 2018). Multifarious studies have confirmed that AgNPs are a potential antiviral substitute against human viruses, including human immunodeficiency virus (Lara et al. 2010), H1N1 influenza A virus, monkeypox virus (Rogers et al. 2008), Tacaribe virus (Speshock et al. 2010), and herpes simplex virus (Baram-Pinto et al. 2009).
Many studies have elucidated wide usage of AgNPs in many intricate details against enveloped viruses and such studies have confirmed that AgNPs preferentially bind to the viral envelope glycoproteins to hinder the binding between viruses and host cell (Lara et al. 2011; Bogdanchikova et al. 2016). Additionally, immune system can recognize the nanoparticles very well and these particles help in inducing immunostimulatory effects in the immune system (Boraschi et al. 2017; Dacoba et al. 2017).
A pivotal step taken in applying nanotechnology in aquaculture was making use of AgNP-based formulations. One such formulation, Argovit R, was inoculated in the form of intramuscular injections into L. vannamei (Romo-Quiñonez et al. 2020). An observation was deduced that silver nanoparticle single dosages can alone enhance the shrimp survival tremendously without toxic effects, thus proving its antiviral activity against WSSV. Another study evaluated that Argovit R promoted the immune response of shrimp infected with WSSV even under unfavorable circumstances, for example elevated concentration of Fe+2 ions in the medium (Ochoa-Meza et al. 2019). Thus, AgNPs can be used as an effective agent against a large heterogeneous form of viruses, especially against WSSV, although intramuscular administration of AgNPs is practically not feasible due to large volumes of organisms that need to be treated. A realistic substitute for intramuscular administration of AgNPs is incorporating AgNPs in the form of feed (Dananjaya et al. 2016). This will assure the applicability of AgNP to evade WSSV infection in shrimps. A new formulation named Argovit-4 (that essentially retains same design of Argovit R) has been recently reported as a propitious antiviral additive in feed to interfere with WSSV infection in shrimp. The most characteristic attributes that contribute to make Argovit-4 as a potential candidate for antiviral therapy in shrimps are (i) its decreased/increased concentrations do not cause toxicity to shrimps, (ii) intramuscular administration of a mixture of WSSV-Argovit-4 potentially decreased shrimp mortality at least by 50%, and (iii) it shows unaltered expression of PEN4, PAP, Crustin, and Rab6 genes suggesting that it does not interfere with immune system of shrimps (Romo-Quiñonez et al. 2020).
Immunostimulants from nutritional factors
There was a marked increase in resistance of shrimps against the pathogens when they are injected with vitamins A, C, and E supplements (Lee and Shiau 2004). Similarly, carotenoids depict functions such as lymphocyte blastogenesis, lymphocyte cytotoxicity activity, stimulate the production of certain cytokines, and aggravate the phagocytic and bacterial killing ability of neutrophils and macrophages. In shrimps, dietary carotenoid supplementation enhanced stress resistance, salinity shock resistance, and antioxidant response before and after viral infection. Trace elements such as incorporation of copper in tiger shrimp resulted in escalated growth, total hemocyte count, and superoxide anion production. In white shrimp, nutritional supplementation with Zn improved immunological responses in a similar way (Lin et al. 2013).
Immunostimulants from hormones
Growth hormone (GH) is a peptide hormone (also known as somatotropin or somatropin) that has numerous beneficial effects such as promotion of lipolysis, gluconeogenesis, increase in protein synthesis and stimulation of immune responses among others. In shrimp larvae, a recombinant bovine growth hormone has been demonstrated to improve development and immunity. In Macrobrachium rosenbergii, dietary supplementation of bovine lactoferrin (LF) improved immunological indices and resistance to Aeromonas hydrophila challenge in crustaceans (Chand et al. 2006).