Introduction

Aquaculture has become an economic and safe source of protein for human consumption around the world. Global food fish production has been increasing at an average annual rate of 6.6% since 1995 (FAO 2017) and reached 80 million tons in 2016 (FAO 2018). The production of Nile tilapia, salmon, and other freshwater species has led to a significant growth in annual per capita consumption, approximately from 1.5 kg in 1961 to 7.8 kg in 2015 (FAO 2018).

In intensive aquaculture, farmed fish can be affected by various infectious diseases worsened by stress factors which may lead to a decrease of fish resistance. Antibiotic prescriptions may be needed to avoid impaired growth performance and significant economic losses due to bacterial disease (Romero et al. 2012). In aquaculture, antibiotics were mainly added to feed supplement into water, resulting in the discharge of drug and their metabolites into the wastewater (Romero et al. 2012). Even when the antibiotic concentrations are well below the minimum inhibitory concentration, the prolonged presence of antibiotics in water, combined with high numbers of bacteria in the polybacterial matrices as the pond, sediment, or biofilm, may put selective pressure on bacterial populations and allow the exchange of antimicrobial resistance genes between bacteria (Baquero et al. 2008; Muziasari et al. 2016; Watts et al. 2017). The passage of antimicrobial residue, antimicrobial-resistant bacteria, and resistance genes from aquatic animals and their environment to terrestrial livestock and humans presents the increasing risk of a widespread emergence of drug-resistant pathogens (Rasul and Majumdar 2017; Santos and Ramos 2018).

Common infections in freshwater fish are caused by the genus Aeromonas. These bacteria are common inhabitants of aquatic animals (fish and shellfish) and aquatic environments such as freshwater, estuarine waters, marine waters, and sediments (Swann and White 1989). In fish farms, the two most frequently encountered species are Aeromonas hydrophila and Aeromonas salmonicida. A. salmonicida subsp. salmonicida mainly affects salmonids and is the causative agent of furunculosis. This disease is responsible for severe economic losses by haemorrhagic septicaemia in the acute form and by fish depreciation due to the development of boils in the muscles in the chronic form (Austin and Austin 2012). A. hydrophila is a ubiquitous bacterium which is commonly isolated from freshwater ponds and which is a normal inhabitant of the gastrointestinal tract of aquatic animals. It may also cause a disease in fish known as “haemorrhagic septicaemia” (Randy White 1991). A. hydrophila is also a zoonotic pathogen that infects humans via foodborne infections or through aquaculture facilities and is a public health hazard (Okocha et al. 2018). Aeromonas are opportunistic environmental pathogens of animals and humans. Genotyping analyses and antibiotic resistance profiles of the two main species A. salmonicida subsp. salmonicida and A. hydrophila demonstrated the presence of multidrug resistance plasmids with a high level of interspecies transfer, including human bacteria (Del Castillo et al. 2013; Vincent et al. 2014). Aeromonas may persist being attached to biofilms on biotic or abiotic surfaces, and the presence of these bacteria with E. coli in polybacterial mixed biofilms promotes the exchange and dissemination of antimicrobial resistance genes (Talagrand-Reboul et al. 2017). Limiting the emergence of antibioresistant Aeromonas and the transfer of their resistance genes by decreasing the antibiotic uses in aquaculture is therefore an issue for fish and public health.

To decrease the use of antibiotics, alternative strategies have been developed to improve fish health and aquaculture systems while reducing the potential for the spread of antimicrobial resistance. These include: (i) vaccination, by considering the difficulty of its application and its controversial effectiveness in fish populations (Gudmundsdóttir and Björnsdóttir 2007; Plant and LaPatra 2011), (ii) immune stimulation by using products derived from plants, bacteria or algae with effects on the microbiome and the immunity of the farmed host, (iii) phage therapy, and (iv) biosecurity approaches such as disinfection of water system (Watts et al. 2017).

In this review, we summarize the promising functional feed alternatives, such as probiotics, prebiotics, plants, essential oils, algae, and phages to reduce antibiotics consumption in aquaculture. The focus of this paper is mainly on their protective efficacies against the most frequent ubiquitous organism (Aeromonas spp.) when delivered in vivo in the three major families of freshwater fish, salmonids, cyprinids, and cichlids.

Methodology analysis to evaluate alternative products against Aeromonas spp. infection

The survey showed that the majority of studied cases of alternatives were carried on probiotics, plants, and prebiotics, respectively. The other alternatives studied are synbiotic (mixture of prebiotics and probiotics) essential oils, algae, bacteriophage, and other non-classified alternative families, like as mineral and nanoparticles. Alternative products were tested mainly on Nile tilapia (Oreochromis niloticus), rainbow trout (Oncorhynchus mykiss), rohu (Labeo rohita), and common carp (Cyprinus carpio).

Mostly, alternative products were tested for their preventive and protective effects. Some studies have also evaluated the curative effect of alternatives like probiotic or triherbal extract-enriched diets (Harikrishnan et al. 2010), aqueous methanolic extracts of tetra (Cotinus coggygria) (Bilen and Elbeshti 2019), and therapeutic phages (Imbeault et al. 2006; Kim et al. 2015). All investigations presented a comparative study in the present review which in the test groups, fish were treated with the alternative candidates and in the control/negative group, fish were not treated. Moreover, alternative products efficacies were sometimes compared with antibiotics (oxytetracycline) (Park et al. 2017; Won et al. 2017; Lee et al. 2016b).

To evaluate the preventive efficacy of functional feed alternative against Aeromonas spp. infections, pathogen was injected by the intraperitoneal (IP) route but fish were also exposed to Aeromonas spp. by immersion (Bandyopadhyay and Das Mohapatra 2009; Liu et al. 2013b), by cohabitation (Irianto and Austin 2003; Hoque et al. 2018; Menanteau-Ledouble et al. 2017), or by oral intubation (Ngamkala et al. 2010; Dong et al. 2018). A. hydrophila was mainly used to infect freshwater fish, with the exception of rainbow trout mainly infected with A. salmonicida. Various infection doses were investigated in challenge experiments that depended mainly on bacterial strain, fish species, administration routes and the survival rate required by the authors. Indeed, different infectious doses could lead to a same RPS. For example, A. hydrophila infection dose at 103 and 109 CFU ml−1 injected by IP route in Mozambique tilapia induced a RPS of 10% (Rajeswari et al. 2016; Suguna et al. 2014). In contrast, a similar infectious dose could lead to very different RPS (A. salmonicida doses at 2.4 107 and 2.107 CFU ml−1 induced a RPS of 80 and 12%, respectively, in rainbow trout (Kim and Austin 2006; Park et al. 2017). The post-infection day duration after Aeromonas challenge should be also taken to account for the mortality rate records which might vary from hours to weeks, depending on the investigation conditions.

Probiotics

In an expert consensus document, the definition of a probiotic has been recently clarified as: “live microorganisms that, when administered in adequate amounts, confer a health benefit on the host” (Hill et al. 2014). In the interest of probiotics use in aquaculture, it was proposed to extend the definition to “living microbial additives that benefit the health of hydrobionts and therefore increase productivity” (Martínez Cruz et al. 2012). In aquatic species, the microbial community in gastrointestinal tract depends on the external environment including water and feed. Potential probiotic bacteria need to tolerate the temperature of pond water in addition to the bile salts and low pH detected in fish intestines. Potential probiotics must also improve feed utilization and growth by considering their viability under processing conditions when added to fish feed (Irianto and Austin 2002; Lacroix and Yildirim 2007). Moreover, other essential properties are defined relative to safety as a non-pathogenic microorganism and to the absence of plasmid-encoded antibiotic resistance (Martínez Cruz et al. 2012). The mechanistic basis and beneficial activities of probiotics previously were explained as being due to a modification of intestinal microbiota, production of antibacterial or antitoxin substances (bacteriocins and organic acids), modulation of the immune system and competition with pathogens for nutrients, and adhesion to intestines (Myers 2007).

The efficacy of potential probiotic bacteria has been extensively studied in which lactic acid bacteria (Lactobacillus spp., Lactococcus spp.) and Bacillus spp. were the most commonly used probiotic (Table 1). Saccharomyces cerevisiae yeasts have also a great promise as a potential probiotic substance (Abdel-Tawwab et al. 2008; Abdel-Tawwab 2012; Ran et al. 2015, 2016; Abass et al. 2018).

Table 1 Summary of in vivo studies in three freshwater fish species for probiotics and bacterial secondary metabolites or enzymes
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Among lactic acid bacteria and Bacillus spp., a large diversity of bacterial species and strains were evaluated. For example, for Bacillus spp., 11 strains belonging to 7 species have been investigated in this review (Table 1). Potential probiotic bacteria were provided from various sources, either bacteria isolated from fish in a local laboratory, commercial strains that were directly purchased like feed additives as Lactococcus lactis (Suprayudi et al. 2017) or even, final commercial product as “Organic green” composed of Lactobacillus acidophilus, Bacillus subtilis, Saccharomyces, and Aspergillus oryzae (Aly et al. 2008).

Probiotic products were administered orally as a feed supplement except some cellular components of probiotic bacteria which were injected intraperitoneally (Ramesh et al. 2015; Giri et al. 2015a, b, c; Ramesh and Souissi 2018). They were administered in a very wide range of dosages and durations, from milligrams to grams per kilogramg of feed, and for days to months before the infectious challenge. Generally, for S. cerevisiae yeasts, the optimal probiotic dose was proposed to be 1 to 2 g kg−1 diet from 56 to 84 days to protect Nile tilapia (O. niloticus) against A. hydrophila infections (Abdel-Tawwab et al. 2008; Abdel-Tawwab 2012; Ran et al. 2015, 2016) but increased to 70 g kg−1 under stress condition (Abass et al. 2018). For potential probiotic bacteria, the optimal dose varied between 107 and 1010 cfu g−1 diet for 2 to 3 months, depending on the species and strain of probiotic and the fish species (Table 1).

The increase of the survival rate and protection effect in the probiotic feeding group compared with the control group was a result of the preventive effect of these probiotics against Aeromonas spp. However, the amplitude of the survival rate between the probiotic and control groups varied greatly and depended on the probiotic species, the feeding dosages and durations, and the experimental infection (dose of bacteria, administration route, duration) (Table 1). In Catla catla, the effect of B. circulans depended on the probiotic dosage: the survival rate was 96.7% with 2 × 105 CFU 100 g−1 feed whether 40.0% with 2 × 106 CFU 100 g−1 feed and 6.7% in the control group (Bandyopadhyay and Das Mohapatra 2009). Besides dose-dependent effects, the duration of feeding fish with probiotics seemed to be an important matter to achieve a higher protection. For example, the relative level of protection against A. hydrophila in Nile tilapia for each probiotic agent Bacillus pumilus or mixture of Lactobacillus acidophilus, Bacillus subtilis, Saccharomyces, and Aspergillus oryzae showed to be higher at the end of the 2nd month than at the end of the 1st month of the feeding trial (Aly et al. 2008).

Due to the influence of many different factors on experimental results, it is difficult to compare the preventive effect of the different probiotics tested against Aeromonas spp. infection. However, some publications compared several probiotics under the same experimental conditions. P. polymyxa MTCC122 seemed to have a better protective effect against A. hydrophila than B. coagulans MTCC9872 or Bacillus licheniformis MTCC6824 in common carp (Gupta et al. 2014). Similarly, Lactobacillus brevis JCM1170 had a better efficacy than L. acidophilus JCM1132 against A. hydrophila in tilapia (Liu et al. 2013b). Furthermore, it has been demonstrated that the incorporation of multispecies probiotics of Saccharomyces cerevisiae, B. subtilis, and Lactococcus lactis (Mohapatra et al. 2014) or B. subtilis, L. plantarum, and P. aeruginosa (Giri et al. 2014) improves health status more effectively than the incorporation of a monospecies probiotic in the diet.

The preventive effect of probiotics against Aeromonas spp. could be explained in part by their immunostimulant effect. Paenibacillus polymyxa had a better immunostimulant effect than Bacillus coagulans MTCC9872 or Bacillus licheniformis MTCC 6824, which could explain the better protective efficacy of P. polymyxa against A. hydrophila (Gupta et al. 2014). However, in contrast with the survival rates, a combination of several probiotics did not seem to significantly increase the immunostimulant effect compared with a single probiotic (Park et al. 2017; Aly et al. 2008).

The duration of time that fish are fed probiotics seemed to be also an important factor on influencing the immunological parameters in fish. Several immunological parameters measured in mucus and serum were improved after 28 days but not after 14 days of B. licheniformis Dahb1 feeding (Gobi et al. 2018). Similarly, administering Bacillus aerophilus KADR3 over a 6-week period resulted in a slightly higher immunostimulant effect than over a three-week period (Ramesh et al. 2017). However, some studies have concluded that immunostimulation can be observed after a 30-day period of probiotic feeding, which is then followed by a declining trend (Giri et al. 2012, 2013, 2014; Mohammadian et al. 2016).

In addition to an immunostimulant effect, the administration of probiotics might protect against tissue lesions induced by Aeromonas. Histological analysis demonstrated that the severity of lesions in intestines and gills was less in rohu fish (L. rohita) fed with B. subtilis, L. lactis, and S. cerevisiae after the A. hydrophila challenge (Mohapatra et al. 2014). In addition, the intestines of Nile tilapia (O. niloticus) exposed orally to L. rhamnosus GG showed an increased inflammatory cell infiltration and reduced intestinal damages from A. hydrophila (Ngamkala et al. 2010). L. lactis 16-7 could also reduce intestinal mucosal barrier damage and inflammation induced by A. hydrophila by antagonizing the colonization of A. hydrophila in crucian carp intestine (Dong et al. 2018). Probiotics could also fortify the intestinal structure. Live baker’s S. cerevisiae yeast and Lactobacillus plantarum AH 78 increased microvilli length of fish intestine (Ran et al. 2015, 2016; Hamdan et al. 2016) and L. plantarum JCM1149 and AHL lactonase enzyme had a synergistic effect on the microvilli density (Liu et al. 2016).

Some studies have found that probiotics could also modify freshwater fish microbiota (Carnevali et al. 2017; Akhter et al. 2015; Dimitroglou et al. 2011). Dietary administration of the grass carp (C. idella) with Shewanella xiamenensis A-1, Aeromonas veronii A-7, and Bacillus subtilis for 28 days or Nile tilapia with Rummeliibacillus stabekisii for 8 weeks, induced benefic alteration of intestinal microbiota by increasing the abundance of Cetobacterium genus with potential immunity function, by reducing the abundance of the potential pathogenic bacteria and by promoting the reproduction of potential probiotics (Hao et al. 2017; Tan et al. 2019). In contrast, feed supplementation by either heat-inactivated or live commercial preparation of the baker’s yeast S. cerevisiae did not influence Nile tilapia (O. niloticus) gut microbiota markedly (Ran et al. 2016).

In addition, probiotics or their secondary metabolites might increase the health status of fish by increasing feed conversion and growth performance (Table 1). Among the different studies analyzed in this review which resulted to higher growth performance after probiotic feeding, there is only one report mentioned that administration of a S. cerevisiae, Bacillus subtilis, and Aspergillus oryzae mixture had no significant effect on growth rates while feed conversion was increased (Iwashita et al. 2015). Probiotic treatments can also have influence on body or organ content. A higher level of proteins and lipids was found in the carcass of fish fed with Bacillus circulans PB7 (Bandyopadhyay and Das Mohapatra 2009). Enterococcus faecalis supplementation also significantly enhanced the production of digestive enzymes in Javanese carp (Puntius gonionotus) intestine as well as the level of propionic and butyric acids (short-chain fatty acids) while no significant difference (P > 0.05) in acetic acid production was observed (Allameh et al. 2017).

Finally, probiotics could participate to stress control. S. cerevisiae-exposed Nile tilapia showed greater tolerance to stress induced by elevated water temperature (40 °C for 48 h) or by a 24-h hypoxia exposure compared with the control group (Abass et al. 2018).

Prebiotics

Prebiotics are non-digestible fibers that are selectively utilized by host microorganisms to confer health benefits and enhance growth performance due to the byproducts generated from their fermentation by gut commensal bacteria, such as changing the composition of the microbiota, inhibiting pathogens, stimulating immune responses and improving stress resistance (Gibson and Roberfroid 1995; Gibson et al. 2017, Ringø et al. 2010, 2014a, b; Patel and Goyal 2012). Prebiotics are defined by three criteria: (a) resistance to gastric acidity, hydrolysis by host enzymes and gastrointestinal absorption; (b) fermentation by intestinal microbiota; and (c) selective stimulation of the growth and/or activity of intestinal bacteria (Gibson et al. 2004). Prebiotics can be classified according to their molecular size or degree of their carbohydrates polymerization into oligosaccharides (inulin, fructooligosaccharides (FOS), mannanoligosaccharides (MOS)) or polysaccharides such as β-glucans (Ringø et al. 2010, 2014a, b; Patel and Goyal 2012).

Among prebiotics investigated to prevent disease in freshwater fish species by Aeromonas spp., β-glucan (β-1,3-glucan or β-1,6-glucan) have been paid attention extendingly (Anjugam et al. 2018; Ji et al. 2017; Douxfils et al. 2017; Falco et al. 2012; Barros et al. 2014; Ngamkala et al. 2010; Zheng et al. 2011), which is mostly isolated from the cell wall of the yeast S. cerevisiae. Commercial products which consisted of a mixture of β-glucan and MOS were also tested (Gupta et al. 2008; Yarahmadi et al. 2014, 2016; Ebrahimi et al. 2012). MOS (Liu et al. 2013a) and microbial levan as a fructan-polysaccharide (Rairakhwada et al. 2007; Gupta et al. 2008) have also been studied.

Generally, β-glucan products were administered orally and added to the basal diet as a feed supplement and seemed to be efficient in preventing the mortality associated with Aeromonas spp. infection, as represented by significant differences (p ≤ 0.05) in survival rate and protection effect between the prebiotic and control groups (Table 2). However, as seen with probiotics, the level of preventive effects depends on several factors such as dose and duration. However, feeding fish with β glucan at 1 to 2 g kg−1 diet for at least 2 weeks seemed to be optimal to high protection and immune response in different Aeromonas infected freshwater fish species including rainbow trout (O. mykiss), common carp (C. carpio), and Nile tilapia (O. niloticus) (Douxfils et al. 2017; Falco et al. 2012; Barros et al. 2014; Ji et al. 2017). In addition, combination of β-glucan and MOS (commercial product) resulted also in high disease resistance against A. hydrophila in rainbow trout (O. mykiss) (Yarahmadi et al. 2014, 2016) and common carp (C. carpio) (Ebrahimi et al. 2012); however, application of β-glucan and MOS alone has not been demonstrated by the authors.

Table 2 Summary of in vivo studies in three freshwater fish species for prebiotics
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The immunostimulant effect of prebiotics is well-known and many studies have indicated that immunosaccharides as β-glucan FOS, MOS, or inulin are beneficial to aquatic animals (Das et al. 2017; Ringø et al. 2010, 2014a, b; Merrifield et al. 2010; Song et al. 2014). However, the underlying mechanisms of prebiotics in enhancement of fish immunity need to be further explored. Some studies shown that a diet supplemented with β-glucan could display the gene expression levels of some immune and inflammation-related cytokines in Aeromonas spp. infected fish but the response depends on the organ, with an upregulation in the spleen and head kidney but a downregulation in the gut (Ji et al. 2017; Douxfils et al. 2017; Falco et al. 2012; Yarahmadi et al. 2014). Furthermore, in some investigations, no significant effect of dietary β-glucan on immune parameters (leucocyte subpopulations, lysozyme activity, ACH50) assessed in serum of rainbow trout and Nile tilapia has been proved despite a preventive effect against Aeromonas infection (Barros et al. 2014; Ji et al. 2017; Douxfils et al. 2017); even more, overdoses and/or prolonged of β-glucan (0.5% for 30 days rather than 2% for 15 days)) led to a poor immune response (Douxfils et al. 2017).

The preventive effect of β-glucan could also be explained by promoting a rapid healing of the intestinal damage and increasing neutrophil infiltration induced by Aeromonas spp. (Ngamkala et al. 2010). The improvement of intestinal morphology has been demonstrated with supplementation of β-glucan and MOS by increasing villi height and tunica muscularis thickness as well as gut protease and lipase activities resulting to higher trout (O. mykiss) growth and feed efficiency (Khodadadi et al. 2018). In addition, higher intestinal villi and improvement of intestinal morphology were observed in MOS-fed (1.5–2 g/kg diet) rainbow trout fish (Yilmaz et al. 2007; Dimitroglou et al. 2009).

Synbiotics

Synbiotics are nutritional supplements, combining a mixture of probiotics and prebiotics in the form of synergism as health-enhancing functional ingredients (Gibson and Roberfroid 1995). In aquaculture, synbiotics can be used in supplementation form or external bath in order to improve growth performance and feed utilization as well as increasing disease resistance, digestibility, and stimulation of the immune system of aquatic organisms (Cerezuela et al. 2011; Ringø and Song 2016; Das et al. 2017). In this paper, synbiotics beneficial effect intended to protect freshwater fish against Aeromonas infections have been reviewed like L. plantarum JCM1149 and scFOS (Liu et al. 2017), B. subtilis and MOS (Kumar et al. 2018), inactivated E. faecalis and MOS (Rodriguez-Estrada et al. 2013), Bacillus spp. (B. coagulans or B. subtilis) and Chitooligosaccharide (COS) (Lin et al. 2012; Devi et al. 2019), L. rhamnosus GG, and natural source of oligofructose-enrich inulin from Jerusalem artichoke or Kantawan (Helianthus tuberosus) (Sewaka et al. 2019) (Table 3). Prior studies revealed that dietary administration of synbiotic induced higher immune modulation (Sewaka et al. 2019; Devi et al. 2019; Kumar et al. 2018; Rodriguez-Estrada et al. 2013; Lin et al. 2012) and disease protection (Sewaka et al. 2019; Devi et al. 2019; Kumar et al. 2018; Rodriguez-Estrada et al. 2013; Liu et al. 2017; Lin et al. 2012), as well as growth rate (Sewaka et al. 2019; Rodriguez-Estrada et al. 2013; Lin et al. 2012) compared with probiotic or prebiotic diets in singular preparations. However, administration of 2 g COS kg−1 diet and B. coagulans 109 CFU g−1 separately for 56 days resulted to identical protection in A. veronii-infected koi (C. carpio koi) in comparison with the combination preparation (survival rate, 60–64% in all treatment groups vs. 33% in control) (Lin et al. 2012).

Table 3 Summary of in vivo studies in three freshwater fish species for synbiotics
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Synbiotic preparations could have the effects on fish intestinal morphometry. B. licheniformis and FOS could improve microvilli length of triangular bream (Zhang et al. 2013) and L. rhamnosus GG and oligofructose-enriched inulin increased absorptive area in juvenile red tilapia (Oreochromis spp.) intestine fish probably leading to higher absorption of available nutrients and better growth performance (Sewaka et al. 2019).

Plants

Medicinal plants and their secondary metabolites, phytochemical compounds, fractions, and plant extracts have attracted much attention as substitutes for antibiotics in controlling the outbreak of diseases in aquaculture due to their eco-friendly and cost-effectiveness benefits. Plant products have a natural origin and most of these medicinal plants do not represent a hazard for human health, animal health, or the environment (Stratev et al. 2018). Medicinal plants can produce various favorable effects due to their active principles such as alkaloids, terpenoids, tannins, saponins, and flavonoids. They can be used for their anti-stress and antioxidant properties, for their growth performance and appetite stimulation enhancement as well as their immunostimulation effect against fish diseases. They also can have antibacterial, antiviral, antifungal, and antiparasitic activities against fish and shellfish pathogens (Reverter et al. 2017).

In this review, phytochemical compounds included a wide range of medicinal plant families which were purchased or collected locally. Whole plants, parts of plants (leaf, seed, fruit), or secondary metabolites extracted with different solvents (water, methanol, chloroform, ethyl acetate) were tested (Table 4).

Table 4 Summary of in vivo studies in three freshwater fish species for phytochemical compounds
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Plant products generally were added to feed in a wide range of dosages and durations depending on various phytochemical substances tested in different fish species in previous studies. However, in some studies, plant extracts were injected intraperitoneally (Divyagnaneswari et al. 2007; Alexander et al. 2010; Devasree et al. 2014; Kirubakaran et al. 2016) or fish were immersed in plant extract (Rather et al. 2017). Investigations demonstrated a significant preventive effect of the majority of herbal extracts against Aeromonas spp., but the effect depends on the phytochemical products and their administration. For example, the survival rate in Mozambique tilapia (O. mossambicus) was higher in fish fed with a chloroform form of Nyctanthes arbortristis seed extract at 1 g kg−1 diet for 21 days (Kirubakaran et al. 2010) than in fish injected intraperitoneally at 20 mg kg−1 with a methanol form of the same seed (Kirubakaran et al. 2016), around 70 and 55%, respectively. However, some plant extracts seem to have no protective effect against Aeromonas spp. infection as methanolic extract of black cumin (Nigella sativa) (Celik Altunoglu et al. 2017) and oyster mushroom (Pleurotus ostreatus) in feeding trials (Bilen et al. 2016a, b) or the mixture of propolis and Aloe barbadensis (aloe) (Dotta et al. 2018).

Some combinations of herbal extracts showed a synergistic effect. For example, combination of two Chinese herbs (Astragalus membranaceus; Lonicera japonica) and boron (Ardó et al. 2008), Astragalus radix Chinese herb and Ganoderma lucidum fungi (Yin et al. 2009), or Satureja khuzestanica Iranian herb mixed with Oliviera decumbens (Alishahi et al. 2016) were more efficient in controlling Aeromonas infection than applying each plant alone. However, guava or mango ethanolic leaf extract alone resulted in a higher protection of rohu (L. rohita) against A. hydrophila than feeding them with both at the same level (Fawole et al. 2016).

The protective effect of phytochemical products could be due to their immunostimulant effect. Indeed, in all publications presenting a protective effect of the products, immune responses and oxidative status were enhanced significantly compared with the control groups. In contrast, the lack of protective effect of black cumin (Nigella sativa) methanolic extract could be linked to the absence of an immunostimulant effect (Celik Altunoglu et al. 2017). Herbal extracts enhanced fish immunity through different patterns. For example, a higher humoral immune responses of Mozambique tilapia (O. mossambicus) was noticed after 3 weeks of Eclipta alba leaf aqueous extract feeding in contrast with no significant modulation in the cellular immune responses (Christybapita et al. 2007). Two Chinese herbs (Astragalus membranaceus; Lonicera japonica) enhanced blood phagocytic cell functions but had a moderate effect on the plasma lysozyme level and no effect on plasma total protein and total immunoglobulin level (Ardó et al. 2008). As result of immunocompetence is increased by plant products, their applications were also studied to enhance the efficacy of some vaccines in farmed fish. Astragalus radix Chinese herb could be used in order to obtain higher survival rate in vaccinated common carp (C. carpio) after an A. hydrophila infection (Yin et al. 2009). However, Aloe vera powder did not enhance immune responses against a formalin-killed atypical A. salmonicida in rainbow trout (Zanuzzo et al. 2015).

Furthermore, the consumption of a diet containing Rehmannia glutinosa RG led to the accumulation of more beneficial microorganisms while inhibiting the growth of potential pathogens as Aeromonas sp. in the intestine of common carp (C. carpio) and which could have positive effects on the immune response of carp (Chang et al. 2018).

Essentials oils

Essential oils (EOs) are volatile, lipophilic, odoriferous, and liquid substances derived from plants for the food, hygiene, cleaning products, perfumery, and also pharmaceutical industries for their potential therapeutic effects (Edris 2007). Over the past two decades, several studies have evaluated the application EOs as a dietary additive in aquaculture due to their diverse properties (e.g., anesthetic, antioxidant, and antimicrobial) that can improve health, growth, and welfare of fish (Souza et al. 2019). The main biochemical compounds of some EOs may play a major role by acting as an anti-pathogen (Perricone et al. 2015). It has been reported that EOs can protect fish from pathogens by enhancing fish immunity, improving fish growth and feed utilization (Vaseeharan and Thaya 2014), and gut bacterial community modulation (Sutili et al. 2017; Ngugi et al. 2017; Al-Sagheer et al. 2018).

In this paper, the application of EOs to protect freshwater fish from Aeromonas infection were analyzed in Table 5 including EOs of lemongrass (Cymbopogon citratus) or geranium (Pelargonium graveolens) (Al-Sagheer et al. 2018), bitter lemon (Citrus limon) (Ngugi et al. 2017), Litsea cubeba leaf (Nguyen et al. 2016), and a commercial product (encapsulated oregano, anise, and citrus EOs) (Menanteau-Ledouble et al. 2015) which demonstrated effective protection against Aeromonas spp. infection in Nile tilapia (O. niloticus), ningu (L. victorianus), common carp (C. carpio), and rainbow trout (O. mykiss) respectively by improving immunological response, oxidative status, or growth performance.

Table 5 Summary of in vivo studies in three freshwater fish species for essential oil
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Satureja thymbra EO was also tested in rainbow trout (O. mykiss) against A. salmonicida but effective doses of S. thymbra EO determined in vitro caused toxic effects and total mortality shortly after injection and doses with low or no toxic effect did not increase the bactericidal activity of fish blood (Okmen et al. 2012). All of the EOs tested in this paper were administered orally as a feed additive except Satureja thymbra EO, which was injected intraperitoneally (Okmen et al. 2012).

Algae

Algae, including both macroalgae (seaweed) and microalgae (unicellular), are fast growing photosynthetic organisms which are potentially good sources of energy because of their high lipid content. They also contain amino acids, minerals, vitamins, chlorophyll, and some substances that have antioxidant effects (Sirakov et al. 2015; Kent et al. 2015). Several advantages of algae as an additive in aquaculture have attracted much attention, such as the positive effect on growth performance, increased triglycerides and protein deposition in muscle, protection of fish from disease, decreased nitrogen output into the environment, increased fish digestibility, physiological activity, starvation tolerance, and carcass quality (Halima 2017; Becker 2004; Mustafa and Nakagawa 1995).

In this review, the efficacy of microalgae as green algae (Chlorella vulgari) or blue-green algae (Spirulina platensis) were revealed in Nile tilapia (O. niloticus) (Abdel-Tawwab and Ahmad 2009; Fadl et al. 2017) (Table 6). The efficacy of polysaccharide fraction of a marine macroalga (Caulerpa scalpelliformi or Padina gymnospora) was also presented in Nile tilapia (O. niloticus) and common carp (C. carpio) (Rajendran et al. 2016; Yengkhom et al. 2018). In addition, the favorable protective efficacy of microencapsulated seaweed extracts was revealed against A. salmonicida in O. mossambicus (Thanigaivel et al. 2019) (Table 6). Algae treatments were administered orally as a feed supplement except the polysaccharide fraction of a marine macroalga (Caulerpa scalpelliformi), which was injected intraperitoneally (Yengkhom et al., 2018). All treatments demonstrated significant differences (p ≤ 0.05) in survival rate and protection effect between algae groups and control groups against Aeromonas infection. In addition, a significant increase of non-specific immune responses has been showed in Aeromonas challenge due to algal alternatives (Abdel-Tawwab and Ahmad 2009; Rajendran et al. 2016; Fadl et al. 2017; Yengkhom et al. 2018; Thanigaivel et al. 2019). Furthermore, Chlorella and Spirulina could improve growth performance of fish, and the proteins and lipids contents in Nile tilapia (O. niloticus) (Abdel-Tawwab and Ahmad 2009; Fadl et al., 2017).

Table 6 Summary of in vivo studies in three freshwater fish species for algae
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Bacteriophages

Use of phages, virulent virus which infect and destroy bacteria, would be a highly promising option to control diseases. However, it has not yet been fully investigated in aquaculture (Oliveira et al. 2012). In the present review, few studies evaluated the efficacy of bacteriophage in treating Aeromonas infection in farmed freshwater fish. It was seen that bacteriophage HER 110 can protect 90% of brook trout (S. fontinalis) in comparison with total mortality in the control group after 4 days of A. salmonicida infection (Imbeault et al. 2006). In addition, Aeromonas Phage PAS-1 can be applied as a biological control of A. salmonicida subsp. salmonicida infection with increased survival rates and mean times to death in rainbow trout (O. mykiss) (Kim et al. 2015).

Others functional products

As mentioned previously, many research studies have focused on the development of functional feed alternatives, examining probiotics, prebiotics and plant-derived compounds or extracts to maintain fish health and performance. There is also a growing interest in nanoparticles due to their antimicrobial effects and as drug delivery systems (Shaalan et al. 2016). For example, 100 μl intraperitoneal injection of fucoidan-coated (marine polysaccharide) gold nanoparticle (Fu-AuNPs) resulted to higher survival rate in treatment group in comparison with control group after 72 h (70 vs. 10%) against A. hydrophila in Mozambique tilapia (O. mossambicus) (Vijayakumar et al. 2017) (Table 7). However, its mode of action has not been studied in vivo while the synthesized Fu-AuNPs at 100 μg ml−1 showed effective inhibition of A. hydrophila, which is much higher than that of chloramphenicol in vitro assay (Vijayakumar et al. 2017).

Table 7 Summary of in vivo studies in three freshwater fish species for non-classified group
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The incorporation of rare earth elements such as azomite, mineral ore (Musthafa et al. 2016) and minerals such as yellow loess (sedimentary deposit of mineral particles) (Lee et al. 2016b; Won et al. 2017) in fish feed has been assessed as a means to control Aeromonas infection (Table 7). The efficacy of yellow loess against A. salmonicida in rainbow trout represented an improved growth performance, non-specific immune responses, and a furunculosis resistance (Lee et al. 2016b; Won et al. 2017).

Furthermore, the utilization of organic acids has attracted considerable attention recently due to their antimicrobial properties and role in enhancing nutrient availability in aquaculture (Ng and Koh 2017). It has been found that a commercial product which contains formic, propionic, and lactic acids and cinnamaldehyde, may be effective as an alternative method to control the impact of furunculosis in rainbow trout. However, significant difference was not found in the feed conversion ratio with the control group in this assay (Menanteau-Ledouble et al. 2017).

Main perspective

In this review, the efficiency of functional alternative products against Aeromonas infection and their potential mechanisms of action in freshwater fish were analyzed and compared. The selected studies tested highly diverse products with wide ranges of doses and durations of administration in different species of freshwater fish which were experimentally infected by Aeromonas. Furthermore, the experimental design of Aeromonas infection was also varied by the species and the strains of Aeromonas bacteria, the infectious doses, and the administration routes. It consequently was almost impossible to compare the studies or to determine whether one product is more effective than another. However, most of these alternatives were added to the basal diet as a feed supplement and were effective in inducing a preventive effect against mortality caused by Aeromonas spp. and in increasing growth performance. First, immunostimulation was the main mechanism of action investigated in the studies reviewed; nevertheless, in some studies, the protective effect of the product is clearly linked to the immunostimulant effect, but in other studies, a protective effect was observed without an increase of fish immunocompetence. Second, products feeding could also induce modifications of the gut microbiota (e.g., increase of the beneficial micro-organisms and decrease of the pathogen bacteria) as well as of the intestine morphometry (e.g., beneficial effects on the structure and decrease of tissue lesions induced by bacteria). All these mechanisms of action need to be described and explained in fish because they are clearly gaps that need to be filled in order to draw conclusions concerning their role in the protective effect of the products. Furthermore, alternative-to-antibiotics researches need to benefit from greater access to expertise in pharmacokinetics and pharmacodynamics, formulation and toxicology, for example, by creation of partnerships with biotechnology companies.

Although there are numerous clinical trials on alternative products in experimental conditions in order to reduce antibiotic use in aquaculture, there is a clear need for careful clinical trial designs in experimental conditions with relevant endpoints: primary endpoints such as reduction of morbidity and mortality but also secondary or surrogate endpoints such as changes in cytokine levels or changes in imaging of infections. Finally, in our knowledge, no evaluation of the functional feed alternative efficiency has been carried out in fish farms, where Aeromonas infection could be heterogeneous between fish, in contrast with the experimental conditions and where the environmental bacterial flora and the quality of water could influence the effect of the product. So, there is also a clear need for careful clinical trial designs in fish farm conditions, especially in order to ensure their benefits and their technical feasibility but also to improve the economic models.