A Review of Probiotics in Shrimp Aquaculture

Development of shrimp aquaculture has been associated with increases in infectious diseases and environmental degradation. An effective alternative to chemicals and antibiotics used for the prevention of these problems is to administer probiotics into the rearing system. Three bacterial genera, Bacillus, Vibrio, and Pseudomonas, are commonly administered as probiotics in shrimp aquaculture. Candidate probiotics are species specific and need to be tested for their effectiveness for certain species in in vitro and in vivo. Supplementation into feed is more effective in conveying probiotics into animals compared to direct application into rearing systems. Overdosage or prolonged administration of probiotics can induce immunosuppression. A cell-density of 10⁵ colony-forming units (CFU) per ml is widely recommended. A combination of probiotics results in better outcomes for the host than individual probiotics. Probiotics improve water quality while reducing pathogenic bacteria. Probiotics show positive effects through an improvement in the physiological and immune responses of shrimps. Probiotics are increasingly becoming important and more common in any organic shrimp farming.     

Probiotics for shrimp larviculture: review of field data from Asia and Latin America

Disease problems have emerged as major constraints in aquaculture production. The prophylactic application of antibiotics is expensive and detrimental, i.e. selection of bacteria that are drug‐resistant or more virulent and the prevalence of drug residues in reared animals. Probiotics, which compete with bacterial pathogens for nutrients and/or inhibit the growth of pathogens, could be a valid alternative to the prophylactic application of chemicals. A mixture of specific Bacillus strains was designed following a research programme on the ability of numerous Bacillus strains to inhibit a range of pathogenic Vibrio strains, to grow under conditions prevailing in shrimp hatcheries and to degrade waste products. These strains were then included in bioassays and challenge tests in order to confirm the lack of toxin production and pathogenicity to humans, target organisms and the environment. Here, we report on the performance of a commercially available mixture of Bacillus strains (SANOLIFE^® MIC), using data from Asian and Latin‐American hatcheries, with Penaeus monodon (Fabricius 1798) and Litopenaeus vannamei (Boone 1931). These results show that probiotics may be a suitable alternative to the prophylactic use of antibiotics. Obviously, minimizing the risk of vibriosis demands a multi‐disciplinary approach, including good hygiene and sanitation measures to reduce the input of potential pathogens, as well as a suitable farm management.     

Revealing the compositions of the intestinal microbiota of three Anguillid eel species using 16S rDNA sequencing

Anguilla japonica, A. marmorata and A. bicolor pacifica are important eel species for aquaculture in Taiwan. In the past, seldom studies focused on the intestinal microbiome of them or the probiotics used in their cultivation. Probiotics are microbes that can improve the health of organisms, and the microbes isolated from the intestines of organisms might be the potential probiotics. Therefore, the aim of this study was to collect the intestinal mucus samples of these three eel species for investigating the autochthonous microbiome using 16S rDNA sequencing. The diversity, abundance of the intestinal microbiome and compositions of all the libraries were identified. Moreover, the composition of intestinal microbiome of eel was suggested that it would be affected by the waters and the characteristics of different eel species. Furthermore, most of the identified predominant genera of the intestinal microbiome of these three eel species, such as Cetobacterium, Clostridium, Shewanella, Acinetobacter and Bacteroides, seemed to have great potential as the probiotics. In the future, more studies shall be conducted to promote the practical application of these candidate probiotics for improving the techniques of eel farming.     

Aqualase®, a yeast‐based in‐feed probiotic,modulates intestinal microbiota,immunity and growth of rainbow trout Oncorhynchus mykiss

Yeast probiotics have great promise, yet they received little attention in fish. This study investigated the influence of Aqualase^®, a yeast‐based commercial probiotic composed of Saccharomyces cerevisiae and Saccharomyces elipsoedas, on health and performance of rainbow trout (Oncorhynchus mykiss). Probiotics were incorporated in the diets at three different inclusion levels (1%, 1.5% and 2%) and administered to the fish for a period of 8 weeks. After the feeding trial, intestinal total viable aerobic bacterial count was significantly higher in fish group that received 2% in‐feed probiotics. In addition, a significant increase in at least 11% in intestinal lactic acid bacteria population was observed in all probiotic‐fed groups. Total protein level and lysozyme activity in skin mucus were significantly elevated following probiotic feeding. Inhibitory potential of skin mucus against fish pathogens was significantly enhanced by at least 50% in probiotic‐fed groups. Humoral and cellular immune parameters were influenced by probiotic feeding and the effects were dependent on inclusion level. Digestive physiology was affected by in‐feed probiotics through improvement of intestinal enzyme activities. All growth performance parameters were significantly improved following probiotic administration specifically at inclusion rate 1.5% and above. Taken together, the results revealed that Aqualase^® is a promising yeast‐based probiotic for rainbow trout with the capability of modulating the intestinal microbiota, immunity and growth.     

Evaluating the probiotic potential and adhesion characteristics of Bacillus spp. isolated from the intestine of Rhynchocypris lagowskii Dybowski

Probiotics have been widely used in aquaculture worldwide especially in China as antibiotics have been banned in aquaculture. In the present study, 27 potential strains were isolated from Rhynchocypris lagowskii Dybowski intestinal tract towards the assessment of their probiotic potential. Two potential probiotics were finally screened from the 27 candidate strains according to the results of enzyme-producing ability, in vitro against pathogens ability and antibiotic sensitivity test. The 2 potential probiotics were identified and confirmed on the basis of their colony morphology, biochemical characteristics, and 16S rRNA sequencing. The probiotic strains, LSG2-3–2 and LSG3-6, were identified to be Bacillus methylotrophicus and B. tequilensis, respectively. Further studies showed that LSG2-3–2 and LSG3-6 had excellent tolerance to high temperature (80℃), low pH (3–5), bile salts (0.3%), intestinal juice (10%), and gastric juice (0.5%). The adhesion rates of LSG2-3–2 and LSG3-6 in the in vitro intestinal mucosal adhesion model were 17.74% and 24.04%, respectively. Analysis of their bacteria surface adhesive proteins revealed that the lectins on the bacterial surface of LSG2-3–2 and LSG3-6 were mainly protein and glycoprotein properties, respectively. The adhesion receptor components in the mucus proteins of the two strains were all protein properties. The results of the inhibitory adhesion test indicated that LSG3-6 had a higher inhibitory effect on Aeromonas hydrophila and LSG2-3–2 had a better inhibitory effect on A. veronii. The biosafety assay confirmed that the isolates were not pathogenic to the host fish. Based on the presently observed characteristic features, it can be concluded that LSG2-3–2 (B. methylotrophicus) and LSG3-6 (B. tequilensis) retrieved from the digestive tract of R. lagowskii can be used for the healthy breeding and disease prevention of R. lagowskii.

     

Competition for attachment of aquaculture candidate probiotic and pathogenic bacteria on fish intestinal mucus

Probiotics for aquaculture are generally only selected by their ability to produce antimicrobial metabolites; however, attachment to intestinal mucus is important in order to remain within the gut of its host. Five candidate probiotics (AP1-AP5), isolated from the clownfish, Amphiprion percula (Lacepéde), were examined for their ability to attach to fish intestinal mucus and compete with two pathogens, Aeromonas hydrophila and Vibrio alginolyticus. Two different radioactive isotopes were used to quantify competition between pathogens and probionts. Attachment of the pathogens was enhanced by the presence of the candidate probiotics. However, the addition of the candidate probiotics after the pathogens resulted in reduced pathogen attachment. Only AP5 caused lower attachment success of V. alginolyticus when added before the pathogen. When AP5 was added first, the average attachment change was 41% compared with 72% when added after V. alginolyticus, suggesting that the probiotic is displaced but that enhanced attachment of the pathogen does not occur. Conversely, when V. alginolyticus was added first, followed by AP5, attachment change was 37% while AP5 had 92% attachment change when added second. This implies that the pathogen was displaced by the candidate probiotic and therefore it appeared that, based on the ability of probiont AP5 to attach to mucus, the growth of the pathogen in the digestive tract might be suppressed by the candidate probiont's presence.     

In vitro selection and characterization of putative probiotics isolated from the gut of Acipenser baerii (Brandt, 1869)

To select and characterize potential probiotic bacteria from the gut microbiota of Siberian sturgeon (Acipenser baerii), 129 strains isolated from the hindgut were screened for antagonistic activity against five fish pathogens. Ten isolates showed antagonism towards three or more pathogens. Nine of these isolates were Gram‐positive, belonging to Lactococcus (seven) and Bacillus (two), and a single strain belonging to the Gram‐negative Citrobacter. These inhibitory isolates were identified using genetic, phentotypic and biochemical traits, and further characterized by in vitro tests assessing the adhesion and growth in mucus and resistance to gastric and intestinal fluids. The candidate probiotics were determined to be non‐pathogenic through an in vivo study. Based on these assays, Lactococcus lactis ssp. lactis STG45 and STG81 showed the broadest inhibitory potential, a high viability in simulated gastrointestinal juice and the highest adhesion capacity to mucus. They were therefore selected as the most promising candidate probiotics. This is the first study screening probiotics among the gut microflora of Siberian sturgeon.     

Encapsulation capacity of Artemia nauplii with customized probiotics for use in the cultivation of western king prawns (Penaeus latisulcatus Kishinouye, 1896)

The encapsulation capacity of Artemia nauplii with customized probiotics Pseudomonas synxantha and Pseudomonas aeruginosa for use in the cultivation of western king prawns (Penaeus latisulcatus) was investigated. Seven trials were conducted to investigate this encapsulation capacity in terms of Artemia survival and probiotic load in Artemia. Newly hatched Artemia nauplii at 250 nauplii mL^?1 were fed individual probiotics at 0, 10³, 10⁵ and 10⁷ colony‐forming units (CFU) per millilitre, and mixtures of these two probiotics (10⁵ CFU mL^?1) at 30:70, 50:50 or 70:30 v/v in a medium of ozonated water (OW), tryptone soya broth (TSB), and a mixture of these media. The appropriate medium for encapsulation of probiotics by Artemia nauplii was the mixture of OW and TSB at 75:25 v/v; whereas, the use of OW or TSB alone was not effective. Artemia nauplii most effectively encapsulated the customized probiotics at 10⁵ CFU mL^?1. The results indicates that the encapsulation of Artemia nauplii is optimized by using a combination of P. synxantha and P. aeruginosa at 50:50 v/v in a media mixture of OW and TSB at 75: 25 v/v. Artemia should be harvested at 48 h when survival is still high (78%) and the probiotic load in Artemia is high (3 × 10⁴ CFU nauplius^?1).     

Effects of Dietary Probiotics on the Growth and Feeding Efficiency of Red Hybrid Tilapia,Oreochromis sp., and Subsequent Resistance to Streptococcus agalactiae

An eight-week trial was performed to evaluate three commercial/prototype probiotics supplemented in red hybrid tilapia, Oreochromis sp. diets. Triplicate groups of tilapia were measured for growth performance, feeding efficiencies, and whole-body composition. After the feeding trial, duplicate groups of tilapia were assessed for their resistance to Streptococcus agalactiae over 23 days. Six diets were supplemented with 0.1% or 0.3% PB₁ consisting of Bacillus subtilis, 0.1% or 0.3% PB₂ consisting of B. licheniformis or 0.1% MPB consisting of Bacillus sp. and Pediococcus sp. Probiotics had no effect (p > 0.05) on growth or feeding efficiencies, although whole-body crude protein was significantly higher in the PB₁ 0.3% treatment. Tilapia in the probiotic treatments had a higher resistance to S. agalactiae and, with the exception of the PB₂ 0.1% diet, were all significantly higher than the control treatment. Although the tested probiotics were not growth promoters, dietary B. subtilis was the most effective prophylactic against pathogenic bacteria.     

Potential environmental effects of probiotics used in aquaculture

Probiotics are a live microbial feed supplement that beneficially affects the host animal. As aquaculture is one of the main areas where probiotics can be applied, the environmental suitability of using probiotics in hatching and nursing facilities for turbot farming was evaluated through the life cycle assessment (LCA) methodology. This LCA study showed that the potential environmental impacts associated with probiotic production are generally offset by the impact reductions linked to lower consumption levels and reduced waste and emission generation rates within the hatching and nursing subsystem. Thus, the use of probiotics is recommended in order to enhance the performance of turbot aquaculture systems provided that operational and economic advantages are attained.