WWW.WAS.ORG • WORLD AQUACULTURE • SEPTEMBER 2016 35 Types of Probiotics Various types of probiotics have been tested for crustacean culture, including Gram negative and Gram positive bacteria, bacteriophages, yeast (Saccharomyces) and phytoplankton. The use of Bacillus and Lactobacillus has shown tremendous potential, related to their ability to synthesize proteases and other enzymes that aid digestion in cultured animals. Pseudomonas, Acinetobacter, and Cellulomonas reduce the accumulation of organic matter and Nitrosomonas and Nitrobacter can control inorganic nitrogen. Bacillus Probiotics Bacillus is the prominent probiotic used in aquaculture to promote health and growth rates through optimization of the immune and digestive systems of cultured animals and its antibacterial activities against many pathogenic microorganisms. Bacillus produces endospores that are resistant to unfavorable environmental conditions. Bacillus inhibits pathogens in the culture water and can enhance water quality. Bacillus can synthesize polypeptides that are effective against a wide variety of bacteria, thus elucidating their inhibitory effects on pathogenic vibrios. The inhibitory effects of Bacillus are attributed to the synthesis of antibacterial proteins, antibiotics, competition for vital nutrients, and surface area. Some recommend using more than three Bacillus species in aquaculture feed. Isolation and description of the antibiotic compounds in Bacillus will improve our understanding of their functions in bacteria. Relatively few bacteria-inhibiting toxins of Bacillus have been characterized, although they have tremendous potential benefits in pathogen exclusion. Zokaeifar et al. (2012) isolated colonies of Bacillus from fermented cucumbers, acetic acid, Allium plants, and Brassica oleracea capitata plants. Colonies were analyzed for their capacity to inhibit pathogenic vibrios. Two strains produced high levels of bactericidal effects toward V. harveyi and V. parahaemolyticus. There are several commonly used Bacillus species for aquaculture purposes. Bacillus subtilis is a Gram positive, catalase positive, spore forming (the toughness of spores allows for digestive tract colonization), non-pathogenic bacteria. Bacillus subtilis can survive at temperatures of 11 - 52° C, 0-9 percent NaCl, and pH of 5-10. It can produce enzymes and subtilisin, which is toxic to certain bacteria. Bacillus subtilis thrives in the gut of crustaceans and synthesizes a range of antibacterials on agar plates. Bacillus is non pathogenic to cultured animals, used to improve survival of cultured animals, enhances water quality, acts as an ingested probiotic treatment, and a prophylactic supplement for digestive issues. Bacillus subtilis also synthesizes gut proteases and other enzymes that aid in digestion. Bacillus megaterium is used for the production of several α-amylases, β-amylases and neutral proteases, including P-amylase, P-glucanase, and megacins. It can synthesise vitamin B12 aerobically and anaerobically and is the major aerobic source for vitamin B12 for cultured animals. Lactobacillus Probiotics Lactobacillus is Gram-positive, non-motile and non-sporulating bacteria that produces lactic acid, has a wide range of physiology, and are catalase negative. They produce antibacterials and bacteriocins that inhibit pathogens, preventing intoxication and infections. Lactobacillus synthesizes protease and other enzymes that enhance the innate digestive system of cultured animals. The growth of certain Lactobacillus species is relatively slow because they require nutrients, a supply of carbon, nucleotides, fatty acids, amino acids and vitamins. (CONTINUED ON PAGE 36) FIGURE 3. Spore stained Bacillus subtilis (100 x magnification). FIGURE 4. Colony morphology of (A) Vibrio cincinnatiensis and (B) Vibrio parahaemolyticus isolated from larval culture water. (A) (B)
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