Enhancing growth and resistance to Vibrio alginolyticus disease in catarina scallop (Argopecten ventricosus) with Bacillus and Lactobacillus probiotic strains during early development

In bivalve aquaculture, selecting suitable probiotic treatments can be crucial for improving hatchery‐rearing of larvae and juveniles. We assessed the potential of five bacterial strains, previously selected in vitro, to improve survival, growth and resistance of catarina scallop Argopecten ventricosus during early and late larval and juvenile developmental stages, as well as during exposure to the pathogen Vibrio alginolyticus. Hatchery‐reared larvae and juveniles were treated with eight treatments of single or combined strains of Bacillus and Lactobacillus at 1 × 10⁶ CFU mL^?1 every 48 h for 9 days (larvae) and 21 days (juveniles). Compared with the control, significantly higher survival and growth in size and weight of early veliger larvae occurred with the antibiotic and the RL5 (Lactobacillus graminis) treatments. Significantly enhanced settlement of pediveliger larvae occurred with a different probiotic strain, the mix of Lactobacillus and Bacillus (MIX‐LB), while higher survival and growth of early juveniles occurred with C3 (Lactobacillus plantarum). The mix of Bacillus (MIX‐B) significantly increased survival of juveniles from V. alginolyticus after 120‐h infection, consistent with maximum activity of superoxide dismutase enzyme. In contrast, all untreated and infected scallops died by 96 h. The three Bacillus strains performed poorly when used as single treatments and when given to early developing larvae. Our results indicate that the action mechanism of probiotic strains is stage specific and strain specific, generating different responses by the host, including improved survival and growth (likely from better nutrient assimilation) and higher resistance against pathogens (possibly from strengthening the immune system).     

Isolation and evaluation of putative probiotic strains from different teleost to prevent Pseudomonas aeruginosa infection in Cyprinus carpio

Probiotics renowned as valuable microbes serve as a potential alternative to control diseases in aquaculture and are considered as an efficient and environment‐friendly approach to reduce the use of antibiotics. The present study aims at the isolation of putative probiotic bacteria from the intestinal tract of different fish species from the Doaba region of Punjab, India. In this study, isolated bacterial strains were characterized based on their morphological, biochemical and molecular characterization by 16S rRNA gene sequencing, followed by in vitro evaluation of different selection parameters described in FAO/WHO guidelines. A total of 169 different bacterial strains were isolated from the gastrointestinal tract of 52 different fish species. After in vitro evaluation, out of 169 bacterial strains only five bacteria (S3, S7, BDK2', BDK7 and BDK9) identified as Enterococcus and Bacillus species showed antagonistic activity against the fish pathogen Pseudomonas aeruginosa (MTCC 4 673). These isolates were screened based on their response towards bile tolerance, pH tolerance, adhesion and drug susceptibility to different antibiotic discs. And, the in vivo results indicated improved growth and survival against the infection (P. aeruginosa) after oral administration of the probiotics. The observations of in vitro and in vivo evaluation indicate that these isolated probiotic strains serve as effective probiotics and can be used as a novel and safe treatment to cure current issues in aquaculture.     

Tilapia recirculating aquaculture systems as a source of plant growth promoting bacteria

A recirculating aquaculture system with farmed tilapia is the most popular combination in aquaponics, an integration of aquaculture and hydroponics. Despite nutrient‐rich fish‐rearing water being regarded as a valuable resource for aquaponics, the quality and value of inhabitant microorganisms are certainly understudied. Our present research illustrates the feasibility of the tilapia‐rearing water as a valuable source of beneficial microorganisms called plant growth promoting bacteria (PGPB). Microbial communities were examined with a combination of culture‐independent high‐throughput 16S rRNA gene sequencing and cultivation methods. Microbial communities determined using high‐throughput sequencing indicated the usefulness of Bacteroidetes and Alphaproteobacteria as beneficial microbial indicators to assess the health condition of recirculating aquaculture systems. Siderophore production, ammonia production and phosphate solubilization assays were used for screening and 41% of isolates were identified as plant growth promoting bacteria. These bacteria were classified as Actinobacteria (eight strains [32% in total], Dietzia, Gordonia, Microbacterium, Mycobacterium and Rhodococcus), Bacilli (six strains [24%], Bacillus and Paenibacillus), Flavobacteriia (one strain [4%], Myroides), Betaproteobacteria (two strains [8%], Acidovorax and Chromobacterium) and Gammaproteobacteria (eight strains [32%], Aeromonas, Plesiomonas and Pseudomonas). We found that the tilapia‐rearing water naturally contained various lineages of PGPB and could be esteemed as a worthy seed bank of PGPB. Because aquaponics is a difficult system to use pesticides and herbicides, the role of PGPB to prevent plant pathogens and maintain healthy root system may be more important than traditional agricultural settings.     

Screening of gastrointestinal bacteria of Indian major carps for a candidate probiotic species for aquaculture practices

Probotic selection could be detrimental for the success of aquaculture practices. While, in most of the cases emphasis is given to the production of antibacterial compounds, and in vitro inhibitory activities of a bacterium for choosing a probiotic strain, however, other criteria like non‐antibiotic resistant, non‐haemolytic nature and non‐pathogenic potentiality of probiotic strain are often overlooked. We have screened gastrointestinal bacteria isolated from all the three species of Indian major carps viz., Labeo rohita, Catla catla and Cirrhinus mrigala for a suitable probiotic. Bacteria like Aeromonas (Aeromonas species, A. hydrophila), Micrococcus, Corynebacterium, Plesiomonas, Bacillus (Bacillus species, B. subtilis) and Pseudomonas (Pseudomonas species, P. aeruginosa) exhibited antibacterial activities against different pathogens. Among these, B. subtilis was found to grow at a wider range of temperatures, pH and salt concentrations and was non‐haemolytic, non‐antibiotic resistant, non‐pathogenic and inhibited all the tested primary and secondary pathogens. Dietary supplementation of the bacteria at 10⁸ CFU g^?1 feed also enhanced the growth of Indian major carp, L. rohita. Based upon these characteristics, we aimed to develop a suitable probiotic for aquaculture practices.     

Isolation and screening of probiotic candidates from marron,Cherax cainii (Austin, 2002) gastrointestinal tract (GIT) and commercial probiotic products for the use in marron culture

Six strains of bacteria including Bacillus mycoides (A10) and Shewanella species (A12) isolated from healthy marron intestine, Bacillus species (PM1), Bacillus subtilis (PM3), Bacillus sp. (PM4) and Bacillus sp. (AQ) from commercial probiotic products were investigated for probiotic potential in marron culture. Antibiotic susceptibility tests indicated PM3 and PM4 were susceptible to all nine antibiotics evaluated. A10, A12 and AQ were resistant to class penicillins, whereas PM1 was resistant to class penicillin and macrolides. All strains were non‐pathogenic for marron. Strong inhibition against Vibrio mimicus and Vibrio cholerae non‐01 was exhibited by PM4 and PM3. A10 inhibited V. mimicus at 72 h of growth, but not V. cholerae non‐01, whereas A12 inhibited V. cholerae non‐01 but not V. mimicus, and AQ showed no inhibition activity. A wide range of enzymes were produced by A10 and AQ using the API ZYM test. Protease enzymes were produced by PM3, PM4, AQ and PM1. In order of effectiveness, the following bacteria have probiotic potential: B. subtilis (PM3), Bacillus sp. (PM4) and B. mycoides (A10). Further study is required to determine the bacterium or any combination that gives a multibeneficial effect on marron.     

Effect of combined use of Bacillus and molasses on microbial communities in shrimp cultural enclosure systems

To assess the effect of addition of probiotics and carbon source on the microbial community structure within the shrimp culture, an enclosure system simulating a high density, intensive aquaculture of Litopenaeus vannamei was set up in Dianbai, Maoming, Guangdong province, China. The phylogenetic profiles of microbial communities in the enclosure system after addition of Bacillus and molasses were studied in the middle and final culture season (on the 45th, 75th and 95th day) using polymerase chain reaction‐denaturing gradient gel electrophoresis. A total of four groups, each containing a triplicate were set up: Group A (control), Group B (64% of daily feed was provided as molasses with Bacillus sp.), Group C (50% of daily feed amount provided as molasses with Bacillus sp.) and Group D (64% of daily feed amount as molasses alone). The results showed that α‐Proteobacteria, δ‐Proteobacteria, Bacteroidetes and Actinobacteria were the dominant bacteria. Moreover, the relative proportion of α‐Proteobacteria increased and that of Bacteroidetes decreased gradually with time. Pathogenic Flexibacter was the dominant genus in Group A on the 45th day, while the functional strains (Sphingomonas sp., Erythrobacter sp., Roseobacter sp., and Rhodobacter sp.) were detected multiple times in Groups B and C. Bacillus sp. was the cultivable, dominant bacterium in Groups B and C on the 95th day. Therefore, the combined use of Bacillus and molasses helped increase diversity of the microbial community, effectively inhibit pathogens, and promote the formation and development of a beneficial microbial community structure in the high density, intensive aquaculture of L. vannamei.     

Identification of bacterial agents causing mortality in postlarvae of giant freshwater prawn (Macrobrachium rosenbergii) in south‐west coastal districts of Bangladesh

Following an incidence of freshwater prawn Macrobrachium rosenbergii postlarvae (PL) mortality in hatcheries in summer 2012, samples from dead PL, rearing water and prawn feed from two south‐west coastal districts of Bangladesh were collected to isolate, identify and characterize the agents causing PL mortality. Antibiogram profile of sixteen randomly selected bacteria, isolated from dead PL, that grew on TCBS, to 20 different antibiotics belonging to 12 major groups revealed that the drug resistance pattern varied from moderate (56% to the drugs: ciprofloxacin, cefotaxime, nitrofurantoin, kanamycin) to complete (to penicillin, ceftazidime and oxacillin) level. To identify the isolates, amplified rDNA restriction analysis (ARDRA) classified them in to four groups, and RAPD (randomly amplified polymorphic DNA) typing yielded nine different types of isolates within these four ARDRA groups. The 16S rDNA gene sequences identified that the groups were genotypically diverse belonging to the bacterial species: Enterobacter cloacae, Klebsiella pneumoniae, Exiguobacterium profundum and Enterococcus casseliflavus, respectively, that all demonstrated their killing potential to PLs in a simulated environment. The study therefore identified four different bacterial pathogens, one of which, Exiguobacterium profundum is reported for the first here in Bangladesh, that demand special consideration for disease management strategy.     

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.     

Infectious pancreatic necrosis virus in fish by‐products is inactivated with inorganic acid (pH 1) and base (pH 12)

The aquaculture industry needs a simple, inexpensive and safe method for the treatment of fish waste without heat. Microbial inactivation by inorganic acid (HCl) or base (KOH) was determined using infectious pancreatic necrosis virus (IPNV) as a model organism for fish pathogens. Salmonella and spores of Clostridium perfringens were general hygiene indicators in supplementary examinations. IPNV, which is considered to be among the most chemical‐ and heat‐resistant fish pathogens, was reduced by more than 3 log in 4 h at pH 1.0 and pH 12.0. Salmonella was rapidly inactivated by the same treatment, whereas spores of C. perfringens were hardly affected. The results indicate that low and high pH treatment could be particularly suitable for fish waste destined for biogas production. pH treatment at aquaculture production sites could reduce the spread of fish pathogens during storage and transportation without disturbing the anaerobic digestion process. The treatment could also be an alternative to the current energy‐intensive steam pressure sterilization of fish waste to be used by the bioenergy, fertilizer and soil improver industries.     

Biological performance of Crassostrea gigas stocks produced at different hatcheries and cultivated under same environmental conditions

Production protocols as well as the origin of broodstock in hatcheries are the important factors determining the attributes of Crassostrea gigas oyster spat. Spat from diverse hatcheries may have different attributes for commercial activities. We described the biological performance (growth and condition index – CI) of spat stocks produced at different hatcheries (H1, H2, H3 and H4), and simultaneously cultivated in La Cruz lagoon (Mexico). Analysis of pathogens and genetic variability was performed for each stock. Temperature, salinity, chlorophyll a and seston were measured during samplings. Stocks showed similar growth patterns but each one showed a distinctive curve. Temperature was the governing factor on growth but its effect was variable depending on the stock and whether the temperature increases, stays high or decreases. Availability of food was high producing elevated CI. No presence of pathogens was detected and it was not possible to obtain amplicons of the mitochondrial regions. Despite differences in performance among stocks, all of them showed higher values (growth rates, CI, survival) than the standards reported for the Gulf of California. These findings are useful for the production and cultivation of C. gigas, but more studies are needed to understand the influence of hatchery protocols on spat quality.