Skip to main content

Advertisement

SpringerLink
Log in

Effectiveness of Probiotic Phaeobacter Bacteria Grown in Biofilters Against Vibrio anguillarum Infections in the Rearing of Turbot (Psetta maxima) Larvae

  • Original Article
  • Published:
Marine Biotechnology Aims and scope Submit manuscript

Abstract

The rearing environment of first-feeding turbot larvae, usually with high larvae densities and organic matter concentrations, may promote the growth of opportunistic pathogenic Vibrionaceae bacteria, compromising the survival of the larvae. The aim of this study was to assess the effectiveness of the biofilm-forming probiotic Phaeobacter 27-4 strain grown on a ceramic biofilter (probiofilter) in preventing Vibrio anguillarum infections in turbot larvae. In seawater with added microalgae and maintained under turbot larvae rearing conditions, the probiofilter reduced the total Vibrionaceae count and the concentration of V. anguillarum, which was undetectable after 144 h by real-time PCR. The probiofilter also improved the survival of larvae challenged with V. anguillarum, showing an accumulated mortality similar to that of uninfected larvae (35–40 %) and significantly (p < 0.05) lower than that of infected larvae with no probiofilter (76 %) due to a decrease in the pathogen concentration and in total Vibrionaceae. Furthermore, the probiofilter improved seawater quality by decreasing turbidity. Phaeobacter 27-4 released from the probiofilters was able to survive in the seawater for at least 11 days. The bacterial diversity in the larvae, analysed by denaturing gradient gel electrophoresis, was low, as in the live prey (rotifers), and remained unchanged in the presence of V. anguillarum or the probiofilter; however, the probiofilter reduced the bacterial carrying capacity of the seawater in the tanks. Phaeobacter-grown biofilters can constantly inoculate probiotics into rearing tanks and are therefore potentially useful for bacterial control in both open and recirculating industrial units.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

Similar content being viewed by others

References

  • Ardjosoediro I, Rammarine IW (2002) The influence of turbidity on growth, feed conversion and survivorship of the Jamaica red tilapia strain. Aquaculture 212:159–165

    Article  Google Scholar 

  • Blanch AR, Alsina M, Simon M, Jofre J (1997) Determination of bacteria associated with reared turbot (Scophthalmus maximus) larvae. J Appl Microbiol 82:729–734

    Article  Google Scholar 

  • Brinkhoff T, Bach G, Heidorn T, Liang L, Schlingloff A, Simon M (2004) Antibiotic production by a Roseobacter clade-affiliated species from the German Wadden Sea and its antagonistic effects on indigenous isolates. Appl Environ Microbiol 70:2560–2565

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Bruhn JB, Nielsen KF, Hjelm M, Hansen M, Bresciani J, Schulz S, Gram L (2005) Ecology, inhibitory activity and morphogenesis of a marine antagonistic bacterium belonging to the Roseobacter clade. Appl Environ Microbiol 71:7263–7270

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Bruhn JB, Haagensen JAJ, Bagge-Ravn D, Gram L (2006) Culture conditions of Roseobacter strain 27-4 affect its attachment and biofilm formation as quantified by real-time PCR. Appl Environ Microbiol 72:3011–3015

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Buchan A, Gonzalez JM, Moran MA (2005) Overview of the marine Roseobacter lineage. Appl Environ Microbiol 71:5665–5677

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Cabello FC (2006) Heavy use of prophylactic antibiotics in aquaculture: a growing problem for human and animal health and for the environment. Environ Microbiol 8:1137–1144

    Article  CAS  PubMed  Google Scholar 

  • Crab R, Avnimelech Y, Defoirdt T, Bossier P, Verstraete W (2007) Nitrogen removal techniques in aquaculture for a sustainable production. Aquaculture 270:1–14

    Article  CAS  Google Scholar 

  • D’Alvise PW, Melchiorsen J, Porsby CH, Nielsen KF, Gram L (2010) Inactivation of Vibrio anguillarum by attached and placktonic Roseobacter cells. Appl Environ Microbiol 76:2366–2370

    Article  PubMed Central  PubMed  Google Scholar 

  • Dalmin G, Kathiresan K, Purushotaman A (2001) Effect of probiotics on bacterial population and health status of shrimp in culture pond ecosystem. J Exp Biol 39:939–942

    CAS  Google Scholar 

  • Dang HY, Lovell CR (2002) Seasonal dynamics of particle-associated and free-living marine Proteobacteria in a salt march tidal creek as determined using fluorescence in situ hybridization. Environ Microbiol 4:287–295

    Article  CAS  PubMed  Google Scholar 

  • Fjellheim AJ, Playfoot KJ, Skjermo J, Vadstein O (2007) Vibrionaceae dominates the microflora antagonistic towards Listonella anguillarum in the intestine of cultured Atlantic cod (Gadus morhua L.) larvae. Aquaculture 269:98–106

    Article  Google Scholar 

  • Gram L, Melchiorsen J, Bruhn JB (2010) Antibacterial activity of marine culturable bacteria collected from a global sampling of ocean surface waters and surface swabs of marine organisms. Mar Biotechnol 12:439–451

    Article  CAS  PubMed  Google Scholar 

  • Hjelm M, Bergh Ø, Riaza A, Nielsen J, Melchiorsen J, Jensen S, Duncan H, Ahrens P, Birkbeck H, Gram L (2004a) Selection and identification of autochthonous potential probiotic bacteria from turbot larvae (Scolphthalmus maximus) rearing units. Syst Appl Microbiol 27:360–371

    Article  PubMed  Google Scholar 

  • Hjelm M, Riaza A, Formoso F, Melchiorsen J, Gram L (2004b) Seasonal incidence of autochthonous antagonistic Roseobacter spp. and Vibrionaceae strains in a turbot larva (Scolphthalmus maximus) rearing system. Appl Environ Microbiol 70:7288–7294

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Kennedy SB, Tucker JW, Neidig CL, Vermeer GK, Cooper VR, Jarrell JL, Sennett DG (1998) Bacterial management strategies for stock enhancement of warm water marine fish: a case study with common snook (Centropomus undecimalis). Bull Mar Sci 62:573–588

    Google Scholar 

  • Makridis P, Bergh Ø, Skjermo J, Vadstein O (2001) Addition of bacteria bioencapsulted in Artemia metanauplii to a rearing system for halibut larvae. Aquac Int 9:225–235

    Article  Google Scholar 

  • Mallekh R, Lagardere JP, Anras MLB, Lafaye JY (1998) Variability in appetite of turbot, Scophthalmus maximus under intensive rearing conditions: the role of environmental factors. Aquaculture 165:123–138

    Article  Google Scholar 

  • Marzorati M, Wittebolle L, Boon N, Daffonchio D, Verstraete W (2008) How to get more out of molecular fingerprints: practical tools for microbial ecology. Environ Microbiol 10:1571–1581

    Article  CAS  PubMed  Google Scholar 

  • Mayali X, Franks PJS, Azam F (2008) Cultivation and ecosystem role of a marine Roseobacter clade-affiliated cluster bacterium. Appl Environ Microbiol 74:2595–2603

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Michaud L, Blancheton JP, Bruni V, Piedrahita R (2006) Effect of particulate organic carbon on heterotrophic bacterial populations and nitrification efficiency in biological filters. Aquac Int 34:224–233

    Google Scholar 

  • Michaud L, Giudice AL, Troussellier M, Smedile F, Bruni V, Blancheton JP (2009) Phylogenetic characterization of the heterotrophic bacterial communities inhabiting a marine recirculating aquaculture system. J Appl Microbiol 107:1935–1946

    Article  CAS  PubMed  Google Scholar 

  • Munro PD, Barbour A, Birkbeck TH (1993) Bacterial flora of rotifers Brachionus plicatilis; evidence for a major location on the external surface and methods for reducing the rotifer bacterial load. In: Reinertsen H, Dahle LA, Jørgensen L, Tyinnereim K (eds) Fish farming technology. A.A. Balkema, Rotterdam, pp 93–100

    Google Scholar 

  • Muyzer G, de Waal EC, Uitterlinden AG (1993) Profiling of complex microbial populations by denaturing gradient gel electrophoresis analysis of polymerase chain reaction-amplified genes coding for 16S rRNA. Appl Environ Microbiol 59:695–700

    PubMed Central  CAS  PubMed  Google Scholar 

  • Olafsen JA (2001) Interactions between fish larvae and bacteria in marine aquaculture. Aquaculture 200:223–247

    Article  Google Scholar 

  • Olsson JC, Jöborn A, Westerdahl A, Blomberg L, Kjelleberg S, Conway PL (1998) Survival, persistence and proliferation of Vibrio anguillarum in juvenile turbot, Scopththalmus maximus (L.), intestine and faeces. J Fish Dis 21:1–9

    Article  Google Scholar 

  • Pintado J, Pérez-Lorenzo M, Luna-González A, Sotelo CG, Prol MJ, Planas M (2010) Monitoring of the bioencapsulation of a probiotic Phaeobacter strain in the rotifers Brachionus plicatilis using denaturing gradient gel electrophoresis. Aquaculture 302:182–194

    Article  Google Scholar 

  • Planas M, Pérez-Lorenzo M, Vázquez A, Pintado J (2005) A model for experimental infections with Vibrio (Listonella) anguillarum in first feeding turbot (Scophthalmus maximus) larvae under hatchery conditions. Aquaculture 250:232–243

    Article  Google Scholar 

  • Planas M, Pérez-Lorenzo M, Hjelm M, Gram L, Fiksdal IU, Bergh Ø, Pintado J (2006) Probiotic effect in vivo of Roseobacter strain 27-4 against Vibrio (Listonella) anguillarum infections in turbot (Scophthalmus maximus L.) larvae. Aquaculture 255:323–333

    Article  Google Scholar 

  • Porsby CH, Nielsen KF, Gram L (2008) Phaeobacter and Ruegeria species of the Roseobacter clade colonize separate niches in a Danish turbot (Scophthalmus maximus)-rearing farm and antagonize Vibrio anguillarum under different growth conditions. Appl Environ Microbiol 74:7356–7364

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Porsby CH, Webber MA, Nielsen KF, Piddock LJV, Gram L (2011) Resistance and tolerance to tropodithietic acid, an antimicrobial in aquaculture, is hard to select. Antimicrob Agents Chemother 55:1332–1337

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Prol MJ, Bruhn JB, Pintado J, Gram L (2009) Real-time PCR detection and quantification of fish probiotic Phaeobacter strain 27-4 and fish pathogenic Vibrio in microalgae, rotifer, Artemia and first feeding turbot (Psetta maxima) larvae. J Appl Microbiol 106:1292–1303

    Article  CAS  PubMed  Google Scholar 

  • Prol-García MJ, Planas M, Pintado J (2010) Different colonization and residence time of Listonella anguillarum and Vibrio splendidus in the rotifer Brachionus plicatilis determined by real-time PCR and DGGE. Aquaculture 302:26–35

    Article  Google Scholar 

  • Prol-García MJ, Gómez M, Sánchez L, Pintado J (2012) Phaeobacter grown in biofilters: a new strategy for the control of Vibrionaceae in aquaculture. Aquac Res. doi:10.1111/are.12046

    Google Scholar 

  • Qi Z, Dierckens K, Defoirdt T, Sorgeloos P, Boon N, Bao Z, Bossier P (2009) Effects of feeding regime and probionts on the diverting microbial communities in rotifer Brachionus culture. Aquac Int 17:303–315

    Article  Google Scholar 

  • Rao D, Webb JS, Kjelleberg S (2006) Microbial colonization and competition on the marine alga Ulva australis. Appl Environ Microbiol 72:5547–5555

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Reid HI, Treasurer JW, Adam B, Birkbeck TH (2009) Analysis of bacterial populations in the gut of developing cod larvae and identification of Vibrio logei, Vibrio anguillarum and V. splendidus as pathogens of cod larvae. Aquaculture 288:36–43

    Article  Google Scholar 

  • Reitan KI, Natvik CM, Vadstein O (1998) Drinking rate, uptake of bacteria and microalgae in turbot larvae. J Fish Biol 53:1145–1154

    Article  Google Scholar 

  • Salvesen I, Skjermo J, Vadstein O (1999) Growth of turbot (Scophthalmus maximus L.) during first feeding in relation to the proportion of r/K-strategists in the bacterial community of the rearing water. Aquaculture 175:337–350

    Article  Google Scholar 

  • Sandlund N, Bergh Ø (2008) Screening and characterisation of potentially pathogenic bacteria associated with Atlantic cod Gadus morhua larvae: bath challenge trials using a multidish system. Dis Aquat Org 81:203–217

    Article  PubMed  Google Scholar 

  • Skjermo J, Vadstein O (1993) Characterization of the bacterial flora of mass cultivated Brachionus plicatilis. Hydrobiologia 255(256):185–191

    Article  Google Scholar 

  • Skjermo J, Salvesen I, Oie G, Olsen Y, Vadstein O (1997) Microbially matured water: A technique for selection of a non-opportunistic bacterial flora in water that may improve performance of marine larvae. Aquac Int 5:13–28

    Article  Google Scholar 

  • Skov MN, Pedersen K, Larsen JL (1995) Comparison of pulsed-field gel electrophoresis, ribotyping, and plasmid profiling for typing of Vibrio anguillarum serovar O1. Appl Environ Microbiol 61:1540–1545

    PubMed Central  CAS  PubMed  Google Scholar 

  • Sugita H, Mizuki H, Shiro I (2008) Prevalence of a fish pathogen, Listonella anguillarum, in the intestinal tract of fish collected off the coast of Japan. Aquac Res 39:103–105

    Article  Google Scholar 

  • Thomson R, Macpherson HL, Riaza A, Birkbeck TH (2005) Vibrio splendidus biotype 1 as a cause of mortalities in hatchery-reared larval turbot, Scophthalmus maximus (L.). J Appl Microbiol 99:243–250

    Article  CAS  PubMed  Google Scholar 

  • Verdonck L, Grisez L, Sweetman E, Minkoff G, Sorgeloos P, Ollevier F, Swings J (1997) Vibrios associated with routine production of Brachionus plicatilis. Aquaculture 149:203–214

    Article  Google Scholar 

  • Verschuere L, Dhont J, Sorgeloos P, Verstraete W (1997) Monitoring Biolog patterns and r/K-strategists in the intensive culture of Artemia juveniles. J Appl Microbiol 83:603–612

    Article  Google Scholar 

  • Verschuere L, Rombaut G, Sorgeloos P, Verstraete W (2000) Probiotic bacteria as biological control agents in aquaculture. Microbiol Mol Biol Rev 64:655–671

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Vine NG, Leukes WD, Kaiser H (2006) Probiotics in marine larviculture. FEMS Microbiol Rev 30:404–427

    Article  CAS  PubMed  Google Scholar 

  • Wagner-Döbler I, Biebl H (2006) Environmental biology of the marine Roseobacter lineage. Annu Rev Microbiol 60:255–280

    Article  PubMed  Google Scholar 

  • Zhang XH, Austin B (2000) Pathogenicity of Vibrio harveyi to salmonids. J Fish Dis 23:93–102

    Article  Google Scholar 

Download references

Acknowledgments

We thank Professor Lone Gram for kindly providing the strains used in this study. We are grateful to Dr Miquel Planas for critical revision of the manuscript and to Alexandro Chamorro for technical assistance. We also thank Elisabeth Heseltine for editing the manuscript.

María J. Prol-García obtained a grant from the I3P Program of CSIC, co-financed by European Social Fund. Research funding was also provided by INIA (ACU03-003) and by the I3 Program of the Spanish Ministry of Education and Science (CSIC-PIE 2006 7 01067).

Author information

Author notes

  1. María J. Prol-García

    Present address: Technical University of Denmark-DTU Systems Biology, Søltofts Plads, Building 221, 2800, Kongens Lyngby, Denmark

Authors and Affiliations

  1. Instituto de Investigacións Mariñas (IIM-CSIC), Eduardo Cabello 6, 36208, Vigo, Spain

    María J. Prol-García & José Pintado

Authors
  1. María J. Prol-García
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. José Pintado
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to María J. Prol-García.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary figure 1

PCR-DGGE profile of 16S rDNA fragments of A) turbot larvae from control, BPh, Va and VaBPh tanks; B) rotifer samples from tanks on day 10 post-hatching (rotifer LD10), of rotifers enriched with I. galbana (rotifers Ig) and of rotifers loaded with V. anguillarum (rotifers Va); and C) seawater samples taken from control, BPh, Va and VaBPh tanks. M: marker (A: Kordia algicida, B: Tenacibaculum discolor, C: Vibrio anguillarum 90-11-287, D: Phaeobacter 27-4, E: Ruegeria mobilis, F: Flexibacter sp). Va: band with a migration pattern similar to V. anguillarum, Ph: band with a migration pattern similar to Phaeobacter 27-4. (DOCX 6544 kb)

Supplementary figure 2

Functional organization based on Pareto–Lorenz distribution curves obtained from DGGE profiles on days 3 (×), 5 (◊), 7 (Δ), 9 (○) and 10 (□) after larval hatching of larvae and seawater from control, BPh, Va and VaBPh tanks. The dashed vertical line at the 0.2x axis is plotted to evaluate the range of the Pareto values. (DOCX 701 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Prol-García, M.J., Pintado, J. Effectiveness of Probiotic Phaeobacter Bacteria Grown in Biofilters Against Vibrio anguillarum Infections in the Rearing of Turbot (Psetta maxima) Larvae. Mar Biotechnol 15, 726–738 (2013). https://doi.org/10.1007/s10126-013-9521-4

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10126-013-9521-4

Keywords

Search

Navigation