March | April 2013

The role of bioremediation in water quality management

The International magazine for the aquaculture feed industry

International Aquafeed is published six times a year by Perendale Publishers Ltd of the United Kingdom.All data is published in good faith, based on information received, and while every care is taken to prevent inaccuracies, the publishers accept no liability for any errors or omissions or for the consequences of action taken on the basis of information published. ©Copyright 2013 Perendale Publishers Ltd. All rights reserved. No part of this publication may be reproduced in any form or by any means without prior permission of the copyright owner. Printed by Perendale Publishers Ltd. ISSN: 1464-0058

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Withtheexpansionanddevel-opment of the aquacultureindustry, several challengesarise.The intensification of

production systems increases the pressureon the environment, which can severelyaffect water quality and as a consequencefish or shrimp performance and the inci-denceofdiseases.

Inaquaculture,theapplicationofbeneficialbacteria (probiotics) is not only associatedwith gut health (feed probiotics), but alsowith bioremediation improving the environ-ment (water and soil) in which the animalsarereared.Theeffectsofbiodegradingstrains(suchasBacillus sp.,Paracoccus sp.,Thiobacillus sp.) added directly to the water involve themodulation of the microbiology profile inponds,degradationofundesirablewastecom-pounds (ammonia, nitrite, hydrogen sulfide),enhanced mineralization of organic matter,

decreased anaerobic conditions in pond soilandreducedsludgeaccumulation.

Moreover, enzymes can be an effectivetool in the degradation of organic matterin very intensive production systems. Thesepositivechangesintheenvironmentaresup-ported by proven benefits for the perform-anceandsurvivalofshrimpfromthelarvaltogrow-outstages.

Toxicity of nitrogenous compounds

Nitrogen compounds, such as nitrite,nitrate and ammonium ions / ammonia aretoxic when their concentrations exceed acertainlevelintherearingwater.Ammoniumnitrogen that occurs partly in the form ofammonium ion (NH4+) and ammonia(NH3+)originatesfromdecomposingorganicwasteandanimalexcretionsinthefarm.Thesensitivity to ammonium nitrogen dependslargelyonthespecies.

Some fish have developed strategies, for

example, the formation of glutamine in thebrain to detoxify ammonium to urea, toprotectthemselvesfromtoxicammonialevels(Randall and Tsui, 2002). Nitrite (NO2-) isusually present below dangerous concentra-tionsinfreshandmarinewater.

However, prolonged exposure to highnitritelevels,especiallywhenoxygenislimited,leads to anoxia and slow suffocation of theanimals, because nitrite changes hemoglobinintomethemoglobin,aformthatisnotabletobindoxygen(LewisandMorris,1986).

Nitrate (NO3-) is the least dangerouscompound and low concentrations are notproblematic.Similartonitrite,nitrateconvertshemoglobin, into a non-binder for oxygen.Permanent exposure to high nitrate levelscausesweightlossandahigheroccurrenceofinfectiousdiseases.Toavoid thesecomplica-tions, excess nitrate needs to be removedto reach lower, non-toxic concentrations(Camargaet al.,2005).Thisisoftenachievedbywaterrenewalatthefarms.

Bioremediation in aquacultureWatewater management in aquaculture

systems is crucial to maintain a good healthstatusof theanimalsaswell as tocounteractthe negative impacts on the environment.Bioremediation, the application of micro-organisms like bacteria to remove dangerouswasteproducts, isapromising tool foronsitetreatment of watewater and contaminatedsediments. For the bioremediation of nitrog-enous compounds, bacteria have to performnitrificationanddenitrification.Bacterialnitrifica-tionistheoxidationofammonium/ammonia(NH4+,NH3+)tonitrate(NO3-)viahydroxylamine and nitrite (NO2-). Denitrification

The role of bioremediation in water quality managementby Goncalo A. Santos, MSc, technical manager – aquaculture, Biomin Holding GmbH, Austria

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Figure 1: Nitrification and denitrification processes in aquatic environments

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describes the reduction of nitrate to nitrousoxideandfinallytonitrogengas,whichreturnsinto the atmosphere (Chávez-Crooker andObreque-Contreras, 2010).Although a rangeofbacterial speciesarecapableofnitrificationand/ordenitrification,notallspeciesareappli-cable for bioremediation products. RecentlystrainssuchasParacoccus sp.andThiobacillus sp.havegainedinterestduetoitsdegradingcapa-bilities. Also Bacillus sp. is also well suited toperformseveralfunctionsinthewatercleanupapplication(Nakanoet al.,1998).

A single strain, rarelyharbours all desiredqualitiesnecessaryforanefficientdegradationoftoxiccompounds,thereforeacombinationofstrainsthatperformbestforoneorseveral

compounds ismore likely toensurea stableperformance.

Beneficial bacteria and enzymes to improve water and soil quality in aquaculture ponds

A key factor for successful aquaculture isto understand the interactions between themicrobialenvironment,gutfloraandimmunesystem of the shrimp, as well as the factorsthat determine the persistence of microbialspecies inthe internalandexternalmicrobialecosystems. While natural environments arebalanced, the farming environment favoursthe growth of micro-organisms as it is richin nutrients and feed waste. Farmed species

are constantly exposed to and challengedby micro-organisms from the surroundingenvironment.

These environmental challenges areobviously influenced by different factors,including farm management and rearingmethods.Aquacultureoperationsgenerallyinvolvethestockingandfeedingofshrimpin open or semi-closed water systems.Semi-closed pond systems have a lowwater turnoverandcanaccumulategases,nutrients, metabolites, waste, etc., whichcan deteriorate the water quality andcreate anoxic conditions in the soil. Thiscanstronglyaffecttheperformanceofthefarmedspecies.Thus,goodpondmanage-

Figure 2: Pond interactions without the addition of beneficial bacteria

Figure 3: Pond interactions with the addition of beneficial bacteria

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-

duced on a large scale it could bring fishproduction to the place where the fish isactuallyconsumed.

The Marine Harvest Group, the world'slargestprivate fishproducer, isbuildingever-largertanks.AtKårstø,Norway,thecompanyhas plans to produce 6,000 tons of salmonper year, onshore, to an average size of 1kilogram, in fish tanks40metres indiameterand10metreshigh,withvolumesofwaterashighas12,000cubicmetres.MarineHarvestalso intends to explore the possibility ofbuildinga landbasedfish farminaquarryatMjølkevikvarden, in Askøy, Norway, wherethe company believes there is potential tobuild a plant big enough to produce 50,000tonsperyearof1kilogramsalmon.

This is equal to 5 percent of the totalbiomass production of salmon and troutin Norway today — currently about 1 mil-lion tons per year. Theoretically, if 20 oftheseplantswhere inoperation today,allofNorway’ssalmonproductiontofullslaughtersize could be accomplished on land. Keyparameters associated with this installationare20,000cubicmetrefishtanks,35metresindiameter.

Pioneering technologyOneof themostpioneering technologies

to be introduced to the industry has beenLinde’s SOLVOX® OxyStream system. The

uniquenessof thetechnology isbasedon itsability to perform three critical functions inonesystem-dissolvingoxygen in thewater,producingthecorrectmarinehydrodynamicsandstrippingoutpotentiallyharmfulnitrogen- and all this via a very low energy require-ment.Thesystemiseasilyinstalled,asanewset-up or as a retrofit to existing fish farmtanks, and is maintenance-free because it isnot associated with any ancillary equipmenttomanagewaterpressure.

SOLVOX® OxyStream significantlyincreases fish production volume, optimisesfish meat quality and considerably improvesoperationsfromanenvironmentalstandpoint.Itisacombinedoxygenationandflowsystemthat not only dissolves the optimal amountof oxygen in the inlet water flow, but alsodistributesitevenlyatanadjustableflowpat-tern throughout the tank, ensuring that thefish stock benefit from the physical exerciseinvolved in swimming against the flow. Theflow regime can be fully tailored accordingto fish size, stock density and fish species,suchassalmonorcod.Thesystemcomprisesa standalone unit, allowing water flow andoxygen dosing to be individually controlledforeachtank.

Themicro-bubblescreatedbySOLVOX®OxyStream create the additional benefit ofhelping to reduce the concentration of dis-solvedinertgasessuchasnitrogen,argonand

carbon dioxide. In particular, oversaturationofnitrogen,eveninrelativelysmallquantities,canendangerthewellbeingoffishstock,slow-ing growth and increasing the possibility ofdisease, and ultimately, even mortality. WiththeinstallationofOxyStream,externaldegas-singunitstopreventinertgasbuild-upwill,inmanycases,becomeobsolete.

Depending on the application, pumpingpressuresas lowas0.05 to0.2bararenor-mally sufficient to oxygenate the incomingwater,stripnitrogenandcreateoptimaltankhydrodynamics. This low operating pressuremakesthesystemveryenergyefficient.

The capabilities of this technology wereproved during trials conducted at a MarineHarvest facility in2011.Results showed thatOxyStreamwastheonlyoxygenationsourcesuitable for rearingyoung salmonhatched intanksrunningonfreshwater,beforegraduallytransitioning them to seawater. This createsan optimum environment in which to rearsalmon, ensuring the correct oxygen levelsthroughouttheentireproductionperiodandkeeping fish stress levels to an absoluteminimum.

Importantly, thetechnologymakes itpos-sible to precisely predict flow velocity andto adjust this velocity in the circular on-landtanks, Depending on their state of matu-rity, fish need a certain water velocity toremainhealthy. If the velocity is not correct,

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Extruder OEE for the Production of Fish FeedExtruder OEE for the Production of Fish Feed

AMANDUS KAHL GmbH & Co. KG, Dieselstrasse 5-9, D-21465 Reinbek / Hamburg, Phone: +49 40 727 71 0, Fax: +49 40 727 71 100, info@amandus-kahl-group.de www.akahl.de

Volume 16 / Issue 2 / March-April 2013 / © Copyright Perendale Publishers Ltd 2013 / All rights reserved

ment is crucial for high production anda healthy crop. Since water quality playsan important role, it is of great value tounderstand the various interactions takingplace within the ponds. These are quitecomplexanddependdirectlyonthepondenvironment, stocked biomass, input ofnutrientsandpondmanagement.

AscanbeseeninFigure2,theaccumula-tionanddegradationoforganicwaste inthepondwillresult inan increasedconsumptionof oxygen (O2) and production of wastecompoundssuchasammonia(NH3),nitrites(NO2 -) and hydrogen sulfide (H2S), whichcan lead to aphytoplanktonbloom.Massivegrowthofphytoplanktoncanfurtherdepleteoxygenduring thenight and contribute to aphytoplanktonbloomcrash.All these factorscontribute to the contamination of waterand soil, creating favourable conditions for

pathogenstogrowandaffectingtheconditionof the shrimp.Under thesepoor conditions,the shrimp faces higher levels of stress andismore susceptible to diseases,which couldresultinpoorgrowthorafailedcropthroughdiseaseoutbreaks.

With the inclusion of beneficial bacteria(Figure 3), organic matter is utilized as asource of nutrients by the bioremediationbacteria,whichreducestheamountofwasteaccumulatinginthepond.Additionally,specificnitrifyinganddenitrifyingbacteriawillconvertNH3 and NO2- into nitrogen gas, reducing

the level of such toxic compounds. SomebeneficialbacteriacanalsodegradetoxicH2S,improvingwaterqualityandodor.Thecom-binationofallthesefactorswillimprovewaterquality and the condition of the pond soil,resulting in a better environment for shrimpwithbettergrowthandhealthstatus.

In the bioremediation process, enzymesplay the role of catalysts that acceleratebiochemicalreactionsinpondsoilandwater.Whenaddedtotheculturewaterorspreadon top of the pond soil, enzymes are ableto degrade the major organic constituentsnormallyfoundinshrimpandfishponds.Eachenzyme has its mode of action and is veryspecific in the chemical reaction it catalyzes(Table1).

Enzymes are also naturally produced andexcreted by some microbes. These extracel-lular enzymes, such as cellulase, protease andamylase,areproducedduringtheaerobicfer-mentation of organic matter by micro-organ-isms, for example by some Bacillus species.Bacilliarecommonlyfoundinpondsedimentsandcanalsobeaddedtothepondwaterforbioremediationpurposes.SomeBacillus sp.arealsoabletodegradenitrogenouscompounds.In addition, their large variety of excreted(extracellular)enzymeshelpstospeedupthedegradationoforganicmatterandtoxiccom-poundssuchasammonia.TheefficientremovalofnitrogenouscompoundscanalsobecarriedoutbynitrifyinganddenitrifyingbacteriasuchasThiobacillus andParacoccus.

Table 1: A diverse range of enzymes used as bioremediation agents in aquaculture

enzyme substrate

amy;lase ß-Glucoside

Cellulase Cellulose

lipase lipids and fat

Protease Protien

Xylanase Xylan, Hemicellulose

Peectinase Pectin

Figure 4: Average growth rate (g/day) of shrimp during the production period

30 | InternAtIonAl AquAFeed | March-April 2013

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March-April 2013 | InternAtIonAl AquAFeed | 31

Whilesomemicro-organismsproliferateina narrow range of environmental conditions(pH,oxygen,availability,etc.),certainenzymesareabletoactinmultipleenvironments.

Nevertheless, some products combiningthepositiveeffectsofbeneficialbacteria andenzymesarealreadybeingusedasbioreme-diationagentsinaquaculture.

Efficacy of enzymes in bioremediation

Enzymes have the capacity to stabilizethe soil organic matter and can be usedeffectively tomanage soil quality and rear-ing conditions for aquatic species.There isnot one specific enzyme that works bestinallcases.Ablendcontainingavarietyofenzymesmaybe themosteffectivemeansforbioremediationinaquaculture.Enzymesgreatly reduce sludge accumulation andanaerobic conditions in pond bottoms.They promote a faster degradation ofthe accumulated organic matter especiallyunder intensive production conditions.This organic matter comprises uneatenfeed, dead plankton, mineral soils, faecesandpathogenicmicro-organisms in thesoilwhere the conditions are often anaerobic.However,forallthesebioremediationproc-esses catalyzed by enzymes, the presenceof beneficial bacteria is important as well.Enzymes acceleratemicrobial processes by

breaking apart large sludge particles, thuscreatingwidersurfaceareaswhichcanthenbe fermented by microbes. This reductionof sludge and dead organic matter can beseenvisuallynotonlythroughbetterwaterquality,butalsothroughbettersoilquality.

Field trialInafieldstudy inChina, itwasobserved

that the combined application of thebioremediation products AquaStar® Pond(Bacillus sp., Enterococcus sp.,Pediococcus sp., Paracoccus sp., Thiobacillus sp) andAquaStar® PondZyme (beneficial bacteriaandablendofamylases,xylanases,cellulasesand proteases) to the water, according toa specific application programme, improvedwater quality, soil condition and ultimately,shrimpperformance.

Four earth shrimpponds (0.7 – 0.8 ha/pond) with a depth of 1 – 1.2 m werestockedwithjuvenileshrimp(approximately1.4 g/shrimp)with a density of 50 shrimp/m². The trial was carried out for a periodof 57 days with a dosage of 500 g/ha ofproduct applied once a month to thetreatment group (twoponds).The controlpondsconsistedoftwopondswithnormalproductionoperations.

The soil of the AquaStar® ponds inPicture 1 was of yellow colour which isregarded as the best bottom type, while

the soil of the control ponds in Picture 2exhibited a dark black colour, an indica-tion of the accumulation of dead organicmatter.

Resultssuggestedthatwiththecombineduseofbeneficialbacteriaandenzymes,pondsoils containing black and glutinous organicsludgeturnedintoamoreyellowsoil.

Intermsofperformance,theaveragedailyweightgainofshrimpintheAquaStar®groupincreasedby36percentandfeedconversionratio improvedby9percentcomparedwiththe control (no probiotic inclusion). TheresultsareshowninFigure4and5.

Basedon theseresults, itwasconcludedthat in the search for more effective andenvironmentally-friendly treatments, benefi-cialbacteriahaveemergedasaviablealter-native. The application of bioremediationsolutions in aquaculture can also benefitfrom the inclusionof enzymes, especially inintensive productions. AquaStar® positivelyaffects the performance of shrimp whilemaintaining a stable environment in thepond, proving to be an effective manage-menttoolinaquaculture.

More InforMatIon:Website: www.biomin.net

30 | InternAtIonAl AquAFeed | March-April 2013 March-April 2013 | InternAtIonAl AquAFeed | 31

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