CAPTURE, ADAPTATION AND ARTIFICIAL CONTROL OF
AnimalReproduction
Science
j o u r n a l ho me p ag e:ww w . e l s e v i e r . c o m / l o c a t e / a n i r e p r o s c i
Capture,adaptation
andartificialcontrol
ofreproductionof
Lophiosilurusalexandri:
Acarnivorousfreshwaterspecies
DelianeCristinaCostaa,WalissondeSouzaeSilvaa,ReinaldoMelilloFilhoa,
KleberCamposMirandaFilhoa,JoséClaudioEpaminondasdosSantosb,
RonaldKennedyLuza,∗
aLaboratóriodeAquaculturadaEscoladeVeterináriadaUniversidadeFederaldeMinasGerais,Av.AntônioCarlos,6627BeloHorizonte,
MG,Brazil
bCentroIntegradodeRecursosPesqueiroseAquiculturadeTrêsMarias–CODEVASF,CaixaPostal11,TrêsMarias,MG,Brazil
a r t i c l e i n f o
Articlehistory: Received18March2015
Receivedinrevisedform8June2015 Accepted9June2015
Availableonline12June2015 Keywords: Breeding Captivity Conservation Parentalcare Transport Lophiosilurusalexandri a b s t r a c t
ThepresentstudydescribesthecaptureadaptationandreproductionofwildLophiosilurus alexandribroodstockinlaboratoryconditions.Thereweretwoperiodswhencapturing wasperformedinnaturalhabitats.Theanimalswereplacedinfourtanksof5m3with watertemperaturesat28◦Cwithtwotankshavingsandbottoms.Thirtydaysafterthe temperatureincreased(duringthewinter)thefirstspawningoccurrednaturally,butonly intankswithsandonthebottom.Duringthebreedingseason,therewere24spawning boutswitheggmasscollectionsoccurringasaresultofthespawningboutsthatoccurred inthetanks.Thehatchingratesforeggsvariedfrom0%to95%.Thespawningboutswere mainlyatnightandonweekends.Inthesecondreproductiveperiod,theanimalswere sexedbycannulationanddistributedinfourtankswithallanimalsbeingmaintainedin tankswithsandonthebottomat28◦C.Duringthisphase,therewere36spawningbouts. Findingsinthepresentstudycontributetotheunderstandingofthereproductivebiology ofthisendangeredspeciesduringcaptivity.
©2015ElsevierB.V.Allrightsreserved.
1. Introduction
Thereproductionofwildspeciesincaptivityiswidely
used around the world as a conservation strategy of
endangered fish, and is a useful tool for the preserva- tionorrestorationofwildpopulations(Kelleyetal.,2006; Blanchetet al.,2008; Saravanan et al.,2013).However, thetaskofadapting wildbroodstocktolaboratorycon- ditions involvesmany processes, rangingfrom capture, transport,acclimationandmaintenanceoftheanimalsin
∗ Correspondingauthorat:LaboratóriodeAquaculturadaEscolade VeterináriadaUniversidadeFederaldeMinasGerais,Av.AntônioCarlos, 6627BeloHorizonte,MG,CEP.31270-901,Brazil.Tel.:+553134092218.
E-mailaddress:luzrk@yahoo.com(R.KennedyLuz).
captivity(Moorheadand Zeng,2010;Maricchiolo etal., 2014).Impropermanagementoftheseactionsmayresult
in gonadal atresia and even lead to mortality of cap-
turedanimalsasa resultof thestressresponse(Barton etal.,2003; Ashley,2007; Murchieetal., 2009).Breed- ingin captivity also requires the study of reproductive biologyofthespeciesandthedevelopmentofstrategies formaintaininganimalsinconfinedconditions(Hunting Ford,2004;PankhurstandFitzgibbon,2006;Moorheadand Zeng,2011).Thisisanimportantconsideration because confinedconditionscanpromotedifferenceswithrespect tothenaturalbehaviorofthespecies.
The “pacamã” Lophiosilurus alexandri (Siluriforme:
PseudopimelodidaeSteindachner,1876)isaspeciesthat hasbeenused for restockingthe “São Francisco” River, Brazil.Thereproductionof“pacamã”hasbeenmadeinfish
http://dx.doi.org/10.1016/j.anireprosci.2015.06.009
reduced wildpopulationsof“pacamã” whichis nowan endangeredspecies(Brasil,2014).Itisanendemicfishof the“São Francisco”river (Travassos, 1960), carnivorous, benthicinhabitatpreferenceandhasnocturnalbehavioral patterns(Shibatta,2003;Tenórioetal.,2006).Reproduc- tioninthisspeciesoccursbetweenOctoberandFebruary (Satoetal.,2003a;Barrosetal.,2007)withthemalesof thespecies caring fortheeggmass (Satoet al.,2003b). Thefecundityofthisspeciesislessthanformanyaquatic speciesbutconsistentwithfecundityratesofspecieswith thesedentarynatureofthisspecies(Santosetal.,2013). ThesizeofvitelogenicoocytesofL.alexandriarelargeand havehighlyadhesivecharacteristics(BazzoliandGodinho, 1997;Rizzoetal.,2002;Guimarães-Cruzetal.,2009;Santos etal.,2013).However,reproductionincontrolledcondi- tionshasnotbeenstudiedforthisspecies.
Thepresentstudywasconductedtoassessreproduc- tivecapacityofL.alexandriwildbroodstockaftercapture, transport,andadaptationinlaboratoryconditions.
2. Materialsandmethods
The present research hasfollowed the methodology
approvedbytheEthicsCommitteeonAnimalUse,Protocol.
25/2010–CEUA/UFMG.
2.1. Firstcaptureofwildanimals
Tocapturetheanimals,alicenseofCaptureandTrans-
port was obtained by the State Institute of Forest-MG
ScientificFishinglicensen◦105/09.Thefirstgroupofani-
mals werecapturedintheregionof“PontaldoAbaeté” near “Três Marias-Minas Gerais”, Brazil (18◦ 20′ 16′′S,
45◦ 49′ 58′′W). The captureperiod was initiated atthe
1800hbetween16 and 26 September 2009.Amesh of
8–20mm wasusedfor thecapturingand animalswere
collectedthenextmorning(0600h).Specimenscaptured weretransportedina190Lboxboattothe“Companhiade
DesenvolvimentodoValedoSãoFrancisco”(CODEVASF)
where animalswere keptinconcrete tanksfor a week.
Thefishweresubsequently transportedtotheAquacul- tureLaboratory(Laqua)ofthe“UniversidadeFederalMinas Gerais,BeloHorizonte,MinasGerais”,Brazil.Thistrans- portwasconductedina1000Lboxwithaeration,water temperatureat22◦C,during4h(380km)oftransportation.
Inthelaboratory,theanimalsweresubmittedtoacclima- tion.Animalswerehousedinasingletankof7m3covered
withgeomembranelinerwithwaterconditionsasfollow: temperature between25◦ and 27◦C and dissolvedoxy-
gen>5.5mgL−1.Atthistime,eachanimalhadamicrochip
implanted(MicrochipParteners)inthelumbarregionfor identificationanddatacollection.Onceaweekabout50% ofthewaterinthetankwasexchanged.
InNovemberof2009,theanimalsweretransferredtoa newtankwiththesamemanagementspecificationsbeing imposed.Atthistime,twomoreanimalswerecapturedand submittedtothesameproceduresaspreviouslydescribed. Theanimalswerefeddailywithfrozenfiletsoftilapiaand theremainswerecollectedafter24h.
Thesecondcaptureoccurredintheregionof“SaoJosédo Buriti”inthereservoirof“TrêsMariasintheSãoFrancisco” RiverBasin(18◦ 42′ 33.2′′S45◦ 09′ 13′′W)in Januaryof
2011.Forthesecondcaptureperiod,changesinpractices occurredtoimprovetheefficiencyoftheprocessincom- parisonwiththefirstcaptureperiod.
Netswith1500mandmeshrangingfrom7to15cm
among knots wereused starting at the 1800hand the
animalswerecaptured thenextmorningat the0600h.
The captured animals were transported in a 190L box
with aeration. At the time of landing, the fish were
weighed,subjectedtoprophylacticantibiotics(sulfadoxine
and trimethoprim 75mgkg−1) and immediately trans-
ported to the Laqua as described for the first capture period.Thetransportlastedabout3h(distanceof269km). UponreachingtheLaqua,theanimalsweresubmittedto
a 30-min acclimation and treated with potassium per-
manganatebath(6mgL−1)for15min.Theanimalswere
subsequentlyallocatedtotwotanksof5m3coveredwith
ageomembraneliner,maintainedinastaticsystem,with additionalaeration(dissolvedoxygen>5.5mgL−1)andat
a controlled temperature (26.0±1.0◦C). The water was
salinized(1gL−1)inthefirst2days.Treatmentwiththe
potassiumpermanganatebathwasconductedagain2days aftercapture.
Theanimalsweremonitoreddaily.Onceaweek,about 50%ofthevolumeofwaterwasexchangedineachtank.
Theanimalswerefedtwotothreetimesperweekwith
tilapiafiletandavitamin–mineralpremix(600mgVitamin Cand30mgofpremix).Thefoodwasoffereddirectlytothe mouthoftheanimalsutilizingahook.
After 15 days, the animals were weighed on a dig-
ital scale (Digital instruments FG5020). At this time, a microchipwasplaceineachspecimenasperformedinthe firstgroupofcapturedanimals.
Duetothelackofmethodologyforsexualdifferenti- ation,thefishofthesecondcapturegroup(17animals) andsurvivinganimalsfromthefirstcapturegroup(seven
animals) were combined in March of 2011 and then
placed in one of four 5m3 tanks. The following condi-
tionsanddensities wereimposed:Tanks1 and4, eight
animals with an average weight of 2.304±1.33kg and
2.55±1.10kg,respectively;Tanks2and 3;fouranimals weighing1.67±0.36kgand1.860±0.43kg,respectively. InTanks2and4,therewasapproximately5–10cmofsand (poolfiltertype)addedtothebottomofthetank.
Tanks of 5m3 were maintained with a volume of
2–2.5m3. The volumewas reduced toavoid fishjump-
ingoutofthetank.Initiallythetemperatureofthetank wasmaintainedat26.0±1.0◦C witha dissolvedoxygen >5.5mgL−1andaphotoperiodof10hlight.About50%of
thevolumeofwaterwasexchangedweekly. 2.2.1. Firstbreedingperiod
InlateJuneof2011(5monthsaftercapturingthesecond
groupofanimals)thewatertemperaturewasincreased
to28◦C.DuringtheperiodfromJune2011toApril2012,
thetankswereinspecteddailyinthemorningandafter-
masswassubsequentlyweighedandplacedin50Ltanks maintainedat28.0±0.5◦Catanoxygenconcentrationof >5.0mgL−1forhatchingwithprocesslasting24–48h.
2.2.2. Restingperiod
Due to the lack of spawning between January and
April of 2012, a reduced temperature was employed
(22.0±1.0◦C)inalltanksinMayof2012.Theanimalswere
fedoncea weekasdescribed previously.Thisperiodof lesserwatertemperaturewasmaintainedfor1yearand 3monthsbecauseofreconstructioninthelaboratory.In addition,thewaterinthetankscontinuedtobereplaced at50%ofthetotalvolumeonceaweek.
2.2.3. Secondbreedingperiod
InAugustof2013,theanimalsweresexedusingthe methodologydescribed byLopesetal.(2013)bydetec- tingtheoviductonthefemales.Forthesecondbreeding
period, the animals were prepared as follows: Tank 1:
fourfemales andtwomalesweighing2.47±0.60kgand 2.53±0.83kg,respectively;Tank2:threefemalesweigh- ing2.93±0.74kgandtwomalesweighing3.28±0.02kg;
Tank 3:three females weighing 3.21±1.23kg and two
malesweighing3.0±0.62kg;Tank4:fourfemalesandtwo malesweighing2.84±1.27kgand1.56±0.98kg,respec- tively.InTanks1and3,sandwasaddedonthebottom asdescribedfortheTanks2and4.Thebiomassofeach individualtankrangedfrom14to15kg.
Thewater temperaturein thetankwasincreasedto
approximately28◦C.Theanimalsstartedtofeedtwicea
weekand50%ofthewaterwasreplacedonceaweekas
previouslydescribed.DuringtheperiodofAugust2013to April2014,thetankswereinspecteddailyinthemorning andafternoontoverifytheoccurrenceofspawning. 3. Results
3.1. Firstcapture
In the first campaign were captured a total of 12
fish (between 3.6 and 4.7kg). Twoanimals died in the CODEVASFstationandtwoanimalsdiedinthefirstweek after being captured after being placed in the labora- tory.Thesefishweredissectedandidentifiedasfemales. Anotherinterestingpointwastheobservationofalarge amountofvisceralfat.For20days,allthefoodofferedwas collected,indicatingthatanimalswerenotfeeding.From thistime,theanimalsgraduallystartedtofeed.After33 daysofbeinghousedinthelaboratory,theanimalswere fedonthetanksurface,eatingoutofthehandoftheanimal caretaker.
Afterbeingtransferredtothenewtanks,theanimals didnoteatforalmost6monthsandoneadditionaldeath occurredduringthistime.Afterthe6monthperiod,there wasa returnofthefeedingbehaviorintheanimals.No spawningboutswereobservedduringthistime(fromthe firstcaptureinSeptemberof2009toMarchof2011).
Twentyanimalswerecapturedin2daysofcollection (15onthefirstdayandfiveinthesecondday)withanimals weighingbetween0.8and2.5kg.Onceinthelaboratory, threeanimalsdiedjumpingoutofthetank.Forthisrea- sonthevolumeofwaterinthetankwasreduced.Animals begantofeed4daysafterarrivingatthelaboratory. 3.2.1. Firstbreedingperiod
Aftercombiningthefishfromthetwocaptureperiods
and having the water temperature increased, the first
spawningoccurred. Nests were established by theani-
malsonthesandybottomof thetanks.Afterspawning
bouts,animalwithdrawalsfromthenestswereattempted andtheanimalsexpressingparentalbehaviorreactedby attemptingtocontinuetoresideonthenest.Thisbehavior wasrepeatedinallsubsequentspawnbouts.Onlyasingle andatnotimewastheremultipleanimalsobservedcaring forasinglemassofeggs.
ThefirstspawningoccurredinTank2,30daysafterthe increaseintemperature(Table1).Itshouldbenotedthat thisoccurredduringthewinter(inJuly).ByearlyDecember of2011,therehad been11 spawningboutsinthis tank witheggmassesweighingfrom46to191g.Thenumberof larvaevariedfrom240to4600.Allspawningboutswere protectedbyasingleindividualidentifiedbymicrochip.
Theaveragetimebetweenspawningboutswasapproxi-
mately14days(6–28days).
InTank4,thefirstspawningoccurredinearlyAugustof 2011,approximately60daysafterthewatertemperature increased.Inthistank,therewere13spawningboutswith theeggmassweighing55–150gandcontaining117–3500 larvae.Fromthetotal,12oftheeggmasseswereprotected bythesameanimalasidentifiedbymicrochipobservation. Theintervalbetweenspawningboutswas9.2days(4–27 days).
Some spawning bouts resulted in a 0% hatching for
unknownreasons.Thespawningboutswerealwaysfound
inthemorning,suggestingthateggfertilizationoccurred atnightandespeciallyonweekends,whentheactivityin thelaboratoryandespeciallyaroundthetankswasless.
Inbothtanks,itwasnotedthataftertheremovalofegg masses,thefishremainedonthenestfor1or2daysand becamemoreaggressive.Inthecourseofspawningperiod,
whentheanimalthatwasprotecting theeggmass was
removedfromthenest,otherfishimmediatelymovedto thenesttoprotecttheeggmass.
Anotherimportantpointisthattherewerespawning boutsonlyinthetankswithsandonthebottom(Tanks 2and4).InTanks1and3,withoutsand,therewerenot spawningboutsduringtheobservationperiod.
Duetotheadherencenatureoftheeggsandthelarge amountofsandintheeggmass,itwasnotpossibletoestab- lishtherelationshipbetweenfecundityandeggweight, andweightoftheeggmassandeggnumber.
3.3. Restingperiod
Theanimalsremainedmorethanayearatthelesser
Months/year Tank1/withoutsand Tank2/withsand Tank3/withoutsand Tank4/withsand June/2011 – – – – July/2011 – 1 – – August/2011 – 3 – 2 September/2011 – 2 – 2 October/2011 – 2 – 4 November/2011 – 2 – 5 December/2011 – 1 – –
January,February,March,April/2012 – – – –
Averageweightofeggmasses(g) – 102.9±36.6 – 126.9±42.4
Tank1:eightanimalswithaverageweightof2.30±1.33kg. Tank2:fouranimalswithaverageweightof1.67±0.36kg. Tank3:fouranimalswithaverageweightof1.86±0.43kg. Tank4:eightanimalswithaverageweightof2.55±1.10kg.
aInthisbreedingperiodtheanimalswasnotsexed.
Furthermore,althoughevenwithaconstantsupplyoffood, thebreedersremained6monthswithoutsearchforfood andnomortalitywasregistered.
3.4. Secondbreedingperiod
InAugust2013,afterarevisedarrangementofanimals inthetanks(accordingtothesex,sandaddedinTanks1and 3andanincreaseinthetemperatureto28◦C),spawning boutswererecordedinalltanks(Table2).
The firstspawningoccurredin Tank3,24 daysafter increasing thetemperature. Inthis tank, there were 12 spawningbouts(25–95g)andtheeggmasseswerealways protectedbythesameanimal.Theaveragenumberofdays betweenspawningboutswas11days(5–23days).
The first spawning took place in Tank 4, 54 days
afterincreasingthetemperature.Inthistank,therewere
18 spawning bouts (8–153g) recorded with three ani-
mals being identified as having spawned. The first six consecutiveneststhatresultedfromthespawningwere protectedbythesameindividual(chipnumber:40612). Thenestsfromthethreesubsequentspawningboutswere protected bya differentanimalthan theoneprotecting theinitialsix eggmassesthat werespawnedin Tank4
(chip number: 4237). The fish number 40612 returned
to protect theegg masses resulting fromthree consec-
utive spawning bouts. Subsequently, fish number 4237
alsoprotectedaneggmassspawning.Inthe14spawn-
ing,themalefishwasnotidentifiedbymicrochipreading.
Thesubsequenteggmassresultingfromaspawningwas
protected by anotheranimal (chip number 19227). The
last threenestswereprotectedbytheanimalwithchip
number 40612. The average number of days between
spawning bouts was 8.2 days (ranging from 3 to 16
days).
Thefirstspawningbout(Tank1)wasinJanuary2014
when there were five spawning bouts and all of the
nestsresultingfromthesespawnswereprotectedbythe sameindividual(30–145g).Theaveragenumberofdays
betweenspawningbouts was11.5days (8–16 days).In
Tank2,therewasonlyonespawning(42g)inNovember 2013.
4. Discussion
ResultsofthepresentstudyindicatewildL.alexandri canbeadaptedtofullycontrolledconditionsincaptivity wheretheyeatnormallyandreproduce.Findingsindicate thepossibilityofmanipulatingthebreedingperiodbycon- trollingthewatertemperatureandfeeding.
The management adopted in the second capture by
transportingonthesamedayafterthetimeofcaptureand applicationofprophylactictreatmentsenhancedtheadap- tationofanimalstolaboratoryconditionsasevidencedby themstartingtofeedin4dayswhileafterthefirsttime ofcapturefishittookmorethan20daysbeforefeeding wasinitiatedafterthetimeofcapture.AccordingtoBernier (2006),appetite reduction in fish occursin response to acuteorchronicstress.Thetransferofanimalsfromtheir naturalhabitattocaptivityinvolvesvariousmanagement practicesthat promotephysiological changesinthefish inducinganimal stress(Iversenetal.,2005; Portzetal., 2006;Harmon,2009).Themagnitudeanddurationofstress duringtheseeventscanlastforhourstodays(Bolasina, 2011;Fanourakietal.,2011).Fishweightatthetimeoffirst andsecondcapturesdiffered(3.6–4.7kg,and0.80–2.5kg, respectively).AccordingtoMekaandMcCormick(2005), largeranimalsmaybemoresusceptibletostress.
Inthepresentstudy,fishspentlongperiodswithout foodwhentemperaturewasmaintainedat22◦C.Fasting
inlowtemperatureswasalsodemonstratedinotherfish species(Sunetal.,2006;Oyugietal.,2012;Wuetal.,2015). InL.alexandri,thelargeamountofvisceralfatcouldbean energyresovoironwhichanimalsdrawduringperiodsof fasting.Thesestoresoffatweredetectedinanimalsthat died and were dissected in the present study. In other species, the lipidsstored as perivisceral fat function as asourceofenergyand canensurethefishsurvivaldur- ingfasting(VanDijket al.,2005;Arringtonetal.,2006; Ibarzetal.,2010).Inaddition,prolongedfastingoccursin responsetothermoregulatorymechanismsoffish(McCue, 2010).
Inrelationtowatertemperatureandsandpresencein thetankforbreeding,theimportanceofthesefactorswere evidentinthepresentstudy.Thespawningboutswerenot observedwhenfishweremaintainedintankswithwater temperaturesof26◦Cafterthefirstcaptureandwithout
thetanksforspawning.
Months/year Tank1 Tank2 Tank3 Tank4
August/2013 – – – – September/2013 – – 1 – October/2013 – – 2 1 November/2013 – 1 4 4 December/2013 – – 4 4 January/2014 1 – 1 5 February/2014 3 – – 2 March/2014 1 – – 2 April/2014 – – – –
Averageweightofeggmasses(g) 81.0±48.6 42.0 69.4±23.1 71.2±45.1
Tank1:fourfemalesweighing2.47±0.60kgandtwomaleswith2.53±0.83kg. Tank2:threefemalesweighing2.93±0.74kgandtwomaleswith3.28±0.02kg. Tank3:threefemalesweighing3.21±1.23kgandtwomaleswith3.0±0.62kg. Tank4:fourfemalesweighing2.84±1.27kgandtwomaleswith1.56±0.98kg.