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Anemia in Experimental African Trypanosomiasis
Ogbadoyi, Emmanuel O., Ukoha, Agwu I.,
Kyewalabe, Elizabeth K.
The Journal of Protozoology Research, 9(2): 5563
1999-04
http://ir.obihiro.ac.jp/dspace/handle/10322/317
The Research Center for Protozoan Molecular
Immunology
帯広畜産大学学術情報リポジトリOAK:Obihiro university Archives of Knowledge
J．Prot（）ZOOI．Rcs．，9，55−6ユ（1朔）
C叩γrfg加⑥Jタワ夕．mど月ど∫gd′dC〝眈rβrPr〃加㈹柁〟bJど川血rJmm〟〃ロわgγ
AnemiainExperimentalAfricaLnTiypaJ10SOmiasis
EMMANUEL O．OGBADOYIl），AGWUI．UKOHA2）ANr）
ELIZAl∃ETH K．KYEWALA13E3）
／舶那加鋸肋止血椚叫′Z〟γれ好打d】川〟5上月fg′ⅣJgビrれ2D叩招rfmど”′げ別oJ咽fc〟J5c椚−Cど5′
Fビ加r〟川′∼加r5和好花eJ川OJpgy′旭〃闇′Nfgどγれヱ翫pf亡〟JDf5ビ〟5ビS尺ど5ビβrr7−Cピ〃恒ノ
P．0．月8エ7コ7占g′N由れZ爪肌あ㍍
Rccej−−edlM町Ch】9りウノAl℃ePledlユAprilL9粥
Kcywords：丁わp【1〃0∫∂〝MリルdJ．■anemia；meChanjsm＝r）’panOSOmiasi5．
ABSTRACT
Anemiaisarecognizedfeatureofbothanimalandhumantrypanosomiasis・
Themechanismoftrypanosomalanemiaishoweverunknown．Understandingthe
TrleChanism of anemiaisimportant for the effeetjve management of
trypanosomiasispatients，eSpeCiallylnanimalswhereanemiacouldbeextremely
SeVere，Anemia was monitoredin five goats experimenta］lyinfectedwith
Tbpanosoma vivax by measuring the packed ce11volume（PCV）and the
hemoglobinlevel（Hb）upto26dayspostinfection．Parasitemiaandserum free
fattyacid（FFA）1evelswerealsoestimated．Massiveparasitemiawasobservedon
days5and12postinfection・Therewa545−59％decreaseinthePCVand33−63％
decreaseinthe Hblevel．Theserum Fl仏1evelproBreSSivelylnCreaSedthrough
Outthecourseofinfection，Anemiaintheearlystagesoftheinfectionisinitiated
andmaintainedbylivlngtryPanOSOmeS，the severltyOftheanemiadependingon
thelevelofparasitemia．Persistent anemiaasthe disease processprogressesis
CauSedbyfactor（S）otherthanlivingtrypanosomes．Itissuggestedthaterythrocyte
haemolysIS and erythrophagocytosis are the underlylng CauSeS Of trypanosomal
anem】a．
INTRODUCTION
Africantrypanosomiasesare aspectrum ofdiseasesin mananddomestic
animalscausedbythetsetsetransmittedprotozoanparasite，tryPanOSOme．Huma
African trypanosomiasis（Sleeping sickness）is caused by TbpanoL”ma bruceE
ま∫
ANEMlA 1N AFRlCAN TRYPANOSOMlASlS
gambienseandT．b・rhodesiense・Animaltrypanosomiasis（naganaincattle）onthe
Other handiscausedbythreespeciesoftrypanosomes，namely T．b．brucei，T．
COngOZenseandT．vivax．Ofthesethreetrypanosomespecjes，T．vivaxisthemos
important trypanosome of veterinary importance in West Africa. Animal
trypanosomiasissti11constitutesamaJOrCOnStraintinthegrowthanddevelopment
Of thelivestockindustryin many eountries of sub−Saharan Africa，thereby
limiting10talfoodproduction．AnimaltrypanosomiasisandEastCoastfeverare
believed toaccountforthe deathof3mi11ion cattle ayearand oneor bothOf
these diseases occur（S）across morethan one third of the African continent
（ILRAD1987）．The estimated annuallossinlivestock due to African
trypanosomiasisisputatUS＄5，000mi11ion（WHO1997）．
Anemiahassincebeenrecognizedasam叫OrpathologlCalfeatureofclinical
trypanOSOmiasisintx）thmanandanimals（fiennes1954；1970）andhasbeenused
as anindicator of trypanotolerancein animalsinftcted with trypaT10SOmeS
（Akinbamijoetal．1998；drIterenetal．1998）．However，tOdate，themechanism｛）f
the anemiais unknown．Rather，hypo血eses whichhave failed t0月nd common
grollndofacceptablespecincityhavebeenputforward．Theseinclllde：hypoplasia
Ofthe bonemarrow，hydraemia，immunologlCalreactions，erythrophagocytosis，
hemorrhagesyndromeandseverehaemolysis（reviewedbyMurrayandI）exter
1988）．AmorerecentadditiontothislistofhypothesesisthereportofIgbokweet
al．（1994）thatperOXidativeipjury to erythrocytes may play a rolein the
pathogenesis of anemia in trypanosomiasis. A good understanding of the
PathogenesisoftheanemiaisimportantforabettermanagementofthepatieIltS，
espeCla11ylnanimaltrypanosomiasiswheretheanemiacouldbeverysevere・This
Studyis part ofour contribution to the efforts towardsthe elucidation ofthe
mechanismofanemiaintrypanosomiasis・
MATERIALS AND METHODS
什り柚町、・岬‥、−
Tbpanosomavivaxstabilate，StOCkKabam／84／NITR／7・4wasobtainedfrom
the NigerianInstitute for Trypanosomiasis Research，Vom，Nigeria．This was
inoculatedinsheep■sbloodandkeptlnliquidnitrogenat−196bcbeforebeingused
ingoats．
（JIMJ∫．・
GoatswerepurchasedfromlocalmarketsinZaria，NorthernNigeriaand
keptinfly−prOOf perISin the facu］ty of veterinary medici11e，Ahmadu Bello
Universlty，Zaria・ApproprlateStepSWeretakentoensurethatallthegoatswere
ingoodhealthandapparentlyfreefromtrypanosomiasisandanyotherformof
ぎ首
ANEMlA IN AFRlCAN TRYPANOSOMlASlS
ParaSiticinfectionbeforetheexperimentalinfection・
Cんβ椚fc（】血．・
Unless otherwise stated，a］1chemicals used were obtained from Sigma，
England，andwereallofanaJytlCalgrade．
均匹雨脚舶用抑血肌
Tengoatsweredividedinto2groupsof5each・Onegroupof5wereeach
infectedwithlO7T．vivaxce11swhilethesecondgroupof5wereuninfectedand
servedascontrol．
PdJⅥ∫fJg椚Jdα〃dク〟CたどdcβJJt，OJ〟用ビrPCVノご
Parasitemia and PCV were deterrnined usJng the haematocrit centrifuge
technique（W〔）0】97（））・
〃g椚〃gJぴぁf〃“ノ〃Jピ〃J．■
ThehemoglobincontentwasestimatedbysimplytakingalittlequantltyOf
bloodsampleandthenreadingoffthehemoglobinlevelinaCounter
haemoglobinometer．
J、、〃〃J・い用い、い‥、州、川・声い・い．トJJl・∫・・バ・
FFAlevelswereestimatedasdescribedbyFalhoILetal．（1973）・Allthe
parametersweremeasuredessentiallytwiceaweek，beginnlngfromdayOtoday
26postinfection・
REStTLTS
PりJ・り∫情，川扇．、
Therewerecyclicalnuctuationsinthenumberofparasitesincirculation
whichresultedin3successivewavesofparasitemiawithpeaksondays5，12and
22postinfection（Fig．1）．Between days5and8，the number oIlparasitesin
Circulation decreasedsharply，reaChingaverylowlevel（1essthan halfthepeak
Value）by day8・The second phase ofdeclining parasitemia whjch was rnore
drasticthanthefirstocc11rred betweendays12and19postinfection・Byday19，
thelevelofparasitemiawasthesameasthatonday2（6timeslessthanthepeak
Value on day12）．Thepeak valueson days5and12were the sal一一e（about4
trypanosomes per microscopefield），While the value for day22was aboutl
lrypanosomeperfield．Theva］uesg】VenareanaVerageOfthevaluesforthefive
gOatS・
ANEMtA TN AFRICAN TRYfnNOSOMIASIS
Figurel・DeveJopmentofparasitemiathrough
山e course orin托ction．Ani11uslmtわn of 仇e
CyClical fluctuatJOnSin parasitemia duriTlg the
VeJticalbars represent standard errors of mean
つL
吋叫EO︼叫S已dh
COurSeOftheex匹hmen血infecltonwhich】a＄ted
払ー払days．臨chpointon仙eve摘録1axisisthe
m組nOf也eYalueso一也efiveh触短dgoats．The
（SEM）．
Ul
∈）
れ
⊂〉
■・→r−M
レ「
r】
Q
｛
Day5（pDStimfecLion）
J，油・Å．・．J．・lJ／い巾〃Iけり一（l−Jご
Ⅶth the exception of goat number3808which showed no apparent
decreaseinPCVbyday2postinfection，thePCVofalltheinfectedgoatsshowed
Signi鮎antdecreasesbyday2，withasmuchas25％decreaseingoatnumber3845
（Fig．2）．Thedecreaseattheendoftheexperimentranged between45−59％．In
general，therewasaprogressivedecreaseinPCVofa11theinfectedgoatsuntilthe
terminationoftheexperiment．
Figure2．The packedcelJvolum芦Ofdle
im飴ctedgoatsdurlngthepeddofinfection
This shows the decliningPCV duringthe
26rday period of expenmentalinfection．
Eachpo）ntOntheverticalaxisistheneanof
仙e valu8SOfthe flveinfヒc便d go且暮SWhile
the SEM are represented by the ve鵬cal
bars．
C
＞「
【＝）
r【†・・」N
Ln
「】
く＝〉
rl
D往）・S（POStinf￠C暮ipn）
〃g椚￠gわぁ血「励ノ励gJ．・
LikethePCV，theHblevelsofa11theinfectedgoatsdecreasedsignificantly
朗
ANEMIA TN AFRTCAN TRYPANOSOMTASTS
duringthecourseofinfectionwithanaverageof48％reductionexceptforgoat
3844whichhaditshemoglobinlevelreducedbya5muChas63％beforeitfinal1y
died（Fig．3）．
Figure 3．The hemoglobinlevels ofthe
infected goatsduTlr［gtheperiodofinfeetion．
Thisillustrates（he decreasein Hb］evels with
VeJticalbarsrepresentSEM．
ムり
︵一pも︶芯已t上H
progressiorlthroughthediseaseproces5■7hch
poJntOr）dle Vertic且1axisIsthe mean ofthe
Ya山es of山e fjYeinfected goats while the
く︶
⊂〉
ln
⊂⊃
レ1
⊂コ ー・｛−・｛M
「】
r′
ロays（pos【infec【i8n〉
∫gr〟椚／佗gβrJγ〟Cfdr錯弓りJgveJニ
TherewasaprogressiveincreaseintheserumFFAlevelofalltheinfected
goats（Fig・4）・Thevaluesofalltheparametersmeasuredina11thecontrolanimals
did not show any appreciable change but remained essentially constantwitll
∝CaSionaldroporrisebylunit（datanotincluded）．
Figure4・Serumfreel加yacid（FFA）1evels
Of血ein托ctedgoats．Ser11mFEAlevelsoflhe
infected goats measured over the 26−day
PerlOdofinfection・EachpoJn10ntheverticaI
axisis the mean of the va］ues of the nve
ex匹nmenta11yinfected goats，Vellicalbars
r叩reSentSEM・
⊂⊃l（
（つレ1
く⊃
「」
lハ
⊂⊃
「l r’￣
D8yS（postinfec【ion〉
且9
ANEMlA 1N AFRlCAN TRYPANOSOMlASlS
DISCIJSSION
AnemiaintrypanosomiasisproceedsintwodistinctbutoverlapplngPhases，
eachphasebeing mediatedbydifferentmechanisms（Murray andDexter1988）・
Thefirst phaseis believedto bemediated byfactors releasedby degeneratln
trypanOSOmeS（Tizzardetal．1978b；ILRAD1987）andredcelldamagebyliving
trypanosomes（Banks1979；Esievo1983）．Ourobservationsinthisstudyarein
SuppOtt Oflivlng tryPanOSOmeS Playlng a key rolein theinitiation and
maintenanceoftheprogressivedevelopment ofanemiaintrypanosomiasis・The
onset ofanemia andthe severltyimthe nrst phase strongly correlate witht
appearance，intensity．and durationofparasitemia・Theobserved considerable
declineinthePCVandtheHbcontentbetweendaysOand5postinfectionandthe
relativelystablevaluesbetweendays5and8（Figs・2and3）whichcoincidedwith
increasinganddecreasingparasitemiarespeCtively（Fig．1）couldnothavebeena
merecoincidence．ThePCVandHbcontentagaindecreasedsharplybetweendays
8and12which wasanOther period ofincreaslng paraSitemia・ThesefindiTlgS
SuggeStthatlivlngtrypanOSOmeSareeSSentialfortheprogressivedevelopmen10f
trypanosomalanemiaininfectedamimals・Ikedeetal・（1977）reportedtheabsence
OfhaemolyslnintheplasmaofT．congolenseinfectedmiceatleastupto15days
POStinfectionbutobservedmarkedreductioninerythrocytehalf−1ifeandmarked
dropin PCVwithin the same period．In cattleinfected with T・COngOlense
increasedred cellbreakdownisbelievedtocommence withthedevelopmentof
parasitemia（HoImes1976；Prestonetal・1979）・Thereforecontrarytothewidely
heldviewthatanemiaintrypanosomiasisisinducedbyfactorsreleasedbydying
trypanosomes，Webelievethattheonsetofanemiaanditsprogression，atleastin
the early stages ofinfectioT），dependslargely on the presence oflivlng
trypanosomesincirculation・
ThepersistentanemiaevenwhenparasitesarebarelydetectabIeisthephase
IIoftheanemia・Webelievethatthisstageismediatedbyfactorsotherthanthose
producedbylivlngtryPanOSOmeS・FFAsandLysophospholipidsarebelievedtobe
hemolytlC factors released by dying trypanosomes and are responsible fo
trypanosomalanemia（Tizzardetal・1978a；Noketal・1993）・Weobservedsteady
andsharplnCreaSeSintheFFAlevelsofalltheinfectedgoatsthrough outthe
period ofinfection（Fig・4）and this could have played a key roIeiTlthe
pathogenesisoftheperSistentanemia・Althoughserumalbuminisknowntobind
toFFAstoneutralizetheirhemolyticeffects（StarinskyandShafrir1970；Tizzard
etal．1978b），itispossiblethatthema5Siveparasitemiathatcharacterizedthefirst
twosuccessivewavesofparasitemiaproducedsufficientFFAstosaturateavai1abl
serumalbumin and someleft unbound to exert their hemolytlC effect on
erythrocytes・
甜
ANEMIAIN AFRICAN TRYP4NOSOMTASIS
Elucidationofthemechanismofanemiaintrypanosomiasishasremainedan
unsoIvedproblem・Jenningsetal，（1974）suggestedthatanemiaintrypanosomiasis
WaSmainlyduetoextravascularhaemolysIS andruledoutirnmunemechanisms
Sinee（heonsetofanemiawasevidentwithinafewdaysofinfection・Murrayand
Dexter（1988）explainedthaLimmunologicalcompetencewasnotessentialforthe
developmentofanemia．DargieetaJ．（1979a；1979b）andDargie（1980）ruledout
haemodilution as a contributory factor to the development of anemia・
Hemorrhagic syndrome whichis characteristic of acuteinfections caused by
Certainstrains ofT・Vivaxinfections wasnotevidentin thepresent study．The
markeddropinPCVandthemarkedreductjonintheHbconten（Observedin［his
WOrkaregoodindicatorsofhaemolysis・Wethereforebelievethatthefirstphase
Ofanemiaislargelyduetoerythrocytehaemolysis．Erythrophagocytosis resultin
frominducedalterationsinerythrocytesurfacestructure（Durocheretal．1975；
Esievo et al・1982；Esievo1983）is alsoapossibility as acauseofanemiain
trypanosomiasis. We therefore conclude that trypanosomal anemia is
multifactoriallyinduced butthe predominantfactorsdependonthe stageofthe
diseaseprocess．
ACKNOWLEDGEMENTS
This study was funded by a research grant from the Ahmadu Bello
Universlty，Zaria，Nigeria，tOAgwuI．Ukoha．The authors are gratefulto：Mr．
RaphaelandMa11amBalla，bothof（heDepartmentofVtterinaryParasitology；Dr・
Jagun of the Nationa］AnimalProduction ResearchInstitute，Zaria for their
technical assistance and Professor K. A. N. Esievo for allowing us to use
haemoglobinometer．
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