DTIC ADA270015: Plasmodium Coatneyi Ring-Infected Erythrocyte Surface Antigens

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MAY 27 '93 09:32PM US6A. MED&, , ,6 476034 ". .
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4. PTLITALES MAONDD IUSUMO TitCLEO ATNMYI RING INFECTED ERYTHIOCYTE SURFACE Uwi( US~
ANTIGENS
S. AUTHOR($)
Eacblanee Udomsangpetch, Arthur E. Brown, C. Dahlem Smit
and H. Kyle Webster
7. PE-RFOgMING QqGA,.,ZATION NAMI(S) ANG ADORES5(ti) 6. P(AFOAMI4NG Q~r4ANIZAION
Walter Reed Army Institute of Research o dIJUMBIR
-oP
Washington D.C. 20307z5100
9. SPONSORJIA1/MONITORiNG AGENCY NAMV,5) AND ADORMS(ES) 10, SPOI5ORJNO/MQNiTOAIdG
U-S. Army Research And Development Cammand AGIN aRIt&"
Ft. Detrick. Frederick, MD. 21702-5012
11. SUPPL~iEMETARY NOTES
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Approved for piblic release
Distribution unlimi-tted
13. AISTLA7 (Mazimurn 00 otasJ
Plasmodium coatneyi produced rilg-infected erythrocyte surface antigen (USA) during
infection of the rhesus monkey. This antigen was immunogenic ýind elicited an antibody
response that awe not persistent but was boosted by repeated infecrtions iMa manner similar
to that seen in P. falciparum infectious in humans. Preliminary data showed that the
appearance and increasing titexof antibodies to P. coatueyi RESA-like antigen were
associated with prolongation of intervals from innocualation to potency and with controL
of parasitemia. Studies using both imunofluorescence assay and western blot analysis
shoved that P. coatneyi-inmune rhesus serum cross-reached with P. falciparum antigens,
but P. falciparum immune human serum Uid not recognize .P. Coatneyi antigen in either
assay. These results show that P. Coatneyi express RUSA-like antigen that elecits an
antibody response similar to that observed for human antibody to P. falciparuz RESA.
However, antibodies to P. coatneyUi did not cross-react with P. falcipatum RESA in erythrocyt,
membrane imnuncfluorescence assay and dot inimunoblot analysis, suggesting taht different
immunogenic epitopes are present on the two molecules. Our observations support the use
of this primate model in RESA-based vaccine development.
14. SU&iECI TERMS 15, MIMSIRA Of ,
Plasmodium coatneyi, ring-infected erythrocyte surface antigen.
P. falciparum -r. PRicE"C Oo .
", IS(cid:127)CvV1 't (%,ASSIFICATION lIo, .. SCuRITY CLASSIFICATION 19. SICURITY CLASSIFICATION 2(cid:127)0. ,M,1AT'ON OF ASQT'AV
Op AREPORT ! Of THIS PAG| OF ARSTRACTW
N11E ~S0.Ob2S~SjtO tidj~d form ?48 'ne"
J. .ol.. 11.h. IlIv,. 49(1). 1993, pp, 127-134
k 0l1right A' 1 I99b fth e American Socletv of1Tropical filedicle aid ||v'gicne
PLAS3IODIUM CO..! TNEYI RING-INFECTED
ERYTHROCYTE SURFACE ANTIGENS
RACHANEE UDOMSAN(MPETCH. ARTHUR E. BROWN.
J C. DAHLEM SMITH. AND H. KYLE WEBSTER
Depart ment olIPathlohiokhlp. Faculty of'.Science. .tahidol t'nii-ersui'. Bangkok. Thai/and: ow n
IC
lDepartments o' immnunolo.- and I etertnarr .l ledictne, .irned Forces Research Insti tute of h'.edical m
Sciences, Bangkok, Thailand
-Abstract. Plasmodium coatnevi produced ring-infected erythrocyte surface antigen
(RESA) during infection of the rhesus monkev. This antigen was immunogenic and elicited
an antibody response that was not persistent but was boosted by repeated infections in
a manner similar to that seen in P..fa/ciparum infections in humans. Preliminary data V
showed that the appearance and increasing titer of antibodies to P. coatne'i RESA-like
antigen were associated with prolongation of intervals from inoculation to patency and
with control of parasitemia. Studies using both immunofluorcscence assay and Western
blot analysis showed that P. coatneyi-immune rhesus serum cross-reacted with P..fal-
ciparum antigens, but P .falciparitui mmune huma,, serum did not recognize P. coatnevti
antigens in either assay. These results show that P. coatney'i expresses RESA-like antigen
that elicits an antibody response similar to that observed for human antibody to P.
falciparum RESA. However. antibodies to P. coatnevi did not cross-react with P. falcip-
arum RESA in erythrocyte membrane immunofluorescence assay and dot immunoblot
analysis, suggesting that different immunogenic epitopes are present on the two molecules.
Our observations support the use of this primate model in RESA-based vaccine devel-
opment.
Plasmodium coatneyi is a non-relapsing ma- MATERIALS AND METHODS
laria parasite of macaque monkeys with a num-
ber of falciparum-like characteristics. Mature Infection of rhesus monkeys with P. coatneyi
asexual blood-stage forms sequester in deep tis-
sue vasculaturel 2 in association with the for-
mation of knob-like protrusions on the surface Four adult rhesus monkeys without prior ex-
of parasitized red blood cells (PRBC)-' and cy- posure to P. coatnevi were selected from a pri-
toadhere to microvascular endothelium.4 Fur- mate colony for study using a protocol approved
thermore, P. coatneyi trophozoite/schizont (T/S)- by the Committee of Laboratory Animal Care
stage PRBC have been shown to cytoadhere to and Use at the Armed Forces Research Institute
uninfected red blood cells (RBC) to form to- of Medical Sciences. Each animal was inocu-
settes., lated intravenously with blood-stage parasites
Little is known about the antigen character- of the type strain' of P. coatneyi. Three mon-
istics of P. coatneyi or of the specificity of elicited keys (AF-B298. AF-B464. and AF-G425) were
immune responses. Using rhesus monkeys (Ala- infected using I ml of a previously frozen sta-
caca ,nulatta) as the laboratory host, we inves- bilate (1.6 x 101 PRBC/pl. 90% hematocrit).
tigated whether P. coatneyvi produces ring-in- One monkey (AF-B425) was repeatedly infected
fected erythrocyte surface antigen (RESA) with fresh heparinized blood (1-2 ml) from do-
analogous to that of P.falciparum".I and whether nor animals with low level parasitemias (140-
there is immune cross-reactivity between these 520 PRBC/Pl). The other three animals were
antigens. Since RESA is a candidate molecule for followed through the course of infection, had
a blood-stage malaria vaccine,'-10 identifying a sera collected, and were killed for tissue stud-
nonhuman primate malaria model with these ies.4 while monkey AF-B425 was given curative
characteristics may facilitate the process of vac- treatment with chloroquine and twice rechal-
cine development. lenged.
127
128
UDOMSAN(PETCIH AND )TIHERS
Parasite and antigcn peparationsrl an eryvthrocyte membrane immunolluorescence
i'lasniodiumP tl'oaosaaiottndeut*y m1t w,zaws atsatkaeknn ffroomm inntfibectteedd (EiMth IuFn) fiaxsesda.y, a idr-edtericetdin. gT /RS-EsStaAg e wPeRreB Cp earnfodr mGeAd-
Shesus monkeys when the parasitemia (ring-stage) w
was 7-10%. Blood was collected in RPMI 1640 fixed, ring-stage PRBC. respectively, as de-
medium containing heparin. pH 7.4. and washed scribed by Perlmann and others.' Sera were se-
three times in a serum-free RPMI 16,10 medium. rially diluted from 1:5 to 1:3,125 using five-fold
Ater the uAffye rctoeaut wryaso arewmoavreedo, eaa 11%cec ellsl ussu-- diluti-oiendrsn,- tagpapnileiesodr 3tom .bwoathh. air-dried T/S-stage and
pension in phosphate-buffered saline (PBS) was GA-fixed ring-stage antigens tbr 30 min. washed.
stained with anti-hiuman immunogiobulin con-
made and used to prepare glutaraldehyde (GA)- stained with anti-human imo globuln con-
lixed. ring-stadgees cPibReB C bmyoPneorllaaynenrasn dfoort hReEsS."AT haesr e-mjuoguatnetde dw iwthi thfl u5o0r%es cgeilnyc eirsool thini oPcByS,a nanadtw eeaxsxh aemd-m
describedined using ultraviolet light microscopy. Rhesus
maining parasites were cultured in RPMI 1640 aed usin ultrol light miro resu
medim spplmenietdh 10%hea-mati- and! human control or immune sera were also
m\are tde diu mA B 's uh pu pmle a mn esen treud m , w2 i5t hm M10 %H E hPe Ea St- .in2a cmti SM uas ned i fieo r ss tari nsipn gc ofvP .ly fcitp oa rtu um y annd i Ib cdo atneoi s -
glutamine. and 20 tigiml of gentamicin. pH 7.4. antigens. respectively, to study antibody cross-
Cultures were kept for at least 30 hr in an in- reactivity elicited by the two parasites in their
cubator at 370C and in an atmosphere of 5% CO, naiural hosts.
plus air by continuous flow. Maturation of par-
asites to the Ti'S-stage in the first schizogonic IIcstern and dot blot anu!ysms
cycle after isolation was confirmed by exami- Frozen samples from infected (P. coatncti and
nation of Giemsa-stained thin blood film-sv-aiter
which RBC were harvested by washing twice with Pfjalciparum)a nd uninfected rhesus monkey and
serum-free RPM! 1640 medium.Asmallaliquot human RBC were diluted 1:10 in sample buffer
ofrpacked RBC was resuspended in an equal vol- (0.1 M Tris buffer. 10% sodium dodecyl sulfate
ume ofp 1% bovine serum albumin (BSA) in PBS [SDS], 9% glycerine. 0.1 M dithiothreitol, pH
and used for preparation of unfixed, air-dried 6.8) and boiled for 3 min. After the samples were
thin blood films. The remaining T/S-stage par- caopoplleiedd into a ann icSeD bSa-tpho, ly2a0c ruyl loamf eidaec h gseal manpdle pwroa-s
asites were resuspended in an equal volume of teins were separated by electrophoresis. The sep-
PBS and stored at - 70"C for Western blot anal- arated proteins were transferred onto nitrocel-
ysis. .lulose paper and incubated with 1% BSA in PBS
ai. P.d iciparumT hai isolate. BR 15, was main- for I hr. Proteins on the nitrocellulose paper were
taired in vitro as previously described'2 for one probed with immune or control sera diluted at
month in human group 0* RBC using the same 1:100 in 1% BSA-PBS for I hr, washed five times
culture medium as above. The PRBC were har-
vested when the parasitemia reached 8-10%. ei- ashinu m05an ITge nc2onn jugated with hocrawsteietdrha dish peraonxtii--
ther ring- or T/S-stage parasites, and handled dase for I hr. washed five times in Tween-PBS.
above. Uninfected RBC from group 0* human incubated with peroxidase substrate (4-chloro- I-
and rhesus monkey donors were prepared and naphthol in 15% methanol in PBS containing
fforor zWene satse ranb obvleo tt oa nparlyosveisd.e control preparations 00.0a1pi%ht hh ydor ogge1en5n% p eroxxidieteh)an of or !155i nm ai. aanndd rirninsseedd
with distilled
water.
A-lntibodies For dot blotting, 10 Ml of a solution containing
0.2 mg/ml of synthetic peptide (EENVEHDA)
Sera from P. coalneyvi-infected rhesus monkeys corresponding to the 3' repeat sequence of P1.
during one or multiple infections, pooled sera falciparum RESA (a gift from Dr. Troye-Blom-
from humans who had a P.falciparum infection, berg, Department of Immunoiogy, University of
sera from uninfected rhesus monkeys, and sera Stockholm, Stockholm, Sweden) was spotted onto
from malaria-naive human volunteers were col- a nitrocellulose paper and dried. As a control
lected and stored at -20*C. protein, BSA at a concentration of 0.5 mg/ml
was also spotted onto the paper. The nitrocel-
Immunofluorescence assay ilose was then incubated with 1% BSA and
A conventional immunofluorescence assay :,robed with immune human or rhesus monkey
(IFA) detecting intracellular parasite antigens and ý,.!ra and stained as described above.
REISA-LIKL ANTIGEN IN P' (o0 ,I' 129
RESULTS cured on three occasions. had serum collected
after the second and belore and after the third
Inhctton of rhesus inonket's with P. coaetfCi inlections. Antibodies to air-dried T/S-stagc an-
tigens increased with the second and third infec-
Four rhesus monkeys were infe~cted with blood- lions reaching titers of 1:625 and 1:3.125. re-
stage parasites of P. coamne 'vi. All four 1'. coat- spectively. Positivity by the EMIF assay. which
m'yi-naive animals developed patent parasitem- as not seen in three aniimals after one infection.
ias on days 2-4, which then increased to 3-10% was demonstrated to GA-fixed ring-stage anti-
in less than two weeks. On days 12-16. sera were gens (Figure 2) at a titer of 1:125 after the second
collected from three animals prior to their being infection and increased to 1:625 after the third
killed for pathologic study. Monkey AF-B425 inlection.
was followed through three separate infections
with P. coatneyi (Figure 1). After the first inoc-
ill'iion. a low level parasitemia (60 PRBC/MI o'f (ro's-rcactvfty of antibodies to P. coatneyi iith
bloud) wa.. 1:cz:ed on day 2. 1b e parasitemia P. rum proteins
became su'patent or day 6 and was again de- Sera collected from P. coatnevi-naive and -
tectable on diy 10. The highest parasitemia, 1.7 immune rhesus monkeys were assayed with P.
10 ' PRBC/Iml of blood, was detected on day .(,Jt l'ti and 1'. Walciparum antigens in the IFA
16 and the monkey was bled for serum collection and EMIF assay with air-dried T.'S-stage and
and given curative treatment with chloroquine GA-fixed ring-stage preparations. The results are
every day for three days. The monkey was in- summarized in Table 2. Naive monkey serum at
travenously reinoculated with P. coatneu"i 84 days a 1:50 dilution did not react with P. coatnevi or
later. On this occasion. parasitemia was detected P. falciparuin antigens. Rhesus immune serum
on day 9 and the highest parasitemia. 8 x 104 (RIS) from the third P. coatney'i infection at a
PRBC/,uI of blood, was observed on day 16. Se- 1:250 dilution reacted with both air-dried T/S-
rum was then collected from the monkey and the stage and GA-fixed ring-stage P. coatneyvi and
animal was given curative treatment as described also with air-dried T/S-stage P. falciparum an-
above. After a third infectious inoculation 75 tigens. but the RIS did not react with GA-fixed.
days after the second infection, parasitemia was ring-stage P fialciparuin antigens. Human im-
detected on day 13 and reached a peak of 2.8 x mune serum (HIS) at a 1:250 dilution reacted
104 PRBC/ul of blood on day 18. The monkey with both air-dried T/S-stage and GA-fixed ring-
was bled on the day of inoculation and again 22 stage A. falciparum antigens. but did not react
days later when the parasitemia had decreased with either antigen preparation of P. coatne*yi. In
to 40 PRBC/ul of blood, three days before the an EMIF assay using P.f alciparum. the HIS had
animal became aparasitemic and prior to the ini- an antibody titer of 1:6,250. Control. nonim-
Stiation of chemotherapy. mune, human serum at a 1:50 dilution did not
rtc,.ct with either P. coatnevi or P. falciparwn
Antibody responses to P. coatneyi in infected antigens.
Srhesus mnonkeyvs
Cross-reactivityo f antibodiest o P. coatneyi with
Plasmodium coatneyi-naive. rhesus monkey denatured P. falciparum proteins
sera at a 1:25 dilution did not react with P. coat-
neyi antigens using the IFA and EMIF assay with Plasinodium coatne.vi-immune rhesus mon-
air-dried T/S-stage and GA-fixed ring-stage P. key and P. falciparurn-immune human sera (RIS
coatneyi antigen preparations, respectively. An- and HIS) were analyzed using Western blotting
tibody titers to P. coatne.vi of infected animals under reducing conditions with crude antigens
are shown in Table 1. Sera collectedfrom three prepared from T/S-stagc P. coatneyi- and P.fal-
monkeys after their primary infections with P. ciparum-infected blood. Uninfected rhesus
coatne.vi, were assayed; a low positive anti-P. monkey and human blood were included as con-
coatneviantibody titer, 1:25, was detected in onri trol antigens. By probing the proteins with RIS
monkeywiththeair-driedT/S-stagebutnotwit i at a 1:100 dilution, P. coatneyi proteins with M,
the GA-fixed ring-stage antigen preparatior. values of 120. 100, 97, 80, and 45 kD were de-
Monkey AF-B425, which was challenged ar tected (Figure 3A, lane c). The RIS also detected
13 0 LUDOMSANUPETCII AND) O'rlERS
Rx Rx
2001000-
100,000
10,000
E 1,000
100-
o
0
10
I0 - • . . . . . .
04 12 1620 0 4 8 12 1620 0 4 8 12 16 2024
First Infection Second Infection Third Infection
TIME FROM INOCULATION (days)
FIGURE I. Plasmodium coatneyt parasitemia in rhesus monkey AF-B425 during the course of a primary and
two subsequent infections. Intravenous inoculations of blood-stage parasites were done on day 0. Each dot
represents parasitemia determined daily. The arrows indicate days or treatment (Rx) with chloroquine. PRBC
- parasiti-ed red blood cells.
a group ofP.falciparum proteins with M, values not detect any proteins ofP. coatney'iin the West.
of 100. 90, 80, and 70 kD (Figure 3A, lane h). etn blot (Figure 3B. lane 0. The HIS did not react
but it did not react with control antigens at the with control nntigens at the Mr values indicated
M, values indicated above (Figure 3A. lanes a above (Figure 3B, lanes e and h).
and d). Probing with HIS at a 1:100 dilution A dot immunoblot assay using the trimer ofa
detected several P.f alciparum proteins. includ- synthetic peptide (EENVEHDA) that corre-
ing the polypeptide with an M, of 155 kD (Figure sponded to the RESA of P.f alciparum showed
3B. lane g). In contrast to the RIS. the HIS did that HIS but not RIS reacted with the peptide.
RESA-LIKE ANTIGEN IN 1'. (V..TNEYI 131
TrARLE I
1111t11odt' tters to Plasmiodjumn coatnevi tit ,ih(P U t lilrnkei-s dh itit i. priimarya Ild ITpeated tit 'ctio,,. astdetermined
I,% ijnlyflitolhnoriscence assai-s*
First S,%ondl I hird
\1-mnke~sl~s IN IDav 16 D~a\ 0i Da\ 12
AF-13298t 05
AF-CP425t 0 (25)
AF-B464t 0(5)
AF-B42S NT 125 (625) 25 (625) 625 (3.125)
*Values are reciprocal titers at antibodies in an mrthroc~tc membrane immunulluorescence assav and an immunoifluorescenice assav (values in
t These monkeys were killed alter the tirst infection.
Neither HIS nor RIS reacted with spots of con- syphilitic patients treated with induced malaria.
trol protein. Thcse antisera to human malarias were tested
against a panel of' intra-erythrocytic antigens of
DISCUSSION simian malaria parasites. Occasional low-titer
reactivity was observed against the P. coatnevi
Plasmnoditon coatnevi infcction of~ macaque preparation. In this study. these immunologic
monkeys has attracted attention as an animal relationships were further explored. Our prelim-
model for thc study of falciparum malaria. The inary results showed that rhesus monkeys de-
similarities previously identified between P. velop antibody reactivity against the RESA-like
coatne~vi infections of rhesus monkeys and P. fal- antigen of P. coatnevi when GA-fixed ring-stage
ciparum infections of humans involve the patho- PRBC are used, and that some P. falciparum
physiology of the infection. Serologic cross-reac- proteins other than RESA are recognized by P.
tivity between PAfalciparun?a nd P. coatne' vi was coalnevyi-immune rhesus sera.
assessed by Collins and others' using sera from During the course of P. coatne~vi infections in
FlOuRs 2. Ring-infected erythrocyte surface antigen of Plasmodluns coatneyf-infected rhesus monkey red
blood cells shown by an erythrocyte membrane immunofluorescenee assay using immune serum from a P.
coatneyi-inlected rhesus monkey. Parasites were visualized by counterstaining with ethidium bromide (arrow-
heads).
I 3,2 ')OMSANpI1-rcII ANt) OTHERS
FAB\ILE 2
.\, p ij' l f" P(cid:1)~'r.'llitt /i(v, !tlI't'Elasmodi-mn coal.- a b c d e f g h
ne'yi and P. falciparum studied hIl jinininohlorlres-
I' *,',,, l' I' i,?, 'u~ruti ;,. 205-
,Iwm ldution 11A LSIIF IA I %M. _11
RIS 1:250
-7
Ills 1:250 -
11 Ni m unolluorescence as~i; iii an unhwcl. air-dried Lriiplo.
/oile'shiloni.stagc parasite preparation: EMIF - imnunolluorescence
assy of*a glularaldehvdeoixed ringl-stage parasite preparation: RIS - 6 #
rhesus immune serum from monke" AF-11425: HIS - human immune (cid:127)45
"filum
rhesus monkeys, the development of specific an-
tibodies to RESA-like antigen was observed. Al-
though the results were obtained from a small
number of infected monkeys and kinetic studies
have yet to be done, the data shown in Table I
suggest that this antibody response required
stimulation by repeated infections and was non-
persistent, which is consistent with the properties
of human antibodies to P..firlciparum RESA. ',
In view of our preliminary results, the time to ,
patency in the multiply infected rhesus monkey
appeared progressively prolonged. Furthermore, A B
the peak parasitemias in the second and third FtoURE 3. Cross-reactivity of antibodies to Plas-
infections were two-fold and six-fold lower. re- modium coatneyi with P. falciparum proteins studied
spectively, than those observed during the pri- by Western blot analysis. Protein extracts of uninfected
mary infection. This is probably a reflection of human red blood cells (lanes a and h). trophozoite.'
schizont (T/S)-stage P..falciparum-infectedh uman red
the increasing levels of malaria-specific antibod- blood cells (lanes b and g), T/S-stage P. coatneyi-in-
ies, including antibodies to RESA. as has been fected rhesus red blood cells (lanes c and f), and un-
shown by the other investigators in P. falciparuin infected rhesus red blood cells (lanes d and e) were
infections.' .1' Since antibodies to RESA are as- electrophoresed and probed with rhesus immune se-
sociated with falciparum malaria, but not other rum from monkey AF-B425 (A) and human immune
serum (B). Arrowheads indicate P. coatne 'i polypep-
human malarias, these observations add an im- tides with M, values of 120. 100, 97, 80 and 45 kD
munologic characteristic to the similarities be- recognized by monkey serum AF-B425 (lane c) and P.
tween P. coatneyi and P. falciparum. falciparum RESA. M, of 155 kDMre cognized by human
Plasmodiumn coalneyi-immune rhesus serum immune serum (lane g). The range of standard molec-
ular masses are indicated by the numbers between the
recognized native P. falciparumn antigens as as- two blots.
sayed by IFA of air-dried T/S-stage parasites. On
the other hand, immune rhesus serum-did not
recognize the RESA of P. falciparum nor did the the P. coatneyi RESA-like antigen is immuno-
immune human serum recognize the RESA-like genie in the rhesus monkey. Although the amino
antigen of P. coatnevi. This suggests that the an- acid sequence of the P. coalnevi RESA-like an-
tigenic portion of the P. coatneyi RESA-like an- tigen is not known and the existence of homology
tigen exposed on the ring-infected rhesus RBC with the P.falciparumR ESA has not been shown.
is different from that of the P.falciparumR ESA it is unlikely that the RESA-like antigen of P.
on infected human RBC. The potent immuno- coatneyi will contain sequences similar to the
genie B cell epitopes of the P..falciparum RESA repeat region nf the P. falciparum RESA. In ad-
have been shown to be localized predominantly dition. it was shown in the EMIF assay and by
in the repetitive amino acid sequences (EEN- dot immunoblotting that the immune rhesus se-
VEHDA and EENV) in the 3' portion of the rum did not recognize the RESA of P. falcipa-
molecule.. "Our preliminary results showed that rum.
RiESA.LIKE ANTrIGEN INI1'. COAiTNPI-1 133
We fuarther investigated the serologic cross- .1ialartasv. Washington: U. S,G overnment Print-
reactivity of antibodies to 11. cot'nael with deC- ing Oitlice. 289-299.
natured P.,h/2lciparum proteins by Western blot- 1. Rudrzeinsokat oMpA. oTiratgsenr Wd. g19m68.o cThttcs fnineP stlr-uc-
ting uinder reducing conditions. The immune modiumn coatflCvi. J Protozool I1"57:3 -88.
rhesus serum recognized several 1'. coazm't'l 4. Aikawa Mi. Brown AE. Smith CD. Tegoshi T.
polypep tides as well as a group of P. fWicipartim Howard R.I. Hlasler TH. Ito Y. Perry G. Collins
polypeptides and thus confirmed the eross-reac- WE. Webster HK. 1992. A primate model for
tivitv of antibodies to P. coatn evi w'ith P. ýcip human cerebral malaria: Plasmodiumn coatneyi-
infected rhesus monkevs. Am~ J Trop ,tied 11t'g
arimi antigens detected by IFA of air-dried T/S- 46: 391-397.
stage parasites. The polypeptide with an M, value 5. Udomsangpetch R, Brown AE. Smith CD. Web-
of 155 MDth e predominant antigen on the sur- %ter HK. 1991. Rosette formation by Plasmo-
face of P. ./'alciparum ring-stage PRBC (Pfl 55/' diumn coatneyi-infected red blood cells. AmnJ
RESA)," was not detected by the immune rhe- Trop tied Iltvg 44. 399-40 1.
ti. Perlmann H-.B er'tins K. Wahlgren MI.C arlsson C.
sus serum. This confirmed the negative result in 13jorkman A. Patarro o ME. Perlniann P. 1984.
EMIIF assay with P. tWeiparitm ring-stage PRBC Antibodies in malarial sera to parasite antigens
and indicated that these twvo proteins. A. c)WUCew in the membrane of ervthirocvts inf~ected with
RESA-like antigen and P. lWciparuni RESA. dif- early asexual stages of Plasmodium Ialciparum.
fer in their natural immunogenic epitopes. .1E xt7.vple1d 159. 1686-1704.
Coppel RL. Cowman AE. Anders AF. Bianco AE.
Immune responses to the RESA have been as-it B L inttbch KR. Kemp DJ. Brown GYV.
sociated with resistance to clinical falciparum 1984. Immune sera recognized on erythrocytes
malaria. 4 II''". " leading to the proposal of RESA a Plasmodiumn fialciparumn antigen co .mposed of
as a candidate vaccine. This report identifies a repeated amino acid sequences. Naur 310:
primmtde elianla ia hic an ntiodyre-789-792.
primteo dea lianiaw hic anantbod re 8.Kabilan L. Troye-Blomberg M, Patarroyo ME,
sponse to RESA occurs and may therefore be Bjorkman A. Perlmann P. 1987. Regulation of
used in the development of RESA-based vac- the immune response in Plasmodium, falcipa-
cines. rum malaria. IV. T cell-dependent production
of immunoglobulin and anti-Plasmodiumn fal-
Acknowledgments: The P. coatne 'vii solate used in this ciparumn antibodies in vitro. ('tin Exvp mmunol
study was generously provided by Dr. William E. Col- 68: 288-297.
lins (Division of Parasitic Diseases. Centers for Disease 9. Kabilan L. Troye-Blomberg M, Perlmann H. An-
Control and Prevention, Atlanta. GA). We thank C. dersson G. Hogh B. Petersen E. Bjorkman A.
Tessanaboon for skillful assistance. Perlmann P. 1988. T cell epitopes in PI`155/
Financial support: This work was part of the MalariaREAamjocndatfraPamoima-
Research Program of the Walter Reed Army Institute ciparumn malaria vaccine. Proc .Vad Acad Sci
of Research. Rachanee Udomsangpetch is supported USA 85: 5659-5663.
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University. Rama VI Road. Bangkok 10400, Thailand. in the repeat regions of the Plasmodiumn falcip-
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