Parasitology Today, vol. 8, no.
7, 1992
13 Foote, S.J. and Kemp, D.J. "(1989) Trends Genet. 5,337-342
14 Corcoran, L.M. et al. (1988) Cell 53, 807-813
15 Patarapotikul, S. and Langsley, G. (1988) Nucleic Acids Res. 16,
4331--4340
16 Foote, S.J. et al. (1989) Cell 57, 921-930
17 Wellems, T.E., Walker-Jonah, A. and Panton, L.J. (1991) Proc.
Natl Acad. Sci. USA 88, 3382-3386
18 Shirley, M.W. et al. (1990) Mol. Biochem. Parasitol. 40, 137-146
19 Kemp, D.J. et al. Proceedings of the I V International Congress on
Malaria and Babesiosis, Instituto Oswaldo Cruz (in press)
20 Limpaiboon, T. et al. (1991) Mol. Biochem. Parasitol. 47, 197-206
21 Kohara, Y., Akiyama, K. and Isono, K. (1987) Cell 50, 495-508
22 Olson, M.V. et al. (1986)Proc. NatIAcad. Sci. USA 83, 7826--7830
23 Coulson, A. et al. (1986) Proc. Natl Acad. Sci. USA 83, 7821-7825
24 Burke, D.T., Carle, G.F. and Olson, M.V. (1987) Science 236,
806-812
25 Kuspa, A. et al. (1989)Proc. Natl Acad. Sci. USA 86, 8917-8921
26 Guzman, P. and Ecker, J.R. (1988) Nucleic Acids Res. 16,
11091-11095
27 Rubock, M.J. et al. (1990) Proc. Natl Acad. Sci. USA 87,
4751-4755
28 Garza, D. et al. (1989) Science 246, 641-646
29 Coulson, A. et al. (1988) Nature 335, 184-186
Surface Antigen Variability and Variation
in Giardia larnblia
T. Nash
Recent studies show that Giardia isolates are hetero-
geneous but fall into at least three groups as determined
by a number of complementary techniques. Giardia
undergoes surface antigenic variation, both in vitro,
and in humans and other animal model infections.
Many of the characteristics of antigenic variation and
the proteins involved, called variant-specific surface
proteins (VSPs), are unique. The sequences of five
VSPs reveal a family of cysteine-rich proteins. Here
Theodore Nash reviews the relationship between anti-
genic variation and Giardia heterogeneity.
Giardia lamblia is one of the most common parasites
of humans. Ingested cysts excyst in the small
intestine and multiply as motile, flagellated, bi-
nucleated trophozoites. These encyst in the lower
intestine and colon and are then excreted in the
feces, sometimes in enormous quantities (approach-
ing 107 per gram). Molecular comparison of cul-
tured isolates 1 and clones has led to an understand-
ing of the genetic relationship among isolates and
the finding that Giardia trophozoites undergo sur-
face antigenic variation.
In this review I shall refer to major surface
proteins of Giardia that undergo antigenic variation
as VSPs. Most clones rapidly become heterologous
Theodore Nash is at the Laboratory of Parasitic Diseases, National
Institute of Allergy and Infectious Diseases, Building 4, Room 126,
Bethesda, MD 20892, USA.
229
30 Triglia, T. and Kemp, D.J. (1991) Mol. Biochem. Parasitol. 44,
207-212
31 Lanzer, M., De Bruin, D. and Ravetch, J.V. E M B O J . (in press)
32 Triglia, T. et al. (1991) Mol. Cell. Biol. 11, 5244-5250
33 Smith, D.R., Smyth, A.P. and Moir, D.T. (1990)Proc. NatlAcad.
Sci. USA 87, 8242-8246
34 Schlessinger, D. (1990) Trends Genet. 6, 248--258
35 Olson, M.V. et al. (1989) Science 245, 1434-1440
36 Green, E.D. et al. (1991) Genomics 11,548-564
37 Green, E.D. and Green, P. (1991) P C R Meth. Appl. 1, 77-90
38 Warburg, A. and Miller, L.H. (1992) Science 255,448-450
39 Kemp, D.J., Cowman, A.F. and Walliker, D. (1990) Adv.
Parasitol. 29, 75-149
40 Li, W-B. et al. (1989) Nucleic Acids Res. 17, 9621-9636
41 Holloway, S.P. et al. (1990) Mol. Biochem. Parasitol. 43,257-270
42 Hicks, K.E. et al. (1991) Gene 100, 123-129
43 Fox, B.A. and Bzik, D.J. (1991) Mol. Biochem. Parasitol. 49,
289-296
44 Li, W-B. et al. (1991) Mol. Biochem. Parasitol. 46, 229-240
45 Robson, K.J.H. and Jennings, M.W. (1991) Mol. Biochem.
Parasitol. 46, 19-34
46 Nolte, D. et al. (1991) Mol. Biochem. Parasitol. 49, 253-264
47 Wilson, R.J.M. et al. (1991) Parasitology Today 7, 134-136
with regard to VSPs but, if not kept in culture over
prolonged periods, consist of a predominant vari-
ant. GS/H7 refers to the H7 clone of isolate GS. The
predominant VSP (VSPH7) is a 58kDa surface
antigen. VSPs have also been called cysteine-rich
proteins (CRPs) 2, but this nomenclature is mis-
leading because there are most likely a number of
cysteine-rich molecules in Giardia with different
functions; I prefer the term VSP. The gene
encoding a particular VSP is designated by italics,
as VSPH7.
Analysis of 29 Giardia lamblia isolates and clones
from some isolates reveals marked surface antigen
variability3-5. Heterogeneity occurs primarily for
two reasons. First, there are heritable differences
between isolates3-5, and second, isolates undergo
antigenic variation that results in heterogeneity
even in clonaUy derived populations2'6'7. Although
the underlying mechanisms of antigenic variation in
Giardia are not known, it should be remembered
that the nature of the proteins involved, the location
of the parasite in the intestine and the biology of the
parasite are very different compared with the
African trypanosomes and other micro-organisms.
Heterogeneity among isolates
A number of characteristics have been used to
differentiate isolates. These include differences in
isoenzymes8-1°, restriction fragment length poly-
morphisms (RFLPs) 3, phenotypic surface antigen
expression4-6, susceptibility to chemotherapeutic
230 Parasitology Today, vol. 8, no. 7, 1992

agents 11, 18S ribosomal D N A (rDNA) sequences 12,
chromosome size and staining characteristics 13,
differences in homologous genes 14, the presence or
absence of certain genes or transcripts (T.E. Nash
and M.R. Mowatt, and A. Aggarwal and T.E.
Nash, unpublished), susceptibility to Giardia RNA
virus ~5 and the ability to grow, in vitro 16.
Over the past decade my colleagues and I have
systematically compared 29 Giardia isolates; a
summary of the findings are shown in Table 1. Our
original grouping is based on RFLPs using un-
defined probes 3 and the patterns of surface re-
activity using a panel of isolate-specific antisera 3'4.
As we learned more about antigenic variation and

Table I. Comparison of 29 Giardia isolatesa

Isolate Group mAb6E7 b mAb5C| b mAb3F6 b mAbGI0 b VSPA6 c VSPH7 c VSPI267 ~ C4 d Giardin e RNA f

WB + + + O E+ O WB O 23 I
LT + + + O S+ ND WB O 23 ND

Isr + + + O S+ O WB O 23 I

Be2 + + + O S+ O WB O 23 I

CAT + + + O S+ O WB O 23 I

RS + + + O E+ ND ND O 23 ND

P-I II + + + O S+ O WB O 23 ND

G2M ND + + + O S+ ND WB O 23 I

Dan II + + + O E+ ND WB O 23 ND

GP II + + + O S+ ND WB O 23 I
G3M ND + + + O S+ ND WB O 23 I
E2 ND + O O O P+ ND P O 23 II

N II O O + O P+ ND P O 23 II

Ac - O O + O P+ ND P O 23 ND

DH II O O + O P+ ND P O 23 II

JH II O O + O P+ O ND O 23 II
Sug ND O O + O O ND ? O ND ND
AB II O O + O P+ ND P O ND II
Nat ND ND ND ND ND P+ ND ND O ND ND
Andi ND O O O O P+ ND P O 23 ND

Mar ND O O O O P+ ND P O 23 ND

Erin ND O O + O P+ ND P O ? ND

GIM ND O O O O O ND O + 4 III

PM ND O O O O O ND O + 4 ND
Bel III O O O O O ND O + ND III

E9 ND O O O O O O O + 4 III

E4 ND O O O O O ND O + 4 II
GS III O O O + O + O + 4 III

CM III O O O + O + O + 4 III
aThe features studied are shown across the top of the table and the various isolates are designated in the far left column. +, presence of a particular feature;
O, absence of the characteristic; ND, not done; ?, uninterpretable results; WB, identical to the WB isolate; E, banding pattern identical to WB; S, similar
banding pattern, containing some bands in common with WB; P, banding pattern dissimilar to WB.
b Presence of any trophozoites expressing surface epitopes recognized by the indicated mAbs. mAbs 6E7 and 5C I were made to isolate WB and mAb G 10/4
to GS. mAb 3F6 was most likely produced to a WB-like organism, which contaminated isolate GS(18).
c Hybridization to internal fragments of VSPA6 and VSP 1267 or a VSPH7-specific oligonucleotide.
tiC4 is a gene that was detected by differential screening of an expression library with two isolate-specific antisera. Under high stringency it recognizes
fragments in Group 3 isolates only (T. Nash and M.R. Mowatt, unpublished).
e Giardin refers to the size of the fragment in kilobases (kb) detected in Southern blots after digestion of genomic DNA with EcoRI and probed with l~-I
giardin.
fGrouping based on the sequence of a 3' fragment of the 18S rDNA 12. Isolates in each group have identical sequences.
Parasitology Today, vol. 8, no. 7, 1992
developed other reagents and techniques, other
methods of comparison were used. These include
the reactivity of trophozoites in populations to
mAbs that recognize specific epitopes in V S P s 6, the
ability of probes encoding fragments of VSP pro-
teins to detect the presence of hybridizing bands in
Southern blots6, the use of conserved and non-
conserved genes as probes in Southern blots (T.E.
Nash and M.R. Mowatt, unpublished) and the
presence of rDNA sequence differences12. Despite
the use of a number of diverse methodologies, the
isolates roughly fall into three groups17: Group 1
resembles isolate WB, Group 2 shares some of the
characteristics of Group 1 but clearly differs, and
Group 3 consists of heterogeneous isolates that have
few features in common with Group 1. There is a
clear relationship between the ability to express
certain epitopes and the other classifying charac-
teristics, indicating that the type and nature of the
VSP repertoires are at least partly dependent on the
Group. For instance, Group 1 isolates express VSPs
recognized by monoclonal antibodies (mAbs) 6E7,
5C1 and 3F6, but Group 2 isolates are unable to
express epitopes recognized by mAbs 6E7 and 5C1
but express mAb 3F6-binding epitopes. In contrast,
Group 3 isolates express none of the epitopes
expressed in Groups 1 or 2 and a subset expresses
epitopes recognized by mAb G10/4. RFLP analysis
using [5-1 giardin and the presence of C4-hybridizing
fragments (see legend for Table 1) clearly separates
Group 3 from the other groups (T.E. Nash and M.R.
Mowatt, unpublished). The genetic differences
between the groups i,; additionally strengthened by
group-specific rDNA sequence changes 12. Sequence
divergence of rDNA has not been recognized within
a species and, together with the other characteristic
differences among the groups, leads to the con-
clusion that these groups branched off from a
common ancestor millions of years ago. Further
support for this conclusion comes from comparison
of the conserved ADP-ribosylation factor 14 and
triosphosphate isomerase genes (M.R. Mowatt,
T.C. Howard and T.E. Nash, unpublished) in the
prototype Group 1 isolate, WB, compared to the
Group 3 isolate, GS. ]Both sequences show a level of
nucleotide difference unusual for members of the
same species.
Antigenic variation, in vitro
Antigenic variation has been shown convincingly
both by the spontaneous appearance of variants in
clones6 and by depletion experiments of clones
using cytotoxic mAbs and the analysis of surviving
trophozoites7'18. Confirmation of these findings
includes an exact correlation of the presence, loss
and subsequent appearance of specific transcripts in
clones with the phenotypic expression of corre-
sponding V S P s 19-2! and sequence analysis of these
transcripts and their corresponding genes 19-21.
Most likely, antigenic variation is a common, if not
universal, phenomenon in Giardia lamblia and has
231
been formally demonstrated in all five isolates
where reagents were available to allow verifi-
cation7,18.
The rates of antigenic variation differ and appear
to be mostly isolate and, to a lesser extent, epitope
dependent22. Clones of isolates will predictably re-
express epitopes characteristic of their parent 6'z2.
We took advantage of this to measure the rate of
appearance of non-expressed epitopes using direct
immunofluorescence. The rates of appearance of
VSPs vary dramatically in two different isolates. In
WB, three specific VSPs appear about once in every
12-13 generations (depending on the VSP), com-
pared to once in every 6.5 generations in GS for the
one VSP we are able to detect. These rapid rates of
change predicted the development of considerable
heterogeneity in continuously maintained cultures,
a finding that has been repeatedly demonstrated,
particularly with GS cultures (T.E. Nash, unpub-
lished). Rates of appearance also differed signifi-
cantly among epitopes expressed by the same isolate.
Although differences in the rates of change were
relatively small (about 0.7 generations), this sug-
gested a bias toward expression of certain epitopes.
Differing rates of appearance of epitopes, in viva,
could have profound biological consequences, such
as a mechanism which could result in prolonged
infections. Also, in these experiments we were
unable to detect two VSPs expressed on the same
trophozoite; only one VSP appears to be expressed at
any given time.
Antigenic variation, in viva
Antigenic variation was initially described, in
vitro, and it was therefore essential to show rel-
evance, in viva. Unlike many of the organisms that
undergo antigenic variation, there was little or no
suggestion from the clinical course in human or
animal infections of the phenomenon. Further-
more, antigenic variation had not been described in
organisms inhabiting the gastrointestinal tract. In
experimental infections of humans 23, gerbils24 and
neonatal mice25 with clones of Giardia lamblia, the
initial VSP is lost and replaced by a heterologous
mixture of VSPs in the two models analyzed
(humans and gerbils). Replacement occurs between
14 and 22 days in mice25 and humans 23 and by the
seventh day in gerbils24. Since antibodies to the
initial VSP are detected by day 14, humoral
antibodies could play a role in the selection of VSPs
in mice and humans but not in gerbils because
change occurs before antibodies are detected. A role
for antibody response is further suggested by the
continued ability of Giardia to undergo antigenic
variation in nude mice and the failure of parasites in
scid mice to do SO26. Trophozoites in the latter
continue to express the initial VSP throughout the
course of infection.
In contrast to the African trypanosomes, cyclical
changes in Giardia VSP expression have not been
found in infections in gerbils24 or humans23; instead
232
there is an expansion of the number of VSPs
expressed. Antigenic variation could cause chronic,
prolonged or normally undetected infections. In the
experimental human infections, the volunteers were
treated at 3weeks, and further analysis was not
possible but in gerbils trophozoites are no longer
seen during microscopic examination in the in-
testines by day 28 (Ref. 24). However, others have
documented the presence of low numbers of organ-
isms after apparent 'self-cure' in gerbils by detect-
ing trophozoites following immunosuppression 27.
The nature of VSPs
The VSPs of Giardia are a unique family of
proteins as judged by the deduced amino acid
sequences from both genomic and complementary
DNA (cDNA) clones 2'19-21'28. They vary in size
from 30 kDa to 200 kDa or more and, for the most
part, are antigenically distinct and cover the surface
of the parasite 5'6'29. Electron micrographs using
specific labeled mAbs showed that VSPs formed a
surface coat that covered the entire trophozoite
including the flagellum (Fig. 1 and Ref. 30) and that
specific mAbs bound to this coat. Despite their
surface location, carbohydrates have not been de-
tected in VSPs. To date, the sequences of five VSP
genes have been determined, and parts of a number
of unpublished sequences appear to resemble the
more fully characterized sequences. An alignment
of three VSP amino acid sequences is shown in
Fig. 2. A striking feature is the high cysteine content
(11-12%) which commonly occurs as Cys-X-X-Cys
motifs. The carboxyl terminus is well conserved
among the VSPs thus far studied and is hydro-
phobic except for the last five amino acids. The
amino terminus is also hydrophobic. Small
stretches of identical amino acids appear in all three
VSPs, and slightly larger areas of homology occur
between sequences of any pair of the three.
A repeating sequence is a feature of a subset of
VSPs 2,6,20 . mAb 6E7 reacts with the surface of the
WB A6. Analysis has shown that the 170 kDa VSP
of WB A6 (or CRP 170) contains a 195 nucleotide
fragment that is repeated about 20 times and
encodes the epitope recognized by the mAb 6E7
(M.R. Mowatt et al., unpublished and Refs 2, 20).
The VSPs of other isolates are also recognized by
this mAb but vary in size, and in at least one
instance the difference in size can be attributed to a
difference in the number of repetitive units (M.R.
Mowatt et al., unpublished). Only one repeating
sequence has been described and it is not clear how
common this feature is or if other repetitive
sequences exist.
Motifs that occur in lin-ll, Isl-1 and mec-3
proteins 31 are known as LIM motifs; one such
m o t i f 32-34 is present in four of five published
(absent in CRP72) and two unpublished VSP
sequences. The LIM motif binds heavy metals,
particularly zinc, and this motif is also found in a
cysteine-rich mammalian intestinal protein that is a
Parasitology Today, vol. 8, no. 7, 1992
Fig. I. Irnrnunogold labeling of a trophozoite using a VSP-specific
rnAb. An 18nrn cell coat covers the entire parasite and this is
specifically labeled (closed arrows). The plasma membrane is
shown by the open arrowheads. Scale bar = 125 nrn. (Reproduced,
with permission, from Ref. 30.)
putative intracellular intestinal zinc transporter 34.
The spacing of the cysteines is well conserved
around the histidine of the LIM-like motif and can
be summarized as CX2_3CX9_14CX2CHisX2CX2-
CX7_8CX2CX15_16CX2_5 where X is any amino
acid. This compares with the LIM motif
C X 2 CXI7_19HisX2CXzCX7_ 11(C)X8C. Other putative
zinc-binding motifs are found sporadically in some
VSPs. If VSPs are able to bind relatively large
amounts of zinc, they could inhibit a number of
important intestinal enzymes, a notion that is
supported by the intestinal symptoms of clinical
giardiasis resembling those of zinc deficiency.
The control of antigenic variation
Although the molecular mechanisms controlling
antigenic variation have not been defined, tran-
scriptional regulatory mechanism(s) appear import-
ant to the control of VSP expression. Stable
transcripts of expressed VSPs are present while
stable transcripts of non-expressed VSPs are absent
in well-characterized clones 2'19-21. Comparison of
VSP sequences of parent and progeny do not reveal
large fragments of parent sequences in progeny
VSP, as one would expect if gene conversion
mechanisms were operative. In African trypano-
somes, expressed variant antigen genes commonly
undergo duplication and are expressed in telomere-
associated sites. V S P genes in Giardia do not
appear to be telomere associated 2 and, with one
possible exception 2°, no expression-linked copies
have been described. It may be that entire genes are
switched on or off by an as yet undefined process.
In one VSP where both genomic and cDNA genes
had been sequenced 19, they were found to be
identical, and in the genome there were two
identical copies arranged tail-to-tail. Where specific
VSP probes have been used, different-sized bands
are seen in Southern blots for two different VSPs,
suggesting two V S P gene types for each V S P 19'21 .
However, in another study four copies of each V S P
 -m
{3
~<< ~

M m ~ ~

o~

• C) C] (D LO ~q m ~U ~ ~D
t~J 5q ® 6] &'% [~
9o
m t e Z ~ Z ~ 6%
*636369 z 5u
"-4
"3 C] 6] 6] (~
<
6] <
< H ~ C] 09 -- C~ < [E
w3 6]
z O • [m 6D 6D
[D a~
c? C] • 63 LO ~
Z • 63 O~ 63
D~ O9 z ~'q
og ~q n~ 63 z
%~.~ Z U] .In ~ Z .~ u'~ 6] ~6] &] u'] . D9 t9 ~-] .<]to[.,] U~ O9 69
-3 ('3 ~q
• Z zq • ~ ~ ~
.~q ~ U9
('3 c~
w3 ul

z • > ~ 6")
~'~ . ~-] G] ~-~ . L9 V] [-a .~=~ Z ~ ~< ~ t~ , [~ &D u-] ,~U'D u'] ~ ~J ~ MO 6]
Z o~ ~ ~
(3 • ~ 63 63 C) (D • <P Z 09
U] , to -- G) C) C)
z • v-] 6"] 69
('3 ~ ~ O Z ru
<2 63 6] {'3 D [~ ~ 0"3
('3 Z 63 • ~ Z ~ D9 L,~
O ~-~ ~< . ( D O 0
" O
O D [~ fD :x
O C] 63 <
(D > 6] ~-u~ &D 62, ~C) (D C)
~z O 6% .(D O (D
. [.o ~-3 09 • to ~-3 [I]
(D 69 6]
Z ~< z z z ,u ~D z
< m < ~U ~ .6] 09 Z
63 • ,--3 59 D9
69 -- [~ 69
o 6D In &D ('3 63 to ~ ~
~ ~ 6D < J] q]
• Z {~ L9 • LO Z
.63 LO ~ z ¢3 ('3
• [:] Z CO .(D C) (D c] 69 63
[3 < >
.(DO) C) oZ Z 6"~
~ eO9 ~ ~ ('3
.(D C] (D e~ 09 ~ .09 ~ ~

01 01 to
b4
234
gene per haploid genome were estimated, suggest-
ing two copies of each type of VSP gene per haploid
genome 21.
Repertoire size was estimated in two ways and
both give roughly equivalent estimates. By deter-
mining the rate of appearance of newly expressed
VSPs in clones 22, the median number of VSPs was
estimated to be between 20.5 and 184.5. Another
method compared the amount of all VSP D N A and
one specific VSP D N A , to yield a repertoire of 130-
150 VSP genes (Fig. 3 and Ref. 21). As mentioned
above, additional calculations indicate that there are
four copies of each VSP gene per haploid genome.
Therefore, the VSP repertoire size is approximately
30-40 (Ref. 21), which is considerably less than the
estimated repertoire size for the African trypano-
somes.
There are few studies of the state of VSPs on the
surface of the parasite. The earliest study of the
170 kDa VSP of WB shows that it is released from
the surface into the culture medium35; neither the
mechanism of release nor the means of attachment
to the membrane is known. In contrast to the
African trypanosomes, at least two studies have
shown that VSPs are not linked by glycosylphos-
phatidylinositol anchors 36 (T.E. Nash, unpub-
lished), neither have free sulfhydral groups been
found on the VSPs of live trophozoites (T.E. Nash,
unpublished).
The biological significance of antigenic variation
in Gmrdia is speculative although escape from the
host's immunological response is commonly men-
tioned. However, although the chronicity of some
infections could be due to antigenic variation, there
are no data directly supporting this idea. Antigenic
variation could be a process that leads to increased
kb
23.1 - -
9.9--
66--
4.4 - -
2.3 --
Fig. 3. Southern blot of H7
2.0-- DNA restricted with Hindlll
and hybridized to MM-16,
an oligonucleotide probe
specific to the conserved 3'
region, or to MM- 17, an
oligonucleotide probe specific
to VSPH7. MM-16 hybridized
to many bands while MM- I 7
hybridized to only two.
(Reproduced, with permis-
MM-I6 MM-17
sion, from Ref. 21.)
Parasitology Today, vol. 8, no. 7, 1992
diversity in a population and thus allows other
selection processes to come into play. Certain
epitopes may or may not be recognized by the host,
and some VSPs may be able to survive better or
contribute to virulence, or permit broadening of the
host range. Implicit in this argument is the fact that
different VSPs have different biological properties.
Indeed, it is known that VSPs differ in their
susceptibility to intestinal proteases, in vitro 37.
Whether immunological/nonimmunological selection
processes are important remains a vexing question.
Although some questions have been answered, it
is obvious that much needs to be learnt to under-
stand not only antigenic variation, but also the
biology of the parasite and host-parasite inter-
actions as well. Giardia is a very successful parasite
and unraveling its survival mechanism(s) will likely
lead not only to control of infection and disease but
also to a better understanding of its cellular biology
and evolution.
References
1 Meyer, E.A. and Pope, B.L. (1965) Nature 207, 1417-1418
2 Adam, R.D. et al. (1988)J. Exp. Med. 167, 109-118
3 Nash, T.E. et al. (1985)J. Infect. Dis. 152, 64-73
4 Nash, T.E. et al. (1985)J. Infect. Dis. 152, 1166-1171
5 Ungar, B.L.P. and Nash, T.E. (1987) Am. J. Trop. Med. Hyg. 37,
283-289
6 Nash, T.E., Conrad, J.T. and Merritt, J.W., Jr (1990) Mol.
Biochem. Parasitol. 42, 125-132
7 Nash, T.E. et al. (1988)J. Immunol. 141,636-641
8 Bertram, S.H. et al. (1983)J. Parasitol. 69, 793-801
9 Meloni, B.P. et al. (1991) Acta Trop. 50, 114-124
10 Korman, S.H. et al. (1986) Z. Parasitenkd. 72, 173-180
11 McIntyre, P. et al. (1986)J. Pediatr. 108, 1005-1010
12 Weiss, J.B., VanKeulen, H. and Nash, T.E. Mol. Biochem.
Parasitol. (in press)
13 Adam, R.D., Nash, T.E. and Wellems, T.E. (1988) Nucleic Acids
Res. 16, 4555--4567
14 Murtagh, J.J., Jr et al. J . Biol. Chem. (in press)
15 Miller, R.L., Wang, A.L. and Wang, C.C. (1988) Exp. Parasitol.
66, 118-123
16 Woo, P.T.K. and Paterson, W.B. (1986) Trans. R. Soc Trop. Med.
Hyg. 80, 56-59
17 Nash, T. (1985) Microecol. Ther. 15, 121-132
18 Aggarwal, A., Merritt, J.W., Jr and Nash, T.E. (1989) Mol.
Biochem. Parasitol. 32, 39-48
19 Mowatt, M.R., Aggarwal, A. and Nash, T.E. (1991)Mol. Biochem.
Parasitol. 49, 215-228
20 Adam, R.D., Yang, Y. and Nash, T.E. (1992) Mol. Cell. Biol. 12,
1194-1201
21 Nash, T.E. and Mowatt, M.R. (1992)Mol. Biochem. Parasitol. 51,
21%228
22 Nash, T.E. et al. (1990) Exp. Parasitol. 71,415-421
23 Nash, T.E. et al. (1990)J. Immunol. 144, 4362-4369
24 Aggarwal, A. and Nash, T.E. (1988) Infect. Immun. 45, 700-707
25 Gottstein, B. et al. (1990) Parasite Immunol. 12, 659-673
26 Gottstein, B. and Nash, T.E. (1991)Parasite Immunol. 13,649-659
27 Lewis, P.D., Jr et al. (1987) Am. J . Trop. Med. Hyg. 36, 33-40
28 Gillin, F.D. et al. (1990) Proc. Natl Acad. Sci. USA 87, 4463-4467
29 Nash, T.E. and Aggarwal, A. (1986)J. Immunol. 136, 2628-2632
30 Pimenta, F.P., DaSilva, P.P. and Nash, T. (1991) Infect. Immun.
59, 3989-3996
31 Li, P.M. et al. (1991) Proc Natl Acad. Sci. USA 88, 9210-9213
32 Freyd, G., Kim, S.K. and Horvitz, R.H. (1990) Nature 344,
876-879
33 Karlsson, S.T. et al. (1990) Nature 344, 87%882
34 Hempe, J.M. and Cousins, R.J. (1991) Proc. Natl Acad. Sci. USA
88, 9671-9674
35 Nash, T.E., Gillin, F.D. and Smith, P.D. (1983)J. Immunol. 131,
2004-2010
36 Das, S. et al. (1991)J. Biol. Chem. 266, 21318-21325
37 Nash, T.E., Merritt, J.W., Jr and Conrad, J.T. (1991) Infect.
lmmun. 59, 1334-1340