Download Endemic Infection with Human T Cell Leukemia/Lymphoma Virus

944
Endemic Infection with Human T Cell Leukemia/Lymphoma Virus Type lIB in
Argentinean and Paraguayan Indians: Epidemiology and Molecular
Characterization
Jorge F. Ferrer, Eduardo Esteban, Syamalima Dube,
Miquel A. Basombrio, Amalia Segovia,
Monica Peralta-Ramos, Dipak K. Dube, Keith Sayre,
Nicolas Aguayo, James Hengst, and Bernard J. Poiesz
Comparative Leukemia and Retrovirus Unit, New Bolton Center,
University of Pennsylvania, Kennett Square, Pennsylvania; Area de
Virologia, Facultad de Ciencias Veterinarias, Universidad Nacional del
Centro de la Provincia de Buenos Aires, Tandil, and Laboratorio de
Patologia Experimental, Facultad de Ciencias de la Salud, Universidad
Nacional de Salta, Salta, Argentina; Departments of Medicine and
Microbiology, State University of New York Health Science Center,
Syracuse, and Cellular Products Inc., Buffalo, New York; AIDS/
Hepatitis Unit, Ortho Diagnostic Systems, Inc., Raritan, New Jersey;
Programa Nacional de SIDA, Asuncion, Paraguay
Human T cell leukemia/lymphoma virus (HTLV-II) type II infection was detected by polymerase
chain reaction or serologic analyses (or both) in 22% of 697 Indians of six different ethnic backgrounds inhabiting the Argentinean and Paraguayan Gran Chaco. None was infected with HTLVI. The prevalence of HTLV-II increased with age (14% in those <13 years and 23% in those ~13
years). HTLV-II infection was found in all 20 Gran Chaco communities studied, but marked
differences (44%-4%) in the rate of infection were observed even in communities separated by only
a few miles. These variations correlated closely with ethnicity. In the high-incidence communities,
infection clustered within families, with evidence for both sexual and perinatal transmission, primarily via breast-feeding. By contrast, only 2% of 94 Mapuche Indians from southern Argentina were
positive for HTLV-II. Analyses of pol and long terminal repeat sequences from 15 Gran Chaco
HTLV-II strains indicated that they constitute a highly conserved branch of the HTLV-lIB substrain.
The primate T cell leukemia/lymphoma viruses (PTLVs),
which include human T cell leukemia/lymphoma virus (HTLV)
types I and II and simian T cell leukemia/lymphoma virus type
I (STLV-I), plus bovine leukemia virus (BLV), make up a
unique genus of oncogenic retroviruses (reviewed in [1]). The
PTtVsand BLV encode for regulatory proteins that transactivate both viral and cellular gene transcription, a phenomenon
believed to be involved in disease pathogenesis [1].
HTLV-I is regarded as the etiologic agent of adult T cell
leukemia/lymphoma, HTLV-I-associated myelopathy/tropical
spastic paraparesis, polymyositis, uveitis, and arthritis (reviewed in [1, 2]). The etiologic roles of STLV-I [3] and BLV
[4] in neoplastic diseases of their respective species of origin
have been well documented. Although HTLV-II is yet to be
definitively associated with any disorder, suggestive evidence
for its etiologic involvement in certain lymphomas/leukemias
[5-8] and neurodegenerative diseases [9-11] has been re-
Received 3 March 1996; revised 27 June 1996.
Verbal informed consent was obtained from individuals in accordance to
US government guidelines in place at the time of this field work.
Grant support: Fundacion Hermanos Agustin y Enrique Rocca and Fundacion Antorchas, Buenos Aires; Secretaria de Ciencia y Tecnica de la Universidad Nacional de Centro de la Provincia de Buenos Aires, Tandil, Argentina;
NIH (HB-67021).
Reprints or correspondence: Dr. Jorge F. Ferrer, New Bolton Center, 382
W. Street Rd., University of Pennsylvania, Kennett Square, PA 19348.
The Journal of Infectious Diseases 1996; 174:944-53
© 1996 by The University of Chicago. All rights reserved.
0022-1899/96/7405-0007$01.00
ported. This evidence and the results of seroepidemiologic surveys showing that infection with HTLV-II is on the rise, particularly among intravenous drug users (IVDUs) of the United
State and Europe [12-18], emphasize the importance of thoroughly studying its natural history and pathogenicity. The
availability of human populations with a high incidence of
infection is essential for these studies, because if HTLV-II
causes disease, it probably does so in a small percentage of
infected hosts and after a long latency period.
The two major subtypes of HTLV-II, designated A and B,
differ genetically by ,....,7% [19-21]. While both subtypes have
been demonstrated in the IVDU population ofthe United States,
subtype A is the most prevalent [1,2]. We have reported serologic and molecular data showing HTL V-II infection in '" 13%
of adult Toba and Wichi Indians living in isolated communities
of northem Argentina [22]. Sequence analyses of DNA isolated
from peripheral blood leukocyte (PBL) samples demonstrated
that the virus present in these Indians belongs to the HTLVIIB subtype [20, 22], which is the predominant subtype found
among other Paleo-Amerindian populations [20, 22-25].
The reliability of epidemiologic data requires the use of
detection assays of proven sensitivity and specificity in the
particular populations studied. Current commercially available
ELISAs or Western blot (WB) assays for anti-PTLV antibodies
are based on HTLV-I antigens or antigens of the HTLV-IIA
substrain, or both. However, the sensitivities and specificities
of these ELISAs and WB assays for the detection of the HTLVlIB subtype have not been adequately evaluated. Polymerase
chain reaction (Pf.R) assays for HTLV-I1II pol DNA have
Jll) 1996; 174 (November)
HTL V-II in South American Indians
been shown to be the most sensitive and specific procedures
available for the diagnosis of PTL V infections [I, 12, 26 - 29].
Thus, as a first step for studying the epidemiology of HTL VII in Gran Chaco Indian populations, we compared the results
of PCR analyses with those of several serologic assays among
246 Indians. These comparisons enabled us to establish more
definitive criteria for identification of Indians infected with
HTL V-IIB [29]. Applying these criteria, we have examined
the distribution ofHTLV-II infection in Indians residing in the
Argentinean and Paraguayan sectors of the Gran Chaco as well
as in southern Argentina. In addition, HTLV-II strains from 15
Indians were subtyped by sequencing and oligomer restriction
analyses and compared phylogenetically to other PTLV strains.
Materials and Methods
Study populations. The Indian populations studied inhabit two
areas of South America that are separated by ~ 1400 km (figure
1). The Gran Chaco is a partially forested tropical flat land occupying > 1 million km2 in northeastern Argentina, central and southern Paraguay, and southeastern Bolivia. It is believed that this
plain began to be populated 6000 years B.P. by the Proto-MacroGuaycuruan Indians, who underwent divisions and subdivisions
into several linguistic families, including the Mataco-Mataguayan,
Mascoian, and Guaycuru proper. Other ethnic groups, such as the
Areas in South America in which Indian communities
studied are located.
Figure 1.
945
Zamucoan, Chiriguanos, and Arawak, also migrated to the Gran
Chaco from the north [30, 31].
The Indians studied in the Argentinean sector of the Gran Chaco
belong to ethnic groups called Chorote, Wichi, and Chulupi, which
are members of the Mataco-Mataguayan linguistic family, and
Toba, Lengua, and Ayoreo, which are members ofthe Guaycuruan,
Mascoian, and Zamucoan linguistic families, respectively. They
live in relatively small and isolated communities located in the
northeastern Argentinean province of Salta, along the Pilcomayo
river, in the areas called Santa Victoria Este and Los Blancos. The
other Indians studied live in communities adjacent to the farming
Mennonite colonies of Femheim and Salve Yanga in central Paraguay. Many of these Indians are employed by the Mennonites. By
and large the subjects studied still obtain their food by hunting,
fishing, and gathering, but many of those working for the Mennonites have incorporated farm products into their diet. None of these
Indians were known IVDUs, nor have they received blood transfusions.
Intermarriages between tribes belonging to the same linguistic
families, particularly between Chorote and Chulupi, have occurred.
Admixtures have been less frequent between tribes of different
linguistic families and are very rare between Indians and nonIndian people. The large majority of the non-Indian people living in
the Gran Chaco are of Spanish (Argentina) or German (Paraguay)
descent. There have been no black populations in these areas.
The Alumine is a mountainous region adjacent to the Andes in
the eastern part of Neuken, a province of southwestern Argentina.
The subjects studied in this region are the Mapuche. These Indians,
who migrated from southern Chile in the 17th century, raise sheep
and cattle and practice some agriculture [31]. Admixtures between
the Mapuche and non-Indian people occur with considerable frequency.
Collection and processing of blood specimens. Specimens
were collected during several trips to the Indian communities. The
selection of the subjects was based primarily on their accessibility
and cooperation. The name, sex, age, ethnicity, community of
residence, and family relationships of each Indian sampled were
recorded.
Heparinized and clotted blood samples were centrifuged on site
at 1500 g for 20 min. Plasma and sera were stored at 4°C. PBL
were obtained as described [22] and stored in liquid nitrogen.
Because of space limitations, we were unable to store PBL from
all of the Indians sampled.
Serology. Sera or plasma from the 791 Indians included in
this study (70 < 13 years' and 721 ~ 13 years) were tested for
HTLV-I and -II antibodies using either the Cambridge Biotech
(rp21 enhanced) ELISA (distributed by Ortho Diagnostic Systems,
Raritan, NJ) or the Vironostika (Organon Teknika, Durham, NC)
ELISA [32] or both. These two ELISAs use an HTLV-I lysate as
the antigen; the Cambridge Biotech ELISA also includes recombinant HTLV-I p21E in the antigen preparation. All of these samples
were also tested with the differential Select HTLV ELISA kit
(Biochem ImmunoSystems, Montreal) [33], which uses synthetic
type-specific HTLV-I and HTLV-II antigens in separate wells.
Samples from 30 I of the Indians, including many with negative
ELISA results, were also tested in the HTLV blot 2.3 WB assay
(Diagnostic Biotechnology; distributed by Cellular Products, Buffalo, NY) [34], in which the antigen preparation includes, in addition to whole HTLV-I viral lysate, both type-specific (rgp46
946
Ferrer et al.
HTLV-I env and rgp46 HTLV-II env) and shared (HTLV-I recombinant p21E) recombinant proteins.
All of the above ELISAs were done following the manufacturer's instructions. Samples that were initially positive were tested
in duplicate and those reacting in at least one of the duplicates
were considered to be positive. Samples that in the WB assays
reacted with HTLV p24, rgp21E, and rgp46-I were considered to
be HTLV-I - positive and those that reacted with HTLV p24,
rgp21E, and rgp46-II were considered to be HTLV-II- positive.
All other reactivities were considered indeterminate or negative.
peR and sequence analyses. DNA was organically extracted
from PBL as described [20]. All samples were first determined to
contain amplifiable DNA by PCR with the human ,8-g1obin primers
PC03 and PC04 [35]. One microgram of each DNA sample was
amplified with the HTLV-I/II pol primers SKIIO/SKlll for 45
cycles of PCR. Specific HTLV-I or HTLV-II sequences were detected using a commercial assay (Cellular Products) [36]. Three
HTLV-II-positive Indian samples (W175, Ch61O, and W43) were
also amplified with the HTLV-II-specific long terminal repeat
(L TR) primers HTIIL(26-4 7) + /HTIIL(351-329) - and
HTIIL(257-278)+/HTIIL(624-606)-, and the amplified products were detected by Southern blot hybridization using oligonucleotides HTIIL(257-278)+d and HTIIL9(351-329)-d, respectively [20].
In order to prevent false-positive reactions, all PCR assays were
done with dUTP and were subjected to uracil DNA glycosylase
sterilization [37]. Pre- and post-PCR operations were done by
separate personnel in separate facilities. Further, all primers had
additional 5' nonviral, nonhuman sequences added to facilitate
sequencing, and all positive retroviral samples were reamplified
with "signature primers" comprising these same nonviral, nonhuman sequences [38]. Failure to amplify with the signature primers
indicates the absence of contaminating amplified DNA.
Amplified pol and LTR DNAs were cloned and sequenced as
described [20], and HTLV-II pol sequences were subtyped as either
A or B by oligomer restriction analysis using the enzymes HinfI
or MseI as described [39]. Sequences were aligned using a commercial software package [40]. The neighbor-joining method using
the maximum likelihood technique to determine distance matrices
[41] was used to compare new and previously published BLV,
PTLV-I, and PTLV-II pol sequences (GenBank accession numbers
K02120, D00647, U45264-U45284, M99067-M99088, L27561L27571, L20351-L20361, U49798-U49820, L08408-L08409,
L1l456, MlO060, M76755, L02534, M92846, U12117, U12102,
L20651, D00294, M76751, 102020, U12114, U12111, L20641,
L20639, 20664, L20656, L20646, L20660, Z46900; also see [4244]) and HTLV-II LTR sequences (GenBank accession numbers
L37129-L37146, U10252-U10266, U46555-U46558, M99091,
M10060, L06856, L06860, U25135, Ll1456; also see [21, 4547]). Both distance matrix and bootstrap (l00 replications) trees
were generated.
Determination of HTL V-II positivity. In our previous report
on a subset of the Indian population [29], the sensitivities and
specificities of the screening HTLV ELISA used herein were 73%74% and 96%-99%, respectively. The sensitivity and specificity
for the HTLV Select ELISA were 72% and 98%, respectively, and
for the 2.3 HTLV Western blot were 70% and 100%. In contrast,
the sensitivity and specificity of the HTLV-II PCR assay were
97% and 100%, respectively. Hence, in the current study, any
sample that was positive for HTLV-II by WB or PCR was deemed
JID 1996; 174 (November)
to be positive. In the absence of WB or PCR analyses, any sample
that was positive in the Cambridge Biotech or Vironostika ELISA
(or both) and also positive for HTLV-II in the Select ELISA was
deemed HTLV-II-positive.
Results
peR and phylogenetic analyses. PCR analyses were done
on DNA samples from 246 Indians. All were positive for human ,8-globin. None was positive for HTLV-I. Ninety-four
samples were positive for HTLV-II pol DNA. When these
HTLV-II - positive samples were reamplified with signature
primers, all were negative, indicating that they were not contaminated with previously amplified DNA. Three of the 246
samples were negative for HTLV-II by PCR, yet the subjects
were positive for HTLV-II antibodies by both Select and WB
assays. Hence, these were deemed false-negative PCR results.
In contrast, among the 94 PCR-positive samples, only 67 (71%)
were confirmed seropositive. Eleven of the HTLV-II -seronegative, PCR-positive Indians were retested 19 months later and
2 had seroconverted. Amplified HTL V-II pol DNA samples
from 15 infected Indians (including 2 who were seronegative)
belonging to all six ethnic groups and all but one of the communities studied in the Gran Chaco were subtyped via cloning
and sequencing and liquid hybridization oligomer restriction
(figure 2). Similarly, LTR sequences were obtained from 3
HTLV-IIB-infected Indians (figure 3). All of these samples
were found to belong to the HTL V-IIB subtype. If the Gran
Chaco HTLV-IIB (T-1) strain is used as a reference, then 0
changes in 2100 bases were observed among the 15 Gran Chaco
pol sequences, while 15 changes in 1400 bases were observed
among the 10 non-Gran Chaco HTLV-IIB strains (P < .001,
Fisher's exact t test). Similarly, the 3 Gran Chaco HTLV-IIB
LTR sequences were significantly more homologous to each
other than to other HTLV-IIB strains (data not shown). Hence,
the Gran Chaco HTLV-lIB strains cluster as a highly conserved
subgroup that differs from most other North and Central American and European HTLV-IIB strains.
Distribution of HTLV-Il according to area, community, and
ethnic background. Using the criteria for positivity as outlined in Methods, 158 (20%) of 791 Indians examined were
positive for HTLV-IIB. None was positive for HTLV-I. Samples from 1008 non-Indian, 19-year-old Argentinian military
conscripts from the Gran Chaco were screened in the HTLVI Cambridge Biotech ELISA. Two were repeatedly reactive
but were negative when tested in the Vironostika and Select
ELISA and in the 2.3 WB assays. None of82 adult non-Indians,
including 10 with leukemia, seen in health clinics in the Gran
Chaco were positive for HTLV antibodies. In addition, 50
volunteer blood donors from Syracuse, New York, were serologically and PCR-negative for HTLV infection, while 20 each
of North American HTLV-I -, HTLV-II -, or HIV-1- infected
patient samples were positive in their respective serologic or
PCR assays.
lID 1996;174 (November)
HTLV-II in South American Indians
947
PTLV-I
Australasian
Figure 2. Bootstrap consensus tree (l00
replicates) of pol sequences (140 bases) of
bovine leukemia virus (BLV) and primate T
cell lymphoma/leukemia viruses (PTLVs)
analyzed by neighbor-joining method using
maximum likelihood technique to calculate
distance matrices. Branch lengths shown do
not correlate with distance. Bootstrap confidence values> 50% are shown on respective branches. Major subtypes of PTLV-I
and PTLV-II and the clustering of HTLVlIB strains identified in Gran Chaco Indians
are indicated. High bootstrap values for
PTLV-IIA and -lIB branches indicate that
Gran Chaco strains clearly belong to
HTLV-lIB subtype. However, bootstrap
value separating them from other HTLVlIB strains is not significant.
West Africa
BLV
West Africa
Gran Chaco
PTLV-II
Tables 1 and 2 summarize the distribution of HTLV-II in
Indians ~ 13 years old among the 20 communities and various
ethnic groups examined. As can be seen, the prevalence of
HTLV-II infection was highest in the Chorote and Chulupi,
somewhat less in the Toba, and considerably less in the communities composed of Wichi, Lengua, Ayoreo, or Mapuche. Infection was demonstrated by serology and PCR in all of the communities, except one inhabited by Mapuches. Although La
Merced and San Luis, inhabited exclusively by Chorote and
Wichi, respectively, have marked differences in the prevalence
ofHTLV-II (43.9% and 3.7%, respectively), they are separated
by <3 km. One of the positive Mapuche had received a blood
transfusion at a local hospital; however, the donor is unknown.
Table 3 shows the prevalence of HTLV-II in the Gran Chaco
Indians according to age and sex. As can be seen, there was
increased infection with age (14.2% in the cohort < 13 years
and 23.2% in those 13-89 years; P = .05 by Fisher's exact t
test), particularly for subjects ~40. Also, in those 13-89 years
old, there was an increased prevalence of HTLV-II in women
compared with men (26.0% vs. 19.2%; P = .03 by Fisher's
exact t test). These data suggested both mother-to-child and
sexual modes of viral transmission. The fact that infection
tended to aggregate in families supported these hypotheses. In
the high-incidence families studied (figure 4), infection was
demonstrated in the offspring of 9 of the 11 positive mothers,
whereas the spouses of only 3 of 17 infected married members
of these families were positive. The data in table 4 indicate
that the maternal transmission rate of HTLV-II was ,. . ., 30%. It
should be noted that 1 of the infected children was born to a
noninfected mother but was nursed from birth by an infected
foster mother. We could not obtain reliable information to
rule out the possibility that the 3 infected children born to
noninfected or nontested mothers had been nursed by an infected foster mother.
Among age- and sex-matched Indians > 13 years old,
HTLV-II infection was demonstrated in 30 (48%) of 62 born
to infected mothers but in only 9 (11%) of78 born to uninfected
mothers. Our study includes 19 HTLV-II - infected Indians
whose spouses and mothers were tested. As shown in table 5,
infection was significantly more frequent among the mothers
(89%) than among the spouses (37%) of these subjects. There
were 115 married couples of which both spouses were tested
for HTLV-II. In 68 of these couples, both spouses were negative, and in 17, both spouses were positive. There were 30
couples with discordant HTLV-II status; in 19 of them, the
wife was positive and the husband was negative, and in 11 the
wife was negative and the husband positive. Thirteen of the
16 tested mothers of the positive spouses in the HTLV-IIdiscordant couples were infected. Data further supporting both
maternal and sexual transmission of HTLV-II are presented in
table 6, which compares prevalence rates of infection in
spouses, mothers, and fathers of infected Indians.
Discussion
Our previous work indicated that PCR, WB, or a combination of screening ELISA followed by Select ELISA testing
were highly specific (100%) testing formats for HTLV-II infec-
Ferrer et aI.
948
lID 1996; 174 (November)
FUC
lib
C23723
PENN7a
W43
,~~~...-SPAN130
CAM--~
SPAN129
ITA47a
Figure 3. Neighbor-joining distance tree
of long terminal repeat sequences (576
bases) from HTLV-IIA and HTLV-lIB isolates worldwide. Branch lengths correlate
with distance between strains. Bootstrap
confidence values (100 replicates) >50%
are shown. High bootstrap value (88%) on
branch separating HTLV-IIA and -liB
strains is significant. Gran Chaco Indian
strains W43, W175, and CH610 belong to
branch of HTLV-liB strains that also includes Seminole strains SEM1050 and
SEM1051. GI2 is strain from Guaymi Indian from Panama. WYU I and WYU2 are
strains from Wayu Indians living in Colombia. Gran Chaco strains group together as
clade, but bootstrap value (38%) is not significant given amount of sequence analyzed. Kayapo Indian strains Braz A21 and
Kayapo form distinct subgroup of HTLVIIA sequences.
88%
KAYAPO
FLN
Iia
HTLV-II in South American Indians
lID 1996; 174 (November)
Table 1.
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Prevalence of HTLV-II infection by community among Gran Chaco Indians ~ 13 years old.
Area
Community
La Merced
La Paz, Kilometro I,
and Kilometro 2
La Esperanza, Pozo del
Tigre, and La Gracia
Monte Carmelo and La
Curb ita
Santa Maria
San Luis
La Vertiente
San Patricio
Chulupi aldeas
Chulupi aldeas
Lengua colonies
Lengua colonies
Ayoreo colonies
Pupenhuen
Ruccahroi
Ethnic
composition
No. HTLVII-positivel
total (%)
No. positive
by PCRlno.
tested
Santa Victoria Este
Chorote
36/82 (43.9)
11/27
Santa Victoria Este
Chorote, Wichi
47/144 (32.6)
31/59
Santa Victoria Este
Chorote, Wi chi
13/39 (33.2)
6/25
Santa Victoria Este
Santa Victoria Este
Santa Victoria Este
Santa Victoria Este
Los Blancos
Paraguay-Femheim
Paraguay-Salve Yanga
Paraguay-Femheim
Paraguay-Salve Yanga
Paraguay-Salve Yanga
Alumine
Alumine
Toba, Wichi
Wichi
Wichi
Wichi
Wichi
Chulupi
Chulupi
Lengua
Lengua
Ayoreo
Mapuche
Mapuche
5/21 (23.8)
2/48 (4.1)
2/53 (3.7)
4/33 (12.1)
2/17 (11.7)
4112 (33.3)
24178 (30.7)
4/37 (10.8)
1/12 (8.3)
2/51 (3.9)
2/33 (6.0)
0/61
5/20
2111
2/22
4/31
2/3
1/1
13113
2/2
III
1/2
1/1
NOTE. Closely related communities of common ethnicity and comparable incidences of HTLV-II infection have
been combined. PCR, polymerase chain reaction.
tion in Indians of the Gran Chaco [29]. PCR testing, however,
was more sensitive than a purely serologic approach (97% vs.
~ 72%). In the present study, the diagnosis of HTLV-II infection was made in 43 Indians by ELISA testing alone, whereas
115 were diagnosed as infected by additional PCR or WB
analyses (or both). Hence, we believe that only a minority of
false-negative results were obtained and that the overwhelming
majority of HTLV-II-positive samples were true-positives.
The extensive precautions and controls that were used should
preclude the possibility that the PCR results were due to contamination. This assumption was verified by the fact that analyses of the HTLV-II sequences obtained from 15 of the HTLV-
Table 2. Prevalence ofHTLV-II in Indians ~ 13 years old according
to ethnic groups.
Ethnic group
Present study
Chorote
Toba
Wichi
Chulupi
Chorote/Wichi'
Chorote/Chulupi"
Lengua
Ayoreo
Mapuche
611171 (35.6)
5115 (25.0)
26/204 (12.7)
32/94 (34.0)
4/14 (28.5)
8110 (80)
5/49 (10.2)
2/51 (3.9)
2/94 (2.1)
NOTE. Data are no. positive/total (%).
* [32].
, Ethnicity of mother/ethnicity of father.
Previous
study*
181121 (14.8)
2/40 (5.0)
Both studies
61/171 (35.6)
23/136 (16.9)
28/244 (11.5)
32/94 (34.0)
4/14 (28.5)
8110 (80.0)
5/49 (10.2)
2/51 (3.9)
2/94 (2.1)
II-positive Indian samples (including 2 from seronegative subjects) typed them as unique and highly related HTLV-lIB substrains. The observation that 19 months later, only 2 of 11
HTLV-II PCR-positive, serologically negative Indians had seroconverted indicates that the seroconversion "window" following HTLV-II infection may be quite prolonged or that some
subjects may harbor a defective provirus.
The data allow for several conclusions regarding the epidemiology ofHTLV-Il infection among Paleo-Amerindians. Our
observations show that the prevalence rates of HTLV-II infection vary markedly among the various ethnic communities,
even those in close proximity to each other. This is unlikely
to be due to genetic factors in the hosts, because when communities that are cohabited by high-prevalence (e.g., the Chorotes)
and low-prevalence (e.g., the Wichi) ethnic groups are tested,
the frequency of infection in the two ethnic groups is virtually
Table 3. Prevalence ofHTLV-II in Gran Chaco Indians according
to age and sex.
No. postive/total (%)
Age (years)
<13
13-19
20-33
34-47
48-61
;;.62
13-89
Female
Male
All
3/32 (9.3)
12/69 (17.3)
221127 (17.3)
31/97 (31.9)
19/55 (34.5)
12/26 (46.1)
97/373 (26.0)
7/38 (18.4)
3/41 (7.3)
15/87 (17.2)
7/47 (14.8)
13/52 (25.0)
11126 (42.3)
49/254 (19.2)
10170 (14.2)
151110 (13.6)
37/214 (17.2)
38/144 (26.3)
32/107 (29.9)
23/52 (44.2)
146/627 (23.2)
950
Ferrer et al.
JID 1996; 174 (November)
Family 1
Family 2
Family 3
Family 4
Figure 4. Pedigrees of 4
families in which presence of
HTLV-II was examined in
members of 3 generations.
Black circles and black
squares denote infected females and males, respectively. Numbers within circles and squares indicate ages
of subjects. NT, not tested.
HTLV-II in South American Indians
JID 1996; 174 (November)
Table 4. HTLV-II infection in children < 13 years old born to or
breast-fed (or both) by infected women.
HTLV-II infection of
mother or foster mother
Positive
Negative or unknown
951
Table 6. HTLV-II infection in spouses, mothers, and fathers of
infected Indians of whom at least one of these relatives was tested.
No. of infected
children/total (%)
Ages of infected
children (years)
Sex of infected
subject
Spouse
Mother
Father
7*/24 (29.1)t
3/46 (6.5)t
6, 7, 8, 8, 8, 9, 9
8,9, 12
Female
Male
Female + male
17/36 (47.2)
17/28 (60.7)
34/64 (53.1)*
22/29 (75.8)
14/19 (73.6)
36/48 (75.0)t
7/20 (35.0)
4/10 (40.0)
11/30 (36.6)t
* 5 were PeR-positive.
t
p < .01 (x 2 ) .
the same. Rather, the high prevalence of HTLV-II in certain
communities would seem to be due to a founder effect or a
result of certain disparate behavioral customs.
Previously reported data on the natural mode of transmission
of HTL V-II in Indian populations have been obtained in studies
in which the subjects were examined only by serology [4951]. However, as shown by our present and previous [29]
studies, in high-incidence populations, serologic tests fail to
identify up to 25% of the HTLV-II-infected persons who are
PCR-positive; thus, it is evident that extensive PCR testing is
required to obtain more definitive information on the natural
mode of transmission of the virus. Therefore, in the present
study a large number of serologically negative and serologically
positive subjects were examined by PCR. Our data indicate a
very high rate (""""30%) of mother-to-child transmission, presumably by breast feeding. Results supporting significant levels
of perinatal HTLV-II transmission are evident in all age groups
studied, particularly in the high-incidence families. Likewise,
a reasonable case can be made for sexual transmission, as the
prevalence of infection increases after puberty. By age 34, the
prevalence rate was almost double that observed in children
< 13 years old. The fact that after puberty, the prevalence of
BTLV-II is significantly greater in women than in men suggests
that transmission from male to female via intercourse is more
Table 5. HTLV-II infection in spouses and mothers of 19 infected
Indians of whom both were tested.
HTLV-II status
Sex ofHTLV-IIpositive subject
Female
Female
Female
Female
Male
Male
Male
Male
Female
Female
Female
Female
+ male
+ male
+ male
+ male
Mother
Spouse
No.
Positive
Positive
Negative
Negative
Positive
Positive
Negative
Negative
Positive
Positive
Negative
Negative
Positive
Negative
Positive
Negative
Positive
Negative
Positive
Negative
Positive
Negative
Positive
Negative
3
6
* 6 of 10 seronegative spouses were PCR-negative.
o
1
4
4
o
1
7
10*
o
2
NOTE. Data are no. positive/total (%). P < .02, Fisher's exact t test *
vs. t; P < .001, Fisher's exact t test, * vs. t and t vs. i.
'
* 14 of 30 seronegative spouses were tested by PCR and all were found
negative.
t 7 of 12 seronegative mothers were tested by PCR and all were found
negative.
i 11 of 19 seronegative fathers were tested by PCR and all were found
negative.
efficient than vice versa. These observations are all consistent
with those previously reported for the PTLV (reviewed in [1,
2, 48-51]).
The data herein also strengthen the hypothesis that HTLVlIB is endemic throughout Paleo-Amerindian groups and may
have been either brought by early migrants from Asia or transmitted to them from an animal source after they crossed Beringia 15,000-30,000 years ago. Because 1% of divergence
within the SKllO/SKlll pol sequence of the PTLV is thought
to represent """"500-1000 years of separation of the host populations [1, 2, 19, 25, 28], our genetic data would suggest that the
Gran Chaco Indians have been separated from the Guaymi
(Panama), Wayu (Colombia), and Seminole Indians (North
America) for 500-3000 years and from the Kayapo Indians
(Brazil) and other HTLV-IIA-infected groups (e.g., the Navajo
and Pueblo in North America) by 3500 to > 7000 years.
Whether these theoretical time frames represent points of genetic divergence or cessation of contacts that could lead to
retroviral infections among Indian groups is uncertain. There
has been some debate as to the origin of HTLV-II strains among
modem people. The above data, plus the fact that there is a
higher prevalence of both HTLV-IIA and HTLV-lIB in humans
of European or African descent living in the New World compared with the prevalence of these infections in current populations from their ancestral continents, suggest that both HTLV IIA and HTLV-IIB were originally endemic in the Americas
and only relatively recently were disseminated throughout the
world [1, 2, 12-19].
Additional molecular comparisons ofPTLV strains from different native populations from around the world will be crucial
to gain new insights into the origin and evolutionary relationships of HTLV-II variants. Clinical and epidemiologic studies
of these infected populations would also be warranted to discern if the various HTLV-II subtypes in their particular hosts
manifest different biologic properties and diseases.
Acknowledgments
We thank N. Auza and P. Soto (Facultad de Ciencias Veterinarias, Universidad del Centro de la Provincia de Buenos Aires,
952
Ferrer et a1.
Argentina) for their invaluable support; C. M. Julia (Ministerio de
Salud y Accion Social, Buenos Aires), J. C. Gomez Alvarenga, 1.
Nunez Burgos, C. Marquez, and A. Aleman (Ministerio de Salud
Publica De Salta, Argentina), D. Lucero (Ministerio de Salud Publica de Neuquen, Argentina), W. Kaethler and A. Stahl (Programa
Pro-Salud Indigena ASCIM, Filadelfia, Paraguay), and L. C. Estigaribia (Ministerio de Asistencia Social y Salud Publica of Paraguay) for facilitating our access to the Indian communities and the
transportation of equipment; D. Campos for assistance in several
logistic aspects of our trips to the communities; and R. Boston for
suggestions regarding the analysis of data.
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