Coinfeccion y Superinfeccion
Coinfeccion y Superinfeccion
Coinfeccion y Superinfeccion
ment gap were excluded from the analysis. Best-fit models of clade and unrelated HIV Spanish isolates was 2.2%–17.8% for
nucleotide substitution were selected according to the Akaike clade a and 2.0%–18.3% for clade b; the corresponding values
information criterion in Modeltest (version 3.6; David Posada for patient 2 were 6.3%–19.2% for clade a and 7.4%–20.4%
Web page, http://www.darwin.uvigo.es). Maximum-likelihood for clade b. The presence of 2 statistically differentiated clusters
trees were estimated under the best-fit model by use of the within these 2 patients’ quasispecies, together with the greater
algorithm implemented in Phyml (version 2.4.1; http:// genetic distance between clusters a and b compared with that
www.atge.lirmm-fr/phyml) and by starting the search from a in unrelated viruses, indicates that HIV dual infection with
BIONJ tree. Phylogenetic confidence was assessed by bootstrap subtype B has occurred in these 2 patients.
analysis of 1000 replicates [10]. Identical sequences derived Because the 2 groups (i.e., a and b) of nucleotide sequences
from the same or different samples were used only once to were present in all the analyzed samples from each patient and
reconstruct the phylogenetic tree shown in figure 2. because no samples close to the seroconversion time were avail-
To perform the viral dating of the different viral populations able, it was not possible to determine whether coinfections or
in each patient, the genetic distance of the reconstructed most superinfections had occurred. To address this problem, we per-
recent common ancestor (MRCA) for each clade was compared formed viral dating of the different patients’ clusters, according
with that of a reconstructed MRCA for the Spanish epidemic. to the method that our laboratory had established for Spanish
The viral dating time was estimated by use of a linear-corre- isolates and that has been explained in the “Patients and meth-
lation equation previously developed on the basis of a large set ods” section above. According to this method, we interpolated
of Spanish samples, an equation that correlates the V3 nucle- the year of the nucleotide sequence of each of the different
otide-sequence divergence to the Spanish-epidemic MRCA and patient clades (table 1). The years obtained for clades a and b
the sampling time [11, 12]. (Genbank accession numbers are for patient 1 were identical (i.e., 1992), whereas those obtained
EF517970–EF518214.) for clades a and b for patient 2 were different; for the latter
Results. Phylogenetic analysis of viral sequences obtained patient, the dating year for clade a was 1987, close to the se-
from Spanish patients with LTNP HIV-1 disease and of nucle- roconversion time, and the dating year for cluster b was 1996
otide sequences derived from unrelated Spanish patients with (i.e., 9 years later). The viral dating indicates that a superin-
HIV-1 disease (either LTNP or typically progressive) showed fection had occurred in patient 2, whereas analysis of the first
that all isolates were subtype B. Nucleotide sequences obtained sample from patient 1 showed that he already was coinfected,
from each patient formed monophyletic groups with high boot- although a previous superinfection could not be ruled out.
strap values (175%), except for patients 1 and 2, each of whose Discussion. In the present report, we have presented data
nucleotide sequences were separated into 2 groups (a and b in on dual infections, with subtype B variants, in 2 patients with
figure 2), situated in different branches of the tree, and sup- LTNP HIV-1 disease who had no clinical manifestations. Al-
ported by bootstrap values 185%. though both patients had LTNP characteristics, 115 years of
The mean genetic distances between groups a and b—5.4% infection, and no clinical symptoms and had not received an-
for patient 1 and 11.2% for patient 2 (table 1)—were higher tiretroviral therapy, they showed different levels of CD4+ T cell
than the genetic distances to other nonepidemiologically related counts, RNA viral load, and risk practices. Whereas patient 1
Spanish HIV-1 isolates (table 1), as indicated in other studies showed multiple peaks in viral load and a continuous but slow
[3]. For patient 1, the range of genetic distances between each decline in CD4+ T cell count, patient 2 showed a more constant
CD4+ T cell count, undetectable or low RNA l load, and 2 mitted estimation of the time of origin of either the global
sporadic blips. Both individuals were and remain classified as epidemic or the epidemic in individual countries [13], although
having LTNP disease. the approach depends on the method used. Along this line, we
We used a phylogenetic approach to distinguish between have previously demonstrated that a molecular clock operates
coinfection and superinfection events. It is known that a mo- in the Spanish HIV-1 epidemic [12]. This approach allowed us
lecular clock operates in HIV-1 evolution, and this has per- to estimate the viral dates for different isolates and to detect
sequences with different dates within individual quasispecies the need for samples to be obtained close to the superinfection
[11, 14]. Using this methodological approach, we obtained the event, could contribute to the underdetection of superinfection.
same date for the 2 viruses in patient 1 (1992) and different In general, superinfection has been detected only in patients
dates (1987 for clade a and 1996 for clade b) for the 2 viruses in whom an increase in viral load set point or in other path-
in patient 2. These results strongly suggest that patient 2 was ogenic markers has occurred.
infected with 1 virus and then, 9 years later, became superin- Previous reports in the literature [3–6, 9, 15] have generally
fected with another virus, which showed an 11.2% genetic dis- assumed that superinfection is associated with disease pro-
tance from the first virus. In contrast, patient 1 was coinfected, gression. Therefore, patients with LTNP disease who have not
for at least 18 years, with 2 viruses differing by a genetic distance experienced disease progression for 110 years could be ex-
of 5.4%, although it is not possible to rule out a previous pected, a priori, to have no superinfections—or at least a low
superinfection (table 1). The present study indicates the use- rate of superinfection. Moreover, if superinfection occurs in 2
fulness of a viral dating strategy to clarify situations that occur groups of patients with HIV-1 with different characteristics
in HIV-1 viral evolution in infected patients [11, 14]. These (e.g., typical and LTNP), it can be considered a general phe-
data show that in these 2 patients there was control of the nomenon that probably occurs in every group of infected pa-
replication of not only a single virus but also of a second, tients. In summary, in the present study we have found that
divergent strain. Moreover, the superinfection in patient 2 oc- dual infections also occur in patients with LTNP HIV-1 disease
curred 9 years after the primoinfection and in the presence of and are not associated with clinical manifestations.
an effective immune control of viral replication.
In most previous reports in the literature, dual infection has
Acknowledgments
been associated with a clinical deterioration of patients [5].
Only 1 case of superinfection in a long-term survivor has been We thank Ingrid Outschoorn for review of the manuscript.
described, and this individual showed a clear progression of
disease after an acute febrile illness and the detection of a
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