Our results suggest a positive relationship between maintenance of viral compartments at the C2V3 region of the envelope gene within the host and the integrity of the CD4+
T cell population. Our group previously found a loss of viral compartmentalization between vaginal fluid and plasma for the pol gene in a report of an individual with a CD4+
T cell count under 50 cells/μl that showed drug resistance mutations in the absence of ART(Tirado et al., 2004a
). Several other studies have identified interesting patterns of viral load concordance between vaginal fluid and plasma. For example, analysis of over seventy women found detectable virus in 85% of plasma samples, while virus was detected in only 39% of the same individuals at coincident vaginal fluid sampling(Uvin and Caliendo, 1997
). However, when the data were stratified by CD4+
counts, 67% of patients exhibited concordant detection of virus in plasma and vaginal fluid from a single time point. This study was unable to differentiate the role of loss of immune control of viral replication within compartments versus mixing of virus across compartments in driving the concordant detection of virus in plasma and vaginal fluid with lower CD4+
counts. However the findings are consistent with our data showing loss of compartmentalization with lower CD4+
T cells. In addition, only 50% of compartmentalized patients versus 75% of intermingled participants, showed detectable viral loads across plasma and vaginal fluid samples ().
Several other groups have reported a direct relationship between higher CD4+
counts and viral compartmentalization in the female genital tract compared with plasma. Kemal, et al
. found correlation between higher CD4+
levels and distinct viral populations in plasma and vaginal tract cell-free virus(Kemal et al., 2003
). Their study used cervico-vaginal lavage, which obtains virus from endocervix, vagina and fornix and represents a larger portion of the lower genital tract than our vaginal swab sampling assessed, indicating a greater extent of tissue is compartmentalized. A later study by the same group analyzed full length RNA genomes from plasma and female genital tract and found strong evidence for compartmentalization(Philpott et al., 2005
). Interestingly, the association between CD4+
T cell levels and compartmentalization was less clear, as only a single study subject showed intermingled viral forms in the two compartments and this study subject had counts of 306 cells/μl. In contrast, three participants had lower counts and showed compartmentalization. There was evidence for recombination between plasma and vaginal tract virus in the patient with the lowest CD4 counts (6 cells/μl). This suggested mixing of virus between plasma and the female genital tract had occurred and perhaps the viral forms represented influence of recent replication as this individual had a genital tract viral load of 2.5million copies/mL(Philpott et al., 2005
). A different study of clade A infection demonstrated clear blood and genital fluid viral compartmentalization in four women with clinically asymptomatic HIV-1 infection, again arguing for a strong role of the intact immune response in maintaining divergence of virus across compartments(Andreoletti et al., 2007
). Sullivan et al
., have also reported an association between CD4+
levels and viral divergence using heteroduplex mobility shift analysis of env
C2V4, which they suggested was evidence for immune status in maintenance of compartmentalization(Sullivan et al., 2005
). Thus our data further support the suggestion that the immune response impacts the compartmentalization of viral forms in genital and plasma fluids.
The important role of an intact immune system in maintaining selective pressure on HIV is a clear function of the CD4+ T cell sub-set; however, the role of the CD4+ T cells in this phenomenon in our dataset is more complex. Only a single participant, 001M, had evidence for immune selection according to calculation of the synonymous:non-synonymous change ratio. Yet the viral load for this patient demonstrated loss of control of the virus at time of sampling, suggesting the selection either occurred prior to sampling or was driven by other factors like target cell repertoire within the compartment. Alternatively, this may represent emergence of an archival form.
Initial comparison of plasma-vaginal fluid compartment divergence showed a rough correlation with CD4+ T cell level; however, patient 006M clearly showed high intracompartmental diversity even with a very low cell count, again indicating a complex role. Upon closer examination, this patient was remarkable in having high diversity in both plasma and vaginal fluid, particularly in comparison with the overall plasma-vaginal fluid divergence, which was quite low at 0.5%. In and , we clearly observe a direct relationship between compartmental separation and CD4+ T cells over 350 cells/μl. Thus divergence between compartments is only maintained at higher CD4+ levels. This may suggest that T cells are important for establishing or at least maintaining distinctions in the niches of viral replication in different anatomical regions of the host, whether or not the diversity within those niches is controlled. Thus the breakdown in the CD4+ T cell compartment by loss below 200 may result in loss of host compartmental selective forces thus allowing the systemic viral forms to become more uniform. In the case of 006M, since the intracompartment diversity is already high, the intercompartmental divergence represents greater diversity of forms within the entire host, rather than maintenance of unique viral niches, thus the net divergence is quite low. In contrast, no other patient has high diversity in both plasma and vaginal fluid. Among these five study participants, the intercompartmental divergence more closely follows a direct comparison with CD4+ levels.