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Presence of Hepatitis C Virus (HCV) RNA in the Genital Tracts of HCV/HIV-1-Coinfected Women
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The Journal of Infectious Diseases Nov 1 2005;192:1557-1565
HCV RNA can be detected in almost 30% of HCV/HIV-1-coinfected women
Marek J. Nowicki,1 Tomasz Laskus,2,3 Georgia Nikolopoulou,1 Marek Radkowski,2,4 Jeffrey Wilkinson,2 Wenbo B. Du,1 Jorge Rakela,2 and Andrea Kovacs1
1Maternal-Child and Adolescent Center for Infectious Diseases and Virology, University of Southern California, Los Angeles; 2Mayo Clinic, Scottsdale, and 3St. Joseph's Hospital and Medical Center, Phoenix, Arizona; 4Medical Academy, Warsaw, Poland
....Findings from our study suggest that a local HCV genital tract reservoir may exist and that this may be the source of infection for those suspected to have been infected sexually, a possibility further supported by the analysis of HCV quasispecies isolated from plasma, PBMCs, and CVL fluid..... Our findings may explain a comparatively higher rate of HCV vertical transmission by HIV-1-coinfected women reported in several studies..... The mechanism by which local HIV-1 facilitates the presence of HCV in the genital tract is unclear.... concomitant infection of the same cells by both viral pathogens with subsequent enhancement of HCV replication, possibly through transactivating properties of the HIV-1 Tat protein... Another possibility is local immune dysfunction, which would allow both viruses to replicate locally....
".....To our knowledge, our study is the first to demonstrate compartmentalization of HCV in the genital tracts of HCV/HIV-1-coinfected women and possible local replication in a large proportion of HCV/HIV-1-coinfected women.... These findings have important implications for both sexual and perinatal transmission of HCV. Increased mother-to-infant and sexual HCV transmission in HCV/HIV-1-coinfected women makes it especially urgent to study and understand the dynamics of HCV in this subset of patients.... Our study also suggests that, among HIV-1-infected women who are HCV viremic, there is an association between shedding of both viruses and that local control of both viruses may be impaired in those found to be shedding. This may explain the increased rate of perinatal HCV transmission to HIV-1-infected newborns and the observation that sexual transmission may be increased in coinfected patients....
INTRODUCTION
Hepatitis C virus (HCV) infection is common among HIV-1-infected patients, with 50%-90% being coinfected [1]. Thus, HCV coinfection has emerged as a major public health problem that contributes to significant morbidity and mortality in HIV-1-infected patients. HIV-1 coinfection accelerates the development of severe liver disease attributable to HCV [1, 2], whereas HCV coinfection has been reported to accelerate the progression of HIV-1 disease [3, 4]. Since the introduction of effective measures to screen blood and blood products for HCV, injection drug use has become the predominant mode of HCV acquisition. However, exposure via injection drugs cannot account for up to 20% of new infections [5, 6]. Several reports have suggested that HCV may be transmitted through sexual intercourse [5, 7, 8], during childbirth [9, 10], and even during casual contacts between household members [11]. The probability of mother-to-child and female-to-male transmission appears to increase in the presence of coinfection with HIV-1 [10, 12-14]. Similar findings have been reported for HIV-1-positive men who engage in high-risk sexual behaviors (such as unprotected sex) with other men [15]. However, there are conflicting results with regard to female-to-male HCV transmission [16, 17]. The mechanism of increased HCV replication in HIV-1-infected patients has been attributed to immunosuppression, as evidenced by high viral loads in patients receiving immunosuppressive drugs after transplantation [18]. However, there is also evidence suggesting that HCV replication may be directly enhanced by the presence of HIV-1 [19].
Surprisingly, despite mounting evidence for the existence of female-to-male and mother-to-child transmission of HCV, very little is known about vaginal and cervical shedding of HCV in HIV-1-positive and HIV-1-negative women. Very few studies have assessed HCV RNA in vaginal secretions. Furthermore, HCV load and quasispecies distribution in the genital tract compartment has not been analyzed previously [20-22]. This could be explained by the difficulties of obtaining cervicovaginal specimens, the methods for which have only recently been standardized in HIV-1 settings [23]. In this article, we present our findings on HCV detection, viral load, and quasispecies composition in the female genital tract using cervicovaginal lavage (CVL) fluid from HCV/HIV-1-coinfected women.
ABSTRACT
Background. Hepatitis C virus (HCV)-infected women-in particular, those coinfected with human immunodeficiency virus type 1 (HIV-1)-can transmit infection to their children and sex partners.
Methods. The present study was conducted to analyze the presence of HCV RNA in cervicovaginal lavage (CVL) fluid from 71 women (58 HCV/HIV-1-coinfected women and 13 HCV-infected, HIV-1-uninfected women) enrolled in the Women's Interagency HIV Study.
Results.
HCV RNA was detected (by a commercial polymerase chain reaction assay) in CVL fluid from 18 (29%) of the HIV-1-infected women and from none of the HIV-1-uninfected women (P < .05).
Multivariate analysis revealed that risk factors for the presence of HCV RNA in CVL fluid were HCV viremia (odds ratio [OR], 16.81; P = .02) and HIV-1 RNA in CVL fluid (OR, 19.87; P = .02).
This observation suggests local interactions between HIV-1 and HCV in the genital tract compartment.
There was no correlation between HCV RNA in CVL fluid and CD4, CD8, or CD3 cell counts, HIV-1 RNA viremia, the number of leukocytes in CVL fluid, or HIV-1 therapy.
Furthermore, in 3 of 5 analyzed patients who had a detectable CVL HCV RNA load, we found viral variants differing in the 5' untranslated region that were present neither in plasma nor in peripheral-blood mononuclear cells.
Conclusions. Our observations point to the importance of the genital tract compartment, in which local HCV replication could be facilitated by local HIV-1 replication.
Demographic and clinical information for the 58 HCV/HIV-1-coinfected women who were and were not found to be shedding HCV in the genital tract can be seen in table 1. The majority of the women were >35 years old, were black or Hispanic, and were unemployed. More than 40% reported having >10 lifetime sex partners, and 79% reported a history of injection drug use. Seventy percent had received highly active antiretroviral therapy within the previous 6 months, 52% had HIV-1 RNA loads <1000 copies/mL, and 80% had CD4 cell counts >200 cells/mL. HCV RNA was detected in plasma from 67% (39/58) of the HCV/HIV-1-coinfected women, compared with only 46% (6/13) of the HCV-infected, HIV-1-uninfected women, and they also had higher plasma HCV RNA loads (median, 1.5 x 106 copies/mL [range 1.5 x 104 to >4.0 x 106 copies/mL] vs. 0.3 x 106 copies/mL [range, undetectable to 1.3 x 106 copies/mL]) (data not shown).
HCV RNA was detected in CVL fluid from 31% (18/58) of the HCV/HIV-1-coinfected women, although the loads were relatively low (median, 1.5 x 103 copies/mL; range, undetectable to 0.4 x 104 copies/mL); 16 of the 58 women had loads <0.8 x 103 copies/mL. None of the HCV-infected, HIV-1-uninfected women (P = .03) had detectable HCV RNA in CVL fluid when tested by a commercial HCV RNA assay (Roche Diagnostic Systems), although 2 had detectable HCV RNA by our in-house qualitative HCV PCR assay. In general, the coinfected women with detectable HCV RNA in CVL fluid were similar to those with undetectable HCV RNA in CVL fluid, except that they had higher plasma HCV RNA loads (P < .04). For the relationship between log10 HIV-1 RNA load in plasma and CVL fluid, Pearson's R = 0.50 (P = .0003); for the relationship between log10 HCV RNA load in plasma and CVL fluid, Pearson's R = 0.33 (P = .0111).
AUTHOR DISCUSSION
Despite the mounting evidence supporting the sexual transmission of HCV, very few reports have addressed the issue of HCV shedding in the female genital tract. In an early study, Caldwell et al. [39] reported the occasional presence of HCV RNA in multiple body fluids (including vaginal fluid collected by use of swabs), whereas, in more recent reports, Manavi et al. [21, 22] found that HCV RNA was present in cervical swabs from 8 of 22 HCV-infected women. Similar results were reported by Belec el al. [20], who detected HCV in cervicovaginal secretions from 4 of 19 women. However, none of these studies included HIV-1-positive patients. In the present study, we found that none of the HCV-infected, HIV-1-uninfected women were positive for HCV RNA in CVL fluid when tested by a commercial quantitative assay. However, both HIV-1-uninfected women in an intensely studied group were positive when tested by our sensitive in-house qualitative HCV PCR assay. These results suggest that HCV may be present at very low levels in the genital tract of some women infected with HCV but not with HIV-1. However, significant levels (>600 IU/mL) of HCV RNA in genital secretions were present only in the genital tracts of the HCV/HIV-1-coinfected women in our study. These data contrast with those published recently by Belec et al. [20, 40], who reported that up to 27% of the cellular fractions of CVL specimens from HCV-infected, HIV-1-uninfected women were HCV RNA positive. This discrepancy is most likely due to the assays used, because 2 of 3 HIV-1-uninfected women had detectable HCV RNA by our in-house assay, compared with none by a commercial assay. This may also be related to a more concentrated CVL specimen used by Belec et al. and the relatively small number of HCV-infected and HIV-1-uninfected women studied by us. The HCV/HIV-1-coinfected women in our study had rates of HCV shedding similar to those noted in the study of HIV-1-negative women by Belec et al. [40]. Additionally, inhibitors of enzymatic amplification in serum, and especially those in mucosal sites such as vaginal and oral compartments, may cause false-negative results for direct detection of HCV RNA by PCR. However, our inclusion of the ultracentrifugation step during HCV RNA extraction from CVL specimens was likely to reduce the effect of soluble PCR inhibitors. In our own experience, the ultracentrifugation step is able to eliminate the inhibitory effect in -50% of specimens that are initially negative for HIV-1 RNA due to inhibitors of the amplification reaction (data not shown). However, it is plausible that some of the women with undetectable HCV RNA in CVL fluid may shed intermittently, and multiple sampling may yield positive results, as seen in HIV-1-infected women [27]. Thus, our data may underestimate the overall prevalence of HCV cervicovaginal shedding.
HIV-1 may locally affect HCV shedding on a molecular level or by immunologic mechanisms such as cytokines and immune activation. The latter possibility is in agreement with recently published data from Manavi et al. [21, 22] that suggest that HCV RNA in the genital tract is associated with lymphocytes and that the virus could possibly replicate there. Although high levels of HCV RNA in CVL fluid from 2 women correlated with traces of blood, the more frequently observed low-level viremia in CVL fluid of -1000 copies/mL appears to be unrelated to blood contamination, as was shown by our multivariate analysis. Furthermore, the detection of unique CVL-derived viral variants different from those detectable either in plasma or PBMCs suggests the presence of local extrahepatic replication. This observation is in agreement with the findings of Belec et al. in HIV-1-uninfected women [40]. Their data strongly suggest an association between HCV and cells in the genital tract compartment. At present, it is unknown which cells harbor or support HCV replication in the cervix or vagina. Data from Manavi et al. point toward cervicovaginal lymphocytes as the likely source [21, 22]. Macrophages have also been reported to harbor replicating HCV in HIV-1-coinfected patients [41] and seem to play a role in mother-to-infant and sexual HIV-1 transmission [42, 43].
The mechanism by which local HIV-1 facilitates the presence of HCV in the genital tract is unclear. One possibility is concomitant infection of the same cells by both viral pathogens with subsequent enhancement of HCV replication, possibly through transactivating properties of the HIV-1 Tat protein [44]. We have recently shown that the same cell could indeed be infected by both viral pathogens [45]. Another possibility is local immune dysfunction, which would allow both viruses to replicate locally.
In summary, we have found that HCV RNA can be detected in almost 30% of HCV/HIV-1-coinfected women and that viral diversity does exist between local HCV and plasma HCV extracted from HCV/HIV-1-coinfected women. Our findings may explain a comparatively higher rate of HCV vertical transmission by HIV-1-coinfected women reported in several studies. The relationship between HIV-1 and HCV shedding is intriguing and suggests a unique local interaction between these 2 viruses in the genital tract.
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