Since 2000, cross-sectional and cohort studies have reported an increasing incidence of acute hepatitis C virus (HCV) (AHCV) infection and reinfection among HIV-positive MSM in many regions worldwide including France [1–3]. Transmission of HCV in this population has been associated with high-risk practices such as unprotected mucosal-traumatic sex (fisting), group sex, but also nasal or intravenous drug use . At the same time, the spread of hepatitis C in HIV-negative MSM appeared to be limited , although AHCV infections in this population may have been underestimated due to less frequent HCV screening and alanine aminotransferase (ALT) assessment. Indeed, sexual transmission of HCV also occurs among HIV-negative MSM, especially those who are included in preexposure prophylaxis (PrEP) programs across the world [6–8]. In Europe, recent reports showed that the prevalence of HCV infection among MSM at enrollment in PrEP ranges from 1.8 to 4.4% [9,10]. In the French Dat’AIDS cohort, the overall incidence of HCV infections in HIV-negative MSM on PrEP was 1.2/100 person-years (including 0.2/100 person-years of reinfection), similar to the rate observed in HIV-positive MSM [2,9].
Currently, international guidelines recommend that HIV-negative MSM starting PrEP should be tested for HCV infection as part of baseline laboratory assessment with an anti-HCV antibody test . However, evidence regarding the frequency of further HCV screening in MSM at risk for ongoing exposure to HCV is lacking. Due to the high incidence of HCV infection in MSM on PrEP, at least annual HCV testing and ALT assessment are recommended in this subgroup . Recent French guidelines propose a quarterly testing for high-risk individuals , even though this is not systematically implemented. Another point that remains a matter of debate is which is the best test to use for AHCV screening. Diagnosis of AHCV is complicated by the lack of well defined criteria, and the fact that it is frequently asymptomatic . Currently, the anti-HCV antibody immunoassay (EIA for ELISA immunoassay) remains the gold standard for HCV screening in laboratories, but the use of HCV antibody rapid tests is spreading in places of point-of-care (POC) testing (POCT). However, because of the possibility of recurrent HCV infections, a nucleic acid test for HCV RNA should be performed when AHCV is suspected . HCV viral load tests are currently performed on automated platforms with high throughput, but their use is limited by a high cost and the requirement of an experienced laboratory. POC molecular testing has recently emerged, as the GeneXpert HCV RNA (Cepheid, Sunnyvale, California, USA) demonstrated excellent performance in the diagnosis and monitoring of HCV infection . Finally, the anti-HCV antigen EIA is a specific and sensitive assay for the diagnosis of AHCV in HIV-positive MSM [17,18], but its diagnostic performances have not been evaluated in HIV-negative MSM.
In the changing landscape of treatment as prevention and PrEP, reliable and up-to-date epidemiological data on HCV among HIV-uninfected MSM are needed, to help developing evidence-based testing policies. The aims of our study were the following: first, assess the incidence of the AHCV among high-risk MSM in the ANRS IPERGAY PrEP trial; second, identify the risk factors for HCV infection in this population; third, determine the sensitivity of both indirect (EIA and antibody rapid tests) and direct (EIA Antigen and HCV RNA) tests available for an early diagnosis of AHCV in HIV-negative MSM on PrEP.
Materials and methods
Study design and population
All the participants from the ANRS IPERGAY PrEP trial enrolled during blinded and/or open phases were analyzed. Inclusion and exclusion criteria have been previously described [6,19]. Briefly, eligible participants were HIV-negative MSM, aged 18 years or older who were at high risk for HIV acquisition, defined as reporting unprotected anal sex with at least two different partners over the previous 6 months. They were enrolled at seven different sites (six in France and one in Canada) from February 2012 to June 2016. All participants provided written informed consent at inclusion in the trial.
Follow-up study visits were scheduled at enrollment, 4 and 8 weeks later, and every 2 months thereafter until 30 June 2016. Before each visit, participants were asked to complete a questionnaire to collect the following information: sociodemographic characteristics, alcohol and/or drug consumption (although using and/or sharing injecting equipment was not systematically recorded), sexual behavior (number of partners over the past 2 months, number of sexual acts over the past 4 weeks, condomless receptive anal sex on the last sexual act) and factors associated with a high-risk of HCV acquisition (fisting or bleeding after sexual intercourse). Each visit included serum testing for HIV infection with a fourth-generation antibody and antigen assay, and biochemical analyses (ALT, serum creatinine level). Stored sera were kept for all participants at each visit. Participants were also tested every 6 months for the following sexually transmitted infections (STI): syphilis (serology), gonorrhea and chlamydial infection (specific PCR done on rectal and throat swabs, and on urine samples).
For the diagnosis of HCV infection, a third-generation antibody immunoassay (EIA ARCHITECT HCV Ab; Abbott, Chicago, Illinois, USA) was performed: first, at screening; second, every 12 months thereafter; third, if ALT (performed every 2 months) raised more than 2.5 times the upper limit of normal.
Acute hepatitis C virus diagnosis and calculation of incidence rate
AHCV infection during follow-up was defined as the time of first positive HCV antibody test, with a negative serology at screening, according to the methodology used in the literature. One participant had a positive HCV serology at the screening visit and was excluded from the IPERGAY trial. Person time for participants with acquired HCV infection was estimated by use of the time from study entry in the trial (at inclusion and not at screening) to the date of diagnosis (based on the first HCV-positive serology and not based on past serum showed infection date). For participants without HCV infection, person time was estimated time from study entry to the time of last visit follow-up in the study. The calculation of the exact 95% confidence intervals (CIs) for incidence estimates was based on the number of events modeled as a Poisson variable for the total person-years. For each participant diagnosed with AHCV, plasma HCV RNA and HCV subtype were determined.
Evaluation of the different tests for the diagnosis of acute hepatitis C virus
In participants with a positive EIA ARCHITECT HCV Ab, we used stored sera to perform the following tests at the time of diagnosis and at previous visits until HCV RNA was negative: rapid tests for detection of HCV antibodies [OraQuick (Bethlehem, Pennsylvania, USA), Meridian Bioscience (Paris, France) and Toyo, Neprhotek (Boulogne-Billancourt, France)], HCV viral load tests [AmpliPrep/COBAS TaqMan HCV Test, Roche (Basel, Switzerland) called ‘Cobas HCV’; Xpert HCV Viral Load, Cepheid (Maurens-Scopont, France) called ‘Xpert HCV’] and HCV antigen immunoassay (EIA ARCHITECT HCV Ag; Abbott). We evaluated the sensitivity of each test, including serum ALT, for the diagnosis of AHCV.
Baseline characteristics were compared between HCV-uninfected participants and those diagnosed with AHCV during the trial, using Wilcoxon test for continuous variables and Chi-square test or Fisher's exact test for quantitative variables. McNemar's test for paired proportions was used to compare rates of participants with positive HCV RNA (using Cobas HCV) and positive alanine aminotransferase at the prior visit before diagnosis. All analyses were conducted with the use of SAS software, version 9.3 (SAS Institute, Cary, North Carolina, USA); P values and 95% CIs are two-sided.
From 1 February 2012 to 30 June 2016, 429 participants were enrolled in the ANRS IPERGAY PrEP trial, and were followed up for a median duration of 2.1 [interquartile range (IQR), 1.5–2.8] years. During the study period on the basis of the first positive HCV serology testing, 14 participants [median age: 30 (IQR, 27–35) years, all men] were diagnosed with AHCV infection, including one coinfected with HIV. One case was diagnosed at entry in the study (first visit after screening) and it was therefore considered as a prevalent case (no. 7, Supplementary Table S1, http://links.lww.com/QAD/B535). For this participant, serum testing at the previous visit (screening) was positive for HCV RNA (and negative for HCV serology). The 13 other cases were diagnosed during follow-up (incident cases), with a total of 932 person-years. The HCV incidence was 13/932 = 1.40 per 100 person-years (95% CI: 0.74–2.39). No case of HCV reinfection was observed during the study. Genomic analysis identified HCV genotype 1a in six patients (43%), 3a in one (7%) and 4d in the other seven (50%).
Behavioral characteristics of all participants at inclusion in the IPERGAY trial are listed in Table 1. Patients diagnosed with AHCV during follow-up reported a significantly higher number of sexual partners over the past 2 months [median number of 17 (IQR, 8–30) versus eight (IQR, 5–16), P = 0.03] and a higher number of sexual acts over the past 4 weeks [median number of 18 (IQR, 12–20) versus 10 (IQR, 5–16), P = 0.02] compared with uninfected patients at baseline. Oral or snorting drug use during the last sexual act (defined as ‘chemsex’) was also statistically more frequent in HCV-infected patients (57 versus 11% in HCV-uninfected individuals, P = 0.0001), especially the consumption of gamma-hydroxybutyric acid or gamma-butyrolactone.
To determine the sensitivity of the different diagnostic tests for AHCV, stored sera were available in all 14 cases at diagnosis. Compared with the EIA ARCHITECT HCV Ab, rapid tests for detection of HCV antibodies had a lower sensitivity: 93% (95% CI: 66–99) for the OraQuick assay, and 79% (95% CI: 49–95) for the Toyo assay (Table 2). At diagnosis, the EIA HCV Antigen and viral load tests (Cobas HCV, Xpert HCV) had a sensitivity of 100%. All patients had significantly elevated ALT (median 451 IU/l; IQR, 103–597).
We then analyzed available stored sera for 13 of the 14 cases on previous visits until HCV RNA was negative (Supplementary Table S1, http://links.lww.com/QAD/B535). Median time between AHCV diagnosis (first time with a positive EIA ARCHITECT HCV Ab) and the prior visit with a negative HCV RNA was 3.5 (IQR, 2.3–4.1) months. One participant coinfected with HIV (no. 14, Supplementary Table S1, http://links.lww.com/QAD/B535) had only one previous sample available 6 months before.
The previous closest visit before AHCV diagnosis (with a negative EIA ARCHITECT HCV Ab) occurred within a median of 1.8 (IQR, 1.3–2.1) months earlier. At that time, none of the participants had positive rapid tests for the detection of HCV antibodies (Table 2). Among the 13 participants tested for HCV RNA using Cobas HCV, 11/13 (85%) were positive. The two participants with negative results (nos. 6 and 9, Supplementary Table S1, http://links.lww.com/QAD/B535), were tested positive simultaneously for both EIA HCV antibody and RNA testing at the visit of diagnosis, 8 and 10 weeks later, respectively. At the prior visit before AHCV diagnosis, the Xpert HCV test, the Cobas HCV test and the EIA HCV Ag had a sensitivity of 100, 85 and 89%, respectively, for the diagnosis of HCV infection. Only 25% (3/12) of the cases had elevated ALT (median 291 IU/l; IQR, 83–381) at the visit preceding AHCV diagnosis. Overall, at that time, among 12 participants who were tested with both Cobas HCV and ALT, seven had a detectable HCV RNA and no increased serum ALT increase (P = 0.008, McNemar's test).
In this study, we report a high incidence of AHCV infection in HIV-negative MSM on PrEP in the ANRS IPERGAY trial, with an estimated incidence rate of 1.4 cases for 100 person-years follow-up (PYFU). This is consistent with other reports which showed that the spread of HCV infection observed since 2000 is not limited to HIV-positive MSM, but also affects HIV-negative MSM especially those with high-risk sexual practices . Another recent French study reported an overall HCV incidence rate among HIV-negative PrEP users of 1.2 per 100 PYFU, which is similar to the rate observed in the ANRS IPERGAY trial and in HIV-positive MSM [2,9].
Genomic analysis identified mainly HCV genotypes 1a (43%) and 4d (50%), which are the two main genotypes identified among HIV-positive MSM in Europe . The 14 individuals of our study population have been included in a larger cohort of HIV-positive and negative MSM in Paris to perform a cluster analysis of HCV strains. Ultradeep sequencing techniques showed a high clustering rate between HIV-positive and HIV-negative MSM communities. In particular, 8/13 (61.5%) individuals diagnosed with HCV in our study belonged to shared transmission chains with HIV-positive individuals in Paris, France . The high incidence of HCV infection observed in PrEP users is likely to be due to the overlap between sexual networks of HIV-positive and HIV-negative MSM, which might facilitate introduction of HCV in HIV-negative MSM on PrEP and results in expanding the epidemic [10,22].
Previous studies in MSM have identified several high-risk sexual practices associated with HCV infection, such as unprotected anal intercourse, fisting, group sex and/or use of nasal recreational drugs during sex [8,20,23]. Our study confirm these data among MSM on PrEP, as those diagnosed with AHCV infection during follow-up reported a significantly higher number of partners and sexual acts, and more frequent use of chemsex. Screening of such practices at enrollment in PrEP programs appears crucial to identify the most at-risk individuals, and to improve HCV screening strategies, prevention interventions and immediate treatment. In HIV-positive MSM, the use of a risk-score based on self-reported known risk factors (MOSAIC questionnaire) to assist HCV testing has proven useful to identify people at risk for AHCV infection . Considering the high HCV incidence in this population, such tools should be also implemented in persons taking PrEP for HIV, and we further plan to validate this MOSAIC questionnaire in ongoing prospective cohort of MSM on PrEP, to identify the most at-risk individuals.
Current guidelines recommend routine screening for HCV infection in HIV-negative MSM on PrEP at least once a year [11,15], with more frequent testing for high-risk individuals (every 3 months) . However, to date, only HCV serology is currently recommended for HCV screening, although the positivity of antibodies can be delayed up to 3 months leading to missed AHCV infections . In our study, at the time of the first positive EIA ARCHITECT HCV Ab test, rapid tests for detection of HCV antibodies had a lower sensitivity, especially for the Toyo assay (79%). The use of these rapid tests is currently spreading in places of POCT for STI screening, but they have not been evaluated in the context of seroconversion during acute infection. Our results suggest that their use cannot be recommended in case of AHCV suspicion. In contrast, we found that the HCV Ag immunoassay and plasma HCV RNA tests were positive in most cases within a median of 2 months before the detection of antibodies, when participants were still asymptomatic and only 25% had elevated ALT. When laboratory testing is available, the use of one of these direct tests may allow shortening the delay of AHCV diagnosis in comparison with an anti-HCV antibody-based screening. HCV RNA regular screening is much more and may not be cost-effective if patients are regularly tested for ALT and HCVB antibodies (every 3 months as recommended since 2018 in France). However, our study demonstrates that HCV antigen has excellent diagnostic performances for the early diagnosis of AHCV. This test is much less expensive than HCV RNA and may be a good alternative for HCV screening in patients with less frequent testing. These findings are important since an early diagnosis of HCV infection is crucial to prevent the ongoing epidemic. Once diagnosed with AHCV infection, people can now benefit from specific anti-HCV cure therapy. Rapid introduction of anti-HCV therapy following diagnosis is expected to reduce HCV incidence among high-risk MSM on PrEP as it has been observed in HIV-positive individuals [25,26].
Our study presents some limitations. First, it was not prospectively designed to analyze risk factors; missing data on behavioral practices (which were self-reported) limit the results’ analysis, even though our data fit what had already been described in previous reports. In particular, the information about using and/or sharing injecting equipment was not systematically recorded. After checking, two individuals among the 14 cases specifically reported injecting drugs over the weeks preceding AHCV diagnosis. The information was not available for the others, and we cannot exclude the fact that they used intravenous drugs which may have contributed to AHCV acquisition. However, the information collected regarding sexual practices were really precise and vary significantly between the two groups, underlining the fact that high-risk sexual behaviors are a major factor driving the HCV epidemic. Second, we only assessed risk behavior at inclusion, whereas sexual practices and/or drug consumption may change following PrEP initiation. Evaluating sexual behavior and drug use over the past few weeks is probably a more accurate method to identify the more susceptible participants to acquire HCV infection. Third, our study lacks statistical power due to the small number of participants and missing samples of stored sera for some cases, which did not allow us to evaluate dynamic of positivity of the different tests. Third, rapid tests for the detection of HCV antibodies were performed on sera and not on whole blood as recommended; the diagnostic performances of the OraQuick and Toyo assays could therefore be overestimated as it has already been demonstrated for HIV-antibodies rapid tests . Finally, we excluded patients with prior HCV infection and could not assess performance of tests for the diagnostic of reinfection.
Overall, our study identifies HIV-negative MSM on PrEP with a high number of partners and/or sexual acts and a frequent use of chemsex as the most at-risk individuals for AHCV infection. We suggest that these high-risk individuals should be screened for HCV infection with the use of direct tests (HCV Ag or plasma HCV RNA). This could help to shorten the diagnosis delay, and initiate immediate anti-HCV therapy to prevent ongoing transmission.
Author contributions: J.G., J.M.M., M.L.C. and C.D. led the study and wrote the article. N.M. and N.G. performed the HCV tests. I.C. and B.G. analyzed the data. L.C., C.T., J.C., F.R., E.C. and J.M.M. conducted the study at their sites. All the authors critically reviewed and approved the article.
Conflicts of interest
There are no conflicts of interest.
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