Study population and sampling
A total of 31 participants established on ART provided at least one paired semen and blood sample over the 12 months of the study. Their baseline characteristics are summarised in Table 1. At study entry, participants had received ART for a median of 7.5 years (IQR 1.9–13.1). Most were receiving coformulated tenofovir disoproxil fumarate/lamivudine/efavirenz (TDF/3TC/EFV). A total of 72 paired samples were collected, with a median of 2 paired samples per participant (range 1–5), including 30 samples at study entry (baseline), and 10, 12, 9 and 11 samples at 1, 3, 6 and 12 months. Eleven participants donated only one set of paired samples. Overall, based on the date of the last paired samples, the median duration of follow-up was 10.2 months (IQR 4.0–14.2). All except 6 participants (one with MGS and 5 without) had received PZQ in the 12 months prior to recruitment. None showed symptoms suggestive of sexually transmitted infections (STIs) and no samples were collected to investigate asymptomatic STIs.
Details of parasitology testing are summarised in Table 2. At baseline, 8 participants tested MGS positive by semen microscopy, 4 tested positive only by real-time PCR of seminal sediment, and 20 tested negative by all tests. Among the 20 participants with a negative baseline test, 3 had a positive test during follow-up (1 by real-time PCR only; 1 by PCR and POC-CCA test and 1 by urine filtration, PCR and POC-CCA test) yielding a total of 15 participants who were classed as MGS positive (A01–A15), whereas 16 were classed as MGS negative (A16–A31). There were no significant differences when comparing the baseline characteristics of the two groups (Table 1). During the study period, 42 paired samples were provided by the MGS positive participants while 30 samples were provided by MGS negative participants.
HIV-1 RNA testing
At baseline, 5/30 (16.7%) participants showed quantifiable plasma HIV-1 RNA (\(\ge\) 40 copies/mL) including 3/14 (21.4%) in the MGS-positive group (A09, A11, A15) and 2/16 (12.5%) in the MGS-negative group (A21, A30) (Table 3). HIV-1 RNA levels ranged from 41 to 56,000 copies/mL During follow-up, 1/5 participants (A09) achieved suppression < 40 copies/mL at 6 months, while continuing to show detectable HIV-1 RNA below the assay LLOQ of 40 copies/mL (estimated 22–39 copies/mL); 1/5 (A11) showed persistent viraemia between 816 and 277 copies/mL at 6 and 12 months; and 3/5 had no follow-up samples collected. A further 4 participants showed detectable plasma HIV-1 RNA below the assay LLOQ of 40 copies/mL (estimated 22–39 copies/mL) at baseline, including 2/14 (14.3%) in the MGS-positive group (A07, A13) and 2/16 (12.5%) in the MGS-negative group (A16, A23). Among 21 participants with undetectable HIV-1 RNA at baseline, two in the MGS-negative group showed plasma viral load rebound > 40 copies/mL at 1 month (A17; 64 copies/mL) and 3 months (A20; 107,000 copies/mL) respectively, followed by resuppression < 40 copies/mL) but detectable HIV-1 RNA at 12 months (Table 3).
Among participants with baseline samples, 3/30 (10.0%) showed detectable HIV-1 RNA in seminal fluid (Table 3). In the MGS-positive group, 2/14 (14.3%) participants (A03, A04) showed detectable HIV-1 RNA (estimated levels between 55 and 400 copies/mL) in seminal fluid, while plasma HIV-1 RNA was undetectable (< 22 copies/mL). One of the two had no follow-up samples. The other showed undetectable HIV-1 RNA in both plasma and seminal fluid at months 1, 3 and 12. In the MGS-negative group, 1/16 (6.3%) participants (A21) showed seminal HIV-1 RNA levels of 4840 copies/mL, but in this case plasma HIV-1 RNA levels were also high at 26,900 copies/mL. A total of 20 participants had at least one follow-up sample (Table 3). Among these, 2 participants (A05, A20) showed newly detectable HIV-1 RNA in seminal fluid. One participant (A05) in the MGS-positive group had persistent HIV-1 RNA in seminal fluid at months 1, 3 and 6, with levels ranging up to 123 copies/mL, while plasma HIV-1 RNA remained persistently undetectable (< 22 copies/mL). The second participant (A20) was in the MGS-negative group. At 3 months, HIV-1 RNA levels were 528 copies/mL in seminal fluid while plasma HIV-1 RNA had rebounded from undetectable to 107,000 copies/mL. Overall, 7/72 (9.7%) seminal fluid samples had detectable or quantifiable HIV-1 RNA, comprising 3 samples at baseline (2 in the MGS-positive group) and 1, 2 and 1 samples at 1, 3 and 6 months, respectively (3 in the MGS-positive group).
Patterns of HIV-1 RNA detection by MGS status
When comparing the 72 paired plasma and seminal samples, HIV-1 RNA detection (with or without quantification) was concordant in 44/72 (61.1%) samples (Table 4). The remaining 28/72 (38.9%) sample pairs were discordant; most had HIV-1 RNA detected in plasma only. In the MGS-negative group, 2/16 participants had detectable HIV-1 RNA in 2 seminal samples, and in both cases the paired plasma showed high HIV-1 RNA levels. In contrast, in the MGS-positive group, 3/15 participants had detectable HIV-1 RNA in 5 seminal samples, and in all cases the paired plasma showed undetectable HIV-1 RNA. The characteristics of the 3 participants are detailed in the Table 5. They had been established on ART long-term, ranging between 8 and 12 years at study entry. Overall, 4 of the 5 seminal samples with detectable HIV-1 RNA had concomitant Schistosoma test positivity.