Quantification of Female Chimeric Cells in the Tonsils of Male Children and Their Determinants.

The factors influencing mother-to-child cell trafficking and persistence over children's lives have yet to be established. The quantification of maternal microchimerism was previously reported through HLA-based approaches, which introduced bias regarding the tolerogenic environment. We aimed to identify cells of maternal origin irrespective of the HLA repertoire and to ascertain the determinants of microchimeric cells. This case-control study enrolled 40 male infants attending pediatric surgery from January 2022 to October 2022. Female cells were quantified in infants' tonsil tissue by using cytogenetic fluorescent in situ hybridization (FISH) coupled with optimized automated microscopy. Out of the 40 infants, half (47.4%) had been breastfed for more than one month, a quarter for less a month, and 10 children (26.3%) were never breastfed. XX cells were observed in male tonsils in two-thirds of participants at a median density of 5 cells per 100,000 cells. In univariate analyses, child age was negatively associated with a high female cell density. In exploratory multivariate analyses, previous breastfeeding is a likely determinant of the persistence of these cells in the host, as well as the rank among siblings. Part of the benefit of breastmilk for child health may therefore be driven by breastfeeding-related microchimerism.

"Maintaining HIV and HCV prevention and care for people who inject drugs despite COVID-19 in Hai Phong, Vietnam".

BACKGROUND: After the emergence of COVID-19, a one-month strict lockdown was imposed in April 2020 in Vietnam, followed by lighter social distancing restrictions over the year. We investigated whether those measures affected people who inject drugs (PWID) in terms of risk behaviors for HIV and HCV and access to prevention and care in the city of Haiphong, a historic hotspot for HIV and drug use. METHODOLOGY: We carried out a 'before-after' study from 2019 to 2020 using respondent-driven sampling method to enroll PWID. They were interviewed on their socioeconomic situation, drug use and sexual behaviors, relations to care services and tested for drugs and methadone in the urine, for HIV, HCV, and HIV plasma viral load when HIV-positive. Changes following the restrictions were assessed by comparing 'before' to 'after' data. RESULTS: 780 PWID were enrolled. Mean age was 44 years; 94% were male. All were actively injecting heroin 'before', versus 56% 'after'. Among those, frequency of consumption decreased from 24 to 17 days per month. No changes were observed in the frequency and practices of methamphetamine smoking. The proportion of PWID on MMT increased from 68.7% to 75.3%, and that of PWID engaging in risky behaviors related to drug injection decreased from 6.0% to 1.5%. No HIV seroconversions were observed; HCV incidence was 2.6/100 person-years (95% CI [0.7-6.7]). 9% of PWID reported a monthly income of less than 130USD 'before' versus 53% 'after'. CONCLUSION: The case of Hai Phong shows that it is possible, during times of COVID-19 pandemic, to maintain access to harm reduction and care and to prevent HIV and HCV transmission among PWID in a resource-limited setting where severe social distancing restrictions are implemented. Further research is needed to assess the consequences of long-term economic difficulties and the impact of actual spread of SARS-Cov2 that has since emerged in Haiphong.

Long-Term Persistence of Mitochondrial DNA Instability among HCV-Cured People Who Inject Drugs.

People who inject drugs (PWID) are a population exposed to many genotoxicants and with a high prevalence of HCV infection. Direct-acting antiviral (DAA) regimens are now widely used to treat chronic HCV infection. Although side effects to treatment are currently rare, the long-term effects such as suspicions of de novo hepatocellular carcinoma (HCC) occurrence or HCC recurrence and cardiac defects are still up for debate. Given the structure of DAAs, the molecules have a potential mitochondrial DNA (mtDNA) genotoxicity. We have previously reported acute mtDNA toxicity of three DAA regimens among PWID with a strong impact on the rate of mtDNA deletion, less on the quantity of mtDNA copy per cell at sustained viral response at 12 weeks (SVR12). Herein, we report the mtDNA parameters nine months after drug discontinuation. We observed that the percentage of the deleted mtDNA genome increased over time. No exposure to any other genotoxicants during this period was associated with a high deletion percentage, suggesting that the replicative advantage of the deleted molecules outweighed their elimination processes. Such observation calls for longer-term follow-up and may contribute to the molecular basis of subclinical side effects of DAA treatments.

Recurrent Severe Subclinical Mastitis and the Risk of HIV Transmission Through Breastfeeding.

Subclinical mastitis (SCM) is an important risk factor of postnatal HIV-1 transmission that is still poorly understood. A longitudinal sub-study of the ANRS12174 trial including 270 breastfeeding mothers in Lusaka, Zambia measured sodium (Na+) and potassium (K+) in archived paired breast milk samples collected at week 14, 26 and 38 postpartum to determine cumulative incidence of SCM and the effects of recurrent severe SCM on HIV-1 shedding in breast milk. A nested retrospective cohort study including 112 mothers was also done to determine longitudinal effects of SCM on four pro-inflammatory cytokines; IL6, IL8, IP10 and RANTES. The cumulative incidence for any SCM (Na + /K + ratio > 0.6) and severe SCM (Na + /K + ratio > 1) were 58.6% (95%CI: 52.7 - 64.5) and 27.8% (95%CI: 22.5 - 33.1), respectively. In majority of affected mothers (51.4%) severe SCM was recurrent. Both breasts were involved in 11.1%, 33.3% and 70% of the mothers with a single episode, 2 and 3 episodes respectively. In affected breasts, an episode of severe SCM resulted in steep upregulation of the four cytokines considered (IL8, IP10, RANTES and IL6) compared to: before and after the episode; contralateral unaffected breasts; and SCM negative control mothers. Recurrent severe SCM significantly increased the odds of shedding cell-free HIV-1 in breast milk (OR: 5.2; 95%CI: 1.7 - 15.6) whereas single episode of severe SCM did not (OR: 1.8; 95%CI: 0.8 - 4.2). A Na+/K+ ratio > 1 indicative of severe SCM is an excellent indicator of breast inflammation characterized by a steep, localized and temporal upregulation of several pro-inflammatory cytokines that favor HIV-1 shedding in mature breast milk and may facilitate postnatal HIV-1 transmission through breastfeeding.

Alarming Tuberculosis Rate Among People Who Inject Drugs in Vietnam.

BACKGROUND: The tuberculosis (TB) epidemic is not homogeneous in the general population but presents high-risk groups. People who inject drugs (PWID) are such a group. However, TB among PWID remains largely undocumented. Our goal was to assess the prevalence of TB and the risk factors associated with TB among PWID in Vietnam. METHODS: We implemented a cross-sectional survey among 2 community-based cohorts of human immunodeficiency virus (HIV)-positive and HIV-negative PWID in Hai Phong. Participants were screened for TB using questions on TB symptoms. Those who reported any symptom were accompanied by peers to the TB clinic for chest x-ray. If the latter was abnormal, a sputum was collected to perform an Xpert MTB/RIF test. RESULTS: A total of 885 PWID were screened for TB. For both cohorts, most PWID were male (>90.0%), with a median age of 42 years. Beside heroin injection, 52.5% of participants reported smoking methamphetamine, and 63.2% were on methadone. Among HIV-positive PWID (N = 451), 90.4% were on antiretroviral therapy and 81.6% had a viral load <1000 copies/mL. Using a complete-case analysis, the estimated TB prevalence was 2.3% (95% confidence interval [CI], 1.0-4.5) and 2.1% (95% CI, 0.8-4.2) among HIV-positive and HIV-negative people, respectively. Living as a couple, arrest over the past 6 months, homelessness, and smoking methamphetamine were independently associated with TB but not HIV infection. CONCLUSIONS: In the context of very large antiretroviral therapy coverage, this extremely high rate of TB among PWID requires urgent actions.

Development and assessment of a community-based screening tool for mental health disorders among people who inject drugs.

INTRODUCTION: The prevalence of mental health disorders among people who use drugs is high and well documented. This hard-to-reach population faces a very low awareness and access to mental health care, especially in developing countries. The objectives of this study were to design and assess a quick screening tool (QST) that community-based organisations (CBO) could routinely apply to a Vietnamese population of people who inject drugs (PWID), in order to refer them appropriately to mental health specialists. METHODS: We devised a tool that included nine questions covering anxiety, depression, suicide risk and psychotic symptomatology. Its use required no specific background and 2 h training. Specificity and sensitivity of the QST were assessed in a population of 418 PWID recruited via respondent driven sampling, using the Mini International Neuropsychiatric Interview questionnaire plus clinical evaluation as a reference standard. Acceptability was assessed using a self-administered anonymous questionnaire submitted to all CBO members who used the QST. RESULTS: CBO members considered the QST easy to use, relevant and helpful to deal with mental health issues. Area under the curve for detection of any symptom using the QST was 0.770. The maximum sensitivity and specificity were reached with a cut-off of 2 [sensitivity was 71.1% (95% confidence interval 62.4, 78.8), specificity was 75.9% (70.5, 80.7)]. DISCUSSION AND CONCLUSIONS: The QST appeared to be both efficient and well accepted. Given the burden of mental health problems among hard-to-reach PWID in developing countries, community-based screenings such as this one could be a particularly appropriate response.

Magnetic field-enhanced agglutination as a readout for rapid serologic assays with human plasma.

Recent virus outbreaks have revealed a critical need for large scale serological assays. However, many available tests either require a cumbersome, costly apparatus or lack the availability of full automation. In order to address these limitations, we describe a homogeneous assay for antibody detection via measurement of superparamagnetic particles agglutination. Application of a magnetic field permits to overcome the limitations governed by Brownian translational diffusion in conventional assays and results in an important acceleration of the aggregation process as well as an improvement of the limit of detection. Furthermore, the use of protein-concentrated fluid such as 5 times-diluted human plasma does not impair the performances of the method. Screening of human plasma samples shows a strict discrimination between seropositive and seronegative samples in an assay duration as short as 14 s. The sensitivity of this method, combined with its quickness and simplicity, makes it a promising diagnostic tool.

Alarmin S100A9 restricts retroviral infection by limiting reverse transcription in human dendritic cells.

Dendritic cells (DC) subsets, like Langerhans cells (LC), are immune cells involved in pathogen sensing. They express specific antimicrobial cellular factors that are able to restrict infection and limit further pathogen transmission. Here, we identify the alarmin S100A9 as a novel intracellular antiretroviral factor expressed in human monocyte-derived and skin-derived LC. The intracellular expression of S100A9 is decreased upon LC maturation and inversely correlates with enhanced susceptibility to HIV-1 infection of LC. Furthermore, silencing of S100A9 in primary human LC relieves HIV-1 restriction while ectopic expression of S100A9 in various cell lines promotes intrinsic resistance to both HIV-1 and MLV infection by acting on reverse transcription. Mechanistically, the intracellular expression of S100A9 alters viral capsid uncoating and reverse transcription. S100A9 also shows potent inhibitory effect against HIV-1 and MMLV reverse transcriptase (RTase) activity in vitro in a divalent cation-dependent manner. Our findings uncover an unexpected intracellular function of the human alarmin S100A9 in regulating antiretroviral immunity in Langerhans cells.

Mitochondrial DNA Instability Is Common in HIV-Exposed Uninfected Newborns.

Worldwide, one million HIV-exposed uninfected (HEU) children are born yearly, and chronic health impairments have been reported in these children. Mitochondrial DNA (mtDNA) instability and altered mtDNA content have been evidenced in these children, but an exhaustive characterization of altered mitochondrial genomes has never been reported. We applied deep mtDNA sequencing coupled to the deletion identification algorithm eKLIPse to the blood of HEU neonates (n = 32), which was compared with healthy controls (n = 15). Dried blood spots (DBS) from African HEU children were collected seven days after birth between November 2009 and May 2012. DBS from French healthy controls were collected at birth (or <3 days of life) in 2012 and in 2019. In contrast to the absence of mtDNA instability observed at the nucleotide level, we identified significant amounts of heteroplasmic mtDNA deletions in 75% of HEU children and in none of controls. The heteroplasmy rate of the 62 mtDNA deletions identified varied from 0.01% to up to 50%, the highest rates being broadly compatible with bioenergetic defect and clinical expression. mtDNA integrity is commonly affected in HEU neonates. The nature of the deletions suggests a mechanism related to aging or tumor-associated mtDNA instability. This child population may be at risk of additional mtDNA genetic alterations considering that they will be exposed to other mitotoxic drugs including antiretroviral or anti-tuberculosis treatment.

The methamphetamine epidemic among persons who inject heroin in Hai Phong, Vietnam.

AIMS: To describe the current methamphetamine (MA) use epidemic among persons who inject heroin (PWID) in Hai Phong, Vietnam, and consider possibilities for mitigating adverse effects of methamphetamine use. METHODS: This study conducted surveys of PWID in 2016, 2017, and 2018 (N = 1383, 1451, and 1445, respectively). Trained interviewers administered structured interviews covering drug use histories, current drug use, and related risk behaviors. The study used urinalysis to confirm current drug use, and conducted HIV and HCV testing. RESULTS: Participants were predominantly male (95%), mean age of 40, and all reported injecting heroin. Respondents' reports of initiating MA use were rare up through early 2000s but increased exponentially through the mid-2010s. MA use was predominantly "smoking," heating the drug and inhaling the vapor using a pipe; injecting MA was rare. Current (past 30 day) MA use appears to have plateaued in 2016-2018 with 53-58% of participants reporting no use in the last 30 days, 37-41% reporting low to moderate use (1 to 19 days in last 30 days), and 5-7% reporting very frequent use (20 or more days in last 30 days). This plateau reflects a rough balance between new users and individuals ceasing use. CONCLUSIONS: MA use has become a substantial public health problem among PWID in Hai Phong. Initiation into MA use rose exponentially from 2005 to about 2015. Use of MA will likely continue for a substantial number of PWID. Currently, no medication is approved for treating MA disorders in Vietnam. Current psychosocial treatment requires highly trained counselors and months of treatment, so that psychosocial treatment for all PWID with MA disorders is likely beyond the resources available in a middle-income country such as Vietnam. Harm reduction programs implemented by community-based organization staff may provide a way to rapidly address aspects of the current MA epidemic. Such programs could emphasize social support for reducing use where possible and for avoiding escalation of use among persons continuing to use.

Growth, clinical and neurodevelopmental outcomes at school age are similar for children who received 1-year lamivudine or lopinavir/ritonavir HIV prophylaxis in early life.

In the ANRS 12174 trial, HIV-exposed uninfected African neonates who received lopinavir-ritonavir (LPV/r) prophylaxis for 1 year exhibited slower growth from birth to week 50 compared with those receiving lamivudine (3TC). We assessed whether this difference in growth persisted over time, and was accompanied by differences in neuropsychological and clinical outcomes. Between February 2017 and February 2018, we conducted a cross-sectional clinical evaluation among former trial participants who completed the 50-week follow-up and who were not HIV-infected. In addition to clinical examination, neuropsychological outcomes were assessed using the tests Kaufman-ABCII, Test of Variables of Attention, Movement Assessment Battery for Children and the Strengths and Difficulties questionnaire, parent version. Of 1101 eligible children, aged 5-7 years, 553 could be traced and analysed (274 in the LPV/r and 279 in the 3TC groups). Growth, clinical and neuropsychological outcomes did not differ between treatment groups. At school age, children exposed to LPV/r and 3TC at birth for 1 year had comparable growth and neuropsychological outcomes without evidence of long-term side-effects of LPV/r. It provides reassuring data on clinical outcomes for all HIV-infected children treated with this antiretroviral drug in early life.

Clinical and Biological Factors Associated With Early Epstein-Barr Virus Infection in Human Immunodeficiency Virus-Exposed Uninfected Infants in Eastern Uganda.

BACKGROUND: Immune control of Epstein-Barr virus (EBV) infection is impaired in individuals with HIV. We explored maternal factors associated with EBV acquisition in HIV-exposed uninfected (HEU) infants and the relationship between EBV infection and serious adverse events (SAEs) during the first year of life. METHODS: 201 HEU infants from Uganda enrolled in the ANRS 12174 trial were tested for antiviral capsid antigen (anti-VCA) antibodies at week 50. Date of infection was estimated by testing EBV DNA at weeks 1, 6, 14, 26, 38, and 50 postpartum on dried blood spots. RESULTS: Eighty-seven (43%) infants tested positive for anti-VCA IgG at week 50. Among the 59 infants positive for EBV DNA, 25% were infected within the first 26 weeks. Almost half (12%) were infected before week 14. Shedding of EBV in breast milk was associated with EBV DNA in maternal plasma (P = .009), HIV RNA detection (P = .039), and lower CD4 count (P = .001) and correlated with plasma EBV DNA levels (P = .002). EBV infant infection at week 50 was associated with shedding of EBV in breast milk (P = .009) and young maternal age (P = .029). Occurrence of a clinical SAE, including malaria and pneumonia, was associated with higher levels of EBV DNA in infants (P = .010). CONCLUSIONS: By assessing EBV infection in HEU infants we observed that infection during the first year is determined by HIV and EBV maternal factors and that EBV DNA levels were higher among infants with clinical SAEs. CLINICAL TRIALS REGISTRATION: NCT00640263.

The CD133+ Stem/Progenitor-Like Cell Subset Is Increased in Human Milk and Peripheral Blood of HIV-Positive Women.

Human milk is a significant source of different CD133+ and/or CD34+ stem/progenitor-like cell subsets in healthy women but their cell distribution and percentages in this compartment of HIV-positive women have not been explored. To date, a decrease of CD34+ hematopoietic stem and progenitor cell frequencies in peripheral blood and bone marrow of HIV-positive patients has been reported. Herein, human milk and peripheral blood samples were collected between day 2-15 post-partum from HIV-positive and HIV-negative women, and cells were stained with stem cell markers and analyzed by flow cytometry. We report that the median percentage of CD45+/highCD34-CD133+ cell subset from milk and blood was significantly higher in HIV-positive than in HIV-negative women. The percentage of CD45dimCD34-CD133+ cell subset from blood was significantly higher in HIV-positive than HIV-negative women. Moreover, percentages of CD45dimCD34+, CD45dimCD34+CD133-, and CD45+highCD34+CD133- cell subsets from blood were significantly lower in HIV-positive than HIV-negative women. The CD133+ stem/progenitor-like cell subsets are increased in early human milk and blood of HIV-positive women and are differentially distributed to CD34+ cell subset frequencies which are decreased in blood.

Daily heroin injection and psychiatric disorders: A cross-sectional survey among People Who Inject Drugs (PWID) in Haiphong, Vietnam.

BACKGROUND: Psychiatric comorbidities are frequent among people who inject drugs, they are associated with a poorer prognosis and need to be addressed. Their interaction with daily heroin injection requires clarification. METHODS: A cross-sectional survey was conducted among PWID recruited in the city of Haiphong, Vietnam, by respondent-driven sampling. The inclusion criteria were age 18 or older and current injection drug use, verified by skin marks and positive urine tests for heroin or methamphetamine. Data on socio-demographic characteristics, drug use, sexual behaviour and access to treatment were collected using face-to-face questionnaires by trained interviewers. PWID were screened by trained psychiatrists for depression, psychotic disorder and suicidality, using the MINI questionnaire. RESULTS: 418 participants were included in the analyses. All were injected heroin users, 21 % were diagnosed with a current major depressive disorder, 15 % with a current psychotic disorder and 12 % presented a suicide risk. In the bivariate analyses, regular meth use, cannabis use and ketamine use were positively associated with presenting at least one psychiatric condition while daily heroin injection and being currently treated with methadone were negatively associated. In the multivariate model, poly-substance use was positively associated with depression (methamphetamine and drinking in addition to heroin) and psychotic disorder (methamphetamine and/or hazardous drinking in addition to heroin) while daily heroin injection and current methadone treatment were negatively and independently associated with depression and psychotic syndrome. CONCLUSIONS: Our survey confirms the burden of methamphetamine use and the protective effect of methadone but also a possible protective effect of daily heroin injection.

Large Stem/Progenitor-Like Cell Subsets can Also be Identified in the CD45- and CD45+/High Populations in Early Human Milk.

BACKGROUND: Stem/progenitor cells have been identified in human milk. However, characterization and percentages of cell subsets in human milk using hematopoietic stem and progenitor cell markers according to the differential expression of CD45, i.e., as CD45dim/+ (mainly hematopoietic stem/progenitor cells) and CD45- (mainly non-hematopoietic stem/progenitor cells), have not been assessed to date. RESEARCH AIM: To characterize stem/progenitor-like cell phenotypes in human milk and to report the percentages of these cells at two different lactation stages compared to peripheral blood. METHODS: Human milk samples paired with peripheral blood samples (N = 10) were analyzed by flow cytometry using CD45, CD34, CD133, CD38, and lineage-negative markers. The percentage of cell subsets was analyzed in colostrum (Day 3 postpartum) and transitional milk (Day 5/6 postpartum) and compared with the peripheral blood counterpart. RESULTS: The percentage of CD45-CD34+ cells was predominant in both colostrum and transitional milk. The percentage of CD45+/highCD133+ cells was high in colostrum while the percentage of CD45-CD133+ cells was high in transitional milk. Furthermore, the median percentages of the CD45-CD34+, CD45-CD133+, and CD45dimCD133+ cell subsets were higher in colostrum than its peripheral blood counterpart (0.11% vs. 0.002%; 0.17% vs. 0.0005%; 0.09% vs. 0.05%, p = .04, respectively); also CD45-CD34-CD133+ and CD45dimCD34-CD133+ cell subsets were higher in colostrum than peripheral blood (1.32% vs. 0.0% and 2.4% vs. 0.06%, p = .04), respectively). CONCLUSION: Early human milk is an abundant reservoir of hematopoietic stem/progenitor-like cells in the CD45+/high population and non-hematopoietic stem/progenitor-like cells in the CD45- population.

Increased Methamphetamine Use among Persons Who Inject Drugs in Hai Phong, Vietnam, and the Association with Injection and Sexual Risk Behaviors.

Methamphetamine use has increased significantly in Southeast Asia in the last 5-10 years, but there is little research on risk behaviors associated with the increase in this region. We evaluate injection and sexual risk behaviors associated with current methamphetamine use among heterosexual persons who inject drugs (PWID) in Hai Phong, Vietnam. We recruited 1336 PWID and assessed associations between methamphetamine use, injection and sexual risk behaviors, stratified by HIV serostatus. There were several statistically significant associations between methamphetamine use and sexual risk behaviors among HIV-positive PWID, but we did not find any associations between methamphetamine use and injection risk behaviors. Methamphetamine may increase the likelihood of PWID transmitting HIV to non-injecting primary sexual partners, as it can reduce ART adherence and can lead to weakening of the immune system and increased HIV viral loads. All participants were injecting heroin, and methamphetamine use may interfere with opiate use treatment. Public health efforts should focus on the large increase in methamphetamine use and the associated sexual risk behaviors. Emphasis on sexual risk behavior, particularly among those who are HIV-positive, in conjunction with continued monitoring of ART adherence and HIV viral loads, is critical.

Human papillomavirus (HPV) shedding in breast milk from African women living with HIV.

BACKGROUND: Little is known about human papillomavirus (HPV) shedding in human breast milk. OBJECTIVE: To investigate HPV shedding in mature breast milk specimens collected from breastfeeding African women living with HIV-1 and not receiving antiretroviral treatment. DESIGN: 62 African women enrolled in the ANRS 12174 trial participated in this study. 79 lactoserum specimens obtained from right and/or left breasts from 42 Zambian women as well as lactosera and cell pellets from 40 milk samples collected from right and left breasts among 20 Ugandan women were tested for HPV using the INNO-LiPA HPV Genotyping Extra II assay. RESULTS: HPV DNA was detected in 9 (11.4%) lactoserum specimens collected from 8 (19.0%) Zambian women. Fourteen (17.5%) samples from 5 (25%) Ugandan women were positive for HPV detection. Differences in HPV type identification between the two breasts as well as between lactoserum and cell pellet were oberved. Overall, 13 (21.0%) of the 62 women included in this study had detectable HPV DNA in their breast milk, representing 11 HPV types, including high-risk, probable high-risk and low-risk types. CONCLUSION: This study confirms that HPV can be frequently detected in breast milk in HIV-infected women. Further studies are needed to understand the way by which maternal milk can shed HPV.

Using dual capture/recapture studies to estimate the population size of persons who inject drugs (PWID) in the city of Hai Phong, Vietnam.

BACKGROUND: Good estimates of key population sizes are critical for appropriating resources to prevent HIV infection. We conducted two capture/recapture studies to estimate the number of PWID currently in Hai Phong, Vietnam. METHODS: A 2014 respondent-driven sampling (RDS) survey served as one capture, and distribution of cigarette lighters at drug use "hotspots" in 2016 served as another "capture." A 2016 survey using RDS, conducted 1 week after lighter distribution, served as "recapture" for both captures. Recaptured participants in the two surveys were identified with a computerized fingerprint reader. Recaptured participants from the lighter distribution were asked to show their lighters. RESULTS: 1385 participants were included in the "recapture" survey. They were 94% male and had a median age of 39. All (100%) injected heroin, and HIV prevalence was 30%. 144 of the 603 participants in the 2014 survey and 152 of the 600 PWID who had received lighters were "recaptured" in the 2016 survey. After adjusting for police suppression of drug use hotspots and conducting sensitivity analyses, our best estimate of the population size from the lighter recapture was 4617 (95% CI: 4090-5143), and our best estimate from the 2014 survey recapture was 5220 (95% CI: 4568-5872). A combined best estimate of the PWID population in Hai Phong is 5000, range 4000-6000. CONCLUSIONS: The capture/recapture studies produced consistent estimates. Adding a lighter/token distribution to planned RDS surveys may provide an inexpensive method for estimating PWID population size. Analyses of the estimates should include contextual information about the local drug scene.