Estimating the prevalence of hepatitis C among intravenous drug users in upper middle income countries: A systematic review and meta-analysis

Aim This systematic review and meta-analysis characterizes the prevalence of hepatitis C virus (HCV) infection among intravenous drug users (IDUs) in upper middle-income countries. Methods Five databases were searched from 1990–2016 for studies that took place in countries with a GDP per capita of $7,000 to $13,000 USD. The data extraction was performed based on information regarding prevalence, sample size, age of participants, duration of intravenous drug use (IDU), recruitment location, dates of data collection, study design, sampling scheme, type of tests used in identifying antibody reactivity to HCV, and the use of confirmatory tests. The synthesis was performed with a random effects model. The Cochrane statistical Q-test was used to evaluate the statistical heterogeneity of the results. Results The 33 studies included in the analysis correspond to a sample of seven countries and 23,342 observations. The point prevalence value estimates and confidence intervals of the random effects model were 0.729 and 0.644–0.800, respectively for all seven countries, and were greatest for China (0.633; 0.522–0.732) as compared to Brazil (0.396; 0.249–0.564). Prevalence for Montenegro (0.416; 0.237–0.621) and Malaysia (0.475; 0.177–0.792) appear to be intermediate. Mexico (0.960) and Mauritania (0.973) had only one study with the largest prevalence. A clear association was not observed between age or duration of IDU and prevalence of HCV, but the data from some groups may indicate a possible relationship. The measures of heterogeneity (Q and I2) suggest a high level of heterogeneity in studies conducted at the country level and by groups of countries. Conclusions In this systematic review and meta-analysis, we found that the pooled prevalence of HCV was high (0.729) among a group of seven upper middle income countries. However, there was significant variation in the prevalence of HCV observed in China (0.633) and Brazil (0.396).


Introduction
Infection with the hepatitis C virus (HCV) is a serious public health problem due to its association with diseases like chronic hepatitis, cirrhosis, and hepatocellular carcinoma [1][2][3][4]. Evidence suggests that the prevalence and number of infected patients has decreased in higher income countries [5]. On the other hand, this disease has increased significantly in certain low-income countries in Africa and Asia [6]. Intravenous drug users (IDU) are one of the groups with a higher prevalence of HCV infection. This group has a significantly higher risk of infection compared with non-injection drug users, or individuals who do not use illegal drugs, due to the sharing of contaminated needles [7][8][9][10][11]. This has been documented in a number of systematic reviews that report the prevalence of HCV among IDUs, which have mostly focused on European countries [7,[12][13][14][15][16][17][18][19]. Other reviews have examined specific countries or regions like China, Latin America, Iran, Australia, the Middle East, and North Africa [20][21][22][23][24]. Of the eight reviews that have been published, four were conducted worldwide and the other four represent middle and low-income countries. In some instances, the country-specific prevalence data reported in the worldwide reviews is limited and it is not clear how the authors determined the point or interval estimates [14][15][16]19]. Additionally, it can be difficult to assess how certain estimates of HCV prevalence were determined since the sources reported in webappendices are not always available, especially for the older publications [14].
Also relevant, is the fact that more studies on IDUs have been conducted in high-income countries, which may suggest that high-income countries have a larger proportion of IDUs. An alternative explanation could be that some high-income countries are also likely to have a larger budget to estimate the prevalence of HCV infection among IDUs. Conversely, research regarding the prevalence of HCV infection in middle-middle and low-middle income countries is limited to a few countries [20].
Estimating the national prevalence of HCV infection among high-risk individuals, such as IDUs, in middle-and low-income countries is important to help guide interventions to reduce the burden of disease and its economic consequences. To the best of our knowledge, there are no systematic reviews that have investigated the prevalence of HCV infection among IDUs in upper-middle income countries. Therefore, our research question is what is the prevalence of HCV infection (measured by HCV antibodies) among IDU population in upper-middleincome countries? The purpose of this systematic review is to estimate the prevalence of HCV infection among IDUs in several upper-middle income countries (UMIC) and as a group, by means of meta-analysis techniques; and to provide an analysis of prevalence by age and duration of IDU use.

Methods
An electronic literature search was conducted to identify English and Spanish articles published between 1990 and 2016 that reported the prevalence of HCV among IDUs in upper middle income countries. The following five databases were used to conduct this search: PubMed, SCOPUS, Medic Latina, LILACS and Scielo Citation Index (Thompson Reuters), and duplicate articles were eliminated. We used five Medical Subject Headings (Mesh): "Prevalence", "Hepatitis C", "Hepatitis C Antibodies", "Substance-Related Disorders", and "Substance Abuse and Intravenous". We also used the following keywords: "prevalence", "hepatitis C", "HCV", "drug abuse", "drug users", "prevalencia", "abuso de drogas" and "VHC" (Table 1A). We did not review the articles listed in the reference sections of the manuscripts we identified. Our search strategy included 26 countries classified as upper middle income, with a gross domestic product (GDP) per capita income between $7,000 and $13,000 US dollars (USD) in 2015, as defined by the World Bank using the Atlas method [25].
The main outcome of the search was the prevalence of HCV infection. This was defined as the proportion of positive cases identified from the total number of individuals who were tested with any of the following Anti-HCV assays: ELISA, chemiluminescence, or any other test that detects antibody response to HCV antibody reactivity [26]. HCV cases were identified when the initial anti-HCV antibody immunoassay was positive. This test provides evidence of present or past infection. Additionally, a positive RNA-HCV test was conducted to indicate the presence of a current infection. If the results of both tests (anti-HCV and RNA-HCV) are available, the result of the first test is confirmed with a RNA-HCV test [27]. We defined "injection drug user" (IDU) as an individual who has been injecting any type of drug for more than six months prior to the study interview.
All studies that reported a prevalence of HCV infection and met the inclusion criteria were included in this analysis. Studies were included if they met the following criteria: 1) the study reported the prevalence of HCV in an IDU population, 2) the study was published after January 1990, and 3) the study was conducted in one of the upper middle income countries included in the search. The exclusion criteria were the following: 1) the purpose of the study was to identify individuals with HIV infection and not chronic HCV infection, and 2) the study only had male or only female subjects. Two members of the research team (VGG, LMS) independently read the published titles and abstracts, and selected the manuscripts that were reviewed in full. Differences in selected papers were resolved by consensus. One member of the team (VGG) read all the papers in full and excluded those with duplicated information based on the title and abstract, in agreement with the previously selected criteria.
We followed the STROBE statement guidelines for cross-sectional studies to retrieve and report information on studies included in the meta-analysis. The retrieval of information from observational studies tried to identify characteristics to the acronym PEOS (Population, Exposure, Outcome and Study type). Organization to report materials was made by information on study, methodology, and results. The study information we report includes dates and sites of Table 1. Characteristics of studies included in the meta-analysis that report HCV prevalence data among IDUs in upper middle income countries. Drug injection in the last 6 months, subjects >16 y, and provided informed consent to enter the study.

Study
Anti-HCV by ELISA/ RNA reflex not conducted.
Interviews conducted at usual meeting places. All IDUs who could be found and met inclusion criteria where recruited. Snowballing sampling was used.
WHO multicity questionnaire with 8 sections (including drug use pattern, sharing injection equipment and sexual behavior).
Data specific on type of drugs and route was not reported. Period of drug injection 9.4 (7.5) (range 1-37) y. [52] First project data collection (1994)(1995)(1996)(1997) in (Continued ) data collection, while the methodology includes: eligibility criteria, type of tests for identifying HCV antibodies, use of RNA reflex testing for confirmation, sample size, and the analytical methods used to account for sampling strategy. In terms of results, we include whether studies report the demographic, clinical and social characteristics of participants, as well as information on type of drug use and mode of administration, time injecting drugs, and risks behavior for HCV infection.
The pooled HCV prevalence and its 95% confidence intervals (CIs) were estimated for each country and for all countries combined. The method used to estimate the pooling prevalence is to transform the prevalence to a variable that is not constrained to the 0-1 range and has an approximately normal distribution. The meta-analysis is conducted on the transformed proportions, using the inverse of the variance of the transformed proportion as the study weight, then the pooled transformed proportion and its CI are back transformed [28]. The Cochrane statistical Q-test was used with a level of <0.1 to evaluate the statistical heterogeneity of the results and the I^2 statistic with a range of values of 0 (no heterogeneity) to 100 (significant heterogeneity) [29]. We used the I^2 statistic to assess the level of heterogeneity within country Years of injecting Median ICR (8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19). Age at first injection. Risk behavior factors: sharing injecting devices. [61] September 2009-December 2010. Two survey sites one in each city, but a description of them was not made.
EC were not stated specifically. Consultations were conducted previous to Survey. RDS was used to recruit IDUs.
Anti-HCV Abbot/ RNA was not conducted.
Multiplier method was used to estimate the size of IDU population. Two approaches were used to determine the multiplier.
Sampling used seeds identified and selected to reflect diversity in residence, gender, levels of risk behavior, age and education.
Age distribution of participants % of males. Marital status and education were also collected.  and between countries. We also analyzed the pooled estimates of HCV prevalence according to the participants' age and duration of IDU. The synthesis was performed with a random effects model (REM). The random effects model is considered more appropriate to estimate prevalence because it considers that the variability of the effect size is affected by the heterogeneity between studies and other aspects besides the variability within each study. The graphic representations of the results of the pooling results and their CIs were reported for each country in a Forest plots. The estimations were carried out using the Comprehensive meta-analysis V.3.3 software.

Results
Our search strategy identified 33 manuscripts from 202 potentially relevant records that were located based on the information provided in the title and abstract. A total of 156 documents were excluded because they did not meet the inclusion criteria (reviews, studies that were not conducted in upper middle income countries, did not include IDUs, or the article was not written in English or Spanish). The complete texts of 46 articles were reviewed to select those that would be included in the study. Thirteen articles were excluded due to the following: 1) duplicate information (n = 6), 2) a different study population (n = 4), 3) a focus on chronic HCV or HIV infection cases (n = 2), or 4) the study was conducted prior to 1990 (n = 1) (Fig  1). Table 1 reports the study characteristics of the 33 publications we selected for this metaanalysis, including the prevalence of HCV infection among IDUs in seven upper middle income countries, from 1995 to 2015. Most studies were conducted in China (n = 16) [30][31][32][33][34][35][36][37][38][39][40][41][42][43][44][45], followed by Brazil (n = 9) [46][47][48][49][50][51][52][53][54], Malaysia (n = 2) [55,56], and Montenegro (n = 3) [57][58][59], while only one study was identified for each of the following countries: Mexico, Mauritania, and Bulgaria (n = 3) [60][61][62]. The part of information corresponding to non-injection drug users (NIDUs) that was reported in two studies [48,49], was excluded from the meta-analysis. But the agreeing with IDU was included. Additionally, we excluded the fragment of information regarding the prevalence of IDUs in the "Ajude-Brazil I" project [53] because it was previously reported in another article [50]. The part which was not repeated was included in the pooled analysis.
Most studies (n = 22) specified that they used a cross-sectional design, and even though nine studies did not explicitly state the design, it can be inferred as a cross-sectional survey. One study is described as a case-control design and another study conducted two surveys at different times. For some studies, the authors provide a brief description of the sampling strategy that was used. For example, six studies used respondent driven sampling, three used snow ball sampling, three used convenience sampling, and two applied a random selection of participants. One study used an existing database of serum samples and three studies did not provide details regarding the sampling strategy that was employed. In-person interviews with participants were conducted in 28 studies. Most studies (n = 19) provided information about the location where interviews were conducted and blood samples were collected. The most common recruitment sites were methadone clinics (n = 5), followed by detox centers, drug treatment centers, and study clinics (n = 9). Other recruitment areas, including program facilities or public streets, were described in five studies. Among the 29 studies that reported the proportion of males and females of the study sample 89% were males. Most of the studies included in our analysis report HCV prevalence based on results from the ELISA test (n = 20), followed by the chemiluminescence assays (Anti-HCV third generation) (n = 6). Seven studies do not report the type of assay used for HCV antibody reactivity testing. Only 8 studies report that confirmation with RNA-HCV test was conducted ( Table 1).
The point prevalence estimates and CIs of the random effects model are 0.73 and 0.64-0.80, respectively, for all seven countries, and are highest for China (0.633; CI95%: 0.522-0.732), as compared to Brazil (0.396; CI95%:0.249-0.564), but they are not significantly different (pvalue >0.05) ( Table 2). Mexico (0.96) and Mauritania (0.973) also have a high prevalence, although these estimates are based on only one study per country. Malaysia (0.475) and Montenegro (0.416) have an intermediate prevalence ( Table 2). The I^2 heterogeneity statistics indicate that heterogeneity was high in all countries, with a high associated p-value. The estimated Q statistic and corresponding p-value for the seven countries also shows that the heterogeneity among groups was high ( Table 2).
The HCV prevalence results by country indicate that the measure of heterogeneity (Q statistic) for China is high, with a considerable range among studies (0.371-0.964) (Fig 2). In Brazil, the dispersion measure Q is lower, although in both cases the p-value is <0.001, which suggests that they do not share a common effect size (Fig 3). The estimate for Montenegro (0.416), which was calculated from three studies, is closer to the HCV prevalence in Brazil (0.396) than in China (0.633) and with a lower heterogeneity (Fig 4). The estimated prevalence of HCV in Malaysia (0.475) is similar to the one observed in Montenegro (Fig 5).
Although there is limited information regarding the age of participants in the 33 studies we included in this meta-analysis, Table 3 reports a possible association between age and prevalence of HCV. Of the six studies that indicate the participants' age, over 50% are 40 years or older (SD 27%), with an average HCV prevalence of 70.5% (95%CI: 0.538-0.830). In another group of 17 studies that report mean age and standard deviation, the average age was 32 years (SD 8 years) and the corresponding prevalence is lower (0.536; 95%CI: 0.441-0.628). These findings suggest that the risk of becoming infected with HCV increases with age, which is to be expected. Table 3 also presents our findings regarding the relationship between time of IDU and HCV prevalence. Based on the information reported in nine studies, the average duration of

Discussion
Based on this systematic review of the literature, only two upper middle income countries (China and Brazil) have conducted nine or more studies that report the prevalence of HCV among IDU. Only two other countries have more than one study that reports the prevalence of HCV infection among IDU (Montenegro and Malaysia). The prevalence of HCV among IDU in China (0.633) is higher than in Brazil (0.396). The Montenegro and Malaysia studies have a similar prevalence (0.416) and (0.475), respectively, but a greater number of studies are needed to confirm the prevalence of HCV among IDUs in these countries. The overall HCV prevalence in the seven countries included in this review is high (0.729; 95%CI: 0.644-0.800), and we also found high heterogeneity in all our analyses. The values of the I2 statistics were high (> 95%) and the Q statistic among these countries was also significant. Our results suggest that most of the participants in the studies we reviewed were male, they began to inject drugs in their early 20s, and they continued to inject drugs for a long time. Our analysis also indicates that when IDUs are older and have injected drugs for a longer time, the prevalence of HCV is expected to be higher. Variability within and between groups can be caused by different reasons such as the study location and methods of recruitment, sampling, diagnostic tests and time of data collection. However, we consider that the random effect model is an adequate analysis method to address the heterogeneity among studies.
The studies included in this review represent a more restricted group of countries, as compared to the reports that were included in other systematic reviews on the prevalence of HCV  among IDU. In their review of the literature, Aceijas and Rhodes estimated the prevalence of HCV among IDU in different regions of the world, and found a greater variability [16]. The authors report the crude average of anti-HCV prevalence among IDUs: Latin America 0.08-0.9, Central Asia 0.1-1.0, South and Southeast Asia 0.34-0.9, East Asia and the Pacific 0.05-0.6. Brazil 0.39 to 0.69 [16]. In the particular case of Brazil our study found that REM estimates of prevalence are more similar than the crude prevalence range which is larger (0.05-0.75). Estimates with raw data in countries such as Mexico are based on a single study of patients with HIV and who in turn are IDUs [63]. In our review, we excluded studies that include IDU cases who are HIV positive, because we considered this may generate a bias in the prevalence estimate.
In another systematic review that included 77 countries from different areas of the world, Nelson et al. reported an overall prevalence of HCV among IDU in the range of 0.098-0.97 based on the midpoint reported by the studies [14]. The authors found that 26 of the 77 countries had a prevalence ranging from 0.6-0.8, while 12 countries had a prevalence greater than 0.8. The results of our study are not comparable to the review by Nelson et al., because we restricted our analysis to upper middle income countries, which narrows the range of HCV prevalence. Additionally, the estimates reported by Nelson et al. [14] are not the result of a synthesis model. Although we identified a very low HCV prevalence for a city in Brazil (0.058) and a very high prevalence in Bulgaria (0.973), the overall pooled prevalence estimated by the REM was 0.729 (CI95%: 0.644-0.800). Nonetheless each country or region may have different conditions that could result in a lower, intermediate or high prevalence.
Another systematic review that examined studies conducted in China, reports the prevalence of HCV among IDUs (the review considered only two studies written in English). Although this study included meta-analysis estimates, it does not report the prevalence estimates based on a random-effects model [21]. The authors report a measure of prevalence summary by region. The variations for the provinces were significant and the overall HCV prevalence was 0.614 (95% CI: 0.557-0.672). That estimate is similar to the one we predicted for this country (0.633; CI95%: 0.522-0.732). The authors reported a REM for HCV infection differences by sex, ethnic origin or being IDU vs non-IDU. In our study we could not conduct such type of analysis because of limited information regarding the participants' age and duration of IDU or other characteristics. Instead we conducted a REM for prevalence estimates and we found a great heterogeneity despite the concentration on selected countries by income level. This might be caused by factors such as: (1) age and sex of the participants, (2) availability of drugs, (3) quality of the data collection and information reported in each study, (4) IDU treatment practice, and (5) policies concerning IDU and treatment.
This study has some limitations, including that we did not have complete information about the age of participants and duration of IDU, which did not allow for an accurate analysis of these variables. Most studies only report the percentage of people older or younger than a https://doi.org/10.1371/journal.pone.0212558.g005 specific age cutoff, and a limited number of studies report the average age and its standard deviation for the study population. Additionally, the duration of time as an IDU is reported for different periods and for different percentages of drug users. A second limitation is that the quality of the studies included in this review was not rated, as was done in a previous study [11], which allows for the evaluation of each study's impact on the reported information. Another limitation is that we were unable to include studies that were published in languages other than English and Spanish, such as Chinese or Portuguese, which could have been relevant for this review. Our systematic review and meta-analysis provides relevant information to characterize the prevalence of HCV infection among IDUs in upper middle income countries. Our study highlights that HCV infection is common among the IDU population, with a pooled prevalence of 0.729 (95% CI 0.644-0.800). The results of our systematic review and meta-analysis suggest that the prevalence of HCV in upper middle income countries can vary greatly, from a lower prevalence in Brazil to a higher prevalence in China. Despite the high heterogeneity in the meta-analysis, our results are relevant because they highlight the variability in the characteristics of the studies examined. This variability can be also a consequence of different methods and analysis techniques to estimate the prevalence of HCV among IDU in UMIC. Our study attempts to address some of the issues in measuring and estimating the prevalence of HCV among IDUs in UMIC. Although some studies in low-or middle-income countries may have limitations that prevent a more accurate characterization of HCV prevalence, we only included studies that specifically reported the prevalence of HCV in an IDU population. Having precise estimates of the prevalence of HCV infection in high risk groups is important so that national health systems can plan to meet the specific health needs of these vulnerable populations.
Supporting information S1