Global and Regional Prevalence and Burden of Latent and Acute Toxoplasmosis in People Living With HIV: an Updated Systematic Review and Meta-analysis


 Background: Toxoplasma gondii infection is one of the most prevalent opportunistic and life-threatening infections in people living with HIV (PLWH). Here, we undertook a meta-analysis to estimate the global prevalence of latent (LT) and acute (AT) toxoplasmosis in PLWH.Methods: Eligible studies reporting the prevalence of LT or AT in PLWH were searched from January 1980 to July 2020, using PubMed/MEDLINE, Scopus, Web of Sciences, SciELO and Embase databases. We used a random effects model to calculate pooled prevalence estimates with 95% confidence intervals (CI) and evaluated overall burden of co-infection worldwide. Countries were categorised based on World Health Organization regions. Multiple subgroup and meta-regression analyses were performed.Results: From 4,024 studies identified, 150 (involving 44,473 PLWH) and 65 (involving 17,705 PLWH) studies met the inclusion criteria, for LT and AT in PLWH, respectively. The overall prevalence rates of LT and AT in PLWH were 37.4% (95% CI, 33.4–4.4) and 1.3% (95%CI, 0.9–1.8%), respectively. We estimated that, worldwide, approximately 14.2 and 0.5 million PLWH are affected by LT and AT, respectively. The highest and lowest burden of LT and AT were seen in the African and Western Pacific regions, respectively. Moreover prevalence rates were highest in countries with low levels of income and human development indexes. We indicated that eating raw/undercooked meat, frequent contact with soil, low numbers of CD4+ T lymphocytes and older age were significant risk factors of toxoplasmosis in PLWH.Conclusion: Our findings revealed that, a high number of PLWH are exposed to or infected with T. gondii. These findings suggest a need for more routine testing, care of, and treatment for T. gondii infection in all PLWH, and educating these patients about risk factors and preventive measures to reduce the burden of both latent and acute toxoplasmosis.


Introduction
Toxoplasma gondii infection or toxoplasmosis is one of the most prevalent opportunistic and lifethreatening infections in people living with human immunode ciency virus (PLWH) [1]. Toxoplasmic avidity; (b) Toxoplasma seropositivity for both IgG and IgM, or (c) seroconversion from a Toxoplasma IgG negative status to an IgG positive status.

Data extraction and quality assessment
We extracted the following variables from each study: rst author's name, country, city, year of publication, study design, the number of participants, number of PLWH seropositive for LT or AT, and the type of diagnostic methods. Countries were grouped according to the WHO-de ned regions [17], per capita income [18] and the human development index [HDI] [19]. Moreover, we extracted patient data on the number of CD4+ T cells (<200, 200-500, and > 500 cells/μL) and number of patients in distinct age groups (<20, 20-40, 41-60, >60), if available. In order to identify associated risk factors for toxoplasmosis in PLWH, we extracted data including place of residence (urban/rural), gender (male or female), close contact with dogs or cats, contact with soil, consumption of raw/undercooked meat, consumption of raw/unwashed vegetables, and drinking untreated water. To evaluate the methodological quality of included studies, we used the JBI (Joanna Briggs Institute) Prevalence Critical Appraisal Tool, as described by Munn et al [20].

Statistical analysis
Analyses were performed using Stata statistical software (v. 16 Stata Corp., College Station, TX, USA). For each individual study, the prevalence rate was de ned as the number of PLWH with LT or AT divided by the total number of PLWH screened. We used the variance of the study-speci c prevalence estimates with the Freeman-Tukey double arcsine transformation before pooling the data using the DerSimonian-Laird random-effects meta-analysis model to minimise the effect of studies with extremely small or extremely large prevalence [21,22]. The random-effects model was selected in anticipation of substantial variation in prevalence estimates of LT or AT across the included studies [23]. The between-studies heterogeneity was evaluated by Cochran's Q test and I 2 index. I 2 values of 25%, 50% and 75% represented low, medium and substantial heterogeneity, respectively [24]. To explore potential drivers of heterogeneity, several subgroup and meta-regression analyses were performed considering the following study features: (i) WHO region, (ii country, (iii) type of diagnostic method, (iv) type of study design, (v) country income level, and (vi) country HDI level. Moreover, to calculate the global burden of LT and AT in PLWH, we followed the method used in the most recently published meta-analysis on this subject [14] and used data from WHO showing the number of PLWH in 2018 [25]. We then multiplied this number by our calculated percentages of PLWH with LT or AT (with a 95% CI) at the global and national levels. We did not undertake an assessment of publication bias, as it is not relevant for prevalence studies [26]. A P value < 0.0 5 was considered statistically signi cant.  Table 2). Meta-regression analyses revealed a signi cant decreasing trend in prevalence in countries with increasing per capita income (C = -2.97 e − 06 ; P-value = 0.004) and HDI levels (C = -0.473; Pvalue < 0.001) ( Fig. 2A and B). Studies that were performed after 2005 showed slightly higher prevalence rates than other periods, although this increasing trend (1980-2020) was non-signi cant in meta-regression analysis (C = 0.0016; P value = 0.46) (   . With respect to other risk factors associated with LT, our results showed that female patients, those who lived in rural areas, those who were cat or dog owners, and those who consumed raw/unwashed vegetables or consumed untreated water were at more, but nonsigni cant greater, risk to acquire infection. More details are given in Table 3 and S10-15 Figs. In Egger's test we did not identify any signi cant publication bias (Table 3).

Global prevalence of acute Toxoplasmosis
The global prevalence of AT in PLWH, when data for all 65 datasets representing 32 countries were pooled, was 1.3% (95%CI, 0.9-1.8%; 289/17,705). The heterogeneity between studies was signi cant (I 2 = 79.1%, P < 0.001). With respect to WHO-epidemiological regions, the highest prevalence rates were found in South America (2.0%; 0.1-5.4%), and then the Eastern Mediterranean region (1.8%; 0.7-3.3%), and the lowest prevalence rate was found in the European region (0.6%; 0.2-1.3%). The pooled prevalence rates in other WHO regions were: 1.6% (0.5-3.1%) in North America, 1.3% (0.9-1.8%) in South-East Asia, 1.2% (0.2-2.6%) in the Western Paci c and 0.9% (0.2-1.2%) in Africa (Table 4 and Fig. 1B). Moreover, we estimated that approximately 492,700 (341,100-682,200) PLWH worldwide were affected by AT. Our estimates demonstrated that countries in the African region, has the largest number of PLWH with AT (231,300; 51,400-308,400), accounting for approximately 47% of cases of HIV-AT co-infection worldwide. Additional details pertaining to the prevalence and burden of AT in PLWH in WHO-regions and individual countries are given in Table 4 and Fig. 1B. Subgroup and meta-regression analyses according to socio-demographic and study characteristics In subgroup analyses, when the pooled prevalence was strati ed according to the income-level of a country, the highest prevalence rates of AT were estimated for countries with lower-middle income-levels (1.8%, 0.7-3.2%) and the lowest for those with low income-levels (0.4%, 0.0-1.9%). Based on HDI level, the highest prevalence rates were seen in countries with low HDI-levels (1.4%, 0.1-1.3%) and the lowest prevalence rates in countries with high HDI-levels (1.3%, 0.7-2.0%) ( Table 2). Moreover meta-regression analyses revealed a non-signi cant decreasing trend in prevalence in countries with increasing per capita income (C = -0.00082; P-value = 0.88) and HDI levels (C = -0.0056; P-value = 0.92) in a country ( Fig. 2D and E). Sub-group analysis on year of study showed higher prevalence rates after 2010. This increasing trend was nonsigni cant in meta-regression analysis (C = 0.0002; P-value = 0.7) (Fig. 2F). In subgroup analyses, according to type of study, the highest prevalence rates were estimated in the case-control (2.6%, 0.8-5.0%), and then in prospective cohort (2.6%, 0.8-5.0%) studies, and the lowest prevalence rates were estimated in retrospective cohorts (1.0%, 0.3-2.0%). Subgroup analysis based on diagnostic methods showed that prevalence rates were similar when studies used both IgG-IgM tests (1.2%, 0.7-1.8%) and seroconversion (1.2%, 0.8-1.7%). More subgroup analyses and details are given in Table 2.

Discussion
This systematic review and meta-analysis is the rst global assessment of the prevalence of AT and the most comprehensive assessment of the prevalence of LT among HIV-infected people. Our ndings highlight the high prevalence and burden of LT (37.4%, approximately 14.2 million people) and AT (1.3%, approximately 0.5 million people) in PLWH. Estimates reported here are similar to the prevalence rates for LT and AT observed in recent meta-analyses that we have conducted among pregnant women [10,11], and also corroborate a general assumption that one-third of humans are infected with Toxoplasma [27]. Our estimate for LT (37.4%) is slightly higher than a previous meta-analysis by Wang et al. (35.8%) in the same population [14], although they included only 74 studies (49% of those included in our meta-analysis) and 25,989 HIV-infected people (approximately 58% of individuals in our meta-analysis) [14].
In the present study, the overall prevalence and burden of LT and AT varied across regions, with the highest prevalence in African and South-American countries and the lowest prevalence in the Western Paci c region. These ndings are in agreement with previous global studies in pregnant women [10,11]. In a study by Wang et al. among PLWH, the prevalence of LT was highest in countries located at sub-Saharan Africa, Latin America and the Caribbean regions [14]. The variability in prevalence in these different regions could be related to differences in climate (e.g. temperature and humidity) [10,28], the degree of contamination of the environment (e.g., soil and water) with cat feces and T. gondii oocysts, cultural or culinary habits, particularly the consumption of semi-cooked or raw meat [7,9], infectivity of the diversity of T. gondii genotypes present, accessibility to public health and sanitary services, control measures for stray cats, income and HDI status of a country [10,11]. A detailed explanation about effects of these variables on the prevalence of toxoplasmosis in different populations is available in previous publications [10,11,27].
Considering risk factors for exposure to T. gondii, our analyses found that eating raw/undercooked meat, frequent contact with soil, older age, and low number of CD4 + lymphocytes were associated with increased seropositivity for LT. The risk factors for acquisition of toxoplasmosis are well documented and have been reviewed in previous studies [29,30,31,32,33]. Consumption of raw/undercooked meat is assumed to be a major source of T. gondii infection, however, as a risk factor, this depends on the kind of meat consumed and local culinary habits of meat preparation; for example in China who used duck and beef meat had higher seropositivity for T. gondii infection compared to those who used other kinds of meat including pork, lamb and chicken [34]. Lamb and pork are major dietary risk factors in European and American countries and lamb is a major risk factor in Middle Eastern countries [30,31,35,36]. Contact with soil is an important source of exposure to infectious oocysts [37,38,39,40,41,42,43,44,45,46]. We identi ed an increasing prevalence with age, which can be explained by the fact that risk of Toxoplasma infection is constant over time and older PLWH have had a longer period of time for exposure to Toxoplasma [7,47]. This observation is in agreement with our previous study in pregnant women [11]. Our ndings indicated that another, and maybe most important risk factor for toxoplasmosis in PLWH, is a low number of CD4 + lymphocytes. It is well known that decline in CD4 + T cells is the main reason for reactivation of latent infection in PLWH and can lead to severe toxoplasmosis and TE in these patients [48,49].
The ndings from this study must be interpreted in consideration of certain limitations. First, and common to systematic reviews, we found a high statistical heterogeneity between studies, which could not be explained by the subgroup and meta-regression analyses; the major sources of heterogeneity might result from the study characteristics including differences in study design, geographical distribution, sample size, publication year, and detection methods, including differences in the performance of these methods and differences in cut-off values for test-positivity. Second, most studies in the literature have used serological methods to detect latent or acute toxoplasmosis in PLWH; serological methods are unreliable in PLWH due to profound immunode ciency. While bioassays (in mice or cats) are considered as the gold standard for the de nitive diagnosis of toxoplasmosis, these methods are not applicable in humans. Moreover due to variation in the sensitivity and speci city (and, thus, positive and negative predictive values) of serological methods, the ndings from the present study might underestimate or overestimate the prevalence of latent or active toxoplasmosis in PLWH. In addition our estimates of acute toxoplasmosis might not be the true prevalence, since relatively few studies were included; we did not include studies using data on DNA, due to a paucity of such data and the fact that the majority of molecular studies were done in HIV patients suspected to TE or pulmonary toxoplasmosis. Thus, the estimated rates reported here must be considered as apparent prevalence rates of latent or acute toxoplasmosis in PLWH. We lastly acknowledge that despite our comprehensive systematic search, the eligible studies and data were not available for many countries and some regions of the world were not appropriately covered, which may hinder the translatability of our results at a global scale.

Conclusion
In conclusion, our ndings revealed that, globally, more than one third of PLWH are exposed to, or infected with. Toxoplasma. International guidelines recommend toxoplasmosis screening for all PLWH and also prophylactic measures (e.g. co-trimoxazole) should be implemented for all T. gondii-seropositive PLWH with < 100 CD4 + cells/µL. This approach is currently not well implemented in many countries.
In our study, the prevalence rates was higher in less developed countries and regions; therefore these countries, in particular, should promote routine testing, care, and treatment for T. gondii infection in all PLWH.
We also determined potential risk factors related to toxoplasosis in PLWH. T. gondii-seronegative PLWH should be aware and educated about these risk factors and related preventive measures to reduce the risk of health problems arising from both latent and acute toxoplasmosis.

Declarations
Compliance with ethical standards Figure 1 Prevalence of latent (A) and acute (B) toxoplasmosis in people living with HIV in different countries using geographic information system (GIS). Note: The designations employed and the presentation of the material on this map do not imply the expression of any opinion whatsoever on the part of Research Square concerning the legal status of any country, territory, city or area or of its authorities, or concerning the delimitation of its frontiers or boundaries. This map has been provided by the authors.