Latent tuberculosis infection in HIV-infected adults of Kinshasa, DR Congo

The World Health Organization (WHO) End Tuberculosis Strategy calls for a 90% reduction in tuberculosis (TB) deaths and an 80% reduction in the incidence rate between 2015 and 2035. The 2018 WHO guidelines recognize the need for high-burden countries to implement outreach and treatment for the most vulnerable patients with latent TB infection (LTBI) in addition to treating patients with the active disease. The Democratic Republic of Congo (DR Congo) is among the countries bearing the highest burden of TB. However, additional data on LTBI is required for effective policy and strategy against the disease. Accordingly, the aim of this study is to estimate the prevalence of LTBI in HIV-infected adults in Kinshasa, DR Congo.


Background The diagnosis and treatment of latent tuberculosis infection (LTBI) is part of the World
Health Organization's End Tuberculosis Strategy to control and eliminate tuberculosis (TB) worldwide [1]. The goal of this strategy is to reduce the incidence of tuberculosis worldwide to less than 10 per 100,000 by 2035 [1,2]. The World Health Organization (WHO) guidelines for 2018 highlight the need for high-burden countries to implement awareness-raising and treatment activities for the most vulnerable patients with latent TB infection in addition to treating patients with active TB [3].
The World Health Organization estimates that 10 million new cases and 1.3 million deaths from TB occurred in 2017 [4]. Of the 10 million cases of TB, 9% were co-infected with the human immunodeficiency virus (HIV) [4]. In 2015, an estimated 10.4 million new TB cases and 1.8 million deaths worldwide were recorded [5], and people living with HIV accounted for 1.2 million (11%) of all new TB cases [5]. Tuberculosis remains the leading cause of death in people living with HIV, particularly in sub-Saharan Africa (SSA), where it accounts for 50% of HIV-related deaths [6], [7]. It is estimated that about one third of the population worldwide is estimated to be latently infected with Mycobacterium tuberculosis [8,9] and thus have the potential of developing active TB. Latent TB infection is highly widespread in developing countries and constitutes a major obstacle to global TB control [10]. Identifying individuals with LTBI will increase screening rates and assist TB control [11].
Several factors increase the risk of active TB by facilitating the reactivation of remote LTBI or by promoting the progression of an infection recently contracted due to active disease [12]. These factors include HIV infection, recent contact with an infectious patient, initiation of anti-tumor necrosis factor (TNF) treatment, receiving dialysis, receiving an organ or hematologic transplantation, silicosis, incarceration, homelessness, drug addiction, and malnutrition [6,7]  Consequently, infection with M. marinum or M. kansasii, which express ESAT-6 or CFP-10, has been shown to produce positive results in IGRAs, as with the TST [18], [19]. There is some evidence of cross-reactivity between ESAT-6 and CFP-10 of M. tuberculosis and M.
leprae, but the clinical significance of this in settings where leprosy and TB are endemic (e.g., India and Brazil) is poorly characterized [20], [21].
In the DR Congo, as in most SSA countries, the high burden imposed by TB infection contrasts with the paucity of data on the prevalence of LTBI, which is increasingly considered a key step for TB control. This situation highlights the need to conduct studies in SSA countries in order to provide information for efficient TB control.
The aim of the present study is to determine the prevalence of LTBI in HIV-infected adults in Kinshasa, DR Congo, and to assess the concordance between the QuantiFERON TB Gold Plus ® (QFT-Plus®) interferon gamma release assay and the TUBERTEST ® skin test. People with a previous history of TB disease and treatment were excluded from this study.

Study population, design, and inclusion criteria
Also, those who reported any of the symptoms of current cough, fever, weight loss, or night sweats were excluded as well as individuals who were too ill to consent or unable to understand or comply with the study protocol.

Data collection
A detailed standard questionnaire was used to gather socio-demographic and clinical information and to record previous laboratory results. The information included date of birth, history of exposure to TB, history of TST test, BCG vaccination history, presence of a BCG scar, and symptoms suggestive of TB. Information on HIV infection was obtained from patient files.
Collection of samples 6 A qualified phlebotomist collected one ml of blood from each patient for laboratory tests.
The blood was collected aseptically by venipuncture directly into each of the four QFT-Plus blood collection tubes.

Tuberculin Skin Test (TST)
The standardized TST was performed using TUBERTEST® (Sanofi, Paris, France), which consists of 0.1 ml to 5 IU of TST. The test results were read 72 hours later. A positive TST was defined as an induration of more than 5 mm in horizontal diameter on the forearm. A negative TST was defined as an induration of less than 5 mm. Anergy is defined as the absence of skin reaction with delayed hypersensitivity following the injection of an antigen to which an individual is known to have developed a cell-mediated immune response [22].

Interferon gamma release assay (IGRA)
The QFT-Plus test was performed according to the manufacturer's instructions (Qiagen, Hilden, Germany) using the four-tube version comprising a negative control, a positive control, and two different TB antigen tubes.

Data management and statistical analysis
The data was entered into an Excel spreadsheet (Microsoft, Redmond, WA) and analyzed using Excel Microsoft and STATA software (Stata Statistical Software, Version 11, College Station, TX, Stata Corp LP, 2009). For the TST, we used the standard 5 mm cut-off points, while for QFT-Plus, we used the QFT-Plus algorithm ( Table 4). All individuals who had positive QFT-Plus and/or positive TST results were considered to have LTBI. The prevalence of LTBI was calculated as the number of positives by either the TST or QFT-Plus test divided by the total number of HIV positive adults tested [23]. The indeterminate results of QFT-Plus were considered negative results. Active TB was ruled out using the WHO symptom screen [3].

Baseline study population
In total, 248 HIV-infected adults under antiretroviral therapy during a mean of 4 years were tested with both the TST and QFT-Plus. Seventy-five of the participants were females, 63% were aged between 30-49 years, 27% had close contact with patients who had active TB, and 2% were diabetics. The main characteristics of the study participants are summarized in Table 1.  (Table 2). Twenty-four percent positive TST results were female while twenty percent of positive QFT Plus results were also female.  (Table 3). Furthermore, 21% of the positive results were from females while 5% from men. (Table 4). Table 4

Discussion
This was a pilot study conducted in the DR Congo to estimate the prevalence of LTBI in HIV-infected adults. A larger study will be conducted all over the country.
Using the IGRA alone to diagnose LTBI, the prevalence of LTBI was found to be 25.8%. While using positivity by the TST, as requested by the guidelines of the WHO and suggested by some authors [24,25], the prevalence of LTBI rose to 30.7%.
Our results are lower than those of the meta-analysis done by Ayubi et al. [26] who found the prevalence of LTBI among HIV infected adults to be 59% (95% CI: 49, 69).
The prevalence of LTBI estimated by the TST was greater than that estimated by QFT-Plus, which is similar to other observations [27]. The higher prevalence of LTBI seen in the TST is likely due to false-positive results attributed by nontuberculous mycobacterium (NTM) infection and prior BCG vaccination [15, [28][29][30]. In Sub-Saharan Africa, where the Bacillus Calmette-Guérin vaccine is administered at birth, the TST will be affected by vaccination, which is not the case with IGRAs [31,32].
There are currently two tests for diagnosing latent tuberculosis infection (LTBI): the TST and the IGRA. We found the results of the TST and QFT-Plus tests to be in poor agreement (0.225), which is similar to what others have found [31,[33][34][35][36][37][38]. by T-SPOT.TB and QFN-G-IT in HIV positive patients were higher for patients with a CD4 count >350 cells/µl than <350 cells/µl (28%, 68% and 39%, which is 3% versus 20% and 10%, respectively) [32]. We believe the reason for this is that gamma interferon in the IGRA is produced after stimulation of CD4 and CD8. The TST is less influenced by the change of CD4 (p<1), while the IGRA is more influenced (p<0,000). Furthermore, even in Consent for publication (Not applicable) Availability of data and material The datasets generated during and/or analyzed during the current study are available from the corresponding author upon reasonable request.

Competing interests
We declare that we have no conflict of interest.