The Latent Tuberculin Test after 1-year Therapy with Anti-TNF in Babylon , Iraq

Background: Rheumatoid arthritis (RA) patients receiving receive anti-TNF agents are at increased risk of reactivation of latent tuberculosis infection (LTBI). The tuberculin skin test (TST) is widely used to screen LTBI and providing preventive treatment, in an effort to meet the WHO target of a 90% reduction in TB by 2035. Objectives: To determine the proportion of TST conversion among RA patients after 1 year of anti-TNF treatment, and association of positive TST result with patients’ socio-demographic characteristics and medical history. Methods: This community-based cross-sectional study was conducted at the department of Rheumatology of Marjan Teaching Hospital in Iraq, for a period of 1 year. Patients with RA/and spondyloarthropathy, and who received antiTNF therapy for >1 year, underwent TST. Their demographic data and medical history were also obtained. All statistical analysis was performed using SPSS (Version 20) and, p < 0.05 was considered as a significant. Data from the baseline and 1 year follow-up was subjected to the Kolmogorov-Smirnov test to determine whether they were normally distributed. Chi-Square test used to test significance of TST among etanrecept and infliximab at the end of the study. Results: A total of 96 patients were enrolled, including 55 (57.3%) males and 41 (42.3%) females with average age of 41.1, and mostly 68 (70.8%) from Babylon Governorate of Iraq. A total of 40 (41.7%) patients had rheumatoid arthritis alone, and the remaining 56 (58.3%) had a comorbidity of spondyloarthropathy. Majority of the patients 65 (67.7%) received the biological agent infliximab, while 31 (32.3%) patients received Etanercept for RA for a period of 1 year. There was a statistically significant decreasing in the median ESR and disease activity from the baseline to the end of the study (p-value <0.01). There was no significant difference in TST results based on gender or age. Both infliximab and etanrecept were significantly associated with a decreasing in ESR and disease activity Conclusion: This study has shown that there was very low TST conversion among RA patients after 1 year of antiTNF treatment and, age and gender were not associated with TST. 1Introduction Rheumatoid arthritis (RA) and spondyloarthropathy are autoimmune diseases characterized by persistent synovitis and systemic inflammation due to the release of a potent cytokine TNF-α, which in turn releases other cytokines (IL1 and IL6). Blockade of TNF-α using anti-TNF agents has revolutionized the treatment of RA and other systemic inflammatory diseases. The efficacy of the anti‐TNF agents such as Etanercept and Infliximab for RA has been demonstrated in large‐scale trials 00]. However, patients receiving anti-TNF agents are at an increased risk of fungal and bacterial infection, particularly reactivation latent tuberculosis infection (LTBI) 0]. LTBI is a state of persistent immune response to stimulation by Mycobacterium tuberculosis (M. tuberculosis) antigens without evidence of clinically manifested active TB. About one-third of the world’s population is estimated to Journal of University of Babylon for Pure and Applied Sciences (JUBAS) by University of Babylon is licened under a Creative Commons Attribution 4.0 International License. 2018.

have LTBI.The lifetime risk of reactivation for a person with documented LTBI is estimated to be 5-10% 0].The reactivation of TB can be averted by screening for high risk individuals and preventive treatment.
In the absence of a standard diagnosis for LTBI, the tuberculin skin test (TST) an easy and inexpensive screening test available worldwide, despite concerns over its sensitivity and specificity.TST is a measure of the delayed-type hypersensitivity reaction to intradermal inoculation of purified protein derivative, a crude mixture of >200 M. tuberculosis proteins 0].Due to the inability of PPD to distinguish between TB infection and Bacille Calmette-Guerin (BCG) induced reactivity, non-infected person may develop a false-positive TST result.Thus, it is recommended that a previous history of BCG vaccination should be considered while interpreting skin test results 0].
Tuberculosis is highly endemic in Iraq, with an estimated incidence of 45 /100000 population (i.e.estimated total new TB cases is around 15000 per year), prevalence of 74/100000 and mortality of 3/100000 0].
The WHO has developed guidelines for the management of LTBI to facilitate achievement of a 90% reduction in TB incidence and a 95% reduction in TB deaths by 2035 0].Thus this community-based cross-sectional study conducted in Iraq to determine the proportion of TST conversion among patients after one year of starting biological treatment.In addition, we assessed the association of positive TST result with patients , socio-demographic characteristics and medical history.

2-Materials and Methods
This was a community-based cross-sectional study conducted at the department of rheumatology of Marjan teaching hospital in Iraq, for a period of 1 year (1st January 2016 to 30th December 2016).

3-Population of the Study
Male and female outpatients aged 18 years diagnosed with rheumatoid arthritis or/and spondyloarthropathy, and who received anti-TNF therapy with biological agents (etanrecept, infliximab) for more than one year, were randomly selected for this study.
Patients with active TB, positive TST before starting biological treatment, and serious medical illness (diabetes mellitus, uremia, malignancy, and HIV infection) were excluded from this study.

4-Screening
All patients were screened for the following before enrollment in this study:  Negative TST before the start of anti-TNF therapy  Normal X-ray results  No evidence of active and latent TB at time of starting the anti-TNF therapy.

5-Data Collection
Demographic data (age, residence area, occupational state) and medical history (history of TB, history of contact with TB patient, medication history, ESR results before (Baseline) and after (New) biological treatment, BCG vaccination and scar status) was obtained from patients.

6-Tuberculin Skin Test
TST was performed in the respiratory unit of Marjan Teaching Hospital using the Mantoux technique.Five international units of Purified Protein Derivative (PPD RT23, Staten Serum Institute, Copenhagen, Denmark) (kept refrigerated and away from light at The Chest and Respiratory Disease Institute), were applied by an intradermal injection in the middle third of the inner forearm.The skin reaction was quantified 48 to 72 hours after the injection by a pulmonologist.Using a transparent ruler, the largest transverse diameter of indurations was measured in millimeters.

7-Interpreting TST Results:
 If no indurations developed, it was considered as 0 mm.Erythema of any size without indurations was considered as 0 mm. If indurations developed it was reported as either ≥10 mm or <10 mm.Indurations ≥10 mm were considered as positive TST.Only patients with positive TST were sent for chest radiography. Patients who did not complete the test were excluded from the study.

8-Statistical Analysis
Categorical variables such as sex were presented in frequency tables (number and percent), while continuous variables such as age were provided as descriptive statistics (mean, standard deviation, median, minimum, maximum and interquartile range).Data from the baseline and 1 year follow-up evaluations was subjected to the Kolmogorov-Smirnov test to determine whether they were normally distributed.Chi-Square test used to test significance of TST among etanrecept and infliximab at the end of the study.ESR was calculated among etanrecept and infliximab at baseline and at end of the study.Significant group differences over time were calculated using the Mann-Whitney test.Wilcoxon's rank sum test was used to compare ESR values between baseline and at the end of the study by using etanrecept and infliximab.
All statistical analysis was performed using SPSS (Version 20) and, p < 0.05 was regarded as significant.
A total of 40 (41.7%)patients had rheumatoid arthritis alone, and the remaining 56 (58.3%) had a comorbidity of spondyloarthropathy.Majority of the patients 65 (67.7%) received the biological agent etanrecept, while 31 (32.3%)patients received infliximab for RA for a period of 1 year.

2-9 Medical History
Of the 96 patients that took the TST, 8 (8.3%) patients tested positive for LTBI but had normal findings on X-ray.None of the patients had a history of TB and all patients tested negative for hepatitis.Majority of the patients 87 (92.6%) had the BCG scar.

3-9 Erythrocyte Sedimentation Rate
There was a statistically significant decreasing in the median ESR from the baseline to the end of the study (pvalue <0.01) (Table 1).

4-9 Anti-TNF
Of the 31 patients receiving etanrecept, none were positive for TST.Whereas, 8 (12.3%) of the 65 patients receiving infliximab were positive and 57 (87.7%) were negative for TST.There was a statistically significant difference in the TST results between etanrecept and infliximab (p-value = 0.041) (Table 2).None of the patients developed active TB during the 1 year follow-up.

5-9 TST results stratified by age group (<40 years and > 40 years)
A total of 48 patients were <40 years old, of which 3 patients tested positive and 45 tested negative for TST.A total of 48 patients were >40 years old, of which 5 patients tested positive and the remaining 43 tested negative for TST.The difference in TST results between the 2 age groups was not statistically significant (p-value=0.465)(Table 3).

6-9 TST results stratified by gender
The TST results were similar in males and females and not statistically significant (p-value=0.290) was ( Table 5).

7-9 TST results stratified by anti-TNF
There was a statistically significant decreasing in the median ESR between Baseline and New visit associated with the use of both etanrecept (p<0.001,Wilcoxon W Test) and infliximab (p=0.014,Wilcoxon W Test) (Table 5).
There was a statistically significant difference in median ESR at Baseline visit between etanrecept and infliximab (p=0.014,Mann-Whitney U test), while there was no statistically significant difference in the median ESR at New visit (p=0.164,Mann-Whitney U test) (Table 5).

10-Conclusion
This study has shown that there was very low TST conversion among RA patients after 1 year of anti-TNF treatment and, age and gender were not associated with TST.

11-Discussion
The diagnostic accuracy of TST in the detection of LTBI is high among patients with inflammatory diseases even in the setting of immunosuppression 0.
ESR is a non-specific marker of inflammation and is elevated in a number of noninfectious (RA) and infectious conditions, including TB.The findings from our study show a significantly decreased ESR after 1 year of treatment with anti-TNF agents, thus indicating efficacy of both biological agents (etanrecept and infliximab) in the treatment of RA.This was in agreement with the clinical efficacy of these agents in several clinical trials[ 0], 0].In addition, there was no statistically significant difference in outcome between the 2 biological agents.
Anti-TNF treatment has been documented as one of the factors associated with increased risk of progression of TB.The risk of TB increased up to 20-fold with 43% of TB cases within the first 90 days of administration of anti-TNF therapy, namely infliximab 0].The findings from our study showed that both infliximab and etanrecept were significantly associated with a decrease in ESR Unlike the study by Borgdorff MW et al which showed an association between males and increased age with TB, our study did not show any significant differences in TST results based on gender or age 0].TB did not develop in any of the 96 patients in this study who were on anti-TNF therapy was for year.The 8 patients who tested positive upon TSTS, showed no abnormality on X-ray.
Limitation of the study is that, although the TST approach increases the detection of remote infection, it may also increase the false-positive rate, leading to a unnecessary prophylactic treatments.

12-Acknowledgement
The authors would like to thanks ClinArt MENA for providing data analysis and medical writing services.