IP-10 dried blood spots assay monitoring treatment efficacy in extrapulmonary tuberculosis in a low-resource setting

Abstract

Treatment efficacy is difficult to evaluate in extrapulmonary tuberculosis (EPTB) patients. Interferon-γ inducible protein (IP-)10 has been suggested as a biomarker for response to treatment. We have investigated if IP-10 from dried plasma spots (DPS) or dried blood spots (DBS) can be used in treatment monitoring of EPTB patients in a low-resource setting of Zanzibar. IP-10 levels in plasma, DPS and DBS samples collected before, during (2 months) and after TB treatment of 36 EPTB patients (6 culture and/or Xpert MTB/RIF positive and 30 clinically diagnosed) and 8 pulmonary tuberculosis (PTB) patients, were quantified by an enzyme-linked immunosorbent assay. There was a high positive correlation between IP-10 measured in plasma and DPS and DBS, respectively. We found a significant decline in IP-10 levels from baseline to end of treatment in plasma, DPS and DBS, both in EPTB and PTB patients. The declines were observed already after 2 months in HIV negative patients. In conclusion, the DPS/DBS IP-10 assay allows for easy and manageable monitoring in low-resource settings and our findings suggest that IP-10 may serve as a biomarker for treatment efficacy in EPTB patients, albeit further studies in cohorts of patients with treatment failure and relapse are needed.

Figure 1

Flow chart of patient inclusion and data collection during the study. Abbreviations: TB, tuberculosis; EPTB, extrapulmonary TB; PTB, pulmonary TB; DPS, dried plasma spot; DBS, dried blood spot.

Figure 2

Changes in IP-10 levels in plasma, dried plasma spots (DPS) and dried blood spots (DBS) during tuberculosis (TB) treatment. IP-10 concentrations measured using enzyme-linked immunosorbent assay from (A) patients with pulmonary TB before treatment (n = 8), at 2 months (n = 6) and at the end of treatment (n = 6) and from (B) patients with extrapulmonary TB before treatment (n = 36), at 2 months of treatment (n = 32) and at completion of 6–10 months of treatment (n = 24). Boxes represent the median and interquartile range, and the whisker show min/max values. Dotted lines indicate statistically significant differences. Statistical significance was evaluated by Wilcoxon Signed Ranks test using a significance level of 0.05.

Figure 3

Comparison of IP-10 levels in plasma, dried plasma spots (DPS) and dried blood spots (DBS) for individual extrapulmonary tuberculosis (EPTB) patients during TB treatment. IP-10 levels during 6–10 months of TB treatment in plasma, DPS and DBS in patients with extrapulmonary TB at baseline (n = 32), at 2 months of treatment (n = 32) and at the end of treatment (n = 24). IP-10 levels in baseline samples from the four patients who died shortly after start of TB therapy, are not presented in this figure. ¹TB pleuritis, ²Combined ascites and TB pleuritis, ^3–4Pregnant during treatment, ^5–7Persistant or increasing lymph node swelling at 2 months of treatment, ^8–9Lymph node TB, ¹⁰Abdominal TB. *HIV-infected patient.

Figure 4

IP-10 levels in different clinical groups of extrapulmonary tuberculosis (EPTB) patients during treatment in plasma, dried plasma spots (DPS) and dried blood spots (DBS). IP-10 concentrations were measured using enzyme-linked immunosorbent assay. EPTB sub-grouped according to (A) HIV status; before treatment (n = 35), at 2 months of treatment (n = 32) and at the end of treatment (n = 24), and according to (B) TB category; before treatment (n = 36), at 2 months (n = 32) and at the end of treatment (n = 24). HIV status was unknown for 1/36 EPTB patients. Boxes represent the median and interquartile range, and the whisker show min/max values. Dotted lines indicate statistically significant differences. Statistical significance was evaluated by Mann-Whitney U test, using a significance level of 0.05.