Diagnostic Value of Anti-HTLV-1-Antibody Quantification in Cerebrospinal Fluid for HTLV-1-Associated Myelopathy

Abstract

The diagnostic accuracy of cerebrospinal fluid (CSF) anti-human T-cell leukemia virus type I (HTLV-1) antibody testing for HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM) remains unclear. Therefore, we measured the anti-HTLV-1 antibody levels in CSF using various test kits, evaluated the stability of CSF antibodies, and performed a correlation analysis using the particle agglutination (PA) method, as well as a receiver operating characteristic (ROC) analysis between patients with HAM and carriers. The CSF anti-HTLV-1 antibody levels were influenced by freeze-thaw cycles but remained stable when the CSF was refrigerated at 4 °C for up to 48 h. Measurements from 92 patients (69 patients with HAM and 23 carriers) demonstrated a strong correlation (r > 0.9) with the PA method across all six quantifiable test kits. All six test kits, along with CSF neopterin and CXCL10, exhibited areas under the ROC curve greater than 0.9, indicating a high diagnostic performance for HAM. Among these, five test kits, Lumipulse and Lumipulse Presto HTLV-I/II, HISCL-UD (a kit under development), HTLV-Abbott, and Elecsys HTLV-I/II, established a cutoff with 100% sensitivity and maximum specificity, achieving a sensitivity of 100% and a specificity ranging from 43.5% to 56.5%. This cutoff value, in combination with clinical findings, will aid in the accurate diagnosis of HAM.

Keywords: HTLV-1-associated myelopathy; anti-HTLV-1 antibody; cerebrospinal fluid; diagnosis; human T-cell leukemia virus type 1.

Figure 1

Effect of freeze–thaw cycles on the measurement of anti-HTLV-1 antibody quantitative data in CSF. Six cases of CSF—two low, two medium, and two high titers—were employed in the assay for each test kit (see the Section 2.5 for details). The CSF underwent one, two, or three freeze–thawing cycles, and the anti-HTLV-1 antibody titers were assessed. Measurements for each time point are displayed as the mean ± SD. A one-way repeated-measures ANOVA was used to compare the corresponding data among the three conditions. p ≤ 0.05 was considered statistically significant. COI, cutoff index; S/CO, signal-to-cutoff ratio. Information on each test kit is shown in Table 1.

Figure 2

Effect of storage time at 4 °C on the measurement of anti-HTLV-1 antibody quantitative data in CSF. Six cases of CSF—two low, two medium, and two high titers—were employed in the assay for each test kit (see the Section 2.5 for details). The CSF was stored at 4 °C for 0 h, 24 h, and 48 h, and the anti-HTLV-1 antibody titers were assessed. Measurements for each time point are displayed as the mean ± SD. A one-way repeated-measures ANOVA was used to compare the corresponding data among the three conditions. p ≤ 0.05 was considered statistically significant. COI, cutoff index; S/CO, signal-to-cutoff ratio. Information on each test kit is shown in Table 1.

Figure 3

Distribution of CSF anti-HTLV-1 antibody titers in patients with HAM. The vertical axis represents the number of patients, and the horizontal axis represents the antibody titer measured using the PA method. Of the 322 patients with HAM, 248 were not on steroid therapy (black), and 74 were on steroid therapy (gray).

Figure 4

ROC analysis to assess the diagnostic performance of the five markers in discriminating patients with HAM from HTLV-1 carriers. We collected the past data of five known markers, the PVL in PBMCs, the PVL in CSF cells, the ratio of these PVLs, CSF CXCL10, and CSF neopterin, in 92 individuals (69 patients with HAM and 23 HTLV-1 carriers). PVL, HTLV-1 proviral load; PBMCs, peripheral blood mononuclear cells; AUC, area under the curve.

Figure 5

ROC analysis of CSF anti-HTLV-1 antibody levels. CSF anti-HTLV-1 antibody levels in 92 individuals (69 patients with HAM and 23 HTLV-1 carriers) were measured using seven different quantifiable anti-HTLV-1 antibody test kits, and an ROC analysis was performed for each. Information on each test kit is shown in Table 1. AUC, area under the curve.

Figure 6

Correlations with the PA method. Using the results of anti-HTLV-1 antibody titers from 92 CSF samples (69 patients with HAM [red] and 23 carriers [blue]), we examined the correlations between the PA method and six other test kits. Antibody titers for the PA method are indicated as Log₂ values, and antibody levels (COI and S/CO) for all other test kits are demonstrated as Log₁₀ values. The linear equation, Spearman’s rank correlation coefficient (rs), 95% confidence interval, and p-value are shown in the figure. COI, cutoff index; S/CO, signal-to-cutoff ratio. Information on each test kit is shown in Table 1.