A novel rapid test for detecting antibody responses to <i>Loa loa</i> infections

<div><p>Ivermectin-based mass drug administration (MDA) programs have achieved remarkable success towards the elimination of onchocerciasis and lymphatic filariasis. However, their full implementation has been hindered in Central Africa by the occurrence of ivermectin-related severe adverse events (SAEs) in a subset of individuals with high circulating levels of <i>Loa loa</i> microfilariae. Extending MDA to areas with coincident <i>L</i>. <i>loa</i> infection is problematic, and inexpensive point-of-care tests for <i>L</i>. <i>loa</i> are acutely needed. Herein, we present a lateral flow assay (LFA) to identify subjects with a serological response to <i>Ll</i>-SXP-1, a specific and validated marker of <i>L</i>. <i>loa</i>. The test was evaluated on serum samples from patients infected with <i>L</i>. <i>loa</i> (n = 109) and other helminths (n = 204), as well as on uninfected controls (n = 77). When read with the naked eye, the test was 94% sensitive for <i>L</i>. <i>loa</i> infection and was 100% specific when sera from healthy endemic and non-endemic controls or from those with <i>S</i>. <i>stercoralis</i> infections were used as the comparators. When sera of patients with <i>O</i>. <i>volvulus</i>, <i>W</i>. <i>bancrofti</i>, or <i>M</i>. <i>perstans</i> were used as the comparators, the specificity of the LFA was 82%, 87%, and 88%, respectively. A companion smartphone reader allowed measurement of the test line intensities and establishment of cutoff values. With a cutoff of 600 Units, the assay sensitivity decreased to 71%, but the specificity increased to 96% for <i>O</i>. <i>volvulus</i>, 100% for <i>W</i>. <i>bancrofti</i>, and 100% for <i>M</i>. <i>perstans</i>-infected individuals. The LFA may find applications in refining the current maps of <i>L</i>. <i>loa</i> prevalence, which are needed to eliminate onchocerciasis and lymphatic filariasis from the African continent.</p></div

Reproducibility of the smartphone reader.

<p>Pooled <i>L</i>. <i>loa</i> sera were tested at different dilutions on a given cassette, and the test line intensity was measured 25 consecutive times. The table reports the mean value and standard deviation in reader units, and coefficient of variation in %. Calculating a CV on negative serum is not meaningful since the denominator is close to zero.</p

Sensitivity and specificity versus <i>O</i>. <i>volvulus</i> as a function of the cutoff value.

<p>Sensitivity and specificity versus <i>O</i>. <i>volvulus</i> as a function of the cutoff value.</p

Analytical sensitivity of the Loa Antibody Rapid Test.

<p>The test was run as per the General Procedure, with 5 μL of undiluted pooled <i>L</i>. <i>loa</i> sera, (left), 5 μL pooled <i>L</i>. <i>loa</i> sera serially diluted in negative serum from uninfected North American individuals, with dilution factors of up to 1:1600 (center), or 5 μL pure negative delipidized serum (right). The test lines were quantified with the smartphone reader. The data is reported in reader units (RUs).</p

Control and test line intensities by disease.

<p>The scatter plot shows individual data points, with median values as blue horizontal bars. The median value for the <i>L</i>. <i>loa</i> samples was 845. A possible cutoff value of 600 RUs is shown as a dotted line. Ov = <i>O</i>. <i>volvulus</i>, Wb = <i>W</i>. <i>bancrofti</i>, Mp = <i>M</i>. <i>perstans</i>, Ss = <i>S</i>. <i>stercoralis</i>, EN = Endemic Normals, NEN = Non-Endemic Normals.</p

Sensitivity and specificity at different cutoff values.

<p>The cutoff values are expressed in Reader Units (RUs). Sensitivity and specificity values are reported in %, with the 95% confidence intervals are reported in brackets. Ov = <i>O</i>. <i>volvulus</i>, Wb = <i>W</i>. <i>bancrofti</i>, Mp = <i>M</i>. <i>perstans</i>, Ss = <i>S</i>. <i>stercoralis</i>, EN = Endemic Normals, NEN = Non-Endemic Normals.</p

Reproducibility of the test strips.

<p>Pooled <i>L</i>. <i>loa</i> sera were tested at different dilutions, each time on 10 different cassettes randomly selected from the same manufacturing lot. The test lines were read after 20–30 minutes with the smartphone reader.</p