A) An overview of ELISA seropositivity and arbitrary units (AU) as well as B) Ll-SXP-1-RDT positivity and absolute intensity of Reader Units (RU) by loiasis subgroups.

*Arbitrary Antibody Units, ** Interquartile range, *** Wilcoxon-rank sum test comparing median AU or RU. ‡Participants were defined by microfilaremia density as low (1–7,999 mf/mL), high (8,000–19,999 mf/mL) and hyper-microfilaremic (≥20,000 mf/mL).</p

Database of the study.

BackgroundThe parasitic disease loiasis is associated with significant morbidity and mortality. Individuals with hyper-microfilaremia (greater than 20,000 microfilariae per mL of blood) may suffer from serious treatment-related or spontaneous adverse events. Diagnosing loiasis remains complex and primarily relies on direct parasite detection. In this study, we analyzed the performance of various diagnostic tests and the influence of parasitological and clinical factors on test outcomes in samples from individuals living in an endemic region.MethodsData and samples were collected from rural Gabon. Loiasis was defined as either detectable microfilaremia, or a positive history of eyeworm as assessed by the RAPLOA questionnaire. Diagnostic testing included a quantitative PCR (qPCR) for detection of Loa loa DNA in blood samples, an in-house crude L. loa antigen IgG ELISA, and a rapid test for antibodies against the Ll-SXP-1 antigen (RDT). Sensitivity and specificity were determined for each test and factors potentially influencing outcomes were evaluated in an exploratory analysis.ResultsELISA, RDT and qPCR results were available for 99.8%, 78.5%, and 100% of the 1,232 participants, respectively. The ELISA and RDT had only modest diagnostic accuracy. qPCR was specific for L. loa microfilaremia and Cycle threshold values correlated with microfilarial density. Anti-L. loa IgG levels were highest in occult loiasis, and antibody levels correlated inversely with L. loa microfilarial density as did RDT line intensities. Only 84.6% and 16.7% of hyper-microfilaremic individuals tested positive by ELISA (11/13) and RDT (2/12), respectively.ConclusionNone of the tests demonstrated high sensitivity and specificity for loiasis. Indirect diagnostic assays were characterized by low specificity. Additionally, hyper-microfilaremic individuals often tested negative by RDT and ELISA, indicating that these tests are not suitable for individual case management in endemic populations.</div

Saponin lysis laboratory protocol.

BackgroundThe parasitic disease loiasis is associated with significant morbidity and mortality. Individuals with hyper-microfilaremia (greater than 20,000 microfilariae per mL of blood) may suffer from serious treatment-related or spontaneous adverse events. Diagnosing loiasis remains complex and primarily relies on direct parasite detection. In this study, we analyzed the performance of various diagnostic tests and the influence of parasitological and clinical factors on test outcomes in samples from individuals living in an endemic region.MethodsData and samples were collected from rural Gabon. Loiasis was defined as either detectable microfilaremia, or a positive history of eyeworm as assessed by the RAPLOA questionnaire. Diagnostic testing included a quantitative PCR (qPCR) for detection of Loa loa DNA in blood samples, an in-house crude L. loa antigen IgG ELISA, and a rapid test for antibodies against the Ll-SXP-1 antigen (RDT). Sensitivity and specificity were determined for each test and factors potentially influencing outcomes were evaluated in an exploratory analysis.ResultsELISA, RDT and qPCR results were available for 99.8%, 78.5%, and 100% of the 1,232 participants, respectively. The ELISA and RDT had only modest diagnostic accuracy. qPCR was specific for L. loa microfilaremia and Cycle threshold values correlated with microfilarial density. Anti-L. loa IgG levels were highest in occult loiasis, and antibody levels correlated inversely with L. loa microfilarial density as did RDT line intensities. Only 84.6% and 16.7% of hyper-microfilaremic individuals tested positive by ELISA (11/13) and RDT (2/12), respectively.ConclusionNone of the tests demonstrated high sensitivity and specificity for loiasis. Indirect diagnostic assays were characterized by low specificity. Additionally, hyper-microfilaremic individuals often tested negative by RDT and ELISA, indicating that these tests are not suitable for individual case management in endemic populations.</div

The PCR settings for the <i>Loa loa</i> qPCR and <i>Mansonella</i> real-time FRET assay.

The PCR settings for the Loa loa qPCR and Mansonella real-time FRET assay.</p

Sensitivity analysis with a loiasis case definition of eyeworm history during the previous year and/or detectable microfilaremia.

Sensitivity analysis with a loiasis case definition of eyeworm history during the previous year and/or detectable microfilaremia.</p

Sensitivity, specificity, positive and negative predictive values of the <i>Loa loa</i> qPCR, in-house ELISA and Ll-SXP-1-RDT to detect loiasis defined by either a positive life-time history of eyeworm assessed by the RAPLOA questionnaire and/or detectable microfilaremia.

Sensitivity, specificity, positive and negative predictive values of the Loa loa qPCR, in-house ELISA and Ll-SXP-1-RDT to detect loiasis defined by either a positive life-time history of eyeworm assessed by the RAPLOA questionnaire and/or detectable microfilaremia.</p

Results of exploratory test analysis of microfilaremic loiasis.

Linear regression models showing the association between A) Ct values of the qPCR, B) arbitrary antibody units (AU) of the IgG ELISA and C) reader units (RU) of the Ll-SXP-1-RDT, and L. loa microfilarial density assessed by microscopy. Percentages of test positivity are shown for microfilaremic subgroups in IgG ELISA (D) and the Ll-SXP-1-RDT (E). In A to D, blue dots represent microscopy results of thick blood smears and orange triangles represent results from leukocyte concentration. In Fig 2A, the negative qPCR Ct values were displayed as a value of 42, which is outside of the detection area. Cut-off values for test-positivity are marked by dashed lines.</p

Overview of employed diagnostic procedures.

(A) Diagnostic scheme. Procedures included history of eyeworm (B) assessed by the RAPLOA questionnaire, an in-house Loa loa IgG ELISA using crude adult worm antigen (C), a rapid diagnostic test detecting antibodies against the Ll-SXP-1 antigen (D), direct microfilaria detection including microscopy of Giemsa-stained thick blood smears (E) and saponin lysis (F), as well as a L. loa qPCR (G).</p