Optimization of the Helmintex method for schistosomiasis diagnosis

' A diagnostic test that is reliable, sensitive, and applicable in the field is extremely important in epidemiological surveys, during medical treatment for schistosomiasis, and for the control and elimination of schistosomiasis. The Helmintex (HTX) method is based on the use of magnetic beads to trap eggs in a magnetic field. This technique is highly sensitive, but the screening of fecal samples consumes lots of time, thus delaying the results, especially in field studies. The objective of this work was to determine the effects of incorporation of the detergent Tween-20 into the method in an attempt to decrease the final pellet volume produced by the HTX method as well as the use of ninhydrin to stain the Schistosoma mansoni eggs. We showed that these modifications reduced the final volume of the fecal sediment produced in the last step of the HTX method by up to 69% and decreased the screening time to an average of 10.1 min per sample. The use of Tween 20 and ninhydrin led to a high percentage of egg recovery (27.2%). The data obtained herein demonstrate that the addition of detergent and the use of ninhydrin to the HTX process can optimize the screening step and also improve egg recovery, thus justifying the insertion of these steps into the HTX method. (C) 2017 Elsevier Inc. All rights reserved

A new diagnostic strategy which uses a luminol-H2O2 system to detect helminth eggs in fecal sediments processed by the Helmintex method

Schistosomiasis remains a serious public health problem in tropical regions, affecting more than 250 million people. Sensitive diagnostic methods represent key tools for disease elimination, in particular in areas with low endemicity. Advances in the use of luminol-based chemiluminescent techniques have enabled greater sensitivity and speed in obtaining results in different diagnostic settings. In this study, we developed a luminol-H2O2 chemiluminescence (CL) method to detect Schistosoma mansoni eggs in human fecal sediments processed by the Helmintex (HTX) method. After S. mansoni eggs were incubated with a solution of luminol-H2O2 the light emission was detected and measured by spectrophotometry at 431 nm for 5 min, using detection and counts of eggs by bright field optical microscopy as a reference. CL intensity was found to correlate with different sources and numbers of eggs. Furthermore, our results showed that the CL method can distinguish positive from negative samples with 100% sensitivity and 71% specificity. To our knowledge, this is the first study to report the use of CL for the diagnosis of helminths from fecal samples. The combination of the HTX method with CL represents an important advance in providing a reference method with the highest standards of sensitivity

Study of diagnostic accuracy of Helmintex, Kato-Katz, and POC-CCA methods for diagnosing intestinal schistosomiasis in Candeal, a low intensity transmission area in northeastern Brazil

<div><p>Control initiatives have successfully reduced the prevalence and intensity of schistosomiasis transmission in several localities around the world. However, individuals that release low numbers of eggs in their feces may not be detected by classical methods that are limited by low sensitivity. Given that accurate estimates of prevalence are key to implementing planning control actions for the elimination of schistosomiasis, new diagnostic tools are needed to effectively monitor infections and confirm transmission interruption. The World Health Organization recommends the Kato-Katz (KK) thick smear as a parasitological test for epidemiological surveys, even though this method has been demonstrated to underestimate prevalence when egg burdens are low. The point-of-care immunodiagnostic for detecting schistosome cathodic circulating antigen (POC-CCA) method has been proposed as a more sensitive substitute for KK in prevalence estimations. An alternative diagnostic, the Helmintex (HTX) method, isolates eggs from fecal samples with the use of paramagnetic particles in a magnetic field. Here, a population-based study involving 461 individuals from Candeal, Sergipe State, Brazil, was conducted to evaluate these three methods comparatively by latent class analysis (LCA). The prevalence of schistosomiasis mansoni was determined to be 71% with POC-CCA, 40.% with HTX and 11% with KK. Most of the egg burdens of the individuals tested (70%) were < 1 epg, thereby revealing a dissociation between prevalence and intensity in this locality. Therefore, the present results confirm that the HTX method is a highly sensitive egg detection procedure and support its use as a reference method for diagnosing intestinal schistosomiasis and for comparative evaluation of other tests.</p></div

Distribution of POC-CCA method results according to egg burden (epg <1 and ≥1) in 187 samples collected from Candeal, Estância, Sergipe, northeastern Brazil, November 2015.

<p>Distribution of POC-CCA method results according to egg burden (epg <1 and ≥1) in 187 samples collected from Candeal, Estância, Sergipe, northeastern Brazil, November 2015.</p

A comparison of point-of-care cathodic circulating antigen detection in urine (POC-CCA) and Helmintex (HTX) diagnostic methods that were applied to samples collected from the community of Candeal, Estância, Sergipe, northeastern Brazil, November 2015 (n = 461).

<p>For this analysis, POC-CCA resulting in a faint line with at least part of its limits not defined or absent (“trace”) was considered a positive result.</p

A comparison of point-of-care cathodic circulating antigen detection in urine (POC-CCA) and Kato-Katz (KK) diagnostic methods that were applied to samples collected from the community of Candeal, Estância, Sergipe, northeastern Brazil, November 2015 (n = 461).

<p>For this analysis, POC-CCA resulting in a faint line with at least part of its limits not defined or absent (“trace”) was considered a negative result.</p

A comparison of point-of-care cathodic circulating antigen detection in urine (POC-CCA) and Helmintex (HTX) diagnostic methods that were applied to samples collected from the community of Candeal, Estância, Sergipe, northeastern Brazil, November 2015 (n = 461).

<p>For this analysis, POC-CCA resulting in a faint line with at least part of its limits not defined or absent (“trace”) was considered a negative result.</p

A comparison of point-of-care cathodic circulating antigen detection in urine (POC-CCA) and Kato-Katz (KK) diagnostic methods that were applied to samples collected from the community of Candeal, Estância, Sergipe, northeastern Brazil, November 2015 (n = 461).

<p>For this analysis, POC-CCA resulting in a faint line with at least part of its limits not defined or absent (“trace”) was considered a positive result.</p

Schematic illustration of procedures in Helmintex method.

<p>It includes concentration steps from 1 to 8, followed by detection steps (9 and 10): ninhydrin staining and microscopic examination.</p

Egg counting by Helmintex (HTX) and Kato-Katz (KK) methods.

<p>(A) Distribution of HTX-measured faecal egg burdens in the 187 subjects who tested positive and (B) distribution of the KK-measured faecal egg burdens in the 55 subjects who tested positive. Note the logarithmic scale on the horizontal axis.</p