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Comparative Study
. 2017 Jan;101(1):381-390.
doi: 10.1007/s00253-016-7925-6. Epub 2016 Oct 31.

Semi-quantitative measurement of asymptomatic L. infantum infection and symptomatic visceral leishmaniasis in dogs using Dual-Path Platform® CVL

Affiliations
Comparative Study

Semi-quantitative measurement of asymptomatic L. infantum infection and symptomatic visceral leishmaniasis in dogs using Dual-Path Platform® CVL

Mandy Larson et al. Appl Microbiol Biotechnol. 2017 Jan.

Abstract

Infection with Leishmania causes diseases with variable presentation. The most severe form is visceral leishmaniasis (VL), caused by either L. donovani or L. infantum. Despite efforts to eliminate VL, to date, molecular detection in resource-poor settings have lacked the accuracy and rapidity that would enable widespread field use and the need for accurate, sensitive assays to detect asymptomatic Leishmania infection has become apparent. The domestic dog serves as the primary reservoir host of L. infantum. Study of this reservoir population provides an opportunity to evaluate the sensitivity and specificity of diagnostics for well-defined, symptomatic, canine visceral leishmaniasis (CVL) and asymptomatic L. infantum infection. Blood samples from an L. infantum-endemic population of US hunting dogs were evaluated with Dual-Path Platform (DPP®) CVL compared to those obtained via direct detection methods (culture- and Leishmania-specific quantitative polymerase chain reaction, qPCR) and immunofluorescence anti-Leishmania antibody test (IFAT). Statistically significant correlations were found between DPP® CVL development time and clinical status, culture status, circulating DNA levels, and IFAT titer. DPP® CVL results correlated with both clinical severity of disease and serological evidence of asymptomatic L. infantum infection. By precisely documenting the minimum time required for the development of a clear positive result in DPP® CVL, this test could be used in a rapid, semi-quantitative manner for the evaluation of asymptomatic and symptomatic CVL. Our results also indicate that a similar test could be used to improve our understanding of human VL.

Keywords: Diagnosis; Leishmania; Public health; Quantitative; Serology; Zoonotic.

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Conflict of interest statement

Conflict of interest The authors declare that they have no conflict of interest.

Figures

Fig. 1
Fig. 1
Representative DPP®CVL images. DPP® CVL was developed by the addition of blood drops, then running buffer, to a sample portal. Within the cassette test window, a negative sample develops only a single control line (C), while a positive sample develops both the test line (T) and the control line (C)
Fig. 2
Fig. 2
DPP® CVL results correlate with Leishmania culture. Comparison of culture results from eight hunting hounds with known clinical status (scored as 1 if parasites were observed, as 0 if not) against DPP® CVL development time. The plot represents a linear regression fit to the data. Black, polysymptomatic; gray, infected asymptomatic; white, uninfected
Fig. 3
Fig. 3
DPP® CVL development times do not differentiate symptomatic from asymptomatic infected animals. DPP® CVL development time is plotted against clinical status (n = 48). AS asymptomatic, SY symptomatic
Fig. 4
Fig. 4
Faster DPP® CVL development time correlates with circulating parasite DNA levels. Whole blood from 91 hunting hounds was tested with DPP® CVL and parasite DNA quantified by qPCR. DPP® CVL development time is plotted against qPCR Ct value. Black, polysymptomatic; gray, infected asymptomatic; white, uninfected control. The solid line indicates the linear fit, with the hatched lines representing the 95 % confidence intervals
Fig. 5
Fig. 5
DPP® CVL development time correlates with IFAT titer. A panel of a known and b previously unknown canine whole blood samples were subjected to a timed DPP® CVL evaluation. In a, comparison between IFAT result and time to positive DPP® CVL was made using preselected samples from eight dogs with known progression histories across the clinical and IFAT diagnostic continuum. The plot represents linear regression fit to the data. In b, sera from 36 previously unknown status dog samples were analyzed via IFAT and DPP® CVL. The plot represents exponential growth curve fit for the resulting data. IFAT positive cutoff value of 1:64 is designated by the horizontal line. IFAT results stratified to c below the positive threshold (<64) or d above the threshold; comparison was made with DPP® CVL development time. Each plot represents a linear regression fit to the data. Black, polysymptomatic; gray, infected asymptomatic; white, uninfected. The solid line indicates the linear fit, with the hatched lines representing the 95 % confidence intervals

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