Dataset Information

Molecular diagnosis of scabies using a novel probe-based polymerase chain reaction assay targeting high-copy number repetitive sequences in the Sarcoptes scabiei genome.

ABSTRACT:

Background

The suboptimal sensitivity and specificity of available diagnostic methods for scabies hampers clinical management, trials of new therapies and epidemiologic studies. Additionally, parasitologic diagnosis by microscopic examination of skin scrapings requires sample collection with a sharp scalpel blade, causing discomfort to patients and difficulty in children. Polymerase chain reaction (PCR)-based diagnostic assays, combined with non-invasive sampling methods, represent an attractive approach. In this study, we aimed to develop a real-time probe-based PCR test for scabies, test a non-invasive sampling method and evaluate its diagnostic performance in two clinical settings.

Methodology/principal findings

High copy-number repetitive DNA elements were identified in draft Sarcoptes scabiei genome sequences and used as assay targets for diagnostic PCR. Two suitable repetitive DNA sequences, a 375 base pair microsatellite (SSR5) and a 606 base pair long tandem repeat (SSR6), were identified. Diagnostic sensitivity and specificity were tested using relevant positive and negative control materials and compared to a published assay targeting the mitochondrial cox1 gene. Both assays were positive at a 1:100 dilution of DNA from a single mite; no amplification was observed in DNA from samples from 19 patients with other skin conditions nor from house dust, sheep or dog mites, head and body lice or from six common skin bacterial and fungal species. Moderate sensitivity of the assays was achieved in a pilot study, detecting 5/7 (71.4% [95% CI: 29.0% - 96.3%]) of clinically diagnosed untreated scabies patients). Greater sensitivity was observed in samples collected by FLOQ swabs compared to skin scrapings.

Conclusions/significance

This newly developed qPCR assay, combined with the use of an alternative non-invasive swab sampling technique offers the possibility of enhanced diagnosis of scabies. Further studies will be required to better define the diagnostic performance of these tests.

SUBMITTER: Chng L

PROVIDER: S-EPMC7939366 | biostudies-literature | 2021 Feb

REPOSITORIES: biostudies-literature

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Publications

Molecular diagnosis of scabies using a novel probe-based polymerase chain reaction assay targeting high-copy number repetitive sequences in the Sarcoptes scabiei genome.

Chng Lena L Holt Deborah C DC Field Matt M Francis Joshua R JR Tilakaratne Dev D Dekkers Milou H MH Robinson Greg G Mounsey Kate K Pavlos Rebecca R Bowen Asha C AC Fischer Katja K Papenfuss Anthony T AT Gasser Robin B RB Korhonen Pasi K PK Currie Bart J BJ McCarthy James S JS Pasay Cielo C

PLoS neglected tropical diseases 20210224 2

<h4>Background</h4>The suboptimal sensitivity and specificity of available diagnostic methods for scabies hampers clinical management, trials of new therapies and epidemiologic studies. Additionally, parasitologic diagnosis by microscopic examination of skin scrapings requires sample collection with a sharp scalpel blade, causing discomfort to patients and difficulty in children. Polymerase chain reaction (PCR)-based diagnostic assays, combined with non-invasive sampling methods, represent an at ...[more]

PMID: 33626043

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Project description:BackgroundDebilitating skin infestations caused by the mite, Sarcoptes scabiei, have a profound impact on human and animal health globally. In Australia, this impact is evident across different segments of Australian society, with a growing recognition that it can contribute to rapid declines of native Australian marsupials. Cross-host transmission has been suggested to play a significant role in the epidemiology and origin of mite infestations in different species but a chronic lack of genetic resources has made further inferences difficult. To investigate the origins and molecular epidemiology of S. scabiei in Australian wildlife, we sequenced the mitochondrial genomes of S. scabiei from diseased wombats (Vombatus ursinus) and koalas (Phascolarctos cinereus) spanning New South Wales, Victoria and Tasmania, and compared them with the recently sequenced mitochondrial genome sequences of S. scabiei from humans.ResultsWe found unique S. scabiei haplotypes among individual wombat and koala hosts with high sequence similarity (99.1% - 100%). Phylogenetic analysis of near full-length mitochondrial genomes revealed three clades of S. scabiei (one human and two marsupial), with no apparent geographic or host species pattern, suggestive of multiple introductions. The availability of additional mitochondrial gene sequences also enabled a re-evaluation of a range of putative molecular markers of S. scabiei, revealing that cox1 is the most informative gene for molecular epidemiological investigations. Utilising this gene target, we provide additional evidence to support cross-host transmission between different animal hosts.ConclusionsOur results suggest a history of parasite invasion through colonisation of Australia from hosts across the globe and the potential for cross-host transmission being a common feature of the epidemiology of this neglected pathogen. If this is the case, comparable patterns may exist elsewhere in the 'New World'. This work provides a basis for expanded molecular studies into mange epidemiology in humans and animals in Australia and other geographic regions.

Project description:Scabies is a highly contagious parasitic disease associated with long-term residence in nursing homes, and it is a public health burden worldwide. However, atypical skin manifestations are frequent and the widely used diagnostic test based on microscopic examinations has limited sensitivity. We evaluated the diagnostic value of polymerase chain reaction (PCR) from skin scraping in patients with suspected scabies. Adult patients with suspected scabies, unrelated diseases or healthy volunteers were enrolled at a tertiary hospital, in Seoul, South Korea, from December 2017 through October 2018. We classified participants based on the consensus criteria established by the International Alliance for the Control of Scabies in 2018; confirmed (microscopic mite detection), clinical (scabies burrow or typical lesions with two history features including itch and close contact with scabies patients), suspected scabies (typical lesion with one history feature or atypical lesion with two history features), or no scabies. PCR was performed on the skin scrapings to target the cytochrome c oxidase subunit 1 (cox1) gene of Sarcoptes scabiei. A total of 47 participants, 33 with suspected scabies, 10 with unrelated diseases, and 4 healthy volunteers were enrolled. Of the 33 patients, 22 were classified as confirmed scabies, 2 as clinical scabies, 6 as suspected scabies, and 3 as no scabies. The sensitivities of the microscopic examination were 100%, 92%, and 73% in confirmed scabies; confirmed and clinical scabies; and confirmed, clinical, and suspected scabies, respectively (p = 0.006). The sensitivities of PCR were 86%, 83%, and 80% in confirmed scabies; confirmed and clinical scabies; and confirmed, clinical, and suspected scabies, respectively (p = 0.59). The specificity of the scabies PCR in the no scabies control was 100% (95% CI = 80-100).PCR testing for scabies may be helpful in the improvement of sensitivity for the diagnosis of scabies by clinical criteria.

Dataset Information

Molecular diagnosis of scabies using a novel probe-based polymerase chain reaction assay targeting high-copy number repetitive sequences in the Sarcoptes scabiei genome.

Background

Methodology/principal findings

Conclusions/significance

Publications

Molecular diagnosis of scabies using a novel probe-based polymerase chain reaction assay targeting high-copy number repetitive sequences in the Sarcoptes scabiei genome.

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