Targeted Sequencing of Genomic Repeat Regions Detects Circulating Cell-free Echinococcus DNA

Zhengqing Wan,Binrong Ding,Jieqiong Tan,Lu Ma,Zhuohua Zhang,Jie Ling,Xiaoqing Peng,Qingshan Tian,Weigang Lv,Shizheng Wu,Junqi Li,Klaus Brehm

doi:10.1371/journal.pntd.0008147

Abstract

BackgroundEchinococcosis is a chronic zoonosis caused by tapeworms of the genus Echinococcus. Treatment of the disease is often expensive and complicated, sometimes requiring extensive surgery. Ultrasonographic imaging is currently the main technique for diagnosis, while immunological analysis provides additional information. Confirmation still needs pathological analysis. However, these diagnostic techniques generally detect infection in late stages of the disease. An accurate, early and non-invasive molecular diagnostic method is still unavailable.Methodology/Principal findingsWe sequenced the cell-free DNA (cfDNA) from plasma of echinococcosis patients and confirmed the presence of Echinococcus DNA. To improve detection sensitivity, we developed a method based on targeted next-generation sequencing of repeat regions. Simulation experiments demonstrate that the targeted sequencing is sensitive enough to detect as little as 0.1% of an Echinococcus genome in 1 mL of plasma. Results obtained using patient plasma shows that the Area Under the Curve (AUC) of the method is 0.862, with a detection sensitivity of 62.50% and specificity of 100%, corresponding to a Youden-index of 0.625.Conclusions/SignificanceThis study provides evidence that hydatid cysts release cfDNA fragments into patient plasma. Using the repeat region targeted sequencing method, highly specific detection of Echinococcus infection was achieved. This study paves a new avenue for potential non-invasive screening and diagnosis of echinococcosis.

Highlights

Echinococcosis is a severe parasitic disease that predominantly affects agricultural and pastoral areas, especially in South America, Africa, and Asia [1, 2]
Echinococcosis is a severe chronic parasitic disease caused by tapeworms of the genus Echinococcus
We showed the presence of Echinococcus-derived cell-free DNA in plasma of echinococcosis patients

Summary

Introduction

Echinococcosis is a severe parasitic disease that predominantly affects agricultural and pastoral areas, especially in South America, Africa, and Asia [1, 2]. The etiological agent of echinococcosis are tapeworms of the genus Echinococcus [3]. There are mainly two types of the disease spread in the world, cystic echinococcosis (CE) caused by dog-transmitted E. granulosus and alveolar echinococcosis (AE) caused by fox-transmitted E. multilocularis [2]. Once an individual is infected by E. granulosus, the larvae of the tapeworm develop into fluid-filled cysts in various organs [4]. As cysts grow larger over a period of time, symptoms caused by compression will arise, and permanent damage to affected organs will occur [2, 5]. Echinococcosis is a chronic zoonosis caused by tapeworms of the genus Echinococcus. These diagnostic techniques generally detect infection in late stages of the disease. Early and non-invasive molecular diagnostic method is still unavailable

Methods

Results

Discussion

Conclusion