Tm-Shift Detection of Dog-Derived Ancylostoma ceylanicum and A. caninum.

Yeqi Fu,Auwalu Yusuf Abdullahi,Yunqiu Liu,Guoqing Li,Xinxin Yan,Jianxiong Hang,Mingwei Wang,Yue Huang,Rongkun Ran,Pan Zhang,Yongxiang Sun

doi:10.1155/2018/7617094

Yeqi Fu, Auwalu Yusuf Abdullahi + Show 9 more

Open Access

https://doi.org/10.1155/2018/7617094

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Abstract

To develop a Tm-shift method for detection of dog-derived Ancylostoma ceylanicum and A. caninum, three sets of primers were designed based on three SNPs (ITS71, ITS197, and ITS296) of their internal transcribed spacer 1 (ITS1) sequences. The detection effect of the Tm-shift was assessed through the stability, sensitivity, accuracy test, and clinical detection. The results showed that these three sets of primers could distinguish accurately between A. ceylanicum and A. caninum. The coefficient of variation in their Tm values on the three SNPs was 0.09% and 0.15% (ITS71), 0.18% and 0.14% (ITS197), and 0.13% and 0.07% (ITS296), respectively. The lowest detectable concentration of standard plasmids for A. ceylanicum and A. caninum was 5.33 × 10−6 ng/μL and 5.03 × 10−6 ng/μL. The Tm-shift results of ten DNA samples from the dog-derived hookworms were consistent with their known species. In the clinical detection of 50 fecal samples from stray dogs, the positive rate of hookworm detected by Tm-shift (42%) was significantly higher than that by microscopic examination (34%), and the former can identify the Ancylostoma species. It is concluded that the Tm-shift method is rapid, specific, sensitive, and suitable for the clinical detection and zoonotic risk assessment of the dog-derived hookworm.

Highlights

Hookworms are common intestinal parasites distributed globally, which can cause serious harm to the health of dogs, cats, and human beings
This study developed a Tm-shift method for detecting A
The amplified internal transcribed spacer 1 (ITS1) fragment from two genomic DNAs of dog-derived A. ceylanicum and A. caninum was 404 bp long (Figure 1) and the generated sequence data were submitted to GenBank

Summary

Introduction

Hookworms are common intestinal parasites distributed globally, which can cause serious harm to the health of dogs, cats, and human beings. There are four species of hookworms, that is, Ancylostoma caninum, A. ceylanicum, A. braziliense, and Uncinaria stenocephala, that can infect dogs [1]. It was reported that A. ceylanicum has become the second most common species of hookworm that can infect humans in Asia [3, 4], especially in Southeast Asian countries such as China [5, 6], Japan [7], Malaysia [8], Laos [9], Thailand [10], and India [11]. A. caninum can reach adulthood in people [1] These two hookworms are zoonotic parasites that can parasitize the host’s intestinal tract and cause severe symptoms such as intestinal bleeding, anemia, malnutrition, and dermatitis [12, 13]. It is very important to establish a rapid and accurate identification method for the two dog-derived hookworms for the prevention and control of hookworm disease

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Conclusion