The use of High Resolution Melting analysis for reliable
differentiation of trichostrongylid nematodes Haemonchus
contortus and Ashworthius sidemi

a 2020

The use of High Resolution Melting analysis for reliable differentiation of trichostrongylid nematodes Haemonchus contortus and Ashworthius sidemi

ŠKORPÍKOVÁ, Lucie, Nikol RESLOVÁ, Jan MAGDÁLEK, Jaroslav VADLEJCH, Martin KAŠNÝ et. al.

Basic information

Original name

The use of High Resolution Melting analysis for reliable differentiation of trichostrongylid nematodes Haemonchus contortus and Ashworthius sidemi

Authors

ŠKORPÍKOVÁ, Lucie, Nikol RESLOVÁ, Jan MAGDÁLEK, Jaroslav VADLEJCH and Martin KAŠNÝ

Edition

4th COMBAR joint Working Group meeting “Anthelmintic Resistance in Ruminants: from Research to Recommendations” 2020

Other information

Language

English

Type of outcome

Konferenční abstrakt

Field of Study

10613 Zoology

Confidentiality degree

není předmětem státního či obchodního tajemství

References:

abstract

Organization unit

Faculty of Science

Keywords (in Czech)

gastrointestinální nematoda; přežvýkavci; haemonchus contortus; ashworthius sidemi; druhová determinace

Keywords in English

gastrointestinal nematodes; ruminants; haemonchus contortus; ashworthius sidemi; species differentiation

Změněno: 18/1/2021 14:44, Mgr. Lucie Škorpíková, Ph.D.

Abstract

V originále

Infection of ruminants by helminth parasites, especially gastrointestinal nematodes, has worldwide a considerable economic and social impact. From this perspective, correct taxonomical identification is a key prerequisite of initiation of appropriate measures. However, differentiation of some nematode species according to morphological features is complicated, e.g. in the case of important haematophagous abomasal parasites, Haemonchus contortus and Ashworthius sidemi. In this study, we employed qPCR followed by High Resolution Melting (HRM) analysis as an advanced molecular method. This approach enables to detect sequence alterations by increasing temperature. Differences in melting profiles can be visualised as the fluorescence of saturating dye that is gradually disassociated from the dsDNA amplicons. As an output, a difference plot representing the most transparent expression of specific matrix curves were used. The aim of this study was to develop a fast and usable qPCR-HRM assay, which uses polymorphisms in internal transcribed spacer 1 (ITS-1) to distinguish both nematodes without the necessity of additional confirmation by electrophoretic separation and/or sequencing. Based on the specific melting curves, we identified a total of 45 specimens of adult nematodes of H. contortus and A. sidemi that came from a wide range of domestic and wild ruminant hosts that occur in the Czech Republic. We also confirmed that qPCR-HRM analysis is applicable for determination of the infective larval stages of the nematodes, which promises a significant improvement in intravital diagnostics.

Links

LTC19018, research and development project

Name: Pilotní strategický plán boje s rezistentními kmeny parazitických červů v českých hospodářských chovech přežvýkavců (Acronym: Rezistence HELM)

Investor: Ministry of Education, Youth and Sports of the CR, INTER-COST

Citovat

ŠKORPÍKOVÁ, Lucie, Nikol RESLOVÁ, Jan MAGDÁLEK, Jaroslav VADLEJCH and Martin KAŠNÝ. The use of High Resolution Melting analysis for reliable differentiation of trichostrongylid nematodes Haemonchus contortus and Ashworthius sidemi. In 4th COMBAR joint Working Group meeting “Anthelmintic Resistance in Ruminants: from Research to Recommendations”. 2020.

@proceedings{1730057,
   author = {Škorpíková, Lucie and Reslová, Nikol and Magdálek, Jan and Vadlejch, Jaroslav and Kašný, Martin},
   booktitle = {4th COMBAR joint Working Group meeting “Anthelmintic Resistance in Ruminants: from Research to Recommendations”},
   keywords = {gastrointestinal nematodes; ruminants; haemonchus contortus; ashworthius sidemi; species differentiation},
   language = {eng},
   title = {The use of High Resolution Melting analysis for reliable differentiation of trichostrongylid nematodes Haemonchus contortus and Ashworthius sidemi},
   url = {https://www.combar4r.com},
   year = {2020}
}

TY  - CONF
ID  - 1730057
AU  - Škorpíková, Lucie - Reslová, Nikol - Magdálek, Jan - Vadlejch, Jaroslav - Kašný, Martin
PY  - 2020
TI  - The use of High Resolution Melting analysis for reliable differentiation of trichostrongylid nematodes Haemonchus contortus and Ashworthius sidemi
KW  - gastrointestinal nematodes
KW  - ruminants
KW  - haemonchus contortus
KW  - ashworthius sidemi
KW  - species differentiation
UR  - https://www.combar4r.com
N2  - Infection of ruminants by helminth parasites, especially gastrointestinal nematodes, has worldwide a considerable economic and social impact. From this perspective, correct taxonomical identification is a key prerequisite of initiation of appropriate measures. However, differentiation of some nematode species according to morphological features is complicated, e.g. in the case of important haematophagous abomasal parasites, Haemonchus contortus and Ashworthius sidemi. In this study, we employed qPCR followed by High Resolution Melting (HRM) analysis as an advanced molecular method. This approach enables to detect sequence alterations by increasing temperature. Differences in melting profiles can be visualised as the fluorescence of saturating dye that is gradually disassociated from the dsDNA amplicons. As an output, a difference plot representing the most transparent expression of specific matrix curves were used. The aim of this study was to develop a fast and usable qPCR-HRM assay, which uses polymorphisms in internal transcribed spacer 1 (ITS-1) to distinguish both nematodes without the necessity of additional confirmation by electrophoretic separation and/or sequencing. Based on the specific melting curves, we identified a total of 45 specimens of adult nematodes of H. contortus and A. sidemi that came from a wide range of domestic and wild ruminant hosts that occur in the Czech Republic. We also confirmed that qPCR-HRM analysis is applicable for determination of the infective larval stages of the nematodes, which promises a significant improvement in intravital diagnostics.
ER  -

ŠKORPÍKOVÁ, Lucie, Nikol RESLOVÁ, Jan MAGDÁLEK, Jaroslav VADLEJCH and Martin KAŠNÝ. The use of High Resolution Melting analysis for reliable differentiation of trichostrongylid nematodes Haemonchus contortus and Ashworthius sidemi. In \textit{4th COMBAR joint Working Group meeting “Anthelmintic Resistance in Ruminants: from Research to Recommendations”}. 2020.

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