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Development and evaluation of molecular tools for detecting and differentiating intestinal amoebae in healthy individuals

Published online by Cambridge University Press:  14 January 2019

Amal Chihi ,
Christen R. Stensvold Open the ORCID record for Christen R. Stensvold [Opens in a new window] ,
Imene Ben-abda ,
Rania Ben-Romdhane ,
Karim Aoun ,
Emna Siala  and
Aïda Bouratbine
Amal Chihi
Affiliation:
Laboratoire de recherche ‘Parasitologie Médicale, Biotechnologies et Biomolécules’, LR 16-IPT-06, Université Tunis El-Manar, Institut Pasteur de Tunis, 13 place Pasteur, B.P. 74 1002 Tunis Belvédère, Tunisia Faculté des Sciences de Bizerte, Université de Carthage, Jarzouna, 7021, Bizerte, Tunisia
Christen R. Stensvold
Affiliation:
Department of Bacteria, Unit of Mycology and Parasitology, Parasites & Fungi, Statens Serum Institut, Artillerivej 5, DK-2300 Copenhagen S, Denmark
Imene Ben-abda
Affiliation:
Laboratoire de recherche ‘Parasitologie Médicale, Biotechnologies et Biomolécules’, LR 16-IPT-06, Université Tunis El-Manar, Institut Pasteur de Tunis, 13 place Pasteur, B.P. 74 1002 Tunis Belvédère, Tunisia Laboratoire de Parasitologie-Mycologie, Institut Pasteur de Tunis, 13, place Pasteur, B.P. 74 1002 Tunis Belvédère, Tunisia
Rania Ben-Romdhane
Affiliation:
Laboratoire de recherche ‘Parasitologie Médicale, Biotechnologies et Biomolécules’, LR 16-IPT-06, Université Tunis El-Manar, Institut Pasteur de Tunis, 13 place Pasteur, B.P. 74 1002 Tunis Belvédère, Tunisia
Karim Aoun
Affiliation:
Laboratoire de recherche ‘Parasitologie Médicale, Biotechnologies et Biomolécules’, LR 16-IPT-06, Université Tunis El-Manar, Institut Pasteur de Tunis, 13 place Pasteur, B.P. 74 1002 Tunis Belvédère, Tunisia
Emna Siala
Affiliation:
Laboratoire de Parasitologie-Mycologie, Institut Pasteur de Tunis, 13, place Pasteur, B.P. 74 1002 Tunis Belvédère, Tunisia
Aïda Bouratbine*
Affiliation:
Laboratoire de recherche ‘Parasitologie Médicale, Biotechnologies et Biomolécules’, LR 16-IPT-06, Université Tunis El-Manar, Institut Pasteur de Tunis, 13 place Pasteur, B.P. 74 1002 Tunis Belvédère, Tunisia Laboratoire de Parasitologie-Mycologie, Institut Pasteur de Tunis, 13, place Pasteur, B.P. 74 1002 Tunis Belvédère, Tunisia
*
Author for correspondence: Aïda Bouratbine, E-mail: aida.bouratbine@pasteur.rns.tn
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Abstract

Amoebae are single-celled parasites frequently colonizing human gut. However, few molecular tools are available for accurate identification. Here, we evaluated a panel of polymerase chain reactions (PCRs) targeting Entamoeba histolytica, Entamoeba dispar, Entamoeba coli, Entamoeba hartmanni, Entamoeba polecki, Endolimax nana and Iodamoeba bütschlii. Thirty-six faecal samples (18 containing at least one amoeba species by microscopy and 18 microscopy negative for amoebae) were tested. Real-time PCRs were used for detection and differentiation of E. histolytica and E. dispar. Conventional PCR with Sanger sequencing were applied for detection and differentiation of E. coli, E. hartmanni, E. polecki, E. nana and I. bütschlii. All microscopy results were confirmed by DNA-based methods. However, more samples were positive for single and mixed amoebic species by DNA-based assays than by microscopy (22 vs 18 and 7 vs 1, respectively). DNA sequencing allowed identification of E. coli subtypes (ST1 and ST2), showed low intra-specific variation within E. hartmanni, identified two phylogenetically distinct groups within E. nana, and identified Iodamoeba at the ribosomal lineage level. Taking into account the high intra-genetic diversity within some of the species at the small subunit (SSU) rRNA gene level, amplification of SSU rRNA genes with subsequent sequencing represents a useful method for detecting, differentiating and subtyping intestinal amoebae.

Keywords

AmoebadiagnosisEndolimaxEntamoebahuman gutIodamoebaparasitePCR
Type
Research Article
Information
Parasitology , Volume 146 , Issue 6 , May 2019 , pp. 821 - 827
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Copyright
Copyright © Cambridge University Press 2019 

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