Assessment of a commercially available multiplex real-time PCR kit against direct immunofluorescence and nested PCRs for the detection of Giardia lamblia, Cryptosporidium spp., and Entamoeba histolytica in sewage

Constantine M. Vassalos; Andre Charlett; Evdokia Vassalou; Anastasia Mpimpa; George Dounias; Georgina Tzanakaki; Athena Mavridou; Gregory Spanakos

doi:10.5004/dwt.2017.20803

Assessment of a commercially available multiplex real-time PCR kit against direct immunofluorescence and nested PCRs for the detection of Giardia lamblia, Cryptosporidium spp., and Entamoeba histolytica in sewage

Constantine M. Vassalos^*, Andre Charlett, Evdokia Vassalou, Anastasia Mpimpa, George Dounias, Georgina Tzanakaki, Athena Mavridou, Gregory Spanakos

^*Corresponding author for this work

Statistics Unit

Research output: Contribution to journal › Article › peer-review

Abstract

The major waterborne protozoan diseases are those caused by Giardia lamblia, Cryptosporidium spp., and Entamoeba histolytica. We studied the performance of a commercial multiplex real-time polymerase chain reaction (MRT-PCR) kit – applied in fecal samples – for the detection of intestinal protozoa in sewage. The MRT-PCR was assessed against direct immunofluorescence assay (DFA); and separate, nested PCRs (nPCRs) for the detection of G. lamblia, Cryptosporidium spp., and E. histolytica. MRT-PCR proved to be highly specific, enabling the detection of E. histolytica and a subset of Cryptosporidium spp. including those mainly responsible for human infections. MRT-PCR was also highly sensitive, finding 10 times more samples contaminated with G. lamblia than DFA. Compared with nPCR for G. lamblia, MRT-PCR was highly accurate. At a cutoff cycle threshold value of 37.6, it showed high sensitivity and specificity in detecting G. lamblia, while reaching substantial agreement with nPCR. Despite variable sensitivity by target DNA, its high specificity made the test a suitable alternative for fast, simultaneous screening for intestinal protozoa of public health importance, revealing co-contamination in five sewage samples. Its high throughput capacity may facilitate informed decision-making for drawing up a sewage monitoring plan and taking appropriate public health measures to minimize the public health risk posed by sewage reuse.

Original language	English
Pages (from-to)	101-111
Number of pages	11
Journal	Desalination and Water Treatment
Volume	91
DOIs	https://doi.org/10.5004/dwt.2017.20803
Publication status	Published - Oct 2017

Bibliographical note

Funding Information:
The authors would like to acknowledge Aftab Jasir, European Centre for Disease Prevention and Control, Stockholm, and Silvia Herrera-León, Instituto de Salud Carlos III, Madrid, for their input to the project proposal and reviewing the manuscript. We would also like to thank Fotis Tsagkarakis, Greek Educational System, Athens, for reviewing mathematical equations and formulas. We also take the opportunity to thank Ioannis G. Koutelekos, Technological Educational Institute, Athens, for his support in preparing the manuscript.

Keywords

Cryptosporidium spp
Cutoff
Entamoeba histolytica
Giardia lamblia
Multiplex real-time PCR
Sewage monitoring

UN SDGs

This output contributes to the following UN Sustainable Development Goals (SDGs)

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10.5004/dwt.2017.20803

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Vassalos, C. M., Charlett, A., Vassalou, E., Mpimpa, A., Dounias, G., Tzanakaki, G., Mavridou, A., & Spanakos, G. (2017). Assessment of a commercially available multiplex real-time PCR kit against direct immunofluorescence and nested PCRs for the detection of Giardia lamblia, Cryptosporidium spp., and Entamoeba histolytica in sewage. Desalination and Water Treatment, 91, 101-111. https://doi.org/10.5004/dwt.2017.20803

@article{aeced52686a847c2bf1afb6d5ef89827,

title = "Assessment of a commercially available multiplex real-time PCR kit against direct immunofluorescence and nested PCRs for the detection of Giardia lamblia, Cryptosporidium spp., and Entamoeba histolytica in sewage",

abstract = "The major waterborne protozoan diseases are those caused by Giardia lamblia, Cryptosporidium spp., and Entamoeba histolytica. We studied the performance of a commercial multiplex real-time polymerase chain reaction (MRT-PCR) kit – applied in fecal samples – for the detection of intestinal protozoa in sewage. The MRT-PCR was assessed against direct immunofluorescence assay (DFA); and separate, nested PCRs (nPCRs) for the detection of G. lamblia, Cryptosporidium spp., and E. histolytica. MRT-PCR proved to be highly specific, enabling the detection of E. histolytica and a subset of Cryptosporidium spp. including those mainly responsible for human infections. MRT-PCR was also highly sensitive, finding 10 times more samples contaminated with G. lamblia than DFA. Compared with nPCR for G. lamblia, MRT-PCR was highly accurate. At a cutoff cycle threshold value of 37.6, it showed high sensitivity and specificity in detecting G. lamblia, while reaching substantial agreement with nPCR. Despite variable sensitivity by target DNA, its high specificity made the test a suitable alternative for fast, simultaneous screening for intestinal protozoa of public health importance, revealing co-contamination in five sewage samples. Its high throughput capacity may facilitate informed decision-making for drawing up a sewage monitoring plan and taking appropriate public health measures to minimize the public health risk posed by sewage reuse.",

keywords = "Cryptosporidium spp, Cutoff, Entamoeba histolytica, Giardia lamblia, Multiplex real-time PCR, Sewage monitoring",

author = "Vassalos, {Constantine M.} and Andre Charlett and Evdokia Vassalou and Anastasia Mpimpa and George Dounias and Georgina Tzanakaki and Athena Mavridou and Gregory Spanakos",

note = "Funding Information: The authors would like to acknowledge Aftab Jasir, European Centre for Disease Prevention and Control, Stockholm, and Silvia Herrera-Le{\'o}n, Instituto de Salud Carlos III, Madrid, for their input to the project proposal and reviewing the manuscript. We would also like to thank Fotis Tsagkarakis, Greek Educational System, Athens, for reviewing mathematical equations and formulas. We also take the opportunity to thank Ioannis G. Koutelekos, Technological Educational Institute, Athens, for his support in preparing the manuscript.",

year = "2017",

month = oct,

doi = "10.5004/dwt.2017.20803",

language = "English",

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Vassalos, CM, Charlett, A, Vassalou, E, Mpimpa, A, Dounias, G, Tzanakaki, G, Mavridou, A & Spanakos, G 2017, 'Assessment of a commercially available multiplex real-time PCR kit against direct immunofluorescence and nested PCRs for the detection of Giardia lamblia, Cryptosporidium spp., and Entamoeba histolytica in sewage', Desalination and Water Treatment, vol. 91, pp. 101-111. https://doi.org/10.5004/dwt.2017.20803

Assessment of a commercially available multiplex real-time PCR kit against direct immunofluorescence and nested PCRs for the detection of Giardia lamblia, Cryptosporidium spp., and Entamoeba histolytica in sewage. / Vassalos, Constantine M.; Charlett, Andre; Vassalou, Evdokia et al.
In: Desalination and Water Treatment, Vol. 91, 10.2017, p. 101-111.

Research output: Contribution to journal › Article › peer-review

TY - JOUR

T1 - Assessment of a commercially available multiplex real-time PCR kit against direct immunofluorescence and nested PCRs for the detection of Giardia lamblia, Cryptosporidium spp., and Entamoeba histolytica in sewage

AU - Vassalos, Constantine M.

AU - Charlett, Andre

AU - Vassalou, Evdokia

AU - Mpimpa, Anastasia

AU - Dounias, George

AU - Tzanakaki, Georgina

AU - Mavridou, Athena

AU - Spanakos, Gregory

N1 - Funding Information: The authors would like to acknowledge Aftab Jasir, European Centre for Disease Prevention and Control, Stockholm, and Silvia Herrera-León, Instituto de Salud Carlos III, Madrid, for their input to the project proposal and reviewing the manuscript. We would also like to thank Fotis Tsagkarakis, Greek Educational System, Athens, for reviewing mathematical equations and formulas. We also take the opportunity to thank Ioannis G. Koutelekos, Technological Educational Institute, Athens, for his support in preparing the manuscript.

PY - 2017/10

Y1 - 2017/10

N2 - The major waterborne protozoan diseases are those caused by Giardia lamblia, Cryptosporidium spp., and Entamoeba histolytica. We studied the performance of a commercial multiplex real-time polymerase chain reaction (MRT-PCR) kit – applied in fecal samples – for the detection of intestinal protozoa in sewage. The MRT-PCR was assessed against direct immunofluorescence assay (DFA); and separate, nested PCRs (nPCRs) for the detection of G. lamblia, Cryptosporidium spp., and E. histolytica. MRT-PCR proved to be highly specific, enabling the detection of E. histolytica and a subset of Cryptosporidium spp. including those mainly responsible for human infections. MRT-PCR was also highly sensitive, finding 10 times more samples contaminated with G. lamblia than DFA. Compared with nPCR for G. lamblia, MRT-PCR was highly accurate. At a cutoff cycle threshold value of 37.6, it showed high sensitivity and specificity in detecting G. lamblia, while reaching substantial agreement with nPCR. Despite variable sensitivity by target DNA, its high specificity made the test a suitable alternative for fast, simultaneous screening for intestinal protozoa of public health importance, revealing co-contamination in five sewage samples. Its high throughput capacity may facilitate informed decision-making for drawing up a sewage monitoring plan and taking appropriate public health measures to minimize the public health risk posed by sewage reuse.

AB - The major waterborne protozoan diseases are those caused by Giardia lamblia, Cryptosporidium spp., and Entamoeba histolytica. We studied the performance of a commercial multiplex real-time polymerase chain reaction (MRT-PCR) kit – applied in fecal samples – for the detection of intestinal protozoa in sewage. The MRT-PCR was assessed against direct immunofluorescence assay (DFA); and separate, nested PCRs (nPCRs) for the detection of G. lamblia, Cryptosporidium spp., and E. histolytica. MRT-PCR proved to be highly specific, enabling the detection of E. histolytica and a subset of Cryptosporidium spp. including those mainly responsible for human infections. MRT-PCR was also highly sensitive, finding 10 times more samples contaminated with G. lamblia than DFA. Compared with nPCR for G. lamblia, MRT-PCR was highly accurate. At a cutoff cycle threshold value of 37.6, it showed high sensitivity and specificity in detecting G. lamblia, while reaching substantial agreement with nPCR. Despite variable sensitivity by target DNA, its high specificity made the test a suitable alternative for fast, simultaneous screening for intestinal protozoa of public health importance, revealing co-contamination in five sewage samples. Its high throughput capacity may facilitate informed decision-making for drawing up a sewage monitoring plan and taking appropriate public health measures to minimize the public health risk posed by sewage reuse.

KW - Cryptosporidium spp

KW - Cutoff

KW - Entamoeba histolytica

KW - Giardia lamblia

KW - Multiplex real-time PCR

KW - Sewage monitoring

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U2 - 10.5004/dwt.2017.20803

DO - 10.5004/dwt.2017.20803

M3 - Article

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SN - 1944-3994

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JO - Desalination and Water Treatment

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ER -

Vassalos CM, Charlett A, Vassalou E, Mpimpa A, Dounias G, Tzanakaki G et al. Assessment of a commercially available multiplex real-time PCR kit against direct immunofluorescence and nested PCRs for the detection of Giardia lamblia, Cryptosporidium spp., and Entamoeba histolytica in sewage. Desalination and Water Treatment. 2017 Oct;91:101-111. doi: 10.5004/dwt.2017.20803

Assessment of a commercially available multiplex real-time PCR kit against direct immunofluorescence and nested PCRs for the detection of Giardia lamblia, Cryptosporidium spp., and Entamoeba histolytica in sewage

Abstract

Bibliographical note

Keywords

UN SDGs

Access to Document

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