A Thermostabilized, One-Step PCR Assay for Simultaneous Detection of Klebsiella pneumoniae and Haemophilus influenzae
Klebsiella pneumoniae and Haemophilus influenzae are two common pathogens associated with respiratory tract infections. The identification of these pathogens using conventional molecular diagnostic tests requires trained personnel, cold-chain transportation, and storage-dependance, which does not re...
Published in: | Journal of Tropical Medicine |
---|---|
Main Authors: | , , , , , |
Format: | Article in Journal/Newspaper |
Language: | English |
Published: |
Wiley
2017
|
Subjects: | |
Online Access: | https://doi.org/10.1155/2017/7210849 https://doaj.org/article/8249fd7578824a189a55d1960421a5fb |
id |
ftdoajarticles:oai:doaj.org/article:8249fd7578824a189a55d1960421a5fb |
---|---|
record_format |
openpolar |
spelling |
ftdoajarticles:oai:doaj.org/article:8249fd7578824a189a55d1960421a5fb 2024-09-09T19:26:12+00:00 A Thermostabilized, One-Step PCR Assay for Simultaneous Detection of Klebsiella pneumoniae and Haemophilus influenzae Nur Amalina Khazani Nik Zuraina Nik Mohd Noor Chan Yean Yean Habsah Hasan Siti Suraiya Suharni Mohamad 2017-01-01T00:00:00Z https://doi.org/10.1155/2017/7210849 https://doaj.org/article/8249fd7578824a189a55d1960421a5fb EN eng Wiley http://dx.doi.org/10.1155/2017/7210849 https://doaj.org/toc/1687-9686 https://doaj.org/toc/1687-9694 1687-9686 1687-9694 doi:10.1155/2017/7210849 https://doaj.org/article/8249fd7578824a189a55d1960421a5fb Journal of Tropical Medicine, Vol 2017 (2017) Arctic medicine. Tropical medicine RC955-962 article 2017 ftdoajarticles https://doi.org/10.1155/2017/7210849 2024-08-05T17:48:43Z Klebsiella pneumoniae and Haemophilus influenzae are two common pathogens associated with respiratory tract infections. The identification of these pathogens using conventional molecular diagnostic tests requires trained personnel, cold-chain transportation, and storage-dependance, which does not render them user-friendly. The aim of this study was to develop a thermostabilized, cold-chain-free, one-step multiplex PCR for simultaneous detection of K. pneumoniae and H. influenzae. The multiplex PCR assay was designed to amplify the php gene of K. pneumoniae (202 bp) and p6 gene of H. influenzae (582 bp). In addition, the specific primer to amplify glm gene of Helicobacter pylori (105 bp) was included as an internal amplification control. Subsequently, the designed primers and all PCR reagents were thermostabilized by lyophilization. The stability of the thermostabilized PCR was evaluated using the Q10 method. The sensitivity and specificity of performances for thermostabilized PCR were evaluated using 127 clinical isolates and were found to be 100% sensitive and specific. The thermostabilized PCR mix was found to be stable for 30 days and the Q10 accelerated stability was found to be 3.02 months. A cold-chain-free, PCR assay for easy, rapid, and simultaneous detection of K. pneumoniae and H. influenzae was successfully developed in this study. Article in Journal/Newspaper Arctic Directory of Open Access Journals: DOAJ Articles Arctic Journal of Tropical Medicine 2017 1 8 |
institution |
Open Polar |
collection |
Directory of Open Access Journals: DOAJ Articles |
op_collection_id |
ftdoajarticles |
language |
English |
topic |
Arctic medicine. Tropical medicine RC955-962 |
spellingShingle |
Arctic medicine. Tropical medicine RC955-962 Nur Amalina Khazani Nik Zuraina Nik Mohd Noor Chan Yean Yean Habsah Hasan Siti Suraiya Suharni Mohamad A Thermostabilized, One-Step PCR Assay for Simultaneous Detection of Klebsiella pneumoniae and Haemophilus influenzae |
topic_facet |
Arctic medicine. Tropical medicine RC955-962 |
description |
Klebsiella pneumoniae and Haemophilus influenzae are two common pathogens associated with respiratory tract infections. The identification of these pathogens using conventional molecular diagnostic tests requires trained personnel, cold-chain transportation, and storage-dependance, which does not render them user-friendly. The aim of this study was to develop a thermostabilized, cold-chain-free, one-step multiplex PCR for simultaneous detection of K. pneumoniae and H. influenzae. The multiplex PCR assay was designed to amplify the php gene of K. pneumoniae (202 bp) and p6 gene of H. influenzae (582 bp). In addition, the specific primer to amplify glm gene of Helicobacter pylori (105 bp) was included as an internal amplification control. Subsequently, the designed primers and all PCR reagents were thermostabilized by lyophilization. The stability of the thermostabilized PCR was evaluated using the Q10 method. The sensitivity and specificity of performances for thermostabilized PCR were evaluated using 127 clinical isolates and were found to be 100% sensitive and specific. The thermostabilized PCR mix was found to be stable for 30 days and the Q10 accelerated stability was found to be 3.02 months. A cold-chain-free, PCR assay for easy, rapid, and simultaneous detection of K. pneumoniae and H. influenzae was successfully developed in this study. |
format |
Article in Journal/Newspaper |
author |
Nur Amalina Khazani Nik Zuraina Nik Mohd Noor Chan Yean Yean Habsah Hasan Siti Suraiya Suharni Mohamad |
author_facet |
Nur Amalina Khazani Nik Zuraina Nik Mohd Noor Chan Yean Yean Habsah Hasan Siti Suraiya Suharni Mohamad |
author_sort |
Nur Amalina Khazani |
title |
A Thermostabilized, One-Step PCR Assay for Simultaneous Detection of Klebsiella pneumoniae and Haemophilus influenzae |
title_short |
A Thermostabilized, One-Step PCR Assay for Simultaneous Detection of Klebsiella pneumoniae and Haemophilus influenzae |
title_full |
A Thermostabilized, One-Step PCR Assay for Simultaneous Detection of Klebsiella pneumoniae and Haemophilus influenzae |
title_fullStr |
A Thermostabilized, One-Step PCR Assay for Simultaneous Detection of Klebsiella pneumoniae and Haemophilus influenzae |
title_full_unstemmed |
A Thermostabilized, One-Step PCR Assay for Simultaneous Detection of Klebsiella pneumoniae and Haemophilus influenzae |
title_sort |
thermostabilized, one-step pcr assay for simultaneous detection of klebsiella pneumoniae and haemophilus influenzae |
publisher |
Wiley |
publishDate |
2017 |
url |
https://doi.org/10.1155/2017/7210849 https://doaj.org/article/8249fd7578824a189a55d1960421a5fb |
geographic |
Arctic |
geographic_facet |
Arctic |
genre |
Arctic |
genre_facet |
Arctic |
op_source |
Journal of Tropical Medicine, Vol 2017 (2017) |
op_relation |
http://dx.doi.org/10.1155/2017/7210849 https://doaj.org/toc/1687-9686 https://doaj.org/toc/1687-9694 1687-9686 1687-9694 doi:10.1155/2017/7210849 https://doaj.org/article/8249fd7578824a189a55d1960421a5fb |
op_doi |
https://doi.org/10.1155/2017/7210849 |
container_title |
Journal of Tropical Medicine |
container_volume |
2017 |
container_start_page |
1 |
op_container_end_page |
8 |
_version_ |
1809895868089761792 |