Comparison of microscopy and Alamar blue reduction in a larval based assay for schistosome drug screening.

<h4>Background</h4>In view of the current widespread use of and reliance on a single schistosomicide, praziquantel, there is a pressing need to discover and develop alternative drugs for schistosomiasis. One approach to this is to develop High Throughput in vitro whole organism screens (...

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Autores principales:	Nuha R Mansour, Quentin D Bickle
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2010
Materias:	Arctic medicine. Tropical medicine RC955-962 Public aspects of medicine RA1-1270
Acceso en línea:	https://doaj.org/article/76e8253b9086463cbe87591b86e571ae
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spelling	oai:doaj.org-article:76e8253b9086463cbe87591b86e571ae2021-11-18T09:12:23ZComparison of microscopy and Alamar blue reduction in a larval based assay for schistosome drug screening.1935-27271935-273510.1371/journal.pntd.0000795https://doaj.org/article/76e8253b9086463cbe87591b86e571ae2010-08-01T00:00:00Zhttps://www.ncbi.nlm.nih.gov/pmc/articles/pmid/20706580/pdf/?tool=EBIhttps://doaj.org/toc/1935-2727https://doaj.org/toc/1935-2735<h4>Background</h4>In view of the current widespread use of and reliance on a single schistosomicide, praziquantel, there is a pressing need to discover and develop alternative drugs for schistosomiasis. One approach to this is to develop High Throughput in vitro whole organism screens (HTS) to identify hits amongst large compound libraries.<h4>Methodology/principal findings</h4>We have been carrying out low throughput (24-well plate) in vitro testing based on microscopic evaluation of killing of ex-vivo adult S. mansoni worms using selected compound collections mainly provided through the WHO-TDR Helminth Drug Initiative. To increase throughput, we introduced a similar but higher throughput 96-well primary in vitro assay using the schistosomula stage which can be readily produced in vitro in large quantities. In addition to morphological readout of viability we have investigated using fluorometric determination of the reduction of Alamar blue (AB), a redox indicator of enzyme activity widely used in whole organism screening. A panel of 7 known schistosome active compounds including praziquantel, produced diverse effects on larval morphology within 3 days of culture although only two induced marked larval death within 7 days. The AB assay was very effective in detecting these lethal compounds but proved more inconsistent in detecting compounds which damaged but did not kill. The utility of the AB assay in detecting compounds which cause severe morbidity and/or death of schistosomula was confirmed in testing a panel of compounds previously selected in library screening as having activity against the adult worms. Furthermore, in prospective library screening, the AB assay was able to detect all compounds which induced killing and also the majority of compounds designated as hits based on morphological changes.<h4>Conclusion</h4>We conclude that an HTS combining AB readout and image-based analysis would provide an efficient and stringent primary assay for schistosome drug discovery.Nuha R MansourQuentin D BicklePublic Library of Science (PLoS)articleArctic medicine. Tropical medicineRC955-962Public aspects of medicineRA1-1270ENPLoS Neglected Tropical Diseases, Vol 4, Iss 8, p e795 (2010)
institution	DOAJ
collection	DOAJ
language	EN
topic	Arctic medicine. Tropical medicine RC955-962 Public aspects of medicine RA1-1270
spellingShingle	Arctic medicine. Tropical medicine RC955-962 Public aspects of medicine RA1-1270 Nuha R Mansour Quentin D Bickle Comparison of microscopy and Alamar blue reduction in a larval based assay for schistosome drug screening.
description	<h4>Background</h4>In view of the current widespread use of and reliance on a single schistosomicide, praziquantel, there is a pressing need to discover and develop alternative drugs for schistosomiasis. One approach to this is to develop High Throughput in vitro whole organism screens (HTS) to identify hits amongst large compound libraries.<h4>Methodology/principal findings</h4>We have been carrying out low throughput (24-well plate) in vitro testing based on microscopic evaluation of killing of ex-vivo adult S. mansoni worms using selected compound collections mainly provided through the WHO-TDR Helminth Drug Initiative. To increase throughput, we introduced a similar but higher throughput 96-well primary in vitro assay using the schistosomula stage which can be readily produced in vitro in large quantities. In addition to morphological readout of viability we have investigated using fluorometric determination of the reduction of Alamar blue (AB), a redox indicator of enzyme activity widely used in whole organism screening. A panel of 7 known schistosome active compounds including praziquantel, produced diverse effects on larval morphology within 3 days of culture although only two induced marked larval death within 7 days. The AB assay was very effective in detecting these lethal compounds but proved more inconsistent in detecting compounds which damaged but did not kill. The utility of the AB assay in detecting compounds which cause severe morbidity and/or death of schistosomula was confirmed in testing a panel of compounds previously selected in library screening as having activity against the adult worms. Furthermore, in prospective library screening, the AB assay was able to detect all compounds which induced killing and also the majority of compounds designated as hits based on morphological changes.<h4>Conclusion</h4>We conclude that an HTS combining AB readout and image-based analysis would provide an efficient and stringent primary assay for schistosome drug discovery.
format	article
author	Nuha R Mansour Quentin D Bickle
author_facet	Nuha R Mansour Quentin D Bickle
author_sort	Nuha R Mansour
title	Comparison of microscopy and Alamar blue reduction in a larval based assay for schistosome drug screening.
title_short	Comparison of microscopy and Alamar blue reduction in a larval based assay for schistosome drug screening.
title_full	Comparison of microscopy and Alamar blue reduction in a larval based assay for schistosome drug screening.
title_fullStr	Comparison of microscopy and Alamar blue reduction in a larval based assay for schistosome drug screening.
title_full_unstemmed	Comparison of microscopy and Alamar blue reduction in a larval based assay for schistosome drug screening.
title_sort	comparison of microscopy and alamar blue reduction in a larval based assay for schistosome drug screening.
publisher	Public Library of Science (PLoS)
publishDate	2010
url	https://doaj.org/article/76e8253b9086463cbe87591b86e571ae
work_keys_str_mv	AT nuharmansour comparisonofmicroscopyandalamarbluereductioninalarvalbasedassayforschistosomedrugscreening AT quentindbickle comparisonofmicroscopyandalamarbluereductioninalarvalbasedassayforschistosomedrugscreening
_version_	1718420972214681600

Comparison of microscopy and Alamar blue reduction in a larval based assay for schistosome drug screening.

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