Detection of Dehalococcoides spp. by Peptide Nucleic Acid Fluorescent in situ Hybridization
Chlorinated solvents including tetrachloroethene (perchloroethene and trichloroethene), are widely used industrial solvents. Improper use and disposal of these chemicals has led to a widespread contamination. Anaerobic treatment technologies that utilize Dehalococcoides spp. can be an effective tool...
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| Veröffentlicht in: | Journal of molecular microbiology and biotechnology 2014-01, Vol.24 (3), p.142-149 |
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| creator | Danko, Anthony S. Fontenete, Silvia J. de Aquino Leite, Daniel Leitão, Patrícia O. Almeida, Carina Schaefer, Charles E. Vainberg, Simon Steffan, Robert J. Azevedo, Nuno F. |
| description | Chlorinated solvents including tetrachloroethene (perchloroethene and trichloroethene), are widely used industrial solvents. Improper use and disposal of these chemicals has led to a widespread contamination. Anaerobic treatment technologies that utilize Dehalococcoides spp. can be an effective tool to remediate these contaminated sites. Therefore, the aim of this study was to develop, optimize and validate peptide nucleic acid (PNA) probes for the detection of Dehalococcoides spp. in both pure and mixed cultures. PNA probes were designed by adapting previously published DNA probes targeting the region of the point mutations described for discriminating between the Dehalococcoides spp. strain CBDB1 and strain 195 lineages. Different fixation, hybridization and washing procedures were tested. The results indicated that the PNA probes hybridized specifically and with a high sensitivity to their corresponding lineages, and that the PNA probes developed during this work can be used in a duplex assay to distinguish between strain CBDB1 and strain 195 lineages, even in complex mixed cultures. This work demonstrates the effectiveness of using PNA fluorescence in situ hybridization to distinguish between two metabolically and genetically distinct Dehalococcoides strains, and they can have strong implications in the monitoring and differentiation of Dehalococcoides populations in laboratory cultures and at contaminated sites. |
| doi_str_mv | 10.1159/000362790 |
| format | Article |
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Improper use and disposal of these chemicals has led to a widespread contamination. Anaerobic treatment technologies that utilize Dehalococcoides spp. can be an effective tool to remediate these contaminated sites. Therefore, the aim of this study was to develop, optimize and validate peptide nucleic acid (PNA) probes for the detection of Dehalococcoides spp. in both pure and mixed cultures. PNA probes were designed by adapting previously published DNA probes targeting the region of the point mutations described for discriminating between the Dehalococcoides spp. strain CBDB1 and strain 195 lineages. Different fixation, hybridization and washing procedures were tested. The results indicated that the PNA probes hybridized specifically and with a high sensitivity to their corresponding lineages, and that the PNA probes developed during this work can be used in a duplex assay to distinguish between strain CBDB1 and strain 195 lineages, even in complex mixed cultures. This work demonstrates the effectiveness of using PNA fluorescence in situ hybridization to distinguish between two metabolically and genetically distinct Dehalococcoides strains, and they can have strong implications in the monitoring and differentiation of Dehalococcoides populations in laboratory cultures and at contaminated sites.</description><identifier>ISSN: 2673-1665</identifier><identifier>ISSN: 1464-1801</identifier><identifier>EISSN: 2673-1673</identifier><identifier>EISSN: 1660-2412</identifier><identifier>DOI: 10.1159/000362790</identifier><identifier>PMID: 24970105</identifier><language>eng</language><publisher>Basel, Switzerland: S. Karger AG</publisher><subject>Anaerobic digestion ; Anaerobic treatment ; Bacteriological Techniques - methods ; Chloroflexi - classification ; Chloroflexi - genetics ; Chloroflexi - isolation & purification ; Dehalococcoides ; Hybridization ; In Situ Hybridization, Fluorescence - methods ; Nucleic acids ; Peptide Nucleic Acids ; Probes ; Research Article ; Sensitivity and Specificity ; Solvents ; Tetrachloroethylene</subject><ispartof>Journal of molecular microbiology and biotechnology, 2014-01, Vol.24 (3), p.142-149</ispartof><rights>2014 S. Karger AG, Basel</rights><rights>2014 S. Karger AG, Basel.</rights><rights>Copyright (c) 2014 S. Karger AG, Basel</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c429t-c18915d81136723b2d34c0ad013f3ff3f4991119d6a3387e5896a95104c04ee13</citedby><cites>FETCH-LOGICAL-c429t-c18915d81136723b2d34c0ad013f3ff3f4991119d6a3387e5896a95104c04ee13</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>314,780,784,2429,27924,27925</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/24970105$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Danko, Anthony S.</creatorcontrib><creatorcontrib>Fontenete, Silvia J.</creatorcontrib><creatorcontrib>de Aquino Leite, Daniel</creatorcontrib><creatorcontrib>Leitão, Patrícia O.</creatorcontrib><creatorcontrib>Almeida, Carina</creatorcontrib><creatorcontrib>Schaefer, Charles E.</creatorcontrib><creatorcontrib>Vainberg, Simon</creatorcontrib><creatorcontrib>Steffan, Robert J.</creatorcontrib><creatorcontrib>Azevedo, Nuno F.</creatorcontrib><title>Detection of Dehalococcoides spp. by Peptide Nucleic Acid Fluorescent in situ Hybridization</title><title>Journal of molecular microbiology and biotechnology</title><addtitle>Microb Physiol</addtitle><description>Chlorinated solvents including tetrachloroethene (perchloroethene and trichloroethene), are widely used industrial solvents. 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This work demonstrates the effectiveness of using PNA fluorescence in situ hybridization to distinguish between two metabolically and genetically distinct Dehalococcoides strains, and they can have strong implications in the monitoring and differentiation of Dehalococcoides populations in laboratory cultures and at contaminated sites.</description><subject>Anaerobic digestion</subject><subject>Anaerobic treatment</subject><subject>Bacteriological Techniques - methods</subject><subject>Chloroflexi - classification</subject><subject>Chloroflexi - genetics</subject><subject>Chloroflexi - isolation & purification</subject><subject>Dehalococcoides</subject><subject>Hybridization</subject><subject>In Situ Hybridization, Fluorescence - methods</subject><subject>Nucleic acids</subject><subject>Peptide Nucleic Acids</subject><subject>Probes</subject><subject>Research Article</subject><subject>Sensitivity and Specificity</subject><subject>Solvents</subject><subject>Tetrachloroethylene</subject><issn>2673-1665</issn><issn>1464-1801</issn><issn>2673-1673</issn><issn>1660-2412</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2014</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><sourceid>ABUWG</sourceid><sourceid>AFKRA</sourceid><sourceid>AZQEC</sourceid><sourceid>BENPR</sourceid><sourceid>CCPQU</sourceid><sourceid>DWQXO</sourceid><sourceid>GNUQQ</sourceid><recordid>eNqF0b1rGzEYBnBRUpqQeuheiiBLOjjRq6-TxuB8Qkg6tFOHQ9a9lyg9n67S3eD89ZWx46FLQEhC_PQg8RDyBdgZgLLnjDGheWXZB3LEdSXmUKaD_V6rQzLL-aUwLoGbSn0ih1zaigFTR-T3JY7oxxB7Glt6ic-uiz56H0ODmeZhOKPLNf2Bw1gO6MPkOwyeXvjQ0Otuigmzx36koac5jBO9XS9TaMKr2yR-Jh9b12Wc7dZj8uv66ufidn7_eHO3uLife8ntOPdgLKjGAAhdcbHkjZCeuYaBaEVbhrQWAGyjnRCmQmWsdlYBK0oigjgmp9vcIcW_E-axXoXyrK5zPcYp11BpowwXwr5PlQLDmQRd6Ml_9CVOqS8fKUpLLbgRsqjvW-VTzDlhWw8prFxa18DqTT_1vp9iv-0Sp-UKm718a6OAr1vwx6UnTHuwu_8PdleQFA</recordid><startdate>20140101</startdate><enddate>20140101</enddate><creator>Danko, Anthony S.</creator><creator>Fontenete, Silvia J.</creator><creator>de Aquino Leite, Daniel</creator><creator>Leitão, Patrícia O.</creator><creator>Almeida, Carina</creator><creator>Schaefer, Charles E.</creator><creator>Vainberg, Simon</creator><creator>Steffan, Robert J.</creator><creator>Azevedo, Nuno F.</creator><general>S. 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Academic</collection><jtitle>Journal of molecular microbiology and biotechnology</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Danko, Anthony S.</au><au>Fontenete, Silvia J.</au><au>de Aquino Leite, Daniel</au><au>Leitão, Patrícia O.</au><au>Almeida, Carina</au><au>Schaefer, Charles E.</au><au>Vainberg, Simon</au><au>Steffan, Robert J.</au><au>Azevedo, Nuno F.</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Detection of Dehalococcoides spp. by Peptide Nucleic Acid Fluorescent in situ Hybridization</atitle><jtitle>Journal of molecular microbiology and biotechnology</jtitle><addtitle>Microb Physiol</addtitle><date>2014-01-01</date><risdate>2014</risdate><volume>24</volume><issue>3</issue><spage>142</spage><epage>149</epage><pages>142-149</pages><issn>2673-1665</issn><issn>1464-1801</issn><eissn>2673-1673</eissn><eissn>1660-2412</eissn><abstract>Chlorinated solvents including tetrachloroethene (perchloroethene and trichloroethene), are widely used industrial solvents. Improper use and disposal of these chemicals has led to a widespread contamination. Anaerobic treatment technologies that utilize Dehalococcoides spp. can be an effective tool to remediate these contaminated sites. Therefore, the aim of this study was to develop, optimize and validate peptide nucleic acid (PNA) probes for the detection of Dehalococcoides spp. in both pure and mixed cultures. PNA probes were designed by adapting previously published DNA probes targeting the region of the point mutations described for discriminating between the Dehalococcoides spp. strain CBDB1 and strain 195 lineages. Different fixation, hybridization and washing procedures were tested. The results indicated that the PNA probes hybridized specifically and with a high sensitivity to their corresponding lineages, and that the PNA probes developed during this work can be used in a duplex assay to distinguish between strain CBDB1 and strain 195 lineages, even in complex mixed cultures. This work demonstrates the effectiveness of using PNA fluorescence in situ hybridization to distinguish between two metabolically and genetically distinct Dehalococcoides strains, and they can have strong implications in the monitoring and differentiation of Dehalococcoides populations in laboratory cultures and at contaminated sites.</abstract><cop>Basel, Switzerland</cop><pub>S. Karger AG</pub><pmid>24970105</pmid><doi>10.1159/000362790</doi><tpages>8</tpages><oa>free_for_read</oa></addata></record> |
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| identifier | ISSN: 2673-1665 |
| ispartof | Journal of molecular microbiology and biotechnology, 2014-01, Vol.24 (3), p.142-149 |
| issn | 2673-1665 1464-1801 2673-1673 1660-2412 |
| language | eng |
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| source | MEDLINE; Karger Journals; Alma/SFX Local Collection |
| subjects | Anaerobic digestion Anaerobic treatment Bacteriological Techniques - methods Chloroflexi - classification Chloroflexi - genetics Chloroflexi - isolation & purification Dehalococcoides Hybridization In Situ Hybridization, Fluorescence - methods Nucleic acids Peptide Nucleic Acids Probes Research Article Sensitivity and Specificity Solvents Tetrachloroethylene |
| title | Detection of Dehalococcoides spp. by Peptide Nucleic Acid Fluorescent in situ Hybridization |
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