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Use of green fluorescent protein and luciferase biomarkers to monitor survival and activity of Arthrobacter chlorophenolicus A6 cells during degradation of 4-chlorophenol in soil
Stockholms universitet.
Stockholms universitet.
Södertörns högskola, Avdelning Naturvetenskap. Karolinska Institutet.
Södertörns högskola, Avdelning Naturvetenskap.
2001 (Engelska)Ingår i: Environmental Microbiology, ISSN 1462-2912, E-ISSN 1462-2920, Vol. 3, nr 1, s. 32-42Artikel i tidskrift (Refereegranskat) Published
Abstract [en]

The recently isolated novel species Arthrobacter chlorophenolicus A6 is capable of growth on and degradation of high concentrations of 4-chlorophenol (up to 350 mug ml(-1)) as the sole carbon and energy source, This strain shows promise for bioremediation of environmental sites contaminated with high levels of chlorophenols. In this study, green fluorescent protein (gfp) or luciferase (luc) genes were used as biomarkers for monitoring cell number and activity, respectively, during degradation of 4-chlorophenol by A. chlorophenolicus cells. The individual marked strains, Arthrobacter chlorophenolicus A6L (luc-tagged) and Arthrobacter chlorophenolicus A6G (gfp-tagged), were monitored during degradation of 250 mug ml(-1) 4-chlorophenol in pure culture and 175 mug g(-1) 4-chlorophenol in soil microcosms. Both gene-tagged strains were capable of cleaning up the contaminated soil during 9 d incubation. During the bioremediation experiments, the luc-tagged cells were monitored using luminometry and the gfp tagged cells using flow cytometry, in addition to selective plate counting for both strains. The cells remained at high population levels in the soil (evidenced by GFP-fluorescent cell counts) and the A. chlorophenolicus A6L population was metabolically active (evidenced by luciferase activity measurements). These results demonstrate that the Arthrobacter chlorophenolicus A6 inoculum is effective for cleaning-up soil containing high concentrations of 4-chlorophenol.

Ort, förlag, år, upplaga, sidor
2001. Vol. 3, nr 1, s. 32-42
Nationell ämneskategori
Mikrobiologi
Identifikatorer
URN: urn:nbn:se:sh:diva-17374DOI: 10.1046/j.1462-2920.2001.00156.xISI: 000167056100004PubMedID: 11225721Scopus ID: 2-s2.0-0035202047OAI: oai:DiVA.org:sh-17374DiVA, id: diva2:571373
Tillgänglig från: 2012-11-22 Skapad: 2012-11-19 Senast uppdaterad: 2017-12-07Bibliografiskt granskad
Ingår i avhandling
1. Use of microbiomics to study human impacts on complex microbial communities
Öppna denna publikation i ny flik eller fönster >>Use of microbiomics to study human impacts on complex microbial communities
2006 (Engelska)Doktorsavhandling, sammanläggning (Övrigt vetenskapligt)
Abstract [en]

The study of bacterial communities in nature is currently a challenge. The majority of bacteria in clinical and environmental samples have not yet been cultured and therefore we cannot fully understand their roles in nature and how the ecological balance in a specific microbial ecosystem can be disrupted. For example, exposure to pollutants in soil and antibiotics in the human gut can have large consequences on microbial populations but the magnitude of these impacts is difficult to assess. In this thesis, a combination of molecular techniques, microbiomics, were used to assess complex microbial communities in soil and the human gut. One goal of this thesis was to study the impact of the toxic compound, 4-chlorophenol, on the soil microbiota. In addition, a specific 4-chlorophenol degrading bacterium, Arthrobacter chlorophenolicus, was monitored in soil. In order to monitor the cells they were chromosomally tagged with marker genes encoding either the green fluorescent protein (the gfp gene) or firefly luciferase (the luc gene). During degradation of high levels of 4-chlorophenol in soil, total cells counts of A. chlorophenolicus cells could be measured by flow cytometry (GFP protein) and the metabolic activity could be measured by lurninometry (luciferase activity). In addition, the relative abundance of A. chlorophenolicus in soil could be measured by terminal restriction fragment length polymorphism (T-RFLP) and a higher relative abundance was detected in soil contaminated with 4chlorophenol compared with non-treated soil. The impacts of 4-chlorophenol and A. chlorophenolicus on the dominant members of the soil microbiota were also assessed by T-RFLP. Another goal of this thesis was to study the impact of a short term antibiotic administration in a long term perspective, using either clindamycin, in a two year study or a triple therapy for eradication of Helicobacter pylori containing clarithromycin and metronidazole, in a four year study, on the human fecal microbiota. Both the total bacterial community and specific populations, i.e. Bacteroides spp. and Enterococcus spp., were monitored by T-RFLP. The Bacteroides populations never returned to their pre-treatment composition after clindamycin exposure during the two year study period. Selection and persistence of resistant Bacteroides clones up to two years after treatment was furthermore detected. In the four year study, Enterococcus populations increased as a response to the clarithromycin and metronidazole treatment. An increase in the levels of antibiotic resistance genes, specific erm genes, conferring resistance to macrolides and lincosamides were detected for up to 2 and 4 years after both types of antibiotic treatments in the respective studies. It was also possible to specifically monitor two probiotic Lactobacillus strains and their transient colonization by T-RFLP. In conclusion, the use of a polyphasic approach with complementary analytical tools made it possible to obtain a comprehensive picture of complex microbial communities. In addition, specific bacteria of interest in complex soil and fecal samples could be monitored using microbiomics approaches.

Ort, förlag, år, upplaga, sidor
Stockholm: Karolinska instiutet, 2006. s. 37
Nationell ämneskategori
Biologiska vetenskaper
Identifikatorer
urn:nbn:se:sh:diva-31990 (URN)91-7140-960-2 (ISBN)
Disputation
2006-12-15, MB416, Alfred Nobels allé 7, Huddinge, 10:00 (Engelska)
Opponent
Handledare
Tillgänglig från: 2017-02-08 Skapad: 2017-02-08 Senast uppdaterad: 2017-02-08Bibliografiskt granskad

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Elväng, Annelie M.Westerberg, KarolinaJernberg, CeciliaJansson, Janet K

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