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Genetic approach for the fast discovery of phenazine producing bacteria.

Schneemann I, Wiese J, Kunz AL, Imhoff JF - Mar Drugs (2011)

Bottom Line: The genetic potential for phenazine production was shown for four type strains belonging to the genera Streptomyces and Pseudomonas as well as for 13 environmental isolates from marine habitats.Phenazine production was demonstrated for the type strains known to produce endophenazines, 2-hydroxy-phenazine, phenazine-1-carboxylic acid, phenazine-1,6-dicarboxylic acid, and chlororaphin as well as for members of marine Actinobacteria.Interestingly, a number of unidentified phenazines possibly represent new phenazine structures.

View Article: PubMed Central - PubMed

Affiliation: Kieler Wirkstoff-Zentrum (KiWiZ) am Leibniz-Institut für Meereswissenschaften (IFM-GEOMAR), Am Kiel-Kanal 44, 24106 Kiel, Germany.

ABSTRACT
A fast and efficient approach was established to identify bacteria possessing the potential to biosynthesize phenazines, which are of special interest regarding their antimicrobial activities. Sequences of phzE genes, which are part of the phenazine biosynthetic pathway, were used to design one universal primer system and to analyze the ability of bacteria to produce phenazine. Diverse bacteria from different marine habitats and belonging to six major phylogenetic lines were investigated. Bacteria exhibiting phzE gene fragments affiliated to Firmicutes, Alpha- and Gammaproteobacteria, and Actinobacteria. Thus, these are the first primers for amplifying gene fragments from Firmicutes and Alphaproteobacteria. The genetic potential for phenazine production was shown for four type strains belonging to the genera Streptomyces and Pseudomonas as well as for 13 environmental isolates from marine habitats. For the first time, the genetic ability of phenazine biosynthesis was verified by analyzing the metabolite pattern of all PCR-positive strains via HPLC-UV/MS. Phenazine production was demonstrated for the type strains known to produce endophenazines, 2-hydroxy-phenazine, phenazine-1-carboxylic acid, phenazine-1,6-dicarboxylic acid, and chlororaphin as well as for members of marine Actinobacteria. Interestingly, a number of unidentified phenazines possibly represent new phenazine structures.

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(a) UV/MS-chromatogram (black line: MS; green line: UV at 250 nm) of a 17 day-old GYM agar plate of strain Streptomyces cinnamonensis DSM 1042T. Endophenazine A-C, phenazine-1-carboxylic acid, phenazine-1,6-dicarboxylic acid and an unknown phenazine substance were detected as well as naphterpin and furanonaphthachinon I; (b) UV/MS-chromatogram (black line: MS; green line: UV at 250 nm) of a 17 day-old GYM agar plate of Streptomyces strain LB129. 1-carboxymethyl phenazine and phencomycin methyl ester were detected; (c) UV/MS-chromatogram (black line: MS; green line: UV at 250 nm) of a 17 day-old GYM agar plate of strain HB202. Different streptophenazines and the aromatic polyketide mayamycin were detected.
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f4-marinedrugs-09-00772: (a) UV/MS-chromatogram (black line: MS; green line: UV at 250 nm) of a 17 day-old GYM agar plate of strain Streptomyces cinnamonensis DSM 1042T. Endophenazine A-C, phenazine-1-carboxylic acid, phenazine-1,6-dicarboxylic acid and an unknown phenazine substance were detected as well as naphterpin and furanonaphthachinon I; (b) UV/MS-chromatogram (black line: MS; green line: UV at 250 nm) of a 17 day-old GYM agar plate of Streptomyces strain LB129. 1-carboxymethyl phenazine and phencomycin methyl ester were detected; (c) UV/MS-chromatogram (black line: MS; green line: UV at 250 nm) of a 17 day-old GYM agar plate of strain HB202. Different streptophenazines and the aromatic polyketide mayamycin were detected.

Mentions: To demonstrate the synthesis of phenazines in all phzE positive strains, cultures of these strains were extracted and analyzed by HPLC-UV/MS analyses. 14 out of 17 of these strains were able to produce one or more substances with molecular masses and UV-spectra similar to known phenazines (Table 3, Figure 4a–c). In S. cinnamonensis DSM 1042T the production of endophenazines A–C (Figure 5) and phenazine-1,6-dicarboxylic acid [15] could be demonstrated (Figure 3a). The metabolite chlororaphin was discovered from Pseudomonas chlororaphis subsp. chlororaphis DSM 50083T. 2-hydroxy-phenazine (Figure 5) and phenazine-1-carboxylic acid were produced by Pseudomonas chlororaphis subsp. aureofaciens DSM 6698T and Pseudomonas chlororaphis subsp. aurantiaca DSM 19603T. In addition, the presence of senacarcin A (strain Streptomyces sp. HB117), saphenyl ester D, aestivophoenin C and a derivative thereof (strains Streptomyces sp. HB122 and HB291) as well as phencomycin methyl ester and 1-carboxymethyl phenazine from strain Streptomyces sp. LB129 (Figure 3b) were identified.


Genetic approach for the fast discovery of phenazine producing bacteria.

Schneemann I, Wiese J, Kunz AL, Imhoff JF - Mar Drugs (2011)

(a) UV/MS-chromatogram (black line: MS; green line: UV at 250 nm) of a 17 day-old GYM agar plate of strain Streptomyces cinnamonensis DSM 1042T. Endophenazine A-C, phenazine-1-carboxylic acid, phenazine-1,6-dicarboxylic acid and an unknown phenazine substance were detected as well as naphterpin and furanonaphthachinon I; (b) UV/MS-chromatogram (black line: MS; green line: UV at 250 nm) of a 17 day-old GYM agar plate of Streptomyces strain LB129. 1-carboxymethyl phenazine and phencomycin methyl ester were detected; (c) UV/MS-chromatogram (black line: MS; green line: UV at 250 nm) of a 17 day-old GYM agar plate of strain HB202. Different streptophenazines and the aromatic polyketide mayamycin were detected.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC3111181&req=5

f4-marinedrugs-09-00772: (a) UV/MS-chromatogram (black line: MS; green line: UV at 250 nm) of a 17 day-old GYM agar plate of strain Streptomyces cinnamonensis DSM 1042T. Endophenazine A-C, phenazine-1-carboxylic acid, phenazine-1,6-dicarboxylic acid and an unknown phenazine substance were detected as well as naphterpin and furanonaphthachinon I; (b) UV/MS-chromatogram (black line: MS; green line: UV at 250 nm) of a 17 day-old GYM agar plate of Streptomyces strain LB129. 1-carboxymethyl phenazine and phencomycin methyl ester were detected; (c) UV/MS-chromatogram (black line: MS; green line: UV at 250 nm) of a 17 day-old GYM agar plate of strain HB202. Different streptophenazines and the aromatic polyketide mayamycin were detected.
Mentions: To demonstrate the synthesis of phenazines in all phzE positive strains, cultures of these strains were extracted and analyzed by HPLC-UV/MS analyses. 14 out of 17 of these strains were able to produce one or more substances with molecular masses and UV-spectra similar to known phenazines (Table 3, Figure 4a–c). In S. cinnamonensis DSM 1042T the production of endophenazines A–C (Figure 5) and phenazine-1,6-dicarboxylic acid [15] could be demonstrated (Figure 3a). The metabolite chlororaphin was discovered from Pseudomonas chlororaphis subsp. chlororaphis DSM 50083T. 2-hydroxy-phenazine (Figure 5) and phenazine-1-carboxylic acid were produced by Pseudomonas chlororaphis subsp. aureofaciens DSM 6698T and Pseudomonas chlororaphis subsp. aurantiaca DSM 19603T. In addition, the presence of senacarcin A (strain Streptomyces sp. HB117), saphenyl ester D, aestivophoenin C and a derivative thereof (strains Streptomyces sp. HB122 and HB291) as well as phencomycin methyl ester and 1-carboxymethyl phenazine from strain Streptomyces sp. LB129 (Figure 3b) were identified.

Bottom Line: The genetic potential for phenazine production was shown for four type strains belonging to the genera Streptomyces and Pseudomonas as well as for 13 environmental isolates from marine habitats.Phenazine production was demonstrated for the type strains known to produce endophenazines, 2-hydroxy-phenazine, phenazine-1-carboxylic acid, phenazine-1,6-dicarboxylic acid, and chlororaphin as well as for members of marine Actinobacteria.Interestingly, a number of unidentified phenazines possibly represent new phenazine structures.

View Article: PubMed Central - PubMed

Affiliation: Kieler Wirkstoff-Zentrum (KiWiZ) am Leibniz-Institut für Meereswissenschaften (IFM-GEOMAR), Am Kiel-Kanal 44, 24106 Kiel, Germany.

ABSTRACT
A fast and efficient approach was established to identify bacteria possessing the potential to biosynthesize phenazines, which are of special interest regarding their antimicrobial activities. Sequences of phzE genes, which are part of the phenazine biosynthetic pathway, were used to design one universal primer system and to analyze the ability of bacteria to produce phenazine. Diverse bacteria from different marine habitats and belonging to six major phylogenetic lines were investigated. Bacteria exhibiting phzE gene fragments affiliated to Firmicutes, Alpha- and Gammaproteobacteria, and Actinobacteria. Thus, these are the first primers for amplifying gene fragments from Firmicutes and Alphaproteobacteria. The genetic potential for phenazine production was shown for four type strains belonging to the genera Streptomyces and Pseudomonas as well as for 13 environmental isolates from marine habitats. For the first time, the genetic ability of phenazine biosynthesis was verified by analyzing the metabolite pattern of all PCR-positive strains via HPLC-UV/MS. Phenazine production was demonstrated for the type strains known to produce endophenazines, 2-hydroxy-phenazine, phenazine-1-carboxylic acid, phenazine-1,6-dicarboxylic acid, and chlororaphin as well as for members of marine Actinobacteria. Interestingly, a number of unidentified phenazines possibly represent new phenazine structures.

Show MeSH
Related in: MedlinePlus