This report describes phenanthrene uptake as well as the effect of phenanthrene on the membrane phospholipid and fatty acid composition in a newly isolated bacterial strain, Sphe3, that we taxonomically identified as Arthrobacter sp. Strain Sphe3 is able to utilize phenanthrene as a carbon source at high rates and appears to internalize phenanthrene with two mechanisms: a passive diffusion when cells are grown on glucose, and an inducible active transport system when cells are grown on phenanthrene as a sole carbon source. Active transport followed Michaelis-Menten kinetics, and it was amenable to inhibition by 2,4-dinitrophenol and sodium azide. Evidence provided here indicates that apart from inducing an active PAH uptake, the presence of phenanthrene elicits significant changes in membrane fluidity.

A novel phenanthrene-degrading bacterium, designated strain Sphe3 T , was isolated from a creosote-contaminated soil in Greece. Cells were non-motile, Gram-positive, aerobic, and rod-to coccus-shaped. The strain was isolated on the basis of formation of a clear zone on agar plates sprayed with phenanthrene. Optimal growth occurred at 30 6C. The G+C content of the DNA was 65.7 mol%. The polar lipid pattern of strain Sphe3 T consisted of phosphatidylethanolamine, phosphatidylglycerol and diphosphatidylglycerol. The dominant fatty acids were iso-C 15 : 0 , anteiso-C 15 : 0 , iso-C 16 : 0 , C 16 : 0 and anteiso-C 17 : 0 , representing .86 % of the total fatty acids. The predominant isoprenoid quinone of strain Sphe3 T was menaquinone-8 (MK-8). Based on 16S rRNA gene sequence analysis, strain Sphe3 T showed 99 and 98.9 % similarity to the type strains of Arthrobacter oxydans and Arthrobacter polychromogenes, respectively. Strain Sphe3 T showed 91 % similarity to homologues of A. oxydans and A. polychromogenes based on recA gene sequence analysis. Based on 16S rRNA and recA gene sequence analysis and DNA-DNA hybridization analysis, as well as physiological and chemotaxonomic characteristics, it is concluded that strain Sphe3 T represents a novel species of the genus Arthrobacter, for which the name Arthrobacter phenanthrenivorans sp. nov. is proposed. The type strain is Sphe3 T (5DSM 18606 T 5LMG 23796 T ).Polycyclic aromatic hydrocarbons (PAHs) are environmental pollutants that are found in many polluted soils as a result of natural or industrial activities, including those of creosote wood-treatment facilities (Mueller et al., 1989). PAHs have attracted considerable attention because of their potential toxicity for higher organisms and resistance to microbial degradation (Kanaly & Harayama, 2000). A wide range of micro-organisms have been discovered that are able to degrade highly stable, toxic organic compounds such as polycyclic and aliphatic hydrocarbons (Habe & Omori, 2003;Kanaly & Harayama, 2000;Van Hamme et al., 2003). Among these micro-organisms, several Arthrobacter species are able to degrade PAHs (Grifoll et al., 1992;Seo et al., 2006). We have previously reported the phenanthrene uptake activity and membrane lipid alterations of a PAH-degrading Arthrobacter strain, Sphe3, isolated from a creosote-contaminated soil in Greece (Kallimanis et al., 2007). In the present study, on the basis of phylogenetic analysis of 16S rRNA and recA gene sequences, together with physiological and chemotaxonomic characteristics and DNA-DNA hybridization analysis, we demonstrate that strain Sphe3 represents a novel species of the genus Arthrobacter.Strain Sphe3 T was isolated from Perivleptos, a creosotepolluted site in Epirus, Greece (12 km north of the city of Ioannina), in which a wood-preserving industry had operated for over 30 years (Kallimanis et al., 2007). Soil samples (10 g) were mixed with 100 ml of minimal medium M9 (Sambrook et al., 1989) supplemented with 0.01 % (w/v) phenanthrene (in crystal form) as the sole ca...

Three bacterial strains, designated as Wphe1, Sphe1, and Ophe1, were isolated from Greek soils contaminated with polycyclic aromatic hydrocarbon (PAH)-containing waste from the wood processing, steel, and oil refinery industries. Wphe1, Sphe1, and Ophe1 were characterized and identified as species of Pseudomonas, Microbacterium, and Paracoccus, respectively, based on Gram staining, biochemical tests, phospholipid analysis, FAME analysis, G+C content and 16S rRNA gene sequence analysis. The results of gas chromatography showed that strain Wphe1 degraded naphthalene, phenanthrene, and m-cresol over a wide temperature range; strain Sphe1 was a degrader of phenanthrene and n-alkanes; most interestingly, strain Ophe1 degraded anthracene, phenanthrene, fluorene, fluoranthene, chrysene, and pyrene, as well as cresol compounds and n-alkanes as sole carbon source. This is the first report of a representative of the genus Paracoccus capable of degrading PAHs with such versatility. These three strains may be useful for bioremediation applications.

Arthrobacter phenanthrenivorans is the type species of the genus, and is able to metabolize phenanthrene as a sole source of carbon and energy. A. phenanthrenivorans is an aerobic, non-motile, and Gram-positive bacterium, exhibiting a rod-coccus growth cycle which was originally isolated from a creosote polluted site in Epirus, Greece. Here we describe the features of this organism, together with the complete genome sequence, and annotation.

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Mycobacterium sp.Spyr1 is a newly isolated strain that occurs in a creosote contaminated site in Greece. It was isolated by an enrichment method using pyrene as sole carbon and energy source and is capable of degrading a wide range of PAH substrates including pyrene, fluoranthene, fluorene, anthracene and acenapthene. Here we describe the genomic features of this organism, together with the complete sequence and annotation. The genome consists of a 5,547,747 bp chromosome and two plasmids, a larger and a smaller one with sizes of 211,864 and 23,681 bp, respectively. In total, 5,588 genes were predicted and annotated.

A protein fraction exhibiting 1-hydroxy-2-naphthoic acid (1-H2NA) dioxygenase activity was purified via ion exchange, hydrophobic interactions, and gel filtration chromatography from Arthrobacter phenanthrenivorans sp. nov. strain Sphe3 isolated from a Greek creosote-oil-polluted site. Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) and tandem MS (MS-MS) analysis revealed that the amino acid sequences of oligopeptides of the major 45-kDa protein species, as analyzed by SDS-PAGE and silver staining, comprising 29% of the whole sequence, exhibited strong homology with 1-H2NA dioxygenase of Nocardioides sp. strain KP7. A BLAST search of the recently sequenced Sphe3 genome revealed two putative open reading frames, named diox1 and diox2, showing 90% nucleotide identity to each other and 85% identity at the amino acid level with the Nocardia sp. homologue. diox1 was found on an indigenous Sphe3 plasmid, whereas diox2 was located on the chromosome. Both genes were induced by the presence of phenanthrene used as a sole carbon and energy source, and as expected, both were subject to carbon catabolite repression. The relative RNA transcription level of the chromosomal (diox2) gene was significantly higher than that of its plasmid (diox1) homologue. Both diox1 and diox2 putative genes were PCR amplified, cloned, and overexpressed in Escherichia coli. Recombinant E. coli cells expressed 1-H2NA dioxygenase activity. Recombinant enzymes exhibited Michaelis-Menten kinetics with an apparent K m of 35 M for Diox1 and 29 M for Diox2, whereas they showed similar kinetic turnover characteristics with K cat /K m values of 11 ؋ 10 6 M ؊1 s ؊1 and 12 ؋ 10 6 M ؊1 s ؊1 , respectively. Occurrence of two diox1 and diox2 homologues in the Sphe3 genome implies that a replicative transposition event has contributed to the evolution of 1-H2NA dioxygenase in A. phenanthrenivorans.

BACKGROUND The effects were studied of different inoculation strategies for selected starters –yeasts and lactic acid bacteria (LAB) – used for the fermentation process of two Greek olive cultivars, Conservolea and Kalamàta. The LAB strains applied were Leuconostoc mesenteroides K T5‐1 and L. plantarum A 135–5; the selected yeast strains were S. cerevisiae KI 30–16 and Debaryomyces hansenii A 15–44 for Kalamàta and Conservolea olives, respectively. RESULTS Table olive fermentation processes were monitored by performing microbiological analyses, and by monitoring changes in pH, titratable acidity and salinity, sugar consumption, and the evolution of volatile compounds. Structural modifications occurring in phenolic compounds of brine were investigated during the fermentation using liquid chromatography / diode array detection / electrospray ion trap tandem mass spectrometry (LC/DAD/ESI‐MSn) and quantified by high‐performance liquid chromatography (HPLC) using a diode array detector. Phenolic compounds in processed Kalamàta olive brines consisted of phenolic acids, verbascoside, caffeoyl‐6‐secologanoside, comselogoside, and the dialdehydic form of decarboxymethylelenolic acid linked to hydroxytyrosol, whereas oleoside and oleoside 11‐methyl ester were identified only in Conservolea olive brines. CONCLUSION Volatile profile and sensory evaluation revealed that the ‘MIX’ (co‐inoculum of yeast and LAB strain) inoculation strategy led to the most aromatic and acceptable Kalamàta olives. For the Conservolea table olives, the ‘YL’ treatment gave the most aromatic and the overall most acceptable product. © 2019 Society of Chemical Industry