Assessing the Effect of Chemical Dispersant Nokomis 3-F4 on the Degradation of a Heavy Crude Oil in Water by a Marine Microbial Consortium.

Degradation efficiency of a heavy crude oil by a marine microbial consortium was evaluated in this study, with and without the addition of a chemical dispersant (Nokomis 3-F4). 15.50% of total petroleum hydrocarbons (TPH) were removed after 15 days of incubation without dispersant, with a degradation rate of 2.39 ± 0.22 mg L-1 day-1. In contrast, the addition of Nokomis 3-F4 increased TPH degradation up to 30.81% with a degradation rate of 5.07 ± 0.37 mg L-1 day-1. 16S rRNA gene sequencing indicated a dominance of the consortium by Achromobacter and Alcanivorax. Nonetheless, significant increases in the relative abundance of Martelella and Ochrobactrum were observed with the addition of Nokomis 3-F4. These results will contribute to further environmental studies of the Gulf of Mexico, where Nokomis 3-F4 can be used as chemical dispersant.

Martelella alba sp. nov., isolated from mangrove rhizosphere soil within the Beibu Gulf.

Strain BGMRC 2036T was isolated from rhizosphere soil of Bruguiear gymnorrhiza collected from the Beibu Gulf of China. Optimum growth occurred at 28 °C, pH 7.0, and under the conditions of 3-5% (w/v) NaCl. The phylogenetic comparisons of 16S rRNA gene sequences displayed that strain BGMRC 2036T was closely related to Martelella limonii NBRC109441T (96.6% sequence similarity), M. mediterranea CGMCC 1.12224T (96.5%), M. lutilitoris GH2-6T (96.5%), M. radicis BM5-7T (96.2%), and M. mangrove BM9-1T (95.9%), M. suaedae NBRC109440T (95.8%). The phylogenomic tree based on the up-to-date bacterial core gene set indicated that the strain BGMRC 2036T form a clade formed with members of the genera Martelella. The major polar lipids include phosphatidylmethylethanolamine, phosphatidylglycerol, phosphatidylcholine, phosphotidylinositol, two unidentified phospholipids, and three unidentified ninhydrin positive phospholipids. The major respiratory quinone is Q-10, which is similar to those of genera Martelella. The main cellular fatty acids are C18:1 ω7c, C16:0, and C12:0 aldehyde. Genome sequencing revealed a genome size of 4.99 Mbp and a G + C content of 62.3 mol%. Pairwise comparison of the genomes of the new strain BGMRC 2036T and the three reference strains M. endophytica YC 6887T, M. mediterranea CGMCC 1.12224T, and M. mangrovi USBA-857 indicated that gANI value was lower than 81% and a digital DNA-DNA hybridization value was lower than 27%. The strain BGMRC 2036T possessed genes putatively encoding riboflavin synthesis and flavodoxin A polyphasic taxonomic study suggested that strain BGMRC 2036T represented a novel species belonging to the genus Martelella, and it was named Martelella alba sp. nov. The type strain is BGMRC 2036T (=KCTC 52121T =NBRC 111908T).

Insights from comparative proteomic analysis into degradation of phenanthrene and salt tolerance by the halophilic Martelella strain AD-3.

A halophilic PAHs-degrading strain, Martelella AD-3, was previously isolated from highly saline petroleum-contaminated soil. In this study, label-free proteomics were performed to identify differentially expressed proteins (DEPs) under Group P (phenanthrene +5% salinity) and Group G (glycerol +1% salinity), which would help to reveal the mechanism of phenanthrene degradation and salt tolerance. A total of 307 up-regulated DEPs were found in Group P, including 17 phenanthrene degradation proteins. Among these phenanthrene-degrading proteins, the ferredoxin of aromatic ring-hydroxylating dioxygenase (RHD) was up-regulated by 110-fold and gentisate 1,2-dioxygenases (GDOs) were only expressed in Group P. Besides, we also found nine high salt stress response proteins, including ectoine synthase and transport protein of compatible (osmoprotectant) solutes, were differentially up-regulated. These results indicate that strain AD-3 mainly relied on RHD and dihydrodiol dehydrogenase to degrade phenanthrene, and accumulated compatible solutes for resistance to salt stress. This study provides strong theoretical guidance for understanding the degradation of phenanthrene by strain AD-3 in high salt environments.

Martelella caricis sp. nov., isolated from a rhizosphere mudflat.

A novel marine bacterium, designated GH2-8T, was isolated from a rhizosphere mudflat of a halophyte (Carexscabrifolia) in Gangwha Island, Republic of Korea and its taxonomic status was investigated by a polyphasic approach. Cells of strain GH2-8T were Gram-stain-negative, strictly aerobic, oxidase-positive, catalase-positive and non-motile rods that showed growth at 10-30 °C, pH 5-10 and 0-11 % (w/v) NaCl. The predominant respiratory quinone was Q-10. The polar lipids were phosphatidylcholine, phosphatidylethanolamine, phosphatidylglycerol, an unidentified aminolipid, an unidentified glycolipid and two unidentified lipids. The major fatty acids were summed feature 8, C16 : 0, C19 : 0cyclo ω8c, C18 : 1ω7c 11-methyl and summed feature 2. The G+C content of the genomic DNA was 53.4 mol%. Comparative analysis of the 16S rRNA gene sequences revealed that the organism belonged to the order 'Rhizobiales' and formed a distinct subline at the root of radiation encompassing members of the genus Martelella. The 16S rRNA gene sequence similarities to the phylogenetic neighbours were Martelella mediterranea (97.1 %), Martelella suaedae (96.9 %), Martelella endophytica (96.6 %), Martelella limonii (96.3 %), Martelella mangrovi (96.1 %) and Martelella radicis (95.5 %). Strain GH2-8T showed low 16S rRNA gene sequence similarities (<93.8 %) to other representatives of the order 'Rhizobiales'. On the basis of the results of phenotypic and phylogenetic analyses, strain GH2-8T (=KACC 19402T=KCCM 90275T=KCTC 62126T=NBRC 113213T) is considered to represent a novel species of the genus Martelella for which the name Martelellacaricis sp. nov. is proposed.

Genomic Analysis of Halotolerant Bacterial Strains Martelella soudanensis NC18T and NC20.

Two novel, halotolerant strains of Martelella soudanensis, NC18T and NC20, were isolated from deep subsurface sediment, deeply sequenced, and comparatively analyzed with related strains. Based on a phylogenetic analysis using 16S rRNA gene sequences, the two strains grouped with members of the genus Martelella. Here, we sequenced the complete genomes of NC18T and NC20 to understand the mechanisms of their halotolerance. The genome sizes and G+C content of the strains were 6.1 Mb and 61.8 mol%, respectively. Moreover, NC18T and NC20 were predicted to contain 5,849 and 5,830 genes, and 5,502 and 5,585 protein-coding genes, respectively. Both strains contain the identically predicted 6 rRNAs and 48 tRNAs. The harboring of halotolerant-associated genes revealed that strains NC18T and NC20 might tolerate high salinity through the accumulation of potassium ions in a "salt-in" strategy induced by K+ uptake protein (kup) and the K+ transport system (trkAH and kdpFABC). These two strains also use the ectoine transport system (dctPQM), the glycine betaine transport system (proVWX), and glycine betaine uptake protein (opu) to accumulate "compatible solutes," such as ectoine and glycine betaine, to protect cells from salt stress. This study reveals the halotolerance mechanism of strains NC18T and NC20 in high salt environments and suggests potential applications for these halotolerant and halophilic strains in environmental biotechnology.

Halophilic Martelella sp. AD-3 enhanced phenanthrene degradation in a bioaugmented activated sludge system through syntrophic interaction.

Polycyclic aromatic hydrocarbons (PAHs) are a group of common recalcitrant pollutant in industrial saline wastewater that raised significant concerns, whereas traditional activated sludge (AS) has limited tolerance to high salinity and PAHs toxicity, restricting its capacity to degrade PAHs. It is therefore urgent to develop a bioaugmented sludge (BS) system to aid in the effective degradation of these types of compounds under saline condition. In this study, a novel bioaugmentation strategy was developed by using halophilic Martelella sp. AD-3 for effectively augmented phenanthrene (PHE) degradation under 3% salinity. It was found that a 0.5∼1.5% (w/w) ratio of strain AD-3 to activated sludge was optimal for achieving high PHE degradation activity of the BS system with degradation rates reaching 2.2 mg⋅gVSS-1⋅h-1, nearly 25 times that of the AS system. Although 1-hydroxy-2-naphthoic acid (1H2N) was accumulated obviously, the mineralization of PHE was more complete in the BS system. Reads-based metagenomic coupled metatranscriptomic analysis revealed that the expression values of ndoB, encoding a dioxygenase associated with PHE ring-cleavage, was 5600-fold higher in the BS system than in the AS system. Metagenome assembly showed the members of the Corynebacterium and Alcaligenes genera were abundant in the strain AD-3 bioaugmented BS system with expression of 10.3±1.8% and 1.9±0.26%, respectively. Moreover, phdI and nahG accused for metabolism of 1H2N have been annotated in both above two genera. Degradation assays of intermediates of PHE confirmed that the activated sludge actually possessed considerable degradation capacity for downstream intermediates of PHE including 1H2N. The degradation capacity ratio of 1H2N to PHE was 87% in BS system, while it was 26% in strain AD-3. These results indicated that strain AD-3 contributed mainly in transforming PHE to 1H2N in BS system, while species in activated sludge utilized 1H2N as substrate to grow, thus establishing a syntrophic interaction with strain AD-3 and achieving the complete mineralization of PHE. Long-term continuous experiment confirmed a stable PHE removal efficiency of 93% and few 1H2N accumulation in BS SBR system. This study demonstrated an effective bioaugmented strategy for the bioremediation of saline wastewater containing PAHs.

Isolation and Characterization of an Acidic, Salt-Tolerant Endoglucanase Cel5A from a Bacterial Strain Martelella endophytica YC6887 Genome.

A Martelella endophytica (M. endophytica) strain YC6887 was previously isolated from the roots of a halophyte, Rosa rugosa, which was sequenced and characterized. The genomic and proteomic analysis showed a carbohydrate-degrading enzyme, endoglucanase Cel5A which was further characterized. The protein analysis revealed that this endoglucanase belongs to glycosidic hydrolase family 5 (GH5) with catalytic domain. This gene encodes 349-residue polypeptide and shows closest similarity with cellulases of other Martelella species. The protein was purified to homogeneity and shown that it was a 39 kDa protein. The purified recombinant Cel5A endoglucanase exhibited maximum activity at 50 °C and pH 4.5. The enzyme was salt tolerant and retained more than 50% residual activity up to 15% NaCl. The homology model structure of Cel5A displayed that it is stable and compact protein structure consisting of eleven α-helical structures and eight ß-sheets. According to the predicted ligand binding site after superimposition with Pseudomonas stutzeri endoglucanase Cel5A (PDB ID: 4LX4), it consisted of five amino acid Asn157, Tyr116, Glu158, Glu270 and Trp303 that may be the expected active site of Cel5A from YC6887. This presented that our strain M. endophytica YC6887 that produces cellulase partially degrade the insoluble polysaccharides into reducing sugars.

Characterization of Martelella soudanensis sp. nov., Isolated from a Mine Sediment.

Gram-stain-negative, strictly aerobic, non-spore-forming, non-motile, and rod-shaped bacterial strains, designated NC18T and NC20, were isolated from the sediment near-vertical borehole effluent originating 714 m below the subsurface located in the Soudan Iron Mine in Minnesota, USA. The 16S rRNA gene sequence showed that strains NC18T and NC20 grouped with members of the genus Martelella, including M. mediterranea DSM 17316T and M. limonii YC7034T. The genome sizes and G + C content of both NC18T and NC20 were 6.1 Mb and 61.8 mol%, respectively. Average nucleotide identity (ANI), the average amino acid identity (AAI), and digital DNA-DNA hybridization (dDDH) values were below the species delineation threshold. Pan-genomic analysis showed that NC18T, NC20, M. mediterranea DSM 17316T, M. endophytica YC6887T, and M. lutilitoris GH2-6T had 8470 pan-genome orthologous groups (POGs) in total. Five Martelella strains shared 2258 POG core, which were mainly associated with amino acid transport and metabolism, general function prediction only, carbohydrate transport and metabolism, translation, ribosomal structure and biogenesis, and transcription. The two novel strains had major fatty acids (>5%) including summed feature 8 (C18:1 ω7c and/or C18:1 ω6c), C19:0 cyclo ω8c, C16:0, C18:1 ω7c 11-methyl, C18:0, and summed feature 2 (C12:0 aldehyde and/or iso-C16:1 I and/or C14:0 3-OH). The sole respiratory quinone was uniquinone-10 (Q-10). On the basis of polyphasic taxonomic analyses, strains NC18T and NC20 represent novel species of the genus Martelella, for which the name Martelella soudanensis sp. nov. is proposed. The type strain is NC18T (=KTCT 82174T = NBRC 114661T).

Martelella lutilitoris sp. nov., isolated from a tidal mudflat.

A novel, Gram-stain-negative marine bacterium, designated GH2-6T, was isolated from a rhizosphere mudflat of a halophyte (Carex scabrifolia) collected in Gangwha Island, the Republic of Korea. The cells of the organism were strictly aerobic, oxidase- and catalase-positive, non-flagellated rods. Growth occurred at 20-45°C, pH 5-10, and 0.5-9 (w/v) NaCl. The requirement of Na+ for growth (0.5-3%) was observed. The major respiratory quinone was Q-10. The major polar lipids were phosphatidylcholine, phosphatidylethanolamine, phosphatidylglycerol, an aminolipid and a glycolipid. The predominant fatty acids were C18:1ω7c, C18:0, C16:0, C19:0 cyclo ω8c, C18:1ω7c 11-methyl and summed feature 2 (C14:0 3-OH and/or C16:1 iso I). The genome size was 4.45 Mb and the G+C content of the genomic DNA was 61.9 mol%. Phylogenetic analyses based on 16S rRNA gene sequences revealed that strain GH2-6T belonged to genus Martelella and formed a tight cluster with M. radicis BM5-7T and M. endophytica YC6887T. Levels of 16S rRNA gene sequence similarity between the novel isolate and members of the genus were 99.3-95.5%, but strain GH2-6T possessed an extended loop (49 nucleotides in length) between positions 187 and 213 of the 16S rRNA gene sequence (E. coli numbering). DDH values in vitro between the novel isolate and the closest relatives were 23.2±12.8-46.3±5.2%. On the basis of polyphasic data presented in this study, the type strain GH2-6T (= KACC 19403T = KCTC 62125T = NBRC 113212T) represents a novel species of the genus Martelella for which the name Martelella lutilitoris sp. nov. is proposed.