Reclassification of Some Exiguobacterium Species Based on Genome Analysis.

The Exiguobacterium genus comprises Gram-stain-positive and facultatively anaerobic bacteria. Some Exiguobacterium species have previously shown significant high 16S rRNA gene sequence similarities with each other. This study evaluates the taxonomic classification of those Exiguobacterium species through comprehensive genome analysis. Average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) values were determined for various Exiguobacterium species pairs. The ANI and dDDH values between Exiguobacterium enclense and Exiguobacterium indicum, Exiguobacterium aquaticum and Exiguobacterium mexicanum, Exiguobacterium soli and Exiguobacterium antarcticum, and Exiguobacterium sibiricum and Exiguobacterium artemiae were above the cut-off level (95-96% for ANI and 70% for dDDH) for species delineation. Based on the findings, we propose to reclassify Exiguobacterium enclense as a later heterotypic synonym of Exiguobacterium indicum, Exiguobacterium aquaticum as a later heterotypic synonym of Exiguobacterium mexicanum, Exiguobacterium soli as a later heterotypic synonym of Exiguobacterium antarcticum and Exiguobacterium sibiricum as a later heterotypic synonym of Exiguobacterium artemiae.

Energetics of the H-Bond Network in Exiguobacterium sibiricum Rhodopsin.

Exiguobacterium sibiricum rhodopsin (ESR) functions as a light-driven proton pump utilizing Lys96 for proton uptake and maintaining its activity over a wide pH range. Using a combination of methodologies including the linear Poisson-Boltzmann equation and a quantum mechanical/molecular mechanical approach with a polarizable continuum model, we explore the microscopic mechanisms underlying its pumping activity. Lys96, the primary proton uptake site, remains deprotonated owing to the loss of solvation in the ESR protein environment. Asp85, serving as a proton acceptor group for Lys96, does not form a low-barrier H-bond with His57. Instead, deprotonated Asp85 forms a salt-bridge with protonated His57, and the proton is predominantly located at the His57 moiety. Glu214, the only acidic residue at the end of the H-bond network exhibits a pKa value of ∼6, slightly elevated due to solvation loss. It seems likely that the H-bond network [Asp85···His57···H2O···Glu214] serves as a proton-conducting pathway toward the protein bulk surface.

Genomic basis of environmental adaptation in the widespread poly-extremophilic Exiguobacterium group.

Delineating cohesive ecological units and determining the genetic basis for their environmental adaptation are among the most important objectives in microbiology. In the last decade, many studies have been devoted to characterizing the genetic diversity in microbial populations to address these issues. However, the impact of extreme environmental conditions, such as temperature and salinity, on microbial ecology and evolution remains unclear so far. In order to better understand the mechanisms of adaptation, we studied the (pan)genome of Exiguobacterium, a poly-extremophile bacterium able to grow in a wide range of environments, from permafrost to hot springs. To have the genome for all known Exiguobacterium type strains, we first sequenced those that were not yet available. Using a reverse-ecology approach, we showed how the integration of phylogenomic information, genomic features, gene and pathway enrichment data, regulatory element analyses, protein amino acid composition, and protein structure analyses of the entire Exiguobacterium pangenome allows to sharply delineate ecological units consisting of mesophilic, psychrophilic, halophilic-mesophilic, and halophilic-thermophilic ecotypes. This in-depth study clarified the genetic basis of the defined ecotypes and identified some key mechanisms driving the environmental adaptation to extreme environments. Our study points the way to organizing the vast microbial diversity into meaningful ecologically units, which, in turn, provides insight into how microbial communities adapt and respond to different environmental conditions in a changing world.

Improved production of alkaline and solvent-stable proteases from a halotolerant Exiguobacterium isolate through heterologous expression.

Halophiles are excellent sources of detergent proteases that are attributed to stability in alkaline pH, salts, surfactants, and hydrophobic solvents. The lower enzymatic yields and tedious downstream processes necessitate the search for newer halophilic sources. We have previously reported a halotolerant Exiguobacterium sp. TBG-PICH-001, which secretes solvent-tolerant alkaline protease/s. The present study describes the heterologous expression of two protease genes, namely, rsep metalloprotease (WP_195864791, 1.23 Kb) and tpa serine protease (WP_195864453, 0.879 Kb) genes. These were cloned into the pET 22b + plasmid vector and expressed in Escherichia coli BL21(DE3). The recombinant proteases rsep and tpa showed respective yields of 6.3 and 6.7 IU/mg, 11 and 12-fold higher than the crude native protease/s from TBG-PICH-001. These showed soluble expression at 46 and 32 KDa, respectively. These were purified to homogeneity through Ni-NTA-affinity chromatography. The purified proteases were characterized for properties like pH & temperature optima and stability, substrate specificity, kinetic parameters, and detergent attributes. They showed affinity towards various substrates with a respective Km of 392 and 301 µM towards casein. The recombinant proteases exhibited stability in the alkaline pH (7-10), surfactants, metal ions, detergents, and hydrophobic solvents, rendering their suitability as detergent additives.

Evaluation of a novel purified and characterized alkaline protease from the extremophile Exiguobacterium alkaliphilum VLP1 as a detergent additive.

This study focused on the isolation and identification of a novel alkaline protease-producing strain from Lake Van, the largest soda lake on Earth. The objective was to purify, characterize, and investigate the potential application of protease in the detergent industry. Through a combination of classical and molecular methods, the most potent protease producer was identified as Exiguobacterium alkaliphilum VLP1. The purification process, involving ammonium sulfate precipitation, ultrafiltration, and anion exchange chromatography, resulted in a 45-fold purification with a yield of 6.4% and specific activity of 1169 U mg-1 protein. The enzyme exhibited a molecular weight of 69 kDa, a Km value of 0.4 mm, and a maximal velocity (Vmax ) value of 2000 U mg-1 . The optimum activity was observed at 40°C and potential of hydrogen (pH) 9, while the enzyme also exhibited remarkable stability in the ranges of 30-60°C and pH 9-12. Notably, this study represents the first report of an alkaline protease isolated and characterized from E. alkaliphilum. This study also highlighted the potential of the enzyme as a detergent additive, as it showed compatibility with commercial detergents and effectively removed blood and chocolate stains from fabrics.

Pigmented Microbial Extract (PMB) from Exiguobacterium Species MB2 Strain (PMB1) and Bacillus subtilis Strain MB1 (PMB2) Inhibited Breast Cancer Cells Growth In Vivo and In Vitro.

Breast cancer (BC) continues to be one of the major causes of cancer deaths in women. Progress has been made in targeting hormone and growth factor receptor-positive BCs with clinical efficacy and success. However, little progress has been made to develop a clinically viable treatment for the triple-negative BC cases (TNBCs). The current study aims to identify potent agents that can target TNBCs. Extracts from microbial sources have been reported to contain pharmacological agents that can selectively inhibit cancer cell growth. We have screened and identified pigmented microbial extracts (PMBs) that can inhibit BC cell proliferation by targeting legumain (LGMN). LGMN is an oncogenic protein expressed not only in malignant cells but also in tumor microenvironment cells, including tumor-associated macrophages. An LGMN inhibition assay was performed, and microbial extracts were evaluated for in vitro anticancer activity in BC cell lines, angiogenesis assay with chick chorioallantoic membrane (CAM), and tumor xenograft models in Swiss albino mice. We have identified that PMB from the Exiguobacterium (PMB1), inhibits BC growth more potently than PMB2, from the Bacillus subtilis strain. The analysis of PMB1 by GC-MS showed the presence of a variety of fatty acids and fatty-acid derivatives, small molecule phenolics, and aldehydes. PMB1 inhibited the activity of oncogenic legumain in BC cells and induced cell cycle arrest and apoptosis. PMB1 reduced the angiogenesis and inhibited BC cell migration. In mice, intraperitoneal administration of PMB1 retarded the growth of xenografted Ehrlich ascites mammary tumors and mitigated the proliferation of tumor cells in the peritoneal cavity in vivo. In summary, our findings demonstrate the high antitumor potential of PMB1.

Characterization of biofilm formation by Exiguobacterium strains in response to arsenic exposure.

IMPORTANCE: In this work, we characterized the composition, structure, and functional potential for biofilm formation of Exiguobacterium strains isolated from the Salar de Huasco in Chile in the presence of arsenic, an abundant metalloid in the Salar that exists in different oxidation states. Our results showed that the Exiguobacterium strains tested exhibit a significant capacity to form biofilms when exposed to arsenic, which would contribute to their resistance to the metalloid. The results highlight the importance of biofilm formation and the presence of specific resistance mechanisms in the ability of microorganisms to survive and thrive under adverse conditions.

Exiguobacterium sp. as a bioinoculant for plant-growth promotion and Selenium biofortification strategies in horticultural plants.

Plant growth-promoting rhizobacteria (PGPR) have a positive effect on plant development and being a promising way to enhance crop productivity and as substitution of chemical fertilizers. Selenium (Se) is an important trace element and its intake is usually lower than the daily minimum amount required for humans; hence, there is a demand on the design of Se biofortification strategies. Here, the genetic traits known to be associated with Plant-Growth Promotion (PGP) and Se biotransformation of Exiguobacterium sp. S17 were evaluated through genome analysis. Its growth-promoting capacity was tested through plant-growth promotion assays in laboratory and field conditions, using Brassica juncea (indian mustard), Beta vulgaris (chard), and Lactuca sativa (lettuce). Additionally, the Se biotransformation ability of Exiguobacterium sp. S17 was evaluated and the obtained selenized bacteria were tested in mustard plants. The sequenced bacteria genome revealed the presence of multiple genes involved in important functions regarding soil and plant colonization, PGP and Se biotransformation. Moreover, it was demonstrated that Exiguobacterium sp. S17 enhanced plant growth and could be useful to produce Se accumulation and biofortification in accumulator plants such as mustard. Thereby, Exiguobacterium sp. S17 might be used for developing new, sustainable, and environmentally friendly agro-technological strategies.

Valorization of biowastes as fermentative substrate for production of Exiguobacterium sp. GM010 pigment and toxicity effect in rats.

Bioconversion of biowastes chicken feather (CF), prawn carapace (PC), fish scale (FS), and corncob (CC) were used for Exiguobacterium sp. GM010 pigment production to reduce environmental pollution. Maximum pigment was produced in 4 % PC hydrolysate medium at pH 8 and 30 °C (0.831 Absorption Unit-AUmL-1) compared to other hydrolysate. Biomass (1061.19 ± 26.14 mg/100 mL) and pigment yield (34.26 ± 0.62 mg/100 mL) were higher in PC medium. In CF + PC hydrolysate combination, biomass and pigment yield was 890.58 ± 11.5 mg/100 mL and 13.94 ± 0.17 mg/100 mL, respectively. Carbon and nitrogen ratio in the medium influenced pigment production. The UV-visible spectrum showed absorption peak at 357, 466, and 491 nm. Further hue angle (77-72) and chroma values (8.68-11.38) distributed over yellowish-orange region of CIELAB spectrum indicated carotenoid like characteristics. Wistar rats fed with pigment (2000 mg/kg bw) did not show sign of toxicity in haematological, biochemical and histopathological analysis. Therefore, pigment produced by recycling the biowastes promotes sustainable bioprocess and circular bioeconomy.

Production of Endoglucanase by Exiguobacterium mexicanum OB24 Using Waste Melon Peels as Substrate.

Millions of tons of agricultural waste are produced globally every year. A practical solution to this global problem is to convert this waste into value-added products. In this study, endoglucanase enzyme production was carried out by using waste melon peels as a carbon source. To use this important resource, its stubborn structure must be broken down. Rumen bacteria are regarded as unique for this job. Therefore, firstly endoglucanase producing rumen bacteria was isolated and the bacteria with the best activity (OB24) were identified by molecular methods (16S rRNA gene squencing). As a result of the sequence analysis, it was determined that isolate belonged to Exiguobacterium mexicanum. Then, by optimizing the culture conditions, the enzyme production potential was increased. The optimal conditions were determined as 50 g/L MPP, 2g/L yeast extract, 60 h incubation time, pH: 6.0, and 40°C temperature. Under optimized conditions the enzyme activity increased approximately 3.8-fold.

Microbial degradation of low density polyethylene by Exiguobacterium sp. strain LM-IK2 isolated from plastic dumped soil.

Low-Density Polyethylene (LDPE) is one of the significant environmental pollutants as it is resistant to natural degradation. In this study, we reported the LDPE-degrading bacterial strain i.e., Exiguobacterium sp. strain LM-IK2 isolated from plastic dumped soil which shows potential degradation capability. The percent weight loss of LDPE was calculated as - 5.70 ± 0.7 after 90 days of incubation in a carbon-free MSM medium. The Field Emission Scanning Electron Microscopy (FE-SEM) analysis shows that LDPE films show slight surface disruption after treatment with bacteria. The Fourier Transform Infrared Spectroscopy (FTIR) revealed the chemical changes in LDPE films e.g., formation and reduction of typical carbonyl peaks after incubation with bacteria. The X-Ray Diffraction (XRD) analysis displayed an increase in percent crystallinity, with a slight change in total carbon content. Genetic analysis showed the presence of Laccase (167 bp) and Alkane Hydroxylase (330 bp) genes that are responsible for LDPE degradation. Thus, Exiguobacterium sp. strain LM-IK2 has the potential to degrade LDPE and could be further explored to improve its efficiency in the bioremediation of LDPE.

Proton transfer reactions in donor site mutants of ESR, a retinal protein from Exiguobacterium sibiricum.

Light-driven proton transport by microbial retinal proteins such as archaeal bacteriorhodopsin involves carboxylic residues as internal proton donors to the catalytic center which is a retinal Schiff base (SB). The proton donor, Asp96 in bacteriorhodopsin, supplies a proton to the transiently deprotonated Schiff base during the photochemical cycle. Subsequent proton uptake resets the protonated state of the donor. This two step process became a distinctive signature of retinal based proton pumps. Similar steps are observed also in many natural variants of bacterial proteorhodopsins and xanthorhodopsins where glutamic acid residues serve as a proton donor. Recently, however, an exception to this rule was found. A retinal protein from Exiguobacterium sibiricum, ESR, contains a Lys residue in place of Asp or Glu, which facilitates proton transfer from the bulk to the SB. Lys96 can be functionally replaced with the more common donor residues, Asp or Glu. Proton transfer to the SB in the mutants containing these replacements (K96E and K96D/A47T) is much faster than in the proteins lacking the proton donor (K96A and similar mutants), and in the case of K96D/A47T, comparable with that in the wild type, indicating that carboxylic residues can replace Lys96 as proton donors in ESR. We show here that there are important differences in the functioning of these residues in ESR from the way Asp96 functions in bacteriorhodopsin. Reprotonation of the SB and proton uptake from the bulk occur almost simultaneously during the M to N transition (as in the wild type ESR at neutral pH), whereas in bacteriorhodopsin these two steps are well separated in time and occur during the M to N and N to O transitions, respectively.

Study of the Ca2+-dependent gene expression of EuPrt, an extracellular metalloprotease produced by the psychro-tolerant bacterium Exiguobacterium undae Su-1.

The effect of a Ca2+ ion on the gene expression of an on-demand type of metalloprotease from psychrotrophic Exiguobacterium undae Su-1 (EuPrt) was studied. We first established a modified m m9 medium for strain Su-1 to examine its effect in more detail. Then, when the strain was cultured in m m9 medium and 1.0 m m CaCl2 was added, we detected the mature EuPrt and its precursor proteins via Western blotting analysis and found the relative protease activity and its transcription increased by 50-fold and 7-fold, respectively, at the peak. Furthermore, the intracellular concentration of Ca2+ ions was analyzed using inductively coupled plasma atomic emission spectroscopy (ICP-AES) with other metal ions along the growth of strain Su-1. The intracellular concentration of Ca2+ ion was found to increase as much as 3-fold in response to the addition of an extracellular Ca2+ ions, indicating that euPrt gene expression is regulated by sensing its intracellular concentration.

Characterization of chitinase from Exiguobacterium antarcticum and its bioconversion of crayfish shell into chitin oligosaccharides.

Crayfish have become a heavily consumed food and its chitin-rich shell is of great value in terms of waste conversion. This study found a novel chitinase (EaChi40) from a marine bacterium Exiguobacterium antarcticum. The gene was cloned and expressed as a soluble protein of 40 kDa, having optimal activity at pH 6.0 and 30 °C. EaChi40 showed good stability and high specific activity, and kinetic studies found Km and Vmax were 0.86 mg/mL and 13.66 µmol/min/mg. For conversion crayfish shell into oligosaccharides, ball milling and ultrasound-assisted hydrogen peroxide decolorization were applied to pretreat crayfish shell to facilitate its hydrolysis. After the enzymatic conversion, the hydrolysis products of chitobiose and N-acetyl-D-glucosamine were 9.09 mg/mL and 9.21 mg/mL, respectively. EaChi40 efficiently degraded crayfish with a high hydrolysis rate of 76.1%. It is expected to be a good candidate for the production of chitin oligosaccharides in the food and biological fields.

Producción de plásticos biodegradables a partir de bacterias de hábitats salinos aisladas de la Laguna de Ayarza / Production of biodegradable plastics from saline habitats bacteria isolated from Laguna de Ayarza

La contaminación por plásticos petroquímicos es una grave amenaza para el medio ambiente que requiere im-plementar alternativas como los bioplásticos para lograr un desarrollo sostenible. Los polihidroxialcanoatos (PHA) son polímeros utilizados para la producción de plásticos biodegradables y que han llamado la atención como sustitutos de los plásticos de base fósil. Sin embargo, el costo de producción de los PHA constituye una barrera para su producción industrial a gran escala. Las de bacterias de hábitats salinos son microorganismos prometedores para la síntesis de PHA debido a sus características tales como altos requisitos de salinidad que previenen la contaminación microbiana, la alta presión osmótica intracelular que permite una fácil lisis celular para purificar los PHA y la capacidad para usar un amplio espectro de sustratos. La presente investigación planteó determinar las cepas nativas de bacterias halófilas y halotolerantes de la Laguna de Ayarza capaces de producir PHA, establecer la capacidad que tienen de utilizar residuos agrícolas para la producción de PHA y determinar su eficiencia. Esto se logró a través de la inoculación de las cepas productoras de PHA en medios de fermentación con pulpa de café, cáscaras de plátanos y salvado de trigo lo que permitió determinar las cepas más eficientes. Se encontró que las bacterias productoras de PHA pertenecen a las especies: Alcaligenes faecalis, Bacillus idriensis, Bacillus megaterium, Exiguobacterium acetylicum, E. aurantiacum, Pseudomonas cuatrocienegasensis y Sta-phylococcus capitis y que las cepas AP21-14, AP21-10 y AP21-03 mostraron los mejores resultados que podrían ser prometedores para la producción a nivel industrial.

Pollution by petrochemical plastics is a serious threat to the environment that requires the implementation of al-ternatives such as bioplastics to achieve sustainable development. Polyhydroxyalkanoates (PHAs) are polymers used for the production of biodegradable plastics and have drawn attention as substitutes for fossil-based plastics. However, the cost of producing PHAs constitutes a barrier to their large-scale industrial production. Bacteria from saline environments bacteria are promising microorganisms for PHA synthesis due to their characteristics such as high salinity requirements that prevent microbial contamination, high intracellular osmotic pressure that allows easy cell lysis to purify PHAs, and the ability to use a broad spectrum of substrates. This research project aimed to determine the native strains of halophilic and halotolerant bacteria from Laguna de Ayarza capable of producing PHA, establish their ability to use agricultural residues for the production of PHA, and determine their efficiency. This was achieved through the inoculation of the PHA-producing strains in fermentation media with coffee pulp, banana peels and wheat bran, which allowed determining the most efficient strains. It was found that the PHA-producing bacteria belong to the species: Alcaligenes faecalis, Bacillus idriensis, Bacillus mega-terium, Exiguobacterium acetylicum, E. aurantiacum, Pseudomonas cuatrocienegasensis and Staphylococcus capitis and that the strains AP21-14, AP21-10 and AP21-03 showed the best results that could be promising for production at an industrial level.

First Isolation of Exiguobacterium aurantiacum in Serbia.

Exiguobacterium aurantiacum is isolated from a variety of environmental samples but rarely from patients. The aim of the study was to represent isolation of unusual bacterial strains that could cause infection in patients. Final identification was performed using matrix-assisted description/ionization time-of-flight mass spectrometry (MALDI-TOF). Two isolates strains of E. aurantiacum were isolated, one isolate from distilled water used during surgical treatment and the second one from a patient with bacteremia after radical prostatectomy, both sensitive to all tested antimicrobials. Environmental strains could cause infection, especially in immunocompromised patients; therefore, rare bacteria testing is required, in which identification special assistance is provided by an automated system MALDI-TOF.

Reduction of hexavalent chromium by Exiguobacterium mexicanum isolated from chromite mines soil.

Hexavalent chromium is a highly toxic element generated due to indiscriminate chromite mining in Sukinda, Odisha. In the present research investigation a relatively higher Cr(VI) resistant (900 mg L-1) bacterium CWB-54 was isolated from the chromite mine water. Based on the biochemical and molecular analysis the strain (CWB-54) was identified as Exiguobacterium mexicanum. When this bacterium was grown at 35 °C, 100 rpm, pH~8.0, and fructose as an electron donor, it could reduce the total hexavalent chromium (100 mg L-1) supplemented in the medium within 33 h of incubation period. Though experiment was carried out to study the effect of Mn, Ni, Cd, Hg and Zn on Cr(VI) reduction by the strain E. mexicanum it has been observed that in the presence of Cd and Hg, Cr(VI) reduction drastically decreased. Characterization of Cr(VI) reduced product by SEM-EDX and TEM analysis revealed intracellular and extracellular Cr(III) deposition in the bacterium, which is assumed to be Cr(OH)3 precipitate in nanometric size. But the extracellular chromate reductase enzyme production is found to be negligible as compared to the intracellular enzyme production. The increased concentration of Cr(VI) above (1000 mg L-1) also showed the genotoxic effect on the DNA. Several reports have been published on Exiguobacterium sp. on different scientific aspect but the current report on the reduction of toxic Cr(VI) by a new species E. mexicanum is a novel one which established the potentiality of this microorganism for a broad area of application.

Engineering of Thermal Stability in a Cold-Active Oligo-1,6-Glucosidase from Exiguobacterium sibiricum with Unusual Amino Acid Content.

A gene coding for a novel putative amylase, oligo-1,6-glucosidase from a psychrotrophic bacterium Exiguobacterium sibiricum from Siberian permafrost soil was cloned and expressed in Escherichia coli. The amino acid sequence of the predicted protein EsOgl and its 3D model displayed several features characteristic for the cold-active enzymes while possessing an unusually high number of proline residues in the loops-a typical feature of thermophilic enzymes. The activity of the purified recombinant protein was tested with p-nitrophenyl α-D-glucopyranoside as a substrate. The enzyme displayed a plateau-shaped temperature-activity profile with the optimum at 25 °C and a pronounced activity at low temperatures (50% of maximum activity at 5 °C). To improve the thermal stability at temperatures above 40 °C, we have introduced proline residues into four positions of EsOgl by site-directed mutagenesis according to "the proline rule". Two of the mutants, S130P and A109P demonstrated a three- and two-fold increased half-life at 45 °C. Moreover, S130P mutation led to a 60% increase in the catalytic rate constant. Combining the mutations resulted in a further increase in stability transforming the temperature-activity profile to a typical mesophilic pattern. In the most thermostable variant A109P/S130P/E176P, the half-life at 45 °C was increased from 11 min (wild-type) to 129 min.

Exiguobacterium algae sp. nov. and Exiguobacterium qingdaonense sp. nov., two novel moderately halotolerant bacteria isolated from the coastal algae.

Two Gram-stain-positive, facultatively anaerobic, rod-shaped bacterial strains, S126T and S82T, were isolated from coastal algae of China. Strains S126T and S82T are halotolerant and could grow in the presence of 0-13% NaCl and 0-14% NaCl, respectively. The two strains shared 98.9% 16S rRNA gene sequence similarity with each other and 93.4-99.8% similarity with type strains of Exiguobacterium species. The major fatty acids (> 10%) of strains S126T and S82T were iso-C17:0, iso-C13:0, anteiso-C13:0 and iso-C15:0. The predominant quinones of strains S126T and S82T were MK-7 and MK-8. The polar lipid profiles of strain S126T and S82T contained diphosphatidylglycerol, phosphatidylglycerol and phosphatidylethanolamine. The cell-wall peptidoglycans of both strains S126T and S82T were of the A3α L-Lys-Gly type. The average nucleotide identity (ANI) and average nucleotide index (AAI) between strains S126T and S82T and type strains of Exiguobacterium species were all below the thresholds to discriminate bacterial species, indicating that they constitute two novel species in the genus Exiguobacterium. Based on polyphasic taxonomy characterization and genomic aspects, the names Exiguobacterium algae sp. nov. and Exiguobacterium qingdaonense sp. nov. are proposed for the two novel species, with type strains being S126T (= CGMCC 1.17116T = KCTC 43079 T) and S82T (= CGMCC 1.17115T = KCTC 43078T), respectively.

Biodegradation of crude oil by immobilized Exiguobacterium sp. AO-11 and shelf life evaluation.

Exiguobacterium sp. AO-11 was immobilized on bio-cord at 109 CFU g-1 carrier for the removal of crude oil from marine environments. To prepare a ready-to-use bioremediation product, the shelf life of the immobilized cells was calculated. Approximately 90% of 0.25% (v/v) crude oil removal was achieved within 9 days when the starved state of immobilized cells was used. The oil removal activity of the immobilized cells was maintained in the presence of oil dispersant (89%) and at pH values of 7-9. Meanwhile, pH, oil concentration and salinity affected the oil removal efficacy. The immobilized cells could be reused for at least 5 cycles. The Arrhenius equation describing the relationship between the rate of reaction and temperature was validated as a useful model of the kinetics of retention of activity by an immobilized biocatalyst. It was estimated that the immobilized cells could be stored in a non-vacuum bag containing phosphate buffer (pH 7.0) at 30 °C for 39 days to retain the cells at 107 CFU g-1 carrier and more than 50% degradation activity. These results indicated the potential of using bio-cord-immobilized crude oil-degrading Exiguobacterium sp. AO-11 as a bioremediation product in a marine environment.