Halophilic Bacterium Halomonas Research Articles

Moderately halophilic bacterium Halomonas alkalicola strain Ext as a platform for poly(3-hydroxybutyrate-co-3-hydroxyvalerate) copolymer production with fruit peels residues as sole carbon source

Polyhydroxyalkanoates (PHAs) are synthesized by a variety of microorganisms as intracellular storage granules under imbalanced nutrition and excess carbon. Carbon sources are key contributors to the high cost of PHA production, hence the need for exploration of cheap and sustainable raw materials for bacteria fermentation. In the present study, Halomonas alkalicola strain Ext isolated from a hypersaline lake in Kenya was assessed for its ability to utilize fruit peels hydrolysates (FPH) as sole carbon sources for PHA production. Sugars were extracted from dried peels of banana, mango, orange, and pineapple fruits through mechanical pretreatment and dilute acid hydrolysis. Fruit peels pretreated with 3 % H2SO4 at 121℃ were utilized for shake flask fermentation to produce PHAs by Halomonas alkalicola Ext. At optimal C:N ratios of between 20:1 and 30:1, the bacterium could produce up to 0.45±0.03, 0.394±0.12, 0.39±0.05, and 0.28±0.0 g/L of PHAs from hydrolysates of orange peels, mango peels, banana peels, and pineapple peels, respectively. A maximum PHA content of 16.92 % was achieved on orange peels hydrolysates with 4 % substrate loading. Monomer analysis with gas chromatography-mass spectrometry (GC-MS) revealed that the bacterium produced a copolymer Poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) with 3-hydroxyvalerate (3HV) contents of 5.77 %, 6.08 %, 6.79 %, 6.845 %, for orange peels, pineapple peels, mango peels and banana peels substrates, respectively. The findings of this study suggest that fruit peels waste is a potential feedstock for sustainable production of PHAs by Halomonas alkalicola.

Complete genome sequence of a haloalkaliphilic heterotrophic bacterium Halomonas salifodinae IM328.

The genome of a halophilic bacterium Halomonas salifodinae IM328 was completely sequenced in order to offer convenience for the research such as the synthesis of compatible solutes. The genome contains a circular chromosome which was sequenced by PacBio system.

Arsenite tolerance and removal potential of the indigenous halophilic bacterium, Halomonas elongata SEK2.

The indigenous halophilic arsenite-resistant bacterium Halomonas elongata strain SEK2 isolated from the high saline soil of Malek Mohammad hole, Lut Desert, Iran, could tolerate high concentrations of arsenate (As5+) and arsenite (As3+) up to 800 and 40 mM in the SW-10 agar medium, respectively. The isolated strain was able to tolerate considerable concentrations of other toxic heavy metals and oxyanions, including Cadmium (Cd2+), Chromate (Cr6+), lead (Pb2+), and selenite (Se4+), regarding the high salinity of the culture media (with a total salt concentration of 10% (w/v)), the tolerance potential of the isolate SEK2 was unprecedented. The bioremoval potential of the isolate SEK2 was examined through the Silver diethyldithiocarbamate (SDDC) method and demonstrated that the strain SEK2 could remove 60% of arsenite from arsenite-containing growth medium after 48 h of incubation without converting it to arsenate. The arsenite adsorption or uptake by the halophilic bacterium was investigated and substantiated through Fourier-transform infrared spectroscopy (FTIR), Scanning Electron Microscope (SEM), and Energy Dispersive X-ray (EDX) analyses. Furthermore, Transmission electron microscope (TEM) analysis revealed ultra-structural alterations in the presence of arsenite that could be attributed to intracellular accumulation of arsenite by the bacterial cell. Genome sequencing analysis revealed the presence of arsenite resistance as well as other heavy metals/oxyanion resistance genes in the genome of this bacterial strain. Therefore, Halomonas elongata strain SEK2 was identified as an arsenite-resistant halophilic bacterium for the first time that could be used for arsenite bioremediation in saline arsenite-polluted environments.

Halophilic Plant Growth-Promoting Rhizobacteria as Producers of Antifungal Metabolites under Salt Stress

Salinity is one of the main factors causing soil deterioration, making it unsuitable for agriculture. It is well documented that the application of halotolerant and halophilic plant growth-promoting bacteria (PGPR: plant growth-promoting rhizobacteria) with biological control activities as an inoculant of cultivated plants offers a biological alternative to the use of agrochemicals, particularly when subjected to salt stress. From this perspective, 70 bacterial strains were isolated from saline soils (sebkha) in arid and semi-arid areas of Eastern Algeria. Three isolates were selected based on their ability to produce bioactive molecules allowing them to promote plant growth, such as hydrolytic enzymes, indole acetic acid (auxin-phytohormone), HCN, NH3, etc. Two of these isolates belonged to the genus Serratia and the third was a halophilic Halomonas bacteria. These bacteria were identified based on their 16S rDNA sequences. Antagonism tests against phytopathogenic fungi were carried out. The identification of the antifungal molecules produced by these bacteria was determined using high-performance liquid chromatography. These bacteria can inhibit mycelial development against phytopathogenic fungi with rates reaching 80.67% against Botrytis cinerea, 76.22% against Aspergillus niger, and 66.67% against Fusarium culmorum for Serratia sp. The strain Halomonas sp. inhibited mycelial growth through the production of volatile substances of Aspergillus niger at 71.29%, Aspergillus flavus at 75.49%, and Penicillium glabrum at a rate of 72.22%. The identification of the antifungal molecules produced by these three bacteria using HPLC revealed that they were polyphenols, which makes these strains the first rhizobacteria capable of producing phenolic compounds. Finally, pot tests to determine the effectiveness of these strains in promoting wheat growth under salinity stress (125 mM, 150 mM, and 200 mM) was carried out. The results revealed that a consortium of two isolates (Serratia sp. and Halomonas sp.) performed best at 125 mM. However, at higher concentrations, it was the halophilic bacteria Halomonas sp. that gave the best result. In all cases, there was a significant improvement in the growth of wheat seedlings inoculated with the bacteria compared to non-inoculated controls.

Temporal dynamics of stress response in Halomonas elongata to NaCl shock: physiological, metabolomic, and transcriptomic insights

BackgroundThe halophilic bacterium Halomonas elongata is an industrially important strain for ectoine production, with high value and intense research focus. While existing studies primarily delve into the adaptive mechanisms of this bacterium under fixed salt concentrations, there is a notable dearth of attention regarding its response to fluctuating saline environments. Consequently, the stress response of H. elongata to salt shock remains inadequately understood.ResultsThis study investigated the stress response mechanism of H. elongata when exposed to NaCl shock at short- and long-time scales. Results showed that NaCl shock induced two major stresses, namely osmotic stress and oxidative stress. In response to the former, within the cell’s tolerable range (1–8% NaCl shock), H. elongata urgently balanced the surging osmotic pressure by uptaking sodium and potassium ions and augmenting intracellular amino acid pools, particularly glutamate and glutamine. However, ectoine content started to increase until 20 min post-shock, rapidly becoming the dominant osmoprotectant, and reaching the maximum productivity (1450 ± 99 mg/L/h). Transcriptomic data also confirmed the delayed response in ectoine biosynthesis, and we speculate that this might be attributed to an intracellular energy crisis caused by NaCl shock. In response to oxidative stress, transcription factor cysB was significantly upregulated, positively regulating the sulfur metabolism and cysteine biosynthesis. Furthermore, the upregulation of the crucial peroxidase gene (HELO_RS18165) and the simultaneous enhancement of peroxidase (POD) and catalase (CAT) activities collectively constitute the antioxidant defense in H. elongata following shock. When exceeding the tolerance threshold of H. elongata (1–13% NaCl shock), the sustained compromised energy status, resulting from the pronounced inhibition of the respiratory chain and ATP synthase, may be a crucial factor leading to the stagnation of both cell growth and ectoine biosynthesis.ConclusionsThis study conducted a comprehensive analysis of H. elongata’s stress response to NaCl shock at multiple scales. It extends the understanding of stress response of halophilic bacteria to NaCl shock and provides promising theoretical insights to guide future improvements in optimizing industrial ectoine production.

Optimized Poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) Production by Moderately Haloalkaliphilic Bacterium Halomonas alkalicola Ext

Polyhydroxyalkanoates (PHAs) are biodegradable and biocompatible polymers that are produced by microorganisms as storage materials under limited nutrition and excess carbon. These PHAs have been found to be ideal for replacing synthetic plastics for use in packaging and biomedical applications. In this study, an alkaliphilic and moderately halophilic bacterium Halomonas alkalicola Ext was isolated from Lake Simbi Nyaima in western Kenya and investigated for PHA production. Sudan Black B and Nile Red A staining showed that bacterium had distinct ability for accumulation of PHAs. To optimize PHA production, the bacterium was grown in submerged fermentation under varying culture conditions and different sources and concentrations of carbon and nitrogen. With one-factor-at-a-time (OFTA) approach, optimal PHA yields were obtained after 72 hours at a pH of 10.0, temperature of 35°C, and 2.5% (w/v) NaCl. The bacterium yielded the highest biomass, and PHA amounts on 2% galactose and 0.1% ammonium sulfate as sources of carbon and nitrogen, respectively. A record PHA yield of 0.071 g g-1 with a titer of 1.419±0.09 g/L was achieved from 3.397 g/L of biomass, equivalent to 41.8% PHA content. Using response surface methodology, PHA titer was increased by 1.5% to 1.44 g/L, while PHA content was improved 1.1-fold to 45.57%. Polymer analysis revealed that the extracted PHA was a poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) (3−HB:3−HV=92:8) with two copolymer subunits of 3-hydroxyvaryrate (3-HB) and 3-hydroxybutyrate (3-HV). Halomonas alkalicola Ext attained efficient galactose conversion into PHBV under high salinity and alkalinity conditions.

Structure of the O-polysaccharide from the moderately halophilic bacterium Halomonas fontilapidosi KR26

Lipopolysaccharide was obtained from the aerobic moderately halophilic bacterium Halomonas fontilapidosi KR26. The O-polysaccharide was isolated by mild acid degradation of the lipopolysaccharide and was examined by chemical methods and by 1H and 13C NMR spectroscopy, including 1H,1H COSY, TOCSY, ROESY, and 1H,13C HSQC, and HMBC experiments. The following structure of the linear tetrasaccharide repeating unit was deduced.→2)-α-l-Rhap-(1→2)-α-l-Rhap-(1→3)-α-l-Rhap-(1→3)-β-d-Galp-(1→

Simultaneous conversion of chromium and malachite green coexists in halophilic bacterium Halomonas xianhensis SUR308 isolated from a solar saltern.

Many industries are known to use heavy metals like chromium (Cr) to fix dyes in the fabrication processes and malachite green (MG) as colorant. Alkalinity, elevated temperature, or salinity of the industrial effluents makes conventional physicochemical removal of MG and hexavalent chromium [Cr(VI)] more difficult to apply and demands to perceive potential cost-effective and environment-friendly treatment methods to eliminate or convert them into less toxic compounds. Here, we report simultaneous removal and bioconversion of MG and Cr(VI) by a halophilic biofilm-forming bacterium Halomonas xianhensis SUR308. It can efficiently produce exopolysaccharides as extracellular polymeric substances (EPS) and form biofilm under oxygen limiting condition. The reduction of hexavalent chromium [Cr(VI)] to trivalent chromium [Cr(III)] is about 100%, and 95% after 84 h of growth in shaken and stagnant culture, respectively. The strain completely decolorizes MG after 48 h of growth in shaken culture. Furthermore, we found that strain SUR308 can efficiently detoxify chromium by reduction and degrades MG via producing various intermediate products simultaneously. Most interestingly, such conversions can also take place in alkaline environment and in environment where substantial amount of salt is present. These unique features of strain SUR308 make it suitable for the simultaneous remediation of toxic heavy metals and hazardous dye even from the environment having higher pH and salinity. The detail molecular mechanism of the bioconversion with its application in open environment would be the future research focus for bioprospecting strain SUR308.

Biosynthesis, characterization and optimization of TiO2 nanoparticles by novel marine halophilic Halomonas sp. RAM2: application of natural dye-sensitized solar cells

BackgroundMetal oxide nanoparticles (NPs) are becoming valuable due to their novel applications. The green synthesis of TiO2 NPs is more popular as a flexible and eco-friendly method compared to traditional chemical synthesis methods. TiO2 NPs are the most commonly used semiconductor in dye-sensitized solar cells (DSSCs).ResultsThe biogenic TiO2 NPs were produced extracellularly by the marine halophilic bacterium Halomonas sp. RAM2. Response surface methodology (RSM) was used to optimize the biosynthesis process, resulting in a starting TiO2 concentration of 0.031 M and a pH of 5 for 92 min (⁓15 nm). TiO2 NPs were well-characterized after the calcination process at different temperatures of 500, 600, 700 and 800 °C. Anatase TiO2 NPs (calcined at 500 °C) with a smaller surface area and a wider bandgap were nominated for use in natural dye-sensitized solar cells (NDSSCs). The natural dye used as a photosensitizer is a mixture of three carotenoids extracted from the marine bacterium Kocuria sp. RAM1. NDSSCs were evaluated under standard illumination. After optimization of the counter electrode, NDSSCBio(10) (10 layers) demonstrated the highest photoelectric conversion efficiency (η) of 0.44%, which was almost as good as NDSSCP25 (0.55%).ConclusionThe obtained results confirmed the successful green synthesis of TiO2 NPs and suggested a novel use in combination with bacterial carotenoids in DSSC fabrication, which represents an initial step for further efficiency enhancement studies.

Unsterile production of a polyhydroxyalkanoate copolymer by Halomonas cupida J9

Microbial production of bioplastics polyhydroxyalkanoates (PHA) has opened new avenues to resolve “white pollution” caused by petroleum-based plastics. PHAs consisting of short- and medium-chain-length monomers, designated as SCL-co-MCL PHAs, exhibit much better thermal and mechanical properties than PHA homopolymers. In this study, a halophilic bacterium Halomonas cupida J9 was isolated from highly saline wastewater and proven to produce SCL-co-MCL PHA consisting of 3-hydroxybutyrate (3HB) and 3-hydroxydodecanoate (3HDD) from glucose and glycerol. Whole-genome sequencing and functional annotation suggest that H. cupida J9 may possess three putative PHA biosynthesis pathways and a class I PHA synthase (PhaCJ9). Interestingly, the purified His6-tagged PhaCJ9 from E. coli BL21 (DE3) showed polymerizing activity towards 3HDD-CoA and a phaCJ9-deficient mutant was unable to produce PHA, which indicated that a low-substrate-specificity PhaCJ9 was exclusively responsible for PHA polymerization in H. cupida J9. Docking simulation demonstrated higher binding affinity between 3HB-CoA and PhaCJ9 and identified the key residues involved in hydrogen bonds formation between 3-hydroxyacyl-CoA and PhaCJ9. Furthermore, His489 was identified by site-specific mutagenesis as the key residue for the interaction of 3HDD-CoA with PhaCJ9. Finally, PHA was produced by H. cupida J9 from glucose and glycerol in shake flasks and a 5-L fermentor under unsterile conditions. The open fermentation mode makes this strain a promising candidate for low-cost production of SCL-co-MCL PHAs. Especially, the low-specificity PhaCJ9 has great potential to be engineered for an enlarged substrate range to synthesize tailor-made novel SCL-co-MCL PHAs.

Physiological metabolic topology analysis of Halomonas elongata DSM 2581T in response to sodium chloride stress.

The halophilic bacterium Halomonas elongata DSM 2581T generally adapts well to high level of salinity by biosynthesizing ectoine, which functions as an important compatible solute protecting the cell against external salinity environment. Halophilic bacteria have specific metabolic activities under high-salt conditions and are gradually applied in various industries. The present study focuses on investigating the physiological and metabolic mechanism of H. elongata DSM 2581T driven by the external salinity environment. The physiological metabolic dynamics under salt stress were investigated to evaluate the effect of NaCl stress on the metabolism of H. elongata. The obtained results demonstrated that ectoine biosynthesis transited from a nongrowth-related process to a growth-related process when the NaCl concentration varied from 1% to 13% (w/v). The maximum biomass (Xm = 41.37 g/L), and highest ectoine production (Pm = 12.91 g/L) were achieved under 8% NaCl. Moreover, the maximum biomass (Xm ) and the maximum specific growth rates (μm ) showed a first rising and then declining trend with the increased NaCl stress. Furthermore, the transcriptome analysis of H. elongata under different NaCl concentrations demonstrated that both 8% and 13% NaCl conditions resulted in increased expressions of genes involved in the pentose phosphate pathway, Entner-Doudoroffpathway, flagellar assembly pathway, and ectoine metabolism, but negatively affected the tricarboxylic acidcycle and fatty acid metabolism. At last, the proposed possible adaptation mechanism under the optimum NaCl concentration in H. elongata was described.

Simultaneous nitrification, denitrification and electricity recovery of Halomonas strains in single chamber microbial fuel cells for seawater sewage treatment

Simultaneous nitrification and denitrification (SND) by microbial fuel cell (MFC) holds great promise in the nitrogen removal from seawater sewage due to the harvest of electricity and low organic carbon requirement. However, this remains very difficult owing to the inhibition of microbial growth and metabolism by the high salinity. In this work, a halophilic bacterium Halomonas sp. B01 with both exoelectrogenic and SND capability in hypersaline environment was reported. Halomonas sp. B01 was screened from several Halomonas species considering their growth and electricity production abilities. Halomonas sp. B01 generated a high power density of 24.2 mW/m2 (0.61 W/m3) and efficient N removal rate of 93.8% in single chamber MFCs on seawater substrate. The nitrogen removal pathway of Halomonas sp. B01 was demonstrated to include (1) nitrification process coupled with electron-donating to the anode, and (2) denitrification process coupled with electron-accepting from the cathode. The present work represents a successful attempt to accomplish simultaneous SND and electricity production in MFCs by a single halophilic strain, which provides a theoretical and method support for the convenient treatment of seawater sewage.

Optimization of Nutritional and Operational Conditions for Producing PHA by the Halophilic Bacterium Halomonas boliviensis from Oil Palm Empty Fruit Bunch and Gluten Hydrolysates

Optimization of Nutritional and Operational Conditions for Producing PHA by the Halophilic Bacterium Halomonas boliviensis from Oil Palm Empty Fruit Bunch and Gluten Hydrolysates

Genome analysis of a halophilic bacterium Halomonas malpeensis YU-PRIM-29T reveals its exopolysaccharide and pigment producing capabilities

Halomonas malpeensis strain YU-PRIM-29T is a yellow pigmented, exopolysaccharide (EPS) producing halophilic bacterium isolated from the coastal region. To understand the biosynthesis pathways involved in the EPS and pigment production, whole genome analysis was performed. The complete genome sequencing and the de novo assembly were carried out using Illumina sequencing and SPAdes genome assembler (ver 3.11.1) respectively followed by detailed genome annotation. The genome consists of 3,607,821 bp distributed in 18 contigs with 3337 protein coding genes and 53% of the annotated CDS are having putative functions. Gene annotation disclosed the presence of genes involved in ABC transporter-dependent pathway of EPS biosynthesis. As the ABC transporter-dependent pathway is also implicated in the capsular polysaccharide (CPS) biosynthesis, we employed extraction protocols for both EPS (from the culture supernatants) and CPS (from the cells) and found that the secreted polysaccharide i.e., EPS was predominant. The EPS showed good emulsifying activities against the petroleum hydrocarbons and its production was dependent on the carbon source supplied. The genome analysis also revealed genes involved in industrially important metabolites such as zeaxanthin pigment, ectoine and polyhydroxyalkanoate (PHA) biosynthesis. To confirm the genome data, we extracted these metabolites from the cultures and successfully identified them. The pigment extracted from the cells showed the distinct UV–Vis spectra having characteristic absorption peak of zeaxanthin (λmax 448 nm) with potent antioxidant activities. The ability of H. malpeensis strain YU-PRIM-29T to produce important biomolecules makes it an industrially important bacterium.

Evaluation of mesophilic Burkholderia sacchari, thermophilic Schlegelella thermodepolymerans and halophilic Halomonas halophila for polyhydroxyalkanoates production on model media mimicking lignocellulose hydrolysates

In this work, the mesophilic bacterium Burkholderia sacchari, the halophilic bacterium Halomonas halophila, and the thermophilic bacterium Schlegelella thermodepolymerans were evaluated with regards to their suitability for polyhydroxyalkanoates (PHA) production from model media mimicking lignocellulose hydrolysates. B. sacchari was capable of utilizing all the tested “model hydrolysates”, yielding comparable PHA titers and turning out as very robust against lignocellulose-derived microbial inhibitors. On the contrary, H. halophila reached substantially higher PHA titers on hexoses-rich media, while S. thermodepolymerans preferred media rich in pentoses. Both extremophiles were more sensitive to microbial inhibitors than B. sacchari. Nevertheless, considering substantially higher PHA productivity of both extremophiles even in the presence of microbial inhibitors and also other positive factors associated with utilization of extremophiles, such as the reduced risk of microbial contamination, both H. halophila and S. thermodepolymerans are auspicious candidates for sustainable PHA production from abundantly available, inexpensive lignocelluloses.

Halomonas venusta Coupled Operation of Nitrogen Removal and Electricity Generation under High Salt

In this paper, the moderately halophilic bacterium Halomonas venusta DSM 4743T was selected to optimize the N removal conditions and the electricity generation conditions in the MFC, and conduct the coupled operation of the electricity generation and denitrification of MFC high-salt wastewater. The results showed when sodium succinate was used as carbon source, C/N was 5:1, pH was 9, and NaCl was 30 g/L, 93.22% N removal rate was obtained at 168 h. The maximum output voltage of MFC is 346 mV under the conditions of 6 g/L NaAc, 50 μm riboflavin, pH=8 and 30 g/L NaCl. H. venusta DSM 4743T MFC generates electric denitrification coupled operation, with the maximum output voltage up to 224 mV and the denitrification rate of 53.5%.

Effect of impact shock on extremophilic Halomonas gomseoemensis EP-3 isolated from hypersaline sulphated lake Laguna de Peña Hueca, Spain

The geologic histories of planetary surfaces reveal that Earth and Mars have been pummeled by cataclysmic impact events. The surface of Mars has been perused to have an impact origin for its hemispheric dichotomy. The spallation during impact events causes the interplanetary transfer of material from Mars to Earth or Mars to Phobos/Deimos. Assessing the survival of micro-organisms in impact conditions is critical for the development of planetary protection strategies for future missions. Shock waves are generated during such major impact events. The objective of the present investigation was to explore the microbial diversity of the hypersaline sulphated Laguna de Peña Hueca, Spain and to study the effect of shock waves on extremophilic bacteria isolated from the lake. Peña Hueca is a hypersaline sulphated lagoon rich in Mg–Na–SO4–Cl, epsomite and hexahydrate and it potentially serves as Planetary field analogue site for Martian chloride deposits and salt-rich subsurface brines of Ocean worlds like Enceladus and Europa. The microbial community structure of the lagoon was studied by 16S rRNA metagenomic sequencing. The phylogenetic studies indicated the presence of phyla Euryarchaeota, Proteobacteria, and Bacteroides in the hypersaline brines of the lagoon. The anoxic sediments of Peña Hueca showed the presence of Haloanaerobiaeta and Hadesarchaeota including the anoxic genus of Haloanaerobium, Desulfosalsimonas and Desulfovermiculum. The effect of impact shock on the halophilic bacterium Halomonas gomseomensis EP-3 isolated from Laguna de Peña Hueca was studied in a Reddy shock tube. The halophilic bacterium was exposed to shock waves at a peak shock pressure of 300 ​kPa and a temperature of 400 ​K. The results of shock recovery experiments of halophilic bacteria reveal 97% killing at 300 ​kPa and Mach number of 1.47 in comparison with Bacillus sp. This study indicates that gram-positive spore-forming Bacillus sp. are better adapted to survival in impact shock waves in comparison to non-sporulating halophiles. In the current study, we present the first report on response of halophiles in impact shock. Furthermore, we demonstrate a novel application of the simple handheld Reddy shock tube in astrobiology. The survival studies of halophiles isolated from terrestrial analogue sites in impact shock can provide valuable insights in astrobiology and microbial physiology in impact shock stress.

Contribution of mechanosensitive channels to osmoadaptation and ectoine excretion in Halomonas elongata

For osmoadaptation the halophilic bacterium Halomonas elongata synthesizes as its main compatible solute the aspartate derivative ectoine. H. elongata does not rely entirely on synthesis but can accumulate ectoine by uptake from the surrounding environment with the help of the osmoregulated transporter TeaABC. Disruption of the TeaABC-mediated ectoine uptake creates a strain that is constantly losing ectoine to the medium. However, the efflux mechanism of ectoine in H. elongata is not yet understood. H. elongata possesses four genes encoding mechanosensitive channels all of which belong to the small conductance type (MscS). Analysis by qRT-PCR revealed a reduction in transcription of the mscS genes with increasing salinity. The response of H. elongata to hypo- and hyperosmotic shock never resulted in up-regulation but rather in down-regulation of mscS transcription. Deletion of all four mscS genes created a mutant that was unable to cope with hypoosmotic shock. However, the knockout mutant grew significantly faster than the wildtype at high salinity of 2 M NaCl, and most importantly, still exported 80% of the ectoine compared to the wildtype. We thus conclude that a yet unknown system, which is independent of mechanosensitive channels, is the major export route for ectoine in H. elongata.

PHA Production and PHA Synthases of the Halophilic Bacterium Halomonas sp. SF2003.

Among the different tools which can be studied and managed to tailor-make polyhydroxyalkanoates (PHAs) and enhance their production, bacterial strain and carbon substrates are essential. The assimilation of carbon sources is dependent on bacterial strain’s metabolism and consequently cannot be dissociated. Both must wisely be studied and well selected to ensure the highest production yield of PHAs. Halomonas sp. SF2003 is a marine bacterium already identified as a PHA-producing strain and especially of poly-3-hydroxybutyrate (P-3HB) and poly-3-hydroxybutyrate-co-3-hydroxyvalerate (P-3HB-co-3HV). Previous studies have identified different genes potentially involved in PHA production by Halomonas sp. SF2003, including two phaC genes with atypical characteristics, phaC1 and phaC2. At the same time, an interesting adaptability of the strain in front of various growth conditions was highlighted, making it a good candidate for biotechnological applications. To continue the characterization of Halomonas sp. SF2003, the screening of carbon substrates exploitable for PHA production was performed as well as production tests. Additionally, the functionality of both PHA synthases PhaC1 and PhaC2 was investigated, with an in silico study and the production of transformant strains, in order to confirm and to understand the role of each one on PHA production. The results of this study confirm the adaptability of the strain and its ability to exploit various carbon substrates, in pure or mixed form, for PHA production. Individual expression of PhaC1 and PhaC2 synthases in a non-PHA-producing strain, Cupriavidus necator H16 PHB¯4 (DSM 541), allows obtaining PHA production, demonstrating at the same time, functionality and differences between both PHA synthases. All the results of this study confirm the biotechnological interest in Halomonas sp. SF2003.

Growth and nitrogen removal characteristics of Halomonas sp. B01 under high salinity

PurposeAt present, the nitrogen (N) removal efficiency of the microbial treatment in the high-salinity nitrogenous wastewaters is relatively low. Study on the N removal behavior and properties of moderately halophilic bacteria Halomonas under high salinity is of great significance for the microbial treatment of high-salinity nitrogenous wastewater.MethodsThe response mechanism of Halomonas sp. B01 to high osmotic pressure stress was investigated by measuring the compatible solute ectoine concentration and superoxide dismutase (SOD) activity. The salt tolerance during growth and N removal of the strain was evaluated by measuring the activities of growth-related and N removal–related enzymes and the mRNA expression abundance of ammonia monooxygenase-encoding gene (amoA). The process of simultaneous heterotrophic nitrification and aerobic denitrification (SND) under high salinity was described by measuring the concentration of inorganic N.ResultHalomonas sp. B01 synthesized ectoine under NaCl stress, and the intracellular ectoine concentration increased with increased NaCl concentration in the growth medium. When the NaCl concentration of the medium reached 120 g L−1, the malondialdehyde concentration and SOD activity were significantly increased to 576.1 μg mg−1 and 1.7 U mg−1, respectively. The growth-related and N removal–related enzymes of the strain were active or most active in medium with 30–60 g L−1 NaCl. The amoA of the strain cultured in medium with 60 g L−1 NaCl had the highest mRNA expression abundance. In the N removal medium containing 60 g L−1 NaCl and 2121 mg L−1 NH4+-N, SND by Halomonas sp. B01 was performed over 96 h and the N removal rate reached 98.8%.ConclusionIn addition to the protective mechanism of synthetic compatible solutes, Halomonas sp. B01 had the repair mechanism of SOD for lipid peroxidation. The growth-related and N removal–related enzymes of the strain were most active at a certain salt concentration; amoA also had the highest mRNA expression abundance under high salinity. Halomonas sp. B01 could efficiently perform N removal by SND under high salinity.