Identification and characterization of magnetotactic Gammaproteobacteria from a salt evaporation pool,Bohai Bay,China

Magnetotactic bacteria (MTB) are phylogenetically diverse prokaryotes that can produce intracellular chain-assembled nanocrystals of magnetite (Fe₃O₄) or greigite (Fe₃S₄). Compared with their wide distribution in the Alpha-, Eta- and Delta-proteobacteria classes, few MTB strains have been identified in the Gammaproteobacteria class, resulting in limited knowledge of bacterial diversity and magnetosome biomineralization within this phylogenetic branch. Here, we identify two magnetotactic Gammaproteobacteria strains (tentatively named FZSR-1 and FZSR-2 respectively) from a salt evaporation pool in Bohai Bay, at the Fuzhou saltern, Dalian City, eastern China. Phylogenetic analysis indicates that strain FZSR-2 is the same species as strains SHHR-1 and SS-5, which were discovered previously from brackish and hypersaline environments respectively. Strain FZSR-1 represents a novel species. Compared with strains FZSR-2, SHHR-1 and SS-5 in which magnetite particles are assembled into a single chain, FZSR-1 cells form relatively narrower magnetite nanoparticles that are often organized into double chains. We find a good relationship between magnetite morphology within strains FZSR-2, SHHR-1 and SS-5 and the salinity of the environment in which they live. This study expands the bacterial diversity of magnetotactic Gammaproteobacteria and provides new insights into magnetosome biomineralization within magnetotactic Gammaproteobacteria.     

Identification of Anaerobic Selenate-Respiring Bacteria from Aquatic Sediments

The diversity population of microorganisms with the capability to use selenate as a terminal electron acceptor, reducing it to selenite and elemental selenium by the process known as dissimilatory selenate reduction, is largely unknown. The overall objective of this study was to gain an in-depth understanding of anaerobic biotransformation of selenium in the environment, particularly anaerobic respiration, and to characterize the microorganisms catalyzing this process. Here, we demonstrate the isolation and characterization of four novel anaerobic dissimilatory selenate-respiring bacteria enriched from a variety of sources, including sediments from three different water bodies in Chennai, India, and a tidal estuary in New Jersey. Strains S5 and S7 from India, strain KM from the Meadowlands, NJ, and strain pn1, categorized as a laboratory contaminant, were all phylogenetically distinct, belonging to various phyla in the bacterial domain. The 16S rRNA gene sequence shows that strain S5 constitutes a new genus belonging to Chrysiogenetes, while strain S7 belongs to the Deferribacteres, with greater than 98% 16S rRNA gene similarity to Geovibrio ferrireducens. Strain KM is related to Malonomonas rubra, Pelobacter acidigallici, and Desulfuromusa spp., with 96 to 97% 16S rRNA gene similarity. Strain pn1 is 99% similar to Pseudomonas stutzeri. Strains S5, S7, and KM are obligately anaerobic selenate-respiring microorganisms, while strain pn1 is facultatively anaerobic. Besides respiring selenate, all these strains also respire nitrate.     

A biogeographic distribution of magnetotactic bacteria influenced by salinity

Magnetotactic bacteria (MTB), which synthesize intracellular ferromagnetic magnetite and/or greigite magnetosomes, have significant roles in global iron cycling in aquatic systems, as well as sedimentary magnetism. The occurrence of MTB has been reported in aquatic environments from freshwater to marine ecosystems; however, the distribution of MTB across heterogeneous habitats remains unclear. Here we examined the MTB communities from diverse habitats across northern and southern China, using comprehensive transmission electron microscopy and comparison of 16S rRNA gene analyses. A total of 334 16S rRNA gene sequences were analyzed, representing the most comprehensive analysis on the diversity and distribution of MTB to date. The majority (95%) of sequences belong to the Alphaproteobacteria, whereas a population of giant magnetotactic rod is affiliated with the Nitrospirae phylum. By a statistical comparison of these sequence data and publicly available MTB sequences, we infer for the first time that the composition of MTB communities represents a biogeographic distribution across globally heterogeneous environments, which is influenced by salinity.     

Culture‐independent characterization of a novel magnetotactic member affiliated to the Beta class of the Proteobacteria phylum from an acidic lagoon

Magnetotactic bacteria (MTB) comprise a group of motile microorganisms common in most mesothermal aquatic habitats with pH values around neutrality. However, during the last two decades, a number of MTB from extreme environments have been characterized including: cultured alkaliphilic strains belonging to the Deltaproteobacteria class of the Proteobacteria phylum; uncultured moderately thermophilic strains belonging to the Nitrospirae phylum; cultured and uncultured moderately halophilic or strongly halotolerant bacteria affiliated with the Deltaproteobacteria and Gammaproteobacteria classes and an uncultured psychrophilic species belonging to the Alphaproteobacteria class. Here, we used culture‐independent techniques to characterize MTB from an acidic freshwater lagoon in Brazil (pH ～ 4.4). MTB morphotypes found in this acidic lagoon included cocci, rods, spirilla and vibrioid cells. Magnetite (Fe₃O₄) was the only mineral identified in magnetosomes of these MTB while magnetite magnetosome crystal morphologies within the different MTB cells included cuboctahedral (present in spirilla), elongated prismatic (present in cocci and vibrios) and bullet‐shaped (present in rod‐shaped cells). Intracellular pH measurements using fluorescent dyes showed that the cytoplasmic pH was close to neutral in most MTB cells and acidic in some intracellular granules. Based on 16S rRNA gene phylogenetic analyses, some of the retrieved gene sequences belonged to the genus Herbaspirillum within the Betaproteobacteria class of the Proteobacteria phylum. Fluorescent in situ hybridization using a Herbaspirillum‐specific probe hybridized with vibrioid MTB in magnetically‐enriched samples. Transmission electron microscopy of the Herbaspirillum‐like MTB revealed the presence of many intracellular granules and a single chain of elongated prismatic magnetite magnetosomes. Diverse populations of MTB have not seemed to have been described in detail in an acid environment. In addition, this is the first report of an MTB phylogenetically affiliated with Betaproteobacteria class.     

Evidence for Autotrophy via the Reverse Tricarboxylic Acid Cycle in the Marine Magnetotactic Coccus Strain MC-1

Strain MC-1 is a marine, microaerophilic, magnetite-producing, magnetotactic coccus phylogenetically affiliated with the α-Proteobacteria. Strain MC-1 grew chemolithotrophically with sulfide and thiosulfate as electron donors with HCO₃⁻/CO₂ as the sole carbon source. Experiments with cells grown microaerobically in liquid with thiosulfate and H¹⁴CO₃⁻/¹⁴CO₂ showed that all cell carbon was derived from H¹⁴CO₃⁻/¹⁴CO₂ and therefore that MC-1 is capable of chemolithoautotrophy. Cell extracts did not exhibit ribulose-1,5-bisphosphate carboxylase-oxygenase (RubisCO) activity, nor were RubisCO genes found in the draft genome of MC-1. Thus, unlike other chemolithoautotrophic, magnetotactic bacteria, strain MC-1 does not appear to utilize the Calvin-Benson-Bassham cycle for autotrophy. Cell extracts did not exhibit carbon monoxide dehydrogenase activity, indicating that the acetyl-coenzyme A pathway also does not function in strain MC-1. The ¹³C content of whole cells of MC-1 relative to the ¹³C content of the inorganic carbon source (Δδ¹³C) was −11.4 . Cellular fatty acids showed enrichment of ¹³C relative to whole cells. Strain MC-1 cell extracts showed activities for several key enzymes of the reverse (reductive) tricarboxylic acid (rTCA) cycle including fumarate reductase, pyruvate:acceptor oxidoreductase and 2-oxoglutarate:acceptor oxidoreductase. Although ATP citrate lyase (another key enzyme of the rTCA cycle) activity was not detected in strain MC-1 using commonly used assays, cell extracts did cleave citrate, and the reaction was dependent upon the presence of ATP and coenzyme A. Thus, we infer the presence of an ATP-dependent citrate-cleaving mechanism. These results are consistent with the operation of the rTCA cycle in MC-1. Strain MC-1 appears to be the first known representative of the α-Proteobacteria to use the rTCA cycle for autotrophy.     

Potential Interactions of Particle-Associated Anammox Bacteria with Bacterial and Archaeal Partners in the Namibian Upwelling System

Recent studies have shown that the anaerobic oxidation of ammonium by anammox bacteria plays an important role in catalyzing the loss of nitrogen from marine oxygen minimum zones (OMZ). However, in situ oxygen concentrations of up to 25 μM and ammonium concentrations close to or below the detection limit in the layer of anammox activity are hard to reconcile with the current knowledge of the physiology of anammox bacteria. We therefore investigated samples from the Namibian OMZ by comparative 16S rRNA gene analysis and fluorescence in situ hybridization. Our results showed that “Candidatus Scalindua” spp., the typical marine anammox bacteria, colonized microscopic particles that were likely the remains of either macroscopic marine snow particles or resuspended particles. These particles were slightly but significantly (P < 0.01) enriched in Gammaproteobacteria (11.8% ± 5.0%) compared to the free-water phase (8.1% ± 1.8%). No preference for the attachment to particles could be observed for members of the Alphaproteobacteria and Bacteroidetes, which were abundant (12 to 17%) in both habitats. The alphaproteobacterial SAR11 clade, the Euryarchaeota, and group I Crenarchaeota, were all significantly depleted in particles compared to their presence in the free-water phase (16.5% ± 3.5% versus 2.6% ± 1.7%, 2.7% ± 1.9% versus <1%, and 14.9% ± 4.6% versus 2.2% ± 1.8%, respectively, all P < 0.001). Sequence analysis of the crenarchaeotal 16S rRNA genes showed a 99% sequence identity to the nitrifying “Nitrosopumilus maritimus.” Even though we could not observe conspicuous consortium-like structures of anammox bacteria with particle-enriched bacterioplankton groups, we hypothesize that members of Gammaproteobacteria, Alphaproteobacteria, and Bacteroidetes play a critical role in extending the anammox reaction to nutrient-depleted suboxic water layers in the Namibian upwelling system by creating anoxic, nutrient-enriched microniches.     

Cultivation and Growth Characteristics of a Diverse Group of Oligotrophic Marine Gammaproteobacteria

Forty-four novel strains of Gammaproteobacteria were cultivated from coastal and pelagic regions of the Pacific Ocean using high-throughput culturing methods that rely on dilution to extinction in very low nutrient media. Phylogenetic analysis showed that the isolates fell into five rRNA clades, all of which contained rRNA gene sequences reported previously from seawater environmental gene clone libraries (SAR92, OM60, OM182, BD1-7, and KI89A). Bootstrap analyses of phylogenetic reliability did not support collapsing these five clades into a single clade, and they were therefore named the oligotrophic marine Gammaproteobacteria (OMG) group. Twelve cultures chosen to represent the five clades were successively purified in liquid culture, and their growth characteristics were determined at different temperatures and dissolved organic carbon concentrations. The isolates in the OMG group were physiologically diverse heterotrophs, and their physiological properties generally followed their phylogenetic relationships. None of the isolates in the OMG group formed colonies on low- or high-nutrient agar upon their first isolation from seawater, while 7 of 12 isolates that were propagated for laboratory testing eventually produced colonies on 1/10 R2A agar. The isolates grew relatively slowly in natural seawater media (1.23 to 2.63 day⁻¹), and none of them grew in high-nutrient media (>351 mg of C liter⁻¹). The isolates were psychro- to mesophilic and obligately oligotrophic; many of them were of ultramicrobial size (<0.1 μm³). This cultivation study revealed that sporadically detected Gammaproteobacteria gene clones from seawater are part of a phylogenetically diverse constellation of organisms mainly composed of oligotrophic and ultramicrobial lineages that are culturable under specific cultivation conditions.     

Desulfamplus magnetovallimortis gen. nov., sp. nov., a magnetotactic bacterium from a brackish desert spring able to biomineralize greigite and magnetite,that represents a novel lineage in the Desulfobacteraceae

A magnetotactic bacterium, designated strain BW-1^T, was isolated from a brackish spring in Death Valley National Park (California, USA) and cultivated in axenic culture. The Gram-negative cells of strain BW-1^T are relatively large and rod-shaped and possess a single polar flagellum (monotrichous). This strain is the first magnetotactic bacterium isolated in axenic culture capable of producing greigite and/or magnetite nanocrystals aligned in one or more chains per cell. Strain BW-1^T is an obligate anaerobe that grows chemoorganoheterotrophically while reducing sulfate as a terminal electron acceptor. Optimal growth occurred at pH 7.0 and 28 °C with fumarate as electron donor and carbon source. Based on its genome sequence, the G + C content is 40.72 mol %. Phylogenomic and phylogenetic analyses indicate that strain BW-1^T belongs to the Desulfobacteraceae family within the Deltaproteobacteria class. Based on average amino acid identity, strain BW-1^T can be considered as a novel species of a new genus, for which the name Desulfamplus magnetovallimortis is proposed. The type strain of D. magnetovallimortis is BW-1^T (JCM 18010^T–DSM 103535^T).     

Phylogeny of the Defined Murine Microbiota: Altered Schaedler Flora

The “altered Schaedler flora” (ASF) was developed for colonizing germfree rodents with a standardized microbiota. The purpose of this study was to identify each of the eight ASF strains by 16S rRNA sequence analysis. Three strains were previously identified as Lactobacillus acidophilus (strain ASF 360), Lactobacillus salivarius (strain ASF 361), and Bacteroides distasonis (strain ASF 519) based on phenotypic criteria. 16S rRNA analysis indicated that each of the strains differed from its presumptive identity. The 16S rRNA sequence of strain ASF 361 is essentially identical to the 16S rRNA sequences of the type strains of Lactobacillus murinis and Lactobacillus animalis (both isolated from mice), and all of these strains probably belong to a single species. Strain ASF 360 is a novel lactobacillus that clusters with L. acidophilus and Lactobacillus lactis. Strain ASF 519 falls into an unnamed genus containing [Bacteroides] distasonis, [Bacteroides] merdae, [Bacteroides] forsythus, and CDC group DF-3. This unnamed genus is in the Cytophaga-Flavobacterium-Bacteroides phylum and is most closely related to the genus Porphyromonas. The spiral-shaped strain, strain ASF 457, is in the Flexistipes phylum and exhibits sequence identity with rodent isolates of Robertson. The remaining four ASF strains, which are extremely oxygen-sensitive fusiform bacteria, group phylogenetically with the low-G+C-content gram-positive bacteria (Firmicutes, Bacillus-Clostridium group). ASF 356, ASF 492, and ASF 502 fall into Clostridium cluster XIV of Collins et al. Morphologically, ASF 492 resembles members of this cluster, Roseburia cecicola, and Eubacterium plexicaudatum. The 16S rRNA sequence of ASF 492 is identical to that of E. plexicaudatum. Since the type strain and other viable original isolates of E. plexicaudatum have been lost, strain ASF 492 is a candidate for a neotype strain. Strain ASF 500 branches deeply in the low-G+C-content gram-positive phylogenetic tree but is not closely related to any organisms whose 16S rRNA sequences are currently in the GenBank database. The 16S rRNA sequence information determined in the present study should allow rapid identification of ASF strains and should permit detailed analysis of the interactions of ASF organisms during development of intestinal disease in mice that are coinfected with a variety of pathogenic microorganisms.     

Isolation and Characterization of Strains CVO and FWKO B, Two Novel Nitrate-Reducing, Sulfide-Oxidizing Bacteria Isolated from Oil Field Brine

Bacterial strains CVO and FWKO B were isolated from produced brine at the Coleville oil field in Saskatchewan, Canada. Both strains are obligate chemolithotrophs, with hydrogen, formate, and sulfide serving as the only known energy sources for FWKO B, whereas sulfide and elemental sulfur are the only known electron donors for CVO. Neither strain uses thiosulfate as an energy source. Both strains are microaerophiles (1% O₂). In addition, CVO grows by denitrification of nitrate or nitrite whereas FWKO B reduces nitrate only to nitrite. Elemental sulfur is the sole product of sulfide oxidation by FWKO B, while CVO produces either elemental sulfur or sulfate, depending on the initial concentration of sulfide. Both strains are capable of growth under strictly autotrophic conditions, but CVO uses acetate as well as CO₂ as its sole carbon source. Neither strain reduces sulfate; however, FWKO B reduces sulfur and displays chemolithoautotrophic growth in the presence of elemental sulfur, hydrogen, and CO₂. Both strains grow at temperatures between 5 and 40°C. CVO is capable of growth at NaCl concentrations as high as 7%. The present 16s rRNA analysis suggests that both strains are members of the epsilon subdivision of the division Proteobacteria, with CVO most closely related to Thiomicrospira denitrifcans and FWKO B most closely related to members of the genus Arcobacter. The isolation of these two novel chemolithotrophic sulfur bacteria from oil field brine suggests the presence of a subterranean sulfur cycle driven entirely by hydrogen, carbon dioxide, and nitrate.     

Isolation of obligately alkaliphilic magnetotactic bacteria from extremely alkaline environments

Large numbers of magnetotactic bacteria were discovered in mud and water samples collected from a number of highly alkaline aquatic environments with pH values of ≈ 9.5. These bacteria were helical in morphology and biomineralized chains of bullet-shaped crystals of magnetite and were present in all the highly alkaline sites sampled. Three strains from different sites were isolated and cultured and grew optimally at pH 9.0-9.5 but not at 8.0 and below, demonstrating that these organisms truly require highly alkaline conditions and are not simply surviving/growing in neutral pH micro-niches in their natural habitats. All strains grew anaerobically through the reduction of sulfate as a terminal electron acceptor and phylogenetic analysis, based on 16S rRNA gene sequences, as well as some physiological features, showed that they could represent strains of Desulfonatronum thiodismutans, a known alkaliphilic bacterium that does not biomineralize magnetosomes. Our results show that some magnetotactic bacteria can be considered extremophilic and greatly extend the known ecology of magnetotactic bacteria and the conditions under which they can biomineralize magnetite. Moreover, our results show that this type of magnetotactic bacterium is common in highly alkaline environments. Our findings also greatly influence the interpretation of the presence of nanometer-sized magnetite crystals, so-called magnetofossils, in highly alkaline environments.     

Identification and Isolation of a Castellaniella Species Important during Biostimulation of an Acidic Nitrate- and Uranium-Contaminated Aquifer

Immobilization of uranium in groundwater can be achieved through microbial reduction of U(VI) to U(IV) upon electron donor addition. Microbial community structure was analyzed in ethanol-biostimulated and control sediments from a high-nitrate (>130 mM), low-pH, uranium-contaminated site in Oak Ridge, TN. Analysis of small subunit (SSU) rRNA gene clone libraries and polar lipid fatty acids from sediments revealed that biostimulation resulted in a general decrease in bacterial diversity. Specifically, biostimulation resulted in an increase in the proportion of Betaproteobacteria (10% of total clones in the control sediment versus 50 and 79% in biostimulated sediments) and a decrease in the proportion of Gammaproteobacteria and Acidobacteria. Clone libraries derived from dissimilatory nitrite reductase genes (nirK and nirS) were also dominated by clones related to Betaproteobacteria (98% and 85% of total nirK and nirS clones, respectively). Within the nirK libraries, one clone sequence made up 59 and 76% of sequences from biostimulated sediments but only made up 10% of the control nirK library. Phylogenetic analysis of SSU rRNA and nirK gene sequences from denitrifying pure cultures isolated from the site indicate that all belong to a Castellaniella species; nearly identical sequences also constituted the majority of biostimulated SSU rRNA and nirK clone libraries. Thus, by combining culture-independent with culture-dependent techniques, we were able to link SSU rRNA clone library information with nirK sequence data and conclude that a potentially novel Castellaniella species is important for in situ nitrate removal at this site.     

Cold-Active Chemoorganotrophic Bacteria from Permanently Ice-Covered Lake Hoare, McMurdo Dry Valleys, Antarctica

Eight strains of chemoorganotrophic bacteria were isolated from the water column of Lake Hoare, McMurdo Dry Valleys, Antarctica, using cold enrichment temperatures. The isolates were Alpha-, Beta-, and Gammaproteobacteria and Actinobacteria spp. All isolates grew at 0°C, and all but one grew at subzero temperatures characteristic of the water column of Lake Hoare. Growth temperature optima varied among isolates, but the majority showed optima near 15°C, indicative of cold-active phenotypes. One isolate was truly psychrophilic, growing optimally around 10°C and not above 20°C. Half of the isolates grew at 2% salt while the other half did not, and all but one isolate grew at 2 atm of O₂. Our isolates are the first prokaryotes from the water column of Lake Hoare to be characterized phylogenetically and physiologically and show that cold-active species of at least two major phyla of Bacteria inhabit Lake Hoare.     

Evidence for autotrophy via the reverse tricarboxylic acid cycle in the marine magnetotactic coccus strain MC-1

Strain MC-1 is a marine, microaerophilic, magnetite-producing, magnetotactic coccus phylogenetically affiliated with the alpha-Proteobacteria. Strain MC-1 grew chemolithotrophically with sulfide and thiosulfate as electron donors with HCO3-/CO2 as the sole carbon source. Experiments with cells grown microaerobically in liquid with thiosulfate and H14CO3-/14CO2 showed that all cell carbon was derived from H14CO3-/14CO2 and therefore that MC-1 is capable of chemolithoautotrophy. Cell extracts did not exhibit ribulose-1,5-bisphosphate carboxylase-oxygenase (RubisCO) activity, nor were RubisCO genes found in the draft genome of MC-1. Thus, unlike other chemolithoautotrophic, magnetotactic bacteria, strain MC-1 does not appear to utilize the Calvin-Benson-Bassham cycle for autotrophy. Cell extracts did not exhibit carbon monoxide dehydrogenase activity, indicating that the acetyl-coenzyme A pathway also does not function in strain MC-1. The 13C content of whole cells of MC-1 relative to the 13C content of the inorganic carbon source (Deltadelta13C) was -11.4 per thousand. Cellular fatty acids showed enrichment of 13C relative to whole cells. Strain MC-1 cell extracts showed activities for several key enzymes of the reverse (reductive) tricarboxylic acid (rTCA) cycle including fumarate reductase, pyruvate:acceptor oxidoreductase and 2-oxoglutarate:acceptor oxidoreductase. Although ATP citrate lyase (another key enzyme of the rTCA cycle) activity was not detected in strain MC-1 using commonly used assays, cell extracts did cleave citrate, and the reaction was dependent upon the presence of ATP and coenzyme A. Thus, we infer the presence of an ATP-dependent citrate-cleaving mechanism. These results are consistent with the operation of the rTCA cycle in MC-1. Strain MC-1 appears to be the first known representative of the alpha-Proteobacteria to use the rTCA cycle for autotrophy.     

Identification of Dekkera bruxellensis (Brettanomyces) from Wine by Fluorescence In Situ Hybridization Using Peptide Nucleic Acid Probes

A new fluorescence in situ hybridization method using peptide nucleic acid (PNA) probes for identification of Brettanomyces is described. The test is based on fluorescein-labeled PNA probes targeting a species-specific sequence of the rRNA of Dekkera bruxellensis. The PNA probes were applied to smears of colonies, and results were interpreted by fluorescence microscopy. The results obtained from testing 127 different yeast strains, including 78 Brettanomyces isolates from wine, show that the spoilage organism Brettanomyces belongs to the species D. bruxellensis and that the new method is able to identify Brettanomyces (D. bruxellensis) with 100% sensitivity and 100% specificity.     

Anaerobic Mineralization of Quaternary Carbon Atoms: Isolation of Denitrifying Bacteria on Dimethylmalonate

The microbial capacity to degrade simple organic compounds with quaternary carbon atoms was demonstrated by enrichment and isolation of five denitrifying strains on dimethylmalonate as the sole electron donor and carbon source. Quantitative growth experiments showed a complete mineralization of dimethylmalonate. According to phylogenetic analysis of the complete 16S rRNA genes, two strains isolated from activated sewage sludge were related to the genus Paracoccus within the α-Proteobacteria (98.0 and 98.2% 16S rRNA gene similarity to Paracoccus denitrificans^T), and three strains isolated from freshwater ditches were affiliated with the β-Proteobacteria (97.4 and 98.3% 16S rRNA gene similarity to Herbaspirillum seropedicae^T and Acidovorax facilis^T, respectively). Most-probable-number determinations for denitrifying populations in sewage sludge yielded 4.6 × 10⁴ dimethylmalonate-utilizing cells ml⁻¹, representing up to 0.4% of the total culturable nitrate-reducing population.     

Isolation and Distribution of a Novel Iron-Oxidizing Crenarchaeon from Acidic Geothermal Springs in Yellowstone National Park

Novel thermophilic crenarchaea have been observed in Fe(III) oxide microbial mats of Yellowstone National Park (YNP); however, no definitive work has identified specific microorganisms responsible for the oxidation of Fe(II). The objectives of the current study were to isolate and characterize an Fe(II)-oxidizing member of the Sulfolobales observed in previous 16S rRNA gene surveys and to determine the abundance and distribution of close relatives of this organism in acidic geothermal springs containing high concentrations of dissolved Fe(II). Here we report the isolation and characterization of the novel, Fe(II)-oxidizing, thermophilic, acidophilic organism Metallosphaera sp. strain MK1 obtained from a well-characterized acid-sulfate-chloride geothermal spring in Norris Geyser Basin, YNP. Full-length 16S rRNA gene sequence analysis revealed that strain MK1 exhibits only 94.9 to 96.1% sequence similarity to other known Metallosphaera spp. and less than 89.1% similarity to known Sulfolobus spp. Strain MK1 is a facultative chemolithoautotroph with an optimum pH range of 2.0 to 3.0 and an optimum temperature range of 65 to 75°C. Strain MK1 grows optimally on pyrite or Fe(II) sorbed onto ferrihydrite, exhibiting doubling times between 10 and 11 h under aerobic conditions (65°C). The distribution and relative abundance of MK1-like 16S rRNA gene sequences in 14 acidic geothermal springs containing Fe(III) oxide microbial mats were evaluated. Highly related MK1-like 16S rRNA gene sequences (>99% sequence similarity) were consistently observed in Fe(III) oxide mats at temperatures ranging from 55 to 80°C. Quantitative PCR using Metallosphaera-specific primers confirmed that organisms highly similar to strain MK1 comprised up to 40% of the total archaeal community at selected sites. The broad distribution of highly related MK1-like 16S rRNA gene sequences in acidic Fe(III) oxide microbial mats is consistent with the observed characteristics and growth optima of Metallosphaera-like strain MK1 and emphasizes the importance of this newly described taxon in Fe(II) chemolithotrophy in acidic high-temperature environments of YNP.     

(Per)chlorate Reduction by the Thermophilic Bacterium Moorella perchloratireducens sp. nov., Isolated from Underground Gas Storage

A thermophilic bacterium, strain An10, was isolated from underground gas storage with methanol as a substrate and perchlorate as an electron acceptor. Cells were gram-positive straight rods, 0.4 to 0.6 μm in diameter and 2 to 8 μm in length, growing as single cells or in pairs. Spores were terminal with a bulged sporangium. The temperature range for growth was 40 to 70°C, with an optimum at 55 to 60°C. The pH optimum was around 7. The salinity range for growth was between 0 and 40 g NaCl liter⁻¹ with an optimum at 10 g liter⁻¹. Strain An10 was able to grow on CO, methanol, pyruvate, glucose, fructose, cellobiose, mannose, xylose, and pectin. The isolate was able to respire with (per)chlorate, nitrate, thiosulfate, neutralized Fe(III) complexes, and anthraquinone-2,6-disulfonate. The G+C content of the DNA was 57.6 mol%. On the basis of 16S rRNA analysis, strain An10 was most closely related to Moorella thermoacetica and Moorella thermoautotrophica. The bacterium reduced perchlorate and chlorate completely to chloride. Key enzymes, perchlorate reductase and chlorite dismutase, were detected in cell extracts. Strain An10 is the first thermophilic and gram-positive bacterium with the ability to use (per)chlorate as a terminal electron acceptor.     

Staphylococcus lipolyticus sp. nov., a new cold-adapted lipase producing marine species

A cold-adapted lipase producing bacterium, designated SS-33^T, was isolated from sea sediment collected from the Bay of Bengal, India, and subjected to a polyphasic taxonomic study. Strain SS-33^T exhibited the highest 16S rRNA gene sequence similarity with Staphylococcus cohnii subsp. urealyticus (97.18 %), Staphylococcus saprophyticus subsp. bovis (97.16 %) and Staphylococcus cohnii subsp. cohnii (97.04 %). Phylogenetic analysis based on the 16S rRNA gene sequences showed that strain SS-33^T belongs to the genus Staphylococcus. Cells of strain SS-33^T were Gram-positive, coccus-shaped, non-spore-forming, non-motile, catalase-positive and oxidase-negative. The major fatty acid detected in strain SS-33^T was anteiso-C15:0 and the menaquinone was MK-7. The genomic DNA G + C content was 33 mol%. The DNA-DNA hybridization among strain SS-33^T and the closely related species indicated that strain SS-33^T represents a novel species of the genus Staphylococcus. On the basis of the morphological, physiological and chemotaxonomic characteristics, the results of phylogenetic analysis and the DNA-DNA hybridization, a novel species is proposed for strain SS-33^T, with the name Staphylococcus lipolyticus sp. nov. The strain type is SS-33^T (=MTCC 10101^T?=?JCM 16560^T). Staphylococcus lipolyticus SS-33^T hydrolyzed various substrates including tributyrin, olive oil, Tween 20, Tween 40, Tween 60, and Tween 80 at low temperatures, as well as mesophilic temperatures. Lipase from strain SS-33^T was partially purified by acetone precipitation. The molecular weight of lipase protein was determined 67 kDa by SDS-PAGE. Zymography was performed to monitor the lipase activity in Native-PAGE. Calcium ions increased lipase activity twofold. The optimum pH of lipase was pH 7.0 and optimum temperature was 30 °C. However, lipase exhibited 90 % activity of its optimum temperature at 10 °C and became more stable at 10 °C as compared to 30 °C. The lipase activity and stability at low temperature has wide ranging applications in various industrial processes. Therefore, cold-adapted mesophilic lipase from strain SS-33^T may be used for industrial applications. This is the first report of the production of cold-adapted mesophilic lipase by any Staphylococcus species.     

Comparative genomic analysis of magnetotactic bacteria from the Deltaproteobacteria provides new insights into magnetite and greigite magnetosome genes required for magnetotaxis

Magnetotactic bacteria (MTB) represent a group of diverse motile prokaryotes that biomineralize magnetosomes, the organelles responsible for magnetotaxis. Magnetosomes consist of intracellular, membrane‐bounded, tens‐of‐nanometre‐sized crystals of the magnetic minerals magnetite (Fe₃O₄) or greigite (Fe₃S₄) and are usually organized as a chain within the cell acting like a compass needle. Most information regarding the biomineralization processes involved in magnetosome formation comes from studies involving Alphaproteobacteria species which biomineralize cuboctahedral and elongated prismatic crystals of magnetite. Many magnetosome genes, the mam genes, identified in these organisms are conserved in all known MTB. Here we present a comparative genomic analysis of magnetotactic Deltaproteobacteria that synthesize bullet‐shaped crystals of magnetite and/or greigite. We show that in addition to mam genes, there is a conserved set of genes, designated mad genes, specific to the magnetotactic Deltaproteobacteria, some also being present in Candidatus Magnetobacterium bavaricum of the Nitrospirae phylum, but absent in the magnetotactic Alphaproteobacteria. Our results suggest that the number of genes associated with magnetotaxis in magnetotactic Deltaproteobacteria is larger than previously thought. We also demonstrate that the minimum set of mam genes necessary for magnetosome formation in Magnetospirillum is also conserved in magnetite‐producing, magnetotactic Deltaproteobacteria. Some putative novel functions of mad genes are discussed.