Chryseobacterium metallicongregator, sp. nov., a bacterium possessing metallophore activity towards rare earth elements.

A polyphasic taxonomic study was carried out on strain ES2T, isolated from sediment of a wetland created to remediate acid drainage from a coal mine. The rod-shaped bacterium formed yellow/orange pigmented colonies and produced the pigment flexirubin. The 16S rRNA gene sequence results assigned the strain to Chryseobacterium, with 98.9 and 98.3 % similarity to Chryseobacterium vietnamense and Chryseobacterium cucumeris, respectively. Computation of the average nucleotide identity and digital DNA-DNA hybridization values with the closest phylogenetic neighbours of ES2T revealed genetic differences at the species level, which were further substantiated by differences in several physiological characteristics. The dominant fatty acids of strain ES2T were iso-C15 : 0, iso-C17 : 1 ω9c, iso C17 : 0 3-OH, and iso-C15 : 0 2-OH. The DNA G+C content was 35.5 mol%. The major polar lipid was phosphatidylethanolamine while menaquinone-6 was the only menaquinone found. This bacterium has been previously shown to possess metallophore activity towards rare earth elements, and based on genome sequencing, possesses all required genes for siderophore production/activity, possibly identifying the source of this unique ability. On the basis of the results obtained here, this bacterium is assigned to the genus Chryseobacterium as representing a new species with the name Chryseobacterium metallicongregator sp. nov., type strain ES2T (=NRRL B-65679T=KCTC 102120T).

Metallophore Activity toward the Rare Earth Elements by Bacteria Isolated from Acid Mine Drainage Due to Coal Mining.

The field of microbe-metal interactions has been gaining significant attention. While the direct impact of metal oxyanions on bacteria has been investigated, significantly less attention has been placed on the ability of certain microbes to 'collect' such metal ions via secreted proteins. Many bacteria possess low-weight molecules called siderophores, which collect Fe from the environment to be brought back to the cell. However, some appear to have additional roles, including binding other metals, termed 'metallophores'. Microbes can remove/sequester these from their surroundings, but the breadth of those that can be removed is still unknown. Using the Chromeazurol S assay, we identified eight isolates, most belonging to the genus Pseudomonas, possessing siderophore activity, mainly from sites impacted by coal mine drainage, also possessing a metallophore activity toward the rare earth elements that does not appear to be related to ionic radii or previously reported EC50 concentrations for E. coli. We found the strength of metallophore activity towards these elements was as follows: Pr > Sc > Eu > Tm > Tb > Er > Yb > Ce > Lu > Sm > Ho > La > Nd > Dy > Gd > Y. This is the first study to investigate such activity and indicates bacteria may provide a means of removal/recovery of these critical elements.

Gordonia metallireducens sp. nov., a tellurite- and selenite-resistant bacterium isolated from the sediment of an acid mine drainage stream.

A polyphasic taxonomic study was carried out on strain TSed Te1T, isolated from sediment of a stream contaminated with acid drainage from a coal mine. The bacterium forms pink-pigmented colonies and has a rod-coccus growth cycle, which also includes some coryneform arrangements. This bacterium is capable of growing in the presence of up to 750 µg ml-1 tellurite and 5000 µg ml-1 selenite, reducing each to elemental form. Nearly complete 16S rRNA gene sequence analysis associated the strain with Gordonia, with 99.5 and 99.3 % similarity to Gordonia namibiensis and Gordonia rubripertincta, respectively. Computation of the average nucleotide identity and digital DNA-DNA hybridization comparisons with the closest phylogenetic neighbour of TSed Te1T revealed genetic differences at the species level, which were further substantiated by differences in several physiological characteristics. The dominant fatty acids were C16 : 0, C18 : 1, C16 : 1 and tuberculostearic acid. The DNA G+C content was 67.6 mol%. Major polar lipids were diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylinositol and phosphatidylinositol mannoside, while MK-9(H2) was the only menaquinone found. Mycolic acids of C56-C60 were present. Whole-cell hydrolysates contained meso-diaminopimelic acid along with arabinose and galactose as the major cell-wall sugars. On the basis of the results obtained in this study, the bacterium was assigned to the genus Gordonia and represents a new species with the name Gordonia metallireducens sp. nov. The type strain is TSed Te1T (=NRRL B-65678T=DSM 114093T).

Shewanella metallivivens sp. nov., a deep-sea hydrothermal vent tube worm endobiont capable of dissimilatory anaerobic metalloid oxyanion reduction.

A polyphasic taxonomic study was carried out on a Gram-stain-negative and rod-shaped strain, ER-Te-42B-LightT, isolated from the tissue of a tube worm, Riftia pachyptila, collected near a deep-sea hydrothermal vent of the Juan de Fuca Ridge in the Pacific Ocean. This bacterium was capable of performing anaerobic respiration using tellurite, tellurate, selenite and orthovanadate as terminal electron acceptors. While facultatively anaerobic, it could aerobically resist tellurite, selenite and orthovanadate up to 2000, 7000 and 10000 µg ml-1, respectively, reducing each oxide to elemental forms. Nearly complete 16S rRNA gene sequence similarity related the strain to Shewanella, with 98.8 and 98.7 % similarity to Shewanella basaltis and Shewanella algicola, respectively. The dominant fatty acids were C16 : 0 and C16 : 1. The major polar lipids were phosphatidylethanolamine and phosphatidylglycerol and MK-7 was the predominant quinone. DNA G+C content was 42.5 mol%. Computation of average nucleotide identity and digital DNA-DNA hybridization values with the closest phylogenetic neighbours of ER-Te-42B-LightT revealed genetic divergence at the species level, which was further substantiated by differences in several physiological characteristics. Based on the obtained results, this bacterium was assigned to the genus Shewanella as a new species with the name Shewanella metallivivens sp. nov., type strain ER-Te-42B-LightT (=VKM B-3580T=DSM 113370T).

Prosthecate aerobic anoxygenic phototrophs Photocaulis sulfatitolerans gen. nov. sp. nov. and Photocaulis rubescens sp. nov. isolated from alpine meromictic lakes in British Columbia, Canada.

Seven Gram-negative flagellated and subsequent prosthecate bacteria were isolated from meromictic Mahoney Lake and Blue Lake in British Columbia, Canada. Each became pink-red after 1-2 weeks of incubation, containing bacteriochlorophyll a incorporated into light harvesting and reaction center pigment-protein complexes. They did not grow anaerobically under illuminated conditions, supporting their identification as obligate aerobic anoxygenic phototrophs (AAP). All isolates preferred high salinity and BL14T tolerated up to 6.5% NaCl or 16.0% Na2SO4. In addition to phenotypic differences, analysis of 16S rRNA gene sequences found both strains BL14T and ML37T were related to Alkalicaulis satelles, G-192T by 98.41 and 98.84%, respectively, and distantly associated to members of the non-phototrophic genus Glycocaulis profundi, ZYF765T (95.59 and 95.36%, respectively) within the newly recognized Maricaulales order of α-Proteobacteria. BL14T and ML37T contained photosynthetic operons of 46,143 and 46,315 bp, where genes of BL14T were uniquely split into two distal operons. Furthermore, A. satelles was not originally published as an AAP, but was also found in this work to contain a similar 45,131 bp fragment. The distinct morphological features, physiological traits and genomic analysis including average nucleotide identity and digital DNA:DNA hybridization of circularized genomes supported the proposal of new genus and species Photocaulis sulfatitolerans gen. nov. sp. nov., type strain BL14T and Photocaulis rubescens sp. nov. type strain ML37T.

Extreme Environments and High-Level Bacterial Tellurite Resistance.

Bacteria have long been known to possess resistance to the highly toxic oxyanion tellurite, most commonly though reduction to elemental tellurium. However, the majority of research has focused on the impact of this compound on microbes, namely E. coli, which have a very low level of resistance. Very little has been done regarding bacteria on the other end of the spectrum, with three to four orders of magnitude greater resistance than E. coli. With more focus on ecologically-friendly methods of pollutant removal, the use of bacteria for tellurite remediation, and possibly recovery, further highlights the importance of better understanding the effect on microbes, and approaches for resistance/reduction. The goal of this review is to compile current research on bacterial tellurite resistance, with a focus on high-level resistance by bacteria inhabiting extreme environments.

Bioremediation potential of bacteria able to reduce high levels of selenium and tellurium oxyanions.

Environmental contamination by Te and Se oxyanions has become a serious concern, with the search for green, ecologically friendly methods for removal gaining ground. Bacteria capable of reducing these highly toxic compounds to a virtually non-toxic elemental form could provide a solution. In this study, four strains of bacteria with potential for bioremediation of Te and Se oxyanions were investigated. Under aerobic conditions over 48 h, Erythromicrobium ramosum, strain E5 removed 244 µg/ml tellurite and 98 µg/ml selenite, Erythromonas ursincola, KR99 203 µg/ml tellurite and 100 µg/ml selenite, AV-Te-18 98 µg/ml tellurite and 103 µg/ml selenite and ER-V-8 93 µg/ml tellurite and 103 µg/ml selenite. In the absence of oxygen, AV-Te-18 and ER-V-8 removed 10 µg/ml tellurite after 24 and 48 h, respectively and 46 and 25 µg/ml selenite, respectively, over 48 h. ER-V-8 removed 14 µg/ml selenate after 5 days. This highlights the great potential of these microbes for use in bioremediation.

Tellurite and Tellurate Reduction by the Aerobic Anoxygenic Phototroph Erythromonas ursincola, Strain KR99 Is Carried out by a Novel Membrane Associated Enzyme.

Erythromonas ursincola, strain KR99 isolated from a freshwater thermal spring of Kamchatka Island in Russia, resists and reduces very high levels of toxic tellurite under aerobic conditions. Reduction is carried out by a constitutively expressed membrane associated enzyme, which was purified and characterized. The tellurite reductase has a molecular weight of 117 kDa, and is comprised of two subunits (62 and 55 kDa) in a 1:1 ratio. Optimal activity occurs at pH 7.0 and 28 °C. Tellurite reduction has a Vmax of 5.15 µmol/min/mg protein and a Km of 3.36 mM. The enzyme can also reduce tellurate with a Vmax and Km of 1.08 µmol/min/mg protein and 1.44 mM, respectively. This is the first purified membrane associated Te oxyanion reductase.

Two distinct periplasmic enzymes are responsible for tellurite/tellurate and selenite reduction by strain ER-Te-48 associated with the deep sea hydrothermal vent tube worms at the Juan de Fuca Ridge black smokers.

Strain ER-Te-48 isolated from a deep-ocean hydrothermal vent tube worm is capable of resisting and reducing extremely high levels of tellurite, tellurate, and selenite, which are used for respiration anaerobically. Tellurite and tellurate reduction is accomplished by a periplasmic enzyme of 215 kDa comprised of 3 subunits (74, 42, and 25 kDa) in a 2:1:1 ratio. The optimum pH and temperature for activity is 8.0 and 35 °C, respectively. Tellurite reduction has a V max of 5.6 µmol/min/mg protein and a K m of 3.9 mM. In the case of the tellurate reaction, V max and K m were 2.6 µmol/min/mg protein and 2.6 mM, respectively. Selenite reduction is carried out by another periplasmic enzyme with a V max of 2.8 µmol/min/mg protein, K m of 12.1 mM, and maximal activity at pH 6.0 and 38 °C. This protein is 165 kDa and comprised of 3 subunits of 98, 44, and 23 kDa in a 1:1:1 ratio.

Aerobic anoxygenic phototrophs in gold mine tailings in Nopiming Provincial Park, Manitoba, Canada.

A sampling trip to Central Gold Mine, Nopiming Provincial Park, Canada, was taken in September 2011. Abundance, distribution, and physiology of aerobic anoxygenic phototrophs (AAP) from 4 locations were studied. Enumeration revealed 14.6% of culturable microbes were AAP. Five strains (NM4.16, NM4.18, C4, C9, C11) were chosen for analysis. All grow best on complex media without vitamin requirements and with an optimal pH 7.0-8.0, with strain C4 preferring pH 6.0. Strain NM4.18 tolerates the highest pH 11.0. Optimal temperature for all is 28 °C (range of 2-37 °C except NM4.16, which survives 45 °C). Strains C9, C11, and NM4.18 grew in 1.0%, 2.0%, and 5.0% NaCl, respectively, while NM4.16 and C4 grew only without NaCl. Isolates were all highly resistant to toxic metal(oid) oxides: tellurite (1500 µg/mL, all), tellurate (1500 µg/mL, C11), selenite (5000 µg/mL, C9, C11, and NM4.18), selenate (1000 µg/mL, C9 and C11), and orthometavanadate and metavanadate (5000 µg/mL, C11 and NM4.18). They could reduce tellurite to the less toxic elemental tellurium. Full 16S rRNA gene sequencing revealed all strains are Alphaproteobacteria, with C4 and NM4.16 closely related to Porphyrobacter colymbi (99.4% and 99.7% sequence similarity, respectively), C9 to Brevundimonas variabilis (99.1%), C11 to Brevundimonas bacteroides (98.6%), and NM4.18 to Erythromonas ursincola (98.5%).

A Diverse Community of Metal(loid) Oxide Respiring Bacteria Is Associated with Tube Worms in the Vicinity of the Juan de Fuca Ridge Black Smoker Field.

Epibiotic bacteria associated with tube worms living in the vicinity of deep sea hydrothermal vents of the Juan de Fuca Ridge in the Pacific Ocean were investigated for the ability to respire anaerobically on tellurite, tellurate, selenite, selenate, metavanadate and orthovanadate as terminal electron acceptors. Out of 107 isolates tested, 106 were capable of respiration on one or more of these oxides, indicating that metal(loid) oxide based respiration is not only much more prevalent in nature than is generally believed, but also is an important mode of energy generation in the habitat. Partial 16S rRNA gene sequencing revealed the bacterial community to be rich and highly diverse, containing many potentially new species. Furthermore, it appears that the worms not only possess a close symbiotic relationship with chemolithotrophic sulfide-oxidizing bacteria, but also with the metal(loid) oxide transformers. Possibly they protect the worms through reduction of the toxic compounds that would otherwise be harmful to the host.

Tellurite-, tellurate-, and selenite-based anaerobic respiration by strain CM-3 isolated from gold mine tailings.

The newly discovered strain CM-3, a Gram-negative, rod-shaped bacterium from gold mine tailings of the Central Mine in Nopiming Provincial Park, Canada, is capable of dissimilatory anaerobic reduction of tellurite, tellurate, and selenite. CM-3 possesses very high level resistance to these oxides, both aerobically and anaerobically. During aerobic growth, tellurite and tellurate resistance was up to 1500 and 1000 µg/ml, respectively. In the presence of selenite, growth occurred at the highest concentration tested, 7000 µg/ml. Under anaerobic conditions, resistance was decreased to 800 µg/ml for the Te oxides; however, much like under aerobic conditions, growth with selenite still took place at 7000 µg/ml. In the absence of oxygen, CM-3 couples oxide reduction to an increase in biomass. Following an initial drop in viable cells, due to switching from aerobic to anaerobic conditions, there was an increase in CFU/ml greater than one order of magnitude in the presence of tellurite (6.6 × 10(3)-8.6 × 10(4) CFU/ml), tellurate (4.6 × 10(3)-1.4 × 10(5) CFU/ml), and selenite (2.7 × 10(5)-5.6 × 10(6) CFU/ml). A control culture without metalloid oxides showed a steady decrease in CFU/ml with no recovery. ATP production was also increased in the presence of each oxide, further indicating anaerobic respiration. Partial 16S rRNA gene sequencing revealed a 99.0 % similarity of CM-3 to Pseudomonas reactans.

The Effect of Tellurite on Highly Resistant Freshwater Aerobic Anoxygenic Phototrophs and Their Strategies for Reduction.

Six fresh water aerobic anoxygenic phototrophs (Erythromicrobium ezovicum, strain E1; Erythromicrobium hydrolyticum, E4(1); Erythromicrobium ramosum, E5; Erythromonas ursincola, KR99; Sandaracinobacter sibiricus, RB 16-17; and Roseococcus thiosulfatophilus, RB3) possessing high level resistance to TeO3(2-) and the ability to reduce it to elemental Te were studied to understand their interaction with this highly toxic oxyanion. Tested organic carbon sources, pH, and level of aeration all had an impact on reduction. Physiological and metabolic responses of cells to tellurite varied among strains. In its presence, versus absence, cellular biomass either increased (KR99, 66.6% and E5, 21.2%) or decreased (RB3, 66.1%, E1, 57.8%, RB 16-17, 41.5%, and E4(1), 21.3%). The increase suggests a possible benefit from tellurite. Cellular ATP production was similarly affected, resulting in an increase (KR99, 15.2% and E5, 38.9%) or decrease (E4(1), 31.9%; RB 16-17, 48.8%; RB3, 55.9%; E1, 35.9%). Two distinct strategies to tellurite reduction were identified. The first, found in E4(1), requires de novo protein preparations as well as an undisturbed whole cell. The second strategy, in which reduction depended on a membrane associated constitutive reductase, was used by the remaining strains.