Widespread use of proton-pumping rhodopsin in Antarctic phytoplankton.

Photosynthetic carbon (C) fixation by phytoplankton in the Southern Ocean (SO) plays a critical role in regulating air-sea exchange of carbon dioxide and thus global climate. In the SO, photosynthesis (PS) is often constrained by low iron, low temperatures, and low but highly variable light intensities. Recently, proton-pumping rhodopsins (PPRs) were identified in marine phytoplankton, providing an alternate iron-free, light-driven source of cellular energy. These proteins pump protons across cellular membranes through light absorption by the chromophore retinal, and the resulting pH energy gradient can then be used for active membrane transport or for synthesis of adenosine triphosphate. Here, we show that PPR is pervasive in Antarctic phytoplankton, especially in iron-limited regions. In a model SO diatom, we found that it was localized to the vacuolar membrane, making the vacuole a putative alternative phototrophic organelle for light-driven production of cellular energy. Unlike photosynthetic C fixation, which decreases substantially at colder temperatures, the proton transport activity of PPR was unaffected by decreasing temperature. Cellular PPR levels in cultured SO diatoms increased with decreasing iron concentrations and energy production from PPR photochemistry could substantially augment that of PS, especially under high light intensities, where PS is often photoinhibited. PPR gene expression and high retinal concentrations in phytoplankton in SO waters support its widespread use in polar environments. PPRs are an important adaptation of SO phytoplankton to growth and survival in their cold, iron-limited, and variable light environment.

Identification of a Functionally Efficient and Thermally Stable Outward Sodium-Pumping Rhodopsin (BeNaR) from a Thermophilic Bacterium.

Rhodopsins are transmembrane proteins with retinal chromophores that are involved in photo-energy conversion and photo-signal transduction in diverse organisms. In this study, we newly identified and characterized a rhodopsin from a thermophilic bacterium, Bellilinea sp. Recombinant Escherichia coli cells expressing the rhodopsin showed light-induced alkalization of the medium only in the presence of sodium ions (Na+), and the alkalization signal was enhanced by addition of a protonophore, indicating an outward Na+ pump function across the cellular membrane. Thus, we named the protein Bellilinea Na+-pumping rhodopsin, BeNaR. Of note, its Na+-pumping activity is significantly greater than that of the known Na+-pumping rhodopsin, KR2. We further characterized its photochemical properties as follows: (i) Visible spectroscopy and HPLC revealed that BeNaR has an absorption maximum at 524 nm with predominantly (>96%) the all-trans retinal conformer. (ii) Time-dependent thermal denaturation experiments revealed that BeNaR showed high thermal stability. (iii) The time-resolved flash-photolysis in the nanosecond to millisecond time domains revealed the presence of four kinetically distinctive photointermediates, K, L, M and O. (iv) Mutational analysis revealed that Asp101, which acts as a counterion, and Asp230 around the retinal were essential for the Na+-pumping activity. From the results, we propose a model for the outward Na+-pumping mechanism of BeNaR. The efficient Na+-pumping activity of BeNaR and its high stability make it a useful model both for ion transporters and optogenetics tools.

Solar-panel and parasol strategies shape the proteorhodopsin distribution pattern in marine Flavobacteriia.

Proteorhodopsin (PR) is a light-driven proton pump that is found in diverse bacteria and archaea species, and is widespread in marine microbial ecosystems. To date, many studies have suggested the advantage of PR for microorganisms in sunlit environments. The ecophysiological significance of PR is still not fully understood however, including the drivers of PR gene gain, retention, and loss in different marine microbial species. To explore this question we sequenced 21 marine Flavobacteriia genomes of polyphyletic origin, which encompassed both PR-possessing as well as PR-lacking strains. Here, we show that the possession or alternatively the lack of PR genes reflects one of two fundamental adaptive strategies in marine bacteria. Specifically, while PR-possessing bacteria utilize light energy ("solar-panel strategy"), PR-lacking bacteria exclusively possess UV-screening pigment synthesis genes to avoid UV damage and would adapt to microaerobic environment ("parasol strategy"), which also helps explain why PR-possessing bacteria have smaller genomes than those of PR-lacking bacteria. Collectively, our results highlight the different strategies of dealing with light, DNA repair, and oxygen availability that relate to the presence or absence of PR phototrophy.

Pelagitalea pacifica gen. nov., sp. nov., a new marine bacterium isolated from seawater.

A strictly aerobic, Gram-negative, beige-pigmented, short-rod-shaped, non-motile and chemoheterotrophic bacteria, designated K2-48(T) was isolated from seawater collected in the Western North Pacific Ocean near Japan. Preliminary analysis based on the 16S rRNA gene sequence revealed that the novel isolate was affiliated with the family Oceanospirillaceae within the class Gammaproteobacteria and that it showed the highest sequence similarity (93.7 %) to Neptunomonas qingdaonensis P10-2-4(T). The strain could be differentiated phenotypically from recognized members of the family Oceanospirillaceae. The major fatty acids of strain K2-48(T) were identified as summed feature 3 (C16:1 ω7c and/or iso-C15:0 2-OH) and C16:0 as defined by the MIDI system. The DNA G+C content was determined to be 43.2 mol%, the major respiratory quinone was identified as ubiquinone 9 and a polar lipid profile was present consisting of phosphatidylethanolamine, a phosphatidylglycerol and an unidentified phospolipid. On the basis of polyphasic taxonomic studies, it was concluded that strain K2-48(T) represents a novel genus sp. We propose the name Pelagitalea pacifica gen. nov., sp. nov. for this strain; its type strain is K2-48(T) (=KCCM 90119(T)).

A light-driven sodium ion pump in marine bacteria.

Light-driven proton-pumping rhodopsins are widely distributed in many microorganisms. They convert sunlight energy into proton gradients that serve as energy source of the cell. Here we report a new functional class of a microbial rhodopsin, a light-driven sodium ion pump. We discover that the marine flavobacterium Krokinobacter eikastus possesses two rhodopsins, the first, KR1, being a prototypical proton pump, while the second, KR2, pumps sodium ions outward. Rhodopsin KR2 can also pump lithium ions, but converts to a proton pump when presented with potassium chloride or salts of larger cations. These data indicate that KR2 is a compatible sodium ion-proton pump, and spectroscopic analysis showed it binds sodium ions in its extracellular domain. These findings suggest that light-driven sodium pumps may be as important in situ as their proton-pumping counterparts.

Rubrivirga marina gen. nov., sp. nov., a member of the family Rhodothermaceae isolated from deep seawater.

Two aerobic, Gram-stain-negative, pale-red-pigmented and rod-shaped bacterial strains, designated SAORIC-26 and SAORIC-28(T), were isolated from seawater (3000 m depth) from the Pacific Ocean. Phylogenetic analysis based on their 16S rRNA gene sequences revealed that the novel isolates could be affiliated with the family Rhodothermaceae of the class Cytophagia. Strains SAORIC-26 and SAORIC-28(T) shared 99.7% pairwise sequence similarity with each other and showed less than 92.6% similarity with other cultivated members of the class Cytophagia. The strains were found to be non-motile, oxidase-positive, catalase-negative and able to hydrolyse gelatin and aesculin. The DNA G+C contents were determined to be 64.8-65.8 mol% and MK-7 was the predominant menaquinone. Summed feature 9 (iso-C17:1ω9c and/or C16:0 10-methyl), summed feature 3 (C16:1ω6c and/or C16:1ω7c) and iso-C15:0 were found to be the major cellular fatty acids. On the basis of this taxonomic study using a polyphasic approach, it was concluded that strains SAORIC-26 and SAORIC-28(T) represent a novel species of a new genus in the family Rhodothermaceae, for which the name Rubrivirga marina gen. nov., sp. nov. is proposed. The type strain of the type species of is SAORIC-28(T) (=KCTC 23867(T)=NBRC 108816(T)). An additional strain of the species is SAORIC-26.

Nonlabens marinus [corrected] sp. nov., a novel member of the Flavobacteriaceae isolated from the Pacific Ocean.

Two aerobic, Gram-negative, orange pigmented and irregular rod-shaped bacteria, designated S1-05 and S1-08(T), were isolated from seawater from the Pacific Ocean. Phylogenetic analysis based on their 16S rRNA gene sequences revealed that the novel isolates could be affiliated with the genus Nonlabens of the family Flavobacteriaceae. The strains S1-05 and S1-08(T) shared 100 % pairwise sequences similarity with each other and showed less than 96.8 % similarity with the cultivated members of the genus Nonlabens. The novel isolates are phenotypically and physiologically different from strains described previously. The strains were found to be non-motile, oxidase positive, catalase positive and hydrolyzed gelatin and aesculin. The G+C contents of the DNA were determined to 41.4 and 41.7 mol% and MK-6 the predominant menaquinone. Anteiso-C15:0 and iso-C15:0 were found to be the major two cellular fatty acids. On the basis of polyphasic taxonomic studies, it was concluded that strains S1-05 and S1-08(T) represent a novel species within the genus Nonlabens, for which the name Nonlabens marina sp. nov. is proposed. The type strain of N. marina is S1-08(T) (=KCTC 23432(T) = NBRC 107738(T)).

Oceanicoccus sagamiensis gen. nov., sp. nov., a gammaproteobacterium isolated from sea water of Sagami Bay in Japan.

A gram-negative, motile, coccoid- and amorphous-shaped, non-pigmented chemoheterotrophic bacterium, designated strain PZ-5(T), was isolated from sea water of Sagami Bay in Japan and subjected to a polyphasic taxonomic study. Phylogenetic analysis based on 16S rRNA gene sequences revealed that the novel isolate could be affiliated with the class Gammaproteobacteria. Strain PZ-5(T) showed below 93.9% similarity with validly published bacteria and demonstrated the highest sequence similarity to Dasania marina KOPRI 20902(T) (93.9%). Strain PZ-5(T) formed a monophyletic group with D. marina KOPRI 20902(T). The DNA G+C content of strain PZ-5(T) was 49.8 mol%. The major isoprenoid quinone was Q-8 and predominant cellular fatty acids were C(15:0) ISO 20H (19%), C(16:1) ω7c (17.4%), C(17;1) ω8c (16.2%), C(11:0) 3OH (7.5%), and C(15:1) ω8c (6.5%). Based on evidence from a polyphasic taxonomical study, it was concluded that the strain should be classified as representing a new genus and species of the class Gammaproteobacteria, for which the name Oceanicoccus sagamiensis gen. nov., sp. nov., (type strain PZ-5(T) =NBRC 107125(T) =KCTC 23278(T)) is proposed.

Psychrosphaera saromensis gen. nov., sp. nov., within the family Pseudoalteromonadaceae, isolated from Lake Saroma, Japan.

Three Gram-negative, motile, coccoid- and ellipsoidal-shaped, non-pigmented, chemoheterotrophic bacteria, designated strains SA4-31, SA4-46 and SA4-48(T), were isolated from Lake Saroma in Japan and subjected to a polyphasic taxonomical study. 16S rRNA gene sequence analysis revealed that the novel isolates could be affiliated to the family Pseudoalteromonadaceae of the order Alteromonadales. The strains shared approximately 99.7-100% sequence similarity with each other and showed 89.5-93.2% similarity with members of the family Pseudoalteromonadaceae with validly published names. The DNA-DNA relatedness among the strains SA 4-31, SA 4-46 and SA 4-48(T) was higher than 80%, a value that is accepted as a phylogenetic definition of one species. The DNA G+C contents of the three strains were 38.7-39.6 mol%. The major isoprenoid quinone was Q-8 and C16:0, C16:1 ω7c, C18:1 ω7c and C12:1 3OH were the major fatty acids. Based on the evidence from the polyphasic taxonomical study, it was concluded that the three strains should be classified as representing a new genus and species of the family Pseudoalteromonadaceae, for which the name Psychrosphaera saromensis gen. nov., sp. nov. (type strain SA4-48(T) =NBRC 107123(T)= KCTC 23240(T)) is proposed.

Vibrio azureus sp. nov., a luminous marine bacterium isolated from seawater.

Two luminous marine bacterial strains, LC2-005(T) and LC2-102, were isolated from seawater at Kuroshio Region and Sagami Bay in Japan, respectively. These bacteria were Gram-negative, oxidase-positive, catalase-positive, motile and rod-shaped. On the basis of 16S rRNA gene sequence analysis, strains LC2-005(T) and LC2-102 formed a cluster within the Vibrio harveyi species group. However, multilocus sequence analysis using five loci (pyrH, ftsZ, mreB, gyrB and gapA) and DNA-DNA hybridization experiments indicated that these strains were distinct from the currently known Vibrio species. Additionally, these strains differ from related Vibrio species in utilization of glucose, mannitol, inositol, sorbitol, rhamnose, sucrose, melibiose and arabinose, production of lysine decarboxylase, ornithine decarboxylase, tryptophan deaminase, esterase (C4), lipase (C4), chymotrypsin, acid phosphatase, alpha-glucosidase, beta-glucosidase and N-acetyl-beta-glucosaminidase and the ability to reduce nitrate to nitrite. The major fatty acids were C(15 : 0) iso 2-OH and/or C(16 : 1)omega7c, C(16 : 0), C(18 : 1)omega7c and C(14 : 0). The DNA G+C contents of strains LC2-005(T) and LC2-102 were 45.2 and 45.5 mol%, respectively. On the basis of the polyphasic taxonomic evidence presented in this study, it can be concluded that strains LC2-005(T) and LC2-102 belong to the same genospecies and represent a novel species of the genus Vibrio, for which the name Vibrio azureus sp. nov. is proposed. The type strain is LC2-005(T) (=NBRC 104587(T) =KCTC 22352(T)).