A bacterium from a mountain lake harvests light using both proton-pumping xanthorhodopsins and bacteriochlorophyll-based photosystems.

Photoheterotrophic bacteria harvest light energy using either proton-pumping rhodopsins or bacteriochlorophyll (BChl)-based photosystems. The bacterium Sphingomonas glacialis AAP5 isolated from the alpine lake Gossenköllesee contains genes for both systems. Here, we show that BChl is expressed between 4°C and 22°C in the dark, whereas xanthorhodopsin is expressed only at temperatures below 16°C and in the presence of light. Thus, cells grown at low temperatures under a natural light-dark cycle contain both BChl-based photosystems and xanthorhodopsins with a nostoxanthin antenna. Flash photolysis measurements proved that both systems are photochemically active. The captured light energy is used for ATP synthesis and stimulates growth. Thus, S. glacialis AAP5 represents a chlorophototrophic and a retinalophototrophic organism. Our analyses suggest that simple xanthorhodopsin may be preferred by the cells under higher light and low temperatures, whereas larger BChl-based photosystems may perform better at lower light intensities. This indicates that the use of two systems for light harvesting may represent an evolutionary adaptation to the specific environmental conditions found in alpine lakes and other analogous ecosystems, allowing bacteria to alternate their light-harvesting machinery in response to large seasonal changes of irradiance and temperature.

The Centres for Disease Control light trap (CDC-LT) and the human decoy trap (HDT) compared to the human landing catch (HLC) for measuring Anopheles biting in rural Tanzania.

BACKGROUND: Vector mosquito biting intensity is an important measure to understand malaria transmission. Human landing catch (HLC) is an effective but labour-intensive, expensive, and potentially hazardous entomological surveillance tool. The Centres for Disease Control light trap (CDC-LT) and the human decoy trap (HDT) are exposure-free alternatives. This study compared the CDC-LT and HDT against HLC for measuring Anopheles biting in rural Tanzania and assessed their suitability as HLC proxies. METHODS: Indoor mosquito surveys using HLC and CDC-LT and outdoor surveys using HLC and HDT were conducted in 2017 and in 2019 in Ulanga, Tanzania in 19 villages, with one trap/house/night. Species composition, sporozoite rates and density/trap/night were compared. Aggregating the data by village and month, the Bland-Altman approach was used to assess agreement between trap types. RESULTS: Overall, 66,807 Anopheles funestus and 14,606 Anopheles arabiensis adult females were caught with 6,013 CDC-LT, 339 indoor-HLC, 136 HDT and 195 outdoor-HLC collections. Indoors, CDC-LT caught fewer An. arabiensis (Adjusted rate ratio [Adj.RR] = 0.35, 95% confidence interval [CI]: 0.27-0.46, p < 0.001) and An. funestus (Adj.RR = 0.63, 95%CI: 0.51-0.79, p < 0.001) than HLC per trap/night. Outdoors, HDT caught fewer An. arabiensis (Adj.RR = 0.04, 95%CI: 0.01-0.14, p < 0.001) and An. funestus (Adj.RR = 0.10, 95%CI: 0.07-0.15, p < 0.001) than HLC. The bias and variability in number of mosquitoes caught by the different traps were dependent on mosquito densities. The relative efficacies of both CDC-LT and HDT in comparison to HLC declined with increased mosquito abundance. The variability in the ratios was substantial for low HLC counts and decreased as mosquito abundance increased. The numbers of sporozoite positive mosquitoes were low for all traps. CONCLUSIONS: CDC-LT can be suitable for comparing mosquito populations between study arms or over time if accuracy in the absolute biting rate, compared to HLC, is not required. CDC-LT is useful for estimating sporozoite rates because large numbers of traps can be deployed to collect adequate mosquito samples. The present design of the HDT is not amenable for use in large-scale entomological surveys. Use of HLC remains important for estimating human exposure to mosquitoes as part of estimating the entomological inoculation rate (EIR).

Unique double concentric ring organization of light harvesting complexes in Gemmatimonas phototrophica.

The majority of life on Earth depends directly or indirectly on the sun as a source of energy. The initial step of photosynthesis is facilitated by light-harvesting complexes, which capture and transfer light energy into the reaction centers (RCs). Here, we analyzed the organization of photosynthetic (PS) complexes in the bacterium G. phototrophica, which so far is the only phototrophic representative of the bacterial phylum Gemmatimonadetes. The isolated complex has a molecular weight of about 800 ± 100 kDa, which is approximately 2 times larger than the core complex of Rhodospirillum rubrum. The complex contains 62.4 ± 4.7 bacteriochlorophyll (BChl) a molecules absorbing in 2 distinct infrared absorption bands with maxima at 816 and 868 nm. Using femtosecond transient absorption spectroscopy, we determined the energy transfer time between these spectral bands as 2 ps. Single particle analyses of the purified complexes showed that they were circular structures with an outer diameter of approximately 18 nm and a thickness of 7 nm. Based on the obtained, we propose that the light-harvesting complexes in G. phototrophica form 2 concentric rings surrounding the type 2 RC. The inner ring (corresponding to the B868 absorption band) is composed of 15 subunits and is analogous to the inner light-harvesting complex 1 (LH1) in purple bacteria. The outer ring is composed of 15 more distant BChl dimers with no or slow energy transfer between them, resulting in the B816 absorption band. This completely unique and elegant organization offers good structural stability, as well as high efficiency of light harvesting. Our results reveal that while the PS apparatus of Gemmatimonadetes was acquired via horizontal gene transfer from purple bacteria, it later evolved along its own pathway, devising a new arrangement of its light harvesting complexes.

Temperature dependence of photosynthetic reaction centre activity in Rhodospirillum rubrum.

The influence of temperature on photosynthetic reactions was investigated by a combination of time-resolved bacteriochlorophyll fluorescence, steady-state and differential absorption spectroscopy, and polarographic respiration measurements in intact cells of purple non-sulphur bacterium Rhodospirillum rubrum. Using variable bacteriochlorophyll fluorescence, it was found that the electron-transport activity increased with the increasing temperature up to 41 °C. The fast and medium components of the fluorescence decay kinetics followed the ideal Arrhenius equation. The calculated activation energy for the fast component was Ea1 = 16 kJ mol-1, while that of the medium component was more than double, with Ea2 = 38 kJ mol-1. At temperatures between 41 and 59 °C, the electron transport was gradually, irreversibly inhibited. Interestingly, the primary charge separation remained fully competent from 20 to 59 °C as documented by both BChl fluorescence and differential absorption spectroscopy of the P870+ signal. At temperatures above 60 °C, the primary photochemistry became reversibly inhibited, which was manifested by an increase in minimal fluorescence, F0, whereas maximal fluorescence, FM, slowly declined. Finally, above 71 °C, the photosynthetic complexes began to disassemble as seen in the decline of all fluorometric parameters and the disappearance of the LH1 absorption band at 880 nm. The extended optimal temperature of photosynthetic reaction centre in a model species of Rhodospirillales adds on the evidence that the good thermostability of the photosynthetic reaction centres is present across all Alphaproteobacteria.

High diversity of thermophilic cyanobacteria in Rupite hot spring identified by microscopy, cultivation, single-cell PCR and amplicon sequencing.

Genotypic and morphological diversity of cyanobacteria in the Rupite hot spring (Bulgaria) was investigated by means of optical microscopy, cultivation, single-cell PCR, and 16S rRNA gene amplicon sequencing. Altogether, 34 sites were investigated along the 71-39 °C temperature gradient. Analysis of samples from eight representative sites shown that Illumina, optical microscopy, and Roche 454 identified 72, 45 and 19% respective occurrences of all cumulatively present taxa. Optical microscopy failed to detect species of minor occurrence; whereas, amplicon sequencing technologies suffered from failed primer annealing and the presence of species with extensive extracellular polysaccharides production. Amplicon sequencing of the 16S rRNA gene V5-V6 region performed by Illumina identified the cyanobacteria most reliably to the generic level. Nevertheless, only the combined use of optical microscopy, cultivation and sequencing methods allowed for reliable estimate of the cyanobacterial diversity. Here, we show that Rupite hot-spring system hosts one of the richest cyanobacterial flora reported from a single site above 50 °C. Chlorogloeopsis sp. was the most abundant at the highest temperature (68 °C), followed by Leptolyngbya boryana, Thermoleptolyngbya albertanoae, Synechococcus bigranulatus, Oculatella sp., and Desertifilum sp. thriving above 60 °C, while Leptolyngbya geysericola, Geitlerinema splendidum, and Cyanobacterium aponinum were found above 50 °C.

Nonlinear effect of irradiance on photoheterotrophic activity and growth of the aerobic anoxygenic phototrophic bacterium Dinoroseobacter shibae.

Aerobic anoxygenic photosynthetic bacteria are an important component of marine microbial communities. They produce energy in light using bacteriochlorophyll a containing photosystems. This extra energy provides an advantage over purely heterotrophic bacteria. One of the most intensively studied AAP bacteria is Dinoroseobacter shibae, a member of the environmentally important Roseobacter clade. Light stimulates its growth and metabolism, but the effect of light intensity remains unclear. Here, we show that an increase in biomass along an irradiance gradient followed the exponential rise to the maximum curve, with saturation at about 300 µmol photons m-2 s-1 , without any inhibition at light intensities up to 600 µmol photons m-2 s-1 . The cells adapted to higher irradiance by reducing pigmentation and increasing the electron transfer rate. This additional energy allowed D. shibae to redirect the metabolism of organic carbon sources such as glucose, leucine, glutamate, acetate and pyruvate toward anabolism, resulting in a twofold increase of their assimilation rates. We provide equations that can be feasibly incorporated into the existing model of D. shibae metabolism to further advance our understanding of the role of photoheterotrophy in the ocean.

Characterization of the microaerophilic, bacteriochlorophyll a-containing bacterium Gemmatimonas phototrophica sp. nov., and emended descriptions of the genus Gemmatimonas and Gemmatimonas aurantiaca.

A red-pigmented, bacteriochlorophyll (BChl) a-producing strain, AP64T, was isolated previously from the freshwater Swan Lake located in the western Gobi Desert. Based on its 16S rRNA gene sequence identity (96.1%) to the type strain Gemmatimonas aurantiaca T-27T, the new isolate was tentatively classified as a member of the bacterial phylum Gemmatimonadetes. Here, we report its formal description and polyphasic characterization. Strain AP64T grew best on agar media under 9.8-15.2% atmospheric oxygen. The cells were rods, dividing by symmetrical or asymmetrical binary fission. Budding structures were also observed. Its genomic DNA G+C content was 64.4% (from the draft genome sequence). Phylogenetic analysis based on the 16S rRNA gene sequence clearly separated AP64T from related species. Its genotypic differentiation from phylogenetically close relatives was further supported by performing in silico DNA-DNA hybridization and calculating average nucleotide identity, whereas the high percentage (67.3%) of shared conserved proteins between strain AP64T and Gemmatimonas aurantiaca T-27T supports the classification of the two strains into the same genus. Strain AP64T contained C16 : 1, C14 : 1 and C18 : 1ω9c as predominant fatty acids. The main respiratory quinone was menaquinone 8 (MK-8). The most distinctive feature of strain AP64T was the presence of fully functional purple bacterial photosynthetic reaction centres. The main CO2-fixation pathways were absent. Strain AP64T was capable of growth and BChl production in constant darkness. Thus, strain AP64T is a facultatively photoheterotrophic organism. It represents a novel species of the genus Gemmatimonas, for which the name Gemmatimonasphototrophica sp. nov. is proposed. The type strain is AP64T ( = DSM 29774T = MCCC 1K00454T). Emended descriptions of the genus Gemmatimonas and Gemmatimonas aurantiaca are also provided.

A novel tumor necrosis factor-mediated mechanism of direct epithelial sodium channel activation.

RATIONALE: Alveolar liquid clearance is regulated by Na(+) uptake through the apically expressed epithelial sodium channel (ENaC) and basolaterally localized Na(+)-K(+)-ATPase in type II alveolar epithelial cells. Dysfunction of these Na(+) transporters during pulmonary inflammation can contribute to pulmonary edema. OBJECTIVES: In this study, we sought to determine the precise mechanism by which the TIP peptide, mimicking the lectin-like domain of tumor necrosis factor (TNF), stimulates Na(+) uptake in a homologous cell system in the presence or absence of the bacterial toxin pneumolysin (PLY). METHODS: We used a combined biochemical, electrophysiological, and molecular biological in vitro approach and assessed the physiological relevance of the lectin-like domain of TNF in alveolar liquid clearance in vivo by generating triple-mutant TNF knock-in mice that express a mutant TNF with deficient Na(+) uptake stimulatory activity. MEASUREMENTS AND MAIN RESULTS: TIP peptide directly activates ENaC, but not the Na(+)-K(+)-ATPase, upon binding to the carboxy-terminal domain of the α subunit of the channel. In the presence of PLY, a mediator of pneumococcal-induced pulmonary edema, this binding stabilizes the ENaC-PIP2-MARCKS complex, which is necessary for the open probability conformation of the channel and preserves ENaC-α protein expression, by means of blunting the protein kinase C-α pathway. Triple-mutant TNF knock-in mice are more prone than wild-type mice to develop edema with low-dose intratracheal PLY, correlating with reduced pulmonary ENaC-α subunit expression. CONCLUSIONS: These results demonstrate a novel TNF-mediated mechanism of direct ENaC activation and indicate a physiological role for the lectin-like domain of TNF in the resolution of alveolar edema during inflammation.

A photoheterotrophic bacterium from Iceland has adapted its photosynthetic machinery to the long days of polar summer.

During their long evolution, anoxygenic phototrophic bacteria have inhabited a wide variety of natural habitats and developed specific strategies to cope with the challenges of any particular environment. Expression, assembly, and safe operation of the photosynthetic apparatus must be regulated to prevent reactive oxygen species generation under illumination in the presence of oxygen. Here, we report on the photoheterotrophic Sediminicoccus sp. strain KRV36, which was isolated from a cold stream in north-western Iceland, 30 km south of the Arctic Circle. In contrast to most aerobic anoxygenic phototrophs, which stop pigment synthesis when illuminated, strain KRV36 maintained its bacteriochlorophyll synthesis even under continuous light. Its cells also contained between 100 and 180 chromatophores, each accommodating photosynthetic complexes that exhibit an unusually large carotenoid absorption spectrum. The expression of photosynthesis genes in dark-adapted cells was transiently downregulated in the first 2 hours exposed to light but recovered to the initial level within 24 hours. An excess of membrane-bound carotenoids as well as high, constitutive expression of oxidative stress response genes provided the required potential for scavenging reactive oxygen species, safeguarding bacteriochlorophyll synthesis and photosystem assembly. The unique cellular architecture and an unusual gene expression pattern represent a specific adaptation that allows the maintenance of anoxygenic phototrophy under arctic conditions characterized by long summer days with relatively low irradiance.IMPORTANCEThe photoheterotrophic bacterium Sediminicoccus sp. KRV36 was isolated from a cold stream in Iceland. It expresses its photosynthesis genes, synthesizes bacteriochlorophyll, and assembles functional photosynthetic complexes under continuous light in the presence of oxygen. Unraveling the molecular basis of this ability, which is exceptional among aerobic anoxygenic phototrophic species, will help to understand the evolution of bacterial photosynthesis in response to changing environmental conditions. It might also open new possibilities for genetic engineering of biotechnologically relevant phototrophs, with the aim of increasing photosynthetic activity and their tolerance to reactive oxygen species.

Cost-effectiveness of the dual prevention pill for contraception and HIV pre-exposure prophylaxis.

Introduction: Women in sub-Saharan Africa (SSA) experience the world's highest rates of both HIV infection and unintended pregnancy. The Dual Prevention Pill (DPP) is a novel multipurpose prevention technology (MPT) that co-formulates HIV pre-exposure prophylaxis (PrEP) and combined hormonal oral contraception into a single daily pill. As a dual indication product, the DPP may be preferred by women facing these overlapping health risks. However, most SSA countries face severe healthcare resource constraints. Research is needed to assess whether, in what populations, and in what use cases the DPP would be cost-effective. Methods: We augmented an agent-based SSA HIV model with maternal health parameters including unintended pregnancy, abortion, and maternal mortality. Based on a previous market analysis, we assumed a primary DPP user population of current oral contraceptive users ages 25-49, and alternative user populations in different risk groups (age 15-24, sex workers, HIV-serodiscordant couples) and baseline product use profiles (unmet need for contraception, oral PrEP use, condom use). In three geographies (western Kenya, Zimbabwe, South Africa), we estimated HIV infections averted, pregnancies averted, disability-adjusted life-years (DALYs) averted, and the incremental cost-effectiveness ratio (ICER) over a 30-year time horizon, assuming equivalent adherence to the DPP as to oral contraceptives, higher adherence, or lower adherence. Results: The DPP is likely to be a cost-effective alternative to oral PrEP among users in need of contraception. Among women not already using PrEP, the DPP is likely to be cost-saving in sex workers and serodiscordant couples. The DPP is unlikely to be cost-effective in oral contraceptive users in the general population. Switching from oral contraception to the DPP could be net harmful in some settings and populations if it were to substantially reduces adherence to oral contraception. Results were robust to a range of time horizons or discount rates. Conclusion: The DPP has the potential to be cost-effective and cost-saving in populations at substantial HIV risk. Outcomes are sensitive to adherence, implying that effective counseling and decision-making tools for users considering the DPP will be essential. More research is needed to understand real-life adherence patterns and ensure health benefits achieved from contraception alone are not lost.

Protein flexibility acclimatizes photosynthetic energy conversion to the ambient temperature.

Adjustment of catalytic activity in response to diverse ambient temperatures is fundamental to life on Earth. A crucial example of this is photosynthesis, where solar energy is converted into electrochemical potential that drives oxygen and biomass generation at temperatures ranging from those of frigid Antarctica to those of scalding hot springs. The energy conversion proceeds by concerted mobilization of electrons and protons on photoexcitation of reaction centre protein complexes. Following physicochemical paradigms, the rates of imperative steps in this process were predicted to increase exponentially with rising temperatures, resulting in different yields of solar energy conversion at the distinct growth temperatures of photosynthetic mesophiles and extremophiles. In contrast, here we show a meticulous adjustment of energy conversion rate, resulting in similar yields from mesophiles and thermophiles. The key molecular players in the temperature adjustment process consist of a cluster of hitherto unrecognized protein cavities and an adjacent packing motif that jointly impart local flexibility crucial to the reaction centre proteins. Mutations within the packing motif of mesophiles that increase the bulkiness of the amino-acid side chains, and thus reduce the size of the cavities, promote thermophilic behaviour. This novel biomechanical mechanism accounts for the slowing of the catalytic reaction above physiological temperatures in contradiction to the classical Arrhenius paradigm. The mechanism provides new guidelines for manipulating the acclimatization of enzymes to the ambient temperatures of diverse habitats. More generally, it reveals novel protein elements that are of potential significance for modulating structure-activity relationships in membrane and globular proteins alike.

2.4-Å structure of the double-ring Gemmatimonas phototrophica photosystem.

Phototrophic Gemmatimonadetes evolved the ability to use solar energy following horizontal transfer of photosynthesis-related genes from an ancient phototrophic proteobacterium. The electron cryo-microscopy structure of the Gemmatimonas phototrophica photosystem at 2.4 Å reveals a unique, double-ring complex. Two unique membrane-extrinsic polypeptides, RC-S and RC-U, hold the central type 2 reaction center (RC) within an inner 16-subunit light-harvesting 1 (LH1) ring, which is encircled by an outer 24-subunit antenna ring (LHh) that adds light-gathering capacity. Femtosecond kinetics reveal the flow of energy within the RC-dLH complex, from the outer LHh ring to LH1 and then to the RC. This structural and functional study shows that G. phototrophica has independently evolved its own compact, robust, and highly effective architecture for harvesting and trapping solar energy.

Naturally zinc-containing bacteriochlorophyll a ([Zn]-BChl a) protects the photosynthetic apparatus of Acidiphilium rubrum from copper toxicity damage.

In almost all photosynthetic organisms the photosynthetic pigments chlorophyll and bacteriochlorophyll (BChl) are Mg2+ containing complexes, but Mg2+ may be exchanged against other metal ions when these are present in toxic concentrations, leading to inactivation of photosynthesis. In this report we studied mechanisms of copper toxicity to the photosynthetic apparatus of Acidiphilium rubrum, an acidophilic purple bacterium that uses Zn2+ instead of Mg2+ as the central metal in the BChl molecules ([Zn]-BChl) of its reaction centres (RCs) and light harvesting proteins (LH1). We used a combination of in vivo measurements of photosynthetic activity (fast fluorescence and absorption kinetics) together with analysis of metal binding to pigments and pigment-protein complexes by HPLC-ICP-sfMS to monitor the effect of Cu2+ on photosynthesis of A. rubrum. Further, we found that its cytoplasmic pH is neutral. We compared these results with those obtained from Rhodospirillum rubrum, a purple bacterium for which we previously reported that the central Mg2+ of BChl can be replaced in vivo in the RCs by Cu2+ under environmentally realistic Cu2+ concentrations, leading to a strong inhibition of photosynthesis. Thus, we observed that A. rubrum is much more resistant to copper toxicity than R. rubrum. Only slight changes of photosynthetic parameters were observed in A. rubrum at copper concentrations that were severely inhibitory in R. rubrum and in A. rubrum no copper complexes of BChl were found. Altogether, the data suggest that [Zn]-BChl protects the photosynthetic apparatus of A. rubrum from detrimental insertion of Cu2+ (trans-metallation) into BChl molecules of its RCs.

Characterization of the Aerobic Anoxygenic Phototrophic Bacterium Sphingomonas sp. AAP5.

An aerobic, yellow-pigmented, bacteriochlorophyll a-producing strain, designated AAP5 (=DSM 111157=CCUG 74776), was isolated from the alpine lake Gossenköllesee located in the Tyrolean Alps, Austria. Here, we report its description and polyphasic characterization. Phylogenetic analysis of the 16S rRNA gene showed that strain AAP5 belongs to the bacterial genus Sphingomonas and has the highest pairwise 16S rRNA gene sequence similarity with Sphingomonas glacialis (98.3%), Sphingomonas psychrolutea (96.8%), and Sphingomonas melonis (96.5%). Its genomic DNA G + C content is 65.9%. Further, in silico DNA-DNA hybridization and calculation of the average nucleotide identity speaks for the close phylogenetic relationship of AAP5 and Sphingomonas glacialis. The high percentage (76.2%) of shared orthologous gene clusters between strain AAP5 and Sphingomonas paucimobilis NCTC 11030T, the type species of the genus, supports the classification of the two strains into the same genus. Strain AAP5 was found to contain C18:1ω7c (64.6%) as a predominant fatty acid (>10%) and the polar lipid profile contained phosphatidylglycerol, diphosphatidylglycerol, phosphatidylethanolamine, sphingoglycolipid, six unidentified glycolipids, one unidentified phospholipid, and two unidentified lipids. The main respiratory quinone was ubiquinone-10. Strain AAP5 is a facultative photoheterotroph containing type-2 photosynthetic reaction centers and, in addition, contains a xathorhodopsin gene. No CO2-fixation pathways were found.

Utilization of light energy in phototrophic Gemmatimonadetes.

Gemmatimonas phototrophica is, so far, the only described phototrophic species of the bacterial phylum Gemmatimonadetes. Its cells contain a unique type of photosynthetic complex with the reaction center surrounded by a double ring antenna, however they can also grow in the dark using organic carbon substrates. Its photosynthesis genes were received via horizontal gene transfer from Proteobacteria. This raises two questions; how the horizontally transferred photosynthesis apparatus has integrated into the cellular machinery, and how much light-derived energy actually contributes to the cellular metabolism? To address these points, the photosynthetic reactions were studied on several levels, from photophysics of the reaction center to cellular growth. Flash photolysis measurements and bacteriochlorophyll fluorescence kinetic measurements documented the presence of fully functional type-2 reaction centers with a large light harvesting antenna. When illuminated, the bacterial cells reduced their respiration rate by 58 ±â€¯5%, revealing that oxidative phosphorylation was replaced by photophosphorylation. Moreover, illumination also more than doubled the assimilation rates of glucose, a sugar that is mostly used for respiration. Finally, light increased the growth rates of Gemmatimonas phototrophica colonies on agar plates. All the presented data provide evidence that photosynthetic complexes are fully integrated into cellular metabolism of Gemmatimonas phototrophica, and are able to provide a substantial amount of energy for its metabolism and growth.

Diversity of cyanobacteria at the Alaska North Slope with description of two new genera: Gibliniella and Shackletoniella.

The diversity of cyanobacteria along the Alaskan North Slope was investigated. We isolated and cultivated 57 strains of cyanobacteria and sequenced a section of their rRNA operon containing a fragment of the 16S rRNA gene. Here, we describe 17 found species belonging mainly to families Coleofasciculaceae, Microcoleaceae, Oculatellaceae, Leptolyngbyaceae and to the order Synechococcales. In pursuing a conservative polyphasic approach, we utilized suggested thresholds in 16S rRNA gene differences in parallel with morphological differences between new and already described taxa for the description of new species and genera. Based on a combination of morphological, molecular and ecological analysis of collected and cultured strains we describe two genera Gibliniella and Shackletoniella as well as six cyanobacterial species; Cephalothrix alaskaensis, Tildeniella alaskaensis, Pseudophormidium americanum, Leptodesmis alaskaensis, Albertania alaskaensis and Nodosilinea alaskaensis. Here, a polyphasic approach was used to identify eight novel and nine established cyanobacterial taxa from a previously non-investigated region that uncovered a high degree of biodiversity in extreme polar environments.

Targeting mutations to the plastidial psbA gene of Chlamydomonas reinhardtii without direct positive selection.

Targeting mutations to specific genomic loci is invaluable for assessing in vivo the effect of these changes on the biological role of the gene in study. Here, we attempted to introduce a mutation that was previously implicated in an increased heat stability of the mesophilic cyanobacterium Synechocystis sp. PCC6803 via homologous recombination to the psbA gene of Chlamydomonas reinhardtii. For that, we established a strategy for targeted mutagenesis that was derived from the efficient genome-wide homologous-recombination-based methodology that was used to target individual genes of Saccharomyces cerevisiae. While the isolated mutants did not show any benefit under elevated temperature conditions, the new strategy proved to be efficient for C. reinhardtii even in the absence of direct positive selection.

Analyzing carotenoids of snow algae by Raman microspectroscopy and high-performance liquid chromatography.

We tested the potential of Raman microspectroscopy to determine carotenoid pigments - both primary (lutein, beta-carotene) and secondary (astaxanthin) carotenoids - in the different species and life-cycle stages of snow algae from the order Chlamydomonadales (Chlorophyta). We compared the performance of Raman spectrometry to a reference method of biological pigment analysis, high-performance liquid chromatography (HPLC). The three main carotenoid Raman bands of the astaxanthin-rich red cysts were located at 1520, 1156 and 1006 cm-1. The shifts (orange aplanozygotes and green motile cells with flagella) in the position of the ν1(CC) Raman band of the polyenic chain is consistent with the expected changes in the ratios of the various carotenoid pigments. Flagellated green cells commonly contain lutein as a major carotenoid, together with minor amounts of ß­carotene and varying amounts of antheraxanthin, violaxanthin and neoxanthin. Aplanozygotes contain mixtures of both primary and secondary carotenoids. In most cases, the ν1(CC) band is an overlapping set of bands, which is due to the signal of all carotenoid pigments in the sample, and a deconvolution along with the band position shifts (mainly ν1) could be used to characterize the mixture of carotenoids. However, the ability of Raman spectroscopy to discriminate between structurally slightly differing carotenoid pigments or several carotenoids in an admixture in an unknown biological system remains limited.

Mechanisms of sublethal copper toxicity damage to the photosynthetic apparatus of Rhodospirillum rubrum.

Magnesium (Mg2+) is the ubiquitous metal ion present in chlorophyll and bacteriochlorophyll (BChl), involved in photosystems in photosynthetic organisms. In the present study we investigated targets of toxic copper binding to the photosynthetic apparatus of the anoxygenic purple bacterium Rhodospirillum rubrum. This was done by a combination of in vivo measurements of flash photolysis and fast fluorescence kinetics combined with the analysis of metal binding to pigments and pigment-protein complexes isolated from Cu-stressed cells by HPLC-ICPMS (ICP-sfMS). This work concludes that R. rubrum is highly sensitive to Cu2+, with a strong inhibition of the photosynthetic reaction centres (RCs) already at 2 µM Cu2+. The inhibition of growth and of RC activity was related to the formation of Cu-containing BChl degradation products that occurred much more in the RC than in LH1. These results suggest that the shift of metal centres in BChl from Mg2+ to Cu2+ can occur in vivo in the RCs of R. rubrum under environmentally realistic Cu2+ concentrations, leading to a strong inhibition of the function of these RCs.