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Temperature is one of the most important physical factors affecting microbial and biochemical processes. We investigated the performance of photosynthetic apparatus of marine photoheterotrophic bacterium Dinoroseobacter shibae under various temperatures. The primary photochemistry and electron transport was measured using variable infra-red fluorometry in the cells grown between 8-35°C. It was found that the photosynthetic electron transport had a broad temperature optimum between 25-50°C. Moreover, the primary charge separation stayed functional even after rising temperature up to 55°C. The same phenomenon was observed also in other phototrophic Rhodobacterales. The psychrotolerant bacterium Roseisalinus antarcticus reached its maximum electron transport rate at 48°C, 30°C above its growth temperature. We propose that the extended temperature stability may be crucial to maintain photosynthetic function under situation when photosynthetic membranes heat up above their ambient temperature due to the heat dissipation of the excess light energy.

This study demonstrates the use of nanoparticles prepared by a gas aggregation source for fabricating structures by combining laser sintering and ablation. At first, the morphology and optical properties of prepared nanoparticle coatings were characterized. Then, the response of coatings to laser irradiation at different powers or exposure times was studied by in situ time-of-flight mass spectrometry, followed by scanning electron microscopy measurements of the resulting structures. By comparing the numbers of detected Ag ions, that were ablated and desorbed, with changes in morphology after irradiation, the optimum conditions for laser sintering and ablation of Ag nanoparticle coatings were found. As a proof of concept we fabricated micromirrors from sintered metal, microwires from both sintered metal and interconnected nanoparticles, and arbitrary metallic bulky or nanoparticle patterns. Vacuum compatibility and the possibility of fabrication of both metallic and nanoparticle structures in one step predetermines applications of developed method in electronics or sensing.

Background: Injectable lenacapavir administered every 6 months is a promising product for HIV pre-exposure prophylaxis (PrEP). We aimed to estimate the health and budget impacts and threshold price at which lenacapavir could be cost-effective in eastern and southern Africa., Methods: We adapted an agent-based network model, EMOD-HIV, to simulate lenacapavir scale-up in Zimbabwe, South Africa, and western Kenya from 2026 to 2035. Uptake assumptions were informed by a literature review of PrEP product preferences. In the main analysis, we varied lenacapavir coverage by subgroup: female sex workers (40% coverage); male clients of female sex workers (40%); adolescent girls and young women aged 15-24 years with more than one sexual partner (32%); women aged 25 years and older with more than one sexual partner (36%); and males with more than one sexual partner (32%). We also assessed a higher coverage scenario (64-76% across subgroups) and scenarios of expanding lenacapavir use, varying from concentrated among those at highest HIV risk to broader coverage including those at medium HIV risk. We estimated the maximum per-dose lenacapavir price that achieved cost-effectiveness (

The first phototrophic member of the bacterial phylum Gemmatimonadota , Gemmatimonas phototrophica AP64 T , received all its photosynthesis genes via distant horizontal gene transfer from a purple bacterium. Here, we investigated how these acquired genes, which are tightly controlled by oxygen and light in the ancestor, are integrated into the regulatory system of its new host. G. phototrophica grew well under aerobic and semiaerobic conditions, with almost no difference in gene expression. Under aerobic conditions, the growth of G. phototrophica was optimal at 80 µmol photon m -2 s -1 , while higher light intensities had an inhibitory effect. The transcriptome showed only a minimal response to the dark-light shift at optimal light intensity, while the exposure to a higher light intensity (200 µmol photon m -2 s -1 ) induced already stronger but still transient changes in gene expression. Interestingly, a singlet oxygen defense was not activated under any conditions tested. Our results indicate that G. phototrophica possesses neither the oxygen-dependent repression of photosynthesis genes known from purple bacteria nor the light-dependent repression described in aerobic anoxygenic phototrophs. Instead, G. phototrophica has evolved as a low-light species preferring reduced oxygen concentrations. Under these conditions, the bacterium can safely employ its photoheterotrophic metabolism without the need for complex regulatory mechanisms., Importance: Horizontal gene transfer is one of the main mechanisms by which bacteria acquire new genes. However, it represents only the first step as the transferred genes have also to be functionally and regulatory integrated into the recipient's cellular machinery. Gemmatimonas phototrophica , a member of bacterial phylum Gemmatimonadota, acquired its photosynthesis genes via distant horizontal gene transfer from a purple bacterium. Thus, it represents a unique natural experiment, in which the entire package of photosynthesis genes was transplanted into a distant host. We show that G. phototrophica lacks the regulation of photosynthesis gene expressions in response to oxygen concentration and light intensity that are common in purple bacteria. This restricts its growth to low-light habitats with reduced oxygen. Understanding the regulation of horizontally transferred genes is important not only for microbial evolution but also for synthetic biology and the engineering of novel organisms, as these rely on the successful integration of foreign genes., Competing Interests: The authors declare no conflict of interest.

In mid-2022, New York City (NYC) became the epicenter of the US mpox outbreak. We provided real-time mpox case forecasts to the NYC Department of Health and Mental Hygiene to aid in outbreak response. Forecasting methodologies evolved as the epidemic progressed. Initially, lacking knowledge of at-risk population size, we used exponential growth models to forecast cases. Once exponential growth slowed, we used a Susceptible-Exposed-Infectious-Recovered (SEIR) model. Retrospectively, we explored if forecasts could have been improved using an SEIR model in place of our early exponential growth model, with or without knowing the case detection rate. Early forecasts from exponential growth models performed poorly, as 2-week mean absolute error (MAE) grew from 53 cases/week (July 1-14) to 457 cases/week (July 15-28). However, when exponential growth slowed, providing insight into susceptible population size, an SEIR model was able to accurately predict the remainder of the outbreak (7-week MAE: 13.4 cases/week). Retrospectively, we found there was not enough known about the epidemiological characteristics of the outbreak to parameterize an SEIR model early on. However, if the at-risk population and case detection rate were known, an SEIR model could have improved accuracy over exponential growth models early in the outbreak., (© 2024 The Author(s). Journal of Medical Virology published by Wiley Periodicals LLC.)

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., Competing Interests: The authors declare no conflict of interest.

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., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (© 2023 Milali, Resar, Kaftan, Campbell, Olowu, Edwards, Platais, Kim, Jenkins and Bershteyn.)

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.

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)., (© 2022. The Author(s).)

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.

In almost all photosynthetic organisms the photosynthetic pigments chlorophyll and bacteriochlorophyll (BChl) are Mg 2+ containing complexes, but Mg 2+ 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 Zn 2+ instead of Mg 2+ 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 Cu 2+ 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 Mg 2+ of BChl can be replaced in vivo in the RCs by Cu 2+ under environmentally realistic Cu 2+ 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 Cu 2+ (trans-metallation) into BChl molecules of its RCs., (Copyright © 2021. Published by Elsevier B.V.)

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 11030 T , the type species of the genus, supports the classification of the two strains into the same genus. Strain AAP5 was found to contain C 18:1 ω 7 c (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 CO 2 -fixation pathways were found.

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 ± 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., (Copyright © 2020 The Authors. Published by Elsevier B.V. All rights reserved.)

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., (© FEMS 2019.)

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 E a1  = 16 kJ mol -1 , while that of the medium component was more than double, with E a2  = 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 P 870 + signal. At temperatures above 60 °C, the primary photochemistry became reversibly inhibited, which was manifested by an increase in minimal fluorescence, F 0 , whereas maximal fluorescence, F M , 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.

Magnesium (Mg 2+ ) 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 Cu 2+ , with a strong inhibition of the photosynthetic reaction centres (RCs) already at 2 μM Cu 2+ . 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 Mg 2+ to Cu 2+ can occur in vivo in the RCs of R. rubrum under environmentally realistic Cu 2+ concentrations, leading to a strong inhibition of the function of these RCs., (Copyright © 2019 Elsevier B.V. All rights reserved.)

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.

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 ν 1 (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 β‑carotene and varying amounts of antheraxanthin, violaxanthin and neoxanthin. Aplanozygotes contain mixtures of both primary and secondary carotenoids. In most cases, the ν 1 (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 ν 1 ) 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., (Copyright © 2019 Elsevier B.V. All rights reserved.)

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.

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., (© 2017 Society for Applied Microbiology and John Wiley & Sons Ltd.)

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.

Interquinone Q A -  → Q B electron-transfer (ET) in isolated photosystem II reaction centers (PSII-RC) is protein-gated. The temperature-dependent gating frequency "k" is described by the Eyring equation till levelling off at T ≥ 240 °K. Although central to photosynthesis, the gating mechanism has not been resolved and due to experimental limitations, could not be explored in vivo. Here we mimic the temperature dependency of "k" by enlarging V D1-208 , the volume of a single residue at the crossing point of the D1 and D2 PSII-RC subunits in Synechocystis 6803 whole cells. By controlling the interactions of the D1/D2 subunits, V D1-208 (or 1/T) determines the frequency of attaining an ET-active conformation. Decelerated ET, impaired photosynthesis, D1 repair rate and overall cell physiology upon increasing V D1-208 to above 130 Å 3 , rationalize the >99% conservation of small residues at D1-208 and its homologous motif in non-oxygenic bacteria. The experimental means and resolved mechanism are relevant for numerous transmembrane protein-gated reactions.

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.

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.

Here, we report an effect of short acclimation to a wide span of temperatures on photosynthetic electron transfer, lipid and fatty acid composition in the snow alga Chlamydomonas cf. nivalis. The growth and oxygen evolution capacity were low at 2 °C yet progressively enhanced at 10 °C and were significantly higher at temperatures from 5 to 15 °C in comparison with the mesophilic control Chlamydomonas reinhardtii. In search of the molecular mechanisms responsible for the adaptation of photosynthesis to low temperatures, we have found unprecedented high rates of QA to QB electron transfer. The thermodynamics of the process revealed the existence of an increased structural flexibility that we explain with the amino acid changes in the D1 protein combined with the physico-chemical characteristics of the thylakoid membrane composed of > 80% negatively charged phosphatidylglycerol., (© 2014 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.)

Light-harvesting complexes ensure necessary flow of excitation energy into photosynthetic reaction centers. In the present work, transient absorption measurements were performed on LH1-RC complexes isolated from two aerobic anoxygenic phototrophs (AAPs), Roseobacter sp. COL2P containing the carotenoid spheroidenone, and Erythrobacter sp. NAP1 which contains the carotenoids zeaxanthin and bacteriorubixanthinal. We show that the spectroscopic data from the LH1-RC complex of Roseobacter sp. COL2P are very similar to those previously reported for Rhodobacter sphaeroides, including the transient absorption spectrum originating from the intramolecular charge-transfer (ICT) state of spheroidenone. Although the ICT state is also populated in LH1-RC complexes of Erythrobacter sp. NAP1, its appearance is probably related to the polarity of the bacteriorubixanthinal environment rather than to the specific configuration of the carotenoid, which we hypothesize is responsible for populating the ICT state of spheroidenone in LH1-RC of Roseobacter sp. COL2P. The population of the ICT state enables efficient S1/ICT-to-bacteriochlorophyll (BChl) energy transfer which would otherwise be largely inhibited for spheroidenone and bacteriorubixanthinal due to their low energy S1 states. In addition, the triplet states of these carotenoids appear well-tuned for efficient quenching of singlet oxygen or BChl-a triplets, which is of vital importance for oxygen-dependent organisms such as AAPs.

The lectin-like domain of Tumor Necrosis Factor (TNF), mimicked by the TIP peptide, activates amiloride-sensitive sodium uptake in type II alveolar epithelial cells and as such increases alveolar liquid clearance in dysfunctional lungs. This protective effect is blunted upon mutation of residues T105, E107 and E110 in human TNF into alanine or upon pre-incubation of the cytokine with the disaccharide N,N'-diacetylchitobiose. In this study, we used molecular docking and molecular dynamics simulation to predict the binding sites for N,N'-diacetylchitobiose and trimannose-O-ethyl in the lectin-like domain of TNF and in the TIP peptide. Specific sites (K98, S99, P100, Q102 and E116) in the three loops of the lectin-like domain provide specific binding for both oligosaccharides, but none of the residues crucial for anti-edema activity are involved in hydrogen bonding with oligosaccharides or are subjected to steric hindrance by them. These results thus suggest that neither chitobiose nor trimannose affect crucial amino acids, while they occupy the cavity in the lectin-like domain. Consequently, both crucial amino acids and the emptiness of the cavity in the lectin-like domain may be critical for TNF's lectin-like activity. Analogously, the R4, E5, P7, Y16 amino acids of the TIP peptide are involved in forming hydrogen bonds with both oligosaccharides, whereas residues T6, E8 and E11 (corresponding to T105, E107 and E110 in hTNF) play an important role in stabilizing the peptide-oligosaccharide complex, supporting the hypothesis that amino acids in the polar region (TPEGAE) of the TIP peptide represent only a partial binding motif for sugars.

Raman microscopy permits structural analysis of protein crystals in situ in hanging drops, allowing for comparison with Raman measurements in solution. Nevertheless, the two methods sometimes reveal subtle differences in structure that are often ascribed to the water layer surrounding the protein. The novel method of drop-coating deposition Raman spectropscopy (DCDR) exploits an intermediate phase that, although nominally "dry," has been shown to preserve protein structural features present in solution. The potential of this new approach to bridge the structural gap between proteins in solution and in crystals is explored here with extrinsic protein PsbP of photosystem II from Spinacia oleracea. In the high-resolution (1.98 Å) x-ray crystal structure of PsbP reported here, several segments of the protein chain are present but unresolved. Analysis of the three kinds of Raman spectra of PsbP suggests that most of the subtle differences can indeed be attributed to the water envelope, which is shown here to have a similar Raman intensity in glassy and crystal states. Using molecular dynamics simulations cross-validated by Raman solution data, two unresolved segments of the PsbP crystal structure were modeled as loops, and the amino terminus was inferred to contain an additional beta segment. The complete PsbP structure was compared with that of the PsbP-like protein CyanoP, which plays a more peripheral role in photosystem II function. The comparison suggests possible interaction surfaces of PsbP with higher-plant photosystem II. This work provides the first complete structural picture of this key protein, and it represents the first systematic comparison of Raman data from solution, glassy, and crystalline states of a protein.

Photosynthetic biomass production rapidly declines in mesophilic cyanobacteria grown above their physiological temperatures largely due to the imbalance between degradation and repair of the D1 protein subunit of the heat susceptible Photosystem II reaction centers (PSIIRC). Here we show that simultaneous replacement of two conserved residues in the D1 protein of the mesophilic Synechocystis sp. PCC 6803, by the analogue residues present in the thermophilic Thermosynechococcus elongatus, enables photosynthetic growth, extensive biomass production and markedly enhanced stability and repair rate of PSIIRC for seven days even at 43 °C but only at elevated CO(2) (1%). Under the same conditions, the Synechocystis control strain initially presented very slow growth followed by a decline after 3 days. Change in the thylakoid membrane lipids, namely the saturation of the fatty acids is observed upon incubation for the different strains, but only the double mutant shows a concomitant major change of the enthalpy and entropy for the light activated Q(A)(-)→Q(B) electron transfer, rendering them similar to those of the thermophilic strain. Following these findings, computational chemistry and protein dynamics simulations we propose that the D1 double mutation increases the folding stability of the PSIIRC at elevated temperatures. This, together with the decreased impairment of D1 protein repair under increased CO(2) concentrations result in the observed photothermal tolerance of the photosynthetic machinery in the double mutant., (© 2011 Dinamarca et al.)

Higher harmonic contributions in the movement of an oscillating atomic force microscopy (AFM) cantilever are generated by nonlinear tip-sample interactions, yielding additional information on structure and physical properties such as sample stiffness. Higher harmonic amplitudes are strongly enhanced in liquid compared to the operation in air, and were previously reported to result in better structural resolution in highly organized lattices of proteins in bacterial S-layers and viral capsids [J. Preiner, J. Tang, V. Pastushenko, P. Hinterdorfer, Phys. Rev. Lett. 99 (2007) 046102]. We compared first and second harmonics AFM imaging of live and fixed human lung epithelial cells, and microvascular endothelial cells from mouse myocardium (MyEnd). Phase-distance cycles revealed that the second harmonic phase is 8 times more sensitive than the first harmonic phase with respect to variations in the distance between cantilever and sample surface. Frequency spectra were acquired at different positions on living and fixed cells with second harmonic amplitude values correlating with the sample stiffness. We conclude that variations in sample stiffness and corresponding changes in the cantilever-sample distance, latter effect caused by the finite feedback response, result in second harmonic images with improved contrast and information that is not attainable in the fundamental frequency of an oscillating cantilever.

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.

Progress in various fields of microscopy techniques brought up enormous possibilities to study the photosynthesis down to the level of individual pigment-protein complexes. The aim of this review is to present recent developments in the photosynthesis research obtained using such highly advanced techniques. Three areas of microscopy techniques covering optical microscopy, electron microscopy and scanning probe microscopy are reviewed. Whereas the electron microscopy and scanning probe microscopy are used in photosynthesis mainly for structural studies of photosynthetic pigment-protein complexes, the optical microscopy is used also for functional studies.

Several million macromolecules are exchanged each minute between the nucleus and cytoplasm by receptor-mediated transport. Most of this traffic is controlled by the small GTPase Ran, which regulates assembly and disassembly of the receptor-cargo complexes in the appropriate cellular compartment. Here we applied dynamic force spectroscopy to study the interaction of Ran with the nuclear import receptor importin beta1 (impbeta) at the single-molecule level. We found that the complex alternates between two distinct conformational states of different adhesion strength. The application of an external mechanical force shifts equilibrium toward one of these states by decreasing the height of the interstate activation energy barrier. The other state can be stabilized by a functional Ran mutant that increases this barrier. These results support a model whereby functional control of Ran-impbeta is achieved by a population shift between pre-existing alternative conformations.

Envelope-free chloroplasts were imaged in situ by contact and tapping mode scanning force microscopy at a lateral resolution of 3-5 nm and vertical resolution of approximately 0.3 nm. The images of the intact thylakoids revealed detailed structural features of their surface, including individual protein complexes over stroma, grana margin and grana-end membrane domains. Structural and immunogold-assisted assignment of two of these complexes, photosystem I (PS I) and ATP synthase, allowed direct determination of their surface density, which, for both, was found to be highest in grana margins. Surface rearrangements and pigment- protein complex redistribution associated with salt-induced membrane unstacking were followed on native, hydrated specimens. Unstacking was accompanied by a substantial increase in grana diameter and, eventually, led to their merging with the stroma lamellae. Concomitantly, PS IIalpha effective antenna size decreased by 21% and the mean size of membrane particles increased substantially, consistent with attachment of mobile light-harvesting complex II to PS I. The ability to image intact photosynthetic membranes at molecular resolution, as demonstrated here, opens up new vistas to investigate thylakoid structure and function.

Plants respond to excess light by a photoprotective reduction of the light harvesting efficiency. The notion that the non-photochemical quenching of chlorophyll fluorescence can be reliably used as an indicator of the photoprotection is put to a test here. The technique of the repetitive flash fluorescence induction is employed to measure in parallel the non-photochemical quenching of the maximum fluorescence and the functional cross-section (sigma(PS II)) which is a product of the photosystem II optical cross-section a(PS II) and of its photochemical yield Phi(PS II) (sigma (PS II) = a(PS II) Phi(PS II)). The quenching is measured for both, the maximum fluorescence found in a single-turnover flash (F(M) (ST)) and in a multiple turnover light pulse (F(M) (MT)). The experiment with the diatom Phaeodactylum tricornutum confirmed that, in line with the prevalent model, the PS II functional cross-section sigma (PS II) is reduced in high light and restored in the dark with kinetics and amplitude that are closely matching the changes of the F(M) (ST) and F(M) (MT) quenching. In contrast, a poor correlation between the light-induced changes in the PS II functional cross-section sigma (PS II) and the quenching of the multiple-turnover F(M) (MT) fluorescence was found in the green alga Scenedesmus quadricauda. The non-photochemical quenching in Scenedesmus quadricauda was further investigated using series of single-turnover flashes given with different frequencies. Several mechanisms that modulate the fluorescence emission in parallel to the Q(A) redox state and to the membrane energization were resolved and classified in relation to the light harvesting capacity of Photosystem II.

Fluorometers that measure the kinetics of chlorophyll fluorescence have become invaluable tools for determining the photosynthetic performance of plants. Many of these instruments use high frequency modulated light to measure the rate, efficiency and regulation of photosynthesis. The technique is non-invasive and is effective under diverse environmental conditions. Recently, imaging fluorometers have been introduced that reveal variability in photosynthesis over the surface of a leaf or between individual plants. Most imaging instruments depend on continuous light or low frequency modulated light for fluorescence excitation, which imposes serious limitations on measurements of the fluorescence parameters, especially the minimum fluorescence (F(0)) and variable fluorescence (F(V)). Here, we describe a new instrument that combines the advantage of high frequency modulated light with two-dimensional imaging of chlorophyll fluorescence. The fluorometer produces dynamic images of chlorophyll fluorescence from leaves or plants, providing accurate mapping of F(0) and F(V), and non-photochemical quenching. A significant feature of the instrument is that it can record fluorescence images of leaves in daylight under field conditions.

The photosynthetic activity of the green alga Scenedesmus quadricauda was investigated during synchronous growth in light/dark cycles. The rate of O2 evolution increased 2-fold during the first 3 to 4 h of the light period, remained high for the next 3 to 4 h, and then declined during the last half of the light period. During cell division, which occurred at the beginning of the dark period, the ability of the cells to evolve O2 was at a minimum. To determine if photosystem II (PSII) controls the photosynthetic capacity of the cells during the cell cycle we measured PSII activity and heterogeneity. Measurements of electron-transport activity revealed two populations of PSII, active centers that contribute to carbon reduction and inactive centers that do not. Measurements of PSII antenna sizes also revealed two populations, PSIIalpha and PSIIbeta, which differ from one another by their antenna size. During the early light period the photosynthetic capacity of the cells doubled, the O2-evolving capacity of PSII was nearly constant, the proportion of PSIIbeta centers decreased to nearly zero, and the proportion of inactive PSII centers remained constant. During the period of minimum photosynthetic activity 30% of the PSII centers were insensitive to the inhibitor 3-(3,4-dichlorophenyl)-1,1-dimethylurea, which may be related to reorganization of the thylakoid membrane. We conclude from these results that PSII does not limit the photosynthetic activity of the cells during the first half of the light period. However, the decline in photosynthetic activity observed during the last half of the light period can be accounted for by limited PSII activity.