Genomic and phenotypic characterization of Thermosynechococcus-like strains reveals eight species within the genus Thermosynechococcus and a novel genus Parathermosynechococcus gen. nov.

Thermophilic unicellular cyanobacteria of the family Thermosynechococcaceae are essential primary producers and integral components of many microbial mats found in hot springs of Asia and North America. Historically, based on their simple morphology, these organisms, along with members of taxonomically unrelated thermophilic Thermostichaceae have been described with a generic term, "Synechococcus", used for elongated unicellular cyanobacteria. This has created significant misperception in the scientific literature regarding the taxonomic status of these essential thermophilic primary producers and their relationship with Synechococcus sensu stricto. In this manuscript, we attempted a genome-driven taxonomic reevaluation of the family Thermosynechococcaceae. Application of genomic analyses such as GTDB classification, ANI/AAI and phylogenomics support the delineation of eight species within genus Thermosynechococcus. Two subspecies were further identified within T. taiwanensis by dDDH and phylogenomics. Moreover, the results also suggest the presence of two putative new genera phylogenetically alongside genus Thermosynechococcus, a thermophilic genus Parathermosynechococcus represented by PCC 6715 and a non-thermophilic genus represented by PCC 6312. The proposed genospecies and new genera were further integrated with morphological and/or ecological information. Interestingly, the phylogeny of 16S-23S ITS achieved a better taxonomic relationship than that of 16S rRNA and supported the genome-based classification of Thermosynechococcus spp. Finally, the pan-genome analysis indicated a conserved pattern of genomic core among known members of Thermosynechococcus.

Thermophilic cyanobacteria-exciting, yet challenging biotechnological chassis.

Thermophilic cyanobacteria are prokaryotic photoautotrophic microorganisms capable of growth between 45 and 73 °C. They are typically found in hot springs where they serve as essential primary producers. Several key features make these robust photosynthetic microbes biotechnologically relevant. These are highly stable proteins and their complexes, the ability to actively transport and concentrate inorganic carbon and other nutrients, to serve as gene donors, microbial cell factories, and sources of bioactive metabolites. A thorough investigation of the recent progress in thermophilic cyanobacteria reveals a significant increase in the number of newly isolated and delineated organisms and wide application of thermophilic light-harvesting components in biohybrid devices. Yet despite these achievements, there are still deficiencies at the high-end of the biotechnological learning curve, notably in genetic engineering and gene editing. Thermostable proteins could be more widely employed, and an extensive pool of newly available genetic data could be better utilised. In this manuscript, we attempt to showcase the most important recent advances in thermophilic cyanobacterial biotechnology and provide an overview of the future direction of the field and challenges that need to be overcome before thermophilic cyanobacterial biotechnology can bridge the gap with highly advanced biotechnology of their mesophilic counterparts. KEY POINTS: • Increased interest in all aspects of thermophilic cyanobacteria in recent years • Light harvesting components remain the most biotechnologically relevant • Lack of reliable molecular biology tools hinders further development of the chassis.

Complete genome sequence and transcriptome response to vitamin C supplementation of Novacetimonas hansenii SI1 - producer of highly-stretchable cellulose.

Novacetimonas hansenii SI1, previously known as Komagataeibacter hansenii, produces bacterial nanocellulose (BNC) with unique ability to stretch. The addition of vitamin C in the culture medium increases the porosity of the membranes and their stretchability making them highly moldable. To better understand the genetic background of this strain, we obtained its complete genome sequence using a hybrid sequencing and assembly strategy. We described the functional regions in the genome which are important for the synthesis of BNC and acetan-like II polymer. We next investigated the effect of 1% vitamin C supplementation on the global gene expression profile using RNA sequencing. Our transcriptomic readouts imply that vitamin C functions mainly as a reducing agent. We found that the changes in cellular redox status are balanced by strong repression of the sulfur assimilation pathway. Moreover, in the reduced conditions, glucose oxidation is decreased and alternative pathways for energy generation, such as acetate accumulation, are activated. The presence of vitamin C negatively influences acetan-like II polymer biosynthesis, which may explain the lowered yield and changed mechanical properties of BNC. The results of this study enrich the functional characteristics of the genomes of the efficient producers of the N. hansenii species. Improved understanding of the adaptation to the presence of vitamin C at the molecular level has important guiding significance for influencing the biosynthesis of BNC and its morphology.

Genome-scale identification and comparative analysis of transcription factors in thermophilic cyanobacteria.

BACKGROUND: The transcription factors (TFs) in thermophilic cyanobacteria might represent a uniquely evolved gene repertoire in light of the strong selective pressure caused by hostile habitats. Understanding the molecular composition of the TF genes in thermophilic cyanobacteria will facilitate further studies regarding verifying their exact biochemical functions and genetic engineering. However, limited information is available on the TFs of thermophilic cyanobacteria. Herein, a thorough investigation and comparative analysis were performed to gain insights into the molecular composition of the TFs in 22 thermophilic cyanobacteria. RESULTS: The results suggested a fascinating diversity of the TFs among these thermophiles. The abundance and type of TF genes were diversified in these genomes. The identified TFs are speculated to play various roles in biological regulations. Further comparative and evolutionary genomic analyses revealed that HGT may be associated with the genomic plasticity of TF genes in Thermostichus and Thermosynechococcus strains. Comparative analyses also indicated different pattern of TF composition between thermophiles and corresponding mesophilic reference cyanobacteria. Moreover, the identified unique TFs of thermophiles are putatively involved in various biological regulations, mainly as responses to ambient changes, may facilitating the thermophiles to survive in hot springs. CONCLUSION: The findings herein shed light on the TFs of thermophilic cyanobacteria and fundamental knowledge for further research regarding thermophilic cyanobacteria with a broad potential for transcription regulations in responses to environmental fluctuations.

Microalgae are not an umbrella solution for power industry waste abatement but could play a role in their valorization.

Microalgae have long been regarded as a promising solution for biological carbon abatement from the power industry, offering renewable biomass without competing for land or water resources used for food crops. In this study, we extensively examined the application of photosynthetic microorganisms for closing carbon, nitrogen, and micronutrient loops in the power industry. Subsequently, we explored the bottom-up integration of algal biorefineries into power industry waste streams for increased economic benefits and reduced environmental impacts. Analysis of the available data indicated that microalgae integration with the power industry is primarily performed using flue-gas-assisted cultivation. This approach allows for carbon sequestration typically below one gram per liter per day, too low to significantly impact carbon abatement at achievable scales of microalgae cultivation. Alternative approaches are also being explored. For example, soluble bicarbonate platforms allow for higher biomass productivity and temporary carbon storage. Meanwhile, the use of ashes and waste heat and thermophilic strains can result in lower cultivation costs and better control of cultivation conditions. These approaches offer further incremental improvement to microalgae-based carbon abatement systems in the power industry but are unlikely to be an umbrella solution for carbon reduction. Consequently, in the near term, microalgae-based carbon valorization systems are likely to be limited to niche applications involving the synthesis of high-value products. For microalgae to truly transform carbon abatement processes radical improvements in both biology and engineering approaches are urgently needed.

Polyphasic characterization of a novel hot-spring cyanobacterium Thermocoleostomius sinensis gen et sp. nov. and genomic insights into its carbon concentration mechanism.

Thermophilic cyanobacteria play a crucial role as primary producers in hot spring ecosystems, yet their microbiological, taxonomic, and ecological characteristics are not extensively studied. This study aimed to characterize a novel strain of thermophilic cyanobacteria, PKUAC-SCTA174 (A174), using a combination of traditional polyphasic methods and modern genomic-based approaches. The study included 16S rRNA-based phylogeny, ITS secondary structure prediction, morphological and habitat analyses, as well as high-quality genome sequencing with corresponding phylogenomic analyses. The results of the 16S rRNA, 16S-23S ITS secondary structure, morphology, and habitat analyses supported the classification of the strain as a member of a novel genus within the family Oculatellaceae, closely related to Albertania and Trichotorquatus. Genomic analysis revealed the presence of a sophisticated carbon-concentrating mechanism (CCM) in the strain, involving two CO2 uptake systems NDH-I3, and NDH-I4, three types of bicarbonate transporters (BCT1, bicA, sbtA,) and two distinct putative carboxysomal carbonic anhydrases (ccaA1 and ccaA2). The expression of CCM genes was investigated with a CO2 shift experiment, indicating varying transcript abundance among different carbon uptake systems. Based on the comprehensive characterization, the strain was delineated as Thermocoleostomius sinensis, based on the botanical code. The study of the complete genome of strain A174 contributes valuable insights into the genetic characteristics of the genus Thermocoleostomius and related organisms and provides a systematic understanding of thermophilic cyanobacteria. The findings presented here offer valuable data that can be utilized for future research in taxogenomics, ecogenomics, and geogenomics.

Characterization of a novel thermophilic cyanobacterium within Trichocoleusaceae, Trichothermofontia sichuanensis gen. et sp. nov., and its CO2-concentrating mechanism.

Thermophiles from extreme thermal environments have shown tremendous potential regarding ecological and biotechnological applications. Nevertheless, thermophilic cyanobacteria remain largely untapped and are rarely characterized. Herein, a polyphasic approach was used to characterize a thermophilic strain, PKUAC-SCTB231 (hereafter B231), isolated from a hot spring (pH 6.62, 55.5°C) in Zhonggu village, China. The analyses of 16S rRNA phylogeny, secondary structures of 16S-23S ITS and morphology strongly supported strain B231 as a novel genus within Trichocoleusaceae. Phylogenomic inference and three genome-based indices further verified the genus delineation. Based on the botanical code, the isolate is herein delineated as Trichothermofontia sichuanensis gen. et sp. nov., a genus closely related to a validly described genus Trichocoleus. In addition, our results suggest that Pinocchia currently classified to belong to the family Leptolyngbyaceae may require revision and assignment to the family Trichocoleusaceae. Furthermore, the complete genome of Trichothermofontia B231 facilitated the elucidation of the genetic basis regarding genes related to its carbon-concentrating mechanism (CCM). The strain belongs to ß-cyanobacteria according to its ß-carboxysome shell protein and 1B form of Ribulose bisphosphate Carboxylase-Oxygenase (RubisCO). Compared to other thermophilic strains, strain B231contains a relatively low diversity of bicarbonate transporters (only BicA for HCO3- transport) but a higher abundance of different types of carbonic anhydrase (CA), ß-CA (ccaA) and γ-CA (ccmM). The BCT1 transporter consistently possessed by freshwater cyanobacteria was absent in strain B231. Similar situation was occasionally observed in freshwater thermal Thermoleptolyngbya and Thermosynechococcus strains. Moreover, strain B231 shows a similar composition of carboxysome shell proteins (ccmK1-4, ccmL, -M, -N, -O, and -P) to mesophilic cyanobacteria, the diversity of which was higher than many thermophilic strains lacking at least one of the four ccmK genes. The genomic distribution of CCM-related genes suggests that the expression of some components is regulated as an operon and others in an independently controlled satellite locus. The current study also offers fundamental information for future taxogenomics, ecogenomics and geogenomic studies on distribution and significance of thermophilic cyanobacteria in the global ecosystem.

Characterization of Molecular Diversity and Organization of Phycobilisomes in Thermophilic Cyanobacteria.

Thermophilic cyanobacteria are cosmopolitan and abundant in the thermal environment. Their light-harvesting complexes, phycobilisomes (PBS), are highly important in photosynthesis. To date, there is limited information on the PBS composition of thermophilic cyanobacteria whose habitats are challenging for survival. Herein, genome-based methods were used to investigate the molecular components of PBS in 19 well-described thermophilic cyanobacteria. These cyanobacteria are from the genera Leptolyngbya, Leptothermofonsia, Ocullathermofonsia, Thermoleptolyngbya, Trichothermofonsia, Synechococcus, Thermostichus, and Thermosynechococcus. According to the phycobiliprotein (PBP) composition of the rods, two pigment types are observed in these thermophiles. The amino acid sequence analysis of different PBP subunits suggests several highly conserved cysteine residues in these thermophiles. Certain amino acid contents in the PBP of thermophiles are significantly higher than their mesophilic counterparts, highlighting the potential roles of specific substitutions of amino acid in the adaptive thermostability of light-harvesting complexes in thermophilic cyanobacteria. Genes encoding PBS linker polypeptides vary among the thermophiles. Intriguingly, motifs in linker apcE indicate a photoacclimation of a far-red light by Leptolyngbya JSC-1, Leptothermofonsia E412, and Ocullathermofonsia A174. The composition pattern of phycobilin lyases is consistent among the thermophiles, except for Thermostichus strains that have extra homologs of cpcE, cpcF, and cpcT. In addition, phylogenetic analyses of genes coding for PBPs, linkers, and lyases suggest extensive genetic diversity among these thermophiles, which is further discussed with the domain analyses. Moreover, comparative genomic analysis suggests different genomic distributions of PBS-related genes among the thermophiles, indicating probably various regulations of expression. In summary, the comparative analysis elucidates distinct molecular components and organization of PBS in thermophilic cyanobacteria. These results provide insights into the PBS components of thermophilic cyanobacteria and fundamental knowledge for future research regarding structures, functions, and photosynthetic improvement.

Distinct Molecular Patterns of Two-Component Signal Transduction Systems in Thermophilic Cyanobacteria as Revealed by Genomic Identification.

Two-component systems (TCSs) play crucial roles in sensing and responding to environmental signals, facilitating the acclimation of cyanobacteria to hostile niches. To date, there is limited information on the TCSs of thermophilic cyanobacteria. Here, genome-based approaches were used to gain insights into the structure and architecture of the TCS in 17 well-described thermophilic cyanobacteria, namely strains from the genus Leptodesmis, Leptolyngbya, Leptothermofonsia, Thermoleptolyngbya, Thermostichus, and Thermosynechococcus. The results revealed a fascinating complexity and diversity of the TCSs. A distinct composition of TCS genes existed among these thermophilic cyanobacteria. A majority of TCS genes were classified as orphan, followed by the paired and complex cluster. A high proportion of histidine kinases (HKs) were predicted to be cytosolic subcellular localizations. Further analyses suggested diversified domain architectures of HK and response regulators (RRs), putatively in association with various functions. Comparative and evolutionary genomic analyses indicated that the horizontal gene transfer, as well as duplications events, might be involved in the evolutionary history of TCS genes in Thermostichus and Thermosynechococcus strains. A comparative analysis between thermophilic and mesophilic cyanobacteria indicated that one HK cluster and one RR cluster were uniquely shared by all the thermophilic cyanobacteria studied, while two HK clusters and one RR cluster were common to all the filamentous thermophilic cyanobacteria. These results suggested that these thermophile-unique clusters may be related to thermal characters and morphology. Collectively, this study shed light on the TCSs of thermophilic cyanobacteria, which may confer the necessary regulatory flexibility; these findings highlight that the genomes of thermophilic cyanobacteria have a broad potential for acclimations to environmental fluctuations.

Construction of an artificial consortium of Escherichia coli and cyanobacteria for clean indirect production of volatile platform hydrocarbons from CO2.

Ethylene and isoprene are essential platform chemicals necessary to produce polymers and materials. However, their current production methods based on fossil fuels are not very efficient and result in significant environmental pollution. For a successful transition more sustainable economic model, producing these key polymeric building blocks from renewable and sustainable resources such as biomass or CO2 is essential. Here, inspired by the symbiotic relationship of natural microbial communities, artificial consortia composed of E. coli strains producing volatile platform chemicals: ethylene and isoprene and two strains of cyanobacteria phototrophically synthesizing and exporting sucrose to feed these heterotrophs were developed. Disaccharide produced by transgenic cyanobacteria was used as a carbon and electron shuttle between the two community components. The E. coli cscB gene responsible for sucrose transport was inserted into two cyanobacterial strains, Thermosynechococcus elongatus PKUAC-SCTE542 and Synechococcus elongatus PCC7942, resulting in a maximal sucrose yield of 0.14 and 0.07 g/L, respectively. These organisms were co-cultured with E. coli BL21 expressing ethylene-forming enzyme or isoprene synthase and successfully synthesized volatile hydrocarbons. Productivity parameters of these co-cultures were higher than respective transgenic cultures of E. coli grown individually at similar sucrose concentrations, highlighting the positive impact of the artificial consortia on the production of these platform chemicals.

Generation of miniploid cells and improved natural transformation procedure for a model cyanobacterium Synechococcus elongatus PCC 7942.

The biotechnologically important and naturally transformable cyanobacterium, Synechococcus elongatus PCC 7942, possesses multiple genome copies irrespective of its growth rate or condition. Hence, segregating mutations across all genome copies typically takes several weeks. In this study, Synechococcus 7942 cultivation on a solid growth medium was optimised using different concentrations of agar, the addition of antioxidants, and overexpression of the catalase gene to facilitate the rapid acquisition of colonies and fully segregated lines. Synechococcus 7942 was grown at different temperatures and nutritional conditions. The miniploid cells were identified using flow cytometry and fluorimetry. The natural transformation was carried out using miniploid cells and validated with PCR and high performance liquid chromatography (HPLC). We identified that 0.35% agar concentration and 200 IU of catalase could improve the growth of Synechococcus 7942 on a solid growth medium. Furthermore, overexpression of a catalase gene enhanced the growth rate and supported diluted culture to grow on a solid medium. Our results reveal that high temperature and phosphate-depleted cells contain the lowest genome copies (2.4 ± 0.3 and 1.9 ± 0.2) and showed the potential to rapidly produce fully segregated mutants. In addition, higher antibiotic concentrations improve the selection of homozygous transformants while maintaining similar genome copies at a constant temperature. Based on our observation, we have an improved cultivation and natural transformation protocol for Synechococcus 7942 by optimising solid media culturing, generating low-ploidy cells that ultimately reduced the time required for the complete segregation of engineered lines.

Discovery of Five New Ethylene-Forming Enzymes for Clean Production of Ethylene in E. coli.

Ethylene is an essential platform chemical with a conjugated double bond, which can produce many secondary chemical products through copolymerisation. At present, ethylene production is mainly from petroleum fractionation and cracking, which are unsustainable in the long term, and harmful to our environment. Therefore, a hot research field is seeking a cleaner method for ethylene production. Based on the model ethylene-forming enzyme (Efe) AAD16440.1 (6vp4.1.A) from Pseudomonas syringae pv. phaseolicol, we evaluated five putative Efe protein sequences using the data derived from phylogenetic analyses and the conservation of their catalytic structures. Then, pBAD expression frameworks were constructed, and relevant enzymes were expressed in E. coli BL21. Finally, enzymatic activity in vitro and in vivo was detected to demonstrate their catalytic activity. Our results show that the activity in vitro measured by the conversion of α-ketoglutarate was from 0.21-0.72 µmol ethylene/mg/min, which varied across the temperatures. In cells, the activity of the new Efes was 12.28-147.43 µmol/gDCW/h (DCW, dry cellular weight). Both results prove that all the five putative Efes could produce ethylene.

Comparative Genomic Analysis Revealed Distinct Molecular Components and Organization of CO2-Concentrating Mechanism in Thermophilic Cyanobacteria.

Cyanobacteria evolved an inorganic carbon-concentrating mechanism (CCM) to perform effective oxygenic photosynthesis and prevent photorespiratory carbon losses. This process facilitates the acclimation of cyanobacteria to various habitats, particularly in CO2-limited environments. To date, there is limited information on the CCM of thermophilic cyanobacteria whose habitats limit the solubility of inorganic carbon. Here, genome-based approaches were used to identify the molecular components of CCM in 17 well-described thermophilic cyanobacteria. These cyanobacteria were from the genus Leptodesmis, Leptolyngbya, Leptothermofonsia, Thermoleptolyngbya, Thermostichus, and Thermosynechococcus. All the strains belong to ß-cyanobacteria based on their ß-carboxysome shell proteins with 1B form of Rubisco. The diversity in the Ci uptake systems and carboxysome composition of these thermophiles were analyzed based on their genomic information. For Ci uptake systems, two CO2 uptake systems (NDH-13 and NDH-14) and BicA for HCO3 - transport were present in all the thermophilic cyanobacteria, while most strains did not have the Na+/HCO3 - Sbt symporter and HCO3 - transporter BCT1 were absent in four strains. As for carboxysome, the ß-carboxysomal shell protein, ccmK2, was absent only in Thermoleptolyngbya strains, whereas ccmK3/K4 were absent in all Thermostichus and Thermosynechococcus strains. Besides, all Thermostichus and Thermosynechococcus strains lacked carboxysomal ß-CA, ccaA, the carbonic anhydrase activity of which may be replaced by ccmM proteins as indicated by comparative domain analysis. The genomic distribution of CCM-related genes was different among the thermophiles, suggesting probably distinct expression regulation. Overall, the comparative genomic analysis revealed distinct molecular components and organization of CCM in thermophilic cyanobacteria. These findings provided insights into the CCM components of thermophilic cyanobacteria and fundamental knowledge for further research regarding photosynthetic improvement and biomass yield of thermophilic cyanobacteria with biotechnological potentials.

Reevaluation of Parasynechococcus-like Strains and Genomic Analysis of Their Microsatellites and Compound Microsatellites.

Morphologically similar to Synechococcus, a large number of Parasynechococcus strains were misclassified, resulting in extreme underestimation of their genetic diversity. In this study, 80 Synechococcus-like strains were reevaluated using a combination of 16S rRNA phylogeny and genomic approach, identifying 54 strains as Parasynechococcus-like strains and showing considerably intragenus genetic divergence among the subclades identified. Further, bioinformatics analysis disclosed diversified patterns of distribution, abundance, density, and diversity of microsatellites (SSRs) and compound microsatellites (CSSRs) in genomes of these Parasynechococcus-like strains. Variations of SSRs and CSSRs were observed amongst phylotypes and subclades. Both SSRs and CSSRs were in particular unequally distributed among genomes. Dinucleotide SSRs were the most widespread, while the genomes showed two patterns in the second most abundant repeat type (mononucleotide or trinucleotide SSRs). Both SSRs and CSSRs were predominantly observed in coding regions. These two types of microsatellites showed positive correlation with genome size (p < 0.01) but negative correlation with GC content (p < 0.05). Additionally, the motif (A)n, (AG)n and (AGC)n was a major one in the corresponding category. Meanwhile, distinctive motifs of CSSRs were found in 39 genomes. This study characterizes SSRs and CSSRs in genomes of Parasynechococcus-like strains and will be useful as a prerequisite for future studies regarding their distribution, function, and evolution. Moreover, the identified SSRs may facilitate fast acclimation of Parasynechococcus-like strains to fluctuating environments and contribute to the extensive distribution of Parasynechococcus species in global marine environments.

Polyphasic Identification and Genomic Insights of Leptothermofonsia sichuanensis gen. sp. nov., a Novel Thermophilic Cyanobacteria Within Leptolyngbyaceae.

Thermal environments are an important reservoir of thermophiles with significant ecological and biotechnological potentials. However, thermophilic isolates remain largely unrecovered from their habitats and are rarely systematically identified. In this study, we characterized using polyphasic approaches a thermophilic strain, PKUAC-SCTAE412 (E412 hereafter), recovered from Lotus Lake hot spring based in Ganzi prefecture, China. The results of 16S rRNA/16S-23S ITS phylogenies, secondary structure, and morphology comparison strongly supported that strain E412 represent a novel genus within Leptolyngbyaceae. This delineation was further confirmed by genome-based analyses [phylogenomic inference, average nucleotide/amino-acid identity, and the percentages of conserved proteins (POCP)]. Based on the botanical code, the isolate is herein delineated as Leptothermofonsia sichuanensis gen. sp. nov, a genus adjacent to recently delineated Kovacikia and Stenomitos. In addition, we successfully obtained the first complete genome of this new genus. Genomic analysis revealed its adaptations to the adverse hot spring environment and extensive molecular components related to mobile genetic elements, photosynthesis, and nitrogen metabolism. Moreover, the strain was capable of modifying the composition of its light-harvesting apparatus depending on the wavelength and photoperiod, showing chromatic adaptation capacity characteristic for T1 and T2 pigmentation types. Other physiological studies showed the strain's ability to utilize sodium bicarbonate and various sulfur compounds. The strain was also shown to be diazotrophic. Interestingly, 24.6% of annotated protein-coding genes in the E412 genome were identified as putatively acquired, hypothesizing that a large number of genes acquired through HGT might contribute to the genome expansion and habitat adaptation of those thermophilic strains. Most the HGT candidates (69.4%) were categorized as metabolic functions as suggested by the KEGG analysis. Overall, the complete genome of strain E412 provides the first insight into the genomic feature of the genus Leptothermofonsia and lays the foundation for future global ecogenomic and geogenomic studies.

Enhanced removal of tetracycline from synthetic wastewater using an optimal ratio of co-culture of Desmodesmus sp. and Klebsiella pneumoniae.

A sustainable approach of Desmodesmus sp. GIEC-179: Klebsiella pneumoniae (DUT-XJR-t-1.2) co-culture ratios were optimized to remove tetracycline (TET) from synthetic wastewater. To enhance the tetracycline removal performance, the effect of microalgae-bacterial co-culture ratio, maximum TET concentration, effective inoculum amount, growth temperature and pH were studied. The optimized ratio 1:2 of Desmodesmus sp.: K. pneumoniae showed the optimal removal percentage at the temperature of 25 °C, pH 7 and 10% inoculum amount; and the removal of TET was recorded as 95%. Moreover, this study explored the Desmodesmus sp.: K. pneumoniae (1:2) nutrient (COD, NH4+ and PO43-) exchange relationship and their interaction of TET removal to better understand their fundamental mechanism. According to the results of this study, Desmodesmus sp.: K. pneumoniae co-culture could be a green option for bio-removal of tetracycline from wastewater.

Comparative Genomics and Physiological Investigation of a New Arthrospira/Limnospira Strain O9.13F Isolated from an Alkaline, Winter Freezing, Siberian Lake.

Cyanobacteria from the genus Arthrospira/Limnospira are considered haloalkalotolerant organisms with optimal growth temperatures around 35 °C. They are most abundant in soda lakes in tropical and subtropical regions. Here, we report the comprehensive genome-based characterisation and physiological investigation of the new strain O9.13F that was isolated in a temperate climate zone from the winter freezing Solenoye Lake in Western Siberia. Based on genomic analyses, the Siberian strain belongs to the Arthrospira/Limnospira genus. The described strain O9.13F showed the highest relative growth index upon cultivation at 20 °C, lower than the temperature 35 °C reported as optimal for the Arthrospira/Limnospira strains. We assessed the composition of fatty acids, proteins and photosynthetic pigments in the biomass of strain O9.13F grown at different temperatures, showing its potential suitability for cultivation in a temperate climate zone. We observed a decrease of gamma-linolenic acid favouring palmitic acid in the case of strain O9.13F compared to tropical strains. Comparative genomics showed no unique genes had been found for the Siberian strain related to its tolerance to low temperatures. In addition, this strain does not possess a different set of genes associated with the salinity stress response from those typically found in tropical strains. We confirmed the absence of plasmids and functional prophage sequences. The genome consists of a 4.94 Mbp with a GC% of 44.47% and 5355 encoded proteins. The Arthrospira/Limnospira strain O9.13F presented in this work is the first representative of a new clade III based on the 16S rRNA gene, for which a genomic sequence is available in public databases (PKGD00000000).

Genome-Wide Investigation and Analysis of Microsatellites and Compound Microsatellites in Leptolyngbya-like Species, Cyanobacteria.

Microsatellites (simple sequence repeats, SSRs) are ubiquitously distributed in almost all known genomes. Here, the first investigation was designed to examine the SSRs and compound microsatellites (CSSRs) in genomes of Leptolyngbya-like strains. The results disclosed diversified patterns of distribution, abundance, density, and diversity of SSRs and CSSRs in genomes, indicating that they may be subject to rapid evolutionary change. The numbers of SSRs and CSSRs were extremely unevenly distributed among genomes, ranging from 11,086 to 24,000 and from 580 to 1865, respectively. Dinucleotide SSRs were the most abundant category in 31 genomes, while the other 15 genomes followed the pattern: mono- > di- > trinucleotide SSRs. The patterns related to SSRs and CSSRs showed differences among phylogenetic groups. Both SSRs and CSSRs were overwhelmingly distributed in coding regions. The numbers of SSRs and CSSRs were significantly positively correlated with genome size (p < 0.01) and negatively correlated with GC content (p < 0.05). Moreover, the motif (A/C)n and (AG)n was predominant in mononucleotide and dinucleotide SSRs, and unique motifs of CSSRs were identified in 39 genomes. This study provides the first insight into SSRs and CSSRs in genomes of Leptolyngbya-like strains and will be useful to understanding their distribution, predicting their function, and tracking their evolution. Additionally, the identified SSRs may provide an evolutionary advantage of fast adaptation to environmental changes and may play an important role in the cosmopolitan distribution of Leptolyngbya strains to globally diverse niches.

Effects of Low Temperature, Nitrogen Starvation and Their Combination on the Photosynthesis and Metabolites of Thermosynechococcus E542: A Comparison Study.

Both low temperature and nitrogen starvation caused chlorosis of cyanobacteria. Here, in this study, for the first time, we compared the effects of low temperature, nitrogen starvation, and their combination on the photosynthesis and metabolites of a thermophilic cyanobacterium strain, Thermosynechococcus E542. Under various culture conditions, the growth rates, pigment contents, and chlorophyll fluorescence were monitored, and the composition of alkanes, lipidomes, and carbohydrates were determined. It was found that low temperature (35 °C) significantly suppressed the growth of Thermosynechococcus E542. Nitrogen starvation at 45 °C and 55 °C did not affect the growth; however, combined treatment of low temperature and nitrogen starvation led to the lowest growth rate and biomass productivity. Both low temperature and nitrogen starvation caused significantly declined contents of pigments, but they resulted in a different effect on the OJIP curves, and their combination led to the lowest pigment contents. The composition of fatty acids and alkanes was altered upon low-temperature cultivation, while nitrogen starvation caused reduced contents of all lipids. The low temperature did not affect carbohydrate contents, while nitrogen starvation greatly enhanced carbohydrate content, and their combination did not enhance carbohydrate content, but led to reduced productivity. These results revealed the influence of low temperature, nitrogen starvation, and their combined treatment for the accumulation of phycobiliproteins, lipids, and carbohydrates of a thermophilic cyanobacterium strain, Thermosynechococcus E542.