Comprehensive understanding of the mutant 'giant' Arthrospira platensis developed via ultraviolet mutagenesis.

Introduction: Cyanobacteria are typically of a size that can be observed under a microscope. Here, we present cyanobacteria of a size that can be observed with the naked eye. Arthrospira platensis NCB002 strain showed differentiated morphological characteristics compared to previously reported Arthrospira spp. Methods: Arthrospira platensis NCB002 was obtained by the UV irradiation of Arthrospira sp. NCB001, which was isolated from freshwater and owned by NCell Co., Ltd. A. platensis NIES-39 was obtained from the National Institute for Environmental Studies (Tsukuba, Japan). We used various analytical techniques to determine its overall characteristics. Results and discussion: The draft genome of strain NCB002 consists of five contigs comprising 6,864,973 bp with a G+C content of 44.3 mol%. The strain NCB002 had an average length of 11.69 ± 1.35 mm and a maximum of 15.15 mm, which is 23.4-50.5 times longer than the length (0.3-0.5 mm) of previously known Arthrospira spp., allowing it to be harvested using a thin sieve. Transcriptome analysis revealed that these morphological differences resulted from changes in cell wall formation mechanisms and increased cell division. Our results show that NCB002 has outstanding industrial value and provides a comprehensive understanding of it.

Green and efficient method to acquire high-value phycobiliprotein from microalgal biomass involving deep eutectic solvent-based ultrasound-assisted extraction.

Phycoerythrin (PE) is a phycobiliprotein holding great potential as a high-value food colorant and medicine. Deep eutectic solvent (DES)-based ultrasound-assisted extraction (UAE) was applied to extract B-PE by disrupting the resistant polysaccharide cell wall of Porphyridium purpureum. The solubility of cell wall monomers in 31 DESs was predicted using COSMO-RS. Five glycerol-based DESs were tested for extraction, all of which showed significantly higher B-PE yields by up to 13.5 folds than water. The DES-dependent B-PE extraction efficiencies were proposedly associated with different cell disrupting capabilities and protein stabilizing effects of DESs. The DES-based UAE method could be considered green according to a metric assessment tool, AGREEprep. The crude extract containing DES was further subjected to aqueous two-phase system, two-step ammonium sulfate precipitation, and ultrafiltration processes. The final purified B-PE had a PE purity ratio of 3.60 and a PC purity ratio of 0.08, comparable to the purity of commercial products.

Endophyte Bacillus tequilensis improves the growth of microalgae Haematococcus lacustris by regulating host cell metabolism.

This study identified an endosymbiotic bacterium, Bacillus tequilensis, residing within the cells of the microalga Haematococcus lacustris through 16S rRNA analysis. To confirm the optimal interactive conditions between H. lacustris and B. tequilensis, the effects of different ratios of cells using H. lacustris of different growth stages were examined. Under optimized conditions, the cell density, dry weight, chlorophyll content, and astaxanthin content of H. lacustris increased significantly, and the fatty acid content improved 1.99-fold. Microscopy demonstrated the presence of bacteria within the H. lacustris cells. The interaction upregulated amino acid and nucleotide metabolism in H. lacustris. Interestingly, muramic and phenylacetic acids were found exclusively in H. lacustris cells in the presence of B. tequilensis. Furthermore, B. tequilensis delayed pigment degradation in H. lacustris. This study reveals the impact of the endosymbiont B. tequilensis on the metabolism of H. lacustris and offers new perspectives on the symbiotic relationship between them.

Establishment of ultrasonic stimulation to enhance growth of Haematococcus lacustris.

In this study, ultrasonication at a frequency of 40 kHz was used to shorten the sonication period and enhance the growth of Haematococcus lacustris. To confirm the optimal conditions, the effects of ultrasound output and treatment interval were examined. Under optimal conditions (20 W and 15-day cycle), the maximum cell density and chlorophyll content were 66.75 × 104 cells mL-1 and 36.54 mg g-1, respectively, which were increased by 50.00% and 39.01%, respectively, compared to the control. Transmission electron microscopy analysis showed that ultrasonication caused tiny cracks in the W4 and W6 strata but did not disrupt the inner W2 layer. Additionally, RT-qPCR analysis showed that ultrasonication upregulated both cell division and nitrogen uptake. No difference were detected in the composition or quantity of fatty acids. This study demonstrates a novel ultrasonic approach for enhancing the growth of H. lacustris.

Microalgal secondary metabolite productions as a component of biorefinery: A review.

The interest in developing microalgae for industrial use has been increasing because of concerns about the depletion of petroleum resources and securing sustainable energy sources. Microalgae have high biomass productivity and short culture periods. However, despite these advantages, various barriers need to be overcome for industrial applications. Microalgal cultivation has a high unit price, thus rendering industrial application difficult. It is indispensably necessary to co-produce their primary and secondary metabolites to compensate for these shortcomings. In this regard, this article reviews the following aspects, (1) co-production of primary and secondary metabolites in microalgae, (2) induction methods for the promotion of the biosynthesis of secondary metabolites, and (3) perspectives on the co-production and co-extraction of primary and secondary metabolites. This paper presents various approaches for producing useful metabolites from microalgae and suggests strategies that can be utilized for the co-production of primary and secondary metabolites.

Complete chloroplast genome of Gypsophila oldhamiana Miq. (Caryophyllales: Caryophyllaceae).

The complete chloroplast genome sequence of Gypsophila oldhamiana Miq., a species of the Caryophyllaceae family, was assembled and analyzed from the de novo assembly of Illumina paired-end sequencing data. The total length of the chloroplast genome of G. oldhamiana was 152,675 bp including a large single-copy (LSC) region of 83,552 bp, a small single-copy (SSC) region of 17,349 bp, and a pair of identical inverted repeat regions (IRs) of 25,887 bp. The genome possessed a total of 130 genes, including 85 protein-coding genes, 37 transfer RNA (tRNA) genes, and 8 ribosomal RNA (rRNA) genes. The phylogenetic analysis of G. oldhamiana with 14 related species discovered the closest taxonomical relationship with Gypsophila vaccaria voucher in the Caryophyllaceae family.

Semi-continuous immobilized cultivation of Porphyridium cruentum for sulfated polysaccharides production.

In this study, semi-continuous immobilized cultivation of Porphyridium cruentum through calcium alginate beads was performed for sulfated polysaccharides (SPs) production. The cell biomass and daily SPs productivity in the calcium alginate bead immobilized culture were increased by up to 79 ± 3.4% and 45.6 ± 3.2%, compared to those in the control, respectively. Furthermore, simultaneous application of immobilization and blue wavelength illumination further increased the phycobiliproteins content by 260 ± 9%, compared to those in the control. Similarly, nutrient deficiencies in combination with immobilization increased daily SPs productivity by about twice that of the control. The chemical composition and biological activity of the extracellular polymeric substances produced through immobilization were similar to those of the control. This study suggests the potential application of calcium alginate beads-based immobilization for continuous and high-efficiency SPs production using P. cruentum.

Association between Novel Marker (Platelet-Lymphocyte Ratio, Neutrophil-Lymphocyte Ratio, and Delta Neutrophil Index) and Outcomes in Sudden Cardiac Arrest Patients.

PURPOSE: It is important that clinicians accurately predict the outcome of patients with sudden cardiac arrest (SCA). The complete blood count (CBC) is an easy and inexpensive test that provides information on blood content. Platelet-lymphocyte ratio (PLR), neutrophil-lymphocyte ratio (NLR), and delta neutrophil index (DNI) are relatively novel biomarkers that have been used in the prognosis of various diseases. We aimed to determine the usefulness of PLR, NLR, and DNI in predicting the outcomes of SCA. MATERIALS AND METHODS: This retrospective observational study was performed on patients with SCA. Patients who visited the tertiary university hospital from January 2015 to December 2019 were targeted. The inclusion criteria were all nontraumatic adult out-hospital cardiac arrest patients. We analyzed DNI, PLR, and NLR based on the CBC results of all enrolled patients. The exclusion criteria were as follows: no data on laboratory study, traumatic arrest, age < 18 years, and a history of leukemia, myelodysplastic syndrome, and myelofibrosis. The primary outcome was assessed as return of spontaneous circulation (ROSC), the secondary outcome as survival to discharge, and the tertiary outcome as neurological outcome. RESULTS: From January 1, 2015, to December 31, 2019, 739 patients were enrolled. ROSC was seen in 324 patients, of whom 60 had survival to discharge and 24 had good neurological outcome at the time of discharge (cerebral performance categories (CPCs) 1-2). The PLR of the ROSC group was 42.41 (range: 4.21-508.7), which was higher than that of the No-ROSC group (p=0.006). The DNI value of the survival group was 0.00 (range: 0.00-40.9), which was lower than that of the nonsurvival group. CONCLUSIONS: Patients with SCA and subsequent ROSC had higher PLR and NLR, while those with survival to discharge had lower DNI values than those with nonsurvival to discharge (p=0.005).

Enhancement of phycoerythrin productivity in Porphyridium purpureum using the clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9 ribonucleoprotein system.

Genetic manipulation of the Porphyridium sp. may increase the production of phycoerythrin. Since phycobiliproteins capture and transfer energy to both photosystems (PS I and PS II), it was hypothesized that the gene mutation involved increases phycoerythrin synthesis. The gene encoding chlorophyll synthase (CHS1) was selected as chlorophyll synthase plays an important role in photosynthesis, mediating the final process of chlorophyll synthesis. Clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9 ribonucleoprotein (CRISPR/Cas9 RNP) delivery system was used to generate the chlorophyll synthase loss-of-function mutants (Δchs1). Independent Δchs1 showed no differences in the growth and production of sulfated polysaccharide compared to control. Phycoerythrin contents of the two independent mutants substantially increased regardless of light source. This study provides a novel applicability for the CRISPR/Cas9 RNP method in red microalgae toward a bio-product of interest. The obtained mutants could serve as potential producers of phycoerythrin if Porphyridium is selected as a natural source.

Astaxanthin biosynthesis promotion with pH shock in the green microalga, Haematococcus lacustris.

In this study, the use of pH shock to improve astaxanthin synthesis in Haematococcus lacustris was investigated. It has been found that pH shock (pH = 4.5, 60 s) imposes stress in the cells and induces physiological changes, which result in astaxanthin accumulation. The optimal acid-base combination of pH shock was H2SO4-KOH, which increased the astaxanthin content per cell to 39 ± 6.92% than those of the control. In addition, pH shock can be applied simultaneously with the other inductive strategies such as high irradiance and carbon source supply. When high irradiance was applied simultaneously with pH shock, astaxanthin yield was increased 65 ± 0.541% than control. In addition, astaxanthin content per cell was increased 105 ± 6.66% than those of the control, with the concomitant application of carbon source addition with pH shock. Herein, these novel findings provide a useful technique for producing astaxanthin using H. lacustris.

Effect of Pseudoalteromonas sp. MEBiC 03485 on biomass production and sulfated polysaccharide biosynthesis in Porphyridium cruentum UTEX 161.

The effect of co-cultivation of Porphyridium cruentum UTEX 161 with Pseudoalteromonas sp. MEBiC 03485 on P. cruentum growth and its sulfated polysaccharide (EPS) production were examined. The strain MEBiC 03485 had beneficial effects on P. cruentum growth, EPS production, and EPS quality. These effects were due to a compound secreted by the strain MEBiC 03485. Notably, secretory compound treatment also increased intracellular phycoerythrin and phycocyanin content by 89.4% and 161%, respectively. In addition, the biological activities of EPS extracted from MEBiC 03485 treatment tended to be higher than the control without treatment. Our results suggest a novel approach for potentially enhancing the growth of P. cruentum and its EPS production and quality by co-culturing with the symbiotic strain MEBiC 03485.

Control of a toxic cyanobacterial bloom species, Microcystis aeruginosa, using the peptide HPA3NT3-A2.

Microcystis aeruginosa, a species of freshwater cyanobacteria, is known to be one of the dominant species causing cyanobacterial harmful algal blooms (CyanoHABs). M. aeruginosa blooms have the potential to produce neurotoxins and peptide hepatotoxins, such as microcystins and lipopolysaccharides (LPSs). Currently, technologies for CyanoHAB control do not provide any ultimate solution because of the secondary pollution associated with the control measures. In this study, we attempted to use the peptide HPA3NT3-A2, which has been reported to be nontoxic and has antimicrobial properties, for the development of an eco-friendly control against CyanoHABs. HPA3NT3-A2 displayed significant algicidal effects against M. aeruginosa cells. HPA3NT3-A2 induced cell aggregation and flotation (thereby facilitating harvest), inhibited cell growth through sedimentation, and eventually destroyed the cells. HPA3NT3-A2 had no algicidal effect on other microalgal species such as Haematococcus pluvialis and Chlorella vulgaris. Additionally, HPA3NT3-A2 was not toxic to Daphnia magna. The algicidal mechanism of HPA3NT3-A2 was intracellular penetration. The results of this study suggest the novel possibility of controlling CyanoHABs using HPA3NT3-A2.

Enhancement of growth and paramylon production of Euglena gracilis by co-cultivation with Pseudoalteromonas sp. MEBiC 03485.

This study investigated the putative effects of co-cultivation of Euglena gracilis with Pseudoalteromonas sp. MEBiC 03485 on the growth of E. gracilis and its paramylon production. The strain MEBiC 03485 had beneficial effects on the growth and paramylon contents of E. gracilis. To determine the optimal conditions for co-cultivation, the effects of algal to bacterial inoculum ratios and E. gracilis growth stages were examined. Under optimal conditions, the biomass productivity and paramylon production were increased by more than 23% and 34%, respectively. These effects were attributed to the extracellular polymeric substances (EPS) from the strain MEBiC 03485. GC-MS and HPAEC were carried out to analyze the composition of EPS. It was found that the EPS consisted of rhamnose, galactose, glucose, and mannose. These results suggest a novel approach for potentially enhancing the growth of E. gracilis as well as its paramylon production, via co-culturing with the symbiotic strain MEBiC 03485.

Blue-Red LED wavelength shifting strategy for enhancing beta-carotene production from halotolerant microalga, Dunaliella salina.

In the present study, to improve the photosynthetic betacarotene productivity of Dunaliella salina, a blue-red LED wavelength-shifting system (B-R system) was investigated. Dunaliella salina under the B-R system showed enhanced density and beta-carotene productivity compared to D. salina cultivated under single light-emitting diode light wavelengths (blue, white, and red light-emitting diode). Additionally, we developed blue light-adapted D. salina (ALE-D. salina) using an adaptive laboratory evolution (ALE) approach. In combination with the B-R system applied to ALE-D. salina (ALE B-R system), the beta-carotene concentration (33.94 ± 0.52 µM) was enhanced by 19.7% compared to that observed for the non-ALE-treated wild-type of D. salina (intact D. salina) under the B-R system (28.34 ± 0.24 µM).

Chemical and genomic diversity of six Lonicera species occurring in Korea.

Lonicera spp. (Caprifoliaceae) are important not only as a common medicinal herb in East Asia but also as one of the most problematic invasive species in North America. In the present study, we performed a systemic analysis of genomic and chemical diversity among six Lonicera species occurring in Korea, L. japonica, L. maackii, L. insularis, L. sachalinensis, L. praeflorens, and L. vesicaria, using chloroplast DNA whole genome shotgun (WGS) sequencing and LC-MS analyses. The phylogenetic and phylochemical relationships did not coincide with each other, but partial consistency could be found among them. InDel-based cDNA marker for authentication was developed based on the genome sequences. Flavonoids, iridoids, and organic acids were identified in the LC-MS analyses, and their inter-species distribution and localization were also revealed.

The complete chloroplast genome sequence of Korean Lonicera japonica and intra-species diversity.

Lonicera japonica is a traditional medicinal plant well known for its anti-inflammatory effect. The complete chloroplast genome sequence of L. japonica collected from Korea was obtained by de novo assembly using whole genome sequence data. The chloroplast genome is 155,060 bp in length, containing 88,853 bp in a large single copy (LSC), 18,653 bp in a small single copy (SSC) and 23,777 bp in a pair of inverted repeats (IRs). A total of 112 genes including 78 protein-coding genes and 34 structural RNA genes were identified. The sequence comparison of two L. japonica collected from Korea and China revealed 48 single nucleotide polymorphisms (SNPs) and 45 insertions/deletions (InDels). In addition, phylogenetic analysis represented intraspecific diversity within L. japonica species collected in Korea and China.

Effect of the ethylene precursor, 1-aminocyclopropane-1-carboxylic acid on different growth stages of Haematococcus pluvialis.

In this study, we explored the effects of ACC on other stages of H. pluvialis. Interestingly, even though ACC displayed a dose-dependent effect on astaxanthin production, it is evident that astaxanthin production could be facilitated whenever the cells were treated at the early red stage. The transcriptional levels of BKT, CHY, SOD, and CAT genes supported enhanced astaxanthin biosynthesis upon ACC treatment at the early red stage. The combinatorial synergistic effect of ACC and light intensity was also confirmed. Finally, two-step application of ACC at the vegetative phase to increase biomass production and at the early-red stage to promote astaxanthin biosynthesis was proposed to maximize the efficiency of ACC treatment.

CLAGen: a tool for clustering and annotating gene sequences using a suffix tree algorithm.

Most multiple gene sequence alignment methods rely on conventions regarding the score of a multiple alignment in pairwise fashion. Therefore, as the number of sequences increases, the runtime of sequencing expands exponentially. In order to solve the problem, this paper presents a multiple sequence alignment method using a linear-time suffix tree algorithm to cluster similar sequences at one time without pairwise alignment. After searching for common subsequences, cross-matching common subsequences were generated, and sometimes inexact matching was found. So, a procedure aimed at masking the inexact cross-matching pairs was suggested here. In addition, BLAST was combined with a clustering tool in order to annotate the clusters generated by suffix tree clustering. The proposed method for clustering and annotating genes consists of the following steps: (1) construction of a suffix tree; (2) searching and overlapping common subsequences; (3) grouping subsequence pairs; (4) masking cross-matching pairs; (5) clustering gene sequences; (6) annotating gene clusters by the BLAST search. The performance of the proposed system, CLAGen, was successfully evaluated with 42 gene sequences in a TCA cycle (a citrate cycle) of bacteria. The system generated 11 clusters and found the longest subsequences of each cluster, which are biologically significant.

Biodegradable aliphatic polyester ionomers.

A series of polyester-based ionomers containing dimethyl 5-sulfoisophthalate sodium salt (DMSI) with up to 5 mol-% diacid units was synthesized by two-step polycondensation. Furthermore a polylactic acid (PLA) was modified by introducing poly(ethylene-co-methacrylic acid) ionomers with different ion groups, such as Na, Li, and Zn, using the melt blending method. This review examines the effect of the ionic group on the dynamic mechanical properties, melt rheology, crystallization behavior, degradation behavior, and biocompatibility with human dermal cells, as well as the nucleating effect of poly(ethylene-co-methacrylic acid) ionomer in PLA.

Cellular interactions and degradation of aliphatic poly(ester amide)s derived from glycine and/or 4-amino butyric acid.

A series of poly(ester amide)s derived from amino acid (glycine or 4-amino butyric acid), diol (1,6-hexanediol or 1,4-butanediol) and sebacoyl chloride were prepared by interfacial polymerization. FT-IR analysis indicated that for poly(ester amide)s derived from glycine, only amide-amide hydrogen bonds and hydrogen-bonded C=O ester groups were established, whereas the poly(ester amide)s derived from 4-amino butyric acid contained amide-amide hydrogen bonds and amide-ester hydrogen bonds, including NH groups and C=O ester groups in free state. The biodegradability was estimated by weight residue of poly(ester amide) films in pH 6 buffer solution with papain at 37 degrees C. The poly(ester amide) films derived from glycine demonstrated significantly improved degradability compared to the poly(ester amide) films derived from 4-amino butyric acid. This difference of degradation rate could be explained by the bonding state in C=O ester groups. The cellular interaction of the poly(ester amide)s was studied by measuring the proliferation of human dermal fibroblasts on the polymer films. The cells proliferated significantly faster on poly(ester amide) films derived from 4-amino butyric acid than on poly(ester amide) films derived from glycine. These results suggest that the poly(ester amide) prepared in this study may serve as a potential cell-compatible biomedical material.