Genome-wide analysis and expression profile of the bZIP gene family in Neopyropia yezoensis.

The basic leucine zipper (bZIP) family consists of conserved transcription factors which are widely present in eukaryotes and play important regulatory roles in plant growth, development, and stress responses. Neopyropia yezoensis is a red marine macroalga of significant economic importance; however, their bZIP family members and functions have not been systematically identified and analyzed. In the present study, the bZIP gene family in Ny. yezoensis was characterized by investigating gene structures, conserved motifs, phylogenetic relationships, chromosomal localizations, gene duplication events, cis-regulatory elements, and expression profiles. Twenty-three Ny. yezoensis bZIP (NyybZIP) genes were identified and sorted into 13 out of 30 groups, which were classified based on the bZIPs of Ny. yezoensis and 15 other red algae species. Phylogenetic analysis revealed that bZIP genes may have a complex evolutionary pattern in red algae. Cross-species collinearity analysis indicated that the bZIP genes in Ny. yezoensis, Neoporphyra haitanensis, and Porphyra umbilicalis are highly evolutionarily conserved. In addition, we identified four main categories of cis-elements, including development-related, light-responsive, phytohormone-responsive and stress-responsive promoter sequences in NyybZIP genes. Finally, RNA sequencing data and quantitative real-time PCR (qRT-PCR) showed that NyybZIP genes exhibited different expression patterns depending on the life stage. NyybZIP genes were also found to be involved in the nitrogen stress response. We thought that bZIP genes may be involved in Ny. yezoensis growth and development, and play a significant role in nitrogen deficiency response. Taken together, our findings provide new insights into the roles of the bZIP gene family and provide a basis for additional research into its evolutionary history and biological functions.

Characteristics of upper respiratory tract rhinovirus in children with allergic rhinitis and its role in disease severity.

Allergic rhinitis (AR) is a global health challenge that particularly affects the quality of life of children. Human rhinovirus (HRV) infection usually causes common cold in the upper respiratory tract (URT) and can also affect airway allergy development, such as asthma exacerbation, but its relationship with AR is poorly understood. The study aimed to gain insight into the characteristics of HRV that is prevalent in AR children and its role in AR severity. A total of 362 children with symptomatic AR were enrolled from southwestern China during 2022-2023, and nasal lavage samples were collected for HRV molecular characterization and cytokine measurement. HRV was detected in 40% of the AR children, with peak detection in autumn. The positive rate was not correlated with whether the subjects were under allergen-specific immunotherapy (AIT). Among the detected HRVs, 42% were species A, 36% were species B, and 22% were species C, involving 21 A genotypes, 6 B genotypes, and 7 C genotypes. HRV positivity was significantly associated with symptom severity (visual analog scale [VAS] score) and elevated levels of local nasal IgE, interleukin-25 (IL-25), IL-4, and CXCL13 in AR children who did not receive antiallergic treatment. All three species of HRV strains (A1B, A21, B27, B70, and C17) had been isolated and were able to infect respiratory epithelial tissue in vitro. Complete genome sequencing showed that the antigenic epitopes of the isolated HRVs had certain variations. Our work reveals the etiological characteristics of URT-HRV in AR children and suggests a role of HRV infection in the pathogenesis of childhood AR. IMPORTANCE: Our study revealed high human rhinovirus (HRV) detection rate in children with allergic rhinitis (AR), and HRV infection (A, B, or C species) is positively associated with the symptom severity in AR children. Elevated nasal IgE, interleukin-25 (IL-25), IL-4, and CXCL13 levels suggest a potential pathogenic mechanism by which HRV infection induces nasal type 2 immune/inflammation responses and local IgE production in AR patients. In addition, etiological analysis found that the main prevalent HRV species in AR children are A and B (~80%), which is different from acute respiratory infection and asthma exacerbation, where species A and C are dominant. The data reveal the distinct species prevalence characteristics of HRV infection in AR. Finally, we isolated all three species of HRV strains from nasal cavity of AR children with varying degrees of antigenic epitope mutations and in vitro infectivity, highlighting the importance of strengthening monitoring and intervention for respiratory HRV infection in AR children.

Chromosome-level genome assembly of humpback grouper using PacBio HiFi reads and Hi-C technologies.

The humpback grouper (Cromileptes altivelis), a medium-sized coral reef teleost, is a naturally rare species distributed in the tropical waters of the Indian and Pacific Oceans. It has high market value, but artificial reproduction and breeding remain limited and need to be improved. Here, we assembled the genome with 1.08 Gb, with a contig N50 of 43.78 Mb. A total of 96.59% of the assembly anchored to 24 pseudochromosomes using Hi-C technology. It contained 24,442 protein-coding sequences, of which 99.3% were functionally annotated. The completeness of the assembly was estimated to be 97.3% using BUSCO. The phylogenomic analysis suggested that humpback grouper should be classified into the genus Epinephelus rather than Cromileptes. The comparative genomic analysis revealed that the gene families related to circadian entrainment were significantly expanded. The high-quality reference genome provides useful genomic tools for exploiting the genomic resource of humpback grouper and supports the functional genomic study of this species in the future.

Characterization of the basic leucine zipper transcription factor family of Neoporphyra haitanensis and its role in acclimation to dehydration stress.

BACKGROUND: Neoporphyra haitanensis, a major marine crop native to southern China, grows in the harsh intertidal habitats of rocky coasts. The thallus can tolerate fluctuating and extreme environmental stresses, for example, repeated desiccation/rehydration due to the turning tides. It is also a typical model system for investigating stress tolerance mechanisms in intertidal seaweed. The basic leucine zipper (bZIP) transcription factors play important roles in the regulation of plants' responses to environmental stress stimuli. However, little information is available regarding the bZIP family in the marine crop Nh. haitanensis. RESULTS: We identified 19 bZIP genes in the Nh. haitanensis genome and described their conserved domains. Based on phylogenetic analysis, these 19 NhhbZIP genes, distributed unevenly on the 11 superscaffolds, were divided into four groups. In each group, there were analogous exon/intron numbers and motif compositions, along with diverse exon lengths. Cross-species collinearity analysis indicated that 17 and 9 NhhbZIP genes were orthologous to bZIP genes in Neopyropia yezoensis and Porphyra umbilicalis, respectively. Evidence from RNA sequencing (RNA-seq) data showed that the majority of NhhbZIP genes (73.68%) exhibited transcript abundance in all treatments. Furthermore, genes NN 2, 4 and 5 showed significantly altered expression in response to moderate dehydration, severe dehydration, and rehydration, respectively. Gene co-expression network analysis of the representative genes was carried out, followed by gene set enrichment analysis. Two NhhbZIP genes collectively responding to dehydration and rehydration and their co-expressing genes mainly participated in DNA repair, DNA metabolic process, and regulation of helicase activity. Two specific NhhbZIP genes responding to severe dehydration and their corresponding network genes were mainly involved in macromolecule modification, cellular catabolic process, and transmembrane transport. Three specific NhhbZIP genes responding to rehydration and their co-expression gene networks were mainly involved in the regulation of the cell cycle process and defense response. CONCLUSIONS: This study provides new insights into the structural composition, evolution, and function of the NhhbZIP gene family. Our results will help us to further study the functions of bZIP genes in response to dehydration and rehydration in Nh. haitanensis and improve Nh. haitanensis in southern China.

Genome-Wide Identification and Analysis of MYB Transcription Factors in Pyropia yezoensis.

MYB transcription factors are one of the largest transcription factor families in plants, and they regulate numerous biological processes. Red algae are an important taxonomic group and have important roles in economics and research. However, no comprehensive analysis of the MYB gene family in any red algae, including Pyropia yezoensis, has been conducted. To identify the MYB gene members of Py. yezoensis, and to investigate their family structural features and expression profile characteristics, a study was conducted. In this study, 3 R2R3-MYBs and 13 MYB-related members were identified in Py. yezoensis. Phylogenetic analysis indicated that most red algae MYB genes could be clustered with green plants or Glaucophyta MYB genes, inferring their ancient origins. Synteny analysis indicated that 13 and 5 PyMYB genes were orthologous to Pyropia haitanensis and Porphyra umbilicalis, respectively. Most Bangiaceae MYB genes contain several Gly-rich motifs, which may be the result of an adaptation to carbon limitations and maintenance of important regulatory functions. An expression profile analysis showed that PyMYB genes exhibited diverse expression profiles. However, the expression patterns of different members appeared to be diverse, and PyMYB5 was upregulated in response to dehydration, low temperature, and Pythium porphyrae infection. This is the first comprehensive study of the MYB gene family in Py. Yezoensis and it provides vital insights into the functional divergence of MYB genes.

ChIP-Based Nuclear DNA Isolation for Genome Sequencing in Pyropia to Remove Cytosol and Bacterial DNA Contamination.

Contamination from cytosolic DNA (plastid and mitochondrion) and epiphytic bacteria is challenging the efficiency and accuracy of genome-wide analysis of nori-producing marine seaweed Pyropia yezoensis. Unlike bacteria and organellar DNA, Pyropia nuclear DNA is closely associated with histone proteins. In this study, we applied Chromatin Immunoprecipitation (ChIP) of histone H3 to isolate nuclear DNA, followed by high-throughput sequencing. More than 99.41% of ChIP-sequencing data were successfully aligned to the reference nuclear genome; this was remarkably higher than those from direct extraction and direct extraction data, in which 40.96% to 42.95% are from plastids. The proportion of data that were mapped to the bacterial database when using ChIP extraction was very low. Additionally, ChIP data can cover up to 89.00% of the nuclear genome, higher than direct extraction data at equal data size and comparable to the latter at equal sequencing depth. The uncovered regions from the three methods are mostly overlapping, suggesting that incomplete sequencing accounts for the missing data, rather than failed chromatin-antibody binding in the ChIP extraction method. This ChIP extraction method can successfully separate nuclear DNA from cytosolic DNA and bacterial DNA, thus overwhelmingly reducing the sequencing cost in a genome resequencing project and providing strictly purified reference data for genome assembly. The method's applicability to other macroalgae makes it a valuable contribution to the algal research community.

Cells and Fugu Response to Capsid of BFNNV Genotype.

The nervous necrosis virus (NNV) of the BFNNV genotype is the causative agent of viral encephalopathy and retinopathy (VER) in cold water fishes. Similar to the RGNNV genotype, BFNNV is also considered a highly destructive virus. In the present study, the RNA2 of the BFNNV genotype was modified and expressed in the EPC cell line. The subcellular localization results showed that the capsid and N-terminal (1-414) were located in the nucleus, while the C-terminal (415-1014) of the capsid was located in the cytoplasm. Meanwhile, cell mortality obviously increased after expression of the capsid in EPC. EPC cells were transfected with pEGFP-CP and sampled at 12 h, 24 h and 48 h for transcriptome sequencing. There are 254, 2997 and 229 up-regulated genes and 387, 1611, and 649 down-regulated genes post-transfection, respectively. The ubiquitin-activating enzyme and ubiquitin-conjugating enzyme were up-regulated in the DEGs, indicating that cell death evoked by capsid transfection may be related to ubiquitination. The qPCR results showed that heat stock protein 70 (HSP70) is extremely up-regulated after expression of BFNNV capsid in EPC, and N-terminal is the key region to evoke the high expression. For further study, the immunoregulation of the capsid in fish pcDNA-3.1-CP was constructed and injected into the Takifugu rubripes muscle. pcDNA-3.1-CP can be detected in gills, muscle and head kidney, and lasted for more than 70 d post-injection. The transcripts of IgM and interferon inducible gene Mx were up-regulated after being immunized in different tissues, and immune factors, such as IFN-γ and C3, were also up-regulated in serum, while C4 was down-regulated one week after injection. It was suggested that pcDNA-3.1-CP can be a potential DNA vaccine in stimulating the immune system of T. rubripes; however, NNV challenge needs to be conducted in the following experiments.

Quantification of Photosynthetic Pigments in Neopyropia yezoensis Using Hyperspectral Imagery.

Phycobilisomes and chlorophyll-a (Chla) play important roles in the photosynthetic physiology of red macroalgae and serve as the primary light-harvesting antennae and reaction center for photosystem II. Neopyropia is an economically important red macroalga widely cultivated in East Asian countries. The contents and ratios of 3 main phycobiliproteins and Chla are visible traits to evaluate its commercial quality. The traditional analytical methods used for measuring these components have several limitations. Therefore, a high-throughput, nondestructive, optical method based on hyperspectral imaging technology was developed for phenotyping the pigments phycoerythrin (PE), phycocyanin (PC), allophycocyanin (APC), and Chla in Neopyropia thalli in this study. The average spectra from the region of interest were collected at wavelengths ranging from 400 to 1000 nm using a hyperspectral camera. Following different preprocessing methods, 2 machine learning methods, partial least squares regression (PLSR) and support vector machine regression (SVR), were performed to establish the best prediction models for PE, PC, APC, and Chla contents. The prediction results showed that the PLSR model performed the best for PE (R Test 2 = 0.96, MAPE = 8.31%, RPD = 5.21) and the SVR model performed the best for PC (R Test 2 = 0.94, MAPE = 7.18%, RPD = 4.16) and APC (R Test 2 = 0.84, MAPE = 18.25%, RPD = 2.53). Two models (PLSR and SVR) performed almost the same for Chla (PLSR: R Test 2 = 0.92, MAPE = 12.77%, RPD = 3.61; SVR: R Test 2 = 0.93, MAPE = 13.51%, RPD =3.60). Further validation of the optimal models was performed using field-collected samples, and the result demonstrated satisfactory robustness and accuracy. The distribution of PE, PC, APC, and Chla contents within a thallus was visualized according to the optimal prediction models. The results showed that hyperspectral imaging technology was effective for fast, accurate, and noninvasive phenotyping of the PE, PC, APC, and Chla contents of Neopyropia in situ. This could benefit the efficiency of macroalgae breeding, phenomics research, and other related applications.

Histone acetylation functions in the wound-induced spore formation in nori.

The red macroalgae Pyropia yezoensis is one of the most economically important marine crops. In the asexual reproduction process, released archeospores could provide secondary seedling resources in nori farming and be used to establish asexual seeding strategies. We previously found that wounds could induce the somatic cells in sectioned Pyropia thalli to develop into large number of asexual wound-induced spores (WIS) in a short time. Many genes involved in signaling pathways, cell division, cell wall remodeling, etc. exhibited transcriptional variation in this cell fate transition process. However, the regulatory mechanisms controlling gene transcription remain elusive. In this study, we found that suberoylanilide hydroxamic acid (SAHA), the inhibitor of histone deacetylase, strongly repressed WIS formation after wounding. The lack of a sharp increase in HDAC activity after wounding, as well as the hyperacetylated status of histone H3 and H4, were observed in SAHA-treated thalli fragments, thus confirming a histone deacetylation-related epigenetic mechanism of wound-induced cell fate reprogramming. Moreover, histone deacetylation is required in the whole process of WIS formation and release. We further compared the genome-wide transcriptional variations after SAHA treatment. SAHA-responsive genes were identified, including some transcriptional factors, chromatin remodeling complex proteins, protein kinases, etc. Transcription of RBOH genes was also altered by SAHA, and moreover, ROS signals in cut fragments were attenuated, both indicating that the ROS systematic signaling pathway is closely associated with histone deacetylation. Our findings provide insights into the biological significance of dynamic histone acetylation states in WIS formation in P. yezoensis.

Cloning, Exogenous Expression and Function Analysis of Interferon-γ from Gadus macrocephalus.

Interferon γ (IFN-γ) is now considered to be one of the key molecules in the regulation of innate and adaptive immunity. The function of IFN-γ is best described in humans, but less of IFN-γ in fish species has been described at protein level. In the present study, IFN-γ from Gadus macrocephalus (GmIFN-γ) has been examined in terms of bioinformatics, prokaryotic expression, yeast expression, antiviral activity and immune regulatory function. The cDNA of GmIFN-γ contains an open reading frame of 570 nucleotides, coding 189 amino acids. The mature protein contains a nuclear localization signal motif and an obvious IFN-γ signature sequence at the C-terminal. GmIFN-γ is very similar to that of Atlantic cod, with homology up to 89.89%, but less than 32% to other species. GmIFN-γ can be detected in the gills, spleen, intestine, brain and kidney. Interestingly, during early development, a strong signal of GmIFN-γ was not detected until 40 days post hatching. Prokaryotic expression plasmid pET-32a-GmIFN-γ was constructed, and the expression products in BL21 were confirmed by Mass Spectrometry. Meanwhile, the plasmid pGAPZA-GmIFN-γ with Myc tag was constructed and transmitted into Pichia pastoris yeast GS115, and the products were tested using Western blot. The purified GmIFN-γ from either BL21 or yeast has a strong antivirus (Spring viremia of carp virus) effect. The vector of pcDNA3.1-GmIFN-γ was expressed in EPC cell lines; high transcript levels of MHC class I chain-related protein A (MICA) gene were detected; and the exogenous GmIFN-γ protein could also induce MICA expression, indicating that GmIFN-γ could stimulate immune response. The yeast GS115 with GmIFN-γ protein, which is an inclusion body, was given to zebrafish orally, and the transcript of zebrafish IFN-γ was upregulated significantly; however, genes of the interferon type-I signal pathway were not well stimulated.

Heat Shock Protein 20 Gene Superfamilies in Red Algae: Evolutionary and Functional Diversities.

Heat shock protein 20 (Hsp20) genes play important roles in plant growth, development, and response to environmental stress. However, the Hsp20 gene family has not yet been systematically investigated, and its function in red algae (Rhodophyta) remains poorly understood. Herein, we characterized Hsp20 gene families in red algae by studying gene structure, conserved motifs, phylogenetic relationships, chromosome location, gene duplication, cis-regulatory elements, and expression profiles. In this study, 97 Hsp20 genes were identified using bioinformatic methods and classified into 13 subfamilies based on phylogenetic relationships. Phylogenetic analysis revealed that Hsp20 genes might have a polyphyletic origin and a complex evolutionary pattern. Gene structure analysis revealed that most Hsp20 genes possessed no introns, and all Hsp20 genes contained a conserved α-crystalline domain in the C-terminal region. Conserved motif analysis revealed that Hsp20 genes belonging to the same subfamily shared similar motifs. Gene duplication analysis demonstrated that tandem and segmental duplication events occurred in these gene families. Additionally, these gene families in red algae might have experienced strong purifying selection pressure during evolution, and Hsp20 genes in Pyropia yezoensis, Pyropia haitanensis, and Porphyra umbilicalis were highly evolutionarily conserved. The cis-elements of phytohormone-, light-, stress-responsive, and development-related were identified in the red algal Hsp20 gene promoter sequences. Finally, using Py. yezoensis, as a representative of red algae, the Hsp20 gene expression profile was explored. Based on the RNA-seq data, Py. yezoensis Hsp20 (PyyHsp20) genes were found to be involved in Py. yezoensis responses against abiotic and biotic stresses and exhibited diverse expression patterns. Moreover, PyyHsp20 is involved in Py. yezoensis growth and development and revealed spatial and temporal expression patterns. These results provide comprehensive and valuable information on Hsp20 gene families in red algae and lay a foundation for their functional characterization. In addition, our study provides new insights into the evolution of Hsp20 gene families in red algae and will help understand the adaptability of red algae to diverse environments.

Comparative Gene Expression and Physiological Analyses Reveal Molecular Mechanisms in Wound-Induced Spore Formation in the Edible Seaweed Nori.

Genetic reprogramming of differentiated cells is studied broadly in multicellular Viridiplantae as an adaptation to herbivory or damage; however, mechanisms underlying cell development and redifferentiation are largely unknown in red algae, their nearest multicellular relatives. Here we investgate cell reprogramming in the widely cultivated, edible seaweed Neopyropia yezoesis ("nori"), where vegetative cells in wounded blades differentiate and release as large numbers of asexual spores. Based upon physiological changes and transcriptomic dynamics after wound stress in N. yezoensis and its congener Neoporphyra haitanensis, another cultivar that does not differentiate spores after wounding, we propose a three-phase model of wound-induced spore development in N. yezoensis. In Phase I, propagation of ROS by RBOH and SOD elicites systematic transduction of the wound signal, while Ca2+ dependent signaling induces cell reprogramming. In Phase II, a TOR signaling pathway and regulation of cyclin and CDK genes result in cell divisions that spread inward from the wound edge. Once sporangia form, Phase III involves expression of proteins required for spore maturation and cell wall softening. Our analyses not only provide the first model for core molecular processes controlling cellular reprogramming in rhodophytes, but also have practical implications for achieving greater control over seeding in commercial nori farming.

Metagenome-Assembled Genomes From Pyropia haitanensis Microbiome Provide Insights Into the Potential Metabolic Functions to the Seaweed.

Pyropia is an economically important edible red alga worldwide. The aquaculture industry and Pyropia production have grown considerably in recent decades. Microbial communities inhabit the algal surface and produce a variety of compounds that can influence host adaptation. Previous studies on the Pyropia microbiome were focused on the microbial components or the function of specific microbial lineages, which frequently exclude metabolic information and contained only a small fraction of the overall community. Here, we performed a genome-centric analysis to study the metabolic potential of the Pyropia haitanensis phycosphere bacteria. We reconstructed 202 unique metagenome-assembled genomes (MAGs) comprising all major taxa present within the P. haitanensis microbiome. The addition of MAGs to the genome tree containing all publicly available Pyropia-associated microorganisms increased the phylogenetic diversity by 50% within the bacteria. Metabolic reconstruction of the MAGs showed functional redundancy across taxa for pathways including nitrate reduction, taurine metabolism, organophosphorus, and 1-aminocyclopropane-1-carboxylate degradation, auxin, and vitamin B12 synthesis. Some microbial functions, such as auxin and vitamin B12 synthesis, that were previously assigned to a few Pyropia-associated microorganisms were distributed across the diverse epiphytic taxa. Other metabolic pathways, such as ammonia oxidation, denitrification, and sulfide oxidation, were confined to specific keystone taxa.

Discrepancy in photosynthetic responses of the red alga Pyropia yezoensis to dehydration stresses under exposure to desiccation, high salinity, and high mannitol concentration.

Macroalgae that inhabit intertidal zones are exposed to the air for several hours during low tide and must endure desiccation and high variations in temperature, light intensity, and salinity. Pyropia yezoensis (Rhodophyta, Bangiales), a typical intertidal red macroalga that is commercially cultivated in the northwestern Pacific Ocean, was investigated under different dehydration stresses of desiccation, high salinity, and high mannitol concentration. Using chlorophyll fluorescence imaging, photosynthetic activities of P. yezoensis thalli were analyzed using six parameters derived from quenching curves and rapid light curves. A distinct discrepancy was revealed in photosynthetic responses to different dehydration stresses. Dehydration caused by exposure to air resulted in rapid decreases in photosynthetic activities, which were always lower than two other stresses at the same water loss (WL) level. High salinity only reduced photosynthesis significantly at its maximum WL of 40% but maintained a relatively stable maximum quantum yield of photosystem II (PSII) (Fv/Fm). High mannitol concentration induced maximum WL of 20% for a longer time (60 min) than the other two treatments and caused no adverse influences on the six parameters at different WL except for a significant decrease in non-photochemical quenching (NPQ) at 20% WL. Illustrated by chlorophyll fluorescence images, severe spatial heterogeneities were induced by desiccation with lower values in the upper parts than the middle or basal parts of the thalli. The NPQ and rETRmax (maximum relative electron transport rate) demonstrated clear distinctions for evaluating photosynthetic responses, indicating their sensitivity and applicability. The findings of this study indicated that the natural dehydration of exposure to air results in stronger and more heterogeneous effects than those of high salinity or high mannitol concentration.

Floridean Starch and Floridoside Metabolic Pathways of Neoporphyra haitanensis and Their Regulatory Mechanism under Continuous Darkness.

Floridean starch and floridoside are the main storage carbohydrates of red algae. However, their complete metabolic pathways and the origin, function, and regulatory mechanism of their pathway genes have not been fully elucidated. In this study, we identified their metabolic pathway genes and analyzed the changes in related gene expression and metabolite content in Neoporphyra haitanensis under continuous dark conditions. Our results showed that genes from different sources, including eukaryotic hosts, cyanobacteria, and bacteria, were combined to construct floridean starch and floridoside metabolic pathways in N. haitanensis. Moreover, compared with those in the control, under continuous dark conditions, floridean starch biosynthesis genes and some degradation genes were significantly upregulated with no significant change in floridean starch content, whereas floridoside degradation genes were significantly upregulated with a significant decrease in floridoside content. This implies that floridean starch content is maintained but floridoside is consumed in N. haitanensis under dark conditions. This study elucidates the "floridean starch-floridoside" metabolic network and its gene origins in N. haitanensis for the first time.

Genome-wide analysis of HSP70 gene superfamily in Pyropia yezoensis (Bangiales, Rhodophyta): identification, characterization and expression profiles in response to dehydration stress.

BACKGROUND: Heat shock proteins (HSPs) perform a fundamental role in protecting plants against abiotic stresses. Individual family members have been analyzed in previous studies, but there has not yet been a comprehensive analysis of the HSP70 gene family in Pyropia yezoensis. RESULTS: We investigated 15 putative HSP70 genes in Py. yezoensis. These genes were classified into two sub-families, denoted as DnaK and Hsp110. In each sub-family, there was relative conservation of the gene structure and motif. Synteny-based analysis indicated that seven and three PyyHSP70 genes were orthologous to HSP70 genes in Pyropia haitanensis and Porphyra umbilicalis, respectively. Most PyyHSP70s showed up-regulated expression under different degrees of dehydration stress. PyyHSP70-1 and PyyHSP70-3 were expressed in higher degrees compared with other PyyHSP70s in dehydration treatments, and then expression degrees somewhat decreased in rehydration treatment. Subcellular localization showed PyyHSP70-1-GFP and PyyHSP70-3-GFP were in the cytoplasm and nucleus/cytoplasm, respectively. Similar expression patterns of paired orthologs in Py. yezoensis and Py. haitanensis suggest important roles for HSP70s in intertidal environmental adaptation during evolution. CONCLUSIONS: These findings provide insight into the evolution and modification of the PyyHSP70 gene family and will help to determine the functions of the HSP70 genes in Py. yezoensis growth and development.

Structural and Functional Impacts of Microbiota on Pyropia yezoensis and Surrounding Seawater in Cultivation Farms along Coastal Areas of the Yellow Sea.

Pyropia yezoensis is the most important commercial edible red algae in China, carrying a variety of resident microbes at its surface. To understand microbiome diversity, community structure, interactions and functions with hosts in this regard, thalli and seawater sampleswere collected from Yantai and Rizhao cultivation farms in the Yellow Sea. The thalli and seawater samples (n = 12) were collected and studied using an Illumina NovaSeq 6000 platform and 16S ribosomal RNA (rRNA) gene sequencing, along with the consideration of environmental factors. Bacterial communities in association with P. yezoensis and surrounding seawater were predominated by Cyanobacteria, Proteobacteria, and Bacteroidetes. The variability of bacterial communities related to P. yezoensis and seawater were predominantly shaped by nitrate (NO3), ammonium (NH4), and temperature. Cluster analysis revealed a close relationship between thalli (RTH and YTH) and seawater (RSW and YSW) in terms of the residing bacterial communities, respectively. PICRUSt analysis revealed the presence of genes associated with amino acid transportation and metabolism, which explained the bacterial dependence on algal-provided nutrients. This study reveals that the diversity of microbiota for P. yezoensis is greatly influenced by abiotic factors and algal organic exudates which trigger chemical signaling and transportation responses from the bacterial community, which in turn activates genes to metabolize subsequent substrates.

Comparative Transcriptome Analysis Provides Insights into Response of Ulva compressa to Fluctuating Salinity Conditions.

Ulva compressa, a green tide-forming species, can adapt to hypo-salinity conditions, such as estuaries and brackish lakes. To understand the underlying molecular mechanisms of hypo-salinity stress tolerance, transcriptome-wide gene expression profiles in U. compressa were created using digital gene expression profiles. The RNA-seq data were analyzed based on the comparison of differently expressed genes involved in specific pathways under hypo-salinity and recovery conditions. The up-regulation of genes in photosynthesis and glycolysis pathways may contribute to the recovery of photosynthesis and energy metabolism, which could provide sufficient energy for the tolerance under long-term hyposaline stress. Multiple strategies, such as ion transportation and osmolytes metabolism, were performed to maintain the osmotic homeostasis. Additionally, several long noncoding RNA were differently expressed during the stress, which could play important roles in the osmotolerance. Our work will serve as an essential foundation for the understanding of the tolerance mechanism of U. compressa under the fluctuating salinity conditions.

Biomass estimation of cultivated red algae Pyropia using unmanned aerial platform based multispectral imaging.

BACKGROUND: Pyropia is an economically advantageous genus of red macroalgae, which has been cultivated in the coastal areas of East Asia for over 300 years. Realizing estimation of macroalgae biomass in a high-throughput way would great benefit their cultivation management and research on breeding and phenomics. However, the conventional method is labour-intensive, time-consuming, manually destructive, and prone to human error. Nowadays, high-throughput phenotyping using unmanned aerial vehicle (UAV)-based spectral imaging is widely used for terrestrial crops, grassland, and forest, but no such application in marine aquaculture has been reported. RESULTS: In this study, multispectral images of cultivated Pyropia yezoensis were taken using a UAV system in the north of Haizhou Bay in the midwestern coast of Yellow Sea. The exposure period of P. yezoensis was utilized to prevent the significant shielding effect of seawater on the reflectance spectrum. The vegetation indices of normalized difference vegetation index (NDVI), ratio vegetation index (RVI), difference vegetation index (DVI) and normalized difference of red edge (NDRE) were derived and indicated no significant difference between the time that P. yezoensis was completely exposed to the air and 1 h later. The regression models of the vegetation indices and P. yezoensis biomass per unit area were established and validated. The quadratic model of DVI (Biomass = - 5.550DVI2 + 105.410DVI + 7.530) showed more accuracy than the other index or indices combination, with the highest coefficient of determination (R2), root mean square error (RMSE), and relative estimated accuracy (Ac) values of 0.925, 8.06, and 74.93%, respectively. The regression model was further validated by consistently predicting the biomass with a high R2 value of 0.918, RMSE of 8.80, and Ac of 82.25%. CONCLUSIONS: This study suggests that the biomass of Pyropia can be effectively estimated using UAV-based spectral imaging with high accuracy and consistency. It also implied that multispectral aerial imaging is potential to assist digital management and phenomics research on cultivated macroalgae in a high-throughput way.

Pyropia yezoensis genome reveals diverse mechanisms of carbon acquisition in the intertidal environment.

Changes in atmospheric CO2 concentration have played a central role in algal and plant adaptation and evolution. The commercially important red algal genus, Pyropia (Bangiales) appears to have responded to inorganic carbon (Ci) availability by evolving alternating heteromorphic generations that occupy distinct habitats. The leafy gametophyte inhabits the intertidal zone that undergoes frequent emersion, whereas the sporophyte conchocelis bores into mollusk shells. Here, we analyze a high-quality genome assembly of Pyropia yezoensis to elucidate the interplay between Ci availability and life cycle evolution. We find horizontal gene transfers from bacteria and expansion of gene families (e.g. carbonic anhydrase, anti-oxidative related genes), many of which show gametophyte-specific expression or significant up-regulation in gametophyte in response to dehydration. In conchocelis, the release of HCO3- from shell promoted by carbonic anhydrase provides a source of Ci. This hypothesis is supported by the incorporation of 13C isotope by conchocelis when co-cultured with 13C-labeled CaCO3.