Evolutionary Patterns of the Chloroplast Genome in Vanilloid Orchids (Vanilloideae, Orchidaceae).

The Vanilloideae (vanilloids) is one of five subfamilies of Orchidaceae and is composed of fourteen genera and approximately 245 species. In this study, the six new chloroplast genomes (plastomes) of vanilloids (two Lecanorchis, two Pogonia, and two Vanilla species) were decoded, and then the evolutionary patterns of plastomes were compared to all available vanilloid plastomes. Pogonia japonica has the longest plastome, with 158,200 bp in genome size. In contrast, Lecanorchis japonica has the shortest plastome with 70,498 bp in genome size. The vanilloid plastomes have regular quadripartite structures, but the small single copy (SSC) region was drastically reduced. Two different tribes of Vanilloideae (Pogonieae and Vanilleae) showed different levels of SSC reductions. In addition, various gene losses were observed among the vanilloid plastomes. The photosynthetic vanilloids (Pogonia and Vanilla) showed signs of stage 1 degradation and had lost most of their ndh genes. The other three species (one Cyrotsia and two Lecanorchis), however, had stage 3 or stage 4 degradation and had lost almost all the genes in their plastomes, except for some housekeeping genes. The Vanilloideae were located between the Apostasioideae and Cypripedioideae in the maximum likelihood tree. A total of ten rearrangements were found among ten Vanilloideae plastomes when compared to the basal Apostasioideae plastomes. The four sub-regions of the single copy (SC) region shifted into an inverted repeat (IR) region, and the other four sub-regions of the IR region shifted into the SC regions. Both the synonymous (dS) and nonsynonymous (dN) substitution rates of IR in-cooperated SC sub-regions were decelerated, while the substitution rates of SC in-cooperated IR sub-regions were accelerated. A total of 20 protein-coding genes remained in mycoheterotrophic vanilloids. Almost all these protein genes show accelerated base substitution rates compared to the photosynthetic vanilloids. Two of the twenty genes in the mycoheterotrophic species faced strong "relaxed selection" pressure (p-value < 0.05).

Ancient Horizontal Gene Transfers from Plastome to Mitogenome of a Nonphotosynthetic Orchid, Gastrodia pubilabiata (Epidendroideae, Orchidaceae).

Gastrodia pubilabiata is a nonphotosynthetic and mycoheterotrophic orchid belonging to subfamily Epidendroideae. Compared to other typical angiosperm species, the plastome of G. pubilabiata is dramatically reduced in size to only 30,698 base pairs (bp). This reduction has led to the loss of most photosynthesis-related genes and some housekeeping genes in the plastome, which now only contains 19 protein coding genes, three tRNAs, and three rRNAs. In contrast, the typical orchid species contains 79 protein coding genes, 30 tRNAs, and four rRNAs. This study decoded the entire mitogenome of G. pubilabiata, which consisted of 44 contigs with a total length of 867,349 bp. Its mitogenome contained 38 protein coding genes, nine tRNAs, and three rRNAs. The gene content of G. pubilabiata mitogenome is similar to the typical plant mitogenomes even though the mitogenome size is twice as large as the typical ones. To determine possible gene transfer events between the plastome and the mitogenome individual BLASTN searches were conducted, using all available orchid plastome sequences and flowering plant mitogenome sequences. Plastid rRNA fragments were found at a high frequency in the mitogenome. Seven plastid protein coding gene fragments (ndhC, ndhJ, ndhK, psaA, psbF, rpoB, and rps4) were also identified in the mitogenome of G. pubilabiata. Phylogenetic trees using these seven plastid protein coding gene fragments suggested that horizontal gene transfer (HGT) from plastome to mitogenome occurred before losses of photosynthesis related genes, leading to the lineage of G. pubilabiata. Compared to species phylogeny of the lineage of orchid, it was estimated that HGT might have occurred approximately 30 million years ago.

Plastome evolution and phylogeny of subtribe Aeridinae (Vandeae, Orchidaceae).

Subtribe Aeridinae (Vandeae, Epidendroideae, Orchidaceae) consists of 83 genera and 2,345 species. The present study completely decoded the plastomes and nuclear ribosomal (nr) RNA gene clusters of seven species of Aeridinae belonging to Gastrochilus, Neofinetia, Pelatantheria, and Thrixspermum and compared them with existing data to investigate their genome evolution and phylogeny. Although no large structural variations were observed among the Aeridinae plastomes, 14 small inversions (SI) were found in Orchidaceae for the first time. Therefore, the evolutionary trends and usefulness of SI as molecular identification markers were evaluated. Since all 11 ndh genes in the Aeridinae plastome were lost or pseudogenized, the evolutionary trends of ndh genes are discussed at the tribe and family levels. In the maximum likelihood tree reconstructed from 83 plastome genes, the five Orchidaceae subfamilies were shown to have diverged in the following order: Apostasioideae, Vanilloideae, Cypripedioideae, Orchioideae, Epidendroideaeae. Divergence times for major lineages were found to be more recent, 5-10 Mya, than previous studies, which only used two or three genes. Vandeae, which includes Aeridinae, formed a sister group with Cymbidieae and Epidendreae. The Vandeae, Cymbidieae, and Epidendreae lineages were inferred to have diverged at 25.31 Mya; thus, numerous speciation events within Aeridineae occurred since then. Furthermore, the present study reconstructed a phylogenetic tree from 422 nrITS sequences belonging to Aerdinae and allied taxa and uses it to discuss the phylogenetic positions and species identities of five endangered species.

The influence of host genotype and salt stress on the seed endophytic community of salt-sensitive and salt-tolerant rice cultivars.

BACKGROUND: Inherent characteristics and changes in the physiology of rice as it attains salt tolerance affect the colonizing bacterial endophytic communities of the rice seeds. These transmissible endophytes also serve as a source of the plant's microbial community and concurrently respond to the host and environmental conditions. This study explores the influence of the rice host as well as the impact of soil salinity on the community structure and diversity of seed bacterial endophytes of rice with varying tolerance to salt stress. Endophytic bacterial diversity was studied through culture-dependent technique and Terminal-Restriction Fragment Length Polymorphism (T-RFLP) analysis. RESULTS: Results revealed considerably diverse communities of bacterial endophytes in the interior of rice seeds. The overall endophytic bacterial communities of the indica rice seeds based on 16S rRNA analysis of clones and isolates are dominated by phylum Proteobacteria followed by Actinobacteria and Firmicutes. Community profiles show common ribotypes found in all cultivars of the indica subspecies representing potential core microbiota belonging to Curtobacterium, Flavobacterium, Enterobacter, Xanthomonas, Herbaspirillum, Microbacterium and Stenotrophomonas. Clustering analysis shows that the host genotype mainly influences the seed endophytic community of the different rice cultivars. Under salt stress conditions, endophytic communities of the salt-sensitive and salt-tolerant rice cultivars shift their dominance to bacterial groups belonging to Flavobacterium, Pantoea, Enterobacter, Microbacterium, Kosakonia and Curtobacterium. CONCLUSION: The endophytic communities of rice indica seeds are shaped by the hosts' genotype, their physiological adaptation to salt stress and phylogenetic relatedness. Under salt stress conditions, a few groups of bacterial communities become prominent causing a shift in bacterial diversity and dominance.

Characterizing endophytic competence and plant growth promotion of bacterial endophytes inhabiting the seed endosphere of Rice.

BACKGROUND: Rice (Oryza sativa L. ssp. indica) seeds as plant microbiome present both an opportunity and a challenge to colonizing bacterial community living in close association with plants. Nevertheless, the roles and activities of bacterial endophytes remain largely unexplored and insights into plant-microbe interaction are compounded by its complexity. In this study, putative functions or physiological properties associated with bacterial endophytic nature were assessed. Also, endophytic roles in plant growth and germination that may allow them to be selectively chosen by plants were also studied. RESULTS: The cultivable seed endophytes were dominated by Proteobacteria particularly class Gammaproteobacteria. Highly identical type strains were isolated from the seed endosphere regardless of the rice host's physiological tolerance to salinity. Among the type strains, Flavobacterium sp., Microbacterium sp. and Xanthomonas sp. were isolated from the salt-sensitive and salt-tolerant cultivars. PCA-Biplot ordination also showed that specific type strains isolated from different rice cultivars have distinguishing similar characteristics. Flavobacterium sp. strains are phosphate solubilizers and indole-3-acetic acid producers with high tolerance to salinity and osmotic stress. Pseudomonas strains are characterized as high siderophore producers while Microbacterium sp. and Xanthomonas sp. strains have very high pectinase and cellulase activity. Among the physiological traits of the seed endophytes, bacterial pectinase and cellulase activity are positively correlated as well as salt and osmotic tolerance. Overall characterization shows that majority of the isolates could survive in 4-8% salt concentration as well as in 0.6 M and 1.2 M sucrose solution. The activities of catalase, pectinase and cellulase were also observed in almost all of the isolates indicating the importance of these characteristics for survival and colonization into the seed endosphere. Seed bacterial endophytes also showed promising plant growth promoting activities including hormone modulation, nitrogen fixation, siderophore production and phosphate solubilization. CONCLUSION: Though many of the isolates possess similar PGP and endophytic physiological traits, this study shows some prominent and distinguishing traits among bacterial groups indicating key determinants for their success as endophytes in the rice seed endosphere. Rice seeds are also inhabited by bacterial endophytes that promote growth during early seedling development.

Inhibition of Tolaasin Cytotoxicity Causing Brown Blotch Disease in Cultivated Mushrooms Using Tolaasin Inhibitory Factors.

Tolaasin, a pore-forming bacterial peptide toxin secreted by Pseudomonas tolaasii, causes brown blotch disease in cultivated mushrooms by forming membrane pores and collapsing the membrane structures. Tolaasin is a lipodepsipeptide, MW 1985, and pore formation by tolaasin molecules is accomplished by hydrophobic interactions and multimerizations. Compounds that inhibit tolaasin toxicity have been isolated from various food additives. Food detergents, sucrose esters of fatty acids, and polyglycerol esters of fatty acids can effectively inhibit tolaasin cytotoxicity. These chemicals, named tolaasin-inhibitory factors (TIF), were effective at concentrations ranging from 10-4 to 10-5 M. The most effective compound, TIF 16, inhibited tolaasin-induced hemolysis independent of temperature and pH, while tolaasin toxicity increased at higher temperatures. When TIF 16 was added to tolaasin-pretreated erythrocytes, the cytotoxic activity of tolaasin immediately stopped, and no further hemolysis was observed. In the artificial lipid bilayer, the single-channel activity of the tolaasin channel was completely and irreversibly blocked by TIF 16. When TIF 16 was sprayed onto pathogen-treated oyster mushrooms growing on the shelves of cultivation houses, the development of disease was completely suppressed, and normal growth of oyster mushrooms was observed. Furthermore, the treatment with TIF 16 did not show any adverse effect on the growth of oyster mushrooms. These results indicate that TIF 16 is a good candidate for the biochemical control of brown blotch disease.

Enhancing Microalgae Content in Biocomposites through a Mechanical Grinding Method.

Microalgae-based biocomposites are gaining traction as ecofriendly and cost-effective alternatives to conventional petroleum-based plastics. However, achieving a homogeneous dispersion of microalgae within a biocomposite matrix remains a challenge. In this study, we investigated the effect of the size of dried microalgae (Chlorella sp.) on the quality of biocomposites. Ball milling, a mechanical grinding process, was used to control the size of the pretreated dried microalgae. Our results demonstrate that the microalgae size strongly depends on the total weight of the stainless-steel balls, rather than the number of balls used in the milling process. Poly(ethylene-vinyl acetate) (EVA), with functional groups resembling those of Chlorella sp., was incorporated into the ball-milled microalgae to produce homogeneous biocomposites. Smaller Chlorella sp. particles improved the ratio of microalgae and the mechanical properties of the biocomposites. Dried Chlorella sp. particles up to 161.43 µm, which were 72.84% smaller than the untreated microalgae, were obtained after 6 h of ball milling using 3/8-inch balls. This enabled the production of biocomposites with 60 wt.% microalgae and 61.02% of the tensile strength of pure EVA, comparable to traditional polymers. Our findings suggest that controlling the microalgae size through ball milling can improve the quality of microalgae-based biocomposites.

Effect of selenium and tungsten on cell growth and metabolite production in syngas fermentation using "Clostridium autoethanogenum".

The effect of tungsten and selenium on cell growth and production of metabolites such as acetic acid and ethanol when fermenting syngas using "Clostridium autoethanogenum" was investigated to improve the process efficiency. General concentrations of selenium and tungsten in the medium are 0.01 µM during acetogenic syngas fermentation. We conducted culture experiments at concentrations of 0, 0.001, 0.01 and 0.1 µM for each heavy metal. The effect of selenium on cell growth and total metabolite production was greater than that of tungsten as the effect of selenium on formate dehydrogenase, an important enzyme of the Wood-Ljungdahl pathway, is greater than that of tungsten. Although an increase in tungsten had a marginal effect on total metabolite production, the ethanol/acetic acid production ratio increased significantly due to a decrease in acetic acid and an increase in ethanol production. Thus, tungsten plays a key role in activating aldehyde:ferredoxin oxidoreductase, a key enzyme in the reduction of acetate to ethanol. A specific ethanol productivity of 0.462 g ethanol/g DCW∙d was obtained in a culture using 0.01 µM selenium and 0.1 µM tungsten, which was 2.18 times higher than when using 0.01 µM of both selenium and tungsten.

Optimization of the Bacteriophage Cocktail for the Prevention of Brown Blotch Disease Caused by Pseudomonas tolaasii.

Brown blotch disease, caused by Pseudomonas tolaasii, is one of the most serious diseases in mushroom cultivation, and its control remains an important issue. This study isolated and evaluated pathogen-specific bacteriophages for the biological control of the disease. In previous studies, 23 varieties of P. tolaasii were isolated from infected mushrooms with disease symptoms and classified into three subtypes, Ptα, Ptß, and Ptγ, based on their 16S rRNA gene sequences analysis and pathogenic characters. In this study, 42 virulent bacteriophages were isolated against these pathogens and tested for their host range. Some phages could lyse more than two pathogens only within the corresponding subtype, and no phage exhibited a wide host range across different pathogen subtypes. To eliminate all pathogens of the Ptα, Ptß, and Ptγ subtype, corresponding phages of one, six, and one strains were required, respectively. These phages were able to suppress the disease completely, as confirmed by the field-scale on-farm cultivation experiments. These results suggested that a cocktail of these eight phages is sufficient to control the disease induced by all 23 P. tolaasii pathogens. Additionally, the antibacterial effect of this phage cocktail persisted in the second cycle of mushroom growth on the cultivation bed.

How high-energy physics plans its future.

Past and present chairs of the Division of Particles and Fields of the American Physical Society explain how the high-energy physics community in the US decides the priorities for research through regular planning exercises that started 40 years ago at Snowmass, Colorado.

The Chloroplast Phylogenomics and Systematics of Zoysia (Poaceae).

The genus Zoysia Willd. (Chloridoideae) is widely distributed from the temperate regions of Northeast Asia-including China, Japan, and Korea-to the tropical regions of Southeast Asia. Among these, four species-Zoysia japonica Steud., Zoysia sinica Hance, Zoysia tenuifolia Thiele, and Zoysia macrostachya Franch. & Sav.-are naturally distributed in the Korean Peninsula. In this study, we report the complete plastome sequences of these Korean Zoysia species (NCBI acc. nos. MF953592, MF967579~MF967581). The length of Zoysia plastomes ranges from 135,854 to 135,904 bp, and the plastomes have a typical quadripartite structure, which consists of a pair of inverted repeat regions (20,962~20,966 bp) separated by a large (81,348~81,392 bp) and a small (12,582~12,586 bp) single-copy region. In terms of gene order and structure, Zoysia plastomes are similar to the typical plastomes of Poaceae. The plastomes encode 110 genes, of which 76 are protein-coding genes, 30 are tRNA genes, and four are rRNA genes. Fourteen genes contain single introns and one gene has two introns. Three evolutionary hotspot spacer regions-atpB~rbcL, rps16~rps3, and rpl32~trnL-UAG-were recognized among six analyzed Zoysia species. The high divergences in the atpB~rbcL spacer and rpl16~rpl3 region are primarily due to the differences in base substitutions and indels. In contrast, the high divergence between rpl32~trnL-UAG spacers is due to a small inversion with a pair of 22 bp stem and an 11 bp loop. Simple sequence repeats (SSRs) were identified in 59 different locations in Z. japonica, 63 in Z. sinica, 62 in Z. macrostachya, and 63 in Z. tenuifolia plastomes. Phylogenetic analysis showed that the Zoysia (Zoysiinae) forms a monophyletic group, which is sister to Sporobolus (Sporobolinae), with 100% bootstrap support. Within the Zoysia clade, the relationship of (Z. sinica, Z japonica), (Z. tenuifolia, Z. matrella), (Z. macrostachya, Z. macrantha) was suggested.

Turn-key constrained parameter space exploration for particle accelerators using Bayesian active learning.

Particle accelerators are invaluable discovery engines in the chemical, biological and physical sciences. Characterization of the accelerated beam response to accelerator input parameters is often the first step when conducting accelerator-based experiments. Currently used techniques for characterization, such as grid-like parameter sampling scans, become impractical when extended to higher dimensional input spaces, when complicated measurement constraints are present, or prior information known about the beam response is scarce. Here in this work, we describe an adaptation of the popular Bayesian optimization algorithm, which enables a turn-key exploration of input parameter spaces. Our algorithm replaces  the need for parameter scans while minimizing prior information needed about the measurement's behavior and associated measurement constraints. We experimentally demonstrate that our algorithm autonomously conducts an adaptive, multi-parameter exploration of input parameter space, potentially orders of magnitude faster than conventional grid-like parameter scans, while making highly constrained, single-shot beam phase-space measurements and accounts for costs associated with changing input parameters. In addition to applications in accelerator-based scientific experiments, this algorithm addresses challenges shared by many scientific disciplines, and is thus applicable to autonomously conducting experiments over a broad range of research topics.

Three complete plastome sequences from the families of Lamiaceae, Mazaceae, and Phrymaceae (Lamiales).

In this study, we report the first complete plastome sequence of Vitex rotundifolia (Lamiaceae) (MT937186). In addition, the plastome sequences of Phryma leptostachya subsp. asiatica (Phrymaceae) (153,324 bp; MT948145) and Mazus pumilus (Mazaceae) (152,847 bp; MT937187) are also included. The gene orders and structures of the three plastomes are collinear with those of the typical plastome of angiosperm. The plastome size of V. rotundifolia is 154,370 bp in length and consists of a large single-copy region of 85,079 bp and a small single-copy region of 17,917 bp, which are separated by a pair of 25,687 bp-long inverted repeat regions. In addition, the plastome sizes of P. leptostachya subsp. asiatica and M. pumilus are 153,324 bp and 152,847 bp, respectively. The three plastomes contain 113 genes, including 79 protein-coding, 30 tRNA, and four rRNA genes. Sixteen genes contain one intron and two genes have two introns. A total of 41 simple sequence repeat loci was identified in the V. rotundifolia plastome. Phylogenetic analysis shows that Viticoideae is a sister group of the last of Lamiaceae except Nepetoideae. The Mazaceae are a sister group of Lamiaceae, while Phrymaceae form a sister group to the Paulowniaceae-Orobanchaceae clade.

Temperature-dependent hemolytic activity of membrane pore-forming peptide toxin, tolaasin.

Tolaasin, a pore-forming peptide toxin produced by Pseudomonas tolaasii, causes brown blotch disease on cultivated mushrooms. Hemolysis using red blood cells was measured to evaluate the cytotoxicity of tolaasin. To investigate the mechanism of tolaasin-induced cell disruption, we studied the effect of temperature on the hemolytic process. At 4 degrees Celsius, poor binding of the tolaasin molecules to the erythrocyte membrane was observed and most of the tolaasin molecules stayed in the solution. However, once tolaasin bound to erythrocytes at 37 degrees Celsius and the temperature was decreased, complete hemolysis was observed even at 4 degrees Celsius. These results indicate that tolaasin binding to cell membrane is temperature-sensitive while tolaasin-induced membrane disruption is less sensitive to temperature change. The effect of erythrocyte concentration was measured to understand the membrane binding and pore-forming properties of tolaasin. The percentage of hemolysis measured by both hemoglobin release and cell lysis decreased as erythrocyte concentration increased in the presence of a fixed amount of tolaasin. The result shows that hemolysis is dependent on the amount of tolaasin and multiple binding of tolaasin is required for the hemolysis of a single cell. In analysis of dose-dependence, the hemolysis was proportional to the tenth power of the amount of tolaasin, implying that tolaasin-induced hemolysis can be explained by a multi-hit model.

Plastome Evolution and Phylogeny of Orchidaceae, With 24 New Sequences.

In order to understand the evolution of the orchid plastome, we annotated and compared 124 complete plastomes of Orchidaceae representing all the major lineages in their structures, gene contents, gene rearrangements, and IR contractions/expansions. Forty-two of these plastomes were generated from the corresponding author's laboratory, and 24 plastomes-including nine genera (Amitostigma, Bulbophyllum, Dactylorhiza, Dipodium, Galearis, Gymnadenia, Hetaeria, Oreorchis, and Sedirea)-are new in this study. All orchid plastomes, except Aphyllorchis montana, Epipogium aphyllum, and Gastrodia elata, have a quadripartite structure consisting of a large single copy (LSC), two inverted repeats (IRs), and a small single copy (SSC) region. The IR region was completely lost in the A. montana and G. elata plastomes. The SSC is lost in the E. aphyllum plastome. The smallest plastome size was 19,047 bp, in E. roseum, and the largest plastome size was 178,131 bp, in Cypripedium formosanum. The small plastome sizes are primarily the result of gene losses associated with mycoheterotrophic habitats, while the large plastome sizes are due to the expansion of noncoding regions. The minimal number of common genes among orchid plastomes to maintain minimal plastome activity was 15, including the three subunits of rpl (14, 16, and 36), seven subunits of rps (2, 3, 4, 7, 8, 11, and 14), three subunits of rrn (5, 16, and 23), trnC-GCA, and clpP genes. Three stages of gene loss were observed among the orchid plastomes. The first was ndh gene loss, which is widespread in Apostasioideae, Vanilloideae, Cypripedioideae, and Epidendroideae, but rare in the Orchidoideae. The second stage was the loss of photosynthetic genes (atp, pet, psa, and psb) and rpo gene subunits, which are restricted to Aphyllorchis, Hetaeria, Hexalectris, and some species of Corallorhiza and Neottia. The third stage was gene loss related to prokaryotic gene expression (rpl, rps, trn, and others), which was observed in Epipogium, Gastrodia, Lecanorchis, and Rhizanthella. In addition, an intermediate stage between the second and third stage was observed in Cyrtosia (Vanilloideae). The majority of intron losses are associated with the loss of their corresponding genes. In some orchid taxa, however, introns have been lost in rpl16, rps16, and clpP(2) without their corresponding gene being lost. A total of 104 gene rearrangements were counted when comparing 116 orchid plastomes. Among them, many were concentrated near the IRa/b-SSC junction area. The plastome phylogeny of 124 orchid species confirmed the relationship of {Apostasioideae [Vanilloideae (Cypripedioideae (Orchidoideae, Epidendroideae))]} at the subfamily level and the phylogenetic relationships of 17 tribes were also established. Molecular clock analysis based on the whole plastome sequences suggested that Orchidaceae diverged from its sister family 99.2 mya, and the estimated divergence times of five subfamilies are as follows: Apostasioideae (79.91 mya), Vanilloideae (69.84 mya), Cypripedioideae (64.97 mya), Orchidoideae (59.16 mya), and Epidendroideae (59.16 mya). We also released the first nuclear ribosomal (nr) DNA unit (18S-ITS1-5.8S-ITS2-28S-NTS-ETS) sequences for the 42 species of Orchidaceae. Finally, the phylogenetic tree based on the nrDNA unit sequences is compared to the tree based on the 42 identical plastome sequences, and the differences between the two datasets are discussed in this paper.

Corrigendum: Plastome Evolution and Phylogeny of Orchidaceae, With 24 New Sequences.

[This corrects the article DOI: 10.3389/fpls.2020.00022.].

Identification of compounds exhibiting inhibitory activity toward the Pseudomonas tolaasii toxin tolaasin I using in silico docking calculations, NMR binding assays, and in vitro hemolytic activity assays.

Using in silico docking calculations, NMR analysis of target-ligand binding, and hemolytic activity assays, we searched a 30,000-compound library for an effective inhibitor of tolaasin I, a Pseudomonas tolaasii toxin that causes virulent infection in mushrooms. Of more than 30,000 compounds screened in silico, two compounds were selected. One of these compounds, sorbitololeic acid, bound to tolaasin I and inhibited its hemolytic activity in vitro. Therefore, sorbitololeic acid can be a potential inhibitor of tolaasin I.

Aeration control of thermophilic aerobic digestion using fluorescence monitoring.

The thermophilic aerobic digestion (TAD) process is recognized as an effective method for rapid waste activated sludge (WAS) degradation and the deactivation of pathogenic microorganisms. Yet, high energy costs due to heating and aeration have limited the commercialization of economical TAD processes. Previous research on autothermal thermophilic aerobic digestion (ATAD) has already reduced the heating cost. However, only a few studies have focused on reducing the aeration cost. Therefore, this study applied a two-step aeration control strategy to a fill-and-draw mode semicontinuous TAD process. The NADH-dependent fluorescence was monitored throughout the TAD experiment, and the aeration rate shifted according to the fluorescence intensity. As a result, the simple two-step aeration control operation achieved a 20.3% reduction in the total aeration, while maintaining an effective and stable operation. It is also expected that more savings can be achieved with a further reduction of the lower aeration rate or multisegmentation of the aeration rate.

Characterization of 20 complete plastomes from the tribe Laureae (Lauraceae) and distribution of small inversions.

Lindera Thunb. (Lauraceae) consists of approximately 100 species, mainly distributed in the temperate and tropical regions of East Asia. In this study, we report 20 new, complete plastome sequences including 17 Lindera species and three related species, Actinodaphne lancifolia, Litsea japonica and Sassafras tzumu. The complete plastomes of Lindera range from 152,502 bp (L. neesiana) to 154,314 bp (L. erythrocarpa) in length. Eleven small inversion (SI) sites are documented among the plastomes. Six of the 11 SI sites are newly reported and they locate in rpoB-trnC, psbC-trnS, petA-psbJ, rpoA and ycf2 regions. The distribution patterns of SIs are useful for species identification. An average of 83 simple sequence repeats (SSRs) were detected in each plastome. The mono-SSRs accounted for 72.7% of total SSRs, followed by di- (12.4%), tetra- (9.4%), tri- (4.2%), and penta-SSRs (1.3%). Of these SSRs, 64.6% were distributed in an intergenic spacer (IGS) region. In addition, 79.8% of the SSRs are located in a large single copy (LSC) region. In contrast, almost no SSRs are distributed in inverted repeat (IR) regions. The SSR loci are useful to identifying species but the phylogenetic value is low because the majority of them show autapomorphic status or highly homoplastic characteristics. The nucleotide diversity (Pi) values also indicated the conserved nature of the IR region compared to LSC and small single copy (SSC) regions. Five spacer regions with high Pi values, trnH-psbA, petA-psbJ and ndhF-rpl32, rpl32-trnL and Ψycf1-ndhF, have a potential use for the molecular identification study of Lindera and related species. Lindera species form a paraphyletic group in the plastome tree because of the inclusion of related genera such as Actinodaphne, Laurus, Litsea and Neolitsea. A former member of tribe Laureae, Sassafras, forms a clade with the tribe Cinnamomeae. The SIs do not affect the phylogenetic relationship of Laureae. This result indicated that ancient plastome captures may have contribute to the mixed intergeneric relationship of Laureae. Alternatively, the result may indicate that the morphological characters defined the genera of Lauraceae originated for several times.

Extensive Losses of Photosynthesis Genes in the Plastome of a Mycoheterotrophic Orchid, Cyrtosia septentrionalis (Vanilloideae: Orchidaceae).

Cyrtosia septentrionalis is an achlorophyllous mycoheterotrophic orchid in the subfamily Vanilloideae (Orchidaceae). This article reports C. septentrionalis's complete plastome sequence and compare it with other orchid plastomes with a same mycoheterotrophic nutritional mode. The C. septentrionalis plastome has decreased to 96,859 bp in length, but it still maintains a quadripartite structure. The C. septentrionalis plastome contains 38 protein-coding genes, 25 tRNA genes, and four ribosomal RNA genes. Most genes related to photosynthesis have been lost, whereas the majority of housekeeping genes remain; this pattern corresponds to the end of stage 3 gene degradation. The inverted repeat regions of the C. septentrionalis plastome have decreased to 10,414 bp and mainly contain the gene ycf2. A block consisting of four rrn genes and rps7 and rps12 has shifted to a small single-copy region. As a result, the small single-copy region was found to be expanded, despite the loss of all ndh genes in the region. Three inversion mutations are required to explain the C. septentrionalis plastome's current gene order. The species is endangered, and these results have implications for its conservation.