Split Sites Research Articles

Comprehensive Screening of a Light-Inducible Split Cre Recombinase with Domain Insertion Profiling.

Splitting proteins with light- or chemically inducible dimers provides a mechanism for post-translational control of protein function. However, current methods for engineering stimulus-responsive split proteins often require significant protein engineering expertise and the laborious screening of individual constructs. To address this challenge, we use a pooled library approach that enables rapid generation and screening of nearly all possible split protein constructs in parallel, where results can be read out by using sequencing. We perform our method on Cre recombinase with optogenetic dimers as a proof of concept, resulting in comprehensive data on the split sites throughout the protein. To improve the accuracy in predicting split protein behavior, we develop a Bayesian computational approach to contextualize errors inherent to experimental procedures. Overall, our method provides a streamlined approach for achieving inducible post-translational control of a protein of interest.

The Crystal Structure of Pb10(PO4)6O Revisited: The Evidence of Superstructure

The crystal structure of Pb10(PO4)6O, the proposed matrix for the potential room-temperature superconductor LK-99, Pb10−xCux(PO4)6O (x = 0.9–1.0), has been reinvestigated via single-crystal X-ray diffraction using crystals prepared by Merker and Wondratschek (Z. Anorg. Allg. Chem. 1960, 306, 25–29). The crystal structure is trigonal, P3¯, a = 9.8109(6), c = 14.8403(12) Å, V = 1237.06(15), R1 = 0.0413 using 3456 unique observed reflections. The crystal structure of Pb10(PO4)6O is a superstructure with regard to the ‘standard’ P63/m apatite structure type. The doubling of the c parameter is induced through the ordering of the split sites of ‘additional’ O’ atoms within the structure channels running parallel to the c axis and centered at (00z). The O’ atoms form short bonds to the Pb1 atoms, resulting in splitting the Pb1 site into two, Pb1A and Pb1B. The structural distortions are further transmitted to the Pb phosphate framework formed by four Pb2 sites and PO4 groups. The structure data previously reported by Krivovichev and Burns (Z. Kristallogr. 2003, 218, 357–365) may either correspond to the Pb10(PO4)6Ox(OH)2−2x (x ~ 0.4) member of the Pb10(PO4)6O—Pb10(PO4)6(OH)2 solid solution series, or to the high-temperature polymorph of Pb10(PO4)6O (with the phase with doubled c parameter being the low-temperature polymorph).

New site Power Transmitter Analysis of 4G LTE FDD 1800 MHz Using Cell Splitting Method

The advancement of cellular telecommunications is growing day by day. Some theories, including 4G LTE FDD 1800 MHz, are becoming irrelevant to the current technology. The theory of power reduction by the cell splitting method is unsuitable for the actual condi-tions in the field. Theoretically, the cell splitting method is carried out on a new site with half of the main site's coverage, in which the transmitter power at the split site is reduced by 12 dB from the main site. This results in the power generated from the calculation be-ing small so that service coverage is limited. This study aimed to test the theory of trans-mitter power reduction in the cell splitting method (scenario 1) and compare it with other power reduction scenarios (Scenario 1: reduced by 15 dB, and Scenario 2: reduced by half from the main site power). The analysis is done by conducting simulations using Atoll RF Planning software. The study results revealed that in Scenario 3, the value of Key Perfor-mance Indicators is better than in other scenarios. The average throughput is 11045 kbps, the RSRP value is -108.58dBm, the range is 1.045 Km, and the average SINR is reduced to 8.52 dB. In addition, the data revealed that the coverage and signal quality of the site cell splitting is better when the transmitter power is 43 dBm (scenario 3). In conclusion, a site with higher transmitter power tends to have better signal strength, quality, and speed, as well as a wider coverage area. Reducing the transmitter cell splitting power will be more effective by reducing the new power site to half of the main site.

Dorsal bony spur in pediatric split cord malformations: eight-year experience from a tertiary care hospital.

Dorsal spurs in Type I split cord malformations (SCM-I) are infrequent findings. The pathogenesis of the same is debatable. The objective of this study is to analyze our experience with SCM-I patients having dorsal bony spurs. Retrospective analysis of SCM patients operated from 2010 to 2017 was performed. Their demographic profile, clinic-radiological features, operative findings, and outcome following surgery were recorded. Twenty-four cases of Type I SCM harboring dorsal bony spurs were identified with mean age of 4.96years. The commonest split site was lumbar, documented in 62.5%. Scoliosis was observed in 58.3%. Pre-operative neurological deficits were seen in 66.6% cases with asymmetric weakness of limbs seen in 16.6%. There was no new neurological deficit observed post-operatively. This is the largest series of dorsal spurs occurring in SCM, reported in literature so far. Meticulous pre-operative evaluation and imaging are important to identify dorsal spurs for appropriate management and good clinical outcome. Differentiating dorsal spur from ventral spur is important as it has a bearing on surgical approach.

Characteristics, Recombination Methods, and Applications Progresses of Split-Cas9 System.

CRISPR technology has been used to revolutionize various facets of life sciences because of its potent gene editing capabilities. In particular, CRISPR technology is anticipated to be used to cure congenital disorders, and malignant cancers brought on by gene mutation. In this article, we introduce a Split-Cas9 system, in which Cas9 protein is split into two or more parts and recombined in cells to function specific induction circumstances. Split-Cas9 system can improve the therapeutic index of CRISPR technology by splitting Cas9 proteins into small fragments, thus enhancing their compatibility with virus vectors and precise temporal and spatial control. This article examined the combination mode of Split-Cas9 system, contrasted the differences in its split sites and activity efficiency, and discussed the use and clinical transformation in vivo and in vitro.

High-throughput library transgenesis in Caenorhabditis elegans via Transgenic Arrays Resulting in Diversity of Integrated Sequences (TARDIS)

High-throughput transgenesis using synthetic DNA libraries is a powerful method for systematically exploring genetic function. Diverse synthesized libraries have been used for protein engineering, identification of protein–protein interactions, characterization of promoter libraries, developmental and evolutionary lineage tracking, and various other exploratory assays. However, the need for library transgenesis has effectively restricted these approaches to single-cell models. Here, we present Transgenic Arrays Resulting in Diversity of Integrated Sequences (TARDIS), a simple yet powerful approach to large-scale transgenesis that overcomes typical limitations encountered in multicellular systems. TARDIS splits the transgenesis process into a two-step process: creation of individuals carrying experimentally introduced sequence libraries, followed by inducible extraction and integration of individual sequences/library components from the larger library cassette into engineered genomic sites. Thus, transformation of a single individual, followed by lineage expansion and functional transgenesis, gives rise to thousands of genetically unique transgenic individuals. We demonstrate the power of this system using engineered, split selectable TARDIS sites in Caenorhabditis elegans to generate (1) a large set of individually barcoded lineages and (2) transcriptional reporter lines from predefined promoter libraries. We find that this approach increases transformation yields up to approximately 1000-fold over current single-step methods. While we demonstrate the utility of TARDIS using C. elegans, in principle the process is adaptable to any system where experimentally generated genomic loci landing pads and diverse, heritable DNA elements can be generated.

High-throughput library transgenesis in Caenorhabditis elegans via Transgenic Arrays Resulting in Diversity of Integrated Sequences (TARDIS).

High-throughput transgenesis using synthetic DNA libraries is a powerful method for systematically exploring genetic function. Diverse synthesized libraries have been used for protein engineering, identification of protein-protein interactions, characterization of promoter libraries, developmental and evolutionary lineage tracking, and various other exploratory assays. However, the need for library transgenesis has effectively restricted these approaches to single-cell models. Here, we present Transgenic Arrays Resulting in Diversity of Integrated Sequences (TARDIS), a simple yet powerful approach to large-scale transgenesis that overcomes typical limitations encountered in multicellular systems. TARDIS splits the transgenesis process into a two-step process: creation of individuals carrying experimentally introduced sequence libraries, followed by inducible extraction and integration of individual sequences/library components from the larger library cassette into engineered genomic sites. Thus, transformation of a single individual, followed by lineage expansion and functional transgenesis, gives rise to thousands of genetically unique transgenic individuals. We demonstrate the power of this system using engineered, split selectable TARDIS sites in Caenorhabditis elegans to generate (1) a large set of individually barcoded lineages and (2) transcriptional reporter lines from predefined promoter libraries. We find that this approach increases transformation yields up to approximately 1000-fold over current single-step methods. While we demonstrate the utility of TARDIS using C. elegans, in principle the process is adaptable to any system where experimentally generated genomic loci landing pads and diverse, heritable DNA elements can be generated.

Structure, site symmetry and spin-orbit coupled magnetism of a Ca12Al14O33 mayenite single crystal substituted with 0.26 at.% Ni

A Ca12Al14O33 mayenite single crystal with 0.26(1) at.% of the metals substituted by Ni was synthesized. Single crystal XRD confirms that about 10 at.% of Al and Ca partially occupy split crystallographic sites with lowered point symmetry. Ni K-edge XAS reveals that Ni is present as Ni2+ and substitutes Al on tetrahedral oxygen coordinated sites. UV-VIS analysis further points to the partial occupation of the trigonally distorted Al(1) and Al(1B) sites by Ni2+. In accordance with the results from the analytical methods, the magnetic model consists of Ni2+ with spin S = 1 on a trigonally distorted tetrahedral coordination site. The J-mulitplet splitting due to spin-orbit coupling with a non-magnetic J = 0 ground state determines the characteristic thermomagnetic properties which show a maximum of the effective paramagnetic moment of 3.61(1) μB at about 191 K and an uniaxial anisotropy (crystal field parameter B02 = 69.5(2) cm−1).

Screening an Effective Dual-Adeno-Associated Virus Split-Cytosine Base Editor System for C-to-T Conversion In Vivo.

The cytosine base editor (CBE) has shown promise as a gene editing tool for gene therapy, as it can convert cytidine to thymidine. Adeno-associated virus (AAV) has been widely used for in vivo gene therapy, but its limited 4.7 kb packing capacity presents challenges in delivering CBE by a single AAV. To address this, one feasible solution is to split CBE into two sections for dual-AAV delivery. In this study, we utilized BE3 as an example and constructed 22 potential split-BE3 pairs with the combination of 11 splitting sites and two split-inteins (Npu and Rma). These split-BE3 pairs were initially screened in the green fluorescent protein (GFP) reporter system, with six split-BE3 pairs selected for further evaluation. The subsequent screening of split-BE3 pairs was performed at two endogenous sites in 293T and HeLa cells, revealing that the split-BE3-Rma674, split-BE3-Rma713, and split-BE3-Rma1005 displayed effective C-to-T conversion after transfection. The effectiveness of dual-AAV split-BE3 was further validated in culture cells and adult mouse eyes. Of note, the split-BE3-Rma674 demonstrated the most efficient C-to-T editing after AAV infection, with a maximal editing efficiency of 23.29% ± 10.98% in the mouse retinal pigment epithelium cells in vivo. Overall, our study presents a novel split-BE3 system with effective C-to-T conversion, which could be applied to CBE-based in vivo gene therapy.

Light-Up Aptameric Sensor of Serotonin for Point-of-Care Use.

Serotonin is a vital neurotransmitter for regulating organism functions, and its abnormal level indicates multiple diseases. Aptamer has emerged as an innovative tool for serotonin analysis very recently; however, the current aptameric sensing platform lacks design flexibility and portability. Here, we introduce a light-up aptameric sensor using designer DNA molecules with tunable affinity and dynamic response and achieve mobile phone-based detection for point-of-care use. We develop a type of allosteric DNA sensor through flanking the serotonin recognition domain with split fluorogenic sequences, where both linker lengths and split sites of the aptamer affect its function. In addition, we design a series of molecular constructs that contain nucleotide mutations and systematically investigate the structure folding and ligand binding of the aptameric molecules. The results show distinct effects of variant mutation sites on conformation change and sensing responses. Notably, the variable aptameric molecules allow affinity and dynamic response regulation, which are adaptable to diverse sensing applications that require different threshold levels. Furthermore, we demonstrate a simple surface-based assay that can use smartphone imaging to visualize results for diagnosis. In a portable and simple manner, highly sensitive and selective serotonin assay is achieved in different biofluids, with detection limits in the low nanomolar range. This study offers an alternative approach for serotonin assay using engineered aptameric molecular probes. We expect that the practical utility may make the method promising in resource-limited settings.

Split AAV8 Mediated ABCA4 Expression for Gene Therapy of Mouse Stargardt Disease (STGD1).

Adeno-associated virus (AAV)-based gene therapy has been shown to be safe and effective in numerous animal models and clinical trials for various ophthalmic diseases. Stargardt disease (STGD1; MIM #248200) is the most common autosomal recessive macular dystrophy disease, and the most common form is caused by mutations in the ABCA4 gene, a gene with 6.8 kb coding sequence. Split intein approaches increase the capacity of dual AAV gene therapy, but at the cost of reduced protein expression, which may be insufficient to achieve a therapeutic effect. In this study, we designed various dual split intein ABCA4 vectors and showed that the efficiency of expression of full-length ABCA4 protein is dependent on combinations of types and split sites of the intein system. The most efficient vectors were identified through in vitro screening, and a novel dual AAV8-ABCA4 vector was constructed and subsequently proven to express full-length ABCA4 protein at a high level, reducing bisretinoid formation and correcting the visual function of ABCA4-knockout mice. Furthermore, we evaluated therapeutic effects of different dosages by subretinal injection in mice model. Both therapeutic effects and safety were guaranteed under the treatment of 1.00 × 109 GC/eye. These results support the optimized dual AAV8-ABCA4 approach in future clinical translation for treatment of Stargardt disease.

Boronic Acid Assisted Self-Assembly of Functional RNAs.

Boronate esters formed by reaction of an oligonucleotide carrying a 5'-boronic acid moiety with the 3'-terminal cis-diol of another have been shown previously to assist assembly of fragmented DNAzymes. Here we demonstrate that boronate esters replacing the natural phosphodiester linkage at selected sites of two functional RNAs, the hairpin ribozyme and the Mango aptamer, allow assembly of functional structures. The hairpin ribozyme, a small naturally occurring RNA that supports the reversible cleavage of appropriate RNA substrates, is very sensitive to fragmentation. Splitting the ribozyme at four different sites led to a significant decrease or even loss of cleavage and ligation activity. Ribozymes assembled from fragments capable of boronate ester formation showed restoration of cleavage activity in some but not all cases, dependent on the split site. Ligation proved to be more challenging, no supportive effect of the boronate ester was observed. Split variants of the Mango aptamer also showed a dramatic loss of functionality, which however, was restored when 5'-boronic acid modified fragments were used for assembly. These studies show for the first time that boronate esters as internucleoside linkages can act as surrogates of natural phosphodiesters in functional RNA molecules.

Preparation, crystal structure and Li ion conductivity of Li10.8Ba1.85Al3.5O12F

Polycrystalline ceramics comprising the newly discovered compound Li10.8Ba1.85Al3.5O12F were synthesized by heating a compacted mixture of Li2O, BaO and Al2O3 together with LiF as a fluorine source at 1023 K for 6 h in an Ar atmosphere. Single crystals of this oxide were subsequently obtained from a melt-solidified sample prepared by heating and slow cooling of the compact from 1133 K. Single-crystal X-ray diffraction established that this compound had a trigonal crystal structure (a = 6.03346(10) Å, c =10.13780(17) Å, space group P-3) containing Li split sites around the (001) plane at z = 0 with approximately 1/3 occupancy and large anisotropic atomic displacement parameters. The Li ion conductivities for the samples prepared by reaction sintering at 1023 K (relative density 55%) and by sintering of a powder compact at 1123 K (relative density 87%) were determined to be 1.6(3) × 10−8 and 2.8(2) × 10−8 S·cm−1 at 305 K and 3.9(2) × 10−6 and 1.0(2) × 10−5 S·cm−1 at 423 K, respectively, while the activation energies for conduction of the samples was found to be 0.52(2) and 0.55(3) eV, respectively. Cyclic voltammetry data acquired using Li and Ta electrodes indicated that this compound was stable in contact with Li metal and demonstrated a Li ion transport number >99%.

Polyhedral Distortions and Unusual Magnetic Order in Spinel FeMn2O4.

Spinel compounds AB2X4 consist of both tetrahedral (AX4) and octahedral (BX6) environments with the former forming a diamond lattice and the latter a geometrically frustrated pyrochlore lattice. Exploring the fascinating physical properties and their correlations with structural features is critical in understanding these materials. FeMn2O4 has been reported to exhibit one structural transition and two successive magnetic transitions. Here, we report the polyhedral distortions and their correlations to the structural and two magnetic transitions in FeMn2O4 by employing the high-resolution neutron powder diffraction. The cation distribution is found to be (Mn0.92+Fe0.13+)A(Mn3+Fe0.93+Mn0.12+)BO4. While large trigonal distortion is found even in the high-temperature cubic phase, the first-order cubic-tetragonal structural transition associated with the elongation of both tetrahedra and octahedra with shared oxygen atoms along the c axis occurs at TS ≈ 750 K, driven by the Jahn-Teller effect of the orbital active B-site Mn3+ cation. Strong magnetoelastic coupling is unveiled at TN1 ≈ 400 K as manifested by the appearance of Néel-type collinear ferrimagnetic order, an anomaly in both tetrahedral and octahedral distortions, as well as an anomalous decrease of the lattice constants c and a weak anomaly of a. Upon cooling to TN2 ≈ 65 K, it evolves to a noncollinear ferrimagnetic order accompanied by the different moments at the split magnetic sites B1 and B2. Only one-half of the B-site Mn3+/Fe3+ spins, i.e., the B2-site spins in the pyrochlore lattice, are canted, which is a unique magnetic order among spinels. The canting angle between A-site and B2-site moments is ∼25°, but the B1-site moment stays antiparallel to the A-site moment even at 10 K. This noncollinear order is accompanied by a modification of the O-B-O bond angles in the octahedra without significant change in lattice constants or tetrahedral/octahedral distortion parameters, indicating a distinct magnetoelastic coupling. We demonstrate distinct roles of the A-site and B-site magnetic cations in the structural and magnetic properties of FeMn2O4. Our study indicates that FeMn2O4 is a wonderful platform to unveil interesting magnetic order and to investigate their correlations with polyhedral distortions and lattice.

Dual-AAV split prime editor corrects the mutation and phenotype in mice with inherited retinal degeneration

The prime editor (PE) can edit genomes with almost any intended changes, including all 12 possible types of base substitutions, small insertions and deletions, and their combinations, without the requirement for double strand breaks or exogenous donor templates. PE demonstrates the possibility of correcting a variety of disease-causing mutations and might expand the therapeutic application of gene editing. In this study, PE was optimized based on a dual-adeno-associated virus (AAV) split-intein system in vitro by screening different split sites and split inteins. We found that splitting PE before amino acid 1105(Ser) of SpCas9 with Rma intein resulted in the highest on-target editing. The orientations of pegRNA and nicking sgRNA in the AAV vector were further optimized. To test the in vivo performance of the optimized dual-AAV split-PE3, it was delivered by subretinal injection in rd12 mice with inherited retinal disease Leber congenital amaurosis. The prime editors corrected the pathogenic mutation with up to 16% efficiency in a precise way, with no detectable off-target edits, restored RPE65 expression, rescued retinal and visual function, and preserved photoceptors. Our findings establish a framework for the preclinical development of PE and motivate further testing of PE for the treatment of inherited retinal diseases caused by various mutations.

Hydrous wadsleyite crystal structure up to 32 GPa

Abstract Hydroxylation of wadsleyite, β-(Mg,Fe)2SiO4, is associated with divalent cation defects and well known to affect its physical properties. However, an atomic-scale understanding of the defect structure and hydrogen bonding at high pressures is needed to interpret the influence of water on the behavior of wadsleyite in the mantle transition zone. We have determined the pressure evolution of the wadsleyite crystal symmetry and structure, including all O∙∙∙O interatomic distances, up to 32 GPa using single-crystal X-ray diffraction on two well-characterized, Fe-bearing (Fo90) samples containing 0.25(4) and 2.0(2) wt% H2O. Both compositions undergo a pressure-dependent monoclinic distortion from orthorhombic symmetry above 9 GPa, with the less hydrous sample showing a larger increase in distortion at increased pressures due to the difference in compressibility of the split M3 site in the monoclinic setting arising from preferred vacancy ordering at the M3B site. Although hydrogen positions cannot be modeled from the X-ray diffraction data, the pressure evolution of the longer O1∙∙∙O4 distance in the structure characterizes the primary hydrogen bond length. We observe the hydrogen-bonded O1∙∙∙O4 distance shorten gradually from 3.080(1) Å at ambient pressure to about 2.90(1) Å at 25 GPa, being still much longer than is defined as strong hydrogen bonding (2.5–2.7 Å). Above 25 GPa and up to the maximum pressure of the experiment at 32.5 GPa, the hydrogen-bonded O1∙∙∙O4 distance decreases no further, despite the fact that previous spectroscopic studies have shown that the primary O-H stretching frequencies continuously drop into the regime of strong hydrogen bonding (<3200 cm–1) above ~15 GPa. We propose that the primary O1-H∙∙∙O4 hydrogen bond in wadsleyite becomes highly nonlinear at high pressures based on its deviation from frequency-distance correlations for linear hydrogen bonds. One possible explanation is that the hydrogen position shifts from being nearly on the long O1-O4 edge of the M3 site to a position more above O1 along the c-axis.

Multiple Sclerosis: Enzymatic Cross Site-Specific Recognition and Hydrolysis of H3 Histone by IgGs against H3, H1, H2A, H2B, H4 Histones, Myelin Basic Protein, and DNA.

Histones have a specific key role in the remodeling of chromatin and gene transcription. In the blood, free histones are damage-connected proteins. Myelin basic protein (MBP) is the major component of the myelin-proteolipid sheath of axons. Antibodies possessing enzymatic activities (abzymes, ABZs) are the specific features of several autoimmune pathologies. IgGs against five histones, MBP, and DNA were obtained from the sera of multiple sclerosis (MS) patients using several affinity chromatographies. The sites of H3 histone splitting by Abs against five individual histones, MBP, and DNA were revealed by MALDI mass spectrometry. It was shown that the number of H3 splitting sites by IgGs against five various histones is different (number of sites): H3 (11), H1 (14), H2A (11), H4 (17), MBP (22), and DNA (29). IgGs against five different histones hydrolyze H3 at different sites, and only a few them coincide. The main reason for the enzymatic cross-reactivity of Abs against H3 and four other histones, as well as MBP, might be the high level of these proteins’ homology. The effective hydrolysis of the H3 histone at 29 sites with IgGs against DNA can be explained by the formation of chimeric abzymes against hybrid antigenic determinants formed by different histones and MBP at the junction of these protein sequences with DNA. The active centers of such abzymes contain structural elements of canonical DNases and proteases. Since free histones are pernicious proteins, antibodies–ABZs against five histones, MBP, and DNA could have a negative role in the pathogenesis of MS and probably other various autoimmune diseases.

Anomalous resistivity upturn in the van der Waals ferromagnet Fe5GeTe2

FenGeTe2 (n = 3, 4, and 5) has recently attracted increasing attention due to its two-dimensional van der Waals characteristic and high temperature ferromagnetism, which makes promises for spintronic devices. A Fe(1) split site is an important structural characteristic of Fe5GeTe2, which makes it very different from other FenGeTe2 (n = 3 and 4) systems. The local atomic disorder and short-range order can be induced by the split site. In this work, high-quality van der Waals ferromagnet Fe5GeTe2 single crystals were grown to study low-temperature transport properties. We found a resistivity upturn below 10 K. The temperature and magnetic field dependence of the resistivity are in good agreement with a combination of the theory of disorder-enhanced three-dimensional electron–electron and single-channel Kondo effect. The Kondo effect exists only at low magnetic fields B<3 T, while electron–electron interaction dominates the appearance for the low-temperature resistivity upturn. We believe that the enhanced three-dimensional electron–electron interaction in this system is induced by the local atomic structural disorder due to the split site of Fe(1). Our results indicate that the split site of Fe plays an important role for the exceptional transport properties.

Application of long-term passive acoustic monitoring to evaluate spatial and temporal patterns of multi-species acoustic assemblages of marine mammals in the western North Atlantic Ocean

Passive acoustic monitoring (PAM) is an efficient method for large-scale marine mammal monitoring. PAM technologies concurrently sample multiple soniferous marine mammal species, and when coupled with verified detectors, provide information that can be used to evaluate community composition. This analysis used data collected by an array of high-frequency PAM recorders at eight sites between 30°0’N and 42°0’N along the shelf-break in the western North Atlantic Ocean. Daily acoustic presence of 13 known marine mammal species and grouped unknown odontocetes was determined for all months between June 2016 and May 2019. Classification trees were generated from monthly summaries of daily detections to identify community composition dissimilarities. Detections of Gervais’s beaked whale (Mesoplodon europaeus) represented the root node and split sites by latitude, grouping the three sites south of 34°0’N and the five sites north of 38°0’N. Tree nodes were further divided by other odontocetes. The presence of mysticetes also varied by site and season. Distinctive communities were identified for each site, with odontocetes being resident and mysticetes more migratory. The root and secondary splits were all driven by beaked whale species, demonstrating the importance of identifying these whales to species instead of aggregating them as is common for visual survey data.

Structures of Ca5(VO4)3Cl and Ca4.78(1)Na0.22(PO4)3Cl0.78: positions of channel anions and repulsion on the anion in apatite-type compounds

Single crystal specimens of apatite-type compounds Ca5(VO4)3Cl and Ca4.78 (1)Na0.22(PO4)3Cl0.78 were prepared with a flux-growth technique and their structures were examined with single-crystal X-ray diffraction. The anion channel is defined by a face-sharing array of nearly regular Ca octahedra, which run along c, together with flat O3 trigonal antiprisms that are concentric with the Ca octahedra and highly oblate in [001]. The position of the channel anion in Ca5(VO4)3Cl is split into two at [0, 0, ±0.1691 (6)] with half occupancies. Dynamic disorder among these two positions is suggested from a saddle-shaped electron density distribution through the Ca regular triangle, i.e. the shared face of the Ca octahedra at z = ¼. The position of Cl− was too close to Ca2+ under bond-valence consideration. This is due to repulsion on Cl− from the flat O3 trigonal antiprism which is located at z = 0, namely, in between pairs of split Cl site positions. Ca4.78 (1)Na0.22(PO4)3Cl0.78 crystallizes as a disordered hexagonal structure in which the crystallographic pattern is not an intermediate state but a projection of two distinct halves of the monoclinic pattern [doubled in b; Mackie, Elliot & Young (1972). Acta Cryst. B28, 1840–1848] in one hexagonal cell. In spite of quite different environments, the bond-valence sums for Cl− in these structures are large and similar to each other. The repulsion between O2− and Cl− and a demand for keeping Ca2+ and Cl− apart are balanced at the positions where bond-valence sums for Cl− are around 1.2.