Publications by authors named "Kazuaki Tanaka"

63 Publications

Isolation and structural elucidation of novel fusicoccan dehydroxypericonicin A from Roussoella sp.

Biosci Biotechnol Biochem 2021 Jul;85(8):1798-1801

Faculty of Agriculture and Life Science, Hirosaki University, Hirosaki, Japan.

A potent antifungal fusicoccane dehydroxypericonicin A (1) was isolated from Roussoella sp. along with known allantofuranone (2). The relative structure of 1 was fully elucidated by NMR spectroscopic manner. The suggested relative structure was confirmed by density functional theory (DFT)-based 13C NMR chemical shift calculations. The absolute configuration was investigated by a spectral comparison of the experimental electronic circular dichroism spectrum with that based on the DFT calculations.
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http://dx.doi.org/10.1093/bbb/zbab103DOI Listing
July 2021

L-NAME, a nitric oxide synthase inhibitor, increases the protein expression of both executioner and inhibitor of apoptosis in the placental bed of mid-to-late pregnant rats.

Congenit Anom (Kyoto) 2021 Sep 27;61(5):183-187. Epub 2021 Apr 27.

Graduate School of Veterinary Medicine, Azabu University, Sagamihara, Japan.

The involvement of nitric oxide (NO) signaling in apoptosis was examined in the placental bed of mid-to-late pregnant rats. Pregnant rats were treated with l-NAME, a nitric oxide synthase inhibitor, by subcutaneous infusion for 48 hours before the examination at day 13.5, 17.5, or 21.5. l-NAME induced apoptosis in the placental bed to a limited extent at days 13.5 and 17.5, but not at day 21.5. When the placental bed was examined at day 17.5, the protein expression of both executioner (C-Cas3) and inhibitor (XIAP) of apoptosis was increased by l-NAME, but they did not co-localized with apoptosis. It was presumed that placental bed apoptosis induced by l-NAME is regulated through the expression of both executioner and inhibitor, possibly involving protein S-nitrosylation.
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http://dx.doi.org/10.1111/cga.12420DOI Listing
September 2021

Cyclohelminthol CPs: Scope and Limitations of Density Functional Theory-Based Structural Elucidation of Natural Products.

J Org Chem 2021 01 7;86(2):1505-1515. Epub 2021 Jan 7.

Faculty of Agriculture and Bioscience, Hirosaki University, 3-Bunkyo-cho, Hirosaki 036-8561, Japan.

The effectiveness and limitations of density functional theory (DFT) calculations in the structural determination of complexed and conformationally flexible natural products were demonstrated using the cyclohelminthols CP-1 () CP-2 (), CP-3 (), and CP-4 () newly isolated from yone96. Prior to DFT calculations, the structures were tentatively assigned using conventional spectroscopic analyses. The structures were verified with reference to DFT-derived C and H NMR chemical shifts, and values, and electronic circular dichroism (ECD) spectra. The C chemical shift calculations were very effective for verifying the ring-structure moieties but less effective for verifying the geometry of the side chain in which the juncture asymmetric carbon (C-16) was apart from the ring-structure moiety. However, H chemical shift calculations compensated for the imperfection of the latter. ECD spectral calculations were used to determine the absolute configurations. Calculations for virtual simple model molecules enabled us to evaluate the reliability of the ECD spectral calculation and derive the chiral torsion responsible for the characteristic Cotton effects.
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http://dx.doi.org/10.1021/acs.joc.0c02378DOI Listing
January 2021

Valproic acid up-regulates the whole NO-citrulline cycle for potent iNOS-NO signaling to promote neuronal differentiation of adipose tissue-derived stem cells.

Nitric Oxide 2021 01 28;106:35-44. Epub 2020 Oct 28.

Graduate School of Veterinary Medicine, Azabu University, Fuchinobe, Chuo-ku, Sagamihara, 252-5201, Japan. Electronic address:

Valproic acid (VPA) remarkably promotes the differentiation of adipose tissue-derived stem cells (ASCs) to mature neuronal cells through nitric oxide (NO) signaling due to up-regulated inducible NO synthase (iNOS) as early as within 3 days. Here, we investigated mechanisms of VPA-promoted neuronal differentiation of ASCs concerning the NO-citrulline cycle, the metabolic cycle producing NO. Cultured rat ASCs were differentiated to mature neuronal cells rich in dendrites and expressing a neuronal marker by treatments with VPA at 2 mM for 3 days and subsequently with the neuronal induction medium for 2 h. Inhibitor (α-methyl-d, l-aspartic acid, MDLA) of arginosuccinate synthase (ASS), a key enzyme of the NO-citrulline cycle, abolishes intracellular NO increase and VPA-promoted neuronal differentiation in ASCs. l-Arginine, the substrate of iNOS, restores the promotion effect of VPA, being against MDLA. Immunocytochemistry showed that ASS and iNOS were increased in ASCs expressing neurofilament medium polypeptide (NeFM), a neuronal marker, by VPA and NIM synergistically. Real-time RT-PCR analysis showed that mRNAs of Ass and arginosuccinate lyase (Asl) in the NO-citrulline cycle were increased by VPA. Chromatin immunoprecipitation assay indicated that Ass and Asl were up-regulated by VPA through the acetylation of their associated histone. From these results, it was considered that VPA up-regulated the whole NO-citrulline cycle, which enabled continuous NO production by iNOS in large amounts for potent iNOS-NO signaling to promote neuronal differentiation of ASCs. This may also indicate a mechanism enabling short-lived NO to function conveniently as a potent signaling molecule that can disappear quickly after its role.
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http://dx.doi.org/10.1016/j.niox.2020.10.006DOI Listing
January 2021

Genetic Variation in Y-Chromosome Genes of Sika Deer () in Japan.

Zoolog Sci 2020 Oct;37(5):411-416

School of Veterinary Medicine, Azabu University, Chuo-Ku, Sagamihara 252-5201, Japan.

Sika deer () in Japan are classified into southern and northern groups. However, previous studies primarily relied on maternally inherited mitochondrial DNA (mtDNA). The paternally inherited Y-chromosome is useful for analyzing the contribution of males to the population genetic history of sika deer. In total, approximately 16 kb of partial sequences of four Y-chromosomal genes, Y-linked, sex-determining region Y, DEAD-box helicase 3 Y-linked, and Zinc finger protein Y-linked, were sequenced to investigate intraspecific variation. As a result, we identified nine intronic single nucleotide polymorphisms (SNPs) in 478 sika deer samples collected over the entire Japanese archipelago from Hokkaido to Kyushu. SNP genotyping revealed 10 distinct haplotypes (SYH1-SYH10). The most common haplotype (SYH1) was present in all populations and was the most abundant haplotype, identified in 80.3% of the sampled individuals. The remaining haplotypes were unique to a single locality. SYH1 was also central to all other haplotypes that diverged by a SNP, resulting in this haplotype being the core of a star-like cluster topography. We found that contrary to mtDNA patterns, there was no clear differentiation of Y-chromosome markers between the southern and the northern populations. Due to the female philopatry of sika deer, mtDNA may provide a highly structured differentiation of populations. On the other hand, the male-biased gene flow may provide a reduced differentiation of populations. Our findings revealed that the genetic structure of the Japanese sika deer is more complex than previously thought based on mtDNA-based phylogeographic studies.
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http://dx.doi.org/10.2108/zs200043DOI Listing
October 2020

Association analysis of non-synonymous polymorphisms of interleukin-4 receptor-α and interleukin-13 genes in canine atopic dermatitis.

J Vet Med Sci 2020 Sep 15;82(9):1253-1259. Epub 2020 Jul 15.

School of Veterinary Medicine, Azabu University, 1-17-71, Fuchinobe, Chuo-ku, Sagamihara, Kanagawa 252-5201, Japan.

Interleukin-4 (IL4) and interleukin-13 (IL13) are involved in the initial response of T helper 2 lymphocytes through the activation of the IL4 receptor alpha (IL4RA), which is a common receptor chain for these cytokines. In humans, several single-nucleotide polymorphisms (SNPs) identified in the IL4R and in interleukin coding genes were associated with atopic disorders. However, the association between canine IL4R polymorphisms and atopic disorders has not been investigated yet. This study aimed to determine the associations between four non-synonymous SNPs and canine atopic dermatitis (CAD) in shiba inu and miniature dachshund populations. Polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP) analysis were used to genotype four polymorphisms of canine IL4R and IL13 in 34 shiba inu and 19 miniature dachshund patients with CAD, as well as 29 shiba inu and 39 miniature dachshund patients without the condition. Results from miniature dachshunds revealed a potential association between the presence of minor A allele rs24378020 and CAD (odds ratio, 0.10; 95% confidence interval, 0.01-0.85; P=0.0062). This CAD resistance allele led to an amino acid substitution (Arg688Cys) that could impair IL4 and IL13 signaling. In shiba inu patients, rs24378020 was fixed by homozygosity of the major G allele. No association was found between the remaining three evaluated SNPs and CAD. Nevertheless, the study suggests that the IL4R Cys688 variant reduces the risk of CAD in miniature dachshunds.
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http://dx.doi.org/10.1292/jvms.20-0301DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7538314PMC
September 2020

Valproic acid promotes mature neuronal differentiation of adipose tissue-derived stem cells through iNOS-NO-sGC signaling pathway.

Nitric Oxide 2019 12 1;93:1-5. Epub 2019 Sep 1.

Graduate School of Veterinary Medicine, Azabu University, Fuchinobe, Chuo-ku, Sagamihara, 252-5201, Japan. Electronic address:

Valproic acid (VPA) remarkably promotes the differentiation of adipose tissue-derived stem cells (ASCs) to mature neuronal cells, enabling neuronal induction within only three days. Here, we investigated the involvement of NO-signaling in the VPA-promoted neuronal differentiation of ASCs as a possible mechanism. Cultured rat ASCs were differentiated to matured neuronal cells rich in dendrites and expressing βIII-tubulin protein, a neuronal marker, by treatments with VPA at 2 mM for 3 days and subsequently with the neuronal induction medium (NIM) containing cAMP-elevating agents for 2 h. Increased intracellular NO was detected in neuronal cells differentiated from ASCs treated with VPA by a fluorescence NO-specific probe, diaminofluorescein-FM diacetate. However, a NO donor (NOC18) increased the incidence of neuronal cells only to a lesser extent than VPA, indicating the insufficiency of exogenous NO. RT-PCR analysis of ASCs treated with VPA showed increased mRNA expression of inducible nitric oxide synthase (iNOS) with the acetylation of its associated histone H3K9. iNOS inhibitors (1400 W and dexamethasone) or a soluble guanylate cyclase (sGC) inhibitor (ODQ) decreased the incidence of neuronal cells differentiated from ASCs treated with VPA. These inhibitors also decreased the mRNA expression of mature neuronal markers, neurofilament medium polypeptide (NeFM) and microtubule-associated protein 2 (MAP2), as well as βIII-tubulin (TUBB3), to various extents. It was considered from these results that VPA promoted mature neuronal differentiation of ASCs through the iNOS-NO-sGC signaling pathway. This provided insights into the regulated neuronal differentiation of ASCs in clinical applications.
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http://dx.doi.org/10.1016/j.niox.2019.08.008DOI Listing
December 2019

Negative air pressure treatment accelerates the penetration of permeable cryoprotectants into bovine ovarian tissue in vitrification protocol and improves cell density after vitrification.

Cryobiology 2019 06 9;88:92-97. Epub 2019 Mar 9.

Graduate School of Veterinary Medicine, Azabu University, Sagamihara, Kanagawa, 252-5201, Japan. Electronic address:

Effects of additional physical treatments during vitrification of the bovine ovarian tissue were examined for increasing of permeability of ethylene glycol (EG) and dimethyl sulfoxide (Me2SO). The concentrations of EG and Me2SO and histological changes in the ovarian tissue were evaluated. In the first equilibration step (7.5% EG and 7.5% Me2SO), all the 10-min physical treatments, i.e., negative (679 hPa) or positive (1347 hPa) air pressure applied with a disposable syringe, and shaking (60 rpm) applied with a laboratory shaker, were comparable to 25-min non-physical treatment (plain) vitrification. When effects of the negative air pressure were examined in the second equilibration step (20% EG and 20% Me2SO), its 10-min treatment was equivalent to 15-min plain vitrification (140-170 mg/g tissue). It was thus indicated that the negative air pressure treatment accelerates the penetration of permeable cryoprotectants into the ovarian tissue slices. Histological examination showed that the cell density and the amount of pan-cadherin in the tunica albuginea of the ovary was reduced by the vitrification, but was improved by the negative air pressure treatment. The amount of pan-cadherin in the tunica albuginea was recommended as a biomarker for evaluation of effectiveness of protocol for cryopreservation of bovine ovarian tissue and considered to be a candidate biomarker for human ovarian tissue cryopreservation.
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http://dx.doi.org/10.1016/j.cryobiol.2019.03.001DOI Listing
June 2019

Isolation of Peribysins O, P, and Q from Periconia macrospinosa KT3863 and Configurational Reinvestigation of Peribysin E Diacetate from Periconia byssoides OUPS-N133.

J Nat Prod 2019 04 15;82(4):911-918. Epub 2019 Feb 15.

Faculty of Agriculture and Life Science , Hirosaki University , 3-Bunkyo-cho , Hirosaki , 036-8561 , Japan.

Peribysins O (1), P (3), and Q (4) were isolated from Periconia macrospinosa KT3863. The relative configuration of the 6,7-epoxide of 1 was elucidated by performing quantitative NOE experiments. The structure of 2, which is a tautomer of 1 present in CDCl solutions in 5% abundance, was also fully characterized by NMR analysis. Their absolute configurations were independently determined by the modified Mosher's method (for 1 and 3), the electronic circular dichroism (ECD) exciton coupling theory after conversion into dibenzoate 9 (for 3), and theoretical ECD calculations (for 1, 3, and 4). The obtained relative structures 1, 3, and 4 were verified by calculating their C chemical shifts using density functional theory (DFT). Although the established (4 S)-enantiomer for 1-4 is in accordance with that of other peribysins isolated from the related fungus Periconia byssoides OUPS-N133, Danishefsky's total synthesis of peribysin E (5) led to the subsequent revision of the (2 R,4 S,5 R,6 S,7 S,8 R,10 S)-enantiomer to the (2 S,4 R,5 S,6 R,7 R,8 S,10 R)-enantiomer. This discordance led us to reinvestigate the configuration using time-dependent DFT-based ECD spectral calculations, which supported the original (4 S)-enantiomer.
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http://dx.doi.org/10.1021/acs.jnatprod.8b01001DOI Listing
April 2019

Identification and antifungal sensitivity of two new species of Diaporthe isolated.

J Infect Chemother 2019 Feb 10;25(2):96-103. Epub 2018 Nov 10.

Faculty of Agriculture and Life Science, Hirosaki University, Aomori, Japan.

Diaporhte species are plant pathogens rarely involved in human diseases, especially eye diseases. We report our findings in two undescribed Diaporhte species. Both were identified by their morphological characteristics and by DNA sequence analyses. In Case 1, an 81-year-old male farmer who had pterygium surgery 7 years earlier developed keratitis and the causal fungus was identified as a new species of Diaporthe, D. oculi. This species can be distinguished from the closely related D. limonicola on Citrus limon (Rutaceae) by the ITS, tef1, and TUB (515/520 = 99.0% in ITS, 315/324 = 97.2% in tef1, and 601/614 = 97.9% in TUB). The isolate from Case 2, a 68-year-old man with a rose thorn injury, was also identified as a new Diaporthe species, D. pseudooculi. Phylogenetically, D. pseudooculi is different from the closely related D. podocarpi-macrophylli by the ITS, tef1, and TUB (525/531 = 98.9% in ITS, 314/333 = 94.3% in tef1, and 436/442 = 98.6% in TUB). We report on the identification, drug sensitivity, and treatment outcomes for these two new species of Diaporthe, D. oculi and D. pseudooculi.
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http://dx.doi.org/10.1016/j.jiac.2018.10.008DOI Listing
February 2019

Cyclohelminthols Y1-Y4 Metabolites Possessing Two Spirocyclopropanes in Their Structure.

J Org Chem 2018 05 10;83(10):5688-5697. Epub 2018 May 10.

Faculty of Agriculture and Life Science , Hirosaki University , 3-Bunkyo-cho , Hirosaki 036-8561 , Japan.

Cyclohelminthols Y1-Y4 (1-4) were isolated from the culture broth of Helminthosporium velutinum yone96. These compounds are diastereomers to each other featuring 3-azabicyclo[3.1.0]hexane-6-spirocyclopentane linked with a cyclopentanespirocyclopropane framework. Their planar structures were established via the comparison of their spectra with the simpler analogue cyclohelminthol X as well as analysis of their HMBC spectra. Although the proton-deficient core frameworks of 1-4 prevented us from obtaining configurational information via conventional NMR analysis, their total structures involving the relative and absolute configurations were established using density functional theory (DFT)-based molecular modeling calculations. The present study demonstrates the effectiveness of the comparison between the theoretical and experimental δC values for stereochemical analysis by focusing on the carbons that show relatively large δC deviations among the isomers. The G-ring of these molecules most likely originates from the cyclopropanation of the C6C7 double bond with the carbene equivalent 6 derived from cyclohelminthol IV (7), which was isolated from the same producer fungus. Preliminary biological experiments revealed the potent cytotoxicity of the (6 S)-isomers against COLO201 cells, whereas the (6 R)-isomers exhibited weak activity. The antifungal assay with Cochiobolus miyabeanus showed a slightly different profile.
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http://dx.doi.org/10.1021/acs.joc.8b00727DOI Listing
May 2018

Estimation of Sobolev embedding constant on a domain dividable into bounded convex domains.

J Inequal Appl 2017 29;2017(1):299. Epub 2017 Nov 29.

Faculty of Science and Engineering, Waseda University, 3-4-1, Okubo Shinjyuku-ku, Tokyo, 169-8555 Japan.

This paper is concerned with an explicit value of the embedding constant from [Formula: see text] to [Formula: see text] for a domain [Formula: see text] ([Formula: see text]), where [Formula: see text]. We previously proposed a formula for estimating the embedding constant on bounded and unbounded Lipschitz domains by estimating the norm of Stein's extension operator. Although this formula can be applied to a domain Ω that can be divided into a finite number of Lipschitz domains, there was room for improvement in terms of accuracy. In this paper, we report that the accuracy of the embedding constant is significantly improved by restricting Ω to a domain dividable into bounded convex domains.
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http://dx.doi.org/10.1186/s13660-017-1571-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5707241PMC
November 2017

Brevicollum, a new genus in Neohendersoniaceae, Pleosporales.

Mycologia 2017 Jul-Aug;109(4):608-619. Epub 2017 Nov 17.

d Genetic Resources Center , National Agriculture and Food Research Organization, 2-1-2 Kannondai, Tskuba, Ibaraki 305-8602, Japan.

A new genus, Brevicollum, is established for two new species, B. hyalosporum on Syzygium samarangense and B. versicolor on Volkameria inermis, in Japan. This genus is characterized by the presence of immersed ascomata with an excentric to central, short cylindrical ostiolar neck, thin ascomatal wall, clavate asci with a shallow ocular chamber, and broadly fusiform, hyaline to pale brown ascospores with a sheath. Molecular phylogenetic analyses based on a combined data set of sequences of 18S and 28S nuc rDNA genes and the second largest subunit of RNA polymerase II (RPB2) revealed that Brevicollum is a member of Neohendersoniaceae (Pleosporales, Dothideomycetes). The genera Brevicollum, Crassiparies, Medicopsis, and Neohendersonia are accepted in Neohendersoniaceae. Revised descriptions of Crassiparies and its type species, C. quadrisporus, are provided based on the holotype and a newly obtained specimen and isolate. An asexual morph of C. quadrisporus is reported for the first time. The nuc rDNA internal transcribed spacer (ITS1-5.8S-ITS2 = ITS) phylogenetic tree of the newly collected species (B. hyalosporum, B. versicolor, and C. quadrisporus), and the unnamed or provisionally identified endophytic species, suggests a broader distribution of Brevicollum and Crassiparies species.
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http://dx.doi.org/10.1080/00275514.2017.1387432DOI Listing
November 2018

Cyclohelminthol X, a Hexa-Substituted Spirocyclopropane from Helminthosporium velutinum yone96: Structural Elucidation, Electronic Circular Dichroism Analysis, and Biological Properties.

J Org Chem 2017 06 16;82(11):5574-5582. Epub 2017 May 16.

Faculty of Agriculture and Life Science, Hirosaki University , 3-Bunkyo-cho, Hirosaki 036-8561, Japan.

Helminthosporium velutinum yone96 produces cyclohelminthol X (1), a unique hexa-substituted spirocyclopropane. Although its molecular formula and NMR spectral data resemble those of AD0157, being isolated from marine fungus Paraconiothyrium sp. HL-78-gCHSP3-B005, our detailed analyses disclosed a totally different structure. Chemical shift calculations and electronic circular dichroism spectral calculations were quite helpful to establish the structure, when those were performed based on density functional theory. The carbon framework of cyclohelminthols I-IV is found at the C1-C8 propenylcyclopentene substructure of 1. Thus, 1 is assumed to be biosynthesized by cyclopropanation between an oxidized form of cyclohelminthol IV and a succinic anhydride derivative 4. Cytotoxicity for two cancer cell lines and proteasome inhibition efficiency are measured.
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http://dx.doi.org/10.1021/acs.joc.7b00393DOI Listing
June 2017

A convenient technique for live-cell observation on the surface of polytetrafluoroethylene with a phase-contrast microscope.

Microscopy (Oxf) 2017 Apr;66(2):136-142

Graduate School of Veterinary Medicine, Azabu University, 1-17-71 Fuchinobe, Chuo-ku, Sagamihara, Kanagawa 252-5201, Japan.

Phase-contrast microscopy is a convenient technique for live-cell observation on the surface of materials with high optical transmittance. Here, we demonstrate a novel technique to observe living cells on the surface of materials with low optical transmittance, such as polytetrafluoroethylene (PTFE), which are widely used in biomaterials for blood-contacting devices. The surface of a cover glass was coated with a thin PTFE layer with sufficient transmittance, thereby enabling the observation of living cells on the PTFE surface with a phase-contrast microscope.
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http://dx.doi.org/10.1093/jmicro/dfw042DOI Listing
April 2017

Roussoella solani causing keratomycosis, with an observed both sexual and asexual morphs.

J Infect Chemother 2017 Sep 25;23(9):651-654. Epub 2017 Apr 25.

Faculty of Agriculture and Life Science, Hirosaki University, Aomori, Japan.

We describe an 82-year-old male farmer who had diabetes mellitus with no history of ocular trauma by soil or plants and who developed a corneal infection due to a fungus. The organism was identified as Roussoella solani based on both the morphological characteristics and phylogenetic analysis using LSU and ITS nrDNA sequences. The sexual stage of R. solani is described and illustrated for the first time. The patient was treated successfully with a combination of topical and systemic voriconazole and micafungin. This case is the first report of keratomycosis caused by R. solani.
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http://dx.doi.org/10.1016/j.jiac.2017.03.005DOI Listing
September 2017

Noninvasive sample collection for the genotyping of neonatal rats using adhesive tape.

J Vet Med Sci 2017 May 27;79(5):827-829. Epub 2017 Mar 27.

Laboratory of Animal Biotechnology, School of Veterinary Medicine, Azabu University, 1-17-71 Fuchinobe, Chuo-ku, Sagamihara, Kanagawa 252-5201, Japan.

To develop a noninvasive sample collection method for genotyping, we compared PCR products from samples collected from neonates using five different brands of adhesive tape. Next, the youngest application age to distinguish genotypes was established. The tapes were applied on the backs of rats on postnatal day (PND) 10. DNA extracts from two brands provided clear PCR products that enabled genotype identification. The youngest age for distinguishing genotypes was PND 5; however, the youngest age that provided accurate results was PND 7. Thus, the present method allows for genotyping during the neonatal period without invasive burden and may improve animal welfare by refining.
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http://dx.doi.org/10.1292/jvms.16-0645DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5447967PMC
May 2017

Phylogenetic Analysis of the Synnema-Producing Genus Synnemapestaloides.

J Fungi (Basel) 2016 Nov 7;2(4). Epub 2016 Nov 7.

Graduate School of Agriculture, Tamagawa University, 6-1-1 Tamagawa-gakuen, Machida, Tokyo 194-8610, Japan.

, the type species of the genus , is a pathogen of . This fungus produces six-celled conidia with appendages at both end cells, and are generated by annellidic conidiogenous cells on the synnema. These conidial structures are similar to those of the genus . The monotypic genus is currently classified in the family Amphisphaeriaceae solely based on conidial morphology. Here we demonstrate that represents a distinct genus in the family Sporocadaceae (Amphisphaeriales) based on differences in the nucleotide sequences of the partial large subunit rDNA gene, the rDNA internal transcribed spacer, and the partial The genus most closely related to is and the species most similar to is which produces short synnema-like conidiomata (sporodochia). These results demonstrate that should be transferred to , and also demonstrate that Sporocadaceae can have synnematal in addition to pycnidial and acervular conidiomata.
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http://dx.doi.org/10.3390/jof2040028DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5715930PMC
November 2016

Differentiation of rat adipose tissue-derived stem cells into neuron-like cells by valproic acid, a histone deacetylase inhibitor.

Exp Anim 2016 25;65(1):45-51. Epub 2015 Sep 25.

Graduate School of Veterinary Medicine, Azabu University, Fuchinobe, Chuo-ku, Sagamihara, Kanagawa 252-5201, Japan.

Valproic acid (VPA) is a widely used antiepileptic drug, which has recently been reported to modulate the neuronal differentiation of adipose tissue-derived stem cells (ASCs) in humans and dogs. However, controversy exists as to whether VPA really acts as an inducer of neuronal differentiation of ASCs. The present study aimed to elucidate the effect of VPA in neuronal differentiation of rat ASCs. One or three days of pretreatment with VPA (2 mM) followed by neuronal induction enhanced the ratio of immature neuron marker βIII-tubulin-positive cells in a time-dependent manner, where the majority of cells also had a positive signal for neurofilament medium polypeptide (NEFM), a mature neuron marker. RT-PCR analysis revealed increases in the mRNA expression of microtubule-associated protein 2 (MAP2) and NEFM mature neuron markers, even without neuronal induction. Three-days pretreatment of VPA increased acetylation of histone H3 of ASCs as revealed by immunofluorescence staining. Chromatin immunoprecipitation assay also showed that the status of histone acetylation at H3K9 correlated with the gene expression of TUBB3 in ASCs by VPA. These results indicate that VPA significantly promotes the differentiation of rat ASCs into neuron-like cells through acetylation of histone H3, which suggests that VPA may serve as a useful tool for producing transplantable cells for future applications in clinical treatments.
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http://dx.doi.org/10.1538/expanim.15-0038DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4783650PMC
November 2016

Minutisphaerales (Dothideomycetes, Ascomycota): a new order of freshwater ascomycetes including a new family, Minutisphaeraceae, and two new species from North Carolina, USA.

Mycologia 2015 Jul-Aug;107(4):845-62

Las Muros, 09420 Rimont, France.

Minutisphaera is a recently established genus of freshwater Dothideomycetes characterized by small, globose to subglobose or apothecioid, erumpent to superficial, brown ascomata; fissitunicate, eight-spored, ovoid to obclavate asci; and 1-2-septate, clavate to broadly fusiform, hyaline to pale brown ascospores with or without a gelatinous sheath and filamentous appendages. The genus currently contains two species: M. fimbriatispora, the type species, and M. japonica. The higher-level phylogenetic relationship of Minutisphaera within the Dothideomycetes currently is unresolved. To establish the phylogenetic position of Minutisphaera within the Dothideomycetes and evaluate the phylogenetic affinities of newly collected Minutisphaera-like taxa, we sequenced three rDNA regions-18S, ITS1-5.8SITS2 (ITS) and 28S nuc rDNA, and a protein-coding gene, MCM7, for newly collected strains of Minutisphaera. Based on maximum likelihood and Bayesian analyses of a combined dataset (18S and 28S) composed of 167 taxa, a more refined dataset (28S and MCM7) comprising 52 taxa and a separate ITS dataset, and an examination of morphology, we describe and illustrate two new species of Minutisphaera. The Minutisphaera clade was strongly supported within the Dothideomycetes with likelihood and Bayesian statistics but did not share phylogenetic affinities with any existing taxonomic group within the Dothideomycetes. We therefore establish a new order, Minutisphaerales, and new family, Minutisphaeraceae, for this monophyletic clade of freshwater ascomycetes. Chemical analysis of the organic extract M. aspera (G427) resulted in isolation and characterization of five known secondary metabolites, of which four were dipeptides (1-4) and one an aromatic polyketide (5). Conversely, two aromatic polyketides (5, 6) were isolated and identified from the organic extract of M. parafimbriatispora (G156-4). The isolated compounds were tested for their antimicrobial activity against an array of bacteria and fungi. Compound 6 showed promising activity against Staphylococcus aureus and Mycobacterium smegmatis with minimal inhibitory concentration values of 30 and 60 μg/mL, respectively.
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http://dx.doi.org/10.3852/15-013DOI Listing
September 2015

Computation-Assisted Structural Elucidation of Epoxyroussoeone and Epoxyroussoedione Isolated from Roussoella japanensis KT1651.

J Nat Prod 2015 Jul 25;78(7):1505-10. Epub 2015 Jun 25.

†Faculty of Agriculture and Bioscience, Hirosaki University, 3-Bunkyo-cho, Hirosaki, Aomori 036-8561, Japan.

The structures of epoxyroussoenone (1) and epoxyroussoedione (3) isolated from a culture broth of Roussoella japanensis KT1651 were determined. Although NMR spectra provided insufficient structural information, computation of the theoretical chemical shifts with DFT EDF2/6-31G* enabled us to elucidate not only the planar structure, but also the relative configuration. Their ECD (electric circular dichroism) spectra suggested the absolute configurations, which were confirmed with time-dependent DFT calculations employing BHandHLYP/TZVP. The ECD calculations for other stereoisomers yielded obviously different spectral profiles, thus confirming the relative structures of 1 and 3.
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http://dx.doi.org/10.1021/np500924nDOI Listing
July 2015

Taxonomic revision of Pseudolachnea and Pseudolachnella and establishment of Neopseudolachnella and Pseudodinemasporium gen. nov.

Mycologia 2015 Mar-Apr;107(2):383-408. Epub 2015 Jan 8.

Faculty of Agriculture and Life Science, Hirosaki University, 3 Bunkyo-cho, Hirosaki, Aomori 036-8561, Japan

The taxonomy of Pseudolachnea and Pseudolachnella is controversial. Some authors have regarded them as congeneric, whereas others have considered them to be distinct genera differentiated merely on the number of conidial septa. A total of 26 isolates of Pseudolachnea-like fungi were subjected to morphological examination and phylogenetic analyses of nuc rDNA internal transcribed spacers 1 and 2 and partial 28S sequences and partial sequence of the translation elongation factor 1α gene. The results indicated that our materials should be classified in four genera: Pseudolachnea, Pseudolachnella, and two new genera, Neopseudolachnella and Pseudodinemasporium. Although the monophyly of both Pseudolachnea and Pseudolachnella was confirmed, it was concluded that differences observed in the conidiomatal structure, such as thickness of basal stroma and the excipulum, were more reliable for their circumscription, instead of conidial septation. Neopseudolachnella was similar to Pseudolachnea and Pseudolachnella in conidial morphology but was characterized by the conidiomata lacking an excipulum, unlike members of the latter two genera. Pseudodinemasporium bore conidia morphologically similar to those of Dinemasporium but was differentiated from the latter by the conidiomata, which was composed of a well developed peridial structure. A total of 12 new species, namely three in Neopseudolachnella (N. acutispora, N. magnispora, N. uniseptata), one in Pseudodinemasporium (P. fabiforme) and eight in Pseudolachnella (P. asymmetrica, P. botulispora, P. brevicoronata, P. campylospora, P. complanata, P. falcatispora, P. fusiformis and P. pachyderma) are described and illustrated.
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http://dx.doi.org/10.3852/14-171DOI Listing
August 2015

Naming and outline of -2014 including proposals for the protection or suppression of generic names.

Fungal Divers 2014 Nov 4;69(1):1-55. Epub 2014 Nov 4.

ABL Herbarium, G.v.d.Veenstraat 107, NL-3762 XK Soest, The Netherlands.

Article 59.1, of the International Code of Nomenclature for Algae, Fungi, and Plants (ICN; Melbourne Code), which addresses the nomenclature of pleomorphic fungi, became effective from 30 July 2011. Since that date, each fungal species can have one nomenclaturally correct name in a particular classification. All other previously used names for this species will be considered as synonyms. The older generic epithet takes priority over the younger name. Any widely used younger names proposed for use, must comply with Art. 57.2 and their usage should be approved by the Nomenclature Committee for Fungi (NCF). In this paper, we list all genera currently accepted by us in (belonging to 23 orders and 110 families), including pleomorphic and nonpleomorphic genera. In the case of pleomorphic genera, we follow the rulings of the current ICN and propose single generic names for future usage. The taxonomic placements of 1261 genera are listed as an outline. Protected names and suppressed names for 34 pleomorphic genera are listed separately. Notes and justifications are provided for possible proposed names after the list of genera. Notes are also provided on recent advances in our understanding of asexual and sexual morph linkages in . A phylogenetic tree based on four gene analyses supported 23 orders and 75 families, while 35 families still lack molecular data.
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http://dx.doi.org/10.1007/s13225-014-0309-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4896388PMC
November 2014

Finding needles in haystacks: linking scientific names, reference specimens and molecular data for Fungi.

Database (Oxford) 2014 30;2014. Epub 2014 Jun 30.

National Center for Biotechnology Information, National Library of Medicine, National Institutes of Health, Bethesda, Maryland, USA, CBS-KNAW Fungal Biodiversity Centre, P.O. Box 85167, 3508 AD Utrecht, The Netherlands, Department of Pharmaceutical Sciences - Microbiology, Università degli Studi di Perugia, Perugia, Italy, Molecular Mycology Research Laboratory, Centre for Infectious Diseases and Microbiology, Marie Bashir Institute for Infectious Diseases and Biosecurity, Sydney Medical School-Westmead Hospital, The University of Sydney, Westmead Millennium Institute, Westmead, Australia, Department of Biological and Environmental Sciences, University of Gothenburg, Box 461, 405 30 Göteborg, Sweden, Ecology and Evolutionary Biology, University of Tennessee, Knoxville, TN 37920, USA, Illinois Natural History Survey, University of Illinois, 1816 South Oak Street, Champaign, IL 61820, USA, Mycology Section, Jodrell Laboratory, Royal Botanic Gardens, Kew, Richmond, Surrey, TW9 3DS, UK, Natural History Museum, University of Tartu, 46 Vanemuise, 51014 Tartu, Estonia, Purdue University, Department of Botany and Plant Pathology, 915 W. State Street, West Lafayette, IN 47907, USA, Institute of Excellence in Fungal Research, and School of Science, Mae Fah Luang University, Chiang Rai 57100, Thailand, Imperial College London, Royal Botanic Gardens, Kew TW9 3DS, England, UK, Muséum National d'Histoire Naturelle, Dépt. Systématique et Evolution CP39, UMR7205, 12 Rue Buffon, F-75005 Paris, France, Kunming Institute of Botany, Chinese Academy of Sciences, Kunming 650201, Yunnan, P. R. China, Departamento de Biología Vegetal II, Facultad de Farmacia, Universidad Complutense de Madrid, Madrid 28040, Spain, Senckenberg Museum of Natural History Görlitz, PF 300 154, 02806 Görlitz, Germany, Department of Microbiology and Plant Pathology, Forestry Agricultural Biotechnology Institute (FABI), University of Pretoria, Pretoria 0001, South Africa, Real Jardín Botánico, RJB-CSIC,

DNA phylogenetic comparisons have shown that morphology-based species recognition often underestimates fungal diversity. Therefore, the need for accurate DNA sequence data, tied to both correct taxonomic names and clearly annotated specimen data, has never been greater. Furthermore, the growing number of molecular ecology and microbiome projects using high-throughput sequencing require fast and effective methods for en masse species assignments. In this article, we focus on selecting and re-annotating a set of marker reference sequences that represent each currently accepted order of Fungi. The particular focus is on sequences from the internal transcribed spacer region in the nuclear ribosomal cistron, derived from type specimens and/or ex-type cultures. Re-annotated and verified sequences were deposited in a curated public database at the National Center for Biotechnology Information (NCBI), namely the RefSeq Targeted Loci (RTL) database, and will be visible during routine sequence similarity searches with NR_prefixed accession numbers. A set of standards and protocols is proposed to improve the data quality of new sequences, and we suggest how type and other reference sequences can be used to improve identification of Fungi. Database URL: http://www.ncbi.nlm.nih.gov/bioproject/PRJNA177353.
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http://dx.doi.org/10.1093/database/bau061DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4075928PMC
February 2015

Lambertellin system, the mechanism for fungal replacement of Monilinia fructigena with Lambertella corni-maris without competitive inhibition on agar media.

Bioorg Med Chem 2014 Apr 5;22(8):2489-95. Epub 2014 Mar 5.

Faculty of Agriculture and Life Science, Hirosaki University, 3-Bunkyo-cho, Hirosaki 036-8561, Japan. Electronic address:

The 'Lambertellin system' was disclosed which rationally explains the fungal replacement (mycoparasitism) of Monilinia fructigena (M. f., the host) with Lambertella corni-maris (L. corni-maris, the parasite) without competitive inhibition in the simultaneous incubations on agar media. The 'Lambertellin system' involves; (a) L. corni-maris secretes nontoxic lambertellols (1, 2) as the diffusible precursors of the authentic responsible substance 3 regardless of existence of the host M. f.; (b) In the absence of the host, the environment around the parasite is kept under neutral condition, and both 1 and 2 are readily transformed into 3; (c) Lambertellin (3) inhibits not only the host but also the parasite. The parasite degrades 3 for detoxification; and (d) Upon the host M. f. approaching closely to the area where the parasite inhabits, the environment around the parasite becomes acidic to stabilize 1 and 2, which gives them a chance to diffuse into the host area. Then these are gradually transformed into 3 to inhibit the host without damaging the parasite. This mechanism also accords with the progress of 'Natsu-Nenju' disease on apple fruits, which is known to be a mysterious phyto-disease because of two unique stages of its lifecycle, anamorphic (asexual) and teleomorphic (sexual). The 'Lambertellin system' would be categorized as a novel class of alleropathies.
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http://dx.doi.org/10.1016/j.bmc.2014.02.051DOI Listing
April 2014

Evaluation of effects of multiple candidate genes (LEP, LEPR, MC4R, PIK3C3, and VRTN) on production traits in Duroc pigs.

Anim Sci J 2014 Mar 15;85(3):198-206. Epub 2013 Oct 15.

Central Research Institute for Feed and Livestock ZEN-NOH, National Federation of Agricultural Co-operative Associations, Kamishihoro, Hokkaido, Japan.

We evaluated multiple effects of genetic variations of five candidate loci (LEP, LEPR, MC4R, PIK3C3 and VRTN) on four production traits (average daily weight gain (ADG); backfat thickness (BFT); loin eye muscle area (EMA); and intramuscular fat content (IMF)) in a closed nucleus herd of pure Duroc pigs. Polymorphisms in LEPR, MC4R and PIK3C3 had significant single gene effects on ADG and BFT. The additive genetic variance in ADG and BFT (16.99% and 22.51%, respectively) was explained by genetic effects of these three loci. No correlations were observed between the LEP genotype and production traits in this study. Although we detected marginally epistatic interactions between LEPR and PIK3C3 on the eye muscle area, there were no significant epistatic effects on any traits among all loci pairs. These results suggest that LEPR, MC4R, PIK3C3 and VRTN may independently influence growth rate and fat deposition. Furthermore, the statistical models for predicting the breeding values of each trait had the lowest Akaike's information criterion values when considering the effect of the MC4R, LEPR, PIK3C3 and VRTN genotype simultaneously. These results suggest that LEPR, MC4R, PIK3C3 and VRTN are useful markers for accurately predicting breeding values in Duroc pigs.
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http://dx.doi.org/10.1111/asj.12134DOI Listing
March 2014

Characterization of a canine tetranucleotide microsatellite marker located in the first intron of the tumor necrosis factor alpha gene.

J Vet Med Sci 2014 Jan 13;76(1):119-22. Epub 2013 Sep 13.

Laboratory of Veterinary Internal Medicine II, Faculty of Veterinary Medicine, Azabu University, Chuo-ku, Sagamihara, Kanagawa 252-5201, Japan.

A polymorphic tetranucleotide (GAAT)n microsatellite in the first intron of the canine tumor necrosis factor alpha (TNFA) gene was characterized in this study; 139 dogs were analyzed: 22 Beagles, 26 Chihuahuas, 20 Miniature Dachshunds, 24 Miniature Poodles, 22 Pembroke Welsh Corgis and 25 Shiba Inus. We detected the presence of the 4 alleles (GAAT)5, (GAAT)6, (GAAT)7 and (GAAT)8, including 9 of the 10 expected genotypes. The expected heterozygosity (He) and the polymorphic information content (PIC) value of this microsatellite locus varied from 0.389 to 0.749 and from 0.333 to 0.682, respectively, among the 6 breeds. The allelic frequency differed greatly among breeds, but this microsatellite marker was highly polymorphic and could be a useful marker for the canine TNFA gene.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3979939PMC
http://dx.doi.org/10.1292/jvms.13-0316DOI Listing
January 2014

Valproic acid, a histone deacetylase inhibitor, decreases proliferation of and induces specific neurogenic differentiation of canine adipose tissue-derived stem cells.

J Vet Med Sci 2014 Jan 27;76(1):15-23. Epub 2013 Aug 27.

Graduate School of Veterinary Medicine, Azabu University, Fuchinobe, Chuo-ku, Sagamihara 252-5201, Japan.

Adipose tissue-derived stem cells (ADSCs) isolated from adult tissue have pluripotent differentiation and self-renewal capability. The tissue source of ADSCs can be obtained in large quantities and with low risks, thus highlighting the advantages of ADSCs in clinical applications. Valproic acid (VPA) is a widely used antiepileptic drug, which has recently been reported to affect ADSC differentiation in mice and rats; however, few studies have been performed on dogs. We aimed to examine the in vitro effect of VPA on canine ADSCs. Three days of pretreatment with VPA decreased the proliferation of ADSCs in a dose-dependent manner; VPA concentrations of 4 mM and above inhibited the proliferation of ADSCs. In parallel, VPA increased p16 and p21 mRNA expression, suggesting that VPA attenuated the proliferative activity of ADSCs by activating p16 and p21. Furthermore, the effects of VPA on adipogenic, osteogenic or neurogenic differentiation were investigated morphologically. VPA pretreatment markedly promoted neurogenic differentiation, but suppressed the accumulation of lipid droplets and calcium depositions. These modifications of ADSCs by VPA were associated with a particular gene expression profile, viz., an increase in neuronal markers, that is, NSE, TUBB3 and MAP2, a decrease in the adipogenic marker, LPL, but no changes in osteogenic markers, as estimated by reverse transcription-PCR analysis. These results suggested that VPA is a specific inducer of neurogenic differentiation of canine ADSCs and is a useful tool for studying the interaction between chromatin structure and cell fate determination.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3979952PMC
http://dx.doi.org/10.1292/jvms.13-0219DOI Listing
January 2014

Aqueous extracts of Rhizopus oryzae induced nitric oxide production in rat hepatocyte cell line RLN-10.

Biosci Biotechnol Biochem 2013 7;77(7):1384-9. Epub 2013 Jul 7.

Graduate School of Veterinary Medicine, Azabu University, Sagamihara 252-5201, Japan.

Aqueous extracts of Rhizopus oryzae (Aq-ROU) have a broad range of physiological activity. Here we identified a new physiological effect of Aq-ROU in rat hepatocyte cell line RLN-10. Aq-ROU induced the accumulation of nitrite, a stable metabolite nitric oxide (NO), in cell culture medium and induced potent diaminofluorescein-FM diacetate staining in the cells. Real-time reverse transcriptase (RT)-PCR analysis showed marked inducible NO synthase gene expression. Additionally, markedly enhanced expression of p22(phox) and temporally increased expression of NADPH oxidase1 indicated that superoxide was produced. Nuclear translocation of nuclear factor-kappa (NF-κ) B p65 increased remarkably following Aq-ROU and following lipopolysaccharide treatment, a potent activator of NF-κB. Ammonium pyrrolidine-1-carbodithioate, an inhibitor of NF-κB, inhibited NO production following Aq-ROU treatment. Our data indicate that Aq-ROU induces NO production and potentially the production of superoxide, which may contribute to the broad range of physiological effects observed for Aq-ROU ingested by animals.
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http://dx.doi.org/10.1271/bbb.120951DOI Listing
February 2014

Freshwater ascomycetes: Minutisphaera (Dothideomycetes) revisited, including one new species from Japan.

Mycologia 2013 Jul-Aug;105(4):959-76. Epub 2013 May 24.

Department of Chemistry and Biochemistry, University of North Carolina, Greensboro, NC, USA.

During investigations of freshwater ascomycetes we found one interesting taxon from Aomori (Japan), as well as three additional taxa from North Carolina (USA), which were morphologically similar to Minutisphaera, a recently described freshwater fungus in the Dothideomycetes. The ascomata of all the collections bore dark hair-like structures around the ostiolar region, obovoid to obclavate bitunicate asci, and one to three septate hyaline to brown ascospores with a sheath (in material from Japan), and with both sheath and appendages (in material from the USA). The apothecial ascomata of these taxa, however, differ from those of the type species of the genus, which are perithecial. Two collections of Minutisphaera-like fungi from the USA were morphologically quite similar but differed in ascospore size. To assess the phylogenetic affinities of Minutisphaera-like taxa with the type species, M. fimbriatispora, we sequenced 18S and 28S nrDNA of five newly collected strains of Minutisphaera. We also sequenced the nrDNA for the entire internal transcribed spacer region of 10 strains to assess interspecific and intraspecific variation with M. fimbriatispora. Additionally we examined the secondary metabolite profiles of two strains from USA. Based on maximum likelihood and Bayesian analyses of combined 18S and 28S, and separate ITS sequences, as well as examination of morphology, we describe and illustrate a new species, M. japonica. One collection from North Carolina is confirmed as M. fimbriatispora, while two other collections are Minutisphaera-like fungi that had a number of similar diagnostic morphological characters but differed only slightly in ascospore sizes. The phylogeny inferred from the internal transcribed spacer region suggested that two out of the three North Carolina collections may be novel and perhaps cryptic species within Minutisphaera. Organic extracts of Minutisphaera from USA, M. fimbriatispora (G155-1) and Minutisphaera-like taxon (G156-1), revealed the presence of palmitic acid and (E)-hexadec-9-en-1-ol as major chemical constituents. We discuss the placement of the Minutisphaera clade within the Dothideomycetes. The description of the genus Minutisphaera is emended to accommodate M. japonica within Minutisphaera.
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http://dx.doi.org/10.3852/12-313DOI Listing
September 2013
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