Characterization of arylsulfatase activity in brine shrimp, Artemia salina

Arylsulfatase from Artemia salina exists in at least two forms (AS I and AS II). The paper presents characterization of the AS II form of the arylsulfatase. The enzyme was able to hydrolyze p-nitrocatechol sulfate (pNCS) as well as ascorbate sulfate. It exhibited maximum activity at temperature of 50 °C and was stable for 2 h at 4-10 °C. Optimum pH shifted from 6.2 at 4 mM pNCS (substrate) to 4.8 at 20 mM pNCS. The enzyme displayed linear kinetics. AS II arylsulfatase exists in two molecular forms (349 and 460 kDa) composed of identical subunits with molecular mass of 53 kDa. Sulfite and phosphate ions were the most potent inhibitors of the enzyme. Cyanide proved to be a weak inhibitor. Sulfate and low concentrations of silver ions had no effect on the enzyme activity. Based on the above results, modifications in the assay for determination of enzyme activity are proposed.     

A comparative study of the fatty acid profile of Artemia franciscana and A. persimilis cultured at mesocosm scale

The goal of this study was to examine the fatty acid (FA) profile of two Artemia species, A. persimilis (Argentina) and A. franciscana (Great Salt Lake,Utah; USA) in coexistence at mesocosm scale. The experiment was carried out to 1) evaluate putative differences in the fatty acid composition of both species while they share resources and 2) to investigate the causes of such differences. Although the coexistence of these species in nature has not yet been observed, it remains possible that this situation may arise in the future mainly due to the invasive ability of A. franciscana. FA analyses were performed on individuals as well as on pooled biomasses of each species, and integrated in multivariate principal components analysis (PCA). Comparison of the relative abundance of FA between the two species revealed that interspecific differences in FA composition are greater than intraspecific variability. Higher percentages of unsaturation were found in the fatty acids of A. persimilis compared to A. franciscana, demonstrating that aside from a high phenotypic effect of diet on the FA composition of the animals, a species-specific genotypic effect should not be discarded.     

Larsenia salina gen. nov., sp. nov., a new member of the family Halomonadaceae based on multilocus sequence analysis

Two Gram-staining-negative, moderately halophilic bacteria, strains M1-18^T and L1-16, were isolated from a saltern located in Huelva (Spain). They were motile, strictly aerobic rods, growing in the presence of 3–25% (w/v) NaCl (optimal growth at 7.5–10% [w/v] NaCl), between pH 4.0 and 9.0 (optimal at pH 6.0–7.0) and at temperatures between 15 and 40 °C (optimal at 37 °C). Phylogenetic analysis based on 16S rRNA gene sequence comparison showed that both strains showed the higher similarity values with Chromohalobacter israelensis ATCC 43985^T (95.2–94.8%) and Chromohalobacter salexigens DSM 3043^T (95.0–94.9%), and similarity values lower than 94.6% with other species of the genera Chromohalobacter, Kushneria, Cobetia or Halomonas. Multilocus sequence analysis (MLSA) based on the partial sequences of atpA, rpoD and secA housekeeping genes indicated that the new isolates formed an independent and monophyletic branch that was related to the peripheral genera of the family Halomonadaceae, Halotalea, Carnimonas and Zymobacter, supporting their placement as a new genus of the Halomonadaceae. The DNA–DNA hybridization between both strains was 82%, whereas the values between strain M1-18^T and the most closely related species of Chromohalobacter and Kushneria were equal or lower to 48%. The major cellular fatty acids were C_18:1ω7c/C_18:1ω6c, C_16:0, and C_16:1ω7c/C_16:1ω6c, a profile that differentiate this new taxon from species of the related genera. We propose the placement of both strains as a novel genus and species, within the family Halomonadaceae, with the name Larsenia salina gen. nov., sp. nov. The type strain is M1-18^T (= CCM 8464 = CECT 8192^T = IBRC-M 10767^T = LMG 27461^T).     

Cryptic microsporidian parasites differentially affect invasive and native Artemia spp.

We investigated the host specificity of two cryptic microsporidian species (Anostracospora rigaudi and Enterocytospora artemiae) infecting invasive (Artemia franciscana) and native (Artemia parthenogenetica) hosts in sympatry. Anostracospora rigaudi was on average four times more prevalent in the native host, whereas E. artemiae was three times more prevalent in the invasive host. Infection with An. rigaudi strongly reduced female reproduction in both host species, whereas infection with E. artemiae had weaker effects on female reproduction. We contrasted microsporidian prevalence in native A. franciscana populations (New World) and in both invaded and non-invaded Artemia populations (Old World). At a community level, microsporidian prevalence was twice as high in native compared with invasive hosts, due to the contrasting host-specificity of An. rigaudi and E. artemiae. At a higher biogeographical level, microsporidian prevalence in A. franciscana did not differ between the invaded populations and the native populations used for the introduction. Although E. artemiae was the only species found both in New and Old World populations, no evidence of its co-introduction with the invasive host was found in our experimental and phylogeographic tests. These results suggest that the success of A. franciscana invasion is probably due to a lower susceptibility to virulent microsporidian parasites rather than to decreased microsporidian prevalence compared with A. parthenogenetica or to lower microsporidian virulence in introduced areas.     

CK2 activity is modulated by growth rate in Saccharomyces cerevisiae

CK2 is a highly conserved protein kinase controlling different cellular processes. It shows a higher activity in proliferating mammalian cells, in various types of cancer cell lines and tumors. The findings presented herein provide the first evidence of an in vivo modulation of CK2 activity, dependent on growth rate, in Saccharomyces cerevisiae. In fact, CK2 activity, assayed on nuclear extracts, is shown to increase in exponential growing batch cultures at faster growth rate, while localization of catalytic and regulatory subunits is not nutritionally modulated. Differences in intracellular CK2 activity of glucose- and ethanol-grown cells appear to depend on both increase in molecule number and k_cat. Also in chemostat cultures nuclear CK2 activity is higher in faster growing cells providing the first unequivocal demonstration that growth rate itself can affect CK2 activity in a eukaryotic organism.     

The deubiquitinase emperor's thumb is a regulator of apoptosis in Drosophila

We have characterized the gene emperor's thumb (et) and showed that it is required for the regulation of apoptosis in Drosophila. Loss-of-function mutations in et result in apoptosis associated with a decrease in the concentration of DIAP1. Overexpression of one form of et inhibits apoptosis, consistent with et having an anti-apoptotic function; however, overexpression of a second form of et induces apoptosis, indicating that the two forms of et may have competing functions. et encodes a protein deubiquitinase, suggesting it regulates apoptosis by controlling the stability of apoptotic regulatory proteins.     

Toxocara canis: Potential activity of natural products against second-stage larvae in vitro and in vivo

The anthelmintic activity of extracts from Chenopodiumambrosioides, Pycnanthusangolensis and Nutridesintox® was in vitro and in vivo investigated, against Toxocaracanis larvae. The in vitro assays results showed that the aqueous extract of Nutridesintox® was the most effective, followed by C. ambrosioides extracts, hexane, dichloromethane and the infusion. P. angolensis extracts showed a lower anthelmintic activity compared to the other natural products. For the in vivo assays, Nutridesintox®, the hexane extract and the infusion of C. ambrosioides were administered orally to T. canis-infected mice, in single doses, during three consecutive days. The efficacy was evaluated on the 17th day post-infection, not only by counting T. canis larvae in the tissues but also by ELISA detection of IgM and IgG antibodies and histological analysis of liver and lungs. The different treatments did not reduce the larvae burden and had no influence on the antibodies dynamic. Interestingly, a reduction on the inflammatory infiltrates was observed in the liver and lung sections of the group treated with the hexane extract of C. ambrosioides. In conclusion, the hexane extract of C. ambrosioides is of further research interest, as it showed an anthelmintic activity in vitro and a reduction on the inflammatory reaction produced by the infection of T. canis larvae in vivo.     

施氮对亚热带樟树林土壤呼吸的影响

人类活动引起全球范围内大气氮沉降量的升高,增加了陆地生态系统氮输入,从而影响土壤CO2排放.为揭示生态系统氮输入升高对土壤呼吸的影响,2010年6月至2012年1月,对亚热带樟树林(Cinnamomum camphora)进行模拟氮添加试验,每月上、下旬采用红外分析法测定4种氮输入水平(CK,0 gm-2a-1;低氮LN,5gm-2a-1;中氮MN,15 g m-2 a-1;高氮HN,30gm-2a-1)下的土壤呼吸速率.结果表明:(1)樟树林土壤呼吸存在明显的季节动态,最高值出现在6月,最小值出现在1月.氮添加处理显著抑制了樟树林的土壤呼吸,LN、MN、HN处理土壤呼吸年累积量分别较对照CK下降37.66％、30.62％、38.95％,各施氮处理间无显著差异,施氮对土壤呼吸的抑制作用随时间推移而减弱;(2)氮添加不影响土壤呼吸昼夜波动特征,但显著抑制土壤呼吸速率;(3)土壤呼吸与土壤温度间存在极显著的指数关系,与土壤湿度相关性不显著,CK、LN处理Q10相近,MN处理最小:(4)氮添加处理促进了土壤中氮的淋失,且随施氮水平的升高而增大.     

Genetic differentiation of Artemia urmiana from various ecological populations of Urmia Lake assessed by PCR amplified RFLP analysis

This study concentrated on the mitochondrial DNA diversity in adult Artemia urmiana populations. A total of 210 individual specimens were collected from the surface and bottom layers from three different geographical areas, comprising six sampling sites in Urmia Lake (Iran). The mitochondrial rDNA gene region was amplified using the PCR technique followed by RFLP analysis based on using eleven restriction endonucleases. Analysis at the intrapopulation level indicated that the two median and south area bottom layers have somewhat higher proportion of polymorphic sites compare to others. Although floating and sinking cysts did not consistently show genetic differences, there was significant genetic variation among all samples (F_ST = 0.019, P = 0.03) and 98% of the variation was within samples. Our results clearly distinguished nucleotide divergence and genetic structuring patterns, suggest that may be genetic differentiation existed in Artemia populations from Urmia Lake. Pairwise comparisons of the samples showed that the south area surface layer location was the most genetically divergent of the six sampling sites. Genetic characterizations of the various Artemia populations in two defined depths (surface and bottom) revealed differentiation that may be important in understanding the ecology of this commercially important species.     

Acanthamoeba polyphaga is a possible host for Vibrio cholerae in aquatic environments

Acanthamoeba is a genus of free-living amoebae found to be able to host many bacterial species living in the environment. Acanthamoebae and Vibrio cholerae are found in the aquatic environments of cholera endemic areas. Previously it has been shown that V. cholerae O1 and O139 can survive and grow in Acanthamoeba castellanii. The aim of this study was to examine the ability of Acanthamoeba polyphaga to host V. cholerae O1 and O139. The interaction between A. polyphaga and V. cholerae strains was studied by means of viable amoeba cell counts and viable count of the bacteria in the absence and presence of amoebae. The viable count of intracellularly growing bacteria was estimated by utilizing gentamicin assay. Electron microscopy was used to determine the localization of V. cholerae inside A. polyphaga. The results showed that A. polyphaga enhanced growth and survival of V. cholerae, which grew and survived inside the amoeba cells for 2 weeks. The electron microscopy showed that A. polyphaga hosted intracellular V. cholerae localized in the vacuoles of amoeba cell. Neither the presence of V. cholerae together with A. polyphaga nor the intracellular localization of the bacteria inhibited growth and survival of A. polyphaga. The outcome of the interaction between these microorganisms may support strongly the role of A. polyphaga as host for V. cholerae O1 and O139.     

More constraint on ParaHox than Hox gene families in early metazoan evolution

Hox and ParaHox (H/P) genes belong to evolutionary-sister clusters that arose through duplication of a ProtoHOX cluster early in animal evolution. In contrast to bilaterians, cnidarians express, beside PG1, PG2 and Gsx orthologs, numerous Hox-related genes with unclear origin. We characterized from marine hydrozoans three novel Hox-related genes expressed at medusa and polyp stages, which include a Pdx/Xlox ParaHox ortholog induced 1 day later than Gsx during embryonic development. To reconstruct H/P genes' early evolution, we performed multiple systematic comparative phylogenetic analyses, which identified derived sequences that blur the phylogenetic picture, recorded dramatically different evolutionary rates between ParaHox and Hox in cnidarians and showed the unexpected grouping of [Gsx-Pdx/Xlox-PG2-PG3] families in a single metagroup distinct from PG1. We propose a novel more parsimonious evolutionary scenario whereby H/P genes originated from a [Gsx-Pdx/Xlox-PG2-PG3]-related ProtoHox gene, the «posterior» and «anterior» H/P genes appearing secondarily. The ProtoHOX cluster would have contained the three Gsx/PG2, Pdx/PG3, Cdx/PG9 paralogs and produced through tandem duplication the primordial HOX and ParaHOX clusters in the Cnidaria-Bilateria ancestor. The stronger constraint on cnidarian ParaHox genes suggests that the primary function of pre-bilaterian H/P genes was to drive cellular evolutionary novelties such as neurogenesis rather than axis specification.     

Validating the in vitro antimicrobial activity of Artemisia afra in polyherbal combinations to treat respiratory infections

Artemisia afra is one of the most widely used medicinal plants in African traditional medicine and is commonly administered in polyherbal combinations to treat respiratory infections. Focussing on plant volatiles, the aim of this study was to provide scientific evidence for the antimicrobial activity of A. afra (principle plant) in combination with essential oils from three medicinal aromatic plants; Agathosma betulina, Eucalyptus globulus and Osmitopsis asteriscoides. In vitro minimum inhibitory concentration (MIC) assays were undertaken on four pathogens (Enterococcus faecalis ATCC 29212, Moraxella catarrhalis ATCC 23246, Klebsiella pneumoniae NCTC 9633 and Cryptococcus neoformans ATCC 90112) to determine antimicrobial efficacy of the oils and their combinations. The fractional inhibitory concentration (FIC) and isobolograms were used to interpret pharmacodynamic interactions such as synergy, antagonism or additive profiles. The antimicrobial activity of the individual oils mostly displayed moderate activity. Predominantly, additive interactions were noted. The most prominent synergistic interaction (FIC value of 0.5) was observed when A. afra was combined with O. asteriscoides in the 8:2 ratio (eight parts A. afra with two parts O. asteriscoides) against C. neoformans. No antagonistic interactions were evident.     

Upregulation of PHLDA2 in Dicer knockdown HEK293 cells

It has been reported that RNAi-dependent chromatin silencing in vertebrates is not restricted to the centromeres. To address whether RNAi machinery could regulate the chromatin structure of imprinted genes, we knocked down Dicer in HEK293 cells and found that the expression of PHLDA2, one of the several genes in the imprinted gene domain of 11p15.5, was specifically upregulated. This was accompanied by a shift towards more activated chromatin at PHLDA2 locus as indicated by change in H3K9 acetylation, however, the methylation state at this locus was not affected. Furthermore, we found that PHLDA2 was downregulated in growth-arrested HEK293 cells induced by either serum deprivation or contact inhibition. This suggests that PHLDA2 upregulation might be a direct result of Dicer depletion rather than the consequence of growth arrest induced by Dicer knockdown. Considering the reports that there is consistent placental outgrowth in PHLDA2 knockout mice and that PHLDA2 overexpression in mice causes growth inhibition, we speculate that PHLDA2 may be a candidate for contributing to the reduced growth rate of Dicer-deficient cells and the very early embryonic lethality in Dicer knockout mice.     

Filtration and respiration of the deep living bivalve Acesta excavata (J.C. Fabricius, 1779) (Bivalvia; Limidae)

Here we report the first study of clearance rate and respiration rate of a deep living bivalve, Acesta excavata (J.C. Fabricius, 1779) (Mollusca: Limidae). We found that A. excavata had extreme values both for clearance and respiration rates compared to other bivalves. It has the second largest clearance rate ever reported, 13.36 l h^− 1 g^− 1, and the second lowest value of respiration rate, 0.12 ml O₂ h^− 1 g^− 1. The gill area of 7063 mm² g^− 1 is one of the largest found in bivalves so far. We suggest that these values indicate a physiological adaptation to the low and irregular food supply in the deep sea rather that a specific adaptation to depth.     

RPB2 sequences reveal a close phylogenetic relationship between tetraploid Hordelymus and diploid Hordeum species in Triticeae (Poaceae)

The origin of Hordelymus genome has been debated for years, and no consensus conclusion was reached. In this study, we sequenced and analyzed the RPB2 (RNA polymerase subunit II) gene from Hordelymus europaeus (L.) Harz, and its potential diploid ancestor species those were suggested in previous studies. The focus of this study was to examine the phylogenetic relationship of Hordelymus genomes with its potential donor Hordeum, Psathyrostachys, and Taeniatherum species. Two distinguishable copies of sequences were obtained from H. europaeus. The obvious difference between the two copies of sequences is a 24 bp indel (insertion/deletion). Phylogenetic analysis showed a strong affinity between Hordeum genome and Hordelymus with 85% bootstrap support. These results suggested that one genome in tetraploid H. europaeus closely related to the genome in Hordeum species. Another genome in H. europaeus is sister to the genomes in Triticeae species examined here, which corresponds well with the recently published EF-G data. No obvious relationship was found between Hordelymus and either Ta genome donor, Taeniatherum caput-medusae or Ns genome donor, Psathyrostachys juncea. Our data does not support the presence of Ta and Ns genome in H. europaeus, and further confirms that H. europaeus is allopolyploid.     

The MerE protein encoded by transposon Tn21 is a broad mercury transporter in Escherichia coli

In order to clarify the physiological role of the merE gene of transposon Tn21, a pE4 plasmid that contained the merR gene of plasmid pMR26 from Pseudomonas strain K-62, and the merE gene of Tn21 from the Shigella flexneri plasmid NR1 (R100) was constructed. Bacteria with plasmid pE4 (merR-o/p-merE) were more hypersensitive to CH₃Hg(I) and Hg(II), and took up significantly more CH₃Hg(I) and Hg(II), than the isogenic strain. The MerE protein encoded by pE4 was localized in the membrane cell fraction, but not in the soluble fraction. Based on these experimental results, we suggest for the first time that the merE gene is a broad mercury transporter mediating the transport of both CH₃Hg(I) and Hg(II) across the bacterial membrane.     

Centrosema is a promiscuous legume nodulated by several new putative species and symbiovars of Bradyrhizobium in various American countries

Centrosema is an American indigenous legume that can be used in agroecosystems for recovery of acidic and degraded soils. In this study, a Centrosema-nodulating rhizobial collection of strains isolated in a poor acid savanna soil from Venezuela was characterized, and the members of the collection were compared to other Centrosema strains from America. The analysis of the rrs gene showed that the strains nodulating Centrosema in American countries were closely related to different species of the genus Bradyrhizobium. However, the analysis of the atpD and recA genes, as well as the 16S–23S ITS region, showed that they formed several new phylogenetic lineages within this genus. The Venezuela strains formed three lineages that were divergent among themselves and with respect to those formed by Centrosema strains isolated in other countries, as well as to the currently described species and genospecies of Bradyrhizobium. In addition, the symbiotic genes nodC and nifH carried by Centrosema-nodulating strains were analyzed for the first time, and it was shown that they belonged to three new phylogenetic lineages within Bradyrhizobium. The nodC genes of the Centrosema strains were divergent among themselves and with respect to the genistearum and glycinearum symbiovars, indicating that Centrosema is a promiscuous legume. According to these results, the currently known Centrosema-nodulating strains represent several new putative species and symbiovars of the genus Bradyrhizobium.