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Sample records for 4-reductase dfr anthocyanidin

  1. Characterization of dihydroflavonol 4-reductase (DFR) genes and their association with cold and freezing stress in Brassica rapa.

    PubMed

    Ahmed, Nasar Uddin; Park, Jong-In; Jung, Hee-Jeong; Yang, Tae-Jin; Hur, Yoonkang; Nou, Ill-Sup

    2014-10-15

    Flavonoids including anthocyanins provide flower and leaf colors, as well as other derivatives that play diverse roles in plant development and interactions with the environment. Dihydroflavonol 4-reductase (DFR) is part of an important step in the flavonoid biosynthetic pathway of anthocyanins. This study characterized 12 DFR genes of Brassica rapa and investigated their association with anthocyanin coloration, as well as cold and freezing stress in several genotypes of B. rapa. Comparison of sequences of these genes with DFR gene sequences from other species revealed a high degree of homology. Constitutive expression of the genes in several pigmented and non-pigmented lines of B. rapa demonstrated correlation with anthocyanin accumulation for BrDFR8 and 9. Conversely, BrDFR2, 4, 8 and 9 only showed very high responses to cold stress in pigmented B. rapa samples. BrDFR1, 3, 5, 6 and 10 responded to cold and freezing stress treatments, regardless of pigmentation. BrDFRs were also shown to be regulated by two transcription factors, BrMYB2-2 and BrTT8, contrasting with anthocyanin accumulation and cold and freezing stress. Thus, the above results suggest that these genes are associated with anthocyanin biosynthesis and cold and freezing stress tolerance and might be useful resources for development of cold and/or freezing stress resistant Brassica crops with desirable colors as well. These findings may also facilitate exploration of the molecular mechanism that regulates anthocyanin biosynthesis and its response to abiotic stresses.

  2. Characterization of dihydroflavonol 4-reductase (DFR) genes and their association with cold and freezing stress in Brassica rapa.

    PubMed

    Ahmed, Nasar Uddin; Park, Jong-In; Jung, Hee-Jeong; Yang, Tae-Jin; Hur, Yoonkang; Nou, Ill-Sup

    2014-10-15

    Flavonoids including anthocyanins provide flower and leaf colors, as well as other derivatives that play diverse roles in plant development and interactions with the environment. Dihydroflavonol 4-reductase (DFR) is part of an important step in the flavonoid biosynthetic pathway of anthocyanins. This study characterized 12 DFR genes of Brassica rapa and investigated their association with anthocyanin coloration, as well as cold and freezing stress in several genotypes of B. rapa. Comparison of sequences of these genes with DFR gene sequences from other species revealed a high degree of homology. Constitutive expression of the genes in several pigmented and non-pigmented lines of B. rapa demonstrated correlation with anthocyanin accumulation for BrDFR8 and 9. Conversely, BrDFR2, 4, 8 and 9 only showed very high responses to cold stress in pigmented B. rapa samples. BrDFR1, 3, 5, 6 and 10 responded to cold and freezing stress treatments, regardless of pigmentation. BrDFRs were also shown to be regulated by two transcription factors, BrMYB2-2 and BrTT8, contrasting with anthocyanin accumulation and cold and freezing stress. Thus, the above results suggest that these genes are associated with anthocyanin biosynthesis and cold and freezing stress tolerance and might be useful resources for development of cold and/or freezing stress resistant Brassica crops with desirable colors as well. These findings may also facilitate exploration of the molecular mechanism that regulates anthocyanin biosynthesis and its response to abiotic stresses. PMID:25108127

  3. Anthocyanidin synthase in non-anthocyanin-producing Caryophyllales species.

    PubMed

    Shimada, Setsuko; Inoue, Yoriko T; Sakuta, Masaaki

    2005-12-01

    Red colors in flowers are mainly produced by two types of pigments: anthocyanins and betacyanins. Although anthocyanins are widely distributed in higher plants, betacyanins have replaced anthocyanins in the Caryophyllales. There has been no report so far to find anthocyanins and betacyanins existing together within the same plant. This curious phenomenon has been examined from genetic and evolutionary perspectives, however nothing is known at the molecular level about the mutual exclusion of anthocyanins and betacyanins in higher plants. Here, we show that spinach (Spinacia oleracea) and pokeweed (Phytolacca americana), which are both members of the Caryophyllales, have functional anthocyanidin synthases (ANSs). The ability of ANSs of the Caryophyllales to oxidize trans-leucocyanidin to cyanidin is comparable to that of ANSs in anthocyanin-producing plants. Expression profiles reveal that, in spinach, dihydroflavonol 4-reductase (DFR) and ANS are not expressed in most tissues and organs, except seeds, in which ANS may contribute to proanthocyanidin synthesis. One possible explanation for the lack of anthocyanins in the Caryophyllales is the suppression or limited expression of the DFR and ANS.

  4. Expression analysis of dihydroflavonol 4-reductase genes in Petunia hybrida.

    PubMed

    Chu, Y X; Chen, H R; Wu, A Z; Cai, R; Pan, J S

    2015-01-01

    Dihydroflavonol 4-reductase (DFR) genes from Rosa chinensis (Asn type) and Calibrachoa hybrida (Asp type), driven by a CaMV 35S promoter, were integrated into the petunia (Petunia hybrida) cultivar 9702. Exogenous DFR gene expression characteristics were similar to flower-color changes, and effects on anthocyanin concentration were observed in both types of DFR gene transformants. Expression analysis showed that exogenous DFR genes were expressed in all of the tissues, but the expression levels were significantly different. However, both of them exhibited a high expression level in petals that were starting to open. The introgression of DFR genes may significantly change DFR enzyme activity. Anthocyanin ultra-performance liquid chromatography results showed that anthocyanin concentrations changed according to DFR enzyme activity. Therefore, the change in flower color was probably the result of a DFR enzyme change. Pelargonidin 3-O-glucoside was found in two different transgenic petunias, indicating that both CaDFR and RoDFR could catalyze dihydrokaempferol. Our results also suggest that transgenic petunias with DFR gene of Asp type could biosynthesize pelargonidin 3-O-glucoside. PMID:25966276

  5. Characterization of three active transposable elements recently inserted in three independent DFR-A alleles and one high-copy DNA transposon isolated from the Pink allele of the ANS gene in onion (Allium cepa L.).

    PubMed

    Kim, Sunggil; Park, Jee Young; Yang, Tae-Jin

    2015-06-01

    Intact retrotransposon and DNA transposons inserted in a single gene were characterized in onions (Allium cepa) and their transcription and copy numbers were estimated in this study. While analyzing diverse onion germplasm, large insertions in the DFR-A gene encoding dihydroflavonol 4-reductase (DFR) involved in the anthocyanin biosynthesis pathway were found in two accessions. A 5,070-bp long terminal repeat (LTR) retrotransposon inserted in the active DFR-A (R4) allele was identified from one of the large insertions and designated AcCOPIA1. An intact ORF encoded typical domains of copia-like LTR retrotransposons. However, AcCOPIA1 contained atypical 'TG' and 'TA' dinucleotides at the ends of the LTRs. A 4,615-bp DNA transposon was identified in the other large insertion. This DNA transposon, designated AcCACTA1, contained an ORF coding for a transposase showing homology with the CACTA superfamily transposable elements (TEs). Another 5,073-bp DNA transposon was identified from the DFR-A (TRN) allele. This DNA transposon, designated AchAT1, belonged to the hAT superfamily with short 4-bp terminal inverted repeats (TIRs). Finally, a 6,258-bp non-autonomous DNA transposon, designated AcPINK, was identified in the ANS-p allele encoding anthocyanidin synthase, the next downstream enzyme to DFR in the anthocyanin biosynthesis pathway. AcPINK also possessed very short 3-bp TIRs. Active transcription of AcCOPIA1, AcCACTA1, and AchAT1 was observed through RNA-Seq analysis and RT-PCR. The copy numbers of AcPINK estimated by mapping the genomic DNA reads produced by NextSeq 500 were predominantly high compared with the other TEs. A series of evidence indicated that these TEs might have transposed in these onion genes very recently, providing a stepping stone for elucidation of enormously large-sized onion genome structure.

  6. Characterization of three active transposable elements recently inserted in three independent DFR-A alleles and one high-copy DNA transposon isolated from the Pink allele of the ANS gene in onion (Allium cepa L.).

    PubMed

    Kim, Sunggil; Park, Jee Young; Yang, Tae-Jin

    2015-06-01

    Intact retrotransposon and DNA transposons inserted in a single gene were characterized in onions (Allium cepa) and their transcription and copy numbers were estimated in this study. While analyzing diverse onion germplasm, large insertions in the DFR-A gene encoding dihydroflavonol 4-reductase (DFR) involved in the anthocyanin biosynthesis pathway were found in two accessions. A 5,070-bp long terminal repeat (LTR) retrotransposon inserted in the active DFR-A (R4) allele was identified from one of the large insertions and designated AcCOPIA1. An intact ORF encoded typical domains of copia-like LTR retrotransposons. However, AcCOPIA1 contained atypical 'TG' and 'TA' dinucleotides at the ends of the LTRs. A 4,615-bp DNA transposon was identified in the other large insertion. This DNA transposon, designated AcCACTA1, contained an ORF coding for a transposase showing homology with the CACTA superfamily transposable elements (TEs). Another 5,073-bp DNA transposon was identified from the DFR-A (TRN) allele. This DNA transposon, designated AchAT1, belonged to the hAT superfamily with short 4-bp terminal inverted repeats (TIRs). Finally, a 6,258-bp non-autonomous DNA transposon, designated AcPINK, was identified in the ANS-p allele encoding anthocyanidin synthase, the next downstream enzyme to DFR in the anthocyanin biosynthesis pathway. AcPINK also possessed very short 3-bp TIRs. Active transcription of AcCOPIA1, AcCACTA1, and AchAT1 was observed through RNA-Seq analysis and RT-PCR. The copy numbers of AcPINK estimated by mapping the genomic DNA reads produced by NextSeq 500 were predominantly high compared with the other TEs. A series of evidence indicated that these TEs might have transposed in these onion genes very recently, providing a stepping stone for elucidation of enormously large-sized onion genome structure. PMID:25515665

  7. The potato R locus codes for dihydroflavonol 4-reductase.

    PubMed

    Zhang, Yongfei; Cheng, Shuping; De Jong, Darlene; Griffiths, Helen; Halitschke, Rayko; De Jong, Walter

    2009-09-01

    The potato R locus is required for the production of red pelargonidin-based anthocyanin pigments in potato (Solanum tuberosum L.). Red color also requires tissue-specific regulatory genes, such as D (for expression in tuber skin) and F (expression in flowers). A related locus, P, is required for production of blue/purple anthocyanins; P is epistatic to R. We have previously reported that the dihydroflavonol 4-reductase gene (dfr) co-segregates with R. To test directly whether R corresponds to dfr, we placed the allele of dfr associated with red color under the control of the CaMV 35S promoter and introduced it into the potato cultivar Prince Hairy (genotype dddd rrrr P-), which has white tubers and pale blue flowers. Transgenic Prince Hairy tubers remained white, but flower color changed to purple. Three independent transgenic lines, as well as a vector-transformed line, were then crossed with the red-skinned variety Chieftain (genotype D-R-pppp), to establish populations that segregated for D, R, P, and the dfr transgene or empty vector. Markers were used to genotype progeny at D and R. Progeny carrying the empty vector in the genetic background D-rrrr produced white or purple tubers, while progeny with the same genotype and the dfr transgene produced red or purple tubers. HPLC and LC-MS/MS analyses of anthocyanins present in Chieftain and in a red-skinned progeny clone with the dfr transgene in a D-rrrr background revealed no qualitative differences. Thus, dfr can fully complement R, both in terms of tuber color and anthocyanin composition.

  8. Molecular Cloning and Characterization of Two Genes Encoding Dihydroflavonol-4-Reductase from Populus trichocarpa

    PubMed Central

    Jia, Zhichun; Yang, Li; Sun, Yimin; Xiao, Xunyan; Song, Feng; Luo, Keming

    2012-01-01

    Dihydroflavonol 4-reductase (DFR, EC 1.1.1.219) is a rate-limited enzyme in the biosynthesis of anthocyanins and condensed tannins (proanthocyanidins) that catalyzes the reduction of dihydroflavonols to leucoanthocyanins. In this study, two full-length transcripts encoding for PtrDFR1 and PtrDFR2 were isolated from Populus trichocarpa. Sequence alignment of the two PtrDFRs with other known DFRs reveals the homology of these genes. The expression profile of PtrDFRs was investigated in various tissues of P. trichocarpa. To determine their functions, two PtrDFRs were overexpressed in tobacco (Nicotiana tabacum) via Agrobacterium-mediated transformation. The associated color change in the flowers was observed in all 35S:PtrDFR1 lines, but not in 35S:PtrDFR2 lines. Compared to the wild-type control, a significantly higher accumulation of anthocyanins was detected in transgenic plants harboring the PtrDFR1. Furthermore, overexpressing PtrDFR1 in Chinese white poplar (P. tomentosa Carr.) resulted in a higher accumulation of both anthocyanins and condensed tannins, whereas constitutively expressing PtrDFR2 only improved condensed tannin accumulation, indicating the potential regulation of condensed tannins by PtrDFR2 in the biosynthetic pathway in poplars. PMID:22363429

  9. Molecular cloning and characterization of three genes encoding dihydroflavonol-4-reductase from Ginkgo biloba in anthocyanin biosynthetic pathway.

    PubMed

    Hua, Cheng; Linling, Li; Shuiyuan, Cheng; Fuliang, Cao; Feng, Xu; Honghui, Yuan; Conghua, Wu

    2013-01-01

    Dihydroflavonol-4-reductase (DFR, EC1.1.1.219) catalyzes a key step late in the biosynthesis of anthocyanins, condensed tannins (proanthocyanidins), and other flavonoids important to plant survival and human nutrition. Three DFR cDNA clones (designated GbDFRs) were isolated from the gymnosperm Ginkgo biloba. The deduced GbDFR proteins showed high identities to other plant DFRs, which form three distinct DFR families. Southern blot analysis showed that the three GbDFRs each belong to a different DFR family. Phylogenetic tree analysis revealed that the GbDFRs share the same ancestor as other DFRs. The expression of the three recombinant GbDFRs in Escherichia coli showed that their actual protein sizes were in agreement with predictions from the cDNA sequences. The recombinant proteins were purified and their activity was analyzed; both GbDFR1 and GbDFR3 could catalyze dihydroquercetin conversion to leucocyanidin, while GbDFR2 catalyzed dihydrokaempferol conversion to leucopelargonidin. qRT-PCR showed that the GbDFRs were expressed in a tissue-specific manner, and transcript accumulation for the three genes was highest in young leaves and stamens. These transcription patterns were in good agreement with the pattern of anthocyanin accumulation in G.biloba. The expression profiles suggested that GbDFR1 and GbDFR2 are mainly involved in responses to plant hormones, environmental stress and damage. During the annual growth cycle, the GbDFRs were significantly correlated with anthocyanin accumulation in leaves. A fitted linear curve showed the best model for relating GbDFR2 and GbDFR3 with anthocyanin accumulation in leaves. GbDFR1 appears to be involved in environmental stress response, while GbDFR3 likely has primary functions in the synthesis of anthocyanins. These data revealed unexpected properties and differences in three DFR proteins from a single species. PMID:23991027

  10. Functional Characterization of a Dihydroflavanol 4-Reductase from the Fiber of Upland Cotton (Gossypium hirsutum).

    PubMed

    Wang, Le; Zhu, Yue; Wang, Peng; Fan, Qiang; Wu, Yao; Peng, Qing-Zhong; Xia, Gui-Xian; Wu, Jia-He

    2016-01-26

    Dihydroflavanol 4-reductase (DFR) is a key later enzyme involved in two polyphenols' (anthocyanins and proanthocyanidins (PAs)) biosynthesis, however it is not characterized in cotton yet. In present reports, a DFR cDNA homolog (designated as GhDFR1) was cloned from developing fibers of upland cotton. Silencing GhDFR1 in cotton by virus-induced gene silencing led to significant decrease in accumulation of anthocyanins and PAs. More interestingly, based on LC-MS analysis, two PA monomers, (-)-epicatachin and (-)-epigallocatachin, remarkably decreased in content in fibers of GhDFR1-silenced plants, but two new monomers, (-)-catachin and (-)-gallocatachin were present compared to the control plants infected with empty vector. The ectopic expression of GhDFR1 in an Arabidopsis TT3 mutant allowed for reconstruction of PAs biosynthesis pathway and led to accumulation of PAs in seed coat. Taken together, these data demonstrate that GhDFR1 contributes to the biosynthesis of anthocyanins and PAs in cotton.

  11. Functional Characterization of a Dihydroflavanol 4-Reductase from the Fiber of Upland Cotton (Gossypium hirsutum).

    PubMed

    Wang, Le; Zhu, Yue; Wang, Peng; Fan, Qiang; Wu, Yao; Peng, Qing-Zhong; Xia, Gui-Xian; Wu, Jia-He

    2016-01-01

    Dihydroflavanol 4-reductase (DFR) is a key later enzyme involved in two polyphenols' (anthocyanins and proanthocyanidins (PAs)) biosynthesis, however it is not characterized in cotton yet. In present reports, a DFR cDNA homolog (designated as GhDFR1) was cloned from developing fibers of upland cotton. Silencing GhDFR1 in cotton by virus-induced gene silencing led to significant decrease in accumulation of anthocyanins and PAs. More interestingly, based on LC-MS analysis, two PA monomers, (-)-epicatachin and (-)-epigallocatachin, remarkably decreased in content in fibers of GhDFR1-silenced plants, but two new monomers, (-)-catachin and (-)-gallocatachin were present compared to the control plants infected with empty vector. The ectopic expression of GhDFR1 in an Arabidopsis TT3 mutant allowed for reconstruction of PAs biosynthesis pathway and led to accumulation of PAs in seed coat. Taken together, these data demonstrate that GhDFR1 contributes to the biosynthesis of anthocyanins and PAs in cotton. PMID:26821011

  12. Excision of transposable elements from the chalcone isomerase and dihydroflavonol 4-reductase genes may contribute to the variegation of the yellow-flowered carnation (Dianthus caryophyllus).

    PubMed

    Itoh, Yoshio; Higeta, Daisuke; Suzuki, Akane; Yoshida, Hiroyuki; Ozeki, Yoshihiro

    2002-05-01

    In the "Rhapsody" cultivar of the carnation, which bears white flowers variegated with red flecks and sectors, a transposable element, dTdic1, belonging to the Ac/Ds superfamily, was found within the dihydroflavonol 4-reductase (DFR) gene. The red flecks and sectors of "Rhapsody" may be attributable to a reversion to DFR activity after the excision of dTdic1. The yellow color of the carnation petals is attributed to the synthesis and accumulation of chalcone 2'-glucoside. In several of the carnation cultivars that bear yellow flowers variegated with white flecks and sectors, both the chalcone isomerase (CHI) and DFR genes are disrupted by dTdic1.

  13. Dihydroflavonol 4-reductase genes encode enzymes with contrasting substrate specificity and show divergent gene expression profiles in Fragaria species.

    PubMed

    Miosic, Silvija; Thill, Jana; Milosevic, Malvina; Gosch, Christian; Pober, Sabrina; Molitor, Christian; Ejaz, Shaghef; Rompel, Annette; Stich, Karl; Halbwirth, Heidi

    2014-01-01

    During fruit ripening, strawberries show distinct changes in the flavonoid classes that accumulate, switching from the formation of flavan 3-ols and flavonols in unripe fruits to the accumulation of anthocyanins in the ripe fruits. In the common garden strawberry (Fragaria×ananassa) this is accompanied by a distinct switch in the pattern of hydroxylation demonstrated by the almost exclusive accumulation of pelargonidin based pigments. In Fragaria vesca the proportion of anthocyanins showing one (pelargonidin) and two (cyanidin) hydroxyl groups within the B-ring is almost equal. We isolated two dihydroflavonol 4-reductase (DFR) cDNA clones from strawberry fruits, which show 82% sequence similarity. The encoded enzymes revealed a high variability in substrate specificity. One enzyme variant did not accept DHK (with one hydroxyl group present in the B-ring), whereas the other strongly preferred DHK as a substrate. This appears to be an uncharacterized DFR variant with novel substrate specificity. Both DFRs were expressed in the receptacle and the achenes of both Fragaria species and the DFR2 expression profile showed a pronounced dependence on fruit development, whereas DFR1 expression remained relatively stable. There were, however, significant differences in their relative rates of expression. The DFR1/DFR2 expression ratio was much higher in the Fragaria×ananassa and enzyme preparations from F.×ananassa receptacles showed higher capability to convert DHK than preparations from F. vesca. Anthocyanin concentrations in the F.×ananassa cultivar were more than twofold higher and the cyanidin:pelargonidin ratio was only 0.05 compared to 0.51 in the F. vesca cultivar. The differences in the fruit colour of the two Fragaria species can be explained by the higher expression of DFR1 in F.×ananassa as compared to F. vesca, a higher enzyme efficiency (Kcat/Km values) of DFR1 combined with the loss of F3'H activity late in fruit development of F.×ananassa.

  14. Dihydroflavonol 4-reductase genes encode enzymes with contrasting substrate specificity and show divergent gene expression profiles in Fragaria species.

    PubMed

    Miosic, Silvija; Thill, Jana; Milosevic, Malvina; Gosch, Christian; Pober, Sabrina; Molitor, Christian; Ejaz, Shaghef; Rompel, Annette; Stich, Karl; Halbwirth, Heidi

    2014-01-01

    During fruit ripening, strawberries show distinct changes in the flavonoid classes that accumulate, switching from the formation of flavan 3-ols and flavonols in unripe fruits to the accumulation of anthocyanins in the ripe fruits. In the common garden strawberry (Fragaria×ananassa) this is accompanied by a distinct switch in the pattern of hydroxylation demonstrated by the almost exclusive accumulation of pelargonidin based pigments. In Fragaria vesca the proportion of anthocyanins showing one (pelargonidin) and two (cyanidin) hydroxyl groups within the B-ring is almost equal. We isolated two dihydroflavonol 4-reductase (DFR) cDNA clones from strawberry fruits, which show 82% sequence similarity. The encoded enzymes revealed a high variability in substrate specificity. One enzyme variant did not accept DHK (with one hydroxyl group present in the B-ring), whereas the other strongly preferred DHK as a substrate. This appears to be an uncharacterized DFR variant with novel substrate specificity. Both DFRs were expressed in the receptacle and the achenes of both Fragaria species and the DFR2 expression profile showed a pronounced dependence on fruit development, whereas DFR1 expression remained relatively stable. There were, however, significant differences in their relative rates of expression. The DFR1/DFR2 expression ratio was much higher in the Fragaria×ananassa and enzyme preparations from F.×ananassa receptacles showed higher capability to convert DHK than preparations from F. vesca. Anthocyanin concentrations in the F.×ananassa cultivar were more than twofold higher and the cyanidin:pelargonidin ratio was only 0.05 compared to 0.51 in the F. vesca cultivar. The differences in the fruit colour of the two Fragaria species can be explained by the higher expression of DFR1 in F.×ananassa as compared to F. vesca, a higher enzyme efficiency (Kcat/Km values) of DFR1 combined with the loss of F3'H activity late in fruit development of F.×ananassa. PMID:25393679

  15. Isolation of dihydroflavonol 4-reductase cDNA clones from Angelonia x angustifolia and heterologous expression as GST fusion protein in Escherichia coli.

    PubMed

    Gosch, Christian; Nagesh, Karthik Mudigere; Thill, Jana; Miosic, Silvija; Plaschil, Sylvia; Milosevic, Malvina; Olbricht, Klaus; Ejaz, Shaghef; Rompel, Annette; Stich, Karl; Halbwirth, Heidi

    2014-01-01

    Blue Angelonia × angustifolia flowers can show spontaneous mutations resulting in white/blue and white flower colourations. In such a white line, a loss of dihydroflavonol 4-reductase (DFR) activity was observed whereas chalcone synthase and flavanone 3-hydroxylase activity remained unchanged. Thus, cloning and characterization of a DFR of Angelonia flowers was carried out for the first time. Two full length DFR cDNA clones, Ang.DFR1 and Ang.DFR2, were obtained from a diploid chimeral white/blue Angelonia × angustifolia which demonstrated a 99% identity in their translated amino acid sequence. In comparison to Ang.DFR2, Ang.DFR1 was shown to contain an extra proline in a proline-rich region at the N-terminus along with two exchanges at the amino acids 12 and 26 in the translated amino acid sequence. The recombinant Ang.DFR2 obtained by heterologous expression in yeast was functionally active catalyzing the NADPH dependent reduction of dihydroquercetin (DHQ) and dihydromyricetin (DHM) to leucocyanidin and leucomyricetin, respectively. Dihydrokaempferol (DHK) in contrast was not accepted as a substrate despite the presence of asparagine in a position assumed to determine DHK acceptance. We show that substrate acceptance testing of DFRs provides biased results for DHM conversion if products are extracted with ethyl acetate. Recombinant Ang.DFR1 was inactive and functional activity could only be restored via exchanges of the amino acids in position 12 and 26 as well as the deletion of the extra proline. E. coli transformation of the pGEX-6P-1 vector harbouring the Ang.DFR2 and heterologous expression in E. coli resulted in functionally active enzymes before and after GST tag removal. Both the GST fusion protein and purified DFR minus the GST tag could be stored at -80°C for several months without loss of enzyme activity and demonstrated identical substrate specificity as the recombinant enzyme obtained from heterologous expression in yeast.

  16. The Balance of Expression of Dihydroflavonol 4-reductase and Flavonol Synthase Regulates Flavonoid Biosynthesis and Red Foliage Coloration in Crabapples.

    PubMed

    Tian, Ji; Han, Zhen-yun; Zhang, Jie; Hu, YuJing; Song, Tingting; Yao, Yuncong

    2015-01-01

    Red leaf color is an attractive trait of Malus families, including crabapple (Malus spp.); however, little is known about the molecular mechanisms that regulate the coloration. Dihydroflavonols are intermediates in the production of both colored anthocyanins and colorless flavonols, and this current study focused on the gene expression balance involved in the relative accumulation of these compounds in crabapple leaves. Levels of anthocyanins and the transcript abundances of the anthocyanin biosynthetic gene, dihydroflavonol 4-reductase (McDFR) and the flavonol biosynthetic gene, flavonol synthase (McFLS), were assessed during the leaf development in two crabapple cultivars, 'Royalty' and 'Flame'. The concentrations of anthocyanins and flavonols correlated with leaf color and we propose that the expression of McDFR and McFLS influences their accumulation. Further studies showed that overexpression of McDFR, or silencing of McFLS, increased anthocyanin production, resulting in red-leaf and red fruit peel phenotypes. Conversely, elevated flavonol production and green phenotypes in crabapple leaves and apple peel were observed when McFLS was overexpressed or McDFR was silenced. These results suggest that the relative activities of McDFR and McFLS are important determinants of the red color of crabapple leaves, via the regulation of the metabolic fate of substrates that these enzymes have in common. PMID:26192267

  17. The Balance of Expression of Dihydroflavonol 4-reductase and Flavonol Synthase Regulates Flavonoid Biosynthesis and Red Foliage Coloration in Crabapples

    PubMed Central

    Tian, Ji; Han, Zhen-yun; Zhang, Jie; Hu, YuJing; Song, Tingting; Yao, Yuncong

    2015-01-01

    Red leaf color is an attractive trait of Malus families, including crabapple (Malus spp.); however, little is known about the molecular mechanisms that regulate the coloration. Dihydroflavonols are intermediates in the production of both colored anthocyanins and colorless flavonols, and this current study focused on the gene expression balance involved in the relative accumulation of these compounds in crabapple leaves. Levels of anthocyanins and the transcript abundances of the anthocyanin biosynthetic gene, dihydroflavonol 4-reductase (McDFR) and the flavonol biosynthetic gene, flavonol synthase (McFLS), were assessed during the leaf development in two crabapple cultivars, ‘Royalty’ and ‘Flame’. The concentrations of anthocyanins and flavonols correlated with leaf color and we propose that the expression of McDFR and McFLS influences their accumulation. Further studies showed that overexpression of McDFR, or silencing of McFLS, increased anthocyanin production, resulting in red-leaf and red fruit peel phenotypes. Conversely, elevated flavonol production and green phenotypes in crabapple leaves and apple peel were observed when McFLS was overexpressed or McDFR was silenced. These results suggest that the relative activities of McDFR and McFLS are important determinants of the red color of crabapple leaves, via the regulation of the metabolic fate of substrates that these enzymes have in common. PMID:26192267

  18. Disequilibrium of Flavonol Synthase and Dihydroflavonol-4-Reductase Expression Associated Tightly to White vs. Red Color Flower Formation in Plants

    PubMed Central

    Luo, Ping; Ning, Guogui; Wang, Zhen; Shen, Yuxiao; Jin, Huanan; Li, Penghui; Huang, Shasha; Zhao, Jian; Bao, Manzhu

    2016-01-01

    Flower color is the main character throughout the plant kingdom. Though substantial information exists regarding the structural and regulatory genes involved in anthocyanin and flavonol biosynthesis, little is known that what make a diverse white vs. red color flower in natural species. Here, the contents of pigments in seven species from varied phylogenetic location in plants with red and white flowers were determined. Flavonols could be detected in red and white flowers, but anthocyanins were almost undetectable in the white cultivar. Comparisons of expression patterns of gene related to the flavonoid biosynthesis indicated that disequilibrium expression of flavonol synthase (FLS) and dihydroflavonol-4-reductase (DFR) genes determined the accumulation of flavonols and anothcyanins in both red and white flowers of seven species. To further investigate the role of such common regulatory patterns in determining flower color, FLS genes were isolated from Rosa rugosa (RrFLS1), Prunus persica (PpFLS), and Petunia hybrida (PhFLS), and DFR genes were isolated from Rosa rugosa (RrDFR1) and Petunia hybrida (PhDFR). Heterologous expression of the FLS genes within tobacco host plants demonstrated conservation of function, with the transgenes promoting flavonol biosynthesis and inhibiting anthocyanin accumulation, so resulting in white flowers. Conversely, overexpression of DFR genes in tobacco displayed down-regulation of the endogenous NtFLS gene, and the promotion of anthocyanin synthesis. On this basis, we propose a model in which FLS and DFR gene-products compete for common substrates in order to direct the biosynthesis of flavonols and anthocyanins, respectively, thereby determining white vs. red coloration of flowers. PMID:26793227

  19. Disequilibrium of Flavonol Synthase and Dihydroflavonol-4-Reductase Expression Associated Tightly to White vs. Red Color Flower Formation in Plants.

    PubMed

    Luo, Ping; Ning, Guogui; Wang, Zhen; Shen, Yuxiao; Jin, Huanan; Li, Penghui; Huang, Shasha; Zhao, Jian; Bao, Manzhu

    2015-01-01

    Flower color is the main character throughout the plant kingdom. Though substantial information exists regarding the structural and regulatory genes involved in anthocyanin and flavonol biosynthesis, little is known that what make a diverse white vs. red color flower in natural species. Here, the contents of pigments in seven species from varied phylogenetic location in plants with red and white flowers were determined. Flavonols could be detected in red and white flowers, but anthocyanins were almost undetectable in the white cultivar. Comparisons of expression patterns of gene related to the flavonoid biosynthesis indicated that disequilibrium expression of flavonol synthase (FLS) and dihydroflavonol-4-reductase (DFR) genes determined the accumulation of flavonols and anothcyanins in both red and white flowers of seven species. To further investigate the role of such common regulatory patterns in determining flower color, FLS genes were isolated from Rosa rugosa (RrFLS1), Prunus persica (PpFLS), and Petunia hybrida (PhFLS), and DFR genes were isolated from Rosa rugosa (RrDFR1) and Petunia hybrida (PhDFR). Heterologous expression of the FLS genes within tobacco host plants demonstrated conservation of function, with the transgenes promoting flavonol biosynthesis and inhibiting anthocyanin accumulation, so resulting in white flowers. Conversely, overexpression of DFR genes in tobacco displayed down-regulation of the endogenous NtFLS gene, and the promotion of anthocyanin synthesis. On this basis, we propose a model in which FLS and DFR gene-products compete for common substrates in order to direct the biosynthesis of flavonols and anthocyanins, respectively, thereby determining white vs. red coloration of flowers. PMID:26793227

  20. Molecular cloning, substrate specificity of the functionally expressed dihydroflavonol 4-reductases from Malus domestica and Pyrus communis cultivars and the consequences for flavonoid metabolism.

    PubMed

    Fischer, Thilo C; Halbwirth, Heidrun; Meisel, Barbara; Stich, Karl; Forkmann, Gert

    2003-04-15

    Treatment with the dioxygenase inhibitor prohexadione-Ca leads to major changes in the flavonoid metabolism of apple (Malus domestica) and pear (Pyrus communis) leaves. Accumulation of unusual 3-deoxyflavonoids is observed, which have been linked to an enhanced resistance toward fire blight. The committed step in this pathway is the reduction of flavanones. Crude extracts from leaves are able to perform this reaction. There was previous evidence that DFR enzymes of certain plants possess additional flavanone 4-reductase (FNR) activity. Such an FNR activity of DFR enzymes is proved here by heterologous expression of the enzymes. The heterologously expressed DFR/FNR enzymes of Malus and Pyrus possess distinct differences in substrate specificities despite only minor differences of the amino acid sequences. Kinetic studies showed that dihydroflavonols generally are the preferred substrates. However, with the observed substrate specificities the occurrence of 3-deoxyflavonoids in vivo after application of prohexadione-Ca can be explained.

  1. Isolation of Dihydroflavonol 4-Reductase cDNA Clones from Angelonia x angustifolia and Heterologous Expression as GST Fusion Protein in Escherichia coli

    PubMed Central

    Gosch, Christian; Nagesh, Karthik Mudigere; Thill, Jana; Miosic, Silvija; Plaschil, Sylvia; Milosevic, Malvina; Olbricht, Klaus; Ejaz, Shaghef; Rompel, Annette; Stich, Karl; Halbwirth, Heidi

    2014-01-01

    Blue Angelonia × angustifolia flowers can show spontaneous mutations resulting in white/blue and white flower colourations. In such a white line, a loss of dihydroflavonol 4-reductase (DFR) activity was observed whereas chalcone synthase and flavanone 3-hydroxylase activity remained unchanged. Thus, cloning and characterization of a DFR of Angelonia flowers was carried out for the first time. Two full length DFR cDNA clones, Ang.DFR1 and Ang.DFR2, were obtained from a diploid chimeral white/blue Angelonia × angustifolia which demonstrated a 99% identity in their translated amino acid sequence. In comparison to Ang.DFR2, Ang.DFR1 was shown to contain an extra proline in a proline-rich region at the N-terminus along with two exchanges at the amino acids 12 and 26 in the translated amino acid sequence. The recombinant Ang.DFR2 obtained by heterologous expression in yeast was functionally active catalyzing the NADPH dependent reduction of dihydroquercetin (DHQ) and dihydromyricetin (DHM) to leucocyanidin and leucomyricetin, respectively. Dihydrokaempferol (DHK) in contrast was not accepted as a substrate despite the presence of asparagine in a position assumed to determine DHK acceptance. We show that substrate acceptance testing of DFRs provides biased results for DHM conversion if products are extracted with ethyl acetate. Recombinant Ang.DFR1 was inactive and functional activity could only be restored via exchanges of the amino acids in position 12 and 26 as well as the deletion of the extra proline. E. coli transformation of the pGEX-6P-1 vector harbouring the Ang.DFR2 and heterologous expression in E. coli resulted in functionally active enzymes before and after GST tag removal. Both the GST fusion protein and purified DFR minus the GST tag could be stored at −80°C for several months without loss of enzyme activity and demonstrated identical substrate specificity as the recombinant enzyme obtained from heterologous expression in yeast. PMID:25238248

  2. Functional Characterization of Dihydroflavonol-4-Reductase in Anthocyanin Biosynthesis of Purple Sweet Potato Underlies the Direct Evidence of Anthocyanins Function against Abiotic Stresses

    PubMed Central

    Wang, Hongxia; Fan, Weijuan; Li, Hong; Yang, Jun; Huang, Jirong; Zhang, Peng

    2013-01-01

    Dihydroflavonol-4-reductase (DFR) is a key enzyme in the catalysis of the stereospecific reduction of dihydroflavonols to leucoanthocyanidins in anthocyanin biosynthesis. In the purple sweet potato (Ipomoea batatas Lam.) cv. Ayamurasaki, expression of the IbDFR gene was strongly associated with anthocyanin accumulation in leaves, stems and roots. Overexpression of the IbDFR in Arabidopsis tt3 mutants fully complemented the pigmentation phenotype of the seed coat, cotyledon and hypocotyl. Downregulation of IbDFR expression in transgenic sweet potato (DFRi) using an RNAi approach dramatically reduced anthocyanin accumulation in young leaves, stems and storage roots. In contrast, the increase of flavonols quercetin-3-O-hexose-hexoside and quercetin-3-O-glucoside in the leaves and roots of DFRi plants is significant. Therefore, the metabolic pathway channeled greater flavonol influx in the DFRi plants when their anthocyanin and proanthocyanidin accumulation were decreased. These plants also displayed reduced antioxidant capacity compared to the wild type. After 24 h of cold treatment and 2 h recovery, the wild-type plants were almost fully restored to the initial phenotype compared to the slower recovery of DFRi plants, in which the levels of electrolyte leakage and hydrogen peroxide accumulation were dramatically increased. These results provide direct evidence of anthocyanins function in the protection against oxidative stress in the sweet potato. The molecular characterization of the IbDFR gene in the sweet potato not only confirms its important roles in flavonoid metabolism but also supports the protective function of anthocyanins of enhanced scavenging of reactive oxygen radicals in plants under stressful conditions. PMID:24223813

  3. Functional characterization of Dihydroflavonol-4-reductase in anthocyanin biosynthesis of purple sweet potato underlies the direct evidence of anthocyanins function against abiotic stresses.

    PubMed

    Wang, Hongxia; Fan, Weijuan; Li, Hong; Yang, Jun; Huang, Jirong; Zhang, Peng

    2013-01-01

    Dihydroflavonol-4-reductase (DFR) is a key enzyme in the catalysis of the stereospecific reduction of dihydroflavonols to leucoanthocyanidins in anthocyanin biosynthesis. In the purple sweet potato (Ipomoea batatas Lam.) cv. Ayamurasaki, expression of the IbDFR gene was strongly associated with anthocyanin accumulation in leaves, stems and roots. Overexpression of the IbDFR in Arabidopsis tt3 mutants fully complemented the pigmentation phenotype of the seed coat, cotyledon and hypocotyl. Downregulation of IbDFR expression in transgenic sweet potato (DFRi) using an RNAi approach dramatically reduced anthocyanin accumulation in young leaves, stems and storage roots. In contrast, the increase of flavonols quercetin-3-O-hexose-hexoside and quercetin-3-O-glucoside in the leaves and roots of DFRi plants is significant. Therefore, the metabolic pathway channeled greater flavonol influx in the DFRi plants when their anthocyanin and proanthocyanidin accumulation were decreased. These plants also displayed reduced antioxidant capacity compared to the wild type. After 24 h of cold treatment and 2 h recovery, the wild-type plants were almost fully restored to the initial phenotype compared to the slower recovery of DFRi plants, in which the levels of electrolyte leakage and hydrogen peroxide accumulation were dramatically increased. These results provide direct evidence of anthocyanins function in the protection against oxidative stress in the sweet potato. The molecular characterization of the IbDFR gene in the sweet potato not only confirms its important roles in flavonoid metabolism but also supports the protective function of anthocyanins of enhanced scavenging of reactive oxygen radicals in plants under stressful conditions. PMID:24223813

  4. The Gene Encoding Dihydroflavonol 4-Reductase Is a Candidate for the anthocyaninless Locus of Rapid Cycling Brassica rapa (Fast Plants Type)

    PubMed Central

    Wendell, Douglas L.; Vaziri, Anoumid; Shergill, Gurbaksh

    2016-01-01

    Rapid cycling Brassica rapa, also known as Wisconsin Fast Plants, are a widely used organism in both K-12 and college science education. They are an excellent system for genetics laboratory instruction because it is very easy to conduct genetic crosses with this organism, there are numerous seed stocks with variation in both Mendelian and quantitative traits, they have a short generation time, and there is a wealth of educational materials for instructors using them. Their main deficiency for genetics education is that none of the genetic variation in RCBr has yet been characterized at the molecular level. Here we present the first molecular characterization of a gene responsible for a trait in Fast Plants. The trait under study is purple/nonpurple variation due to the anthocyaninless locus, which is one of the Mendelian traits most frequently used for genetics education with this organism. We present evidence that the DFR gene, which encodes dihyroflavonol 4-reductase, is the candidate gene for the anthocyaninless (ANL) locus in RCBr. DFR shows complete linkage with ANL in genetic crosses with a total of 948 informative chromosomes, and strains with the recessive nonpurple phenotype have a transposon-related insertion in the DFR which is predicted to disrupt gene function. PMID:27548675

  5. The Gene Encoding Dihydroflavonol 4-Reductase Is a Candidate for the anthocyaninless Locus of Rapid Cycling Brassica rapa (Fast Plants Type).

    PubMed

    Wendell, Douglas L; Vaziri, Anoumid; Shergill, Gurbaksh

    2016-01-01

    Rapid cycling Brassica rapa, also known as Wisconsin Fast Plants, are a widely used organism in both K-12 and college science education. They are an excellent system for genetics laboratory instruction because it is very easy to conduct genetic crosses with this organism, there are numerous seed stocks with variation in both Mendelian and quantitative traits, they have a short generation time, and there is a wealth of educational materials for instructors using them. Their main deficiency for genetics education is that none of the genetic variation in RCBr has yet been characterized at the molecular level. Here we present the first molecular characterization of a gene responsible for a trait in Fast Plants. The trait under study is purple/nonpurple variation due to the anthocyaninless locus, which is one of the Mendelian traits most frequently used for genetics education with this organism. We present evidence that the DFR gene, which encodes dihyroflavonol 4-reductase, is the candidate gene for the anthocyaninless (ANL) locus in RCBr. DFR shows complete linkage with ANL in genetic crosses with a total of 948 informative chromosomes, and strains with the recessive nonpurple phenotype have a transposon-related insertion in the DFR which is predicted to disrupt gene function. PMID:27548675

  6. The Gene Encoding Dihydroflavonol 4-Reductase Is a Candidate for the anthocyaninless Locus of Rapid Cycling Brassica rapa (Fast Plants Type).

    PubMed

    Wendell, Douglas L; Vaziri, Anoumid; Shergill, Gurbaksh

    2016-01-01

    Rapid cycling Brassica rapa, also known as Wisconsin Fast Plants, are a widely used organism in both K-12 and college science education. They are an excellent system for genetics laboratory instruction because it is very easy to conduct genetic crosses with this organism, there are numerous seed stocks with variation in both Mendelian and quantitative traits, they have a short generation time, and there is a wealth of educational materials for instructors using them. Their main deficiency for genetics education is that none of the genetic variation in RCBr has yet been characterized at the molecular level. Here we present the first molecular characterization of a gene responsible for a trait in Fast Plants. The trait under study is purple/nonpurple variation due to the anthocyaninless locus, which is one of the Mendelian traits most frequently used for genetics education with this organism. We present evidence that the DFR gene, which encodes dihyroflavonol 4-reductase, is the candidate gene for the anthocyaninless (ANL) locus in RCBr. DFR shows complete linkage with ANL in genetic crosses with a total of 948 informative chromosomes, and strains with the recessive nonpurple phenotype have a transposon-related insertion in the DFR which is predicted to disrupt gene function.

  7. Estrogenic activity of naturally occurring anthocyanidins.

    PubMed

    Schmitt, E; Stopper, H

    2001-01-01

    Anthocyanins, which are natural plant pigments from the flavonoid family, represent substantial constituents of the human diet. Because some other bioflavonoids are known to have estrogenic activity, the aim of this study was to determine the estrogenic activity of the anthocyanine aglycones. Binding affinity to the estrogen receptor-alpha was 10,000- to 20,000-fold lower than that of the endogenous estrogen estradiol. In the estrogen receptor-positive cell line MCF-7, the anthocyanidins induced expression of a reporter gene. The tested anthocyanidins showed estrogen-inducible cell proliferation in two cell lines (MCF-7 and BG-1), but not in the receptor-negative human breast cancer cell line MDA-MB-231. The phytoestrogen-induced cell proliferation could be blocked by addition of the receptor antagonist 4-hydroxytamoxifen. Combination treatments with the endogenous estrogen estradiol resulted in a reduction of estradiol-induced cell proliferation. Overall, the tested anthocyanidins exert estrogenic activity, which might play a role in altering the development of hormone-dependent adverse effects.

  8. The miR156-SPL9-DFR pathway coordinates the relationship between development and abiotic stress tolerance in plants.

    PubMed

    Cui, Long-Gang; Shan, Jun-Xiang; Shi, Min; Gao, Ji-Ping; Lin, Hong-Xuan

    2014-12-01

    Young organisms have relatively strong resistance to diseases and adverse conditions. When confronted with adversity, the process of development is delayed in plants. This phenomenon is thought to result from the rebalancing of energy, which helps plants to coordinate the relationship between development and stress tolerance; however, the molecular mechanism underlying this phenomenon remains mysterious. In this study, we found that miR156 integrates environmental signals to ensure timely flowering, thus enabling the completion of breeding. Under stress conditions, miR156 is induced to maintain the plant in the juvenile state for a relatively long period of time, whereas under favorable conditions, miR156 is suppressed to accelerate the developmental transition. Blocking the miR156 signaling pathway in Arabidopsis thaliana with 35S::MIM156 (via target mimicry) increased the sensitivity of the plant to stress treatment, whereas overexpression of miR156 increased stress tolerance. In fact, this mechanism is also conserved in Oryza sativa (rice). We also identified downstream genes of miR156, i.e. SQUAMOSA PROMOTER BINDING PROTEIN-LIKE 9 (SPL9) and DIHYDROFLAVONOL-4-REDUCTASE (DFR), which take part in this process by influencing the metabolism of anthocyanin. Our results uncover a molecular mechanism for plant adaptation to the environment through the miR156-SPLs-DFR pathway, which coordinates development and abiotic stress tolerance. PMID:25345491

  9. The miR156-SPL9-DFR pathway coordinates the relationship between development and abiotic stress tolerance in plants.

    PubMed

    Cui, Long-Gang; Shan, Jun-Xiang; Shi, Min; Gao, Ji-Ping; Lin, Hong-Xuan

    2014-12-01

    Young organisms have relatively strong resistance to diseases and adverse conditions. When confronted with adversity, the process of development is delayed in plants. This phenomenon is thought to result from the rebalancing of energy, which helps plants to coordinate the relationship between development and stress tolerance; however, the molecular mechanism underlying this phenomenon remains mysterious. In this study, we found that miR156 integrates environmental signals to ensure timely flowering, thus enabling the completion of breeding. Under stress conditions, miR156 is induced to maintain the plant in the juvenile state for a relatively long period of time, whereas under favorable conditions, miR156 is suppressed to accelerate the developmental transition. Blocking the miR156 signaling pathway in Arabidopsis thaliana with 35S::MIM156 (via target mimicry) increased the sensitivity of the plant to stress treatment, whereas overexpression of miR156 increased stress tolerance. In fact, this mechanism is also conserved in Oryza sativa (rice). We also identified downstream genes of miR156, i.e. SQUAMOSA PROMOTER BINDING PROTEIN-LIKE 9 (SPL9) and DIHYDROFLAVONOL-4-REDUCTASE (DFR), which take part in this process by influencing the metabolism of anthocyanin. Our results uncover a molecular mechanism for plant adaptation to the environment through the miR156-SPLs-DFR pathway, which coordinates development and abiotic stress tolerance.

  10. Surface-enhanced Raman scattering (SERS) study of anthocyanidins

    NASA Astrophysics Data System (ADS)

    Zaffino, Chiara; Russo, Bianca; Bruni, Silvia

    2015-10-01

    Anthocyanins are an important class of natural compounds responsible for the red, purple and blue colors in a large number of flowers, fruits and cereal grains. They are polyhydroxy- and polymethoxy-derivatives of 2-phenylbenzopyrylium (flavylium) salts, which are present in nature as glycosylated molecules. The aim of the present study is to assess the identification of anthocyanidins, i.e. anthocyanins without the glycosidic moiety, by means of surface-enhanced Raman spectroscopy (SERS), a very chemically-specific technique which is moreover sensitive to subtle changes in molecular structures. These features can lead to elect SERS, among the spectroscopic tools currently at disposal of scientists, as a technique of choice for the identification of anthocyanidins, since: (1) anthocyanidins structurally present the same benzopyrylium moiety and differentiate only for the substitution pattern on their phenyl ring, (2) different species are present in aqueous solution depending on the pH. It will be demonstrated that, while resonance Raman spectra of anthocyanidins are very similar to one another, SER spectra show greater differences, leading to a further step in the identification of such important compounds in diluted solutions by means of vibrational spectroscopy. Moreover, the dependence on the pH of the six most common anthocyanidins, i.e. cyanidin, delphinidin, pelargonidin, peonidin, malvidin and petunidin, is studied. To the best of the authors' knowledge, a complete SERS study of such important molecules is reported in the present work for the first time.

  11. Emergence of Plasmid-Borne dfrA14 Trimethoprim Resistance Gene in Shigella sonnei.

    PubMed

    Miranda, Alfonso; Ávila, Bárbara; Díaz, Patricia; Rivas, Lina; Bravo, Karen; Astudillo, Javier; Bueno, Constanza; Ulloa, María T; Hermosilla, Germán; Del Canto, Felipe; Salazar, Juan C; Toro, Cecilia S

    2016-01-01

    The most common mechanism of trimethoprim (TMP)-resistance is the acquisition of dihydrofolate reductase enzyme resistant to this drug. Previous molecular characterization of TMP-genes resistance in Chilean isolates of Shigella sonnei searching for dfrA1 and dfrA8, showed solely the presence of dfrA8 (formerly dhfrIIIc). However, these genetic markers were absent in S. sonnei strains further isolated during an outbreak in 2009. To identify the TMP-resistance gene in these strains, a genomic DNA library from a TMP-resistant (TMP(R)) S. sonnei representative strain for the outbreak was used to clone, select and identify a TMP-resistance marker. The TMP(R) clone was sequenced by primer walking, identifying the presence of the dfrA14 gene in the sul2-strA'-dfrA14-'strA-strB gene arrangement, harbored in a native 6779-bp plasmid. The same plasmid was isolated by transforming with a ~4.2 MDa plasmid extracted from several TMP(R) S. sonnei strains into Escherichia coli. This plasmid, named pABC-3, was present only in dfrA14-positive strains and was homologous to a previously described pCERC-1, but different due to the absence of an 11-bp repetitive unit. The distribution of dfrA1, dfrA8, and dfrA14 TMP-resistance genes was determined in 126 TMP(R) S. sonnei isolates. Most of the strains (96%) carried only one of the three TMP-resistance genes assessed. Thus, all strains obtained during the 2009-outbreak harbored only dfrA14, whereas, dfrA8 was the most abundant gene marker before outbreak and, after the outbreak dfrA1 seems have appeared in circulating strains. According to PFGE, dfrA14-positive strains were clustered in a genetically related group including some dfrA1- and dfrA8-positive strains; meanwhile other genetic group included most of the dfrA8-positive strains. This distribution also correlated with the isolation period, showing a dynamics of trimethoprim genetic markers prevalent in Chilean S. sonnei strains. To our knowledge, dfrA14 gene associated to a small

  12. Emergence of Plasmid-Borne dfrA14 Trimethoprim Resistance Gene in Shigella sonnei

    PubMed Central

    Miranda, Alfonso; Ávila, Bárbara; Díaz, Patricia; Rivas, Lina; Bravo, Karen; Astudillo, Javier; Bueno, Constanza; Ulloa, María T.; Hermosilla, Germán; Del Canto, Felipe; Salazar, Juan C.; Toro, Cecilia S.

    2016-01-01

    The most common mechanism of trimethoprim (TMP)-resistance is the acquisition of dihydrofolate reductase enzyme resistant to this drug. Previous molecular characterization of TMP-genes resistance in Chilean isolates of Shigella sonnei searching for dfrA1 and dfrA8, showed solely the presence of dfrA8 (formerly dhfrIIIc). However, these genetic markers were absent in S. sonnei strains further isolated during an outbreak in 2009. To identify the TMP-resistance gene in these strains, a genomic DNA library from a TMP-resistant (TMPR) S. sonnei representative strain for the outbreak was used to clone, select and identify a TMP-resistance marker. The TMPR clone was sequenced by primer walking, identifying the presence of the dfrA14 gene in the sul2-strA'-dfrA14-‘strA-strB gene arrangement, harbored in a native 6779-bp plasmid. The same plasmid was isolated by transforming with a ~4.2 MDa plasmid extracted from several TMPR S. sonnei strains into Escherichia coli. This plasmid, named pABC-3, was present only in dfrA14-positive strains and was homologous to a previously described pCERC-1, but different due to the absence of an 11-bp repetitive unit. The distribution of dfrA1, dfrA8, and dfrA14 TMP-resistance genes was determined in 126 TMPR S. sonnei isolates. Most of the strains (96%) carried only one of the three TMP-resistance genes assessed. Thus, all strains obtained during the 2009-outbreak harbored only dfrA14, whereas, dfrA8 was the most abundant gene marker before outbreak and, after the outbreak dfrA1 seems have appeared in circulating strains. According to PFGE, dfrA14-positive strains were clustered in a genetically related group including some dfrA1- and dfrA8-positive strains; meanwhile other genetic group included most of the dfrA8-positive strains. This distribution also correlated with the isolation period, showing a dynamics of trimethoprim genetic markers prevalent in Chilean S. sonnei strains. To our knowledge, dfrA14 gene associated to a small non

  13. Emergence of Plasmid-Borne dfrA14 Trimethoprim Resistance Gene in Shigella sonnei.

    PubMed

    Miranda, Alfonso; Ávila, Bárbara; Díaz, Patricia; Rivas, Lina; Bravo, Karen; Astudillo, Javier; Bueno, Constanza; Ulloa, María T; Hermosilla, Germán; Del Canto, Felipe; Salazar, Juan C; Toro, Cecilia S

    2016-01-01

    The most common mechanism of trimethoprim (TMP)-resistance is the acquisition of dihydrofolate reductase enzyme resistant to this drug. Previous molecular characterization of TMP-genes resistance in Chilean isolates of Shigella sonnei searching for dfrA1 and dfrA8, showed solely the presence of dfrA8 (formerly dhfrIIIc). However, these genetic markers were absent in S. sonnei strains further isolated during an outbreak in 2009. To identify the TMP-resistance gene in these strains, a genomic DNA library from a TMP-resistant (TMP(R)) S. sonnei representative strain for the outbreak was used to clone, select and identify a TMP-resistance marker. The TMP(R) clone was sequenced by primer walking, identifying the presence of the dfrA14 gene in the sul2-strA'-dfrA14-'strA-strB gene arrangement, harbored in a native 6779-bp plasmid. The same plasmid was isolated by transforming with a ~4.2 MDa plasmid extracted from several TMP(R) S. sonnei strains into Escherichia coli. This plasmid, named pABC-3, was present only in dfrA14-positive strains and was homologous to a previously described pCERC-1, but different due to the absence of an 11-bp repetitive unit. The distribution of dfrA1, dfrA8, and dfrA14 TMP-resistance genes was determined in 126 TMP(R) S. sonnei isolates. Most of the strains (96%) carried only one of the three TMP-resistance genes assessed. Thus, all strains obtained during the 2009-outbreak harbored only dfrA14, whereas, dfrA8 was the most abundant gene marker before outbreak and, after the outbreak dfrA1 seems have appeared in circulating strains. According to PFGE, dfrA14-positive strains were clustered in a genetically related group including some dfrA1- and dfrA8-positive strains; meanwhile other genetic group included most of the dfrA8-positive strains. This distribution also correlated with the isolation period, showing a dynamics of trimethoprim genetic markers prevalent in Chilean S. sonnei strains. To our knowledge, dfrA14 gene associated to a small

  14. In vitro effects of anthocyanidins on sinonasal epithelial nitric oxide production and bacterial physiology

    PubMed Central

    Hariri, Benjamin M.; Payne, Sakeena J.; Chen, Bei; Mansfield, Corrine; Doghramji, Laurel J.; Adappa, Nithin D.; Palmer, James N.; Kennedy, David W.; Niv, Masha Y.

    2016-01-01

    Background: T2R bitter taste receptors play a crucial role in sinonasal innate immunity by upregulating mucociliary clearance and nitric oxide (NO) production in response to bitter gram-negative quorum-sensing molecules in the airway surface liquid. Previous studies showed that phytochemical flavonoid metabolites, known as anthocyanidins, taste bitter and have antibacterial effects. Our objectives were to examine the effects of anthocyanidins on NO production by human sinonasal epithelial cells and ciliary beat frequency, and their impact on common sinonasal pathogens Pseudomonas aeruginosa and Staphylococcus aureus. Methods: Ciliary beat frequency and NO production were measured by using digital imaging of differentiated air-liquid interface cultures prepared from primary human cells isolated from residual surgical material. Plate-based assays were used to determine the effects of anthocyanidins on bacterial swimming and swarming motility. Biofilm formation and planktonic growth were also assessed. Results: Anthocyanidin compounds triggered epithelial cells to produce NO but not through T2R receptors. However, anthocyanidins did not impact ciliary beat frequency. Furthermore, they did not reduce biofilm formation or planktonic growth of P. aeruginosa. In S. aureus, they did not reduce planktonic growth, and only one compound had minimal antibiofilm effects. The anthocyanidin delphinidin and anthocyanin keracyanin were found to promote bacterial swimming, whereas anthocyanidin cyanidin and flavonoid myricetin did not. No compounds that were tested inhibited bacterial swarming. Conclusion: Results of this study indicated that, although anthocyanidins may elicited an innate immune NO response from human cells, they do not cause an increase in ciliary beating and they may also cause a pathogenicity-enhancing effect in P. aeruginosa. Additional studies are necessary to understand how this would affect the use of anthocyanidins as therapeutics. This study emphasized the

  15. [Anthocyanidin inhibits immunoglobulin E-mediated allergic response in mast cells].

    PubMed

    Jin, Guang-Ri; Hong, Hai; Jin, Guang-Yu; Li, Ying-Zhe; Li, Guang-Zhao; Yan, Guang-Hai

    2012-01-01

    This study is to investigate the anti-allergic effect of anthocyanidin and to explore its possible mechanism. The experiments of passive cutaneous anaphylaxis reaction (PCA) and colorimetry were used to determine the effect of anthocyanidin on degranulation of mast cells in vivo. For in vitro study, various concentrations of anthocyanidin (100, 50 and 25 micromol x L(-1)) were added to the culture medium of mast cells cultured with 100 microg x L(-1) of dinitrophenyl (DNP) specific IgE overnight. The azelastine (100 micromol x L(-1)) was selected as the positive control. The antigen (DNP-human serum albumin, DNP-HAS)-induced release of degranulation was measured by enzymatic assay, histamine was determined by EIA, and interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-alpha) were measured by Western blotting, separately. In addition, the effects of anthocyanidin on phosphorylation of NF-kappaB, p38MAPK and Akt were observed by Western blotting. The results showed that treatments with anthocyanidin (100 and 50 mg x kg(-1)) were followed by a decrease in PCA of rats. Anthocyanidin (100 and 50 micromol x L(-1)) obviously suppressed the degranulation from mast cells, whereas results from anthocyanidin (100 and 50 micromol x L(-1)) group indicated significant inhibitory effect on histamine, the calcium uptake, TNF-alpha, IL-6, phosphorylation of NF-kappaB, p38MAPK and Akt of mast cells induced by antigen. Anthocyanidin may suppress the anaphylactic reaction by inhibiting the action of mast cells. NF-kappaB, p38MAPK and Akt at least in part contribute to this event. PMID:22493802

  16. Determination of anthocyanidins in berries and red wine by high-performance liquid chromatography.

    PubMed

    Nyman, N A; Kumpulainen, J T

    2001-09-01

    A high-performance liquid chromatographic (HPLC) method for the determination of anthocyanidins from berries and red wine is described. Delphinidin, cyanidin, petunidin, pelargonidin, peonidin, and malvidin contents of bilberry (Vaccinium myrtillus), black currant (Ribes nigrum), strawberry (Fragaria ananassa cv. Jonsok), and a Cabernet sauvignon (Vitis vinifera) red wine were determined. The aglycon forms of the anthocyanins present in the samples were revealed by acid hydrolysis. A reversed phase analytical column was employed to separate the anthocyanidins before identification by diode array detection. The suitability of the method was tested by determining the recovery (95-102% as aglycons and 69-104% from glycosides) for each anthocyanidin. Method repeatability was tested by charting the total aglycon content of two samples over a period of 14 analyses and determining the coefficients of variation (1.41% for bilberry and 2.56% for in-house reference material). The method developed proved thus to be effective for reliable determination of anthocyanidins from freeze-dried berry samples and red wine. The total anthocyanidin content of the tested samples was as follows: in-house reference material, 447 +/- 8 mg/100 g; strawberry, 23.8 +/- 0.4 mg/100 g; black currant, 135 +/- 3 mg/100 g; bilberry, 360 +/- 3 mg/100 g; and Cabernet sauvignon red wine, 26.1 +/- 0.1 mg/100 mL. PMID:11559107

  17. Predominance of dfrG as determinant of trimethoprim resistance in imported Staphylococcus aureus.

    PubMed

    Nurjadi, D; Schäfer, J; Friedrich-Jänicke, B; Mueller, A; Neumayr, A; Calvo-Cano, A; Goorhuis, A; Molhoek, N; Lagler, H; Kantele, A; Van Genderen, P J J; Gascon, J; Grobusch, M P; Caumes, E; Hatz, C; Fleck, R; Mockenhaupt, F P; Zanger, P

    2015-12-01

    To investigate the global occurrence of trimethoprim-sulfamethoxazole resistance and the genetic mechanisms of trimethoprim resistance, we analysed Staphylococcus aureus from travel-associated skin and soft-tissue infections treated at 13 travel clinics in Europe. Thirty-eight per cent (75/196) were trimethoprim-resistant and 21% (41/196) were resistant to trimethoprim-sulfamethoxazole. Among methicillin-resistant S. aureus, these proportions were 30% (7/23) and 17% (4/23), respectively. DfrG explained 92% (69/75) of all trimethoprim resistance in S. aureus. Travel to South Asia was associated with the highest risk of acquiring trimethoprim-sulfamethoxazole-resistant S. aureus. We conclude that globally dfrG is the predominant determinant of trimethoprim resistance in human S. aureus infection.

  18. Effect of anthocyanidins on myogenic differentiation in induced and non-induced primary myoblasts from rainbow trout (Oncorhynchus mykiss).

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A study was conducted to test whether an anthocyanidin mixture (peonidin, cyanidin and pelargonidin chloride) modulates myogenesis in both induced and non-induced myogenic cells from juvenile rainbow trout (Oncorhynchus mykiss). We evaluated three different anthocyanidin concentrations (1X, 2.5X and...

  19. Effect of anthocyanidins on myogenic differentiation in induced and non-induced primary myoblasts from rainbow trout (Oncorhynchus mykiss).

    PubMed

    Villasante, Alejandro; Powell, Madison S; Murdoch, Gordon K; Overturf, Ken; Cain, Kenneth; Wacyk, Jurij; Hardy, Ronald W

    2016-01-01

    A study was conducted to test whether an anthocyanidin mixture (peonidin, cyanidin and pelargonidin chloride) modulates myogenesis in both induced and non-induced myogenic cells from juvenile rainbow trout (Oncorhynchus mykiss). We evaluated three different anthocyanidin concentrations (1×, 2.5× and 10×) at two sampling times (24 and 36h). To test for treatment effects, we analyzed the expression of myoD and pax7 as well as two target genes of the Notch signaling pathway, hey2 and her6. In induced myogenic cells, the lowest and middle anthocyanidin doses caused significantly greater expression of myoD after 24h of treatment compared to control. A significantly higher expression of pax7 in cells exposed to either anthocyanidin treatment during 36h compared was observed. Similarly, the pax7/myoD ratio was significantly lower in cells exposed to the lowest anthocyanidin doses during 24h compared to control. No significant effect of anthocyanidin treatments on the expression of hey2 and her6 at either sampling point was detected. In non-induced cells, we observed no effect of anthocyanidins on myoD expression and significant down-regulation on pax7 expression in cells exposed to either anthocyanidin mixture concentrations after 24 and 36h of treatment compared to control. Further, the pax7/myoD ratio was significantly lower in cells exposed to either anthocyanidin doses at both sampling time. In non-induced cells, the highest anthocyanidin dose provoked significantly greater expression of hey2 after 24h of treatment compared to control. We detected no such effect in non-induced cells exposed to the lowest and middle anthocyanidin doses during 24h of treatment. The expression of her6 was unaffected by anthocyanidin treatments at either sampling time or doses compared to control. Collectively, these findings provide evidence that anthocyanidins modulate specific components of the myogenic programming in fish, thereby potentially affecting somatic growth in fish fed

  20. Effects of latitude-related factors and geographical origin on anthocyanidin concentrations in fruits of Vaccinium myrtillus L. (bilberries).

    PubMed

    Akerström, Andreas; Jaakola, Laura; Bång, Ulla; Jäderlund, Anders

    2010-11-24

    Two data sets are presented to identify the effect of growth location and origin of parental plant on anthocyanidin concentrations in Vaccinium myrtillus fruits. Bilberries were collected from wild populations growing at different latitudes and from cultivated plants originating from different geographical locations but grown in the same location for over 10 years. High-performance liquid chromatography analysis showed that anthocyanidin concentrations varied significantly with latitude and with geographical origin, with higher values form northern latitudes or from a more northerly origin of parent plants. The results show that anthocyanidin concentrations in bilberries are under strong genetic control but are also influenced by climatic factors. Furthermore, the proportions of specific anthocyanidins differed between latitudes and between plants with different parental origins. The diversity in anthocyanidin concentration and composition has important implications for plant breeders and for future development of varieties with high antioxidant capacity.

  1. Purification and characterization of (+)dihydroflavonol (3-hydroxyflavanone) 4-reductase from flowers of Dahlia variabilis.

    PubMed

    Fischer, D; Stich, K; Britsch, L; Grisebach, H

    1988-07-01

    Individual flowers from inflorescences of Dahlia variabilis (cv Scarlet Star) in young developmental stages contained relatively high activity of (+)-dihydroflavonol (DHF) 4-reductase. The DHF reductase was purified from such flowers to apparent homogeneity by a five-step procedure. This included affinity adsorption on Blue Sepharose and elution of the enzyme with NADP+. By gel filtration and by sodium dodecyl sulfate-polyacrylamide gel electrophoresis it was shown that DHF reductase contains only one polypeptide chain with a Mr of about 41,000. The reductase required NADPH as cofactor and catalyzed transfer of the pro-S hydrogen of NADPH to the substrate. Flavanones and dihydroflavonols (3-hydroxyflavanones) were substrates for DHF reductase with pH optima of about 6.0 for flavanones and of about 6.8 for dihydroflavonols. Flavanones were reduced to the corresponding flavan-4-ols and (+)-dihydroflavonols to flavan-3,4-cis-diols. Apparent Michaelis constants determined for (2S)-naringenin, (2S)-eriodicytol, (+)-dihydrokaempferol, (+)-dihydroquercetin, and NADPH were, respectively, 2.3, 2, 10, 15, and 42 microM. V/Km values were higher for dihydroflavonols than for flavanones. Conversion of dihydromyricetin to leucodelphinidin was also catalyzed by the enzyme at a low rate, whereas flavones and flavonols were not accepted as substrates. DHF reductase was not inhibited by metal chelators. PMID:3293532

  2. Anthocyanidins inhibit activator protein 1 activity and cell transformation: structure-activity relationship and molecular mechanisms.

    PubMed

    Hou, De-Xing; Kai, Keiko; Li, Jian-Jian; Lin, Shigang; Terahara, Norihiko; Wakamatsu, Mika; Fujii, Makoto; Young, Mattew R; Colburn, Nancy

    2004-01-01

    Anthocyanins are the chemical components that give the intense color to many fruits and vegetables, such as blueberries, red cabbages and purple sweet potatoes. Extensive studies have indicated that anthocyanins have strong antioxidant activities. To investigate the mechanism of anthocyanidins as an anticancer food source, six kinds of anthocyanidins representing the aglycons of most anthocyanins, were used to examine their effects on tumor promotion in mouse JB6 cells, a validated model for screening cancer chemopreventive agents and elucidating the molecular mechanisms. Of the six anthocyanins tested, only those with an ortho-dihydroxyphenyl structure on the B-ring suppressed 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced cell transformation and activator protein-1 transactivation, suggesting that the ortho-dihydroxyphenyl may contribute to the inhibitory action. Delphinidin, but not peonidin, blocked the phosphorylation of protein kinases in the extracellular signal-regulated protein kinase (ERK) pathway at early times and the c-Jun N-terminal kinase (JNK) signaling pathway at later times. p38 kinase was not inhibited by delphinidin. Furthermore, two mitogen-activated protein kinase (MAPK) specific inhibitors (SP600125 for JNK and UO126 for ERK) could specifically block the activation of JNK and ERK and cell transformation. Those results demonstrate that anthocyanidins contribute to the inhibition of tumorigenesis by blocking activation of the MAPK pathway. These findings provide the first molecular basis for the anticarcinogenic action of anthocyanidins. PMID:14514663

  3. Changes of anthocyanins, anthocyanidins, and antioxidant activity in bilberry extract during dry heating.

    PubMed

    Yue, X; Xu, Z

    2008-08-01

    Thermal stability of 10 anthocyanins found in a bilberry extract was studied at different heating temperatures and times. Degradation of the 10 anthocyanins, delphinidin, cyanidin, petunidin, peonidin, and malvidin derivat with different conjugated sugars, followed 1st-order reaction kinetics at heating temperatures 80, 100, and 125 degrees C. Though the degradation rate constants of anthocyanins were not significantly different from each other at the same heating temperature, they increased drastically when heating temperature was increased to 125 degrees C. At that temperature, the half-lives for all anthocyanins were less than 8 min. The degradation rate constants followed the Arrhenius equation. The trend of lower activation energy of the anthocyanins with arabinoside than with galactoside or glucoside was observed. These conjugated sugars were cleaved from the anthocyanins to produce their corresponding anthocyanidins or aglycones during heating. The production of anthocyanidins increased con stantly and was converted from approximately 30% of the degraded anthocyanins when heated at 100 degrees C for up to 30 min. At 125 degrees C, the increase of anthocyanidins lasted for 10 min, after which the degradation rate of anthocyanidins exceeded the production rate. Antioxidant activities of the heated extracts were estimated by measuring DPPH (2, 2'-diphenyl-1-picrylhydrazyl) free radical scavenging activity. The extracts heated at 80 degrees C for 30 min, 100 degrees C for 10 and 20 min, and 125 degrees C for 10 min had higher free radical scavenging capability than unheated extract.

  4. Anthocyanidins inhibit activator protein 1 activity and cell transformation: structure-activity relationship and molecular mechanisms.

    PubMed

    Hou, De-Xing; Kai, Keiko; Li, Jian-Jian; Lin, Shigang; Terahara, Norihiko; Wakamatsu, Mika; Fujii, Makoto; Young, Mattew R; Colburn, Nancy

    2004-01-01

    Anthocyanins are the chemical components that give the intense color to many fruits and vegetables, such as blueberries, red cabbages and purple sweet potatoes. Extensive studies have indicated that anthocyanins have strong antioxidant activities. To investigate the mechanism of anthocyanidins as an anticancer food source, six kinds of anthocyanidins representing the aglycons of most anthocyanins, were used to examine their effects on tumor promotion in mouse JB6 cells, a validated model for screening cancer chemopreventive agents and elucidating the molecular mechanisms. Of the six anthocyanins tested, only those with an ortho-dihydroxyphenyl structure on the B-ring suppressed 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced cell transformation and activator protein-1 transactivation, suggesting that the ortho-dihydroxyphenyl may contribute to the inhibitory action. Delphinidin, but not peonidin, blocked the phosphorylation of protein kinases in the extracellular signal-regulated protein kinase (ERK) pathway at early times and the c-Jun N-terminal kinase (JNK) signaling pathway at later times. p38 kinase was not inhibited by delphinidin. Furthermore, two mitogen-activated protein kinase (MAPK) specific inhibitors (SP600125 for JNK and UO126 for ERK) could specifically block the activation of JNK and ERK and cell transformation. Those results demonstrate that anthocyanidins contribute to the inhibition of tumorigenesis by blocking activation of the MAPK pathway. These findings provide the first molecular basis for the anticarcinogenic action of anthocyanidins.

  5. A Fruit-Specific Putative Dihydroflavonol 4-Reductase Gene Is Differentially Expressed in Strawberry during the Ripening Process1

    PubMed Central

    Moyano, Enriqueta; Portero-Robles, Ignacio; Medina-Escobar, Nieves; Valpuesta, Victoriano; Muñoz-Blanco, Juan; Luis Caballero, José

    1998-01-01

    A cDNA clone encoding a putative dihydroflavonol 4-reductase gene has been isolated from a strawberry (Fragaria × ananassa cv Chandler) DNA subtractive library. Northern analysis showed that the corresponding gene is predominantly expressed in fruit, where it is first detected during elongation (green stages) and then declines and sharply increases when the initial fruit ripening events occur, at the time of initiation of anthocyanin accumulation. The transcript can be induced in unripe green fruit by removing the achenes, and this induction can be partially inhibited by treatment of de-achened fruit with naphthylacetic acid, indicating that the expression of this gene is under hormonal control. We propose that the putative dihydroflavonol 4-reductase gene in strawberry plays a main role in the biosynthesis of anthocyanin during color development at the late stages of fruit ripening; during the first stages the expression of this gene could be related to the accumulation of condensed tannins. PMID:9625725

  6. Volatile, anthocyanidin, quality and sensory changes in rabbiteye blueberry from whole fruit through pilot plant juice processing.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    BACKGROUND: High antioxidant content and keen marketing have increased blueberry demand and increased local production which in turn mandates new uses for abundant harvests. Pilot scale processes were employed to investigate the anthocyanidin profiles, qualitative volatile compositions, and sensori...

  7. Contribution of dfrA and inhA mutations to the detection of isoniazid-resistant Mycobacterium tuberculosis isolates.

    PubMed

    Ho, Yu Min; Sun, Yong-Jiang; Wong, Sin-Yew; Lee, Ann S G

    2009-09-01

    Screening of 127 isoniazid (INH)-resistant Mycobacterium tuberculosis isolates from Singapore for mutations within the dfrA and inhA genes revealed mutations in 0 and 5 (3.9%) isolates respectively, implying that mutations in dfrA do not contribute to the detection of INH-resistant M. tuberculosis and that mutations within inhA are rare. Thirty-seven (29%) of the 127 isolates had no mutations in any of the genes implicated in INH resistance (katG, kasA, and ndh; inhA and ahpC promoters), suggesting that there are new INH targets yet to be discovered. PMID:19581462

  8. Anthocyanidins modulate the activity of human DNA topoisomerases I and II and affect cellular DNA integrity.

    PubMed

    Habermeyer, Michael; Fritz, Jessica; Barthelmes, Hans U; Christensen, Morten O; Larsen, Morten K; Boege, Fritz; Marko, Doris

    2005-09-01

    In the present study, we investigated the effect of anthocyanidins on human topoisomerases I and II and its relevance for DNA integrity within human cells. Anthocyanidins bearing vicinal hydroxy groups at the B-ring (delphinidin, DEL; cyanidin, CY) were found to potently inhibit the catalytic activity of human topoisomerases I and II, without discriminating between the IIalpha and the IIbeta isoforms. However, in contrast to topoisomerase poisons, DEL and CY did not stabilize the covalent DNA-topoisomerase intermediates (cleavable complex) of topoisomerase I or II. Using recombinant topoisomerase I, the presence of CY or DEL (> or = 1 microM) effectively prohibited the stabilization of the cleavable complex by the topoisomerase I poison camptothecin. We furthermore investigated whether the potential protective effect vs topoisomerase I poisons is reflected also on the cellular level, affecting the DNA damaging properties of camptothecin. Indeed, in HT29 cells, low micromolar concentrations of DEL (1-10 microM) significantly diminished the DNA strand breaking effect of camptothecin (100 microM). However, at concentrations > or = 50 microM, all anthocyanidins tested (delphinidin, cyanidin, malvidin, pelargonidin, and paeonidin), including those not interfering with topoisomerases, were found to induce DNA strand breaks in the comet assay. All of these analogues were able to compete with ethidium bromide for the intercalation into calf thymus DNA and to replace the minor groove binder Hoechst 33258. These data indicate substantial affinity to double-stranded DNA, which might contribute at least to the DNA strand breaking effect of anthocyanidins at higher concentrations (> or = 50 microM).

  9. Structural basis for acceptor-substrate recognition of UDP-glucose: anthocyanidin 3-O-glucosyltransferase from Clitoria ternatea.

    PubMed

    Hiromoto, Takeshi; Honjo, Eijiro; Noda, Naonobu; Tamada, Taro; Kazuma, Kohei; Suzuki, Masahiko; Blaber, Michael; Kuroki, Ryota

    2015-03-01

    UDP-glucose: anthocyanidin 3-O-glucosyltransferase (UGT78K6) from Clitoria ternatea catalyzes the transfer of glucose from UDP-glucose to anthocyanidins such as delphinidin. After the acylation of the 3-O-glucosyl residue, the 3'- and 5'-hydroxyl groups of the product are further glucosylated by a glucosyltransferase in the biosynthesis of ternatins, which are anthocyanin pigments. To understand the acceptor-recognition scheme of UGT78K6, the crystal structure of UGT78K6 and its complex forms with anthocyanidin delphinidin and petunidin, and flavonol kaempferol were determined to resolutions of 1.85 Å, 2.55 Å, 2.70 Å, and 1.75 Å, respectively. The enzyme recognition of unstable anthocyanidin aglycones was initially observed in this structural determination. The anthocyanidin- and flavonol-acceptor binding details are almost identical in each complex structure, although the glucosylation activities against each acceptor were significantly different. The 3-hydroxyl groups of the acceptor substrates were located at hydrogen-bonding distances to the Nε2 atom of the His17 catalytic residue, supporting a role for glucosyl transfer to the 3-hydroxyl groups of anthocyanidins and flavonols. However, the molecular orientations of these three acceptors are different from those of the known flavonoid glycosyltransferases, VvGT1 and UGT78G1. The acceptor substrates in UGT78K6 are reversely bound to its binding site by a 180° rotation about the O1-O3 axis of the flavonoid backbones observed in VvGT1 and UGT78G1; consequently, the 5- and 7-hydroxyl groups are protected from glucosylation. These substrate recognition schemes are useful to understand the unique reaction mechanism of UGT78K6 for the ternatin biosynthesis, and suggest the potential for controlled synthesis of natural pigments. PMID:25556637

  10. Structural basis for acceptor-substrate recognition of UDP-glucose: anthocyanidin 3-O-glucosyltransferase from Clitoria ternatea

    PubMed Central

    Hiromoto, Takeshi; Honjo, Eijiro; Noda, Naonobu; Tamada, Taro; Kazuma, Kohei; Suzuki, Masahiko; Blaber, Michael; Kuroki, Ryota

    2015-01-01

    UDP-glucose: anthocyanidin 3-O-glucosyltransferase (UGT78K6) from Clitoria ternatea catalyzes the transfer of glucose from UDP-glucose to anthocyanidins such as delphinidin. After the acylation of the 3-O-glucosyl residue, the 3′- and 5′-hydroxyl groups of the product are further glucosylated by a glucosyltransferase in the biosynthesis of ternatins, which are anthocyanin pigments. To understand the acceptor-recognition scheme of UGT78K6, the crystal structure of UGT78K6 and its complex forms with anthocyanidin delphinidin and petunidin, and flavonol kaempferol were determined to resolutions of 1.85 Å, 2.55 Å, 2.70 Å, and 1.75 Å, respectively. The enzyme recognition of unstable anthocyanidin aglycones was initially observed in this structural determination. The anthocyanidin- and flavonol-acceptor binding details are almost identical in each complex structure, although the glucosylation activities against each acceptor were significantly different. The 3-hydroxyl groups of the acceptor substrates were located at hydrogen-bonding distances to the Nε2 atom of the His17 catalytic residue, supporting a role for glucosyl transfer to the 3-hydroxyl groups of anthocyanidins and flavonols. However, the molecular orientations of these three acceptors are different from those of the known flavonoid glycosyltransferases, VvGT1 and UGT78G1. The acceptor substrates in UGT78K6 are reversely bound to its binding site by a 180° rotation about the O1–O3 axis of the flavonoid backbones observed in VvGT1 and UGT78G1; consequently, the 5- and 7-hydroxyl groups are protected from glucosylation. These substrate recognition schemes are useful to understand the unique reaction mechanism of UGT78K6 for the ternatin biosynthesis, and suggest the potential for controlled synthesis of natural pigments. PMID:25556637

  11. Crystal Structures of Trimethoprim-Resistant DfrA1 Rationalize Potent Inhibition by Propargyl-Linked Antifolates.

    PubMed

    Lombardo, Michael N; G-Dayanandan, Narendran; Wright, Dennis L; Anderson, Amy C

    2016-02-12

    Multidrug-resistant Enterobacteriaceae, notably Escherichia coli and Klebsiella pneumoniae, have become major health concerns worldwide. Resistance to effective therapeutics is often carried by class I and II integrons that can confer insensitivity to carbapenems, extended spectrum β-lactamases, the antifolate trimethoprim, fluoroquinolones, and aminoglycosides. Specifically of interest to the study here, a prevalent gene (dfrA1) coding for an insensitive dihydrofolate reductase (DHFR) confers 190- or 1000-fold resistance to trimethoprim for K. pneumoniae and E. coli, respectively. Attaining inhibition of both the wild-type and resistant forms of the enzyme is critical for new antifolates. For several years, we have been developing the propargyl-linked antifolates (PLAs) as effective inhibitors against trimethoprim-resistant DHFR enzymes. Here, we show that the PLAs are active against both the wild-type and DfrA1 DHFR proteins. We report two high-resolution crystal structures of DfrA1 bound to potent PLAs. The structure-activity relationships and crystal structures will be critical in driving the design of broadly active inhibitors against wild-type and resistant DHFR. PMID:27624966

  12. First report on class 1 integrons and Trimethoprim-resistance genes from dfrA group in uropathogenic E. coli (UPEC) from the Aleppo area in Syria.

    PubMed

    Al-Assil, Bodour; Mahfoud, Maysa; Hamzeh, Abdul Rezzak

    2013-05-01

    Horizontal gene transfer (HGT) introduces advantageous genetic elements into pathogenic bacteria using tools such as class1 integrons. This study aimed at investigating the distribution of these integrons among uropathogenic E. coli (UPEC) isolated from patients in Aleppo, Syria. It also set to uncover the frequencies of the clinically relevant DfrA1 and DfrA17,7, as well as various associations leading to reduced susceptibility. This study involved 75 Trimethoprim-resistant E. coli isolates from in- and outpatients with urinary tract infections (UTIs) from 3 major hospitals in Aleppo. Bacterial identification, resistance and extended-spectrum-β-lactamase (ESBL) production testing were performed according to Clinical Laboratory Standards Institute guidelines. Detection of integrons and DfrA genes was done using PCR and statistical significance was inferred through χ2 (Fisher's) test. Class1 integrons were detected in 54.6% of isolates while DfrA1 and DfrA17,7 were found in 16% and 70.6% of tested samples respectively. Furthermore, only DfrA17,7 were strongly associated with class1 integrons, as were reduced susceptibility to the majority of individual antibiotics, multidrug resistance and ESBL production. This study demonstrated the high prevalence of class1 integrons among UPEC strains in Aleppo, Syria, as well as their significant associations with MDR. This data give information for local healthcare provision using antibiotic chemotherapy.

  13. Anthocyanidins and Flavonols, Major nod Gene Inducers from Seeds of a Black-Seeded Common Bean (Phaseolus vulgaris L.) 1

    PubMed Central

    Hungria, Mariangela; Joseph, Cecillia M.; Phillips, Donald A.

    1991-01-01

    Eleven compounds released from germinating seeds of a black-seeded bean (Phaseolus vulgaris L., cv PI165426CS) induce transcription of nod genes in Rhizobium leguminosarum biovar phaseoli. Aglycones from 10 of those compounds were identified by spectroscopic methods (ultraviolet/visible, proton nuclear magnetic resonance, and mass spectroscopy), and their biological activities were demonstrated by induction of β-galactosidase activity in R. leguminosarum strains containing nodA-lacZ or nodC-lacZ fusions controlled by R. leguminosarum biovar phaseoli nodD genes. By making comparisons with authentic standards, the chemical structures for aglycones from the 10 molecules were confirmed as being anthocyanidins (delphinidin, petunidin, and malvidin) and flavonols (myricetin, quercetin, and kaempferol). All anthocyanidins and flavonols had 3-O-glycosylation and free hydroxyl groups at the 4′, 5, and 7 positions. Hydrolysis experiments showed that the mean concentration required for half-maximum nod gene induction (I50) by the 10 glycosides was about half that of the corresponding aglycones. The mean I50 value for the three anthocyanidins (360 nanomolar) was less (P ≤ 0.05) than that of the three flavonol aglycones (980 nanomolar). Each seed released approximately 2500 nanomoles of anthocyanidin and 450 nanomoles of flavonol nod gene inducers in conjugated forms during the first 6 hours of imbibition. Based on amounts and activities of the compounds released, anthocyanins contributed approximately 10-fold more total nod-inducing activity than flavonol glycosides. These anthocyanidins from bean seeds represent the first nod-inducing compounds identified from that group of flavonoids. PMID:16668462

  14. Anthocyanidins and Flavonols, Major nod Gene Inducers from Seeds of a Black-Seeded Common Bean (Phaseolus vulgaris L.).

    PubMed

    Hungria, M; Joseph, C M; Phillips, D A

    1991-10-01

    Eleven compounds released from germinating seeds of a black-seeded bean (Phaseolus vulgaris L., cv PI165426CS) induce transcription of nod genes in Rhizobium leguminosarum biovar phaseoli. Aglycones from 10 of those compounds were identified by spectroscopic methods (ultraviolet/visible, proton nuclear magnetic resonance, and mass spectroscopy), and their biological activities were demonstrated by induction of beta-galactosidase activity in R. leguminosarum strains containing nodA-lacZ or nodC-lacZ fusions controlled by R. leguminosarum biovar phaseoli nodD genes. By making comparisons with authentic standards, the chemical structures for aglycones from the 10 molecules were confirmed as being anthocyanidins (delphinidin, petunidin, and malvidin) and flavonols (myricetin, quercetin, and kaempferol). All anthocyanidins and flavonols had 3-O-glycosylation and free hydroxyl groups at the 4', 5, and 7 positions. Hydrolysis experiments showed that the mean concentration required for half-maximum nod gene induction (I(50)) by the 10 glycosides was about half that of the corresponding aglycones. The mean I(50) value for the three anthocyanidins (360 nanomolar) was less (P anthocyanidin and 450 nanomoles of flavonol nod gene inducers in conjugated forms during the first 6 hours of imbibition. Based on amounts and activities of the compounds released, anthocyanins contributed approximately 10-fold more total nod-inducing activity than flavonol glycosides. These anthocyanidins from bean seeds represent the first nod-inducing compounds identified from that group of flavonoids.

  15. Transcripts of anthocyanidin reductase and leucoanthocyanidin reductase and measurement of catechin and epicatechin in tartary buckwheat.

    PubMed

    Kim, Yeon Bok; Thwe, Aye Aye; Kim, Yeji; Li, Xiaohua; Cho, Jin Woong; Park, Phun Bum; Valan Arasu, Mariadhas; Abdullah Al-Dhabi, Naif; Kim, Sun-Ju; Suzuki, Tastsuro; Hyun Jho, Kwang; Park, Sang Un

    2014-01-01

    Anthocyanidin reductase (ANR) and leucoanthocyanidin reductase (LAR) play an important role in the monomeric units biosynthesis of proanthocyanidins (PAs) such as catechin and epicatechin in several plants. The aim of this study was to clone ANR and LAR genes involved in PAs biosynthesis and examine the expression of these two genes in different organs under different growth conditions in two tartary buckwheat cultivars, Hokkai T8 and T10. Gene expression was carried out by quantitative real-time RT-PCR, and catechin and epicatechin content was analyzed by high performance liquid chromatography. The expression pattern of ANR and LAR did not match the accumulation pattern of PAs in different organs of two cultivars. Epicatechin content was the highest in the flowers of both cultivars and it was affected by light in only Hokkai T8 sprouts. ANR and LAR levels in tartary buckwheat might be regulated by different mechanisms for catechin and epicatechin biosynthesis under light and dark conditions.

  16. Transcripts of Anthocyanidin Reductase and Leucoanthocyanidin Reductase and Measurement of Catechin and Epicatechin in Tartary Buckwheat

    PubMed Central

    Kim, Yeon Bok; Thwe, Aye Aye; Kim, YeJi; Li, Xiaohua; Cho, Jin Woong; Park, Phun Bum; Valan Arasu, Mariadhas; Abdullah Al-Dhabi, Naif; Kim, Sun-Ju; Suzuki, Tastsuro; Hyun Jho, Kwang; Park, Sang Un

    2014-01-01

    Anthocyanidin reductase (ANR) and leucoanthocyanidin reductase (LAR) play an important role in the monomeric units biosynthesis of proanthocyanidins (PAs) such as catechin and epicatechin in several plants. The aim of this study was to clone ANR and LAR genes involved in PAs biosynthesis and examine the expression of these two genes in different organs under different growth conditions in two tartary buckwheat cultivars, Hokkai T8 and T10. Gene expression was carried out by quantitative real-time RT-PCR, and catechin and epicatechin content was analyzed by high performance liquid chromatography. The expression pattern of ANR and LAR did not match the accumulation pattern of PAs in different organs of two cultivars. Epicatechin content was the highest in the flowers of both cultivars and it was affected by light in only Hokkai T8 sprouts. ANR and LAR levels in tartary buckwheat might be regulated by different mechanisms for catechin and epicatechin biosynthesis under light and dark conditions. PMID:24605062

  17. Transcripts of anthocyanidin reductase and leucoanthocyanidin reductase and measurement of catechin and epicatechin in tartary buckwheat.

    PubMed

    Kim, Yeon Bok; Thwe, Aye Aye; Kim, Yeji; Li, Xiaohua; Cho, Jin Woong; Park, Phun Bum; Valan Arasu, Mariadhas; Abdullah Al-Dhabi, Naif; Kim, Sun-Ju; Suzuki, Tastsuro; Hyun Jho, Kwang; Park, Sang Un

    2014-01-01

    Anthocyanidin reductase (ANR) and leucoanthocyanidin reductase (LAR) play an important role in the monomeric units biosynthesis of proanthocyanidins (PAs) such as catechin and epicatechin in several plants. The aim of this study was to clone ANR and LAR genes involved in PAs biosynthesis and examine the expression of these two genes in different organs under different growth conditions in two tartary buckwheat cultivars, Hokkai T8 and T10. Gene expression was carried out by quantitative real-time RT-PCR, and catechin and epicatechin content was analyzed by high performance liquid chromatography. The expression pattern of ANR and LAR did not match the accumulation pattern of PAs in different organs of two cultivars. Epicatechin content was the highest in the flowers of both cultivars and it was affected by light in only Hokkai T8 sprouts. ANR and LAR levels in tartary buckwheat might be regulated by different mechanisms for catechin and epicatechin biosynthesis under light and dark conditions. PMID:24605062

  18. Crystal structure of UDP-glucose:anthocyanidin 3-O-glucosyltransferase from Clitoria ternatea.

    PubMed

    Hiromoto, Takeshi; Honjo, Eijiro; Tamada, Taro; Noda, Naonobu; Kazuma, Kohei; Suzuki, Masahiko; Kuroki, Ryota

    2013-11-01

    Flowers of the butterfly pea (Clitoria ternatea) accumulate a group of polyacylated anthocyanins, named ternatins, in their petals. The first step in ternatin biosynthesis is the transfer of glucose from UDP-glucose to anthocyanidins such as delphinidin, a reaction catalyzed in C. ternatea by UDP-glucose:anthocyanidin 3-O-glucosyltransferase (Ct3GT-A; AB185904). To elucidate the structure-function relationship of Ct3GT-A, recombinant Ct3GT-A was expressed in Escherichia coli and its tertiary structure was determined to 1.85 Å resolution by using X-ray crystallography. The structure of Ct3GT-A shows a common folding topology, the GT-B fold, comprised of two Rossmann-like β/α/β domains and a cleft located between the N- and C-domains containing two cavities that are used as binding sites for the donor (UDP-Glc) and acceptor substrates. By comparing the structure of Ct3GT-A with that of the flavonoid glycosyltransferase VvGT1 from red grape (Vitis vinifera) in complex with UDP-2-deoxy-2-fluoro glucose and kaempferol, locations of the catalytic His-Asp dyad and the residues involved in recognizing UDP-2-deoxy-2-fluoro glucose were essentially identical in Ct3GT-A, but certain residues of VvGT1 involved in binding kaempferol were found to be substituted in Ct3GT-A. These findings are important for understanding the differentiation of acceptor-substrate recognition in these two enzymes. PMID:24121335

  19. Mechanistic study on the oxidation of anthocyanidin synthase by quantum mechanical calculation.

    PubMed

    Nakajima, Jun-ichiro; Sato, Yoshiharu; Hoshino, Tyuji; Yamazaki, Mami; Saito, Kazuki

    2006-07-28

    Anthocyanidin synthase (ANS), a member of the 2-oxoglutarate-dependent dioxygenase family in flavonoid biosynthesis, catalyzes the conversion of leucoanthocyanidins (e.g. 2R,3S,4S-cis-leucocyanidin, LCD) to flav-2-en-3,4-diols, a direct precursor of colored anthocyanidins via flavan-3,3,4-triols. The detailed oxygenation mechanism of 2R,3S,4S-cis-LCD to flav-2-en-3,4-diols was investigated using the density functional theory method. An initial model for the calculation was constructed from a structure obtained by a 100-ps molecular dynamics simulation of Arabidopsis ANS under physiological conditions. This model consisted of an LCD molecule as the substrate together with an iron atom, two histidine residues, an aspartic acid residue, a succinate, and an oxygen atom as ligands of the iron atom. The results of the calculation indicated that both the C-3 and C-4 positions of LCD can be oxidized, although C-4 oxidation is preferable. The C-3 oxidation required several steps to form flavan-3,3,4-triol: 1) formation of Fe(III)-OH and a substrate C-3 radical via hydrogen atom abstraction by Fe(IV)=O, 2) formation of a C-3 ketone and a water molecule, 3) addition of OH(-) into the C-3 position of the ketone, and 4) addition of H(+) to form flavan-3,3,4-triol. On the other hand, C-4 oxidation of 2R,3S,4S-cis-LCD resulted in the direct formation of 2R,3R-trans-dihydroquercetin. These results suggest that the oxidation at C-3 of LCD, a key reaction for coloring in anthocyanin biosynthesis, can be regarded as a "side reaction" from the viewpoint of quantum mechanics of enzymatic reactions. Molecular evolutional implications of ANS and related proteins are discussed in terms of reaction dynamics.

  20. Ground and excited state properties of high performance anthocyanidin dyes-sensitized solar cells in the basic solutions

    SciTech Connect

    Prima, Eka Cahya; Yuliarto, Brian; Suyatman; Dipojono, Hermawan Kresno

    2015-09-30

    The aglycones of anthocyanidin dyes were previously reported to form carbinol pseudobase, cis-chalcone, and trans-chalcone due to the basic levels. The further investigations of ground and excited state properties of the dyes were characterized using density functional theory with PCM(UFF)/B3LYP/6-31+G(d,p) level in the basic solutions. However, to the best of our knowledge, the theoretical investigation of their potential photosensitizers has never been reported before. In this paper, the theoretical photovoltaic properties sensitized by dyes have been successfully investigated including the electron injections, the ground and excited state oxidation potentials, the estimated open circuit voltages, and the light harvesting efficiencies. The results prove that the electronic properties represented by dyes’ LUMO-HOMO levels will affect to the photovoltaic performances. Cis-chalcone dye is the best anthocyanidin aglycone dye with the electron injection spontaneity of −1.208 eV, the theoretical open circuit voltage of 1.781 V, and light harvesting efficiency of 56.55% due to the best HOMO-LUMO levels. Moreover, the ethanol solvent slightly contributes to the better cell performance than the water solvent dye because of the better oxidation potential stabilization in the ground state as well as in the excited state. These results are in good agreement with the known experimental report that the aglycones of anthocyanidin dyes in basic solvent are the high potential photosensitizers for dye-sensitized solar cell.

  1. Ground and excited state properties of high performance anthocyanidin dyes-sensitized solar cells in the basic solutions

    NASA Astrophysics Data System (ADS)

    Prima, Eka Cahya; Yuliarto, Brian; Suyatman, Dipojono, Hermawan Kresno

    2015-09-01

    The aglycones of anthocyanidin dyes were previously reported to form carbinol pseudobase, cis-chalcone, and trans-chalcone due to the basic levels. The further investigations of ground and excited state properties of the dyes were characterized using density functional theory with PCM(UFF)/B3LYP/6-31+G(d,p) level in the basic solutions. However, to the best of our knowledge, the theoretical investigation of their potential photosensitizers has never been reported before. In this paper, the theoretical photovoltaic properties sensitized by dyes have been successfully investigated including the electron injections, the ground and excited state oxidation potentials, the estimated open circuit voltages, and the light harvesting efficiencies. The results prove that the electronic properties represented by dyes' LUMO-HOMO levels will affect to the photovoltaic performances. Cis-chalcone dye is the best anthocyanidin aglycone dye with the electron injection spontaneity of -1.208 eV, the theoretical open circuit voltage of 1.781 V, and light harvesting efficiency of 56.55% due to the best HOMO-LUMO levels. Moreover, the ethanol solvent slightly contributes to the better cell performance than the water solvent dye because of the better oxidation potential stabilization in the ground state as well as in the excited state. These results are in good agreement with the known experimental report that the aglycones of anthocyanidin dyes in basic solvent are the high potential photosensitizers for dye-sensitized solar cell.

  2. Cloning of an anthocyanidin synthase gene homolog from blackcurrant (Ribes nigrum L.) and its expression at different fruit stages.

    PubMed

    Li, X-G; Wang, J; Yu, Z-Y

    2015-01-01

    Anthocyanidin synthase (ANS), a 2-oxoglutarate (2OG) and Fe(II)-dependent oxygenase, catalyzes the penultimate step in anthocyanin biosynthesis, from leucoanthocyanidins to anthocyanidins, the first colored compound in the anthocyanin pathway. In this study, a full-length, 1427-bp long cDNA named RnANS1, which is homologous to the anthocyanidin synthase gene, was cloned from blackcurrant using a homologous cloning strategy. RnANS1 is highly homologous to other plant ANS genes at both the nucleotide and amino acid sequence levels. The deduced protein contains domains conserved in the 2OG and Fe(II)-dependent oxygenase, and is phylogenetically closely related to Paeonia suffruticosa and Paeonia lactiflora. The expression of RnANS1 was upregulated during fruit maturation, and correlated with the accumulation of anthocyanins and soluble carbohydrates in the fruit. Further characterization of the structure and expression patterns of RnANS1 will clarify our understanding of anthocyanin biosynthesis in blackcurrant, and support the development of molecular approaches to manipulate anthocyanin production in this plant. PMID:25867421

  3. Molecular cloning, computational and expression analysis of anthocyanidin reductase in tea (Camellia sinensis).

    PubMed

    Thirugnanasambantham, Krishnaraj; Muralidaran, Senguttuvan; Mandal, Abul Kalam Azad

    2014-09-01

    Tea [Camellia sinensis (L.) O. Kuntze] is one of the most popular non-alcoholic beverages rich in phenolic compounds, which includes epigallocatechin gallate (EGCG), epigallocatechin (EGC), epicatechin gallate (ECG), epicatechin (EC) and catechin (C). Anthocyanidin reductase (ANR) is responsible for catechin biosynthesis in plants, and analysis of its protein sequences and structures will be valuable for further research in the field. We have screened our dormant bud-specific complementary DNA (cDNA) library and reported 1,322-bp cDNA encoding CsANR. Analysis of the sequence revealed the presence of 1,011-bp open reading frame with coding capacity for a polypeptide of 337 amino acids, flanked by 1,123- and 196-bp 5' and 3' untranslated regions, respectively. Theoretical molecular weight (MW) and isoelectric point (pI) of the deduced ANR protein were predicted (using ProtParam) to be 36.4 kDa and 6.54. For the first time, we have reported 3D model of ANR from C. sinensis. Quality of the predicted model was analysed with PROCHECK analysis. Molecular docking of modelled ANR revealed similar binding pockets for both substrates and products. Expression analyses of CsANR and accumulation pattern of catechins were observed to be varied with developmental age of tissue and seasonal condition. Variation in accumulation pattern of catechins and its fractions was found to be correlated with expression pattern of ANR.

  4. Premature and ectopic anthocyanin formation by silencing of anthocyanidin reductase in strawberry (Fragaria × ananassa).

    PubMed

    Fischer, Thilo C; Mirbeth, Beate; Rentsch, Judith; Sutter, Corina; Ring, Ludwig; Flachowsky, Henryk; Habegger, Ruth; Hoffmann, Thomas; Hanke, Magda-Viola; Schwab, Wilfried

    2014-01-01

    Strawberry (Fragaria × ananassa) is a fruit crop with a distinct biphasic flavonoid biosynthesis. Whereas, in the immature receptacle, high levels of proanthocyanidins accumulate, which are associated with herbivore deterrence and pathogen defense, the prominent color-giving anthocyanins are primarily produced in ripe 'fruits' helping to attract herbivores for seed dispersal. Here, constitutive experimental down-regulation of one branch of proanthocyanidin biosynthesis was performed. As a result, the proportion of epicatechin monomeric units within the proanthocyanidin polymer chains was reduced, but this was not the case for the epicatechin starter unit. Shortened chain lengths of proanthocyanidins were also observed. All enzymatic activities for the production of color-giving anthocyanins were already present in unripe fruits at levels allowing a striking red anthocyanin phenotype in unripe fruits of the RNAi silencing lines. An immediately recognizable phenotype was also observed for the stigmata of flowers, which is another epicatechin-forming tissue. Thus, the down-regulation of anthocyanidin reductase (ANR) induced a redirection of the proanthocyanidin pathway, leading to premature and ectopic anthocyanin biosynthesis via enzymatic glycosylation as the alternative pathway. This redirection is also seen in flavonol biosynthesis, which is paralleled by higher pollen viability in silencing lines. ANRi transgenic lines of strawberry provide a versatile tool for the study of the biological functions of proanthocyanidins.

  5. Isolation and Characterization of cDNAs Encoding Leucoanthocyanidin Reductase and Anthocyanidin Reductase from Populus trichocarpa

    PubMed Central

    Lu, Wanxiang; Yang, Li; Karim, Abdul; Luo, Keming

    2013-01-01

    Proanthocyanidins (PAs) contribute to poplar defense mechanisms against biotic and abiotic stresses. Transcripts of PA biosynthetic genes accumulated rapidly in response to infection by the fungus Marssonina brunnea f.sp. multigermtubi, treatments of salicylic acid (SA) and wounding, resulting in PA accumulation in poplar leaves. Anthocyanidin reductase (ANR) and leucoanthocyanidin reductase (LAR) are two key enzymes of the PA biosynthesis that produce the main subunits: (+)-catechin and (−)-epicatechin required for formation of PA polymers. In Populus, ANR and LAR are encoded by at least two and three highly related genes, respectively. In this study, we isolated and functionally characterized genes PtrANR1 and PtrLAR1 from P. trichocarpa. Phylogenetic analysis shows that Populus ANR1 and LAR1 occurr in two distinct phylogenetic lineages, but both genes have little difference in their tissue distribution, preferentially expressed in roots. Overexpression of PtrANR1 in poplar resulted in a significant increase in PA levels but no impact on catechin levels. Antisense down-regulation of PtrANR1 showed reduced PA accumulation in transgenic lines, but increased levels of anthocyanin content. Ectopic expression of PtrLAR1 in poplar positively regulated the biosynthesis of PAs, whereas the accumulation of anthocyanin and flavonol was significantly reduced (P<0.05) in all transgenic plants compared to the control plants. These results suggest that both PtrANR1 and PtrLAR1 contribute to PA biosynthesis in Populus. PMID:23741362

  6. Direct and indirect inactivation of tumor cell protective catalase by salicylic acid and anthocyanidins reactivates intercellular ROS signaling and allows for synergistic effects.

    PubMed

    Scheit, Katrin; Bauer, Georg

    2015-03-01

    Salicylic acid and anthocyanidins are known as plant-derived antioxidants, but also can provoke paradoxically seeming prooxidant effects in vitro. These prooxidant effects are connected to the potential of salicylic acid and anthocyanidins to induce apoptosis selectively in tumor cells in vitro and to inhibit tumor growth in animal models. Several epidemiological studies have shown that salicylic acid and its prodrug acetylsalicylic acid are tumor-preventive for humans. The mechanism of salicylic acid- and anthocyanidin-dependent antitumor effects has remained enigmatic so far. Extracellular apoptosis-inducing reactive oxygen species signaling through the NO/peroxynitrite and the HOCl signaling pathway specifically induces apoptosis in transformed cells. Tumor cells have acquired resistance against intercellular reactive oxygen species signaling through expression of membrane-associated catalase. Here, we show that salicylic acid and anthocyanidins inactivate tumor cell protective catalase and thus reactive apoptosis-inducing intercellular reactive oxygen species signaling of tumor cells and the mitochondrial pathway of apoptosis Salicylic acid inhibits catalase directly through its potential to transform compound I of catalase into the inactive compound II. In contrast, anthocyanidins provoke a complex mechanism for catalase inactivation that is initiated by anthocyanidin-mediated inhibition of NO dioxygenase. This allows the formation of extracellular singlet oxygen through the reaction between H(2)O(2) and peroxynitrite, amplification through a caspase8-dependent step and subsequent singlet oxygen-mediated inactivation of catalase. The combination of salicylic acid and anthocyanidins allows for a remarkable synergistic effect in apoptosis induction. This effect may be potentially useful to elaborate novel therapeutic approaches and crucial for the interpretation of epidemiological results related to the antitumor effects of secondary plant compounds.

  7. Mass spectrometric and enzymatic evidence confirm the existence of anthocyanidin 3,5-O-diglucosides in cabernet sauvignon (Vitis vinifera L.) grape berries.

    PubMed

    Xing, Ran-Ran; Li, Si-Yu; He, Fei; Yang, Zhe; Duan, Chang-Qing; Li, Zheng; Wang, Jun; Pan, Qiu-Hong

    2015-04-01

    It has been widely accepted that anthocyanidin 3,5-O-diglucosides do not exist in Vitis vinifera L. Cabernet Sauvignon (CS) berries. However, our anthocyanin analyses using HPLC-ESI-MS/MS detected the existence of a low level of anthocyanidin 3,5-O-diglucosides in the Cabernet Sauvignon grape berries grown in China. The authenticity of these samples was confirmed with microsatellite markers. The existence of anthocyanidin 3,5-O-diglucoside was further verified by the enzymatic evidence for the first time. Four putative 5-O-glucosyltransferase (5GT) genes were isolated from the Cabernet Sauvignon berries. The enzymatic analysis showed that a recombinant protein (designated as Vv5GT3) glucosylated the 3-O- and 5-O-positions of anthocyanidins and flavonols. A phylogenetic analysis revealed that this bifunctional enzyme belongs to the 5GT subfamily of UDP-glycosyltransferases. This finding brought a new understanding of the anthocyanins' profile and their biosynthesis in V. vinifera and would be helpful for further investigations of the mechanism of accumulation of anthocyanidin diglucosides in Cabernet Sauvignon berries in China's wine-producing regions.

  8. Crystal structure of UDP-glucose:anthocyanidin 3-O-glucosyltransferase from Clitoria ternatea

    PubMed Central

    Hiromoto, Takeshi; Honjo, Eijiro; Tamada, Taro; Noda, Naonobu; Kazuma, Kohei; Suzuki, Masahiko; Kuroki, Ryota

    2013-01-01

    Flowers of the butterfly pea (Clitoria ternatea) accumulate a group of polyacylated anthocyanins, named ternatins, in their petals. The first step in ternatin biosynthesis is the transfer of glucose from UDP-glucose to antho­cyanidins such as delphinidin, a reaction catalyzed in C. ternatea by UDP-glucose:anthocyanidin 3-O-glucosyltransferase (Ct3GT-A; AB185904). To elucidate the structure–function relationship of Ct3GT-A, recombinant Ct3GT-A was expressed in Escherichia coli and its tertiary structure was determined to 1.85 Å resolution by using X-ray crystallography. The structure of Ct3GT-A shows a common folding topology, the GT-B fold, comprised of two Rossmann-like β/α/β domains and a cleft located between the N- and C-domains containing two cavities that are used as binding sites for the donor (UDP-Glc) and acceptor substrates. By comparing the structure of Ct3GT-A with that of the flavonoid glycosyltransferase VvGT1 from red grape (Vitis vinifera) in complex with UDP-2-deoxy-2-fluoro glucose and kaempferol, locations of the catalytic His-Asp dyad and the residues involved in recognizing UDP-2-deoxy-2-fluoro glucose were essentially identical in Ct3GT-A, but certain residues of VvGT1 involved in binding kaempferol were found to be substituted in Ct3GT-A. These findings are important for understanding the differentiation of acceptor-substrate recognition in these two enzymes. PMID:24121335

  9. De Novo Transcriptome Analysis to Identify Anthocyanin Biosynthesis Genes Responsible for Tissue-Specific Pigmentation in Zoysiagrass (Zoysia japonica Steud.)

    PubMed Central

    Ahn, Jong Hwa; Kim, June-Sik; Kim, Seungill; Soh, Hye Yeon; Shin, Hosub; Jang, Hosung; Ryu, Ju Hyun; Kim, Ahyeong; Yun, Kil-Young; Kim, Shinje; Kim, Ki Sun; Choi, Doil; Huh, Jin Hoe

    2015-01-01

    Zoysiagrass (Zoysia japonica Steud.) is commonly found in temperate climate regions and widely used for lawns, in part, owing to its uniform green color. However, some zoysiagrass cultivars accumulate red to purple pigments in their spike and stolon tissues, thereby decreasing the aesthetic value. Here we analyzed the anthocyanin contents of two zoysiagrass cultivars ‘Anyang-jungji’ (AJ) and ‘Greenzoa’ (GZ) that produce spikes and stolons with purple and green colors, respectively, and revealed that cyanidin and petunidin were primarily accumulated in the pigmented tissues. In parallel, we performed a de novo transcriptome assembly and identified differentially expressed genes between the two cultivars. We found that two anthocyanin biosynthesis genes encoding anthocyanidin synthase (ANS) and dihydroflavonol 4-reductase (DFR) were preferentially upregulated in the purple AJ spike upon pigmentation. Both ANS and DFR genes were also highly expressed in other zoysiagrass cultivars with purple spikes and stolons, but their expression levels were significantly low in the cultivars with green tissues. We observed that recombinant ZjDFR1 and ZjANS1 proteins successfully catalyze the conversions of dihydroflavonols into leucoanthocyanidins and leucoanthocyanidins into anthocyanidins, respectively. These findings strongly suggest that upregulation of ANS and DFR is responsible for tissue-specific anthocyanin biosynthesis and differential pigmentation in zoysiagrass. The present study also demonstrates the feasibility of a de novo transcriptome analysis to identify the key genes associated with specific traits, even in the absence of reference genome information. PMID:25905914

  10. De Novo Transcriptome Analysis to Identify Anthocyanin Biosynthesis Genes Responsible for Tissue-Specific Pigmentation in Zoysiagrass (Zoysia japonica Steud.).

    PubMed

    Ahn, Jong Hwa; Kim, June-Sik; Kim, Seungill; Soh, Hye Yeon; Shin, Hosub; Jang, Hosung; Ryu, Ju Hyun; Kim, Ahyeong; Yun, Kil-Young; Kim, Shinje; Kim, Ki Sun; Choi, Doil; Huh, Jin Hoe

    2015-01-01

    Zoysiagrass (Zoysia japonica Steud.) is commonly found in temperate climate regions and widely used for lawns, in part, owing to its uniform green color. However, some zoysiagrass cultivars accumulate red to purple pigments in their spike and stolon tissues, thereby decreasing the aesthetic value. Here we analyzed the anthocyanin contents of two zoysiagrass cultivars 'Anyang-jungji' (AJ) and 'Greenzoa' (GZ) that produce spikes and stolons with purple and green colors, respectively, and revealed that cyanidin and petunidin were primarily accumulated in the pigmented tissues. In parallel, we performed a de novo transcriptome assembly and identified differentially expressed genes between the two cultivars. We found that two anthocyanin biosynthesis genes encoding anthocyanidin synthase (ANS) and dihydroflavonol 4-reductase (DFR) were preferentially upregulated in the purple AJ spike upon pigmentation. Both ANS and DFR genes were also highly expressed in other zoysiagrass cultivars with purple spikes and stolons, but their expression levels were significantly low in the cultivars with green tissues. We observed that recombinant ZjDFR1 and ZjANS1 proteins successfully catalyze the conversions of dihydroflavonols into leucoanthocyanidins and leucoanthocyanidins into anthocyanidins, respectively. These findings strongly suggest that upregulation of ANS and DFR is responsible for tissue-specific anthocyanin biosynthesis and differential pigmentation in zoysiagrass. The present study also demonstrates the feasibility of a de novo transcriptome analysis to identify the key genes associated with specific traits, even in the absence of reference genome information. PMID:25905914

  11. cDNA-AFLP analysis on bolting or flowering of flowering Chinese cabbage and molecular characteristics of BrcuDFR-like/BrcuAXS gene.

    PubMed

    Xiao, Xufeng; Lei, Jianjun; Cao, Bihao; Chen, Guoju; Chen, Changming

    2012-07-01

    The molecular basis of flower bud differentiation in flowering Chinese cabbage (Brassica rapa L. ssp. Chinensis var. utilis Tsen et Lee) was studied in this work. Samples were taken from two varieties, the early-blooming "Youqin 49" and the late-blooming "Youqingtiancaixin 80", at five different developmental stages and studied via cDNA-AFLP. Nineteen expression sequence tags (ESTs) associated with bolting or flowering were isolated and cloned. Blast results indicated that 15 ESTs were involved in the synthesis of anthocayanins, photosynthesis, signal transduction, and phytochrome production. Two ESTs had high similarity to hypothetical proteins with unknown function. Two other ESTs shared no similarity to any sequence in the NCBI database and potentially may be newly identified genes. The deduced amino acid sequences of EST amplified by primer A6T4 or A8T4 had high similarity to both dihydroflavonol reductase (DFR) and UDP-D: -apiose/UDP-D: -xylose synthase (AXS), thus was named BrcuDFR-like/BrcuAXS. Using the cDNA sequence, a putative BrcuDFR-like/BrcuAXS gene was cloned and characterized from flowering Chinese cabbage via rapid amplification of cDNA ends (RACE). The full-length cDNA has 1332 bp with an open frame of 919 bp which codes for a polypeptide of 313 amino acids. The corresponding genome sequence is 2,046 bp. Comparison of cDNA and its corresponding genomic sequence indicates that BrcuDFR-like/BrcuAXS contains 9 exons and 8 introns. The temporal expression patterns indicated the gene is more likely to encode the DFR protein, which catalyzes the synthesis of anthocayanins, than UDP-D: -apiose/UDP-D: -xylose synthase (AXS), which catalyzes the conversion of UDP-D: -glucuronate to a mixture of UDP-D: -apiose and UDP-D: -xylose. Further work is needed to determine what role BrcuDFR-like/BrcuAXS plays during floral organ development.

  12. Flavor of fresh blueberry juice and the comparison to amount of sugars, acids, anthocyanidins, and physicochemical measurements.

    PubMed

    Bett-Garber, Karen L; Lea, Jeanne M; Watson, Michael A; Grimm, Casey C; Lloyd, Steven W; Beaulieu, John C; Stein-Chisholm, Rebecca E; Andrzejewski, Brett P; Marshall, Donna A

    2015-04-01

    Six cultivars of southern highbush (SHB) and rabbiteye (RE) blueberry samples were harvested on 2 different dates. Each treatment combination was pressed 2 times for repeated measures. Fresh juice was characterized for 18 flavor/taste/feeling factor attributes by a descriptive flavor panel. Each sample was measured for sugars, acids, anthocyanidins, Folin-Ciocalteu, soluble solids (BRIX), titratable acidity (TA), and antioxidant capacity (ORACFL ). Flavors were correlated with the composition and physicochemical data. Blueberry flavor correlated with 3 parameters, and negatively correlated with 2. Strawberry correlated with oxalic acid and negatively correlated with sucrose and quinic acid. Sweet aroma correlated with oxalic and citric acid, but negatively correlated with sucrose, quinic, and total acids. Sweet taste correlated with 11 parameters, including the anthocyanidins; and negatively correlated with 3 parameters. Neither bitter nor astringent correlated with any of the antioxidant parameters, but both correlated with total acids. Sour correlated with total acids and TA, while negatively correlating with pH and BRIX:TA. Throat burn correlated with total acids and TA. Principal component analysis negatively related blueberry, sweet aroma, and sweet to sour, bitter, astringent, tongue tingle, and tongue numbness. The information in this component was related to pH, TA, and BRIX:TA ratio. Another principal component related the nonblueberry fruit flavors to BRIX. This PC, also divided the SHB berries from the RE. This work shows that the impact of juice composition on flavor is very complicated and that estimating flavor with physicochemical parameters is complicated by the composition of the juice.

  13. [Investigation of integrons, sul1-2 and dfr genes in trimethoprim-sulfametoxazole-resistant Stenotrophomonas maltophilia strains isolated from clinical samples].

    PubMed

    Ozkaya, Esra; Aydin, Faruk; Bayramoglu, Gülçin; Buruk, Celal Kurtuluş; Sandalli, Cemal

    2014-04-01

    Stenotrophomonas maltophilia, which is a non-fermentative gram-negative bacillus, has an increasing importance in nosocomial and opportunistic infections. Since it exhibits resistance to numerous broad-spectrum antibiotics such as aminoglycosides, beta-lactams and tetracyclines, it may considerably limit empirical treatment options. Trimethoprim-sulfamethoxazole (SXT) is recommended as the first-line therapy in the treatment of S.maltophilia infections thanks to its high potency and usefulness in a range of patients. In recent years, however, studies in different geographical regions have started to report resistance to SXT. In this study, we aimed to investigate the genes sul1, sul2, dfrA9, dfrA10, dfrA20 and class I, class II integron gene cassettes which are known to play role in SXT resistance among SXT-resistant S.maltophilia strains. A total of 618 S.maltophilia strains isolated from various clinical samples of 339 patients between January 2006 and October 2011 at the laboratory of Medical Microbiology Department, Faculty of Medicine, Karadeniz Technical University, Trabzon, Turkey, were included in the study. The isolates were identified by both conventional methods and the Phoenix automated identification system (Becton Dickinson, USA). SXT resistance was determined in the isolates of 32 patients (32/339, 9.4%) by both the automated system and agar dilution method of them 29 (90.6%) were hospital-acquired, and 3 (9.4%) were community-acquired. The genes which are known as SXT resistance determining genes including sul1, sul2, dfr genes, and class I and class II integron gene cassettes were analyzed by using specific primers with polymerase chain reaction in the 32 SXT-resistant isolates. Subsequently, nucleotide sequence analysis of the amplified materials was performed. As a result of this assay, the presence of class I integron gene cassette and sul1 gene were detected in one isolate. Nucleotide sequence analysis of the gene cassette revealed oxacilinase

  14. The Sorghum Gene for Leaf Color Changes upon Wounding (P) Encodes a Flavanone 4-Reductase in the 3-Deoxyanthocyanidin Biosynthesis Pathway

    PubMed Central

    Kawahigashi, Hiroyuki; Kasuga, Shigemitsu; Sawada, Yuji; Yonemaru, Jun-ichi; Ando, Tsuyu; Kanamori, Hiroyuki; Wu, Jianzhong; Mizuno, Hiroshi; Momma, Mitsuru; Fujimoto, Zui; Hirai, Masami Yokota; Matsumoto, Takashi

    2016-01-01

    Upon wounding or pathogen invasion, leaves of sorghum [Sorghum bicolor (L.) Moench] plants with the P gene turn purple, whereas leaves with the recessive allele turn brown or tan. This purple phenotype is determined by the production of two 3-deoxyanthocyanidins, apigeninidin and luteolinidin, which are not produced by the tan-phenotype plants. Using map-based cloning in progeny from a cross between purple Nakei-MS3B (PP) and tan Greenleaf (pp) cultivars, we isolated this gene, which was located in a 27-kb genomic region around the 58.1 Mb position on chromosome 6. Four candidate genes identified in this region were similar to the maize leucoanthocyanidin reductase gene. None of them was expressed before wounding, and only the Sb06g029550 gene was induced in both cultivars after wounding. The Sb06g029550 protein was detected in Nakei-MS3B, but only slightly in Greenleaf, in which it may be unstable because of a Cys252Tyr substitution. A recombinant Sb06g029550 protein had a specific flavanone 4-reductase activity, and converted flavanones (naringenin or eriodictyol) to flavan-4-ols (apiforol or luteoforol) in vitro. Our data indicate that the Sb06g029550 gene is involved in the 3-deoxyanthocyanidin synthesis pathway. PMID:26994288

  15. The Sorghum Gene for Leaf Color Changes upon Wounding (P) Encodes a Flavanone 4-Reductase in the 3-Deoxyanthocyanidin Biosynthesis Pathway.

    PubMed

    Kawahigashi, Hiroyuki; Kasuga, Shigemitsu; Sawada, Yuji; Yonemaru, Jun-Ichi; Ando, Tsuyu; Kanamori, Hiroyuki; Wu, Jianzhong; Mizuno, Hiroshi; Momma, Mitsuru; Fujimoto, Zui; Hirai, Masami Yokota; Matsumoto, Takashi

    2016-01-01

    Upon wounding or pathogen invasion, leaves of sorghum [Sorghum bicolor (L.) Moench] plants with the P gene turn purple, whereas leaves with the recessive allele turn brown or tan. This purple phenotype is determined by the production of two 3-deoxyanthocyanidins, apigeninidin and luteolinidin, which are not produced by the tan-phenotype plants. Using map-based cloning in progeny from a cross between purple Nakei-MS3B (PP) and tan Greenleaf (pp) cultivars, we isolated this gene, which was located in a 27-kb genomic region around the 58.1 Mb position on chromosome 6. Four candidate genes identified in this region were similar to the maize leucoanthocyanidin reductase gene. None of them was expressed before wounding, and only the Sb06g029550 gene was induced in both cultivars after wounding. The Sb06g029550 protein was detected in Nakei-MS3B, but only slightly in Greenleaf, in which it may be unstable because of a Cys252Tyr substitution. A recombinant Sb06g029550 protein had a specific flavanone 4-reductase activity, and converted flavanones (naringenin or eriodictyol) to flavan-4-ols (apiforol or luteoforol) in vitro Our data indicate that the Sb06g029550 gene is involved in the 3-deoxyanthocyanidin synthesis pathway. PMID:26994288

  16. Targeting cysteine rich C1 domain of Scaffold protein Kinase Suppressor of Ras (KSR) with anthocyanidins and flavonoids - a binding affinity characterization study.

    PubMed

    Karthik, Dhananjayan; Majumder, Pulak; Palanisamy, Sivanandy; Khairunnisa, Kalathil; Venugopal, Varsha

    2014-01-01

    Kinase Suppressor of Ras (KSR) is a molecular scaffold that interacts with the core kinase components of the ERK cascade, Raf, MEK, ERK to provide spatial and temporal regulation of Ras-dependent ERK cascade signaling. Interruption of this mechanism can have a high influence in inhibiting the downstream signaling of the mutated tyrosine kinase receptor kinase upon ligand binding. Still none of the studies targeted to prevent the binding of Raf, MEK binding on kinase suppressor of RAS. In that perspective the cysteine rich C1 domain of scaffold proteins kinase suppressor of Ras-1 was targeted rather than its ATP binding site with small ligand molecules like flavones and anthocyanidins and analyzed through insilico docking studies. The binding energy evaluation shows the importance of hydroxyl groups at various positions on the flavone and anthocyanidin nucleus. Over all binding interaction shows these ligands occupied the potential sites of cysteine rich C1 domain of scaffold protein KSR. PMID:25352726

  17. The anthocyanidin delphinidin mobilizes endogenous copper ions from human lymphocytes leading to oxidative degradation of cellular DNA.

    PubMed

    Hanif, Sarmad; Shamim, Uzma; Ullah, M F; Azmi, Asfar S; Bhat, Showket H; Hadi, S M

    2008-07-10

    Epidemiological and experimental evidence exists to suggest that pomegranate and its juice possess chemopreventive and anticancer properties. The anthocyanidin delphinidin is a major polyphenol present in pomegranates and has been shown to be responsible for these effects. Plant polyphenols are recognized as naturally occurring antioxidants but also catalyze oxidative DNA degradation of cellular DNA either alone or in the presence of transition metal ions such as copper. In this paper we show that similar to various other classes of polyphenols, delphinidin is also capable of causing oxidative degradation of cellular DNA. Lymphocytes were exposed to various concentrations of delphinidin (10, 20, 50 microM) for 1h and the DNA breakage was assessed using single cell alkaline gel electrophoresis (Comet assay). Inhibition of DNA breakage by several scavengers of reactive oxygen species (ROS) indicated that it is caused by the formation of ROS. Incubation of lymphocytes with neocuproine (a cell membrane permeable Cu(I) chelator) inhibited DNA degradation in intact lymphocytes in a dose dependent manner. Bathocuproine, which is unable to permeate through the cell membrane, did not cause such inhibition. We have further shown that delphinidin is able to degrade DNA in cell nuclei and that such DNA degradation is also inhibited by neocuproine suggesting that nuclear copper is mobilized in this reaction. These results indicate that the generation of ROS possibly occurs through mobilization of endogenous copper ions. The results are in support of our hypothesis that the prooxidant activity of plant polyphenols may be an important mechanism for their anticancer properties.

  18. ABCC1, an ATP Binding Cassette Protein from Grape Berry, Transports Anthocyanidin 3-O-Glucosides[W][OA

    PubMed Central

    Francisco, Rita Maria; Regalado, Ana; Ageorges, Agnès; Burla, Bo J.; Bassin, Barbara; Eisenach, Cornelia; Zarrouk, Olfa; Vialet, Sandrine; Marlin, Thérèse; Chaves, Maria Manuela; Martinoia, Enrico; Nagy, Réka

    2013-01-01

    Accumulation of anthocyanins in the exocarp of red grapevine (Vitis vinifera) cultivars is one of several events that characterize the onset of grape berry ripening (véraison). Despite our thorough understanding of anthocyanin biosynthesis and regulation, little is known about the molecular aspects of their transport. The participation of ATP binding cassette (ABC) proteins in vacuolar anthocyanin transport has long been a matter of debate. Here, we present biochemical evidence that an ABC protein, ABCC1, localizes to the tonoplast and is involved in the transport of glucosylated anthocyanidins. ABCC1 is expressed in the exocarp throughout berry development and ripening, with a significant increase at véraison (i.e., the onset of ripening). Transport experiments using microsomes isolated from ABCC1-expressing yeast cells showed that ABCC1 transports malvidin 3-O-glucoside. The transport strictly depends on the presence of GSH, which is cotransported with the anthocyanins and is sensitive to inhibitors of ABC proteins. By exposing anthocyanin-producing grapevine root cultures to buthionine sulphoximine, which reduced GSH levels, a decrease in anthocyanin concentration is observed. In conclusion, we provide evidence that ABCC1 acts as an anthocyanin transporter that depends on GSH without the formation of an anthocyanin-GSH conjugate. PMID:23723325

  19. Peach MYB7 activates transcription of the proanthocyanidin pathway gene encoding leucoanthocyanidin reductase, but not anthocyanidin reductase

    PubMed Central

    Zhou, Hui; Lin-Wang, Kui; Liao, Liao; Gu, Chao; Lu, Ziqi; Allan, Andrew C.; Han, Yuepeng

    2015-01-01

    Proanthocyanidins (PAs) are a group of natural phenolic compounds that have a great effect on both flavor and nutritious value of fruit. It has been shown that PA synthesis is regulated by R2R3-MYB transcription factors (TFs) via activation of PA-specific pathway genes encoding leucoanthocyanidin reductase and anthocyanidin reductase. Here, we report the isolation and characterization of a MYB gene designated PpMYB7 in peach. The peach PpMYB7 represents a new group of R2R3-MYB genes regulating PA synthesis in plants. It is able to activate transcription of PpLAR1 but not PpANR, and has a broader selection of potential bHLH partners compared with PpMYBPA1. Transcription of PpMYB7 can be activated by the peach basic leucine-zipper 5 TF (PpbZIP5) via response to ABA. Our study suggests a transcriptional network regulating PA synthesis in peach, with the results aiding the understanding of the functional divergence between R2R3-MYB TFs in plants. PMID:26579158

  20. MLW, TRU, LLW, MIXED, HAZARDOUS WASTES AND ENVIRONMENTAL RESTORATION. WASTE MANAGEMENT/ENERGY SECURITY AND A CLEAN ENVIRONMENT. DFR Decommissioning: the Breeder Fuel Processing

    SciTech Connect

    Bonnet, C.; Potier, P.; Ashton, Brian Morris

    2003-02-27

    The Dounreay site, in North Scotland, was opened in 1955 and a wide range of nuclear facilities have been built and operated there by UKAEA (The United Kingdom Atomic Energy Authority) for the development of atomic energy research. The Dounreay Fast Reactor (DFR) was built between 1955 and 1957, and operated until 1977 for demonstration purposes and for producing electricity. Today, its decommissioning is a key part of the whole Dounreay Site Restoration Plan that integrates the major decommissioning activities such as the fuel treatment and the waste management. The paper presents the contract strategy and provides an overview of the BFR project which consists in the removal of the breeder elements from the reactor and their further treatment. It mainly provides particular details of the Retrieval and Processing Facilities design.

  1. In silico study for diversing the molecular pathway of pigment formation: an alternative to manual coloring in cotton fibers.

    PubMed

    Ahad, Ammara; Ahmad, Aftab; Din, Salah Ud; Rao, Abdul Q; Shahid, Ahmad A; Husnain, Tayyab

    2015-01-01

    Diversity of colors in flowers and fruits is largely due to anthocyanin pigments. The flavonoid/anthocyanin pathway has been most extensively studied. Dihydroflavonol 4-reductase (DFR) is a vital enzyme of the flavonoid pathway which displays major impact on the formation of anthocyanins, flavan 3-ols and flavonols. The substrate specificity of the DFR was found to play a crucial role in determination of type of anthocyanidins. Altering the flavonoid/anthocyanin pathway through genetic engineering to develop color of our own choice is an exciting subject of future research. In the present study, comparison among four DFR genes (Gossypium hirsutum, Iris × hollandica, Ang. DFRI and DFRII), sequence alignment for homology as well as protein modeling and docking is demonstrated. Estimation of catalytic sites, prediction of substrate preference and protein docking were the key features of this article. For specific substrate uptake, a proline rich region and positions 12 plus 26 along with other positions emphasizing the 26-amino acid residue region (132-157) was tested. Results showed that proline rich region position 12, 26, and 132-157 plays an important role in selective attachment of DFRs with respective substrates. Further, "Expasy ProtParam tool" results showed that Iris × hollandica DFR amino acids (Asn 9: Asp 23) are favorable for reducing DHQ and DHM thus accumulating delphinidin, while Gossypium hirsutum DFR has (Asn 13: Asp 21) hypothesized to consume DHK. Protein docking data showed that amino acid residues in above mentioned positions were just involved in attachment of DFR with substrate and had no role in specific substrate uptake. Advanced bioinformatics analysis has revealed that all above mentioned positions have role in substrate attachment. For substrate specificity, other residues region is involved. It will help in color manipulations in different plant species. PMID:26442064

  2. Efficacy of xylanase purified from Aspergillus niger DFR-5 alone and in combination with pectinase and cellulase to improve yield and clarity of pineapple juice.

    PubMed

    Pal, Ajay; Khanum, Farhath

    2011-10-01

    Pineapple is one of the fruits having xylan rich hemicellulose content more than pectin. Therefore, the efficacy of absolutely purified xylanase from A. niger DFR-5 alone and in combination with pectinase and cellulase on juice yield and clarity was studied. Xylanase provided maximum yield (71.3%) and clarity (64.7%) of juice in comparison to control responses (61.8% yield and 57.8% clarity). When used together, a synergistic effect of xylanase, pectinase and cellulase on process responses was observed indicating the necessity of a cock-tail of hydrolytic enzymes for complete cell wall degradation. Overall, an increase in juice yield by 52.9% was observed. The process was numerically optimized with the constraint of 'minimum' pectinase and cellulase and 'maximum' xylanase and incubation time for 'maximum' juice yield and clarity. The closeness of observed response (90.2% yield and 80.9% clarity) to the predicted one (89.6% yield and 80.3% clarity) indicated the validity of developed model. PMID:23572788

  3. Flower color patterning in pansy (Viola × wittrockiana Gams.) is caused by the differential expression of three genes from the anthocyanin pathway in acyanic and cyanic flower areas.

    PubMed

    Li, Qin; Wang, Jian; Sun, Hai-Yan; Shang, Xiao

    2014-11-01

    The petals of pansy (Viola × wittrockiana Gams.) 'Mengdie' exhibit a cyanic blotched pigmentation pattern. The accumulation of anthocyanins, cyanidin and delphinidin, was detected in the upper epidermal cells of the cyanic blotches. In order to elucidate the mechanism by which cyanic blotches are formed in pansy petal, the expression level of genes involved in anthocyanin synthesis was measured and compared between cyanic blotches and acyanic areas of the flower. The use of primers in conserved regions allowed the successful isolation of six cDNA clones encoding putative anthocyanin enzymes from pansy petals. The clones isolated encoded chalcone synthase (CHS), chalcone isomerase (CHI), flavanone 3-hydroxylase (F3H), flavonoid 3'-hydroxylase (F3'H), dihydroflavonol 4-reductase (DFR) and anthocyanidin synthase (ANS). The transcription patterns of seven genes (VwCHS, VwCHI, VwF3H, VwF3'H, VwDFR, VwF3'5'H, and VwANS) in cyanic blotches and acyanic areas of the petals at seven stages of flower development were determined by real-time quantitative PCR. Transcription of VwF3'5'H, VwDFR and VwANS was significantly increased in cyanic blotches at stages III-V of flower development, implicating these genes in the pigmentation of Viola × wittrockiana Gams. petals.

  4. Flower color patterning in pansy (Viola × wittrockiana Gams.) is caused by the differential expression of three genes from the anthocyanin pathway in acyanic and cyanic flower areas.

    PubMed

    Li, Qin; Wang, Jian; Sun, Hai-Yan; Shang, Xiao

    2014-11-01

    The petals of pansy (Viola × wittrockiana Gams.) 'Mengdie' exhibit a cyanic blotched pigmentation pattern. The accumulation of anthocyanins, cyanidin and delphinidin, was detected in the upper epidermal cells of the cyanic blotches. In order to elucidate the mechanism by which cyanic blotches are formed in pansy petal, the expression level of genes involved in anthocyanin synthesis was measured and compared between cyanic blotches and acyanic areas of the flower. The use of primers in conserved regions allowed the successful isolation of six cDNA clones encoding putative anthocyanin enzymes from pansy petals. The clones isolated encoded chalcone synthase (CHS), chalcone isomerase (CHI), flavanone 3-hydroxylase (F3H), flavonoid 3'-hydroxylase (F3'H), dihydroflavonol 4-reductase (DFR) and anthocyanidin synthase (ANS). The transcription patterns of seven genes (VwCHS, VwCHI, VwF3H, VwF3'H, VwDFR, VwF3'5'H, and VwANS) in cyanic blotches and acyanic areas of the petals at seven stages of flower development were determined by real-time quantitative PCR. Transcription of VwF3'5'H, VwDFR and VwANS was significantly increased in cyanic blotches at stages III-V of flower development, implicating these genes in the pigmentation of Viola × wittrockiana Gams. petals. PMID:25270164

  5. Cloning and characterization of Vitis vinifera UDP-glucose:flavonoid 3-O-glucosyltransferase, a homologue of the enzyme encoded by the maize Bronze-1 locus that may primarily serve to glucosylate anthocyanidins in vivo.

    PubMed

    Ford, C M; Boss, P K; Hoj, P B

    1998-04-10

    We report here the cloning and optimized expression at 16 degrees C and the characterization of a Vitis vinifera UDP-glucose:flavonoid 3-O-glucosyltransferase, an enzyme responsible for a late step in grapevine anthocyanin biosynthesis. The properties of this and other UDP-glucose:flavonoid 3-O-glucosyltransferases, homologues of the product encoded by the maize Bronze-1 locus, are a matter of conjecture. The availability of a purified recombinant enzyme allowed for the unambiguous determination of the characteristics of a flavonoid 3-O-glucosyltransferase. Kinetic analyses showed that kcat for glucosylation of cyanidin, an anthocyanidin substrate, is 48 times higher than for glucosylation of the flavonol quercetin, whereas Km values are similar for both substrates. Activity toward other classes of substrates is absent. Cu2+ ions strongly inhibit the action of this and other glucosyltransferases; however, we suggest that this phenomenon in large part is due to Cu2+-mediated substrate degradation rather than inhibition of the enzyme. Additional lines of complementary biochemical data also indicated that in the case of V. vinifera, the principal, if not only, role of UDP-glucose:flavonoid 3-O-glucosyltransferases is to glucosylate anthocyanidins in red fruit during ripening. Other glucosyltransferases with a much higher relative activity toward quercetin are suggested to glucosylate flavonols in a distinct spatial and temporal pattern. It should be considered whether gene products homologous to Bronze-1 in some cases more accurately should be referred to as UDP-glucose:anthocyanidin 3-O-glucosyltransferases.

  6. Inhibitory effect and mechanisms of an anthocyanins- and anthocyanidins-rich extract from purple-shoot tea on colorectal carcinoma cell proliferation.

    PubMed

    Hsu, Chih-Ping; Shih, Yi-Ting; Lin, Bor-Ru; Chiu, Chui-Feng; Lin, Chih-Cheng

    2012-04-11

    One newly bred variety of tea cultivar, purple-shoot tea, was selected to evaluate its antiproliferative effects on colorectal carcinoma cells, as well as normal colon cells. The phytochemicals and identified catechins of purple-shoot tea extract (PTE) were significantly higher than that of ordinary tea, especially the anthocyanins (surpassed by 135-fold) and anthocyanidins (surpassed by 3.5-fold). PTE inhibited the proliferation of COLO 320DM (IC(50) = 64.9 μg/mL) and HT-29 (IC(50) = 55.2 μg/mL) by blocking cell cycle progression during the G(0)/G(1) phase and inducing apoptotic death. Western blotting indicated that PTE induced cell cycle arrest by reducing the expression of cyclin E and cyclin D1 in COLO 320DM and the upregulation of p21 and p27 cyclin-dependent kinase inhibitors in HT-29. Two cells treated with PTE also indicated the cleavage of PARP, activation of caspase 3, and an increased Bax/Bcl-2 ratio. Our results showed that PTE is a potential novel dietary agent for colorectal cancer chemoprevention.

  7. Molecular identification of 1-Cys peroxiredoxin and anthocyanidin/flavonol 3-O-galactosyltransferase from proanthocyanidin-rich young fruits of persimmon (Diospyros kaki Thunb.).

    PubMed

    Ikegami, Ayako; Akagi, Takashi; Potter, Daniel; Yamada, Masahiko; Sato, Akihiko; Yonemori, Keizo; Kitajima, Akira; Inoue, Kentaro

    2009-09-01

    Fruits of persimmon (Diospyros kaki Thunb.) accumulate large amounts of proanthocyanidins (PAs) in the early stages of development. Astringent (A)-type fruits remain rich in soluble PAs even after they reach full-mature stage, whereas non-astringent (NA)-type fruits lose these compounds before full maturation. As a first step to elucidate the mechanism of PA accumulation in this non-model species, we used suppression subtractive hybridization to identify transcripts accumulating differently in young fruits of A- and NA-type. Interestingly, only a few clones involved in PA biosynthesis were identified in A-NA libraries. Represented by multiple clones were those encoding a novel 1-Cys peroxiredoxin and a new member of family 1 glycosyltransferases. Quantitative RT-PCR analyses confirmed correlation of the amount of PAs and accumulation of transcripts encoding these proteins in young persimmon fruits. Furthermore, the new family 1 glycosyltransferase was produced in Escherichia coli and shown to efficiently catalyze galactosylation at 3-hydroxyl groups of several anthocyanidins and flavonols. These findings suggest a complex mechanism of PA accumulation in persimmon fruits.

  8. Combination of supercritical fluid extraction with counter-current chromatography to isolate anthocyanidins from the petals of Chaenomeles sinensis based on mathematical calculations.

    PubMed

    Li, Sainan; Guo, Liping; Liu, Chunming; Fu, Zi Ao; Zhang, Yuchi

    2013-11-01

    Supercritical fluid extraction (SFE) coupled with high-speed counter-current chromatography (HSCCC) was successfully used for the extraction and on-line isolation of the anthocyanidins from the petals of Chaenomeles sinensis in two stages. The SFE parameters were optimized by an orthogonal test, and the solvent systems of SFE and HSCCC were calculated and optimized with the help of a multiexponential function model. In the first stage, the lower phase of the solvent system of n-butanol/tert-butyl methyl ether/acetonitrile/0.1% aqueous TFA (0.715:1.0:0.134:1.592, v/v/v/v) was used as both the SFE modifier and the HSCCC stationary phase, after extraction, the extractants were pumped into HSCCC column, and then eluted with the corresponding upper phase to isolate the moderately hydrophobic compounds. In the second stage, the upper phase of the solvent system of n-butanol/ethyl acetate/acetonitrile/0.1% aqueous TFA (1.348:1.0:0.605:2.156, v/v/v/v) was used as both the SFE modifier and the HSCCC stationary phase, followed by elution with the corresponding lower phase to separate the hydrophobic compounds. With the help of two-stage SFE/HSCCC, six compounds including delphinidin-3-O-glucoside (Dp3G), cyanidin-3-O-glucoside (Cy3G), peonidin-3-O-glucoside (Pn3G), delphinidin (Dp), peonidin (Pn), and malvidin (Mv) were successfully separated within 300 min. The targeted compounds were identified by UV spectrophotometry, MS, and NMR spectroscopy. This research has opened up great prospects for the industrial application of SFE-HSCCC for the automatic extraction and separation of unstable compounds.

  9. The Reaumuria trigyna leucoanthocyanidin dioxygenase (RtLDOX) gene complements anthocyanidin synthesis and increases the salt tolerance potential of a transgenic Arabidopsis LDOX mutant.

    PubMed

    Zhang, Huirong; Du, Chao; Wang, Yan; Wang, Jia; Zheng, Linlin; Wang, Yingchun

    2016-09-01

    Reaumuria trigyna is a typical, native desert halophyte that grows under extreme conditions in Inner Mongolia. In a previous transcriptomic profiling analysis, flavonoid pathway-related genes in R. trigyna showed significant differences in transcript abundance under salt stress. Leucoanthocyanidin dioxygenase (LDOX, EC 1.14.11.19) is one of three dioxygenases in the flavonoid pathway that catalyzes the formation of anthocyanidins from leucoanthocyanidins. In this study, we cloned the full-length cDNA of R. trigyna LDOX (RtLDOX), and found RtLDOX recombinant protein was able to replace flavanone-3-hydroxylase (F3H, EC 1.14.11.9), another dioxygenase in the flavonoid pathway, to convert naringenin to dihydrokaempferol in vitro. R. trigyna LDOX can complement the Arabidopsis LDOX mutant transparent testa11 (tt11-11), which has reduced proanthocyanin (PA) and anthocyanin levels in seeds, to accumulate these two compounds. Thus, RtLDOX acts as a multifunctional dioxygenase to effect the synthesis of PA and anthocyanins and can perform F3H dioxygenase activities in the flavonoid biosynthesis pathway. The RtLDOX promoter harbored many cis-acting elements that might be recognized and bound by transcription factors related to stress response. RtLDOX expression was strongly increased under salt stress, and RtLDOX transgenic Arabidopsis mutant under NaCl stress accumulated the content of flavonoids leading to an increased antioxidant activities and plant biomass. These results suggest that RtLDOX as a multifunctional dioxygenase in flavonoid biosynthesis involves in enhancing plant response to NaCl stress. PMID:27219053

  10. cDNA cloning, heterologous expressions, and functional characterization of malonyl-coenzyme a:anthocyanidin 3-o-glucoside-6"-o-malonyltransferase from dahlia flowers.

    PubMed

    Suzuki, Hirokazu; Nakayama, Toru; Yonekura-Sakakibara, Keiko; Fukui, Yuko; Nakamura, Noriko; Yamaguchi, Masa-Atsu; Tanaka, Yoshikazu; Kusumi, Takaaki; Nishino, Tokuzo

    2002-12-01

    In the flowers of important ornamental Compositae plants, anthocyanins generally carry malonyl group(s) at their 3-glucosyl moiety. In this study, for the first time to our knowledge, we have identified a cDNA coding for this 3-glucoside-specific malonyltransferase for anthocyanins, i.e. malonyl-coenzyme A:anthocyanidin 3-O-glucoside-6"-O-malonyltransferase, from dahlia (Dahlia variabilis) flowers. We isolated a full-length cDNA (Dv3MaT) on the basis of amino acid sequences specifically conserved among anthocyanin acyltransferases of the versatile plant acyltransferase family. Dv3MaT coded for a protein of 460 amino acids. Quantitative real-time PCR analyses of Dv3MaT showed that the transcript was present in accordance with the distribution of 3MaT activities and the anthocyanin accumulation pattern in the dahlia plant. The Dv3MaT cDNA was expressed in Escherichia coli, and the recombinant enzyme was purified to homogeneity and characterized. The recombinant Dv3MaT catalyzed the regiospecific transfer of the malonyl group from malonyl-coenzyme A (K(m), 18.8 microM) to pelargonidin 3-O-glucoside (K(m), 46.7 microM) to produce pelargonidin 3-O-6"-O-malonylglucoside with a k(cat) value of 7.3 s(-1). The other enzymatic profiles of the recombinant Dv3MaT were closely related to those of native anthocyanin malonyltransferase activity in the extracts of dahlia flowers. Dv3MaT cDNA was introduced into petunia (Petunia hybrida) plants whose red floral color is exclusively provided by cyanidin 3-O-glucoside and 3,5-O-diglucoside. Thirteen transgenic lines of petunia were found to produce malonylated products of these anthocyanins (11-63 mol % of total anthocyanins in the flower). The spectral stability of cyanidin 3-O-6"-O-malonylglucoside at the pHs of intracellular milieus of flowers was significantly higher than that of cyanidin 3-O-glucoside. Moreover, 6"-O-malonylation of cyanidin 3-O-glucoside effectively prevented the anthocyanin from attack of beta

  11. Red Anthocyanins and Yellow Carotenoids Form the Color of Orange-Flower Gentian (Gentiana lutea L. var. aurantiaca).

    PubMed

    Berman, Judit; Sheng, Yanmin; Gómez Gómez, Lourdes; Veiga, Tania; Ni, Xiuzhen; Farré, Gemma; Capell, Teresa; Guitián, Javier; Guitián, Pablo; Sandmann, Gerhard; Christou, Paul; Zhu, Changfu

    2016-01-01

    Flower color is an important characteristic that determines the commercial value of ornamental plants. Gentian flowers occur in a limited range of colors because this species is not widely cultivated as a cut flower. Gentiana lutea L. var. aurantiaca (abbr, aurantiaca) is characterized by its orange flowers, but the specific pigments responsible for this coloration are unknown. We therefore investigated the carotenoid and flavonoid composition of petals during flower development in the orange-flowered gentian variety of aurantiaca and the yellow-flowered variety of G. lutea L. var. lutea (abbr, lutea). We observed minor varietal differences in the concentration of carotenoids at the early and final stages, but only aurantiaca petals accumulated pelargonidin glycosides, whereas these compounds were not found in lutea petals. We cloned and sequenced the anthocyanin biosynthetic gene fragments from petals, and analyzed the expression of these genes in the petals of both varieties to determine the molecular mechanisms responsible for the differences in petal color. Comparisons of deduced amino acid sequences encoded by the isolated anthocyanin cDNA fragments indicated that chalcone synthase (CHS), chalcone isomerase (CHI), anthocyanidin synthase 1 (ANS1) and ANS2 are identical in both aurantiaca and lutea varieties whereas minor amino acid differences of the deduced flavonone 3-hydroxylase (F3H) and dihydroflavonol 4-reductase (DFR) between both varieties were observed. The aurantiaca petals expressed substantially higher levels of transcripts representing CHS, F3H, DFR, ANS and UDP-glucose:flavonoid-3-O-glucosyltransferase genes, compared to lutea petals. Pelargonidin glycoside synthesis in aurantiaca petals therefore appears to reflect the higher steady-state levels of pelargonidin synthesis transcripts. Moreover, possible changes in the substrate specificity of DFR enzymes may represent additional mechanisms for producing red pelargonidin glycosides in petals of

  12. Red Anthocyanins and Yellow Carotenoids Form the Color of Orange-Flower Gentian (Gentiana lutea L. var. aurantiaca).

    PubMed

    Berman, Judit; Sheng, Yanmin; Gómez Gómez, Lourdes; Veiga, Tania; Ni, Xiuzhen; Farré, Gemma; Capell, Teresa; Guitián, Javier; Guitián, Pablo; Sandmann, Gerhard; Christou, Paul; Zhu, Changfu

    2016-01-01

    Flower color is an important characteristic that determines the commercial value of ornamental plants. Gentian flowers occur in a limited range of colors because this species is not widely cultivated as a cut flower. Gentiana lutea L. var. aurantiaca (abbr, aurantiaca) is characterized by its orange flowers, but the specific pigments responsible for this coloration are unknown. We therefore investigated the carotenoid and flavonoid composition of petals during flower development in the orange-flowered gentian variety of aurantiaca and the yellow-flowered variety of G. lutea L. var. lutea (abbr, lutea). We observed minor varietal differences in the concentration of carotenoids at the early and final stages, but only aurantiaca petals accumulated pelargonidin glycosides, whereas these compounds were not found in lutea petals. We cloned and sequenced the anthocyanin biosynthetic gene fragments from petals, and analyzed the expression of these genes in the petals of both varieties to determine the molecular mechanisms responsible for the differences in petal color. Comparisons of deduced amino acid sequences encoded by the isolated anthocyanin cDNA fragments indicated that chalcone synthase (CHS), chalcone isomerase (CHI), anthocyanidin synthase 1 (ANS1) and ANS2 are identical in both aurantiaca and lutea varieties whereas minor amino acid differences of the deduced flavonone 3-hydroxylase (F3H) and dihydroflavonol 4-reductase (DFR) between both varieties were observed. The aurantiaca petals expressed substantially higher levels of transcripts representing CHS, F3H, DFR, ANS and UDP-glucose:flavonoid-3-O-glucosyltransferase genes, compared to lutea petals. Pelargonidin glycoside synthesis in aurantiaca petals therefore appears to reflect the higher steady-state levels of pelargonidin synthesis transcripts. Moreover, possible changes in the substrate specificity of DFR enzymes may represent additional mechanisms for producing red pelargonidin glycosides in petals of

  13. Red Anthocyanins and Yellow Carotenoids Form the Color of Orange-Flower Gentian (Gentiana lutea L. var. aurantiaca)

    PubMed Central

    Gómez Gómez, Lourdes; Veiga, Tania; Ni, Xiuzhen; Farré, Gemma; Capell, Teresa; Guitián, Javier; Guitián, Pablo; Sandmann, Gerhard; Christou, Paul

    2016-01-01

    Flower color is an important characteristic that determines the commercial value of ornamental plants. Gentian flowers occur in a limited range of colors because this species is not widely cultivated as a cut flower. Gentiana lutea L. var. aurantiaca (abbr, aurantiaca) is characterized by its orange flowers, but the specific pigments responsible for this coloration are unknown. We therefore investigated the carotenoid and flavonoid composition of petals during flower development in the orange-flowered gentian variety of aurantiaca and the yellow-flowered variety of G. lutea L. var. lutea (abbr, lutea). We observed minor varietal differences in the concentration of carotenoids at the early and final stages, but only aurantiaca petals accumulated pelargonidin glycosides, whereas these compounds were not found in lutea petals. We cloned and sequenced the anthocyanin biosynthetic gene fragments from petals, and analyzed the expression of these genes in the petals of both varieties to determine the molecular mechanisms responsible for the differences in petal color. Comparisons of deduced amino acid sequences encoded by the isolated anthocyanin cDNA fragments indicated that chalcone synthase (CHS), chalcone isomerase (CHI), anthocyanidin synthase 1 (ANS1) and ANS2 are identical in both aurantiaca and lutea varieties whereas minor amino acid differences of the deduced flavonone 3-hydroxylase (F3H) and dihydroflavonol 4-reductase (DFR) between both varieties were observed. The aurantiaca petals expressed substantially higher levels of transcripts representing CHS, F3H, DFR, ANS and UDP-glucose:flavonoid-3-O-glucosyltransferase genes, compared to lutea petals. Pelargonidin glycoside synthesis in aurantiaca petals therefore appears to reflect the higher steady-state levels of pelargonidin synthesis transcripts. Moreover, possible changes in the substrate specificity of DFR enzymes may represent additional mechanisms for producing red pelargonidin glycosides in petals of

  14. Salmonella enterica serovar Typhimurium virulence-resistance plasmids derived from the pSLT carrying nonconventional class 1 integrons with dfrA12 gene in their variable region and sul3 in the 3' conserved segment.

    PubMed

    Beutlich, Janine; Rodicio, M Rosario; Mendoza, M Carmen; García, Patricia; Kirchner, Miranda; Luzzi, Ida; Mevius, Dik; Threlfall, John; Helmuth, Reiner; Guerra, Beatriz

    2013-12-01

    Drug-resistant derivatives of serovar-specific virulence plasmids, such as pSLT, in clinically-relevant Salmonella enterica serovar Typhimurium strains, represent a threat for human health. We have analysed 14 S. Typhimurium isolates recovered in Italy and the United Kingdom from swine and from cases of human infection for the presence of virulence-resistance (VR) plasmids. They were negative for the multidrug resistance (MDR) region of the Salmonella genomic island 1 (SGI1), but expressed resistance to ampicillin, chloramphenicol, streptomycin/spectinomycin, sulfamethoxazole, and tetracyclines. The isolates were characterised by XbaI pulsed-field gel electrophoresis, multilocus sequence typing, and detection of resistance and virulence determinants (PCR/sequencing). Identification of VR plasmids was accomplished by PCR detection of bla genes (encoding ampicillin resistance), class 1 integrons and the pSLT virulence gene spvC. Plasmid analyses were performed by alkaline lysis, S1-nuclease digestion, replicon typing, conjugation, restriction analyses, and Southern blot/hybridization. Two blaOXA-1 positive isolates contained pSLT-derived plasmids related to pUO-StVR2. In nine isolates, eight from swine and one from a patient, MDR-conferring-IncFII-VR plasmids were detected. They contained the blaTEM-1 gene as well as a nonconventional class 1 integron with dfrA12-aadA2 gene cassettes in its variable region, and a sul3 gene in the 3' conserved segment. Restriction analysis suggested a novel pSLT variant. The results obtained underline the role of swine as a potential reservoir for the blaTEM-1-IncFII-plasmids. The occurrence and spread of virulence- and MDR-conferring plasmids should be considered as a potential public health problem.

  15. Flower color diversity revealed by differential expression of flavonoid biosynthetic genes and flavonoid accumulation in herbaceous peony (Paeonia lactiflora Pall.).

    PubMed

    Zhao, Daqiu; Tao, Jun; Han, Chenxia; Ge, Jintao

    2012-12-01

    Herbaceous peony (Paeonia lactiflora Pall.) is an important ornamental plant which contains different flower colors. In this paper, eight genes encoding phenylalanine ammonialyase (PAL), chalcone synthase (CHS), chalcone isomerase (CHI), flavanone 3-hydroxylase (F3H), flavonoid 3'-hydroxylase (F3'H), dihydroflavonol 4-reductase (DFR), anthocyanidin synthase (ANS), UDP-glucose: flavonoid 3-o-glucosyltransferase (UF3GT) were isolated. Moreover, the expression patterns of these eight genes and UF5GT in the flowers were investigated in three cultivars, that is, 'Hongyanzhenghui', 'Yulouhongxing' and 'Huangjinlun' with purplish-red, white and yellow flower respectively. Furthermore, flavonoid accumulation in the flowers was also analyzed. The results showed that in different organs, most of genes expressed higher in flowers than in other organs. During the development of flowers, all genes could be divided into four groups. The first group (PlPAL) was highly expressed in S1 and S4. The second group (PlCHS and PlCHI) was at a high expression level throughout the whole developmental stages. The third group (PlF3H, PlF3'H, PlDFR, PlANS and PlUF5GT) gradually decreased with the development of flowers. The fourth group (PlUF3GT) gradually increased during the flower development. In addition, anthoxanthins and anthocyanins were detected in 'Hongyanzhenghui' and 'Yulouhongxing', chalcones and anthoxanthins were found in 'Huangjinlun'. When different color flowers were concerned, low expression level of PlCHI induced most of the substrate accumulation in the form of chalcones and displaying yellow, changing a small part of substrates to anthoxanthins, and there was no anthocyanin synthesis in 'Huangjinlun' because of low expression level of DFR. In 'Yulouhongxing', massive expressions of upstream genes and low expression of DFR caused synthesis of a great deal of anthoxanthins and a small amount of colorless anthocyanins. In 'Hongyanzhenghui', a large number of colored

  16. Transposon tagging of a male-sterility, female-sterility gene, St8, revealed that the meiotic MER3 DNA helicase activity is essential for fertility in soybean

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The W4 locus in soybean encodes a dihydroflavonol-4-reductase (DFR2) that regulates pigmentation patterns in flowers and hypocotyl. The mutable w4-m allele that governs variegated flowers has arisen through insertion of a CACTA-type transposable element, Tgm9, in DFR2. In the w4-m line, reversion fr...

  17. Mapping of an anthocyanin-regulating MYB transcription factor and its expression in red and green pear, Pyrus communis.

    PubMed

    Pierantoni, Luca; Dondini, Luca; De Franceschi, Paolo; Musacchi, Stefano; Winkel, Brenda S J; Sansavini, Silviero

    2010-12-01

    'Max Red Bartlett' is a red bud mutation of the yellow pear (Pyrus communis L.) cultivar 'Williams' (known as 'Bartlett' in North America). Anthocyanins are the most important pigments for red colour in fruits. Synthesis of anthocyanins is mediated by a number of well-characterized enzymes that include chalcone synthase (CHS), flavanone-3-hydroxylase (F3H), dihydroflavonol-4-reductase (DFR), anthocyanidin synthase (ANS), and UDP-glucose:flavonoid-3-O-glucosyltransferase (UFGT). Expression of the genes encoding these five enzymes was examined in pear fruit skin in order to elucidate the molecular mechanism for red coloration. In addition, the gene PcMYB10, encoding an R2R3 MYB transcription factor involved in anthocyanin biosynthetic pathway regulation, was isolated from both 'Williams' and 'Max Red Bartlett'. Analysis of the deduced amino acid sequence suggests that this gene is an ortholog of anthocyanin regulators known in other plant species. Its expression level was significantly higher in 'Max Red Bartlett' (red pear) compared with the original yellow variety 'Williams'. Although the map position of PcMYB10 corresponds to that of MdMYBa and MdMYB10, which control pigmentation of apple fruit skin, PcMYB10 is not directly responsible for red versus yellow colour in the two pear varieties, as the mutation underlying this difference maps to a different region of the pear genome.

  18. A bHLH transcription factor, DvIVS, is involved in regulation of anthocyanin synthesis in dahlia (Dahlia variabilis).

    PubMed

    Ohno, Sho; Hosokawa, Munetaka; Hoshino, Atsushi; Kitamura, Yoshikuni; Morita, Yasumasa; Park, Kyeung-Ii; Nakashima, Akiko; Deguchi, Ayumi; Tatsuzawa, Fumi; Doi, Motoaki; Iida, Shigeru; Yazawa, Susumu

    2011-10-01

    Dahlias (Dahlia variabilis) exhibit a wide range of flower colours because of accumulation of anthocyanin and other flavonoids in their ray florets. Two lateral mutants were used that spontaneously occurred in 'Michael J' (MJW) which has yellow ray florets with orange variegation. MJOr, a bud mutant producing completely orange ray florets, accumulates anthocyanins, flavones, and butein, and MJY, another mutant producing completely yellow ray florets, accumulates flavones and butein. Reverse transcription-PCR analysis showed that expression of chalcone synthase 1 (DvCHS1), flavanone 3-hydroxylase (DvF3H), dihydroflavonol 4-reductase (DvDFR), anthocyanidin synthase (DvANS), and DvIVS encoding a basic helix-loop-helix transcription factor were suppressed, whereas that of chalcone isomerase (DvCHI) and DvCHS2, another CHS with 69% nucleotide identity with DvCHS1, was not suppressed in the yellow ray florets of MJY. A 5.4 kb CACTA superfamily transposable element, transposable element of Dahlia variabilis 1 (Tdv1), was found in the fourth intron of the DvIVS gene of MJW and MJY, and footprints of Tdv1 were detected in the variegated flowers of MJW. It is shown that only one type of DvIVS gene was expressed in MJOr, whereas these plants are likely to have three types of the DvIVS gene. On the basis of these results, the mechanism regulating the formation of orange and yellow ray florets in dahlia is discussed. PMID:21765172

  19. Relationship between gene expression and the accumulation of catechin during spring and autumn in tea plants (Camellia sinensis L.)

    PubMed Central

    Liu, Min; Tian, Heng-lu; Wu, Jian-Hua; Cang, Ren-Rong; Wang, Run-Xian; Qi, Xiao-Hua; Xu, Qiang; Chen, Xue-Hao

    2015-01-01

    The tea plant (Camellia sinensis L.) is an important commercial crop with remarkably high catechin concentrations. Tea is popular worldwide given the plant's health benefits. Catechins are the main astringent substance in tea and are synthesized mainly via the phenylpropanoid pathway. In this study, eight cultivars of tea plants harvested both in spring and autumn were used to investigate differences in catechin concentrations by using high-performance liquid chromatography. The expression levels of genes associated with catechin biosynthesis were investigated using reverse transcription-quantitative polymerase chain reaction. The results indicated that the total catechin (TC) concentrations were significantly higher in tea plants harvested in autumn than in those harvested in spring, based on higher concentrations of epigallocatechin (EGC) in autumn tea (P<0.01). The expression of the genes phenylalanine ammonia-lyase (PAL), flavanone 3-hydroxylase (F3H), flavonoid 3′,5′-hydroxylase (F3′5′H), dihydroflavonol 4-reductase (DFR), and anthocyanidin synthase (ANS) is closely related to the TC content of tea plants in both spring and autumn. Positive correlations between PAL, cinnamate 4-hydroxylase (C4H), F3H, and DFR expression and EGC accumulation in autumn tea were identified, with correlation coefficients of 0.710, 0.763, 0.884, and 0.707, respectively. A negative correlation between ANS expression level and EGC concentrations in tea plants harvested in spring was noted (r=−0.732). Additionally, negative correlations between F3H and ANS expression levels and the catechin content were identified in spring tea, whereas the correlations were positive in autumn tea. Significant differences in the F3H and ANS expression levels between spring and autumn tea indicate that F3H and ANS are potentially key genes affecting catechin accumulation in tea plants. PMID:26504566

  20. Relationship between gene expression and the accumulation of catechin during spring and autumn in tea plants (Camellia sinensis L.).

    PubMed

    Liu, Min; Tian, Heng-Lu; Wu, Jian-Hua; Cang, Ren-Rong; Wang, Run-Xian; Qi, Xiao-Hua; Xu, Qiang; Chen, Xue-Hao

    2015-01-01

    The tea plant (Camellia sinensis L.) is an important commercial crop with remarkably high catechin concentrations. Tea is popular worldwide given the plant's health benefits. Catechins are the main astringent substance in tea and are synthesized mainly via the phenylpropanoid pathway. In this study, eight cultivars of tea plants harvested both in spring and autumn were used to investigate differences in catechin concentrations by using high-performance liquid chromatography. The expression levels of genes associated with catechin biosynthesis were investigated using reverse transcription-quantitative polymerase chain reaction. The results indicated that the total catechin (TC) concentrations were significantly higher in tea plants harvested in autumn than in those harvested in spring, based on higher concentrations of epigallocatechin (EGC) in autumn tea (P<0.01). The expression of the genes phenylalanine ammonia-lyase (PAL), flavanone 3-hydroxylase (F3H), flavonoid 3',5'-hydroxylase (F3'5'H), dihydroflavonol 4-reductase (DFR), and anthocyanidin synthase (ANS) is closely related to the TC content of tea plants in both spring and autumn. Positive correlations between PAL, cinnamate 4-hydroxylase (C4H), F3H, and DFR expression and EGC accumulation in autumn tea were identified, with correlation coefficients of 0.710, 0.763, 0.884, and 0.707, respectively. A negative correlation between ANS expression level and EGC concentrations in tea plants harvested in spring was noted (r=-0.732). Additionally, negative correlations between F3H and ANS expression levels and the catechin content were identified in spring tea, whereas the correlations were positive in autumn tea. Significant differences in the F3H and ANS expression levels between spring and autumn tea indicate that F3H and ANS are potentially key genes affecting catechin accumulation in tea plants. PMID:26504566

  1. Ultraviolet A-specific induction of anthocyanin biosynthesis in the swollen hypocotyls of turnip (Brassica rapa).

    PubMed

    Zhou, Bo; Li, Yuhua; Xu, Zhiru; Yan, Haifang; Homma, Shinichiro; Kawabata, Saneyuki

    2007-01-01

    Ultraviolet A (UV-A)-mediated regulation of anthocyanin biosynthesis was investigated in swollen hypocotyls of the red turnip 'Tsuda'. The shaded swollen hypocotyls which contained negligible anthocyanin were exposed to artificial light sources including low fluence UV-B, UV-A, blue, red, far-red, red plus UV-A, far-red plus UV-A, and blue plus red. Among these lights, only UV-A induced anthocyanin biosynthesis and co-irradiation of red or far-red with UV-A did not affect the extent of UV-A-induced anthocyanin accumulation. The expression of phenylalanine ammonia lyase (PAL; EC 4.3.1.5), chalcone synthase (CHS; EC 2.3.1.74), flavanone 3-hydroxylase (F3H; EC 1.14.11.9), dihydroflavonol 4-reductase (DFR; EC 1.1.1.219), and anthocyanidin synthase (ANS; EC 1.14.11.19) genes was increased with time during a 24 h exposure to UV-A. In contrast, irradiation with red, blue, UV-B, and a combination of blue with red failed to induce CHS expression. Microarray analysis showed that only a few genes, including CHS and F3H, were induced significantly by UV-A, while a separate set of many genes was induced by low fluence UV-B. The UV-A-specific induction of anthocyanin biosynthesis and the unique gene expression profile upon UV-A irradiation as compared with blue and UV-B demonstrated that the observed induction of anthocyanin biosynthesis in red turnips was mediated by a distinct UV-A-specific photoreceptor, but not by phytochromes, UV-A/blue photoreceptors, or UV-B photoreceptors.

  2. Comparative Leaves Transcriptome Analysis Emphasizing on Accumulation of Anthocyanins in Brassica: Molecular Regulation and Potential Interaction with Photosynthesis.

    PubMed

    Mushtaq, Muhammad A; Pan, Qi; Chen, Daozong; Zhang, Qinghua; Ge, Xianhong; Li, Zaiyun

    2016-01-01

    The purple leaf pigmentation mainly associated with anthocyanins accumulation is common in Brassica but the mechanisms of its production and its potential physiological functions are poorly understood. Here, we performed the phenotypic, cytological, physiological, and comparative leaves transcriptome analyses of 11 different varieties belonging to five Brassica species with purple or green leaves. We observed that the anthocyanin was accumulated in most of vegetative tissues in all species and also in reproduction organs of B. carinata. Anthocyanin accumulated in different part of purple leaves including adaxial and abaxial epidermal cells as well as palisade and spongy mesophyll cells. Leave transcriptome analysis showed that almost all late biosynthetic genes (LBGs) of anthocyanin, especially Dihydroflavonol 4-Reductase (DFR), Anthocyanidin Synthase (ANS) and Transparent Testa 19 (TT19), were highly up-regulated in all purple leaves. However, only one of transcript factors in anthocyanin biosynthesis pathway, Transparent Testa 8 (TT8), was up regulated along with those genes in all purple leaves, indicating its pivotal role for anthocyanin production in Brassica. Interestingly, with the up-regulation of genes for anthocyanin synthesis, Cytosolic 6-phosphogluconolactonase (PLG5) which involved in the oxidative pentose-phosphate pathway was up-regulated in all purple leaves and three genes FTSH PROTEASE 8 (FTS8), GLYCOLATE OXIDASE 1 (GOX1), and GLUTAMINE SYNTHETASE 1;4 (GLN1;4) related to degradation of photo-damaged proteins in photosystem II and light respiration were down-regulated. These results highlighted the potential physiological functions of anthocyanin accumulation related to photosynthesis which might be of great worth in future. PMID:27047501

  3. Effect of calcium on strawberry fruit flavonoid pathway gene expression and anthocyanin accumulation.

    PubMed

    Xu, Wenping; Peng, Hui; Yang, Tianbao; Whitaker, Bruce; Huang, Luhong; Sun, Jianghao; Chen, Pei

    2014-09-01

    Two diploid woodland strawberry (Fragaria vesca) inbred lines, Ruegen F7-4 (red fruit-bearing) and YW5AF7 (yellow fruit-bearing) were used to study the regulation of anthocyanin biosynthesis in fruit. Ruegen F7-4 fruit had similar total phenolics and anthocyanin contents to commercial octoploid (F. × ananassa) cultivar Seascape, while YW5AF7 exhibited relatively low total phenolics content and no anthocyanin accumulation. Foliar spray of CaCl2 boosted fruit total phenolics content, especially anthocyanins, by more than 20% in both Seascape and RF7-4. Expression levels of almost all the flavonoid pathway genes were comparable in Ruegen F7-4 and YW5AF7 green-stage fruit. However, at the turning and ripe stages, key anthocyanin structural genes, including flavanone 3-hydroxylase (F3H1), dihydroflavonol 4-reductase (DFR2), anthocyanidin synthase (ANS1), and UDP-glucosyltransferase (UGT1), were highly expressed in Ruegen F7-4 compared with YW5AF7 fruit. Calcium treatment further stimulated the expression of those genes in Ruegen F7-4 fruit. Anthocyanins isolated from petioles of YW5AF7 and Ruegen F-7 had the same HPLC-DAD profile, which differed from that of Ruegen F-7 fruit anthocyanins. All the anthocyanin structural genes except FvUGT1 were detected in petioles of YW5AF7 and Ruegen F-7. Taken together, these results indicate that the "yellow" gene in YW5AF7 is a fruit specific regulatory gene(s) for anthocyanin biosynthesis. Calcium can enhance accumulation of anthocyanins and total phenolics in fruit possibly via upregulation of anthocyanin structural genes. Our results also suggest that the anthocyanin biosynthesis machinery in petioles is different from that in fruit. PMID:25036468

  4. Effect of calcium on strawberry fruit flavonoid pathway gene expression and anthocyanin accumulation.

    PubMed

    Xu, Wenping; Peng, Hui; Yang, Tianbao; Whitaker, Bruce; Huang, Luhong; Sun, Jianghao; Chen, Pei

    2014-09-01

    Two diploid woodland strawberry (Fragaria vesca) inbred lines, Ruegen F7-4 (red fruit-bearing) and YW5AF7 (yellow fruit-bearing) were used to study the regulation of anthocyanin biosynthesis in fruit. Ruegen F7-4 fruit had similar total phenolics and anthocyanin contents to commercial octoploid (F. × ananassa) cultivar Seascape, while YW5AF7 exhibited relatively low total phenolics content and no anthocyanin accumulation. Foliar spray of CaCl2 boosted fruit total phenolics content, especially anthocyanins, by more than 20% in both Seascape and RF7-4. Expression levels of almost all the flavonoid pathway genes were comparable in Ruegen F7-4 and YW5AF7 green-stage fruit. However, at the turning and ripe stages, key anthocyanin structural genes, including flavanone 3-hydroxylase (F3H1), dihydroflavonol 4-reductase (DFR2), anthocyanidin synthase (ANS1), and UDP-glucosyltransferase (UGT1), were highly expressed in Ruegen F7-4 compared with YW5AF7 fruit. Calcium treatment further stimulated the expression of those genes in Ruegen F7-4 fruit. Anthocyanins isolated from petioles of YW5AF7 and Ruegen F-7 had the same HPLC-DAD profile, which differed from that of Ruegen F-7 fruit anthocyanins. All the anthocyanin structural genes except FvUGT1 were detected in petioles of YW5AF7 and Ruegen F-7. Taken together, these results indicate that the "yellow" gene in YW5AF7 is a fruit specific regulatory gene(s) for anthocyanin biosynthesis. Calcium can enhance accumulation of anthocyanins and total phenolics in fruit possibly via upregulation of anthocyanin structural genes. Our results also suggest that the anthocyanin biosynthesis machinery in petioles is different from that in fruit.

  5. A bHLH transcription factor, DvIVS, is involved in regulation of anthocyanin synthesis in dahlia (Dahlia variabilis)

    PubMed Central

    Ohno, Sho; Hosokawa, Munetaka; Hoshino, Atsushi; Kitamura, Yoshikuni; Morita, Yasumasa; Park, Kyeung-II; Nakashima, Akiko; Deguchi, Ayumi; Tatsuzawa, Fumi; Doi, Motoaki; Iida, Shigeru; Yazawa, Susumu

    2011-01-01

    Dahlias (Dahlia variabilis) exhibit a wide range of flower colours because of accumulation of anthocyanin and other flavonoids in their ray florets. Two lateral mutants were used that spontaneously occurred in ‘Michael J’ (MJW) which has yellow ray florets with orange variegation. MJOr, a bud mutant producing completely orange ray florets, accumulates anthocyanins, flavones, and butein, and MJY, another mutant producing completely yellow ray florets, accumulates flavones and butein. Reverse transcription–PCR analysis showed that expression of chalcone synthase 1 (DvCHS1), flavanone 3-hydroxylase (DvF3H), dihydroflavonol 4-reductase (DvDFR), anthocyanidin synthase (DvANS), and DvIVS encoding a basic helix–loop–helix transcription factor were suppressed, whereas that of chalcone isomerase (DvCHI) and DvCHS2, another CHS with 69% nucleotide identity with DvCHS1, was not suppressed in the yellow ray florets of MJY. A 5.4 kb CACTA superfamily transposable element, transposable element of Dahlia variabilis 1 (Tdv1), was found in the fourth intron of the DvIVS gene of MJW and MJY, and footprints of Tdv1 were detected in the variegated flowers of MJW. It is shown that only one type of DvIVS gene was expressed in MJOr, whereas these plants are likely to have three types of the DvIVS gene. On the basis of these results, the mechanism regulating the formation of orange and yellow ray florets in dahlia is discussed. PMID:21765172

  6. Comparative Leaves Transcriptome Analysis Emphasizing on Accumulation of Anthocyanins in Brassica: Molecular Regulation and Potential Interaction with Photosynthesis.

    PubMed

    Mushtaq, Muhammad A; Pan, Qi; Chen, Daozong; Zhang, Qinghua; Ge, Xianhong; Li, Zaiyun

    2016-01-01

    The purple leaf pigmentation mainly associated with anthocyanins accumulation is common in Brassica but the mechanisms of its production and its potential physiological functions are poorly understood. Here, we performed the phenotypic, cytological, physiological, and comparative leaves transcriptome analyses of 11 different varieties belonging to five Brassica species with purple or green leaves. We observed that the anthocyanin was accumulated in most of vegetative tissues in all species and also in reproduction organs of B. carinata. Anthocyanin accumulated in different part of purple leaves including adaxial and abaxial epidermal cells as well as palisade and spongy mesophyll cells. Leave transcriptome analysis showed that almost all late biosynthetic genes (LBGs) of anthocyanin, especially Dihydroflavonol 4-Reductase (DFR), Anthocyanidin Synthase (ANS) and Transparent Testa 19 (TT19), were highly up-regulated in all purple leaves. However, only one of transcript factors in anthocyanin biosynthesis pathway, Transparent Testa 8 (TT8), was up regulated along with those genes in all purple leaves, indicating its pivotal role for anthocyanin production in Brassica. Interestingly, with the up-regulation of genes for anthocyanin synthesis, Cytosolic 6-phosphogluconolactonase (PLG5) which involved in the oxidative pentose-phosphate pathway was up-regulated in all purple leaves and three genes FTSH PROTEASE 8 (FTS8), GLYCOLATE OXIDASE 1 (GOX1), and GLUTAMINE SYNTHETASE 1;4 (GLN1;4) related to degradation of photo-damaged proteins in photosystem II and light respiration were down-regulated. These results highlighted the potential physiological functions of anthocyanin accumulation related to photosynthesis which might be of great worth in future.

  7. A De novo Transcriptomic Approach to Identify Flavonoids and Anthocyanins "Switch-Off" in Olive (Olea europaea L.) Drupes at Different Stages of Maturation.

    PubMed

    Iaria, Domenico L; Chiappetta, Adriana; Muzzalupo, Innocenzo

    2015-01-01

    Highlights A de novo transcriptome reconstruction of olive drupes was performed in two genotypesGene expression was monitored during drupe development in two olive cultivarsTranscripts involved in flavonoid and anthocyanin pathways were analyzed in Cassanese and Leucocarpa cultivarsBoth cultivar and developmental stage impact gene expression in Olea europaea fruits. During ripening, the fruits of the olive tree (Olea europaea L.) undergo a progressive chromatic change characterized by the formation of a red-brown "spot" which gradually extends on the epidermis and in the innermost part of the mesocarp. This event finds an exception in the Leucocarpa cultivar, in which we observe a destabilized equilibrium between the metabolisms of chlorophyll and other pigments, particularly the anthocyanins whose switch-off during maturation promotes the white coloration of fruits. Despite its importance, genomic information on the olive tree is still lacking. Different RNA-seq libraries were generated from drupes of "Leucocarpa" and "Cassanese" olive genotypes, sampled at 100 and 130 days after flowering (DAF), and were used in order to identify transcripts involved in the main phenotypic changes of fruits during maturation and their corresponding expression patterns. A total of 103,359 transcripts were obtained and 3792 and 3064 were differentially expressed in "Leucocarpa" and "Cassanese" genotypes, respectively, during 100-130 DAF transition. Among them flavonoid and anthocyanin related transcripts such as phenylalanine ammonia lyase (PAL), cinnamate 4-hydroxylase (C4H), 4-coumarate-CoA ligase (4CL), chalcone synthase (CHS), chalcone isomerase (CHI), flavanone 3-hydroxylase (F3H), flavonol 3'-hydrogenase (F3'H), flavonol 3'5 '-hydrogenase (F3'5'H), flavonol synthase (FLS), dihydroflavonol 4-reductase (DFR), anthocyanidin synthase (ANS), UDP-glucose:anthocianidin: flavonoid glucosyltransferase (UFGT) were identified. These results contribute to reducing the current gap in

  8. Comparative Leaves Transcriptome Analysis Emphasizing on Accumulation of Anthocyanins in Brassica: Molecular Regulation and Potential Interaction with Photosynthesis

    PubMed Central

    Mushtaq, Muhammad A.; Pan, Qi; Chen, Daozong; Zhang, Qinghua; Ge, Xianhong; Li, Zaiyun

    2016-01-01

    The purple leaf pigmentation mainly associated with anthocyanins accumulation is common in Brassica but the mechanisms of its production and its potential physiological functions are poorly understood. Here, we performed the phenotypic, cytological, physiological, and comparative leaves transcriptome analyses of 11 different varieties belonging to five Brassica species with purple or green leaves. We observed that the anthocyanin was accumulated in most of vegetative tissues in all species and also in reproduction organs of B. carinata. Anthocyanin accumulated in different part of purple leaves including adaxial and abaxial epidermal cells as well as palisade and spongy mesophyll cells. Leave transcriptome analysis showed that almost all late biosynthetic genes (LBGs) of anthocyanin, especially Dihydroflavonol 4-Reductase (DFR), Anthocyanidin Synthase (ANS) and Transparent Testa 19 (TT19), were highly up-regulated in all purple leaves. However, only one of transcript factors in anthocyanin biosynthesis pathway, Transparent Testa 8 (TT8), was up regulated along with those genes in all purple leaves, indicating its pivotal role for anthocyanin production in Brassica. Interestingly, with the up-regulation of genes for anthocyanin synthesis, Cytosolic 6-phosphogluconolactonase (PLG5) which involved in the oxidative pentose-phosphate pathway was up-regulated in all purple leaves and three genes FTSH PROTEASE 8 (FTS8), GLYCOLATE OXIDASE 1 (GOX1), and GLUTAMINE SYNTHETASE 1;4 (GLN1;4) related to degradation of photo-damaged proteins in photosystem II and light respiration were down-regulated. These results highlighted the potential physiological functions of anthocyanin accumulation related to photosynthesis which might be of great worth in future. PMID:27047501

  9. A De novo Transcriptomic Approach to Identify Flavonoids and Anthocyanins "Switch-Off" in Olive (Olea europaea L.) Drupes at Different Stages of Maturation.

    PubMed

    Iaria, Domenico L; Chiappetta, Adriana; Muzzalupo, Innocenzo

    2015-01-01

    Highlights A de novo transcriptome reconstruction of olive drupes was performed in two genotypesGene expression was monitored during drupe development in two olive cultivarsTranscripts involved in flavonoid and anthocyanin pathways were analyzed in Cassanese and Leucocarpa cultivarsBoth cultivar and developmental stage impact gene expression in Olea europaea fruits. During ripening, the fruits of the olive tree (Olea europaea L.) undergo a progressive chromatic change characterized by the formation of a red-brown "spot" which gradually extends on the epidermis and in the innermost part of the mesocarp. This event finds an exception in the Leucocarpa cultivar, in which we observe a destabilized equilibrium between the metabolisms of chlorophyll and other pigments, particularly the anthocyanins whose switch-off during maturation promotes the white coloration of fruits. Despite its importance, genomic information on the olive tree is still lacking. Different RNA-seq libraries were generated from drupes of "Leucocarpa" and "Cassanese" olive genotypes, sampled at 100 and 130 days after flowering (DAF), and were used in order to identify transcripts involved in the main phenotypic changes of fruits during maturation and their corresponding expression patterns. A total of 103,359 transcripts were obtained and 3792 and 3064 were differentially expressed in "Leucocarpa" and "Cassanese" genotypes, respectively, during 100-130 DAF transition. Among them flavonoid and anthocyanin related transcripts such as phenylalanine ammonia lyase (PAL), cinnamate 4-hydroxylase (C4H), 4-coumarate-CoA ligase (4CL), chalcone synthase (CHS), chalcone isomerase (CHI), flavanone 3-hydroxylase (F3H), flavonol 3'-hydrogenase (F3'H), flavonol 3'5 '-hydrogenase (F3'5'H), flavonol synthase (FLS), dihydroflavonol 4-reductase (DFR), anthocyanidin synthase (ANS), UDP-glucose:anthocianidin: flavonoid glucosyltransferase (UFGT) were identified. These results contribute to reducing the current gap in

  10. Ultraviolet A-specific induction of anthocyanin biosynthesis in the swollen hypocotyls of turnip (Brassica rapa).

    PubMed

    Zhou, Bo; Li, Yuhua; Xu, Zhiru; Yan, Haifang; Homma, Shinichiro; Kawabata, Saneyuki

    2007-01-01

    Ultraviolet A (UV-A)-mediated regulation of anthocyanin biosynthesis was investigated in swollen hypocotyls of the red turnip 'Tsuda'. The shaded swollen hypocotyls which contained negligible anthocyanin were exposed to artificial light sources including low fluence UV-B, UV-A, blue, red, far-red, red plus UV-A, far-red plus UV-A, and blue plus red. Among these lights, only UV-A induced anthocyanin biosynthesis and co-irradiation of red or far-red with UV-A did not affect the extent of UV-A-induced anthocyanin accumulation. The expression of phenylalanine ammonia lyase (PAL; EC 4.3.1.5), chalcone synthase (CHS; EC 2.3.1.74), flavanone 3-hydroxylase (F3H; EC 1.14.11.9), dihydroflavonol 4-reductase (DFR; EC 1.1.1.219), and anthocyanidin synthase (ANS; EC 1.14.11.19) genes was increased with time during a 24 h exposure to UV-A. In contrast, irradiation with red, blue, UV-B, and a combination of blue with red failed to induce CHS expression. Microarray analysis showed that only a few genes, including CHS and F3H, were induced significantly by UV-A, while a separate set of many genes was induced by low fluence UV-B. The UV-A-specific induction of anthocyanin biosynthesis and the unique gene expression profile upon UV-A irradiation as compared with blue and UV-B demonstrated that the observed induction of anthocyanin biosynthesis in red turnips was mediated by a distinct UV-A-specific photoreceptor, but not by phytochromes, UV-A/blue photoreceptors, or UV-B photoreceptors. PMID:17426056

  11. Identification of flavonoids and expression of flavonoid biosynthetic genes in two coloured tree peony flowers.

    PubMed

    Zhao, Daqiu; Tang, Wenhui; Hao, Zhaojun; Tao, Jun

    2015-04-10

    Tree peony (Paeonia suffruticosa Andr.) has been named the "king of flowers" because of its elegant and gorgeous flower colour. Among these colours, the molecular mechanisms of white formation and how white turned to red in P. suffruticosa is little known. In this study, flower colour variables, flavonoid accumulation and expression of flavonoid biosynthetic genes of white ('Xueta') and red ('Caihui') P. suffruticosa were investigated. The results showed that the flower colours of both cultivars were gradually deepened with the development of flowers. Moreover, two anthoxanthin compositions apigenin 7-O-glucoside together with apigenin deoxyheso-hexoside were identified in 'Xueta' and 'Caihui', but one main anthocyanin composition peonidin 3,5-di-O-glucoside (Pn3G5G) was only found in 'Caihui'. Total contents of anthocyanins in 'Caihui' was increased during flower development, and the same trend was presented in anthoxanthins and flavonoids of these two cultivars, but the contents of these two category flavonoid in 'Caihui' were always higher than those in 'Xueta'. Furthermore, nine structural genes in flavonoid biosynthetic pathway were isolated including the full-length cDNAs of phenylalanine ammonialyase gene (PAL), chalcone synthase gene (CHS) and chalcone isomerase gene (CHI), together with the partial-length cDNAs of flavanone 3-hydroxylase gene (F3H), flavonoid 3'-hydroxylase gene (F3'H), dihydroflavonol 4-reductase gene (DFR), anthocyanidin synthase gene (ANS), UDP-glucose: flavonoid 3-O-glucosyltransferase gene (UF3GT) and UDP-glucose: flavonoid 5-O-glucosyltransferase gene (UF5GT), and PAL, UF3GT and UF5GT were reported in P. suffruticosa for the first time. Their expression patterns showed that transcription levels of downstream genes in 'Caihui' were basically higher than those in 'Xueta', especially PsDFR and PsANS, suggesting that these two genes may play a key role in the anthocyanin biosynthesis which resulted in the shift from white to red in

  12. Identification of flavonoids and expression of flavonoid biosynthetic genes in two coloured tree peony flowers.

    PubMed

    Zhao, Daqiu; Tang, Wenhui; Hao, Zhaojun; Tao, Jun

    2015-04-10

    Tree peony (Paeonia suffruticosa Andr.) has been named the "king of flowers" because of its elegant and gorgeous flower colour. Among these colours, the molecular mechanisms of white formation and how white turned to red in P. suffruticosa is little known. In this study, flower colour variables, flavonoid accumulation and expression of flavonoid biosynthetic genes of white ('Xueta') and red ('Caihui') P. suffruticosa were investigated. The results showed that the flower colours of both cultivars were gradually deepened with the development of flowers. Moreover, two anthoxanthin compositions apigenin 7-O-glucoside together with apigenin deoxyheso-hexoside were identified in 'Xueta' and 'Caihui', but one main anthocyanin composition peonidin 3,5-di-O-glucoside (Pn3G5G) was only found in 'Caihui'. Total contents of anthocyanins in 'Caihui' was increased during flower development, and the same trend was presented in anthoxanthins and flavonoids of these two cultivars, but the contents of these two category flavonoid in 'Caihui' were always higher than those in 'Xueta'. Furthermore, nine structural genes in flavonoid biosynthetic pathway were isolated including the full-length cDNAs of phenylalanine ammonialyase gene (PAL), chalcone synthase gene (CHS) and chalcone isomerase gene (CHI), together with the partial-length cDNAs of flavanone 3-hydroxylase gene (F3H), flavonoid 3'-hydroxylase gene (F3'H), dihydroflavonol 4-reductase gene (DFR), anthocyanidin synthase gene (ANS), UDP-glucose: flavonoid 3-O-glucosyltransferase gene (UF3GT) and UDP-glucose: flavonoid 5-O-glucosyltransferase gene (UF5GT), and PAL, UF3GT and UF5GT were reported in P. suffruticosa for the first time. Their expression patterns showed that transcription levels of downstream genes in 'Caihui' were basically higher than those in 'Xueta', especially PsDFR and PsANS, suggesting that these two genes may play a key role in the anthocyanin biosynthesis which resulted in the shift from white to red in

  13. A R2R3-MYB Transcription Factor Regulates the Flavonol Biosynthetic Pathway in a Traditional Chinese Medicinal Plant, Epimedium sagittatum.

    PubMed

    Huang, Wenjun; Khaldun, A B M; Chen, Jianjun; Zhang, Chanjuan; Lv, Haiyan; Yuan, Ling; Wang, Ying

    2016-01-01

    Flavonols as plant secondary metabolites with vital roles in plant development and defense against UV light, have been demonstrated to be the main bioactive components (BCs) in the genus Epimedium plants, several species of which are used as materials for Herba Epimedii, an important traditional Chinese medicine. The flavonol biosynthetic pathway genes had been already isolated from Epimedium sagittatum, but a R2R3-MYB transcription factor regulating the flavonol synthesis has not been functionally characterized so far in Epimedium plants. In this study, we isolated and characterized the R2R3-MYB transcription factor EsMYBF1 involved in regulation of the flavonol biosynthetic pathway from E. sagittatum. Sequence analysis indicated that EsMYBF1 belongs to the subgroup 7 of R2R3-MYB family which contains the flavonol-specific MYB regulators identified to date. Transient reporter assay showed that EsMYBF1 strongly activated the promoters of EsF3H (flavanone 3-hydroxylase) and EsFLS (flavonol synthase), but not the promoters of EsDFRs (dihydroflavonol 4-reductase) and EsANS (anthocyanidin synthase) in transiently transformed Nicotiana benthamiana leaves. Both yeast two-hybrid assay and transient reporter assay validated EsMYBF1 to be independent of EsTT8, or AtTT8 bHLH regulators of the flavonoid pathway as cofactors. Ectopic expression of EsMYBF1 in transgenic tobacco resulted in the increased flavonol content and the decreased anthocyanin content in flowers. Correspondingly, the structural genes involved in flavonol synthesis were upregulated in the EsMYBF1 overexpression lines, including NtCHS (chalcone synthase), NtCHI (chalcone isomerase), NtF3H and NtFLS, whereas the late biosynthetic genes of the anthocyanin pathway (NtDFR and NtANS) were remarkably downregulated, compared to the controls. These results suggest that EsMYBF1 is a flavonol-specific R2R3-MYB regulator, and involved in regulation of the biosynthesis of the flavonol-derived BCs in E. sagittatum. Thus

  14. Comparative Transcriptome Analysis of Genes Involved in Anthocyanin Biosynthesis in the Red and Yellow Fruits of Sweet Cherry (Prunus avium L.)

    PubMed Central

    Wei, Hairong; Chen, Xin; Zong, Xiaojuan; Shu, Huairui; Gao, Dongsheng; Liu, Qingzhong

    2015-01-01

    Background Fruit color is one of the most important economic traits of the sweet cherry (Prunus avium L.). The red coloration of sweet cherry fruit is mainly attributed to anthocyanins. However, limited information is available regarding the molecular mechanisms underlying anthocyanin biosynthesis and its regulation in sweet cherry. Methodology/Principal Findings In this study, a reference transcriptome of P. avium L. was sequenced and annotated to identify the transcriptional determinants of fruit color. Normalized cDNA libraries from red and yellow fruits were sequenced using the next-generation Illumina/Solexa sequencing platform and de novo assembly. Over 66 million high-quality reads were assembled into 43,128 unigenes using a combined assembly strategy. Then a total of 22,452 unigenes were compared to public databases using homology searches, and 20,095 of these unigenes were annotated in the Nr protein database. Furthermore, transcriptome differences between the four stages of fruit ripening were analyzed using Illumina digital gene expression (DGE) profiling. Biological pathway analysis revealed that 72 unigenes were involved in anthocyanin biosynthesis. The expression patterns of unigenes encoding phenylalanine ammonia-lyase (PAL), 4-coumarate-CoA ligase (4CL), chalcone synthase (CHS), chalcone isomerase (CHI), flavanone 3-hydroxylase (F3H), flavanone 3’-hydroxylase (F3’H), dihydroflavonol 4-reductase (DFR), anthocyanidin synthase (ANS) and UDP glucose: flavonol 3-O-glucosyltransferase (UFGT) during fruit ripening differed between red and yellow fruit. In addition, we identified some transcription factor families (such as MYB, bHLH and WD40) that may control anthocyanin biosynthesis. We confirmed the altered expression levels of eighteen unigenes that encode anthocyanin biosynthetic enzymes and transcription factors using quantitative real-time PCR (qRT-PCR). Conclusions/Significance The obtained sweet cherry transcriptome and DGE profiling data

  15. A R2R3-MYB transcription factor from Epimedium sagittatum regulates the flavonoid biosynthetic pathway.

    PubMed

    Huang, Wenjun; Sun, Wei; Lv, Haiyan; Luo, Ming; Zeng, Shaohua; Pattanaik, Sitakanta; Yuan, Ling; Wang, Ying

    2013-01-01

    Herba epimedii (Epimedium), a traditional Chinese medicine, has been widely used as a kidney tonic and antirheumatic medicine for thousands of years. The bioactive components in herba epimedii are mainly prenylated flavonol glycosides, end-products of the flavonoid pathway. Epimedium species are also used as garden plants due to the colorful flowers and leaves. Many R2R3-MYB transcription factors (TFs) have been identified to regulate the flavonoid and anthocyanin biosynthetic pathways. However, little is known about the R2R3-MYB TFs involved in regulation of the flavonoid pathway in Epimedium. Here, we reported the isolation and functional characterization of the first R2R3-MYB TF (EsMYBA1) from Epimedium sagittatum (Sieb. Et Zucc.) Maxim. Conserved domains and phylogenetic analysis showed that EsMYBA1 belonged to the subgroup 6 clade (anthocyanin-related MYB clade) of R2R3-MYB family, which includes Arabidopsis AtPAP1, apple MdMYB10 and legume MtLAP1. EsMYBA1 was preferentially expressed in leaves, especially in red leaves that contain higher content of anthocyanin. Alternative splicing of EsMYBA1 resulted in three transcripts and two of them encoded a MYB-related protein. Yeast two-hybrid and transient luciferase expression assay showed that EsMYBA1 can interact with several bHLH regulators of the flavonoid pathway and activate the promoters of dihydroflavonol 4-reductase (DFR) and anthocyanidin synthase (ANS). In both transgenic tobacco and Arabidopsis, overexpression of EsMYBA1 induced strong anthocyanin accumulation in reproductive and/or vegetative tissues via up-regulation of the main flavonoid-related genes. Furthermore, transient expression of EsMYBA1 in E. sagittatum leaves by Agrobacterium infiltration also induced anthocyanin accumulation in the wounded area. This first functional characterization of R2R3-MYB TFs in Epimedium species will promote further studies of the flavonoid biosynthesis and regulation in medicinal plants. PMID:23936468

  16. A De novo Transcriptomic Approach to Identify Flavonoids and Anthocyanins “Switch-Off” in Olive (Olea europaea L.) Drupes at Different Stages of Maturation

    PubMed Central

    Iaria, Domenico L.; Chiappetta, Adriana; Muzzalupo, Innocenzo

    2016-01-01

    Highlights A de novo transcriptome reconstruction of olive drupes was performed in two genotypesGene expression was monitored during drupe development in two olive cultivarsTranscripts involved in flavonoid and anthocyanin pathways were analyzed in Cassanese and Leucocarpa cultivarsBoth cultivar and developmental stage impact gene expression in Olea europaea fruits. During ripening, the fruits of the olive tree (Olea europaea L.) undergo a progressive chromatic change characterized by the formation of a red-brown “spot” which gradually extends on the epidermis and in the innermost part of the mesocarp. This event finds an exception in the Leucocarpa cultivar, in which we observe a destabilized equilibrium between the metabolisms of chlorophyll and other pigments, particularly the anthocyanins whose switch-off during maturation promotes the white coloration of fruits. Despite its importance, genomic information on the olive tree is still lacking. Different RNA-seq libraries were generated from drupes of “Leucocarpa” and “Cassanese” olive genotypes, sampled at 100 and 130 days after flowering (DAF), and were used in order to identify transcripts involved in the main phenotypic changes of fruits during maturation and their corresponding expression patterns. A total of 103,359 transcripts were obtained and 3792 and 3064 were differentially expressed in “Leucocarpa” and “Cassanese” genotypes, respectively, during 100–130 DAF transition. Among them flavonoid and anthocyanin related transcripts such as phenylalanine ammonia lyase (PAL), cinnamate 4-hydroxylase (C4H), 4-coumarate-CoA ligase (4CL), chalcone synthase (CHS), chalcone isomerase (CHI), flavanone 3-hydroxylase (F3H), flavonol 3′-hydrogenase (F3′H), flavonol 3′5 ′-hydrogenase (F3′5′H), flavonol synthase (FLS), dihydroflavonol 4-reductase (DFR), anthocyanidin synthase (ANS), UDP-glucose:anthocianidin: flavonoid glucosyltransferase (UFGT) were identified. These results contribute

  17. A R2R3-MYB Transcription Factor Regulates the Flavonol Biosynthetic Pathway in a Traditional Chinese Medicinal Plant, Epimedium sagittatum

    PubMed Central

    Huang, Wenjun; Khaldun, A. B. M.; Chen, Jianjun; Zhang, Chanjuan; Lv, Haiyan; Yuan, Ling; Wang, Ying

    2016-01-01

    Flavonols as plant secondary metabolites with vital roles in plant development and defense against UV light, have been demonstrated to be the main bioactive components (BCs) in the genus Epimedium plants, several species of which are used as materials for Herba Epimedii, an important traditional Chinese medicine. The flavonol biosynthetic pathway genes had been already isolated from Epimedium sagittatum, but a R2R3-MYB transcription factor regulating the flavonol synthesis has not been functionally characterized so far in Epimedium plants. In this study, we isolated and characterized the R2R3-MYB transcription factor EsMYBF1 involved in regulation of the flavonol biosynthetic pathway from E. sagittatum. Sequence analysis indicated that EsMYBF1 belongs to the subgroup 7 of R2R3-MYB family which contains the flavonol-specific MYB regulators identified to date. Transient reporter assay showed that EsMYBF1 strongly activated the promoters of EsF3H (flavanone 3-hydroxylase) and EsFLS (flavonol synthase), but not the promoters of EsDFRs (dihydroflavonol 4-reductase) and EsANS (anthocyanidin synthase) in transiently transformed Nicotiana benthamiana leaves. Both yeast two-hybrid assay and transient reporter assay validated EsMYBF1 to be independent of EsTT8, or AtTT8 bHLH regulators of the flavonoid pathway as cofactors. Ectopic expression of EsMYBF1 in transgenic tobacco resulted in the increased flavonol content and the decreased anthocyanin content in flowers. Correspondingly, the structural genes involved in flavonol synthesis were upregulated in the EsMYBF1 overexpression lines, including NtCHS (chalcone synthase), NtCHI (chalcone isomerase), NtF3H and NtFLS, whereas the late biosynthetic genes of the anthocyanin pathway (NtDFR and NtANS) were remarkably downregulated, compared to the controls. These results suggest that EsMYBF1 is a flavonol-specific R2R3-MYB regulator, and involved in regulation of the biosynthesis of the flavonol-derived BCs in E. sagittatum. Thus

  18. Silencing of flavanone-3-hydroxylase in apple (Malus × domestica Borkh.) leads to accumulation of flavanones, but not to reduced fire blight susceptibility.

    PubMed

    Flachowsky, Henryk; Halbwirth, Heidi; Treutter, Dieter; Richter, Klaus; Hanke, Magda-Viola; Szankowski, Iris; Gosch, Christian; Stich, Karl; Fischer, Thilo C

    2012-02-01

    Transgenic antisense flavanone-3-hydroxylase apple plants were produced to mimic the effect of the agrochemical prohexadione-Ca on apple leaves. This enzyme inhibitor for 2-oxoglutarate dependent dioxygenases is used as a growth retardant and for control of secondary fire blight of leaves. Like using the agent, silencing of flavanone-3-hydroxylase leads to an accumulation of flavanones in leaves, but in contrast not to the formation of 3-deoxyflavonoids. In prohexadione-Ca treated leaves the 3-deoxyflavonoid luteoforol is formed from accumulating flavanones, acting as an antimicrobial compound against the fire blight pathogen Erwinia amylovora. Seemingly, the silencing of just one of the 2-oxoglutarate dependent dioxygenases (in apple also flavonol synthase and anthocyanidin synthase take part downstream in the pathway) does not provide a sufficiently high ratio of flavanones to dihydroflavonols. This seems to be needed to let the dihydroflavonol-4-reductase/flavanone-4-reductase enzyme reduce flavanones to luteoforol, and to let this be reduced by the leucoanthocyanidin-4-reductase/3-deoxyleucoanthocyanidin-4-reductase, each acting with their respective weak secondary activities. Accordingly, also the intended inducible resistance to fire blight by prohexadione-Ca is not observed with the antisense flavanone-3-hydroxylase apple plants. On the other hand, for most transgenic lines with strong flavanone-4-reductase down-regulation, up-regulation of gene expression for the other flavonoid genes was found. This provides further evidence for the feedback regulation of flavonoid gene expression having been previously reported for the prohexadione-Ca inhibited apple plants.

  19. Design-for-reliability (DfR) of aerospace electronics: Attributes and challenges

    NASA Astrophysics Data System (ADS)

    Bensoussan, A.; Suhir, E.

    The next generation of multi-beam satellite systems that would be able to provide effective interactive communication services will have to operate within a highly flexible architecture. One option to develop such flexibility is to employ microwaves and/or optoelectronic components and to make them reliable. The use of optoelectronic devices, equipments and systems will result indeed in significant improvement in the state-of-the-art only provided that the new designs will suggest a novel and effective architecture that will combine the merits of good functional performance, satisfactory mechanical (structural) reliability and high cost effectiveness. The obvious challenge is the ability to design and fabricate equipment based on EEE components that would be able to successfully withstand harsh space environments for the entire duration of the mission. It is imperative that the major players in the space industry, such as manufacturers, industrial users, and space agencies, understand the importance and the limits of the achievable quality and reliability of optoelectronic devices operated in harsh environments. It is equally imperative that the physics of possible failures is well understood and, if necessary, minimized, and that adequate Quality Standards are developed and employed. The space community has to identify and to develop the strategic approach for validating optoelectronic products. This should be done with consideration of numerous intrinsic and extrinsic requirements for the systems' performance. When considering a particular next generation optoelectronic space system, the space community needs to address the following major issues: proof of concept for this system, proof of reliability and proof of performance. This should be done with taking into account the specifics of the anticipated application. High operational reliability cannot be left to the prognostics and health monitoring/management (PHM) effort and stage, no matter how important and - ffective such an effort might be. Reliability should be pursued at all the stages of the equipment lifetime: design, product development, manufacturing, burn-in testing and, of course, subsequent PHM after the space apparatus is launched and operated.

  20. Anthocyanidins and polyphenols in five brassica species microgreens: analysis by UHPLC-PDA-ESI/HRMS/MSn

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Brassica vegetables are known to contain relatively high concentrations of bioactive compounds associated with human health. A comprehensive profiling of polyphenols from five Brassica species microgreens was conducted using ultra high-performance liquid chromatography photo diode array high-resolu...

  1. Improving the Analysis of Anthocyanidins from Blueberries Using Response Surface Methodology

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Background: Recent interest in the health promoting potential of anthocyanins points to the need for robust and reliable analytical methods. It is essential to know that the health promoting chemicals are present in juices and other products processed from whole fruit. Many different methods have be...

  2. Cloning and characterization of a potato StAN11 gene involved in anthocyanin biosynthesis regulation.

    PubMed

    Li, Wang; Wang, Bing; Wang, Man; Chen, Min; Yin, Jing-Ming; Kaleri, Ghullam Murtaza; Zhang, Rui-Jie; Zuo, Tie-Niu; You, Xiong; Yang, Qing

    2014-04-01

    Anthocyanins are a class of products of plant secondary metabolism and are responsible for tubers color in potato. The biosynthesis of anthocyanins is a complex biological process, in which multiple genes are involved including structural genes and regulatory genes. In this study, StAN11, a WD40-repeat gene, was cloned from potato cultivar Chieftain (Solanum tuberosum L.). StAN11 (HQ599506) contained no intron and its open reading frame (ORF) was 1,029 bp long, encoding a putative protein of 342 amino acids. In order to verify its role in anthocyanin biosynthesis, StAN11 was inserted behind the CaMV-35S promoter of pCMBIA1304 and the recombination vector was introduced into the potato cultivar Désirée plants by Agrobacterium-mediated transformation. The color of transgenic tuber skin was significantly deepened, compared to the wild-type control, which was highly consistent with the accumulation of anthocyanin and expression of StAN11 in transgenic lines tuber skin. Further analysis on the expression of Flavonone-3-hydroxylase (F3H), Dihydroflavonol reductase (DFR), Anthocyanidin synthase (ANS), and Flavonoid 3-O-glucosyl transferase (3GT) in transgenic plants revealed that only DFR was upregulated. This result suggested that StAN11 regulated anthocyanin biosynthesis in potato by controlling DFR expression and accumulation of anthocyanin could be increased through overexpression of StAN11 in the tubers with the genetic background of anthocyanin biosynthesis. PMID:24304603

  3. Elucidation of Molecular Identity of the W3 Locus and Its Implication in Determination of Flower Colors in Soybean

    PubMed Central

    Park, Gyu Tae; Sundaramoorthy, Jagadeesh; Lee, Jeong-Dong; Kim, Jeong Hoe; Seo, Hak Soo; Song, Jong Tae

    2015-01-01

    The wide range of flower colors in soybean is controlled by six independent loci (W1, W2, W3, W4, Wm, and Wp). Among these loci, mutations in the W3 locus under the w4 allelic background (i.e., w3w4) produce near-white flowers, while the W3w4 genotype produces purple throat flowers. Although a gene encoding dihydroflavonol 4-reductase, DFR1, has been known to be closely associated with the W3 locus, its molecular identity has not yet been characterized. In the present study, we aimed to determine whether DFR1 is responsible for allelic variations in the W3 locus. On the basis of the sequence of a DFR probe, Glyma.14G072700 was identified as a candidate gene for DFR1, and nucleotide sequences of Glyma.14G072700 from cultivars with previously validated genotypes for the W3 locus were determined. As a result, a number of nucleotide polymorphisms, mainly single-base substitutions, between both coding and 5′-upstream region sequences of the W3 and w3 alleles were identified. Among them, an indel of 311-bp in the 5′-upstream region was noteworthy, since the Glyma.14G072700 in all the w3 alleles examined contained the indel, whereas that in all the W3 alleles did not; the former was barely expressed, but the latter was well expressed. These results suggest that Glyma.14G072700 is likely to correspond to DFR1 for the W3 locus and that its expression patterns may lead to allelic color phenotypes of W3 and w3 alleles under the w4 allelic background. PMID:26555888

  4. Elucidation of Molecular Identity of the W3 Locus and Its Implication in Determination of Flower Colors in Soybean.

    PubMed

    Park, Gyu Tae; Sundaramoorthy, Jagadeesh; Lee, Jeong-Dong; Kim, Jeong Hoe; Seo, Hak Soo; Song, Jong Tae

    2015-01-01

    The wide range of flower colors in soybean is controlled by six independent loci (W1, W2, W3, W4, Wm, and Wp). Among these loci, mutations in the W3 locus under the w4 allelic background (i.e., w3w4) produce near-white flowers, while the W3w4 genotype produces purple throat flowers. Although a gene encoding dihydroflavonol 4-reductase, DFR1, has been known to be closely associated with the W3 locus, its molecular identity has not yet been characterized. In the present study, we aimed to determine whether DFR1 is responsible for allelic variations in the W3 locus. On the basis of the sequence of a DFR probe, Glyma.14G072700 was identified as a candidate gene for DFR1, and nucleotide sequences of Glyma.14G072700 from cultivars with previously validated genotypes for the W3 locus were determined. As a result, a number of nucleotide polymorphisms, mainly single-base substitutions, between both coding and 5'-upstream region sequences of the W3 and w3 alleles were identified. Among them, an indel of 311-bp in the 5'-upstream region was noteworthy, since the Glyma.14G072700 in all the w3 alleles examined contained the indel, whereas that in all the W3 alleles did not; the former was barely expressed, but the latter was well expressed. These results suggest that Glyma.14G072700 is likely to correspond to DFR1 for the W3 locus and that its expression patterns may lead to allelic color phenotypes of W3 and w3 alleles under the w4 allelic background. PMID:26555888

  5. Improvement of catechin production in Escherichia coli through combinatorial metabolic engineering.

    PubMed

    Zhao, Shujuan; Jones, J Andrew; Lachance, Daniel M; Bhan, Namita; Khalidi, Omar; Venkataraman, Sylesh; Wang, Zhengtao; Koffas, Mattheos A G

    2015-03-01

    Reconstruction of highly efficient biosynthesis pathways is essential for the production of valuable plant secondary metabolites in recombinant microorganisms. In order to improve the titer of green tea catechins in Escherichia coli, combinatorial strategies were employed using the ePathBrick vectors to express the committed catechin pathway: flavanone 3β-hydroxylase (F3H), dihydroflavonol 4-reductase (DFR), and leucoanthocyanidin reductase (LAR). Three F3H, three DFR, and two LAR genes originating from different plant species were selected and synthesized, to create 18 pathway variants to be screened in E. coli. Constructs containing F3H(syn) originally from Camellia sinensis, DFR(syn) from Anthurium andraeanum, C. sinensis, or Fragaria ananass, and LAR(syn) from Desmodium uncinatum (p148, p158 and p168) demonstrated high conversion efficiency with either eriodictyol or naringenin as substrate. A highly efficient construct was created by assembling additional copies of DFR(syn) and LAR(syn) enabling a titer of 374.6 ± 43.6 mg/L of (+)-catechin. Improving the NADPH availability via the ΔpgiΔppc mutation, BLΔpgiΔppc-p148 produced the highest titer of catechin at 760.9 ± 84.3 mg/L. After utilizing a library of scaffolding proteins, the strain BLΔpgiΔppc-p168-759 reached the highest titer of (+)-catechin of 910.9 ± 61.3 mg/L from 1.0 g/L of eriodictyol in batch culture with M9 minimal media. The impact of oxygen availability on the biosynthesis of catechin was also investigated. PMID:25527438

  6. Flavor of fresh blueberry juice and the comparison to amount of sugars, acids, anthocyanidins, and physicochemical measurements

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Six cultivars of southern highbush (SHB) and rabbiteye (RE) blueberry samples were harvested twice. Each treatment combination was pressed two times for repeated measures. Fresh juice was characterized for eighteen flavor/taste/feeling factor attributes by a descriptive flavor panel. Each sample ...

  7. Complete assignment of structural genes involved in flavonoid biosynthesis influencing bulb color to individual chromosomes of the shallot (Allium cepa L.).

    PubMed

    Masuzaki, Shin-ichi; Shigyo, Masayoshi; Yamauchi, Naoki

    2006-08-01

    We analyzed Japanese bunching onion (Allium fistulosum L.) - shallot (Allium cepa L. Aggregatum group) alien chromosome addition lines in order to assign the genes involved in the flavonoid biosynthesis pathway to chromosomes of the shallot. Two complete sets of alien monosomic additions (2n = 2x + 1 = 17) were used for determining the chromosomal locations of several partial sequences of candidate genes, CHS, CHI, F3H, DFR, and ANS via analyses of PCR-based markers. The results of DNA marker analyses showed that the CHS-A, CHS-B, CHI, F3H, DFR, and ANS genes should be assigned to chromosomes 2A, 4A, 3A, 3A, 7A, and 4A, respectively. HPLC analyses of 14 A. fistulosum - shallot multiple alien additions (2n = 2x + 2 - 2x + 7 = 18 - 23) were conducted to identify the anthocyanin compounds produced in the scaly leaves. A direct comparison between the genomic constitution and the anthocyanin compositions of the multiple additions revealed that a 3GT gene for glucosylation of anthocyanidin was located on 4A. Thus, we were able to assign all structural genes involved in flavonoid biosynthesis influencing bulb color to individual chromosomes of A. cepa.

  8. Anthocyanin accumulation and expression of anthocyanin biosynthetic genes in radish (Raphanus sativus).

    PubMed

    Park, Nam Il; Xu, Hui; Li, Xiaohua; Jang, In Hyuk; Park, Suhyoung; Ahn, Gil Hwan; Lim, Yong Pyo; Kim, Sun Ju; Park, Sang Un

    2011-06-01

    Radish [Raphanus sativus (Rs)] is an important dietary vegetable in Asian countries, especially China, Japan, and Korea. To elucidate the molecular mechanisms of anthocyanin accumulation in radish, the gene expression of enzymes directly involved in anthocyanin biosynthesis was analyzed. These genes include phenylalanine ammonia lyase (PAL), cinnamate 4-hydroxylase (C4H), 4-coumarate-CoA ligase (4CL), chalcone synthase (CHS), chalcone isomerase (CHI), flavanone 3-hydroxylase (F3H), dihydroflavonol reductase (DFR), and anthocyanidin synthase (ANS). RsDFR and RsANS were found to accumulate in the flesh or skin of two radish cultivars (Man Tang Hong and Hong Feng No.1). Radish skin contained higher CHS, CHI, and F3H transcript levels than radish flesh in all three cultivars. In the red radish, 16 anthocyanins were separated and identified by high-performance liquid chromatography (HPLC) and elctrospray ionization-tandem mass spectrometry (ESI-MS/MS). Some of them were acylated with coumaroyl, malonoyl, feruoyl, and caffeoyl moieties. Furthermore (-)-epicatechin and ferulic acid were also identified in the three cultivars.

  9. Arabidopsis CAPRICE (MYB) and GLABRA3 (bHLH) control tomato (Solanum lycopersicum) anthocyanin biosynthesis.

    PubMed

    Wada, Takuji; Kunihiro, Asuka; Tominaga-Wada, Rumi

    2014-01-01

    In Arabidopsis thaliana the MYB transcription factor CAPRICE (CPC) and the bHLH transcription factor GLABRA3 (GL3) are central regulators of root-hair differentiation and trichome initiation. By transforming the orthologous tomato genes SlTRY (CPC) and SlGL3 (GL3) into Arabidopsis, we demonstrated that these genes influence epidermal cell differentiation in Arabidopsis, suggesting that tomato and Arabidopsis partially use similar transcription factors for epidermal cell differentiation. CPC and GL3 are also known to be involved in anthocyanin biosynthesis. After transformation into tomato, 35S::CPC inhibited anthocyanin accumulation, whereas GL3::GL3 enhanced anthocyanin accumulation. Real-time reverse transcription PCR analyses showed that the expression of anthocyanin biosynthetic genes including Phe-ammonia lyase (PAL), the flavonoid pathway genes chalcone synthase (CHS), dihydroflavonol reductase (DFR), and anthocyanidin synthase (ANS) were repressed in 35S::CPC tomato. In contrast, the expression levels of PAL, CHS, DFR, and ANS were significantly higher in GL3::GL3 tomato compared with control plants. These results suggest that CPC and GL3 also influence anthocyanin pigment synthesis in tomato. PMID:25268379

  10. Complete assignment of structural genes involved in flavonoid biosynthesis influencing bulb color to individual chromosomes of the shallot (Allium cepa L.).

    PubMed

    Masuzaki, Shin-ichi; Shigyo, Masayoshi; Yamauchi, Naoki

    2006-08-01

    We analyzed Japanese bunching onion (Allium fistulosum L.) - shallot (Allium cepa L. Aggregatum group) alien chromosome addition lines in order to assign the genes involved in the flavonoid biosynthesis pathway to chromosomes of the shallot. Two complete sets of alien monosomic additions (2n = 2x + 1 = 17) were used for determining the chromosomal locations of several partial sequences of candidate genes, CHS, CHI, F3H, DFR, and ANS via analyses of PCR-based markers. The results of DNA marker analyses showed that the CHS-A, CHS-B, CHI, F3H, DFR, and ANS genes should be assigned to chromosomes 2A, 4A, 3A, 3A, 7A, and 4A, respectively. HPLC analyses of 14 A. fistulosum - shallot multiple alien additions (2n = 2x + 2 - 2x + 7 = 18 - 23) were conducted to identify the anthocyanin compounds produced in the scaly leaves. A direct comparison between the genomic constitution and the anthocyanin compositions of the multiple additions revealed that a 3GT gene for glucosylation of anthocyanidin was located on 4A. Thus, we were able to assign all structural genes involved in flavonoid biosynthesis influencing bulb color to individual chromosomes of A. cepa. PMID:17038797

  11. New member of the R2R3-MYB transcription factors family in grapevine suppresses the anthocyanin accumulation in the flowers of transgenic tobacco.

    PubMed

    Pérez-Díaz, J Ricardo; Pérez-Díaz, Jorge; Madrid-Espinoza, José; González-Villanueva, Enrique; Moreno, Yerko; Ruiz-Lara, Simón

    2016-01-01

    In grapevine, anthocyanins and proanthocyanidins are the main flavonoids in berries, which are associated to organoleptic properties in red wine such as color and astringency. Flavonoid pathway is specifically regulated at transcriptional level and several R2R3-MYB proteins have shown to act as positive regulators. However, some members of this family have shown to repress the flavonoid biosynthesis. In this work, we present the characterization of VvMYB4-like gene, which encodes a putative transcriptional factor highly expressed in the skin of berries at the pre veraison stage in grapevine. Its over-expression in tobacco resulted in the loss of pigmentation in flowers due a decrease in anthocyanin accumulation. Severity in anthocyanin suppression observed in petals could be associated with the expression level of the VvMYB4-like transgene. Expression analysis of flavonoid structural genes revealed the strong down-regulation of the flavonoid-related genes anthocyanidin synthase (ANS) and dihydroflavonol reductase (DFR) genes and also the reduction of the anthocyanin-related gene UDP glucose:flavonoid 3-O-glucosyl transferase (UFGT), which was dependent of the transgene expression. In addition, expression of VvMYB4-like in the model plant Arabidopsis showed similar results, with the higher down-regulation observed in the AtDFR and AtLDOX genes. These results suggest that VvMYB4-like may play an important role in regulation of anthocyanin biosynthesis in grapevine acting as a transcriptional repressor of flavonoid structural genes.

  12. Arabidopsis CAPRICE (MYB) and GLABRA3 (bHLH) Control Tomato (Solanum lycopersicum) Anthocyanin Biosynthesis

    PubMed Central

    Wada, Takuji; Kunihiro, Asuka; Tominaga-Wada, Rumi

    2014-01-01

    In Arabidopsis thaliana the MYB transcription factor CAPRICE (CPC) and the bHLH transcription factor GLABRA3 (GL3) are central regulators of root-hair differentiation and trichome initiation. By transforming the orthologous tomato genes SlTRY (CPC) and SlGL3 (GL3) into Arabidopsis, we demonstrated that these genes influence epidermal cell differentiation in Arabidopsis, suggesting that tomato and Arabidopsis partially use similar transcription factors for epidermal cell differentiation. CPC and GL3 are also known to be involved in anthocyanin biosynthesis. After transformation into tomato, 35S::CPC inhibited anthocyanin accumulation, whereas GL3::GL3 enhanced anthocyanin accumulation. Real-time reverse transcription PCR analyses showed that the expression of anthocyanin biosynthetic genes including Phe-ammonia lyase (PAL), the flavonoid pathway genes chalcone synthase (CHS), dihydroflavonol reductase (DFR), and anthocyanidin synthase (ANS) were repressed in 35S::CPC tomato. In contrast, the expression levels of PAL, CHS, DFR, and ANS were significantly higher in GL3::GL3 tomato compared with control plants. These results suggest that CPC and GL3 also influence anthocyanin pigment synthesis in tomato. PMID:25268379

  13. Transposon Tagging of a Male-Sterility, Female-Sterility Gene, St8, Revealed that the Meiotic MER3 DNA Helicase Activity Is Essential for Fertility in Soybean

    PubMed Central

    Baumbach, Jordan; Pudake, Ramesh N.; Johnson, Callie; Kleinhans, Kaylin; Ollhoff, Alexandrea; Palmer, Reid G.; Bhattacharyya, Madan K.; Sandhu, Devinder

    2016-01-01

    The W4 locus in soybean encodes a dihydroflavonol-4-reductase (DFR2) that regulates pigmentation patterns in flowers and hypocotyl. The mutable w4-m allele that governs variegated flowers has arisen through insertion of a CACTA-type transposable element, Tgm9, in DFR2. In the w4-m line, reversion from variegated to purple flower indicates excision of Tgm9, and its insertion at a new locus. Previously, we have identified a male-sterile, female-sterile mutant among the selfed progenies of a revertant plant carrying only purple flowers. Co-segregation between Tgm9 and the sterility phenotype suggested that the mutant was generated by insertion of Tgm9 at the St8 locus. The transposon was localized to exon 10 of Glyma.16G072300 that shows high identity to the MER3 DNA helicase involved in crossing over. Molecular analysis of fertile branches from two independent revertant plants confirmed precise excision of Tgm9 from the st8 allele, which restored fertility. In soybean, the gene is expressed in flower-buds, trifoliate leaves and stem. Phylogenetic analysis placed St8 in a clade with the Arabidopsis and rice MER3 suggesting that St8 is most likely the orthologous MER3 soybean gene. This study established the utility of Tgm9 in gene identification as well as in forward and reverse genetics studies. PMID:26930200

  14. Fruit coloration and anthocyanin biosynthesis after bag removal in non-red and red apples (Malus × domestica Borkh.).

    PubMed

    Liu, Yulian; Che, Fei; Wang, Lixin; Meng, Rui; Zhang, Xiaojun; Zhao, Zhengyang

    2013-01-25

    In the present study, evolution of apple color (L* and a/b), the accumulation of anthocyanins and the activity of the related enzymes, phenylalanine ammonia-lyase (PAL), chalcone isomerase (CHI), dihydroflavonol4-reductase (DFR) and UDP-Glucose: flavonoid-3-O-galactosyl transferase (UFGT), were investigated in bagged non-red apple cultivars ('Granny Smith' and 'Golden Delicious') and red apple cultivars ('Starkrimon' and 'Pink Lady'). Young fruits were bagged 40-45 days after flowering (DAF), and fruits of 'Golden Delicious' and 'Starkrimon' were uncovered and exposed to light 120 DAF, while those of 'Granny Smith' and 'Pink Lady' were exposed for 160 DAF. Results showed that cyanidin 3-galactoside (cy3-gal) was the most abundant anthocyanin in both non-red and red cultivars. Level of anthocyanins was higher in 'Granny Smith' than in 'Golden Delicious', indicating that red color was easier to develop in green cultivar 'Granny Smith' than in yellow cultivar 'Golden Delicious' after bag removal. The cy3-gal accumulation of non-red cultivars tested was not significantly correlated with PAL, CHI and DFR activity, but was significantly correlated with UFGT activity. During the reddening of non-red apples, UFGT may be the more important factor in the anthocyanin biosynthesis.

  15. Transposon Tagging of a Male-Sterility, Female-Sterility Gene, St8, Revealed that the Meiotic MER3 DNA Helicase Activity Is Essential for Fertility in Soybean.

    PubMed

    Baumbach, Jordan; Pudake, Ramesh N; Johnson, Callie; Kleinhans, Kaylin; Ollhoff, Alexandrea; Palmer, Reid G; Bhattacharyya, Madan K; Sandhu, Devinder

    2016-01-01

    The W4 locus in soybean encodes a dihydroflavonol-4-reductase (DFR2) that regulates pigmentation patterns in flowers and hypocotyl. The mutable w4-m allele that governs variegated flowers has arisen through insertion of a CACTA-type transposable element, Tgm9, in DFR2. In the w4-m line, reversion from variegated to purple flower indicates excision of Tgm9, and its insertion at a new locus. Previously, we have identified a male-sterile, female-sterile mutant among the selfed progenies of a revertant plant carrying only purple flowers. Co-segregation between Tgm9 and the sterility phenotype suggested that the mutant was generated by insertion of Tgm9 at the St8 locus. The transposon was localized to exon 10 of Glyma.16G072300 that shows high identity to the MER3 DNA helicase involved in crossing over. Molecular analysis of fertile branches from two independent revertant plants confirmed precise excision of Tgm9 from the st8 allele, which restored fertility. In soybean, the gene is expressed in flower-buds, trifoliate leaves and stem. Phylogenetic analysis placed St8 in a clade with the Arabidopsis and rice MER3 suggesting that St8 is most likely the orthologous MER3 soybean gene. This study established the utility of Tgm9 in gene identification as well as in forward and reverse genetics studies. PMID:26930200

  16. Flower colour and cytochromes P450.

    PubMed

    Tanaka, Yoshikazu; Brugliera, Filippa

    2013-02-19

    Cytochromes P450 play important roles in biosynthesis of flavonoids and their coloured class of compounds, anthocyanins, both of which are major floral pigments. The number of hydroxyl groups on the B-ring of anthocyanidins (the chromophores and precursors of anthocyanins) impact the anthocyanin colour, the more the bluer. The hydroxylation pattern is determined by two cytochromes P450, flavonoid 3'-hydroxylase (F3'H) and flavonoid 3',5'-hydroxylase (F3'5'H) and thus they play a crucial role in the determination of flower colour. F3'H and F3'5'H mostly belong to CYP75B and CYP75A, respectively, except for the F3'5'Hs in Compositae that were derived from gene duplication of CYP75B and neofunctionalization. Roses and carnations lack blue/violet flower colours owing to the deficiency of F3'5'H and therefore lack the B-ring-trihydroxylated anthocyanins based upon delphinidin. Successful redirection of the anthocyanin biosynthesis pathway to delphinidin was achieved by expressing F3'5'H coding regions resulting in carnations and roses with novel blue hues that have been commercialized. Suppression of F3'5'H and F3'H in delphinidin-producing plants reduced the number of hydroxyl groups on the anthocyanidin B-ring resulting in the production of monohydroxylated anthocyanins based on pelargonidin with a shift in flower colour to orange/red. Pelargonidin biosynthesis is enhanced by additional expression of a dihydroflavonol 4-reductase that can use the monohydroxylated dihydrokaempferol (the pelargonidin precursor). Flavone synthase II (FNSII)-catalysing flavone biosynthesis from flavanones is also a P450 (CYP93B) and contributes to flower colour, because flavones act as co-pigments to anthocyanins and can cause blueing and darkening of colour. However, transgenic plants expression of a FNSII gene yielded paler flowers owing to a reduction of anthocyanins because flavanones are precursors of anthocyanins and flavones.

  17. New member of the R2R3-MYB transcription factors family in grapevine suppresses the anthocyanin accumulation in the flowers of transgenic tobacco.

    PubMed

    Pérez-Díaz, J Ricardo; Pérez-Díaz, Jorge; Madrid-Espinoza, José; González-Villanueva, Enrique; Moreno, Yerko; Ruiz-Lara, Simón

    2016-01-01

    In grapevine, anthocyanins and proanthocyanidins are the main flavonoids in berries, which are associated to organoleptic properties in red wine such as color and astringency. Flavonoid pathway is specifically regulated at transcriptional level and several R2R3-MYB proteins have shown to act as positive regulators. However, some members of this family have shown to repress the flavonoid biosynthesis. In this work, we present the characterization of VvMYB4-like gene, which encodes a putative transcriptional factor highly expressed in the skin of berries at the pre veraison stage in grapevine. Its over-expression in tobacco resulted in the loss of pigmentation in flowers due a decrease in anthocyanin accumulation. Severity in anthocyanin suppression observed in petals could be associated with the expression level of the VvMYB4-like transgene. Expression analysis of flavonoid structural genes revealed the strong down-regulation of the flavonoid-related genes anthocyanidin synthase (ANS) and dihydroflavonol reductase (DFR) genes and also the reduction of the anthocyanin-related gene UDP glucose:flavonoid 3-O-glucosyl transferase (UFGT), which was dependent of the transgene expression. In addition, expression of VvMYB4-like in the model plant Arabidopsis showed similar results, with the higher down-regulation observed in the AtDFR and AtLDOX genes. These results suggest that VvMYB4-like may play an important role in regulation of anthocyanin biosynthesis in grapevine acting as a transcriptional repressor of flavonoid structural genes. PMID:26497001

  18. Identification and Molecular Analysis of Four New Alleles at the W1 Locus Associated with Flower Color in Soybean

    PubMed Central

    Sundaramoorthy, Jagadeesh; Park, Gyu Tae; Chang, Jeong Ho; Lee, Jeong-Dong; Kim, Jeong Hoe; Seo, Hak Soo; Chung, Gyuhwa; Song, Jong Tae

    2016-01-01

    In soybean, flavonoid 3′5′-hydroxylase (F3′5′H) and dihydroflavonol-4-reductase (DFR) play a crucial role in the production of anthocyanin pigments. Loss-of-function of the W1 locus, which encodes the former, or W3 and W4, which encode the latter, always produces white flowers. In this study, we searched for new genetic components responsible for the production of white flowers in soybean and isolated four white-flowered mutant lines, i.e., two Glycine soja accessions (CW12700 and CW13381) and two EMS-induced mutants of Glycine max (PE1837 and PE636). F3′5′H expression in CW12700, PE1837, and PE636 was normal, whereas that in CW13381 was aberrant and missing the third exon. Sequence analysis of F3′5′H of CW13381 revealed the presence of an indel (~90-bp AT-repeat) in the second intron. In addition, the F3′5′H of CW12700, PE1837, and PE636 harbored unique single-nucleotide substitutions. The single nucleotide polymorphisms resulted in substitutions of amino acid residues located in or near the SRS4 domain of F3′5′H, which is essential for substrate recognition. 3D structure modeling of F3′5′H indicated that the substitutions could interfere with an interaction between the substrate and heme group and compromise the conformation of the active site of F3′5′H. Recombination analysis revealed a tight correlation between all of the mutant alleles at the W1 locus and white flower color. On the basis of the characterization of the new mutant alleles, we discussed the biological implications of F3′5′H and DFR in the determination of flower colors in soybean. PMID:27442124

  19. Transcriptome sequencing and metabolite analysis reveals the role of delphinidin metabolism in flower colour in grape hyacinth.

    PubMed

    Lou, Qian; Liu, Yali; Qi, Yinyan; Jiao, Shuzhen; Tian, Feifei; Jiang, Ling; Wang, Yuejin

    2014-07-01

    Grape hyacinth (Muscari) is an important ornamental bulbous plant with an extraordinary blue colour. Muscari armeniacum, whose flowers can be naturally white, provides an opportunity to unravel the complex metabolic networks underlying certain biochemical traits, especially colour. A blue flower cDNA library of M. armeniacum and a white flower library of M. armeniacum f. album were used for transcriptome sequencing. A total of 89 926 uni-transcripts were isolated, 143 of which could be identified as putative homologues of colour-related genes in other species. Based on a comprehensive analysis relating colour compounds to gene expression profiles, the mechanism of colour biosynthesis was studied in M. armeniacum. Furthermore, a new hypothesis explaining the lack of colour phenotype of the grape hyacinth flower is proposed. Alteration of the substrate competition between flavonol synthase (FLS) and dihydroflavonol 4-reductase (DFR) may lead to elimination of blue pigmentation while the multishunt from the limited flux in the cyanidin (Cy) synthesis pathway seems to be the most likely reason for the colour change in the white flowers of M. armeniacum. Moreover, mass sequence data obtained by the deep sequencing of M. armeniacum and its white variant provided a platform for future function and molecular biological research on M. armeniacum.

  20. Growth inhibition in Chinese cabbage (Brassica rapa var. chinensis) growth exposed to di-n-butyl phthalate.

    PubMed

    Liao, Chien-Sen; Yen, Jui-Hung; Wang, Yei-Shung

    2009-04-30

    The toxicity and effects of di-n-butyl phthalate (DBP), an endocrine disruptor, on the growth of Chinese cabbage (Brassica rapa var. chinensis) were studied. Etiolation occurred on leaves of Chinese cabbage plant treated with 50mg/L of DBP for 42 d. DBP even below 1mg/L had a significant effect on the concentration of chlorophyll in Chinese cabbage and the biomass showed a severe decrease under treatment with more than 30 mg/L of DBP. At a concentration below 1mg/L of DBP, no significant difference in accumulation was found, but treatments with concentration exceeding 10, 30, 50 and 100mg/L all resulted in significant accumulation of DBP. Six protein spots extracted from leaf tissue of DBP-treated Chinese cabbage displaying a differential expression are shown in 2-DE maps. According to proteome level studies, three protein spots were found to increase and were identified, respectively, as acyl-[acyl-carrier-protein] desaturase (acyl-ACP desaturase), root phototropism protein 3 (RPT3) and ferredoxin-nitrite reductase (Fd-NiR). The other three protein spots were found to decrease and were identified respectively as dihydroflavonol-4-reductase (DFR), aminoacyl-tRNA synthetase (aaRS) and ATP synthase subunit beta. The key finding is that the other closely related plant, Bok choy (Brassica rapa subsp. chinensis), the subspecies of Chinese cabbage, respond differently to the same chemicals.

  1. Characterization of two tartary buckwheat R2R3-MYB transcription factors and their regulation of proanthocyanidin biosynthesis.

    PubMed

    Bai, Yue-Chen; Li, Cheng-Lei; Zhang, Jin-Wen; Li, Shuang-Jiang; Luo, Xiao-Peng; Yao, Hui-Peng; Chen, Hui; Zhao, Hai-Xia; Park, Sang-Un; Wu, Qi

    2014-11-01

    Tartary buckwheat (Fagopyrum tataricum Gaertn.) contains high concentrations of flavonoids. The flavonoids are mainly represented by rutin, anthocyanins and proanthocyanins in tartary buckwheat. R2R3-type MYB transcription factors (TFs) play key roles in the transcriptional regulation of the flavonoid biosynthetic pathway. In this study, two TF genes, FtMYB1 and FtMYB2, were isolated from F. tataricum and characterized. The results of bioinformatic analysis indicated that the putative FtMYB1 and FtMYB2 proteins belonged to the R2R3-MYB family and displayed a high degree of similarity with TaMYB14 and AtMYB123/TT2. In vitro and in vivo evidence both showed the two proteins were located in the nucleus and exhibited transcriptional activation activities. During florescence, both FtMYB1 and FtMYB2 were more highly expressed in the flowers than any other organ. The overexpression of FtMYB1 and FtMYB2 significantly enhanced the accumulation of proanthocyanidins (PAs) and showed a strong effect on the target genes' expression in Nicotiana tabacum. The expression of dihydroflavonol-4-reductase (DFR) was upregulated to 5.6-fold higher than that of control, and the expression level was lower for flavonol synthase (FLS). To our knowledge, this is the first functional characterization of two MYB TFs from F. tataricum that control the PA pathway. PMID:24730512

  2. Transcriptome sequencing and metabolite analysis reveals the role of delphinidin metabolism in flower colour in grape hyacinth.

    PubMed

    Lou, Qian; Liu, Yali; Qi, Yinyan; Jiao, Shuzhen; Tian, Feifei; Jiang, Ling; Wang, Yuejin

    2014-07-01

    Grape hyacinth (Muscari) is an important ornamental bulbous plant with an extraordinary blue colour. Muscari armeniacum, whose flowers can be naturally white, provides an opportunity to unravel the complex metabolic networks underlying certain biochemical traits, especially colour. A blue flower cDNA library of M. armeniacum and a white flower library of M. armeniacum f. album were used for transcriptome sequencing. A total of 89 926 uni-transcripts were isolated, 143 of which could be identified as putative homologues of colour-related genes in other species. Based on a comprehensive analysis relating colour compounds to gene expression profiles, the mechanism of colour biosynthesis was studied in M. armeniacum. Furthermore, a new hypothesis explaining the lack of colour phenotype of the grape hyacinth flower is proposed. Alteration of the substrate competition between flavonol synthase (FLS) and dihydroflavonol 4-reductase (DFR) may lead to elimination of blue pigmentation while the multishunt from the limited flux in the cyanidin (Cy) synthesis pathway seems to be the most likely reason for the colour change in the white flowers of M. armeniacum. Moreover, mass sequence data obtained by the deep sequencing of M. armeniacum and its white variant provided a platform for future function and molecular biological research on M. armeniacum. PMID:24790110

  3. Growth inhibition in Chinese cabbage (Brassica rapa var. chinensis) growth exposed to di-n-butyl phthalate.

    PubMed

    Liao, Chien-Sen; Yen, Jui-Hung; Wang, Yei-Shung

    2009-04-30

    The toxicity and effects of di-n-butyl phthalate (DBP), an endocrine disruptor, on the growth of Chinese cabbage (Brassica rapa var. chinensis) were studied. Etiolation occurred on leaves of Chinese cabbage plant treated with 50mg/L of DBP for 42 d. DBP even below 1mg/L had a significant effect on the concentration of chlorophyll in Chinese cabbage and the biomass showed a severe decrease under treatment with more than 30 mg/L of DBP. At a concentration below 1mg/L of DBP, no significant difference in accumulation was found, but treatments with concentration exceeding 10, 30, 50 and 100mg/L all resulted in significant accumulation of DBP. Six protein spots extracted from leaf tissue of DBP-treated Chinese cabbage displaying a differential expression are shown in 2-DE maps. According to proteome level studies, three protein spots were found to increase and were identified, respectively, as acyl-[acyl-carrier-protein] desaturase (acyl-ACP desaturase), root phototropism protein 3 (RPT3) and ferredoxin-nitrite reductase (Fd-NiR). The other three protein spots were found to decrease and were identified respectively as dihydroflavonol-4-reductase (DFR), aminoacyl-tRNA synthetase (aaRS) and ATP synthase subunit beta. The key finding is that the other closely related plant, Bok choy (Brassica rapa subsp. chinensis), the subspecies of Chinese cabbage, respond differently to the same chemicals. PMID:18678443

  4. Flower colour and cytochromes P450†

    PubMed Central

    Tanaka, Yoshikazu; Brugliera, Filippa

    2013-01-01

    Cytochromes P450 play important roles in biosynthesis of flavonoids and their coloured class of compounds, anthocyanins, both of which are major floral pigments. The number of hydroxyl groups on the B-ring of anthocyanidins (the chromophores and precursors of anthocyanins) impact the anthocyanin colour, the more the bluer. The hydroxylation pattern is determined by two cytochromes P450, flavonoid 3′-hydroxylase (F3′H) and flavonoid 3′,5′-hydroxylase (F3′5′H) and thus they play a crucial role in the determination of flower colour. F3′H and F3′5′H mostly belong to CYP75B and CYP75A, respectively, except for the F3′5′Hs in Compositae that were derived from gene duplication of CYP75B and neofunctionalization. Roses and carnations lack blue/violet flower colours owing to the deficiency of F3′5′H and therefore lack the B-ring-trihydroxylated anthocyanins based upon delphinidin. Successful redirection of the anthocyanin biosynthesis pathway to delphinidin was achieved by expressing F3′5′H coding regions resulting in carnations and roses with novel blue hues that have been commercialized. Suppression of F3′5′H and F3′H in delphinidin-producing plants reduced the number of hydroxyl groups on the anthocyanidin B-ring resulting in the production of monohydroxylated anthocyanins based on pelargonidin with a shift in flower colour to orange/red. Pelargonidin biosynthesis is enhanced by additional expression of a dihydroflavonol 4-reductase that can use the monohydroxylated dihydrokaempferol (the pelargonidin precursor). Flavone synthase II (FNSII)-catalysing flavone biosynthesis from flavanones is also a P450 (CYP93B) and contributes to flower colour, because flavones act as co-pigments to anthocyanins and can cause blueing and darkening of colour. However, transgenic plants expression of a FNSII gene yielded paler flowers owing to a reduction of anthocyanins because flavanones are precursors of anthocyanins and flavones. PMID:23297355

  5. De novo transcriptome of Brassica juncea seed coat and identification of genes for the biosynthesis of flavonoids.

    PubMed

    Liu, Xianjun; Lu, Ying; Yuan, Yuhui; Liu, Shuyan; Guan, Chunyun; Chen, Sheyuan; Liu, Zhongsong

    2013-01-01

    Brassica juncea, a worldwide cultivated crop plant, produces seeds of different colors. Seed pigmentation is due to the deposition in endothelial cells of proanthocyanidins (PAs), end products from a branch of flavonoid biosynthetic pathway. To elucidate the gene regulatory network of seed pigmentation in B. juncea, transcriptomes in seed coat of a yellow-seeded inbred line and its brown-seeded near- isogenic line were sequenced using the next-generation sequencing platform Illumina/Solexa and de novo assembled. Over 116 million high-quality reads were assembled into 69,605 unigenes, of which about 71.5% (49,758 unigenes) were aligned to Nr protein database with a cut-off E-value of 10(-5). RPKM analysis showed that the brown-seeded testa up-regulated 802 unigenes and down-regulated 502 unigenes as compared to the yellow-seeded one. Biological pathway analysis revealed the involvement of forty six unigenes in flavonoid biosynthesis. The unigenes encoding dihydroflavonol reductase (DFR), leucoantho-cyanidin dioxygenase (LDOX) and anthocyanidin reductase (ANR) for late flavonoid biosynthesis were not expressed at all or at a very low level in the yellow-seeded testa, which implied that these genes for PAs biosynthesis be associated with seed color of B. juncea, as confirmed by qRT-PCR analysis of these genes. To our knowledge, it is the first time to sequence the transcriptome of seed coat in Brassica juncea. The unigene sequences obtained in this study will not only lay the foundations for insight into the molecular mechanisms underlying seed pigmentation in B.juncea, but also provide the basis for further genomics research on this species or its allies. PMID:23990927

  6. Three R2R3-MYB Transcription Factors Regulate Distinct Floral Pigmentation Patterning in Phalaenopsis spp.1[OPEN

    PubMed Central

    Hsu, Chia-Chi; Chen, You-Yi; Tsai, Wen-Chieh; Chen, Wen-Huei; Chen, Hong-Hwa

    2015-01-01

    Orchidaceae are well known for their fascinating floral morphologic features, specialized pollination, and distinctive ecological strategies. With their long-lasting flowers of various colors and pigmentation patterning, Phalaenopsis spp. have become important ornamental plants worldwide. In this study, we identified three R2R3-MYB transcription factors PeMYB2, PeMYB11, and PeMYB12. Their expression profiles were concomitant with red color formation in Phalaenopsis spp. flowers. Transient assay of overexpression of three PeMYBs verified that PeMYB2 resulted in anthocyanin accumulation, and these PeMYBs could activate the expression of three downstream structural genes Phalaenopsis spp. Flavanone 3-hydroxylase5, Phalaenopsis spp. Dihydroflavonol 4-reductase1, and Phalaenopsis spp. Anthocyanidin synthase3. In addition, these three PeMYBs participated in the distinct pigmentation patterning in a single flower, which was revealed by virus-induced gene silencing. In the sepals/petals, silencing of PeMYB2, PeMYB11, and PeMYB12 resulted in the loss of the full-red pigmentation, red spots, and venation patterns, respectively. Moreover, different pigmentation patterning was regulated by PeMYBs in the sepals/petals and lip. PeMYB11 was responsive to the red spots in the callus of the lip, and PeMYB12 participated in the full pigmentation in the central lobe of the lip. The differential pigmentation patterning was validated by RNA in situ hybridization. Additional assessment was performed in six Phalaenopsis spp. cultivars with different color patterns. The combined expression of these three PeMYBs in different ratios leads to a wealth of complicated floral pigmentation patterning in Phalaenopsis spp. PMID:25739699

  7. AtROS1 overexpression provides evidence for epigenetic regulation of genes encoding enzymes of flavonoid biosynthesis and antioxidant pathways during salt stress in transgenic tobacco

    PubMed Central

    Bharti, Poonam; Mahajan, Monika; Vishwakarma, Ajay K.; Bhardwaj, Jyoti; Yadav, Sudesh Kumar

    2015-01-01

    In plants, epigenetic changes have been identified as regulators of developmental events during normal growth as well as environmental stress exposures. Flavonoid biosynthetic and antioxidant pathways play a significant role in plant defence during their exposure to environmental cues. The aim of this study was to unravel whether genes encoding enzymes of flavonoid biosynthetic and antioxidant pathways are under epigenetic regulation, particularly DNA methylation, during salt stress. For this, a repressor of silencing from Arabidopsis, AtROS1, was overexpressed in transgenic tobacco. Generated transgenics were evaluated to examine the influence of AtROS1 on methylation status of promoters as well as on coding regions of genes encoding enzymes of flavonoids biosynthesis and antioxidant pathways. Overexpression of AtROS1 increases the demethylation levels of both promoters as well as coding regions of genes encoding chalcone synthase, chalcone isomerase, flavanone 3-hydroxylase, flavonol synthase, dihydroflavonol 4-reductase, and anthocyanidin synthase of the flavonoid biosynthetic pathway, and glutathione S-transferase, ascorbate peroxidase, glutathione peroxidase, and glutathione reductase of the antioxidant pathway during control conditions. The level of demethylation was further increased at promoters as well as coding regions of these genes during salt-stress conditions. Transgenic tobacco overexpressing AtROS1 showed tolerance to salt stress that could have been due to the higher expression levels of the genes encoding enzymes of the flavonoid biosynthetic and antioxidant pathways. This is the first comprehensive study documenting the epigenetic regulation of flavonoid biosynthetic and antioxidant pathways during salt-stress exposure of plants. PMID:26116024

  8. Expression of Genes Involved in Anthocyanin Biosynthesis in Relation to Anthocyanin, Proanthocyanidin, and Flavonol Levels during Bilberry Fruit Development1

    PubMed Central

    Jaakola, Laura; Määttä, Kaisu; Pirttilä, Anna Maria; Törrönen, Riitta; Kärenlampi, Sirpa; Hohtola, Anja

    2002-01-01

    The production of anthocyanins in fruit tissues is highly controlled at the developmental level. We have studied the expression of flavonoid biosynthesis genes during the development of bilberry (Vaccinium myrtillus) fruit in relation to the accumulation of anthocyanins, proanthocyanidins, and flavonols in wild berries and in color mutants of bilberry. The cDNA fragments of five genes from the flavonoid pathway, phenylalanine ammonia-lyase, chalcone synthase, flavanone 3-hydroxylase, dihydroflavonol 4-reductase, and anthocyanidin synthase, were isolated from bilberry using the polymerase chain reaction technique, sequenced, and labeled with a digoxigenin-dUTP label. These homologous probes were used for determining the expression of the flavonoid pathway genes in bilberries. The contents of anthocyanins, proanthocyanidins, and flavonols in ripening bilberries were analyzed with high-performance liquid chromatography-diode array detector and were identified using a mass spectrometry interface. Our results demonstrate a correlation between anthocyanin accumulation and expression of the flavonoid pathway genes during the ripening of berries. At the early stages of berry development, procyanidins and quercetin were the major flavonoids, but the levels decreased dramatically during the progress of ripening. During the later stages of ripening, the content of anthocyanins increased strongly and they were the major flavonoids in the ripe berry. The expression of flavonoid pathway genes in the color mutants of bilberry was reduced. A connection between flavonol and anthocyanin synthesis in bilberry was detected in this study and also in previous data collected from flavonol and anthocyanin analyses from other fruits. In accordance with this, models for the connection between flavonol and anthocyanin syntheses in fruit tissues are presented. PMID:12376640

  9. Three R2R3-MYB transcription factors regulate distinct floral pigmentation patterning in Phalaenopsis spp.

    PubMed

    Hsu, Chia-Chi; Chen, You-Yi; Tsai, Wen-Chieh; Chen, Wen-Huei; Chen, Hong-Hwa

    2015-05-01

    Orchidaceae are well known for their fascinating floral morphologic features, specialized pollination, and distinctive ecological strategies. With their long-lasting flowers of various colors and pigmentation patterning, Phalaenopsis spp. have become important ornamental plants worldwide. In this study, we identified three R2R3-MYB transcription factors PeMYB2, PeMYB11, and PeMYB12. Their expression profiles were concomitant with red color formation in Phalaenopsis spp. flowers. Transient assay of overexpression of three PeMYBs verified that PeMYB2 resulted in anthocyanin accumulation, and these PeMYBs could activate the expression of three downstream structural genes Phalaenopsis spp. Flavanone 3-hydroxylase5, Phalaenopsis spp. Dihydroflavonol 4-reductase1, and Phalaenopsis spp. Anthocyanidin synthase3. In addition, these three PeMYBs participated in the distinct pigmentation patterning in a single flower, which was revealed by virus-induced gene silencing. In the sepals/petals, silencing of PeMYB2, PeMYB11, and PeMYB12 resulted in the loss of the full-red pigmentation, red spots, and venation patterns, respectively. Moreover, different pigmentation patterning was regulated by PeMYBs in the sepals/petals and lip. PeMYB11 was responsive to the red spots in the callus of the lip, and PeMYB12 participated in the full pigmentation in the central lobe of the lip. The differential pigmentation patterning was validated by RNA in situ hybridization. Additional assessment was performed in six Phalaenopsis spp. cultivars with different color patterns. The combined expression of these three PeMYBs in different ratios leads to a wealth of complicated floral pigmentation patterning in Phalaenopsis spp.

  10. Jasmonic acid enhancement of anthocyanin accumulation is dependent on phytochrome A signaling pathway under far-red light in Arabidopsis.

    PubMed

    Li, Ting; Jia, Kun-Peng; Lian, Hong-Li; Yang, Xu; Li, Ling; Yang, Hong-Quan

    2014-11-01

    Anthocyanins are critical for plants. It is shown that the expression of genes encoding the key enzymes such as dihydroflavonol 4-reductase (DFR), UDP-Glc: flavonoid 3-O-glucosyltransferase (UF3GT), and leucoanthocyanidin dioxygenase (LDOX) in anthocyanin biosynthesis pathway is regulated by MYB75, a R2R3 MYB transcription factor. The production of anthocyanin is known to be promoted by jasmonic acid (JA) in light but not in darkness. The photoreceptors cryptochrome 1 (CRY1), phytochrome B (phyB), and phytochrome A (phyA) are also shown to mediate light promotion of anthocyanin accumulation, respectively, whereas their downstream factor COP1, a master negative regulator of photomorphogensis, represses anthocyanin accumulation. However, whether JA coordinates with photoreceptors in the regulation of anthocyanin accumulation is unknown. Here, we show that under far-red light, JA promotes anthocyanin accumulation in a phyA signaling pathway-dependent manner. The phyA mutant is hyposensitive to jasmonic acid analog methyl jasmonic acid (MeJA) under far-red light. The dominant mutant of MYB75, pap1-D, accumulates significantly higher levels of anthocyanin than wild type under far-red light, whereas knockdown of MYBs (MYB75, MYB90, MYB113, and MYB114) through RNAi significantly reduces MeJA promotion of anthocyanin accumulation. The phyA pap1-D double mutant shows reduced responsiveness to MeJA, similar to phyA mutant under far-red light. In darkness, a mutant allele of cop1, cop1-4, shows enhanced responsiveness to MeJA, but pap1-D mutant is barely responsive to MeJA. Upon MeJA application, the cop1-4 pap1-D double mutant accumulates considerably higher levels of anthocyanin than cop1-4 in darkness. Protein studies indicate that MYB75 protein is stabilized by white light and far-red light. Further gene expression studies suggest that MeJA promotes the expression of DFR, UF3GT, and LDOX genes in a phyA- and MYB75-dependent manner under far-red light. Our findings suggest

  11. Nitrogen supply affects anthocyanin biosynthetic and regulatory genes in grapevine cv. Cabernet-Sauvignon berries.

    PubMed

    Soubeyrand, Eric; Basteau, Cyril; Hilbert, Ghislaine; van Leeuwen, Cornelis; Delrot, Serge; Gomès, Eric

    2014-07-01

    Accumulation of anthocyanins in grape berries is influenced by environmental factors (such as temperature and light) and supply of nutrients, i.e., fluxes of carbon and nitrogen feeding the berry cells. It is established that low nitrogen supply stimulates anthocyanin production in berry skin cells of red varieties. The present works aims to gain a better understanding of the molecular mechanisms involved in the response of anthocyanin accumulation to nitrogen supply in berries from field grown-plants. To this end, we developed an integrated approach combining monitoring of plant nitrogen status, metabolite measurements and transcript analysis. Grapevines (cv. Cabernet-Sauvignon) were cultivated in a vineyard with three nitrogen fertilization levels (0, 60 and 120 kg ha(-1) of nitrogen applied on the soil). Anthocyanin profiles were analyzed and compared with gene expression levels. Low nitrogen supply caused a significant increase in anthocyanin levels at two ripening stages (26 days post-véraison and maturity). Delphinidin and petunidin derivatives were the most affected compounds. Transcript levels of both structural and regulatory genes involved in anthocyanin synthesis confirmed the stimulation of the phenylpropanoid pathway. Genes encoding phenylalanine ammonia-lyase (PAL), chalcone synthase (CHS), flavonoid-3',5'-hydroxylase (F3'5'H), dihydroflavonol-4-reductase (DFR), leucoanthocyanidin dioxygenase (LDOX) exhibited higher transcript levels in berries from plant cultivated without nitrogen compared to the ones cultivated with 120 kg ha(-1) nitrogen fertilization. The results indicate that nitrogen controls a coordinated regulation of both positive (MYB transcription factors) and negative (LBD proteins) regulators of the flavonoid pathway in grapevine. PMID:24735825

  12. Complexity and robustness of the flavonoid transcriptional regulatory network revealed by comprehensive analyses of MYB-bHLH-WDR complexes and their targets in Arabidopsis seed.

    PubMed

    Xu, Wenjia; Grain, Damaris; Bobet, Sophie; Le Gourrierec, José; Thévenin, Johanne; Kelemen, Zsolt; Lepiniec, Loïc; Dubos, Christian

    2014-04-01

    In Arabidopsis thaliana, proanthocyanidins (PAs) accumulate in the innermost cell layer of the seed coat (i.e. endothelium, chalaza and micropyle). The expression of the biosynthetic genes involved relies on the transcriptional activity of R2R3-MYB and basic helix-loop-helix (bHLH) proteins which form ternary complexes ('MBW') with TRANSPARENT TESTA GLABRA1 (TTG1) (WD repeat protein). The identification of the direct targets and the determination of the nature and spatio-temporal activity of these MBW complexes are essential steps towards a comprehensive understanding of the transcriptional mechanisms that control flavonoid biosynthesis. In this study, various molecular, genetic and biochemical approaches were used. Here, we have demonstrated that, of the 12 studied genes of the pathway, only dihydroflavonol-4-reductase (DFR), leucoanthocyanidin dioxygenase (LDOX), BANYULS (BAN), TRANSPARENT TESTA 19 (TT19), TT12 and H(+) -ATPase isoform 10 (AHA10) are direct targets of the MBW complexes. Interestingly, although the TT2-TT8-TTG1 complex plays the major role in developing seeds, three additional MBW complexes (i.e. MYB5-TT8-TTG1, TT2-EGL3-TTG1 and TT2-GL3-TTG1) were also shown to be involved, in a tissue-specific manner. Finally, a minimal promoter was identified for each of the target genes of the MBW complexes. Altogether, by answering fundamental questions and by demonstrating or invalidating previously made hypotheses, this study provides a new and comprehensive view of the transcriptional regulatory mechanisms controlling PA and anthocyanin biosynthesis in Arabidopsis. PMID:24299194

  13. Relating genes in the biosynthesis of the polyphenol composition of Andean colored potato collection

    PubMed Central

    Tejeda, Leslie; Alvarado, Juan Antonio; Dębiec, Magdalena; Peñarrieta, José Mauricio; Cárdenas, Oscar; Alvarez, Maria Teresa; Chawade, Aakash; Nilsson, Lars; Bergenståhl, Björn

    2014-01-01

    The objective of this study was to evaluate total antioxidant capacity (TAC), total phenolic content (TPH), and the identification of anthocyanidin and polyphenolic compounds in 13 colored potatoes collected from the Andean region of Bolivia, and understand how the chemical composition correlated with the botanical classification and molecular characterization of genes, ans (anthocyanidin synthase) and stan1 (Solanum tuberosum anthocyanidin synthase), associated with the synthesis of anthocyanidins. The results show the existence of a limited correlation between botanical classification, based on morphological identification and polyphenol composition. No association between genetic grouping of the ans and stan genes and botanical classification was found. However, it was possible to identify a correlation between the ans gene clades and the levels of anthocyanidins as well as of other polyphenols. Thus, this result confirms the concept that potato color can be used in the search for high polyphenol potato cultivars. PMID:24804064

  14. Factors that cause trimethoprim resistance in Streptococcus pyogenes.

    PubMed

    Bergmann, René; van der Linden, Mark; Chhatwal, Gursharan S; Nitsche-Schmitz, D Patric

    2014-01-01

    The use of trimethoprim in treatment of Streptococcus pyogenes infections has long been discouraged because it has been widely believed that this pathogen is resistant to this antibiotic. To gain more insight into the extent and molecular basis of trimethoprim resistance in S. pyogenes, we tested isolates from India and Germany and sought the factors that conferred the resistance. Resistant isolates were identified in tests for trimethoprim or trimethoprim-sulfamethoxazole (SXT) susceptibility. Resistant isolates were screened for the known horizontally transferable trimethoprim-insensitive dihydrofolate reductase (dfr) genes dfrG, dfrF, dfrA, dfrD, and dfrK. The nucleotide sequence of the intrinsic dfr gene was determined for resistant isolates lacking the horizontally transferable genes. Based on tentative criteria, 69 out of 268 isolates (25.7%) from India were resistant to trimethoprim. Occurring in 42 of the 69 resistant isolates (60.9%), dfrF appeared more frequently than dfrG (23 isolates; 33.3%) in India. The dfrF gene was also present in a collection of SXT-resistant isolates from Germany, in which it was the only detected trimethoprim resistance factor. The dfrF gene caused resistance in 4 out of 5 trimethoprim-resistant isolates from the German collection. An amino acid substitution in the intrinsic dihydrofolate reductase known from trimethoprim-resistant Streptococcus pneumoniae conferred resistance to S. pyogenes isolates of emm type 102.2, which lacked other aforementioned dfr genes. Trimethoprim may be more useful in treatment of S. pyogenes infections than previously thought. However, the factors described herein may lead to the rapid development and spread of resistance of S. pyogenes to this antibiotic agent.

  15. Molecular Basis of Sulfonamide and Trimethoprim Resistance in Fish-Pathogenic Aeromonas Isolates ▿

    PubMed Central

    Kadlec, Kristina; von Czapiewski, Ellen; Kaspar, Heike; Wallmann, Jürgen; Michael, Geovana Brenner; Steinacker, Ulrike; Schwarz, Stefan

    2011-01-01

    Sulfonamide-trimethoprim-resistant Aeromonas salmonicida and motile Aeromonas spp. from diseased fish of the GERM-Vet study carried the sul1 gene together with mostly cassette-borne trimethoprim resistance genes, including the novel gene dfrA28. The seven dfrA and dfrB genes identified were located mostly in class 1 integrons which commonly harbored other gene cassettes. PMID:21764945

  16. Factors that cause trimethoprim resistance in Streptococcus pyogenes.

    PubMed

    Bergmann, René; van der Linden, Mark; Chhatwal, Gursharan S; Nitsche-Schmitz, D Patric

    2014-01-01

    The use of trimethoprim in treatment of Streptococcus pyogenes infections has long been discouraged because it has been widely believed that this pathogen is resistant to this antibiotic. To gain more insight into the extent and molecular basis of trimethoprim resistance in S. pyogenes, we tested isolates from India and Germany and sought the factors that conferred the resistance. Resistant isolates were identified in tests for trimethoprim or trimethoprim-sulfamethoxazole (SXT) susceptibility. Resistant isolates were screened for the known horizontally transferable trimethoprim-insensitive dihydrofolate reductase (dfr) genes dfrG, dfrF, dfrA, dfrD, and dfrK. The nucleotide sequence of the intrinsic dfr gene was determined for resistant isolates lacking the horizontally transferable genes. Based on tentative criteria, 69 out of 268 isolates (25.7%) from India were resistant to trimethoprim. Occurring in 42 of the 69 resistant isolates (60.9%), dfrF appeared more frequently than dfrG (23 isolates; 33.3%) in India. The dfrF gene was also present in a collection of SXT-resistant isolates from Germany, in which it was the only detected trimethoprim resistance factor. The dfrF gene caused resistance in 4 out of 5 trimethoprim-resistant isolates from the German collection. An amino acid substitution in the intrinsic dihydrofolate reductase known from trimethoprim-resistant Streptococcus pneumoniae conferred resistance to S. pyogenes isolates of emm type 102.2, which lacked other aforementioned dfr genes. Trimethoprim may be more useful in treatment of S. pyogenes infections than previously thought. However, the factors described herein may lead to the rapid development and spread of resistance of S. pyogenes to this antibiotic agent. PMID:24492367

  17. A New Surgical Procedure “Dumbbell-Form Resection” for Selected Hilar Cholangiocarcinomas With Severe Jaundice

    PubMed Central

    Wang, Shuguang; Tian, Feng; Zhao, Xin; Li, Dajiang; He, Yu; Li, Zhihua; Chen, Jian

    2016-01-01

    Abstract The aim of the study is to evaluate the therapeutic effect of a new surgical procedure, dumbbell-form resection (DFR), for hilar cholangiocarcinoma (HCCA) with severe jaundice. In DFR, liver segments I, IVb, and partial V above the right hepatic pedicle are resected. Hemihepatectomy is recognized as the preferred procedure; however, its application is limited in HCCAs with severe jaundice. Thirty-eight HCCA patients with severe jaundice receiving DFR and 70 receiving hemihepatectomy from January 2008 to January 2013 were included. Perioperative parameters, operation-related morbidity and mortality, and post-operative survival were analyzed. A total of 21.1% patients (8/38) in the DFR group received percutaneous transhepatic biliary drainage (PTBD), which was significantly <81.4% (57/70) in the hemihepatectomy group. The TBIL was higher in the DFR group at operation (243.7 vs 125.6 μmol/L, respectively). The remnant liver volume was significantly higher after DFR. The operation-related morbidity was significantly lower after DFR than after hemihepatectomy (26.3% vs 48.6%, respectively). None of the patients died during the perioperative period after DFR, whereas 3 died after hemihepatectomy. There was no difference in margin status, histological grade, lymph-node involvement, and distant metastasis between the 2 groups. The 1-, 3-, and 5-year survival rates after DFR (68.4%, 32.1%, and 21.4%, respectively) showed no significant difference with those after hemihepatectomy (62.7%, 34.6%, and 23.3%, respectively). Kaplan–Meier analysis indicated that overall survival and recurrence after DFR demonstrated no significant difference compared with hemihepatectomy. DFR appears to be feasible for selected HCCA patients with severe jaundice. However, its indications should be restricted. PMID:26765439

  18. 78 FR 53285 - Seagoing Barges

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-08-29

    .... Abbreviations CFR Code of Federal Regulations DFR Direct final rule E.O. Executive Order FR Federal Register... rule (DFR) entitled ``Seagoing Barges'' (76 FR 77712). Following the receipt of an adverse comment on... (77 FR 20727). On January 9, 2013, we published a notice of proposed rulemaking (NPRM)...

  19. Spontaneous symmetry breaking and masses numerical results in Doplicher-Fredenhagen-Roberts noncommutative space-time

    NASA Astrophysics Data System (ADS)

    Neves, M. J.; Abreu, Everton M. C.

    2016-04-01

    With the elements of the Doplicher-Fredenhagen-Roberts (DFR) noncommutative formalism, we have constructed a standard electroweak model. We have introduced the spontaneous symmetry breaking and the hypercharge in DFR framework. The electroweak symmetry breaking was analyzed and the masses of the new bosons were computed.

  20. 78 FR 9828 - Establishment of User Fees for Filovirus Testing of Nonhuman Primate Liver Samples

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-02-12

    ... Federal Register (77 FR 7109) that provided the background, rationale, description of the services and... instructions. On the same date, we published a companion Direct Final Rule (DFR) (77 FR 6981). In both the NPRM... amendment in the Federal Register on June 15, 2012 (77 FR 35878), withdrawing the DFR. II. Public...

  1. 77 FR 35878 - Establishment of User Fees for Filovirus Testing of Nonhuman Primate Liver Samples

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-06-15

    ... (NPRM) (77 FR 7109) that proposed identical filovirus testing and user fee requirements. In both the DFR... HHS/CDC published a Direct Final Rule (DFR) (77 FR 6971) amending 42 CFR 71.53 by adding a new... February 10, 2012, HHS/CDC also published a companion Notice of Proposed Rulemaking (NPRM) (77 FR...

  2. 75 FR 27188 - Revising the Notification Requirements in the Exposure Determination Provisions of the Hexavalent...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-05-14

    ... March 17, 2010, OSHA published a DFR in the Federal Register (75 FR 12681) amending the employee... direct final rulemaking. OSHA noted as much in the DFR notice. (See 75 FR at 12683 (`` he number of work... FR at 12683 (``OSHA will not consider a comment recommending an additional amendment to be...

  3. Effect of hydrothermal processing on antioxidant contents and capacities in pigmented rice (Oryza sativa L.)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Purple and red bran rice cultivars (Oryza sativa L.) are rich sources of antioxidants including lipophilic antioxidants (vitamin E homologues and '-oryzanol), soluble phenolics (including anthocyanidins and proanthocyanidins), and cell-wall-bound phenolics. This study investigated impacts of hydroth...

  4. Endogenous post-transcriptional gene silencing of flavone synthase resulting in high accumulation of anthocyanins in black dahlia cultivars.

    PubMed

    Deguchi, Ayumi; Ohno, Sho; Hosokawa, Munetaka; Tatsuzawa, Fumi; Doi, Motoaki

    2013-05-01

    Black color in flowers is a highly attractive trait in the floricultural industry, but its underlying mechanisms are largely unknown. This study was performed to identify the bases of the high accumulation of anthocyanidins in black cultivars and to determine whether the high accumulation of total anthocyanidins alone leads to the black appearance. Our approach was to compare black dahlia (Dahlia variabilis) cultivars with purple cultivars and a purple flowering mutant of a black cultivar, using pigment and molecular analyses. Black cultivars characteristically exhibited low lightness, high petal accumulation of cyanidin and total anthocyanidins without flavones, and marked suppression of flavone synthase (DvFNS) expression. A comparative study using black and purple cultivars revealed that neither the absence of flavones nor high accumulation of total anthocyanidins is solely sufficient for black appearance, but that cyanidin content in petals is also an important factor in the phenotype. A study comparing the black cultivar 'Kokucho' and its purple mutant showed that suppression of DvFNS abolishes the competition between anthocyanidin and flavone synthesis and leads to accumulation of cyanidin and total anthocyanidins that produce a black appearance. Surprisingly, in black cultivars the suppression of DvFNS occurred in a post-transcriptional manner, as determined by small RNA mapping. PMID:23389674

  5. Molecular analysis of herbivore-induced condensed tannin synthesis: cloning and expression of dihydroflavonol reductase from trembling aspen (Populus tremuloides).

    PubMed

    Peters, Darren J; Constabel, C Peter

    2002-12-01

    In order to study condensed tannin synthesis and its induction by herbivory, a dihydroflavonol reductase (DFR) cDNA was isolated from trembling aspen (Populus tremuloides). Bacterial overexpression demonstrated that this cDNA encodes a functional DFR enzyme, and Southern analysis revealed that DFR likely is a single-copy gene in the aspen genome. Aspen plants that were mechanically wounded showed a dramatic increase in DFR expression after 24 h in both wounded leaves and unwounded leaves on wounded trees. Feeding by forest tent caterpillar (Malacosoma disstria) and satin moth (Leucoma salicis) larvae, and treatment with methyl jasmonate, all strongly induced DFR expression. DFR enzyme activity was also induced in wounded aspen leaves, and phytochemical assays revealed that condensed tannin concentrations significantly increased in wounded and systemic leaves. The expression of other genes involved in the phenylpropanoid pathway were also induced by wounding. Our findings suggest that the induction of condensed tannins, compounds known to be important for defense against herbivores, is mediated by increased expression of DFR and other phenylpropanoid genes.

  6. Antimicrobial resistance in commensal Escherichia coli from pigs during metaphylactic trimethoprim and sulfamethoxazole treatment and in the post-exposure period.

    PubMed

    Mazurek, Justyna; Bok, Ewa; Stosik, Michał; Baldy-Chudzik, Katarzyna

    2015-02-01

    The prevalence of trimethoprim (TMP) and sulfamethoxazole (SMX) resistance in commensal E. coli from pigs was tested in this study. E. coli was derived from three groups of piglets in successive stages of metaphylactic therapy and from two groups of sows 10 and 18 weeks after the treatment. MIC values of TMP and SMX were determined for a total of 352 strains. The presence of resistance genes (dfrA1, dfrA5, dfrA7, dfrA12, dfrA17, sul1, sul2, sul3) and class 1 and 2 integron-associated dfrA gene cassettes was tested. Resistance to TMP was very high during the administration of the antimicrobial (from 97 to 100%) and amounted to 86% and 69% in the post-exposure period; MIC > 32 mg/L. The isolates from all groups of pigs were resistant to sulfamethoxazole, with MIC > 1028 mg/L. The dfrA1 and sul1 genes (as part of integrons) dominated in E. coli from piglets, but the dfrA12 and sul1 genes were prevalent in E. coli from sows. Coexistence of the different dfrA genes was detected in 71 isolates from all groups of swine. Transcription analysis revealed that most of these genes were not transcribed, particularly gene cassettes of class 1 integrons. The research revealed a high level of resistance associated with the metaphylactic treatment, persistence and circulation of resistance in bacterial populations. Diverse genetic background with multiple and not transcribed resistance genes was observed. PMID:25689993

  7. 78 FR 47319 - Fee Schedule for Reference Biological Standards and Biological Preparations

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-08-05

    ... Rule (DFR) titled ``Distribution of Reference Biological Standards and Biological Preparations (78 FR... review should include any adjustment to reflect changes in costs or market value. HHS/CDC has conducted...

  8. Determination of Flavonoids and Anthocyanins in Nitraria tangutorum by High Performance Liquid Chromatography Coupled with Tandem Mass Spectrometry.

    PubMed

    Zhe, Gao; Ying-Chun, Wang; Yan-Xu, Chang

    2016-01-01

    Using high-performance liquid chromatography coupled with diode array detection and electrospray ionization tandem mass spectrometry (HPLC-DAD-MSn) method, qualitative and quantitative analysis of flavonoids of stems, leaves, fruits and seeds, and anthocyanidin of fresh fruits in Nitraria tangutorum were performed. A total of 14 flavonoid components were identified from the seeds of N. tangutorum including three quercetin derivatives, three kaempferol derivatives, and eight isorhamnetin derivatives. A total of 12, 10, and 7 flavonoid components were identified from leaves, stems, and fruits of N. tangutorum, respectively; all were present in seeds also. The total content of flavonoids in leaves was the highest, up to 42.43 mg/g·dry weight. A total of 12 anthocyanidin components were identified from the fresh fruits of N. tangutorum, belonging to five anthocyanidin. The total content of anthocyanidin in fresh fruits was up to 45.83 mg/100 g· fresh weight, of which the acylated anthocyanidin accounted for 65.7%. The HPLC-DAD-MS(n) method can be operated easily, rapidly, and accurately, and is feasible for qualitative and quantitative analysis of flavone glycosides in N. tangutorum. PMID:26972973

  9. Comment: importance of cognitive reserve in traumatic brain injury.

    PubMed

    Bigler, Erin D

    2014-05-01

    The expectation for moderate to severe traumatic brain injury (TBI) is permanent damage and lasting deficits. However, in a multicenter investigation, Schneider et al.(1) show that by 1 year postinjury, one-fourth of patients with TBI achieve disability-free recovery (DFR), defined as a score of zero on the Disability Rating Scale. Of importance, cognitive reserve (CR) in the form of educational attainment was related to DFR.

  10. Transcriptomic analysis of purple leaf determination in birch.

    PubMed

    Lin, Lin; Mu, Huaizhi; Jiang, Jing; Liu, Guifeng

    2013-09-10

    'Purple Rain', a purple cultivar of Betula pendula, has dark purple leaves throughout the vegetative period. In this study, B. pendula 'Purple Rain' was found to have a higher anthocyanidin level compared with B. pendula, Transcriptome analysis revealed numerous changes in gene expression that could be attributed to color change, including the upregulation of 2467 unigenes and the downregulation of 2299 unigenes in 'Purple Rain'. Furthermore, anthocyanidin synthesis and transcriptional regulation were altered in 'Purple Rain', which may have contributed to phenotypic changes. These results provide unique molecular insights into the biochemical pathways and regulatory networks that function in a purple variety of B. pendula.

  11. Genetic redundancy and persistence of plasmid-mediated trimethoprim/sulfamethoxazole resistant effluent and stream water Escherichia coli.

    PubMed

    Suhartono, Suhartono; Savin, Mary; Gbur, Edward E

    2016-10-15

    Antibiotic resistant bacteria may persist in effluent receiving surface water in the presence of low (sub-inhibitory) antibiotic concentrations if the bacteria possess multiple genes encoding resistance to the same antibiotic. This redundancy of antibiotic resistance genes may occur in plasmids harboring conjugation and mobilization (mob) and integrase (intI) genes. Plasmids extracted from 76 sulfamethoxazole-trimethoprim resistant Escherichia coli originally isolated from effluent and an effluent-receiving stream were used as DNA template to identify sulfamethoxazole (sul) and trimethoprim (dfr) resistances genes plus detect the presence of intI and mob genes using PCR. Sulfamethoxazole and trimethoprim resistance was plasmid-mediated with three sul (sul1, sul2 and sul3 genes) and four dfr genes (dfrA12, dfrA8, dfrA17, and dfrA1 gene) the most prevalently detected. Approximately half of the plasmids carried class 1 and/or 2 integron and, although unrelated, half were also transmissible. Sampling site in relationship to effluent input significantly affected the number of intI and mob but not the number of sul and dfr genes. In the presence of low (sub-inhibitory) sulfamethoxazole concentration, isolates persisted regardless of integron and mobilization gene designation, whereas in the presence of trimethoprim, the presence of both integron and mobilization genes made isolates less persistent than in the absence of both or the presence of a gene from either group individually. Regardless, isolates persisted in large concentrations throughout the experiment. Treated effluent containing antibiotic resistant bacteria may be an important source of integrase and mobilization genes into the stream environment. Sulfamethoxazole-trimethoprim resistant bacteria may have a high degree of genetic redundancy and diversity carrying resistance to each antibiotic, although the role of integrase and mobilization genes towards persistence is unclear. PMID:27455416

  12. Prevalence of antibiotic resistance genes in the bacterial flora of integrated fish farming environments of Pakistan and Tanzania.

    PubMed

    Shah, Syed Q A; Colquhoun, Duncan J; Nikuli, Hamisi L; Sørum, Henning

    2012-08-21

    The use of a wide variety of antimicrobials in human and veterinary medicine, including aquaculture, has led to the emergence of antibiotic resistant pathogens. In the present study, bacteria from water, sediments, and fish were collected from fish farms in Pakistan and Tanzania with no recorded history of antibiotic use. The isolates were screened for the presence of resistance genes against various antimicrobials used in aquaculture and animal husbandry. Resistant isolates selected by disk diffusion and genotyped by Southern hybridization were further screened by polymerase chain reaction (PCR) and amplicon sequencing. The prominent resistance genes identified encoded tetracycline [tetA(A) and tetA(G)], trimethoprim [dfrA1, dfrA5, dfrA7, dfrA12, and dfrA15], amoxicillin [bla(TEM)], streptomycin [strA-strB], chloramphenicol [cat-1], and erythromycin resistance [mefA]. The int1 gene was found in more than 30% of the bacterial isolates in association with gene cassettes. MAR indices ranged from 0.2 to 1. The bla(NDM-1) gene was not identified in ertapenem resistant isolates. It is hypothesized that integrated fish farming practices utilizing domestic farm and poultry waste along with antibiotic residues from animal husbandry may have contributed to a pool of resistance genes in the aquaculture systems studied.

  13. Richard Willstätter and the 1915 Nobel Prize in chemistry.

    PubMed

    Trauner, Dirk

    2015-10-01

    One hundred years after his Nobel Prize, Richard Willstätter's achievements and the fascinating role he played in 20th century chemistry are discussed in this Essay. Several of his discoveries, such as the anthocyanidins, cyclooctatetraene, the ortho-quinones, and the structure of cocaine, will forever be associated with his name.

  14. Richard Willstätter and the 1915 Nobel Prize in chemistry.

    PubMed

    Trauner, Dirk

    2015-10-01

    One hundred years after his Nobel Prize, Richard Willstätter's achievements and the fascinating role he played in 20th century chemistry are discussed in this Essay. Several of his discoveries, such as the anthocyanidins, cyclooctatetraene, the ortho-quinones, and the structure of cocaine, will forever be associated with his name. PMID:26291186

  15. Anthocyanin Interactions with DNA: Intercalation, Topoisomerase I Inhibition and Oxidative Reactions

    PubMed Central

    Webb, Michael R.; Min, Kyungmi; Ebeler, Susan E.

    2009-01-01

    Anthocyanins and their aglycone anthocyanidins are pigmented flavonoids found in significant amounts in many commonly consumed foods. They exhibit a complex chemistry in aqueous solution, which makes it difficult to study their chemistry under physiological conditions. Here we used a gel electrophoresis assay employing supercoiled DNA plasmid to examine the ability of these compounds (1) to intercalate DNA, (2) to inhibit human topoisomerase I through both inhibition of plasmid relaxation activity (catalytic inhibition) and stabilization of the cleavable DNA-topoisomerase complex (poisoning), and (3) to inhibit or enhance oxidative single-strand DNA nicking. We found no evidence of DNA intercalation by anthocyan(id)ins in the physiological pH range for any of the compounds used in this study—cyanidin chloride, cyanidin 3-O-glucoside, cyanidin 3,5-O-diglucoside, malvidin 3-O-glucoside and luteolinidin chloride. The anthocyanins inhibited topoisomerase relaxation activity only at high concentrations (> 50 μM) and we could find no evidence of topoisomerase I cleavable complex stabilization by these compounds. However, we observed that all of the anthocyan(id)ins used in this study were capable of inducing significant oxidative DNA strand cleavage (nicking) in the presence of 1 mM DTT (dithiothreitol), while the free radical scavenger, DMSO, at concentrations typically used in similar studies, completely inhibited DNA nicking. Finally, we propose a mechanism to explain the anthocyan(id)in induced oxidative DNA cleavage observed under our experimental conditions. PMID:19924259

  16. Detect and classify faults using neural nets

    SciTech Connect

    Kezunovic, M.; Rikalo, I.

    1996-10-01

    The analysis of transmission line faults is essential to the proper performance of the power system. It is required if protective relays are to take the appropriate action and in monitoring the performance of relays, circuit breakers, and other protective and control elements. The detection and classification of transmission line faults is a fundamental component of such fault analysis. Another application of fault analysis is in software packages for automated analysis of digital fault recorder (DFR) files. Recently, such a package, called DFR Assistant, was developed for substation applications. This program can be installed locally in a substation, in which case it is connected directly to the DFR via a high speed parallel link, or it can be installed at a central station, in which case it can be configured to automatically analyze events coming from all DFRs.

  17. Class 1 and class 2 integrons and plasmid-mediated antibiotic resistance in coliforms isolated from ten rivers in northern Turkey.

    PubMed

    Ozgumus, Osman Birol; Sandalli, Cemal; Sevim, Ali; Celik-Sevim, Elif; Sivri, Nuket

    2009-02-01

    We aimed to determine the molecular mechanisms of antibiotic resistance in coliforms isolated from ten rivers in northern region of Turkey. A total of 183 isolates were tested for antimicrobial susceptibility by disk diffusion and agar dilution methods. Resistance to ampicillin, streptomycin, trimethoprim, tetracycline, and chloramphenicol was detected in 58%, 51.9%, 24%, 28.4%, and 12.5%, respectively. Twelve (6.5%) phylogenetically distant organisms were detected to harbor self-transmissible plasmids ranging 52 to >147 kb in sizes. Resistances to ampicillin, tetracycline, trimethoprim, streptomycin, and nalidixic acid were commonly transferable traits. Transferable nalidixic acid-resistant strains harbored qnrS gene, which was the first report of plasmid-mediated quinolone resistance in bacteria of environmental origin in Turkey. Fourteen and five coliforms harbored class 1 and class 2 integrons, respectively, and some of them were located on transferable plasmids. Sequence analyses of variable regions of the class 1 and 2 integrons harbored various gene cassettes, dfrA1, dfr2d, dfrA7, dfrA16, dfrA17, aadA1, aadA5, bla(oxA-30), and sat1. A gene cassette array, dfrA16 has been demonstrated for the first time in a Citrobacter koseri isolate. Class 1 and class 2-bearing strains were clustered in different groups by BOX-PCR fingerprinting. Rivers in the northern Turkey may act as receptacle for the multi-drug resistant enterobacteria and can serve as reservoirs of the antimicrobial resistance determinants in the environment. The actual risk to public health is the transfer of resistance genes from the environmental bacteria to human pathogens.

  18. Sequence of pNL194, a 79.3-kilobase IncN plasmid carrying the blaVIM-1 metallo-beta-lactamase gene in Klebsiella pneumoniae.

    PubMed

    Miriagou, V; Papagiannitsis, C C; Kotsakis, S D; Loli, A; Tzelepi, E; Legakis, N J; Tzouvelekis, L S

    2010-10-01

    The nucleotide sequence of pNL194, a VIM-1-encoding plasmid, is described in this study. pNL194 (79,307 bp) comprised an IncN-characteristic segment (38,940 bp) and a mosaic structure (40,367 bp) including bla(VIM-1), aacA7, aadA1, aadA2, dfrA1, dfrA12, aphA1, strA, strB, and sul1. Tn1000 or Tn5501 insertion within fipA probably facilitated recruitment of additional mobile elements carrying resistance genes. PMID:20660690

  19. Sequence of pNL194, a 79.3-Kilobase IncN Plasmid Carrying the blaVIM-1 Metallo-β-Lactamase Gene in Klebsiella pneumoniae▿

    PubMed Central

    Miriagou, V.; Papagiannitsis, C. C.; Kotsakis, S. D.; Loli, A.; Tzelepi, E.; Legakis, N. J.; Tzouvelekis, L. S.

    2010-01-01

    The nucleotide sequence of pNL194, a VIM-1-encoding plasmid, is described in this study. pNL194 (79,307 bp) comprised an IncN-characteristic segment (38,940 bp) and a mosaic structure (40,367 bp) including blaVIM-1, aacA7, aadA1, aadA2, dfrA1, dfrA12, aphA1, strA, strB, and sul1. Tn1000 or Tn5501 insertion within fipA probably facilitated recruitment of additional mobile elements carrying resistance genes. PMID:20660690

  20. Sequence of pNL194, a 79.3-kilobase IncN plasmid carrying the blaVIM-1 metallo-beta-lactamase gene in Klebsiella pneumoniae.

    PubMed

    Miriagou, V; Papagiannitsis, C C; Kotsakis, S D; Loli, A; Tzelepi, E; Legakis, N J; Tzouvelekis, L S

    2010-10-01

    The nucleotide sequence of pNL194, a VIM-1-encoding plasmid, is described in this study. pNL194 (79,307 bp) comprised an IncN-characteristic segment (38,940 bp) and a mosaic structure (40,367 bp) including bla(VIM-1), aacA7, aadA1, aadA2, dfrA1, dfrA12, aphA1, strA, strB, and sul1. Tn1000 or Tn5501 insertion within fipA probably facilitated recruitment of additional mobile elements carrying resistance genes.

  1. Deployable Fresnel Rings

    NASA Technical Reports Server (NTRS)

    Kennedy, Timothy F.; Fink, Patrick W.; Chu, Andrew W.; Lin, Gregory Y.

    2014-01-01

    Deployable Fresnel rings (DFRs) significantly enhance the realizable gain of an antenna. This innovation is intended to be used in combination with another antenna element, as the DFR itself acts as a focusing or microwave lens element for a primary antenna. This method is completely passive, and is also completely wireless in that it requires neither a cable, nor a connector from the antenna port of the primary antenna to the DFR. The technology improves upon the previous NASA technology called a Tri-Sector Deployable Array Antenna in at least three critical aspects. In contrast to the previous technology, this innovation requires no connector, cable, or other physical interface to the primary communication radio or sensor device. The achievable improvement in terms of antenna gain is significantly higher than has been achieved with the previous technology. Also, where previous embodiments of the Tri-Sector antenna have been constructed with combinations of conventional (e.g., printed circuit board) and conductive fabric materials, this innovation is realized using only conductive and non-conductive fabric (i.e., "e-textile") materials, with the possible exception of a spring-like deployment ring. Conceptually, a DFR operates by canceling the out-of-phase radiation at a plane by insertion of a conducting ring or rings of a specific size and distance from the source antenna, defined by Fresnel zones. Design of DFRs follow similar procedures to those outlined for conventional Fresnel zone rings. Gain enhancement using a single ring is verified experimentally and through computational simulation. The experimental test setup involves a microstrip patch antenna that is directly behind a single-ring DFR and is radiating towards a second microstrip patch antenna. The first patch antenna and DFR are shown. At 2.42 GHz, the DFR improves the transmit antenna gain by 8.6 dB, as shown in Figure 2, relative to the wireless link without the DFR. A figure illustrates the

  2. Anthocyanin regulatory/structural gene expression in Phalaenopsis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Partial cDNA fragments of Myb, Myc, Wd, Chs and Dfr genes were generated by Reverse Transcription-PCR using total RNA isolated from flowers of P. amabilis (L.) Blume (anthocyanin-free) and P. schilleriana Rchb. f. (anthocyanin-containing) and cloned into a TOPO vector. RT-PCR revealed that the struc...

  3. 77 FR 65497 - Gross Combination Weight Rating (GCWR); Definition

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-10-29

    ... (77 FR 51706) is withdrawn effective October 26, 2012. ] FOR FURTHER INFORMATION CONTACT: Tom Kelly... Federal Register (73 FR 3316). II. Background On August 27, 2012, FMCSA published a DFR to amend the..., 2012 (77 FR 51706). Issued on: October 22, 2012. Larry W. Minor, Associate Administrator, Office...

  4. 40 CFR 80.620 - What are the additional requirements for diesel fuel or distillates produced by foreign...

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... documents that reflect movement and storage of the certified DFR-Diesel from the refinery to the load port... as specified in paragraph (n)(1) of this section, and a description of the diesel fuel's movement and... the United States related to the requirements of this subpart. (3) The forum for any civil or...

  5. Characterization of integron-mediated antimicrobial resistance among Escherichia coli strains isolated from a captive population of Amur tigers in China.

    PubMed

    Xue, Yuan; Chen, Jianfei; Wang, Yulong; Zhang, Yanlong; Liu, Dan; Hua, Yuping

    2013-12-01

    The present study was undertaken to identify and characterize integrons and integrated resistance gene cassettes among multidrug resistant Escherichia coli isolates from a captive population of Amur tigers (Panthera tigris altaica) in China. In addition, the prevalence of antimicrobial resistance and class I integrons was assessed in E. coli strains (n = 61) isolated from a captive population of Amur tigers in Heilongjiang Amur Tiger Park, China. Among the isolates, 52.46% (32 of 61) were positive for intI1, but no isolates carried intI2 or intI3. Most isolates were susceptible to amoxicillin/clavulanic acid, aztreonam, and polymyxin B, while they also exhibited high incidence rates of resistance to ampicillin, doxycycline, chloramphenicol, tetracycline, and dihydrofolate reductase. Sequencing analysis revealed three gene cassettes, which encoded resistance to dihydrofolate reductase (dfrA15), dihydrofolate reductase (dfrA12), and adenyltransferase (aadA2). The gene cassette arrays dfrA15 (31%) and dfrA12-aadA2 (19%) were most prevalent among these isolates.

  6. 78 FR 2147 - Seagoing Barges

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-01-09

    ... Act notice regarding our public dockets in the January 17, 2008, issue of the Federal Register (73 FR... order FR Federal Register NPRM Notice of proposed rulemaking OCMI Officer in Charge, Marine Inspection... December 14, 2011, the Coast Guard published a direct final rule (DFR) entitled ``Seagoing Barges.'' 76...

  7. 77 FR 42988 - Updating OSHA Construction Standards Based on National Consensus Standards; Head Protection...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-07-23

    ... DFR on June 22, 2012 (77 FR 37587). OSHA also is publishing a correction to the proposed rule that it published the same day in the Federal Register (77 FR 37617). DATES: This correction to the direct final... From the Federal Register Online via the Government Publishing Office DEPARTMENT OF...

  8. Occurrence and molecular characteristics of antimicrobial resistance of Escherichia coli from broilers, pigs and meat products in Thailand and Cambodia provinces.

    PubMed

    Trongjit, Suthathip; Angkittitrakul, Sunpetch; Chuanchuen, Rungtip

    2016-09-01

    Nine hundred and forty-one samples were collected in Sa Keao, Thailand (n = 554) and Banteay Meanchey, Cambodia (n = 387) from July 2014 to January 2015. A total of 667 Escherichia coli isolates (381 isolates from Sa Keao and 286 isolates from Banteay Meanchey) were obtained and examined for antimicrobial susceptibility, class 1 integrons, ESBL genes and horizontal transfer of resistance determinants. Prevalence of E. coli in pig and broiler carcass samples from slaughterhouses and fresh markets was 36-85% in Sa Keao and 11-69% in Banteay Meanchey. The majority of these isolates were multidrug resistant (75.3%). Class 1 integrons were common in both Thai (47%) and Cambodian (62%) isolates, of which four resistance gene cassette arrays including aadA1, dfrA1-aadA1, dfrA12-aadA2 and aadA2-linF were identified. Class 1 integrons in two broiler isolates from Sa Keao (dfrA12-aadA2) and one broiler isolate from Banteay Meanchey (dfrA1-aadA1) were horizontally transferable. Sixteen isolates were confirmed to be ESBL-producing strains with ESBL gene blaCTX-M-15 , broad spectrum β-lactamase gene blaTEM-1 and the AmpC gene blaCMY-2 being detected. The blaTEM-1 gene was most prevalent and located on a conjugative plasmid. PMID:27474453

  9. Characterization of integron-mediated antimicrobial resistance among Escherichia coli strains isolated from a captive population of Amur tigers in China.

    PubMed

    Xue, Yuan; Chen, Jianfei; Wang, Yulong; Zhang, Yanlong; Liu, Dan; Hua, Yuping

    2013-12-01

    The present study was undertaken to identify and characterize integrons and integrated resistance gene cassettes among multidrug resistant Escherichia coli isolates from a captive population of Amur tigers (Panthera tigris altaica) in China. In addition, the prevalence of antimicrobial resistance and class I integrons was assessed in E. coli strains (n = 61) isolated from a captive population of Amur tigers in Heilongjiang Amur Tiger Park, China. Among the isolates, 52.46% (32 of 61) were positive for intI1, but no isolates carried intI2 or intI3. Most isolates were susceptible to amoxicillin/clavulanic acid, aztreonam, and polymyxin B, while they also exhibited high incidence rates of resistance to ampicillin, doxycycline, chloramphenicol, tetracycline, and dihydrofolate reductase. Sequencing analysis revealed three gene cassettes, which encoded resistance to dihydrofolate reductase (dfrA15), dihydrofolate reductase (dfrA12), and adenyltransferase (aadA2). The gene cassette arrays dfrA15 (31%) and dfrA12-aadA2 (19%) were most prevalent among these isolates. PMID:24450054

  10. 77 FR 13969 - Revising Standards Referenced in the Acetylene Standard

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-03-08

    ... Association (GGA) acetylene standard (see 76 FR 75782). In the DFR, OSHA deleted reference to CGA G-1-2003 and... final rule published on December 5, 2011 (76 FR 75782), is effective on March 5, 2012. For the purposes....C. 553, Secretary of Labor's Order 1-2012 (77 FR 3912), and 29 CFR part 1911. Signed at...

  11. Precise spatio-temporal regulation of the anthocyanin biosynthetic pathway leads to petal spot formation in Clarkia gracilis (Onagraceae)

    PubMed Central

    Martins, Talline R.; Berg, Jeremy J.; Blinka, Steven; Rausher, Mark D.; Baum, David A.

    2012-01-01

    Summary Petal spots are widespread in Angiosperms and are often implicated in pollinator attraction. Clarkia gracilis petals each have a single red-purple spot that contrasts against a pink background. The position and presence of spots in C. gracilis are determined by the epistatic interaction of alleles at two, as-yet-unidentified loci.We used HPLC to identify the different pigments produced in the petals, and qualitative and quantitative RT-PCR to assay for spatio-temporal patterns of expression of different anthocyanin pathway genes.We found that spots contain different pigments from the remainder of the petal, being composed of cyanidin/peonidin-based, instead of malvidin-based anthocyanins. Expression assays of anthocyanin pathway genes show that Dfr2 has a spot-specific expression pattern and acts as a switch for spot production. Co-segregation analyses implicate the gene products of the P and I loci as trans-regulators of this switch. Spot pigments appear earlier in development due to early expression of Dfr2 and F3′h1. Pigments in the background appear later, due to later expression of Dfr1 and F3′5′h1.The evolution of this spot production mechanism appears to have been facilitated by duplication of the Dfr gene and to have required substantial reworking of the anthocyanin pathway regulatory network. PMID:23231386

  12. Diversity of antimicrobial resistance and virulence genes in methicillin-resistant non-Staphylococcus aureus staphylococci from veal calves.

    PubMed

    Argudín, M Angeles; Vanderhaeghen, Wannes; Butaye, Patrick

    2015-04-01

    In this study we determined whether methicillin-resistant non-Staphylococcus aureus (MRNAS) from veal calves may be a potential reservoir of antimicrobial-resistance and virulence genes. Fifty-eight MRNAS were studied by means of DNA-microarray and PCR for detection of antimicrobial resistance and virulence genes. The isolates carried a variety of antimicrobial-resistance genes [aacA-aphD, aadD, aph3, aadE, sat, spc, ampA, erm(A), erm(B), erm(C), erm(F), erm(T), lnu(A), msr(A)-msr(B), vga(A), mph(C), tet(K), tet(M), tet(L), cat, fexA, dfrA, dfrD, dfrG, dfrK, cfr, fusB, fosB, qacA, qacC, merA-merB]. Some isolates carried resistance genes without showing the corresponding resistance phenotype. Most MRNAS carried typical S. aureus virulence factors like proteases (sspP) and enterotoxins (seg) genes. Most Staphylococcus epidermidis isolates carried the arginine catabolic element, and nearly 40% of the Staphylococcus sciuri isolates carried leukocidins, and/or fibronectin-binding protein genes. MRNAS were highly multi-resistant and represent an important reservoir of antimicrobial resistance and virulence genes.

  13. 78 FR 65932 - Updating OSHA Standards Based on National Consensus Standards; Signage

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-11-04

    ... rule (DFR) (see 78 FR 35585) updating its signage standards for general industry and construction. In... rule published June 13, 2013 (78 FR 35585). FOR FURTHER INFORMATION CONTACT: General information and... 29 U.S.C. 653, 655, and 657, 5 U.S.C. 553, Secretary of Labor's Order 1-2012 (77 FR 3912), and 29...

  14. 78 FR 66642 - Updating OSHA Standards Based on National Consensus Standards; Signage

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-11-06

    ..., 2013, OSHA published in the Federal Register (78 FR 35559) a direct final rule that revised its signage...) signage protection standards (see 78 FR 35559). Specifically, the DFR updated the signage references in... September 11, 2013. DATES: The direct final rule published on June 13, 2013 (78 FR 35559), became...

  15. Diffusion Behavior of Cumulative He Doped in Cu/W Multilayer Nanofilms at Room Temperature

    NASA Astrophysics Data System (ADS)

    Ling, Wang; Wang, Liu; Yue, Li; Yun-Long, Shi; Yuan-Xia, Lao; Xiao-Bo, Lu; Ai-Hong, Deng; Yuan, Wang

    2016-06-01

    Not Available Supported by the National Natural Science Foundation of China under Grant Nos 11275132, 51171124 and 11505121, the International Science and Technology Cooperation Program of China under Grant No 2014DFR50710, and the Scientific and Technical Supporting Programs Funded by the Science and Technology Department of Sichuan Province under Grant No 2014GZ0004.

  16. Resistance and integron characterization of Acinetobacter baumannii in a teaching hospital in Chongqing, China

    PubMed Central

    Huang, C.; Long, Q.; Qian, K.; Fu, T.; Zhang, Z.; Liao, P.; Xie, J.

    2015-01-01

    A total of 189 Acinetobacter baumannii isolates were collected in 2011 from a teaching hospital in Chongqing, China. Susceptibility data showed strains carrying integrons were significantly more resistant to all tested antibiotics that strains lacking integrons. Five types of gene cassettes belonging to class I integrons were identified in this study, and for the first time two types of gene cassettes belonging to class II integrons are reported. Most of the cassettes belong to a class I integron (136/144) encoding arr3, aacA4, dfrA17, aadA5, aadB, cat, blaOXA10, aadA1, aadA2, dfrA and aacC1. Isolates contained a class I gene cassette; AadA2-HP-dfrA was the prevalent strain in this hospital. A class II integron was detected in eight strains, which contained the type IV fimbriae expression regulatory gene pilR and sulfate adenylyltransferase, suggesting a possible role in multidrug resistance. The major epidemic strains from intensive care unit patients belong to international clone 2. In conclusion, the presence of integrons was significantly associated with multiple drug resistance of A. baumannii in this hospital, and class I integron isolates bearing AadA2-HP-dfrA were the prevalent strain in this hospital. PMID:26649184

  17. 75 FR 27239 - Revising the Notification Requirements in the Exposure Determination Provisions of the Hexavalent...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-05-14

    ..., 2010 (75 FR 12485), is withdrawn. FOR FURTHER INFORMATION CONTACT: For general information and press....1026, and 29 CFR 1926.1126 (75 FR 12681). OSHA also published a companion proposed rule proposing the same changes to the Cr(VI) standards. (75 FR 12485, March 16, 2010). In the DFR, OSHA stated that...

  18. Antimicrobial resistance and antimicrobial resistance genes in marine bacteria from salmon aquaculture and non-aquaculture sites.

    PubMed

    Shah, Syed Q A; Cabello, Felipe C; L'abée-Lund, Trine M; Tomova, Alexandra; Godfrey, Henry P; Buschmann, Alejandro H; Sørum, Henning

    2014-05-01

    Antimicrobial resistance (AR) detected by disc diffusion and antimicrobial resistance genes detected by DNA hybridization and polymerase chain reaction with amplicon sequencing were studied in 124 marine bacterial isolates from a Chilean salmon aquaculture site and 76 from a site without aquaculture 8 km distant. Resistance to one or more antimicrobials was present in 81% of the isolates regardless of site. Resistance to tetracycline was most commonly encoded by tetA and tetG; to trimethoprim, by dfrA1, dfrA5 and dfrA12; to sulfamethizole, by sul1 and sul2; to amoxicillin, by blaTEM ; and to streptomycin, by strA-strB. Integron integrase intl1 was detected in 14 sul1-positive isolates, associated with aad9 gene cassettes in two from the aquaculture site. intl2 Integrase was only detected in three dfrA1-positive isolates from the aquaculture site and was not associated with gene cassettes in any. Of nine isolates tested for conjugation, two from the aquaculture site transferred AR determinants to Escherichia coli. High levels of AR in marine sediments from aquaculture and non-aquaculture sites suggest that dispersion of the large amounts of antimicrobials used in Chilean salmon aquaculture has created selective pressure in areas of the marine environment far removed from the initial site of use of these agents.

  19. Estimation of daily human intake of food flavonoids.

    PubMed

    Mullie, Patrick; Clarys, Peter; Deriemaeker, Peter; Hebbelinck, Marcel

    2008-06-01

    The daily intake of food flavonoids was determined using a semiquantitative food frequency questionnaire (FFQ) and a 4-day food record (4DFR) in a group of 45 female Flemish dietitians. The subjects were asked to report their food intake three times: day 1 using the FFQ (FFQ1); between days 2 and 13 using a 4-day non-consecutive food diary (4DFR); and again using the FFQ on day 14 (FFQ2). The total flavonoid intakes (mean and standard deviation) as estimated with the different methods were: for FFQ1, 166.0 +/- 146.6 mg/day; for 4DFR, 203.0 +/- 243.2 mg/day; and for FFQ2, 158.3 +/- 151.8 mg/day. There was a significant different estimate for the amount of flavan-3-ols, flavanones and flavones when comparing the two FFQs with the 4DFR. The two research methods classified 88% of the 45 dietitians in the same or in an adjacent quartile for total flavonoid intake. The findings of this study indicate that the developed FFQ seems to be a simple and reliable method to assign subjects in quartiles of flavonoid intake.

  20. An Escherichia coli strain, PGB01, isolated from feral pigeon Faeces, thermally fit to survive in pigeon, shows high level resistance to trimethoprim.

    PubMed

    Kumar, Arvind; Tiwary, Bipransh Kumar; Kachhap, Sangita; Nanda, Ashis Kumar; Chakraborty, Ranadhir

    2015-01-01

    In this study, of the hundred Escherichia coli strains isolated from feral Pigeon faeces, eighty five strains were resistant to one or more antibiotics and fifteen sensitive to all the antibiotics tested. The only strain (among all antibiotic-resistant E. coli isolates) that possessed class 1 integron was PGB01. The dihydrofolate reductase gene of the said integron was cloned, sequenced and expressed in E. coli JM109. Since PGB01 was native to pigeon's gut, we have compared the growth of PGB01 at two different temperatures, 42°C (normal body temperature of pigeon) and 37°C (optimal growth temperature of E. coli; also the human body temperature), with E. coli K12. It was found that PGB01 grew better than the laboratory strain E. coli K12 at 37°C as well as at 42°C. In the thermal fitness assay, it was observed that the cells of PGB01 were better adapted to 42°C, resembling the average body temperature of pigeon. The strain PGB01 also sustained more microwave mediated thermal stress than E. coli K12 cells. The NMR spectra of the whole cells of PGB01 varied from E. coli K12 in several spectral peaks relating some metabolic adaptation to thermotolerance. On elevating the growth temperature from 37°C to 42°C, susceptibility to kanamycin (both strains were sensitive to it) of E. coli K12 was increased, but in case of PGB01 no change in susceptibility took place. We have also attempted to reveal the basis of trimethoprim resistance phenotype conferred by the dfrA7 gene homologue of PGB01. Molecular Dynamics (MD) simulation study of docked complexes, PGB01-DfrA7 and E. coli TMP-sensitive-Dfr with trimethoprim (TMP) showed loss of some of the hydrogen and hydrophobic interaction between TMP and mutated residues in PGB01-DfrA7-TMP complex compared to TMP-sensitive-Dfr-TMP complex. This loss of interaction entails decrease in affinity of TMP for PGB01-DfrA7 compared to TMP-sensitive-Dfr. PMID:25750990

  1. An Escherichia coli strain, PGB01, isolated from feral pigeon Faeces, thermally fit to survive in pigeon, shows high level resistance to trimethoprim.

    PubMed

    Kumar, Arvind; Tiwary, Bipransh Kumar; Kachhap, Sangita; Nanda, Ashis Kumar; Chakraborty, Ranadhir

    2015-01-01

    In this study, of the hundred Escherichia coli strains isolated from feral Pigeon faeces, eighty five strains were resistant to one or more antibiotics and fifteen sensitive to all the antibiotics tested. The only strain (among all antibiotic-resistant E. coli isolates) that possessed class 1 integron was PGB01. The dihydrofolate reductase gene of the said integron was cloned, sequenced and expressed in E. coli JM109. Since PGB01 was native to pigeon's gut, we have compared the growth of PGB01 at two different temperatures, 42°C (normal body temperature of pigeon) and 37°C (optimal growth temperature of E. coli; also the human body temperature), with E. coli K12. It was found that PGB01 grew better than the laboratory strain E. coli K12 at 37°C as well as at 42°C. In the thermal fitness assay, it was observed that the cells of PGB01 were better adapted to 42°C, resembling the average body temperature of pigeon. The strain PGB01 also sustained more microwave mediated thermal stress than E. coli K12 cells. The NMR spectra of the whole cells of PGB01 varied from E. coli K12 in several spectral peaks relating some metabolic adaptation to thermotolerance. On elevating the growth temperature from 37°C to 42°C, susceptibility to kanamycin (both strains were sensitive to it) of E. coli K12 was increased, but in case of PGB01 no change in susceptibility took place. We have also attempted to reveal the basis of trimethoprim resistance phenotype conferred by the dfrA7 gene homologue of PGB01. Molecular Dynamics (MD) simulation study of docked complexes, PGB01-DfrA7 and E. coli TMP-sensitive-Dfr with trimethoprim (TMP) showed loss of some of the hydrogen and hydrophobic interaction between TMP and mutated residues in PGB01-DfrA7-TMP complex compared to TMP-sensitive-Dfr-TMP complex. This loss of interaction entails decrease in affinity of TMP for PGB01-DfrA7 compared to TMP-sensitive-Dfr.

  2. Anthocyanins present in selected tropical fruits: acerola, jambolão, jussara, and guajiru.

    PubMed

    de Brito, Edy Sousa; de Araújo, Manuela Cristina Pessanha; Alves, Ricardo Elesbão; Carkeet, Colleen; Clevidence, Beverly A; Novotny, Janet A

    2007-11-14

    Many tropical fruits are rich in anthocyanins, though limited information is available about the characterization and quantification of these anthocyanins. The identification and quantification of anthocyanin pigments in four tropical fruits were determined by HPLC-MS/MS. Fruits studied included acerola (Malphigia emarginata), jussara (Euterpe edulis), jambolão (Syzygium cumini), and guajiru (Chrysobalanus icaco). All four fruits were found to contain anthocyanin pigments. Anthocyanidin backbones included cyanidin, delphinidin, peonidin, pelargonidin, petunidin, and malvidin. Guajiru contained several acylated forms, while acerola, jussara, and jambolão contained only nonacylated glycosides. These results demonstrate that these tropical fruits are rich in anthocyanins and that the anthocyanins are widely ranging in anthocyanidin backbone, glycosylation, and acylation.

  3. The influence of tissue handling on the flavonoid content of the aquatic plant Posidonia oceanica.

    PubMed

    Cannac, Magali; Ferrat, Lila; Barboni, Toussaint; Pergent, Gerard; Pasqualini, Vanina

    2007-05-01

    In recent times, more and more studies have focused on flavonoids as biomarkers of environmental quality in aquatic plants, in particular, Posidonia oceanica (Linnaeus) Delile. It is therefore of interest to determine how different prehandling methods can affect flavonoid concentrations. The methods tested were (1) immediate extraction of fresh samples, (2) extraction after 48 hr chilling, (3) freeze-drying, and (4) oven drying. Chilling and freeze-drying considerably altered the quantity of flavonoids measured, but not their profile. The effect of oven drying was not significant. Chilling led to a loss of 57% of total (pro)anthocyanidins, 39% of total flavonols, and 48% of all simple flavonols (myricetin, quercetin, isorhamnetin, and kaempferol). Freeze-drying caused a loss of 71% of total (pro)anthocyanidins, 87% of total flavonols, and 95% of all simple flavonols. PMID:17404815

  4. Occurrence of Cellulose-Producing Gluconacetobacter spp. in Fruit Samples and Kombucha Tea, and Production of the Biopolymer.

    PubMed

    Neera; Ramana, Karna Venkata; Batra, Harsh Vardhan

    2015-06-01

    Cellulose producing bacteria were isolated from fruit samples and kombucha tea (a fermented beverage) using CuSO4 solution in modified Watanabe and Yamanaka medium to inhibit yeasts and molds. Six bacterial strains showing cellulose production were isolated and identified by 16S rRNA gene sequencing as Gluconacetobacter xylinus strain DFBT, Ga. xylinus strain dfr-1, Gluconobacter oxydans strain dfr-2, G. oxydans strain dfr-3, Acetobacter orientalis strain dfr-4, and Gluconacetobacter intermedius strain dfr-5. All the cellulose-producing bacteria were checked for the cellulose yield. A potent cellulose-producing bacterium, i.e., Ga. xylinus strain DFBT based on yield (cellulose yield 5.6 g/L) was selected for further studies. Cellulose was also produced in non- conventional media such as pineapple juice medium and hydrolysed corn starch medium. A very high yield of 9.1 g/L cellulose was obtained in pineapple juice medium. Fourier transform infrared spectrometer (FT-IR) analysis of the bacterial cellulose showed the characteristic peaks. Soft cellulose with a very high water holding capacity was produced using limited aeration. Scanning electron microscopy (SEM) was used to analyze the surface characteristics of normal bacterial cellulose and soft cellulose. The structural analysis of the polymer was performed using (13)C solid-state nuclear magnetic resonance (NMR). More interfibrillar space was observed in the case of soft cellulose as compared to normal cellulose. This soft cellulose can find potential applications in the food industry as it can be swallowed easily without chewing.

  5. Occurrence of Cellulose-Producing Gluconacetobacter spp. in Fruit Samples and Kombucha Tea, and Production of the Biopolymer.

    PubMed

    Neera; Ramana, Karna Venkata; Batra, Harsh Vardhan

    2015-06-01

    Cellulose producing bacteria were isolated from fruit samples and kombucha tea (a fermented beverage) using CuSO4 solution in modified Watanabe and Yamanaka medium to inhibit yeasts and molds. Six bacterial strains showing cellulose production were isolated and identified by 16S rRNA gene sequencing as Gluconacetobacter xylinus strain DFBT, Ga. xylinus strain dfr-1, Gluconobacter oxydans strain dfr-2, G. oxydans strain dfr-3, Acetobacter orientalis strain dfr-4, and Gluconacetobacter intermedius strain dfr-5. All the cellulose-producing bacteria were checked for the cellulose yield. A potent cellulose-producing bacterium, i.e., Ga. xylinus strain DFBT based on yield (cellulose yield 5.6 g/L) was selected for further studies. Cellulose was also produced in non- conventional media such as pineapple juice medium and hydrolysed corn starch medium. A very high yield of 9.1 g/L cellulose was obtained in pineapple juice medium. Fourier transform infrared spectrometer (FT-IR) analysis of the bacterial cellulose showed the characteristic peaks. Soft cellulose with a very high water holding capacity was produced using limited aeration. Scanning electron microscopy (SEM) was used to analyze the surface characteristics of normal bacterial cellulose and soft cellulose. The structural analysis of the polymer was performed using (13)C solid-state nuclear magnetic resonance (NMR). More interfibrillar space was observed in the case of soft cellulose as compared to normal cellulose. This soft cellulose can find potential applications in the food industry as it can be swallowed easily without chewing. PMID:25926011

  6. Abrasive stripping square-wave voltammetry of blackberry, raspberry, strawberry, pomegranate, and sweet and blue potatoes.

    PubMed

    Komorsky-Lovrić, Šebojka; Novak, Ivana

    2011-08-01

    Electro-oxidation potentials of 7 fruits and vegetables were determined by abrasive stripping voltammetry. The responses were characterized by 2 peaks with maxima at 0.45 and 0.55 V compared with Ag/AgCl, respectively. Both electrode reactions appear reversible at a frequency of 8 Hz. They can be ascribed to anthocyanidins and ellagic acid as electroactive compounds. By this method, an antioxidative capacity of a certain plant can be quickly estimated without extraction of active components.

  7. The protective effect and action mechanism of Vaccinium myrtillus L. on gastric ulcer in mice.

    PubMed

    Ogawa, Kenjirou; Oyagi, Atsushi; Tanaka, Junji; Kobayashi, Saori; Hara, Hideaki

    2011-08-01

    Vaccinium myrtillus L. anthocyanoside (VMA) is used as a folk medicine to treat diseases related to gastric ulcers in northern Europe. However, the effects of VMA and its detailed mechanism on gastric ulcer have not been investigated sufficiently. Therefore, the aim of the present study was to investigate the protective effects of VMA on gastric mucosal damage in a murine gastric ulcer model. First the effects of VMA on ethanol-induced gastric ulcers in mice were investigated. Then, the levels of lipid peroxide in murine stomach homogenates were measured to investigate the antioxidative effects of VMA. In addition, the free radical scavenging activity of VMA and its main anthocyanidins were evaluated by electron spin resonance measurement. Oral administration of VMA (10, 30 and 100 mg/kg) significantly protected gastric mucosa against HCl/ethanol-induced gastric ulcers. Furthermore, VMA inhibited lipid peroxide levels in a concentration-dependent manner and showed high scavenging activity against the superoxide anion radical (·O(2) (-) ) and the hydroxyl radical (·OH). Anthocyanidins also showed scavenging activity against the ·O(2) (-) , while only delphinidin showed high scavenging activity against the ·OH. These findings indicate that the protective effects of VMA on HCl/ethanol-induced gastric mucosal injury may be partially due to the antiperoxidative effects of anthocyanidins.

  8. The protective effect and action mechanism of Vaccinium myrtillus L. on gastric ulcer in mice.

    PubMed

    Ogawa, Kenjirou; Oyagi, Atsushi; Tanaka, Junji; Kobayashi, Saori; Hara, Hideaki

    2011-08-01

    Vaccinium myrtillus L. anthocyanoside (VMA) is used as a folk medicine to treat diseases related to gastric ulcers in northern Europe. However, the effects of VMA and its detailed mechanism on gastric ulcer have not been investigated sufficiently. Therefore, the aim of the present study was to investigate the protective effects of VMA on gastric mucosal damage in a murine gastric ulcer model. First the effects of VMA on ethanol-induced gastric ulcers in mice were investigated. Then, the levels of lipid peroxide in murine stomach homogenates were measured to investigate the antioxidative effects of VMA. In addition, the free radical scavenging activity of VMA and its main anthocyanidins were evaluated by electron spin resonance measurement. Oral administration of VMA (10, 30 and 100 mg/kg) significantly protected gastric mucosa against HCl/ethanol-induced gastric ulcers. Furthermore, VMA inhibited lipid peroxide levels in a concentration-dependent manner and showed high scavenging activity against the superoxide anion radical (·O(2) (-) ) and the hydroxyl radical (·OH). Anthocyanidins also showed scavenging activity against the ·O(2) (-) , while only delphinidin showed high scavenging activity against the ·OH. These findings indicate that the protective effects of VMA on HCl/ethanol-induced gastric mucosal injury may be partially due to the antiperoxidative effects of anthocyanidins. PMID:21290441

  9. A genomics investigation of partitioning into and among flavonoid-derived condensed tannins for carbon sequestration in Populus

    SciTech Connect

    Harding, Scott, A; Tsai, Chung-jui; Lindroth, Richard, L

    2013-03-24

    constituted a metabolic carbon drain in developing leaves that was not observed in the roots. We propose that PEA, in addition to other factors, including flavonoid pathway Myb transcription factors, is an important contributor to carbon management and plant defense in Populus. Objective 3: From work related to the first two objectives, it appeared that CT chemistry, at least in terms of the proportions of mono, di and tri hydroxylation at the phenylpropanoid-derived B-ring, changed little if at all when CT accrual per unit time was increased. A large number of transgenic Populus plants with alterations in the expression of flavonoid pathway genes and the potential to produce B-ring, chemically altered CT were generated during the project. Transgenic lines of Populus tremula Michx. Populus alba L. clone 717-1B4, a low CT producer, were produced that over- or under-express several mid and late flavonoid pathway genes including dihydroxyflavonol reductase (DFR-2 isoforms), leucoanthocyanidin reductase (LAR-3 isoforms), anthocyanidin reductase (ANR-2 isoforms), flavonol synthase (FLS-2 isoforms). A large number of additional transformation constructs (chalcone synthases, flavone synthases, and flavanol hydroxylases) were developed that failed to result in transgenic plants. We have purified CT from several of the successful lines and have obtained evidence from pyrolysis GC-MS that CT chemical composition was altered in transgenic lines harboring overexpression constructs for one of the two DFR isoforms. We have also observed increased CT levels in leaves of those lines, but the increases vary substantially in magnitude from experiment to experiment which has led to ongoing efforts to understand the variation before attempting to publish the findings. Preliminary results from some of the transgenic work were presented: An C*, Luo K, El Kayal W, Harding SA, Tsai C-J (2009) Transgenic manipulation of condensed tannins in Populus. IUFRO Tree Biotechnology Conference, Whistler, BC

  10. Basic Performance of the Standard Retrieval Algorithm for the Dual-frequency Precipitation Radar

    NASA Astrophysics Data System (ADS)

    Seto, S.; Iguchi, T.; Kubota, T.

    2013-12-01

    The core satellite of the Global Precipitation Measurement (GPM) mission, which is scheduled to be launched in 2014, will carry the Dual-frequency Precipitation Radar (DPR). DPR is a successor of the single-frequency Precipitation Radar (PR) currently working on the Tropical Rainfall Measuring Mission (TRMM) satellite. The authors are developing a precipitation retrieval algorithm for the DPR based on the standard algorithm of PR. Precipitation radar measurement depends on drop size distribution (DSD) rather than precipitation rates. Generally, DSD is assumed to follow an exponential or a Gamma distribution with two unknown parameters. Here, DSD can be represented on a 2-dimensional plane. For a single-frequency radar measurement, as two parameters cannot be determined, an empirical power law between radar reflectivity factor Z and rain rate R (Z-R relationship) is used. Assuming Z-R relationship is equivalent to constraining DSD on a 1-dimenisonal curve. In the standard algorithm of PR, an empirical power law between specific attenuation k and Z (k-Z relationship) is given to correct attenuation (Hitschfeld-Bordan method; HB method). Assuming k-Z relationship is also equivalent to constraining DSD on another 1-dimensional curve. Fortunately, space-borne or air-borne radars such as PR can measure surface backscattering cross section and surface reference technique (SRT) is applied to estimate path integrated attenuation (PIA). By referring to PIA estimates, k-Z relationship can be adjusted. In a developing algorithm for DPR, k-Z relation is assumed and the HB method is applied for each frequency. Once attenuation-corrected radar reflectivity factor Ze is given at both frequencies, the dual-frequency ratio (DFR) of Ze's is calculated, and DSD parameters are retrieved easily from the DFR (DFR method). However, the retrieved DSD generally does not agree with assumed k-Z relations. Then, k-Z relations are adjusted to fit the DSD. The HB method and DFR method can be

  11. Resistance gene pool to co-trimoxazole in non-susceptible Nocardia strains

    PubMed Central

    Valdezate, Sylvia; Garrido, Noelia; Carrasco, Gema; Villalón, Pilar; Medina-Pascual, María J.; Saéz-Nieto, Juan A.

    2015-01-01

    The soil-borne pathogen Nocardia sp. causes severe cutaneous, pulmonary, and central nervous system infections. Against them, co-trimoxazole (SXT) constitutes the mainstay of antimicrobial therapy. However, some Nocardia strains show resistance to SXT, but the underlying genetic basis is unknown. We investigated the presence of genetic resistance determinants and class 1–3 integrons in 76 SXT-resistant Nocardia strains by PCR and sequencing. By E test, these clinical strains showed SXT minimum inhibitory concentrations of ≥32:608 mg/L (ratio of 1:19 for trimethoprim: sulfamethoxazole). They belonged to 12 species, being the main representatives Nocardia farcinica (32%), followed by N. flavorosea (6.5%), N. nova (11.8%), N. carnea (10.5%), N. transvalensis (10.5%), and Nocardia sp. (6.5%). The prevalence of resistance genes in the SXT-resistant strains was as follows: sul1 and sul2 93.4 and 78.9%, respectively, dfrA(S1) 14.7%, blaTEM-1 and blaZ 2.6 and 2.6%, respectively, VIM-2 1.3%, aph(3′)-IIIa 40.8%, ermA, ermB, mefA, and msrD 2.6, 77.6, 14.4, and 5.2%, respectively, and tet(O), tet(M), and tet(L) 48.6, 25.0, and 3.9%, respectively. Detected amino acid changes in GyrA were not related to fluoroquinolone resistance, but probably linked to species polymorphism. Class 1 and 3 integrons were found in 93.42 and 56.57% strains, respectively. Class 2 integrons and sul3 genes were not detected. Other mechanisms, different than dfrA(S1), dfrD, dfrF, dfrG, and dfrK, could explain the strong trimethoprim resistance shown by the other 64 strains. For first time, resistance determinants commonly found in clinically important bacteria were detected in Nocardia sp. sul1, sul2, erm(B), and tet(O) were the most prevalent in the SXT-resistant strains. The similarity in their resistome could be due to a common genetic platform, in which these determinants are co-transferred. PMID:25972856

  12. Assessing the benefits of design for recycling for plastics inelectronics: A case study of computer enclosures

    SciTech Connect

    Masanet, Eric; Horvath, Arpad

    2007-12-31

    With the emergence of extended producer responsibilityregulations for electronic devices, it is becoming increasingly importantfor electronics manufacturers to apply design for recycling (DFR) methodsin the design of plastic enclosures. This paper presents an analyticalframework for quantifying the environmental and economic benefits of DFRfor plastic computer enclosures during the design process, usingstraightforward metrics that can be aligned with corporate environmentaland financial performance goals. The analytical framework is demonstratedvia a case study of a generic desktop computer enclosure design, which isrecycled using a typical US "take-back" system for plastics from wasteelectronics. The case study illustrates how the analytical framework canbe used by the enclosure designer to quantify the environmental andeconomic benefits of two important DFR strategies: choosing high-valueresins and minimizing enclosure disassembly time. Uncertainty analysis isperformed to quantify the uncertainty surrounding economic conditions inthe future when the enclosure is ultimately recycled.

  13. Natural mutations in two homoeologous TT8 genes control yellow seed coat trait in allotetraploid Brassica juncea (AABB).

    PubMed

    Padmaja, Lakshmi K; Agarwal, Parul; Gupta, Vibha; Mukhopadhyay, Arundhati; Sodhi, Yaspal S; Pental, Deepak; Pradhan, Akshay K

    2014-02-01

    Identification of the candidate gene responsible for the seed coat colour variation in Brassica juncea was undertaken following an earlier study where two independent loci (BjSc1 and BjSc2) were mapped to two linkage groups, LG A9 and B3 (Padmaja et al. in Theor Appl Genet 111:8-14, 2005). The genome search from BRAD data for the presence of flavonoid genes in B. rapa identified three candidate genes namely, DFR, TT1 and TT8 in the LG A9. Quantitative real-time PCR revealed absence of transcript for the late biosynthetic genes (LBGs) and showed significant reduction of transcript in the TT8 from the developing seeds of yellow-seeded line. While mapping of two DFR genes, the BjuA.DFR and BjuB.DFR did not show perfect co-segregation with the seed coat colour loci, that of the two TT8 genes, BjuA.TT8 and BjuB.TT8 showed perfect co-segregation with the seed coat colour phenotype. The BjuA.TT8 allele from the yellow-seeded line revealed the presence of an insertion of 1,279 bp in the exon 7 and did not produce any transcript as revealed by reverse transcriptase PCR. The BjuB.TT8 allele from the yellow-seeded line revealed the presence of an SNP (C→T) in the exon 7 resulting in a stop codon predicting a truncated protein lacking the C-terminal 8 amino acid residues and produced significantly low level of transcript than its wild-type counterpart. Hence, it is hypothesized that the mutations in both the TT8 genes are required for inhibiting the transcription of LBGs in the yellow-seeded mutant of B. juncea.

  14. Comparison of different transvaginal ovum pick-up protocols to optimise oocyte retrieval and embryo production over a 10-week period in cows.

    PubMed

    Chaubal, S A; Molina, J A; Ohlrichs, C L; Ferre, L B; Faber, D C; Bols, P E J; Riesen, J W; Tian, X; Yang, X

    2006-05-01

    The objective was to develop a simple and effective ovum pick-up (OPU) protocol for cows, optimised for oocyte harvest and subsequent in vitro embryo production (IVP). Five protocols differing in collection frequency, dominant follicle removal (DFR) and FSH stimulation were tested on groups of three cows each, over an interval of 10 consecutive weeks. Performance was evaluated on per OPU session, per week and pooled (3 cowsx10weeks) basis. Among the non-stimulated groups, on a per cow per session basis, once- or twice-weekly OPU had no effect on the mean (+/- S.E.M.) number of follicles aspirated, oocytes retrieved and blastocysts produced (0.6+/-0.8 and 0.7 +/- 0.7, respectively). However, DFR 72 h prior to OPU almost doubled blastocyst production (1.2 +/- 1.3). In stimulated groups, FSH treatment (80 mg IM and 120 mg SC) was given once weekly prior to OPU. Treatment with FSH, followed by twice-weekly OPU, failed to show any synergistic effect of FSH and increased aspiration frequency. When FSH was given 36 h after DFR, followed by OPU 48 h later, more (P < 0.05) follicles (16.0 +/- 5.0), oocytes (10.6 +/- 4.5) and embryos (2.1 +/- 1.2) were obtained during each session, but not on a weekly basis. Pooled results over 10 weeks showed an overall improved performance for the treatment groups with twice-weekly OPU sessions, due to double the number of OPU sessions performed. However, the protocol that consisted of DFR, FSH treatment and a subsequent single OPU per week, was the most productive and cost-effective, with potential commercial appeal.

  15. Determinants of exposure to captan in fruit growing.

    PubMed

    de Cock, J; Heederik, D; Kromhout, H; Boleij, J S; Hoek, F; Wegh, H; Tjoe Ny, E

    1998-03-01

    A series of studies investigated occupational exposure to pesticides among fruit growers in The Netherlands during spraying and reentry of orchards between 1990 and 1992 to identify and quantify determinants of exposure. Determinants of exposure are discussed as a starting point for hazard identification and control. Captan was used as a marker for exposure. Cabin use of the tractor was the most prominent determinant of dermal exposure during spraying. For respiratory exposure, factors related to preparation of pesticides were most prominent. A long duration of exposure may reflect a different exposure situation compared with a short duration of exposure. As different determinants of exposure prevailed for each subgroup, consideration should be given to constructing exposure models for each group separately. Dislodgeable foliar residue (DFR) was the most prominent determinant of exposure for both respiratory and dermal exposure during reentry. However, no significant relation between DFR and dermal exposure of forehead and sternal area was found, perhaps because there was no direct contact with foliage here. Therefore, use of a transfer factor based on DFR to estimate total dermal exposure is only a crude estimate. The half-life of captan on crops varied from 10-17 days, so substantial exposure when entering the orchard is very likely, particularly when spraying frequency is high. The main starting points for reduction of exposure are use of a cabin, DFR, and individual time spent on different tasks. Determinants that are constant over time (cabin use) may have an especially great influence on grouping workers, according to long-term exposure in epidemiological studies. As determinants of exposure vary for the different exposure routes and body locations (for dermal exposure), the measure of interest for a specific study design will decide which determinants are most relevant.

  16. Incidence of antimicrobial-resistance genes and integrons in antibiotic-resistant bacteria isolated from eels and aquaculture ponds.

    PubMed

    Lin, Mao; Wu, Xiaomei; Yan, Qingpi; Ma, Ying; Huang, Lixing; Qin, Yingxue; Xu, Xiaojin

    2016-07-01

    The overuse of antimicrobials in aquaculture has promoted the selection of antimicrobial-resistant bacteria. Here we investigated the abundance of antimicrobial-resistance genes and integrons in 108 strains of antibiotic-resistant bacteria isolated from eels and aquaculture ponds in China. Conventional PCR was implemented to examine common antibiotic-resistance genes, integrons, and their gene cassette arrays. The results showed that the antibiotic-resistance genes blaTEM, tetC, sulI, aadA, floR, and qnrB were detected at high percentages, as were a number of other resistance genes. Class I integrons were present in 79.63% of the strains, and 10 out of 108 isolates carried class II integrons. Class III integrons were not detected. Three strains carried both class I and class II integrons, and 73.26% of the class I integron-positive isolates contained the qacEΔ1/sul1 gene. Fourteen types of integron cassette arrays were found among class I integron-positive isolates. A new array, dfrB4-catB3-blaOXA-10-aadA1, was discovered in this study. The gene cassette array dfrA12-orfF-aadA2 was the most widely distributed. In summary, 23 different gene cassettes encoding resistance to 8 classes of antibiotics were identified in the class I integrons, and the main cassettes contained genes encoding resistance to aminoglycosides (aad) and trimethoprim (dfr). All class II integron-positive strains had only a single gene cassette array, viz. dfrA1-catB2-sat2-aadA1. High levels of antimicrobial-resistance genes and integrons in eels and auqauculture ponds suggest that the overuse of antimicrobials should be strictly controlled and that the levels of bacterial antimicrobial-resistance genes in aquaculture should be monitored. PMID:27409235

  17. The structure and competitive substrate inhibition of dihydrofolate reductase from Enterococcus faecalis reveal restrictions to cofactor docking.

    PubMed

    Bourne, Christina R; Wakeham, Nancy; Webb, Nicole; Nammalwar, Baskar; Bunce, Richard A; Berlin, K Darrell; Barrow, William W

    2014-02-25

    We are addressing bacterial resistance to antibiotics by repurposing a well-established classic antimicrobial target, the dihydrofolate reductase (DHFR) enzyme. In this work, we have focused on Enterococcus faecalis, a nosocomial pathogen that frequently harbors antibiotic resistance determinants leading to complicated and difficult-to-treat infections. An inhibitor series with a hydrophobic dihydrophthalazine heterocycle was designed from the anti-folate trimethoprim. We have examined the potency of this inhibitor series based on inhibition of DHFR enzyme activity and bacterial growth, including in the presence of the exogenous product analogue folinic acid. The resulting preferences were rationalized using a cocrystal structure of the DHFR from this organism with a propyl-bearing series member (RAB-propyl). In a companion apo structure, we identify four buried waters that act as placeholders for a conserved hydrogen-bonding network to the substrate and indicate an important role in protein stability during catalytic cycling. In these structures, the nicotinamide of the nicotinamide adenine dinucleotide phosphate cofactor is visualized outside of its binding pocket, which is exacerbated by RAB-propyl binding. Finally, homology models of the TMP(R) sequences dfrK and dfrF were constructed. While the dfrK-encoded protein shows clear sequence changes that would be detrimental to inhibitor binding, the dfrF-encoded protein model suggests the protein would be relatively unstable. These data suggest a utility for anti-DHFR compounds for treating infections arising from E. faecalis. They also highlight a role for water in stabilizing the DHFR substrate pocket and for competitive substrate inhibitors that may gain advantages in potency by the perturbation of cofactor dynamics.

  18. Land cover controls on depression-focused recharge: an example from southern Ontario

    NASA Astrophysics Data System (ADS)

    Buttle, J. M.; Greenwood, W. J.

    2015-12-01

    The Oak Ridges Moraine (ORM) is a critical hydrogeologic feature in southern Ontario. Although previous research has highlighted the implications of spatially-focused recharge in closed topographic depressions for regional groundwater resources, such depression-focused recharge (DFR) has not been empirically demonstrated on the ORM. Permeable surficial sands and gravels mantling much of the ORM imply that water fluxes will largely be vertical recharge rather than lateral downslope transfer into depressions. Nevertheless, lateral fluxes may occur in winter and spring, when concrete frost development encourages surface runoff of rainfall and snowmelt. The potential for DFR was examined under forest and agricultural land cover with similar soils and surficial geology. Soil water contents, soil temperatures and ground frost thickness were measured at the crest and base of closed depressions in two agricultural fields and two forest stands on permeable ORM outcrops. Recharge from late-fall to the end of spring snowmelt was estimated via 1-d water balances and surface-applied bromide tracing. Both forest and agricultural sites experienced soil freezing; however, greater soil water contents prior to freeze-up at the latter led to concrete soil frost development. This resulted in lateral movement of snowmelt and rainfall into topographic depressions and surface ponding, which did not occur in forest depressions. Water balance recharge exceeded estimates from the bromide tracer approach at all locations; nevertheless, both methods indicated DRF exceeded recharge at the depression crest in agricultural areas with little difference in forest areas. Water balance estimates suggest winter-spring DFR (1300 - 2000 mm) is 3-5× recharge on level agricultural sites. Differences in the potential for DFR between agricultural and forest land covers have important implications for the spatial variability of recharge fluxes and the quality of recharging water on the ORM.

  19. Retrieval of Snow and Rain From Combined X- and W-B and Airborne Radar Measurements

    NASA Technical Reports Server (NTRS)

    Liao, Liang; Meneghini, Robert; Tian, Lin; Heymsfield, Gerald M.

    2008-01-01

    Two independent airborne dual-wavelength techniques, based on nadir measurements of radar reflectivity factors and Doppler velocities, respectively, are investigated with respect to their capability of estimating microphysical properties of hydrometeors. The data used to investigate the methods are taken from the ER-2 Doppler radar (X-band) and Cloud Radar System (W-band) airborne Doppler radars during the Cirrus Regional Study of Tropical Anvils and Cirrus Layers-Florida Area Cirrus Experiment campaign in 2002. Validity is assessed by the degree to which the methods produce consistent retrievals of the microphysics. For deriving snow parameters, the reflectivity-based technique has a clear advantage over the Doppler-velocity-based approach because of the large dynamic range in the dual-frequency ratio (DFR) with respect to the median diameter Do and the fact that the difference in mean Doppler velocity at the two frequencies, i.e., the differential Doppler velocity (DDV), in snow is small relative to the measurement errors and is often not uniquely related to Do. The DFR and DDV can also be used to independently derive Do in rain. At W-band, the DFR-based algorithms are highly sensitive to attenuation from rain, cloud water, and water vapor. Thus, the retrieval algorithms depend on various assumptions regarding these components, whereas the DDV-based approach is unaffected by attenuation. In view of the difficulties and ambiguities associated with the attenuation correction at W-band, the DDV approach in rain is more straightforward and potentially more accurate than the DFR method.

  20. Prevalence and characterization of Salmonella species isolated from pigs, ducks and chickens in Sichuan Province, China.

    PubMed

    Li, Ruichao; Lai, Jing; Wang, Yang; Liu, Shuliang; Li, Yun; Liu, Kunyao; Shen, Jianzhong; Wu, Congming

    2013-04-15

    This study aimed to analyze the prevalence of Salmonella isolated from different parts of the food production chain, and to characterize these isolates. A total of 165 Salmonella enterica isolates were identified from 1382 samples taken from conventional farms, abattoirs and retail markets from 2010 to 2011 in Sichuan, China. The Salmonella isolates were assayed for serotype, antimicrobial susceptibility, prevalence of class 1 integrons and β-lactamase genes, and subtyped using pulsed-field gel electrophoresis. Among these isolates, S. enterica serotypes Derby (76 isolates, 46%) and Typhimurium (16 isolates, 10%) were the most prevalent, and high antimicrobial resistance rates were observed for tetracycline (77%), sulfamethoxazole/trimethoprim (43%), nalidixic acid (41%) and spectinomycin (41%). Class 1 integrons were detected in 21% of these isolates, and contained gene cassettes dfrA12-aadA2, dfrA1-aadA1, dfrA1, blaPSE-1 and dfrA1/aadA2. blaOXA-1 was the most commonly identified β-lactamase gene (n=14), followed by blaTEM-1 (n=6), blaPSE-1 (n=4) and blaCMY-2 (n=1). A S. enterica serotype Indiana isolate derived from chicken from a market was positive for both blaOXA-1 and blaCMY-2, and resistant to nine tested antibiotics. The PFGE patterns were diverse. Our findings indicated that most isolates from different sampling sites were phenotypically and genetically diverse, and Salmonella was widespread and may transmit along the food production chain from farm to market. Isolates with decreased susceptibility to fluoroquinolones and extended-spectrum cephalosporins, which are used to fight foodborne Salmonella, pose a serious threat to public health. PMID:23474653

  1. Gender in metropolitan development strategies: the case of Durban.

    PubMed

    Todes, A

    1995-10-01

    The author describes her experiences with the Durban Development Forum (DDF) and the strategies used in the Durban Functional Region (DFR) of South Africa. Policies are generally concerned with low income people, spatial distribution, and the organization of development and only partially concerned with gender issues. This paper discusses general urban development strategies in South Africa, gives an overview of the position of women in the DFR, and the strategies of the DDF. Prior to 1994 urban development plans focused on righting the inequalities and inefficiencies of apartheid cities, promotion of local economic development, and development implementation that was highly political and excluded major inputs from Black people. The DDF was formed in 1993 and aimed to be inclusive of all population groups in development planning. A set of principles were developed to guide development based on consensus decision making. Women's groups were included from the beginning, but little research was and is available on specific conditions and needs of different groups of women. Durban experienced since the 1980s a rise in the number of women in the labor force, a rise in female migrants to the DFR, and movement into "more insecure, poorly serviced newer settlements." Under apartheid, low income housing for Blacks was situated on peripheral areas or more distant locations that were away from employment in commerce and service industries in the central city. Women in urban slums survived with few sanitation amenities and limited access to an adequate water supply or electricity. Most development policy in the DFR ignored gender differences and gender power relations and focused on improving land, housing, and amenities for the poor in the central city. Although the general approach to improve infrastructure and facilities and to remove obstacles to development may have benefits for low income women, there are in fact mixed benefits for women. PMID:12291437

  2. Natural mutations in two homoeologous TT8 genes control yellow seed coat trait in allotetraploid Brassica juncea (AABB).

    PubMed

    Padmaja, Lakshmi K; Agarwal, Parul; Gupta, Vibha; Mukhopadhyay, Arundhati; Sodhi, Yaspal S; Pental, Deepak; Pradhan, Akshay K

    2014-02-01

    Identification of the candidate gene responsible for the seed coat colour variation in Brassica juncea was undertaken following an earlier study where two independent loci (BjSc1 and BjSc2) were mapped to two linkage groups, LG A9 and B3 (Padmaja et al. in Theor Appl Genet 111:8-14, 2005). The genome search from BRAD data for the presence of flavonoid genes in B. rapa identified three candidate genes namely, DFR, TT1 and TT8 in the LG A9. Quantitative real-time PCR revealed absence of transcript for the late biosynthetic genes (LBGs) and showed significant reduction of transcript in the TT8 from the developing seeds of yellow-seeded line. While mapping of two DFR genes, the BjuA.DFR and BjuB.DFR did not show perfect co-segregation with the seed coat colour loci, that of the two TT8 genes, BjuA.TT8 and BjuB.TT8 showed perfect co-segregation with the seed coat colour phenotype. The BjuA.TT8 allele from the yellow-seeded line revealed the presence of an insertion of 1,279 bp in the exon 7 and did not produce any transcript as revealed by reverse transcriptase PCR. The BjuB.TT8 allele from the yellow-seeded line revealed the presence of an SNP (C→T) in the exon 7 resulting in a stop codon predicting a truncated protein lacking the C-terminal 8 amino acid residues and produced significantly low level of transcript than its wild-type counterpart. Hence, it is hypothesized that the mutations in both the TT8 genes are required for inhibiting the transcription of LBGs in the yellow-seeded mutant of B. juncea. PMID:24247234

  3. Simulations of Dual-Frequency Radar Rainfall Retrievals

    NASA Astrophysics Data System (ADS)

    D'Adderio, Leo Pio; Tokay, Ali; Meneghini, Robert; Liao, Liang; Petersen, Walter A.; Porcù, Federico

    2016-04-01

    The retrieval of raindrop size distribution (DSD) is one of the key objectives of National Aeronautics and Space Administration (NASA) Global Precipitation Measurement (GPM) Mission. The dual-frequency precipitation radar (DPR) on board GPM core satellite is the primary resource for the retrieval of DSD. The DPR operates at Ku- and Ka-band and these frequencies have different sensitivities to the precipitation at the surface. Both frequencies are subject to the attenuation but at different magnitude. The high sensitivity of Ka-band measurements intends to detect solid and/or light liquid precipitation, while Ku-band frequency will be able to measure relatively higher intensity precipitation. The data from simultaneous Ka- and Ku-band measurements will allow a more accurate estimation of the DSD. The DSD retrieval algorithm uses three-parameter gamma distribution where mass weighted diameter (Dmass), normalized intercept parameter with respect to the liquid water content, and the shape parameter will be derived from dual-frequency radar measurements. A key problem is the retrieval of three unknown with two measurements. The simulation of the dual frequency ratio (DFR), using disdrometric data collected in different field campaigns of Ground Validation (GV) program of GPM mission, can cast light on this retrieval problem. Furthermore, the use of a third and/or different wavelength in the satellite measurements can be an added value to correctly retrieve both light and heavy rain. This study seeks relationship between the DFR and Dmass in different rain regimes. The DFR based both on Ka-/Ku-band and on frequencies other than Ka-/Ku-band is investigated. The dependence on the gamma distribution shape parameter, which is set to three in the DPR DSD retrieval algorithm, of the DFR-Dmass relationship is also analyzed.

  4. Incidence of antimicrobial-resistance genes and integrons in antibiotic-resistant bacteria isolated from eels and aquaculture ponds.

    PubMed

    Lin, Mao; Wu, Xiaomei; Yan, Qingpi; Ma, Ying; Huang, Lixing; Qin, Yingxue; Xu, Xiaojin

    2016-07-01

    The overuse of antimicrobials in aquaculture has promoted the selection of antimicrobial-resistant bacteria. Here we investigated the abundance of antimicrobial-resistance genes and integrons in 108 strains of antibiotic-resistant bacteria isolated from eels and aquaculture ponds in China. Conventional PCR was implemented to examine common antibiotic-resistance genes, integrons, and their gene cassette arrays. The results showed that the antibiotic-resistance genes blaTEM, tetC, sulI, aadA, floR, and qnrB were detected at high percentages, as were a number of other resistance genes. Class I integrons were present in 79.63% of the strains, and 10 out of 108 isolates carried class II integrons. Class III integrons were not detected. Three strains carried both class I and class II integrons, and 73.26% of the class I integron-positive isolates contained the qacEΔ1/sul1 gene. Fourteen types of integron cassette arrays were found among class I integron-positive isolates. A new array, dfrB4-catB3-blaOXA-10-aadA1, was discovered in this study. The gene cassette array dfrA12-orfF-aadA2 was the most widely distributed. In summary, 23 different gene cassettes encoding resistance to 8 classes of antibiotics were identified in the class I integrons, and the main cassettes contained genes encoding resistance to aminoglycosides (aad) and trimethoprim (dfr). All class II integron-positive strains had only a single gene cassette array, viz. dfrA1-catB2-sat2-aadA1. High levels of antimicrobial-resistance genes and integrons in eels and auqauculture ponds suggest that the overuse of antimicrobials should be strictly controlled and that the levels of bacterial antimicrobial-resistance genes in aquaculture should be monitored.

  5. [GENOTYPING OF THE BURKHOLDERIA MALLEI STRAINS BASED ON DIFFERENT REGION ANALYSIS].

    PubMed

    Bondareva, O S; Savchenko, S S; Tkachenko, G A; Ledeneva, M L; Lemasova, L V; Antonov, V A

    2016-01-01

    Development of the genotyping methods of glanders agent is urgent due to its high pathogenicity, lack of effective preventive measures and threat of the use of Burkholderia mallei as a biological weapon. In this work we proposed a scheme for the typing of the B. mallei strains based on different region analysis (DFR). The choice of variable loci differentially presented in various strains of glanders agents was performed by analyzing annotated whole-genome sequences of the B. mallei strains. Primers and fluorescence probes were designed for 9 selected loci. The amplification conditions for different regions were optimized in two variants: with electrophoretic detection and hybridization-fluorescence detection in the strip format. The possibility of applying the DFR analysis to genetic characterization of strains was assessed in 14 B. mallei strains. The genetic profiles of the studied B. mallei strains revealed that the developed DFR-typing scheme was characterized by high discrimination power (Hunter-Gaston index value was 0.92), reproducibility, rapidity, easy interpretation, and applicability for epidemiological surveillance of glanders.

  6. Characterization of the variable region in the class 1 integron of antimicrobial-resistant Escherichia coli isolated from surface water.

    PubMed

    Canal, Natália; Meneghetti, Karine Lena; de Almeida, Clara Ponzi; da Rosa Bastos, Marina; Otton, Letícia Muner; Corção, Gertrudes

    2016-01-01

    Fecal bacteria are considered to be a potential reservoir of antimicrobial resistance genes in the aquatic environment and could horizontally transfer these genes to autochthonous bacteria when carried on transferable and/or mobile genetic elements. Such circulation of resistance genes constitutes a latent public health hazard. The aim of this study was to characterize the variable region of the class 1 integron and relate its genetic content to resistance patterns observed in antimicrobial-resistant Escherichia coli isolated from the surface waters of Patos Lagoon, Southern Brazil. Genetic diversity of the isolates and presence of the qacEΔ1 gene, which confers resistance to quaternary ammonium compounds, were also investigated. A total of 27 isolates were analyzed. The variable region harbored dfrA17, dfrA1 and dfrA12 genes, which confer resistance to trimethoprim, and aadA1, aadA5 and aadA22 genes that encode resistance to streptomycin/spectinomycin. Most of the isolates were considered resistant to quaternary ammonium compounds and all of them carried the qacEΔ1 gene at the 3' conserved segment of the integron. ERIC-PCR analyses of E. coli isolates that presented the integrons showed great genetic diversity, indicating diverse sources of contamination in this environment. These results suggest that fecal bacteria with class 1 integrons in aquatic environments are potentially important reservoirs of antibiotic-resistance genes and may transfer these elements to other bacteria that are capable of infecting humans. PMID:26991286

  7. Utilization of Putrescine in Tobacco Cell Lines Resistant to Inhibitors of Polyamine Synthesis 1

    PubMed Central

    Hiatt, Andrew; Malmberg, Russell L.

    1988-01-01

    Three tobacco cell lines have been analyzed which are resistant to lethal inhibitors of either putrescine production or conversion of putrescine into polyamines. Free and conjugated putrescine pools, the enzymic activities (arginine, ornithine, and S-adenosylmethionine decarboxylases), and the growth characteristics during acidic stress were measured in suspension cultures of each cell line. One cell line, resistant to difluoromethylornithine (Dfr1) had a very low level of ornithine decarboxylase activity which was half insensitive to the inhibitor in vitro. Intracellular free putrescine in Dfr1 was elevated 10-fold which was apparently due to a 20-fold increase in the arginine decarboxylase activity. The increased free putrescine titer was not reflected in an increased level of spermidine, spermine, or putrescine conjugation. Dfr1 cultures survived acidic stress at molarities which were lethal to wild type cultures. Two other mutants, resistant to methylglyoxal bis(guanylhydrazone) (Mgr3, Mgr12), had near normal levels of the three decarboxylases and normal titers of free putrescine, spermidine, and spermine. Both mutants however had elevated levels of conjugated putrescine. Mgr12 had an increased sensitivity to acidic medium. These results suggest that increased levels of free putrescine production may enhance the ability of tobacco cells to survive acid stress. This was supported by the observation that cytotoxic effects of inhibiting arginine decarboxylase in wild type cell lines were dependent on the acidity of the medium. Images Fig. 1 PMID:16665927

  8. Integron-mediated Multidrug Resistance in a Global Collection of Nontyphoidal Salmonella enterica Isolates

    PubMed Central

    Krauland, Mary G.; Marsh, Jane W.; Paterson, David L.

    2009-01-01

    Salmonella enterica bacteria have become increasingly resistant to antimicrobial agents, partly as a result of genes carried on integrons. Clonal expansion and horizontal gene transfer may contribute to the spread of antimicrobial drug–resistance integrons in these organisms. We investigated this resistance and integron carriage among 90 isolates with the ACSSuT phenotype (resistance to ampicillin, chloramphenicol, streptomycin, sulfamethoxazole, and tetracycline) in a global collection of S. enterica isolates. Four integrons, dfrA12/orfF/aadA2, dfrA1/aadA1, dfrA7, and arr2/blaOXA30/cmlA5/aadA2, were found in genetically unrelated isolates from 8 countries on 4 continents, which supports a role for horizontal gene transfer in the global dissemination of S. enterica multidrug resistance. Serovar Typhimurium isolates containing identical integrons with the gene cassettes blaPSE1 and aadA2 were found in 4 countries on 3 continents, which supports the role of clonal expansion. This study demonstrates that clonal expansion and horizontal gene transfer contribute to the global dissemination of antimicrobial drug resistance in S. enterica. PMID:19239750

  9. [GENOTYPING OF THE BURKHOLDERIA MALLEI STRAINS BASED ON DIFFERENT REGION ANALYSIS].

    PubMed

    Bondareva, O S; Savchenko, S S; Tkachenko, G A; Ledeneva, M L; Lemasova, L V; Antonov, V A

    2016-01-01

    Development of the genotyping methods of glanders agent is urgent due to its high pathogenicity, lack of effective preventive measures and threat of the use of Burkholderia mallei as a biological weapon. In this work we proposed a scheme for the typing of the B. mallei strains based on different region analysis (DFR). The choice of variable loci differentially presented in various strains of glanders agents was performed by analyzing annotated whole-genome sequences of the B. mallei strains. Primers and fluorescence probes were designed for 9 selected loci. The amplification conditions for different regions were optimized in two variants: with electrophoretic detection and hybridization-fluorescence detection in the strip format. The possibility of applying the DFR analysis to genetic characterization of strains was assessed in 14 B. mallei strains. The genetic profiles of the studied B. mallei strains revealed that the developed DFR-typing scheme was characterized by high discrimination power (Hunter-Gaston index value was 0.92), reproducibility, rapidity, easy interpretation, and applicability for epidemiological surveillance of glanders. PMID:27183720

  10. Phytochrome-interacting factors PIF4 and PIF5 negatively regulate anthocyanin biosynthesis under red light in Arabidopsis seedlings.

    PubMed

    Liu, Zhongjuan; Zhang, Yongqiang; Wang, Jianfeng; Li, Ping; Zhao, Chengzhou; Chen, Yadi; Bi, Yurong

    2015-09-01

    Light is an important environmental factor inducing anthocyanin accumulation in plants. Phytochrome-interacting factors (PIFs) have been shown to be a family of bHLH transcription factors involved in light signaling in Arabidopsis. Red light effectively increased anthocyanin accumulation in wild-type Col-0, whereas the effects were enhanced in pif4 and pif5 mutants but impaired in overexpression lines PIF4OX and PIF5OX, indicating that PIF4 and PIF5 are both negative regulators for red light-induced anthocyanin accumulation. Consistently, transcript levels of several genes involved in anthocyanin biosynthesis and regulatory pathway, including CHS, F3'H, DFR, LDOX, PAP1 and TT8, were significantly enhanced in mutants pif4 and pif5 but decreased in PIF4OX and PIF5OX compared to in Col-0, indicating that PIF4 and PIF5 are transcriptional repressor of these gene. Transient expression assays revealed that PIF4 and PIF5 could repress red light-induced promoter activities of F3'H and DFR in Arabidopsis protoplasts. Furthermore, chromatin immunoprecipitation-quantitative PCR (ChIP-qPCR) test and electrophoretic mobility shift assay (EMSA) showed that PIF5 could directly bind to G-box motifs present in the promoter of DFR. Taken together, these results suggest that PIF4 and PIF5 negatively regulate red light-induced anthocyanin accumulation through transcriptional repression of the anthocyanin biosynthetic genes in Arabidopsis. PMID:26259175

  11. Characterization of the variable region in the class 1 integron of antimicrobial-resistant Escherichia coli isolated from surface water.

    PubMed

    Canal, Natália; Meneghetti, Karine Lena; de Almeida, Clara Ponzi; da Rosa Bastos, Marina; Otton, Letícia Muner; Corção, Gertrudes

    2016-01-01

    Fecal bacteria are considered to be a potential reservoir of antimicrobial resistance genes in the aquatic environment and could horizontally transfer these genes to autochthonous bacteria when carried on transferable and/or mobile genetic elements. Such circulation of resistance genes constitutes a latent public health hazard. The aim of this study was to characterize the variable region of the class 1 integron and relate its genetic content to resistance patterns observed in antimicrobial-resistant Escherichia coli isolated from the surface waters of Patos Lagoon, Southern Brazil. Genetic diversity of the isolates and presence of the qacEΔ1 gene, which confers resistance to quaternary ammonium compounds, were also investigated. A total of 27 isolates were analyzed. The variable region harbored dfrA17, dfrA1 and dfrA12 genes, which confer resistance to trimethoprim, and aadA1, aadA5 and aadA22 genes that encode resistance to streptomycin/spectinomycin. Most of the isolates were considered resistant to quaternary ammonium compounds and all of them carried the qacEΔ1 gene at the 3' conserved segment of the integron. ERIC-PCR analyses of E. coli isolates that presented the integrons showed great genetic diversity, indicating diverse sources of contamination in this environment. These results suggest that fecal bacteria with class 1 integrons in aquatic environments are potentially important reservoirs of antibiotic-resistance genes and may transfer these elements to other bacteria that are capable of infecting humans.

  12. Phytochrome-interacting factors PIF4 and PIF5 negatively regulate anthocyanin biosynthesis under red light in Arabidopsis seedlings.

    PubMed

    Liu, Zhongjuan; Zhang, Yongqiang; Wang, Jianfeng; Li, Ping; Zhao, Chengzhou; Chen, Yadi; Bi, Yurong

    2015-09-01

    Light is an important environmental factor inducing anthocyanin accumulation in plants. Phytochrome-interacting factors (PIFs) have been shown to be a family of bHLH transcription factors involved in light signaling in Arabidopsis. Red light effectively increased anthocyanin accumulation in wild-type Col-0, whereas the effects were enhanced in pif4 and pif5 mutants but impaired in overexpression lines PIF4OX and PIF5OX, indicating that PIF4 and PIF5 are both negative regulators for red light-induced anthocyanin accumulation. Consistently, transcript levels of several genes involved in anthocyanin biosynthesis and regulatory pathway, including CHS, F3'H, DFR, LDOX, PAP1 and TT8, were significantly enhanced in mutants pif4 and pif5 but decreased in PIF4OX and PIF5OX compared to in Col-0, indicating that PIF4 and PIF5 are transcriptional repressor of these gene. Transient expression assays revealed that PIF4 and PIF5 could repress red light-induced promoter activities of F3'H and DFR in Arabidopsis protoplasts. Furthermore, chromatin immunoprecipitation-quantitative PCR (ChIP-qPCR) test and electrophoretic mobility shift assay (EMSA) showed that PIF5 could directly bind to G-box motifs present in the promoter of DFR. Taken together, these results suggest that PIF4 and PIF5 negatively regulate red light-induced anthocyanin accumulation through transcriptional repression of the anthocyanin biosynthetic genes in Arabidopsis.

  13. Dietary intake of flavonoids and oesophageal and gastric cancer: incidence and survival in the United States of America (USA)

    PubMed Central

    Petrick, J L; Steck, S E; Bradshaw, P T; Trivers, K F; Abrahamson, P E; Engel, L S; He, K; Chow, W-H; Mayne, S T; Risch, H A; Vaughan, T L; Gammon, M D

    2015-01-01

    Background: Flavonoids, polyphenolic compounds concentrated in fruits and vegetables, have experimentally demonstrated chemopreventive effects against oesophageal and gastric cancer. Few epidemiologic studies have examined flavonoid intake and incidence of these cancers, and none have considered survival. Methods: In this USA multicentre population-based study, case participants (diagnosed during 1993–1995 with oesophageal adenocarcinoma (OEA, n=274), gastric cardia adenocarcinoma (GCA, n=248), oesophageal squamous cell carcinoma (OES, n=191), and other gastric adenocarcinoma (OGA, n=341)) and frequency-matched controls (n=662) were interviewed. Food frequency questionnaire responses were linked with USDA Flavonoid Databases and available literature for six flavonoid classes and lignans. Case participants were followed until 2000 for vital status. Multivariable-adjusted odds ratios (ORs) and hazard ratios (HRs) (95% confidence intervals (CIs)) were estimated, comparing highest with lowest intake quartiles, using polytomous logistic and proportional hazards regressions, respectively. Results: Little or no consistent association was found for total flavonoid intake (main population sources: black tea, orange/grapefruit juice, and wine) and incidence or survival for any tumour type. Intake of anthocyanidins, common in wine and fruit juice, was associated with a 57% reduction in the risk of incident OEA (OR=0.43, 95% CI=0.29–0.66) and OES (OR=0.43, 95% CI=0.26–0.70). The ORs for isoflavones, for which coffee was the main source, were increased for all tumours, except OES. Anthocyanidins were associated with decreased risk of mortality for GCA (HR=0.63, 95% CI=0.42–0.95) and modestly for OEA (HR=0.87, 95% CI=0.60–1.26), but CIs were wide. Conclusions: Our findings, if confirmed, suggest that increased dietary anthocyanidin intake may reduce incidence and improve survival for these cancers. PMID:25668011

  14. Antimicrobial Resources for Disinfection of Potable Water Systems for Future Spacecraft

    NASA Technical Reports Server (NTRS)

    Morford, Megan A.; Birmele, Michele; Roberts, Michael S.

    2012-01-01

    As human exploration adventures beyond low earth orbit, life support systems will require more innovation and research to become self-sustaining and durable. One major concern about future space travel is the ability to store and decontaminate water for consumption and hygiene. This project explores materials and technologies for possible use in future water systems without requiring point-of-use (POU) filtering or chemical additives such as iodine or silver that require multiple doses to remain effective. This experimentation tested the efficacy of a variety of antimicrobial materials against biofilm formation in a high shear CDC Biofilm Reactor (CBR) and some materials in a low shear Drip Flow Reactor (DFR) which(also utilizes ultra violet light emitting diodes (UVLEDs) as an antimicrobial resource. Most materials were tested in the CBR using the ASTM E 2562-07 1method involving the Pseudomonas aeruginosa and coupon samples that vary in their antimicrobial coatings and surface layer topographies. In a controlled environmental chamber (CEC), the CBR underwent a batch phase, continuous flow phase (CFP), and a harvest before analysis. The DFR portion of this experimentation was performed in order to assess the antimicrobial capabilities of ultraviolet-A LEDs (UV-A) in potable water systems. The ASTM E 2647-08 was modified in order to incorporate UV-A LEDs and to operate as a closed, re-circulating system. The modified DFR apparatus that was utilized contains 4 separate channels each of which contain 2 UV-A LEDs (1 chamber is masked off to serve as a control) and each channel is equipped with its own reservoir and peristaltic pump head. The 10 DFR runs discussed in this report include 4 initial experimental runs that contained blank microscope slides to test the UVA LEDs alone, 2 that incorporated solid silver coupons, 2 that utilized titanium dioxide (Ti02) coupons as a photocatalyst, and 2 runs that utilized silver coated acrylic slides. Both the CBR and DFR

  15. Analysis for prevalence and physical linkages amongst integrons, ISEcp1, ISCR1, Tn21 and Tn7 encountered in Escherichia coli strains from hospitalized and non-hospitalized patients in Kenya during a 19-year period (1992–2011)

    PubMed Central

    2013-01-01

    Background We determined the prevalence and evidence for physical linkage amongst integrons, insertion sequences, Tn21 and Tn7 transposons in a collection of 1327 E. coli obtained over a 19-year period from patients in Kenya. Results The prevalence of class 1 integrons was 35%, class 2 integrons were detected in 3 isolates but no isolate contained a class 3 integron. Integron lacking the 3’-CS or those linked to sul3 gene or IS26 or those containing the ISCR1 were only detected in multidrug resistant (MDR) strains. The dfrAs were the most common cassettes and their prevalence was: - dfrA1(28%), dfrA12(20%), dfA17(9%), dfrA7(9%), and dfrA16(5%). The aadA were the second most abundant cassettes and their prevalence was: - aadA1(25%), aadA2(21%), and aadA5(14%). Other cassettes occurred in lower prevalence of below 5%. Prevalence of Tn21, ISEcp1, ISCR1 and IS26 was 22%, 10%, 15%, and 7% respectively. Majority of Tn21 containing integrons carried a complete set of transposition genes while class 2 integrons were borne on Tn7 transposon. The qnrA genes were detected in 34(3%) isolates while 19(1%) carried qnrB. All qnr genes were in MDR strains carrying integrons containing the ISCR1. Close to 88% of blaTEM-52 were linked to IS26 while ≥ 80% of blaCTX-Ms and blaCMYs were linked to ISEcp1. Only a few studies have identified a blaCTX-M-9 containing an ISEcp1 element as reported in this study. Multiple genetic elements, especially those borne on incIl, incFII, and incL/M plasmids, and their associated resistance genes were transferrable en bloc to E. coli strain J53 in mating experiments. Conclusions This is the first detailed study on the prevalence of selected elements implicated in evolution of resistance determinants in a large collection of clinical E. coli in Africa. Proliferation of such strains carrying multiple resistance elements is likely to compromise the use of affordable and available treatment options for majority of poor patients in Africa. There is

  16. Abrasive stripping square-wave voltammetry of blackberry, raspberry, strawberry, pomegranate, and sweet and blue potatoes.

    PubMed

    Komorsky-Lovrić, Šebojka; Novak, Ivana

    2011-08-01

    Electro-oxidation potentials of 7 fruits and vegetables were determined by abrasive stripping voltammetry. The responses were characterized by 2 peaks with maxima at 0.45 and 0.55 V compared with Ag/AgCl, respectively. Both electrode reactions appear reversible at a frequency of 8 Hz. They can be ascribed to anthocyanidins and ellagic acid as electroactive compounds. By this method, an antioxidative capacity of a certain plant can be quickly estimated without extraction of active components. PMID:22417490

  17. Blotting Assisted by Heating and Solvent Extraction for DESI-MS Imaging

    NASA Astrophysics Data System (ADS)

    Cabral, Elaine C.; Mirabelli, Mario F.; Perez, Consuelo J.; Ifa, Demian R.

    2013-06-01

    Imprints of potato sprout ( Solanum tuberosum L.), gingko leaves (Gingko biloba L. ) and strawberries (Fragaria x ananassa Duch. ) were successfully imaged by desorption electrospray ionization mass spectrometry (DESI-MS) on TLC plates through blotting assisted by heating and/or solvent extraction. Ion images showing the distribution of significant compounds such as glycoalkaloid toxins in potato sprout, ginkgolic acids and flavonoids in ginkgo leaves, and sugars and anthocyanidin in strawberry were obtained. Practical implications of this work include analysis of a wide range of irregular or soft materials by different imprinting conditions without requiring the addition of matrices or use of specific kinds of surfaces.

  18. Blotting assisted by heating and solvent extraction for DESI-MS imaging.

    PubMed

    Cabral, Elaine C; Mirabelli, Mario F; Perez, Consuelo J; Ifa, Demian R

    2013-06-01

    Imprints of potato sprout (Solanum tuberosum L.), gingko leaves (Gingko biloba L.) and strawberries (Fragaria x ananassa Duch.) were successfully imaged by desorption electrospray ionization mass spectrometry (DESI-MS) on TLC plates through blotting assisted by heating and/or solvent extraction. Ion images showing the distribution of significant compounds such as glycoalkaloid toxins in potato sprout, ginkgolic acids and flavonoids in ginkgo leaves, and sugars and anthocyanidin in strawberry were obtained. Practical implications of this work include analysis of a wide range of irregular or soft materials by different imprinting conditions without requiring the addition of matrices or use of specific kinds of surfaces. PMID:23605686

  19. Flavonoids: prospective drug candidates.

    PubMed

    Cazarolli, Luisa Helena; Zanatta, Leila; Alberton, Elga Heloisa; Figueiredo, Maria Santos Reis Bonorino; Folador, Poliane; Damazio, Rosangela Guollo; Pizzolatti, Moacir Geraldo; Silva, Fátima Regina Mena Barreto

    2008-11-01

    The purpose of this review is to discuss the recent developments related to the chemistry and medicinal properties of flavonoids. Major flavonoids that show well categorized structures and well defined structure function-relationships are: flavans, flavanones, flavones, flavanonols, flavonols, catechins, anthocyanidins and isoflavone. The biological properties of flavonoids include antioxidant, anti-inflamatory, antitumoral, antiviral and antibacterial, as well as a direct cytoprotective effect on coronary and vascular systems, the pancreas and the liver. These characteristics place them among the most attractive natural substances available to enrich the current therapy options.

  20. Blotting assisted by heating and solvent extraction for DESI-MS imaging.

    PubMed

    Cabral, Elaine C; Mirabelli, Mario F; Perez, Consuelo J; Ifa, Demian R

    2013-06-01

    Imprints of potato sprout (Solanum tuberosum L.), gingko leaves (Gingko biloba L.) and strawberries (Fragaria x ananassa Duch.) were successfully imaged by desorption electrospray ionization mass spectrometry (DESI-MS) on TLC plates through blotting assisted by heating and/or solvent extraction. Ion images showing the distribution of significant compounds such as glycoalkaloid toxins in potato sprout, ginkgolic acids and flavonoids in ginkgo leaves, and sugars and anthocyanidin in strawberry were obtained. Practical implications of this work include analysis of a wide range of irregular or soft materials by different imprinting conditions without requiring the addition of matrices or use of specific kinds of surfaces.

  1. Possible effects of dietary polyphenols on sugar absorption and digestion.

    PubMed

    Williamson, Gary

    2013-01-01

    Excessive post-prandial glucose excursions are a risk factor for developing diabetes, associated with impaired glucose tolerance. One way to limit the excursion is to inhibit the activity of digestive enzymes for glucose production and of the transporters responsible for glucose absorption. Flavonols, theaflavins, gallate esters, 5-caffeoylqunic acid and proanthocyanidins inhibit α-amylase activity. Anthocyanidins and catechin oxidation products, such as theaflavins and theasinsensins, inhibit maltase; sucrase is less strongly inhibited but anthocyanidins seem somewhat effective. Lactase is inhibited by green tea catechins. Once produced in the gut by digestion, glucose is absorbed by SGLT1 and GLUT2 transporters, inhibited by flavonols and flavonol glycosides, phlorizin and green tea catechins. These in vitro data are supported by oral glucose tolerance tests on animals, and by a limited number of human intervention studies on polyphenol-rich foods. Acarbose is a drug whose mechanism of action is only through inhibition of α-amylases and α-glucosidases, and in intervention studies gives a 6% reduction in diabetes risk over 3 years. A lifetime intake of dietary polyphenols, assuming the same mechanism, has therefore a comparable potential to reduce diabetes risk, but more in vivo studies are required to fully test the effect of modulating post-prandial blood glucose in humans.

  2. Hydrogen-rich water reestablishes ROS homeostasis but exerts differential effects on anthocyanin synthesis in two varieties of radish sprouts under UV-A irradiation.

    PubMed

    Su, Nana; Wu, Qi; Liu, Yuanyuan; Cai, Jiangtao; Shen, Wenbiao; Xia, Kai; Cui, Jin

    2014-07-01

    The aims of the study were to investigate whether hydrogen gas (H2) was involved in regulation of anthocyanin biosynthesis in two contrasting radish (Raphanus sativus L.) varieties (low [LA] and high [HA] level of anthocyanin) under UV irradiation. The results showed that hydrogen-rich water (HRW) significantly blocked the UV-A-induced increase of H2O2 and O2(•-) accumulation, and enhanced the UV-A-induced increase of superoxide dismutase (SOD) and ascorbate peroxidase (APX) activities in LA and HA. Furthermore, UV-A-induced increase of anthocyanin and total phenols was further enhanced only in HA sprouts cotreated with HRW. LC-MS/MS analysis showed that five anthocyanidins existed in HA sprouts, but only two in LA sprouts. Meanwhile, the cyanidin was the most abundant anthocyanidin in HA, and the cyanidin was 2-fold higher cotreated with HRW than UV-A. Molecular analyses showed that the anthocyanin biosynthesis-related genes were upregulated significantly in both HA (in particular) and LA sprouts treated with HRW plus UV-A. These data imply that HRW reestablishes reactive oxygen species homeostasis in both LA and HA, but exerts different effects on anthocyanin accumulation between them under UV-A.

  3. Potential natural sensitizers extracted from the skin of Canarium odontophyllum fruits for dye-sensitized solar cells

    NASA Astrophysics Data System (ADS)

    Lim, Andery; Kumara, N. T. R. N.; Tan, Ai Ling; Mirza, Aminul Huq; Chandrakanthi, R. L. N.; Petra, Mohammad Iskandar; Ming, Lim Chee; Senadeera, G. K. R.; Ekanayake, Piyasiri

    2015-03-01

    Possibility of use of dye extract from skin samples of a seasonal, indigenous fruit from Borneo, namely Canarium odontophyllum, in dye sensitized solar cells (DSSCs) are explored. Three main groups of flavonoid pigments are detected and these pigments exhibit different UV-vis absorption properties, and hence showing different light harvesting capabilities. When applied in DSSCs. The detected pigment constituents of the extract consist of aurone (maritimein), anthocyanidin (pelargonidin) and anthocyanidin (cyanidin derivatives). When tested in DSSC, the highest conversion efficiency of 1.43% is exhibited by cyanidin derivatives, and this is followed by conversion efficiencies of 0.51% and 0.79% for aurone and pelargonidin, respectively. It is shown that individual pigments, like cyanidin derivatives and pelargonidin, exhibit higher power conversion efficiency when compared to that of C.odontophyllum skin pigment mixture (with a conversion efficiency of only 0.68%). The results indicate a possibility of masking effects of the pigments when used as a mixture. The acidification of C.odontophyllum skin pigments with concentrated hydrochloric acid improves the conversion efficiency of the mixture from 0.68% to 0.99%. The discussion in this paper will draw data and observations from the variation in absorption and adsorption properties, the HOMO-LUMO levels, the energy band gaps and the functional group compositions of the detected flavonoids.

  4. Phytochemical composition and biological activity of 8 varieties of radish (Raphanus sativus L.) sprouts and mature taproots.

    PubMed

    Hanlon, Paul R; Barnes, David M

    2011-01-01

    Radishes (Raphanus sativus L.) are members of the cruciferous vegetable family that contain many classes of biologically active phytochemicals. This study determined the phytochemical composition of the sprouts and mature taproots of 8 radish varieties. Radish sprouts contained significantly greater concentrations of glucosinolates (3.8-fold) and isothiocyanates (8.2-fold) than the mature radish taproot and also contained significantly greater concentrations of phenolics (on average 6.9-fold). The anthocyanin concentrations of the mature radish taproot were significantly greater than in the sprouts of red, pink, and purple varieties. The primary anthocyanidins present in the red and pink radish varieties were pelargonidin and delphinidin, while the primary anthocyanidin in the purple radish variety was cyanidin. Radish sprouts were between 9- and 59-fold more potent than the corresponding mature taproot at activating the antioxidant response element (ARE) in a stably transfected hepatoma cell line. The ARE activity of the radish sprouts and mature taproots was significantly correlated with the total isothiocyanate concentration of the radishes. Practical Application: Understanding the influence variety and developmental stage has on the biological activity of cruciferous vegetables provides important information for further studies examining the in vivo effects of radish treatment and foundation for providing recommendations to reduce the risk of chronic disease through dietary intervention.

  5. Molecular structures of the stem tuber anthocyanins of colored potatoes and their coloring effects on the tubers.

    PubMed

    Zhao, Chang Ling; Wen, Guo Song; Mao, Zi Chao; Xu, Shao Zhong; Liu, Zheng Jie; Zhao, Ming Fu; Lin, Chun

    2015-03-01

    This paper summarized the important achievements about the general characteristics of the molecular structures of the stem tuber anthocyanins of Colored potatoes and the basic coloring effects of the anthocyanins on the tubers. The various coloration patterns of the skins and/or flesh of Colored potato tubers result from the accumulation of the anthocyanins in the periderms, phelloderms and/or peripheral cortices of the tubers, and the tuber colors are fundamentally determined by the matching profiles of the six naturally occurring anthocyanidins, i.e., cyanidin, delphinidin, malvidin, pelargonidin, peonidin and petunidin. Generally, the tuber anthocyanidins hold an O-glycosidic bond-linked rutinosyl at the C3 site, and either a glucosyl linked by an O-glycosidic bond or no substituent group may exist at the C5 site simultaneously. Furthermore, an E-monoacyl frequently exists at the C3-rutinosyls or at the C5-glucosyls of most tuber anthocyanins, and the phenolic acids acylating the tuber anthocyanins are often p-coumaric, ferulic and caffeic acids. The popular names of the p- coumaric acid derivatives of the malvidin, pelargonidin, peonidin and petunidin of the tubers are Malvanin, Pelanin, Peonanin and Petanin, respectively. This review provides a reference for the exploration of the mechanism of the tuber coloration and the identification of the molecular structures of the stem tuber anthocyanins of Colored potatoes.

  6. Two glycosyltransferases involved in anthocyanin modification delineated by transcriptome independent component analysis in Arabidopsis thaliana

    PubMed Central

    Yonekura-Sakakibara, Keiko; Fukushima, Atsushi; Nakabayashi, Ryo; Hanada, Kousuke; Matsuda, Fumio; Sugawara, Satoko; Inoue, Eri; Kuromori, Takashi; Ito, Takuya; Shinozaki, Kazuo; Wangwattana, Bunyapa; Yamazaki, Mami; Saito, Kazuki

    2012-01-01

    To identify candidate genes involved in Arabidopsis flavonoid biosynthesis, we applied transcriptome coexpression analysis and independent component analyses with 1388 microarray data from publicly available databases. Two glycosyltransferases, UGT79B1 and UGT84A2 were found to cluster with anthocyanin biosynthetic genes. Anthocyanin was drastically reduced in ugt79b1 knockout mutants. Recombinant UGT79B1 protein converted cyanidin 3-O-glucoside to cyanidin 3-O-xylosyl(1→2)glucoside. UGT79B1 recognized 3-O-glucosylated anthocyanidins/flavonols and uridine diphosphate (UDP)-xylose, but not 3,5-O-diglucosylated anthocyanidins, indicating that UGT79B1 encodes anthocyanin 3-O-glucoside: 2′′-O-xylosyltransferase. UGT84A2 is known to encode sinapic acid: UDP-glucosyltransferase. In ugt84a2 knockout mutants, a major sinapoylated anthocyanin was drastically reduced. A comparison of anthocyanin profiles in ugt84a knockout mutants indicated that UGT84A2 plays a major role in sinapoylation of anthocyanin, and that other UGT84As contribute the production of 1-O-sinapoylglucose to a lesser extent. These data suggest major routes from cyanidin 3-O-glucoside to the most highly modified cyanidin in the potential intricate anthocyanin modification pathways in Arabidopsis. PMID:21899608

  7. Potential natural sensitizers extracted from the skin of Canarium odontophyllum fruits for dye-sensitized solar cells.

    PubMed

    Lim, Andery; Kumara, N T R N; Tan, Ai Ling; Mirza, Aminul Huq; Chandrakanthi, R L N; Petra, Mohammad Iskandar; Ming, Lim Chee; Senadeera, G K R; Ekanayake, Piyasiri

    2015-03-01

    Possibility of use of dye extract from skin samples of a seasonal, indigenous fruit from Borneo, namely Canarium odontophyllum, in dye sensitized solar cells (DSSCs) are explored. Three main groups of flavonoid pigments are detected and these pigments exhibit different UV-vis absorption properties, and hence showing different light harvesting capabilities. When applied in DSSCs. The detected pigment constituents of the extract consist of aurone (maritimein), anthocyanidin (pelargonidin) and anthocyanidin (cyanidin derivatives). When tested in DSSC, the highest conversion efficiency of 1.43% is exhibited by cyanidin derivatives, and this is followed by conversion efficiencies of 0.51% and 0.79% for aurone and pelargonidin, respectively. It is shown that individual pigments, like cyanidin derivatives and pelargonidin, exhibit higher power conversion efficiency when compared to that of C.odontophyllum skin pigment mixture (with a conversion efficiency of only 0.68%). The results indicate a possibility of masking effects of the pigments when used as a mixture. The acidification of C.odontophyllum skin pigments with concentrated hydrochloric acid improves the conversion efficiency of the mixture from 0.68% to 0.99%. The discussion in this paper will draw data and observations from the variation in absorption and adsorption properties, the HOMO-LUMO levels, the energy band gaps and the functional group compositions of the detected flavonoids.

  8. Anthocyanin biosynthesis for cold and freezing stress tolerance and desirable color in Brassica rapa.

    PubMed

    Ahmed, Nasar Uddin; Park, Jong-In; Jung, Hee-Jeong; Hur, Yoonkang; Nou, Ill-Sup

    2015-07-01

    Flavonoids are divided into several structural classes, including anthocyanins, which provide flower and leaf colors and other derivatives that play diverse roles in plant development and interactions with the environment. This study characterized four anthocyanidin synthase (ANS) genes of Brassica rapa, a structural gene of the anthocyanin biosynthetic pathway, and investigated their association with pigment formation, cold and freezing tolerance in B. rapa. Sequences of these genes were analyzed and compared with similar gene sequences from other species, and a high degree of homology with their respective functions was found. Organ-specific expression analysis revealed that these genes were only expressed in the colored portion of leaves of different lines of B. rapa. Conversely, B. rapa anthocyanidin synthase (BrANS) genes also showed responses to cold and freezing stress treatment in B. rapa. BrANSs were also shown to be regulated by two transcription factors, BrMYB2-2 and BrTT8, contrasting with anthocyanin accumulation and cold stress. Thus, the above results suggest the association of these genes with anthocyanin biosynthesis and cold and freezing stress tolerance and might be useful resources for development of cold-resistant Brassica crops with desirable colors as well.

  9. Anthocyanin biosynthesis for cold and freezing stress tolerance and desirable color in Brassica rapa.

    PubMed

    Ahmed, Nasar Uddin; Park, Jong-In; Jung, Hee-Jeong; Hur, Yoonkang; Nou, Ill-Sup

    2015-07-01

    Flavonoids are divided into several structural classes, including anthocyanins, which provide flower and leaf colors and other derivatives that play diverse roles in plant development and interactions with the environment. This study characterized four anthocyanidin synthase (ANS) genes of Brassica rapa, a structural gene of the anthocyanin biosynthetic pathway, and investigated their association with pigment formation, cold and freezing tolerance in B. rapa. Sequences of these genes were analyzed and compared with similar gene sequences from other species, and a high degree of homology with their respective functions was found. Organ-specific expression analysis revealed that these genes were only expressed in the colored portion of leaves of different lines of B. rapa. Conversely, B. rapa anthocyanidin synthase (BrANS) genes also showed responses to cold and freezing stress treatment in B. rapa. BrANSs were also shown to be regulated by two transcription factors, BrMYB2-2 and BrTT8, contrasting with anthocyanin accumulation and cold stress. Thus, the above results suggest the association of these genes with anthocyanin biosynthesis and cold and freezing stress tolerance and might be useful resources for development of cold-resistant Brassica crops with desirable colors as well. PMID:25504198

  10. Flavonoids, Flavonoid Subclasses, and Esophageal Cancer Risk: A Meta-Analysis of Epidemiologic Studies

    PubMed Central

    Cui, Lingling; Liu, Xinxin; Tian, Yalan; Xie, Chen; Li, Qianwen; Cui, Han; Sun, Changqing

    2016-01-01

    Flavonoids have been suggested to play a chemopreventive role in carcinogenesis. However, the epidemiologic studies assessing dietary intake of flavonoids and esophageal cancer risk have yielded inconsistent results. This study was designed to examine the association between flavonoids, each flavonoid subclass, and the risk of esophageal cancer with a meta-analysis approach. We searched for all relevant studies with a prospective cohort or case-control study design published from January 1990 to April 2016, using PUBMED, EMBASE, and Web of Science. Pooled odds ratios (ORs) were calculated using fixed or random-effect models. In total, seven articles including 2629 cases and 481,193 non-cases were selected for the meta-analysis. Comparing the highest-intake patients with the lowest-intake patients for total flavonoids and for each flavonoid subclass, we found that anthocyanidins (OR = 0.60, 95% CI: 0.49–0.74), flavanones (OR = 0.65, 95% CI: 0.49–0.86), and flavones (OR = 0.78, 95% CI 0.64–0.95) were inversely associated with the risk of esophageal cancer. However, total flavonoids showed marginal association with esophageal cancer risk (OR = 0.78, 95% CI: 0.59–1.04). In conclusion, our study suggested that dietary intake of total flavonoids, anthocyanidins, flavanones, and flavones might reduce the risk of esophageal cancer. PMID:27338463

  11. Purification and characterization of a betanidin glucosyltransferase from Amaranthus tricolor L catalyzing non-specific biotransformation of flavonoids.

    PubMed

    Das, Shibendu Sekhar; Gauri, Samiran S; Misra, Biswapriya Biswavas; Biswas, Mousumi; Dey, Satyahari

    2013-10-01

    Betacyanins are the major pigments present in Amaranthus tricolor, a leafy vegetable consumed globally. The terminal glycosylation of the aglycone betanidin is an important step in the biosynthesis of this natural red antioxidant pigment. A betanidin 5-O-glucosyltransferase (BGT) was fully purified to 134 folds (specific activity, 265.2 nkat mg(-1)) from the red amaranth by ammonium sulfate precipitation followed by hydrophobic interaction, anion exchange and size exclusion chromatography. Homogeneity of the purified protein was confirmed by 2-dimensional polyacrylamide gel electrophoresis (2D PAGE). The molecular weight of the enzyme determined by liquid chromatography-mass spectrometry (LC-MS) was found to be 62.8 kDa. Furthermore, the enzyme glycosylated flavonoids (kaempferol and quercetin) but not anthocyanidins, presence of which is mutually exclusive to betacyanin accumulating plants. The apparent Km (344±2.34 μM) and Vmax (17.24 μM min(-1)) of the enzyme were determined by LC-MS/MS. Peptide mass fingerprinting of the purified protein showed 38.4% coverage of peptide masses with anthocyanidin 3-O-glucosyltransferase from Zea mays. Study on this purified enzyme, for the first time, revealed its role of glycosylation in biosynthesis of betacyanin in A. tricolor and indicates promiscuous substrate-specificity.

  12. Molecular cloning and biochemical characterization of the UDP-glucose: flavonoid 3-O-glucosyltransferase from Concord grape (Vitis labrusca).

    PubMed

    Hall, Dawn; Yuan, Xiao Xin; Murata, Jun; De Luca, Vincenzo

    2012-02-01

    Glucosylation of anthocyanidin substrates at the 3-O-position is crucial for the red pigmentation of grape berries and wine. The gene that encodes the enzyme involved in this reaction has been cloned from Vitis labrusca cv. Concord, heterologously expressed, and the recombinant enzyme (rVL3GT) was characterized. VL3GT has 96% amino acid sequence identity with Vitis vinifera VV3GT and groups phylogenetically with several other flavonoid 3-O-glycosyltransferases. In vitro substrate specificity studies and kinetic analyses of rVL3GT indicate that this enzyme preferentially glucosylates cyanidin as compared with quercetin. Crude protein extracts from several Concord grape tissues were assayed for glucosyltransferase activity with cyanidin and quercetin as acceptor substrates. A comparison of the VL3GT activities toward with these substrates showed that the 3GT enzyme activity is consistent with the expression of VL3GT in these tissues and is coincident with the biosynthesis of anthocyanins in both location and developmental stages. Enzyme activities in grape mesocarp, pre-veraison exocarp, leaf, flower bud, and flower tissues glucosylated quercetin but not cyanidin at high rates, suggesting the presence of additional enzymes which are able to glucosylate the 3-O-position of flavonols with higher specificity than anthocyanidins.

  13. Characterization of trimethoprim-sulfamethoxazole resistance genes and their relatedness to class 1 integron and insertion sequence common region in gram-negative bacilli.

    PubMed

    Shin, Hae Won; Lim, Jinsook; Kim, Semi; Kim, Jimyung; Kwon, Gye Cheol; Koo, Sun Hoe

    2015-01-01

    Trimethoprim-sulfamethoxazole (TMP-SMX) has been used for the treatment of urinary tract infections, but increasing resistance to TMP-SMX has been reported. In this study, we analyzed TMP-SMX resistance genes and their relatedness with integrons and insertion sequence common regions (ISCRs) in uropathogenic gram-negative bacilli. Consecutive nonduplicate TMP-SMX nonsusceptible clinical isolates of E. coli, K. pneumoniae, Acinetobacter spp., and P. aeruginosa were collected from urine. The minimal inhibitory concentration was determined by Etest. TMP-SMX resistance genes (sul and dfr), integrons, and ISCRs were analyzed by PCR and sequencing. A total of 45 E. coli (37.8%), 15 K. pneumoniae (18.5%), 12 Acinetobacter spp. (70.6%), and 9 Pseudomonas aeruginosa (30.0%) isolates were found to be resistant to TMP-SMX. Their MICs were all over 640. In E. coli and K. pneumoniae, sul1 and dfr genes were highly prevalent in relation with integron1. The sul3 gene was detected in E. coli. However, in P. aeruginosa and Acinetobacter spp., only sul1 was prevalent in relation with class 1 integron; however, dfr was not detected and sul2 was less prevalent than in Enterobacteriaceae. ISCR1 and/or ISCR2 were detected in E. coli, K. pneumoniae, and Acinetobacter spp. but the relatedness with TMP-SMX resistance genes was not prominent. ISCR14 was detected in six isolates of E. coli. In conclusion, resistance mechanisms for TMP-SMX were different between Enterobacteriaceae and glucose non-fermenting gram-negative bacilli. Class 1 integron was widely disseminated in uropathogenic gram-negative baciili, so the adoption of prudent use of antimicrobial agents and the establishment of a surveillance system are needed.

  14. A study of air/space-borne dual-wavelength radar for estimation of rain profiles

    NASA Astrophysics Data System (ADS)

    Liang, Liao; Meneghini, Robert

    2005-11-01

    In this study, a framework is given by which air/space-borne dual-wavelength radar data can be used to estimate the characteristic parameters of hydrometeors. The focus of the study is on the Global Precipitation Measurement (GPM) precipitation radar, a dual-wavelength radar that will operate in the Ku (13.6 GHz) and Ka (35 GHz) bands. A key aspect of the retrievals is the relationship between the differential frequency ratio (DFR) and the median volume diameter, D0, and its dependence on the phase state of the hydrometeors. It is shown that parametric plots of D0 and particle concentration in the plane of the DFR and the radar reflectivity factor in the Ku band can be used to reduce the ambiguities in deriving D0 from DFR. A self-consistent iterative algorithm, which does not require the use of an independent pathattenuation constraint, is examined by applying it to the apparent radar reflectivity profiles simulated from a drop size distribution (DSD) model. For light to moderate rain, the self-consistent rain profiling approach converges to the correct solution only if the same shape factor of the Gamma distributions is used both to generate and retrieve the rain profiles. On the other hand, if the shape factors differ, the iteration generally converges but not to the correct solution. To further examine the dual-wavelength techniques, the selfconsistent iterative algorithm, along with forward and backward rain profiling algorithms, are applied to measurements taken from the 2nd generation Precipitation Radar (PR-2) built by the Jet Propulsion Laboratory. Consistent with the model results, it is found that the estimated rain profiles are sensitive to the shape factor of the size distribution when the iterative, self-consistent approach is used but relatively insensitive to this parameter when the forward- and backward-constrained approaches are used.

  15. Occurrence of antibiotic resistance and characterization of resistant genes and integrons in Enterobacteriaceae isolated from integrated fish farms south China

    USGS Publications Warehouse

    Su, Hao-Chang; Ying, Guang-Guo; Tao, Ran; Zhang, Rui-Quan; Fogarty, Lisa R.; Kolpin, Dana W.

    2011-01-01

    Antibiotics are still widely applied in animal husbandry to prevent diseases and used as feed additives to promote animal growth. This could result in antibiotic resistance to bacteria and antibiotic residues in animals. In this paper, Enterobacteriaceae isolated from four integrated fish farms in Zhongshan, South China were tested for antibiotic resistance, tetracycline resistance genes, sulfonamide resistance genes, and class 1 integrons. The Kirby-Bauer disk diffusion method and polymerase chain reaction (PCR) assays were carried out to test antibiotic susceptibility and resistance genes, respectively. Relatively high antibiotic resistance frequencies were found, especially for ampicillin (80%), tetracycline (52%), and trimethoprim (50%). Out of 203 Enterobacteriaceae isolates, 98.5% were resistant to one or more antibiotics tested. Multiple antibiotic resistance (MAR) was found highest in animal manures with a MAR index of 0.56. Tetracycline resistance genes (tet(A), tet(C)) and sulfonamide resistance genes (sul2) were detected in more than 50% of the isolates. The intI1 gene was found in 170 isolates (83.7%). Both classic and non-classic class 1 integrons were found. Four genes, aadA5, aadA22, dfr2, and dfrA17, were detected. To our knowledge, this is the first report for molecular characterization of antibiotic resistance genes in Enterobacteriaceae isolated from integrated fish farms in China and the first time that gene cassette array dfrA17-aadA5 has been detected in such fish farms. Results of this study indicated that fish farms may be a reservoir of highly diverse and abundant antibiotic resistant genes and gene cassettes. Integrons may play a key role in multiple antibiotic resistances posing potential health risks to the general public and aquaculture.

  16. Cs-miR156 is involved in the nitrogen form regulation of catechins accumulation in tea plant (Camellia sinensis L.).

    PubMed

    Fan, Kai; Fan, Dongmei; Ding, Zhaotang; Su, Yanhua; Wang, Xiaochang

    2015-12-01

    The nitrogen source affects the growth of tea plants and regulates the accumulation of catechins in the leaves. In this report, we assessed the influences of NH4(+) and NO3(-) on plant growth, catechins accumulation and associated gene expression. Compared with the preferential nitrogen source NH4(+), when NO3(-) was supplied as the sole nitrogen source, tea plants showed similar symptoms with the nitrogen-free treatments and showed lower nitrogen, free amino acid accumulation, chlorophyll content and biomass gain, indicating NO3(-) was not efficiently used by these plants. However, the total shoot catechins content was significantly higher for NO3(-) treatments than that for NH4(+) treatment or combined NH4(+)+NO3(-) treatment, suggesting that, in addition to its influence on plant growth, the nitrogen form regulated the accumulation of catechins in tea. The expression of catechins biosynthesis-related genes was associated with the regulation of catechins accumulation and composition changes mediated by nitrogen form. PAL, CHS, CHI, and DFR genes exhibited higher expression levels in plants supplied with NO3(-), in which the transcript level of DFR in the shoots was significantly correlated with the catechins content. In the end, we identified a new function for the Cs-miR156, which was drastically induced through NH4(+). Moreover, a potential mechanism of the Cs-miR156 pathway in regulating catechins biosynthesis in tea plants has been suggested, with particular respect to nitrogen forms. Cs-miR156 might repress the expression of the target gene SPL to regulate the DFR gene, which plays a vital role in catechins biosynthesis.

  17. Occurrence of antibiotic resistance and characterization of resistance genes and integrons in Enterobacteriaceae isolated from integrated fish farms in South China.

    PubMed

    Su, Hao-Chang; Ying, Guang-Guo; Tao, Ran; Zhang, Rui-Quan; Fogarty, Lisa Reynolds; Kolpin, Dana W

    2011-11-01

    Antibiotics are still widely applied in animal husbandry to prevent diseases and used as feed additives to promote animal growth. This could result in antibiotic resistance to bacteria and antibiotic residues in animals. In this paper, Enterobacteriaceae isolated from four integrated fish farms in Zhongshan, South China were tested for antibiotic resistance, tetracycline resistance genes, sulfonamide resistance genes, and class 1 integrons. The Kirby-Bauer disk diffusion method and polymerase chain reaction (PCR) assays were carried out to test antibiotic susceptibility and resistance genes, respectively. Relatively high antibiotic resistance frequencies were found, especially for ampicillin (80%), tetracycline (52%), and trimethoprim (50%). Out of 203 Enterobacteriaceae isolates, 98.5% were resistant to one or more antibiotics tested. Multiple antibiotic resistance (MAR) was found highest in animal manures with a MAR index of 0.56. Tetracycline resistance genes (tet(A), tet(C)) and sulfonamide resistance genes (sul2) were detected in more than 50% of the isolates. The intI1 gene was found in 170 isolates (83.7%). Both classic and non-classic class 1 integrons were found. Four genes, aadA5, aadA22, dfr2, and dfrA17, were detected. To our knowledge, this is the first report for molecular characterization of antibiotic resistance genes in Enterobacteriaceae isolated from integrated fish farms in China and the first time that gene cassette array dfrA17-aadA5 has been detected in such fish farms. Results of this study indicated that fish farms may be a reservoir of highly diverse and abundant antibiotic resistant genes and gene cassettes. Integrons may play a key role in multiple antibiotic resistances posing potential health risks to the general public and aquaculture.

  18. Capillary-driven microfluidic chips with evaporation-induced flow control and dielectrophoretic microbead trapping

    NASA Astrophysics Data System (ADS)

    Temiz, Yuksel; Skorucak, Jelena; Delamarche, Emmanuel

    2014-07-01

    This work reports our efforts on developing simple-to-use microfluidic devices for point-of-care diagnostic applications with recent extensions that include the trapping of microbeads using dielectrophoresis (DEP) and the modulation of the liquid flow using integrated microheaters. DEP serves the purpose of trapping microbeads coated with receptors and analytes for detection of a fluorescent signal. The microheater is actuated once the chip is filled by capillarity, creating an evaporation-induced flow tuned according to assay conditions. The chips are composed of a glass substrate patterned with 50-nm-thick Pd electrodes and microfluidic structures made using a 20-μm-thick dry-film resist (DFR). Chips are covered/sealed by low temperature (50°C) lamination of a 50-μm-thick DFR layer having excellent optical and mechanical properties. To separate cleaned and sealed chips from the wafer, we used an effective chip singulation technique which we informally call the "chip-olate" process. In the experimental section, we first studied dielectrophoretic trapping of 10-μm beads for flow rates ranging from 80 pL s-1 to 2.5 nL s-1 that are generated by an external syringe pump. Then, we characterized the embedded microheater in DFR-covered chips. Flow rates as high as 8 nL s-1 were generated by evaporation-induced flow when the heater was biased by 10 V, corresponding to 270-mW power. Finally, DEP-based trapping and fluorescent detection of functionalized beads were demonstrated as the flow was generated by evaporation-induced flow after the microfluidic structures were filled by capillarity.

  19. Capillary-driven microfluidic chips with evaporation-induced flow control and dielectrophoretic microbead trapping

    NASA Astrophysics Data System (ADS)

    Temiz, Yuksel; Skorucak, Jelena; Delamarche, Emmanuel

    2014-03-01

    This work reports our efforts on developing simple-to-use microfluidic devices for point-of-care diagnostic applications with recent extensions that include the trapping of microbeads using dielectrophoresis (DEP) and the modulation of capillary-driven flow using integrated microheaters. DEP serves the purpose of trapping microbeads coated with receptors and analytes for detection of a fluorescent signal. The microheater is actuated once the chip is filled by capillarity, creating an evaporation-induced flow tuned according to assay conditions. The chips are composed of a glass substrate patterned with 50-nm-thick Pd electrodes and microfluidic structures made using a 20-μm-thick dry-film resist (DFR). Chips are covered/sealed by low-temperature (50 °C) lamination of a 50-μm-thick DFR layer having excellent optical and mechanical properties. To separate cleaned and sealed chips from the wafer, we used an effective chip singulation technique that we informally call the "chip-olate" process. In the experimental section, we first studied dielectrophoretic trapping of 10 μm beads for flow rates ranging from 80 pL s-1 to 2.5 nL s-1 and that are generated by an external syringe pump. Then, we characterized the embedded microheater in DFR-covered chips. Flow rates as high as 8 nL s-1 were generated by evaporation-induced flow when the heater was biased by 10 V, corresponding to 270 mW power. Finally, DEP-based trapping and fluorescent detection of functionalized beads were demonstrated as the flow was generated by the combination of capillary filling and evaporation-induced flow.

  20. Identification and Antimicrobial Resistance of Bacteria Isolated from Probiotic Products Used in Shrimp Culture.

    PubMed

    Noor Uddin, Gazi Md; Larsen, Marianne Halberg; Christensen, Henrik; Aarestrup, Frank M; Phu, Tran Minh; Dalsgaard, Anders

    2015-01-01

    Probiotics are increasingly used in aquaculture to control diseases and improve feed digestion and pond water quality; however, little is known about the antimicrobial resistance properties of such probiotic bacteria and to what extent they may contribute to the development of bacterial resistance in aquaculture ponds. Concerns have been raised that the declared information on probiotic product labels are incorrect and information on bacterial composition are often missing. We therefore evaluated seven probiotics commonly used in Vietnamese shrimp culture for their bacterial species content, phenotypic antimicrobial resistance and associated transferable resistance genes. The bacterial species was established by 16S rRNA sequence analysis of 125 representative bacterial isolates. MIC testing was done for a range of antimicrobials and whole genome sequencing of six multiple antimicrobial resistant Bacillus spp. used to identify resistance genes and genetic elements associated with horizontal gene transfer. Thirteen bacterial species declared on the probiotic products could not be identified and 11 non-declared Bacillus spp. were identified. Although our culture-based isolation and identification may have missed a few bacterial species present in the tested products this would represent minor bias, but future studies may apply culture independent identification methods like pyro sequencing. Only 6/60 isolates were resistant to more than four antimicrobials and whole genome sequencing showed that they contained macrolide (ermD), tetracycline (tetL), phenicol (fexA) and trimethoprim (dfrD, dfrG and dfrK) resistance genes, but not known structures associated with horizontal gene transfer. Probiotic bacterial strains used in Vietnamese shrimp culture seem to contribute with very limited types and numbers of resistance genes compared to the naturally occurring bacterial species in aquaculture environments. Approval procedures of probiotic products must be strengthened

  1. Dynamics of a Class 1 Integron Located on Plasmid or Chromosome in Two Aeromonas spp. Strains

    PubMed Central

    Pérez-Valdespino, Abigail; Lazarini-Martínez, Alfredo; Rivera-González, Alejandro X.; García-Hernández, Normand; Curiel-Quesada, Everardo

    2016-01-01

    Integrons are non-mobile bacterial genetic elements that carry different cassettes conferring antibiotic resistance. Cassettes can excise or integrate by action of an integron-encoded integrase, enabling bacteria to face environmental challenges. In this work, the functionality and dynamics of two integrons carrying the same cassette arrangement (dfrA12–orfF–aadA2), but located on plasmid or chromosome in two different strains were studied. In order to demonstrate the functionality of the Class 1 integrase, circular cassette integration intermediaries were PCR amplified by PCR using extrachromosomal DNA extracted from bacteria grown in the presence or absence of cassette-encoded antibiotics. Circular aadA2 and dfrA12–orfF–aadA2 cassettes were detected in cultures grown either in the presence or absence of antibiotics in both strains. No dfrA12–orfF circular intermediates could be detected under any culture conditions. These results show that both integrons are functional. However, these elements show different dynamics and functionality since the presence of streptomycin led to detectable gene rearrangements in the variable region only in the strain with the plasmid-born integron. In addition, complete integration products were demonstrated using a receptor molecule carrying an empty integron. In this case, integration products were observed in both strains even in the absence of antibiotics, but they were more evident in the strain with the plasmid-located integron when streptomycin was present in the culture medium. This suggests that integrons in the two strains respond differently to streptomycin even though DNA sequences upstream the intI1 gene, including the lexA boxes of both integrons are identical. PMID:27733851

  2. PyMYB10 and PyMYB10.1 Interact with bHLH to Enhance Anthocyanin Accumulation in Pears

    PubMed Central

    Feng, Shouqian; Sun, Shasha; Chen, Xiaoliu; Wu, Shujing; Wang, Deyun; Chen, Xuesen

    2015-01-01

    Color is an important agronomic trait of pears, and the anthocyanin content of fruit is immensely significant for pear coloring. In this study, an anthocyanin-activating R2R3-MYB transcription factor gene, PyMYB10.1, was isolated from fruits of red sand pear (Pyrus pyrifolia cv. Aoguan). Alignments of the nucleotide and amino acid sequences suggested that PyMYB10.1 was involved in anthocyanin regulation. Similar to PyMYB10, PyMYB10.1 was predominantly expressed in red tissues, including the skin, leaf and flower, but it was minimally expressed in non-red fruit flesh. The expression of this gene could be induced by light. Dual-luciferase assays indicated that both PyMYB10 and PyMYB10.1 activated the AtDFR promoter. The activation of AtDFR increased to a greater extent when combined with a bHLH co-factor, such as PybHLH, MrbHLH1, MrbHLH2, or AtbHLH2. However, the response of this activation depended on the protein complex formed. PyMYB10-AtbHLH2 activated the AtDFR promoter to a greater extent than other combinations of proteins. PyMYB10-AtbHLH2 also induced the highest anthocyanin accumulation in tobacco transient-expression assays. Moreover, PybHLH interacted with PyMYB10 and PyMYB10.1. These results suggest that both PyMYB10 and PyMYB10.1 are positive anthocyanin biosynthesis regulators in pears that act via the formation of a ternary complex with PybHLH. The functional characterization of PyMYB10 and PyMYB10.1 will aid further understanding of the anthocyanin regulation in pears. PMID:26536358

  3. A Study on Feasibility of Dual-Wavelength Radar for Identification of Hydrometeor Phases

    NASA Technical Reports Server (NTRS)

    Liao, Liang; Meneghini, Robert

    2010-01-01

    An important objective for the Dual-wavelength Ku-/Ka-band Precipitation Radar (DPR) that will be on board the Global Precipitation Measuring (GPM) core satellite, is to identify the phase state of hydrometeors along the range direction. To assess this, radar signatures are simulated in snow and rain to explore the relation between the differential frequency ratio (DFR), defined as the difference of radar reflectivity factors between Ku- and Ka-bands, and the radar reflectivity factor at Ku-band, ZKu, for different hydrometeor types. Model simulations indicate that there is clear separation between snow and rain in the ZKu-DFR plane assuming that the snow follows the Gunn-Marshall size distribution (1958) and rain follows the Marshall-Palmer size distribution (1948). In an effort to verify the simulated results, the data collected by the Airborne Second Generation Precipitation Radar (APR-2) in the Wakasa Bay AMSR-E campaign are employed. Using the signatures of Linear Depolarization Ratio (LDR) at Ku-band, the APR-2 data can be easily divided into the regions of snow, mixed phase and rain for stratiform storms. These results are then superimposed onto the theoretical curves computed from the model in the ZKu-DFR plane. It has been found that in 90% of the cases, snow and rain can be distinguished if the Ku-band radar reflectivity exceeds 18 dBZ (the minimum detectable level of GPM DPR at Ku-band). This is also the case for snow and mixed-phase hydrometeors. Although snow can be easily distinguished from rain and melting hydrometeors by using Ku- and Ka-band radar, the rain and mixed-phase particles are not always separable. It is concluded that Ku- and Ka-band dual-wavelength radar might provide a potential means to identify the phase state of hydrometeors.

  4. Insights into the structure-function relationship of disease resistance protein HCTR in maize (Zea mays L.): a computational structural biology approach.

    PubMed

    Dehury, Budheswar; Sahu, Mousumi; Patra, Mahesh Chandra; Sarma, Kishore; Sahu, Jagajjit; Sen, Priyabrata; Modi, Mahendra Kumar; Choudhury, Manabendra Dutta; Barooah, Madhumita

    2013-09-01

    The disease resistance gene Hm1 of maize encodes a NADPH-dependent reductase enzyme, HC-toxin reductase (HCTR) that detoxifies the HC toxin secreted by the race specific fungus Cochliobolus carbonum race 1. HCTR enzyme shares 29.6% sequence identity with dihydroflavonol reductase (DFR) of grape, a key enzyme involved in flavonoid biosynthesis. Here we report the comparative modelling, molecular dynamics simulation and docking studies to explain the structure-function relationship and the mode of cofactor (NADPH) binding in HCTR enzyme at the molecular level. The nucleotide binding domain of modelled HCTR adopts a classic Rossmann fold and possesses a consensus glycine rich GxGxxG motif. Molecular simulation studies suggested that HCTR model retained stability throughout the simulation in aqueous solution. HCTR model showed considerable structural identities with the cofactor binding site of DFR, but significant difference in the catalytic site might be the reason of functional divergence between these families of proteins. Similarly electrostatic surface potential analysis of both HCTR and DFR revealed profound variations in the charge distribution over the substrate binding site, which can be correlated with the sequence variability and may suggest distinct substrate-binding patterns and differences in the catalytic mechanism. Docking results indicated Phe19, Gly21, Arg40, Thr90, Gly208, Arg218, Glu221 and Thr222 are important residues for cofactor (NADPH) binding through strong hydrogen bonding and electrostatic interactions. Alanine scanning and analysis of docking energies of mutant proteins suggested that Phe19, and Arg40 are two critical residues for the cofactor binding. The result from the present study is expected to pave the way for exploration of similar genes in other economically important crop varieties. PMID:24004829

  5. Cs-miR156 is involved in the nitrogen form regulation of catechins accumulation in tea plant (Camellia sinensis L.).

    PubMed

    Fan, Kai; Fan, Dongmei; Ding, Zhaotang; Su, Yanhua; Wang, Xiaochang

    2015-12-01

    The nitrogen source affects the growth of tea plants and regulates the accumulation of catechins in the leaves. In this report, we assessed the influences of NH4(+) and NO3(-) on plant growth, catechins accumulation and associated gene expression. Compared with the preferential nitrogen source NH4(+), when NO3(-) was supplied as the sole nitrogen source, tea plants showed similar symptoms with the nitrogen-free treatments and showed lower nitrogen, free amino acid accumulation, chlorophyll content and biomass gain, indicating NO3(-) was not efficiently used by these plants. However, the total shoot catechins content was significantly higher for NO3(-) treatments than that for NH4(+) treatment or combined NH4(+)+NO3(-) treatment, suggesting that, in addition to its influence on plant growth, the nitrogen form regulated the accumulation of catechins in tea. The expression of catechins biosynthesis-related genes was associated with the regulation of catechins accumulation and composition changes mediated by nitrogen form. PAL, CHS, CHI, and DFR genes exhibited higher expression levels in plants supplied with NO3(-), in which the transcript level of DFR in the shoots was significantly correlated with the catechins content. In the end, we identified a new function for the Cs-miR156, which was drastically induced through NH4(+). Moreover, a potential mechanism of the Cs-miR156 pathway in regulating catechins biosynthesis in tea plants has been suggested, with particular respect to nitrogen forms. Cs-miR156 might repress the expression of the target gene SPL to regulate the DFR gene, which plays a vital role in catechins biosynthesis. PMID:26520678

  6. Characterization of trimethoprim-sulfamethoxazole resistance genes and their relatedness to class 1 integron and insertion sequence common region in gram-negative bacilli.

    PubMed

    Shin, Hae Won; Lim, Jinsook; Kim, Semi; Kim, Jimyung; Kwon, Gye Cheol; Koo, Sun Hoe

    2015-01-01

    Trimethoprim-sulfamethoxazole (TMP-SMX) has been used for the treatment of urinary tract infections, but increasing resistance to TMP-SMX has been reported. In this study, we analyzed TMP-SMX resistance genes and their relatedness with integrons and insertion sequence common regions (ISCRs) in uropathogenic gram-negative bacilli. Consecutive nonduplicate TMP-SMX nonsusceptible clinical isolates of E. coli, K. pneumoniae, Acinetobacter spp., and P. aeruginosa were collected from urine. The minimal inhibitory concentration was determined by Etest. TMP-SMX resistance genes (sul and dfr), integrons, and ISCRs were analyzed by PCR and sequencing. A total of 45 E. coli (37.8%), 15 K. pneumoniae (18.5%), 12 Acinetobacter spp. (70.6%), and 9 Pseudomonas aeruginosa (30.0%) isolates were found to be resistant to TMP-SMX. Their MICs were all over 640. In E. coli and K. pneumoniae, sul1 and dfr genes were highly prevalent in relation with integron1. The sul3 gene was detected in E. coli. However, in P. aeruginosa and Acinetobacter spp., only sul1 was prevalent in relation with class 1 integron; however, dfr was not detected and sul2 was less prevalent than in Enterobacteriaceae. ISCR1 and/or ISCR2 were detected in E. coli, K. pneumoniae, and Acinetobacter spp. but the relatedness with TMP-SMX resistance genes was not prominent. ISCR14 was detected in six isolates of E. coli. In conclusion, resistance mechanisms for TMP-SMX were different between Enterobacteriaceae and glucose non-fermenting gram-negative bacilli. Class 1 integron was widely disseminated in uropathogenic gram-negative baciili, so the adoption of prudent use of antimicrobial agents and the establishment of a surveillance system are needed. PMID:25348695

  7. ‘Chip-olate’ and dry-film resists for efficient fabrication, singulation and sealing of microfluidic chips

    NASA Astrophysics Data System (ADS)

    Temiz, Yuksel; Delamarche, Emmanuel

    2014-09-01

    This paper describes a technique for high-throughput fabrication and efficient singulation of chips having closed microfluidic structures and takes advantage of dry-film resists (DFRs) for efficient sealing of capillary systems. The technique is illustrated using 4-inch Si/SiO2 wafers. Wafers carrying open microfluidic structures are partially diced to about half of their thickness. Treatments such as surface cleaning are done at wafer-level, then the structures are sealed using low-temperature (45 °C) lamination of a DFR that is pre-patterned using a craft cutter, and ready-to-use chips are finally separated manually like a chocolate bar by applying a small force (≤ 4 N). We further show that some DFRs have low auto-fluorescence at wavelengths typically used for common fluorescent dyes and that mechanical properties of some DFRs allow for the lamination of 200 μm wide microfluidic structures with negligible sagging (~1 μm). The hydrophilicity (advancing contact angle of ~60°) of the DFR supports autonomous capillary-driven flow without the need for additional surface treatment of the microfluidic chips. Flow rates from 1 to 5 µL min-1 are generated using different geometries of channels and capillary pumps. In addition, the ‘chip-olate’ technique is compatible with the patterning of capture antibodies on DFR for use in immunoassays. We believe this technique to be applicable to the fabrication of a wide range of microfluidic and lab-on-a-chip devices and to offer a viable alternative to many labor-intensive processes that are currently based on wafer bonding techniques or on the molding of poly(dimethylsiloxane) (PDMS) layers.

  8. Identification and Antimicrobial Resistance of Bacteria Isolated from Probiotic Products Used in Shrimp Culture.

    PubMed

    Noor Uddin, Gazi Md; Larsen, Marianne Halberg; Christensen, Henrik; Aarestrup, Frank M; Phu, Tran Minh; Dalsgaard, Anders

    2015-01-01

    Probiotics are increasingly used in aquaculture to control diseases and improve feed digestion and pond water quality; however, little is known about the antimicrobial resistance properties of such probiotic bacteria and to what extent they may contribute to the development of bacterial resistance in aquaculture ponds. Concerns have been raised that the declared information on probiotic product labels are incorrect and information on bacterial composition are often missing. We therefore evaluated seven probiotics commonly used in Vietnamese shrimp culture for their bacterial species content, phenotypic antimicrobial resistance and associated transferable resistance genes. The bacterial species was established by 16S rRNA sequence analysis of 125 representative bacterial isolates. MIC testing was done for a range of antimicrobials and whole genome sequencing of six multiple antimicrobial resistant Bacillus spp. used to identify resistance genes and genetic elements associated with horizontal gene transfer. Thirteen bacterial species declared on the probiotic products could not be identified and 11 non-declared Bacillus spp. were identified. Although our culture-based isolation and identification may have missed a few bacterial species present in the tested products this would represent minor bias, but future studies may apply culture independent identification methods like pyro sequencing. Only 6/60 isolates were resistant to more than four antimicrobials and whole genome sequencing showed that they contained macrolide (ermD), tetracycline (tetL), phenicol (fexA) and trimethoprim (dfrD, dfrG and dfrK) resistance genes, but not known structures associated with horizontal gene transfer. Probiotic bacterial strains used in Vietnamese shrimp culture seem to contribute with very limited types and numbers of resistance genes compared to the naturally occurring bacterial species in aquaculture environments. Approval procedures of probiotic products must be strengthened

  9. Antimicrobial resistance determinants in Acinetobacter baumannii isolates taken from military treatment facilities.

    PubMed

    Taitt, Chris Rowe; Leski, Tomasz A; Stockelman, Michael G; Craft, David W; Zurawski, Daniel V; Kirkup, Benjamin C; Vora, Gary J

    2014-01-01

    Multidrug-resistant (MDR) Acinetobacter baumannii infections are of particular concern within medical treatment facilities, yet the gene assemblages that give rise to this phenotype remain poorly characterized. In this study, we tested 97 clinical A. baumannii isolates collected from military treatment facilities (MTFs) from 2003 to 2009 by using a molecular epidemiological approach that enabled for the simultaneous screening of 236 antimicrobial resistance genes. Overall, 80% of the isolates were found to be MDR, each strain harbored between one and 17 resistant determinants, and a total of 52 unique resistance determinants or gene families were detected which are known to confer resistance to β-lactam (e.g., blaGES-11, blaTEM, blaOXA-58), aminoglycoside (e.g., aphA1, aacC1, armA), macrolide (msrA, msrB), tetracycline [e.g., tet(A), tet(B), tet(39)], phenicol (e.g., cmlA4, catA1, cat4), quaternary amine (qacE, qacEΔ1), streptothricin (sat2), sulfonamide (sul1, sul2), and diaminopyrimidine (dfrA1, dfrA7, dfrA19) antimicrobial compounds. Importantly, 91% of the isolates harbored blaOXA-51-like carbapenemase genes (including six new variants), 40% harbored the blaOXA-23 carbapenemase gene, and 89% contained a variety of aminoglycoside resistance determinants with up to six unique determinants identified per strain. Many of the resistance determinants were found in potentially mobile gene cassettes; 45% and 7% of the isolates contained class 1 and class 2 integrons, respectively. Combined, the results demonstrate a facile approach that supports a more complete understanding of the genetic underpinnings of antimicrobial resistance to better assess the load, transmission, and evolution of MDR in MTF-associated A. baumannii.

  10. Identification and Antimicrobial Resistance of Bacteria Isolated from Probiotic Products Used in Shrimp Culture

    PubMed Central

    Noor Uddin, Gazi Md.; Larsen, Marianne Halberg; Christensen, Henrik; Aarestrup, Frank M.; Phu, Tran Minh; Dalsgaard, Anders

    2015-01-01

    Probiotics are increasingly used in aquaculture to control diseases and improve feed digestion and pond water quality; however, little is known about the antimicrobial resistance properties of such probiotic bacteria and to what extent they may contribute to the development of bacterial resistance in aquaculture ponds. Concerns have been raised that the declared information on probiotic product labels are incorrect and information on bacterial composition are often missing. We therefore evaluated seven probiotics commonly used in Vietnamese shrimp culture for their bacterial species content, phenotypic antimicrobial resistance and associated transferable resistance genes. The bacterial species was established by 16S rRNA sequence analysis of 125 representative bacterial isolates. MIC testing was done for a range of antimicrobials and whole genome sequencing of six multiple antimicrobial resistant Bacillus spp. used to identify resistance genes and genetic elements associated with horizontal gene transfer. Thirteen bacterial species declared on the probiotic products could not be identified and 11 non-declared Bacillus spp. were identified. Although our culture-based isolation and identification may have missed a few bacterial species present in the tested products this would represent minor bias, but future studies may apply culture independent identification methods like pyro sequencing. Only 6/60 isolates were resistant to more than four antimicrobials and whole genome sequencing showed that they contained macrolide (ermD), tetracycline (tetL), phenicol (fexA) and trimethoprim (dfrD, dfrG and dfrK) resistance genes, but not known structures associated with horizontal gene transfer. Probiotic bacterial strains used in Vietnamese shrimp culture seem to contribute with very limited types and numbers of resistance genes compared to the naturally occurring bacterial species in aquaculture environments. Approval procedures of probiotic products must be strengthened

  11. Antibiotic Resistance, Virulence, and Genetic Background of Community-Acquired Uropathogenic Escherichia coli from Algeria.

    PubMed

    Yahiaoui, Merzouk; Robin, Frédéric; Bakour, Rabah; Hamidi, Moufida; Bonnet, Richard; Messai, Yamina

    2015-10-01

    The aim of the study was to investigate antibiotic resistance mechanisms, virulence traits, and genetic background of 150 nonrepetitive community-acquired uropathogenic Escherichia coli (CA-UPEC) from Algeria. A rate of 46.7% of isolates was multidrug resistant. bla genes detected were blaTEM (96.8% of amoxicillin-resistant isolates), blaCTX-M-15 (4%), overexpressed blaAmpC (4%), blaSHV-2a, blaTEM-4, blaTEM-31, and blaTEM-35 (0.7%). All tetracycline-resistant isolates (51.3%) had tetA and/or tetB genes. Sulfonamides and trimethoprim resistance genes were sul2 (60.8%), sul1 (45.9%), sul3 (6.7%), dfrA14 (25.4%), dfrA1 (18.2%), dfrA12 (16.3%), and dfrA25 (5.4%). High-level fluoroquinolone resistance (22.7%) was mediated by mutations in gyrA (S83L-D87N) and parC (S80I-E84G/V or S80I) genes. qnrB5, qnrS1, and aac(6')-Ib-cr were rare (5.3%). Class 1 and/or class 2 integrons were detected (40.7%). Isolates belonged to phylogroups B2+D (50%), A+B1 (36%), and F+C+Clade I (13%). Most of D (72.2%) and 38.6% of B2 isolates were multidrug resistant; they belong to 14 different sequence types, including international successful ST131, ST73, and ST69, reported for the first time in the community in Algeria and new ST4494 and ST4529 described in this study. Besides multidrug resistance, B2 and D isolates possessed virulence factors of colonization, invasion, and long-term persistence. The study highlighted multidrug-resistant CA-UPEC with high virulence traits and an epidemic genetic background.

  12. Clonal dissemination of a single Shigella sonnei strain among Iranian children during Fall 2012 in Tehran, I.R. Iran.

    PubMed

    Alizadeh-Hesar, Mahsa; Bakhshi, Bita; Najar-Peerayeh, Shahin

    2015-08-01

    Shigella species are a common cause of bacterial diarrhea worldwide and the disease is characterized by seasonality. Shigella has been encountered by widespread resistance to commonly used antibiotics which is a serious concern. The aim of this study was to analyze the epidemiological relatedness of Shigella strains isolated from children during one year period by PFGE method and to investigate antimicrobial resistance determinants and cassettes among Shigella species. The occurrence of Shigella spp. in the present study was 1.32% during the study period and the majority of cases (56 (80%)) were occurred during autumn while Shigella sonnei was the most prevalent species identified. Multi-drug resistance phenotype was seen in 98.5% of total isolates with SXT(r)/TE(r)/TMP(r) resistance pattern. Among the 70 Shigella spp. analyzed in this study, 16 isolates were positive for class I integron (int1(+)) with two types of gene cassette arrays (dfrA17/aadA5 and dfrA7).The class 2 integron was more frequently detected among the isolates (85.71%) with dfrA1/sat1/aadA1 (10%) and dfrA1/sat1 (75.71%) gene cassettes. The tetA and tetB determinants were observed in 75.7% and 21.42% of Shigella isolates and tet(A) was the foremost in S. sonnei and Shigella flexneri population. In this study 5 tetracycline resistant isolates had no tetracycline resistance gene (A-D) and no association was recognized between the value of MIC against tetracycline and the tet genes content of isolates. Fifty three of total Shigella isolates (75.7%) showed an identical PFGE patterns. Seven PFGE clusters observed in our study were composed of members with one to three band variations, which is indicative of closely related isolates. The major cluster (cluster C) constituted 75.7% of total isolates, all of which (except eight isolates) consonantly showed identical class 2 integron of 1500 bp which strongly suggests the dissemination of a single S. sonnei clone among the pediatric population in 2012

  13. Quantitation of Rh D antigen sites on weak D and D variant red cells by flow cytometry.

    PubMed

    Jones, J W; Lloyd-Evans, P; Kumpel, B M

    1996-01-01

    Information on the number of D sites on weak D (Du) and D variant cells is limited and incomplete. The aim of this study was to use a simple non-isotopic technique utilising a combination of flow cytometry and ELISA to quantitate the number of D sites on an extensive range of these cells. Five human monoclonal IgG anti-D (BRAD-7 (JAC10), BRAD-5, 2B6, BRAD-3, H27) and one affinity-purified polyclonal IgG anti-D were each used at a saturating concentration of 20 micrograms/ml. In general, BRAD-3, BRAD-5 and 2B6 gave the highest number of D sites per cell (SPC), H27 and the polyclonal anti-D were slightly lower, while BRAD-7 gave the lowest SPC with all D-positive cells tested except DFR. Interestingly, BRAD-7 gave the highest SPC with DFR cells. Rh D antigen density for R2R2 cells was approximately double that seen with either R2r or Rzero (presumed R0r) cells. R1R1 cells gave only moderately higher SPC than R1r cells.Higher SPC were obtained with the R1 haplotype if the CW antigen was present. Weak D, Va and VI cells of the R1 haplotype had higher SPC than those of the Rzero or R2 haplotypes. The majority of D variant cells were found to have lower SPC than normal cells, and for polyclonal anti-D, which was the only anti-D to react with all D variants, SPC decreased in the order IIIc > IIIa > HMii > IVa > Va > DFR > DBT > IVb > VII > II > HMi > VI. The number of molecules of IgG anti-D bound to D variant cells varied by up to 10 times with reactive monoclonal antibodies. The highest SPC on weak D and D variant cells were obtained with BRAD-7 (DFR, 9,500), BRAD-3 (Va, 12,500), BRAD-5 (II, 5,500; VII, 5,300; weak D, 1,300), H27 (III, 24,000; VI, 2,900; HMi, 3,000; HMii, 16,800) and polyclonal anti-D (IVa, 9,300; IVb, 4,000; DBT, 4,300).

  14. Molecular epidemiology of orf513-bearing class 1 integrons in multiresistant clinical isolates from Argentinean hospitals.

    PubMed

    Arduino, Sonia M; Catalano, Mariana; Orman, Betina E; Roy, Paul H; Centrón, Daniela

    2003-12-01

    The spread of orf513-bearing class 1 integrons is associated with bla(CTX-M-2) in gram-negative clinical isolates in Argentina, with In35 being the most frequently found integron (74%). Among 65 isolates without bla(CTX-M-2), only one harbored a novel orf513-bearing class 1 integron with the dfrA3b gene. The finding of orf513 not associated with class 1 integrons in two gram-positive strains indicates the widespread occurrence of this putative site-specific recombinase.

  15. Quantitation of Rh D antigen sites on weak D and D variant red cells by flow cytometry.

    PubMed

    Jones, J W; Lloyd-Evans, P; Kumpel, B M

    1996-01-01

    Information on the number of D sites on weak D (Du) and D variant cells is limited and incomplete. The aim of this study was to use a simple non-isotopic technique utilising a combination of flow cytometry and ELISA to quantitate the number of D sites on an extensive range of these cells. Five human monoclonal IgG anti-D (BRAD-7 (JAC10), BRAD-5, 2B6, BRAD-3, H27) and one affinity-purified polyclonal IgG anti-D were each used at a saturating concentration of 20 micrograms/ml. In general, BRAD-3, BRAD-5 and 2B6 gave the highest number of D sites per cell (SPC), H27 and the polyclonal anti-D were slightly lower, while BRAD-7 gave the lowest SPC with all D-positive cells tested except DFR. Interestingly, BRAD-7 gave the highest SPC with DFR cells. Rh D antigen density for R2R2 cells was approximately double that seen with either R2r or Rzero (presumed R0r) cells. R1R1 cells gave only moderately higher SPC than R1r cells.Higher SPC were obtained with the R1 haplotype if the CW antigen was present. Weak D, Va and VI cells of the R1 haplotype had higher SPC than those of the Rzero or R2 haplotypes. The majority of D variant cells were found to have lower SPC than normal cells, and for polyclonal anti-D, which was the only anti-D to react with all D variants, SPC decreased in the order IIIc > IIIa > HMii > IVa > Va > DFR > DBT > IVb > VII > II > HMi > VI. The number of molecules of IgG anti-D bound to D variant cells varied by up to 10 times with reactive monoclonal antibodies. The highest SPC on weak D and D variant cells were obtained with BRAD-7 (DFR, 9,500), BRAD-3 (Va, 12,500), BRAD-5 (II, 5,500; VII, 5,300; weak D, 1,300), H27 (III, 24,000; VI, 2,900; HMi, 3,000; HMii, 16,800) and polyclonal anti-D (IVa, 9,300; IVb, 4,000; DBT, 4,300). PMID:8912461

  16. Continuous Time Random Walk and Migration-Proliferation Dichotomy of Brain Cancer

    NASA Astrophysics Data System (ADS)

    Iomin, A.

    A theory of fractional kinetics of glial cancer cells is presented. A role of the migration-proliferation dichotomy in the fractional cancer cell dynamics in the outer-invasive zone is discussed and explained in the framework of a continuous time random walk. The main suggested model is based on a construction of a 3D comb model, where the migration-proliferation dichotomy becomes naturally apparent and the outer-invasive zone of glioma cancer is considered as a fractal composite with a fractal dimension Dfr < 3.

  17. Continuous Time Random Walk and Migration-Proliferation Dichotomy of Brain Cancer

    NASA Astrophysics Data System (ADS)

    Iomin, A.

    2015-10-01

    A theory of fractional kinetics of glial cancer cells is presented. A role of the migration-proliferation dichotomy in the fractional cancer cell dynamics in the outer-invasive zone is discussed and explained in the framework of a continuous time random walk. The main suggested model is based on a construction of a 3D comb model, where the migration-proliferation dichotomy becomes naturally apparent and the outer-invasive zone of glioma cancer is considered as a fractal composite with a fractal dimension Dfr < 3.

  18. Molecular Characteristics of Salmonella Genomic Island 1 in Proteus mirabilis Isolates from Poultry Farms in China

    PubMed Central

    Lei, Chang-Wei; Zhang, An-Yun; Liu, Bi-Hui; Guan, Zhong-Bin; Xu, Chang-Wen; Xia, Qing-Qing; Cheng, Han; Zhang, Dong-Dong

    2014-01-01

    Six out of the 64 studied Proteus mirabilis isolates from 11 poultry farms in China contained Salmonella genomic island 1 (SGI1). PCR mapping showed that the complete nucleotide sequences of SGI1s ranged from 33.2 to 42.5 kb. Three novel variants, SGI1-W, SGI1-X, and SGI1-Y, have been characterized. Resistance genes lnuF, dfrA25, and qnrB2 were identified in SGI1 for the first time. PMID:25267683

  19. Two Novel Salmonella Genomic Island 1 Variants in Proteus mirabilis Isolates from Swine Farms in China

    PubMed Central

    Lei, Chang-Wei; Zhang, An-Yun; Liu, Bi-Hui; Yang, Li-Qin; Guan, Zhong-Bin; Xu, Chang-Wen; Zhang, Dong-Dong; Yang, Yong-Qiang

    2015-01-01

    Four different Salmonella genomic island 1 (SGI1) variants, including two novel variants, were characterized in one Salmonella enterica serovar Rissen sequence type ST1917 isolate and three Proteus mirabilis isolates from swine farms in China. One novel variant was derived from SGI1-B with the backbone gene S021 disrupted by a 12.72-kb IS26 composite transposon containing the dfrA17-aadA5 cassettes and macrolide inactivation gene cluster mphA-mrx-mphR. The other one was an integron-free SGI1 and contained a 183-bp truncated S025 next to IS6100 and S044. PMID:25918148

  20. Distribution of class 1 integrons among enteropathogenic Escherichia coli.

    PubMed

    Najibi, S; Bakhshi, B; Fallahzad, S; Pourshafie, M R; Katouli, M; Sattari, M; Alebouyeh, M; Tajbakhsh, M

    2012-05-01

    The aim of this study was to investigate the incidence of and resistance gene content of class 1 integrons among enteropathogenic Escherichia coli (EPEC) and non-EPEC and to investigate intraspecies genetic diversity of EPEC strains isolated from children with diarrhea in Iran. Twenty-eight EPEC and 16 non-EPEC strains isolated from children with diarrhea were tested for the presence of a class 1 integron associated integrase gene (int1). Sequence analysis was performed to identify the resistance gene content of integrons. Genetic diversity and cluster analysis of EPEC isolates were also investigated using enterobacterial repetitive intergenic concensus-polymerase chain reaction (ERIC-PCR) fingerprinting. Twenty-three (82%) EPEC isolates and 11 (68.7%) non-EPEC isolates harbored the int1 gene specific to the conserved integrase region of class 1 integrons. Sequence analysis revealed the dominance of dfrA and aadA gene cassettes among the isolates of both groups. ERIC-PCR fingerprinting of EPEC isolates revealed a high diversity among these isolates. The widespread distribution of 2 resistance gene families (dfrA and aadA) among both groups of EPEC and non-EPEC isolates indicates the significance of integrons in antibiotic resistance transfer among these bacteria. Furthermore, clonal diversity of EPEC isolates harbouring a class 1 integron also suggests the circulation of these mobile elements among a diverse population of EPEC in this country. PMID:22540184

  1. Molecular genetics of para-aminosalicylic acid resistance in clinical isolates and spontaneous mutants of Mycobacterium tuberculosis.

    PubMed

    Mathys, Vanessa; Wintjens, René; Lefevre, Philippe; Bertout, Julie; Singhal, Amit; Kiass, Mehdi; Kurepina, Natalia; Wang, Xiao-Ming; Mathema, Barun; Baulard, Alain; Kreiswirth, Barry N; Bifani, Pablo

    2009-05-01

    The emergence of Mycobacterium tuberculosis resistant to first-line antibiotics has renewed interest in second-line antitubercular agents. Here, we aimed to extend our understanding of the mechanisms underlying para-aminosalicylic acid (PAS) resistance by analysis of six genes of the folate metabolic pathway and biosynthesis of thymine nucleotides (thyA, dfrA, folC, folP1, folP2, and thyX) and three N-acetyltransferase genes [nhoA, aac(1), and aac(2)] among PAS-resistant clinical isolates and spontaneous mutants. Mutations in thyA were identified in only 37% of the clinical isolates and spontaneous mutants. Overall, 24 distinct mutations were identified in the thyA gene and 3 in the dfrA coding region. Based on structural bioinformatics techniques, the altered ThyA proteins were predicted to generate an unfolded or dysfunctional polypeptide. The MIC was determined by Bactec/Alert and dilution assay. Sixty-three percent of the PAS-resistant isolates had no mutations in the nine genes considered in this study, revealing that PAS resistance in M. tuberculosis involves mechanisms or targets other than those pertaining to the biosynthesis of thymine nucleotides. The alternative mechanism(s) or pathway(s) associated with PAS resistance appears to be PAS concentration dependent, in marked contrast to thyA-mutated PAS-resistant isolates.

  2. Noncommutative Geometry of the Moyal Plane: Translation Isometries, Connes' Distance on Coherent States, Pythagoras Equality

    NASA Astrophysics Data System (ADS)

    Martinetti, Pierre; Tomassini, Luca

    2013-10-01

    We study the metric aspect of the Moyal plane from Connes' noncommutative geometry point of view. First, we compute Connes' spectral distance associated with the natural isometric action of on the algebra of the Moyal plane . We show that the distance between any state of and any of its translated states is precisely the amplitude of the translation. As a consequence, we obtain the spectral distance between coherent states of the quantum harmonic oscillator as the Euclidean distance on the plane. We investigate the classical limit, showing that the set of coherent states equipped with Connes' spectral distance tends towards the Euclidean plane as the parameter of deformation goes to zero. The extension of these results to the action of the symplectic group is also discussed, with particular emphasis on the orbits of coherent states under rotations. Second, we compute the spectral distance in the double Moyal plane, intended as the product of (the minimal unitization of) by . We show that on the set of states obtained by translation of an arbitrary state of , this distance is given by the Pythagoras theorem. On the way, we prove some Pythagoras inequalities for the product of arbitrary unital and non-degenerate spectral triples. Applied to the Doplicher- Fredenhagen-Roberts model of quantum spacetime [DFR], these two theorems show that Connes' spectral distance and the DFR quantum length coincide on the set of states of optimal localization.

  3. [Characterization of class 1 and class 2 integron gene cassettes in Escherichia coli strains isolated from urine cultures: a multicenter study].

    PubMed

    Çopur Çiçek, Ayşegül; Sandallı, Cemal; Budak, Emine Esra; Yağmur, Gülhan; Çizmeci, Zeynep; Ak, Sibel; Balcı, Pervin Özlem; Şay Coşkun, Safiye Umut; Ay Altıntop, Yasemin; Fırat, Mehmet; Sarı, Fatma; Çalışkan, Ahmet; Yıldız, Nazan; Sancaktar, Metin; Özgümüş, Osman Birol

    2016-04-01

    %-3.2%), respectively. The frequency of positive IntI1 gene and class 1 integron gene cassettes were found as 25.8% (162/626) and 16.6% (104/626), respectively, whereas the frequency of positive intI2 gene II and class 2 integron gene cassettes were 5.1% (32/626) and 3% (19/626), respectively. The lowest intI1 gene frequency was detected in the isolates from Kayseri (16.6%) and the highest in the isolates from Kahramanmaraş (35.4%) provinces. While there was no intI2 gene in the isolates from Denizli and Kayseri, the highest frequency was 12.1% in the isolates from Şanlıurfa province. dfrA1 gene, the most frequent gene among integron gene cassettes was positive in 31 class 1 integron gene cassette alone, and positive with aadA1 gene in 18 class 1 integron gene cassettes. dfrA1 gene was positive with aadA1a just in one isolate. dfrA17 allele was positive in one isolate alone, in 28 isolates with aadA1, and in 15 isolates with aadA5. aadA1 gene was detected in four isolates. dfrA17-sat-aadA5 co-existence was detected among class 2 integron gene cassette in isolates from six provinces. dfrA1-sat-aadA1 was detected in one isolate from Ankara province and dfrA1 was detected in one isolate in Niğde province only. As a result, dfrA1 and aadA1 genes are the most common types of genes among class 1 and class 2 integron gene cassettes in E.coli isolated from urine cultures. It was concluded that high resistance against streptomycin (31.2%) and SXT (41.2%) supported the dissemination of integron-mediated genes dfr, sul1 and aad in the isolates. PMID:27175490

  4. [Do elderly individuals with a cognitive handicap have worse oral health?].

    PubMed

    Ferreira, Raquel Conceição; Vargas, Andréa Maria Duarte; Fernandes, Neuma Carla Neves; Souza, João Gabriel Silva; Sá, Maria Aparecida Barbosa de; Oliveira, Lorenna Fonseca Braga de; Martins, Andréa Maria Eleutério de Barros Lima

    2014-08-01

    A comparison of the oral health of elderly people with and without a cognitive handicap was assessed. The cognitive condition, the indices of decayed, missing, filled teeth (DMFT), decayed, filled roots (DFR), the need for dental treatment, the presence of plaque (P), calculus (C), the community periodontal index (CPI), the rate of periodontal attachment loss (PAL), edentulism, prosthetic use and the need for prosthetics were evaluated in a complex probabilistic sample by conglomerates of the elderly (65-74 years). PASW(r) 17.0 was used for the statistical analyses with correction for the design effect, applying the Mann Whitney and chi-square test with 95% reliability. A total of 736 elderly individuals were interviewed and examined. Those with cognitive impairment had higher average DMFT, DFR and lower average healthy sextant CPI, a lower prevalence of sextants without plaque/calculus, use of prosthetics and higher prevalence of edentulism and need for prosthetics. Elderly individuals with a cognitive handicap had poorer oral health. PMID:25119081

  5. Free and Cued Selective Reminding Test: an Italian normative study.

    PubMed

    Frasson, P; Ghiretti, R; Catricalà, E; Pomati, S; Marcone, A; Parisi, L; Rossini, P M; Cappa, S F; Mariani, C; Vanacore, N; Clerici, F

    2011-12-01

    The presence of episodic memory impairment is required for the diagnosis of Alzheimer's dementia by all current diagnostic criteria. The new research criteria proposed by Dubois et al. (Lancet Neurol 6:734-746, 2007) require that the impairment should not improve significantly with cueing, recognition testing nor after the control of effective encoding. This is considered to be the core deficit of "prodromal Alzheimer's disease". The Free and Cued Selective Reminding Test (FCSRT) is a memory test that allows in assessing these specific features of memory impairment. Here, we report normative data for an Italian version of the FCSRT. The test is based on the 12 pictorial stimuli, 6 belonging to the living domain, and 6 to the non-living domain. Six scores were derived from the performance of 227 healthy Italian adults, with age, sex and education homogenously distributed across subgroups: immediate free recall (IFR), immediate total recall (ITR), delayed-free recall (DFR), delayed total recall (DTR), Index of Sensitivity of Cueing (ISC), number of intrusions. In multiple regression analyses, age emerged as an influencing factor for both IFR and DFR, with older people obtaining lower scores. Education and gender appear to influence only IFR, with better performance by more educated subjects and females. Adjusted scores were used to determine inferential cutoff scores and to compute equivalent scores. PMID:21594655

  6. A Novel bHLH Transcription Factor Involved in Regulating Anthocyanin Biosynthesis in Chrysanthemums (Chrysanthemum morifolium Ramat.).

    PubMed

    Xiang, Li-li; Liu, Xiao-fen; Li, Xue; Yin, Xue-ren; Grierson, Donald; Li, Fang; Chen, Kun-song

    2015-01-01

    Chrysanthemums (Chrysanthemum morifolium Ramat.) exhibit a variety of flower colors due to their differing abilities to accumulate anthocyanins. One MYB member, CmMYB6, has been verified as a transcription regulator of chrysanthemum genes involved in anthocyanin biosynthesis; however, the co-regulators for CmMYB6 remain unclear in chrysanthemum. Here, the expression pattern of CmbHLH2, which is clustered in the IIIf bHLH subgroup, was shown to be positively correlated with the anthocyanin content of cultivars with red, pink and yellow flower colors, respectively. CmbHLH2 significantly upregulated the CmDFR promoter and triggered anthocyanin accumulation when co-expressed with CmMYB6. Yeast one-hybrid analyses indicated that CmbHLH2 was able to bind directly to the CmDFR promoter. Moreover, yeast two-hybrid assays indicated protein-protein interaction between CmbHLH2 and CmMYB6. These results suggest that CmbHLH2 is the essential partner for CmMYB6 in regulating anthocyanin biosynthesis in chrysanthemum. PMID:26619181

  7. Antibiotic resistance and prevalence of class 1 and 2 integrons in Escherichia coli isolated from two wastewater treatment plants, and their receiving waters (Gulf of Gdansk, Baltic Sea, Poland).

    PubMed

    Kotlarska, Ewa; Łuczkiewicz, Aneta; Pisowacka, Marta; Burzyński, Artur

    2015-02-01

    In this study, antimicrobial-resistance patterns were analyzed in Escherichia coli isolates from raw (RW) and treated wastewater (TW) of two wastewater treatment plants (WWTPs), their marine outfalls (MOut), and mouth of the Vistula River (VR). Susceptibility of E. coli was tested against different classes of antibiotics. Isolates resistant to at least one antimicrobial agent were PCR tested for the presence of integrons. Ampicillin-resistant E. coli were the most frequent, followed by amoxicillin/clavulanate (up to 32 %), trimethoprim/sulfamethoxazole (up to 20 %), and fluoroquinolone (up to 15 %)-resistant isolates. Presence of class 1 and 2 integrons was detected among tested E. coli isolates with rate of 32.06 % (n = 84) and 3.05 % (n = 8), respectively. The presence of integrons was associated with increased frequency of resistance to fluoroquinolones, trimethoprim/sulfamethoxazole, amoxicillin/clavulanate, piperacillin/tazobactam, and presence of multidrug-resistance phenotype. Variable regions were detected in 48 class 1 and 5 class 2 integron-positive isolates. Nine different gene cassette arrays were confirmed among sequenced variable regions, with predominance of dfrA1-aadA1, dfrA17-aadA5, and aadA1 arrays. These findings illustrate the importance of WWTPs in spreading of resistance genes in the environment and the need for inclusion of at least monitoring efforts in the regular WWTP processes.

  8. Biochemical responses in tree foliage exposed to coal-fired power plant emission in seasonally dry tropical environment.

    PubMed

    Sharma, Atul Prakash; Tripathi, B D

    2009-11-01

    A biomonitoring study was conducted to investigate the responses of plants exposed to power plant emission in a dry tropical environment. For this purpose, five sampling sites were selected in the prevailing wind direction (NE) at different distance to thermal power plant (TPP) within 8.0 km range and a reference site was selected in eastern direction at a distance of 22.0 km. The two most common tree species, Ficus benghalensis L. (Evergreen tree) and Dalbergia sisso Roxb. (deciduous tree) were selected as test plants. Ambient sulphur dioxide (SO(2)), nitrogen dioxide (NO(2)), suspended particulate matter (SPM), respirable suspended particulate matter (RSPM), dust-fall rate (DFR) and plant responses such as leaf pigments (chlorophyll a, chlorophyll b and carotenoids), ascorbic acid, sugar and sulphate-sulphur (SO4(2-)-S) contents were measured. Ambient SO(2), NO(2), SPM, RSPM and DFR showed significant spatial and temporal variation at different sites. Considerable reduction in pigment (chlorophyll a, chlorophyll b and carotenoids) and sugar contents were observed at sites receiving higher pollution load. Ascorbic acid exhibited significant positive correlation with pollution load. Accumulation of SO4(2-)-S in leaf tissue showed significant positive correlation with ambient SO(2) concentration at all the sites. At the same time, SO4(2-)-S showed significant negative correlation with pigment and sugar content. D. sisso Roxb. tree was found to be more sensitive as compared to F. benghalensis L. tree.

  9. Accumulation of Phenylpropanoids by White, Blue, and Red Light Irradiation and Their Organ-Specific Distribution in Chinese Cabbage (Brassica rapa ssp. pekinensis).

    PubMed

    Kim, Yeon Jeong; Kim, Yeon Bok; Li, Xiaohua; Choi, Su Ryun; Park, Suhyoung; Park, Jong Seok; Lim, Yong Pyo; Park, Sang Un

    2015-08-01

    This study investigated optimum light conditions for enhancing phenylpropanoid biosynthesis and the distribution of phenylpropanoids in organs of Chinese cabbage (Brassica rapa ssp. pekinensis). Blue light caused a high accumulation of most phenolic compounds, including p-hydroxybenzoic acid, ferulic acid, quercetin, and kaempferol, at 12 days after irradiation (DAI). This increase was coincident with a noticeable increase in expression levels of BrF3H, BrF3'H, BrFLS, and BrDFR. Red light led to the highest ferulic acid content at 12 DAI and to elevated expression of the corresponding genes during the early stages of irradiation. White light induced the highest accumulation of kaempferol and increased expression of BrPAL and BrDFR at 9 DAI. The phenylpropanoid content analysis in different organs revealed organ-specific accumulation of p-hydroxybenzoic acid, quercetin, and kaempferol. These results demonstrate that blue light is effective at increasing phenylpropanoid biosynthesis in Chinese cabbage, with leaves and flowers representing the most suitable organs for the production of specific phenylpropanoids.

  10. A Novel bHLH Transcription Factor Involved in Regulating Anthocyanin Biosynthesis in Chrysanthemums (Chrysanthemum morifolium Ramat.)

    PubMed Central

    Li, Xue; Yin, Xue-ren; Grierson, Donald; Li, Fang; Chen, Kun-song

    2015-01-01

    Chrysanthemums (Chrysanthemum morifolium Ramat.) exhibit a variety of flower colors due to their differing abilities to accumulate anthocyanins. One MYB member, CmMYB6, has been verified as a transcription regulator of chrysanthemum genes involved in anthocyanin biosynthesis; however, the co-regulators for CmMYB6 remain unclear in chrysanthemum. Here, the expression pattern of CmbHLH2, which is clustered in the IIIf bHLH subgroup, was shown to be positively correlated with the anthocyanin content of cultivars with red, pink and yellow flower colors, respectively. CmbHLH2 significantly upregulated the CmDFR promoter and triggered anthocyanin accumulation when co-expressed with CmMYB6. Yeast one-hybrid analyses indicated that CmbHLH2 was able to bind directly to the CmDFR promoter. Moreover, yeast two-hybrid assays indicated protein-protein interaction between CmbHLH2 and CmMYB6. These results suggest that CmbHLH2 is the essential partner for CmMYB6 in regulating anthocyanin biosynthesis in chrysanthemum. PMID:26619181

  11. A differential fluorescent receptor for nucleic acid analysis.

    PubMed

    Bengtson, Hillary N; Kolpashchikov, Dmitry M

    2014-01-24

    Differential receptors use an array of sensors to recognize analytes. Each sensor in the array can recognize not one, but several analytes with different rates, so a single analyte triggers a response of several sensors in the array. The receptor thus produces a pattern of signals that is unique for each analyte, thereby enabling identification of a specific analyte by producing a "fingerprint" pattern. We applied this approach for the analysis of DNA sequences of Mycobacterium tuberculosis strains that differ by single nucleotide substitutions in the 81-bp hot-spot region that imparts rifampin resistance. The technology takes advantage of the new multicomponent, selfassembling sensor, which produces a fluorescent signal in the presence of specific DNA sequences. A differential fluorescent receptor (DFR) contained an array of three such sensors and differentiated at least eight DNA sequences. The approach requires only one molecular-beacon-like fluorescent reporter, which can be used by all three sensors. The DFR developed in this study represents a cost-efficient alternative to molecular diagnostic technologies that use fluorescent hybridization probes.

  12. Genetic environments of the transferable plasmid-mediated blaCTX-M-3 gene in Serratia marcescens isolates.

    PubMed

    Chu, Pei-Yu; Peng, Chien-Fang

    2014-01-01

    In this study, genetic environments of the transferable plasmid-mediated blaCTX-M-3 gene were characterized among 14 isolates of cefotaxime-resistant Serratia marcescens using PCR and BLAST DNA sequence analysis. A total of 3 types of genetic architectures in the regions surrounding this blaCTX-M-3 gene were identified. Type I architecture was characterized by the presence of a complete insertion sequence of tnpA-ISEcp1, identified as interrupting a reverse IS26 sequence in the upstream region of the blaCTX-M-3 gene. A reverse-directional orf477 fragment was located downstream of the blaCTX-M-3 gene, which was in the same direction of the mucA gene. A common region containing the orf513 element was located upstream of the mucA gene. Moreover, a copy of the 3'-CS2 element was located immediately upstream of the orf513 element. A novel complex class 1 integron was characterized by the presence of the dfrA19 gene, which was flanked by two copies of class 1 integrons. This is the first report to describe the dfrA19 gene within a novel complex class 1 integron in S. marcescens isolates from Taiwan. This novel complex class 1 integron structure was located distantly upstream of the blaCTX-M-3 gene.

  13. Development, relative validity, and reliability of a food frequency questionnaire for a case-control study on dietary advanced glycation end products and diabetes complications.

    PubMed

    Luevano-Contreras, Claudia; Durkin, Taylor; Pauls, Maria; Chapman-Novakofski, Karen

    2013-12-01

    Dietary advanced glycation end products (dAGEs) could be involved on diabetes complications, yet their quantification is not standardized. The objective of this study was to design a food frequency questionnaire (FFQ) for dAGEs, and to assess its reliability and validity. For the design, data from 30 subjects was used. The final instrument had 90 food items. To measure reliability and validity, 20 participants with type 2 diabetes filled out twice the FFQ (FFQ-T1, FFQ-T2) and 7-day food records (7-dFR). The Shrout-Fleiss coefficient was 0.98 showing good reliability. For validation, the results for the weighted kappa were 0.55 (moderate agreement) for FFQ-T1 and 0.64 (good agreement) for FFQ-T2, and 75% and 80% of subjects respectively were correctly classified into tertiles; Bland-Altman graphics showed no systematic bias. This FFQ is comparable to 7-dFR for measuring dAGEs. To our knowledge, this is the first questionnaire designed to measure specifically dAGEs.

  14. [Do elderly individuals with a cognitive handicap have worse oral health?].

    PubMed

    Ferreira, Raquel Conceição; Vargas, Andréa Maria Duarte; Fernandes, Neuma Carla Neves; Souza, João Gabriel Silva; Sá, Maria Aparecida Barbosa de; Oliveira, Lorenna Fonseca Braga de; Martins, Andréa Maria Eleutério de Barros Lima

    2014-08-01

    A comparison of the oral health of elderly people with and without a cognitive handicap was assessed. The cognitive condition, the indices of decayed, missing, filled teeth (DMFT), decayed, filled roots (DFR), the need for dental treatment, the presence of plaque (P), calculus (C), the community periodontal index (CPI), the rate of periodontal attachment loss (PAL), edentulism, prosthetic use and the need for prosthetics were evaluated in a complex probabilistic sample by conglomerates of the elderly (65-74 years). PASW(r) 17.0 was used for the statistical analyses with correction for the design effect, applying the Mann Whitney and chi-square test with 95% reliability. A total of 736 elderly individuals were interviewed and examined. Those with cognitive impairment had higher average DMFT, DFR and lower average healthy sextant CPI, a lower prevalence of sextants without plaque/calculus, use of prosthetics and higher prevalence of edentulism and need for prosthetics. Elderly individuals with a cognitive handicap had poorer oral health.

  15. Improved high performance liquid chromatographic separation of anthocyanin compounds from grapes using a novel mixed-mode ion-exchange reversed-phase column.

    PubMed

    McCallum, Jason L; Yang, Raymond; Young, J Christopher; Strommer, Judith N; Tsao, Rong

    2007-04-27

    A novel mixed mode HPLC method using a column combining both ion-exchange and reversed-phase separation mechanisms has been developed to facilitate analysis of anthocyanins in grapes. Chromatographic performance and subsequent analysis of anthocyanidin diglucosides and acylated compounds are significantly improved using the new column, compared to those associated with conventional C18 reversed-phase methods. The mixed mode column produces a distinctive eluting pattern for the different anthocyanin subgroups, avoiding overlaps found with C18 columns. The enhanced chromatographic resolution provides nearly complete separation of 37 anthocyanin types, and permits detection of delphinidin 3-O-(6''-O-caffeoyl) beta-D-glucoside for the first time in extracts of skins from Concord grapes. PMID:17382950

  16. Functional analysis of differentially expressed proteins in Chinese bayberry (Myrica rubra Sieb. et Zucc.) fruits during ripening.

    PubMed

    Chen, Yi-Yong; Zhang, Ze-Huang; Zhong, Can-Yu; Song, Xiao-Min; Lin, Qi-Hua; Huang, Chun-Mei; Huang, Rong-Hui; Chen, Wei

    2016-01-01

    This study developed a proteome reference map of Myrica rubra fruits at the green, pink and red stages during ripening using two-dimensional gel electrophoresis (2-DE). Forty-six differentially expressed proteins were detected in the gel, of which 43 were successfully identified by matrix-assisted laser desorption ionization time-of-flight/time-of-flight mass spectrometry and protein database searching. We found that malic enzyme related to the decrease of organic acid acidity was up-regulated. The high abundance of pyruvate decarboxylase and alcohol dehydrogenase may contribute to fruit peculiar fragrant characteristics. Phenylalanine ammonia-lyase, chalcone synthase 11, UDP-glucose:flavonoid 3-O-glucosyltransferase, and anthocyanidin synthase, enzymes involved in the anthocyanin metabolic pathway, were all up-regulated. The physiological data agree with fruit proteome results. These findings provided insights into the metabolic processes and regulatory mechanisms during Chinese bayberry fruit ripening. PMID:26213036

  17. Chemical, Bioactive, and Antioxidant Potential of Twenty Wild Culinary Mushroom Species

    PubMed Central

    Sharma, S. K.; Gautam, N.

    2015-01-01

    The chemical, bioactive, and antioxidant potential of twenty wild culinary mushroom species being consumed by the people of northern Himalayan regions has been evaluated for the first time in the present study. Nutrients analyzed include protein, crude fat, fibres, carbohydrates, and monosaccharides. Besides, preliminary study on the detection of toxic compounds was done on these species. Bioactive compounds evaluated are fatty acids, amino acids, tocopherol content, carotenoids (β-carotene, lycopene), flavonoids, ascorbic acid, and anthocyanidins. Fruitbodies extract of all the species was tested for different types of antioxidant assays. Although differences were observed in the net values of individual species all the species were found to be rich in protein, and carbohydrates and low in fat. Glucose was found to be the major monosaccharide. Predominance of UFA (65–70%) over SFA (30–35%) was observed in all the species with considerable amounts of other bioactive compounds. All the species showed higher effectiveness for antioxidant capacities. PMID:26199938

  18. Analysis of polar antioxidants in Heartsease (Viola tricolor L.) and Garden pansy (Viola x wittrockiana Gams.).

    PubMed

    Vukics, V; Kery, A; Guttman, A

    2008-10-01

    Heartsease (Viola tricolor L.) is a well-known medicinal plant. Its biological activities are supposed to be related to its antioxidant capacity. Garden pansies (Viola x wittrockiana Gams.) have been crossbred from heartsease and are applied as ornamental plants only. In this study, the mother and the daughter species are compared from a phytochemical point of view. Their flavonoid and anthocyanidin contents are determined by spectroscopic methods recommended by the European Pharmacopoeia 5.0. The compositions of the samples (heartsease and garden pansy varietas of several petal color) are analyzed by high-performance liquid chromatography with UV detection and their antioxidant capacity is determined by trolox equivalent antioxidant capacity assay. Our results suggest that garden pansy, especially its flower, is a promising source of natural antioxidants. In addition, a significant correlation is found between the flavonoid content and antioxidant activity.

  19. In vitro inhibition of cytochrome P450 3A4 by Aronia melanocarpa constituents.

    PubMed

    Bräunlich, Marie; Christensen, Hege; Johannesen, Siri; Slimestad, Rune; Wangensteen, Helle; Malterud, Karl E; Barsett, Hilde

    2013-01-01

    Extracts, subfractions, isolated anthocyanins and procyanidins, and two phenolic acids from aronia [Aronia melanocarpa] were investigated for their CYP3A4 inhibitory effects, using midazolam as the probe substrate and recombinant insect cell microsomes expressing CYP3A4 as the enzyme source. Procyanidin B5 was a considerably stronger CYP3A4 inhibitor in vitro than the isomeric procyanidin B2 and comparable to bergamottin, a known CYP3A4 inhibitor from grapefruit juice. The inhibitory activity of proanthocyanidin-containing fractions was correlated to the degree of polymerization. Among the anthocyanins, cyanidin 3-arabinoside showed stronger CYP3A4 inhibition than cyanidin 3-galactoside and cyanidin 3-glucoside. Thus, the ability to inhibit CYP3A4 in vitro seems to be influenced by the sugar unit linked to the anthocyanidin.

  20. Antimicrobial effects of the stem bark extracts of Parkia biglobosa (Jacq.) Benth. on Shigellae.

    PubMed

    Millogo-Kone, H; Guissou, Ip; Nacoulma, O; Traore, A S

    2007-06-10

    Total and hydroalcoholic extracts of the stem barks of Parkia biglobosa (Jacq) Benth. (Mimosaceae) were tested on strains belonging to three species of Shigellae: S. dysenteriae, S. flexneri and S. boydii collected from hospitals in Ouagadougou, Burkina Faso. The results showed that both extracts were active against Shigellae. The hydroalcoholic extract was more active than the decoction (aqueous one) prescribed by the traditional healer. Both extracts were particularly effective against S. dysenteriae, the most virulent of the three pathogenic species. The effects of the extracts have been compared to that of gentamicin. The phytochemical screening on the extracts revealed the presence of sterols, triterpenes, polyphenolic compounds including tannins, flavonoids, coumarins, anthocyanidins. Other components are saponosides and reducing sugars.

  1. Vaccinium corymbosum L. (blueberry) extracts exhibit protective action against cadmium toxicity in Saccharomyces cerevisiae cells.

    PubMed

    Oprea, Eliza; Ruta, Lavinia L; Nicolau, Ioana; Popa, Claudia V; Neagoe, Aurora D; Farcasanu, Ileana C

    2014-01-01

    Blueberries (Vaccinium corymbosum L.) are a rich source of antioxidants and their consumption is believed to contribute to food-related protection against oxidative stress. In the present study, the chemoprotective action of blueberry extracts against cadmium toxicity was investigated using a cadmium-hypersensitive strain of Saccharomyces cerevisiae. Four varieties of blueberries were used in the study, and it was found that the extracts with high content of total anthocyanidins exhibited significant protective effect against the toxicity of cadmium and H2O2. Both the blueberry extracts and pure cyanidin exhibited protective effects against cadmium in a dose-dependent manner, but without significantly interfering with the cadmium accumulation by the yeast cells. The results imply that the blueberry extracts might be a potentially valuable food supplement for individuals exposed to high cadmium.

  2. Constitutive expression of the maize genes B1 and C1 in transgenic Hi II maize results in differential tissue pigmentation and generates resistance to Helicoverpa zea.

    PubMed

    Johnson, Eric T; Berhow, Mark A; Dowd, Patrick F

    2010-02-24

    Anthocyanin biosynthesis in maize protects tissues from biotic and abiotic stresses. Constitutive expression of the maize B1 and C1 genes, which induces anthocyanin biosynthesis, resulted in transgenic plants with varied phenotypes. Some colored leaves were substantially resistant to thrips damage, while only leaves with the highest levels of cyanidin, the predominant anthocyanidin detected in all colored transgenic tissues, were resistant to corn earworm (CEW) larvae. Colored anthers were resistant to CEW feeding, and reductions in CEW growth were significantly correlated to levels of cyanidin in the anthers. Cyanidin chloride and cyanidin-3-glucoside chloride added to insect diet slowed the growth of CEW larvae. Attempts to produce 3'5'-hydroxylated anthocyanins in colored maize with the expression of a petunia F3'5'H hydroxylase gene were unsuccessful. PMID:20108901

  3. Structural insights into substrate specificity of Feruloyl-CoA 6’-Hydroxylase from Arabidopsis thaliana

    DOE PAGES

    Sun, Xinxiao; Zhou, Dayong; Kandavelu, Palani; Zhang, Hua; Yuan, Qipeng; Wang, Bi -Cheng; Rose, John; Yan, Yajun

    2015-05-20

    Coumarins belong to an important class of plant secondary metabolites. Feruloyl-CoA 6’-hydroxylase (F6’H), a 2-oxoglutarate dependent dioxygenase (2OGD), catalyzes a pivotal step in the biosynthesis of a simple coumarin scopoletin. In this study, we determined the 3-dimensional structure of the F6’H1 apo enzyme by X-ray crystallography. It is the first reported structure of a 2OGD enzyme involved in coumarin biosynthesis and closely resembles the structure of Arabidopsis thaliana anthocyanidin synthase. To better understand the mechanism of enzyme catalysis and substrate specificity, we also generated a homology model of a related ortho-hydroxylase (C2’H) from sweet potato. By comparing these two structures,more » we targeted two amino acid residues and verified their roles in substrate binding and specificity by site-directed mutagenesis.« less

  4. Structural insights into substrate specificity of Feruloyl-CoA 6’-Hydroxylase from Arabidopsis thaliana

    SciTech Connect

    Sun, Xinxiao; Zhou, Dayong; Kandavelu, Palani; Zhang, Hua; Yuan, Qipeng; Wang, Bi -Cheng; Rose, John; Yan, Yajun

    2015-05-20

    Coumarins belong to an important class of plant secondary metabolites. Feruloyl-CoA 6’-hydroxylase (F6’H), a 2-oxoglutarate dependent dioxygenase (2OGD), catalyzes a pivotal step in the biosynthesis of a simple coumarin scopoletin. In this study, we determined the 3-dimensional structure of the F6’H1 apo enzyme by X-ray crystallography. It is the first reported structure of a 2OGD enzyme involved in coumarin biosynthesis and closely resembles the structure of Arabidopsis thaliana anthocyanidin synthase. To better understand the mechanism of enzyme catalysis and substrate specificity, we also generated a homology model of a related ortho-hydroxylase (C2’H) from sweet potato. By comparing these two structures, we targeted two amino acid residues and verified their roles in substrate binding and specificity by site-directed mutagenesis.

  5. Crystal Structures of Glycosyltransferase UGT78G1 Reveal the Molecular Basis for Glycosylation and Deglycosylation of (Iso)flavonoids

    SciTech Connect

    Modolo, Luzia V.; Li, Lenong; Pan, Haiyun; Blount, Jack W.; Dixon, Richard A.; Wang, Xiaoqiang

    2010-09-21

    The glycosyltransferase UGT78G1 from Medicago truncatula catalyzes the glycosylation of various (iso)flavonoids such as the flavonols kaempferol and myricetin, the isoflavone formononetin, and the anthocyanidins pelargonidin and cyanidin. It also catalyzes a reverse reaction to remove the sugar moiety from glycosides. The structures of UGT78G1 bound with uridine diphosphate or with both uridine diphosphate and myricetin were determined at 2.1 {angstrom} resolution, revealing detailed interactions between the enzyme and substrates/products and suggesting a distinct binding mode for the acceptor/product. Comparative structural analysis and mutagenesis identify glutamate 192 as a key amino acid for the reverse reaction. This information provides a basis for enzyme engineering to manipulate substrate specificity and to design effective biocatalysts with glycosylation and/or deglycosylation activity.

  6. Greater flavonoid intake is associated with improved CVD risk factors in US adults.

    PubMed

    Kim, Kijoon; Vance, Terrence M; Chun, Ock K

    2016-04-01

    Epidemiological studies have reported that diets high in flavonoids are associated with a reduced risk of CVD. However, evidence on the association of dietary flavonoid intake with CVD risk factors is still scarce. The present study aimed to investigate the association of dietary flavonoid intake with CVD risk factors among US adults in the National Health and Nutrition Examination Survey (NHANES) 2007-2012. A total of 4042 US adults aged 19 years and older from the NHANES 2007-2012 participated in this cross-sectional, population-based study. Intakes of total and individual flavonoids were estimated from 2-d 24-h diet recall data by matching with the expanded US Department of Agriculture flavonoid, isoflavone and proanthocyanidin databases. After adjusting for covariates, increased HDL-cholesterol was associated with higher total flavonoid intake (0·54 % change). TAG and TAG:HDL-cholesterol ratio were inversely associated with anthocyanidin (-1·25 % change for TAG; -1·60 % change for TAG:HDL-cholesterol ratio) and total flavonoid intakes (-1·31 % change for TAG; -1·83 % change for TAG:HDL-cholesterol ratio), respectively. Insulin and homoeostasis model assessment for insulin resistance (HOMA-IR) were inversely associated with flavone (for insulin, -3·18 % change; 95 % CI -5·85, -0·44; for HOMA-IR, -3·10 % change; 95 % CI -5·93, -0·19) and isoflavone intakes (for insulin, -3·11 % change; 95 % CI -5·46, -0·70; for HOMA-IR, -4·01 % change; 95 % CI -6·67, -1·27). BMI was negatively associated with anthocyanidin intake (-0·60 % change). This study showed that higher flavonoid intake was associated with improved CVD risk factors. Further research is warranted to confirm the findings from this study as these associations were moderate in strength. PMID:26931451

  7. De Novo Sequencing and Analysis of the Safflower Transcriptome to Discover Putative Genes Associated with Safflor Yellow in Carthamus tinctorius L.

    PubMed Central

    Liu, Xiuming; Dong, Yuanyuan; Yao, Na; Zhang, Yu; Wang, Nan; Cui, Xiyan; Li, Xiaowei; Wang, Yanfang; Wang, Fawei; Yang, Jing; Guan, Lili; Du, Linna; Li, Haiyan; Li, Xiaokun

    2015-01-01

    Safflower (Carthamus tinctorius L.), an important traditional Chinese medicine, is cultured widely for its pharmacological effects, but little is known regarding the genes related to the metabolic regulation of the safflower’s yellow pigment. To investigate genes related to safflor yellow biosynthesis, 454 pyrosequencing of flower RNA at different developmental stages was performed, generating large databases.In this study, we analyzed 454 sequencing data from different flowering stages in safflower. In total, 1,151,324 raw reads and 1,140,594 clean reads were produced, which were assembled into 51,591 unigenes with an average length of 679 bp and a maximum length of 5109 bp. Among the unigenes, 40,139 were in the early group, 39,768 were obtained from the full group and 28,316 were detected in both samples. With the threshold of “log2 ratio ≥ 1”, there were 34,464 differentially expressed genes, of which 18,043 were up-regulated and 16,421 were down-regulated in the early flower library. Based on the annotations of the unigenes, 281 pathways were predicted. We selected 12 putative genes and analyzed their expression levels using quantitative real time-PCR. The results were consistent with the 454 sequencing results. In addition, the expression of chalcone synthase, chalcone isomerase and anthocyanidin synthase, which are involved in safflor yellow biosynthesis and safflower yellow pigment (SYP) content, were analyzed in different flowering periods, indicating that their expression levels were related to SYP synthesis. Moreover, to further confirm the results of the 454 pyrosequencing, full-length cDNA of chalcone isomerase (CHI) and anthocyanidin synthase (ANS) were cloned from safflower petal by RACE (Rapid-amplification of cDNA ends) method according to fragment of the transcriptome. PMID:26516840

  8. De Novo Sequencing and Analysis of the Safflower Transcriptome to Discover Putative Genes Associated with Safflor Yellow in Carthamus tinctorius L.

    PubMed

    Liu, Xiuming; Dong, Yuanyuan; Yao, Na; Zhang, Yu; Wang, Nan; Cui, Xiyan; Li, Xiaowei; Wang, Yanfang; Wang, Fawei; Yang, Jing; Guan, Lili; Du, Linna; Li, Haiyan; Li, Xiaokun

    2015-10-26

    Safflower (Carthamus tinctorius L.), an important traditional Chinese medicine, is cultured widely for its pharmacological effects, but little is known regarding the genes related to the metabolic regulation of the safflower's yellow pigment. To investigate genes related to safflor yellow biosynthesis, 454 pyrosequencing of flower RNA at different developmental stages was performed, generating large databases.In this study, we analyzed 454 sequencing data from different flowering stages in safflower. In total, 1,151,324 raw reads and 1,140,594 clean reads were produced, which were assembled into 51,591 unigenes with an average length of 679 bp and a maximum length of 5109 bp. Among the unigenes, 40,139 were in the early group, 39,768 were obtained from the full group and 28,316 were detected in both samples. With the threshold of "log2 ratio ≥ 1", there were 34,464 differentially expressed genes, of which 18,043 were up-regulated and 16,421 were down-regulated in the early flower library. Based on the annotations of the unigenes, 281 pathways were predicted. We selected 12 putative genes and analyzed their expression levels using quantitative real time-PCR. The results were consistent with the 454 sequencing results. In addition, the expression of chalcone synthase, chalcone isomerase and anthocyanidin synthase, which are involved in safflor yellow biosynthesis and safflower yellow pigment (SYP) content, were analyzed in different flowering periods, indicating that their expression levels were related to SYP synthesis. Moreover, to further confirm the results of the 454 pyrosequencing, full-length cDNA of chalcone isomerase (CHI) and anthocyanidin synthase (ANS) were cloned from safflower petal by RACE (Rapid-amplification of cDNA ends) method according to fragment of the transcriptome.

  9. Transcriptomic analysis of Camellia ptilophylla and identification of genes associated with flavonoid and caffeine biosynthesis.

    PubMed

    Li, M M; Xue, J Y; Wen, Y L; Guo, H S; Sun, X Q; Zhang, Y M; Hang, Y Y

    2015-12-29

    Camellia ptilophylla, or cocoa tea, is naturally decaffeinated and its predominant catechins and purine alkaloids are trans-catechins and theobromine Regular tea [Camellia sinensis (L.) O. Ktze.] is evolutionarily close to cocoa tea and produces cis-catechins and caffeine. Here, the transcriptome of C. ptilophylla was sequenced using the 101-bp paired-end technique. The quality of the raw data was assessed to yield 70,227,953 cleaned reads totaling 7.09 Gbp, which were assembled de novo into 56,695 unique transcripts and then clustered into 44,749 unigenes. In catechin biosynthesis, leucoanthocyanidin reductase (LAR) catalyzes the transition of leucoanthocyanidin to trans-catechins, while anthocyanidin synthase (ANS) and anthocyanidin reductase (ANR) catalyze cis-catechin production. Our data demonstrate that two LAR genes (CpLAR1 and CpLAR2) by C. ptilophylla may be advantageous due to the combined effects of this quantitative trait, permitting increased leucoanthocyanidin consumption for the synthesis of trans-catechins. In contrast, the only ANS gene observed in C. sinensis (CsANS) shared high identity (99.2%) to one homolog from C. ptilophylla (CpANS1), but lower identity (~80%) to another (CpANS2). We hypothesized that the diverged CpANS2 might have lost its ability to synthesize cis-catechins. C. ptilophylla and C. sinensis each contain two copies of ANR, which share high identity and may share the same function. Transcriptomic sequencing captured two N-methyl nucleosidase genes named NMT1 and NMT2. NMT2 was highly identical to three orthologous genes TCS2, PCS2, and ICS2, which did not undergo methylation in vitro; in contrast, NMT1 was less identical to TCS, PCS and ICS, indicating that NMT1 may undergo neofunctionalization.

  10. SEEDSTICK is a Master Regulator of Development and Metabolism in the Arabidopsis Seed Coat

    PubMed Central

    Paolo, Dario; Rueda-Romero, Paloma; Guerra, Rosalinda Fiorella; Battaglia, Raffaella; Rogachev, Ilana; Aharoni, Asaph; Kater, Martin M.; Caporali, Elisabetta; Colombo, Lucia

    2014-01-01

    The role of secondary metabolites in the determination of cell identity has been an area of particular interest over recent years, and studies strongly indicate a connection between cell fate and the regulation of enzymes involved in secondary metabolism. In Arabidopsis thaliana, the maternally derived seed coat plays pivotal roles in both the protection of the developing embryo and the first steps of germination. In this regard, a characteristic feature of seed coat development is the accumulation of proanthocyanidins (PAs - a class of phenylpropanoid metabolites) in the innermost layer of the seed coat. Our genome-wide transcriptomic analysis suggests that the ovule identity factor SEEDSTICK (STK) is involved in the regulation of several metabolic processes, providing a strong basis for a connection between cell fate determination, development and metabolism. Using phenotypic, genetic, biochemical and transcriptomic approaches, we have focused specifically on the role of STK in PA biosynthesis. Our results indicate that STK exerts its effect by direct regulation of the gene encoding BANYULS/ANTHOCYANIDIN REDUCTASE (BAN/ANR), which converts anthocyanidins into their corresponding 2,3-cis-flavan-3-ols. Our study also demonstrates that the levels of H3K9ac chromatin modification directly correlate with the active state of BAN in an STK-dependent way. This is consistent with the idea that MADS-domain proteins control the expression of their target genes through the modification of chromatin states. STK might thus recruit or regulate histone modifying factors to control their activity. In addition, we show that STK is able to regulate other BAN regulators. Our study demonstrates for the first time how a floral homeotic gene controls tissue identity through the regulation of a wide range of processes including the accumulation of secondary metabolites. PMID:25521508

  11. Chemical composition of Solanum nigrum linn extract and induction of autophagy by leaf water extract and its major flavonoids in AU565 breast cancer cells.

    PubMed

    Huang, Hsiu-Chen; Syu, Kai-Yang; Lin, Jen-Kun

    2010-08-11

    Solanum nigrum Linn (SN) belongs to the Solanaceae family, is a plant growing widely in south Asia, and has been used in traditional folk medicine. It is believed to have antipyretic, diuretic, anticancer, and hepatoprotective effects. During the summertime, this plant has been heavily used to supplement beverages to quench thirst on hot days in Taiwan and several southern Asian countries. In this study, the polyphenols and anthocyanidin in various parts of the SN plant were analyzed by HPLC. The leaves were found to be richer in polyphenols than stem and fruit. SN leaves contained the highest concentration of gentisic acid, luteolin, apigenin, kaempferol, and m-coumaric acid. However, the anthocyanidin existed only in the purple fruits. Additionally, the cytotoxicity of the leaf, stem, or fruit extract was evaluated against cancer cell lines and normal cells. The results showed that AU565 breast cancer cells were more sensitive to the extract. Furthermore, the results demonstrated a significant cytotoxic effect of SN leaf extract on AU565 cells that was mediated via two different mechanisms depending on the exposure concentrations. A low dose of SN leaf extract induced autophagy but not apoptosis. Higher doses (>100 microg/mL) of SN leaf extract could inhibit the level of p-Akt and cause cell death due to the induction of autophagy and apoptosis. However, these findings indicate that SN leaf extract induced cell death in breast cells via two distinct antineoplastic activities, the abilities to induce apoptosis and autophagy, therefore suggesting that it may provide a useful remedy to treat breast cancer. PMID:20681660

  12. Effects of methyl jasmonate on accumulation of flavonoids in seedlings of common buckwheat (Fagopyrum esculentum Moench).

    PubMed

    Horbowicz, M; Wiczkowski, W; Koczkodaj, Danuta; Saniewski, M

    2011-09-01

    The jasmonates, which include jasmonic acid and its methyl ester (MJ), play a central role in regulating the biosynthesis of many secondary metabolites, including flavonoids, and also are signaling molecules in environmental stresses. Synthesis of anthocyanins pigments is a final part of flavonoids pathway route. Accumulation of the pigments in young seedlings is stimulated by various environmental stresses, such as high-intensity light, wounding, pathogen attack, drought, sugar and nutrient deficiency. The anthocyanins take part in defense system against excess of light and UV-B light, and therefore it is probably main reason why young plant tissues accumulate enlarged levels of the pigments. The effects of exogenously applied MJ on level of anthocyanins, glycosides of apigenin, luteolin, quercetin and proanthocyanidins in seedlings of common buckwheat (Fagopyrum esculentum Moench) were studied. MJ decreased contents of all the found cyanidin glycosides and its aglycone in hypocotyls of buckwheat seedlings. However contents of particular anthocyanins in cotyledons of buckwheat seedlings treated with the plant hormone were not significantly different from the control. Applied doses of MJ did not affect levels of quercetin, apigenin and luteolin glycosides in the analyzed parts of buckwheat seedlings: cotyledons and hypocotyls. On the other hand, treatment of buckwheat seedlings with MJ clearly stimulated of proanthocyanidins biosynthesis in hypocotyls. We suggest that methyl jasmonate induces in hypocotyls of buckwheat seedlings the leucocyanidin reductase or anthocyanidin reductase, possible enzymes in proanthocyanidins synthesis, and/or inhibits anthocyanidin synthase, which transforms leucocyanidin into cyanidin. According to our knowledge this is the first report regarding the effect of methyl jasmonate on enhancing the accumulation of proanthocyanidins in cultivated plants.

  13. De Novo Sequencing and Analysis of the Safflower Transcriptome to Discover Putative Genes Associated with Safflor Yellow in Carthamus tinctorius L.

    PubMed

    Liu, Xiuming; Dong, Yuanyuan; Yao, Na; Zhang, Yu; Wang, Nan; Cui, Xiyan; Li, Xiaowei; Wang, Yanfang; Wang, Fawei; Yang, Jing; Guan, Lili; Du, Linna; Li, Haiyan; Li, Xiaokun

    2015-01-01

    Safflower (Carthamus tinctorius L.), an important traditional Chinese medicine, is cultured widely for its pharmacological effects, but little is known regarding the genes related to the metabolic regulation of the safflower's yellow pigment. To investigate genes related to safflor yellow biosynthesis, 454 pyrosequencing of flower RNA at different developmental stages was performed, generating large databases.In this study, we analyzed 454 sequencing data from different flowering stages in safflower. In total, 1,151,324 raw reads and 1,140,594 clean reads were produced, which were assembled into 51,591 unigenes with an average length of 679 bp and a maximum length of 5109 bp. Among the unigenes, 40,139 were in the early group, 39,768 were obtained from the full group and 28,316 were detected in both samples. With the threshold of "log2 ratio ≥ 1", there were 34,464 differentially expressed genes, of which 18,043 were up-regulated and 16,421 were down-regulated in the early flower library. Based on the annotations of the unigenes, 281 pathways were predicted. We selected 12 putative genes and analyzed their expression levels using quantitative real time-PCR. The results were consistent with the 454 sequencing results. In addition, the expression of chalcone synthase, chalcone isomerase and anthocyanidin synthase, which are involved in safflor yellow biosynthesis and safflower yellow pigment (SYP) content, were analyzed in different flowering periods, indicating that their expression levels were related to SYP synthesis. Moreover, to further confirm the results of the 454 pyrosequencing, full-length cDNA of chalcone isomerase (CHI) and anthocyanidin synthase (ANS) were cloned from safflower petal by RACE (Rapid-amplification of cDNA ends) method according to fragment of the transcriptome. PMID:26516840

  14. Antioxidant and antiproliferative activities of anthocyanin/ellagitannin-enriched extracts from Syzygium cumini L. (‘jamun’, the Indian Blackberry)

    PubMed Central

    Aqil, Farrukh; Gupta, Akash; Munagala, Radha; Jeyabalan, Jeyaprakash; Kausar, Hina; Sharma, Ramjee; Singh, Inder Pal; Gupta, Ramesh C.

    2012-01-01

    Colored fruits, particularly berries, are highly chemoprotective because of their antioxidant, anti-proliferative and anti-inflammatory activities. We report cancer chemoprotective potential of Syzygium cumini L., commonly known as ‘jamun’ or Indian blackberry. Anthocyanins and other polyphenolics were extracted with acidic ethanol, and enriched by amberlite XAD7/HP20 (1:1). The pulp powder was found to contain 0.54% anthocyanins, 0.17% ellagic acid/ellagitannins and 1.15% polyphenolics. Jamun seed contained no detectable anthocyanins, but had higher amounts of ellagic acid/ellagitannins (0.5%) and total polyphenolics (2.7%) than the pulp powder. Upon acid hydrolysis, the pulp extract yielded five anthocyanidins by HPLC: malvidin (44.4%), petunidin (24.2%), delphinidin (20.3%), cyanidin (6.6%), and peonidin (2.2%). Extracts of both jamun pulp (1,445±64 μmol of trolox equivalent (TE)/g) and seeds (3,379±151 μM of TE/g) showed high oxygen radical absorbance capacity (ORAC). Their high antioxidant potential was also reflected by 2,2′-azino-bis(3-ethylbenzthiazoline-6-sulphonic acid) (ABTS)- and 2,2-diphenyl-1-picrylhydrazyl (DPPH)-scavenging, and ferrous ion-chelating activities. We also analyzed anti-proliferative activity of jamun extracts against human lung cancer A549 cells. The hydrolyzed pulp and seed extracts showed significant antiproliferative activity. However, unhydrolyzed extracts showed much less activity. These data showed that in addition to five anthocyanidins, jamun contains appreciable amounts of ellagic acid/ellagitannins, with high antioxidant and antiproliferative activities. PMID:22420901

  15. Antioxidant and antiproliferative activities of anthocyanin/ellagitannin-enriched extracts from Syzygium cumini L. (Jamun, the Indian Blackberry).

    PubMed

    Aqil, Farrukh; Gupta, Akash; Munagala, Radha; Jeyabalan, Jeyaprakash; Kausar, Hina; Sharma, Ram Jee; Singh, Inder Pal; Gupta, Ramesh C

    2012-04-01

    Colored fruits, particularly berries, are highly chemoprotective because of their antioxidant, antiproliferative, and antiinflammatory activities. We report the cancer chemoprotective potential of Syzygium cumini L., commonly known as jamun or Indian blackberry. Anthocyanins and other polyphenolics were extracted with acidic ethanol and enriched by amberlite XAD7/HP20 (1:1). The pulp powder was found to contain 0.54% anthocyanins, 0.17% ellagic acid/ellagitannins, and 1.15% total polyphenolics. Jamun seed contained no detectable anthocyanins but had higher amounts of ellagic acid/ellagitannins (0.5%) and total polyphenolics (2.7%) than the pulp powder. Upon acid hydrolysis, the pulp extract yielded 5 anthocyanidins by HPLC: malvidin (44.4%), petunidin (24.2%), delphinidin (20.3%), cyanidin (6.6%), and peonidin (2.2%). Extracts of both jamun pulp (1,445 ± 64 μmol of trolox equivalent (TE)/g) and seeds (3,379 ± 151 μM of TE/g) showed high oxygen radical absorbance capacity. Their high antioxidant potential was also reflected by 2,2'-azino-bis(3-ethylbenzthiazoline-6-sulphonic acid)- and 2,2-diphenyl-1-picrylhydrazyl-scavenging and ferrous ion-chelating activities. We also analyzed antiproliferative activity of jamun extracts against human lung cancer A549 cells. The hydrolyzed pulp and seed extracts showed significant antiproliferative activity. However, unhydrolyzed extracts showed much less activity. These data showed that in addition to 5 anthocyanidins, jamun contains appreciable amounts of ellagic acid/ellagitannins, with high antioxidant and antiproliferative activities. PMID:22420901

  16. Antimicrobial Resistance, Extended-Spectrum β-Lactamase Productivity, and Class 1 Integrons in Escherichia coli from Healthy Swine.

    PubMed

    Changkaew, Kanjana; Intarapuk, Apiradee; Utrarachkij, Fuangfa; Nakajima, Chie; Suthienkul, Orasa; Suzuki, Yasuhiko

    2015-08-01

    Administration of antimicrobials to food-producing animals increases the risk of higher antimicrobial resistance in the normal intestinal flora of these animals. The present cross-sectional study was conducted to investigate antimicrobial susceptibility and extended-spectrum β-lactamase (ESBL)-producing strains and to characterize class 1 integrons in Escherichia coli in healthy swine in Thailand. All 122 of the tested isolates had drug-resistant phenotypes. High resistance was found to ampicillin (98.4% of isolates), chloramphenicol (95.9%), gentamicin (78.7%), streptomycin (77.9%), tetracycline (74.6%), and cefotaxime (72.1%). Fifty-four (44.3%) of the E. coli isolates were confirmed as ESBL-producing strains. Among them, blaCTX-M (45 isolates) and blaTEM (41 isolates) were detected. Of the blaCTX-M-positive E. coli isolates, 37 carried the blaCTX-M-1 cluster, 12 carried the blaCTX-M-9 cluster, and 5 carried both clusters. Sequence analysis revealed blaTEM-1, blaTEM-135, and blaTEM-175 in 38, 2, and 1 isolate, respectively. Eighty-seven (71%) of the 122isolates carried class 1 integrons, and eight distinct drug-resistance gene cassettes with seven different integron profiles were identified in 43 of these isolates. Gene cassettes were associated with resistance to aminoglycosides (aadA1, aadA2, aadA22, or aadA23), trimethoprim (dfrA5, dfrA12, or dfrA17), and lincosamide (linF). Genes encoding β-lactamases were not found in class 1 integrons. This study is the first to report ESBL-producing E. coli with a class 1 integron carrying the linF gene cassette in swine in Thailand. Our findings confirm that swine can be a reservoir of ESBL-producing E. coli harboring class 1 integrons, which may become a potential health risk if these integrons are transmitted to humans. Intensive analyses of animal, human, and environmental isolates are needed to control the spread of ESBL-producing E. coli strains.

  17. Isolation and molecular characterization of multidrug-resistant Enterobacteriaceae strains from pork and environmental samples in Xiamen, China.

    PubMed

    Liu, Zongbao; Zhang, Zhigang; Yan, He; Li, Jianrong; Shi, Lei

    2015-01-01

    This study was conducted to investigate the prevalence and molecular characterization of multidrug-resistant (MDR) Enterobacteriaceae isolated from swine meat and the breeding environment. A total of 102 MDR Enterobacteriaceae strains belonging to five genera were obtained from 210 samples collected from a large-scale swine farm from March 2012 to June 2013 in Xiamen, People's Republic of China. Among these MDR isolates, Escherichia coli strains were found most frequently in both meat and environmental samples, followed by Citrobacter spp., Klebsiella spp., and Shigella spp. The neighbor-joining phylogenetic tree indicated that 70.3 % of Escherichia and 50 % of Citrobacter isolates from meat samples shared 100 % homology with relevant isolates from environmental samples. Resistance was most frequently observed to sulfonamide, trimethoprim, aminoglycoside, chloramphenicol, β-lactam, and tetracycline. Close correlation was noted between antibiotic resistance phenotype and the genes responsible for resistance to sulfonamide (sulI), trimethoprim (dhfrI), aminoglycoside (aadA, aac(3)-I, aphA-1, and aac(3)-IV), chloramphenicol (catI and cmlA), β-lactam (blaSHV, blaOXA, and blaTEM), florfenicol (floR), and tetracycline (tet(A) and tet(B)), which were widely distributed with prevalences of 72.5, 6.9, 62.7, 14.7, 78.4, 11.8, 25.5, 42.2, 12.7, 14.7, 39.2, 87.2, 68.6, and 34.3 % , respectively. Class 1 integrons carrying aadA22, dfrA17-aadA5, or dfrA12-aadA2 cassette arrays were commonly found in isolates from all samples. The gene cassette aac(6')-Ib-cr-arr-3-dfrA27-aadA16 was first found in an Enterobacter amnigenus isolate. Conjugation experiments revealed the plasmid-mediated transfer of class 1 integrons. Our results indicate that swine meat and the farming environment can be sources of antibiotic-resistant bacteria, which could be potentially transmitted to humans via the meat products industry chain.

  18. Identification of integrons and phylogenetic groups of drug-resistant Escherichia coli from broiler carcasses in China.

    PubMed

    Wu, Hao; Xia, Shibo; Bu, Fanyun; Qi, Jing; Liu, Yuqing; Xu, Hai

    2015-10-15

    The dissemination of drug-resistant Escherichia coli in poultry products is becoming a public concern, as it endangers food security and human health. It is very common for E. coli to exhibit drug resistance in the poultry industry in China due to the excessive use of antibiotics. However, few studies have examined the drug resistance endowed by integrons and integron-associated gene cassettes in different phylogenetic groups of E. coli isolated from broiler carcasses. In this study, 373 antibiotic-resistant E. coli strains were isolated from the surfaces or insides of broiler carcasses from a slaughterhouse in Shandong Province, China. According to phylogenetic assays of chuA, yjaA, and an anonymous DNA fragment, TSPE4-C2, these isolates belong to four phylogenetic groups (A, B1, B2, and D) and seven subgroups (A0, A1, B1, B21, B22, D1, and D2). Of the tested isolates, 95.71% (n=357) are multi-drug resistant, among which group B1 was predominant, accounting for 33.51% (n=125) of the tested isolates. A high percentage of the E. coli isolates were resistant to amoxicillin-clavulanic acid (99.20%, n=370), doxycycline (92.23%, n=344), sulfamethoxazole-trimethoprim (90.88%, n=339), ciprofloxacin, (64.61%, n=241), sulbactam-cefoperazone (51.21%, n=191), and amikacin (33.78%, n=126). Furthermore, among the 373 isolates, class 1 and 2 integrons were identified in 292 (78.28%) and 49 (13.14%) of the isolates, respectively, while no class 3 integrons were detected. The most prevalent gene cassette arrays were dfrA17-aadA5 and dfrA12-orfF-aadA2 in the variable region of class 1 integrons, while only one gene cassette array (dfrA1-sat2-aadA1) was detected in the variable region of class 2 integrons. Class 1 integrons were distributed in various physiological subtypes, whereas no predominant phylogenetic groups could be identified. The presence of class 2 integrons in the B21 subtype was significantly higher than in the other subtypes, and it coexisted with the class 1

  19. Isolation and molecular characterization of multidrug-resistant Enterobacteriaceae strains from pork and environmental samples in Xiamen, China.

    PubMed

    Liu, Zongbao; Zhang, Zhigang; Yan, He; Li, Jianrong; Shi, Lei

    2015-01-01

    This study was conducted to investigate the prevalence and molecular characterization of multidrug-resistant (MDR) Enterobacteriaceae isolated from swine meat and the breeding environment. A total of 102 MDR Enterobacteriaceae strains belonging to five genera were obtained from 210 samples collected from a large-scale swine farm from March 2012 to June 2013 in Xiamen, People's Republic of China. Among these MDR isolates, Escherichia coli strains were found most frequently in both meat and environmental samples, followed by Citrobacter spp., Klebsiella spp., and Shigella spp. The neighbor-joining phylogenetic tree indicated that 70.3 % of Escherichia and 50 % of Citrobacter isolates from meat samples shared 100 % homology with relevant isolates from environmental samples. Resistance was most frequently observed to sulfonamide, trimethoprim, aminoglycoside, chloramphenicol, β-lactam, and tetracycline. Close correlation was noted between antibiotic resistance phenotype and the genes responsible for resistance to sulfonamide (sulI), trimethoprim (dhfrI), aminoglycoside (aadA, aac(3)-I, aphA-1, and aac(3)-IV), chloramphenicol (catI and cmlA), β-lactam (blaSHV, blaOXA, and blaTEM), florfenicol (floR), and tetracycline (tet(A) and tet(B)), which were widely distributed with prevalences of 72.5, 6.9, 62.7, 14.7, 78.4, 11.8, 25.5, 42.2, 12.7, 14.7, 39.2, 87.2, 68.6, and 34.3 % , respectively. Class 1 integrons carrying aadA22, dfrA17-aadA5, or dfrA12-aadA2 cassette arrays were commonly found in isolates from all samples. The gene cassette aac(6')-Ib-cr-arr-3-dfrA27-aadA16 was first found in an Enterobacter amnigenus isolate. Conjugation experiments revealed the plasmid-mediated transfer of class 1 integrons. Our results indicate that swine meat and the farming environment can be sources of antibiotic-resistant bacteria, which could be potentially transmitted to humans via the meat products industry chain. PMID:25581181

  20. Prevalence and characteristics of extended-spectrum β-lactamase genes in Escherichia coli isolated from piglets with post-weaning diarrhea in Heilongjiang province, China

    PubMed Central

    Xu, Guofeng; An, Wei; Wang, Hongdong; Zhang, Xiuying

    2015-01-01

    Objectives: The purpose of this study was to investigate the prevalence of extended spectrum β-lactamase (ESBL) genes in Escherichia coli isolated from post-weaning diarrhea (PWD) piglets in Heilongjiang province, China. Methods: Of 458 E. coli isolated from 589 fecal samples from PWD piglets, a total of 198 isolates were confirmed as ESBL producers by the double-disk synergy test (DDST). Polymerase chain reaction (PCR) and sequencing were performed to identify genes for ESBL, plasmid-mediated quinolone resistance (PMQR), and integrons. Results: Of the 198 isolates, blaCTX−M and blaTEM were detected in 191 and 149 isolates, respectively. Sequencing revealed that 10 blaCTX−M subtypes were detected, and blaCTX−M−14 was the most prevalent, followed by blaCTX−M−55 and blaCTX−M−65. Of the 149 TEM-positive strains, four were blaTEM−52 and the rest were blaTEM−1. Among the 198 ESBL-positive isolates, 173 isolates were found to harbor at least one PMQR gene, with oqxAB, qnrS, qnrB, qepA, and aac(6′)-Ib-cr being detected alone or in combination in 125, 114, 26, 24, and 45 strains, respectively. One hundred and fifty-five ESBL-positive isolates were also positive for class I integron (int1), and eight different gene cassette arrays were confirmed in 110 isolates by restriction fragment length polymorphism (RFLP) and DNA sequencing analyses, with predominance of dfrA17-aadA5, dfrA12-orfF-aadA2, and dfrA1-aadA1 arrays. Conclusion: To the best of our knowledge, this is the first report of the blaTEM−52 gene in pig E. coli isolates in China and this is also the first description of the coexistence of the qnrB, qnrS, aac(6′)-Ib-cr, qepA, and oqxAB genes in one E. coli strain. PMID:26500640

  1. Static and Dynamic Postural Changes after a Mountain Ultra-Marathon of 80 km and 5500 D+

    PubMed Central

    Marcolin, Giuseppe; Grainer, Alessandro; Reggiani, Carlo; Bisiacchi, Patrizia; Cona, Giorgia; Petrone, Nicola

    2016-01-01

    The study aimed to investigate the effect of fatigue on static and dynamic postural stability after completing a mountain ultra-marathon. Twelve male athletes participated in the study. Postural stability was assessed before and immediately after the race. Static postural stability was evaluated on a dynamometric platform with eyes opened (OE) and closed (CE). Dynamic postural stability was assessed with OE on an instrumented plate which allowed medio-lateral oscillations. Stabilometric data were affected by fatigue in the OE condition, concerning sway path velocity (p = 0.0006), sway area velocity (p = 0.0006), area of the confidence ellipse (p = 0.0016), maximal anterior-posterior (AP) (p = 0.0017) and medio-lateral (ML) (p = 0.0039) oscillations. In the CE condition the sway path velocity (p = 0.0334), the maximal ML oscillations (p = 0.0161) and the area of the confident ellipse (p = 0.0180) were also negatively influenced. Stabilogram diffusion analysis showed in the OE condition an increase of short-term diffusion coefficients considering the anterior-posterior direction (Dfys; p = 0.0023) and the combination of the two (Dfr2s; p = 0.0032). Equally, long term diffusion coefficients increased considering the anterior-posterior direction (Dfyl; p = 0.0093) and the combination of the two (Dfr2l; p = 0.0086). In CE condition greater values were detected for medio-lateral direction (Dfxl; p = 0.033), anterior-posterior direction (Dfyl; p = 0.0459) and the combination of the two (Dfr2l; p = 0.0048). The dynamic postural stability test showed an increase of the time spent with the edges of the plate on the floor (p = 0.0152). Our results showed that mountain ultra-marathon altered static stability more than dynamic stability. An involvement of cognitive resources to monitor postural stability after fatiguing could be the explanation of the worsening in the automatic task (quiet standing) and of the positive compensation in the less automatic task (dynamic standing

  2. Two LcbHLH Transcription Factors Interacting with LcMYB1 in Regulating Late Structural Genes of Anthocyanin Biosynthesis in Nicotiana and Litchi chinensis During Anthocyanin Accumulation

    PubMed Central

    Lai, Biao; Du, Li-Na; Liu, Rui; Hu, Bing; Su, Wen-Bing; Qin, Yong-Hua; Zhao, Jie-Tang; Wang, Hui-Cong; Hu, Gui-Bing

    2016-01-01

    Anthocyanin biosynthesis requires the MYB-bHLH-WD40 protein complex to activate the late biosynthetic genes. LcMYB1 was thought to act as key regulator in anthocyanin biosynthesis of litchi. However, basic helix-loop-helix proteins (bHLHs) as partners have not been identified yet. The present study describes the functional characterization of three litchi bHLH candidate anthocyanin regulators, LcbHLH1, LcbHLH2, and LcbHLH3. Although these three litchi bHLHs phylogenetically clustered with bHLH proteins involved in anthcoyanin biosynthesis in other plant, only LcbHLH1 and LcbHLH3 were found to localize in the nucleus and physically interact with LcMYB1. The transcription levels of all these bHLHs were not coordinated with anthocyanin accumulation in different tissues and during development. However, when co-infiltrated with LcMYB1, both LcbHLH1 and LcbHLH3 enhanced anthocyanin accumulation in tobacco leaves with LcbHLH3 being the best inducer. Significant accumulation of anthocyanins in leaves transformed with the combination of LcMYB1 and LcbHLH3 were noticed, and this was associated with the up-regulation of two tobacco endogenous bHLH regulators, NtAn1a and NtAn1b, and late structural genes, like NtDFR and NtANS. Significant activity of the ANS promoter was observed in transient expression assays either with LcMYB1-LcbHLH1 or LcMYB1-LcbHLH3, while only minute activity was detected after transformation with only LcMYB1. In contrast, no activity was measured after induction with the combination of LcbHLH2 and LcMYB1. Higher DFR expression was also oberseved in paralleling with higher anthocyanins in co-transformed lines. LcbHLH1 and LcbHLH3 are essential partner of LcMYB1 in regulating the anthocyanin production in tobacco and probably also in litchi. The LcMYB1-LcbHLH complex enhanced anthocyanin accumulation may associate with activating the transcription of DFR and ANS. PMID:26925082

  3. Identification of integrons and phylogenetic groups of drug-resistant Escherichia coli from broiler carcasses in China.

    PubMed

    Wu, Hao; Xia, Shibo; Bu, Fanyun; Qi, Jing; Liu, Yuqing; Xu, Hai

    2015-10-15

    The dissemination of drug-resistant Escherichia coli in poultry products is becoming a public concern, as it endangers food security and human health. It is very common for E. coli to exhibit drug resistance in the poultry industry in China due to the excessive use of antibiotics. However, few studies have examined the drug resistance endowed by integrons and integron-associated gene cassettes in different phylogenetic groups of E. coli isolated from broiler carcasses. In this study, 373 antibiotic-resistant E. coli strains were isolated from the surfaces or insides of broiler carcasses from a slaughterhouse in Shandong Province, China. According to phylogenetic assays of chuA, yjaA, and an anonymous DNA fragment, TSPE4-C2, these isolates belong to four phylogenetic groups (A, B1, B2, and D) and seven subgroups (A0, A1, B1, B21, B22, D1, and D2). Of the tested isolates, 95.71% (n=357) are multi-drug resistant, among which group B1 was predominant, accounting for 33.51% (n=125) of the tested isolates. A high percentage of the E. coli isolates were resistant to amoxicillin-clavulanic acid (99.20%, n=370), doxycycline (92.23%, n=344), sulfamethoxazole-trimethoprim (90.88%, n=339), ciprofloxacin, (64.61%, n=241), sulbactam-cefoperazone (51.21%, n=191), and amikacin (33.78%, n=126). Furthermore, among the 373 isolates, class 1 and 2 integrons were identified in 292 (78.28%) and 49 (13.14%) of the isolates, respectively, while no class 3 integrons were detected. The most prevalent gene cassette arrays were dfrA17-aadA5 and dfrA12-orfF-aadA2 in the variable region of class 1 integrons, while only one gene cassette array (dfrA1-sat2-aadA1) was detected in the variable region of class 2 integrons. Class 1 integrons were distributed in various physiological subtypes, whereas no predominant phylogenetic groups could be identified. The presence of class 2 integrons in the B21 subtype was significantly higher than in the other subtypes, and it coexisted with the class 1

  4. Two LcbHLH Transcription Factors Interacting with LcMYB1 in Regulating Late Structural Genes of Anthocyanin Biosynthesis in Nicotiana and Litchi chinensis During Anthocyanin Accumulation.

    PubMed

    Lai, Biao; Du, Li-Na; Liu, Rui; Hu, Bing; Su, Wen-Bing; Qin, Yong-Hua; Zhao, Jie-Tang; Wang, Hui-Cong; Hu, Gui-Bing

    2016-01-01

    Anthocyanin biosynthesis requires the MYB-bHLH-WD40 protein complex to activate the late biosynthetic genes. LcMYB1 was thought to act as key regulator in anthocyanin biosynthesis of litchi. However, basic helix-loop-helix proteins (bHLHs) as partners have not been identified yet. The present study describes the functional characterization of three litchi bHLH candidate anthocyanin regulators, LcbHLH1, LcbHLH2, and LcbHLH3. Although these three litchi bHLHs phylogenetically clustered with bHLH proteins involved in anthcoyanin biosynthesis in other plant, only LcbHLH1 and LcbHLH3 were found to localize in the nucleus and physically interact with LcMYB1. The transcription levels of all these bHLHs were not coordinated with anthocyanin accumulation in different tissues and during development. However, when co-infiltrated with LcMYB1, both LcbHLH1 and LcbHLH3 enhanced anthocyanin accumulation in tobacco leaves with LcbHLH3 being the best inducer. Significant accumulation of anthocyanins in leaves transformed with the combination of LcMYB1 and LcbHLH3 were noticed, and this was associated with the up-regulation of two tobacco endogenous bHLH regulators, NtAn1a and NtAn1b, and late structural genes, like NtDFR and NtANS. Significant activity of the ANS promoter was observed in transient expression assays either with LcMYB1-LcbHLH1 or LcMYB1-LcbHLH3, while only minute activity was detected after transformation with only LcMYB1. In contrast, no activity was measured after induction with the combination of LcbHLH2 and LcMYB1. Higher DFR expression was also oberseved in paralleling with higher anthocyanins in co-transformed lines. LcbHLH1 and LcbHLH3 are essential partner of LcMYB1 in regulating the anthocyanin production in tobacco and probably also in litchi. The LcMYB1-LcbHLH complex enhanced anthocyanin accumulation may associate with activating the transcription of DFR and ANS. PMID:26925082

  5. Diffraction Ellipsometry Studies on Insect Flight Muscle

    NASA Astrophysics Data System (ADS)

    Shen, Sui

    Characterization of the orientation and distribution of myosin cross-bridge at rigor, relax, low ionic strength (36 mM) and activation (pCa 4.3) conditions are of great interest since these states have been proposed to be transient steps in the cyclical interaction of myosin heads with actin during contraction. Measurements sensitive to the cross-bridge orientation in chemically skinned single muscle fibers of the insect, Lethocerus collossicus have been performed under various physiological conditions using laser diffraction ellipsometry. Determination of both the total birefringence, Deltan, and the differential field ratio, rm DFR (defined as {E_parallel -E_|over E_parallel-E _|}),is necessary for complete characterization of the optical polarization state. For rigor insect fiber, the birefringence value was close to the value we obtained from chemically skinned frog muscle fibers. However, the differential field ratio, DFR, was a negative value for insect fiber, while we always measured a positive value from frog muscle fibers. Polarization states of light diffracted from fibers exhibited a dependence on configurations of structural proteins at different conditions: fluid index matching using o-toluidine, alpha -chymotrypsin cleavage, KCl myosin extraction, rigor state, relaxed state, exogenous S-1 binding on rigor fiber, low ionic strength state, activation state at resting or stretched length. Results of our data analysis suggested that: (1) the negative DFR value of the insect flight muscle was contributed by alpha-actinin arranged perpendicular to the fiber axis in the Z-line, (2) in rigor fiber, 70% of myosin heads are doubly bound (45^circ and 90^ circ) while the rest of 30% are in single head binding configuration (90^circ), (3) myosin heads are randomly oriented in relaxed fiber, (4) mean axial angle is about 62^ circ for exogenous myosin heads binding on rigor fiber, (5) at low ionic strength, 25% of the total myosin heads are weakly attached to actin

  6. Prevalence and characterization of extended-spectrum beta-lactamase (ESBL)- and CMY-2-producing Escherichia coli isolates from healthy food-producing animals in Tunisia.

    PubMed

    Ben Sallem, Rym; Ben Slama, Karim; Sáenz, Yolanda; Rojo-Bezares, Beatriz; Estepa, Vanesa; Jouini, Ahlem; Gharsa, Haythem; Klibi, Naouel; Boudabous, Abdellatif; Torres, Carmen

    2012-12-01

    The prevalence of extended-spectrum beta-lactamase (ESBL)- and plasmidic AmpC-beta-lactamase (pAmpC-BL)-producing Escherichia coli isolates has been studied in food-producing animals at the farm level in Tunisia, and recovered isolates were characterized for the presence of other resistance genes and integrons. Eighty fecal samples of food-producing animals (23 sheep, 22 chickens, 22 cattle, six horses, five rabbits, and two dromedaries) were obtained from 35 different farms in Tunisia in 2011. Samples were inoculated onto MacConkey agar plates supplemented with cefotaxime (2 mg/L) for cefotaxime-resistant (CTX(R)) E. coli recovery. CTX(R) E. coli isolates were detected in 11 out of 80 samples (13.8%), and one isolate per sample was further characterized (10 from chickens and one from a dromedary). The 11 CTX(R) isolates were distributed into phylogroups: B1 (five isolates), A (two isolates), D (three isolates), and B2 (one isolate). The following beta-lactamase genes were detected: bla(CTX-M-1) (seven isolates), bla(CTX-M-1)+bla(TEM-135) (one isolate), bla(CTX-M-1)+bla(TEM-1b) (one isolate), and bla(CMY-2) (two isolates). All ESBL- and pAmpC-BL-producing E. coli strains showed unrelated pulsed-field gel electrophoresis patterns. Seven isolates contained class 1 integrons with four gene cassette arrangements: dfrA17-aadA5 (three isolates), dfrA1-aadA1 (two isolates), dfrA15-aadA1 (one isolate), and aadA1 (one isolate). All isolates showed tetracycline resistance and contained the tet(A) +/- tet(B) genes. Virulence genes detected were as follows (number of isolates in parentheses): fimA (10); aer (eight); papC (two); and papGIII, hly, cnf, and bfp (none). Chicken farms constitute a reservoir of ESBL- and pAmpC-BL-producing E. coli isolates of the CTX-M-1 and CMY-2 types that potentially could be transmitted to humans via the food chain or by direct contact.

  7. [Expression of the genes involved in anthocyanin biosynthesis of 'Tsuda' turnip].

    PubMed

    Xu, Zhi-Ru; Li, Yu-Hua

    2006-10-01

    'Tsuda' turnip (Brassica campestris L. ssp. rapa), in which roots anthocyanin pigmentation is light-sensitive, was used as the material. 'Tsuda' plants were held in darkness or irradiated with sun light and constant light for different time. Anthocyanins in root peel of 'Tsuda' turnip exposed to constant light were identified and quantified with a UV-visual spectrophotometer. The results demonstrated that the anthocyanins accumulation in 'Tsuda' was related with light-exposure time (Fig.1 and Table 1). Fragments of genes selected from the subtraction library of 'Tsuda' turnip involved in anthocyanin biosynthesis were used as probes. The Northern blotting results showed that the expression of PAL, CHS, F3H, DFR and ANS could be induced by irradiation with light and the expression of these genes was related with light exposure time. The expression of MYB was basically the same whether in darkness or in light (Figs.2,3).

  8. Integron, Plasmid and Host Strain Characteristics of Escherichia coli from Humans and Food Included in the Norwegian Antimicrobial Resistance Monitoring Programs

    PubMed Central

    Sunde, Marianne; Simonsen, Gunnar Skov; Slettemeås, Jannice Schau; Böckerman, Inger; Norström, Madelaine

    2015-01-01

    Antimicrobial resistant Escherichia coli (n=331) isolates from humans with bloodstream infections were investigated for the presence of class 1 and class 2 integrons. The integron cassettes arrays were characterized and the findings were compared with data from similar investigations on resistant E. coli from meat and meat products (n=241) produced during the same time period. All isolates were obtained from the Norwegian monitoring programs for antimicrobial resistance in human pathogens and in the veterinary sector. Methods used included PCR, sequencing, conjugation experiments, plasmid replicon typing and subtyping, pulsed-field-gel-electrophoresis and serotyping. Integrons of class 1 and 2 occurred significantly more frequently among human isolates; 45.4% (95% CI: 39.9-50.9) than among isolates from meat; 18% (95% CI: 13.2 -23.3), (p<0.01, Chi-square test). Identical cassette arrays including dfrA1-aadA1, aadA1, dfrA12-orfF-aadA2, oxa-30-aadA1 (class 1 integrons) and dfrA1-sat1-aadA1 (class 2 integrons) were detected from both humans and meat. However, the most prevalent cassette array in human isolates, dfrA17-aadA5, did not occur in isolates from meat, suggesting a possible linkage between this class 1 integron and a subpopulation of E. coli adapted to a human host. The drfA1-aadA1 and aadA1 class 1 integrons were found frequently in both human and meat isolates. These isolates were subjected to further studies to investigate similarities with regard to transferability, plasmid and host strain characteristics. We detected incF plasmids with pMLST profile F24:A-:B1 carrying drfA1-aadA1 integrons in isolates from pork and in a more distantly related E. coli strain from a human with septicaemia. Furthermore, we showed that most of the class 1 integrons with aadA1 were located on incF plasmids with pMLST profile F51:A-:B10 in human isolates. The plasmid was present in unrelated as well as closely related host strains, demonstrating that dissemination of this

  9. First Report of bla(IMP-8) in Raoultella planticola.

    PubMed

    Tseng, Sung-Pin; Wang, Jann-Tay; Liang, Chih-Yuan; Lee, Pei-Shan; Chen, Yee-Chun; Lu, Po-Liang

    2014-01-01

    Two carbapenem-resistant Raoultella planticola clinical isolates were isolated from patients with pneumonia and Port-A catheter-related bacteremia, respectively, in Taiwan. These isolates remained susceptible to fluoroquinolone, aminoglycoside, and colistin. Though the two isolates had the same antibiogram, plasmidic carbapenemase blaIMP-8, class 1 integron cassette (dfrA12-orfF-aadA2), and qnrB2, they had different pulsed-field gel electrophoresis patterns, plasmid sizes, and outer membrane protein loss profiles. To our knowledge, this is the first report of blaIMP-8 found in R. planticola. Interestingly, blaIMP-8 is the most common carbapenemase found in Klebsiella pneumoniae in Taiwan. In the literature, carbapenemase genes in R. planticola in each country were also found in carbapenem-resistant Enterobacteriaceae in the same country.

  10. IncA/C Plasmid Carrying bla(NDM-1), bla(CMY-16), and fosA3 in a Salmonella enterica Serovar Corvallis Strain Isolated from a Migratory Wild Bird in Germany.

    PubMed

    Villa, L; Guerra, B; Schmoger, S; Fischer, J; Helmuth, R; Zong, Z; García-Fernández, A; Carattoli, A

    2015-10-01

    A Salmonella enterica serovar Corvallis strain was isolated from a wild bird in Germany. This strain carried the IncA/C2 pRH-1238 plasmid. Complete sequencing of the plasmid was performed, identifying the blaNDM-1, blaCMY-16, fosA3, sul1, sul2, strA, strB, aac(6')-Ib, aadA5, aphA6, tetA(A), mphA, floR, dfrA7, and merA genes, which confer clinically relevant resistance to most of the antimicrobial classes, including β-lactams with carbapenems, fosfomycin, aminoglycosides, co-trimoxazole, tetracyclines, and macrolides. The strain likely originated from the Asiatic region and was transferred to Germany through the Milvus migrans migratory route. PMID:26169417

  11. Biofunctional properties of Eruca sativa Miller (rocket salad) hydroalcoholic extract.

    PubMed

    Sultan, Khushbakht; Zakir, Muhammad; Khan, Haroon; Rauf, Abdur; Akber, Noor Ul; Khan, Murad Ali

    2016-01-01

    Eruca sativa Miller is a worldwide common alimentary plant (rocket leaves). The aim of this study was to correlate the potential in vitro scavenging activity of the E. sativa hydroalcoholic extract (HAE) with its in vivo hypoglycaemic effect. In DDPH free radical (DFR) and ferric-reducing antioxidant power assays, HAE in a concentration dependent manner (25-100 μg/mL) displayed a strong scavenging activity with maximum effect of 88% and 75% at 100 μg/mL, respectively. Daily administration of HAE (50 mg/kg; p.o.) in the in vivo model of alloxan-induced diabetic rabbits for 28 days showed significant reduction in glycaemia, also supported by recovery of body weight. In conclusion, our results give preliminary information on the potential use of this plant as a nutraceutical, useful to control and/or prevent a hyperglycaemic status.

  12. Transgenic potato plants with overexpression of dihydroflavonol reductase can serve as efficient nutrition sources.

    PubMed

    Kostyn, Kamil; Szatkowski, Michal; Kulma, Anna; Kosieradzka, Iwona; Szopa, Jan

    2013-07-10

    Potato (Solanum tuberosum) is considered to be one of the most important crops cultivated in Europe and the entire world. The tubers of the potato are characterized by rich starch and protein contents and high concentrations of antioxidants, such as vitamin C and flavonoids. Notably, the presence of the phenolic antioxidants is of high importance as they have health-related properties. They are known to reduce the incidence of atherosclerosis, prevent certain kinds of cancer, and aid with many other kinds of diseases. The aim of this study was to find the most efficient way to increase the content of phenolic antioxidants in potato tubers through transgenesis. The results showed that the most efficacious way to achieve this goal was the overexpression of the dihydroflavonol reductase gene (DFR). The produced transgenic potato plants served as a nutrition source for laboratory rats; the study has confirmed their nontoxicity and nutritional benefits on the tested animals.

  13. Enhancing the natural folate level in wine using bioengineering and stabilization strategies.

    PubMed

    Liu, Yazheng; Walkey, Christopher J; Green, Timothy J; van Vuuren, Hennie J J; Kitts, David D

    2016-03-01

    Folate deficiency is linked to many diseases, some of which may have higher probability in individuals with alcohol-induced alterations in one-carbon metabolism. Our study shows that folate content in commercial wine is not related to white or red varieties, but associated with the yeast that is used to produce the wine. The stability of folate in these wines, once opened for consumption, did not correlate with total phenolic or sulfite content. In addition, we employed yeast bioengineering to fortify wine with folate. We confirmed by overexpression that FOL2 was the key gene encoding the rate-limiting step of folate biosynthesis in wine yeast. In this study, we also show that overexpression of other folate biosynthesis genes, including ABZ1, ABZ2, DFR1, FOL1 and FOL3, had no effect on folate levels in wine. Ensuring stability of the increased natural folate in all wines was achieved by the addition of ascorbate.

  14. Environmental Isolate of Rahnella aquatilis Harbors Class 1 Integron.

    PubMed

    Koczura, Ryszard; Mokracka, Joanna; Makowska, Nicoletta

    2016-01-01

    The paper presents first description of class 1 integron in an environmental strain of Rahnella aquatilis, a rarely isolated Gram-negative bacterium of the family Enterobacteriaceae. The strain was isolated from the Warta river water, Poland. Class 1 integrase gene was detected by a PCR assay. Sequencing of the integron's variable region showed the presence of a dfrA1-aadA1 gene cassette array. The integron was located in a 54-kbp plasmid that was transferable to Escherichia coli J-53 recipient strain in a conjugation assay. The integron-bearing R. aquatilis strain was resistant to aminoglycosides, penicillins, trimethoprim, sulfamethoxazole, and trimethoprim/sulfamethoxazole. This paper confirms that water environment play a major role in the spread of integrons and, consequently, antimicrobial resistance, among bacteria of various genera.

  15. Characterization of antimicrobial resistance in Klebsiella species isolated from chicken broilers.

    PubMed

    Wu, Hao; Wang, Mingyu; Liu, Yuqing; Wang, Xinhua; Wang, Yunkun; Lu, Jinxing; Xu, Hai

    2016-09-01

    The prevalence of antimicrobial resistant Klebsiella pneumoniae in poultry products has been a public concern, as it severely endangers food safety and human health. In this study, we investigated 90 antimicrobial resistant Klebsiella strains that were isolated from a commercial broiler slaughter plant in Shandong province of China. Nearly all (89/90) of the isolates were identified as infectious phylogenetic group KpI-type K. pneumoniae. Out of these 90 strains, 87 (96.7%) were multidrug-resistant isolates, and 87 (96.7%) were extended-spectrum beta-lactamase (ESBL)-producing isolates. An analysis of the prevalence of quinolone resistance genes showed that 7.8%, 77.8%, 26.7%, and 2.2% of the strains carried the qnrA, qnrB, qnrS, and qepA genes, respectively. An analysis of beta-lactam resistance genes showed that a high percentage of the strains contain the blaTEM (76.7%), blaSHV (88.9%), and blaCTX-M (75.6%) genes, among which three blaSHV subtypes (blaSHV-1, n=30; blaSHV-11, n=38; blaSHV-12, n=12) and three blaCTX-M subtypes (blaCTX-M-14, n=14; blaCTX-M-15, n=35; blaCTX-M-55, n=19) were found. A further investigation of mobile genetic elements involved in horizontal multidrug resistance gene transfer showed the presence of class 1 and 2 integrons in 77 (85.6%) and five (5.6%) isolates, respectively, while no class 3 integrons were detected. Four types of class 1 integrons containing specific gene cassette arrays (dfrA12-orfF-aadA2, dfrA17-aadA5, dfrA1-aadA1, and empty) were identified. Only one gene cassette array (dfrA1-sat2-aadA1) was detected in the class 2 integrons. Furthermore, four different types of insertion sequence common region 1 (ISCR1)-mediated downstream structures were successfully identified in 46 class 1 integron-positive isolates, among which ISCR1-sapA-like-qnrB2-qacEΔ1 was the most commonly observed structure. Chi-square tests revealed a significant association between ESBL genes, plasmid-mediated quinolone resistance (PMQR) genes, and

  16. Transgenic potato plants with overexpression of dihydroflavonol reductase can serve as efficient nutrition sources.

    PubMed

    Kostyn, Kamil; Szatkowski, Michal; Kulma, Anna; Kosieradzka, Iwona; Szopa, Jan

    2013-07-10

    Potato (Solanum tuberosum) is considered to be one of the most important crops cultivated in Europe and the entire world. The tubers of the potato are characterized by rich starch and protein contents and high concentrations of antioxidants, such as vitamin C and flavonoids. Notably, the presence of the phenolic antioxidants is of high importance as they have health-related properties. They are known to reduce the incidence of atherosclerosis, prevent certain kinds of cancer, and aid with many other kinds of diseases. The aim of this study was to find the most efficient way to increase the content of phenolic antioxidants in potato tubers through transgenesis. The results showed that the most efficacious way to achieve this goal was the overexpression of the dihydroflavonol reductase gene (DFR). The produced transgenic potato plants served as a nutrition source for laboratory rats; the study has confirmed their nontoxicity and nutritional benefits on the tested animals. PMID:23692339

  17. Detection of blaCTX-M-type genes in complex class 1 integrons carried by Enterobacteriaceae isolated from retail chicken meat in Brazil.

    PubMed

    Casella, Tiago; Rodríguez, María Margarita; Takahashi, Juliana Tiemi; Ghiglione, Barbara; Dropa, Milena; Assunção, Ednei; Nogueira, Maurício Lacerda; Lincopan, Nilton; Gutkind, Gabriel; Nogueira, Mara Corrêa Lelles

    2015-03-16

    CTX-M-type extended-spectrum β-lactamase (ESBL)-producing Enterobacteriaceae have been increasingly identified in humans and animals, and their potential transmission by contaminated food has been highlighted. In this study, we report for the first time the isolation of multidrug-resistant (MDR) Escherichia coli, Klebsiella pneumoniae and Proteus mirabilis strains harboring blaCTXM-2 or blaCTXM-8 gene variants in chicken meat sold in markets in southeast Brazil. In this regard, the genetic environment of the blaCTX-M-2 gene is composed of a complex class 1 integron and an ISCR1-associated sequence with dfr and/or aadA gene cassettes located within the variable region. In summary, chicken meat may be a reservoir of MDR Enterobacteriaceae harboring blaCTX-M-type genes, which is a public health concern.

  18. Complete Sequences of Multidrug Resistance Plasmids Bearing rmtD1 and rmtD2 16S rRNA Methyltransferase Genes.

    PubMed

    Bueno, Maria Fernanda C; Francisco, Gabriela R; de Oliveira Garcia, Doroti; Doi, Yohei

    2016-01-04

    Complete nucleotide sequences were determined for two plasmids bearing rmtD group 16S rRNA methyltransferase genes. pKp64/11 was 78 kb in size, belonged to the IncL/M group, and harbored blaTEM-1b, sul1, qacEΔ1, dfrA22, and rmtD1 across two multidrug resistance regions (MRRs). pKp368/10 was 170 kb in size, belonged to the IncA/C group, and harbored acrB, sul1, qacEΔ1, ant(3″)-Ia, aac(6')-Ib, cat, rmtD2, and blaCTX-M-8 across three MRRs. The rmtD-containing regions shared a conserved motif, suggesting a common origin for the two rmtD alleles.

  19. LMFBR operational and experimental local-fault experience, primarily with oxide fuel elements

    SciTech Connect

    Warinner, D.K.

    1980-01-01

    Case-by-case reviews of selective world experience with severe local faults, particularly fuel failure and fuel degradation, are reviewed for two sodium-cooled thermal reactors, several LMFBRs, and LMFBR-fuels experiments. The review summarizes fuel-failure frequency and illustrates the results of the most damaging LMFBR local-fault experiences of the last 20 years beginning with BR-5 and including DFR, BOR-60, BR2's MFBS- and Mol-loops experiments, Fermi, KNK, Rapsodie, EBR-II, and TREAT-D2. Local-fault accommodation is demonstrated and a need to more thoroughly investigate delayed-neutron and gaseous-fission-product signals is highlighted in view of uranate formation, observed blockages, and slow fuel-element failure-propagation.

  20. Detection of blaCTX-M-type genes in complex class 1 integrons carried by Enterobacteriaceae isolated from retail chicken meat in Brazil.

    PubMed

    Casella, Tiago; Rodríguez, María Margarita; Takahashi, Juliana Tiemi; Ghiglione, Barbara; Dropa, Milena; Assunção, Ednei; Nogueira, Maurício Lacerda; Lincopan, Nilton; Gutkind, Gabriel; Nogueira, Mara Corrêa Lelles

    2015-03-16

    CTX-M-type extended-spectrum β-lactamase (ESBL)-producing Enterobacteriaceae have been increasingly identified in humans and animals, and their potential transmission by contaminated food has been highlighted. In this study, we report for the first time the isolation of multidrug-resistant (MDR) Escherichia coli, Klebsiella pneumoniae and Proteus mirabilis strains harboring blaCTXM-2 or blaCTXM-8 gene variants in chicken meat sold in markets in southeast Brazil. In this regard, the genetic environment of the blaCTX-M-2 gene is composed of a complex class 1 integron and an ISCR1-associated sequence with dfr and/or aadA gene cassettes located within the variable region. In summary, chicken meat may be a reservoir of MDR Enterobacteriaceae harboring blaCTX-M-type genes, which is a public health concern. PMID:25576985

  1. Physiological factors affecting transcription of genes involved in the flavonoid biosynthetic pathway in different rice varieties.

    PubMed

    Chen, Xiaoqiong; Itani, Tomio; Wu, Xianjun; Chikawa, Yuuki; Irifune, Kohei

    2013-01-01

    Flavonoids play an important role in the grain color and flavor of rice. Since their characterization in maize, the flavonoid biosynthetic genes have been extensively studied in grape, Arabidopsis, and Petunia. However, we are still a long way from understanding the molecular features and mechanisms underlying the flavonoid biosynthetic pathway. The present study was undertaken to understand the physiological factors affecting the transcription and regulation of these genes. We report that the expression of CHI, CHS, DFR, LAR, and ANS, the 5 flavonoid biosynthetic genes in different rice varieties, differ dramatically with respect to the stage of development, white light, and sugar concentrations. We further demonstrate that white light could induce the transcription of the entire flavonoid biosynthetic gene pathway; however, differences were observed in the degrees of sensitivity and the required illumination time. Our study provides valuable insights into understanding the regulation of the flavonoid biosynthetic pathway. PMID:24389954

  2. Genetic and environmental effects influencing fruit colour and QTL analysis in raspberry.

    PubMed

    McCallum, Susan; Woodhead, Mary; Hackett, Christine A; Kassim, Angzzas; Paterson, Alistair; Graham, Julie

    2010-08-01

    Raspberry (Rubus idaeus) fruit colour was assessed in the Latham x Glen Moy mapping population using a colour meter and visual scores over three seasons and three environments. The colour measurements were found to be significantly associated with pigment content, have high heritability, and stable QTL were identified across environments and seasons. Anthocyanin content has previously been shown to be the major contributor to fruit colour in red raspberry. Major structural genes (F3'H, FLS, DFR, IFR, OMT and GST) and transcription factors (bZIP, bHLH and MYB) influencing flavonoid biosynthesis have been identified, mapped and shown to underlie QTL for quantitative and qualitative anthocyanin composition. Favourable alleles for the selected traits were identified for the aspects of fruit colour and partitioning of individual pigments.

  3. Complete Sequences of Multidrug Resistance Plasmids Bearing rmtD1 and rmtD2 16S rRNA Methyltransferase Genes

    PubMed Central

    Bueno, Maria Fernanda C.; Francisco, Gabriela R.; de Oliveira Garcia, Doroti

    2016-01-01

    Complete nucleotide sequences were determined for two plasmids bearing rmtD group 16S rRNA methyltransferase genes. pKp64/11 was 78 kb in size, belonged to the IncL/M group, and harbored blaTEM-1b, sul1, qacEΔ1, dfrA22, and rmtD1 across two multidrug resistance regions (MRRs). pKp368/10 was 170 kb in size, belonged to the IncA/C group, and harbored acrB, sul1, qacEΔ1, ant(3″)-Ia, aac(6′)-Ib, cat, rmtD2, and blaCTX-M-8 across three MRRs. The rmtD-containing regions shared a conserved motif, suggesting a common origin for the two rmtD alleles. PMID:26729503

  4. Effective route maintenance and restoration schemes in mobile ad hoc networks.

    PubMed

    Kang, Byung-Seok; Ko, In-Young

    2010-01-01

    This study proposes a location-based hybrid routing protocol to improve data packet delivery and to reduce control message overhead in mobile ad hoc networks. In mobile environments, where nodes move continuously at a high speed, it is generally difficult to maintain and restore route paths. Therefore, this study suggests a new flooding mechanism to control route paths. The essence of the proposed scheme is its effective tracking of the destination's location based on the beacon messages of the main route nodes. Through experiments based on an NS-2 simulator, the proposed scheme shows improvements in the data packet delivery ratio and reduces the amount of routing control message overhead compared with existing routing protocols such as AODV, LAR, ZRP and AODV-DFR.

  5. [Expression of the genes involved in anthocyanin biosynthesis of 'Tsuda' turnip].

    PubMed

    Xu, Zhi-Ru; Li, Yu-Hua

    2006-10-01

    'Tsuda' turnip (Brassica campestris L. ssp. rapa), in which roots anthocyanin pigmentation is light-sensitive, was used as the material. 'Tsuda' plants were held in darkness or irradiated with sun light and constant light for different time. Anthocyanins in root peel of 'Tsuda' turnip exposed to constant light were identified and quantified with a UV-visual spectrophotometer. The results demonstrated that the anthocyanins accumulation in 'Tsuda' was related with light-exposure time (Fig.1 and Table 1). Fragments of genes selected from the subtraction library of 'Tsuda' turnip involved in anthocyanin biosynthesis were used as probes. The Northern blotting results showed that the expression of PAL, CHS, F3H, DFR and ANS could be induced by irradiation with light and the expression of these genes was related with light exposure time. The expression of MYB was basically the same whether in darkness or in light (Figs.2,3). PMID:17075183

  6. Diode laser using narrow bandwidth interference filter at 852 nm and its application in Faraday anomalous dispersion optical filter

    NASA Astrophysics Data System (ADS)

    Jiang, Zhaojie; Zhou, Qi; Tao, Zhiming; Zhang, Xiaogang; Zhang, Shengnan; Zhu, Chuanwen; Lin, Pingwei; Chen, Jingbiao

    2016-08-01

    We demonstrate an 852-nm external cavity diode laser (ECDL) system whose wavelength is mainly determined by an interference filter instead of other wavelength selective elements. The Lorentzian linewidth measured by the heterodyne beating between two identical lasers is 28.3 kHz. Moreover, we test the application of the ECDL in the Faraday atomic filter. Besides saturated absorption spectrum, the transmission spectrum of the Faraday atomic filter at 852 nm is measured by using the ECDL. This interference filter ECDL method can also be extended to other wavelengths and widen the application range of diode laser. Project supported by the National Natural Science Foundation of China (Grant No. 91436210) and the International Science and Technology Cooperation Program of China (Grant No. 2010DFR10900).

  7. Effective Route Maintenance and Restoration Schemes in Mobile Ad Hoc Networks

    PubMed Central

    Kang, Byung-Seok; Ko, In-Young

    2010-01-01

    This study proposes a location-based hybrid routing protocol to improve data packet delivery and to reduce control message overhead in mobile ad hoc networks. In mobile environments, where nodes move continuously at a high speed, it is generally difficult to maintain and restore route paths. Therefore, this study suggests a new flooding mechanism to control route paths. The essence of the proposed scheme is its effective tracking of the destination’s location based on the beacon messages of the main route nodes. Through experiments based on an NS-2 simulator, the proposed scheme shows improvements in the data packet delivery ratio and reduces the amount of routing control message overhead compared with existing routing protocols such as AODV, LAR, ZRP and AODV-DFR. PMID:22315570

  8. Integron, Plasmid and Host Strain Characteristics of Escherichia coli from Humans and Food Included in the Norwegian Antimicrobial Resistance Monitoring Programs.

    PubMed

    Sunde, Marianne; Simonsen, Gunnar Skov; Slettemeås, Jannice Schau; Böckerman, Inger; Norström, Madelaine

    2015-01-01

    Antimicrobial resistant Escherichia coli (n=331) isolates from humans with bloodstream infections were investigated for the presence of class 1 and class 2 integrons. The integron cassettes arrays were characterized and the findings were compared with data from similar investigations on resistant E. coli from meat and meat products (n=241) produced during the same time period. All isolates were obtained from the Norwegian monitoring programs for antimicrobial resistance in human pathogens and in the veterinary sector. Methods used included PCR, sequencing, conjugation experiments, plasmid replicon typing and subtyping, pulsed-field-gel-electrophoresis and serotyping. Integrons of class 1 and 2 occurred significantly more frequently among human isolates; 45.4% (95% CI: 39.9-50.9) than among isolates from meat; 18% (95% CI: 13.2 -23.3), (p<0.01, Chi-square test). Identical cassette arrays including dfrA1-aadA1, aadA1, dfrA12-orfF-aadA2, oxa-30-aadA1 (class 1 integrons) and dfrA1-sat1-aadA1 (class 2 integrons) were detected from both humans and meat. However, the most prevalent cassette array in human isolates, dfrA17-aadA5, did not occur in isolates from meat, suggesting a possible linkage between this class 1 integron and a subpopulation of E. coli adapted to a human host. The drfA1-aadA1 and aadA1 class 1 integrons were found frequently in both human and meat isolates. These isolates were subjected to further studies to investigate similarities with regard to transferability, plasmid and host strain characteristics. We detected incF plasmids with pMLST profile F24:A-:B1 carrying drfA1-aadA1 integrons in isolates from pork and in a more distantly related E. coli strain from a human with septicaemia. Furthermore, we showed that most of the class 1 integrons with aadA1 were located on incF plasmids with pMLST profile F51:A-:B10 in human isolates. The plasmid was present in unrelated as well as closely related host strains, demonstrating that dissemination of this

  9. Molecular Analysis of Antibiotic Resistance Determinants and Plasmids in Malaysian Isolates of Multidrug Resistant Klebsiella pneumoniae

    PubMed Central

    Al-Marzooq, Farah; Mohd Yusof, Mohd Yasim; Tay, Sun Tee

    2015-01-01

    Infections caused by multidrug resistant Klebsiella pneumoniae have been increasingly reported in many parts of the world. A total of 93 Malaysian multidrug resistant K. pneumoniae isolated from patients attending to University of Malaya Medical Center, Kuala Lumpur, Malaysia from 2010-2012 were investigated for antibiotic resistance determinants including extended-spectrum beta-lactamases (ESBLs), aminoglycoside and trimethoprim/sulfamethoxazole resistance genes and plasmid replicons. CTX-M-15 (91.3%) was the predominant ESBL gene detected in this study. aacC2 gene (67.7%) was the most common gene detected in aminoglycoside-resistant isolates. Trimethoprim/sulfamethoxazole resistance (90.3%) was attributed to the presence of sul1 (53.8%) and dfrA (59.1%) genes in the isolates. Multiple plasmid replicons (1-4) were detected in 95.7% of the isolates. FIIK was the dominant replicon detected together with 13 other types of plasmid replicons. Conjugative plasmids (1-3 plasmids of ~3-100 kb) were obtained from 27 of 43 K. pneumoniae isolates. An ESBL gene (either CTX-M-15, CTX-M-3 or SHV-12) was detected from each transconjugant. Co-detection with at least one of other antibiotic resistance determinants [sul1, dfrA, aacC2, aac(6ˊ)-Ib, aac(6ˊ)-Ib-cr and qnrB] was noted in most conjugative plasmids. The transconjugants were resistant to multiple antibiotics including β-lactams, gentamicin and cotrimoxazole, but not ciprofloxacin. This is the first study describing the characterization of plasmids circulating in Malaysian multidrug resistant K. pneumoniae isolates. The results of this study suggest the diffusion of highly diverse plasmids with multiple antibiotic resistance determinants among the Malaysian isolates. Effective infection control measures and antibiotic stewardship programs should be adopted to limit the spread of the multidrug resistant bacteria in healthcare settings. PMID:26203651

  10. Characterization of mannitol-fermenting methicillin-resistant staphylococci isolated from pigs in Nigeria

    PubMed Central

    Ugwu, Clifford C.; Gomez-Sanz, Elena; Agbo, Ifeoma C.; Torres, Carmen; Chah, Kennedy F.

    2015-01-01

    This study was conducted to determine the species distribution, antimicrobial resistance pheno- and genotypes and virulence traits of mannitol-positive methicillin-resistant staphylococci (MRS) isolated from pigs in Nsukka agricultural zone, Nigeria. Twenty mannitol-positive methicillin-resistant coagulase-negative staphylococcal (MRCoNS) strains harboring the mecA gene were detected among the 64 Staphylococcus isolates from 291 pigs. A total of 4 species were identified among the MRCoNS isolates, namely, Staphylococcus sciuri (10 strains), Staphylococcus lentus (6 strains), Staphylococcus cohnii (3 strains) and Staphylococcus haemolyticus (one strain). All MRCoNS isolates were multidrug-resistant. In addition to β-lactams, the strains were resistant to fusidic acid (85%), tetracycline (75%), streptomycin (65%), ciprofloxacin (65%), and trimethoprim/sulphamethoxazole (60%). In addition to the mecA and blaZ genes, other antimicrobial resistance genes detected were tet(K), tet(M), tet(L), erm(B), erm(C), aacA-aphD, aphA3, str, dfrK, dfrG, cat pC221, and cat pC223. Thirteen isolates were found to be ciprofloxacin-resistant, and all harbored a Ser84Leu mutation within the QRDR of the GyrA protein, with 3 isolates showing 2 extra substitutions, Ser98Ile and Arg100Lys (one strain) and Glu88Asp and Asp96Thr (2 strains). A phylogenetic tree of the QRDR nucleotide sequences in the gyrA gene revealed a high nucleotide diversity, with several major clusters not associated with the bacterial species. Our study highlights the possibility of transfer of mecA and other antimicrobial resistance genes from MRCoNS to pathogenic bacteria, which is a serious public health and veterinary concern. PMID:26413075

  11. Antimicrobial Resistance of Escherichia fergusonii Isolated from Broiler Chickens.

    PubMed

    Simmons, Karen; Islam, M Rashedul; Rempel, Heidi; Block, Glenn; Topp, Edward; Diarra, Moussa S

    2016-06-01

    The objective of this study was to investigate the antibiotic resistance of Escherichia fergusonii isolated from commercial broiler chicken farms. A total of 245 isolates from cloacal and cecal samples of 28- to 36-day-old chickens were collected from 32 farms. Isolates were identified using PCR, and their susceptibility to 16 antibiotics was determined by disk diffusion assay. All isolates were susceptible to meropenem, amikacin, and ciprofloxacin. The most common resistances were against ampicillin (75.1%), streptomycin (62.9%), and tetracycline (57.1%). Of the 184 ampicillin-resistant isolates, 127 were investigated using a DNA microarray carrying 75 probes for antibiotic resistance genetic determinants. Of these 127 isolates, the β-lactamase blaCMY2, blaTEM, blaACT, blaSHV, and blaCTX-M-15 genes were detected in 120 (94.5%), 31 (24.4%), 8 (6.3%), 6 (4.7%), and 4 (3.2%) isolates, respectively. Other detected genes included those conferring resistance to aminoglycosides (aadA1, strA, strB), trimethoprims (dfrV, dfrA1), tetracyclines (tetA, tetB, tetC, tetE), and sulfonamides (sul1, sul2). Class 1 integron was found in 35 (27.6%) of the ampicillin-resistant isolates. However, our data showed that the tested E. fergusonii did not carry any carbapenemase blaOXA genes. Pulsed-field gel electrophoresis revealed that the selected ampicillin-resistant E. fergusonii isolates were genetically diverse. The present study indicates that the monitoring of antimicrobial-resistant bacteria should include enteric bacteria such as E. fergusonii, which could be a reservoir of antibiotic resistance genes. The detection of isolates harboring extended-spectrum β-lactamase genes, particularly blaCTX-M-15, in this work suggests that further investigations on the occurrence of such genes in broilers are warranted.

  12. Isolation and Characterization of Antimicrobial-Resistant Nontyphoidal Salmonella enterica Serovars from Imported Food Products.

    PubMed

    Bae, Dongryeoul; Kweon, Ohgew; Khan, Ashraf A

    2016-08-01

    The objective of this study was to determine antimicrobial resistance and elucidate the resistance mechanism in nontyphoidal Salmonella enterica serovars isolated from food products imported into the United States from 2011 to 2013. Food products contaminated with antimicrobial-resistant nontyphoidal S. enterica were mainly imported from Taiwan, Indonesia, Vietnam, and China. PCR, DNA sequencing, and plasmid analyses were used to characterize antimicrobial resistance determinants. Twentythree of 110 S. enterica isolates were resistant to various antimicrobial classes, including β-lactam, aminoglycoside, phenicol, glycopeptide, sulfonamide, trimethoprim, and/or fluoroquinolone antimicrobial agents. Twelve of the isolates were multidrug resistant strains. Antimicrobial resistance determinants blaTEM-1, blaCTX-M-9, blaOXA-1, tetA, tetB, tetD, dfrA1, dfrV, dhfrI, dhfrXII, drf17, aadA1, aadA2, aadA5, orfC, qnrS, and mutations of gyrA and parC were detected in one or more antimicrobial-resistant nontyphoidal S. enterica strains. Plasmid profiles revealed that 12 of the 23 antimicrobial-resistant strains harbored plasmids with incompatibility groups IncFIB, IncHI1, IncI1, IncN, IncW, and IncX. Epidemiologic and antimicrobial resistance monitoring data combined with molecular characterization of antimicrobial resistance determinants in Salmonella strains isolated from imported food products may provide information that can be used to establish or implement food safety programs to improve public health.

  13. DNA fingerprinting and antimicrobial susceptibility pattern of clinical and environmental Acinetobacter baumannii isolates: a multicentre study.

    PubMed

    Salimizand, Himen; Menbari, Shaho; Ramazanzadeh, Rashid; Khonsha, Masomeh; Vahedi, Mohammad Saleh

    2014-11-21

    Backgrounds and aims: The aims of this study were to establish antibiotic profile and the molecular epidemiology of Acinetobacter baumannii isolates, with considering the effectiveness of control infection measures across three hospitals in the Kurdistan, west part of Iran. Methods: Fifty-four A. baumannii isolates were collected from patients and environmental specimens. Antibiotic susceptibility patterns (Antibio-type) were evaluated for 17 different antibiotics and MIC for imipenem was done. Isolates were assessed for the presence of metallo-beta-lactamases (MBLs), class 1 and 2 integrons, and integrated gene cassettes and blaOXA-like family genes. Repetitive-sequence-based PCR (REP-PCR) was done for analysing clonality and relativeness of isolates (REP-type). Results: Antibiotic susceptibility patterns distinguished 11 distinct Antibio-types and REP-PCR showed three clusters with 20 subclusters, mostly belonged to two clonal subgroups, A1 and B1. blaOXA-51 and blaOXA-23 were detected in 100% (54/54) and 52% (28/54), respectively, while blaOXA-24-like and blaOXA-58 were not present in isolates. MBLs were not detected, but, however, high rate of imipenem resistance was observed (52%). MIC90 of imipenem was 16 μg/ml. Class 1 integrons were detected in 11% (6/54) of isolates followed by 24% (13/54) of class 2. Both classes of integron genes were detected in 15% (8/54) of isolates. Integrated gene cassettes were in low level (11% of class 1 harboring isolates). Two arrays of gene cassettes were revealed, dfrA5-like and dfrA17-aadA5. Conclusion: Infection control surveillance should be considered as a serious manner, even the superficial eradication of hospital acquired pathogens. MBL genes were not induced carbapenem resistance in studied hospital settings, but blaOXA-51 & 23 contributed in imipenem resistant. Integrons had a little share in resistance of A. baumannii isolates.

  14. Antimicrobial resistance of Salmonella serovars isolated from beef at retail markets in the north Vietnam.

    PubMed

    Thai, Truong Ha; Hirai, Takuya; Lan, Nguyen Thi; Shimada, Akinori; Ngoc, Pham Thi; Yamaguchi, Ryoji

    2012-09-01

    Approximately 39.9% (63/158) of beef samples collected from retail markets in Hanoi from January to June 2009 were Salmonella-positive. Nine Salmonella serovars, Anatum (28.6%), Rissen (25.4%), Weltevreden (12.7%), Typhimurium (7.9%), Derby (7.9%), Lexington (7.9%), Dublin (4.6%), Newport (3.2%) and London (1.8%), were identified. Thirty-seven (58.7%) of the 63 Salmonella isolates were resistant to at least one antimicrobial tested, of which 29 (46%) isolates showed multidrug resistance (MDR). The isolates were commonly resistant to tetracycline (46.0%), sulphonamide (39.7%), ampicilline (31.7%), streptomycin (30.2%), trimethoprim (28.6%), kanamycin (28.6%) and chloramphenicol (22.2%). Fourteen (bla(TEMV), bla(OXA-1), aadA1, aadA2, sul1, tetA, tetB, tetG, cmlA1, floR, dfrA1, dfrA12, aac (3)-IV and aphA1-1AB) out of 22 antimicrobial resistance genes were detected by PCR from the resistant isolates. The catA1, Kn, blaPSE-1 genes and plasmid-mediated quinolones resistance (PMQR) genes such as qnrA, qnrB, qnrS, qepA and acc (6')-ib-cr were not detected. Mutations in the gyrA gene leading to the amino acid changes Ser83Phe and/or Asp87Asn were found in 6 out of the 11 quinolone-resistant isolates. The data revealed that multidrug resistant Salmonella strains were widely distributed in north Vietnam via the food chain and might contain multiple genes specifying identical resistant phenotypes. Thus, continuous studies are necessary to clarify the mechanisms of MDR in Salmonella and its spread in the livestock market.

  15. Antimicrobial resistance of Escherichia coli isolates from canine urinary tract infections.

    PubMed

    Chang, Shao-Kuang; Lo, Dan-Yuan; Wei, Hen-Wei; Kuo, Hung-Chih

    2015-01-01

    This study determined the antimicrobial resistance profiles of Escherichia coli isolates from dogs with a presumptive diagnosis of urinary tract infection (UTI). Urine samples from 201 dogs with UTI diagnosed through clinical examination and urinalysis were processed for isolation of Escherichia coli. Colonies from pure cultures were identified by biochemical reactions (n=114) and were tested for susceptibility to 18 antimicrobials. The two most frequent antimicrobials showing resistance in Urinary E. coli isolates were oxytetracycline and ampicillin. Among the resistant isolates, 17 resistance patterns were observed, with 12 patterns involving multidrug resistance (MDR). Of the 69 tetracycline-resistant E. coli isolates, tet(B) was the predominant resistance determinant and was detected in 50.9% of the isolates, whereas the remaining 25.5% isolates carried the tet(A) determinant. Most ampicillin and/or amoxicillin-resistant E. coli isolates carried blaTEM-1 genes. Class 1 integrons were prevalent (28.9%) and contained previously described gene cassettes that are implicated primarily in resistance to aminoglycosides and trimethoprim (dfrA1, dfrA17-aadA5). Of the 44 quinolone-resistant E. coli isolates, 38 were resistant to nalidixic acid, and 6 were resistant to nalidixic acid, ciprofloxacin and enrofloxacin. Chromosomal point mutations were found in the GyrA (Ser83Leu) and ParC (Ser80Ile) genes. Furthermore, the aminoglycoside resistance gene aacC2, the chloramphenicol resistant gene cmlA and the florfenicol resistant gene floR were also identified. This study revealed an alarming rate of antimicrobial resistance among E. coli isolates from dogs with UTIs. PMID:25720807

  16. Characterization of Virulence-Associated Genes, Antimicrobial Resistance Genes, and Class 1 Integrons in Salmonella enterica serovar Typhimurium Isolates from Chicken Meat and Humans in Egypt.

    PubMed

    Ahmed, Heba A; El-Hofy, Fatma I; Shafik, Saleh M; Abdelrahman, Mahmoud A; Elsaid, Gamilat A

    2016-06-01

    Foodborne pathogens are leading causes of illness especially in developing countries. The current study aimed to characterize virulence-associated genes and antimicrobial resistance in 30 Salmonella Typhimurium isolates of chicken and human origin at Mansoura, Egypt. The results showed that invA, avrA, mgtC, stn, and bcfC genes were identified in all the examined isolates, while 96.7% and 6.7% were positive for sopB and pef genes, respectively. The highest resistance frequencies of the isolates were to chloramphenicol and trimethoprim-sulfamethoxazole (73.3%, each), followed by streptomycin (56.7%), tetracycline and ampicillin (53.3%, each), and gentamicin (30%). However, only 2.7% of the isolates were resistant to cefotaxime and ceftriaxone each. Different resistance-associated genes, including blaTEM, aadB, aadC, aadA1, aadA2, floR, tetA(A), tetA(B), and sul1, were identified in Salmonella Typhimurium isolates with the respective frequencies of 53.3%, 6.7%, 23.3%, 46.7%, 63.3%, 73.3%, 60%, 20%, and 96.7%. None of the isolates was positive for blaSHV, blaOXA, and blaCMY genes. The results showed that the intI1 gene was detected in 24 (80%) of the examined Salmonella Typhimurium isolates. Class 1 integrons were found in 19 (79.2%) isolates that were intI1 positive. Seven integron profiles (namely: P-I to P-VII) were identified with P-V (gene cassette dfrA15, aadA2), the most prevalent profile. To the best of our knowledge, this is the first study to characterize the unusual gene cassette array dfrA12-OrfF-aadA27 from Salmonella Typhimurium isolates in Egypt. PMID:26977940

  17. Anatomical and biochemical analysis reveal the role of anthocyanins in flower coloration of herbaceous peony.

    PubMed

    Zhao, Da-Qiu; Wei, Meng-Ran; Liu, Ding; Tao, Jun

    2016-05-01

    Herbaceous peony (Paeonia lactiflora Pall.) is particularly appreciated because of its elegant and gorgeous flower color, but little is known about the underlying mechanisms of flower coloration. In this study, three P. lactiflora cultivars 'Xuefeng', 'Fenyulou' and 'Dahonglou' with white, pink and red flower were selected as the materials. Their anatomical structures, cell sap pH and metal elements were investigated, and the colored pigment mainly distributed in palisade mesophyll was only found in 'Fenyulou' and 'Dahonglou', and their shape of epidermal cells, cell sap pH and metal elements were not the key factors deciding phenotype color. Moreover, the qualitative and quantitative analysis of flavonoids were performed, their total anthocyanin, anthoxanthin and flavonoid contents were decreased during flower development, and only anthocyanin content in 'Dahonglou' was always higher than that in 'Xuefeng' and 'Fenyulou'. Subsequently, three anthocyanin compositions were found, and peonidin 3,5-di-O-glucoside (Pn3G5G) was identified as the main anthocyanin composition. In addition, the full-length of flavonol synthase gene (FLS) was isolated with the GenBank accession number KM259902, and the expression patterns of eight flavonoid biosynthetic genes showed that only PlDFR and PlANS basically had the highest levels in 'Dahonglou' and the lowest levels in 'Xuefeng', and they basically displayed a descended trend during flower development especially PlDFR, suggesting that these two genes might play a key role in the anthocyanin biosynthesis which resulted in the shift from white to pink and red in flowers. These results would contribute to understand the underlying molecular mechanisms of flower coloration in P. lactiflora.

  18. MOLECULAR DETECTION OF ANTIBIOTIC-RESISTANCE DETERMINANTS IN ESCHERICHIA COLI ISOLATED FROM THE ENDANGERED AUSTRALIAN SEA LION (NEOPHOCA CINEREA).

    PubMed

    Delport, Tiffany C; Harcourt, Robert G; Beaumont, Linda J; Webster, Koa N; Power, Michelle L

    2015-07-01

    Greater interaction between humans and wildlife populations poses significant risks of anthropogenic impact to natural ecosystems, especially in the marine environment. Understanding the spread of microorganisms at the marine interface is therefore important if we are to mitigate adverse effects on marine wildlife. We investigated the establishment of Escherichia coli in the endangered Australian sea lion (Neophoca cinerea) by comparing fecal isolation from wild and captive sea lion populations. Fecal samples were collected from wild colonies March 2009-September 2010 and from captive individuals March 2011-May 2013. Using molecular screening, we assigned a phylotype to E. coli isolates and determined the presence of integrons, mobile genetic elements that capture gene cassettes conferring resistance to antimicrobial agents common in fecal coliforms. Group B2 was the most abundant phylotype in all E. coli isolates (n = 37), with groups A, B1, and D also identified. Integrons were not observed in E. coli (n = 21) isolated from wild sea lions, but were identified in E. coli from captive animals (n = 16), from which class I integrases were detected in eight isolates. Sequencing of gene cassette arrays identified genes conferring resistance to streptomycin-spectinomycin (aadA1) and trimethoprim (dfrA17, dfrB4). Class II integrases were not detected in the E. coli isolates. The frequent detection in captive sea lions of E. coli with resistance genes commonly identified in human clinical cases suggests that conditions experienced in captivity may contribute to establishment. Identification of antibiotic resistance in the microbiota of Australian sea lions provides crucial information for disease management. Our data will inform conservation management strategies and provide a mechanism to monitor microorganism dissemination to sensitive pinniped populations. PMID:25919463

  19. Effects of in-channel sand excavation on the hydrology of the Pearl River Delta, China

    NASA Astrophysics Data System (ADS)

    Luo, Xian-Lin; Zeng, Eddy Y.; Ji, Rong-Yao; Wang, Chao-Pin

    2007-09-01

    SummaryThe hydrology and morphology of the Pearl River Delta (PRD; South China) water system has been predominantly dictated by human activities over the last 20 years. Uncontrolled sand excavation occurred in all 324 tributaries, largely to meet the construction needs arising from the rapid economic growth and urbanization in the region. It was estimated that >8.7 × 10 8 m 3 of sand were excavated from 1986 to 2003 based on field surveys of excavating activities and the river hypsography, resulting in average downcutting depths of 0.59-1.73 m, 0.34-4.43 m, and 1.77-6.48 m in the main channels of the West River, North River, and East River (three major water networks in the PRD), respectively. Consequently, the water levels in upstream of the PRD were decreased by 1.59-3.12 m (Sanshui Station). Uneven sand dredging also caused changes in the divided flow ratio (DFR) between various water courses. For example, the DFR increased by 8.8% at the Sanshui Station on the upper part of the North River network from the early 1980s to 1999. DFRs also increased almost 7.7% at the four major runoff outlets in the eastern side of the PRD. As a result, present brackish-water has intruded upward 10-20 km more than in the 1980s. Apparently, there are two sides to the effects of sand excavation. The positive effects are decreased chances of flooding damages, improved navigating conditions, and more water inputs to rapid economically growing regions. The negative effects include increased grade slope and instability of the riverbank, disruption of navigation in upstream dredging pits during dry seasons, and brackish-water intrusion.

  20. Molecular Analysis of Antibiotic Resistance Determinants and Plasmids in Malaysian Isolates of Multidrug Resistant Klebsiella pneumoniae.

    PubMed

    Al-Marzooq, Farah; Mohd Yusof, Mohd Yasim; Tay, Sun Tee

    2015-01-01

    Infections caused by multidrug resistant Klebsiella pneumoniae have been increasingly reported in many parts of the world. A total of 93 Malaysian multidrug resistant K. pneumoniae isolated from patients attending to University of Malaya Medical Center, Kuala Lumpur, Malaysia from 2010-2012 were investigated for antibiotic resistance determinants including extended-spectrum beta-lactamases (ESBLs), aminoglycoside and trimethoprim/sulfamethoxazole resistance genes and plasmid replicons. CTX-M-15 (91.3%) was the predominant ESBL gene detected in this study. aacC2 gene (67.7%) was the most common gene detected in aminoglycoside-resistant isolates. Trimethoprim/sulfamethoxazole resistance (90.3%) was attributed to the presence of sul1 (53.8%) and dfrA (59.1%) genes in the isolates. Multiple plasmid replicons (1-4) were detected in 95.7% of the isolates. FIIK was the dominant replicon detected together with 13 other types of plasmid replicons. Conjugative plasmids (1-3 plasmids of ~3-100 kb) were obtained from 27 of 43 K. pneumoniae isolates. An ESBL gene (either CTX-M-15, CTX-M-3 or SHV-12) was detected from each transconjugant. Co-detection with at least one of other antibiotic resistance determinants [sul1, dfrA, aacC2, aac(6')-Ib, aac(6')-Ib-cr and qnrB] was noted in most conjugative plasmids. The transconjugants were resistant to multiple antibiotics including β-lactams, gentamicin and cotrimoxazole, but not ciprofloxacin. This is the first study describing the characterization of plasmids circulating in Malaysian multidrug resistant K. pneumoniae isolates. The results of this study suggest the diffusion of highly diverse plasmids with multiple antibiotic resistance determinants among the Malaysian isolates. Effective infection control measures and antibiotic stewardship programs should be adopted to limit the spread of the multidrug resistant bacteria in healthcare settings.

  1. Detection of antibiotic resistant enterococci and Escherichia coli in free range Iberian Lynx (Lynx pardinus).

    PubMed

    Gonçalves, Alexandre; Igrejas, Gilberto; Radhouani, Hajer; Santos, Tiago; Monteiro, Ricardo; Pacheco, Rui; Alcaide, Eva; Zorrilla, Irene; Serra, Rodrigo; Torres, Carmen; Poeta, Patrícia

    2013-07-01

    Thirty fecal samples from wild specimens of Iberian lynx were collected and analyzed for Enterococcus spp. (27 isolates) and Escherichia coli (18 isolates) recovery. The 45 isolates obtained were tested for antimicrobial resistance, molecular mechanisms of resistance, and presence of virulence genes. Among the enterococci, Enterococcus faecium and Enterococcus hirae were the most prevalent species (11 isolates each), followed by Enterococcus faecalis (5 isolates). High percentages of resistance to tetracycline and erythromycin (33% and 30%, respectively) were detected among enterococcal isolates. The tet(M) and/or tet(L), erm(B), aac(6')-Ie-aph(2″)-Ia, ant(6)-Ia, or aph(3')-IIIa genes were detected among resistant enterococci. Virulence genes were detected in one E. faecalis isolate (cpd, cylB, and cylL) and one E. hirae isolate (cylL). High percentages of resistance were detected in E. coli isolates to tetracycline (33%), streptomycin (28%), nalidixic acid (28%), and sulfamethoxazole-trimethoprim (SXT, 22%). Additionally, the blaTEM, tet(A), aadA, cmlA, and different combinations of sul genes were detected among most ampicillin, tetracycline, streptomycin, chloramphenicol and SXT-resistant isolates, respectively. Two isolates contained a class 1 integron with the gene cassette arrays dfrA1 + aadA1 and dfrA12 + aadA2. The E. coli isolates were ascribed to phylo-groups A (n=5); B1 (n=4); B2 (n=6), and D (n=3), with the virulence gene fimA present in all E. coli isolates. This study found resistance genes in wild specimens of Iberian lynx. Thus, it is important to notice that multiresistant bacteria have reached species as rare and completely non-synanthropic as the Iberian lynx. Furthermore, the susceptibility of this endangered species to bacterial infection may be affected by the presence of these virulence and resistance genes.

  2. Virulence and resistance determinants of German Staphylococcus aureus ST398 isolates from nonhuman sources.

    PubMed

    Argudín, M A; Tenhagen, B-A; Fetsch, A; Sachsenröder, J; Käsbohrer, A; Schroeter, A; Hammerl, J A; Hertwig, S; Helmuth, R; Bräunig, J; Mendoza, M C; Appel, B; Rodicio, M R; Guerra, B

    2011-05-01

    A series of 100 Staphylococcus aureus isolates ascribed to sequence type 398 (ST398) and recovered from different sources (healthy carrier and diseased pigs, dust from pig farms, milk, and meat) in Germany were investigated for their virulence and antimicrobial resistance genetic background. Antimicrobial resistance was determined by the disk diffusion method. Virulence and resistance determinants (37 and 31 genes, respectively) were tested by PCR. Only two virulence profiles, including the accessory gene regulator agrI and three or four hemolysin-encoding genes, were detected. In contrast, 33 resistance profiles were distinguished (only 11 were shown by more than one isolate). Fifty-nine isolates were multiresistant (four or more antimicrobial classes), and 98 were methicillin resistant (mecA positive). All of the ST398 isolates showed resistance to tetracycline [encoded by tet(M) alone or together with tet(K) and/or tet(L)]. In addition, 98% were resistant to other antimicrobials, including macrolide-lincosamine-streptogramin B (70%, encoded by ermA, ermB, and ermC, alone or in combination), trimethoprim (65%, mostly due to dfrK and dfrG), kanamycin and gentamicin [29% and 14%, respectively, mainly related to aac(6')-Ie-aph(2″)-Ia and/or ant(4')-Ia but also to aph(3')-IIIa], chloramphenicol (9%, fexA or cfr), quinupristin-dalfopristin (9%), ciprofloxacin (8%), and trimethoprim-sulfamethoxazole (4%). The heterogeneity of the resistance profiles underlines the ability of the ST398 clone to acquire multiple antimicrobial resistance genes. However, the virulence gene content of the tested isolates was low. Continuous surveillance is needed to clarify whether its pathogenicity potential for animals and humans will increase over time.

  3. A MYB transcription factor regulates anthocyanin biosynthesis in mangosteen (Garcinia mangostana L.) fruit during ripening.

    PubMed

    Palapol, Yossapol; Ketsa, Saichol; Lin-Wang, Kui; Ferguson, Ian B; Allan, Andrew C

    2009-05-01

    Mangosteen (Garcinia mangostana L.) fruit undergo rapid red colour development, both on the tree and after harvest, resulting in high anthocyanin production in the pericarp. Here, we report the isolation of three full-length mangosteen MYB transcription factors (GmMYB1, GmMYB7 and GmMYB10) and all the anthocyanin biosynthetic pathway genes (GmPal to GmUFGT). Phylogenetic analysis at the protein level of the R2R3-MYB transcription factor family showed GmMYB10 had a high degree of similarity with production of anthocyanin pigment1 in Arabidopsis and as well as sequences from other plant species related to the elevation of anthocyanin pigmentation. In transient transactivation assays, GmMYB10, co-expressed with AtbHLH2, strongly activated the GmDFR and AtDFR promoters. Transcripts of GmMYB10 and GmUFGT were highly abundant with onset of pigmentation and subsequently during red colouration. Our results suggest that GmMYB10 plays an important role in regulating anthocyanin biosynthesis both on the tree and after harvest, while GmUFGT may be a key biosynthetic gene in mangosteen pigmentation. The expression patterns of GmMYB10 and GmUFGT correlated with ethylene production that increased linearly until stage 5 (dark purple) and decreased thereafter. 1-Methycyclopropene (1-MCP) clearly delayed red colouration with resulting down-regulation of GmMYB10. These results suggest that the effect of ethylene on anthocyanin biosynthesis may be via the regulation of GmMYB10 expression.

  4. Antimicrobial Resistance, Virulence Profiles and Molecular Subtypes of Salmonella enterica Serovars Typhi and Paratyphi A Blood Isolates from Kolkata, India during 2009-2013

    PubMed Central

    Dutta, Shanta; Das, Surojit; Mitra, Utpala; Jain, Priyanka; Roy, Indranil; Ganguly, Shelley S.; Ray, Ujjwayini; Dutta, Phalguni; Paul, Dilip Kumar

    2014-01-01

    Enteric fever, caused by Salmonella enterica, remains an unresolved public health problem in India and antimicrobial therapy is the main mode of treatment. The objective of this study was to characterize the Salmonella enterica isolates from Kolkata with respect to their antimicrobial resistance (AMR), virulence profiles and molecular subtypes. Salmonella enterica blood isolates were collected from clinically suspected enteric fever patients attending various hospitals in Kolkata, India from January 2009 to June 2013 and were tested for AMR profiles by standard protocols; for resistance gene transfer by conjugation; for resistance and virulence genes profiles by PCR; and for molecular subtypes by Pulsed Field Gel Electrophoresis (PFGE). A total of 77 Salmonella enterica serovar Typhi (S. Typhi) and 25 Salmonella enterica serovar Paratyphi A (S. Paratyphi A) from Kolkata were included in this study. Although multidrug resistance (resistance to chloramphenicol, ampicillin, co-trimoxazole) was decreasing in S. Typhi (18.2%) and absent in S. Paratyphi A, increased resistance to fluoroquinolone, the current drug of choice, caused growing concern for typhoid treatment. A single, non-conjugative non-IncHI1 plasmid of 180 kb was found in 71.4% multidrug resistant (MDR) S. Typhi; the remaining 28.6% isolates were without plasmid. Various AMR markers (blaTEM-1, catA, sul1, sul2, dfrA15, strA-strB) and class 1 integron with dfrA7 gene were detected in MDR S. Typhi by PCR and sequencing. Most of the study isolates were likely to be virulent due to the presence of virulence markers. Major diversity was not noticed among S. Typhi and S. Paratyphi A from Kolkata by PFGE. The observed association between AMR profiles and S. Typhi pulsotypes might be useful in controlling the spread of the organism by appropriate intervention. The study reiterated the importance of continuous monitoring of AMR and molecular subtypes of Salmonella isolates from endemic regions for better

  5. Novel insight into the mechanism underlying light-controlled anthocyanin accumulation in eggplant (Solanum melongena L.).

    PubMed

    Jiang, Mingmin; Ren, Li; Lian, Hongli; Liu, Yang; Chen, Huoying

    2016-08-01

    Eggplant is rich in anthocyanins, which are the major secondary metabolites and beneficial to human health. We discovered that the anthocyanin biosynthesis of eggplant cultivar 'Lanshan Hexian' was regulated by light. In this study, we isolated two blue light receptor genes, SmCRY1 and SmCRY2, and negative/positive anthocyanin regulatory factors SmCOP1 and SmHY5 from eggplant. In terms of transcript levels, SmCRY1, SmCRY2 and SmHY5 were up-regulated by light, while SmCOP1 was down-regulated. Subsequently, the four genes were functionally complemented in phenotype of corresponding mutants, indicating that they act as counterparts of Arabidopsis genes. Yeast two-hybrid and bimolecular fluorescence complementation assays showed that SmCRY1 and SmCRY2 interact with SmCOP1 in a blue-light-dependent manner. It also obtained the result that SmCOP1 interacts with SmHY5 and SmMYB1. Furthermore, using yeast one-hybrid assay, we found that SmHY5 and SmMYB1 both bind the promoters of anthocyanin biosynthesis structural genes (SmCHS and SmDFR). Taken together, blue-light-triggered CRY1/CRY2-COP1 interaction creates the condition that HY5 and MYB1 combine with the downstream anthocyanin synthesis genes (CHS and DFR) in eggplant. Our finding provides a new working model by which light controls anthocyanin accumulation in eggplant. PMID:27297989

  6. A molecular assessment of the genetic model of spathe color inheritance in Anthurium andraeanum (Hort.).

    PubMed

    Gopaulchan, David; Umaharan, Pathmanathan; Lennon, Adrian M

    2014-03-01

    Past genetic studies have shown three independent loci designated O, R and M control spathe color in Anthurium andraeanum (Hort.). To evaluate the genetic model and to understand the control of anthocyanin biosynthesis at the molecular level, the expression of the anthocyanin biosynthetic genes, CHS, F3H, DFR, ANS and F3'H, was examined at the mRNA and protein levels and correlated to anthocyanin content and spathe color in eight genetically characterized anthurium cultivars representing different states of the O, R and M loci. The results showed that the expression of F3H and ANS was co-regulated by a putative transcription factor encoded by the R locus, and the expression of DFR was regulated by a putative transcription factor encoded by the O locus. White cultivars, which were in the homozygous recessive state for either O or R or both, exhibited reduced expression of the anthocyanin biosynthetic genes and hence had negligible levels of anthocyanin. Cultivars that were mm displayed reduced expression of F3'H suggesting that it may either encode a defective form of the F3'H gene or a regulator that controls its expression. Additionally, a correlation between anthocyanin abundance and the expression of F3'H in the red cultivars suggested that F3'H expression may be a key control point in the regulation of anthocyanin biosynthesis in anthurium and hence plays a major role in influencing the shade intensity in red cultivars. These findings provide evidence in support of the genetic model for color inheritance in the spathe. PMID:24363030

  7. Regulation of anthocyanin biosynthesis in Arabidopsis thaliana red pap1-D cells metabolically programmed by auxins.

    PubMed

    Liu, Zhong; Shi, Ming-Zhu; Xie, De-Yu

    2014-04-01

    Red pap1-D cells of Arabidopsis thaliana have been cloned from production of anthocyanin pigmentation 1-Dominant (pap1-D) plants. The red cells are metabolically programmed to produce high levels of anthocyanins by a WD40-bHLH-MYB complex that is composed of the TTG1, TT8/GL3 and PAP1 transcription factors. Here, we report that indole 3-acetic acid (IAA), naphthaleneacetic acid (NAA) and 2,4-dichlorophenoxyacetic acid (2,4-D) regulate anthocyanin biosynthesis in these red cells. Seven concentrations (0, 0.2, 0.4, 2.2, 9, 18 and 27 μM) were tested for the three auxins. IAA and 2,4-D at 2.2-27 μM reduced anthocyanin levels. NAA at 0-0.2 μM or above 9 μM also decreased anthocyanin levels, but from 0.4 to 9 μM, it increased them. HPLC-ESI-MS analysis identified seven cyanin molecules that were produced in red pap1-D cells, and their levels were affected by auxins. The expression levels of ten genes, including six transcription factors (TTG1, EGL3, MYBL2, TT8, GL3 and PAP1) and four pathway genes (PAL1, CHS, DFR and ANS) involved in anthocyanin biosynthesis were analyzed upon various auxin treatments. The resulting data showed that 2,4-D, NAA and IAA control anthocyanin biosynthesis by regulating the expression of TT8, GL3 and PAP1 as well as genes in the anthocyanin biosynthetic pathway, such as DFR and ANS. In addition, the expression of MYBL2, PAL1 and CHS in red pap1-D and wild-type cells differentially respond to the three auxins. Our data demonstrate that the three auxins regulate anthocyanin biosynthesis in metabolically programmed red cells via altering the expression of transcription factor genes and pathway genes. PMID:24370633

  8. Novel insight into the mechanism underlying light-controlled anthocyanin accumulation in eggplant (Solanum melongena L.).

    PubMed

    Jiang, Mingmin; Ren, Li; Lian, Hongli; Liu, Yang; Chen, Huoying

    2016-08-01

    Eggplant is rich in anthocyanins, which are the major secondary metabolites and beneficial to human health. We discovered that the anthocyanin biosynthesis of eggplant cultivar 'Lanshan Hexian' was regulated by light. In this study, we isolated two blue light receptor genes, SmCRY1 and SmCRY2, and negative/positive anthocyanin regulatory factors SmCOP1 and SmHY5 from eggplant. In terms of transcript levels, SmCRY1, SmCRY2 and SmHY5 were up-regulated by light, while SmCOP1 was down-regulated. Subsequently, the four genes were functionally complemented in phenotype of corresponding mutants, indicating that they act as counterparts of Arabidopsis genes. Yeast two-hybrid and bimolecular fluorescence complementation assays showed that SmCRY1 and SmCRY2 interact with SmCOP1 in a blue-light-dependent manner. It also obtained the result that SmCOP1 interacts with SmHY5 and SmMYB1. Furthermore, using yeast one-hybrid assay, we found that SmHY5 and SmMYB1 both bind the promoters of anthocyanin biosynthesis structural genes (SmCHS and SmDFR). Taken together, blue-light-triggered CRY1/CRY2-COP1 interaction creates the condition that HY5 and MYB1 combine with the downstream anthocyanin synthesis genes (CHS and DFR) in eggplant. Our finding provides a new working model by which light controls anthocyanin accumulation in eggplant.

  9. Chemical stability of amorphous materials: specific and general media effects in the role of water in the degradation of freeze-dried zoniporide.

    PubMed

    Luthra, Suman A; Shalaev, Evgenyi Y; Medek, Ales; Hong, Jinyang; Pikal, Michael J

    2012-09-01

    The objective of the present work was to determine whether hydrolysis in a model lyophile was influenced by general media effects with water-changing properties of the medium or via a specific mechanism of water as a reactant. Four formulations of zoniporide and sucrose (1:10) were prepared with variable amounts of sorbitol [0%-25% (w/v) of total solids). These formulations were then equilibrated at 6% and 11% relative humidity using saturated salt solutions. The lyophile cakes were analyzed by differential scanning calorimetery (DSC), (isothermal microcalorimetry (IMC), solid- state nuclear magnetic resonance (ssNMR) spectroscopy, and ultraviolet-visible diffuse reflectance (DFR) spectroscopy. DSC and IMC were used to assess the global molecular mobility. ssNMR relaxation times were measured to access local mobility. The DFR was used to determine the solid-state acidity expressed as the Hammett acidity function. Stability of samples was evaluated at 40°C by monitoring potency and purity by high-performance liquid chromatography (HPLC). Results were interpreted in terms of the various roles of water: media effect, plasticization, polarity, and reactant. The kinetics of hydrolysis was observed to be correlated with either/both specific "chemical" effects, that is, water reactant as well as media effect, specifically global molecular mobility of the matrix. Increase in reaction rate with increase in water content is not linear and is a weaker dependence than in some hydrolytic reactions in organic solvents. A moderate amount of an inert plasticizer, sorbitol, conferred additional stabilization, possibly by restricting the amplitude and frequency of fast motions that are on a small length scale.

  10. A putative functional MYB transcription factor induced by low temperature regulates anthocyanin biosynthesis in purple kale (Brassica Oleracea var. acephala f. tricolor).

    PubMed

    Zhang, Bin; Hu, Zongli; Zhang, Yanjie; Li, Yali; Zhou, Shuang; Chen, Guoping

    2012-02-01

    The purple kale (Brassica Oleracea var. acephala f. tricolor) is a mutation in kales, giving the mutant phenotype of brilliant purple color in the interior. Total anthocyanin analysis showed that the amount of anthocyanins in the purple kale was up to 1.73 mg g(-1) while no anthocyanin was detected in the white kale. To elucidate the molecular mechanism of the anthocyanin biosynthesis in the purple kale, we analyzed the expression of structural genes and some transcription factors associated with anthocyanin biosynthesis in the purple cultivar "Red Dove" and the white cultivar "White Dove". The result showed that nearly all the anthocyanin biosynthetic genes showed higher expression levels in the purple cultivar than in the white cultivar, especially for DFR and ANS, they were barely detected in the white cultivar. Interestingly, the fact that a R2R3 MYB transcription factor named BoPAP1 was extremely up-regulated in the purple kale and induced by low temperature attracted our attention. Further sequence analysis showed that BoPAP1 shared high similarity with AtPAP1 and BoMYB1. In addition, the anthocyanin accumulation in the purple kale is strongly induced by the low temperature stress. The total anthocyanin contents in the purple kale under low temperature were about 50-fold higher than the plants grown in the greenhouse. The expression of anthocyanin biosynthetic genes C4H, F3H, DFR, ANS and UFGT were all enhanced under the low temperature. These evidences strongly suggest that BoPAP1 may play an important role in activating the anthocyanin structural genes for the abundant anthocyanin accumulation in the purple kale.

  11. Antimicrobial resistance of Escherichia coli isolates from canine urinary tract infections

    PubMed Central

    CHANG, Shao-Kuang; LO, Dan-Yuan; WEI, Hen-Wei; KUO, Hung-Chih

    2014-01-01

    This study determined the antimicrobial resistance profiles of Escherichia coli isolates from dogs with a presumptive diagnosis of urinary tract infection (UTI). Urine samples from 201 dogs with UTI diagnosed through clinical examination and urinalysis were processed for isolation of Escherichia coli. Colonies from pure cultures were identified by biochemical reactions (n=114) and were tested for susceptibility to 18 antimicrobials. The two most frequent antimicrobials showing resistance in Urinary E. coli isolates were oxytetracycline and ampicillin. Among the resistant isolates, 17 resistance patterns were observed, with 12 patterns involving multidrug resistance (MDR). Of the 69 tetracycline-resistant E. coli isolates, tet(B) was the predominant resistance determinant and was detected in 50.9% of the isolates, whereas the remaining 25.5% isolates carried the tet(A) determinant. Most ampicillin and/or amoxicillin-resistant E. coli isolates carried blaTEM-1 genes. Class 1 integrons were prevalent (28.9%) and contained previously described gene cassettes that are implicated primarily in resistance to aminoglycosides and trimethoprim (dfrA1, dfrA17-aadA5). Of the 44 quinolone-resistant E. coli isolates, 38 were resistant to nalidixic acid, and 6 were resistant to nalidixic acid, ciprofloxacin and enrofloxacin. Chromosomal point mutations were found in the GyrA (Ser83Leu) and ParC (Ser80Ile) genes. Furthermore, the aminoglycoside resistance gene aacC2, the chloramphenicol resistant gene cmlA and the florfenicol resistant gene floR were also identified. This study revealed an alarming rate of antimicrobial resistance among E. coli isolates from dogs with UTIs. PMID:25720807

  12. Empirical conversion of the vertical profile of reflectivity from Ku-band to S-band frequency

    NASA Astrophysics Data System (ADS)

    Cao, Qing; Hong, Yang; Qi, Youcun; Wen, Yixin; Zhang, Jian; Gourley, Jonathan J.; Liao, Liang

    2013-02-01

    ABSTRACT This paper presents an empirical method for converting reflectivity from Ku-band (13.8 GHz) to S-band (2.8 GHz) for several hydrometeor species, which facilitates the incorporation of Tropical Rainfall Measuring Mission (TRMM) Precipitation Radar (PR) measurements into quantitative precipitation estimation (QPE) products from the U.S. Next-Generation Radar (NEXRAD). The development of empirical dual-frequency relations is based on theoretical simulations, which have assumed appropriate scattering and microphysical models for liquid and solid hydrometeors (raindrops, snow, and ice/hail). Particle phase, shape, orientation, and density (especially for snow particles) have been considered in applying the T-matrix method to compute the scattering amplitudes. Gamma particle size distribution (PSD) is utilized to model the microphysical properties in the ice region, melting layer, and raining region of precipitating clouds. The variability of PSD parameters is considered to study the characteristics of dual-frequency reflectivity, especially the variations in radar dual-frequency ratio (DFR). The empirical relations between DFR and Ku-band reflectivity have been derived for particles in different regions within the vertical structure of precipitating clouds. The reflectivity conversion using the proposed empirical relations has been tested using real data collected by TRMM-PR and a prototype polarimetric WSR-88D (Weather Surveillance Radar 88 Doppler) radar, KOUN. The processing and analysis of collocated data demonstrate the validity of the proposed empirical relations and substantiate their practical significance for reflectivity conversion, which is essential to the TRMM-based vertical profile of reflectivity correction approach in improving NEXRAD-based QPE.

  13. MOLECULAR DETECTION OF ANTIBIOTIC-RESISTANCE DETERMINANTS IN ESCHERICHIA COLI ISOLATED FROM THE ENDANGERED AUSTRALIAN SEA LION (NEOPHOCA CINEREA).

    PubMed

    Delport, Tiffany C; Harcourt, Robert G; Beaumont, Linda J; Webster, Koa N; Power, Michelle L

    2015-07-01

    Greater interaction between humans and wildlife populations poses significant risks of anthropogenic impact to natural ecosystems, especially in the marine environment. Understanding the spread of microorganisms at the marine interface is therefore important if we are to mitigate adverse effects on marine wildlife. We investigated the establishment of Escherichia coli in the endangered Australian sea lion (Neophoca cinerea) by comparing fecal isolation from wild and captive sea lion populations. Fecal samples were collected from wild colonies March 2009-September 2010 and from captive individuals March 2011-May 2013. Using molecular screening, we assigned a phylotype to E. coli isolates and determined the presence of integrons, mobile genetic elements that capture gene cassettes conferring resistance to antimicrobial agents common in fecal coliforms. Group B2 was the most abundant phylotype in all E. coli isolates (n = 37), with groups A, B1, and D also identified. Integrons were not observed in E. coli (n = 21) isolated from wild sea lions, but were identified in E. coli from captive animals (n = 16), from which class I integrases were detected in eight isolates. Sequencing of gene cassette arrays identified genes conferring resistance to streptomycin-spectinomycin (aadA1) and trimethoprim (dfrA17, dfrB4). Class II integrases were not detected in the E. coli isolates. The frequent detection in captive sea lions of E. coli with resistance genes commonly identified in human clinical cases suggests that conditions experienced in captivity may contribute to establishment. Identification of antibiotic resistance in the microbiota of Australian sea lions provides crucial information for disease management. Our data will inform conservation management strategies and provide a mechanism to monitor microorganism dissemination to sensitive pinniped populations.

  14. The Arabidopsis Transcription Factor ANAC032 Represses Anthocyanin Biosynthesis in Response to High Sucrose and Oxidative and Abiotic Stresses

    PubMed Central

    Mahmood, Kashif; Xu, Zhenhua; El-Kereamy, Ashraf; Casaretto, José A.; Rothstein, Steven J.

    2016-01-01

    Production of anthocyanins is one of the adaptive responses employed by plants during stress conditions. During stress, anthocyanin biosynthesis is mainly regulated at the transcriptional level via a complex interplay between activators and repressors of anthocyanin biosynthesis genes. In this study, we investigated the role of a NAC transcription factor, ANAC032, in the regulation of anthocyanin biosynthesis during stress conditions. ANAC032 expression was found to be induced by exogenous sucrose as well as high light (HL) stress. Using biochemical, molecular and transgenic approaches, we show that ANAC032 represses anthocyanin biosynthesis in response to sucrose treatment, HL and oxidative stress. ANAC032 was found to negatively affect anthocyanin accumulation and the expression of anthocyanin biosynthesis (DFR, ANS/LDOX) and positive regulatory (TT8) genes as demonstrated in overexpression line (35S:ANAC032) compared to wild-type under HL stress. The chimeric repressor line (35S:ANAC032-SRDX) exhibited the opposite expression patterns for these genes. The negative impact of ANAC032 on the expression of DFR, ANS/LDOX and TT8 was found to be correlated with the altered expression of negative regulators of anthocyanin biosynthesis, AtMYBL2 and SPL9. In addition to this, ANAC032 also repressed the MeJA- and ABA-induced anthocyanin biosynthesis. As a result, transgenic lines overexpressing ANAC032 (35S:ANAC032) produced drastically reduced levels of anthocyanin pigment compared to wild-type when challenged with salinity stress. However, transgenic chimeric repressor lines (35S:ANAC032-SRDX) exhibited the opposite phenotype. Our results suggest that ANAC032 functions as a negative regulator of anthocyanin biosynthesis in Arabidopsis thaliana during stress conditions. PMID:27790239

  15. Antimicrobial Susceptibility and Mechanisms of Resistance in Shigella and Salmonella Isolates from Children under Five Years of Age with Diarrhea in Rural Mozambique▿

    PubMed Central

    Mandomando, Inácio; Jaintilal, Dinis; Pons, Maria J.; Vallès, Xavier; Espasa, Mateu; Mensa, Laura; Sigaúque, Betuel; Sanz, Sergi; Sacarlal, Jahit; Macete, Eusébio; Abacassamo, Fátima; Alonso, Pedro L.; Ruiz, Joaquim

    2009-01-01

    The antimicrobial susceptibility and mechanisms of resistance of 109 Shigella and 40 Salmonella isolates from children with diarrhea in southern Mozambique were assessed. The susceptibility to seven antimicrobial agents was tested by disk diffusion, and mechanisms of resistance were searched by PCR or colorimetric method. A high proportion of Shigella isolates were resistant to chloramphenicol (Chl) (52%), ampicillin (Amp) (56%), tetracycline (Tet) (66%), and trimethoprim-sulfamethoxazole (Sxt) (84%). Sixty-five percent of the isolates were multidrug resistant. Shigella flexneri isolates were more resistant than those of Shigella sonnei to Amp (66% versus 0.0%, P < 0.001) and Chl (61% versus 0.0%, P < 0.001), whereas S. sonnei isolates presented higher resistance to Tet than S. flexneri isolates (93% versus 64%, P = 0.02). Resistance among Salmonella isolates was as follows: Tet and Chl, 15% each; Sxt, 18%; and Amp, 25%. Only 3% of Salmonella isolates were resistant to nalidixic acid (Nal), and none to ciprofloxacin or ceftriaxone (Cro). Among Salmonella isolates, multiresistance was found in 23%. Among Shigella isolates, antibiotic resistance was related mainly to the presence of oxa-1-like β-lactamases for Amp, dfrA1 genes for Sxt, tetB genes for Tet, and Chl acetyltransferase (CAT) activity for Chl. Among Salmonella isolates, resistance was conferred by tem-like β-lactamases for Amp, floR genes and CAT activity for Chl, tetA genes for Tet, and dfrA1 genes for Sxt. Our data show that Shigella isolates are resistant mostly to the most available, inexpensive antibiotics by various molecular mechanisms but remain susceptible to ciprofloxacin, Cro, and Nal, which is the first line for empirical treatment of shigellosis in the country. PMID:19332670

  16. Exploiting synthetic aperture radar imagery for retrieving vibration signatures of concealed machinery

    NASA Astrophysics Data System (ADS)

    Pérez, Francisco; Campbell, Justin B.; Jaramillo, Monica; Dunkel, Ralf; Atwood, Thomas; Doerry, Armin; Gerstle, Walter H.; Santhanam, Balu; Hayat, Majeed M.

    2016-05-01

    It has been demonstrated that the instantaneous acceleration associated with vibrating objects that are directly imaged by synthetic aperture radar (SAR) can be estimated through the application of the discrete fractional Fourier transform (DFrFT) using the information contained in the complex SAR image. In general, vibration signatures may include, for example, the number of chirped sinusoids as well as their respective base frequencies and chirp rates. By further processing the DFrFT-processed data for clutter-noise rejection by means of pseudo- subspace methods, has been shown that the SAR-vibrometry method can be reliable as long as the signal-to-noise ratio (SNR) and the signal-to-clutter ratio (SCR) of the slow-time SAR signal at the range-line of interest exceeds 15dB. Meanwhile, the Nyquist theorem dictates that the maximum measurable vibration frequency is limited by half of the pulse-repetition frequency. This paper focuses on the detection and estimation of vibrations generated by machinery concealed within buildings and other structures. This is a challenging task in general because the vibration signatures of the source are typically altered by their housing structure; moreover, the SNR at the surface of the housing structure tends to be reduced. Here, experimental results for three different vibrating targets, including one concealed target, are reported using complex SAR images acquired by the General Atomics Lynx radar at resolutions of 1-ft and 4-in. The concealed vibrating target is actuated by a gear motor with an off-balance weight attached to it, which is enclosed by a wooden housing. The vibrations of the motor are transmitted to a chimney that extends above the housing structure. Using the SAR vibrometry approach, it is shown that it is possible to distinguish among the three vibrating objects based upon their vibration signatures.

  17. Characterization of mannitol-fermenting methicillin-resistant staphylococci isolated from pigs in Nigeria.

    PubMed

    Ugwu, Clifford C; Gomez-Sanz, Elena; Agbo, Ifeoma C; Torres, Carmen; Chah, Kennedy F

    2015-01-01

    This study was conducted to determine the species distribution, antimicrobial resistance pheno- and genotypes and virulence traits of mannitol-positive methicillin-resistant staphylococci (MRS) isolated from pigs in Nsukka agricultural zone, Nigeria. Twenty mannitol-positive methicillin-resistant coagulase-negative staphylococcal (MRCoNS) strains harboring the mecA gene were detected among the 64 Staphylococcus isolates from 291 pigs. A total of 4 species were identified among the MRCoNS isolates, namely, Staphylococcus sciuri (10 strains), Staphylococcus lentus (6 strains), Staphylococcus cohnii (3 strains) and Staphylococcus haemolyticus (one strain). All MRCoNS isolates were multidrug-resistant. In addition to β-lactams, the strains were resistant to fusidic acid (85%), tetracycline (75%), streptomycin (65%), ciprofloxacin (65%), and trimethoprim/sulphamethoxazole (60%). In addition to the mecA and blaZ genes, other antimicrobial resistance genes detected were tet(K), tet(M), tet(L), erm(B), erm(C), aacA-aphD, aphA3, str, dfrK, dfrG, cat pC221, and cat pC223. Thirteen isolates were found to be ciprofloxacin-resistant, and all harbored a Ser84Leu mutation within the QRDR of the GyrA protein, with 3 isolates showing 2 extra substitutions, Ser98Ile and Arg100Lys (one strain) and Glu88Asp and Asp96Thr (2 strains). A phylogenetic tree of the QRDR nucleotide sequences in the gyrA gene revealed a high nucleotide diversity, with several major clusters not associated with the bacterial species. Our study highlights the possibility of transfer of mecA and other antimicrobial resistance genes from MRCoNS to pathogenic bacteria, which is a serious public health and veterinary concern.

  18. Anatomical and biochemical analysis reveal the role of anthocyanins in flower coloration of herbaceous peony.

    PubMed

    Zhao, Da-Qiu; Wei, Meng-Ran; Liu, Ding; Tao, Jun

    2016-05-01

    Herbaceous peony (Paeonia lactiflora Pall.) is particularly appreciated because of its elegant and gorgeous flower color, but little is known about the underlying mechanisms of flower coloration. In this study, three P. lactiflora cultivars 'Xuefeng', 'Fenyulou' and 'Dahonglou' with white, pink and red flower were selected as the materials. Their anatomical structures, cell sap pH and metal elements were investigated, and the colored pigment mainly distributed in palisade mesophyll was only found in 'Fenyulou' and 'Dahonglou', and their shape of epidermal cells, cell sap pH and metal elements were not the key factors deciding phenotype color. Moreover, the qualitative and quantitative analysis of flavonoids were performed, their total anthocyanin, anthoxanthin and flavonoid contents were decreased during flower development, and only anthocyanin content in 'Dahonglou' was always higher than that in 'Xuefeng' and 'Fenyulou'. Subsequently, three anthocyanin compositions were found, and peonidin 3,5-di-O-glucoside (Pn3G5G) was identified as the main anthocyanin composition. In addition, the full-length of flavonol synthase gene (FLS) was isolated with the GenBank accession number KM259902, and the expression patterns of eight flavonoid biosynthetic genes showed that only PlDFR and PlANS basically had the highest levels in 'Dahonglou' and the lowest levels in 'Xuefeng', and they basically displayed a descended trend during flower development especially PlDFR, suggesting that these two genes might play a key role in the anthocyanin biosynthesis which resulted in the shift from white to pink and red in flowers. These results would contribute to understand the underlying molecular mechanisms of flower coloration in P. lactiflora. PMID:26922162

  19. Emergence of a Clonal Lineage of Multidrug-Resistant ESBL-Producing Salmonella Infantis Transmitted from Broilers and Broiler Meat to Humans in Italy between 2011 and 2014.

    PubMed

    Franco, Alessia; Leekitcharoenphon, Pimlapas; Feltrin, Fabiola; Alba, Patricia; Cordaro, Gessica; Iurescia, Manuela; Tolli, Rita; D'Incau, Mario; Staffolani, Monica; Di Giannatale, Elisabetta; Hendriksen, Rene S; Battisti, Antonio

    2015-01-01

    We report the spread of a clone of multidrug-resistant (MDR), ESBL-producing (blaCTX-M-1) Salmonella enterica subsp. enterica serovar Infantis, in the Italian broiler chicken industry and along the food-chain. This was first detected in Italy in 2011 and led to human infection in Italy in 2013-2014.A set (n = 49) of extended-spectrum cephalosporin (ESC)-resistant (R) isolates of S. Infantis (2011-2014) from humans, food-producing animals and meat thereof, were studied along with a selected set of earlier and more recent ESC-susceptible (ESC-S) isolates (n = 42, 2001-2014). They were characterized by macrorestriction-PFGE analysis and genetic environment of ESC-resistance. Isolates representative of PFGE-patterns and origin were submitted to Whole Genome Sequencing. The emerging ESC-R clone, detected mainly from broiler chickens, broiler meat and humans, showed a minimum pattern of clinical resistance to cefotaxime, tetracycline, sulfonamides, and trimethoprim, beside ciprofloxacin microbiological resistance (MIC 0.25 mg/L). All isolates of this clone harbored a conjugative megaplasmid (~ 280-320 Kb), similar to that described in ESC-susceptible S. Infantis in Israel (pESI-like) in 2014. This megaplasmid carried the ESBL gene blaCTX-M-1, and additional genes [tet(A), sul1, dfrA1 and dfrA14] mediating cefotaxime, tetracycline, sulfonamide, and trimethoprim resistance. It also contained genes conferring enhanced colonization capability, virulence (fimbriae, yersiniabactin), resistance and fitness (qacE1, mer) in the intensive-farming environment. This emerging clone of S. Infantis has been causing infections in humans, most likely through the broiler industry. Since S. Infantis is among major serovars causing human infections in Europe and is an emerging non-typhoidal Salmonella globally, further spread of this lineage in primary productions deserves quick and thorough risk-management strategies. PMID:26716443

  20. Molecular characterization of enterohemorrhagic Escherichia coli isolates from cattle.

    PubMed

    Bakhshi, Bita; Najibi, Sakine; Sepehri-Seresht, Saeed

    2014-09-01

    A total of 21 (4.3%) enterohemorrhagic E. coli strains were isolated by biochemical tests and identification of the eae(+)stx1(+)stx2(+) genotype from 490 stool samples obtained from calves with diarrhea during 1-year period from a major farm in Tehran, Iran. All of the strains showed resistance to ampicillin, ciprofloxacin, trimethoprim, streptomycin, chloramphenicol and tetracycline, while 19% showed resistance to gentamicin. Out of 21 EHEC strains, 11 (53%) harbored class 1 integron. Two different amplification products, which were approximately 750 and 1,700 bp in size, were obtained from amplified variable regions (in-F/in-R primers) in 3 (14.3%) and 4 (19%) of the EHEC isolates, which corresponded to dfrA7(dihydrofolate reductase type I) and dfrA1/aadA1(dihydrofolate reductase/aminoglycoside adenyltransferase) resistance gene cassettes, respectively, and this was confirmed by sequencing. Genotyping analysis revealed a total of 16 pulsotypes that corresponded to 16 isolates with the similarity indices of 62% and 30% for the most and least similar isolates, respectively, 9 of which harbored class 1 integron. Analysis of pulsotypes showed an extensive diversity among the isolates harboring integron, which is indicative of a lack of any significant genetic relatedness among the isolates. No obvious relation could be deduced between integron content and special pulsotypes. The little data available on the genotyping patterns of EHEC isolates from cattle and their resistance gene contents emphasize the need to establish genotyping databases in order to monitor and source track the source of emergence and spread of new resistant and integron-carrying genotypes. PMID:24920487

  1. Molecular Characterization of Enterohemorrhagic Escherichia coli Isolates from Cattle

    PubMed Central

    BAKHSHI, Bita; NAJIBI, Sakine; SEPEHRI-SERESHT, Saeed

    2014-01-01

    ABSTRACT A total of 21 (4.3%) enterohemorrhagic E. coli strains were isolated by biochemical tests and identification of the eae+stx1+stx2+ genotype from 490 stool samples obtained from calves with diarrhea during 1-year period from a major farm in Tehran, Iran. All of the strains showed resistance to ampicillin, ciprofloxacin, trimethoprim, streptomycin, chloramphenicol and tetracycline, while 19% showed resistance to gentamicin. Out of 21 EHEC strains, 11 (53%) harbored class 1 integron. Two different amplification products, which were approximately 750 and 1,700 bp in size, were obtained from amplified variable regions (in-F/in-R primers) in 3 (14.3%) and 4 (19%) of the EHEC isolates, which corresponded to dfrA7(dihydrofolate reductase type I) and dfrA1/aadA1(dihydrofolate reductase/aminoglycoside adenyltransferase) resistance gene cassettes, respectively, and this was confirmed by sequencing. Genotyping analysis revealed a total of 16 pulsotypes that corresponded to 16 isolates with the similarity indices of 62% and 30% for the most and least similar isolates, respectively, 9 of which harbored class 1 integron. Analysis of pulsotypes showed an extensive diversity among the isolates harboring integron, which is indicative of a lack of any significant genetic relatedness among the isolates. No obvious relation could be deduced between integron content and special pulsotypes. The little data available on the genotyping patterns of EHEC isolates from cattle and their resistance gene contents emphasize the need to establish genotyping databases in order to monitor and source track the source of emergence and spread of new resistant and integron-carrying genotypes. PMID:24920487

  2. Carbohydrate accumulation may be the proximate trigger of anthocyanin biosynthesis under autumn conditions in Begonia semperflorens.

    PubMed

    Zhang, K M; Li, Z; Li, Y; Li, Y H; Kong, D Z; Wu, R H

    2013-11-01

    Many plant leaves appear red in the autumn, and many papers have focused on the environmental factors and role of anthocyanin in this process. However few papers have examined the substances that are induced during this process. We hypothesised that excess sugar accumulation directly induces anthocyanin accumulation under autumn conditions. Using two methods (restricting phloem movement and exogenous sucrose feeding), we found that both surplus photosynthate and exogenous sucrose could induce anthocyanin biosynthesis, corresponding to up-regulation of several enzymes involved in anthocyanin biosynthesis (phenylalanine ammonia lyase, chalcone isomerase, dihydroflavonol 4-reductase and flavonoid 3-O-glucosyl transferase) and in transport (glutathione S-transferase). Our results suggest that excess carbohydrate may be the proximate trigger for induction of anthocyanin biosynthesis in autumn, but only when carbohydrates are accumulated for storage.

  3. Complete 6-Deoxy-d-altro-heptose Biosynthesis Pathway from Campylobacter jejuni

    PubMed Central

    McCallum, Matthew; Shaw, Steven D.; Shaw, Gary S.; Creuzenet, Carole

    2012-01-01

    The Campylobacter jejuni capsule is important for colonization and virulence in various infection models. In most strains, the capsule includes a modified heptose whose biological role and biosynthetic pathway are unknown. To decipher the biosynthesis pathway for the 6-deoxy-d-altro-heptose of strain 81-176, we previously showed that the 4,6-dehydratase WcbK and the reductase WcaG generated GDP-6-deoxy-d-manno-heptose, but the C3 epimerase necessary to form GDP-6-deoxy-d-altro-heptose was not identified. Herein, we characterized the putative C3/C5 epimerase Cjj1430 and C3/C5 epimerase/C4 reductase Cjj1427 from the capsular cluster. We demonstrate that GDP-6-deoxy-d-altro-heptose biosynthesis is more complex than anticipated and requires the sequential action of WcbK, Cjj1430, and Cjj1427. We show that Cjj1430 serves as C3 epimerase devoid of C5 epimerization activity and that Cjj1427 has no epimerization activity and only serves as a reductase to produce GDP-6-deoxy-d-altro-heptose. Cjj1430 and Cjj1427 are the only members of the C3/C5 epimerases and C3/C5 epimerase/C4 reductase families shown to have activity on a heptose substrate and to exhibit only one of their two to three potential activities, respectively. Furthermore, we show that although the reductase WcaG is not part of the main pathway, its presence and its product affect the outcome of the pathway in a complex regulatory loop involving Cjj1427. This work provides the grounds for the elucidation of similar pathways found in other C. jejuni strains and other pathogens. It provides new molecular tools for the synthesis of carbohydrate antigens useful for vaccination and for the screening of enzymatic inhibitors that may have antibacterial effects. PMID:22787156

  4. A new flavan-3,4-diol from Acacia auriculiformis by paper ionophoresis

    PubMed Central

    Drewes, S. E.; Roux, D. G.

    1966-01-01

    1. The heartwood of A. auriculiformis contains a typical mixture of analogues consisting of three isomeric flavan-3,4-diols, a dihydroflavonol, flavanone, flavonol and chalcone based on the 4′,7,8-trihydroxyl pattern. These were resolved by preparative paper chromatography and preparative paper ionophoresis. 2. Crystalline (−)-teracacidin [(2R,3R,4R)-4′,7,8-trihydroxy-2,3-cis-flavan-3,4-cis-diol] was obtained in high (10%) yield, and a new crystalline derivative of (−)-isoteracacidin [(−)-2,3-cis-3,4-trans isomer] was isolated. The crystalline methyl ether of a new (+)-2,3-trans-3,4-cis isomer was isolated. 3. The absolute configurations of (−)-isoteracacidin (2R,3R,4S) and of the (+)-2,3-trans-3,4-cis isomer (2R,3S,4S) were tentatively assigned on the basis of nuclear-magnetic-resonance spectroscopy, paper ionophoresis and paper-chromatographic comparison with the epimerization products of (−)-teracacidin. 4. Possible reasons for the absence of polymeric leuco-anthocyanidin tannins are discussed. 5. (±)-4′,7,8-Trihydroxydihydroflavonol and (±)-4′,7,8-trihydroxyflavanone were isolated for the first time. 6. The bark polyphenols consist mainly of polymeric leuco-delphinidins and leuco-cyanidins which redden exceptionally rapidly to light. The mechanism of this phenomenon is discussed. PMID:5941342

  5. Tuberculosis and nature's pharmacy of putative anti-tuberculosis agents.

    PubMed

    Chinsembu, Kazhila C

    2016-01-01

    Due to the growing problem of drug resistant Mycobacterium tuberculosis strains, coupled with the twinning of tuberculosis (TB) to human immunodeficiency virus/acquired immunodeficiency syndrome (HIV/AIDS), the burden of TB is now difficult to manage. Therefore, new antimycobacterial agents are being sought from natural sources. This review focuses on natural antimycobacterial agents from endophytes and medicinal plants of Africa, Europe, Asia, South America and Canada. In the countries mentioned in this review, numerous plant species display putative anti-TB activity. Several antimycobacterial chemical compounds have also been isolated, including: ellagitannin punicalagin, allicin, anthraquinone glycosides, iridoids, phenylpropanoids, beta-sitosterol, galanthimine, crinine, friedelin, gallic acid, ellagic acids, anthocyanidin, taraxerol, termilignan B, arjunic acid, glucopyranosides, 1-epicatechol, leucopelargonidol, hydroxybenzoic acids, benzophenanthridine alkaloids, neolignans, and decarine. These compounds may provide leads to novel and more efficacious drugs to lessen the global burden of TB and drug-resistant M. tuberculosis strains. If there is a long-term remedy for TB, it must lie in nature's pharmacy of putative antimycobacterial agents.

  6. Development of a fast extraction method and optimization of liquid chromatography-mass spectrometry for the analysis of phenolic compounds in lentil seed coats.

    PubMed

    Mirali, Mahla; Ambrose, Stephen J; Wood, Stephen A; Vandenberg, Albert; Purves, Randy W

    2014-10-15

    A systematic set of optimization experiments was conducted to design an efficient extraction and analysis protocol for screening six different sub-classes of phenolic compounds in the seed coat of various lentil (Lens culinaris Medik.) genotypes. Different compounds from anthocyanidins, flavan-3-ols, proanthocyanidins, flavanones, flavones, and flavonols sub-classes were first optimized for use as standards for liquid chromatography mass spectrometry (LC-MS) with UV detection. The effect of maceration duration, reconstitution solvent, and extraction solvent were investigated using lentil genotype CDC Maxim. Chromatographic conditions were optimized by examining column separation efficiencies, organic composition, and solvent gradient. The results showed that a 1h maceration step was sufficient and that non-acidified solvents were more appropriate; a 70:30 acetone: water (v/v) solvent was ultimately selected. Using a Kinetex PFP column, the organic concentration, gradient, and flow rate were optimized to maximize the resolution of phenolic compounds in a short 30-min analysis time. The optimized method was applied to three lentil genotypes with different phenolic compound profiles to provide information of value to breeding programs.

  7. Antioxidant Activity in Extracts of 27 Indigenous Taiwanese Vegetables

    PubMed Central

    Chao, Pi-Yu; Lin, Su-Yi; Lin, Kuan-Hung; Liu, Yu-Fen; Hsu, Ju-Ing; Yang, Chi-Ming; Lai, Jun-You

    2014-01-01

    The objectives of this study were to identify the antioxidants and antioxidant axtivity in 27 of Taiwan’s indigenous vegetables. Lycium chinense (Lc), Lactuca indica (Li), and Perilla ocymoides (Po) contained abundant quercetin (Que), while Artemisia lactiflora (Al) and Gynura bicolor (Gb) were rich in morin and kaempferol, respectively. Additionally, Nymphoides cristata (Nc) and Sechium edule (Se)-yellow had significantly higher levels of myricetin (Myr) than other tested samples. Cyanidin (Cyan) and malvidin (Mal) were abundant in Gb, Abelmoschus esculentus Moench (Abe), Po, Anisogonium esculentum (Retz.) Presl (Ane), Ipomoea batatas (Ib)-purple, and Hemerocallis fulva (Hf)-bright orange. Relatively high levels of Trolox equivalent antioxidant capacity (TEAC), oxygen radical absorption capacity (ORAC), and 1,1-diphenyl-2-picryl-hydrazyl (DPPH) radical scavenger were generated from extracts of Toona sinensis (Ts) and Po. Significant and positive correlations between antioxidant activity and polyphenols, anthocyanidins, Que, Myr, and morin were observed, indicating that these phytochemicals were some of the main components responsible for the antioxidant activity of tested plants. The much higher antioxidant activity of Po, Ts, and Ib (purple leaf) may be related to their higher Cyan, Que, and polyphenol content. PMID:24858497

  8. Antioxidant activity in extracts of 27 indigenous Taiwanese vegetables.

    PubMed

    Chao, Pi-Yu; Lin, Su-Yi; Lin, Kuan-Hung; Liu, Yu-Fen; Hsu, Ju-Ing; Yang, Chi-Ming; Lai, Jun-You

    2014-05-01

    The objectives of this study were to identify the antioxidants and antioxidant axtivity in 27 of Taiwan's indigenous vegetables. Lycium chinense (Lc), Lactuca indica (Li), and Perilla ocymoides (Po) contained abundant quercetin (Que), while Artemisia lactiflora (Al) and Gynura bicolor (Gb) were rich in morin and kaempferol, respectively. Additionally, Nymphoides cristata (Nc) and Sechium edule (Se)-yellow had significantly higher levels of myricetin (Myr) than other tested samples. Cyanidin (Cyan) and malvidin (Mal) were abundant in Gb, Abelmoschus esculentus Moench (Abe), Po, Anisogonium esculentum (Retz.) Presl (Ane), Ipomoea batatas (Ib)-purple, and Hemerocallis fulva (Hf)-bright orange. Relatively high levels of Trolox equivalent antioxidant capacity (TEAC), oxygen radical absorption capacity (ORAC), and 1,1-diphenyl-2-picryl-hydrazyl (DPPH) radical scavenger were generated from extracts of Toona sinensis (Ts) and Po. Significant and positive correlations between antioxidant activity and polyphenols, anthocyanidins, Que, Myr, and morin were observed, indicating that these phytochemicals were some of the main components responsible for the antioxidant activity of tested plants. The much higher antioxidant activity of Po, Ts, and Ib (purple leaf) may be related to their higher Cyan, Que, and polyphenol content. PMID:24858497

  9. Identification and Quantitation of Anthocyanins in Purple-Fleshed Sweet Potatoes Cultivated in China by UPLC-PDA and UPLC-QTOF-MS/MS.

    PubMed

    He, Wei; Zeng, Maomao; Chen, Jie; Jiao, Yuzhi; Niu, Fuxiang; Tao, Guanjun; Zhang, Shuang; Qin, Fang; He, Zhiyong

    2016-01-13

    The identification and quantitation of the anthocyanins in 12 purple-fleshed sweet potato (PFSP) cultivars ('Jihei 1', 'Xuzi 3', 'Xuzi 6', 'Zhezi 4', 'Ningzi 1', 'Ningzi 2', 'Ningzi 3', 'Ning 2-2', 'Ning 6-8', 'Guangzi 1', 'Ziluolan', and 'Qinzi 1') in China were carried out using a combination of ultraperformance liquid chromatography-photodiode array (UPLC-PDA), quadrupole-time-of-flight mass spectrometry (QTOF-MS), and tandem mass spectrometry (MS/MS) analyses. Thirteen acylated anthocyanins were tentatively characterized, including two new PFSP anthocyanins, cyanidin 3-caffeoyl-vanilloyl sophoroside-5-glucoside and peonidin 3-caffeoyl-vanilloyl sophoroside-5-glucoside. The quantitative analyses of these anthocyanins were conducted using cyanidin 3-O-glucoside as a standard. The total anthocyanin content of the PFSPs depended on the cultivar. The five PFSP cultivars with the highest content of anthocyanins were 'Jihei 1', 'Xuzi 3', 'Zhezi 4', 'Ziluolan', and 'Qinzi 1'. This is the first report of the 'Ningzi 2', 'Ningzi 3', and 'Ning 2-2' PFSP cultivars containing only diacylated anthocyanins and of the 'Xuzi 6' cultivar containing single anthocyanidin-based anthocyanins.

  10. Identification and Quantitation of Anthocyanins in Purple-Fleshed Sweet Potatoes Cultivated in China by UPLC-PDA and UPLC-QTOF-MS/MS.

    PubMed

    He, Wei; Zeng, Maomao; Chen, Jie; Jiao, Yuzhi; Niu, Fuxiang; Tao, Guanjun; Zhang, Shuang; Qin, Fang; He, Zhiyong

    2016-01-13

    The identification and quantitation of the anthocyanins in 12 purple-fleshed sweet potato (PFSP) cultivars ('Jihei 1', 'Xuzi 3', 'Xuzi 6', 'Zhezi 4', 'Ningzi 1', 'Ningzi 2', 'Ningzi 3', 'Ning 2-2', 'Ning 6-8', 'Guangzi 1', 'Ziluolan', and 'Qinzi 1') in China were carried out using a combination of ultraperformance liquid chromatography-photodiode array (UPLC-PDA), quadrupole-time-of-flight mass spectrometry (QTOF-MS), and tandem mass spectrometry (MS/MS) analyses. Thirteen acylated anthocyanins were tentatively characterized, including two new PFSP anthocyanins, cyanidin 3-caffeoyl-vanilloyl sophoroside-5-glucoside and peonidin 3-caffeoyl-vanilloyl sophoroside-5-glucoside. The quantitative analyses of these anthocyanins were conducted using cyanidin 3-O-glucoside as a standard. The total anthocyanin content of the PFSPs depended on the cultivar. The five PFSP cultivars with the highest content of anthocyanins were 'Jihei 1', 'Xuzi 3', 'Zhezi 4', 'Ziluolan', and 'Qinzi 1'. This is the first report of the 'Ningzi 2', 'Ningzi 3', and 'Ning 2-2' PFSP cultivars containing only diacylated anthocyanins and of the 'Xuzi 6' cultivar containing single anthocyanidin-based anthocyanins. PMID:26687974

  11. Cytoprotective Mechanism of Cyanidin and Delphinidin against Oxidative Stress-Induced Tenofibroblast Death

    PubMed Central

    Nam, Dae Cheol; Hah, Young Sool; Nam, Jung Been; Kim, Ra Jeong; Park, Hyung Bin

    2016-01-01

    Age-related rotator cuff tendon degeneration is related to tenofibroblast apoptosis. Anthocyanins reduce oxidative stress-induced apoptotic cell death in tenofibroblasts. The current study investigated the presence of cell protective effects in cyanidin and delphinidin, the most common aglycon forms of anthocyanins. We determined whether these anthocyanidins have antiapoptotic and antinecrotic effects in tenofibroblasts exposed to H2O2, and evaluated their biomolecular mechanisms. Both cyanidin and delphinidin inhibited H2O2-induced apoptosis in a dose-dependent manner. However, at concentrations of 100 μg/ml or greater, delphinidin showed cytotoxicity against tenofibroblasts and a decreased antinecrotic effect. Cyanidin and delphinidin both showed inhibitory effects on the H2O2-induced increase in intracellular ROS formation and the activation of ERK1/2 and JNK. In conclusion, both cyanidin and delphinidin have cytoprotective effects on cultured tenofibroblasts exposed to H2O2. These results suggest that cyanidin and delphinidin are both beneficial for the treatment of oxidative stress-mediated tenofibroblast cell death, but their working concentrations are different. PMID:27098861

  12. Analysis of stereochemistry and biosynthesis of epicatechin in tea plants by chiral phase high performance liquid chromatography.

    PubMed

    Qian, Yumei; Zhao, Xianqian; Zhao, Lei; Cui, Lilan; Liu, Li; Jiang, Xiaolan; Liu, Yajun; Gao, Liping; Xia, Tao

    2015-12-01

    Tea (Camellia sinensis) is rich in flavan-3-ols (catechins), especially epicatechin (EC), which is the predominant extension unit of polymeric proanthocyanidins (PAs). However, studies assessing EC's stereochemistry are scarce. Here, a high performance liquid chromatography column using amylose tris-(3, 5-dimethylphenylcarbamate) immobilized on silica-gel as chiral stationary phases (CSPs) was applied to explore its stereochemistry and biosynthetic pathway in tea plants. The results revealed (-)-epicatechin [(-)-EC] was the predominant di-hyroxy-non-galloylated-catechins, while (+)-epicatechin [(+)-EC] was not detected. Interestingly, (-)-EC was the only product obtained from cyanidin using the partially purified native C. sinensis anthocyanidin reductase (CsANR) in the presence of reduction nicotinamide adenine dinucleotide phosphate (NADPH); meanwhile, (+)-EC was the main product using recombinant CsANR in the same conditions. In addition, (-)-EC could be obtained from (+)-catechin [(+)-C] using recombinant CsANR, which displayed C3-epimerase activity in the presence of oxidation nicotinamide adenine dinucleotide phosphate (NADP(+)). But the partially purified native CsANR did not possess this function. Finally, (-)-EC could result from the de-gallate acid reaction of epicatechin gallate (ECG) catalyzed by a novel partially purified native galloylated catechins hydrolase (GCH) from tea leaves. In summary, (-)-EC is likely the product of native protein from the tea plants, and (+)-EC is only produced in a reaction catalyzed by recombinant CsANR in vitro.

  13. Classification of fruits based on anthocyanin types and relevance to their health effects.

    PubMed

    Fang, Jim

    2015-01-01

    Anthocyanins are a group of water-soluble pigments that confer the blue, purple, and red color to many fruits. Anthocyanin-rich fruits can be divided into three groups based on the types of aglycones of their anthocyanins: pelargonidin group, cyanidin/peonidin group, and multiple anthocyanidins group. Some fruits contain a major anthocyanin type and can serve as useful research tools. Cyanidin glycosides and peonidin glycosides can be metabolically converted to each other by methylation and demethylation. Both cyanidin and peonidin glycosides can be metabolized to protocatechuic acid and vanillic acid. Pelargonidin-3-glucoside is metabolized to 4-hydroxybenoic acid. On the other hand, phenolic acid metabolites of delphinidin, malvidin, and petunidin glycosides are unstable and can be further fragmented into smaller molecules. A literature review indicates berries with higher cyanidin content, such as black raspberries, chokeberries, and bilberries are more likely to produce an antiinflammatory effect. This observation seems to be consistent with the hypothesis that one or more stable phenolic acid metabolites contribute to the antiinflammatory effects of anthocyanin-rich fruits. More studies are needed before we can conclude that fruits rich in cyanidin, peonidin, or pelargonidin glycosides have better antiinflammatory effects. Additionally, fruit polyphenols other than anthocyanins could contribute to their antiinflammatory effects. Furthermore, blueberries could exert their health effects with other mechanisms such as improving intestinal microbiota composition. In summary, this classification system can facilitate our understanding of the absorption and metabolic processes of anthocyanins and the health effects of different fruits.

  14. The Transcriptional Repressor MYB2 Regulates Both Spatial and Temporal Patterns of Proanthocyandin and Anthocyanin Pigmentation in Medicago truncatula.

    PubMed

    Jun, Ji Hyung; Liu, Chenggang; Xiao, Xirong; Dixon, Richard A

    2015-10-01

    Accumulation of anthocyanins and proanthocyanidins (PAs) is limited to specific cell types and developmental stages, but little is known about how antagonistically acting transcriptional regulators work together to determine temporal and spatial patterning of pigmentation at the cellular level, especially for PAs. Here, we characterize MYB2, a transcriptional repressor regulating both anthocyanin and PA biosynthesis in the model legume Medicago truncatula. MYB2 was strongly upregulated by MYB5, a major regulator of PA biosynthesis in M. truncatula and a component of MYB-basic helix loop helix-WD40 (MBW) activator complexes. Overexpression of MYB2 abolished anthocyanin and PA accumulation in M. truncatula hairy roots and Arabidopsis thaliana seeds, respectively. Anthocyanin deposition was expanded in myb2 mutant seedlings and flowers accompanied by increased anthocyanin content. PA mainly accumulated in the epidermal layer derived from the outer integument in the M. truncatula seed coat, starting from the hilum area. The area of PA accumulation and ANTHOCYANIDIN REDUCTASE expression was expanded into the seed body at the early stage of seed development in the myb2 mutant. Genetic, biochemical, and cell biological evidence suggests that MYB2 functions as part of a multidimensional regulatory network to define the temporal and spatial pattern of anthocyanin and PA accumulation linked to developmental processes.

  15. Genus vaccinium: medicine, cosmetics, and coatings.

    PubMed

    Johnson, Brandy J; Lin, Baochuan; Bongard, Jason E

    2010-06-01

    Vaccinium is a genus of shrubs several of which, including cranberry, lingonberry, and blueberry, produce edible fruit. Consumption of the fruit of these plants and juices pressed from it has long been known to have healthful effects. A number of functional compounds have been extracted from the fruits and seeds of these plants. Anthocyanidins and related compounds have been reported to possess antimicrobial, antioxidant, and anti-inflammatory properties. Extracts of the fruits have been applied to the inhibition of non-enzymatic glycosylation in anti-aging preparations. The oil of the cranberry seed is high in antioxidants and is a source of omega-3, -6, and -9 fatty acids making it an attractive cosmetics component. The inhibition of tumor growth, angiogenesis, and metastasis by extracts of these fruits has been described. Extracts of the plants have also been applied to the development of antimicrobial paints and coatings. Here, we provide an overview of the wide range of applications described for various compounds from and preparations of these plants and survey the recent patents related to those applications.

  16. Flavonoid intake and the risk of age-related cataract in China's Heilongjiang Province

    PubMed Central

    Ma, Yingna; Gao, Weiqi; Wu, Kun; Bao, Yongping

    2015-01-01

    Background/objectives Epidemiological evidence suggests that diets rich in flavonoids may reduce the risk of developing age-related cataract (ARC). Flavonoids are widely distributed in foods of plant origin, and the objective of this study was to evaluate retrospectively the association between the intakes of the five flavonoid subclasses and the risk of ARC. Subjects/methods A population-based case-control study (249 cases and 66 controls) was carried out in Heilongjiang province, which is located in the northeast of China, and where intakes and availability of fresh vegetables and fruits can be limited. Dietary data gathered by food-frequency questionnaire (FFQ) were used to calculate flavonoid intake. Adjusted odds ratio (OR) and 95% confidence interval (CI) were estimated by logistic regression. Results No linear associations between risk of developing ARC and intakes of total dietary flavonoids, anthocyanidins, flavon-3-ol, flavanone, total flavones or total flavonols were found, but quercetin and isorhamnetin intake was inversely associated with ARC risk (OR 11.78, 95% CI: 1.62–85.84, p<0.05, and OR 6.99, 95% CI: 1.12–43.44, p<0.05, quartile 4 vs. quartile 1, respectively). Conclusion As quercetin is contained in many plant foods and isorhamnetin in very few foods, we concluded that higher quercetin intake may be an important dietary factor in the reduction of the risk of ARC. PMID:26652740

  17. Low medium pH value enhances anthocyanin accumulation in Malus crabapple leaves.

    PubMed

    Zhang, Yanchen; Zhang, Jie; Song, Tingting; Li, Jinyan; Tian, Ji; Jin, Kaina; Yao, Yuncong

    2014-01-01

    Anthocyanin is a critical factor involved in coloration of plant tissues, but the mechanism how medium pH values affect anthocyanin accumulation in woody plants is unknown. We analyzed anthocyanin composition and the expression of elements encoding anthocyanin and flavonols biosynthesis underlying different medium pH values by using three different leave color type cultivars. HPLC analysis demonstrated that high medium pH values treatment induced a dramatic decrease in the concentration of cyaniding in crabapple leaves. Conversely, the high medium pH values induced up-regulation of the content of flavones and flavonols, suggesting that low pH treatment-induced anthocyanin accumulation. Quantitative real time PCR experiment showed the expression level of anthocyanidin synthase (McANS) and uridine diphosphate glucose flavonoid 3-O-glucosyltransferase (McUFGT) was up-regulated by low pH values treatment, and high medium pH value treatment up-regulate the transcription level of flavonol synthase (McFLS). Meanwhile, several MYB TFs have been suggested in the regulation of pH responses. These results strongly indicate that the low pH treatment-induced anthocyanin accumulation is mediated by the variation of mRNA transcription of the anthocyanin biosynthetic genes. PMID:24914811

  18. The Transcriptional Repressor MYB2 Regulates Both Spatial and Temporal Patterns of Proanthocyandin and Anthocyanin Pigmentation in Medicago truncatula[OPEN

    PubMed Central

    2015-01-01

    Accumulation of anthocyanins and proanthocyanidins (PAs) is limited to specific cell types and developmental stages, but little is known about how antagonistically acting transcriptional regulators work together to determine temporal and spatial patterning of pigmentation at the cellular level, especially for PAs. Here, we characterize MYB2, a transcriptional repressor regulating both anthocyanin and PA biosynthesis in the model legume Medicago truncatula. MYB2 was strongly upregulated by MYB5, a major regulator of PA biosynthesis in M. truncatula and a component of MYB-basic helix loop helix-WD40 (MBW) activator complexes. Overexpression of MYB2 abolished anthocyanin and PA accumulation in M. truncatula hairy roots and Arabidopsis thaliana seeds, respectively. Anthocyanin deposition was expanded in myb2 mutant seedlings and flowers accompanied by increased anthocyanin content. PA mainly accumulated in the epidermal layer derived from the outer integument in the M. truncatula seed coat, starting from the hilum area. The area of PA accumulation and ANTHOCYANIDIN REDUCTASE expression was expanded into the seed body at the early stage of seed development in the myb2 mutant. Genetic, biochemical, and cell biological evidence suggests that MYB2 functions as part of a multidimensional regulatory network to define the temporal and spatial pattern of anthocyanin and PA accumulation linked to developmental processes. PMID:26410301

  19. Bioavailability of Dietary Polyphenols and Gut Microbiota Metabolism: Antimicrobial Properties

    PubMed Central

    Miguélez, Elisa M.; Villar, Claudio J.

    2015-01-01

    Polyphenolic compounds are plant nutraceuticals showing a huge structural diversity, including chlorogenic acids, hydrolyzable tannins, and flavonoids (flavonols, flavanones, flavan-3-ols, anthocyanidins, isoflavones, and flavones). Most of them occur as glycosylated derivatives in plants and foods. In order to become bioactive at human body, these polyphenols must undergo diverse intestinal transformations, due to the action of digestive enzymes, but also by the action of microbiota metabolism. After elimination of sugar tailoring (generating the corresponding aglycons) and diverse hydroxyl moieties, as well as further backbone reorganizations, the final absorbed compounds enter the portal vein circulation towards liver (where other enzymatic transformations take place) and from there to other organs, including behind the digestive tract or via blood towards urine excretion. During this transit along diverse tissues and organs, they are able to carry out strong antiviral, antibacterial, and antiparasitic activities. This paper revises and discusses these antimicrobial activities of dietary polyphenols and their relevance for human health, shedding light on the importance of polyphenols structure recognition by specific enzymes produced by intestinal microbial taxa. PMID:25802870

  20. Preventive and Prophylactic Mechanisms of Action of Pomegranate Bioactive Constituents

    PubMed Central

    Viladomiu, Monica; Hontecillas, Raquel; Lu, Pinyi; Bassaganya-Riera, Josep

    2013-01-01

    Pomegranate fruit presents strong anti-inflammatory, antioxidant, antiobesity, and antitumoral properties, thus leading to an increased popularity as a functional food and nutraceutical source since ancient times. It can be divided into three parts: seeds, peel, and juice, all of which seem to have medicinal benefits. Several studies investigate its bioactive components as a means to associate them with a specific beneficial effect and develop future products and therapeutic applications. Many beneficial effects are related to the presence of ellagic acid, ellagitannins (including punicalagins), punicic acid and other fatty acids, flavonoids, anthocyanidins, anthocyanins, estrogenic flavonols, and flavones, which seem to be its most therapeutically beneficial components. However, the synergistic action of the pomegranate constituents appears to be superior when compared to individual constituents. Promising results have been obtained for the treatment of certain diseases including obesity, insulin resistance, intestinal inflammation, and cancer. Although moderate consumption of pomegranate does not result in adverse effects, future studies are needed to assess safety and potential interactions with drugs that may alter the bioavailability of bioactive constituents of pomegranate as well as drugs. The aim of this review is to summarize the health effects and mechanisms of action of pomegranate extracts in chronic inflammatory diseases. PMID:23737845

  1. Intermolecular binding of blueberry pectin-rich fractions and anthocyanin.

    PubMed

    Lin, Z; Fischer, J; Wicker, L

    2016-03-01

    Pectin was extracted from blueberry powder into three fractions of water soluble (WSF), chelator soluble (CSF) and sodium carbonate soluble (NSF). The fractions were incubated with cyanidin-3-glucoside (C3G), a mixture of five anthocyanidins (cyanidin, pelargonidin, malvidin, petunidin and delphinidin) or blueberry juice at pH 2.0-4.5. Free anthocyanins and bound anthocyanin-pectin mixtures were separated by ultrafiltration. WSF bound the least amount of anthocyanin at all pH values. CSF had stronger anthocyanin binding ability at pH 2.0-3.6, while NSF had stronger anthocyanin binding ability at pH 3.6-4.5. The pectin and anthocyanin binding was lowest at pH 4.5 and higher at pH 2.0-3.6. Nearly doubling C3G pigment content increased bound anthocyanin percentage by 16-23% at pH 3.6, which favored anthocyanin aromatic stacking, compared to 3-9% increase at pH 2.0. Ionic interaction between anthocyanin flavylium cations and free pectic carboxyl groups, and anthocyanin stacking may be two major mechanisms for pectin and anthocyanin binding. PMID:26471644

  2. Contribution of Anthocyanin Composition to Total Antioxidant Capacity of Berries.

    PubMed

    Lee, Sang Gil; Vance, Terrence M; Nam, Tae-Gyu; Kim, Dae-Ok; Koo, Sung I; Chun, Ock K

    2015-12-01

    The present study aimed to evaluate the contribution of anthocyanin composition to the total antioxidant capacity (TAC) of berries having different anthocyanin composition; blackberry, black currant, and blueberry. Blackberry demonstrated the highest TAC, while it had the lowest total anthocyanin content among the three berries in both of the phenolic extract and anthocyanin fractions. On the other hand, black currant had the highest total anthocyanin content, but the lowest TAC. Cyanidin-3-O-glucoside (cya-3-glc) accounted for 94% of blackberry anthocyanins, and as one of the strongest antioxidants present in these three berries, it substantially contributed to the TAC of blackberry anthocyanin fraction (96.0%). Delphinidin-3-O-rutinoside and cyanidin-3-O-rutinoside in black currant had lower antioxidant capacities compared with delphinin-3-O-glucoside and cya-3-glc, resulting in its lowest TAC among berry anthocyanin fractions examined. Malvidin derivatives, major anthocyanins of blueberry, had considerably lower antioxidant capacity than other anthocyanidin derivatives, such as cyanidin or delphinidin, resulting in lower TAC of blueberry compared with blackberry. Our findings indicate that anthocyanin composition as well as the antioxidant capacity of individual anthocyanins contributes to the TAC of berries rich in distinct anthocyanins. PMID:26515081

  3. Investigation of Antioxidant Activity of Pomegranate Juices by Means of Electron Paramagnetic Resonance and UV-Vis Spectroscopy.

    PubMed

    Kozik, Violetta; Jarzembek, Krystyna; Jędrzejowska, Agnieszka; Bąk, Andrzej; Polak, Justyna; Bartoszek, Mariola; Pytlakowska, Katarzyna

    2015-01-01

    Pomegranate fruit (Punica granatum L.) is a source of numerous phenolic compounds, and it contains flavonoids such as anthocyanins, anthocyanidins, cyanidins, catechins and other complexes of flavonoids, ellagitannins, and hydrolyzed tannins. Pomegranate juice shows antioxidant, antiproliferative, and anti-atherosclerotic properties. The antioxidant capacity (TEAC) of the pomegranate juices was measured using electron paramagnetic resonance (EPR) spectroscopy and 1,1-diphenyl-2-picrylhydrazyl (DPPH•) as a source of free radicals, and the total phenolic (TP) content was measured using UV-Vis spectroscopy. All the examined pomegranate juices exhibited relatively high antioxidant properties. The TEAC values determined by means of EPR spectroscopy using Trolox (TE) as a free radical scavenger were in the range of 463.12 to 1911.91 μmol TE/100 mL juice. The TP content measured by the Folin-Ciocalteu method, using gallic acid (GA) as a free radical scavenger, widely varied in the investigated pomegranate juice samples and ranged from 1673.62 to 5263.87 mg GA/1 L juice. The strongest antioxidant properties were observed with the fresh pomegranate juices obtained from the fruits originating from Israel, Lebanon, and Azerbaijan. Correlation analysis of numerical data obtained by means of EPR spectroscopy (TEAC) and UV-Vis spectroscopy (TP) gave correlation coefficient (r)=0.90 and determination coefficient (r2)=0.81 (P<0.05).

  4. Isolation and analysis of bioactive constituents of sour cherry (Prunus cerasus) seed kernel: an emerging functional food.

    PubMed

    Bak, Istvan; Lekli, Istvan; Juhasz, Bela; Varga, Edit; Varga, Balazs; Gesztelyi, Rudolf; Szendrei, Levente; Tosaki, Arpad

    2010-08-01

    A plant-based diet reduces the risk for the development of several chronic diseases, such as ischemic heart disease or cancer due to natural compounds found in plants. Numerous cereals, berries, fruits, and vegetables, including sour cherry (Prunus cerasus), which is a favored fruit worldwide, contain biological active components. The antioxidant components of the sour cherry seed kernel have not been investigated until now. The aim of our study was to isolate and analyze the bioactive constituents of sour cherry seed kernel. We separated the oil fraction of the kernel; then the remaining solid fraction was dried, and the oil-free kernel extract was further analyzed. Our results show that sour cherry seed kernel oil contains vegetable oils including unsaturated fatty acids, oleic acids, alpha-tocopherol, tocotrienols, and tocopherol-like components. The components of the solid fraction include various bioactive structures such as polyphenols, flavonoids, vegetable acids, and pro- and anthocyanidins, which could have useful therapeutic effects in the prevention of various vascular diseases. PMID:20482278

  5. Degradation products of cyanidin glycosides from tart cherries and their bioactivities.

    PubMed

    Seeram, N P; Bourquin, L D; Nair, M G

    2001-10-01

    The bioactive anthocyanins present in tart cherries, Prunus cerasus L. (Rosaceae) cv. Balaton, are cyanidin 3-glucosylrutinoside (1), cyanidin 3-rutinoside (2), and cyanidin 3-glucoside (3). Cyanidin (4) is the major anthocyanidin in tart cherries. In our continued evaluation of the in vivo and in vitro efficacy of these anthocyanins to prevent inflammation and colon cancer, we have added these compounds to McCoy's 5A medium in an effort to identify their degradation products during in vitro cell culture studies. This resulted in the isolation and characterization of protocatechuic acid (5), the predominant degradation product. In addition, 2,4-dihydroxybenzoic acid (6) and 2,4,6-trihydroxybenzoic acid (7) were identified as degradation products. However, these degradation products were not quantified. Compounds 5-7 were also identified as degradation products when anthocyanins were subjected to varying pH and thermal conditions. In cyclooxygenase (COX)-I and -II enzyme inhibitory assays, compounds 5-7 did not show significant activities when compared to the NSAIDs Naproxen, Celebrex, and Vioxx, or Ibuprofen, at 50 microM concentrations. However, at a test concentration of 50 microM, the antioxidant activity of protocatechuic acid (5) was comparable to those of the commercial antioxidants tert-butylhydroquinone (TBHQ), butylated hydroxytoluene (BHT), and butylated hydroxyanisole (BHA), and superior to that of vitamin E at 10 microM concentrations. PMID:11600045

  6. Cloning and Characterization of a Flavonol Synthase Gene from Scutellaria baicalensis

    PubMed Central

    Kim, Yeon Bok; Kim, KwangSoo; Kim, YeJi; Tuan, Pham Anh; Kim, Haeng Hoon; Cho, Jin Woong; Park, Sang Un

    2014-01-01

    Flavonols are the most abundant of all the flavonoids and play pivotal roles in a variety of plants. We isolated a cDNA clone encoding flavonol synthase from Scutellaria baicalensis (SbFLS). The SbFLS cDNA is 1011 bp long, encodes 336 amino acid residues, and belongs to a family of 2-oxoglutarate-dependent dioxygenases. The overall structure of SbFLS is very similar to that of Arabidopsis thaliana anthocyanidin synthase (AtANS), with a β jelly-roll fold surrounded by tens of short and long α-helices. SbFLS was constitutively expressed in the roots, stems, leaves, and flowers, with particularly high expression in the roots and flowers. SbFLS transcript levels in the roots were 376-, 70-, and 2.5-fold higher than in the leaves, stems, and flowers. The myricetin content was significantly higher than that of kaempferol and quercetin. Therefore, we suggest that SbFLS mediates flavonol formation in the different organs of S. baicalensis. Our study may contribute to the knowledge of the role of FLS in S. baicalensis. PMID:24672406

  7. Anthocyanins as Functional Food Colors

    NASA Astrophysics Data System (ADS)

    Motohashi, Noboru; Sakagami, Hiroshi

    Anthocyanins, a proanthocyanidin-type of flavonoid, contain an abundance of functional phytochemicals and occur in fruits such as cranberry, blueberry, orange, apple and in vegetables such as tomato, sweet pepper, spinach, and radishes. Functional and essential diets have been ingested in daily life since the primitive era of history. When anthocyanins are coupled with some water-soluble sugar molecules, their color becomes red, yellow, violet, or blue. It is very intriguing that anthocyanins provide the colorful variety of pigments for pansies, petunias, plums, and other diverse flowers. Chlorophyll in various fruits and vegetables is the main green phyto-component, while anthocyanins are probably the most important visible plant pigments in the natural kingdom having specific colors. Anthocyanins have been clinically used in many folklore medicines worldwide. Anthocyanins could provide health benefits for age-related diseases as well as other diseases. Anthocyanins have higher antioxidant capacity against oxidative stress induced by excess reactive oxygen species (ROS) such as superoxide radicals, hydrogen peroxide, and thus the human body might be protected from oxidative injury by anthocyanins. On the basis of these facts, we review the synthesis of plant flavonoids and their ability to scavenge oxidants, inhibit or activate enzymes, and the safety of proanthocyanidins and anthocyanidins present in common foods.

  8. Persimmon (Diospyros kaki) fruit: hidden phytochemicals and health claims

    PubMed Central

    Butt, Masood Sadiq; Sultan, M. Tauseef; Aziz, Mahwish; Naz, Ambreen; Ahmed, Waqas; Kumar, Naresh; Imran, Muhammad

    2015-01-01

    Currently, nutrition and health linkages focused on emerging strategy of diet based regimen to combat various physiological threats including cardiovascular disorders, oxidative stress, diabetes mellitus, etc. In this context, consumption of fruits and vegetables is gaining considerable importance as safeguard to maintain human health. Likewise, their phytochemicals and bioactive molecules are also becoming popular as promising demulcent against various ailments. The current review is an effort to sum up information regarding persimmon fruit with special reference to its phytochemistry and associated health claims. Accordingly, the role of its certain bioactive molecules like proanthocyanidin, carotenoids, tannins, flavonoids, anthocyanidin, catechin, etc. is highlighted. Owing to rich phytochemistry, persimmon and its products are considered effective in mitigating oxidative damage induced by reactive oxygen species (ROS). The antioxidant potential is too responsible for anti-malignant and anti-melanogenic perspectives of persimmon functional ingredients. Additionally, they are effectual in soothing lifestyle related disparities e.g. cardiovascular disorders and diabetes mellitus. There are proven facts that pharmacological application of persimmon or its functional ingredients like proanthocyanidin may helps against hyperlipidemia and hyperglycemia. Nevertheless, astringent taste and diospyrobezoars formation are creating lacuna to prop up its vitality. In toto, persimmon and its components hold potential as one of effective modules in diet based therapy; however, integrated research and meta-analysis are still required to enhance meticulousness. PMID:27047315

  9. Extracts, anthocyanins and procyanidins from Aronia melanocarpa as radical scavengers and enzyme inhibitors.

    PubMed

    Bräunlich, Marie; Slimestad, Rune; Wangensteen, Helle; Brede, Cato; Malterud, Karl E; Barsett, Hilde

    2013-03-01

    Extracts, subfractions, isolated anthocyanins and isolated procyanidins B2, B5 and C1 from the berries and bark of Aronia melanocarpa were investigated for their antioxidant and enzyme inhibitory activities. Four different bioassays were used, namely scavenging of the diphenylpicrylhydrazyl (DPPH) radical, inhibition of 15-lipoxygenase (15-LO), inhibition of xanthine oxidase (XO) and inhibition of α-glucosidase. Among the anthocyanins, cyanidin 3-arabinoside possessed the strongest and cyanidin 3-xyloside the weakest radical scavenging and enzyme inhibitory activity. These effects seem to be influenced by the sugar units linked to the anthocyanidin. Subfractions enriched in procyanidins were found to be potent α-glucosidase inhibitors; they possessed high radical scavenging properties, strong inhibitory activity towards 15-LO and moderate inhibitory activity towards XO. Trimeric procyanidin C1 showed higher activity in the biological assays compared to the dimeric procyanidins B2 and B5. This study suggests that different polyphenolic compounds of A. melanocarpa can have beneficial effects in reducing blood glucose levels due to inhibition of α-glucosidase and may have a potential to alleviate oxidative stress.

  10. Enzymic and Protein Character of Tonoplast from Hippeastrum Vacuoles 1

    PubMed Central

    Wagner, George J.

    1981-01-01

    The membrane of anthocyanin containing Hippeatrum petal vacuoles was examined for protein and enzyme content after purification by equilibrium density centrifugation. Light scattering, protein, and a Mg2+-dependent nucleotide specific ATPase were associated with membrane having a density of 1.08 to 1.12 grams per cubic centimeter. A small amount of acid phosphatase was also present in this region of the gradient, but this activity peaked at about 1.12 grams per cubic centimeter. A component of yeast tonoplast, α-mannosidase, was not significantly present. UDP-glucose, anthocyanidin-3-O-glucosyltransferase, thought to be a cytosol enzyme in Hippeastrum, was absent from tonoplast of vacuoles isolated by osmotic shock in 0.2 molar K2HPO4 or 0.35 molar mannitol. Vacuolar acid phosphatase was insensitive to ethylenediaminetetraacetate but was 80% inhibited by 10 millimolar KF, while ATPase was inactivated by 2 millimolar ethylenediaminetetraacetate and only 50% inhibited by 10 millimolar KF. Five major and about 9 minor polypeptides were detected by sodium dodecyl sulfate-polyacrylamide gel electrophoresis of membrane protein on 5 to 30 and 6 to 16% gradient gels. Images PMID:16661944

  11. Isolation and analysis of bioactive constituents of sour cherry (Prunus cerasus) seed kernel: an emerging functional food.

    PubMed

    Bak, Istvan; Lekli, Istvan; Juhasz, Bela; Varga, Edit; Varga, Balazs; Gesztelyi, Rudolf; Szendrei, Levente; Tosaki, Arpad

    2010-08-01

    A plant-based diet reduces the risk for the development of several chronic diseases, such as ischemic heart disease or cancer due to natural compounds found in plants. Numerous cereals, berries, fruits, and vegetables, including sour cherry (Prunus cerasus), which is a favored fruit worldwide, contain biological active components. The antioxidant components of the sour cherry seed kernel have not been investigated until now. The aim of our study was to isolate and analyze the bioactive constituents of sour cherry seed kernel. We separated the oil fraction of the kernel; then the remaining solid fraction was dried, and the oil-free kernel extract was further analyzed. Our results show that sour cherry seed kernel oil contains vegetable oils including unsaturated fatty acids, oleic acids, alpha-tocopherol, tocotrienols, and tocopherol-like components. The components of the solid fraction include various bioactive structures such as polyphenols, flavonoids, vegetable acids, and pro- and anthocyanidins, which could have useful therapeutic effects in the prevention of various vascular diseases.

  12. Tuberculosis and nature's pharmacy of putative anti-tuberculosis agents.

    PubMed

    Chinsembu, Kazhila C

    2016-01-01

    Due to the growing problem of drug resistant Mycobacterium tuberculosis strains, coupled with the twinning of tuberculosis (TB) to human immunodeficiency virus/acquired immunodeficiency syndrome (HIV/AIDS), the burden of TB is now difficult to manage. Therefore, new antimycobacterial agents are being sought from natural sources. This review focuses on natural antimycobacterial agents from endophytes and medicinal plants of Africa, Europe, Asia, South America and Canada. In the countries mentioned in this review, numerous plant species display putative anti-TB activity. Several antimycobacterial chemical compounds have also been isolated, including: ellagitannin punicalagin, allicin, anthraquinone glycosides, iridoids, phenylpropanoids, beta-sitosterol, galanthimine, crinine, friedelin, gallic acid, ellagic acids, anthocyanidin, taraxerol, termilignan B, arjunic acid, glucopyranosides, 1-epicatechol, leucopelargonidol, hydroxybenzoic acids, benzophenanthridine alkaloids, neolignans, and decarine. These compounds may provide leads to novel and more efficacious drugs to lessen the global burden of TB and drug-resistant M. tuberculosis strains. If there is a long-term remedy for TB, it must lie in nature's pharmacy of putative antimycobacterial agents. PMID:26464047

  13. Dark purple nectar as a foraging signal in a bird-pollinated Himalayan plant.

    PubMed

    Zhang, Feng-Ping; Cai, Xiang-Hai; Wang, Hong; Ren, Zong-Xin; Larson-Rabin, Zachary; Li, De-Zhu

    2012-01-01

    • Some plants secrete coloured nectar to attract pollinators, but little is known about the chemical origins of nectar colouration and its ecological function. Leucosceptrum canum stands out as the only plant with coloured nectar recorded in the Himalayas. Here, we focused on the compound associated with the dark colour of the nectar, as well as its secretion dynamics during the flowering season and its relationship to pollinators. • Fresh nectar was analysed by semi-preparative reversed-phase high-performance liquid chromatography (HPLC), LC-MS and HRESIMS (high resolution electronspray ionization mass spectroscopy) to determine which compound causes the nectar colouration. Behavioural experiments were conducted with birds and honeybees to elucidate the effect of the nectar colour and volume on pollinators. • We identified a purple anthocyanidin, 5-hydroxyflavylium, as a natural nectar product for the first time. Two short-billed birds were found to pollinate this plant, which employs two nectar-based mechanisms to direct bird pollinators to reproductively active flowers, controlling nectar palatability and presenting a foraging signal for birds by altering nectar volume and colour in a developmental stage-specific manner. • 5-Hydroxyflavylium was found to be the cause of the nectar colouration, the function of which is to act as a foraging signal to increase pollination efficiency through nectar visibility and palatability.

  14. Enzymic and protein character of tonoplast from Hippeastrum vacuoles

    SciTech Connect

    Wagner, G.J.

    1981-01-01

    The membrane of anthocyanin containing Hippeastrum petal vacuoles was examined for protein and enzyme content after purification by equilibrium density centrifugation. Light scattering, protein, and a Mg/sup 2 +/ -dependent nucleotide specific ATPase were associated with membrane having a density of 1.08 to 1.12 grams per cubic centimeter. A small amount of acid phosphatase was also present in this region of the gradient, but this activity peaked at about 1.12 grams per cubic centimeter. A component of yeast tonoplast, ..cap alpha..-mannosidase, was not significantly present. UDP-glucose, anthocyanidin-3-O-glucosyltransferase, thought to be a cytosol enzyme in Hippeastrum, was absent from tonoplast of vacuoles isolated by osmotic shock in 0.2 molar K/sub 2/HPO/sub 4/ or 0.35 molar mannitol. Vacuolar acid phosphatase was insensitive to ethylenediaminetetraacetate but was 80% inhibited by 10 millimolar KF, while ATPase was inactivated by 2 millimolar ethylenediaminetetraacetate and only 50% inhibited by 10 millimolar KF,. Five major and about 9 minor polypeptides were detected by sodium dodecyl sulfate-polyacrylamide gel electrophoresis of membrane protein on 5 to 30 and 6 to 16% gradient gels.

  15. Chemistry of Secondary Polyphenols Produced during Processing of Tea and Selected Foods

    PubMed Central

    Tanaka, Takashi; Matsuo, Yosuke; Kouno, Isao

    2010-01-01

    This review will discuss recent progress in the chemistry of secondary polyphenols produced during food processing. The production mechanism of the secondary polyphenols in black tea, whisky, cinnamon, and persimmon fruits will be introduced. In the process of black tea production, tea leaf catechins are enzymatically oxidized to yield a complex mixture of oxidation products, including theaflavins and thearubigins. Despite the importance of the beverage, most of the chemical constituents have not yet been confirmed due to the complexity of the mixture. However, the reaction mechanisms at the initial stages of catechin oxidation are explained by simple quinone–phenol coupling reactions. In vitro model experiments indicated the presence of interesting regio- and stereoselective reactions. Recent results on the reaction mechanisms will be introduced. During the aging of whisky in oak wood barrels, ellagitannins originating from oak wood are oxidized and react with ethanol to give characteristic secondary ellagitannins. The major part of the cinnamon procyanidins is polymerized by copolymerization with cinnamaldehyde. In addition, anthocyanidin structural units are generated in the polymer molecules by oxidation which accounts for the reddish coloration of the cinnamon extract. This reaction is related to the insolubilization of proanthocyanidins in persimmon fruits by condensation with acetaldehyde. In addition to oxidation, the reaction of polyphenols with aldehydes may be important in food processing. PMID:20161999

  16. Analysis of TTG1 function in Arabis alpina

    PubMed Central

    2014-01-01

    Background In Arabidopsis thaliana (A. thaliana) the WD40 protein TRANSPARENT TESTA GLABRA1 (TTG1) controls five traits relevant for the adaptation of plants to environmental changes including the production of proanthocyanidin, anthocyanidin, seed coat mucilage, trichomes and root hairs. The analysis of different Brassicaceae species suggests that the function of TTG1 is conserved within the family. Results In this work, we studied the function of TTG1 in Arabis alpina (A. alpina). A comparison of wild type and two Aattg1 alleles revealed that AaTTG1 is involved in the regulation of all five traits. A detailed analysis of the five traits showed striking phenotypic differences between A. alpina and A. thaliana such that trichome formation occurs also at later stages of leaf development and that root hairs form at non-root hair positions. Conclusions The evolutionary conservation of the regulation of the five traits by TTG1 on the one hand and the striking phenotypic differences make A. alpina a very interesting genetic model system to study the evolution of TTG1-dependent gene regulatory networks at a functional level. PMID:24406039

  17. Differential expression of flavonoid biosynthesis genes and accumulation of phenolic compounds in common buckwheat (Fagopyrum esculentum).

    PubMed

    Li, Xiaohua; Park, Nam Il; Xu, Hui; Woo, Sun-Hee; Park, Cheol Ho; Park, Sang Un

    2010-12-01

    Common buckwheat (Fagopyrum esculentum) is a short-season grain crop that is a source of rutin and other phenolic compounds. In this study, we isolated the cDNAs of 11 F. esculentum enzymes in the flavonoid biosynthesis pathway, namely, phenylalanine ammonia lyase (PAL), cinnamate 4-hydroxylase (C4H), 4-coumarate:CoA ligase (4CL) 1 and 2, chalcone synthase (CHS), chalcone isomerase (CHI), flavone 3-hydroxylase (F3H), flavonoid 3'-hydroxylase (F3'H), flavonol synthase (FLS) 1 and 2, and anthocyanidin synthase (ANS). Quantitative real-time polymerase chain reaction analysis showed that these genes were most highly expressed in the stems and roots. However, high performance liquid chromatography analysis indicated that their flavonoid products, such as rutin and catechin, accumulated in the flowers and leaves. These results suggested that flavonoids may be transported within F. esculentum. In addition, light and dark growth conditions affected the expression levels of the biosynthesis genes and accumulation of phenolic compounds in F. esculentum sprouts.

  18. Effect of Methyl, Hydroxyl, and Chloro Substituents in Position 3 of 3',4',7-Trihydroxyflavylium: Stability, Kinetics, and Thermodynamics.

    PubMed

    Alejo-Armijo, Alfonso; Salido, Sofía; Altarejos, Joaquín; Parola, A Jorge; Gago, Sandra; Basílio, Nuno; Cabrita, Luis; Pina, Fernando

    2016-08-22

    The effect of methyl, hydroxyl, and chloride substituents in position 3 of the 3',4',7-trihydroxyflavylium core structure was studied. The stability, relative energy of each of chemical species (thermodynamics), and their rates of interconversion (kinetics) are very dependent on these substituents. By comparing the mole fraction distribution at equilibrium of the three multistate systems with the parent 3',4',7-trihydroxyflavylium, introduction of a methyl substituent in position 3 increases the mole fraction of hemiketal at the expense of the trans-chalcone and increases the hydration rate very significantly; a hydroxyl substituent in position 3 gives rise to a degradation process, as observed in anthocyanidins. In the case of 3-chloro-3',4',7-trihydroxyflavylium, a dramatic increase of the flavylium cation acidity was observed and a photochromic system can be operated upon irradiation of the respective trans-chalcone in 1 m HCl. According to the photochromic response of 3,3',4',7-tetrahydroxyflavylium and 3',4',7-trihydroxyflavylium, some requirements for a good photochromic performance are discussed.

  19. Metabolite profiling reveals temperature effects on the VOCs and flavonoids of different plant populations.

    PubMed

    Goh, H-H; Khairudin, K; Sukiran, N A; Normah, M N; Baharum, S N

    2016-01-01

    Temperature is one of the key factors in limiting the distribution of plants and controlling major metabolic processes. A series of simulated reciprocal transplant experiments were performed to investigate the effect of temperature on plant chemical composition. Polygonum minus of different lowland and highland origin were grown under a controlled environment with different temperature regimes to study the effects on secondary metabolites. We applied gas chromatography-mass spectrometry and liquid chromatography time-of-flight mass spectrometry to identify the chemical compounds. A total of 37 volatile organic compounds and 85 flavonoids were detected, with the largest response observed in the compositional changes of aldehydes and terpenes in highland plants under higher temperature treatment. Significantly less anthocyanidin compounds and larger amounts of flavonols were detected under higher temperature treatment. We also studied natural variation in the different plant populations growing under the same environment and identified compounds unique to each population through metabolite fingerprinting. This study shows that the origin of different plant populations influences the effects of temperature on chemical composition. PMID:26417881

  20. Flavonoid Intake in European Adults (18 to 64 Years)

    PubMed Central

    Vogiatzoglou, Anna; Mulligan, Angela A.; Lentjes, Marleen A. H.; Luben, Robert N.; Spencer, Jeremy P. E.; Schroeter, Hagen; Khaw, Kay-Tee; Kuhnle, Gunter G. C.

    2015-01-01

    Background Flavonoids are a group of phenolic secondary plant metabolites that are ubiquitous in plant-based diets. Data from anthropological, observational and intervention studies have shown that many flavonoids are bioactive. For this reason, there is an increasing interest in investigating the potential health effects of these compounds. The translation of these findings into the context of the health of the general public requires detailed information on habitual dietary intake. However, only limited data are currently available for European populations. Objective The objective of this study is to determine the habitual intake and main sources of anthocyanidins, flavanols, flavanones, flavones, flavonols, proanthocyanidins, theaflavins and thearubigins in the European Union. Design We use food consumption data from the European Food Safety Authority (EFSA) and the FLAVIOLA Food Composition Database to estimate intake of flavonoids. Results Mean (±SEM) intake of total flavonoids in Europe was 428±49 mg/d, of which 136±14 mg/d were monomeric compounds. Gallated flavan-3-ols (53±12 mg/d) were the main contributor. The lowest flavonoid intake was observed in Mediterranean countries (monomeric compounds: 95±11 mg/d). The distribution of intake was skewed in many countries, especially in Germany (monomeric flavonoids; mean intake: 181 mg/d; median intake: 3 mg/d). Conclusions The habitual intake of flavonoids in Europe is below the amounts found to have a significant health effect. PMID:26010916

  1. Characterization and estimation of proanthocyanidins and other phenolics in coffee pulp (Coffea arabica) by thiolysis-high-performance liquid chromatography.

    PubMed

    Ramirez-Coronel, Maria Ascencion; Marnet, Nathalie; Kolli, V S Kumar; Roussos, Sevastianos; Guyot, Sylvain; Augur, Christopher

    2004-03-10

    Fresh and 3-day-old coffee pulp of the Arabica variety were analyzed for polyphenol composition followed by characterization by two different methods. The first method consisted in subjecting coffee pulp powder to direct thiolysis. For the second method, coffee pulp was subjected to successive solvent extractions, followed by thiolysis. Quantification of phenolic compounds was then achieved by high-performance liquid chromatography (HPLC) analysis of thiolysis products. Four major classes of polyphenols were identified: flavan-3-ols (monomers and procyanidins), hydroxycinnamic acids, flavonols, and anthocyanidins. Differences in concentration of procyanidins were observed between fresh and 3-day-old coffee pulp. Constitutive units were mainly epicatechin, representing more than 90% of the proanthocyanidin units, with average degrees of polymerization in the range of 3.8-9.1. Monomer to hexamer units of flavan-3-ols from fresh coffee pulp were separated by normal-phase HPLC. Molecular size of oligomeric proanthocyanidins was obtained by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). Results obtained confirm the presence of oligomers of the flavan-3-ol (-)-epicatechin.

  2. Caffeoylquinic acid-rich purple sweet potato extract, with or without anthocyanin, imparts neuroprotection and contributes to the improvement of spatial learning and memory of SAMP8 mouse.

    PubMed

    Sasaki, Kazunori; Han, Junkyu; Shimozono, Hidetoshi; Villareal, Myra O; Isoda, Hiroko

    2013-05-29

    The effects of caffeoylquinic acid (CQA)-rich purple sweet potato (PSP) extract, with (PSPEa) or without (PSPEb) anthocyanin, on the improvement of spatial learning and memory of senescence-accelerated prone mouse strain (SAMP) 8 was determined. SAMP8 was treated with 20 mg/kg/day of PSPEa or PSPEb for 30 days. The effect on spatial learning and memory and the molecular mechanism of this effect were determined in vivo (SAMP8) and in vitro (SH-SY5Y cells). PSPEa or PSPEb reduced the escape latency time of SAMP8 by 17.0 ± 8.0 and 14.2 ± 5.8 s (P < 0.01), respectively. PSPEa administration induced an overexpression of antioxidant-, energy metabolism-, and neuronal plasticity-related proteins in the brain of SAMP8. Additionally, PSPEa and PSPEb increased the cell viability by 141.6 and 133% as compared to Aβ1-42-treated cells. These findings suggest that PSP rich in CQA derivatives with or without anthocyanidine had a neuroprotective effect on mouse brain and can improve the spatial learning and memory of SAMP8.

  3. Methyl jasmonate affects phenolic metabolism and gene expression in blueberry (Vaccinium corymbosum).

    PubMed

    Cocetta, Giacomo; Rossoni, Mara; Gardana, Claudio; Mignani, Ilaria; Ferrante, Antonio; Spinardi, Anna

    2015-02-01

    Blueberry (Vaccinium corymbosum) is a fruit very much appreciated by consumers for its antioxidant potential and health-promoting traits. Its beneficial potential properties are mainly due to a high content of anthocyanins and their amount can change after elicitation with methyl jasmonate. The aim of this work is to evaluate the changes in expression of several genes, accumulation of phenolic compounds and alterations in antioxidant potential in two different blueberry cultivars ('Duke' and 'Blueray') in response to methyl jasmonate (0.1 mM). Results showed that 9 h after treatment, the expression of phenylalanine ammonium lyase, chalcone synthase and anthocyanidin synthase genes was stimulated more in the 'Blueray' variety. Among the phenols measured an increase was recorded also for epicatechin and anthocyanin concentrations. 'Duke' is a richer sourche of anthocyanins compared to 'Blueray', treatment with methyl jasmonate promoted in 'Blueray' an increase in pigments as well as in the antioxidant potential, especially in fully ripe berries, but treated 'Duke' berries had greater levels, which were not induced by methyl jasmonate treatment. In conclusion, methyl jasmonate was, in some cases, an effective elicitor of phenolic metabolism and gene expression in blueberry, though with different intensity between cultivars.

  4. Intermolecular binding of blueberry pectin-rich fractions and anthocyanin.

    PubMed

    Lin, Z; Fischer, J; Wicker, L

    2016-03-01

    Pectin was extracted from blueberry powder into three fractions of water soluble (WSF), chelator soluble (CSF) and sodium carbonate soluble (NSF). The fractions were incubated with cyanidin-3-glucoside (C3G), a mixture of five anthocyanidins (cyanidin, pelargonidin, malvidin, petunidin and delphinidin) or blueberry juice at pH 2.0-4.5. Free anthocyanins and bound anthocyanin-pectin mixtures were separated by ultrafiltration. WSF bound the least amount of anthocyanin at all pH values. CSF had stronger anthocyanin binding ability at pH 2.0-3.6, while NSF had stronger anthocyanin binding ability at pH 3.6-4.5. The pectin and anthocyanin binding was lowest at pH 4.5 and higher at pH 2.0-3.6. Nearly doubling C3G pigment content increased bound anthocyanin percentage by 16-23% at pH 3.6, which favored anthocyanin aromatic stacking, compared to 3-9% increase at pH 2.0. Ionic interaction between anthocyanin flavylium cations and free pectic carboxyl groups, and anthocyanin stacking may be two major mechanisms for pectin and anthocyanin binding.

  5. Ferric ions involved in the flower color development of the Himalayan blue poppy, Meconopsis grandis.

    PubMed

    Yoshida, Kumi; Kitahara, Sayoko; Ito, Daisuke; Kondo, Tadao

    2006-05-01

    The Himalayan blue poppy, Meconopsis grandis, has sky blue-colored petals, although the anthocyanidin nucleus of the petal pigment is cyanidin. The blue color development in this blue poppy involving ferric ions was therefore studied. We analyzed the vacuolar pH, and the organic and inorganic components of the colored cells. A direct measurement by a proton-selective microelectrode revealed that the vacuolar pH value was 4.8. The concentrations of the total anthocyanins in the colored cells were around 5mM, and ca. three times more concentrated flavonols were detected. Fe was detected by atomic analysis of the colored cells, and the ratio of Fe to anthocyanins was ca. 0.8 eq. By mixing the anthocyanin, flavonol and metal ion components in a buffered aq. solution at pH 5.0, we were able to reproduce the same blue color; the visible absorption spectrum and CD were identical to those in the petals, with Fe(3+), Mg(2+) and flavonol being essential for the blue color. The blue pigment in Meconopsis should be a new type of metal complex pigment that is different from a stoichiometric supramolecular pigment such as commelinin or protocyanin.

  6. Evolutionary correlations in flavonoid production across flowers and leaves in the Iochrominae (Solanaceae).

    PubMed

    Berardi, Andrea E; Hildreth, Sherry B; Helm, Richard F; Winkel, Brenda S J; Smith, Stacey D

    2016-10-01

    Plant reproductive and vegetative tissues often use the same biochemical pathways to produce specialized metabolites. In such cases, selection acting on the synthesis of specific products in a particular tissue could result in correlated changes in other products of the pathway, both in the same tissue and in other tissues. This study examined how changes in floral anthocyanin pigmentation affect the production of other compounds of the flavonoid pathway in flowers and in leaves. Focusing on the Iochrominae, a clade of Solanaceae with a wide range of flower colors, liquid chromatography coupled with mass spectrometry and UV detection was used to profile and quantify the variation in two classes of flavonoids, anthocyanins and flavonols. Purple, red, orange and white-flowered Iochrominae produced all of the six common anthocyanidin types, as well as several classes of flavonols. Differences in anthocyanin and flavonol production were significantly correlated in flowers, particularly with respect to B ring hydroxylation pattern. However, these differences in floral flavonoids were not strongly related to differences in leaf chemistry. Specifically, most species made only flavonols (not anthocyanins) in leaves, and these comprised the two most common flavonols, quercetin and kaempferol, regardless of the color of the flower. These results suggest that shifts in flower color may occur without significant pleiotropic consequences for flavonoid production in vegetative tissues. Similar studies in other systems will be important for testing the generality of this pattern in other groups of flowering plants.

  7. Cyanidin suppresses ultraviolet B-induced COX-2 expression in epidermal cells by targeting MKK4, MEK1, and Raf-1.

    PubMed

    Kim, Jong-Eun; Kwon, Jung Yeon; Seo, Sang Kwon; Son, Joe Eun; Jung, Sung Keun; Min, So Yun; Hwang, Mun Kyung; Heo, Yong-Seok; Lee, Ki Won; Lee, Hyong Joo

    2010-05-15

    Skin cancer is the most frequently diagnosed cancer in the United States. Ultraviolet B (UVB) rays (wavelength: 280-320nm) play a pivotal role in the development of skin cancer by inducing the expression of inflammatory proteins such as cyclooxygenase-2 (COX-2). Cyanidin, the most plentiful of the plant pigments known as anthocyanidins, is a potent chemopreventive agent. In the present study, we examined the molecular mechanisms underlying the chemopreventive activity of cyanidin and identified its molecular targets. Cyanidin inhibited UVB-induced COX-2 expression and prostaglandin E(2) secretion in the epidermal skin cell line JB6 P+ by suppressing the transactivation of nuclear factor-kappaB and activator protein-1 which are well-known transcription factors regulated by mitogen-activated protein kinase. Cyanidin markedly inhibited the phosphorylation of JNK1/2, ERK1/2, and MEK1/2 than the of MKK4 and Raf-1, two upstream kinases of JNK1/2, ERK1/2, and MEK1/2. Cyanidin significantly suppressed the activities of MKK4, MEK1, and Raf-1 through direct binding. Transient transfection of a small interfering RNA specific for MKK4 inhibited the UVB-induced expression of COX-2 in JB6 P+ cells, as did the expression of a dominant-negative ERK2 mutant. We conclude that MKK4, MEK1, and Raf-1 are targets of cyanidin for the suppression of UVB-induced COX-2 expression.

  8. Herbicide-induced anthocyanin accumulation in transgenic rice by expression of rice OSB2 under the control of rice CYP72A21 promoter.

    PubMed

    Hirose, Sakiko; Kawahigashi, Hiroyuki; Tagiri, Akemi; Ohkawa, Yasunobu

    2008-02-27

    CYP72A21, a rice cytochrome P450 gene, is induced by chloroacetamide herbicides. OSB2, a rice myc-type transcription factor, induces anthocyanin accumulation in rice leaves. To produce plants for biomonitoring by color change, we combined the CYP72A21 promoter and the OSB2 gene and introduced them into the rice isogenic line Taichung-65 CB A (T65), which contains loci CB and A from the rice cultivar Murasakiine. Leaves of the transgenic plants turned red upon treatment with the chloroacetamide herbicides acetochlor, alachlor, and metolachlor. Seedling shoots reddened upon treatment with alachlor or metolachlor at 10 microM, a concentration slightly higher than that used in the field. Anthocyanin content was increased approximately 200% by the treatment. The color changes were consistent with increased shoot expression of OSB2 and the anthocyanidin synthase gene (ANS). This system promises easy detection of rice plant gene expression. Transgenic plants could be used in the future to biomonitor accumulated herbicides.

  9. Recent advances in understanding the anti-diabetic actions of dietary flavonoids.

    PubMed

    Babu, Pon Velayutham Anandh; Liu, Dongmin; Gilbert, Elizabeth R

    2013-11-01

    Flavonoids are polyphenolic compounds that are abundant in fruits and vegetables, and increasing evidence demonstrates a positive relationship between consumption of flavonoid-rich foods and disease prevention. Epidemiological, in vitro and animal studies support the beneficial effects of dietary flavonoids on glucose and lipid homeostasis. It is encouraging that the beneficial effects of some flavonoids are at physiological concentrations and comparable to clinically-used anti-diabetic drugs; however, clinical research in this field and studies on the anti-diabetic effects of flavonoid metabolites are limited. Flavonoids act on various molecular targets and regulate different signaling pathways in pancreatic β-cells, hepatocytes, adipocytes and skeletal myofibers. Flavonoids may exert beneficial effects in diabetes by (i) enhancing insulin secretion and reducing apoptosis and promoting proliferation of pancreatic β-cells; (ii) improving hyperglycemia through regulation of glucose metabolism in hepatocytes; (iii) reducing insulin resistance, inflammation and oxidative stress in muscle and fat and (iv) increasing glucose uptake in skeletal muscle and white adipose tissue. This review highlights recent findings on the anti-diabetic effects of dietary flavonoids, including flavan-3-ols, flavanones, flavonols, anthocyanidins, flavones and isoflavones, with particular emphasis on the studies that investigated the cellular and molecular mechanisms involved in the beneficial effects of the compounds.

  10. Persimmon (Diospyros kaki) fruit: hidden phytochemicals and health claims.

    PubMed

    Butt, Masood Sadiq; Sultan, M Tauseef; Aziz, Mahwish; Naz, Ambreen; Ahmed, Waqas; Kumar, Naresh; Imran, Muhammad

    2015-01-01

    Currently, nutrition and health linkages focused on emerging strategy of diet based regimen to combat various physiological threats including cardiovascular disorders, oxidative stress, diabetes mellitus, etc. In this context, consumption of fruits and vegetables is gaining considerable importance as safeguard to maintain human health. Likewise, their phytochemicals and bioactive molecules are also becoming popular as promising demulcent against various ailments. The current review is an effort to sum up information regarding persimmon fruit with special reference to its phytochemistry and associated health claims. Accordingly, the role of its certain bioactive molecules like proanthocyanidin, carotenoids, tannins, flavonoids, anthocyanidin, catechin, etc. is highlighted. Owing to rich phytochemistry, persimmon and its products are considered effective in mitigating oxidative damage induced by reactive oxygen species (ROS). The antioxidant potential is too responsible for anti-malignant and anti-melanogenic perspectives of persimmon functional ingredients. Additionally, they are effectual in soothing lifestyle related disparities e.g. cardiovascular disorders and diabetes mellitus. There are proven facts that pharmacological application of persimmon or its functional ingredients like proanthocyanidin may helps against hyperlipidemia and hyperglycemia. Nevertheless, astringent taste and diospyrobezoars formation are creating lacuna to prop up its vitality. In toto, persimmon and its components hold potential as one of effective modules in diet based therapy; however, integrated research and meta-analysis are still required to enhance meticulousness.

  11. Caffeoylquinic acid-rich purple sweet potato extract, with or without anthocyanin, imparts neuroprotection and contributes to the improvement of spatial learning and memory of SAMP8 mouse.

    PubMed

    Sasaki, Kazunori; Han, Junkyu; Shimozono, Hidetoshi; Villareal, Myra O; Isoda, Hiroko

    2013-05-29

    The effects of caffeoylquinic acid (CQA)-rich purple sweet potato (PSP) extract, with (PSPEa) or without (PSPEb) anthocyanin, on the improvement of spatial learning and memory of senescence-accelerated prone mouse strain (SAMP) 8 was determined. SAMP8 was treated with 20 mg/kg/day of PSPEa or PSPEb for 30 days. The effect on spatial learning and memory and the molecular mechanism of this effect were determined in vivo (SAMP8) and in vitro (SH-SY5Y cells). PSPEa or PSPEb reduced the escape latency time of SAMP8 by 17.0 ± 8.0 and 14.2 ± 5.8 s (P < 0.01), respectively. PSPEa administration induced an overexpression of antioxidant-, energy metabolism-, and neuronal plasticity-related proteins in the brain of SAMP8. Additionally, PSPEa and PSPEb increased the cell viability by 141.6 and 133% as compared to Aβ1-42-treated cells. These findings suggest that PSP rich in CQA derivatives with or without anthocyanidine had a neuroprotective effect on mouse brain and can improve the spatial learning and memory of SAMP8. PMID:23647122

  12. Oxygen Consumption by Red Wines. Part II: Differential Effects on Color and Chemical Composition Caused by Oxygen Taken in Different Sulfur Dioxide-Related Oxidation Contexts.

    PubMed

    Carrascon, Vanesa; Fernandez-Zurbano, Purificación; Bueno, Mónica; Ferreira, Vicente

    2015-12-30

    Chemical changes caused by oxidation of red wines during 5 consecutive air-saturation cycles have been assessed. In order to investigate the existing relationship between the effects caused by O2 and the levels and consumption rates of wine SO2, the total oxygen consumed by the wines (16-25 mg/L) was subdivided into different nonmutually exclusive categories. The ones found most influential on chemical changes were the O2 consumed in the first saturation without equivalent SO2 consumption (O2preSO2) and the O2 consumed when levels of free SO2 were below 5 mg/L (radical forming O2). Chromatic changes were strongly related to both O2 categories, even though anthocyanidin degradation was not related to any O2 category. Radical forming O2 prevented both formation of red pigments and reduction of epigallocatechin and other proanthocyanidins, induced accumulation of phenolic acids, and caused losses of β-damascenone and whiskylactone without evidence of acetaldehyde formation. O2preSO2 seemed to play a key role in the formation of blue pigments and in the decrease of Folin index and of many important aroma compounds. PMID:26646423

  13. Recent advances in understanding the anti-diabetic actions of dietary flavonoids

    PubMed Central

    Babu, Pon Velayutham Anandh; Liu, Dongmin; Gilbert, Elizabeth R.

    2013-01-01

    Flavonoids are polyphenolic compounds that are abundant in fruits and vegetables and increasing evidence demonstrates a positive relationship between consumption of flavonoid-rich foods and disease prevention. Epidemiological, in vitro and animal studies support the beneficial effects of dietary flavonoids on glucose and lipid homeostasis. It is encouraging that the beneficial effects of some flavonoids are at physiological concentrations and comparable to clinically-used anti-diabetic drugs; however, clinical research in this field and studies on the anti-diabetic effects of flavonoid metabolites are limited. Flavonoids act on various molecular targets and regulate different signaling pathways in pancreatic β-cells, hepatocytes, adipocytes, and skeletal myofibers. Flavonoids may exert beneficial effects in diabetes by (i) enhancing insulin secretion and reducing apoptosis and promoting proliferation of pancreatic β-cells, (ii) improving hyperglycemia through regulation of glucose metabolism in hepatocytes, (iii) reducing insulin resistance, inflammation and oxidative stress in muscle and fat, and (iv) increasing glucose uptake in skeletal muscle and white adipose tissue. This review highlights recent findings on the anti-diabetic effects of dietary flavonoids, including flavan-3-ols, flavanones, flavonols, anthocyanidins, flavones, and isoflavones, with particular emphasis on the studies that investigated the cellular and molecular mechanisms involved in the beneficial effects of the compounds. PMID:24029069

  14. Genotoxicity of phytoestrogens.

    PubMed

    Stopper, H; Schmitt, E; Kobras, K

    2005-07-01

    Plant extracts containing phytohormones are very popular as 'alternative' medicine for many kinds of diseases. They are especially favored by women who enter menopause and are concerned about the side effects of hormone replacement therapy. However, adverse health effects of phytoestrogens have often been ignored. This review examines the literature on genotoxicity and apoptotic effects of phytohormones. Genistein, coumestrol, quercetin, zearalenone, and resveratrol exerted genotoxic effects in in vitro test systems. Other phytoestrogens such as lignans, the isoflavones daidzein and glycetein, anthocyanidins, and the flavonol fisetin exhibited only weak or no effects in vitro. However, some metabolites of daidzein showed a genotoxic activity in vitro. Practically all of the phytoestrogens exhibit pro-apoptotic effects in some cell systems. Further investigations regarding dose-response-relationships and other aspects relevant for extrapolation to human exposure seem necessary. Until then, care may be advised in taking concentrated phytohormones. Nevertheless, the intake of substantial amounts of plant-food in a normal diet constitutes an important, individual contribution to cancer prevention.

  15. Evolutionary correlations in flavonoid production across flowers and leaves in the Iochrominae (Solanaceae).

    PubMed

    Berardi, Andrea E; Hildreth, Sherry B; Helm, Richard F; Winkel, Brenda S J; Smith, Stacey D

    2016-10-01

    Plant reproductive and vegetative tissues often use the same biochemical pathways to produce specialized metabolites. In such cases, selection acting on the synthesis of specific products in a particular tissue could result in correlated changes in other products of the pathway, both in the same tissue and in other tissues. This study examined how changes in floral anthocyanin pigmentation affect the production of other compounds of the flavonoid pathway in flowers and in leaves. Focusing on the Iochrominae, a clade of Solanaceae with a wide range of flower colors, liquid chromatography coupled with mass spectrometry and UV detection was used to profile and quantify the variation in two classes of flavonoids, anthocyanins and flavonols. Purple, red, orange and white-flowered Iochrominae produced all of the six common anthocyanidin types, as well as several classes of flavonols. Differences in anthocyanin and flavonol production were significantly correlated in flowers, particularly with respect to B ring hydroxylation pattern. However, these differences in floral flavonoids were not strongly related to differences in leaf chemistry. Specifically, most species made only flavonols (not anthocyanins) in leaves, and these comprised the two most common flavonols, quercetin and kaempferol, regardless of the color of the flower. These results suggest that shifts in flower color may occur without significant pleiotropic consequences for flavonoid production in vegetative tissues. Similar studies in other systems will be important for testing the generality of this pattern in other groups of flowering plants. PMID:27291343

  16. Inhibitive Effects of Mulberry Leaf-Related Extracts on Cell Adhesion and Inflammatory Response in Human Aortic Endothelial Cells

    PubMed Central

    Chao, P.-Y.; Lin, K.-H.; Chiu, C.-C.; Yang, Y.-Y.; Huang, M.-Y.; Yang, C.-M.

    2013-01-01

    Effects of mulberry leaf-related extracts (MLREs) on hydrogen peroxide-induced DNA damage in human lymphocytes and on inflammatory signaling pathways in human aortic endothelial cells (HAECs) were studied. The tested MLREs were rich in flavonols, especially bombyx faces tea (BT) in quercetin and kaempferol. Polyphenols, flavonoids, and anthocyanidin also abounded in BT. The best trolox equivalent antioxidant capacity (TEAC) was generated from the acidic methanolic extracts of BT. Acidic methanolic and water extracts of mulberry leaf tea (MT), mulberry leaf (M), and BT significantly inhibited DNA oxidative damage to lymphocytes based on the comet assay as compared to the H2O2-treated group. TNF-α-induced monocyte-endothelial cell adhesion was significantly suppressed by MLREs. Additionally, nuclear factor kappa B (NF-κB) expression was significantly reduced by BT and MT. Significant reductions were also observed in both NF-κB and activator protein (AP)-1 DNA binding by MLREs. Significant increases in peroxisome proliferator-activated receptor (PPAR) α and γ DNA binding by MLREs were also detected in M and MT extracts, but no evidence for PPAR α DNA binding in 50 μg/mL MT extract was found. Apparently, MLREs can provide distinct cytoprotective mechanisms that may contribute to its putative beneficial effects on suppressing endothelial responses to cytokines during inflammation. PMID:24371453

  17. Effects of dietary supplementation with a combination of fish oil, bilberry extract, and lutein on subjective symptoms of asthenopia in humans.

    PubMed

    Kawabata, Fuminori; Tsuji, Tomoko

    2011-12-01

    The aim of this study was to determine the effects of dietary supplementation with a combination of fish oil, bilberry extract, and lutein on subjective symptoms of asthenopia in humans by a double- blind, randomized, parallel-group, and placebo-controlled trial. In the Active group, eleven subjects ingested a supplement containing omega-3 fatty acid-rich fish oil (docosahexaenoic acid 783 mg/day, eicosapentaenoic acid 162 mg/day), bilberry extract (anthocyanidin 59 mg/day), and lutein (17.5 mg/day) in soft gel capsule form, every day for 4 weeks. In the Placebo group, nine subjects ingested placebo capsules. Before and after supplementation, subjects completed a questionnaire to determine their asthenopia symptoms and were also assessed for mental fatigue symptom by the visual analog scale (VAS) test. Asthenopia symptoms such as "stiff shoulder, low back pain", "frustration", "dry-eye", and "stuffy head" were improved in the Active group. Furthermore, a score of mental fatigue was improved after 4 weeks of supplementation, and no side effects were observed after the 4-week supplementation and a 2-week washout period in the Active group. These results suggest that dietary supplementation with the combination of omega-3 fatty acid-rich fish oil, bilberry extract, and lutein may safely improve subjective symptoms of asthenopia and mental fatigue in humans.

  18. Development of a Recombinant Escherichia coli Strain for Overproduction of the Plant Pigment Anthocyanin

    PubMed Central

    Lim, Chin Giaw; Wong, Lynn; Bhan, Namita; Dvora, Hila; Xu, Peng; Venkiteswaran, Sankaranarayanan

    2015-01-01

    Anthocyanins are water-soluble colored pigments found in terrestrial plants and are responsible for the red, blue, and purple coloration of many flowers and fruits. In addition to the plethora of health benefits associated with anthocyanins (cardioprotective, anti-inflammatory, antioxidant, and antiaging properties), these compounds have attracted widespread attention due to their promising potential as natural food colorants. Previously, we reported the biotransformation of anthocyanin, specifically cyanidin 3-O-glucoside (C3G), from the substrate (+)-catechin in Escherichia coli. In the present work, we set out to systematically improve C3G titers by enhancing substrate and precursor availability, balancing gene expression level, and optimizing cultivation and induction parameters. We first identified E. coli transporter proteins that are responsible for the uptake of catechin and secretion of C3G. We then improved the expression of the heterologous pathway enzymes anthocyanidin synthase (ANS) and 3-O-glycosyltransferase (3GT) using a bicistronic expression cassette. Next, we augmented the intracellular availability of the critical precursor UDP-glucose, which has been known as the rate-limiting precursor to produce glucoside compounds. Further optimization of culture and induction conditions led to a final titer of 350 mg/liter of C3G. We also developed a convenient colorimetric assay for easy screening of C3G overproducers. The work reported here constitutes a promising foundation to develop a cost-effective process for large-scale production of plant-derived anthocyanin from recombinant microorganisms. PMID:26150456

  19. Flavonoids and risk of squamous cell esophageal cancer.

    PubMed

    Rossi, Marta; Garavello, Werner; Talamini, Renato; La Vecchia, Carlo; Franceschi, Silvia; Lagiou, Pagona; Zambon, Paola; Dal Maso, Luigino; Bosetti, Cristina; Negri, Eva

    2007-04-01

    The relation between 5 classes of flavonoids (flavanones, flavan-3-ols, flavonols, flavones and anthocyanidines) and esophageal cancer was investigated using data from a case-control study conducted between 1992 and 1997 in 3 areas of northern Italy. The study included 304 cases (275 men, 29 women) with a first diagnosis of squamous-cell carcinoma of the esophagus and 743 controls (593 men, 150 women) with no history of cancer, admitted for acute illnesses, unrelated to tobacco and alcohol consumption, to major hospitals of the areas under surveillance. Dietary habits were investigated using a validated food frequency questionnaire. Odds ratios (ORs) and 95% confidence intervals (CI) were computed after allowance for age, sex, study centre, years of education, alcohol drinking, tobacco smoking, body mass index and energy intake. An inverse association emerged between flavanone intake and esophageal cancer risk (OR=0.38 for the highest vs. the lowest quintile, 95% CI=0.23-0.66). The inverse relation between flavanones and esophageal cancer tended to be stronger in those who drank >or=6 drinks/day. In conclusion, this study suggests that flavanone intake is inversely associated with esophageal cancer risk and may account, with vitamin C, for the protective effect of fruit, especially citrus fruit, on esophageal cancer. PMID:17192901

  20. Role of intestinal microbiota in the generation of polyphenol derived phenolic acid mediated attenuation of Alzheimer’s disease β-amyloid oligomerization

    PubMed Central

    Wang, Dongjie; Ho, Lap; Faith, Jeremiah; Ono, Kenjiro; Janle, Elsa M.; Lachcik, Pamela J.; Cooper, Bruce R.; Jannasch, Amber H.; D’Arcy, Bruce R.; Williams, Barbara A.; Ferruzzi, Mario G.; Levine, Samara; Zhao, Wei; Dubner, Lauren; Pasinetti, Giulio M.

    2015-01-01

    Scope Grape seed polyphenol extract (GSPE) is receiving increasing attention for its potential preventative and therapeutic roles in Alzheimer’s disease (AD) and other age-related neurodegenerative disorders. The intestinal microbiota is known to actively convert many dietary polyphenols, including GSPE, to phenolic acids. There is limited information on the bioavailability and bioactivity of GSPE-derived phenolic acid in the brain. Methods and Results We orally administered GSPE to rats and investigated the bioavailability of 12 phenolic acids known to be generated by microbiota metabolism of anthocyanidins. GSPE treatment significantly increased the content of 2 of the phenolic acids in the brain: 3-hydroxybenzoic acid (3-HBA) and 3-(3′-hydroxyphenyl) propionic acid (3-HPP), resulting in the brain accumulations of the two phenolic acids at μM concentrations. We also provided evidence that 3-HBA and 3-HPP potently interfere with the assembly of β-amyloid (Aβ) peptides into neurotoxic Aβ aggregates that play key roles in AD pathogenesis. Conclusion Our observation suggests important contribution of the intestinal microbiota to the protective activities of GSPE (as well as other polyphenol preparations) in AD. Outcomes from our studies support future preclinical and clinical investigations exploring the potential contributions of the intestinal microbiota in protecting against the onset/progression of AD and other neurodegenerative conditions. PMID:25689033

  1. Cloning and characterization of a flavonol synthase gene from Scutellaria baicalensis.

    PubMed

    Kim, Yeon Bok; Kim, KwangSoo; Kim, Yeji; Tuan, Pham Anh; Kim, Haeng Hoon; Cho, Jin Woong; Park, Sang Un

    2014-01-01

    Flavonols are the most abundant of all the flavonoids and play pivotal roles in a variety of plants. We isolated a cDNA clone encoding flavonol synthase from Scutellaria baicalensis (SbFLS). The SbFLS cDNA is 1011 bp long, encodes 336 amino acid residues, and belongs to a family of 2-oxoglutarate-dependent dioxygenases. The overall structure of SbFLS is very similar to that of Arabidopsis thaliana anthocyanidin synthase (AtANS), with a β jelly-roll fold surrounded by tens of short and long α-helices. SbFLS was constitutively expressed in the roots, stems, leaves, and flowers, with particularly high expression in the roots and flowers. SbFLS transcript levels in the roots were 376-, 70-, and 2.5-fold higher than in the leaves, stems, and flowers. The myricetin content was significantly higher than that of kaempferol and quercetin. Therefore, we suggest that SbFLS mediates flavonol formation in the different organs of S. baicalensis. Our study may contribute to the knowledge of the role of FLS in S. baicalensis. PMID:24672406

  2. Identification of flavonoid 3'-hydroxylase in the yellow flower of Delphinium zalil.

    PubMed

    Miyahara, Taira; Hamada, Arisa; Okamoto, Mitsutoshi; Hirose, Yukio; Sakaguchi, Kimitoshi; Hatano, Shoji; Ozeki, Yoshihiro

    2016-09-01

    The flowers of delphinium cultivars owe their coloration to anthocyanins such as delphinidin or pelargonidin derivatives. To date, no delphinium cultivars have been found with red flowers due to the presence of cyanidin derivatives. This suggests that delphiniums do not have cyanidin biosynthesis ability because of the loss of function of flavonoid 3' hydroxylase (F3'H). Here, we show that the wild delphinium species Delphinium zalil (synonym semibarbatum) can accumulate quercetin 3-glucosides in its sepals, presumably through F3'H activity. We isolated F3'H cDNA from D. zalil (DzF3'H) and produced a recombinant enzyme from a yeast transformant. The recombinant DzF3'H protein could convert naringenin, apigenin, dihydrokaempferol and kaempferol to eriodictyol, luteolin, dihydroquercetin and quercetin, respectively. An expression analysis confirmed that blue flowered D. grandiflorum does not express F3'H, and also showed that flavonoid 3',5'-hydroxylase and anthocyanidin synthase do not function in D. zalil sepals. DzF3'H can act as a flavonoid hydroxylase to produce cyanidin accumulation. The introduction of the DzF3'H gene into other delphinium species by conventional breeding may enable development of cultivars with novel flower colors. PMID:27478933

  3. Development of a Recombinant Escherichia coli Strain for Overproduction of the Plant Pigment Anthocyanin.

    PubMed

    Lim, Chin Giaw; Wong, Lynn; Bhan, Namita; Dvora, Hila; Xu, Peng; Venkiteswaran, Sankaranarayanan; Koffas, Mattheos A G

    2015-09-01

    Anthocyanins are water-soluble colored pigments found in terrestrial plants and are responsible for the red, blue, and purple coloration of many flowers and fruits. In addition to the plethora of health benefits associated with anthocyanins (cardioprotective, anti-inflammatory, antioxidant, and antiaging properties), these compounds have attracted widespread attention due to their promising potential as natural food colorants. Previously, we reported the biotransformation of anthocyanin, specifically cyanidin 3-O-glucoside (C3G), from the substrate (+)-catechin in Escherichia coli. In the present work, we set out to systematically improve C3G titers by enhancing substrate and precursor availability, balancing gene expression level, and optimizing cultivation and induction parameters. We first identified E. coli transporter proteins that are responsible for the uptake of catechin and secretion of C3G. We then improved the expression of the heterologous pathway enzymes anthocyanidin synthase (ANS) and 3-O-glycosyltransferase (3GT) using a bicistronic expression cassette. Next, we augmented the intracellular availability of the critical precursor UDP-glucose, which has been known as the rate-limiting precursor to produce glucoside compounds. Further optimization of culture and induction conditions led to a final titer of 350 mg/liter of C3G. We also developed a convenient colorimetric assay for easy screening of C3G overproducers. The work reported here constitutes a promising foundation to develop a cost-effective process for large-scale production of plant-derived anthocyanin from recombinant microorganisms.

  4. Isolation and characterization of a cDNA clone of UDP-galactose: flavonoid 3-O-galactosyltransferase (UF3GaT) expressed in Vigna mungo seedlings.

    PubMed

    Mato, M; Ozeki, Y; Itoh, Y; Higeta, D; Yoshitama, K; Teramoto, S; Aida, R; Ishikura, N; Shibata, M

    1998-11-01

    Four cDNA clones were isolated from Vigna mungo seedlings by the screening with cDNA encoding UDP-glucose:flavonoid 3-O-glucosyltransferase (UF3GT) of Antirrhinum majus as a probe; the product of the gene corresponding to one cDNA was more highly expressed in the first simple leaves than in stems. Nucleotide sequence analysis revealed 1,691 bp (including 326 bp non-reading) containing an open reading frame of 455 amino acids. The deduced amino acid sequence showed 42% and 23% identity with those of A. majus UDP-glucose:flavonoid 3-O-glucosyltransferase (UF3GT) and Petunia hybrida UDP-rhamnose:anthocyanidin 3-O-glucoside rhamnosyltransferase (RT), respectively. One region of the cDNA (amino acids 325 to 387) showed similarity to ceramide UDP-galactosyltransferases of mice, rats and humans. A crude extract from Escherichia coli, in which the protein was expressed from the cDNA, showed high UF3GaT activity but low UF3GT activity, and was similar in K(m), optimal pH and substrate specificity to UF3GaT from V. mungo. We conclude that we have obtained UDP-galactose:flavonoid 3-O-galactosyltransferase (UF3GaT) cDNA from V. mungo.

  5. Anti-inflammatory properties of red ginger (Zingiber officinale var. Rubra) extract and suppression of nitric oxide production by its constituents.

    PubMed

    Shimoda, Hiroshi; Shan, Shao-Jie; Tanaka, Junji; Seki, Azusa; Seo, Joung-Wook; Kasajima, Naoki; Tamura, Satoru; Ke, Yan; Murakami, Nobutoshi

    2010-02-01

    Red ginger (Zingiber officinale var. Rubra) has been prescribed as an analgesic for arthritis pain in Indonesian traditional medicine. The surface color of the rhizome is purple because of the anthocyanidins in its peel. We prepared 40% ethanolic extract from dried red ginger (red ginger extract [RGE]) and evaluated its anti-inflammatory activity using acute and chronic inflammation models. In an acetic acid-induced mouse writhing model, RGE (10-100 mg/kg) suppressed both the frequency of writhing and the increase in permeability of abdominal capillaries. On the other hand, continuous treatment with RGE (10 mg/kg) significantly (P < .05) suppressed footpad edema in a rat adjuvant arthritis model. To clarify the anti-inflammatory mechanism of RGE, we examined the effect on prostaglandin (PG) and nitric oxide (NO) production from mouse leukemic monocytes (RAW264 cells) stimulated by lipopolysaccharide. RGE (3 and 10 microg/mL) significantly (P < .05) suppressed PGE(2) production, while it also suppressed NO production at 100 microg/mL. After bioassay-guided separation of RGE, we found that [6]-shogaol and gingerdiols suppressed NO production. Red dye fractions presumed to be proanthocyanidins also suppressed NO production at 100 microg/mL. Consequently, we found a potent suppressive effect of RGE on acute and chronic inflammation, and inhibition of macrophage activation seems to be involved in this anti-inflammatory effect. [6]-Shogaol, gingerdiols, and proanthocyanidins were identified as constituents that inhibited NO production.

  6. Molecular and biochemical characterization of the UDP-glucose: Anthocyanin 5-O-glucosyltransferase from Vitis amurensis.

    PubMed

    He, Fei; Chen, Wei-Kai; Yu, Ke-Ji; Ji, Xiang-Nan; Duan, Chang-Qing; Reeves, Malcolm J; Wang, Jun

    2015-09-01

    Generally, red Vitis vinifera grapes only contain monoglucosidic anthocyanins, whereas most non-vinifera red grapes of the Vitis genus have both monoglucosidic and bis-glucosidic anthocyanins, the latter of which are believed to be more hydrophilic and more stable. Although previous studies have established the biosynthetic mechanism for formation of monoglucosidic anthocyanins, less attention has been paid to that of bis-glucosidic anthocyanins. In the present research, the full-length cDNA of UDP-glucose: anthocyanin 5-O-glucosyltransferase from Vitis amurensis Rupr. cv. 'Zuoshanyi' grape (Va5GT) was cloned. After acquisition and purification of recombinant Va5GT, its enzymatic parameters were systematically analyzed in vitro. Recombinant Va5GT used malvidin-3-O-glucoside as its optimum glycosidic acceptor when UDP-glucose was used as the glycosidic donor. Va5GT-GFP was found to be located in the cytoplasm by analyzing its subcellular localization with a laser-scanning confocal fluorescence microscope, and this result was coincident with its metabolic function of modifying anthocyanins in grape cells. Furthermore, the relationship between the transcriptional expression of Va5GT and the accumulation of anthocyanidin bis-glucosides during berry development suggested that Va5GT is a key enzyme in the biosynthesis of bis-glucosidic anthocyanins in V. amurensis grape berries.

  7. Identification and characterisation of F3GT1 and F3GGT1, two glycosyltransferases responsible for anthocyanin biosynthesis in red-fleshed kiwifruit (Actinidia chinensis).

    PubMed

    Montefiori, Mirco; Espley, Richard V; Stevenson, David; Cooney, Janine; Datson, Paul M; Saiz, Anna; Atkinson, Ross G; Hellens, Roger P; Allan, Andrew C

    2011-01-01

    Much of the diversity of anthocyanins is due to the action of glycosyltransferases, which add sugar moieties to anthocyanidins. We identified two glycosyltransferases, F3GT1 and F3GGT1, from red-fleshed kiwifruit (Actinidia chinensis) that perform sequential glycosylation steps. Red-fleshed genotypes of kiwifruit accumulate anthocyanins mainly in the form of cyanidin 3-O-xylo-galactoside. Genes in the anthocyanin and flavonoid biosynthetic pathway were identified and shown to be expressed in fruit tissue. However, only the expression of the glycosyltransferase F3GT1 was correlated with anthocyanin accumulation in red tissues. Recombinant enzyme assays in vitro and in vivo RNA interference (RNAi) demonstrated the role of F3GT1 in the production of cyanidin 3-O-galactoside. F3GGT1 was shown to further glycosylate the sugar moiety of the anthocyanins. This second glycosylation can affect the solubility and stability of the pigments and modify their colour. We show that recombinant F3GGT1 can catalyse the addition of UDP-xylose to cyanidin 3-galactoside. While F3GGT1 is responsible for the end-product of the pathway, F3GT1 is likely to be the key enzyme regulating the accumulation of anthocyanin in red-fleshed kiwifruit varieties.

  8. Antioxidant activities and determination of phenolic compounds isolated from oriental plums (Soldam, Oishiwase and Formosa)

    PubMed Central

    Kim, Mee-Ree; Cho, Soo-Muk; Kim, So-Young; Kim, Jung-Bong; Cho, Young-Sook

    2012-01-01

    The purposes of this study were to determine phenolic compounds and to evaluate antioxidant activities of plums (Soldam, Oishiwase and Formosa). Soldam contains the highest amount of total phenolics among cultivars (Formosa: 4.0%, Oishiwase: 3.3%, Soldam: 6.4% for total phenolic) as well as the total flavonoids of which constituents were mainly myricetin and anthocyanidin. The antioxidant activities were measured by DPPH, ABTS radical scavenging, and SOD-like activities. The DPPH radical scavenging activity of Korean plum extracts (200 µg/mL) showed more than 43%, and the Soldam turned out to be the highest : ID50 value: 160-177 µg/mL for Formosa and Oishiwase; 58-64 µg/mL for Soldam. The ABTS radical scavenging activity of Korean plum extracts (200 µg/mL) was found to be more than 50%. The SOD-like activity of Korean plum extracts (200 µg/mL) showed more than 70%. Among three kinds of cultivars, Soldam had the highest antioxidant activity. The nitrite scavenging activity of Soldam was 61.5%, which is the highest, compared with that of the other cultivars, about 50%. From these results, Korean plums turned out to be phytochemical rich fruit as well as to show high antioxidant activities. PMID:22977680

  9. Relative velocity changes using ambient seismic noise at Okmok and Redoubt volcanoes, Alaska

    NASA Astrophysics Data System (ADS)

    Bennington, N. L.; Haney, M. M.; De Angelis, S.; Thurber, C. H.

    2013-12-01

    Okmok and Redoubt are two of the most active volcanoes in the Aleutian Arc. Leading up to its most recent eruption, Okmok, a shield volcano on Umnak Island, showed precursors to volcanic activity only five hours before it erupted explosively in July 2008. Redoubt, a stratovolcano located along the Cook Inlet, displayed several months of precursory activity leading up to its March 2009 eruption. Frequent activity at both volcanoes poses a major hazard due to heavy traffic along the North Pacific air routes. Additionally, Okmok is adjacent to several of the world's most productive fisheries and Redoubt is located only 110 miles SW of Anchorage, the major population center of Alaska. For these reasons, it is imperative that we improve our ability to detect early signs of unrest, which could potentially lead to eruptive activity at these volcanoes. We take advantage of continuous waveforms recorded on seismic networks at Redoubt and Okmok in an attempt to identify seismic precursors to the recent eruptions at both volcanoes. We perform seismic interferometry using ambient noise, following Brenguier et al. (2008), in order to probe the subsurface and determine temporal changes in relative seismic velocity from pre- through post-eruption, for the 2008 Okmok and 2009 Redoubt eruptions. In a preliminary investigation, we analyzed 6 months of noise cross-correlation functions averaged over 10-day intervals leading up to the 2009 eruption at Redoubt. During February 2009, station pairs RSO-DFR and RDN-RSO showed a decrease in seismic velocity of ~0.02%. By the beginning of March, the relative velocity changes returned to background levels. Stations RSO and RDN are located within the summit breach, and station DFR is to the north. Although these results are preliminary, it is interesting to note that the decrease in seismic velocity at both station pairs overlaps with the time period when Grapenthin et al. (2012) hypothesize magma in the mid-to-deep crustal reservoir was

  10. Factors relating to eating style, social desirability, body image and eating meals at home increase the precision of calibration equations correcting self-report measures of diet using recovery biomarkers: findings from the Women’s Health Initiative

    PubMed Central

    2013-01-01

    Background The extent to which psychosocial and diet behavior factors affect dietary self-report remains unclear. We examine the contribution of these factors to measurement error of self-report. Methods In 450 postmenopausal women in the Women’s Health Initiative Observational Study doubly labeled water and urinary nitrogen were used as biomarkers of objective measures of total energy expenditure and protein. Self-report was captured from food frequency questionnaire (FFQ), four day food record (4DFR) and 24 hr. dietary recall (24HR). Using regression calibration we estimated bias of self-reported dietary instruments including psychosocial factors from the Stunkard-Sorenson Body Silhouettes for body image perception, the Crowne-Marlowe Social Desirability Scale, and the Three Factor Eating Questionnaire (R-18) for cognitive restraint for eating, uncontrolled eating, and emotional eating. We included a diet behavior factor on number of meals eaten at home using the 4DFR. Results Three categories were defined for each of the six psychosocial and diet behavior variables (low, medium, high). Participants with high social desirability scores were more likely to under-report on the FFQ for energy (β = -0.174, SE = 0.054, p < 0.05) and protein intake (β = -0.142, SE = 0.062, p < 0.05) compared to participants with low social desirability scores. Participants consuming a high percentage of meals at home were less likely to under-report on the FFQ for energy (β = 0.181, SE = 0.053, p < 0.05) and protein (β = 0.127, SE = 0.06, p < 0.05) compared to participants consuming a low percentage of meals at home. In the calibration equations combining FFQ, 4DFR, 24HR with age, body mass index, race, and the psychosocial and diet behavior variables, the six psychosocial and diet variables explained 1.98%, 2.24%, and 2.15% of biomarker variation for energy, protein, and protein density respectively. The variations explained are

  11. Nasal carriage of methicillin-resistant Staphylococcus aureus (MRSA) among Swiss veterinary health care providers: detection of livestock- and healthcare-associated clones.

    PubMed

    Wettstein Rosenkranz, K; Rothenanger, E; Brodard, I; Collaud, A; Overesch, G; Bigler, B; Marschall, J; Perreten, V

    2014-07-01

    We screened a total of 340 veterinarians (including general practitioners, small animal practitioners, large animal practitioners, veterinarians working in different veterinary services or industry), and 29 veterinary assistants for nasal carriage of methicillin-resistant Staphylococcus aureus (MRSA) and Staphylococcus pseudintermedius (MRSP) at the 2012 Swiss veterinary annual meeting. MRSA isolates (n = 14) were detected in 3.8 % (95 % CI 2.1 - 6.3 %) of the participants whereas MRSP was not detected. Large animal practitioners were carriers of livestock-associated MRSA (LA-MRSA) ST398-t011-V (n = 2), ST398-t011-IV (n = 4), and ST398-t034-V (n = 1). On the other hand, participants working with small animals harbored human healthcare-associated MRSA (HCA-MRSA) which belonged to epidemic lineages ST225-t003-II (n = 2), ST225-t014-II (n = 1), ST5-t002-II (n = 2), ST5-t283-IV (n = 1), and ST88-t186-IV (n = 1). HCA-MRSA harbored virulence factors such as enterotoxins, β-hemolysin converting phage and leukocidins. None of the MRSA isolates carried Panton-Valentine leukocidin (PVL). In addition to the methicillin resistance gene mecA, LA-MRSA ST398 isolates generally contained additional antibiotic resistance genes conferring resistance to tetracycline [tet(M) and tet(K)], trimethoprim [dfrK, dfrG], and the aminoglycosides gentamicin and kanamycin [aac(6')-Ie - aph(2')-Ia]. On the other hand, HCA-MRSA ST5 and ST225 mainly contained genes conferring resistance to the macrolide, lincosamide and streptogramin B antibiotics [erm(A)], to spectinomycin [ant(9)-Ia], amikacin and tobramycin [ant(4')-Ia], and to fluoroquinolones [amino acid substitutions in GrlA (S84L) and GyrA (S80F and S81P)]. MRSA carriage may represent an occupational risk and veterinarians should be aware of possible MRSA colonization and potential for developing infection or for transmitting these strains. Professional exposure to animals should be reported upon hospitalization and before medical

  12. Molecular Characterization of Enterotoxin-Producing Escherichia coli Collected in 2011–2012, Russia

    PubMed Central

    Kartsev, Nikolay N.; Fursova, Nadezhda K.; Pachkunov, Dmitry M.; Bannov, Vasiliy A.; Eruslanov, Boris V.; Svetoch, Edward A.; Dyatlov, Ivan A.

    2015-01-01

    sulphonamides dfrA17-aadA5 and dfrA12-orfF-aadA2. One isolate ETEC_Ef-6 was found to be a multidrug-resistant (MDR) pathogen that carried both the beta-lactamase gene and class 1 integron. These data suggest the circulation of ETEC in Russia. Further investigations are necessary to study the spread of the revealed ETEC sequence types (STs) and serotypes. Their role in the etiology of diarrhea should be also estimated. PMID:25923803

  13. The Emergence and Spread of Multiple Livestock-Associated Clonal Complex 398 Methicillin-Resistant and Methicillin-Susceptible Staphylococcus aureus Strains among Animals and Humans in the Republic of Ireland, 2010–2014

    PubMed Central

    Brennan, Gráinne I.; Abbott, Yvonne; Burns, Aisling; Leonard, Finola; McManus, Brenda A.; O’Connell, Brian; Coleman, David C.; Shore, Anna C.

    2016-01-01

    Clonal complex (CC) 398 methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-susceptible S. aureus (MSSA) are associated with carriage and infection among animals and humans but only a single case of CC398 MRSA has been reported in the Republic of Ireland (ROI). The present study investigated the molecular epidemiology of CC398 MRSA (n = 22) and MSSA (n = 10) from animals and humans in the ROI from 2010–2014. Isolates underwent antimicrobial susceptibility testing, spa typing, DNA microarray profiling and PCR for CC398-associated resistance genes. All MRSA underwent SCCmec IV or V subtyping. Four distinct CC398-MRSA incidents were identified from (i) a man in a nursing home (spa type t011-SCCmec IVa, immune evasion complex (IEC) negative), (ii) a horse and veterinarian who had recently travelled to Belgium (t011-IVa, IEC positive), (iii) pigs (n = 9) and farm workers (n = 9) on two farms, one which had been restocked with German gilts and the other which was a finisher farm (t034-VT, IEC negative, 3/9 pigs; t011- VT, IEC negative, 6/9 pigs & 9/9 farm workers), and (iv) a child who had worked on a pig farm in the UK (t034-VT, IEC negative). Isolates also carried different combinations of multiple resistance genes including erm(A), erm(B), tet(K), tet(M) & tet(L), fexA, spc, dfrG, dfrK aacA-aphD and aadD further highlighting the presence of multiple CC398-MRSA strains. CC398 MSSA were recovered from pigs (n = 8) and humans (n = 2). CC398 MSSA transmission was identified among pigs but zoonotic transmission was not detected with animal and human isolates exhibiting clade-specific traits. This study highlights the importation and zoonotic spread of CC398 MRSA in the ROI and the spread of CC398 MSSA among pigs. Increased surveillance is warranted to prevent further CC398 MRSA importation and spread in a country that was considered CC398 MRSA free. PMID:26886749

  14. Molecular characterization of multidrug-resistant extended-spectrum β-lactamase-producing Enterobacteriaceae isolated in Antananarivo, Madagascar

    PubMed Central

    2013-01-01

    Background We investigated the molecular characteristics of multidrug-resistant, extended-spectrum β-lactamase (ESBL)-producing Enterobacteriaceae isolated in community settings and in hospitals in Antananarivo, Madagascar. Results Forty-nine E. coli, K. pneumoniae, K. oxytoca and E. cloacae ESBL-producing isolates were studied. In antimicrobial susceptibility analyses, many of the isolates exhibited resistance to aminoglycosides, fluoroquinolones and trimethoprim-sulfamethoxazole. Gene amplification analysis and sequencing revealed that 75.5% (n=37) of the isolates harbored blaCTX-M-15 and 38.7% (n=19) harbored blaSHV-12. The non-ESBLs resistance genes detected were blaTEM-1, blaOXA-1, aac(6′)-Ib,aac(6′)-Ib-cr, tetA, sul-1, sul-2, qnrA, qnrB and catB-3. We found dfrA and aadA gene cassettes in the class 1 integron variable regions of the isolates, and the combination of dfrA17-aadA5 to be the most prevalent. All blaCTX-M-15 positive isolates also contained the ISEcp1 insertion element. Conjugation and transformation experiments indicated that 70.3% of the antibiotic resistance genes resided on plasmids. Through a PCR based replicon typing method, plasmids carrying the blaSHV-12 or blaCTX-M-15 genes were assigned to either the IncFII replicon type or, rarely, to the HI2 replicon type. All isolates were subtyped by the rep-PCR and ERIC-PCR methods. Phylogenetic grouping and virulence genotyping of the E. coli isolates revealed that most of them belonged to group A1. One isolate assigned to group B2 harbored blaCTX-M-15 and five virulence genes (traT, fyuA, iutA, iha and sfa) and was related to the O25b-ST131 clone. Conclusions Our results highlight the dissemination of multidrug resistant Enterobacteriaceae isolates in Antananarivo. These findings underline the need for a rational use of antibiotic and for appropriate methods of screening ESBL in routine laboratories in Antananarivo. PMID:23594374

  15. Complete Nucleotide Sequences of blaKPC-4- and blaKPC-5-Harboring IncN and IncX Plasmids from Klebsiella pneumoniae Strains Isolated in New Jersey

    PubMed Central

    Chen, Liang; Chavda, Kalyan D.; Fraimow, Henry S.; Mediavilla, José R.; Melano, Roberto G.; Jacobs, Michael R.; Bonomo, Robert A.

    2013-01-01

    Klebsiella pneumoniae carbapenemase (KPC)-producing Enterobacteriaceae have emerged as major nosocomial pathogens. blaKPC, commonly located on Tn4401, is found in Gram-negative bacterial strains, with the two most common variants, blaKPC-2 and blaKPC-3, identified in plasmids with diverse genetic backgrounds. In this study, we examined blaKPC-4- and blaKPC-5-bearing plasmids recovered from two K. pneumoniae strains, which were isolated from a single New Jersey hospital in 2005 and 2006, respectively. IncN plasmid pBK31551 is 84 kb in length and harbors blaKPC-4, blaTEM-1, qnrB2, aac(3)-Ib, aph(3′)-I, qacF, qacEΔ1, sul1, and dfrA14, which confer resistance to β-lactams, quinolones, aminoglycosides, quaternary ammonium compounds, and co-trimoxazole. The conserved regions within pBK31551 are similar to those of other IncN plasmids. Surprisingly, analysis of the Tn4401 sequence revealed a large IS110- and Tn6901-carrying element (8.3 kb) inserted into the istA gene, encoding glyoxalase/bleomycin resistance, alcohol dehydrogenase, and S-formylglutathione hydrolase. Plasmid pBK31567 is 47 kb in length and harbors blaKPC-5, dfrA5, qacEΔ1, and sul1. pBK31567 belongs to a novel IncX subgroup (IncX5) and possesses a highly syntenic plasmid backbone like other IncX plasmids; however, sequence similarity at the nucleotide level is divergent. The blaKPC-5 gene is carried on a Tn4401 element and differs from the genetic environment of blaKPC-5 described in Pseudomonas aeruginosa strain P28 from Puerto Rico. This study underscores the genetic diversity of multidrug-resistant plasmids involved in the spread of blaKPC genes and highlights the mobility and plasticity of Tn4401. Comparative genomic analysis provides new insights into the evolution and dissemination of KPC plasmids belonging to different incompatibility groups. PMID:23114770

  16. Diversity of Plasmids Encoding Virulence and Resistance Functions in Salmonella enterica subsp. enterica Serovar Typhimurium Monophasic Variant 4,[5],12:i:- Strains Circulating in Europe

    PubMed Central

    García, Patricia; Hopkins, Katie L.; García, Vanesa; Beutlich, Janine; Mendoza, M. Carmen; Threlfall, John; Mevius, Dik; Helmuth, Reiner; Rodicio, M. Rosario; Guerra, Beatriz

    2014-01-01

    Plasmids encoding resistance and virulence properties in multidrug resistant (MDR) Salmonella enterica (S.) serovar Typhimurium monophasic variant 4,[5],12:i:- isolates recovered from pigs and humans (2006-2008) in Europe were characterised. The isolates were selected based on the detection by PCR-amplification of S. Typhimurium virulence plasmid pSLT genes and were analysed by multi-locus sequence typing (MLST). The resistance genes present in the isolates and the association of these genes with integrons, transposons and insertion sequences were characterised by PCR-sequencing, and their plasmid location was determined by alkaline lysis and by S1-nuclease pulsed-field gel electrophoresis (PFGE) Southern-blot hybridisation. Plasmids were further analysed by replicon typing, plasmid MLST and conjugation experiments. The 10 S. 4,[5],12,i:- selected isolates belonged to ST19. Each isolate carried a large plasmid in which MDR with pSLT-associated virulence genes were located. After analysis, eight different plasmids of three incompatibility groups (IncA/C, IncR and IncF) were detected. Two IncA/C plasmids represented novel variants within the plasmid family of the S. 4,[5],12:i:- Spanish clone, and carried an empty class 1 integron with a conventional qacEΔ1-sul1 3′ conserved segment or an In-sul3 type III with estX-psp-aadA2-cmlA1-aadA1-qacH variable region linked to tnpA440-sul3, part of Tn2, Tn21 and Tn1721 transposons, and ISCR2. Four newly described IncR plasmids contained the resistance genes within In-sul3 type I (dfrA12-orfF-aadA2-cmlA1-aadA1-qacH/tnpA440-sul3) and part of Tn10 [tet(B)]. Two pSLT-derivatives with FIIs-ST1+FIB-ST17 replicons carried cmlA1-[aadA1-aadA2]-sul3-dfrA12 and blaTEM-1 genes linked to an In-sul3 type I integron and to Tn2, respectively. In conclusion, three emerging European clones of S. 4,[5],12:i:- harboured MDR plasmids encoding additional virulence functions that could contribute significantly to their evolutionary success. PMID

  17. Diversity of plasmids encoding virulence and resistance functions in Salmonella enterica subsp. enterica serovar Typhimurium monophasic variant 4,[5],12:i:- strains circulating in Europe.

    PubMed

    García, Patricia; Hopkins, Katie L; García, Vanesa; Beutlich, Janine; Mendoza, M Carmen; Threlfall, John; Mevius, Dik; Helmuth, Reiner; Rodicio, M Rosario; Guerra, Beatriz

    2014-01-01

    Plasmids encoding resistance and virulence properties in multidrug resistant (MDR) Salmonella enterica (S.) serovar Typhimurium monophasic variant 4,[5],12:i:- isolates recovered from pigs and humans (2006-2008) in Europe were characterised. The isolates were selected based on the detection by PCR-amplification of S. Typhimurium virulence plasmid pSLT genes and were analysed by multi-locus sequence typing (MLST). The resistance genes present in the isolates and the association of these genes with integrons, transposons and insertion sequences were characterised by PCR-sequencing, and their plasmid location was determined by alkaline lysis and by S1-nuclease pulsed-field gel electrophoresis (PFGE) Southern-blot hybridisation. Plasmids were further analysed by replicon typing, plasmid MLST and conjugation experiments. The 10 S. 4,[5],12,i:- selected isolates belonged to ST19. Each isolate carried a large plasmid in which MDR with pSLT-associated virulence genes were located. After analysis, eight different plasmids of three incompatibility groups (IncA/C, IncR and IncF) were detected. Two IncA/C plasmids represented novel variants within the plasmid family of the S. 4,[5],12:i:- Spanish clone, and carried an empty class 1 integron with a conventional qacEΔ1-sul1 3' conserved segment or an In-sul3 type III with estX-psp-aadA2-cmlA1-aadA1-qacH variable region linked to tnpA440-sul3, part of Tn2, Tn21 and Tn1721 transposons, and ISCR2. Four newly described IncR plasmids contained the resistance genes within In-sul3 type I (dfrA12-orfF-aadA2-cmlA1-aadA1-qacH/tnpA440-sul3) and part of Tn10 [tet(B)]. Two pSLT-derivatives with FIIs-ST1+FIB-ST17 replicons carried cmlA1-[aadA1-aadA2]-sul3-dfrA12 and blaTEM-1 genes linked to an In-sul3 type I integron and to Tn2, respectively. In conclusion, three emerging European clones of S. 4,[5],12:i:- harboured MDR plasmids encoding additional virulence functions that could contribute significantly to their evolutionary success.

  18. International Spread and Persistence of TEM-24 Is Caused by the Confluence of Highly Penetrating Enterobacteriaceae Clones and an IncA/C2 Plasmid Containing Tn1696::Tn1 and IS5075-Tn21▿

    PubMed Central

    Novais, Ângela; Baquero, Fernando; Machado, Elisabete; Cantón, Rafael; Peixe, Luísa; Coque, Teresa M.

    2010-01-01

    TEM-24 remains one of the most widespread TEM-type extended-spectrum β-lactamases (ESBLs) among Enterobacteriaceae. To analyze the reasons influencing its spread and persistence, a multilevel population genetics study was carried out on 28 representative TEM-24 producers from Belgium, France, Portugal, and Spain (13 Enterobacter aerogenes isolates, 6 Escherichia coli isolates, 6 Klebsiella pneumoniae isolates, 2 Proteus mirabilis isolates, and 1 Klebsiella oxytoca isolate, from 1998 to 2004). Clonal relatedness (XbaI pulsed-field gel electrophoresis [PFGE] and E. coli phylogroups) and antibiotic susceptibility were determined by standard procedures. Plasmid analysis included determination of the incompatibility group (by PCR, hybridization, and/or sequencing) and comparison of restriction fragment length polymorphism (RFLP) patterns. Characterization of genetic elements conferring antibiotic resistance included integrons (classes 1, 2, and 3) and transposons (Tn3, Tn21, and Tn402). Similar PFGE patterns were identified among E. aerogenes, K. pneumoniae, and P. mirabilis isolates, while E. coli strains were diverse (phylogenetic groups A, B2, and D). Highly related 180-kb IncA/C2 plasmids conferring resistance to kanamycin, tobramycin, chloramphenicol, trimethoprim, and sulfonamides were identified. Each plasmid contained defective In0-Tn402 (dfrA1-aadA1, aacA4, or aacA4-aacC1-orfE-aadA2-cmlA1) and In4-Tn402 (aacA4 or dfrA1-aadA1) variants. These integrons were located within Tn21, Tn1696, or hybrids of these transposons, with IS5075 interrupting their IRtnp and IRmer. In all cases, blaTEM-24 was part of an IS5075-ΔTn1 transposon within tnp1696, mimicking other genetic elements containing blaTEM-2 and blaTEM-3 variants. The international dissemination of TEM-24 is fuelled by an IncA/C2 plasmid acquired by different enterobacterial clones which seem to evolve by gaining diverse genetic elements. This work highlights the risks of a confluence between highly

  19. Dissemination of Novel Antimicrobial Resistance Mechanisms through the Insertion Sequence Mediated Spread of Metabolic Genes.

    PubMed

    Furi, Leonardo; Haigh, Richard; Al Jabri, Zaaima J H; Morrissey, Ian; Ou, Hong-Yu; León-Sampedro, Ricardo; Martinez, Jose L; Coque, Teresa M; Oggioni, Marco R

    2016-01-01

    The widely used biocide triclosan selectively targets FabI, the NADH-dependent trans-2-enoyl-acyl carrier protein (ACP) reductase, which is also an important target for the development of narrow spectrum antibiotics. The analysis of triclosan resistant Staphylococcus aureus isolates had previously shown that in about half of the strains, the mechanism of triclosan resistance consists on the heterologous duplication of the triclosan target gene due to the acquisition of an additional fabI allele derived from Staphylococcus haemolyticus (sh-fabI). In the current work, the genomic sequencing of 10 of these strains allowed the characterization of two novel composite transposons TnSha1 and TnSha2 involved in the spread of sh-fabI. TnSha1 harbors one copy of IS1272, whereas TnSha2 is a 11.7 kb plasmid carrying TnSha1 present either as plasmid or in an integrated form generally flanked by two IS1272 elements. The target and mechanism of integration for IS1272 and TnSha1 are novel and include targeting of DNA secondary structures, generation of blunt-end deletions of the stem-loop and absence of target duplication. Database analyses showed widespread occurrence of these two elements in chromosomes and plasmids, with TnSha1 mainly in S. aureus and with TnSha2 mainly in S. haemolyticus and S. epidermidis. The acquisition of resistance by means of an insertion sequence-based mobilization and consequent duplication of drug-target metabolic genes, as observed here for sh-fabI, is highly reminiscent of the situation with the ileS2 gene conferring mupirocin resistance, and the dfrA and dfrG genes conferring trimethoprim resistance both of which are mobilized by IS257. These three examples, which show similar mechanisms and levels of spread of metabolic genes linked to IS elements, highlight the importance of this genetic strategy for recruitment and rapid distribution of novel resistance mechanisms in staphylococci.

  20. Ectopic Expression of the Coleus R2R3 MYB-Type Proanthocyanidin Regulator Gene SsMYB3 Alters the Flower Color in Transgenic Tobacco.

    PubMed

    Zhu, Qinlong; Sui, Shunzhao; Lei, Xinghua; Yang, Zhongfang; Lu, Kun; Liu, Guangde; Liu, Yao-Guang; Li, Mingyang

    2015-01-01

    Proanthocyanidins (PAs) play an important role in plant disease defense and have beneficial effects on human health. We isolated and characterized a novel R2R3 MYB-type PA-regulator SsMYB3 from a well-known ornamental plant, coleus (Solenostemon scutellarioides), to study the molecular regulation of PAs and to engineer PAs biosynthesis. The expression level of SsMYB3 was correlated with condensed tannins contents in various coleus tissues and was induced by wounding and light. A complementation test in the Arabidopsis tt2 mutant showed that SsMYB3 could restore the PA-deficient seed coat phenotype and activated expression of the PA-specific gene ANR and two related genes, DFR and ANS. In yeast two-hybrid assays, SsMYB3 interacted with the Arabidopsis AtTT8 and AtTTG1 to reform the ternary transcriptional complex, and also interacted with two tobacco bHLH proteins (NtAn1a and NtJAF13-1) and a WD40 protein, NtAn11-1. Ectopic overexpression of SsMYB3 in transgenic tobacco led to almost-white flowers by greatly reducing anthocyanin levels and enhancing accumulation of condensed tannins. This overexpression of SsMYB3 upregulated the key PA genes (NtLAR and NtANR) and late anthocyanin structural genes (NtDFR and NtANS), but downregulated the expression of the final anthocyanin gene NtUFGT. The formative SsMYB3-complex represses anthocyanin accumulation by directly suppressing the expression of the final anthocyanin structural gene NtUFGT, through competitive inhibition or destabilization of the endogenous NtAn2-complex formation. These results suggested that SsMYB3 may form a transcription activation complex to regulate PA biosynthesis in the Arabidopsis tt2 mutant and transgenic tobacco. Our findings suggest that SsMYB3 is involved in the regulation of PA biosynthesis in coleus and has the potential as a molecular tool for manipulating biosynthesis of PAs in fruits and other crops using metabolic engineering. PMID:26448466

  1. The sequence diversity and expression among genes of the folic acid biosynthesis pathway in industrial Saccharomyces strains.

    PubMed

    Goncerzewicz, Anna; Misiewicz, Anna

    2015-01-01

    Folic acid is an important vitamin in human nutrition and its deficiency in pregnant women's diets results in neural tube defects and other neurological damage to the fetus. Additionally, DNA synthesis, cell division and intestinal absorption are inhibited in case of adults. Since this discovery, governments and health organizations worldwide have made recommendations concerning folic acid supplementation of food for women planning to become pregnant. In many countries this has led to the introduction of fortifications, where synthetic folic acid is added to flour. It is known that Saccharomyces strains (brewing and bakers' yeast) are one of the main producers of folic acid and they can be used as a natural source of this vitamin. Proper selection of the most efficient strains may enhance the folate content in bread, fermented vegetables, dairy products and beer by 100% and may be used in the food industry. The objective of this study was to select the optimal producing yeast strain by determining the differences in nucleotide sequences in the FOL2, FOL3 and DFR1 genes of folic acid biosynthesis pathway. The Multitemperature Single Strand Conformation Polymorphism (MSSCP) method and further nucleotide sequencing for selected strains were applied to indicate SNPs in selected gene fragments. The RT qPCR technique was also applied to examine relative expression of the FOL3 gene. Furthermore, this is the first time ever that industrial yeast strains were analysed regarding genes of the folic acid biosynthesis pathway. It was observed that a correlation exists between the folic acid amount produced by industrial yeast strains and changes in the nucleotide sequence of adequate genes. The most significant changes occur in the DFR1 gene, mostly in the first part, which causes major protein structure modifications in KKP 232, KKP 222 and KKP 277 strains. Our study shows that the large amount of SNP contributes to impairment of the selected enzymes and S. cerevisiae and S

  2. Dissemination of Novel Antimicrobial Resistance Mechanisms through the Insertion Sequence Mediated Spread of Metabolic Genes.

    PubMed

    Furi, Leonardo; Haigh, Richard; Al Jabri, Zaaima J H; Morrissey, Ian; Ou, Hong-Yu; León-Sampedro, Ricardo; Martinez, Jose L; Coque, Teresa M; Oggioni, Marco R

    2016-01-01

    The widely used biocide triclosan selectively targets FabI, the NADH-dependent trans-2-enoyl-acyl carrier protein (ACP) reductase, which is also an important target for the development of narrow spectrum antibiotics. The analysis of triclosan resistant Staphylococcus aureus isolates had previously shown that in about half of the strains, the mechanism of triclosan resistance consists on the heterologous duplication of the triclosan target gene due to the acquisition of an additional fabI allele derived from Staphylococcus haemolyticus (sh-fabI). In the current work, the genomic sequencing of 10 of these strains allowed the characterization of two novel composite transposons TnSha1 and TnSha2 involved in the spread of sh-fabI. TnSha1 harbors one copy of IS1272, whereas TnSha2 is a 11.7 kb plasmid carrying TnSha1 present either as plasmid or in an integrated form generally flanked by two IS1272 elements. The target and mechanism of integration for IS1272 and TnSha1 are novel and include targeting of DNA secondary structures, generation of blunt-end deletions of the stem-loop and absence of target duplication. Database analyses showed widespread occurrence of these two elements in chromosomes and plasmids, with TnSha1 mainly in S. aureus and with TnSha2 mainly in S. haemolyticus and S. epidermidis. The acquisition of resistance by means of an insertion sequence-based mobilization and consequent duplication of drug-target metabolic genes, as observed here for sh-fabI, is highly reminiscent of the situation with the ileS2 gene conferring mupirocin resistance, and the dfrA and dfrG genes conferring trimethoprim resistance both of which are mobilized by IS257. These three examples, which show similar mechanisms and levels of spread of metabolic genes linked to IS elements, highlight the importance of this genetic strategy for recruitment and rapid distribution of novel resistance mechanisms in staphylococci. PMID:27446047

  3. The sequence diversity and expression among genes of the folic acid biosynthesis pathway in industrial Saccharomyces strains.

    PubMed

    Goncerzewicz, Anna; Misiewicz, Anna

    2015-01-01

    Folic acid is an important vitamin in human nutrition and its deficiency in pregnant women's diets results in neural tube defects and other neurological damage to the fetus. Additionally, DNA synthesis, cell division and intestinal absorption are inhibited in case of adults. Since this discovery, governments and health organizations worldwide have made recommendations concerning folic acid supplementation of food for women planning to become pregnant. In many countries this has led to the introduction of fortifications, where synthetic folic acid is added to flour. It is known that Saccharomyces strains (brewing and bakers' yeast) are one of the main producers of folic acid and they can be used as a natural source of this vitamin. Proper selection of the most efficient strains may enhance the folate content in bread, fermented vegetables, dairy products and beer by 100% and may be used in the food industry. The objective of this study was to select the optimal producing yeast strain by determining the differences in nucleotide sequences in the FOL2, FOL3 and DFR1 genes of folic acid biosynthesis pathway. The Multitemperature Single Strand Conformation Polymorphism (MSSCP) method and further nucleotide sequencing for selected strains were applied to indicate SNPs in selected gene fragments. The RT qPCR technique was also applied to examine relative expression of the FOL3 gene. Furthermore, this is the first time ever that industrial yeast strains were analysed regarding genes of the folic acid biosynthesis pathway. It was observed that a correlation exists between the folic acid amount produced by industrial yeast strains and changes in the nucleotide sequence of adequate genes. The most significant changes occur in the DFR1 gene, mostly in the first part, which causes major protein structure modifications in KKP 232, KKP 222 and KKP 277 strains. Our study shows that the large amount of SNP contributes to impairment of the selected enzymes and S. cerevisiae and S

  4. The Emergence and Spread of Multiple Livestock-Associated Clonal Complex 398 Methicillin-Resistant and Methicillin-Susceptible Staphylococcus aureus Strains among Animals and Humans in the Republic of Ireland, 2010-2014.

    PubMed

    Brennan, Gráinne I; Abbott, Yvonne; Burns, Aisling; Leonard, Finola; McManus, Brenda A; O'Connell, Brian; Coleman, David C; Shore, Anna C

    2016-01-01

    Clonal complex (CC) 398 methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-susceptible S. aureus (MSSA) are associated with carriage and infection among animals and humans but only a single case of CC398 MRSA has been reported in the Republic of Ireland (ROI). The present study investigated the molecular epidemiology of CC398 MRSA (n = 22) and MSSA (n = 10) from animals and humans in the ROI from 2010-2014. Isolates underwent antimicrobial susceptibility testing, spa typing, DNA microarray profiling and PCR for CC398-associated resistance genes. All MRSA underwent SCCmec IV or V subtyping. Four distinct CC398-MRSA incidents were identified from (i) a man in a nursing home (spa type t011-SCCmec IVa, immune evasion complex (IEC) negative), (ii) a horse and veterinarian who had recently travelled to Belgium (t011-IVa, IEC positive), (iii) pigs (n = 9) and farm workers (n = 9) on two farms, one which had been restocked with German gilts and the other which was a finisher farm (t034-VT, IEC negative, 3/9 pigs; t011-VT, IEC negative, 6/9 pigs & 9/9 farm workers), and (iv) a child who had worked on a pig farm in the UK (t034-VT, IEC negative). Isolates also carried different combinations of multiple resistance genes including erm(A), erm(B), tet(K), tet(M) & tet(L), fexA, spc, dfrG, dfrK aacA-aphD and aadD further highlighting the presence of multiple CC398-MRSA strains. CC398 MSSA were recovered from pigs (n = 8) and humans (n = 2). CC398 MSSA transmission was identified among pigs but zoonotic transmission was not detected with animal and human isolates exhibiting clade-specific traits. This study highlights the importation and zoonotic spread of CC398 MRSA in the ROI and the spread of CC398 MSSA among pigs. Increased surveillance is warranted to prevent further CC398 MRSA importation and spread in a country that was considered CC398 MRSA free. PMID:26886749

  5. Ectopic Expression of the Coleus R2R3 MYB-Type Proanthocyanidin Regulator Gene SsMYB3 Alters the Flower Color in Transgenic Tobacco

    PubMed Central

    Zhu, Qinlong; Sui, Shunzhao; Lei, Xinghua; Yang, Zhongfang; Lu, Kun; Liu, Guangde; Liu, Yao-Guang; Li, Mingyang

    2015-01-01

    Proanthocyanidins (PAs) play an important role in plant disease defense and have beneficial effects on human health. We isolated and characterized a novel R2R3 MYB-type PA-regulator SsMYB3 from a well-known ornamental plant, coleus (Solenostemon scutellarioides), to study the molecular regulation of PAs and to engineer PAs biosynthesis. The expression level of SsMYB3 was correlated with condensed tannins contents in various coleus tissues and was induced by wounding and light. A complementation test in the Arabidopsis tt2 mutant showed that SsMYB3 could restore the PA-deficient seed coat phenotype and activated expression of the PA-specific gene ANR and two related genes, DFR and ANS. In yeast two-hybrid assays, SsMYB3 interacted with the Arabidopsis AtTT8 and AtTTG1 to reform the ternary transcriptional complex, and also interacted with two tobacco bHLH proteins (NtAn1a and NtJAF13-1) and a WD40 protein, NtAn11-1. Ectopic overexpression of SsMYB3 in transgenic tobacco led to almost-white flowers by greatly reducing anthocyanin levels and enhancing accumulation of condensed tannins. This overexpression of SsMYB3 upregulated the key PA genes (NtLAR and NtANR) and late anthocyanin structural genes (NtDFR and NtANS), but downregulated the expression of the final anthocyanin gene NtUFGT. The formative SsMYB3-complex represses anthocyanin accumulation by directly suppressing the expression of the final anthocyanin structural gene NtUFGT, through competitive inhibition or destabilization of the endogenous NtAn2-complex formation. These results suggested that SsMYB3 may form a transcription activation complex to regulate PA biosynthesis in the Arabidopsis tt2 mutant and transgenic tobacco. Our findings suggest that SsMYB3 is involved in the regulation of PA biosynthesis in coleus and has the potential as a molecular tool for manipulating biosynthesis of PAs in fruits and other crops using metabolic engineering. PMID:26448466

  6. Characterization of Flavan-3-ols and Expression of MYB and Late Pathway Genes Involved in Proanthocyanidin Biosynthesis in Foliage of Vitis bellula

    PubMed Central

    Zhu, Yue; Peng, Qing-Zhong; Du, Ci; Li, Ke-Gang; Xie, De-Yu

    2013-01-01

    Proanthocyanidins (PAs) are fundamental nutritional metabolites in different types of grape products consumed by human beings. Although the biosynthesis of PAs in berry of Vitis vinifera has gained intensive investigations, the understanding of PAs in other Vitis species is limited. In this study, we report PA formation and characterization of gene expression involved in PA biosynthesis in leaves of V. bellula, a wild edible grape species native to south and south-west China. Leaves are collected at five developmental stages defined by sizes ranging from 0.5 to 5 cm in length. Analyses of thin layer chromatography (TLC) and high performance liquid chromatography-photodiode array detector (HPLC-PAD) show the formation of (+)-catechin, (−)-epicatechin, (+)-gallocatechin and (−)-epigallocatechin during the entire development of leaves. Analyses of butanol-HCl boiling cleavage coupled with spectrometry measurement at 550 nm show a temporal trend of extractable PA levels, which is characterized by an increase from 0.5 cm to 1.5 cm long leaves followed by a decrease in late stages. TLC and HPLC-PAD analyses identify cyanidin, delphinidin and pelargonidin produced from the cleavage of PAs in the butanol-HCl boiling, showing that the foliage PAs of V. bellula include three different types of extension units. Four cDNAs, which encode VbANR, VbDFR, VbLAR1 and VbLAR2, respectively, are cloned from young leaves. The expression patterns of VbANR and VbLAR2 but not VbLAR1 and VbDFR follow a similar trend as the accumulation patterns of PAs. Two cDNAs encoding VbMYBPA1 and VbMYB5a, the homologs of which have been demonstrated to regulate the expression of both ANR and LAR in V. vinifera, are also cloned and their expression profiles are similar to those of VbANR and VbLAR2. In contrast, the expression profiles of MYBA1 and 2 homologs involved in anthocyanin biosynthesis are different from those of VbANR and VbLAR2. Our data show that both ANR and LAR branches are involved in

  7. Dissemination of Novel Antimicrobial Resistance Mechanisms through the Insertion Sequence Mediated Spread of Metabolic Genes

    PubMed Central

    Furi, Leonardo; Haigh, Richard; Al Jabri, Zaaima J. H.; Morrissey, Ian; Ou, Hong-Yu; León-Sampedro, Ricardo; Martinez, Jose L.; Coque, Teresa M.; Oggioni, Marco R.

    2016-01-01

    The widely used biocide triclosan selectively targets FabI, the NADH-dependent trans-2-enoyl-acyl carrier protein (ACP) reductase, which is also an important target for the development of narrow spectrum antibiotics. The analysis of triclosan resistant Staphylococcus aureus isolates had previously shown that in about half of the strains, the mechanism of triclosan resistance consists on the heterologous duplication of the triclosan target gene due to the acquisition of an additional fabI allele derived from Staphylococcus haemolyticus (sh-fabI). In the current work, the genomic sequencing of 10 of these strains allowed the characterization of two novel composite transposons TnSha1 and TnSha2 involved in the spread of sh-fabI. TnSha1 harbors one copy of IS1272, whereas TnSha2 is a 11.7 kb plasmid carrying TnSha1 present either as plasmid or in an integrated form generally flanked by two IS1272 elements. The target and mechanism of integration for IS1272 and TnSha1 are novel and include targeting of DNA secondary structures, generation of blunt-end deletions of the stem-loop and absence of target duplication. Database analyses showed widespread occurrence of these two elements in chromosomes and plasmids, with TnSha1 mainly in S. aureus and with TnSha2 mainly in S. haemolyticus and S. epidermidis. The acquisition of resistance by means of an insertion sequence-based mobilization and consequent duplication of drug-target metabolic genes, as observed here for sh-fabI, is highly reminiscent of the situation with the ileS2 gene conferring mupirocin resistance, and the dfrA and dfrG genes conferring trimethoprim resistance both of which are mobilized by IS257. These three examples, which show similar mechanisms and levels of spread of metabolic genes linked to IS elements, highlight the importance of this genetic strategy for recruitment and rapid distribution of novel resistance mechanisms in staphylococci. PMID:27446047

  8. Microbiological quality of ready-to-eat salads: an underestimated vehicle of bacteria and clinically relevant antibiotic resistance genes.

    PubMed

    Campos, Joana; Mourão, Joana; Pestana, Nazaré; Peixe, Luísa; Novais, Carla; Antunes, Patrícia

    2013-09-16

    The increase demand for fresh vegetables is causing an expansion of the market for minimally processed vegetables along with new recognized food safety problems. To gain further insight on this topic we analyzed the microbiological quality of Portuguese ready-to-eat salads (RTS) and their role in the spread of bacteria carrying acquired antibiotic resistance genes, food products scarcely considered in surveillance studies. A total of 50 RTS (7 brands; split or mixed leaves, carrot, corn) were collected in 5 national supermarket chains in Porto region (2010). They were tested for aerobic mesophilic counts, coliforms and Escherichia coli counts as well as for the presence of Salmonella and Listeria monocytogenes. Samples were also plated in different selective media with/without antibiotics before and after enrichment. The E. coli, other coliforms and Enterococcus recovered were characterized for antibiotic resistance profiles and clonality with phenotypic and genetic approaches. A high number of RTS presented poor microbiological quality (86%--aerobic mesophilic counts, 74%--coliforms, 4%--E. coli), despite the absence of screened pathogens. In addition, a high diversity of bacteria (species and clones) and antibiotic resistance backgrounds (phenotypes and genotypes) were observed, mostly with enrichment and antibiotic selective media. E. coli was detected in 13 samples (n=78; all types and 4 brands; phylogenetic groups A, B1 and D; none STEC) with resistance to tetracycline [72%; tet(A) and/or tet(B)], streptomycin (58%; aadA and/or strA-strB), sulfamethoxazole (50%; sul1 and/or sul2), trimethoprim (50%; dfrA1 or dfrA12), ampicillin (49%; blaTEM), nalidixic acid (36%), ciprofloxacin (5%) or chloramphenicol (3%; catA). E. coli clones, including the widespread group D/ST69, were detected in different samples from the same brand or different brands pointing out to a potential cross-contamination. Other clinically relevant resistance genes were detected in 2 Raoultella

  9. High-intensity focused ultrasound (HIFU) using Sonablate{trade mark, serif} devices for the treatment of localized prostate cancer: 13-year experience

    NASA Astrophysics Data System (ADS)

    Uchida, Toyoaki; Tomonaga, Tetsuro; Shoji, Sunao; Kim, Hakushi; Nagata, Yoshihiro

    2012-11-01

    To report on the long-term results of high-intensity focused ultrasound (HIFU) in the treatment of localized prostate cancer. Eight hundred and eighty-four men with prostate cancer treated with Sonablate® (SB) devices were included. All patients were followed for more than 2 years. The patients were divided into three groups: in the first group, 419 patients were treated with SB200/500 from 1999 to 2006; in the second group, 263 patients were treated with SB 500 ver. 4 from 2005 to 2009: in the third group, 202 patients were treated with SB 500 TCM from 2007 up to present. Biochemical failure was defined according to the Phoenix definition (PSA nadir + 2 ng/ml). The mean age, PSA, Gleason score, operation time, and follow-up period in each group were 68, 66 and 67 years, 11.2, 9.7 and 9.3 ng/ml, 6.2, 6.6 and 6.7, 167, 101 and 106 min, and 56, 48 and 36 months, respectively. The biochemical disease-free rate (bDFR) in each group at 5 years was, respectively, 54%, 61% and 84%, and was 50% at 10 years in the SB200/500 group (p<0.0001). The bDFR in patients in the low-, intermediate-, and high-risk groups in all patients at 10 years were 72% and 58%, 44%, respectively (p<0.0001). The BDFR in patients in the low-, intermediate-, and high-risk groups in the SB500 TCM group at 5 years were 97%, 83%, and 74% (p=0.0056). The negative prostate biopsy rates in 3 groups were 81%, 92% and 88%, respectively. As post HIFU complications, urethral stricture, acute epididymitis and urinary incontinence were noted in 18.0%, 6.2% and 1.9%, respectively. Rectourethral fistula was occurred in 0.6% in the first HIFU cases, Postoperative erectile dysfunction was noted in 27% of patients at 2 years after HIFU. HIFU therapy appears to be minimally invasive, efficacious, and safe for patients with localized prostate cancer. Technological advances as well as cultural and economic vectors have caused a shift from to minimally invasive techniques.

  10. Molecular Characterization of Enterotoxin-Producing Escherichia coli Collected in 2011-2012, Russia.

    PubMed

    Kartsev, Nikolay N; Fursova, Nadezhda K; Pachkunov, Dmitry M; Bannov, Vasiliy A; Eruslanov, Boris V; Svetoch, Edward A; Dyatlov, Ivan A

    2015-01-01

    sulphonamides dfrA17-aadA5 and dfrA12-orfF-aadA2. One isolate ETEC_Ef-6 was found to be a multidrug-resistant (MDR) pathogen that carried both the beta-lactamase gene and class 1 integron. These data suggest the circulation of ETEC in Russia. Further investigations are necessary to study the spread of the revealed ETEC sequence types (STs) and serotypes. Their role in the etiology of diarrhea should be also estimated. PMID:25923803

  11. Molecular Characterization of Enterotoxin-Producing Escherichia coli Collected in 2011-2012, Russia.

    PubMed

    Kartsev, Nikolay N; Fursova, Nadezhda K; Pachkunov, Dmitry M; Bannov, Vasiliy A; Eruslanov, Boris V; Svetoch, Edward A; Dyatlov, Ivan A

    2015-01-01

    sulphonamides dfrA17-aadA5 and dfrA12-orfF-aadA2. One isolate ETEC_Ef-6 was found to be a multidrug-resistant (MDR) pathogen that carried both the beta-lactamase gene and class 1 integron. These data suggest the circulation of ETEC in Russia. Further investigations are necessary to study the spread of the revealed ETEC sequence types (STs) and serotypes. Their role in the etiology of diarrhea should be also estimated.

  12. In Vitro Biosynthesis and Chemical Identification of UDP-N-acetyl-d-quinovosamine (UDP-d-QuiNAc)*

    PubMed Central

    Li, Tiezheng; Simonds, Laurie; Kovrigin, Evgenii L.; Noel, K. Dale

    2014-01-01

    N-acetyl-d-quinovosamine (2-acetamido-2,6-dideoxy-d-glucose, QuiNAc) occurs in the polysaccharide structures of many Gram-negative bacteria. In the biosynthesis of QuiNAc-containing polysaccharides, UDP-QuiNAc is the hypothetical donor of the QuiNAc residue. Biosynthesis of UDP-QuiNAc has been proposed to occur by 4,6-dehydration of UDP-N-acetyl-d-glucosamine (UDP-GlcNAc) to UDP-2-acetamido