Highly enantioselective production of a chiral intermediate of sitagliptin by a novel isolate of <Emphasis Type="Italic">Pseudomonas pseudoalcaligenes</Emphasis>

Objective

To produce (S)-3-hydroxy-1-(3-(trifluoromethyl)-5,6-dihydro[1,2,4]triazolo[4,3-a]pyrazin-7(8H)-yl]-4-(2,4,5-trifluorophenyl)butan-1-one (S)-1 from 4-oxo-4-[3-(trifluoromethyl)-5,6-dihydro [1,2,4]triazolo[4,3-a]pyrazin-7(8H)-yl)-1-(2,4,5-trifluorophenyl)butan-2-one (2) by microbial bioreduction.

Results

A new isolate of Pseudomonas pseudoalcaligenes reduced enantioselectively prochiral ketone 2 to chiral alcohol (S)-1. Whole cells of the bacterium were tolerant towards 20 % (v/v) DMSO and 10 g 2/l. Under the optimal conditions, the preparative-scale bioreduction yielded (S)-1 at 90 % yield and >99 % ee. Cells could be re-used with the yield and ee of product being 45 % and >99 %, respectively, after five cycles.

Conclusion

Bioreduction using whole cells of P. pseudoalcaligenes is an attractive approach to produce (S)-1, as a chiral intermediate of the anti-diabetic drug, sitagliptin.

     

Revision of the carabid genus <Emphasis Type="Italic">Meroctenus</Emphasis> Gemminger et Harold,with a new status of <Emphasis Type="Italic">Xenodochus</Emphasis> Andrewes (Coleoptera,Carabidae: Harpalini)

Originally described as a monotypical genus with unclear taxonomic position from Sudan, Meroctenus Gemminger et Harold, 1868 is treated as a polytypical genus of the Selenophori genus group with two subgenera: Meroctenus s. str. and Xenodochus Andrewes, 1941, stat. n. (the latter was previously considered a distinct genus). Within Meroctenus, two species are recognized: M. (Meroctenus) crenulatus Chaudoir, 1843 (type species) and M. (M.) mediocris (Andrewes, 1936), comb, n., transferred to Meroctenus s. str. from Xenodochus. A new subspecies M. (M.) crenulatus orientalis subsp. n. is described from Pakistan. Diagnoses of the genus Meroctenus in new interpretation as well as of its two subgenera are discussed, and a taxonomic review of the subgenus Meroctenus s. str. with a key to the species and subspecies is provided. The following synonymy is proposed: Meroctenus Gemminger et Harold, 1868 = Paregaploa Müller, 1947, syn. n.; Meroctenus crenulatus (Chaudoir, 1843) = Egaploa (Paregaploa) conviva Müller, 1947, syn. n. Lectotypes are designated for Ctenomerus crenulatus Chaudoir, 1843 and Xenodus mediocris Andrewes, 1936.     

Fine mapping of a male sterility gene <Emphasis Type="Italic">ms</Emphasis>-<Emphasis Type="Italic">3</Emphasis> in a novel cucumber (<Emphasis Type="Italic">Cucumis sativus</Emphasis> L.) mutant

Key message

The cucumber male sterility gene ms - 3 was fine mapped in a 76 kb region harboring an MMD1 -like gene Csa3M006660 that may be responsible for the male sterile in cucumber.

Abstract

A cucumber (Cucumis sativus L.) male sterile mutant (ms-3) in an advanced-generation inbred line was identified, and genetic analysis revealed that the male sterility trait was controlled by a recessive nuclear gene, ms-3, which was stably inherited. Histological studies suggested that the main cause of the male sterility was defective microsporogenesis, resulting in no tetrad or microspores being formed. Bulked segregant analysis (BSA) and genotyping of an F₂ population of 2553 individuals were employed used to fine map ms-3, which was delimited to a 76 Kb region. In this region, a single non-synonymous SNP was found in the Csa3M006660 gene locus, which was predicted to result in an amino acid change. Quantitative RT-PCR analysis of Csa3M006660 was consistent with the fact that it plays a role in the early development of cucumber pollen. The protein encoded by Csa3M006660 is predicted to be homeodomain (PHD) finger protein, and the high degree of sequence conservation with homologs from a range of plant species further suggested the importance of the ms-3 non-synonymous mutation. The data presented here provide support for Csa3M006660 as the most likely candidate gene for Ms-3.

     

The seed dormancy allele <Emphasis Type="Italic">TaSdr</Emphasis>-<Emphasis Type="Italic">A1a</Emphasis> associated with pre-harvest sprouting tolerance is mainly present in Chinese wheat landraces

Key message

We cloned TaSdr - A1 gene, and developed a gene-specific marker for TaSdr - A1 . A QTL for germination index at the TaSdr - A1 locus was identified in the Yangxiaomai/Zhongyou 9507 RIL population.

Abstract

Pre-harvest sprouting (PHS) affects yield and end-use quality in bread wheat (Triticum aestivum L.). In the present study we found an association between the TaSdr-A1 gene and PHS tolerance in bread wheat. TaSdr-A1 on chromosome 2A was cloned using a homologous cloning approach. Sequence analysis of TaSdr-A1 revealed an SNP at position 643, with the G allele being present in genotypes with lower germination index (GI) values and A in those with higher GI. These alleles were designated as TaSdr-A1a and TaSdr-A1b, respectively. A cleaved amplified polymorphism sequence (CAPS) marker Sdr2A based on the SNP was developed, and linkage mapping and QTL analysis were conducted to confirm the association between TaSdr-A1 and seed dormancy. Sdr2A was located in a 2.9 cM interval between SSR markers Xgwm95 and Xgwm372. A QTL for GI at the TaSdr-A1 locus explained 6.6, 7.3, and 8.2 % of the phenotypic variances in a Yangxiaomai/Zhongyou 9507 RIL population grown at Beijing, Shijiazhuang, and the averaged data from the two environments, respectively. Two sets of Chinese wheat cultivars used for validating the TaSdr-A1 polymorphism and the corresponding gene-specific marker Sdr2A showed that TaSdr-A1 was significantly associated with GI. Among 29 accessions with TaSdr-A1a, 24 (82.8 %) were landraces, indicating the importance of Chinese wheat landraces as sources of PHS tolerance. This study identified a novel PHS resistance allele TaSdr-A1a mainly presented in Chinese landraces and it is likely to be the causal gene for QPhs.ccsu-2A.3, providing new information for an understanding of seed dormancy.

     

Pancreatic lipase inhibitory activity of monogalactosyldiacylglycerols isolated from the freshwater microalga <Emphasis Type="Italic">Chlorella sorokiniana</Emphasis>

Chemical investigation of the freshwater microalga Chlorella sorokiniana led to the isolation of a monogalactosyldiacylglycerol (MGDG)-rich fraction possessing dose-dependent inhibitory activity against pancreatic lipase activity. The MGDG-rich fraction contains 12 MGDGs identified by LC/HRMS analysis. Among them, three MGDGs were new compounds, namely, (2S)-1-O-(7Z,10Z-hexadecadienoyl)-2-O-(7Z,10Z,13Z-hexadecatrienoyl)-3-O-β-D-galactopyranosylglycerol (1), (2S)-1-O-linoleoyl-2-O-(7Z,10Z-hexadecadienoyl)-3-O-β-D-galactopyranosylglycerol (6), and (2S)-1-O-oleoyl-2-O-(7Z,10Z-hexadecadienoyl)-3-O-β-D-galactopyranosylglycerol (8). The major galactolipids were isolated by semipreparative HPLC and tested for their effect toward pancreatic lipase inhibitory activity. All the tested MGDGs showed significant reduction of pancreatic lipase activity indicating possible beneficial use for management of lipase-related disorders such as obesity.     

Epigenetic regulation of <Emphasis Type="Italic">MdMYB1</Emphasis> is associated with paper bagging-induced red pigmentation of apples

Main conclusion

Paper-bagging treatment can transform non-transcribed MdMYB1 - 2 and MdMYB1 - 3 alleles into transcribed alleles through epigenetic regulations, resulting in the red pigmentation of a normally non-red apple cultivar ‘Mutsu.’ Anthocyanin biosynthesis in apples is regulated by MdMYB1/A/10, an R2R3-Type MYB gene. ‘Mutsu,’ a triploid apple cultivar harboring non-transcribed MdMYB1-2 and MdMYB1-3 alleles, retains green skin color under field conditions. However, it can show red/pink pigmentation under natural or artificial ultraviolet-B (UV-B) light exposure after paper-bagging and bag removal treatment. In the present study, we found that in ‘Mutsu,’ paper bagging-induced red pigmentation was due to the activation of non-transcribed MdMYB1-2/-3 alleles, which triggered the expression of downstream anthocyanin biosynthesis genes in a UV-B-dependent manner. By monitoring the epigenetic changes during UV-B-induced pigmentation, no significant differences in DNA methylation and histone modifications in the 5′ upstream region of MdMYB1-2/-3 were recorded between the UV-B-treated fruit skin (red) and the fruit skin treated only by white light (green). In contrast, bag treatment lowered the DNA methylation in this region of MdMYB1-2/-3 alleles. Similarly, higher levels of histone H3 acetylation and trimethylation of H3 tail at lysine 4, and lower level of trimethylation of H3 tail at lysine 27 were observed in the 5′ upstream region of MdMYB1-2/-3 in the skin of the fruit immediately after bag removal. These results suggest that bagging treatment can induce epigenetic changes, facilitating the binding of trans factor(s) to MdMYB1-2/-3 alleles, resulting in the activation of these MYBs after bag removal.

     

A consortium of immobilized rhodococci for oilfield wastewater treatment in a column bioreactor

The possible use of a consortium of actinobacteria from the genus Rhodococcus immobilized on a polymeric carrier has been investigated for oilfield wastewater treatment in a bioreactor. It has been found that Rhodococcus opacus IEGM 263 and Rhodococcus ruber IEGM 231 cells remain viable at high concentrations of mineral salts in water and are able to oxidize oil hydrocarbons up to 62–81%. It has been shown that the consortium of rhodococci was more efficient in the elimination of hydrocarbons from wastewater than monocultures.     

Growth of <Emphasis Type="Italic">Dehalococcoides mccartyi</Emphasis> species in an autotrophic consortium producing limited acetate

The dechlorinating Dehalococcoides mccartyi species requires acetate as carbon source, but little is known on its growth under acetate limiting conditions. In this study, we observed growth and dechlorination of a D. mccartyi-containing mixed consortium in a fixed-carbon-free medium with trichloroethene in the aqueous phase and H₂/CO₂ in the headspace. Around 4 mM formate was produced by day 40, while acetate was constantly below 0.05 mM. Microbial community analysis of the consortium revealed dominance by D. mccartyi and Desulfovibrio sp. (57 and 22% 16S rRNA gene copies, respectively). From this consortium, Desulfovibrio sp. strain F1 was isolated and found to produce formate and acetate (1.2 mM and 48 µM, respectively, by day 24) when cultivated alone in the above mentioned medium without trichloroethene. An established co-culture of strain F1 and D. mccartyi strain 195 demonstrated that strain 195 could grow and dechlorinate using acetate produced by strain F1; and that acetate was constantly below 25 µM in the co-culture. To verify that such low level of acetate is utilizable by D. mccartyi, we cultivated strain 195 alone under acetate-limiting conditions and found that strain 195 consumed acetate to below detection (5 µM). Based on the acetate consumption and cell yield of D. mccartyi, we estimated that on average 1.2?×?10⁸ acetate molecules are needed to supply carbon for one D. mccartyi cell. Our study suggests that Desulfovibrio may supply a steady but low amount of fixed carbon to dechlorinating bacteria, exhibiting important implications for natural bio-attenuation when fixed carbon is limited.     

New and little-known crickets of the subfamily Phalangopsinae (Orthoptera,Gryllidae). 11. The genus <Emphasis Type="Italic">Parendacustes</Emphasis> (Part 2)

Discussions concerning the composition of the genus Parendacustes Chop., in particular, its subgenus Minizacla Gor., are continued. Eleven new taxa of this subgenus are described: P. trusmadi sp. n., P. mulu sp. n., P. brevispina sp. n., P. modispina sp. n., P. longispina sp. n., P. forficula sabah subsp. n., P. doloduo sp. n., P. buton sp. n., P. pallescens sp. n., P. kendari sp. n., and P. lindu sp. n. New data on P. makassari Gor. are also provided.     

Characterisation of a novel quantitative trait locus, <Emphasis Type="Italic">GN4</Emphasis>-<Emphasis Type="Italic">1</Emphasis>, for grain number and yield in rice (<Emphasis Type="Italic">Oryza sativa</Emphasis> L.)

Key message

A novel QTL for grain number, GN4-1, was identified and fine-mapped to an ~ 190-kb region on the long arm of rice chromosome 4.

Abstract

Rice grain yield is primarily determined by three components: number of panicles per plant, grain number per panicle and grain weight. Among these traits, grain number per panicle is the major contributor to grain yield formation and is a crucial trait for yield improvement. In this study, we identified a major quantitative trait locus (QTL) responsible for rice grain number on chromosome 4, designated GN4-1 (a QTL for Grain Number on chromosome 4), using advanced segregating populations derived from the crosses between an elite indica cultivar ‘Zhonghui 8006’ (ZH8006) and a japonica rice ‘Wuyunjing 8’ (WYJ8). GN4-1 was delimited to an ~ 190-kb region on chromosome 4. The genetic effect of GN4-1 was estimated using a pair of near-isogenic lines. The GN4-1 gene from WYJ8 promoted accumulation of cytokinins in the inflorescence and increased grain number per panicle by ~ 17%. More importantly, introduction of the WYJ8 GN4-1 gene into ZH8006 increased grain yield by ~ 14.3 and ~ 11.5% in the experimental plots in 2014 and 2015, respectively. In addition, GN4-1 promoted thickening of the culm and may enhance resistance to lodging. These results demonstrate that GN4-1 is a potentially valuable gene for improvement of yield and lodging resistance in rice breeding.

     

Assessing <Emphasis Type="Italic">Azorella</Emphasis> (Apiaceae) and its allies: Phylogenetics and a new classification

Phylogenetic analyses of nuclear rDNA spacers (ITS and ETS) from Azorella and five closely related genera confirm earlier plastid results indicating that Azorella, Huanaca, Mulinum, and Schizeilema are all polyphyletic, and that the monotypic genus Laretia is nested within one of the six subclades of Azorella. Only Stilbocarpa is monophyletic, but that genus is embedded within a larger clade that includes representatives of three other genera (Azorella, Huanaca, and Schizeilema). Both nuclear and plastid datasets identify the same 10 clades, but the placement of these clades remains unstable. A new classification is presented in which these six genera are reduced to a single genus (Azorella) comprising 58 species arranged in 10 sections, one of which is newly described here (Azorella sect. Ranunculus), and one lectotypified (Azorella sect. Glabratae). A total of 13 new combinations are made (A. albovaginata, A. allanii, A. boelckei, A. burkartii, A. colensoi, A. echegarayi, A. echinus, A. hallei, A. lyallii, A. polaris, A. prolifera, A. robusta, A. ulicina), along with three replacement names (A. atacamensis, A. ruizii, A. schizeilema). For each of the 58 accepted species, a full synonymy is provided along with geographic ranges (and nomenclatural notes, where useful).     

A Review of the Fauna of the Planthopper Family Lophopidae (Homoptera,Fulgoroidea) from Vietnam with Descriptions of New Genera and Species and with Taxonomic Notes

Six new species are described from Vietnam, Apia simplexsp. n., Bisma angulatasp. n., Pitambara trypetoidessp. n., Sarmatoca cathemerinagen. et sp. n., Maracota soulieraegen. et sp. n., and Serida castaneasp. n., and Zeleja thoracalissp. n. from Malaysia. Two new genera, Maracotagen. n. (type species M. soulierae sp. n.) and Sarmatocagen. n. (type species S. cathemerina sp. n.), and a new subgenus Zelomachasubgen. n. (type species Zeleja thoracalis sp. n.) in the genus Zeleja are erected. The genus Binaluana Soulier-Perkins et Stroiński is downgraded to a subgenus of Zeleja Melichar. The genus Silvispina Wang et Soulier-Perkins with an uncertain systematic position is attributed to the subfamily Lophopinae and the tribe Lophopini. The placement of the genus Elasmoscelis Spinola (and, correspondingly, the tribe Elasmoscelini) in the subfamily Menoscinae, the genus Epiptyxis Gerstaecker in the tribe Menoscini, and the genus Jivatma Melichar in the tribe Lophopini is substantiated. New records of 11 species are given from Vietnam, some being first records for this country.     

Systematic studies in Neotropical <Emphasis Type="Italic">Rinorea</Emphasis> (Violaceae): Two new sections and a new generic segregate

Rinorea, the second most species-rich genus in the Violaceae, has been shown to be polyphyletic with four separate clades recovered in phylogenetic studies. Among these clades is the Rinorea crenata group, which is composed of three Neotropical species. This group has been shown in family- and genus-level molecular phylogenies to be resolved outside of a large clade representing Rinorea s.str. Based on phylogenetic, morphological, and anatomical evidence, Bribria, a new genus, is segregated from Rinorea s.str. and described, with new combinations made for its three species: Bribria apiculata, Bribria crenata, and Bribria oraria. In addition, two new sections in Rinorea s.str. are described to accommodate the remaining Neotropical species: Rinorea sect. Rinorea and Rinorea sect. Pubiflora, which correspond to Group IIa Rinorea and Group IIc Pubiflora, respectively, in W. H. A. Hekking’s monograph of Neotropical Rinorea.     

Cytogenetic mapping of a major locus for resistance to Fusarium head blight and crown rot of wheat on <Emphasis Type="Italic">Thinopyrum elongatum</Emphasis> 7EL and its pyramiding with valuable genes from a <Emphasis Type="Italic">Th. ponticum</Emphasis> homoeologous arm onto bread wheat 7DL

Key message

A major locus for resistance to different Fusarium diseases was mapped to the most distal end of Th. elongatum 7EL and pyramided with Th. ponticum beneficial genes onto wheat 7DL.

Abstract

Perennial Triticeae species of the Thinopyrum genus are among the richest sources of valuable genes/QTL for wheat improvement. One notable and yet unexploited attribute is the exceptionally effective resistance to a major wheat disease worldwide, Fusarium head blight, associated with the long arm of Thinopyrum elongatum chromosome 7E (7EL). We targeted the transfer of the temporarily designated Fhb-7EL locus into bread wheat, pyramiding it with a Th. ponticum 7el₁L segment stably inserted into the 7DL arm of wheat line T4. Desirable genes/QTL mapped along the T4 7el₁L segment determine resistance to wheat rusts (Lr19, Sr25) and enhancement of yield-related traits. Mapping of the Fhb-7EL QTL, prerequisite for successful pyramiding, was established here on the basis of a bioassay with Fusarium graminearum of different 7EL-7el₁L bread wheat recombinant lines. These were obtained without resorting to any genetic pairing promotion, but relying on the close 7EL-7el₁L homoeology, resulting in 20% pairing frequency between the two arms. Fhb-7EL resided in the telomeric portion and resistant recombinants could be isolated with useful combinations of more proximally located 7el₁L genes/QTL. The transferred Fhb-7EL locus was shown to reduce disease severity and fungal biomass in grains of infected recombinants by over 95%. The same Fhb-7EL was, for the first time, proved to be effective also against F. culmorum and F. pseudograminearum, predominant agents of crown rot. Prebreeding lines possessing a suitable 7EL-7el₁L gene/QTL assembly showed very promising yield performance in preliminary field tests.

     

Drastic anthocyanin increase in response to <Emphasis Type="Italic">PAP1</Emphasis> overexpression in <Emphasis Type="Italic">fls1</Emphasis> knockout mutant confers enhanced osmotic stress tolerance in <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis>

Key message

pap1 - D/fls1ko double mutant plants that produce substantial amounts of anthocyanin show tolerance to abiotic stress.

Abstract

Anthocyanins are flavonoids that are abundant in various plants and have beneficial effects on both plants and humans. Many genes in flavonoid biosynthetic pathways have been identified, including those in the MYB-bHLH-WD40 (MBW) complex. The MYB gene Production of Anthocyanin Pigment 1 (PAP1) plays a particularly important role in anthocyanin accumulation. PAP1 expression in many plant systems strongly increases anthocyanin levels, resulting in a dark purple color in many plant organs. In this study, we generated double mutant plants that harbor fls1ko in the pap1-D background (i.e., pap1-D/fls1ko plants), to examine whether anthocyanins can be further enhanced by blocking flavonol biosynthesis under PAP1 overexpression. We also wanted to examine whether the increased anthocyanin levels contribute to defense against osmotic stresses. The pap1-D/fls1ko mutants accumulated higher anthocyanin levels than pap1-D plants in both control and sucrose-treated conditions. However, flavonoid biosynthesis genes were slightly down-regulated in the pap1-D/fls1ko seedlings as compared to their expression in pap1-D seedlings. We also report the performance of pap1-D/fls1ko seedlings in response to plant osmotic stresses.

     

Map-based cloning of the dominant genic male sterile <Emphasis Type="Italic">Ms</Emphasis>-<Emphasis Type="Italic">cd1</Emphasis> gene in cabbage (<Emphasis Type="Italic">Brassica oleracea</Emphasis>)

Key message

Using map-based cloning, we delimited the Ms - cd1 gene responsible for the male sterile phenotype in B. oleracea to an approximately 39-kb fragment. Expression analysis suggests that a new predicted gene, a homolog of the Arabidopsis SIED1 gene, is a potential candidate gene.

Abstract

A dominant genic male sterile (DGMS) mutant 79-399-3 in Brassica oleracea (B. oleracea) is controlled by a single gene named Ms-cd1, which was genetically mapped on chromosome C09. The derived DGMS lines of 79-399-3 have been successfully applied in hybrid cabbage breeding and commercial hybrid seed production of several B. oleracea cultivars in China. However, the Ms-cd1 gene responsible for the DGMS has not been identified, and the molecular basis of the DGMS is unclear, which then limits its widespread application in hybrid cabbage seed production. In the present study, a large BC₉ population with 12,269 individuals was developed for map-based cloning of the Ms-cd1 gene, and Ms-cd1 was mapped to a 39.4-kb DNA fragment between two InDel markers, InDel14 and InDel24. Four genes were identified in this region, including two annotated genes based on the available B. oleracea annotation database and two new predicted open reading frames (ORFs). Finally, a newly predicted ORF designated Bol357N3 was identified as the candidate of the Ms-cd1 gene. These results will be useful to reveal the molecular mechanism of the DGMS and develop more practical DGMS lines with stable male sterility for hybrid seed production in cabbage.

     

Combined drought and heat stress in <Emphasis Type="Italic">Camellia oleifera</Emphasis> cultivars: leaf characteristics,soluble sugar and protein contents,and Rubisco gene expression

Key message

Leaf relative water content, leaf area, leaf fresh weight, and SPAD chlorophyll meter readings along with Co - rbcL and Co - rbcS expression can be used for evaluating Camellia oleifera responses to combined drought and heat stress and subsequent recovery after rainfall events.

Abstract

Leaf characteristics, soluble protein and total soluble sugar contents as well as Rubisco-related gene expression in three cultivars of C. oleifera were measured during a combined drought and heat stress period and after subsequent rainfall events. Leaf relative water content (RWC) was significantly correlated with leaf area (LA), leaf fresh weight (FW), SPAD chlorophyll meter readings, and the levels of Co-rbcL and Co-rbcS expression. Results suggest that leaf RWC, LA, leaf FW, SPAD readings together with Co-rbcL and Co-rbcS expression can be used for evaluating responses of C. oleifera cultivars to combined drought and heat stress and subsequent recovery after rainfall events. Rubisco activase might be used for evaluating plant recovery after rainfall. This study identified cultivars differing in tolerance to the combined stress and recovery. Information derived from this study should be valuable for improving survivability and productivity of C. oleifera cultivars.