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PubMed1995

The genome organization of banana bunchy top virus: analysis of six ssDNA components.

We have cloned, sequenced and analysed an additional five circular ssDNA components of banana bunchy top virus (BBTV) which we have called components 2, 3, 4, 5 and 6. These components were present in all BBTV infections tested. Four of these components (components 3, 4, 5 and 6) had one large open reading frame (ORF) in the virion sense located 3' of a stem-loop structure. Each ORF had a potential TATA box and one or two potential polyadenylation signals associated with it and each polyadenylation signal had an associated GC-rich region containing the trinucleotide sequence TTG. A number of ORFs were identified in component 2 but none of these had appropriately located potential TATA boxes and polyadenylation signals associated with them. None of the ORF amino acid sequences nor the full DNA sequences of any of the components had significant sequence identity with any known protein or nucleic acid sequences. However, the ORF of component 4 encoded a 30 residue hydrophobic domain which may indicate that this ORF encoded a transmembrane protein. Further, the ORFs of components 3 and 5 potentially encoded proteins of about 20 kDa, the size of the BBTV coat protein. There were two regions of sequence identity between the five components described here and the previously described component 1. Each component contained a conserved stem-loop structure and a nonanucleotide potential TATA box which was 5' of the large virion-sense ORF in five of the components. The stem-loop structures were incorporated in a common region (CR-SL) of 69 nucleotides which was 62% identical between components. All six BBTV components also contained a major common region (CR-M) which was located 5' of the CR-SL in each component, in the non-coding region and was 76% identical over 92 nucleotides. Each CR-M contained a near-complete 16 nucleotide direct repeat and a GC-box which was similar to the rightward promoter element found in wheat dwarf geminivirus. From these results, BBTV appears to belong to an undescribed plant virus group which could also include subterranean clover stunt virus, coconut foliar decay virus, faba bean necrotic yellows virus and milk vetch dwarf virus.

Amino Acid Sequence↗
PubMed1998

Sequences of ten circular ssDNA components associated with the milk vetch dwarf virus genome.

Milk vetch dwarf virus (MDV) is a member of the proposed genus Nanovirus, and its genome is composed of multiple, circular ssDNA components of about 1 kb. We have cloned and sequenced ten ssDNA components and designated them MDV-C1 to C10. Each DNA component contains one potential major open reading frame, and contains a putative stem-loop structure in the non-coding region. Notably, four components (C1, C2, C3 and C10) encode distinct replication-associated (Rep) proteins of 33 kDa, which show only limited (42-57%) amino acid identity. The six other components encode proteins with calculated molecular masses ranging from 12.7 to 19.7 kDa. Comparison of the sequences with those of other nanoviruses reveals that MDV is closely related to faba bean necrotic yellows virus (FBNYV) and subterranean clover stunt virus (SCSV). Six putative MDV genome products, including one Rep and five non-Rep proteins, show high (70.4-90.9%) amino acid identity to the corresponding six FBNYV proteins, whereas two other Rep proteins encoded by MDV-C2 and C3 are 82.3% and 73.0% identical to those encoded by SCSV-C2 and C6, respectively. These results indicate that MDV, FBNYV and SCSV have diverged from a common origin, which had multiple Rep components. In addition, the putative proteins encoded by MDV-C4 and its homologues contain a consensus retinoblastoma-binding motif, suggesting that they may be involved in controlling the host cell cycle.

Amino Acid Sequence↗
PubMed2002

Dual-colour imaging of membrane protein targeting directed by poa semilatent virus movement protein TGBp3 in plant and mammalian cells.

The movement function of poa semilatent hordeivirus (PSLV) is mediated by the triple gene block (TGB) proteins, of which two, TGBp2 and TGBp3, are membrane proteins. TGBp3 is localized to peripheral bodies in the vicinity of the plasma membrane and is able to re-direct TGBp2 from the endoplasmic reticulum (ER) to the peripheral bodies. For imaging of TGBp3-mediated protein targeting, PSLV TGBp3 tagged with a red fluorescent protein (DsRed) was used. Coexpression of DsRed-TGBp3 with GFP targeted to the ER lumen (ER-GFP) demonstrated that ER-GFP was contained in typical ER structures and peripheral bodies formed by TGBp3 protein, suggesting an ER origin for these bodies. In transient coexpression with viral membrane proteins tagged with GFP, DsRed-TGBp3 directed to the peripheral bodies the homologous TGBp2 protein and two unrelated membrane proteins, the 6 kDa movement protein of beet yellows closterovirus and the putative movement protein encoded by the genome component 4 of faba bean necrotic yellows nanovirus. However, coexpression of TGBp3 with GFP derivatives targeted to the ER membranes by artificial hydrophobic tail sequences suggested that targeting to the ER membranes per se was not sufficient for TGBp3-directed protein trafficking to peripheral bodies. TGBp3-induced targeting of TGBp2 also occurred in mammalian cells, indicating the universal nature of the protein trafficking signals and the cotargeting mechanism.

Animals↗
PubMed1992

Regulation of photosynthesis by end-product accumulation in leaves of plants storing starch, sucrose, and hexose sugars.

In the present study, leaves of different plant species were girdled by the hot wax collar method to prevent export of assimilates. Photosynthetic activity of girdled and control leaves was evaluated 3 to 7 days later by two methods: (a) carbon exchange rate (CER) of attached leaves was determined under ambient CO(2) concentrations using a closed gas system, and (b) maximum photosynthetic capacity (A(max)) was determined under 3% CO(2) with a leaf disc O(2) electrode. Starch, hexoses, and sucrose were determined enzymically. Typical starch storers like soybean (Glycine max L.) (up to 87.5 milligrams of starch per square decimeter in girdled leaves), cotton (Gossypium hirsutum L.), and cucumber (Cucumis sativus L.) responded to 7 days of girdling by increased (80-100%) stomatal resistance (r(s)) and decreased A(max) (>50%). On the other hand, spinach (Spinacia oleracea L.), a typical sucrose storer (up to 160 milligrams of sucrose per square decimeter in girdled leaves), showed only a slight reduction in CER and almost no change in A(max). Intermediate plants like tomato (Lycopersicon esculentum Mill.), sunflower (Helianthus annuus L.), broad bean (Vicia faba L.), bean (Phaseolus vulgaris L.), and pea (Pisum sativum L.), which upon girdling store both starch and sucrose, responded to the girdle by a considerable reduction in CER but only moderate inhibition of A(max), indicating that the observed reduction in CER was primarily a stomatal response. Both the wild-type tobacco (Nicotiana sylvestris) (which upon girdling stored starch and hexoses) and the starchless mutant (which stored only hexoses, up to 90 milligrams per square decimeter) showed 90 to 100% inhibition of CER and approximately 50% inhibition of A(max). In general, excised leaves (6 days) behaved like girdled leaves of the respective species, showing 50% reduction of A(max) in wild-type and starchless N. sylvestris but only slight decline of A(max) in spinach. The results of the present study demonstrate the possibility of the occurrence of end-product inhibition of photosynthesis in a large number of crop plants. The long-term inhibition of photosynthesis in girdled leaves is not confined to stomatal responses since the A(max) declined up to 50%. The inhibition of A(max) by girdling was strongest in starch storers, but starch itself cannot be directly responsible, because the starchless mutant of N. sylvestris was also strongly inhibited. Similarly, the inhibition cannot be attributed to hexose sugars either, because soybean, cotton, and cucumber are among the plants most strongly inhibited although they do not maintain a large hexose pool. Spinach, a sucrose storer, showed the least inhibition in both girdled and excised leaf systems, which indicates that sucrose is probably not directly responsible for the end-product inhibition of photosynthesis. The occurrence of strong end-product inhibition appears to be correlated with high acid-invertase activity in fully expanded leaves. The inhibition may be related to the nature of soluble sugar metabolism in the extrachloroplastic compartment and may be caused by a metabolite that has different rates of accumulation and turnover in sucrose storers and other plants.

Journal Article↗
PubMed1990

Two plasmids other than the nodulation plasmid are necessary for formation of nitrogen-fixing nodules by Rhizobium leguminosarum.

A system which allows direct selection for curing of plasmids in Gram-negative bacteria was used to generate derivatives of Rhizobium leguminosarum VF39 cured of each of six plasmids present in this strain. Phenotypes could be correlated with the absence of five of the six plasmids. The smallest plasmid, pRleVF39a, carries genes for the production of a melanin-like pigment as has been previously reported. Plasmid pRleVF39d carries nodulation and nitrogen fixation genes. Curing of the plasmids pRleVF39c and pRleVF39e gave rise to strains which formed Fix- nodules on peas, lentils, and faba beans. The nodules formed by the strains cured of pRleVF39c contained few, if any, bacteria. Analysis of washed cells by SDS-PAGE showed that this strain is defective in lipopolysaccharide (LPS) production; the defect could be complemented by introducing plasmids from several other R. leguminosarum strains, and by the R. leguminosarum biovar phaseoli LPS gene clones pCos126 and pDel27. The nodules formed by the strain cured of pRleVF39e had a reduced symbiotic zone, an enlarged senescence zone, and an abundance of starch granules. This strain grew at a much slower rate than the wild type, was unable to grow on minimal medium, and no longer produced melanin. These defects could be complemented by at least one other Rhizobium plasmid, pRle336e, a plasmid of strain 336 which is distinct from the nodulation plasmid (pRle336c) and the plasmid (pRle336d) which could complement the LPS defect associated with the loss of pRleVF39c. This demonstrates that genes necessary for symbiosis can be carried on at least three different plasmids in R. leguminosarum.

DNA Transposable Elements↗
PubMed1982

Quantitative study of nodulation competitiveness in Rhizobium strains.

We compared the nodulation competitiveness of three strains of Rhizobium leguminosarum by counting the number of nodules formed on faba bean plants after the application at sowing time of different concentrations of the strains to soils already containing Rhizobium strains of the same species. A relationship of type y = ax was found to exist between the ratio of the nodules formed by the applied inoculum strain to the nodules formed by the soil strains and the ratio of Rhizobium cells in the inoculum to the cells in the soil. This relationship was also confirmed in another competition experiment in which two R. meliloti strains of identical competitiveness were mixed in various proportions. The relationship can also be applied to the majority of results reported in the literature. Should it prove to be more widely applicable, it could be used to estimate the relative competitiveness of Rhizobium strains and thus predict the performance of an inoculum in a given soil.

Journal Article↗
PubMed2000

Clink, a nanovirus-encoded protein, binds both pRB and SKP1.

Clink, a 20-kDa protein of faba bean necrotic yellows virus, a single-stranded DNA plant virus, interacts with pRB family members and a SKP1 homologue from Medicago sativa. An LxCxE motif and an F-box of Clink mediate the interactions with the respective proteins. The capacity of Clink to bind pRB correlates with its ability to stimulate viral replication. Interaction of a single protein with the cell cycle regulator pRB and SKP1, a constituent of the ubiquitin-protein turnover pathway, appears to be a novel feature. Hence, Clink may represent a new class of viral cell cycle modulators.

Amino Acid Motifs↗
PubMed2025

Nodules-associated Klebsiella oxytoca complex: genomic insights into plant growth promotion and health risk assessment.

The swift emergence of antibiotic resistance genes (ARGs) across interconnected One Health compartments poses a significant global threat. Although plant growth-promoting (PGP) bacteria possess numerous attributes beneficial to host plants, many of these bacteria also harbor ARGs, necessitating a focused assessment of their negative implications. In this context, here we performed whole genome sequencing of 14 PGP endophytic strains isolated from root nodules of faba beans, belonging to three Klebsiella oxytoca species complex (KoSC): K. grimontii (n = 5), K. michiganensis (n = 5), and K. pasteurii (n = 4). We performed comparative genomics, molecular typing, and pangenome analyses on these strains. We identified significant diversity within the KoSC population, classifying the strains into five sequence types (STs), three of which are novel to this study (ST-542, ST-569, and ST-629). Phylogenomic analysis revealed that the bacterial strains clustered more closely by ST than by their source of isolation. Annotation of gene clusters indicated that all assembled genomes are enriched with genes involved in PGP activities, alongside a robust array of genes conferring tolerance to abiotic stresses. Importantly, our findings disclosed that the 14 assembled genomes harbored multiple ARGs, conferring resistance to various antibiotic classes, with 71% of the population classified as multidrug-resistant based on the in vitro antibiotic susceptibility assay. Furthermore, all genomes contained an array of virulence factors critical for survival, pathogenesis, biofilm formation, and root colonization. In conclusion, this study substantiates the hypothesis that certain PGP bacteria may serve as potential reservoirs of multidrug resistance, posing significant public health risks. Thus, the future advancement of bacteria-based biofertilizers should integrate environmental considerations and monitor their impact on antibiotic resistance dissemination in soil ecosystems.

Klebsiella oxytoca↗
PubMed2026

Microbiota-gut-muscle axis shapes fish muscle texture by regulating collagen synthesis.

BACKGROUND: Increasing studies have emphasized the communication network between the gut microbiome and host organs, revealing that such interactions significantly influence host physiological performances. However, whether a gut-muscle axis exists to regulate muscle quality in animal production is unknown. RESULTS: In two independent cohorts, the muscle hardness of tilapia subjected to a long-term faba bean diet exhibited significant inter-individual variation. RNA-Seq analyses of the high-hardness (H) and low-hardness (L) groups pointed to collagen-based extracellular matrix as a possible factor driving muscle hardness development. Transplantation of gut microbiota from the H donor resulted in enhanced collagen synthesis in gnotobiotic zebrafish. Muscular collagen deposition was featured with an increased abundance of gut Cetobacterium. Gnotobiotic models colonized with live C. somerae exhibited enhanced collagen synthesis. Integrated analyses of microbiome function, bacterial genome, and metabolic profiles identified acetate as a key effector of C. somerae. Acetate incubation upregulated collagen I expression in TGF-β-activated fibroblasts in an acetylation-dependent manner. Mechanistically, acetate promoted the acetylation of SMAD2/3, enhancing its nuclear transport and stability, which ultimately increased collagen expression. An acetate-supplemented feeding experiment corroborated these findings. CONCLUSION: The comprehensive results provided evidences that gut microbes regulated tilapia muscle texture through SMAD2/3 acetylation-driven collagen synthesis. This study expands our understanding of the multifaceted role of the gut-muscle axis in muscle physiology. Furthermore, our findings highlight that targeting gut microbiota and the downstream collagen synthesis pathway could be promising for manipulating muscle quality in animal production. Video Abstract.

Animals↗
PubMed1983

OER and RBE of high energy neutron beams for growth inhibition in Vicia faba.

The radiobiologic characteristics of 15 MeV neutrons produced by the d + T reaction at the TNO of Rijswijk and of neutrons produced by the d(50) + Be and p(75) + Be reactions at the cyclotron Cyclone of Louvain-la- Neuve were compared. Growth inhibition in Vicia faba bean roots was used as biologic system. An OER value of 1.5 +/- 0.1 ws obtained for the neutron beams compared. The RBE of 15 MeV, d(50) + Be and p(75) + Be neutrons was found equal to 3.4 +/- 0.2, 3.2 +/- 0.2 and 2.9 +/- 0.3, respectively, relative to gamma rays, for a total (n + gamma) absorbed dose of 0.6 Gy.

Fabaceae↗
PubMed1980

Methionine and cystine requirements of growing turkeys.

Sulfur amino acid requirements of starting, growing, finishing male Large White turkeys were investigated in three experiments using 3000 turkeys. Requirements were determined by least squares analyses of response curves obtained by supplementing methionine and cystine deficient basal diets with graded levels of DL-methionine and/or L-cystine. Experiments were conducted from 1 to 4, 8 to 12, and 16 to 20 weeks of age. Diets included starch, soybean meal, faba beans, and field peas. Requirements for total sulfur amino acids (TSAA) determined with diets marginally deficient in cystine were .95 to 1.01, .70 to .71, and .43 to .48% of the diet for starting (1 to 4 weeks), growing (8 to 12 weeks), finishing (16 to 20 weeks) turkeys, respectively, in different experiments. On the basis of dietary metabolizable energy, the respective TSAA requirement values (in %/therm) were .298 to .332, .205 to .221, and .128 to .134. Minimum methionine requirements (determined with excess dietary cystine) were .46, .30, and .19% of the diet for starting, growing, and finishing turkeys, respectively. Cystine replacement values, calculated by dividing the maximum usable levels of cystine (TSAA requirements minus methionine requirements) by TSAA requirements were 55, 58, and 57% of the TSAA requirement for starting, growing, and finishing turkeys, respectively.

Animal Nutritional Physiological Phenomena↗
PubMed1999

Chemical characterization of effective and ineffective strains of Rhizobium leguminosarum bv. viciae.

Chemical composition of lipopolysaccharide (LPS) isolated from an effective (97) and ineffective (87) strains of R. l. viciae has been determined. LPS preparations from the two strains contained: glucose, galactose, mannose, fucose, arabinose, heptose, glucosamine, galactosamine, quinovosamine, and 3-N-methyl-3,6-dideoxyhexose, as well as glucuronic, galacturonic and 3-deoxyoctulosonic acid. The following fatty acids were identified: 3-OH 14:0, 3-OH 15:0, 3-OH 16:0, 3-OH 18:0 and 27-OH 28:0. The ratio of 3-OH 14:0 to other major fatty acids in LPS 87 was higher that in LPS 97. SDS/PAGE profiles of LPS indicated that, in lipopolysaccharides, relative content of S form LPS I to that of lower molecular mass (LPS II) was much higher in the effective strain 97 than in 87. All types of polysaccharides exo-, capsular-, lipo, (EPS, CPS, LPS, respectively) examined possessed the ability to bind faba bean lectin. The degree of affinity of the host lectin to LPS 87 was half that to LPS 97. Fatty acids (FA) composition from bacteroids and peribacteroid membrane (PBM) was determined. Palmitic, stearic and hexadecenoic acids were common components found in both strains. There was a high content of unsaturated fatty acids in bacteroids as well as in PBM lipids. The unsaturation index in the PBM formed by strain 87 was lower than in the case of strain 97. Higher ratio of 16:0 to 18:1 fatty acids was characteristic for PMB of the ineffective strain.

Carbohydrates↗
PubMed2000

Rapid Construction of Full-length cDNA Clones of Tobacco Mosaic Virus and the Infectivity Assay of Its in Vitro Transcript.

The full-length cDNA of tobacco mosaic virus faba bean isolate (TMV-B) was amplified with RT-PCR in which T7 promoter sequence was added in the 5' terminus of its upstream primer, so that full-length cDNA was put directly under the control of a T7 promoter. The cDNA was cloned into plasmid pT7Blue and linear DNA was got by digesting the recombinant with KpnI or KpnI and PstI. Using these linear DNA and full-length PCR product as templates, respectively, their in vitro transcripts were inoculated to Nicotiana tabacum and Chenopodium amaranticolor. All of the transcripts had infectivity and produced symptoms similar to that of wild TMV. It was found that transcripts of the full-length PCR product had higher infectious efficiency than those of linear DNA.

Journal Article↗
PubMed2001

[Cloning and sequencing of 16S rRNA gene of Phytoplasma CWB1 strain associated with cactus witches' broom].

A 1.5 kb DNA fragment was amplified in DNA samples extracted from Opuntia salmiana porm showed witches'-broom symptom. The result indicates the existence of phytoplasma associated with this disease and this phytoplasma was designated as CWB1. The amplified fragment was ligated to pGEM-T easy vector and then transformed into JM109 strain of E. coli. Cloned DNA fragments were verified by PCR, restriction endonuclease (EcoRI) digestion and sequence analysis. The result revealed that the 16S rRNA gene of CWB1 consists of 1489 bp and shared 99.7% homology with Faba bean phyllody which belongs to phytoplasma 16S rII-C subgroup. So we can classify this strain into phytoplasma 16S rII-C subgroup.

Cloning, Molecular↗
PubMed1992

RBE and OER measurements on the p(66) + Be neutron beam at Faure, South Africa.

Results reported are for single dose exposures and refer to 60Co-gamma-irradiation. The RBE determined by V79 cell survival and based on the Do ratio was found to be 1.70 +/- 0.4 ranging from 1.5 to 1.8. In the case of the regeneration of mouse jejunal crypts the RBE was calculated at ten cell survival and was found to be 1.68. The maximum acute mouse skin reaction at a skin score of 2.0 was found to be 2.1 while the average skin reaction was 1.7. Growth retardation of Vicia faba bean roots measured at the level of 50% indicated an average RBE of 3.0 and a range of 2.7 to 3.7. The OER obtained for V79 cell survival was found to be 1.7 to 1.8. Comparison is made with the RBE and OER measurements for the neutron facilities at Clatterbridge, Fermilab and Louvain-la-Neuve which produce neutrons by the same nuclear reaction and whose physical specifications closely resemble those of the Faure neutrons. This comparison indicates that the Faure beam shows no unusual biological features and that its biological effectiveness is in line with that expected from its physical characteristics.

Animals↗
PubMed1982

In vitro translation and processing of a precursor form of favin, a lectin from Vicia faba.

Favin, the glucose- and mannose-binding lectin isolated from fava (Vicia faba) beans, consists of two polypeptide chains (alpha, Mr = 5,571; beta, Mr = 20,700). Translation of fava bean mRNA in vitro in a wheat germ-derived system yields a single favin polypeptide chain of Mr = 29,000. This molecule appears to consist of a hydrophobic 29-amino acid residue signal sequence at the NH2 terminus followed by the beta chain sequence; it also includes the alpha chain sequence. These results suggest that the alpha and beta chains arise by post-translational cleavage of a single precursor polypeptide: signal-beta chain-alpha chain. The signal peptide is similar in sequence to those seen in animal and prokaryotic systems, suggesting that translocation mechanisms are highly conserved. Translation of favin mRNA in the presence of dog pancreas microsomal membranes yields at least three polypeptides in addition to the presumed precursor chain. The largest of these molecules is translocated into the lumen of the membrane vesicles and glycosylated but its signal sequence remains intact. The two other species are translocated and glycosylated, but their signal sequences have been removed; they appear to differ from each other in that one begins with the beta chain sequence and the other begins one residue after the NH2-terminal threonine of the beta chain. These three variants could reflect normal features of the processing of the favin precursor but more likely result from aberrant processing of the plant protein by dog pancreas membranes.

Amino Acid Sequence↗
PubMed1981

[Genetic improvement in Vicia faba and favism. I. Distribution and levels of presumably hemolytic metabolites].

Covicine + vicine, L-DOPA-glucoside + L-DOPA and ascorbic acid were determined in different lines of Vicia faba beans throughout the biological cycle of the plant. As the seed matures the levels of convicine + vicine as well as of ascorbic acid decrease with seed maturation in all the lines examined. L-DOPA, which is lacking in cotyledons but present in the tegument, also decrease and is nearly undectable in some lines with white flowers.

Ascorbic Acid↗
PubMed1981

[Genetic improvement in Vicia and favism. II. Glycosidase activity].

Glycosidase activities with starch and PNP-glycosides as substrates can be detected in the cotyledons, leaves, pods and teguments of Vicia faba beans. The mainly hydrolize alpha- and beta-glucosides, alpha- and beta-galactosides as well as starch. The amylase activity arises at very early stages of seed maturation and then rapidly decreases, whereas glycosidases increase up to the 30% of the beans dry weight before decreasing. The beta-glucosidase might play a role in regulating the metabolism of possibly haemolitic glycosides (i.e. vicine, convicine and L-DOPA glucoside) thus explaining the presence of this enzyme.

Fabaceae↗