Rapid and accurate species and genomic species identification and exhaustive population diversity assessment of Agrobacterium spp. using recA-based PCR

Agrobacteria are common soil bacteria that interact with plants as commensals, plant growth promoting rhizobacteria or alternatively as pathogens. Indigenous agrobacterial populations are composites, generally with several species and/or genomic species and several strains per species. We thus developed a recA-based PCR approach to accurately identify and specifically detect agrobacteria at various taxonomic levels. Specific primers were designed for all species and/or genomic species of Agrobacterium presently known, including 11 genomic species of the Agrobacterium tumefaciens complex (G1-G9, G13 and G14, among which only G2, G4, G8 and G14 still received a Latin epithet: pusense, radiobacter, fabrum and nepotum, respectively), A. larrymoorei, A. rubi, R. skierniewicense, A. sp. 1650, and A. vitis, and for the close relative Allorhizobium undicola. Specific primers were also designed for superior taxa, Agrobacterium spp. and Rhizobiaceace. Primer specificities were assessed with target and non-target pure culture DNAs as well as with DNAs extracted from composite agrobacterial communities. In addition, we showed that the amplicon cloning-sequencing approach used with Agrobacterium-specific or Rhizobiaceae-specific primers is a way to assess the agrobacterial diversity of an indigenous agrobacterial population. Hence, the agrobacterium-specific primers designed in the present study enabled the first accurate and rapid identification of all species and/or genomic species of Agrobacterium, as well as their direct detection in environmental samples.     

RNAi-Mediated Silencing of the Flavanone 3-Hydroxylase Gene and Its Effect on Flavonoid Biosynthesis in Strawberry Fruit

Anthocyanin biosynthesis requires the activities of several enzymes in vivo. Flavanone 3-hydroxylase (F3H) converts flavanone into dihydroflavanol at an early step in the anthocyanin biosynthesis pathway. In this study we constructed an RNAi gene-silencing vector that encodes a hairpin F3H RNA. Agrobacterium strain GV3101 harboring the F3H RNAi vector was injected into strawberry fruits which were still attached to the plants 14 days after pollination. The phenotype was observed 10 days postinjection, and fruits were tested by RT-PCR and northern blot assays. The results showed that the F3H gene was downregulated by approximately 70 % in the agroinfiltrated fruits compared with the control. HPLC–MS analysis showed that anthocyanin content was greatly reduced, flavonol was also decreased, and the levels of p-coumaroyl glucoside and p-coumaroyl-1-acetate were markedly increased. We conclude that the precursors were shunted to the phenylpropanoid pathway, and that F3H is one of the key enzymes required for the biosynthesis of flavonoids in strawberry fruit. According to our results, reducing gene function via RNA interference is a rapid, simple, and effective way to identify gene function in strawberry fruit.     

Genetic analysis of barrel field size in the first somatosensory area (SI) in inbred and recombinant inbred strains of mice

We measured the combined area of posterior medial barrel subfield (PMBSF) and anterior lateral barrel subfield (ALBSF) areas in four common inbred strains (C3H/HeJ, A?/J, C57BL?/6J, DBA/2J), B6D2F1, and ten recombinant inbred (RI) strains generated from C57BL/6J and DBA/2J progenitors (BXD) as an initial attempt to examine the genetic influences underlying natural variation in barrel field size in adult mice. These two subfields are associated with the representation of the whisker pad and sinus hairs on the contralateral face. Using cytochrome oxidase labeling to visualize the barrel field, we measured the size of the combined subfields in each mouse strain. We also measured body weight and brain weight in each strain. We report that DBA/2J mice have a larger combined PMBSF/ALBSF area (6.15?±?0.10?mm²,?n?=?7) than C57BL?/6J (5.48?±?0.13?mm²,?n?=?10), C3H/HeJ (5.37?±?0.16?mm²,?n?=?10), and A/J mice (5.04?±?0.09?mm²,?n?=?15), despite the fact that DBA/2J mice have smaller average brain and body sizes. This finding may reflect dissociation between systems that control brain size with those that regulate barrel field area. In addition, BXD strains (average n?=?4) and parental strains showed considerable and continuous variation in PMBSF/ALBSF area, suggesting that this trait is polygenic. Furthermore, brain, body, and cortex weights have heritable differences between inbred strains and among BXD strains. PMBSF/ALBSF pattern appears similar among inbred and BXD strains, suggesting that somatosensory patterning reflects a common plan of organization. This data is an important first step in the quantitative genetic analysis of the parcellation of neocortex into diverse cytoarchitectonic zones that vary widely within and between species, and in identifying the genetic factors underlying barrel field size using quantitative trait locus (QTL) analyses.     

MAIGO2 is involved in abscisic acid‐mediated response to abiotic stresses and Golgi‐to‐ER retrograde transport

The central role of multisubunit tethering complexes in intracellular trafficking has been established in yeast and mammalian systems. However, little is known about their roles in the stress responses and the early secretory pathway in Arabidopsis. In this study, Maigo2 (MAG2), which is equivalent to the yeast Tip20p and mammalian Rad50‐interacting protein, is found to be required for the responses to salt stress, osmotic stress and abscisic acid in seed germination and vegetative growth, and MAG2‐like (MAG2L) is partially redundant with MAG2 in response to environmental stresses. MAG2 strongly interacts with the central region of ZW10, and both proteins are important as plant endoplasmic reticulum (ER)‐stress regulators. ER morphology and vacuolar protein trafficking are unaffected in the mag2, mag2l and zw10 mutants, and the secretory marker to the apoplast is correctly transported in mag2 plants, which indicate that MAG2 functions as a complex with ZW10, and is potentially involved in Golgi‐to‐ER retrograde trafficking. Therefore, a new role for ER–Golgi membrane trafficking in abiotic‐stress and ER‐stress responses is discovered.     

Competitive interaction between the exotic plant Rhus typhina L. and the native tree Quercus acutissima Carr. in Northern China under different soil N:P ratios

Background and aims

Anthropogenic nitrogen (N) and phosphorus (P) input has changed the relative importance of nutrient elements. This study aimed to examine the effects of different nutrient conditions on the interaction between exotic and native plants.

Methods

We conducted a greenhouse experiment with a native species Quercus acutissima Carr. and an exotic species Rhus typhina L. grown in monocultures or mixtures, under three N:P ratios (5, 15 and 45 corresponding to N-limited, basic N and P supply and P-limited conditions, respectively). After 12 weeks of treatment, traits related to biomass allocation, leaf physiology and nutrient absorption were determined.

Results

R. typhina was dominant under competition, with a high capacity for carbon assimilation and nutrient absorption, and the dominance was unaffected by increasing N:P ratios. R. typhina invested more photosynthate in leaves and more nutrients in the photosynthetic apparatus, enabling high biomass production. Q. acutissima invested more photosynthate in roots and more nutrients in leaf persistence at the expense of reduced carbon assimilation capacity.

Conclusions

Different trade-offs in biomass and nutrient allocation of the two species is an important reason for their distinct performances under competition and helps R. typhina to maintain dominance under different nutrient conditions.     

Variation in cadmium accumulation and translocation among peanut cultivars as affected by iron deficiency

Purpose

The current study aimed to test the hypothesis that the variations in shoot Cd accumulation among peanut cultivars was ascribed to the difference in capacity of competition with Fe transport, xylem loading and transpiration.

Methods

A hydroponics experiment was conducted to determine the plant biomass, gas exchange, and Cd accumulation in Fe-sufficient or -deficient plants of 12 peanut cultivars, at low Cd level (0.2 μM CdCl₂).

Results

Peanut varied among cultivars in morpho-physiological response to Cd stress as well as Cd accumulation, translocation and distribution. Qishan 208 and Xvhua 13 showed a higher capacity for accumulating Cd in their shoots. Fe deficiency increased the concentration and amount of Cd in plant organs, but decreased TF _{root to shoot} and TF _{root to stem}, while TF _{stem to leaf} remained unaffected. Fe deficiency-induced increase rates of Cd concentration and total Cd amount in roots and leaves were negatively correlated with the values in Fe-sufficient plants. Transpiration rate was positively correlated with leaf Cd concentration, TF _{root to shoot}, TF _{root to stem} and TF _{stem to leaf}.

Conclusions

The difference in shoot Cd concentration among peanut cultivars was mainly ascribed to the difference in Fe transport system, xylem loading capacity and transpiration.     

Converting paddy fields to Lei bamboo (Phyllostachys praecox) stands affected soil nutrient concentrations, labile organic carbon pools, and organic carbon chemical compositions

Background and aims

Land-use change often markedly alters soil carbon (C) and nitrogen (N) pool sizes with implications for climate change and soil sustainability. The objective of this research was to study the effect of converting paddy fields to Lei bamboo (Phyllostachys praecox) stands on soil C and N and other nutrient pools as well as the chemical structure of soil organic C (SOC) in the soil profile.

Methods

Soils (Anthrosols) from four adjacent paddy field–bamboo forest pairs with a known land-use history were sampled from Lin’an County, Zhejiang Province. Soil water soluble organic C (WSOC), hot water soluble organic C (HWSOC), microbial biomass C (MBC), readily oxidizable C (ROC), water soluble organic N (WSON), and other soil chemical and physical properties were determined. Soil organic C functional group compositions were determined by ¹³C-nuclear magnetic resonance (NMR).

Results

Concentrations of soil available P, available K, and different N forms increased (P?<?0.05) by the land-use conversion. Higher concentrations of SOC and total N (TN) were observed in the subsoil (20–40 and 40–60 cm soil layers) but not in the topsoil (0–20 cm layer) in the bamboo stands than in the paddy fields. The storage of SOC and TN in the entire soil profile (0–60 cm) increased by 56.7 and 70.7 %, respectively, after the land-use change. The increases in the SOC stock of the three soil layers were 11.0, 14.3, and 9.5 Mg C ha^?1, respectively. The conversion decreased WSOC concentrations in the subsoil but increased the ROC concentration in the topsoil. Solid-state NMR spectroscopy of soil samples showed that the conversion increased (P?<?0.05) the O-alkyl C content while decreased the aromatic C content, alkyl C to O-alkyl C ratio (A/O-A), and aromaticity of SOC.

Conclusions

Conversion of paddy fields to bamboo stands increased soil nutrient availability, and SOC and TN stocks. Effects of land-use change on C pools and C chemistry of SOC varied among different soil layers in the profile. The impact of the land-use conversion on soil organic C pools was not restricted to the topsoil, but changes in the subsoil were equally large and should be accounted for.     

Foot-and-Mouth Disease Virus Modulates Cellular Vimentin for Virus Survival

Foot-and-mouth disease virus (FMDV), the causative agent of foot-and-mouth disease, is an Aphthovirus within the Picornaviridae family. During infection with FMDV, several host cell membrane rearrangements occur to form sites of viral replication. FMDV protein 2C is part of the replication complex and thought to have multiple roles during virus replication. To better understand the role of 2C in the process of virus replication, we have been using a yeast two-hybrid approach to identify host proteins that interact with 2C. We recently reported that cellular Beclin1 is a natural ligand of 2C and that it is involved in the autophagy pathway, which was shown to be important for FMDV replication. Here, we report that cellular vimentin is also a specific host binding partner for 2C. The 2C-vimentin interaction was further confirmed by coimmunoprecipitation and immunofluorescence staining to occur in FMDV-infected cells. It was shown that upon infection a vimentin structure forms around 2C and that this structure is later resolved or disappears. Interestingly, overexpression of vimentin had no effect on virus replication; however, overexpression of a truncated dominant-negative form of vimentin resulted in a significant decrease in viral yield. Acrylamide, which causes disruption of vimentin filaments, also inhibited viral yield. Alanine scanning mutagenesis was used to map the specific amino acid residues in 2C critical for vimentin binding. Using reverse genetics, we identified 2C residues that are necessary for virus growth, suggesting that the interaction between FMDV 2C and cellular vimentin is essential for virus replication.