Host-induced silencing of Mi-msp-1 confers resistance to root-knot nematode Meloidogyne incognita in eggplant

RNA interference (RNAi)-based host-induced gene silencing (HIGS) is emerging as a novel, efficient and target-specific tool to combat phytonematode infection in crop plants. Mi-msp-1, an effector gene expressed in the subventral pharyngeal gland cells of Meloidogyne incognita plays an important role in the parasitic process. Mi-msp-1 effector is conserved in few of the species of root-knot nematodes (RKNs) and does not share considerable homology with the other phytonematodes, thereby making it a suitable target for HIGS with minimal off-target effects. Six putative eggplant transformants harbouring a single copy RNAi transgene of Mi-msp-1 was generated. Stable expression of the transgene was detected in T₁, T₂ and T₃ transgenic lines for which a detrimental effect on RKN penetration, development and reproduction was documented upon challenge infection with nematode juveniles. The post-parasitic nematode stages extracted from the transgenic plants showed long-term RNAi effect in terms of targeted downregulation of Mi-msp-1. These findings suggest that HIGS of Mi-msp-1 enhances nematode resistance in eggplant and protect the plant against RKN parasitism at very early stage.

     

Daily dosing of rifapentine cures tuberculosis in three months or less in the murine model

Background

Availability of an ultra-short-course drug regimen capable of curing patients with tuberculosis in 2 to 3 mo would significantly improve global control efforts. Because immediate prospects for novel treatment-shortening drugs remain uncertain, we examined whether better use of existing drugs could shorten the duration of treatment. Rifapentine is a long-lived rifamycin derivative currently recommended only in once-weekly continuation-phase regimens. Moxifloxacin is an 8-methoxyfluoroquinolone currently used in second-line regimens.

Methods and Findings

Using a well-established mouse model with a high bacterial burden and human-equivalent drug dosing, we compared the efficacy of rifapentine- and moxifloxacin-containing regimens with that of the standard daily short-course regimen based on rifampin, isoniazid, and pyrazinamide. Bactericidal activity was assessed by lung colony-forming unit counts, and sterilizing activity was assessed by the proportion of mice with culture-positive relapse after 2, 3, 4, and 6 mo of treatment. Here, we demonstrate that replacing rifampin with rifapentine and isoniazid with moxifloxacin dramatically increased the activity of the standard daily regimen. After just 2 mo of treatment, mice receiving rifapentine- and moxifloxacin-containing regimens were found to have negative lung cultures, while those given the standard regimen still harbored 3.17 log₁₀ colony-forming units in the lungs (p < 0.01). No relapse was observed after just 3 mo of treatment with daily and thrice-weekly administered rifapentine- and moxifloxacin-containing regimens, whereas the standard daily regimen required 6 mo to prevent relapse in all mice.

Conclusions

Rifapentine should no longer be viewed solely as a rifamycin for once-weekly administration. Our results suggest that treatment regimens based on daily and thrice-weekly administration of rifapentine and moxifloxacin may permit shortening the current 6 mo duration of treatment to 3 mo or less. Such regimens warrant urgent clinical investigation.     

Combating biotic stresses in plants by synthetic microbial communities: Principles,applications and challenges

Presently, agriculture worldwide is facing the major challenge of feeding the increasing population sustainably. The conventional practices have not only failed to meet the projected needs, but also led to tremendous environmental consequences. Hence, to ensure a food-secure and environmentally sound future, the major thrust is on sustainable alternatives. Due to challenges associated with conventional means of application of biocontrol agents in the management of biotic stresses in agroecosystems, significant transformations in this context are needed. The crucial role played by soil microbiome in efficiently and sustainably managing the agricultural production has unfolded a newer approach of rhizosphere engineering that shows immense promise in mitigating biotic stresses in an eco-friendly manner. The strategy of generating synthetic microbial communities (SynComs), by integrating omics approaches with traditional techniques of enumeration and in-depth analysis of plant–microbe interactions, is encouraging. The review discusses the significance of the rhizospheric microbiome in plant's fitness, and its manipulation for enhancing plant attributes. The focus of the review is to critically analyse the potential tools for the design and utilization of SynComs as a sustainable approach for rhizosphere engineering to ameliorate biotic stresses in plants. Furthermore, based on the synthesis of reports in the area, we have put forth possible solutions to some of the critical issues that impair the large-scale application of SynComs in agriculture.     

Effect of water stress on proline content and transcript levels in Lathyrus sativus

Response of Lathyrus sativus plants to water stress showed that ABA responsive genes such as PLE 25, TAS 14 and RAB 17 are synthesized constitutively, the levels of which decline gradually with increase in water stress or ABA levels. Proline accumulation was highest in leaves (65-fold) followed by stem (56-fold), root (38-fold) and marginal increase in etiolated seedlings. Proline increase was also observed in plant parts not exposed to light.     

Inactivation of Cyanobacterial Nitrogenase After Exposure to Ultraviolet-B Radiation

Exposure of the N₂-fixing cyanobacterium Anabaena BT2 to ultraviolet-B radiation (2.5 W m⁻²) for 30 min resulted in complete loss of nitrogenase activity but 100% cell killing occurred only after a 90-min exposure. Inactivation of nitrogenase activity was not specific to Anabaena BT2; other species also showed a similar effect. The time required for 100% killing and inactivation of nitrogenase activity differed in various species, and this difference may be ascribed to the presence of different levels of UV-B protection mechanisms in individual species. Inhibition of nitrogenase activity was immediate, since exposure of cultures to UV-B for as little as 5 min elicited some inhibition of activity. The activity of UV-B-inhibited nitrogenase did not recover upon transfer of exposed cells to fluorescent light, suggesting that the inhibition may be due to specific inactivation of the enzyme. By employment of inhibitors of protein synthesis and PS-II activity, it was demonstrated that restoration of nitrogenase activity in a UV-B-treated culture occurred by fresh synthesis of nitrogenase polypeptide. Our findings suggest that estimation of nitrogenase activity in diazotrophic species may be used as a marker enzyme for assessing the impact of UV-B radiation. Received: 13 June 2002 / Accepted: 22 July 2002     

Do the different parental 'heteromes' cause genomic shock in newly formed allopolyploids?

Allopolyploidy, the joining of two parental genomes in a polyploid organism with diploid meiosis, is an important mechanism of reticulate evolution. While many successful long-established allopolyploids are known, those formed recently undergo an instability phase whose basis is now being characterized. We describe observations made with the Arabidopsis system that include phenotypic instability, gene silencing and activation, and methylation changes. We present a model based on the epigenetic destabilization of genomic repeats, which in the parents are heterochromatinized and suppressed. We hypothesize that loss of epigenetic suppression of these sequences, here defined as the heterome, results in genomic instability including silencing of single-copy genes.     

Phlorizin recognition in a C-terminal fragment of SGLT1 studied by tryptophan scanning and affinity labeling

SGLT1 as a sodium/glucose cotransporter is strongly inhibited by phlorizin, a phloretin 2'-glucoside that has strong interactions with the C-terminal loop 13. We have examined phlorizin recognition by the protein by site-directed single Trp scanning mutagenesis experiments. Six mutants (Q581W, E591W, R601W, D611W, E621W, and L630W) of truncated loop 13 (amino acids 564-638) were expressed in Escherichia coli and purified to homogeneity. Changes in Trp quenching and positions of the emission maxima were determined after addition of phlorizin. D611W displayed the largest quenching of 80%, followed by R601W (67%). It also exhibited the maximum red shift in Trp fluorescence ( approximately 14 nm), indicating an exposure of this region to a more hydrophilic environment. Titration experiments performed for each mutant showed a similar affinity for all mutants, except for D611W, which exhibited a significantly lower affinity (Kd approximately 54 microm). Also the maximum change in the collisional quenching constant by acrylamide was noted for D611W (KSV = 11 m-1 in the absence of phlorizin and 55 m-1 in its presence). Similar results were obtained with phloretin. CD measurements and computer modeling revealed that D611W is positioned in a random coil situated between two alpha-helical segments. By combining gel electrophoresis, enzymatic fragmentation, and matrix-assisted laser desorption ionization mass spectrometry, we also analyzed truncated loop 13 photolabeled with 3-azidophlorizin. The attachment site of the ortho-position of aromatic ring B of phlorizin was localized to Arg-602. Taken together, these data indicate that phlorizin binding elicits changes in conformation leading to a less ordered state of loop 13. Modeling suggests an interaction of the 4- and 6-OH groups of aromatic ring A of phlorizin with the region between amino acids 606 and 611 and an interaction of ring B at or around amino acid 602. Phloretin seems to interact with the same region of the protein.