Biotechnological advances in pomegranate (Punica granatum L.)

Pomegranate (Punica granatum L.) is known for its nutritional, medicinal, and ornamental importance. It is conventionally propagated by hardwood and softwood cuttings, but about 1 yr is needed before the rooted cuttings can be transplanted to the field. Propagation by seed is undesirable as populations are heterozygous and seed propagation leads to wide variations in tree and fruit characteristics. Several studies have been conducted on in vitro culture of pomegranate, and protocols have been developed for plant regeneration through organogenesis and embryogenesis from various types of explants. Tissue culture has enabled mass propagation of superior genotypes of both wild and cultivated varieties. However, successful application of tissue culture systems for genetic engineering of pomegranate is still limited. Molecular markers are essential for identification and discrimination of genotypes for genetic conservation, crop improvement, breeding programs, and commercialization of superior genotypes. These techniques may also be applicable to rapid identification and indexing of disease-free planting material. This review focuses on the biotechnological approaches that are being used for pomegranate improvement.     

Plant regeneration in <Emphasis Type="Italic">Robinia pseudoacacia</Emphasis> from cell suspension cultures

A method for plant regeneration in Robinia pseudoacacia L. from cell suspension culture was established. Non regenerative friable callus from hypocotyls and cotyledon explants from in vitro raised seedling induced on solid Murashige and Skoog (MS) medium supplemented with 0.05 mg dm⁻³ 2,4-dichlorophenoxyacetic acid (2,4-D) was used for initiation of cell suspension cultures on same MS medium but without agar. Single cells were isolated after 3 d and the optimum cell density was 1–3 × 10⁴ cells per cm³ of the liquid MS medium. Plating efficiency was 29.6 % and callus formed within 4 weeks was subcultured and transferred to solid MS medium supplemented with 0.6 mg dm⁻³ benzyladenine (BA) along with 0.05 mg dm⁻³ α-naphthalene-1-acetic acid (NAA) for the induction of adventitious bud primordia. The shoots developed were isolated and re-cultured on MS medium containing 0.6 mg dm⁻³ BA. These microshoots after dipping in 1–2 cm³ of 10 mg dm⁻³ indole-3-butyric acid (IBA) for 24 h in dark were cultured on half strength solid MS medium supplemented with 0.05 % charcoal and showed 80–82 % rooting within 4 weeks.     

Why they eat,what they eat: patterns of wild edible plants consumption in a tribal area of Western Himalaya

Background

From time immemorial, wild plants have been used for edible purposes. They still continue to be a major source of nutrition for tribal people. However, unfortunately, their use is now declining. This has implications in food security, narrowing genetic base, and future leads. The present study was, therefore, carried out in Chhota Bhangal region of Western Himalaya to analyze uses of wild edible plants (WEP) and the motivations behind their use or abandonment.

Methods

Field surveys were conducted to the study area from January 2016 to March 2017. Household surveys, group discussions, free listing, and structured questionnaires were used to elicit information on WEP. WEP use was categorized into six categories (vegetables, fruits, chutney, flavoring food, raw food, and local brew). Trends of use (continuing, decreasing, increasing, and not used) and motivations (environmental, economic, sociocultural, agriculture and land use practices, and human-wildlife conflict) behind their use were analyzed.

Results

Fifty plant species were used by the local people for edible purposes under six WEP categories. Mean and median of WEP used per respondent was 22.3 and 21, respectively. Highest number of these were used as vegetable (mean 8.9) while lowest were used as brew (mean 0.4). Out of the 50 WEP used, 20 were prioritized for motivation analyses. Though plant use is still maintained in the area, changes are evident. Almost 50% of the respondents revealed that they still continue the use of WEP while 36% reported trends of declining use as compared to 5–10 years back. Close to 10% respondents have stopped consuming WEP now and ~?3% reported an increase in the use of WEP. Among the WEP categories, use of chutney showed an increasing trend. Sociocultural motivations were found to play a prime role, both, in limiting and promoting WEP use. Taste and aroma were the major sociocultural reasons behind using WEP while modernization and changing lifestyle were the main reasons behind declining use of WEP.

Conclusions

The study concludes that though use of WEP is still maintained in the area, changes in consumption trends are evident. Sociocultural motivations guided use of WEP in the area.

     

Using incentives to recruit physicians into behavioral trials: lessons learned from four studies

Objective

To describe lessons learned from the use of different strategies for recruiting physicians responsible for trauma triage, we summarize recruitment data from four behavioral trials run in the United States between 2010 and 2016.

Results

We ran a series of behavioral trials with the primary objective of understanding the influence of heuristics on physician decision making in trauma triage. Three studies were observational; one tested an intervention. The trials used different methods of recruitment (in-person vs. email), timing of the honorarium (pre-paid vs. conditional on completion), type of honorarium [a $100 gift card (monetary reward) vs. an iPad mini 2 (material incentive)], and study tasks (a vignette-based questionnaire, virtual simulation, and intervention plus virtual simulation). We recruited 989 physicians, asking each to complete a questionnaire or virtual simulation online. Recruitment and response rates were 80% in the study where we approached physicians in person, used a pre-paid material incentive, and required that they complete both an intervention plus a virtual simulation. They were 56% when we recruited physicians via email, used a monetary incentive conditional on completion of the task, and required that they complete a vignette-based questionnaire. Trial registration clinicaltrials.gov; NCT02857348

     

Cytoplasmic Translocation of Polypyrimidine Tract-Binding Protein and Its Binding to Viral RNA during Japanese Encephalitis Virus Infection Inhibits Virus Replication

Japanese encephalitis virus (JEV) has a single-stranded, positive-sense RNA genome containing a single open reading frame flanked by the 5′- and 3′-non-coding regions (NCRs). The virus genome replicates via a negative-sense RNA intermediate. The NCRs and their complementary sequences in the negative-sense RNA are the sites for assembly of the RNA replicase complex thereby regulating the RNA synthesis and virus replication. In this study, we show that the 55-kDa polypyrimidine tract-binding protein (PTB) interacts in vitro with both the 5′-NCR of the positive-sense genomic RNA - 5NCR(+), and its complementary sequence in the negative-sense replication intermediate RNA - 3NCR(-). The interaction of viral RNA with PTB was validated in infected cells by JEV RNA co-immunoprecipitation and JEV RNA-PTB colocalization experiments. Interestingly, we observed phosphorylation-coupled translocation of nuclear PTB to cytoplasmic foci that co-localized with JEV RNA early during JEV infection. Our studies employing the PTB silencing and over-expression in cultured cells established an inhibitory role of PTB in JEV replication. Using RNA-protein binding assay we show that PTB competitively inhibits association of JEV 3NCR(-) RNA with viral RNA-dependent RNA polymerase (NS5 protein), an event required for the synthesis of the plus-sense genomic RNA. cAMP is known to promote the Protein kinase A (PKA)-mediated PTB phosphorylation. We show that cells treated with a cAMP analogue had an enhanced level of phosphorylated PTB in the cytoplasm and a significantly suppressed JEV replication. Data presented here show a novel, cAMP-induced, PTB-mediated, innate host response that could effectively suppress JEV replication in mammalian cells.     

TGF-beta potentiates airway smooth muscle responsiveness to bradykinin

The molecular mechanisms by which bradykinin induces excessive airway obstruction in asthmatics remain unknown. Transforming growth factor (TGF)-beta has been involved in regulating airway inflammation and remodeling in asthma, although it is unknown whether TGF-beta can modulate bradykinin-associated bronchial hyperresponsiveness. To test whether TGF-beta directly modulates airway smooth muscle (ASM) responsiveness to bradykinin, isolated murine tracheal rings were used to assess whether TGF-beta alters ASM contractile responsiveness to bradykinin. Interestingly, we found TGF-beta-treated murine rings (12.5 ng/ml, 18 h) exhibited increased expression of bradykinin 2 (B(2)) receptors and became hyperreactive to bradykinin, as shown by increases in maximal contractile responses and receptor distribution. We investigated the effect of TGF-beta on bradykinin-evoked calcium signals since calcium is a key molecule regulating ASM excitation-contraction coupling. We reported that TGF-beta, in a dose- (0.5-10 ng/ml) and time- (2-24 h) dependent manner, increased mRNA and protein expression of the B(2) receptor in cultured human ASM cells. Maximal B(2) receptor protein expression that colocalized with CD44, a marker of membrane cell surface, occurred after 18 h of TGF-beta treatment and was further confirmed using fluorescence microscopy. TGF-beta (2.5 ng/ml, 18 h) also increased bradykinin-induced intracellular calcium mobilization in fura-2-loaded ASM cells. TGF-beta-mediated enhancement of calcium mobilization was not attenuated with indomethacin, a cyclooxygenase inhibitor. These data demonstrate for the first time that TGF-beta may play a role in mediating airway hyperresponsiveness to bradykinin seen in asthmatics by enhancing ASM contractile responsiveness to bradykinin, possibly as a result of increased B(2) receptor expression and signaling.