Down regulation of virulence factors of Pseudomonas aeruginosa by salicylic acid attenuates its virulence on Arabidopsis thaliana and Caenorhabditis elegans.

Salicylic acid (SA) is a phenolic metabolite produced by plants and is known to play an important role in several physiological processes, such as the induction of plant defense responses against pathogen attack. Here, using the Arabidopsis thaliana-Pseudomonas aeruginosa pathosystem, we provide evidence that SA acts directly on the pathogen, down regulating fitness and virulence factor production of the bacteria. Pseudomonas aeruginosa PA14 showed reduced attachment and biofilm formation on the roots of the Arabidopsis mutants lox2 and cpr5-2, which produce elevated amounts of SA, as well as on wild-type Arabidopsis plants primed with exogenous SA, a treatment known to enhance endogenous SA concentration. Salicylic acid at a concentration that did not inhibit PA14 growth was sufficient to significantly affect the ability of the bacteria to attach and form biofilm communities on abiotic surfaces. Furthermore, SA down regulated three known virulence factors of PA14: pyocyanin, protease, and elastase. Interestingly, P. aeruginosa produced more pyocyanin when infiltrated into leaves of the Arabidopsis transgenic line NahG, which accumulates less SA than wild-type plants. This finding suggests that endogenous SA plays a role in down regulating the synthesis and secretion of pyocyanin in vivo. To further test if SA directly affects the virulence of P. aeruginosa, we used the Caenorhabditis elegans-P. aeruginosa infection model. The addition of SA to P. aeruginosa lawns significantly diminished the bacterium's ability to kill the worms, without affecting the accumulation of bacteria inside the nematodes' guts, suggesting that SA negatively affects factors that influence the virulence of P. aeruginosa. We employed microarray technology to identify SA target genes. These analyses showed that SA treatment affected expression of 331 genes. It selectively repressed transcription of exoproteins and other virulence factors, while it had no effect on expression of housekeeping genes. Our results indicate that in addition to its role as a signal molecule in plant defense responses, SA works as an anti-infective compound by affecting the physiology of P. aeruginosa and ultimately attenuating its virulence.

Plant models for animal pathogenesis.

Several bacteria that are pathogenic to animals also infect plants. Mechanistic studies have proven that some human/animal pathogenic bacteria employ a similar subset of virulence determinants to elicit disease in animals, invertebrates and plants. Therefore, the results of plant infection studies are relevant to animal pathogenesis. This discovery has resulted in the development of convenient, cost-effective, and reliable plant infection models to study the molecular basis of infection by animal pathogens. Plant infection models provide a number of advantages in the study of animal pathogenesis. Using a plant model, mutations in animal pathogenic bacteria can easily be screened for putative virulence factors, a process which if done using existing animal infection models would be time-consuming and tedious. High-throughput screening of plants also provides the potential for unravelling the mechanisms by which plants resist animal pathogenic bacteria, and provides a means to discover novel therapeutic agents such as antibiotics and anti-infective compounds. In this review, we describe the developing technique of using plants as a model system to study Pseudomonas aeruginosa, Enterococcus faecalis and Staphylococcus aureus pathogenesis, and discuss ways to use this new technology against disease warfare and other types of bioterrorism.

Chemical characterization of basil (Ocimum basilicum L.) found in local accessions and used in traditional medicines in Iran.

Ocimum species are used in traditional Iranian medicine, as a culinary herb, and as a well-known source of flavoring principles. Horticultural characteristics, including quantitative and qualitative traits along with the chemical variation of phenolic acids, of 23 accessions of basil (Ocimum basilicum L.) from Iran were studied. Morphological studies of accessions showed a high level of variability in recorded traits. Quantification of phenolic acids was determined using high-performance liquid chromatography and showed drastic variations between accessions. Chemical studies revealed that rosmarinic acid is the predominant phenolic acid present in both flower and leaf tissues. Unusual basil accessions were identified that can serve as genetic sources of phenolic acids for crop improvement.

Permeabilization and in situ adsorption studies during growth and coumarin production in hairy root cultures of Cichorium intybus L.

Effect of addition of a permeabilizing agent dimethyl sulfoxide (DMSO) and a solid adsorbent, XAD -7, on growth and coumarin production in hairy root cultures of C. intybus was studied. Continuous permeabilization of the hairy root cultures of C. intybus with DMSO has been shown to be an effective strategy for enhanced release of coumarins while preserving the root viability. DMSO at 0.2% (v/v) level showed the maximum growth and coumarin production but was less as compared to control on day 28. Treatment of cells with increasing concentrations of DMSO (0.3 - 0.6 % v/v) to hairy root cultures of C. intybus, showed an inverse relationship with growth and coumarin production. Growth and production of coumarins increased with 1% media filtrate (MF) of cultures of Phytopthora parasitica var. nicotiana treatment. It was observed that treatment with DMSO (0.2% v/v) and 1% MF of P. parasitica showed the better growth and coumarin production with an increased release of coumarins as compared to the control and other treatments. It was observed that treatment of hairy root cultures with XAD-7 resulted in lesser growth and coumarin production as compared to control during the culture period. Addition of XAD-7 along with 1% MF of P. parasitica showed enhanced growth, coumarin production and increased adsorption as compared to control and lone XAD-7 treatment. Combined addition of DMSO/XAD-7 with fungal elicitor showed synergistic response in terms of biomass and coumarin production. Excretion of coumarins in both the cases was dependent on the presence of DMSO/XAD-7. These results showed that continuous permeabilization of hairy root cultures of C. intybus by using DMSO at 0.2% (v/v) level coupled with 1% MF of P. parasitica maintained viability of tissues and produced coumarins at higher level.

Biotechnological production of plant-based insecticides.

The demand for natural and nonpersistent insecticides is increasing day by day. Plant cell cultures could be an alternative to conventional methods of production of insecticides from field-grown plants. In vitro cultured plant cells produce a wide array of insecticides as a part of their secondary metabolism. Their ability to synthesize key enzymes and the manipulation of these could lead to the enhanced production of many insecticides of industrial importance. The development of a high-yielding hairy root culture system for thiophenes, nicotine, and phytoecdysones is of considerable interest. In this article, the current literature on various factors that influence the growth, production, and secretion of six insecticidal compounds, namely, pyrethrins, azadirachtin, thiophenes, nicotine, rotenoids, and phytoecdysones which have been prospects for the scale-up of cell cultures, genetic engineering to obtain transgenic plants, and metabolically engineered plants for increased production of bio-molecules, has been discussed. Environmental safety clearance and the future prospects of application of biomolecules for plant-derived insecticides are presented.

Enhancement of growth and coumarin production in hairy root cultures of witloof chicory (Cichorium intybus L.cv. Lucknow local) under the influence of fungal elicitors.

Studies were conducted to determine the possible production of coumarins in hairy root cultures of Cichorium intybus L.cv. Lucknow local under the influence of microbial agents. Pythium aphanidermatum and Phytopthora parasitica var. nicotiana were cultured and their mycelial and medium filtrate were used for the elicitation of coumarin production. The media filtrate of P. parasitica at 1.0% v/v added to the MS basal medium led to the maximum growth of hairy roots of C. intybus, which was 1.57-fold higher than the untreated control on the 28th day of culture, along with maximum esculin and esculetin yields which were 4.06- and 3.71-fold higher than the control on the 28th day. The yields of esculin and esculetin in hairy root cultures of chicory strongly correlated with growth. To check the effect of these fungal elicitors on endogenous polyamine metabolism, titers of total endogenous polyamines were analyzed. It was confirmed that the media filtrate of P. parasitica at 1.0% v/v concentration resulted in maximum accumulation of total endogenous polyamines, wherein endogenous spermine titers were found to be maximum as compared to endogenous spermidine and putrescine titers on the 28th day. Total endogenous spermine in the case of 1.0% MF v/v P. parasitica was 1.3-fold higher than that of the control on the 28th day of culture. Maximum growth index with greater length of primary root (17.61+/-0.18 cm) and greater number of secondary and tertiary roots was recorded for the medium filtrate of P. parasitica at 1.0% v/v concentration. This study also provided an insight into the morphological changes in terms of branching patterns, occurring in roots under the influence of these fungal elicitors.