Control of pyrimidine nucleotide formation in <Emphasis Type=Italic>Pseudomonas</Emphasis><Emphasis Type=Italic>fulva</Emphasis>

Control of pyrimidine formation was examined in Pseudomonas fulva ATCC 31418. Pyrimidine supplementation lowered pyrimidine biosynthetic pathway enzyme activities in cells grown on glucose or succinate as a carbon source indicating possible repression of enzyme synthesis. Pyrimidine limitation experiments were conducted using an orotidine 5′-monophosphate decarboxylase mutant strain isolated in this study. Compared to uracil-supplemented, glucose-grown mutant cells, pyrimidine limitation of this strain caused aspartate transcarbamoylase, dihydroorotase, dihydroorotate dehydrogenase and orotate phosphoribosyltransferase activities to increase about 6-, 13-, 3-, 15-fold, respectively, which confirmed regulation of enzyme synthesis by pyrimidines. At the level of enzyme activity, transcarbamoylase activity in Ps. fulva was strongly inhibited by pyrophosphate, CTP, GTP and GDP under saturating substrate concentrations.     

Quorum sensing regulation in <Emphasis Type=Italic>Pseudomonas</Emphasis>

Expression of many bacterial genes is regulated in a cell density-dependent manner via small signal molecules known as autoinducers; this type of regulation is termed quorum sensing (QS). The QS systems that employ N-acyl-homoserine lactones (HSLs) are best un derstood in Gram-negative bacteria. QS regulates expression of various genes, including the genes responsible for the production of virulence factors, synthesis of exoenzymes and antibiotics, antagonistic properties of bacteria, etc. The QS systems of the genus Pseudomonas are linked to other global regulatory networks of the cell, and their functions are controlled by numerous additional regulatory factors. Such regulators and the QS systems together form an intricate multifactorial cascade regulatory network. The review considers the QS systems of several Pseudomonas species, their interaction with other regulatory systems, and their roles in the regulation of cell processes.     

<Emphasis Type=Italic>Pseudomonas</Emphasis> bacteria and phosphorous fertilization,affecting wheat (<Emphasis Type=Italic>Triticum</Emphasis><Emphasis Type=Italic>aestivum</Emphasis> L.) yield and P uptake under greenhouse and field conditions

We have recently indicated the plant growth promoting activities of Pseudomonas sp. as well as their alleviating effects on some soil stressors such as salinity. This is because in recent years, biological fertilizers have received special attention by scientists in sustainable agriculture. Accordingly, it is pertinent to specify the beneficiary level of such soil bacteria on plant growth including phosphorous (P) uptake. Hence, the objectives were to determine: (1) the plant growth promoting effects of the tested Pseudomonas sp., and (2) its combined effects with different P fertilization rates on the nutrient uptake (N, P, and K) and yield of wheat (Triticum aestivum L.) under greenhouse and field conditions. The experiments were factorially arranged on the basis of a completely randomized block design with three replicates and were conducted at the Research Farm of Agriculture and Natural Resources Research Center of Khorasan, Mashhad, Iran. P was fertilized at three levels including 0, 25 and 50 kg/ha P₂O₅. Pseudomonas sp. including Pseudomonas fluorescens 153, P. fluorescens 169, P. putida 4, and P. putida 108 were tested. Activities such as production of ACC deaminase and IAA-like products, as well as P solubilization were among the most important activities of the tested Pseudomonas sp. Such bacterial effects greatly enhanced wheat growth and yield under greenhouse and field conditions. The results also showed that the effects of Pseudomonas sp. on wheat nutrient uptake and the effects of bacteria as well as P fertilization on wheat yield were significant. P. putida 108 was the most effective strain enhancing wheat P uptake and grain yield under greenhouse (96 and 58%) and field (80 and 37%) conditions, respectively. Hence, although Pseudomonas sp. could be a suitable replacement for high P fertilization, however, the optimum wheat yield resulted when the bioinoculants are combined with 50% (25 kg/ha P₂O₅) P fertilization. This finding has great agricultural and environmental implications.     

<Emphasis Type=Italic>Agrobacterium</Emphasis>-mediated <Emphasis Type=Italic>in planta</Emphasis> transformation of maize germ cells

A transfer DNA (T-DNA) carrying the marker gene nptII was detected in the genomes of diploid and haploid maize plants obtained after the treatment of pistil filaments with a suspension of Agrobacterium during artificial pollination. PCR analysis of total DNA isolated from 155 canamycin-resistant diploid F1 seedlings revealed T-DNA insertions in the genomes of 111 plants (32.7% of the total number of analyzed seeds). The example of matroclinal haploids was used to demonstrate that T-DNA may be transported to the egg cell by the growing pollen tube (PT). Twelve out of 16 analyzed haploid plants contained the T-DNA insertion. The possible mechanism of the transfer of the Agrobacterium T-DNA to the maize genome during pollination is discussed.     

Salicylate degradation by a cold-adapted <Emphasis Type=Italic>Pseudomonas</Emphasis> sp.

Pseudomonas sp. strain MC1 was characterized as a cold-adapted, naphthalene-degrading bacterium that is able to grow in a broad temperature range of 5–30°C. MC1 harbors a catabolic plasmid, designated pYIC1, which is almost identical to the archetypal NAH7 plasmid from the mesophilic bacterium Pseudomonas putida G7. On pYIC1, the catabolic genes for naphthalene degradation are clustered in two operons: nahAa-Ab-Ac-Ad-B-F-C-Q-E-D encoding the conversion of naphthalene to salicylate, and nahG-T-H-I-N-L-O-M-K-J encoding the conversion of salicylate through meta-cleavage pathway to pyruvate and acetyl CoA. NahH, the bona fide extradiol dioxygenase in MC1 salicylate metabolism, is thermolabile and is a cold-adapted enzyme. The thermal profiles of the NahH enzyme activities expressed in different hosts indicate the presence of a factor(s) or mechanism(s) to protect the thermolabile NahH enzyme (100% aa identity with MC1 counterpart) in G7. Overall, the results reported in the present work suggest that the thermolabile NahH might be a product of the cold-adaptation process of MC1 and thus contribute to the survival and growth ability of MC1 on salicylate and naphthalene in cold environments.     

Cryopreservation of <Emphasis Type=Italic>Robinia</Emphasis><Emphasis Type=Italic>pseudoacacia</Emphasis>

Cryopreservation of Robinia pseudoacacia explants by vitrification achieved 78% survival following the stepwise preculture of shoot tips in (0.3 + 0.5 + 0.7 M) sucrose with a 80 min incubation in PVS2; compared to 87% survival after desiccation of explants to 30% water content, following 3 days alginate bead (with glycerol and sucrose treatments) preculture in 0.7 M sucrose.     

Physiological aspects of tolerance in <Emphasis Type=Italic>Atriplex</Emphasis><Emphasis Type=Italic>halimus</Emphasis> L. to NaCl and drought

Forty-day-old seedlings of Atriplex halimus were treated either with NaCl (50, 300 and 550 mM) for the subsequent 30 days or with 15% PEG for the subsequent 10 days. As much as 50 mM of NaCl significantly increased shoot fresh and dry weight and height; nevertheless, 300 or 550 mM NaCl seemed to have no effect. On the other hand, these growth parameters were not affected by drought after 3 or 6 days, but were reduced after 10 days. The gas exchange parameters (photosynthetic rate, stomatal conductance and transpiration rate) were increased by 50 mM NaCl, but decreased by 300 and 550 mM. These parameters were decreased in response to drought only after 10 days of withholding water. In contrast to Na⁺, K⁺ was significantly decreased by NaCl but not by drought. The time course effect revealed that phosphoenol pyruvate carboxylase (PEPC) protein was doubled in response to NaCl after 1 and 5 h and continued thereafter, higher than control, while drought had no significant effect. Rubisco seemed unchanged by NaCl or drought. It could be concluded that the decrease in fresh weight might be attributed to the decrease in water content. Moreover, the decrease in photosynthesis could result from a decrease in stomatal conductance, a protective mechanism against water loss to improve water use efficiency. These findings indicate that Atriplex halimus tolerates NaCl and drought through decreasing growth, reducing gas exchange parameters to improve water use efficiency, uptake Na⁺ and saving, if any, the photosynthetic enzyme particularly PEPC.     

Tissue-culture enhanced transposition of the maize transposable element <Emphasis Type=Italic>Dissociation</Emphasis> in <Emphasis Type=Italic>Brassica oleracea</Emphasis> var. '<Emphasis Type=Italic>Italica</Emphasis>'

To investigate the potential of heterologous transposons as a gene tagging system in broccoli (Brassica oleracea var. Italica), we have introduced a Ds-based two-element transposon system. Ds has been cloned into a 35S-SPT excision-marker system, with transposition being driven by an independent 35S-transposase gene construct (Tpase). In three successive selfed generations of plants there was no evidence of germinal-excision events. To overcome this apparent inability to produce B. oleracea plants with germinal excisions, we performed a novel tissue-culture technique to select for fully green shoots from seed with somatic-excision events. The results showed a very high efficiency of regeneration of fully green plants (up to 65%) and molecular analysis indicated that the plants genetically were like plants that contain a germinal-excision event. Further molecular analysis of these plants showed that 69% exhibited reinsertion of Ds back into the plant genome. Sequencing of donor-site footprints after Ds excision, revealed that there is an indication of more-severe deletions and rearrangements when higher concentrations of streptomycin are used in the tissue-culture selection process. Adapted versions of this regeneration technique have a high potential for providing germinal excision-like events in heterologous plants species which show low transposon activity. Alternatively, there is the potential to increase the proportion of 'germinal' plants in earlier generations of more-active plant species.     

Attenuation of fibrosis <Emphasis Type=Italic>in vitro</Emphasis> and <Emphasis Type=Italic>in vivo</Emphasis> with <Emphasis Type=Italic>SPARC</Emphasis> siRNA

Introduction  

SPARC is a matricellular protein, which, along with other extracellular matrix components including collagens, is commonly over-expressed in fibrotic diseases. The purpose of this study was to examine whether inhibition of SPARC can regulate collagen expression in vitro and in vivo, and subsequently attenuate fibrotic stimulation by bleomycin in mouse skin and lungs.     

Presence of <Emphasis Type=Italic>C. albidus</Emphasis>, <Emphasis Type=Italic>C. laurentii</Emphasis> and <Emphasis Type=Italic>C. uniguttulatus</Emphasis> in Crop and Droppings of Pigeon Lofts (<Emphasis Type=Italic>Columba livia</Emphasis>)

Columba livia is an important reservoir and carrier of Cryptococcus neoformans, Cryptococcus uniguttulatus, Cryptococcus laurentii and Cryptococcus albidus. Upper digestive tract of this species is also known as a habitat for Cryptococcus neoformans. Given the increasing clinical interest of this microorganism, 331 swabs from crop and 174 dropping samples from pigeon lofts in Grand Canary Island have been studied. The obtained results show an extensive presence samples 81 positive (24.47%) of Cryptococcus spp. in analysed crops: 32 (9.66%) for C. neoformans, 24 (7.2%) for C. uniguttulatus, 23 (6.9%) for C. albidus and 2 (0.6%) for C. laurentii. In the same way, Cryptococcus spp was also isolated in 82 (47.13%), dropping samples: C. neoformans in 59 (33.9%), C. uniguttulatus, in 9 (5.17%), C. laurentii in 8 (4.59%) and C. albidus in 6 (3.44%) of the investigated samples, respectively. The cryptococcosis produced by species of cryptococci other than C. neoformans has become more important during the last decade, supporting the study on the role of pigeon in the epidemiology of this disease.     

Efficient separation and purification of allophycocyanin from <Emphasis Type=Italic>Spirulina</Emphasis> (<Emphasis Type=Italic>Arthrospira</Emphasis>) <Emphasis Type=Italic>platensis</Emphasis>

Allophycocyanin (APC) is a minor component of phycobiliproteins in cyanobacteria and red algae. This paper describes a simple and inexpensive extracting method for isolating APC from Spirulina (Arthrospira) platensis with high efficiency. The crude phycobiliprotein extract was pretreated by ammonium sulfate fractionation. Then, by adding hydroxylapatite into crude phycobiliprotein extract dissolved in 20 mM phosphate buffer (pH 7.0), APC was selectively adsorbed by hydroxylapatite but C-phycocyanin (C-PC) was not. The hydroxylapatite was collected and APC was extracted from the crude phycobiliprotein extract. Then, the enriched APC was washed off from the hydroxylapatite using 100 mM phosphate buffer (pH 7.0). In this simple extracting method it was easy to remove C-PC and isolate APC in large amounts. The absorbance ratio A ₆₅₀/A ₂₈₀ of extracted APC reached 2.0. The recovery yield was 70%, representing 4.61 mg · g⁻¹ wet weight. The extracted APC could be further purified by a simple anion-exchange chromatography with a pH gradient from 5.6 to 4.0. The absorbance ratio A ₆₅₀/A ₂₈₀ of the purified APC reached 5.0, and the overall recovery yield was 43%, representing 2.83 mg · g⁻¹ wet weight. Its purity was confirmed by native polyacrylamide gel electrophoresis (PAGE) and sodium dodecyl sulfate-PAGE.     

<Emphasis Type=Italic>In vitro</Emphasis> organogenesis of <Emphasis Type=Italic>Passiflora alata</Emphasis>

Passiflora alata in vitro organogenesis was studied based on explant type, culture medium composition, and incubation conditions. The results indicated that the morphogenic process occurred more efficiently when hypocotyl segment-derived explants were cultured in media supplemented with cytokinin and AgNO₃ incubated under a 16-h photoperiod. The shoot bud elongation and plant development were obtained by transferring the material to MSM culture medium supplemented with GA₃ and incubated in flasks with vented lids. Histological analyses of the process revealed that the difficulties in obtaining plants could be related to the development of protuberances and leaf primordia structures, which did not contain shoot apical meristem. Roots developed easily by transferring elongated shoots to 1/2 MSM culture medium. Plant acclimatization occurred successfully, and somaclonal variation was not visually detected. The efficiency of this organogenesis protocol will be evaluated for genetic transformation of this species to obtain transgenic plants expressing genes that can influence the resistance to Cowpea aphid borne mosaic virus.     

Interaction of <Emphasis Type=Italic>Bdellovibrio bacteriovorus</Emphasis> with bacteria <Emphasis Type=Italic>Campylobacter jejuni</Emphasis> and <Emphasis Type=Italic>Helicobacter pylori</Emphasis>

Interaction of Bdellovibrio bacteriovorus 100NCJB with bacteria Campylobacter jejuni (strains 1, 2, 3, 4, and 5) and Helicobacter pylori, strain TX30a, was confirmed. The results indicate that lytic activity of bdellovibrios both in liquid media and cells attached to a surface was observed. The potential use of the antimicrobial activity of predatory bacteria for environmental bioprotection and public health is discussed.