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Cadmium (Cd) accumulation in crop plants such as tobacco can lead to human exposure to this carcinogenic metal. Therefore, efforts should be made to minimize the Cd content of soils and crops. We review the options for reducing Cd content of plants, with emphasis on tobacco, a plant that can accumulate relatively high levels of this metal in its leaves. Many studies aimed at understanding Cd biology in plants do not reflect field conditions, because, often of necessity, non-field-like conditions were used. Thus, further study is needed to understand which processes govern Cd uptake, accumulation, etc., under field conditions. Numerous factors, such as soil characteristics, agronomic practices, and environmental conditions, impact the uptake of Cd by plants, including tobacco. Identifying anthropogenic sources of Cd and controlling application may limit Cd accumulation in agricultural fields. Soil remediation strategies may be envisaged to reduce Cd availability to the plant (e.g., soil amendments) or to extract Cd from the soil (e.g., phytoextraction). Another approach for reducing Cd in crops involves genetic modification of the plant to reduce Cd uptake or to change its partitioning in the plant. Indeed, more knowledge has been gained in recent decades regarding the mechanisms governing the transport, accumulation, and compartmentalization of Cd by tobacco and other plants. Several types of genes can be considered for genetic engineering to affect these processes. Although no single remedy appears to exist that might drastically reduce the Cd content of crops, including tobacco, an integrated approach may prove useful. [ABSTRACT FROM AUTHOR]

We report the first linkage map of tobacco (Nicotiana tabacum L.) generated through microsatellite markers. The microsatellite markers were predominantly derived from genomic sequences of the Tobacco Genome Initiative (TGI) through bioinformatics screening for microsatellite motives. A total of 684 primer pairs were screened for functionality in a panel of 16 tobacco lines. Of those, 637 primer pairs were functional. Potential parents for mapping populations were evaluated for their polymorphism level through genetic similarity analysis. The similarity analysis revealed that the known groups of tobacco varieties (Burley, Flue-cured, Oriental and Dark) form distinct clusters. A mapping population, based on a cross between varieties Hicks Broad Leaf and Red Russian, and consisting of 186 F2 individuals, was selected for mapping. A total of 282 functional microsatellite markers were polymorphic in this population and 293 loci could be mapped together with the morphological trait flower color. Twenty-four tentative linkage groups spanning 1,920 cM could be identified. This map will provide the basis for the genetic mapping of traits in tobacco and for further analyses of the tobacco genome.

The Cooperation Centre for Scientific Research Relative to Tobacco (CORESTA; Paris, France) "Task Force Genetically Modified Tobacco-Detection Methods" investigated the performance of qualitative and quantitative methods based on the polymerase chain reaction (PCR) for the detection and quantitation of genetically modified (GM) tobacco. In the 4 successful rounds of proficiency testing, the cauliflower mosaic virus 35S RNA promoter (CaMV 35S) and the Agrobacterium tumefaciens nopaline synthase terminator (NOS) were selected as target sequences. Blind-coded reference materials containing from 0.1 to 5.0% and from 0.15 to 4% GM tobacco were used in 2 rounds of qualitative and quantitative PCR, respectively. Eighteen laboratories from 10 countries participated in this study. Considering all methods and 2 rounds, the different laboratories were able to detect GM tobacco at the 0.1% level in 46 out of 58 tests in qualitative assays. The results of the proficiency test indicate that both end point screening and real-time quantitative methods are suitable for the detection of genetically modified organisms in tobacco leaf samples having a GM content of 0.1% or higher. The CORESTA proficiency study represents a first step towards the interlaboratory evaluation of accuracy and precision of PCR-based GM tobacco detection, which may lead to the harmonization of analytical procedures and to the enhancement of comparability of testing results produced by different laboratories.

Every year raw tobacco and manufactured tobacco products are lost to two major storage pests, the cigarette beetle, Lasioderma serricorne (F) and the tobacco moth, Ephestia elutella (Hiibner). Post-harvest management of both insects is achieved through sanitation, insect monitoring and fumigation with phosphine. However, insect resistance to phosphine and control failures have been reported, and fumigants are under constant regulatory pressure. Here we report the evaluation of spinosad, a bioinsecticide derived from the fermentation of the soil micro-organism Saccharopolyspora spinosa Mertz & Yao. Spinosad was first registered in 1997 and is now widely used as a field pest control agent on many crops, including tobacco. The insecticidal activity of the fermentation product (technical spinosad, TS) was measured by diet incorporation assays against L serricorne and E elutella larvae. Mortality levels were determined on newly hatched larvae and over the whole insect life cycle. For both species, no emergence of adult insects was observed in cured tobacco sprayed with 50mg TS kg(-1) and inoculated with eggs or newly hatched larvae. These results indicated that spinosad has potential for the control of both species in stored tobacco, since 100% control of both pests could be achieved at 50 mg TS kg(-1), and with almost full control (90-95%) at 10 mg kg(-1). We also monitored the stability of the product on cured tobacco. The original concentration of the main active component of TS, spinosyn A, did not change significantly over 18 months, indicating no loss of spinosad during a typical leaf storage period of time. Bioassays against larvae confirmed that the bioinsecticidal activity of spinosad was retained.

Experiments were conducted in tobacco (Nicotiana tabacum L.) to investigate the effect of sodium bicarbonate (NaHCO(3)) on the conversion of nicotine to nornicotine, a secondary alkaloid that can form the tobacco-specific nitrosamine N-nitrosonornicotine (NNN). The results showed that, under optimum conditions, NaHCO(3) stimulated nicotine conversion in converter plants to the maximum level predetermined by the genetic background. The conversion level in NaHCO(3)-treated leaves was 2-3 times higher than that in control leaves. For young seedlings the optimum concentration of sodium bicarbonate was 0.8% aqueous solution, and for adult plants the optimum concentration was 1%. Lower concentrations resulted in partial stimulation, whereas higher concentration damaged leaf tissue and resulted in a lower conversion level. Studies with different temperatures (from 22 to 43 degrees C) showed that 37 degrees C was optimal. This temperature allowed the least amount of time, 2-3 days for mature leaves and 4-6 days for green leaves, for the major converters to reach >95% of nicotine conversion. An examination of leaves from different growth stages and stalk positions showed that the amount of time needed for conversion was longer for young leaves and shorter for mature leaves. Treatment of leaves with NaHCO(3) affords a rapid and convenient means of identifying and removing nornicotine converter plants during growth in the greenhouse or field.

A worldwide survey was conducted to evaluate the frequency and distribution of Bacillus thuringiensis populations on cured tobacco leaves during post-harvest storage. In total, 133 tobacco samples of different types and origins were analyzed. Nine percent of the samples showed the presence of B. thuringiensis, and 24 B. thuringiensis strains were isolated and characterized. The majority of the isolates produced bipyramidal crystals, and three fourths of them showed a second type of crystal protein (cuboidal or heterogeneous crystals). Only three isolates showed the rhomboidal crystal morphology characteristic of the anti-coleopteran B. thuringiensis subsp. tenebrionis. PCR analysis with primers specific for cry1 and cry3 genes revealed eight distinct cry gene profiles. The results of this study indicate that B. thuringiensis is naturally present at low frequency on the phylloplane of cured tobacco leaves and that its distribution is worldwide.

Bacillus thuringiensis was isolated from dried tobacco residues and dead tobacco beetles (Lasioderma serricorne (F.); Coleoptera: Anobiidae) collected in a large number of locations worldwide. Eighty-eight samples of stored tobacco were analyzed and yielded 78 B. thuringiensis strains which were characterized on the basis of parasporal crystal morphology, sodium dodecyl sulfate-polyacrylamide gel electrophoresis profiles, and the results of an immunoblot analysis of the insecticidal crystal proteins. Flagellar antigen identification was used to differentiate selected isolates. Strains that produced rhomboidal crystals associated with the Coleoptera-specific pathotype (Cry III group) were the most abundant strains (59% of the isolates). Preliminary toxicity assays were performed with L. serricorne larvae, and the results suggested that activity is not restricted to isolates related to the Coleoptera-specific group. The results of our survey indicate that B. thuringiensis is part of the natural microflora in the stored-tobacco environment and that this special habitat represents a source of B. thuringiensis isolates that may be used to control stored-product pests.