Characterization of <Emphasis Type="Italic">Penicillium</Emphasis> Species by Ribosomal DNA Sequencing and BOX,ERIC and REP-PCR Analysis

The genus Penicillium is one of the largest and widely distributed fungal genera described to date. As a result, its taxonomic classification and species discrimination within this genus has become complicated. In this study, 52 isolates that belonged to the Penicillum genus and other related genera were characterized using two DNA-based methods: (i) analysis of the nucleotide sequences of internal transcribed spacers in ribosomal DNA and (ii) analysis of DNA fingerprints that were generated by polymerase chain reactions with specific primers for enterobacterial repetitive intergenic consensus (ERIC) and repetitive extragenic palindromic (REP) sequences, and BOX elements. Using both methods, Penicillium species were discriminated from other fungal genera. Furthermore, Penicillium species that include strains which are used as biocontrol agents, such as P. glabrum, P. purpurogenum, and P. oxalicum, could be distinguished from other Penicillium species using these techniques. Based on our findings, we propose that a polyphasic approach that includes analysis of the nucleotide sequences of ribosomal DNA and detecting the presence of highly conserved, repeated nucleotide sequences can be used to determine the genetic relationships between different Penicillium species. Furthermore, we propose that our results can be used as a start point to develop a strategy to monitor the environmental presence of particular strains of Penicillium species when they are used as biocontrol agents.     

Comparison of genomes of eight species of sections <Emphasis Type="Italic">Linum</Emphasis> and <Emphasis Type="Italic">Adenolinum</Emphasis> from the genus<Emphasis Type="Italic"> Linum</Emphasis> based on chromosome banding,molecular markers and RAPD analysis

Karyotypes of species sects. Linum and Adenolinum have been studied using C/DAPI-banding, Ag-NOR staining, FISH with 5S and 26S rDNA and RAPD analysis. C/DAPI-banding patterns enabled identification of all homologous chromosome pairs in the studied karyotypes. The revealed high similarity between species L. grandiflorum (2n = 16) and L. decumbens by chromosome and molecular markers proved their close genome relationship and identified the chromosome number in L. decumbens as 2n = 16. The similarity found for C/DAPI-banding patterns between species with the same chromosome numbers corresponds with the results obtained by RAPD-analysis, showing clusterization of 16-, 18- and 30-chromosome species into three separate groups. 5S rDNA and 26S rDNA were co-localized in NOR-chromosome 1 in the genomes of all species investigated. In 30-chromosome species, there were three separate 5S rDNA sites in chromosomes 3, 8 and 13. In 16-chromosome species, a separate 5S rDNA site was also located in chromosome 3, whereas in 18-chromosome species it was found in the long arm of NOR-chromosome 1. Thus, the difference in localization of rDNA sites in species with 2n = 16, 2n = 30 and 2n = 18 confirms taxonomists opinion, who attributed these species to different sects. Linum and Adenolinum, respectively. The obtained results suggest that species with 2n = 16, 2n = 18 and 2n = 30 originated from a 16-chromosome ancestor.     

Differential expression and regulation of expansin gene family members during fruit growth and development of ‘Shijia’ longan fruit

In this work, five expansin cDNAs (DlExp1–5) from ‘Shijia’ longan fruit were isolated and characterized. Moreover, the expression profiles of five expansin genes and the effect of naphthalene acetic acid (NAA) and thidiazuron (TDZ) on their expressions were investigated. The results showed that five expansins exhibited different expression patterns during fruit growth and development. DlExp1 was constitutively expressed in the pericarp while the levels of DlExp1 mRNA in the aril were very high at early stage of fruit development, and decreased gradually from 28 to 77 days after anthesis (DAA). DlExp2 and DlExp4 were related to the growth of pericarp, whereas the expression of DlExp2 and DlExp5 in the aril decreased from 28 to 77 DAA. In addition, NAA and TDZ applied at the stage of rapid pericarp (21 DAA) or aril growth (56 DAA) increased the accumulations of DlExp1 and DlExp2 mRNA in the pericarp and aril, while NAA and TDZ had no or little effect on the accumulations of DlExp3, DlExp4 and DlExp5. DlExp1 and DlExp2 also accumulated highly in rapidly growing tissues, such as young stems and leaves. These findings indicated that Exp genes played a different role in longan fruit growth and showed different response to plant growth substances.     

The effects of diet and caloric restriction on adipose tissue fatty acid signatures of tufted puffin (<Emphasis Type="Italic">Fratercula cirrhata</Emphasis>) nestlings

Fatty acid (FA) signature analysis is a powerful tool to investigate foraging ecology and food web dynamics in marine ecosystems. However, use of FA signatures to qualitatively or quantitatively infer diets is potentially complicated by effects of nutritional state on lipid metabolism. Estimation of diets using the quantitative fatty acid signature analysis (QFASA) model requires the use of calibration coefficients to account for predator metabolism of individual FAs. We conducted a captive feeding experiment to determine the effects of a 50% reduction in food intake on growth rate and adipose tissue FA signatures of tufted puffin (Fratercula cirrhata) nestlings, a species that routinely experiences food restriction during growth. FA signatures of chicks fed low- and high-calorie diets both exhibited a change in composition in response to the dietary shift with the direction of change in the composition of individual FAs matching the direction of change in the dietary FAs. Despite a growth rate in the restricted nestlings that was 38% of those in the well-fed group, rates of FA turnover were not different between high and low-calorie treatments, and turnover was close to, but not entirely complete, after 27 days on both high-calorie and restricted diets. FA signatures of tufted puffin nestlings were significantly affected by caloric restriction, but these effects were much less pronounced than those of dietary turnover, and calibration coefficients of puffins fed low and high-calorie diets were highly correlated. Our results demonstrate that changes in physiological state can affect FA metabolism, but future research is required to better understand whether the size of these effects is sufficient to substantially alter diet estimation using the QFASA model.     

Use of plate-wash samples to monitor the fates of culturable bacteria in mercury- and trichloroethylene-contaminated soils

With the ultimate aim of developing bioremediation technology that use the optimum bacterial community for each pollutant, we performed polymerase chain reaction (PCR)-denaturing gradient gel electrophoresis (DGGE) and phylogenetic analysis and identified communities of culturable bacteria in HgCl(2)- and trichloroethylene (TCE)-contaminated soil microcosms. PCR-DGGE band patterns were similar at 0 and 1 ppm HgCl(2), but changes in specific bands occurred at 10 ppm HgCl(2). Band patterns appearing at 10 and 100 ppm TCE were very different from those at 0 ppm. Phylogenetic analysis showed four bacterial groups in the HgCl(2)-contaminatied cultures: Firmicutes, Actinobacteria, Proteobacteria, and Bacteroidetes. Most high-density bands, decreased-density bands, and common bands were classified into the phyla Proteobacteria, Actinobacteria, and Firmicutes, respectively; the effects of HgCl(2) on culturable bacteria appeared to differ among phyla. Duganella violaceinigra [98.4% similarity to DNA Data Bank of Japan (DDBJ) strain], Lysobacter koreensis (98.2%), and Bacillus panaciterrae (98.6%) were identified as bacteria specific to HgCl(2)-contaminated soils. Bacteria specific to TCE-contaminated soils were distributed into three phyla (Firmicutes, Proteobacteria, and Actinobacteria), but there was no clear relationship between phylum and TCE effects on culturable bacteria. Paenibacillus kobensis (97.3%), Paenibacillus curdlanolyticus (96.3%), Paenibacillus wynnii (99.8%), and Sphingomonas herbicidovorans (99.4%) were identified as bacteria specific to TCE-contaminated soils. These bacteria may be involved in pollutant degradation.     

Inferring angiosperm phylogeny from EST data with widespread gene duplication

Background

Most studies inferring species phylogenies use sequences from single copy genes or sets of orthologs culled from gene families. For taxa such as plants, with very high levels of gene duplication in their nuclear genomes, this has limited the exploitation of nuclear sequences for phylogenetic studies, such as those available in large EST libraries. One rarely used method of inference, gene tree parsimony, can infer species trees from gene families undergoing duplication and loss, but its performance has not been evaluated at a phylogenomic scale for EST data in plants.

Results

A gene tree parsimony analysis based on EST data was undertaken for six angiosperm model species and Pinus, an outgroup. Although a large fraction of the tentative consensus sequences obtained from the TIGR database of ESTs was assembled into homologous clusters too small to be phylogenetically informative, some 557 clusters contained promising levels of information. Based on maximum likelihood estimates of the gene trees obtained from these clusters, gene tree parsimony correctly inferred the accepted species tree with strong statistical support. A slight variant of this species tree was obtained when maximum parsimony was used to infer the individual gene trees instead.

Conclusion

Despite the complexity of the EST data and the relatively small fraction eventually used in inferring a species tree, the gene tree parsimony method performed well in the face of very high apparent rates of duplication.

     

Drought’s impact on Ca,Fe, Mg,Mo and S concentration and accumulation patterns in the plants and soil of a Mediterranean evergreen <Emphasis Type="Italic">Quercus </Emphasis><Emphasis Type="Italic">ilex</Emphasis> forest

We conducted a 6-year field manipulation drought experiment in an evergreen Quercus ilex forest where we simulated the drought predicted by GCM and ecophysiological models for the coming decades (an average of 15% soil moisture reduction). We thereby tested the hypothesis that enhanced drought will change Ca, Fe, Mg, Mo and S availability, concentrations and accumulation patterns in Mediterranean ecosystems. The strongest effects of drought occurred in the soil. Drought increased the total soil concentrations of S, the soil extract concentrations of Fe, Mg and S, the Mg saturation in the soil exchangeable complex and tended to increase the percentage base saturation of the soil exchangeable complex. These increased soil concentrations were related to a decrease of plant uptake capacity and not to an increase of soil enzyme activity, which in fact decreased under drier conditions. Drought increased leaf Mg concentrations in the three dominant species although only significantly in Quercus ilex and Arbutus unedo (20 and 14%, respectively). In contrast, drought tended to decrease Ca in Phillyrea latifolia (18%) and Ca and Fe concentrations in the wood of all three species. Drought increased Ca and Fe concentrations in the roots of Quercus ilex (26 and 127%). There was a slight general trend to decrease total biomass accumulation of nutrients that depend on water flux such as Mg, Fe and S. This effect was related to a decrease of soil moisture that reduced soil flow, and to a decrease in photosynthetic capacity, sap flow, transpiration and growth, and therefore plant uptake capacity under drought observed in Quercus ilex and Arbutus unedo. On the contrary, drought increased Mo accumulation in aboveground biomass in Phillyrea latifolia and reduced Mo accumulation in Arbutus unedo by reducing growth and wood Mo concentrations (51%). Phillyrea latifolia showed a great capacity to adapt to drier conditions, with no decrease in growth, an increase of Mo uptake capacity and a decrease in leaf Ca concentration, which was related to a decrease in transpiration under drought. The results indicate asymmetrical changes in species capacity to accumulate these elements, which are likely to produce changes in inter-specific competitive relations among dominant plant species and in their nutritional quality as food sources. The results also indicate that drought tended to decrease nutrient content in aboveground biomass, mainly through the decrease in growth and transpiration of the most sensitive species and caused an increase in the availability of these nutrients in soil. Thus, drought decreased the ecosystem’s capacity to retain Mg, Fe and S, facilitating their loss in torrential rainfalls.     

Zero erucic acid trait of rapeseed (<Emphasis Type="Italic">Brassica napus</Emphasis> L.) results from a deletion of four base pairs in the <Emphasis Type="Italic">fatty acid elongase 1</Emphasis> gene

The fatty acid elongase 1 (FAE1) gene is a key gene in the erucic acid biosynthesis in rapeseed. The complete coding sequences of the FAE1 gene were isolated separately from eight high and zero erucic acid rapeseed cultivars (Brassica napus L.). A four base pair deletion between T1366 and G1369 in the FAE1 gene was found in a number of the cultivars, which leads to a frameshift mutation and a premature stop of the translation after the 466th amino acid residue. This deletion was predominantly found in the C-genome and rarely in the A-genome of B. napus. Expression of the gene isoforms with the four base pair deletion in a yeast system generated truncated proteins with no enzymatic activity and could not produce very long chain fatty acids as the control with an intact FAE1 gene did in yeast cells. In the developing rape seeds the FAE1 gene isoforms with the four base pair deletion were transcribed normally but failed to translate proteins to form a functional complex. The four base pair deletion proved to be a mutation responsible for the low erucic acid trait in rapeseed and independent from the point mutation reported by Han et al. (Plant Mol Biol 46:229–239, 2001). Gang Wu, Yuhua Wu contribute equally to this article.