Correlation of Phenotype with the Genotype of Egg-Contaminating Salmonella enterica Serovar Enteritidis

The genotype of Salmonella enterica serovar Enteritidis was correlated with the phenotype using DNA-DNA microarray hybridization, ribotyping, and Phenotype MicroArray analysis to compare three strains that differed in colony morphology and phage type. No DNA hybridization differences were found between two phage type 13A (PT13A) strains that varied in biofilm formation; however, the ribotype patterns were different. Both PT13A strains had DNA sequences similar to that of bacteriophage Fels2, whereas the PT4 genome to which they were compared, as well as a PT4 field isolate, had a DNA sequence with some similarity to the bacteriophage ST64b sequence. Phenotype MicroArray analysis indicated that the two PT13A strains and the PT4 field isolate had similar respiratory activity profiles at 37°C. However, the wild-type S. enterica serovar Enteritidis PT13A strain grew significantly better in 20% more of the 1,920 conditions tested when it was assayed at 25°C than the biofilm-forming PT13A strain grew. Statistical analysis of the respiratory activity suggested that S. enterica serovar Enteritidis PT4 had a temperature-influenced dimorphic metabolism which at 25°C somewhat resembled the profile of the biofilm-forming PT13A strain and that at 37°C the metabolism was nearly identical to that of the wild-type PT13A strain. Although it is possible that lysogenic bacteriophage alter the balance of phage types on a farm either by lytic competition or by altering the metabolic processes of the host cell in subtle ways, the different physiologies of the S. enterica serovar Enteritidis strains correlated most closely with minor, rather than major, genomic changes. These results strongly suggest that the pandemic of egg-associated human salmonellosis that came into prominence in the 1980s is primarily an example of bacterial adaptive radiation that affects the safety of the food supply.     

Importance of glucose-6-phosphate dehydrogenase in taxol biosynthesis in <Emphasis Type="Italic">Taxus chinensis</Emphasis> cultures

The roles of glucose-6-phosphate dehydrogenase (G6PDH) in paclitaxel production were investigated in cell suspension cultures of Taxus chinensis. In the normal cultures, the trend of G6PDH activity was similar to that of cell growth. Addition of glutamate increased G6PDH activity, while dehydroepiandrosterone (DHEA) decreased G6PDH activity. In elicitor-treated cultures, cell growth was depressed, while G6PDH activity and taxol production were enhanced compared with the control. Glutamate recovered the depression of cell growth, and resulted in further increase in G6PDH activity and taxol production. Contrarily, DHEA exacerbated the depression of cell growth, and decreased G6PDH activity and taxol production induced by fungal elicítor. The results indicated that G6PDH played a critic role of taxol production by affecting cell viability.     

Phylogenetics of New World Astragalus: Screening of novel nuclear loci for the reconstruction of phylogenies at low taxonomic levels

This study explores methods to use information gathered from genomics technology to understand evolutionary relationships in the hyperdiverse legume group Neo-Astragalus. These species inhabit deserts and mountains of North and South America, and even though the monophyly of the group is well established, relationships within it are still poorly understood. Plastid genes, commonly used to infer phylogenies in plants, are usually not useful for closely related taxa because of low levels of genetic variation. TheMedicago truncatula genome project provided a suite of candidate nuclear loci with high levels of variation that might prove suitable for low-level phylogenetics. This paper reports the development of methods for screening a large number of these nuclear loci, and detailed analysis of four of them. Four different patterns of phylogenetic diversification occur in the loci sampled from these genomes ofAstragalus species. One locus (CNGC4) was single copy and could be directly used in phylogenetic analyses. Two loci (ARG10 and FENR) showed patterns strongly suggestive of duplication events in some taxa, and one locus (tRALS) has apparently undergone a cryptic duplication, making it very difficult to diagnose. Potential methods for using the information provided by these loci are discussed.     

Endonuclease-like activity of heme proteins

Heme proteins, metmyoglobin, methemoglobin, and metcytochrome c showed unusual affinity for double-stranded DNA. Calorimetric studies show that binding of methemoglobin to calf thymus DNA (CTDNA) is weakly endothermic, and the binding constant is 4.9+/-0.7x10(5) M(-1). The Soret absorption bands of the heme proteins remained unchanged, in the presence of excess CTDNA, but a new circular dichroic band appeared at 210 nm. Helix melting studies indicated that the protein-DNA mixture denatures at a lower temperature than the individual components. Thermograms obtained by differential scanning calorimetry of the mixture indicated two distinct transitions, which are comparable to the thermograms obtained for individual components, but there was a reduction in the excess heat capacity. Activation of heme proteins by hydrogen peroxide resulted in the formation of high valent Fe(IV) oxo intermediates, and CTDNA reacted rapidly under these conditions. The rate was first-order in DNA concentration, and this reactivity resulted in DNA strand cleavage. Upon activation with hydrogen peroxide, for example, the heme proteins converted the supercoiled pUC18 DNA into nicked circular and linear DNA. No reaction occurred in the absence of the heme protein, or hydrogen peroxide. These data clearly indicate a novel property of several heme proteins, and this is first report of the endonuclease-like activity of the heme proteins.