Use of the Gram stain in microbiology

The Gram stain differentiates bacteria into two fundamental varieties of cells. Bacteria that retain the initial crystal violet stain (purple) are said to be 'Gram-positive,' whereas those that are decolorized and stain red with carbol fuchsin (or safranin) are said to be 'Gram-negative.' This staining response is based on the chemical and structural makeup of the cell walls of both varieties of bacteria. Gram-positives have a thick, relatively impermeable wall that resists decolorization and is composed of peptidoglycan and secondary polymers. Gram-negatives have a thin peptidoglycan layer plus an overlying lipid-protein bilayer known as the outer membrane, which can be disrupted by decolorization. Some bacteria have walls of intermediate structure and, although they are officially classified as Gram-positives because of their linage, they stain in a variable manner. One prokaryote domain, the Archaea, have such variability of wall structure that the Gram stain is not a useful differentiating tool.     

Replication slippage may cause parallel evolution in the secondary structures of mitochondrial transfer RNAs

Presence of the dihydrouridine (D) stem in the mitochondrial cysteine tRNA is unusually variable among lepidosaurian reptiles. Phylogenetic and comparative analyses of cysteine tRNA gene sequences identify eight parallel losses of the D-stem, resulting in D-arm replacement loops. Sampling within the monophyletic Acrodonta provides no evidence for reversal. Slipped-strand mispairing of noncontiguous repeated sequences during replication or direct replication slippage can explain repeats observed within cysteine tRNAs that contain a D-arm replacement loop. These two mechanisms involving replication slippage can account for the loss of the cysteine tRNA D-stem in several lepidosaurian lineages, and may represent general mechanisms by which the secondary structures of mitochondrial tRNAs are altered.      

The impact of molecular systematics on hypotheses for the evolution of root nodule symbioses and implications for expanding symbioses to new host plant genera

Current taxonomic schemes place plants that can participate in root nodule symbioses among disparate groups of angiosperms. According to the classification scheme of Cronquist (1981) which is based primarily on the analysis of morphological characters, host plants of rhizobial symbionts are placed in subclasses Rosidae and Hamamelidae, and those of Frankia are distributed among subclasses Rosidae, Hamamelidae, Magnoliidae and Dilleniidae. This broad phylogenetic distribution of nodulated plants has engendered the notion that nitrogen fixing endosymbionts, particularly those of actinorhizal plants, can interact with a very broad range of unrelated host plant genotypes. New angiosperm phylogenies based on DNA sequence comparisons reveal a markedly different relationship among nodulated plants and indicate that they form a more coherent group than has previously been thought (Chase et al., 1993; Swensen et al., 1994; Soltis et al., 1995). Molecular data support a single origin of the predisposition for root nodule symbiosis (Soltis et al., 1995) and at the same time support the occurrence of multiple origins of symbiosis within this group (Doyle, 1994; Swensen, 1996; Swensen and Mullin, In Press).     

Genetic organization and transposition properties of IS511

IS511 is an endogenous insertion sequence (IS) of the bacterium Caulobactercrescentus strain CB15 and it is the first Caulobacter IS to be characterized at the molecular level. We determined the 1266-bp nucleotide sequence of IS511 and investigated its genetic organization, relationship to other ISs, and transposition properties. IS511 belongs to a distinct branch of the IS3 family that includes ISRI, IS476, and IS1222, based on nucleotide sequence similarity. The nucleotide sequence of IS511 encodes open reading frames (orfs) designated here as orfA and orfB, and their relative organization and amino acid sequences of the predicted protein products are very similar to those of orfAs and orfBs of other IS3 family members. Nuclease S1 protection assays identified an IS511 RNA, and its 5′ end maps approximately 16 nucleotides upstream of orfA and about six nucleotides downstream of a sequence that is similar to the consensus sequence of C. crescentus housekeeping promoters. Evidence is presented that IS511 is capable of precise excision from the chromosome, and transposition from the chromosome to a plasmid. Transpositional insertions of IS511 occurred within sequences with a relatively high G?+?C content, and they were usually, but not always, flanked by a 4-bp direct repeat that matches a sequence at the site of insertion. We also determined the nucleotide sequence flanking the four endogenous IS511 elements that reside in the chromosome of C. crescentus. Our findings demonstrate that IS511 is a transposable IS that belongs to a branch of the IS3 family.     

Extractable polysaccharides of Panax quinquefolius L. (North American ginseng) root stimulate TNFalpha production by alveolar macrophages.

We have investigated the immunostimulatory activity of the medicinal plant Panax quinquefolius L. (North American ginseng). Rat alveolar macrophages were treated with different extracts from 4-year old roots, and tumour necrosis factor alpha (TNF) production was used as a measure of immunostimulatory activity. Aqueous extracts of P. quinquefolius root (1-100 microg/ml) were found to significantly stimulate alveolar macrophage TNF release. Both a P. quinquefolius methanol extract containing ginsenosides (but no polysaccharides), and pure ginsenoside-Rb1, the major ginsenoside present in P. quinquefolius, were found to be inactive as TNF-stimulating agents. Significant TNF-stimulating activity was found in the extractable polysaccharide fraction, which was hydrolyzed and found to contain glucose, galactose, arabinose, rhamnose, and mannose. This represents the first evidence that North American ginseng exerts cytokine-stimulating activity on macrophages.     

Comparisons of genetic parameters and clonal value predictions from clonal trials and seedling base population trials of radiata pine

Different methods for predicting clonal values were explored for diameter growth (diameter at breast height (DBH)) in a radiata pine clonal forestry program: (1) clones were analyzed with a full model in which the total genetic variation was partitioned into additive, dominance, and epistasis (Clone Only—Full Model); (2) clones were analyzed together with seedling base population data (Clone Plus Seedling (CPS)), and (3) clones were analyzed with a reduced model in which the only genetic term was the total genetic variance (Clone Only—Reduced Model). DBH was assessed at age 5 for clones and between ages 4 to 13 at the seedling trials. Significant additive, dominance, and epistatic genetic effects were estimated for DBH using the CPS model. Nonadditive genetic effects for DBH were 87% as large as additive genetic effects. Narrow-sense () and broad-sense () heritability estimates for DBH using the CPS model were 0.14 ± 0.01 and 0.26 ± 0.01, respectively. Accuracy of predicted clonal values increased 4% by combining the clone and seedling data over using clonal data alone, resulting in greater confidence in the predicted genetic performance of clones. Our results indicate that exploiting nonadditive genetic effects in clonal varieties will generate greater gains than that typically obtainable from conventional family-based forestry of radiata pine. The predicted genetic gain for DBH from deployment of the top 5% of clones was 24.0%—an improvement of more than 100% over family forestry at the same selection intensity. We conclude that it is best practice to predict clonal values by incorporating seedling base population data in the clonal analysis.