Effect of nutrient deprivation on lipid, carbohydrate, DNA, RNA, and protein levels in Vibrio cholerae

The response of Vibrio cholerae to low nutrient levels was determined by measuring the concentrations of lipids, carbohydrates, DNA, RNA, and proteins over a 30-day starvation period. Ultrastructural integrity was observed by transmission electron microscopy. Total lipids and carbohydrates declined rapidly within the first 7 days, while DNA and protein exhibited a more constant decline over the 30 days of starvation. In contrast, RNA showed little decrease upon starvation. Although neutral lipids were lost, the percentage of neutral lipids did not decline as rapidly as the phospholipids. Detectable levels of poly-beta-hydroxybutyrate disappeared completely by 7 days. Carbohydrate profiles revealed the relative loss of the five-carbon sugar ribose and N-acetylglucosamine and a relative increase in the total six-carbon sugars, especially glucose. Morphologically, ribosomes appeared to exhibit no structural change, while inclusion bodies and mesosomelike structures disappeared completely, and cell wall and membrane integrity was lost. The data suggest that V. cholerae differs somewhat from other marine vibrios in its response to low nutrients but shares some characteristics in common with them. The data also suggest that certain lipids and carbohydrates may provide the endogenous energy sources needed for dormancy preparation and cell maintenance under nutrient starvation.     

Model genomes: the benefits of analysing homologous human and mouse sequences.

The human genome initiative has provided the motivating force for launching sequencing projects suitable for testing various DNA-sequencing strategies, as well as motivating the development of mapping and sequencing technologies. In addition to projects targeting selected regions of the human genome, other projects are based on model organisms such as yeast, nematode and mouse. The sequencing of homologous regions of human and mouse genomes is a new approach to genome analysis, and is providing insights into gene evolution, function and regulation which could not be determined so easily from the analysis of just one species.     

Urinary excretion of prostaglandin E2 and prostaglandin F2α in potassium-deficient rats

Potassium-deficiency was induced in rats by dietary deprivation of potassium. The animals became polyuric and urine osmolality decreased more then three-fold compared to controls. Urinary excretion of prostaglandin E₂ (PGE₂) and prostaglandin F_2α (PGF_2α) did not increase during 2 weeks of potassium depletion. Partial inhibition of renal prostaglandin synthesis by meclofenamate did not increase the urine osmolality after water deprivation. These results make unlikely the hypothesis that the polyuria of potassium-deficiency, is the result of enhanced renal synthesis of prostaglandins with subsequent antagonism of the hydro-osmotic effect of vasopressin. Male animals consistently excreted less PGE₂ than female animals.     

Optimization of asymmetric polymerase chain reaction for rapid fluorescent DNA sequencing

A high-throughput method for the preparation of single-stranded template DNA, which is suitable for sequence analysis using fluorescent labeling chemistry, is described here. In this procedure, the asymmetric polymerase chain reaction is employed to amplify recombinant plasmid or bacteriophage DNA directly from colonies or plaques. The use of amplification primers located at least 200 base pairs 5' to the site of sequencing primer annealing removes the need for extensive purification of the asymmetric polymerase chain reaction product. Instead, the single-stranded product DNA is purified by a simple isopropanol precipitation step and then directly sequenced using fluorescent dye-labeled oligonucleotides. This method significantly reduces the time and labor required for template preparation and improves fluorescent DNA sequencing strategies by providing a much more uniform yield of single-stranded DNA.     

The Molecular Evolution of the Vertebrate Trypsinogens

We expand the already large number of known trypsinogen nucleotide and amino acid sequences by presenting additional trypsinogen sequences from the tunicate (Boltenia villosa), the lamprey (Petromyzon marinus), the pufferfish (Fugu rubripes), and the frog (Xenopus laevis). The current array of known trypsinogen sequences now spans the entire vertebrate phylogeny. Phylogenetic analysis is made difficult by the presence of multiple isozymes within species and rates of evolution that vary highly between both species and isozymes. We nevertheless present a Fitch-Margoliash phylogeny constructed from pairwise distances. We employ this phylogeny as a vehicle for speculation on the evolution of the trypsinogen gene family as well as the general modes of evolution of multigene families. Unique attributes of the lamprey and tunicate trypsinogens are noted. Received: 12 July 1997     

A survey of infections in stereotactic surgery

In 365 consecutive intracranial stereotactic procedures over a 6-year period, there were three postoperative infections; 1 case of increased neurological deficit and 1 of fatality. Staphylococcus aureus was the most common organism involved; however, multiple organisms were seen in 2 cases. Multiplicity of organisms necessitated the use of a broad spectrum of antibiotics in the preoperative and postoperative periods. The method described has permitted an infection rate which compares favorably to other reports in the literature. Perioperative antibiotics, antibiotic-containing irrigation solutions, and careful attention to sterile techniques seem to be instrumental in maintaining a low infection rate in our study. Since the new spheroid design has been used, there have been no infections.     

Relationship Estimation from Whole-Genome Sequence Data

The determination of the relationship between a pair of individuals is a fundamental application of genetics. Previously, we and others have demonstrated that identity-by-descent (IBD) information generated from high-density single-nucleotide polymorphism (SNP) data can greatly improve the power and accuracy of genetic relationship detection. Whole-genome sequencing (WGS) marks the final step in increasing genetic marker density by assaying all single-nucleotide variants (SNVs), and thus has the potential to further improve relationship detection by enabling more accurate detection of IBD segments and more precise resolution of IBD segment boundaries. However, WGS introduces new complexities that must be addressed in order to achieve these improvements in relationship detection. To evaluate these complexities, we estimated genetic relationships from WGS data for 1490 known pairwise relationships among 258 individuals in 30 families along with 46 population samples as controls. We identified several genomic regions with excess pairwise IBD in both the pedigree and control datasets using three established IBD methods: GERMLINE, fastIBD, and ISCA. These spurious IBD segments produced a 10-fold increase in the rate of detected false-positive relationships among controls compared to high-density microarray datasets. To address this issue, we developed a new method to identify and mask genomic regions with excess IBD. This method, implemented in ERSA 2.0, fully resolved the inflated cryptic relationship detection rates while improving relationship estimation accuracy. ERSA 2.0 detected all 1^st through 6^th degree relationships, and 55% of 9^th through 11^th degree relationships in the 30 families. We estimate that WGS data provides a 5% to 15% increase in relationship detection power relative to high-density microarray data for distant relationships. Our results identify regions of the genome that are highly problematic for IBD mapping and introduce new software to accurately detect 1^st through 9^th degree relationships from whole-genome sequence data.