Prolonged inactivation of nicotinic acid adenine dinucleotide phosphate-induced Ca2+ release mediates a spatiotemporal Ca2+ memory

Although numerous extracellular stimuli are coupled to increases in intracellular Ca(2+), different stimuli are thought to achieve specificity by eliciting different spatiotemporal Ca(2+) increases. We investigated the effect of nicotinic acid adenine dinucleotide phosphate (NAADP) inactivation on spatiotemporal Ca(2+) signals in intact sea urchin eggs. The photorelease of NAADP but not inositol 1,4,5-trisphosphate or cyclic ADP-ribose resulted in self-inactivation. When NAADP was released first locally and subsequently globally, the spatial pattern of the first response shaped that of the second. Specifically, the local release of NAADP created a Ca(2+) gradient that was reversed during the subsequent global release of NAADP. Neither cyclic ADP-ribose nor inositol 1,4,5-trisphosphate showed a similar effect. In contrast to homogenates, NAADP inactivation was reversible in intact eggs with resensitization occurring in approximately 20 min. Because initial NAADP responses affect later responses, NAADP can serve as a mechanism for a Ca(2+) memory that has both spatial and temporal components. This NAADP-mediated Ca(2+) memory provides a novel mechanism for cells to control spatiotemporal Ca(2+) increases.     

Adaptive evolution and effective population size in wild house mice

Estimates of the proportion of amino acid substitutions that have been fixed by selection (α) vary widely among taxa, ranging from zero in humans to over 50% in Drosophila. This wide range may reflect differences in the efficacy of selection due to differences in the effective population size (N(e)). However, most comparisons have been made among distantly related organisms that differ not only in N(e) but also in many other aspects of their biology. Here, we estimate α in three closely related lineages of house mice that have a similar ecology but differ widely in N(e): Mus musculus musculus (N(e) ～ 25,000-120,000), M. m. domesticus (N(e) ～ 58,000-200,000), and M. m. castaneus (N(e) ～ 200,000-733,000). Mice were genotyped using a high-density single nucleotide polymorphism array, and the proportions of replacement and silent mutations within subspecies were compared with those fixed between each subspecies and an outgroup, Mus spretus. There was significant evidence of positive selection in M. m. castaneus, the lineage with the largest N(e), with α estimated to be approximately 40%. In contrast, estimates of α for M. m. domesticus (α = 13%) and for M. m. musculus (α = 12 %) were much smaller. Interestingly, the higher estimate of α for M. m. castaneus appears to reflect not only more adaptive fixations but also more effective purifying selection. These results support the hypothesis that differences in N(e) contribute to differences among species in the efficacy of selection.     

Characterization of a novel parainfluenza virus,caviid parainfluenza virus 3, from laboratory guinea pigs (Cavia porcellus)

A novel Respirovirus was isolated from nasopharyngeal swab specimens from clinically normal laboratory guinea pigs, and was characterized and named caviid parainfluenza virus 3 (CavPIV-3). The CavPIV-3 is enveloped, is 100 to 300 nm in diameter, and has a characteristic 15-nm-diameter chevron-shaped virus ribonucleocapsid protein. Sequence analysis of the fusion glycoprotein of CavPIV-3 revealed it to be 94% identical to human and guinea pig parainfluenza 3 (PIV-3) viruses and 80% identical to bovine PIV-3. To determine whether CavPIV-3 causes clinical disease in laboratory guinea pigs and to compare the serologic response of guinea pigs to CavPIV-3 and to other paramyxoviruses, an infection study was performed, in which groups of guinea pigs were inoculated with CavPIV-3, Sendai virus, simian virus 5 (SV-5), murine pneumonia virus (PVM), or bovine PIV-3 virus. During the course of the study, guinea pigs were maintained in an infectious disease suite, housed in Micro-Isolator cages, and were only manipulated under a laminar flow hood. Clinical signs of disease were not observed in any of the paramyxovirus-inoculated guinea pigs during the eight-week course of the study, and histologic signs of disease were not evident at necropsy eight weeks after inoculation. Guinea pigs inoculated with CavPIV-3, Sendai virus, PVM, and bovine PIV-3 developed robust homologous or heterologous serologic responses. In contrast, guinea pigs inoculated with SV-5 developed modest or equivocal serologic responses, as assessed by use of an enzyme-linked immunosorbent assay. Further, use of the SV-5 enzyme-linked immunosorbent assay resulted in the highest degree of non-specific reactivity among all of the paramyxovirus assays. In summary, CavPIV-3 is a novel guinea pig Respirovirus that subclinically infects laboratory guinea pigs, resulting in a robust serologic response, but no observed clinical or histologic disease. The CavPIV-3 fusion glycoprotein gene sequence is available from GenBank as accession No. AF394241, and the CavPIV-3 virus is available from the American Type Culture Collection as accession No. DR-1547.     

Changes in Bacterial Species Composition in Enrichment Cultures with Various Dilutions of Inoculum as Monitored by Denaturing Gradient Gel Electrophoresis

Denaturing gradient gel electrophoresis revealed changes in the bacterial species obtained from enrichment cultures with different inoculum dilutions. This inoculum dilution enrichment approach may facilitate the detection and isolation of a greater number of bacterial species than traditional enrichment techniques.     

Optimizing parental selection for genetic linkage maps.

Genetic linkage maps based on restriction fragment length polymorphisms are useful for many purposes; however, different populations are required to fulfill different objectives. Clones from the linkage map(s) are subsequently probed onto populations developed for special purposes such as gene tagging. Therefore, clones contained on the initial map(s) must be polymorphic on a wide range of genotypes to have maximum utility. The objectives of this research were to (i) calculate polymorphism information content values of 51 low-copy DNA clones and (ii) use the resulting values to choose potential mapping parents. Polymorphism information content was calculated using gene diversity by classifying restriction fragment patterns on a diverse set of 18 wheat genotypes. Combinations of potential parents were then compared by examining both the proportion of polymorphic clones and the likelihood that those mapped clones would give a polymorphism when used on other populations. Genotype pairs were identified that would map more highly informative DNA clones compared with a population derived from the most polymorphic potential parents. The methodologies used to characterize clones and rank potential parents should be applicable to other species and types of markers as well.     

Response of protozoans to detergent-enzymes

Summary These studies were divided into two parts: (1) time until death curves in which Paramecium caudatum Ehrenberg was exposed to selected concentrations of the detergent-enzyme Axion, and (2) exposure of fresh-water protozoan communities to selected concentrations of Axion. The former were carried out in test tubes, the latter in plastic troughs with a constantly flowing fluid. Both types of bioassays were carried out with filtered water from Douglas Lake, Michigan, U.S.A. at a temperature of about 25° C. One hundred percent of an exposed population of Paramecium caudatum was killed in 20 to 30 minutes at a concentration of 100 ppm of Axion. No appreciable change in the controls was noted during these tests. The bioassays with fresh-water protozoan communities were carried out in plastic troughs with a constant flow of Douglas Lake water. During the exposure period the lake water flow was stopped and a flow of Axion solution was substituted for approximately three hours. After this exposure period lake water flow were restored. The number of species was determined before and at intervals following exposure. Appropriate controls were maintained. A three hour exposure to 56 ppm (introduced concentration) of Axion caused a 35 percent reduction in the number of species in less than five hours with recovery to the original number in approximately 145 hours; a three hour exposure to 75 ppm caused a 54% reduction in number of species in less than five hours with recovery to the original number in approximately 240 hours; 100 ppm produced a 55 percent reduction with recovery in approximately 200 hours; 125 ppm produced a 48% reduction with recovery in approximately 148 hours; 200 ppm caused a 78% reduction with recovery in about 140 hours. Response patterns vary as do those of bioassays with single species. However, it is important to note that each test community had different types of species present, and it is quite likely that some communities had a higher percentage of sensitive species than others. Since this variation also occurs in nature the tests are probably representative of the differences that exist when natural communities are exposed to wastes. Variation in control troughs was less than ± 10 percent in the number of species.