Uniparental chromosome elimination in the early embryogenesis of the inviable salmonid hybrids between masu salmon female and rainbow trout male

Chromosome elimination through chromosome loss and partial deletion is known to be one of the causes of embryonic inviability in some salmonid interspecific hybrids. Using fluorescence in situ hybridization and related techniques, including whole chromosome painting and comparative genomic hybridization, parental origin of eliminated chromosomes was identified in the inviable hybrids between masu salmon (Ms, Oncorhynchus masou) female and rainbow trout (Rb, O. mykiss) male at the early embryonic stage prior to death. In these hybrids, the haploid Rb chromosome number decreased to nearly half, whereas the Ms chromosomes were retained as one or occasionally two full haploid complements. The Rb chromosomes were also involved in the frequently observed fragments and micronuclei. Whereas the occurrence of fragments was constant throughout the observed period, chromosome loss occurred mainly from just after fertilization to the blastulae stage. In tissue sections and cell spreads of late blastula, some Rb chromosomes were trapped in the midzone from ana- to telophase, resulting in micronuclei at the subsequent interphase. Micronuclei and mitotic abnormalities were also observed in the androgenetic haploid hybrids. However, such abnormalities were seldom or never observed in the viable reciprocal hybrids. The present findings suggest that the paternal Rb chromosomes in the inviable hybrids are preferentially eliminated through mitotic abnormalities during early embryogenesis, owing to a possible incompatibility between the maternal Ms cytoplasm and paternal Rb genome. Received: 22 August 1996; in revised form: 14 November 1996 / Accepted: 20 November 1996     

Essential role of the yolk syncytial layer for the development of isolated blastoderms from medaka embryos

To investigate a possible role of the yolk syncytial layer (YSL) in the development of the medaka embryo, blastoderms were isolated at different stages of embryogenesis either with or without the layer and were incubated in a culture medium. The blastoderms from cleavage stage embryos (stage 8–9), in which the YSL had not yet formed, developed into an irregular mass of cells. But some of the blastoderms isolated with the YSL from the blastula embryos (stage 10) developed into embryo-like structures with apparent body axes and contained differentiated organs, such as the eye, ear, contractile heart, yolk sac-like sphere and posterior body trunk with notochord. The proportion of such explants increased as the developmental stage proceeded. However, the proportion was much smaller when blastoderms were isolated at the blastula stage without the YSL. These results suggest that the YSL is essential for the development of embryonic structures. At stage 12 (early gastrula), the frequency of formation of such structures was the same among blastoderms with or without the YSL, so that these embryos are apparently committed for pattern formation.     

Multiple sequence alignment by parallel simulated annealing

We have developed simulated annealing algorithms to solve theproblem of multiple sequence alignment. The algorithm wns shownto give the optimal solution as confirmed by the rigorous dynamicprogramming algorithm for three-sequence alignment. To overcomelong execution times for simulated annealing, we utilized aparallel computer. A sequential algorithm, a simple parallelalgorithm and the temperature parallel algorithm were testedon a problem. The results were compared with the result obtainedby a conventional tree-based algorithm where alignments weremerged by two-' dynamic programming. Every annealing algorithmproduced a better energy value than the conventional algorithm.The best energy value, which probably represents the optimalsolution, wns reached within a reasonable time by both of theparallel annealing algorithms. We consider the temperature parallelalgorithm of simulated annealing to be the most suitable forfinding the optimal multiple sequence alignment because thealgorithm does not require any scheduling for optimization.The algorithm is also usefiui for refining multiple alignmentsobtained by other hewistic methods.     

Effect of floral organ sizes on female reproductive success in<Emphasis Type="Italic">Erythronium japonicum</Emphasis> (Liliaceae)

We examined the effects of floral organ size on female reproductive success in self-incompatibleErythronium japonicum. We measured tepal size and anther-stigma separation and investigated the relationship between these measurements and fruiting and seeding success. We found that tepal length was positively correlated with fruiting success and the number of seeds per fruit This suggests that pollinator attraction is affected by tepal length in f.japonicum and that the number of pollinator visits affects female reproductive success. Anther-stigma separation was the most variable floral trait measured and was not correlated with either fruiting or seeding success in this species, suggesting that the proportion of outcross pollen deposited on stigmas by pollinators does not increase with anther-stigma separation. This is inconsistent with a previous report onEtyyhtonium. grandiflorum. Pollinator size might explain this interspecific difference in the effect of anther-stigma separation on female reproductive success.     

Chromosome mapping of MHC class I in rainbow trout (Oncorhynchus mykiss)

The major histocompatibility complex (MHC) is well-studied in mammals. Much research has addressed the genomic organisation of MHC genes and it is well established that human MHC class I genes are located on chromosome 6. However, information on the organisation of the MHC complex in rainbow trout is only beginning to become available. In the present study it was determined that rainbow trout MHC class I sequences are located on chromosome 18. This is the first reported use of fluorescence in situ hybridisation (FISH) to identify the chromosomal location of genes involved in the immune system of fish.     

Influence of isoflurane concentration and hypoxia on functional magnetic resonance imaging for the detection of bicuculline-induced neuronal activation

Our present study shows that transient changes in relative cerebral blood volume (rCBV) induced by stimulation with bicuculline under six different conditions of anesthesia can be detected with high spatial resolution functional magnetic resonance imaging (fMRI). Bicuculline was administered at a low dose to induce neural activation, and no seizure activity was noted. Of the six conditions, the maximal reaction to bicuculline was observed under 1.5% isoflurane in 60% nitrogen and 40% oxygen. Our results imply that the rCBV changes under this level of isoflurane anesthesia with body temperature maintained at 37.5 degrees C are probably suitable for further fMRI studies.     

Development and characterization of 24 polymorphic microsatellite markers for bluegill sunfish, <Emphasis Type="Italic">Lepomis macrochirus</Emphasis>

The bluegill sunfish (Lepomis macrochirus) is a notorious exotic fish, which causes serious damages to freshwater ecosystems in the world. Therefore, it is a matter of urgency to control this species for the conservation of ecosystem. New microsatellite loci were developed for L. macrochirus using high throughput 454 GS-FLX+ pyrosequencing. We selected 24 primer pairs that were tested on 47 individuals from Lake Biwa, which encompasses the oldest and largest population of L. macrochirus in Japan. Allele per locus varied 2–17. Expected and observed heterozygosities ranged from 0.185 to 0.888 and 0.120 to 0.872, respectively. Applied to L. macrochirus in Lake Biwa, these markers could detect genetic differentiation among populations, which showed a pattern of isolation by distance. These markers are expected to be useful in the estimation of population structure and ecology of L. macrochirus.     

Two novel gene mutations (Glu174→Lys,Phe383→Tyr) causing the “hepatic” form of carnitine palmitoyltransferase II deficiency

Carnitine palmitoyltransferase II (CPT II) deficiency has two different clinical forms, one with “hepatic” and the other with “muscular” symptoms. We studied the molecular basis of the “hepatic” form in two Japanese siblings. Their CPT II activity in lymphoblasts was reduced to 3% of the level observed in normal controls. cDNA analysis showed that the proband was a compound heterozygote. One allele carried a new mutation, G621→A (Glu174→Lys). The other carried three single-base substitutions; a new mutation, T1249→A (Phe383→Tyr), and two previously reported polymorphisms. The brother had the same four substitutions. Neither of the two new mutations in this study was detected in the 60 alleles of 30 Japanese control subjects. Secondary structure prediction analysis of the mutated CPT II protein was different from that of the normal protein. We concluded that these mutations caused the “hepatic” form of CPT II deficiency in the probands. Received: 16 October 1995