Molecular Evolution of the Duplicated Amy Locus in the Drosophila Melanogaster Species Subgroup: Concerted Evolution Only in the Coding Region and an Excess of Nonsynonymous Substitutions in Speciation

From the analysis of restriction maps of the Amy region in eight sibling species belonging to the Drosophila melanogaster species subgroup, we herein show that the patterns of duplication of the Amy gene are almost the same in all species. This indicates that duplication occurred before speciation within this species subgroup. From the nucleotide sequence data, we show a strong within-species similarity between the duplicated loci in the Amy coding region. This is in contrast to a strong similarity in the 5' and 3' flanking regions within each locus (proximal or distal) throughout the species subgroup. This means that concerted evolution occurred only in the Amy coding region and that differentiated evolution between the duplication occurred in the flanking regions. Moreover, when comparing the species, we also found a significant excess of nonsynonymous substitutions. In particular, all the fixed substitutions specific to D. erecta were found to be nonsynonymous. We thus conclude that adaptive protein evolution occurred in the lineage of D. erecta that is a ``specialist' species for host plants and probably also occurs in the process of speciation in general.     

The molecular analysis ofbrown eye color mutations isolated from geographically discrete populations ofDrosophila melanogaster

A large proportion of spontaneous mutations inDrosophila melanogaster strains of laboratory origin are associated with insertions of mobile DNA elements. As a first step toward determining whether spontaneous laboratory mutations are predictive for mutational events occurring in the wild, recessivebrown (bw) eye color mutants were isolated. By inbreeding the progeny of wild-caughtDrosophila melanogaster females,bw mutations were isolated from seven separate geographic sites distributed among Japan, California, Siberia and Hungary. Among a total of 14 mutations studied, no case of transposon mutagenesis was found. At least 4 mutations are associated with small deletions in thebw gene. The remainder are inseparable from wild-typebw by Southern analysis and are presumed to be basepair changes or very small indels. Although only two spontaneousbw mutants of laboratory origin have been analyzed molecularly, one is a mobile element insertion.     

Detection of dilute antigen in large sample volumes using antibody-coated polyester cloth

Summary A model antibody, goat anti-rabbit IgG antibody, was adsorbed onto a disk of polyester cloth and then fixed into a column apparatus. The macroporosity of the cloth allowed rapid immunoconcentration of a model antigen, rabbit IgG, by passing a large volume of the dilute antigen through the antibody-coated cloth. Such immunoconcentration permitted detection of the dilute antigen which otherwise would have gone undetected.     

Detection of human blood by cloth-based enzyme immunoassay of immunoglobulin G

A specific and sensitive assay for the detection of human blood was developed using polyester cloth coated with goat anti-human IgG antibody to capture human IgG, an abundant and stable protein in blood. The captured IgG was detected by the reaction between goat anti-human IgG antibody-peroxidase conjugate and a chromogenic peroxidase substrate. Because the assay is simple and rapid, and permits simultaneous analysis of multiple samples, it has the potential to be used as a forensic test for human blood.     

Formation of porphyrin pi cation radical in zinc-substituted horseradish peroxidase

Zinc-substituted horseradish peroxidase is oxidized by K2IrCl6 to a characteristic state which retains one oxidizing equivalent more than the zinc peroxidase. The oxidized enzyme gives an optical absorption spectrum similar to that of compound I of peroxidase and catalase, and a g = 2 electron paramagnetic resonance signal which has an intensity corresponding to the porphyrin content. It is reduced back to the zinc peroxidase by a stoichiometric amount of ferrocyanide or by a large excess of K3IrCl6. From the equilibrium data, the value of E0' for the zinc peroxidase couple is estimated to be 0.74 V at pH 6. The oxidized zinc peroxidase is also formed by the addition of H2O2 or upon illumination with white light. The rate constants for the oxidation by K2IrCl6 and H2O2 at pH 8.0 are 8 x 10(5) and 8 x 10(2) M-1 s-1, respectively. No essential spectral change can be observed when K2IrCl6 is added to the metal-free peroxidase (protoporphyrin--apoperoxidase complex) or to zinc-substituted sperm whale myoglobin.     

Test for Selection on Polymorphic Isozyme Genes Using the Population Cage Method

Published experimental data of several species of Drosophila using the population cage method were reexamined. The authors of these works have claimed that selection operates on polymorphic isozyme genes. Since selection coefficients (s) estimated by us for the above data, assuming overdominance, or for some cases, additivity between alleles, were too large (s > 0.1), we conclude that these experimental results do not reflect the effects of the single loci in question; rather, we suggest that careful experimentation and judgment are required for testing with the population cage method whether or not selection actually operates on single genes.     

Random screening of promoters from Escherichia coli and classification based on the promoter strength

Five hundred fifty DNA fragments 100-500 base pairs in length were cloned from total chromosomal DNA of Escherichia coli, each capable of promoting the synthesis of beta-lactamase when inserted upstream of the ampC structural gene without its own promoter in a promoter-probe plasmid. All clones in this library of putative promoters were classified based on the level of resistance to ampicillin, which ranged from 10 to more than 1,500 micrograms/ml. Most of the higher levels of drug resistance (more than 1,000 micrograms/ml) were due not only to an increase in gene expression but also to an increase in plasmid copy number. The DNA fragments which produced the highest level of drug resistance all mapped at 5.7 min on the E. coli chromosome and shared the same nucleotide sequence. In these fragments, a strong promoter was found, which carries an up stream AT-rich sequence in addition to -35 and -10 signals of the promoter consensus.     

Herpangina surveillance in Japan, 1982-1989. A report of the national epidemiological surveillance of infectious agents in Japan.

In Japan, herpangina cases increase every summer with an incidence peak in July. During the past eight years from January, 1982 to December, 1989, a total of 3,974 viruses from herpangina cases were reported from 47 participating laboratories, and coxsackie A (CA) viruses accounted for most of them, 3,055 (76.9%). The major types associated with herpangina were, in order of frequency, CA4, 10, 5, 6, 2 and 3, representing 87.2%-80.8% of total isolations reported for each respective type in this period. Eighty-four point four percent of the total virus isolations were from children at four years of age or younger. More than 80% of total CA virus isolations were from nasopharyngeal specimens and nearly 90% of the viruses were isolated in mice.     

Poliovirus surveillance: isolation of polioviruses in Japan, 1980-1991. A report of the National Epidemiological Surveillance of Infectious Agents in Japan.

This report presents an overall distribution of poliovirus isolations in Japan, where poliomyelitis has been under control over two decades as a result of legal administration of two doses of the trivalent live oral poliovirus vaccine of the Sabin strains (OPV) to children under 48 months of age. During the past 12 years from 1980 through 1991, a total of 1,126 poliovirus isolations from humans and 268 isolations from sewage/river water were reported by respectively 49 and nine of the participating laboratories. Type 2 was most frequently isolated from children after administration of one dose of OPV, followed by type 1 and type 3. On the contrary, after the second dose of OPV, the rate of isolation of type 3 exceeded those of type 2 and type 1. Seasonal and age distribution of poliovirus isolations from both humans and sewage/river water paralleled the OPV vaccination schedule in Japan. One percent of the isolations were, however, from infants younger than the vaccination-scheduled ages and 5% were from children older than those ages, including one each from 15 and 16 years olds. The data indicate that the poliovirus has silently been disseminated from vaccinated children to others and the community, thus suggesting repeated transmission of the viruses. The fact that some elder children had poliovirus colonization in their alimentary tracts indicates a potential risk of infection of such a population when exposed to a wild virus and of becoming a source of transmission to others.     

Thiobacillus novellus cytochrome c oxidase contains one heme alpha molecule and one copper atom per catalytic unit.

The minimal structural unit of cytochrome c oxidase purified from Thiobacillus novellus was composed of one molecule each of two subunits with molecular masses of 32 and 23 kDa, respectively, and the unit had one molecule of heme a and one atom of copper. In the presence of n-octyl-beta-D-thioglucoside, the oxidase existed as the monomeric form of the unit, while it occurred as the dimeric form of the unit in the presence of Tween 20. The monomeric form showed an active cytochrome c oxidizing activity and reduced molecular oxygen to water with ferrocytochrome c. Namely, it has been shown that the bacterial cytochrome c oxidase with one heme a molecule and one copper atom per molecule can catalyze oxidation of ferrocytochrome c with concomitant reduction of molecular oxygen to water.