Resistance Mechanism in a Terbinafine-Resistant Strain of <Emphasis Type="Italic">Microsporum canis</Emphasis>

To clarify the terbinafine (TRF) resistance mechanism in a TRF-resistant strain of Microsporum canis, the expression of the pleiotropic drug resistance (PDR1), multidrug resistance (MDR1), MDR2 and MDR4 genes were investigated by real-time quantitative PCR (RT-qPCR) analysis, given the known interaction of the corresponding proteins with antifungals and with the efflux blocker FK506. The expression of the PDR1, MDR1, MDR2 and MDR4 genes was 2–4 times higher in the TRF-resistant strain grown in the presence of 0.14 µg/mL of TRF than in TRF-susceptible strains cultured in the absence of TRF. The TRF-resistant strain exhibited MICs of > 32 µg/mL for TRF alone; this resistance was attenuated to an MIC of 8 µg/mL in the presence of FK506, indicating that the TRF inhibitory concentration index value was < 0.75. The additive effect of the efflux blocker FK506 on TRF resistance was detected in the TRF-resistant strain. These results indicated that the TRF resistance in this strain reflects overexpression of genes encoding ABC transporter proteins.     

The endosymbiont <Emphasis Type="Italic">Wolbachia</Emphasis> in Eurasian populations of <Emphasis Type="Italic">Drosophila melanogaster</Emphasis>

The endosymbiotic α-proteobacteria Wolbachia is widely spread among arthropods and Filariidae nematodes. This bacterium is transmitted vertically via a transovarian route. Wolbachia is a cause of several reproductive abnormalities in the host species. We analyzed the isofemale lines created using flies collected from Drosophila melanogaster natural populations for infection with the endosymbiont Wolbachia. Wolbachia were genotyped according to five variable markers: the presence of insertion sequence IS5 in two loci, the copy number of two minisatellite repeats, and an inversion. Overall, 665 isofemale lines isolated from the populations of D. melanogaster from Ukraine, Belarus, Moldova, Caucasus, Central Asia, Ural, Udmurtia, Altai, West and East Siberia, and Far East in 1974 through 2005 were used in the work. The samples from Ukrainian, Altaian, and Middle Asian populations were largest. The infection rate of D. melanogaster populations from Middle Asia, Altaian, and Eastern Europe (Ukraine, Moldavia, and Belarus) with Wolbachia amounted to 64, 56, and 39%, respectively. The D. melanogaster population from the Caucasus displayed heterogeneity in the genotypes of this cytoplasmic infection. The Wolbachia genotype wMel, detected in all the populations studied, was the most abundant. The genotype wMelCS2 was always present in the populations from Middle Asia and Altai and was among the rare variants in the D. melanogaster populations from the Eastern Europe. Single instances of the Wolbachia genotype wMelCS occurred in a few flies from the Central Asian and Altai populations, but was not found this genotype in the other regions.     

Specifics of anaphase chromatid segregation in <Emphasis Type="Italic">Drosophila melanogaster</Emphasis> mitotic mutants

Anaphase chromatid segregation defects (CSDs) were quantitatively and qualitatively studied in neural ganglion cells of third-instar larvae of several control wild type Drosophila melanogaster strains and four strains with mutations of the aar ^v158 , ff3, mast ^v40 , and CycB ^2g cell cycle genes. A linear specificity was observed for the CSD frequency, type, determination, and correction probability. The probability of anaphase CSD correction was close to unity in the control strains and lower in the mutant strains. The lower correction probability in the mutant strains was explained in the context of two findings, that the mutations induced the CSDs that were atypical of the wild type strains and were potentially uncorrectable in anaphase and that the mutations negatively affected the relative anaphase time in mitosis.     

Molecular and cytogenic analysis of pericentromeric heterochromatin in ovarian nurse cells of <Emphasis Type="Italic">Drosophila melanogaster</Emphasis> subgroup species

Evolutionary rearrangements of pericentromeric heterochromatin among Drosophila melanogaster subgroup species have been investigated. A region-specific DNA library from Drosophila orena ovarian nurse cell chromocenter was obtained by the microdissection of polythene chromosomes. The probe has been localized on chromosomes of ovarian nurse cells of Drosophila melanogaster subgroup species using fluorescent hybridization in situ. Sequences homologous to the sequences of the DNA probe were detected in the chromocenter and pericentromeric regions of D. orena polythene chromosomes, in all pericentromeric regions of other species with several exceptions. There was no labeling on one of the arms of the D. simulans chromosome 2; however, these sequences were present on the telomere of D. erecta chromosome 3 and in regions adjacent to the brightly DAPI-stained heterochromatin blocks of D. yakuba, D. santomea and D. teissieri chromosomes 2 and 3. At the S₆ stage (secondary reticulate nucleus), labeled chromatin can be found mostly within a restricted territory in D. orena nucleus; no such chromatin can be detected throughout the rest of the nucleus. On the contrary, at this stage, in nuclei of other species, labeled DNA is spread diffusely.     

Polymorphism of <Emphasis Type="Italic">yellow</Emphasis> locus in <Emphasis Type="Italic">Drosophila melanogaster</Emphasis> mutants from natural populations

We have studied the molecular characteristics of the yellow locus (y; 1–0.0), which determines the body color of phenotypically wild-type and mutant alleles isolated in different years from geographically distant populations of Drosophila melanogaster. According to the Southern blot, data restriction maps of the yellow locus of all examined strains differ from one another, as well as from Oregon stock. FISH analysis shows that, in the neighborhood of the yellow locus in the X chromosome, neither P nor hobo elements are found in y^1–775 stock, while only hobo is found in these region in y^1–859 and y^1–866 stocks, only the P element is found in y⁺sn⁸⁴⁹ stock, and both elements are found in y^1–719 stock. Thus, all yellow mutants studied are of independent origin. Locus yellow located on the end of X chromosome (region 1A5–8 on the cytologic map) carries significantly more transposon than retrotransposon induced mutations compared to the white locus (region 3C2). It is possible that, at the ends of Drosophila melanogaster chromosomes, transposons are more active than retrotransposons.     

The Effects of Stress-Related Hormones on the Stress Resistance in <Emphasis Type="Italic">Drosophila melanogaster</Emphasis> Carrying Mutation in the <Emphasis Type="Italic">Dilp6</Emphasis> Gene

The heat stress resistance of Drosophila melanogaster females carrying a hypomorphic mutation of the DILP6 insulin-like protein gene (dilp6 ⁴¹ ) under a change in the level of stress-related hormones (juvenile hormone and octopamine) is studied. It is revealed that the dilp6 ⁴¹ mutation decreases the heat stress resistance of mature D. melanogaster females. An experimental decrease in the level of juvenile hormone is shown to restore the stress resistance of mutant females to the level of stress resistance observed in wild type Canton S females. These data suggest that the effects of the dilp6 ⁴¹ mutation on the stress resistance of females are mediated by an increased level of juvenile hormone. An experimental increase in the octopamine level that causes an increase in juvenile hormone level supports this hypothesis: the resistance to heat stress decreases in females of both lines and this decrease is more significant in mutant females than in the control line. Thus, it is established for the first time that the effect of the hypomorphic dilp6 gene mutation on the heat stress resistance of D. melanogaster females is mediated by juvenile hormone.     

Heterochromatic DNA repeats in <Emphasis Type="Italic">Drosophila</Emphasis> and unusual gene silencing in yeast cells

The 1.25-kb heterochromatic Stellate repeats of Drosophila melanogaster are capable of stably persisting in transgenic constructs and silencing the white reporter gene (mosaic position effect variegation). This system reveals an unusual form of silencing, which is insensitive to known modifiers of position effect variegation. The unusual form of silencing was studied with yeast Saccharomyces cerevisiae, a simple eukaryotic model. To be transferred into yeast cells, the D. melanogaster Stellate repeats were cloned in the pYAC4 centromeric vector (CEN4, URA3, TRP1, HIS3). The HIS3 and/or URA3 genes could be inactive in plasmids consisting of pYAC4 and the Stellate insert in yeast cells. Deletion of D. melanogaster DNA from the plasmid was found to activate the URA3 and HIS3 genes. It was assumed that the genes were repressed rather than damaged in the presence of the Stellate repeats and that a new form of gene silencing was revealed in.     

Hypomorphic mutation of the <Emphasis Type="Italic">dilp6</Emphasis> gene increases DILP3 expression in insulin-producing cells of <Emphasis Type="Italic">Drosophila melanogaster</Emphasis>

The effect of strong hypomorphic mutation of the insulin-like protein DILP6 (dilp6), synthesized in the fat body, on the expression intensity of insulin-like protein DILP3 synthesized in median neurosecretory cells (MNCs) of D. melanogaster brain was examined. The intensity of DILP3 expression in the MNCs of the larvae was evaluated immunohistochemically using antibodies against DILP3 and confocal microscopy. For the first time, it was demonstrated that, in dilp6 ⁴¹ mutants, there was a sharp increase in DILP3 expression level. The data obtained indicate the existence of negative feedback coordinating the expression of insulin-like proteins synthesized in MNCs and peripheral tissues of Drosophila.     

Differential Under-replication of Satellite DNAs during <Emphasis Type="Italic">Drosophila</Emphasis> Development

SATELLITE DNAs are heavily concentrated in the centromeric heterochromatin of metaphase chromosomes^1–3. Satellites and other repeated polynucleotide sequences are under-represented in the polytene, salivary gland cells of Drosophila melanogaster, D. virilis and D. hydei larvae but are fully represented in diploid cells from embryos and imaginal disks^4–6. This under-representation in polytene cells stems from the association of heterochromatin in the chromocentre and the progressive under-replication of the chromocentre during larval development^7,8.     

The endosymbiotic bacterium <Emphasis Type="Italic">Wolbachia</Emphasis> enhances the nonspecific resistance to insect pathogens and alters behavior of <Emphasis Type="Italic">Drosophila melanogaster</Emphasis>

To determine biologically important effects of the cytoplasmic endosymbiont Wolbachia, two substrains of the same Drosophila melanogaster strain have been studied, one of them infected with Wolbachia and the other treated with tetracycline to eliminate the bacterium. Females of D. melanogaster infected with Wolbachia are more resistant to the fungus Blauveria bassiana (an insect pathogen) than uninfected females; infected females also exhibited changes in oviposition substrate preference. Males infected with the bacterium are more competitive than uninfected males. The possible role of Wolbachia in the formation of alternative ecological strategies of D. melanogaster is discussed.     

<Emphasis Type="Italic">OsNHX2</Emphasis>, an Na<Superscript>+</Superscript>/H<Superscript>+</Superscript> antiporter gene,can enhance salt tolerance in rice plants through more effective accumulation of toxic Na<Superscript>+</Superscript> in leaf mesophyll and bundle sheath cells

The Na⁺/H⁺ antiporters play an important role in salt tolerance in plants. However, the functions of OsNHXs in rice except OsNHX1 have not been well studied. Using the gain- and loss-of-function strategies, we studied the potential role of OsNHX2 in salt tolerance in rice. Overexpression of OsNHX2 (OsNHX2-OE) in rice showed the significant tolerance to salt stress than wild-type plants and OsNHX2 knockdown transgenic plants (OsNHX2-KD). Under salt treatments of 300-mM NaCl for 5 days, the plant fresh weights, relative water percentages, shoot heights, Na⁺ contents, K⁺ contents, and K⁺/Na⁺ ratios in leaves of OsNHX2-OE transgenic plants were higher than those in wild-type plants, while no differences were detected in roots. K⁺/Na⁺ ratios in rice leaf mesophyll cells and bundle sheath cells were higher in OsNHX2-OE transgenic plants than in wild-type plants and OsNHX2-KD transgenic plants. Our data indicate that OsNHX2 plays an important role in salt stress based on leaf mesophyll cells and bundle sheath cells and can be served in genetically engineering crop plants with enhanced salt tolerance.