G protein-coupled receptor kinase function is essential for chemosensation in C. elegans

G protein-coupled receptors (GPCRs) mediate diverse signaling processes, including olfaction. G protein-coupled receptor kinases (GRKs) are important regulators of G protein signal transduction that specifically phosphorylate activated GPCRs to terminate signaling. Despite previously described roles for GRKs in GPCR signal downregulation, animals lacking C. elegans G protein-coupled receptor kinase-2 (Ce-grk-2) function are not hypersensitive to odorants. Instead, decreased Ce-grk-2 function in adult sensory neurons profoundly disrupts chemosensation, based on both behavioral analysis and Ca(2+) imaging. Although mammalian arrestin proteins cooperate with GRKs in receptor desensitization, loss of C. elegans arrestin-1 (arr-1) does not disrupt chemosensation. Either overexpression of the C. elegans Galpha subunit odr-3 or loss of eat-16, which encodes a regulator of G protein signaling (RGS) protein, restores chemosensation in Ce-grk-2 mutants. These results demonstrate that loss of GRK function can lead to reduced GPCR signal transduction and suggest an important role for RGS proteins in the regulation of chemosensation.     

Patterns of testicular cytology in men with primary infertility: any change since the Gulf War?

OBJECTIVE: To evaluate the patterns of testicular cytology in men with primary infertility, to compare the morphologic patterns between the periods 1990-1995, immediately after Gulf War II, and 1997-2001 and to determine whether there is a correlation between hormonal profile, testicular volume and morphologic pattern. STUDY DESIGN: Retrospective study of men with primary infertility. History, complete physical examination, hormonal assay and testicular ultrasound were evaluated. A total of 545 patients had samples for testicular cytology obtained from both testes. The patient's consent was obtained in all cases. Smears were interpreted under light microscopy after treatment with Diff-Quik. A total of 104 healthy, fertile subjects were used for comparison of the hormonal profile and testicular volume. RESULTS: The mean (+/- SD) age was 28.66 +/- 4.36 years and duration of marriage 4.4 +/- 4.36 years. There were 11.2% patients with normal cytology, 55.8% with hypospermatogenesis, 28.4% with testicular atrophy, 2.9% with Sertoli cells only and 1.7% with maturation arrest. A significant increase in hypospermatogenesis and decrease in the Sertoli cell-only pattern were noted in 1997-2001 when compared with 1990-1995. The mean left testicular volume was 10.53 +/- 5.51 mL3 in the infertile group vs. 15.2 +/- 4.97 in the fertile group (p < 0.003); right testicular volume was 10.84 +/- 4.77 vs. 15.15 +/- 5.31 (p < 0.003). The hormonal profile revealed higher luteinizing hormone and follicle-stimulating hormone levels in the infertile group vs. control group (8.53 +/- 9.03 and 16.44 +/- 19.243 vs. 6.98 +/- 4.53 and 7.37 +/- 6.63, respectively [p < 0.001]). Free testosterone was higher in the fertile group (39.69 +/- 12.76 vs. 20.28 +/- 8.5 [p < 0.000]). CONCLUSION: The majority of infertile males in our cohort had hypospermatogenesis; testicular atrophy was the next most common disorder. There was no major change in overall absolute numbers since the Gulf War. Testicular cytology by fine needle aspiration is a safe and well-tolerated complementary investigation for unexplained male infertility.     

Microbial Assemblages in Soil Microbial Succession After Glacial Retreat in Svalbard (High Arctic)

Microbial community composition (cyanobacteria and eukaryotic microalgae abundance and diversity, bacterial abundance, and soil respiration) was studied in subglacial and periglacial habitats on five glaciers near Ny-Alesund, Svalbard (79 degrees N). Soil microbial communities from nonvegetated sites (subglacial, recently deglaciated, and cryoconite sediments) and sites with plant cover (deglaciated some hundreds of years ago) were analyzed. Physicochemical analyses (pH, texture, water content, organic matter, total C and N content) were also performed on the samples. In total, 57 taxa of 23 genera of cyanobacteriaand algae were identified. Algae from the class Chlorophyceae (25 species) and cyanobacteria (23 species) were richest in biodiversity. The numbers of identified species in single habitat types were 23 in subglacial, 39 inbarren, 22 in cryoconite, and 24 in vegetated soils. The highest cyanobacterial and algal biovolume and cell numbers, respectively, were present in cryoconite (13x10(4) microm3 mg-1 soil and 508 cells per mg of soil), followed by barren (5.7x10(4) and 188), vegetated (2.6x10(4) and 120), and subglacial (0.1x10(4) and 5) soils. Cyanobacteria prevailed in all soil samples. Algae (mainly green algae) were present only as accessory organisms. The density of bacteria showed a slightly different trend to that of the cyanobacterial and algal assemblages. The highest number of bacteria was present in vegetated (mean: 13,722x10(8) cells per mg of soil dry wt.), followed by cryoconite (3802x10(8)), barren (654x10(8)), and subglacial (78x10(8)) soils. Response of cyanobacteria and algae to physical parameters showed that soil texture and water content are important for biomass development. In addition, it is shown that nitrogen and water content are the main factors affecting bacterial abundance and overall soil respiration. Redundancy analysis (RDA) with forward selection was used to create a model explaining variability in cyanobacterial, algal, and bacterial abundance. Cryoconites accounted for most of the variation in cyanobacteria and algae biovolume, followed by barren soils. Oscillatoriales, desmids, and green coccoid algae preferred cryoconites, whereas Nostocales and Chroococcales occurred mostly in barren soils. From the data obtained, it is evident that of the studied habitats cryoconite sediments are the most suitable ones for the development of microbial assemblages. Although subglacial sediments do not provide as good conditions as cryoconites, they support the survival of microbial communities. Both mentioned habitats are potential sources for the microbial recolonization of freshly deglaciated soil after the glacier retreat.     

An atypical BAR domain protein in autophagy

The sorting nexin Atg20 interacts with the selective macroautophagy/autophagy scaffolding protein Atg11, suggesting an important role for Atg20 in the initiation of selective autophagy. To explore this possibility, we recently investigated the structure and function of Atg20 using a variety of biophysical and yeast genetic approaches. Our data demonstrate that the BAR domain of Atg20 interacts with Snx4/Atg24 to form an asymmetric heterodimeric BAR domain complex. Atg20 also contains a long intrinsically disordered N terminus that facilitates binding to Atg11 and a large 89-amino acid insertion in its BAR domain, which we have termed the BAR-GAP. This BAR-GAP region is a unique feature of Atg20 and has not been observed in other BAR domains. Furthermore, the BAR-GAP of Atg20 contains an amphipathic helix which is required for membrane binding, tubulation and autophagy. Our findings demonstrate the important role of this novel region in autophagy.     

Benzalkonium chloride tolerance of <Emphasis Type="Italic">Listeria monocytogenes</Emphasis> strains isolated from a meat processing facility is related to presence of plasmid-borne <Emphasis Type="Italic">bcr</Emphasis>ABC cassette

Listeria monocytogenes is a serious foodborne pathogen capable of persisting in food processing environments. Tolerance to disinfectants used in industrial settings constitutes an important factor of Listeria survival. In the present study, the mechanism of tolerance to benzalkonium chloride (BAC) was investigated in 77 L. monocytogenes isolates from a meat facility. By PCR approach, the mdrL and lde chromosomal efflux pump genes were detected in all isolates. No isolate was positive for qacH and emrE genes. However, the bcrABC cassette was present in 17 isolates of serogroup IIa possessing the same AscI/ApaI pulsotype, the operon being localized on a plasmid. The significant relation of BAC tolerance with bcrABC presence was confirmed as all bcrABC positive isolates showed the highest minimal inhibitory concentration (MIC) values for BAC and increased sensitivity to BAC was observed after plasmid curing. No effect of the efflux pump inhibitor reserpine on BAC tolerance in bcrABC positive strains was observed in contrast to all bcrABC negative strains. Lower ethidium bromide efflux in bcrABC positive isolates compared to bcrABC negative and plasmid-cured L. monocytogenes isolates was observed. The expression of bcrABC genes was BAC-induced. The confirmed effect of bcrABC to increased BAC tolerance, coupled with its plasmid location, may be an important factor in potential dissemination of the biocide resistance among Listeria species. The understanding of molecular mechanisms of biocide tolerance should help to improve control measures to prevent further spread of L. monocytogenes in food production environments with frequent use of BAC.     

Holocentric chromosomes may be an apomorphy of Droseraceae

Holocentric chromosomes have evolved in various plant and animal taxa, which suggests they may confer a selective advantage in certain conditions, yet their adaptive potential has scarcely been studied. One of the reasons may reside in our insufficient knowledge of the phylogenetic distribution of holocentric chromosomes across eukaryotic phylogeny. In the present study, we focused on Droseraceae, a carnivorous plant family with an unknown chromosomal structure in monotypic genera Dionaea and Aldrovanda, and a closely related monotypic family Drosophyllaceae. We used flow cytometry to detect holocentric chromosomes by measuring changes in the ratio of the number of G2 nuclei to the number of G1 nuclei in response to gamma irradiation and determined chromosomal structures in Aldrovanda vesiculosa, Dionaea muscipula, Drosera tokaiensis, and Drosera ultramafica from Droseraceae and Drosophyllum lusitanicum from Drosophyllaceae. We confirmed monocentric chromosomes in D. lusitanicum and detected holocentric chromosomes in all four Droseraceae. Our novel finding of holocentric chromosomes in monotypic genera Aldrovanda and Dionaea suggests that all Droseraceae may be holocentric, but to confirm that further research is needed due to previously reported conflicting results in Drosera rotundifolia.     

Effect of chemical modificationin situ onl-glycerol-3-phosphate dehydrogenase in brown adipose tissue mitochondria

Mitochondriall-glycerol-3-phosphate dehydrogenase (E.C. 1.1.99.5.) was studied by chemical modificationin situ with different amino acid side chain specific reagents in mitochondria isolated from hamster brown adipose tissue. The SH-modifying reagents have only slight effect on the enzyme activity. The most effective chemicals were tetranitromethane and diazobenzene sulfonic acid. The enzyme activity can be abolished completely by both of them. In the presence of Ca²⁺ and/or glycerol-3-phosphate inhibition was greater at the same electrophilic reagent concentration. The effect of Ca²⁺ and glycerol-3-phosphate is nonadditive on inhibition by these reagents.     

Determination of ochratoxin A in urine and faeces of swine by high-performance liquid chromatography

Sensitive methods for the determination of ochratoxin A in urine and faeces of swine are described. The samples were extracted with chloroform at pH <2, and the extracts were cleaned up by a combination of solid-phase extraction and liquid—liquid partition. High-performance liquid chromatography with fluorescence detection was used for detection and determination. The detection limits were 0.3 ng/ml for urine and 1.5 ng/g for faeces. Recoveries of ochratoxin A from spiked samples of urine and faeces were 93% and 60%, respectively. Because of the low detection limit and the fast and relatively easy performance, the method for the determination of ochratoxin A in urine proved suitable for the estimation of possible contamination of live animals.     

Factors associated with reed decline in a eutrophic fishpond,Rožmberk (South Bohemia,Czech Republic)

Characteristics of the growth and performance ofPhragmites australis as well as sediment characteristics were investigated along the western shore of Ro?mberk fishpond. The reed performance decreased toward the southern end of the shore, proximate to outlets of wastewater effluent and untreated sewage. While the reed stand was closed and looked healthy at the northern end, gaps occurred within the flooded part of the reed belt further southwards; reed was absent in water along the southernmost part of the shore, though dead shoot stubble indicated its presence in earlier times. In the latter site, the surface layer of sediment consisted of fine mud with a high organic matter content and a high oxygen demand. To a smaller extent, patches of partly decomposed reed litter inside the gaps showed the same properties. It is suggested that organic matter accumulating within the flooded part of the reed belt may have reduced plant performance which ultimately lead to the formation of gaps. At a later stage, the lakeside fringe of the reed belt collapsed, thus completing the retreat of reed from water. A protective effect of calcium against the adverse effects of organic matter is suggested.