Investigation of the zinc uptake system of the human fungal pathogen Candida parapsilosis

Candida parapsilosis is the second or third most commonly isolated Candida species from blood cultures and is frequently associated with infections in neonatal intensive care units. Candida species have several virulence factors enabling them to adapt to host environmental conditions and cause infections. These factors include adhesion, biofilm formation, and secretion of hydrolytic enzymes, such as acidic proteinases and lipases. Candida species also obtain heavy metal ions from their environment, such as zinc. Zinc is a cofactor of several proteins and a vital element in cellular mechanisms of the fungi. On the one hand, the host niche represents a zinc-limited environment, that indirectly inhibits microbial growth. In order to survive in such an environment, these pathogens have evolved a zinc transport system that allows them to access bound zinc ions during infection. On the other hand, high zinc ion concentration within the host can also be toxic to microbes e.g. in the phagosomes of Mycobacterium tuberculosis infected macrophages. In case of C. albicans, zinc acquisition processes are intensively studied, but we lack information of the zinc uptake, transfer and homeostasis mechanisms in C. parapsilosis. Here, predicted potential zinc transporters in C. parapsilosis using in silico analyses, generated homozygous knock out mutants and performed their phenotypical characterization by exposing them to various types of stressors and zinc limiting conditions. Furthermore, we analyzed their virulence traits by examining kinetics of fungal cell uptake by macrophages, their killing efficiency and also investigated zinc ion levels in the phagolysosome during in vitro infections

The Potency of Anadara nodifera Shell as Natural Testosterone Booster for Male Canary (Seriunus canaria)

Anadara clam shell contains high Zinc. This microelement is recognised as acting natural aromatase blocker (NAB) that capable of boosting testosterone level. The aim of this study was to evaluates the testosterone level of Canaries following Anadara nodifera administration. Adult male Canaries (n = 27) were group into 3 with 9 repetition of each i.e T0 [control, was given by Carboxymethyl Cellulose Sodium (Na-CMC)], T1 (Zinc Sulphate 0.013 mg/30 g BW), and T2 (NAB 0.3 mg/30 g BW). The T2 treatment was NAB or shell powder of Anadara nodifera which was diluted 1:1 in Na-CMC. The birds were treated orally for 21 days. The complete blood cell count was performed in a haemo analyser except the erythrocyte was calculated manually in the Improved Neubauer counting chamber. Testosterone level was measured in ELISA. Subsequently gen CYP19 aromatase in syrinx, brain and testis were identified employing IHC and descriptively analysed. The result indicated significant elevated level of testosterone (p<0.05). The expression of CYP19 aromatase receptor was depressed in syrinx, brain and testis. However there was no significant changes on the blood profile (p>0.05). To conclude, Anadara nodifera at 0.3 mg/30 g BW was capable of acting as NAB to promote testosterone level. This was confirmed by the reduction of CYP19 aromatase gene receptors in syrinx, brain and testis. However NAB treatment was not significantly affecting routine blood profile.

Cadmium hijacks the high zinc response by binding and activating the HIZR-1 nuclear receptor

Cadmium is an environmental pollutant and significant health hazard that is similar to the physiological metal zinc. In Caenorhabditis elegans, high zinc homeostasis is regulated by the high zinc activated nuclear receptor (HIZR-1) transcription factor. To define relationships between the responses to high zinc and cadmium, we analyzed transcription. Many genes were activated by both high zinc and cadmium, and hizr-1 was necessary for activation of a subset of these genes; in addition, many genes activated by cadmium did not require hizr-1, indicating there are at least two mechanisms of cadmium-regulated transcription. Cadmium directly bound HIZR-1, promoted nuclear accumulation of HIZR-1 in intestinal cells, and activated HIZR-1–mediated transcription via the high zinc activation (HZA) enhancer. Thus, cadmium binding promotes HIZR-1 activity, indicating that cadmium acts as a zinc mimetic to hijack the high zinc response. To elucidate the relationships between high zinc and cadmium detoxification, we analyzed genes that function in three pathways: the pcs-1/phytochelatin pathway strongly promoted cadmium resistance but not high zinc resistance, the hizr-1/HZA pathway strongly promoted high zinc resistance but not cadmium resistance, and the mek-1/sek-1/kinase signaling pathway promoted resistance to high zinc and cadmium. These studies identify resistance pathways that are specific for high zinc and cadmium, as well as a shared pathway.

476 Effect of Holding Period on Viability, Acrosome Status and Intracellular Zinc of Boar Sperm During Liquid Storage

Sperm capacitation results in acrosomal remodeling, increased membrane fluidity, plasma membrane fusability and hyperactivated sperm. Distinct sperm zinc signatures reflect intracellular zinc content, capacitation status, and viability. Therefore, flow cytometric assessment could be a useful tool to evaluate sperm quality and longevity during liquid storage of boar semen. Cooled semen (17°C) from commercial boars (n = 12) were assessed at Day-1, -4 and -7 after collection for motility (IVOS II, Hamilton Thorne, Beverly, MA), intracellular zinc and viability. Samples were stained with 8 µg/mL Hoecsht-33342 (Hoe), 1 µM FluoZinTM-3-AM (Zinc), 10 µM propidium iodide (PI) and 5.6 µg/mL PNA-Alexa647TM (Molecular Probes, Eugene, OR), incubated for 30 min, washed with PBS, and aditionally incubated for 30 min. Acquisition and analysis of 20000 events were performed using Bio-Rad ZE5 Cell Analyzer (Hercules, CA) and FLowJoTM (Ashland, OR). Hoe+ events were gated into Zinc/PI plot to determine viable sperm with high Zinc (Zinc+/PI-). Likewise, Hoe+ events were gated into PI/PNA plot and then into Zinc histogram to detect viable-intact-acrosome with high Zinc (Zn+/PI-/PNA-). Data were analyzed by mixed model for repeated measures, Tukey-adjusted pairwise comparisons and Pearson’s correlation. Viable sperm at Day-1 (86.0±3.1; emmean±SM) and Day-4 (84.7±2.8) showed similar percentage of Zinc+ sperm, and both significantly higher compared to Day-7 (52.2±2.8). Also, viable-intact-acrosome sperm displayed similar percentage of Zinc+ sperm at Day-1 (93.8±3.4) and Day-4 (91.6±3.4) and both higher than sperm at Day-7 (64.5±3.1; P &lt; 0.0001). Percentages of Zinc+ sperm were higher (P &lt; 0.001) when viable sperm had intact acrosome within Day. Percentages of Zinc+ sperm strongly correlated to viable-intact-acrosome sperm (r=0.76, P &lt; 0.0001) and total motility (r=0.65, P &lt; 0.0001). We conclude that sperm having intact plasmatic membrane and acrosome display higher intracellular zinc content which is related to capacitated status, remaining non-capacitated until at least to Day-4 after collection in the tested extender.

Mycobacterial resistance to zinc poisoning requires assembly of P-ATPase-containing membrane metal efflux platforms

Transition metals are toxic at high concentrations. The P1B-ATPase metal exporter CtpC/Rv3270 is required for resistance to zinc poisoning in the human pathogen Mycobacterium tuberculosis. Here, we discovered that zinc resistance also depends on the chaperone-like protein PacL1/Rv3269. PacL1 bound Zn2+, but unlike PacL1 and CtpC, the PacL1 metal-binding motif (MBM) was required only at high zinc concentrations. PacL1 co-localized with CtpC in dynamic microdomains within the mycobacterial plasma membrane. Microdomain formation did not require flotillins nor the PacL1 MBM. Instead, loss of the PacL1 Glutamine/Alanine repeats led to loss of CtpC and sensitivity to zinc. PacL1 and CtpC are within the same operon, and homologous PacL1-P1B-ATPase pairs are widely distributed within and across prokaryotes. PacL1 colocalized and functioned redundantly with PacL orthologs in Mycobacterium tuberculosis. Overall, our study suggests that PacL proteins are scaffolds that assemble P-ATPase-containing metal efflux platforms, a novel type of functional membrane microdomain that underlies bacterial resistance to metal poisoning.

Genotypic and ecological variability of zinc content in the grain of spring bread wheat varieties in the international nursery KASIB

Spring bread wheat is the staple crop in Western Siberia and Kazakhstan, a significant portion of which goes for export. Wheat breeding with a high level of zinc in wheat grain is the most cost-effective and environmentally friendly way to address zinc deficiency in the diet. The purpose of this work was to evaluate the contribution of the factors ‘location’ and ‘genotype’ in the variability of zinc content in wheat grain, and to identify the best varieties as sources of this trait for breeding. The research on screening zinc content in the wheat grain of 49 spring bread wheat varieties from the KazakhstanSiberia Spring Wheat Trial (KASIB) nursery was carried out at 4 sites in Russia (Chelyabinsk, Omsk, Tyumen, Novosibirsk) and 2 sites in Kazakhstan (Karabalyk and Shortandy) in 2017–2018. The content of zinc in wheat grain was evaluated at the Ionomic Facility of University of Nottingham in the framework of the EU project European Plant Phenotyping Network-2020. The analysis of variance showed that the main contribution into the general phenotypic variation of the studied trait, 38.7 %, was made by the factor ‘location’ due to different contents of zinc and moisture in the soil of trial sites; the effect of the factor ‘year’ was 13.5 %, and the effect of the factor ‘genotype’ was 8.0 %. The most favorable environmental conditions for accumulation of zinc in wheat grain were observed in the Omsk region. In Omsk, the average zinc content in all studied varieties was 50.4 mg/kg, with 63.7 mg/kg in the best variety ‘OmGAU 100’. These values are higher than the target values of the international program Harvest Plus. ‘Novosibirskaya 16’ (49.4 mg/kg), ‘Silach’ (48.4 mg/kg), ‘Line 4-10-16’ (47.2 mg/kg), ‘Element 22’ (46.3 mg/kg) and ‘Lutescens 248/01’ (46.0 mg/kg) were identified as being the best varieties. Significant possibilities for the production of wheat grain with high zinc content, which is in demand for the production of bread and pastry products with functional properties, were identified in the Western Siberian region.

Epibrassinolide improves the growth performance of Sedum lineare upon Zn stress through boosting antioxidative capacities

As an essential element, zinc (Zn) can improve or inhibit the growth of plants depending on its concentrations. In this study, the effects of 24-Epibrassinolide (EBR), one well-known steroid phytohormone regulating plant growth and alleviating abiotic stress damage, on morphological parameters and antioxidant capacities of Sedum lineare were investigated under different Zn doses. Compared to plants only exposed to Zn, simultaneously foliar application of 0.75 μM EBR significantly improved multiple morphological characteristics and such growth-improving effects were more significant at high Zn concentrations. At a detrimental 800 μM Zn, EBR benefitted plant growth most prominently, as shown by that the stem length, fresh weight and internode length were increased by 111%, 85% and 157%, respectively; than Zn solely treated plants. EBR spray also enhanced both the activities of antioxidant enzymes such as peroxidase (POD), ascorbate peroxidase (APX), glutathione reductase (GR), and the contents of antioxidative agents including ascorbic acid (AsA) and glutathione (GSH), which in turn decreased the accumulation of reactive oxygen species (ROS) and alleviated the lipid peroxidation in plants. Thus, by demonstrating that EBR could help S. lineare resist high-zinc stress through strengthening the antioxidant system, this work provided a new idea for expanding the planting range of Crassulaceae plants in heavy metal contaminated soil for phytoremediation purpose in the future.