SnTox1, a Parastagonospora nodorum necrotrophic effector, is a dual-function protein that facilitates infection while protecting from wheat-produced chitinases
Liu, Zhaohui ; Gao, Yuanyuan ; Kim, Yong Min ; Faris, Justin D. ; Shelver, Weilin L. ; Wit, Pierre J.G.M. de; Xu, Steven S. ; Friesen, Timothy L. - \ 2016
New Phytologist 211 (2016)3. - ISSN 0028-646X - p. 1052 - 1064.
Parastagonosopora nodorum - Chitin - Host-selective toxin - Necrotroph - Necrotrophic effector - Programmed cell death (PCD) - Wheat (Triticum aestivum) chitinases
SnTox1 induces programmed cell death and the up-regulation of pathogenesis-related genes including chitinases. Additionally, SnTox1 has structural homology to several plant chitin-binding proteins. Therefore, we evaluated SnTox1 for chitin binding and localization. We transformed an avirulent strain of Parastagonospora nodorum as well as three nonpathogens of wheat (Triticum aestivum), including a necrotrophic pathogen of barley, a hemibiotrophic pathogen of sugar beet and a saprotroph, to evaluate the role of SnTox1 in infection and in protection from wheat chitinases. SnTox1 bound chitin and an SnTox1-green fluorescent fusion protein localized to the mycelial cell wall. Purified SnTox1 induced necrosis in the absence of the pathogen when sprayed on the leaf surface and appeared to remain on the leaf surface while inducing both epidermal and mesophyll cell death. SnTox1 protected the different fungi from chitinase degradation. SnTox1 was sufficient to change the host range of a necrotrophic pathogen but not a hemibiotroph or saprotroph. Collectively, this work shows that SnTox1 probably interacts with a receptor on the outside of the cell to induce cell death to acquire nutrients, but SnTox1 accomplishes a second role in that it protects against one aspect of the defense response, namely the effects of wheat chitinases.
Multiplex immunoassay for persistent organic pollutants in tilapia: comparison of imaging- and flow cytometry-based platforms using spectrally encoded paramagnetic microspheres
Meimaridou, A. ; Haasnoot, W. ; Shelver, W.L. ; Franek, M. ; Nielen, M.W.F. - \ 2013
Food Additives & Contaminants. Pt. A, Chemistry, Analysis, Control, Exposure & Risk Assessment 30 (2013)5. - ISSN 1944-0049 - p. 843 - 852.
polybrominated diphenyl ethers - polycyclic aromatic-hydrocarbons - polychlorinated-biphenyls - dietary-intake - farmed salmon - food - fish - contaminants - pcbs - milk
Recent developments in spectrally encoded microspheres (SEMs)-based technologies provide high multiplexing possibilities. Most SEMs-based assays require a flow cytometer with sophisticated fluidics and optics. A new imaging super-paramagnetic SEMs-based alternative platform transports SEMs with considerably less fluid volume into a measuring chamber. Once there SEMs are held in a monolayer by a magnet. Light-emitting diodes (LEDs) are focused on the chamber to illuminate the SEMs – instead of lasers and they are imaged by a charge-coupled device (CCD) detector, offering a more compact sized, transportable and affordable system. The feasibility of utilising this system to develop a 3-plex SEMs-based imaging immunoassay (IMIA) for the screening of persistent organic pollutants (POPs) was studied. Moreover the performance characteristics of 3-plex IMIA were critically compared with the conventional 3-plex flow cytometric immunoassay (FCIA). Both SEM technologies have potential for the multiplex analysis of polychlorinated biphenyls (PCBs), polybrominated diphenyl ethers (PBDEs) and polycyclic aromatic hydrocarbons (PAHs) in buffer and fish extract with insignificant differences in assay sensitivities. Furthermore, we developed a faster and simpler, modified QuEChERS-like generic POPs extraction from tilapia fillet using sodium hydrogen carbonate as one of the salt additives and dispersive solid-phase extraction (dSPE) as a clean-up. Finally, a preliminary in-house validation using 40 different blank and spiked tilapia fillet samples was performed in both systems and the results obtained were critically compared. The lower-cost imaging SEMs-based system performed similarly to the original flow cytometer and, in combination with the new quicker QuEChERS-like extraction, it has high potential for future rapid screening of POPs in several other sample matrices such as other fish species, vegetable refined oils and environmental samples.
Multiplex Screening of Persistent Organic Pollutants in Fish Using Spectrally Encoded Microspheres
Meimaridou, A. ; Kalachova, K. ; Shelver, W.L. ; Franek, M. ; Pulkrabova, J. ; Haasnoot, W. ; Nielen, M.W.F. - \ 2011
Analytical Chemistry 83 (2011)22. - ISSN 0003-2700 - p. 8696 - 8702.
polybrominated diphenyl ethers - dioxin-like compounds - polycyclic aromatic-hydrocarbons - polychlorinated-biphenyls pcbs - magnetic particle immunoassay - health implications - mass-spectrometry - food samples - contamination - congeners
Persistent organic pollutants (POPs) are environmental and food-related contaminants of global public health concern and known to be carcinogenic and endocrine disruptors. Their monitoring is essential, and an easy-to-use, rapid, and affordable multianalyte screening method with simplified sample preparation can be a valuable tool prior to instrumental analysis. For this purpose, a flow cytometric immunoassay (FCIA), based on a spectrally encoded microbeads technology, was developed for the multiplex detection of polycyclic aromatic hydrocarbons (PAHs), polychlorinated biphenyls (PCBs), and polybrominated diphenyl ethers (BDEs) in buffer and fish extracts. The sensitivities of the assays in the three-plex FCIA format were similar to the individual FCIAs for the marker compounds benzo[a]pyrene (BaP), 3,3',4,4'-tetrachlorobiphenyl (PCB77), and 2,2',4,4'-tetrabromodiphenyl ether (BDE47) in buffer with IC50 values of 0.4, 20, and 2 µg L–1, respectively. Apart from the three markers, we could detect at least 14 other POPs. Extracts of fish with different fat content, prepared with a simplified extraction and cleanup procedure, had an insignificant influence on the overall three-plex FCIA performance, with the exception of some impact on the PAHs detection. The performance of the three-plex FCIA, in combination with the simple extraction procedure, is adequate for regulatory control in accordance with the required limits.
Towards a multiplex flow cytometric immunoassay for Persistent Organic Pollutants in food
Meimaridou, A. ; Mintzas, D. ; Franek, M. ; Shelver, W.L. ; Haasnoot, W. ; Nielen, M.W.F. - \ 2011