title: Comparison of Protein Phosphatase Inhibition Assay with LC-MS/MS for Diagnosis of Microcystin Toxicosis in Veterinary Cases.
abstract: Microcystins are acute hepatotoxins of increasing global concern in drinking and recreational waters and are a major health risk to humans and animals. Produced by cyanobacteria, microcystins inhibit serine/threonine protein phosphatase 1 (PP1). A cost-effective PP1 assay using p-nitrophenyl phosphate was developed to quickly assess water and rumen content samples. Significant inhibition was determined via a linear model, which compared increasing volumes of sample to the log-transformed ratio of the exposed rate over the control rate of PP1 activity. To test the usefulness of this model in diagnostic case investigations, samples from two veterinary cases were tested. In August 2013 fifteen cattle died around two ponds in Kentucky. While one pond and three tested rumen contents had significant PP1 inhibition and detectable levels of microcystin-LR, the other pond did not. In August 2013, a dog became fatally ill after swimming in Clear Lake, California. Lake water samples collected one and four weeks after the dog presented with clinical signs inhibited PP1 activity. Subsequent analysis using liquid chromatography-mass spectrometry (LC-MS/MS) detected microcystin congeners -LR, -LA, -RR and -LF but not -YR. These diagnostic investigations illustrate the advantages of using functional assays in combination with LC-MS/MS.
authors.affiliation: Department of Molecular Biosciences, School of Veterinary Medicine, University of California at Davis, 1089 Veterinary Medicine Drive, 2225 VM3B, Davis, CA 95616, USA. cemoore@ucdavis.edu.
authors.forename: Caroline E
authors.initials: CE
authors.lastname: Moore
authors.affiliation: Department of Molecular Biosciences, School of Veterinary Medicine, University of California at Davis, 1089 Veterinary Medicine Drive, 2225 VM3B, Davis, CA 95616, USA. jjuan8042@gmail.com.
authors.forename: Jeanette
authors.initials: J
authors.lastname: Juan
authors.affiliation: Department of Molecular Biosciences, School of Veterinary Medicine, University of California at Davis, 1089 Veterinary Medicine Drive, 2225 VM3B, Davis, CA 95616, USA. yaplin@ucdavis.edu.
authors.forename: Yanping
authors.initials: Y
authors.lastname: Lin
authors.affiliation: Veterinary Diagnostic Laboratory, University of Kentucky, Lexington, KY 40511, USA. cynthia.gaskill@uky.edu.
authors.forename: Cynthia L
authors.initials: CL
authors.lastname: Gaskill
authors.affiliation: Department of Molecular Biosciences, School of Veterinary Medicine, University of California at Davis, 1089 Veterinary Medicine Drive, 2225 VM3B, Davis, CA 95616, USA. bpuschner@ucdavis.edu.
authors.forename: Birgit
authors.initials: B
authors.lastname: Puschner
mesh_terms: D000818:Animals; D002140:California; D002417:Cattle; D002853:Chromatography, Liquid / Q000379:methods*; D004285:Dogs; D005618:Fresh Water; D007629:Kentucky; D016014:Linear Models; D052998:Microcystins / Q000633:toxicity*; D011041:Poisoning / Q000175:diagnosis / Q000662:veterinary; D054645:Protein Phosphatase 1 / Q000037:antagonists & inhibitors*; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D052998:Microcystins; D054645:Protein Phosphatase 1
keywords: blue-green algae; cyanobacteria; diagnosis; microcystins; protein phosphatase inhibition assay; veterinary toxicology