Genetics and Genomics Commons^™

Biolistic Transformation Of Oil Palm Using The Phosphomannose Isomerase (Pmi) Gene As The Positive Selectable Marker., Bahariah Bohari

Student Works (2010-2019)

The selectable marker system based on the Escherichia coli phosphomannose isomerase (pmi) gene was adapted for genetic transformation of oil palm. This system makes use of the pmi gene that encodes phosphomannose isomerase, which converts mannose-6-phosphate to fructose-6-phosphate and uses mannose as the selection agent. This is to anticipate the future requirement of using non-antibiotic resistance genes for the commercialization of transgenic oil palm. The use of antibiotic or herbicide-based selection systems has caused much public concern due to inadequate knowledge of the agents’ impact on the environment and on human health. In this study, four transformation vectors, namely pMI3, …

A Systematic Study Of Fagraea Sensu Lato And A Revision Of The Peninsular Malaysian Taxa., Sugumaran Manickam

Student Works (2010-2019)

systematic study of the Peninsular Malaysian species of Fagraea sensu lato was undertaken. This was done in the light of recent revisionary work done for Borneo that documented 20 new species and demonstrated that previous species concepts for Fagraea were too broad. Parallel to this the distinction of the subgeneric groups recognised as sections, i.e., Cyrtophyllum, Fagraea and Racemosae, were investigated with molecular phylogenetic methods. Representative taxa from the Malay Peninsula and Borneo augmented by sequences from other taxa in the same subtribe and tribe were used in the molecular analysis. Gene sequences from ITS, trnL–F (trnL intron + trnL–F …

Computational Identification Of Micro Rna In Musa Sp Est Sequences., Tan Yew Seong

Student Works (2010-2019)

MicroRNAs (miRNAs) are small single-stranded non-protein-coding RNAs of about 22 nucleotides that play an important role in post-transcriptional gene regulation in animals and plants by targeting mRNAs for direct cleavage of mRNAs or repression of mRNA translation. In this study, a homology search based approach has been used for identifying the novel miRNA genes of Musa sp. using the BLAST program with a total of 42,978 Musa sp. expressed sequence tags (EST) were compared to a total of 959 previously known plant miRNA sequences. Forty-seven candidate miRNA with homology to previously known miRNAs were identified. By using the mFold3.1 program, …

Isolation And Development Of Microsatellite Markers Of Channa Striata., Faiza Omar

Student Works (2010-2019)

Channa striata, locally known as haruan has long been regarded as a valuable edible fish and it is also consumed for its putative effects on wound healing. The main objective of the project was to develop microsatellite markers for use in the study of genetic diversity of the wild Channa striata populations in Malaysia. In this study, seven new microsatellite loci were isolated from Channa striata using a Random Amplified Microsatellite (RAMs) based technique which was an efficient and reliable method. Ten primers were used in population genetic studies of Channa striata.Sixty sets of microsatellite primer pairs were used in …

High-Density Screening Reveals A Different Spectrum Of Genomic Aberrations In Chronic Lymphocytic Leukemia Patients With ‘Stereotyped’ Ighv3-21 And Ighv4-34 B-Cell Receptors, Millaray Marincevic, Nicola Cahill, Rebeqa Gunnarsson, Anders Isaksson, Mahmoud Mansouri, Hanna Göransson, Markus Rasmussen, Mattias Jansson, Fergus Ryan, Karin Karlsson, Hans-Olov Adami, Fred Davi, Jesper Jurlander, Gunnar Juliusson, Kostas Stamatopoulos, Richard Rosenquist

Articles

Background The existence of multiple subsets of chronic lymphocytic leukemia expressing ‘stereotyped’ Bcell receptors implies the involvement of antigen(s) in leukemogenesis. Studies also indicate that ‘stereotypy’ may influence the clinical course of patients with chronic lymphocytic leukemia, for example, in subsets with stereotyped IGHV3-21 and IGHV4-34 B-cell receptors; however, little is known regarding the genomic profile of patients in these subsets. Design and Methods We applied 250K single nucleotide polymorphism-arrays to study copy-number aberrations and copy-number neutral loss-of-heterozygosity in patients with stereotyped IGHV3-21 (subset #2, n=29), stereotyped IGHV4-34 (subset #4, n=17; subset #16, n=8) and non-subset #2 IGHV3-21 (n=13) and …

Anomalous Constitutive Src Kinase Activity Promotes B Lymphoma Survival And Growth, Jiyuan Ke, R. Lakshman Chelvarajan, Vishal Sindhava, Darrell A. Robertson, Lazaros Lekakis, C. Darrell Jennings, Subbarao Bondada

Microbiology, Immunology, and Molecular Genetics Faculty Publications

Background: Previously we have shown that B cell receptor (BCR) expression and B cell receptor signaling pathways are important for the basal growth of B lymphoma cells. In particular we have shown that the activation of Syk, a non-src family protein tyrosine kinase and the mitogen activated protein kinases (MAPK), ERK and JNK that mediate BCR signals are required for the constitutive growth of B lymphoma cells. Since src family protein tyrosine kinases (SFKs) like Lyn are known to be needed for the phosphorylation of BCR co-receptors, Ig-alpha and Ig-beta, we hypothesized that one or more SFKs will be constitutively …

Genetic Structure Of Yonahlossee Salamander Populations., Joshua Andrew Rudd

Electronic Theses and Dissertations

Plethodon yonahlossee is the largest eastern Plethodontid salamander. It has been classified as a species of greatest conservation need by the Tennessee Wildlife Resources Agency (TWRA). Found only in mountainous areas along the borders of Tennessee, North Carolina, and Virginia, populations of the yonahlossee are considered to be rare and local throughout their range. Genetic differentiation among populations of any species is usually attributable to long-standing, extrinsic barriers to gene flow. Because of their disjunct population structure and some observed morphological variation, genetic differentiation among yonahlossee populations is expected. A genetic structure study of yonahlossee was conducted to identify any …

Temperature Compensation Of Auxin Dependent Developmental Patterning, Nicholas J. Kaplinsky

Biology Faculty Works

The establishment of localized auxin gradients plays a central role in developmental patterning in plants. Auxin levels and responses have been shown to increase with temperature although developmental patterning is not affected. This suggests the existence of a homeostatic mechanism that ensures that patterning occurs normally over a range of temperatures. We recently described the cloning and characterization of BOBBER1 (BOB1), an Arabidopsis gene which encodes a small heat shock protein. BOB1 is required for the establishment of auxin gradients and for normal developmental patterning. BOB1 is also required for organismal thermotolerance and localizes to heat shock granules at elevated …

Pot1 Proteins In Green Algae And Land Plants: Dna-Binding Properties And Evidence Of Co-Evolution With Telomeric Dna, Eugene V. Shakirov, Xiangyu Song, Jessica A. Joseph, Dorothy E. Shippen

Biological Sciences Faculty Research

Telomeric DNA terminates with a single-stranded 3′ G-overhang that in vertebrates and fission yeast is bound by POT1 (Protection Of Telomeres). However, no in vitro telomeric DNA binding is associated with Arabidopsis POT1 paralogs. To further investigate POT1–DNA interaction in plants, we cloned POT1 genes from 11 plant species representing major branches of plant kingdom. Telomeric DNA binding was associated with POT1 proteins from the green alga Ostreococcus lucimarinus and two flowering plants, maize and Asparagus. Site-directed mutagenesis revealed that several residues critical for telomeric DNA recognition in vertebrates are functionally conserved in plant POT1 proteins. However, the plant proteins …

Apolipoprotein E But Not B Is Required For The Formation Of Infectious Hepatitis C Virus Particles, Jieyun Jiang, Guangxiang Luo

Microbiology, Immunology, and Molecular Genetics Faculty Publications

Our previous studies have found that hepatitis C virus (HCV) particles are enriched in apolipoprotein E (apoE) and that apoE is required for HCV infectivity and production. Studies by others, however, suggested that both microsomal transfer protein (MTP) and apoB are important for HCV production. To define the roles of apoB and apoE in the HCV life cycle, we developed a single-cycle HCV growth assay to determine the correlation of HCV assembly with apoB and apoE expression, as well as the influence of MTP inhibitors on the formation of HCV particles. The small interfering RNA (siRNA)-mediated knockdown of apoE expression …

Drosophila Adult Eye Model To Teach Scanning Electron Microscopy In An Undergraduate Cell Biology Laboratory, Meghana Tare, Oorvashi Roy Puli, Sarah M. Oros, Amit Singh

Biology Faculty Publications

We have devised an undergraduate laboratory exercise to study tissue morphology using fruit fly, Drosophila melanogaster, as the model organism. Drosophila can be reared in a cost effective manner in a short period of time. This experiment was a part of the undergraduate curriculum of the cell biology laboratory course aimed to demonstrate the use of scanning electron microscopy (SEM) technique to study the morphology of adult eye of Drosophila. The adult eye of Drosophila is a compound eye, which comprises of 800 unit eyes, and serves as an excellent model for SEM studies. We used flies that …

Detection Of Sub-Clinical Cwd Infection In Conventional Test-Negative Deer Long After Oral Exposure To Urine And Feces From Cwd+ Deer, Nicholas J. Haley, Candace K. Mathiason, Mark D. Zabel, Glenn C. Telling, Edward A Hoover

Microbiology, Immunology, and Molecular Genetics Faculty Publications

BACKGROUND: Chronic wasting disease (CWD) of cervids is a prion disease distinguished by high levels of transmissibility, wherein bodily fluids and excretions are thought to play an important role. Using cervid bioassay and established CWD detection methods, we have previously identified infectious prions in saliva and blood but not urine or feces of CWD+ donors. More recently, we identified very low concentrations of CWD prions in urine of deer by cervid PrP transgenic (Tg[CerPrP]) mouse bioassay and serial protein misfolding cyclic amplification (sPMCA). This finding led us to examine further our initial cervid bioassay experiments using sPMCA.

OBJECTIVES: We sought …

It's Not Too Late For The Harpy Eagle (Harpia Harpyja): High Levels Of Genetic Diversity And Differentiation Can Fuel Conservation Programs, Heather R. Lerner, Jeff A. Johnson, Alec R. Lindsay Ph. D., Lloyd F. Kiff, David P. Mindell

Faculty Works

Background: The harpy eagle (Harpia harpyja) is the largest Neotropical bird of prey and is threatened by human persecution and habitat loss and fragmentation. Current conservation strategies include local education, captive rearing and reintroduction, and protection or creation of trans-national habitat blocks and corridors. Baseline genetic data prior to reintroduction of captive-bred stock is essential for guiding such efforts but has not been gathered previously.

Methodology/Findings: We assessed levels of genetic diversity, population structure and demographic history for harpy eagles using samples collected throughout a large portion of their geographic distribution in Central America (n = 32) and South America …

Bobber1 Is A Noncanonical Arabidopsis Small Heat Shock Protein Required For Both Development And Thermotolerance, Dahlia Erin Perez , '08, J. S. Hoyer, Ayanna Iman Johnson , '09, Zachary Ray Moody , '07, J. Lopez, Nicholas J. Kaplinsky

Biology Faculty Works

Plants have evolved a range of cellular responses to maintain developmental homeostasis and to survive over a range of temperatures. Here, we describe the in vivo and in vitro functions of BOBBER1 (BOB1), a NudC domain containing Arabidopsis (Arabidopsis thaliana) small heat shock protein. BOB1 is an essential gene required for the normal partitioning and patterning of the apical domain of the Arabidopsis embryo. Because BOB1 loss-of-function mutants are embryo lethal, we used a partial loss-of-function allele (bob1-3) to demonstrate that BOB1 is required for organismal thermotolerance and postembryonic development. Recombinant BOB1 protein functions as a molecular chaperone and prevents …

Differential Expression Of Genes During Diapause In The Flesh Fly, Sarcophaga Crassipalpis., Puja Karki

Electronic Theses and Dissertations

The objective of this study was to identify genes that are differentially regulated during diapause when compared with nondiapausing pupae in Sarcophaga crassipalpis. The results of a Suppression Subtractive Hybridization procedure was used to indentify genes that are differentially regulated in both diapause and nondiapausing states while suppressing genes that are common to both states. Randomly picked colonies from both subtractive libraries were isolated and the inserts sequenced. The sequences were analyzed using the bioinformatics tools NCBI, BlastX, Clustal W, etc. Out of 384 clones, 59 genes were found to be upregulated during diapause and 37 genes were found …

Assessment Of The Endangered Species Podarcis Carbonelli On A Microgeographic Scale: A Molecular, Morphological And Physiological Approach, Maria Clara Figueirinhas Do Amaral

Masters Theses & Specialist Projects

The lizard Podarcis carbonelli is an endangered species endemic to the Iberian Peninsula. One location where this species occurs is at the Berlengas Natural Preserve, an Atlantic archipelago off the coast of Portugal. These island populations are geographically separated from nearby mainland populations. The fundamental question is, are these insular individuals distinct from the mainland populations? Four localities were chose for comparison: two island populations and two nearby coastal populations. We assessed this question using three distinct approaches: molecular, morphological and physiological approach. We sequenced the 12S RNA, the mtDNA Control Region and the 7th intron of the !-fibrinogen gene …

Evaluation Of Annotation Performances Between Automated And Curated Databases Of E.Coli Using The Correlation Coefficient, Reddysalilaja Marpuri

Masters Theses & Specialist Projects

This project compared the performance of the correlation coefficient to show similarities in annotations between a predictive automated bacterial annotation database and the curated EcoCyc database. EcoCyc is a conservative multidimensional annotation system that is exclusively based on experimentally validated findings by over 15,000 publications. The automated annotation system, used in the comparison was BASys. It is often used as a first pass annotation tool that tries to add as many annotations as possible by drawing upon over 30 information sources. Gene ontology served as one basis of comparison between these databases because of the limited common terms in the …

Cd19 Signaling Is Impaired In Murine Peritoneal And Splenic B-1 B Lymphocytes, Trivikram Dasu, Vishal Sindhava, Stephen H. Clarke, Subbarao Bondada

Microbiology, Immunology, and Molecular Genetics Faculty Publications

B-1 cells reside predominantly within the coelomic cavities, tonsils, Peyer's patches, spleen (a minor fraction – ∼5%) and are absent in the lymph nodes. They are the primary sources of natural IgM in the body. B-1 cells express polyreactive B cell receptors (BCRs) that cross react with self-antigens and are thus implicated in auto-immune disorders. Previously, we reported that peritoneal B-1 cells are deficient in CD19-mediated intracellular signals leading to Ca²⁺ mobilization. Here, we find that splenic B-1 cells, like peritoneal B-1 cells, are defective in Ca²⁺ release upon B cell activation by co-cross-linking BCR and CD19. In …

Rapid Evolution Of Sex-Pheromone-Producing Enzyme In Drosophila, Troy R. Shirangi, Héloïse D. Dufour, Thomas M. Williams, Sean B. Carroll

Biology Faculty Publications

A wide range of organisms use sex pheromones to communicate with each other and to identify appropriate mating partners. While the evolution of chemical communication has been suggested to cause sexual isolation and speciation, the mechanisms that govern evolutionary transitions in sex pheromone production are poorly understood. Here, we decipher the molecular mechanisms underlying the rapid evolution in the expression of a gene involved in sex pheromone production in Drosophilid flies. Long-chain cuticular hydrocarbons (e.g., dienes) are produced female-specifically, notably via the activity of the desaturase DESAT-F, and are potent pheromones for male courtship behavior in Drosophila melanogaster. We …

Lipid Nanoparticles With Accessible Nickel As A Vaccine Delivery System For Single And Multiple His-Tagged Hiv Antigens, Weili Yan, Anekant Jain, Ronan O'Carra, Jerold Woodward, Wenxue Li, Guanhan Li, Avindra Nath, Russell J. Mumper

Microbiology, Immunology, and Molecular Genetics Faculty Publications

Lipid-based nanoparticles (NPs) with a small amount of surface-chelated nickel (Ni-NPs) were developed to easily formulate the HIV his-tagged Tat protein, as well as to formulate and co-deliver two HIV antigens (his-p24 and his-Nef) on one particle. Female BALB/c mice were immunized by s.c. injection with his-Tat/Ni-NP formulation (1.5 μg Tat-his/mouse) and control formulations on day 0 and 14. The day 28 anti-Tat specific IgG titer with his-Tat/Ni-NP was significantly greater than that with Alum/his-Tat. Furthermore, splenocytes from his-Tat/Ni-NP immunized mice secreted significantly higher IFN-γ than those from mice immunized with Alum/his-Tat. Although Ni-NPs did not show better adjuvant activity …

Dna-Binding By Haemophilus Influenzae And Escherichia Coli Ybab, Members Of A Widely-Distributed Bacterial Protein Family, Anne E. Cooley, Sean P. Riley, Keith Kral, M. Clarke Miller, Edward Demoll, Michael G. Fried, Brian Stevenson

Microbiology, Immunology, and Molecular Genetics Faculty Publications

BACKGROUND: Genes orthologous to the ybaB loci of Escherichia coli and Haemophilus influenzae are widely distributed among eubacteria. Several years ago, the three-dimensional structures of the YbaB orthologs of both E. coli and H. influenzae were determined, revealing a novel "tweezer"-like structure. However, a function for YbaB had remained elusive, with an early study of the H. influenzae ortholog failing to detect DNA-binding activity. Our group recently determined that the Borrelia burgdorferi YbaB ortholog, EbfC, is a DNA-binding protein. To reconcile those results, we assessed the abilities of both the H. influenzae and E. coli YbaB proteins to bind DNA …

Partitioning The Apical Domain Of The Arabidopsis Embryo Requires The Bobber1 Nudc Domain Protein, R. J. Jurkuta, Nicholas J. Kaplinsky, Jennifer Elizabeth Spindel , '10, M. K. Barton

Biology Faculty Works

The apical domain of the embryo is partitioned into distinct regions that will give rise to the cotyledons and the shoot apical meristem. In this article, we describe a novel screen to identify Arabidopsis thaliana embryo arrest mutants that are defective in this partitioning, and we describe the phenotype of one such mutant, bobber1. bobber1 mutants arrest at the globular stage of development, they express the meristem-specific SHOOTMERISTEMLESS gene throughout the top half of the embryo, and they fail to express the AINTEGUMENTA transcript normally found in cotyledons. Thus, BOBBER1 is required to limit the extent of the meristem domain …

Contribution Of Ledgf/P75 To Prostate Cancer Chemoresistance, Melanie Mediavilla-Varela

Loma Linda University Electronic Theses, Dissertations & Projects

Prostate cancer (PCa) is the second leading cause of male cancer death in the United States, and is considered a health disparity because of its disproportionate incidence and mortality in African American (AA) males. A major challenge in PCa treatment is its recurrence into hormone refractory prostate cancer (HRPC). Docetaxel is the standard of care for first line chemotherapy to HRPC. Unfortunately, most patients ultimately manifest resistance to DTX and succumb to the disease. Circumventing DTX resistance in HRPC will require better understanding of the mechanisms by which this drug kills prostate tumor cells, and the genes that promote this …

Studies In Prolonged Oxidative Stress Resistance In Porphyromonas Gingivalis, Rachelle Mckenzie

Loma Linda University Electronic Theses, Dissertations & Projects

Porphyromonas gingivalis is a gram negative, anaerobic bacterium implicated as a major etiological agent in chronic adult periodontitis. In the inflammatory environment of the periodontal pocket, the organism must overcome oxidative stress as a result of exposure to air in the oral cavity as well as to reactive oxygen species (ROS) generated by host immune cells. A comprehensive mechanism for oxidative stress resistance in P. gingivalis is yet to be fully elucidated. In this study we examined the modulation of gene expression under conditions of oxidative stress by whole-genome DNA microarray analysis. Our findings indicated that numerous genes are involved …

Localization And Expression Of The Aer Receptor In Escherichia Coli, Daniel Salcedo

Loma Linda University Electronic Theses, Dissertations & Projects

In Escherichia coli, chemotactic movement towards an energetically-favorable environment is mediated by five transmembrane chemoreceptors. These sensory proteins respond to numerous environmental signals, including amino acids, sugars, and pH. The aerotaxis receptor, Aer, is an intracellular sensor that responds to oxygen concentration and other parameters affecting cellular energy levels. Bacterial chemotaxis receptors form dimeric signaling units that organize into tetramers and hexamers and form large clusters at the cell poles. A cluster arrangement could increase receptor sensitivity by lateral communication between dimers, thus amplifying the signal from one dimer. We have shown that Aer forms dimers, trimer of dimers, …

Characterization Of Conserved Properties Of Hemagglutinin Of H5n1 And Human Influenza Viruses: Possible Consequences For Therapy And Infection Control, Veljko Veljkovic, Nevena Veljkovic, Claude P. Muller, Sybille Müller, Sanja Glisic, Vladimir Perovic, Heinz Köhler

Microbiology, Immunology, and Molecular Genetics Faculty Publications

BACKGROUND: Epidemics caused by highly pathogenic avian influenza virus (HPAIV) are a continuing threat to human health and to the world's economy. The development of approaches, which help to understand the significance of structural changes resulting from the alarming mutational propensity for human-to-human transmission of HPAIV, is of particularly interest. Here we compare informational and structural properties of the hemagglutinin (HA) of H5N1 virus and human influenza virus subtypes, which are important for the receptor/virus interaction.

RESULTS: Presented results revealed that HA proteins encode highly conserved information that differ between influenza virus subtypes H5N1, H1N1, H3N2, H7N7 and defined an …

Borrelia Burgdorferi Ebfc Defines A Newly-Identified, Widespread Family Of Bacterial Dna-Binding Proteins, Sean P. Riley, Tomasz Bykowski, Anne E. Cooley, Logan H. Burns, Kelly Babb, Catherine A. Brissette, Amy Bowman, Matthew L. Rotondi, M. Clarke Miller, Edward Demoll, Kap Lim, Michael G. Fried, Brian Stevenson

Microbiology, Immunology, and Molecular Genetics Faculty Publications

The Lyme disease spirochete, Borrelia burgdorferi, encodes a novel type of DNA-binding protein named EbfC. Orthologs of EbfC are encoded by a wide range of bacterial species, so characterization of the borrelial protein has implications that span the eubacterial kingdom. The present work defines the DNA sequence required for high-affinity binding by EbfC to be the 4 bp broken palindrome GTnAC, where ‘n’ can be any nucleotide. Two high-affinity EbfC-binding sites are located immediately 5′ of B. burgdorferi erp transcriptional promoters, and binding of EbfC was found to alter the conformation of erp promoter DNA. Consensus EbfC-binding …

Altered Regulation Of Aquaporin Gene Expression In Allergen And Il-13-Induced Mouse Models Of Asthma, Carissa M. Krane, Bijia Deng, Venkateshwar Mutyam, Casey A. Mcdonald, Stephen Pazdziorko, Lawrence Mason, Samuel Goldman, Marion Kasaian, Divya Chaudhary, Cara Williams, Melisa W.Y. Ho

Biology Faculty Publications

IL-13 is known to affect many processes that contribute to an asthmatic phenotype, including inflammation, fibrosis, and mucus production. Members of the aquaporin (AQP) family of transmembrane water channels are targets of regulation in models of lung injury and inflammation. Therefore, we examined AQP mRNA and protein expression in allergen and IL-13-induced mouse models of asthma. Lungs from ovalbumin sensitized and ovalbumin challenged (OVA/OVA) and IL-13 treated mice showed airway thickening, increased mucus production, and pulmonary eosinophilia. Pulmonary function tests showed a significant increase in methacholine-induced airway hyperreactivity in OVA/OVA and IL-13-treated mice as compared with controls. Quantitative PCR analysis …

Comparison Of Gene Ontology Term Annotations Between E.Coli K12 Databases, Reddysailaja Marpuri

Student Research Conference Select Presentations

The objective of this project was to get up-to-date functional information on all genes of E.coli K12 strains based on Genome Ontology terms. Gene Ontology is described by a defined library of terms related the biological process, cellular components and molecular functions of a gene in an organism. The genome sequence of an organism gains its value when it is annotated with gene ontology terms, which bridges the gap from the sequence to the biology of an organism. Since we use annotated gene database in the prediction of the function of newly sequenced genes, it is important to have databases …

Detection Of Cwd Prions In Urine And Saliva Of Deer By Transgenic Mouse Bioassay, Nicholas J. Haley, Davis M. Seelig, Mark D. Zabel, Glenn C. Telling, Edward A. Hoover

Microbiology, Immunology, and Molecular Genetics Faculty Publications

Chronic wasting disease (CWD) is a prion disease affecting captive and free-ranging cervids (e.g. deer, elk, and moose). The mechanisms of CWD transmission are poorly understood, though bodily fluids are thought to play an important role. Here we report the presence of infectious prions in the urine and saliva of deer with chronic wasting disease (CWD). Prion infectivity was detected by bioassay of concentrated, dialyzed urine and saliva in transgenic mice expressing the cervid PrP gene (Tg[CerPrP] mice). In addition, PrP(CWD) was detected in pooled and concentrated urine by protein misfolding cyclic amplification (PMCA). The concentration of abnormal prion protein …