Dataset Information

Large-scale PRNP genotyping of small ruminants using an automated high-throughput MALDI-TOF MS assay.

ABSTRACT: Scrapie resistance or susceptibility in sheep is associated with single nucleotide polymorphisms (SNPs) at codons 136, 154, and 171 of the prion protein gene (PRNP). In addition, phenylalanine mutation at codon 141 has been recognized as a risk factor for atypical scrapie. In contrast, K222, D146, and S146 alleles confer genetic resistance to classical scrapie in goats. High-throughput genotyping technologies would provide significant benefits in scrapie eradication plans. The ability to resolve oligonucleotides varying in mass by less than a single nucleotide makes MALDI-TOF mass spectrometry (MS) a suitable platform for PRNP genotyping. We evaluated the commercial Myriapod scrapie kit (Diatech Pharmacogenetics), associated with a highly automated processing platform incorporating MALDI-TOF MS technology, to detect SNPs at codons 136, 154, 171, 141, and 222 of small ruminant PRNP. The Myriapod scrapie kit was accredited according to UNI CEI EN ISO/IEC 17025. We present the genotyping results of 10,960 sheep in Sicily and 1,822 goats in Sicily and Calabria (southern Italy) tested during 2017. We found a high frequency (43.9%) of the protective ARR allele in sheep and a promising 12.3% of the resistant K222 variant in goats. This efficient and high-throughput method is suitable for extensive PRNP genotyping, as requested in the European scrapie eradication plan.

SUBMITTER: Migliore S

PROVIDER: S-EPMC6857025 | biostudies-literature | 2019 Jul

REPOSITORIES: biostudies-literature

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Large-scale <i>PRNP</i> genotyping of small ruminants using an automated high-throughput MALDI-TOF MS assay.

Migliore Sergio S Bivona Maurizio M Gagliostro Enrico E Buttitta Onofrio O Lo Mascolo Francesca F Macrì Daniele D

Journal of veterinary diagnostic investigation : official publication of the American Association of Veterinary Laboratory Diagnosticians, Inc 20190625 4

Scrapie resistance or susceptibility in sheep is associated with single nucleotide polymorphisms (SNPs) at codons 136, 154, and 171 of the prion protein gene (<i>PRNP</i>). In addition, phenylalanine mutation at codon 141 has been recognized as a risk factor for atypical scrapie. In contrast, K222, D146, and S146 alleles confer genetic resistance to classical scrapie in goats. High-throughput genotyping technologies would provide significant benefits in scrapie eradication plans. The ability to ...[more]

PMID: 31238800

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Project description:MALDI-TOF mass spectrometry (MS) on whole cells enables the detection of different cell types and cell activation. Here, we wondered whether this approach would be useful to investigate the host response in sepsis.Peripheral blood mononuclear cells (PBMCs) from patients with severe sepsis and healthy donors were analyzed with MALDI-TOF MS. PBMCs from healthy donors were also stimulated with lipopolysaccharide, peptidoglycan, CpG oligonucleotides, polyinosinic polycytidylic acid, and with heat-inactivated bacteria. Averaged spectra of PBMCs stimulated in vitro by different agonists were generated from the database using the Biotyper software and matching scores between each spectrum from patients and averaged spectra from the database were calculated.We show that the MALDI-TOF MS signature of PBMCs from septic patients was specific, compared with healthy controls. As the fingerprints observed in patients may be related to PBMC activation, PBMCs from healthy controls were stimulated with cytokines, soluble Pathogen-Associated Molecular Patterns (PAMPs) and heat-killed bacteria, and we created a database of reference spectra. The MALDI-TOF MS profiles of PBMCs from septic patients were then compared with the database. No differences were found between patients with documented infection (n?=?6) and those without bacteriological documentation (n?=?6). The spectra of PBMCs from septic patients matched with those of interferon-?- and interleukin-10-stimulated PBMCs, confirming that sepsis is characterized by both inflammatory and immunoregulatory features. Interestingly, the spectra of PBMCs from septic patients without documented infection matched with the reference spectrum of PBMCs stimulated by CpG oligonucleotides, suggesting a bacterial etiology in these patients.Despite the limits of this preliminary study, these results indicate that the monitoring of functional status of PBMCs in peripheral blood by whole cell MALDI-TOF MS could provide unique opportunities to assess disease progression or resolution in clinical settings.

Dataset Information

Large-scale PRNP genotyping of small ruminants using an automated high-throughput MALDI-TOF MS assay.

Publications

Large-scale <i>PRNP</i> genotyping of small ruminants using an automated high-throughput MALDI-TOF MS assay.

Similar Datasets

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