Genomics to detect transmission of livestock-associated methicillin-resistant Staphylococcus aureus from UK pigs in abattoirs during slaughter.

BACKGROUND: Livestock-associated MRSA (LA-MRSA) transmission/cross-contamination can occur at abattoir through colonized pigs, increasing occupational hazards and health concerns for workers. To assess this risk we used genomics to identify LA-MRSA lineages present in batches of pigs sent to slaughter and distribution of clones. METHODS: WGS was performed on 85 LA-MRSA previously isolated from six abattoirs from 105 batches of pigs sent from 100 UK farms. spa typing and MLST were performed on all isolates. A mashtree tree was constructed to compare genomes of the LA-MRSA with 1281 global isolates from livestock and humans. A phylogenetic tree and pairwise SNP distance matrices were built from whole genomes of 109 isolates closest to those from abattoirs to compare evolutionary relationships and identify clones. RESULTS: All abattoir isolates belonged to CC398 and were mainly of spa type t011, although other spa types were present. Phylogenetic analysis confirmed the abattoir isolates were most closely related to each other and to pig LA-MRSA from across Europe, indicating a common evolutionary origin with related lineages colonizing UK pigs.Comparison of genomes using SNPs suggested between one and four clones were transferring between pigs from different batches. Transmission likely occurred on farm premises, during transportation, and/or within abattoirs through contact with contaminated surfaces in lairage or post-stunning. CONCLUSIONS: Genomics forensically identified related isolates/clones circulating in pigs at slaughter, showing contamination occurs often. Results suggest that further genomic tracking will identify hotspots, and improvements in measures such as biosecurity and disinfection will help reduce risk for workers.

Improving the laboratory diagnosis of pyruvate kinase deficiency.

Pyruvate kinase (PK) deficiency is an autosomal recessive disease caused by mutations in the PKLR gene, which reduce erythrocyte PK enzyme activity and result in decreased energy synthesis in red cells, causing haemolytic anaemia. Historically, the investigation into pyruvate kinase deficiency (PKD) has been led by a red cell enzyme assay determining PK enzyme activity per unit of haemoglobin. For our laboratory, the reference range was set by Beutler et al. in 1977 when the test was first established. The introduction of genetic testing permitted the creation of reference sample datasets, with positive controls having two pathogenic variants causing disease. This permitted re-assessment of the enzyme assay's sensitivity and specificity, and was used to reassess the reference range of the enzyme assay. Using sequenced samples, we have devised an enzyme assay, DNA testing workflow, which minimises false negative/positive results and improves the diagnostic efficiency. This combined enzyme-DNA testing strategy should improve the diagnostic accuracy whilst limiting the number of expensive DNA tests. During this evaluation, 10 novel genetic variants were identified and are described.