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Saving your bacon: nanopore sequencing of deadly porcine viruses in the Philippines

Amanda Warr

Amanda Warr, The Roslin Institute

Abstract

African Swine Fever Virus (ASFV) is emerging as a serious threat to global pork production. The disease is characterised by haemorrhagic fever leading to up to 100% mortality in domestic pigs and wild boar. There is no treatment or vaccine, and the main strategy for controlling the disease is mass culling and biosecurity perimeters. It is estimated that last year a quarter of the world’s domestic pig population either died of ASFV or were culled to contain it. The virus is a ~180kb DNA virus. Porcine reproductive and respiratory syndrome virus (PRRSV) is a panzootic virus that causes one of the most economically important pig diseases worldwide. It is characterised by respiratory distress, inappetence, and fever, leading to up to 100% mortality in pre-weaned piglets and to death and mummification in utero or abortion in pregnant sows. Treatment is currently limited to stringent biosecurity measures as vaccination only confers partial protection. The virus is a 15kb positive-strand RNA virus, and with nanopore sequencing we can sequence the entire genome in a single read. This allows us not only to track how the virus is spreading but identify quasispecies and recombination events that are missed by methods focussed on the amplification of single open reading frames. Additionally, direct RNA sequencing allows us to characterise the profile of subgenomic RNA, produced during replication, across different viruses. The Philippines are the 8th largest producer of pork in the world, yet 64% of the ~12.5 million domestic animals are farmed in backyard farms. With differing practices between backyard farms and commercial farms, it is important to understand how pathogens spread in order to implement targeted interventions. PRRSV has a huge economic impact in farms around the world and improved surveillance of this virus could reduce the losses Filipino farmers experience and improve the welfare of their animals. ASFV was only introduced to the Philippines towards the end of 2019 and continues to spread throughout the country. If transmission routes are understood, this and potential future outbreaks may be contained better. By bringing the sequencer to the samples we can analyse these viruses locally in the Philippines. Here I will present the methods we have developed to amplify and sequence the genomes of these viruses in the Philippines, including a bioinformatics pipeline that can be run on a standard laptop, to produce highly accurate sequences.

Bio

Amanda Warr recently completed her PhD at The Roslin Institute in Edinburgh, UK. Her PhD research involved using genomics to investigate reproductive traits in pigs and reassembling the pig genome using long-read sequencing. Although this work was primarily in bioinformatics, she also spent time in the lab using the MinION and training others to use the sequencer. She has accumulated a number of MinION-related side projects and collaborations, including work in a variety of species on anti-microbial resistance, viral epidemiology, genome assembly in mammals and microbiomes, and diagnostics. Currently, she is employed as a Postdoctoral Research Fellow at The Roslin Institute with Mick Watson and Christine Tait-Burkard, with main projects focussing on the rumen microbiome, functional genomics in chickens and tracking the spread of porcine reproductive and respiratory syndrome virus in the Philippines.