Advancing livestock pathogen genomics in the Philippines through whole-genome sequencing of african swine fever virus (Asfivirus haemorrhagiae) and porcine reproductive and respiratory syndrome virus 2 (Betaarterivirus americense) using long-read nanopore sequencing

Date

6-2025

Abstract

African swine fever virus (ASFV) and porcine reproductive and respiratory syndrome virus (PRRSV) are two of the most economically devastating viral pathogens affecting swine production globally. ASFV, a large double-stranded DNA virus, causes acute hemorrhagic disease with near 100% mortality, while PRRSV, a positive-sense RNA virus, leads to reproductive failure and respiratory illness, compounded by rapid viral evolution and limited vaccine cross-protection. In the Philippines, both viruses have caused major outbreaks, yet genomic data–essential for surveillance, epidemiology, and control strategies–have remained scarce. This dissertation presents the first comprehensive applications of long-read sequencing technologies for ASFV and PRRSV genomic surveillance in the country, addressing critical gaps in local pathogen genomics by providing valuable baseline data for both research and disease management and by establishing protocols for rapid, field-compatible analysis.

The first study reports the first locally sequenced and assembled coding-complete genome f ASFV from a 2021 outbreak in Pangasinan. Using Oxford Nanopore Technologies' (ONT) MinION platform and host DNA depletion, a 192,377 bp genome was assembled and shown to cluster With genotype I strains circulating in Asia and Europe. This work demonstrated the feasibility of portable long-read sequencing for ASFV whole-genome recovery under field-relevant conditions.

The second study expanded this approach by sequencing ten ASFV genomes from outbreak samples collected between 2021 and 2023 across Luzon, Visayas, and Mindanao, Using a multiplexed, tiled amplicon method, all strains were confirmed as genotype I1, serogroup 8, and closely related to the Georgia 2007/1 reference. While most genomes were highly conserved, single nucleotide polymorphisms and a notable 1.9 kb deletion were identified in select strains. Multi-locus genotyping across 13 viral markers provided high-resolution insights into the genetic diversity and potential microevolution of ASFV in the Philippines. This study is among the first to use region-wide ASFV genome profiling to inform molecular surveillance in the country.

In the third study, two coding-complete genomes of PRRSV-2 from field samples collected in 2021 and 2023 were sequenced. Notably, this is the first study in the Philippines to conduct both full-length ORF5 and whole-genome sequencing of PRRS V-2 using long amplicon targeted sequencing. Through ORF5-targeted and long-amplicon tiling approaches, one strain (BA2021012A) was classified as a lineage L8C field strain, while the other (ME20230008B-2) was identified as a lineage L7 vaccine-like strain closely related to U.S. commercial vaccine strains. These are the first PRRSV-2 whole genomes publicly reported from the Philippines, offering critical insights into circulating lineages and the co-detection of field and vaccine-related strains.

Complementing these genomics work is the development of a prototype molecular diagnostics laboratory_ nformation management system (LIMS) towards an integrated approach to strengthening livestock disease surveillance in the Philippines. The LIMS was designed to modernize molecular diagnostics workflows and it featured modules for sample accessioning, analysis request management, test result nput, approval, and automated reporting. The system was tested in a simulated environment and demonstrated enhanced traceability, reduced manual error, operational efficiency, and improved data management. Future enhancements include integration with laboratory instruments, mobile compatibility, and real-time quality control tracking.

Together, these studies demonstrate the feasibility of integrating long-read sequencing and informatics to support genomics-informed surveillance and disease control in swine populations. This work lays the foundation for responsive, data-driven strategies for managing ASFV and PRRSV, and other animal infectious diseases in the Philippines, contributing to national efforts in safeguarding food security and agricultural resilience.

Document Type

Dissertation

Degree

Doctor of Philosophy in Microbiology

College

Graduate School (GS)

Adviser/Committee Chair

Lucille C. Villegas

Co-adviser

Homer D. Pantua

Committee Member

Nacita B. Lantican, Walter A. Laviña, Ria Mae H. Borromeo, Windell L. Rivera, Cherry P. Fernandez-Colorado

Language

English

Location

UPLB Main Library Special Collections Section (USCS)

En – AGROVOC descriptors

AFRICAN SWINE FEVER VIRUS; PRRS; SWINE; VIRAL GENOMES; DISEASE SURVEILLANCE

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