# Environmental metagenomics enhances detection of circulating viruses from live poultry markets in Cambodia

## TL;DR

Environmental metagenomics in Cambodian live-bird markets detects more viruses than traditional methods, including dangerous H5N1 strains.

## Contribution

Metagenomic analysis of environmental samples outperforms poultry swabs in detecting zoonotic viruses in live-bird markets.

## Key findings

- Environmental samples detected 40 viruses from families like Orthomyxoviridae and Coronaviridae.
- Air samples captured the highest diversity of poultry viruses.
- Environmental metagenomics identified H5N1 clades missed by poultry swabs.

## Abstract

Environmental surveillance has emerged as a pivotal strategy for early detection of pathogens that pose a threat to humans. In Asia, live-bird markets (LBMs) are key human-animal interfaces for zoonotic virus transmission. Traditional sampling strategies are time-consuming, expensive and carry significant biosafety risks. Here, we assess the performance of metagenomics on environmental samples (ES) versus traditional poultry swabs for detecting viral pathogens in two Cambodian LBMs between January 2022 and April 2023. ES, including air (n = 35), cage swabs (n = 17), carcass wash water (n = 17) and drinking water (n = 9) are collected alongside oropharyngeal and cloacal swabs from chickens (n = 30) and ducks (n = 29). ES is sensitive in detecting 40 viruses from pathogen families including Orthomyxoviridae and Coronaviridae. Air samples capture the greatest diversity of poultry viruses. Viral contigs from ES show high sequence identity to poultry swab contigs when aligned to the same gene. We show ES outperforms poultry samples in detecting the highly pathogenic influenza A/H5N1, including clades 2.3.4.4b and 2.3.2.1c, which are found in the environment but are missed by poultry swabs. Our findings show metagenomics on ES replicates traditional surveillance, offering broader coverage and improved pathogen detection. This approach could be pivotal for mitigating zoonotic spillover and enhancing pandemic preparedness.

Authors use metagenomic environmental surveillance in Cambodian live-bird markets and found that air sampling captured the greatest diversity of poultry viruses. It outperformed traditional swabs, identifying undetected H5N1 clades.