Discovering disease-causing pathogens in resource-scarce Southeast Asia using a global metagenomic pathogen monitoring system
Authors: Bohl Jennifer A. , Lay Sreyngim , Chea Sophana , Ahyong Vida , Parker Daniel M. , Gallagher Shannon , Fintzi Jonathan , Man Somnang , Ponce Aiyana , Sreng Sokunthea , Kong Dara , Oliveira Fabiano , Kalantar Katrina , Tan Michelle , Fahsbender Liz , Sheu Jonathan , Neff Norma , Detweiler Angela M. , Ly Sokna , Sath Rathanak , Huch Chea , Kry Hok , Leang Rithea , Huy Rekol , Lon Chanthap , Tato Cristina M. , DeRisi Joseph L. , Manning Jessica E. 
Published: September 2021

Understanding the regional pathogen landscape and surveillance of emerging pathogens is key to mitigating epidemics. Challenges lie in resource-scarce settings, where outbreaks are likely to emerge, but where laboratory diagnostics and bioinformatics capacity are limited. Using unbiased metagenomic next generation sequencing (mNGS), we identified a variety of vector-borne, zoonotic and emerging pathogens responsible for undifferentiated fevers in a peri-urban population in Cambodia. From March 2019 to October 2020, we enrolled 473 febrile patients aged 6 months to 65 years of age presenting to a large peri-urban hospital in Cambodia. We collected sera and prepared sequencing libraries from extracted pathogen RNA for unbiased metagenomic sequencing and subsequent bioinformatic analysis on the global cloud-based platform, IDseq. We employed multivariate Bayesian models to evaluate specific pathogen risk causing undifferentiated febrile illness. mNGS identified vector-borne pathogens as the largest clinical category with dengue virus (124/489) as the most abundant pathogen.

Underappreciated zoonotic pathogens such as Plasmodium knowlesi, leptospirosis, and co-infecting HIV were also detected. Early detection of chikungunya virus presaged a larger national outbreak of more than 6,000 cases. Pathogen-agnostic mNGS investigation of febrile persons in resource-scarce Southeast Asia is feasible and revealing of a diverse pathogen landscape. Coordinated and ongoing unbiased mNGS pathogen surveillance can better identify the breadth of endemic, zoonotic or emerging pathogens and deployment of rapid public health response.

Clinical Trial Numbers

NCT04034264 and NCT03534245.

Significance Statement

Public health authorities recently advocated for global expansion of sequencing capacity worldwide; however, the importance of genomics-based surveillance to detect emerging pathogens or variants in resource-limited settings is paramount, especially in a populous, biodiverse Southeast Asia. From 2019 to 2020, pathogen metagenomic next generation sequencing (mNGS) of febrile patients in Cambodia identified several vector-borne and zoonotic pathogens, both common and underappreciated, and resulted in a variety of actionable health interventions. Understanding these pathogen discoveries, and the attendant challenges of mNGS in these outbreak-prone settings, is critical for today’s global society and decision-makers in order to implement sequencing-based pathogen or variant detection.

Significance Statement

Metagenomic pathogen sequencing offers an unbiased approach to characterizing febrile illness. In resource-scarce settings with high biodiversity, it is critical to identify disease-causing pathogens in order to understand burden and to prioritize efforts for control. Here, mNGS characterization of the pathogen landscape in Cambodia revealed diverse vector-borne and zoonotic pathogens irrespective of age and gender as risk factors. Identification of key pathogens led to changes in national program surveillance. This study provides a recent ‘real world’ example for the use of mNGS surveillance in both identifying diverse microbial landscapes and detecting outbreaks of vector-borne, zoonotic, and other emerging pathogens in resource-scarce settings.

Classification

Biological Sciences; microbiology; medical sciences