Unveiling Viral Diversity with RNA-Seq
for Better Outbreak Defense
Most viruses rarely make headlines until it is too late. The recent H5N1 bird flu outbreak is a stark reminder. Since 2022, this virus has devastated U.S. poultry flocks, affecting nearly 175 million birds, and has spilled over into more than two dozen mammal species1, 2. By the time these stories reach the news, the damage is already well underway.
Viral profiling offers a way to change that narrative. Researchers are using RNA sequencing to explore and characterize viral diversity and abundance, providing insights that enhance both surveillance and diagnostic efforts. Unlike targeted assays that search for a specific virus, total RNA sequencing captures the whole landscape of RNA, revealing both known viruses and novel ones.
With efficient RNA-Seq workflows, scientists can move quickly from samples to answers. Tools like the Zymo-Seq RiboFree® Total RNA Library kit are helping these efforts by providing a universal rRNA depletion method that works across any organism, including those carrying novel viruses. The faster we understand what is circulating, the better chance we have of stopping the next pandemic before it becomes a global threat.
Tracking High-Risk Coronaviruses in Wild Bats
Key Takeaways
Study: Wang J et al. (2023) Individual bat virome analysis reveals co-infection and spillover among bats and virus zoonotic potential. Nat Commun 14: 4079. https://doi.org/10.1038/s41467-023-39835-1
Goal: Profile the individual RNA virome of 149 wild bats from six regions across Yunnan, China, known as a hotspot for emerging bat-borne viruses.
Method: Extract total RNA from rectal samples and perform metatranscriptomic sequencing.
Findings:
- Identified 55 distinct mammal-associated viral species
- 42% of virus-positive bats carried multiple viruses
- Discovered a likely recombinant Bat SARS-like coronavirus CX1 with the potential to bind human ACE2
Bats are known reservoirs for a wide range of viruses, yet how these viruses spread, evolve, and coexist within a single animal is poorly understood. To investigate, researchers profiled the virome of 149 wild bats from six regions in Yunnan Province, a known hotspot for bat-borne coronaviruses.
The team applied metatranscriptomic sequencing to total RNA extracted from rectal samples. Libraries from different bat species were prepared with the Zymo-Seq RiboFree® Total RNA Library Kit. Each bat was analyzed individually to maintain resolution across species, locations, and viral load.
The scientists focused their analysis on 55 mammal-associated viral species belonging to 12 families, predominantly Reoviridae, Picornaviridae, and Coronaviridae. They also found that nearly half of the bats carried at least one virus, and 42% of those carried multiple species. Such high co-infection rates create opportunities for recombination and reassortment, accelerating viral evolution and the risk of new pathogens emerging.
Among the most concerning findings was a likely recombinant bat SARS-like coronavirus CX1 capable of binding to human ACE2 receptors, a possible warning signal of potential future spillover.
Mapping Tick-Borne Viruses in Southwestern China
Key Takeaways
Study: Wang J et al. (2023) Metatranscriptomics reveals diversity of tick-borne viruses in Nujiang, Yunnan Province. Front Cell Infect Microbiol 13: 1283019. doi:10.3389/fcimb.2023.1283019
Goal: Characterize the virome of ticks in Nujiang Prefecture, a region where livestock, wildlife, and humans interact closely.
Method: Metatranscriptomic sequencing of 2,570 ticks pooled by time, location, and morphology.
Findings:
- Identified 13 RNA viruses from 8 viral families
- Identified 6 bunyaviruses distributed among 3 families; this class has been linked to human and livestock febrile illnesses in recent years
- 5 potential new viral species were identified, 4 highly divergent from known viruses
Nujiang Prefecture in northwest Yunnan Province boasts diverse ecosystems and supports a wide range of arthropods. Ticks, an arthropod known for viral transmission, thrive where livestock, wildlife, and humans overlap, making them key players in the spread of emerging zoonotic diseases. In recent years, scientists have contributed significantly to the identification of tick-borne pathogens, and the authors of this paper joined the effort by performing viral profiling on 2,570 ticks.
Between 2017 and 2022, these ticks were collected from cattle and sheep across four counties. Samples were pooled by time, location, and morphology. Total RNA was extracted, and libraries were prepared with the Zymo-Seq RiboFree® Total RNA Library Kit. The researchers performed de novo assembly and phylogenetic analysis to categorize the detected viral sequences.
The study identified 13 RNA viruses across eight known families, most belonging to unclassified genera or species.
Recovering Viral Genomes from Archived Museum Specimens
Key Takeaways
Study: Salazar-Hamm PS et al. (2024) Choclo virus recovered from deep metatranscriptomics of archived frozen tissues. PLoS Negl Trop Dis 18(1): e0011672. https://doi.org/10.1371/journal.pntd.0011672
Goal: Detect and characterize Choclo virus (CHOV) from 20-year-old archived rodent tissues to support hantavirus surveillance in Panama.
Method: Deep metatranscriptomic sequencing of libraries prepared from archived heart and kidney samples.
Findings:
- Assembled a 91% complete CHOV genome at 16X coverage
- Contributed the second publicly available CHOV genome
- Expands knowledge of CHOV evolution and pathogenicity for improved future surveillance
In Panama, the Costa Rican pygmy rice rat (Oligoryzomys costaricensis) serves as a reservoir for Choclo virus (CHOV), a type of hantavirus. CHOV can cause hantavirus cardiopulmonary syndrome (HCPS), a severe disease responsible for nearly all known human cases. It is hypothesized that as commensal rodent populations grow and human contact with infected animals increases, the risk of zoonotic pathogen transmission may rise. Given the broad diversity of hantaviruses, their expanding global impact, and the limited CHOV data available in public databases, it is critical for scientists to continue building our understanding of this virus.
Researchers prepared libraries from rodent tissues archived nearly two decades ago using the Zymo-Seq RiboFree® Total RNA Library Kit, and performed deep metatranscriptomic sequencing. They successfully built a 91% complete CHOV genome with an average of 16X coverage through reference-guided assembly, with nearly complete recovery of the S and M segments (98% and 99%, respectively). Overall, the authors assembled a CHOV genome that varied 1.1% from the reference available resulting in 8 nonsynonymous mutations.
This study demonstrates that deep metatranscriptomic sequencing enables the detection of low-abundance RNA viruses in long-term archived samples, including those preserved in natural history collections. By generating the second publicly available CHOV genome, the study provides critical data to support future surveillance, diagnostics, and evolutionary studies of this zoonotic pathogen.
Advancing Viral Detection Across Species and Sample Types
Understanding how viruses emerge, evolve, and move across species is essential for predicting and preventing outbreaks. The Zymo-Seq RiboFree® Total RNA Library Kit offers a streamlined, metatranscriptomic solution, enabling researchers to profile viruses from any organism and identify both known and novel viruses. Whether tracking vector-borne viruses, surveying wildlife, or analyzing archival tissues, metatranscriptome profiling provides scientists a head start in identifying potential threats before they become headline news.
References:
- CDC. USDA Reported H5N1 Bird Flu Detections in Poultry. Avian Influenza (Bird Flu) https://archive.cdc.gov/#/details?url=https://www.cdc.gov/bird-flu/situation-summary/data-map-commercial.html
- Caserta, L. C. et al. Spillover of highly pathogenic avian influenza H5N1 virus to dairy cattle. Nature 634, 669–676 (2024).