Detection of new world hantaviruses by a broadly reactive rRT-PCR assay

Orthohantaviruses are negative-sense RNA viruses that persistently infect rodents and can cause severe human disease. Orthohantavirus diversity complicates comprehensive surveillance strategies and diagnostic tools. Currently, serological assays and nested PCR with degenerate primers are used for hantavirus diagnosis and rodent surveillance, but these assays have long turnaround-times. Here, a broadly reactive Orthohantavirus real-time PCR assay is presented that can be incorporated into public health and research laboratories across the Americas.

Starting from an existing rRT-PCR primer-probe set originally developed for the detection of Sin Nombre virus, a pan-America Orthohantavirus assay (HantAmeriQ) was developed that can be used for both human and rodent surveillance.

Using synthetic RNA, the HantAmeriQ assay detected all New World Orthohantaviruses known to cause human disease at a limit of detection between 25 (Blue River virus) and 28,000 (Bayou virus) copies/reaction. Genetically diverse hantaviruses from North America (including Sin Nombre, New York, Monongahela, Blue River, and Bayou viruses) and Central/South America (including Catacamas, Carrizal, Choclo, Maciel, and Andes viruses) were detected within similar limits. The HantAmeriQ assay performed as well as or better than a commercially available assay for the detection of North American, but not South American hantaviruses and detected diverse hantaviruses from rodent and human specimens.

Here, the HantAmeriQ assay detected New World Orthohantaviruses in both rodent and human samples from across the United States, and hantaviruses from Central and South America. The assay demonstrates strong potential for diagnostic use in North American clinical laboratories and for surveillance and research applications in rodent-centered field studies.