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. 2016 May;54(5):1335-9.
doi: 10.1128/JCM.00113-16. Epub 2016 Mar 9.

Molecular Diagnosis of Hemorrhagic Fever with Renal Syndrome Caused by Puumala Virus

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Molecular Diagnosis of Hemorrhagic Fever with Renal Syndrome Caused by Puumala Virus

Nina Lagerqvist et al. J Clin Microbiol. 2016 May.

Abstract

Rodent-borne hantaviruses cause two severe acute diseases: hemorrhagic fever with renal syndrome (HFRS) in Eurasia, and hantavirus pulmonary syndrome (HPS; also called hantavirus cardiopulmonary syndrome [HCPS]) in the Americas. Puumala virus (PUUV) is the most common causative agent of HFRS in Europe. Current routine diagnostic methods are based on serological analyses and can yield inconclusive results. Hantavirus-infected patients are viremic during the early phase of disease; therefore, detection of viral RNA genomes can be a valuable complement to existing serological methods. However, the high genomic sequence diversity of PUUV has hampered the development of molecular diagnostics, and currently no real-time reverse transcription-quantitative (RT)-PCR assay is available for routine diagnosis of HFRS. Here, we present a novel PUUV RT-PCR assay. The assay was validated for routine diagnosis of HFRS on samples collected in Sweden during the winter season from 2013 to 2014. The assay allowed detection of PUUV RNA in 98.7% of confirmed clinical HFRS samples collected within 8 days after symptomatic onset. In summary, this study shows that real-time RT-PCR can be a reliable alternative to serological tests during the early phase of HFRS.

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Figures

FIG 1
FIG 1
Conservation map of PUUV nucleocapsid protein ORF from Swedish isolates (list of sequences in Table A1 in the supplemental material). Blue arrows represent the position of forward (F) and reverse (R) primers and probe (P) in the PUUV RT-PCR.
FIG 2
FIG 2
Limit of detection. The number of positives per total number of replicates tested is given above each box. The mean values are indicated by horizontal lines; boxes denote the 25th to 75th percentiles and whiskers, the 5th to 95th percentiles; dots represent outliers. GCE, genome copy equivalents.
FIG 3
FIG 3
CT value of positive samples from PUUV-infected patients (n = 86) sampled at day 1 to 14 after onset of HFRS. Four patients, sampled at days 5, 9, 10, and 14, were negative by RT-PCR and not included in the graph.

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