Diagnostic Assays for Crimean-Congo Hemorrhagic Fever

Jessica Vanhomwegen

, Maria João Alves, Tatjana Avšič Županc, Silvia Bino, Sadegh Chinikar, Helen Karlberg, Gülay Korukluoğlu, Miša Korva, Masoud Mardani, Ali Mirazimi, Mehrdad Mousavi, Anna Papa, Ana Saksida, Batool Sharifi-Mood, Persofoni Sidira, Katerina Tsergouli, Roman Wölfel, Hervé Zeller, and Philippe Dubois

Author affiliations: Author affiliations: Institut Pasteur, Paris, France (J. Vanhomwegen, P. Dubois); European Centre for Disease Control, Stockholm, Sweden (J. Vanhomwegen, H. Zeller); National Institute of Health Dr. Ricardo Jorge, Lisbon, Portugal (M.J. Alves); Faculty of Medicine, Ljubljana, Slovenia (T. Avšič Županc, M. Korva, A. Saksida); Institute of Public Health, Tirana, Albania (S. Bino); Pasteur Institute of Iran, Tehran, Iran (S. Chinikar); Swedish Institute for Infectious Disease Control, Solna, Sweden (H. Karlberg, A. Mirazimi, M. Mousavi); Refik Saydam National Public Health Agency, Ankara, Turkey (G. Korukluoğlu); Shahid Beheshti University of Medical Sciences, Tehran, Iran (M. Mardani, B. Sharifi-Mood); Aristotle University of Thessaloniki, Thessaloniki, Greece (A. Papa, P. Sidira, K. Tsergouli); and Bundeswehr Institute of Microbiology, Munich, Germany (R. Wölfel)

Abstract

Crimean-Congo hemorrhagic fever (CCHF) is a highly contagious viral tick-borne disease with case-fatality rates as high as 50%. We describe a collaborative evaluation of the characteristics, performance, and on-site applicability of serologic and molecular assays for diagnosis of CCHF. We evaluated ELISA, immunofluorescence, quantitative reverse transcription PCR, and low-density macroarray assays for detection of CCHF virus using precharacterized archived patient serum samples. Compared with results of local, in-house methods, test sensitivities were 87.8%–93.9% for IgM serology, 80.4%–86.1% for IgG serology, and 79.6%–83.3% for genome detection. Specificity was excellent for all assays; molecular test results were influenced by patient country of origin. Our findings demonstrate that well-characterized, reliable tools are available for CCHF diagnosis and surveillance. The on-site use of such assays by health laboratories would greatly diminish the time, costs, and risks posed by the handling, packaging, and shipping of highly infectious biologic material.

Crimean-Congo hemorrhagic fever (CCHF) is a tick-borne zoonotic disease caused by a virus (CCHFV) belonging to the Nairovirus genus (1). The disease is asymptomatic in infected animals but can develop into severe illness in humans, with case-fatality rates as high as 50% in some outbreaks (2,3). The incubation period is typically 3–7 days, with sudden onset of myalgia, headache, and fever that can develop into a severe hemorrhagic syndrome (4,5). CCHFV is transmitted by tick bite (from mainly Hyalomma spp. ticks) or by contact with blood or tissues from infected livestock or patients with CCHF (2,6).
Sporadic cases of CCHF and community and nosocomial outbreaks have been increasingly reported, and the disease’s geographic distribution is the most extensive among tick-borne diseases. Currently, CCHFV is enzootic in southeastern Europe (Bulgaria, Albania, Kosovo, and Greece), southern Russia, and several countries in the Middle East, Africa, and Asia (7–9). Given the abundance of vectors, potential hosts, favorable climate and ecology, and intensified human travel, emergence and rapid establishment of new CCHF foci in other countries are substantial risks (10). Emergence or reemergence of CCHF poses a serious public health threat because it is highly contagious and highly lethal, has the potential to cause nosocomial infection, and is difficult to treat, prevent, and control. In addition to enhanced surveillance and development of therapeutics, access to early, sensitive, and specific laboratory diagnosis is a key factor in increasing preparedness in Europe and other countries at risk (11–13).
Although viral isolation is the standard for CCHF diagnosis, because it has to be done in high-containment biosafety level 4 facilities, the number of laboratories that can perform this technique is limited. Moreover, because cell cultures lack sensitivity and usually only detect the relatively high viremia level encountered during the first 5 days of illness, viral isolation is not without error or uncertainty. As a consequence, reference laboratories have been using the best available practicable methods to determine the presence or absence of infection (11). These methods include conventional and real-time quantitative reverse transcription PCR (RT-PCR and qRT-PCR) for detection of the viral genome (14–18) and indirect immunofluorescence assays (IFAs) or ELISAs for detection of specific IgM and IgG antibodies (19–22). No consensus on the most efficient molecular and serologic testing method has been reached.
In this context, a working group of experts from reference laboratories was constituted under the initiative of the European Network for Diagnostics of Imported Viral Diseases to take part in a multicenter study of CCHF diagnostic tests. The aim of this study was to evaluate and compare the performance of, and review the operational characteristics of, available CCHF diagnostic tests by using panels of well-characterized, archived serum samples from patients from geographically diverse settings.