Antibodies against Severe Fever with Thrombocytopenia Syndrome Virus in Healthy Persons, China, 2013

In June 2013, a subclinical infection with severe fever with thrombocytopenia syndrome virus (SFTSV) was detected in Zhejiang Province, China, prompting seroprevalence studies in 6 districts within the province. Of 986 healthy persons tested, 71 had IgG antibodies against SFTSV. This finding suggests that most natural infections with SFTSV are mild or subclinical.


The Study
A human case of SFTS was identified in a rural area of the Pujiang district ( Figure 1). The patient, a 60-yearold local male subsistence farmer, sought treatment at the Zhejiang Provincial People's Hospital on June 1, 2012, after a 6-day history of fever (maximum axillary temperature 40°C), malaise, chills, gingival bleeding, hyperemia of conjunctivae, and diarrhea (10 or fewer times per day). Initial laboratory testing revealed thrombocytopenia (13 × 10 9 platelets/L; reference range 100-300 × 10 9 platelets/L) and leukocytopenia (0.93 × 10 9 leukocytes/L; reference range 4-10 × 10 9 /L). Supportive therapy was provided, and the patient's condition seemed to improve on the second day: platelet count rose to 34 × 10 9 platelets/L, and leukocyte count rose to 7.28 × 10 9 leukocytes/L. However, on the third day, the patient became weak and died of multiple organ failure.
Serum samples from the patient were tested for the presence of SFTSV RNA by quantitative real-time reverse transcription PCR as previously described (1). The QIAamp Viral RNA Mini Kit (QIAGEN, Hilden, Germany) was used for RNA extraction. Detection of all 3 viral RNA segments by quantitative real-time reverse transcription PCR and isolation of the virus from Vero cell culture confirmed the association between the clinical syndrome and SFTSV infection. In addition, SFTSV-specific IgG and low levels of viral RNA were detected in a blood sample from a family member of the patient. The family member did not report exposure to potential animal hosts or vectors, so SFTSV transmission is believed to have occurred through personal contact when the family member was caring for the patient. Signs of illness did not develop in the family member.
To investigate if additional mild subclinical infections occurred, we, with the support of the local disease control department, conducted a seroprevalence study in the patient's village in Pujiang district. A total of 54 blood samples were collected from 54 healthy volunteers. We used an ELISA kit provided by the National Center for Disease Control and Prevention to prepare and test serum samples for the presence of SFTSV-specific IgM and IgG (5,6). This ELISA compares well with serum neutralization assays for SFTSV (6). All serum samples were negative for SFTSVspecific IgM, whereas 4 (7.4%) of the serum samples were positive for SFTSV-specific IgG (Figure 2). None of the IgG-positive participants reported any disease symptoms that are associated with SFTSV infections.
To further investigate the occurrence of mild or subclinical SFTSV infections, we collected serum samples from healthy volunteers in 5 additional districts in Zhejiang Province and tested the samples for SFTSV-specific IgG. The percentages of positive samples, by district, follow (the no. positive/total no. tested are shown in parentheses): Lishui, 10.2% (18/176); Jinhua, 3.5% (7/200); Ningbo, 10.9 (28/256); Taizhou, 3% (3/100); and Jiaxing, 5.5% (11/200) ( Figure 2). Results were confirmed by immunofluorescence assay conducted as previously reported (1). In brief, SFTSV-infected Vero cells were fixed with cold acetone, washed with distilled water, air-dried, and then stored at -70°C. Serum samples were diluted 1:20 in phosphate-buffered saline supplemented with 0.01% Evans blue (Sigma-Aldrich Co. LLC, St. Louis, MO, USA). Slides were then incubated with the diluted serum samples in a humidified chamber at 37°C for 30 min and then washed with phosphate-buffered saline. The wells were incubated with fluorescein isothiocyanate-labeled goat anti-human IgG conjugate (Boshide, Wuhan, China), washed, and analyzed by the use of fluorescence microscopy. The results of these experiments confirmed the presence of SFTSV-specific IgG in 84.5% (60/71) of the samples with ELISA-positive results. Serum samples that were negative for SFTSV by ELISA were also negative by immunofluorescence assay.

Conclusions
SFTSV can cause severe disease and high rates of death among infected hospitalized patients. The virus also has the limited ability to be transmited from person to person through contact with contaminated blood, but secondary cases are generally less severe and have so far not resulted in fatalities (7)(8)(9). Nonetheless, there is great public health concern regarding SFTSV.
Our seroprevalence study was prompted by the identification of a subclinical, secondary infection that was most likely caused by person-to-person transmission of the virus from an infected family member with a fatal case of SFTS. We found an overall SFTSV seroprevalence of 7.2% among 986 healthy persons who reported no symptoms associated with SFTS. Because the seropositive participants in our study did not have contact with persons with diagnosed cases of SFTS, their infections most likely occurred through natural exposure. From this, we conclude that SFTSV infections are widespread in rural areas of Zhejiang Province, and only a small percentage of the infections result in clinical disease. Emerging Infectious Diseases • www.cdc.gov/eid • Vol. 20, No. 8, August 2014