Comparison Between the SFTS-QS Kit and the PowerChek SFTSV Real-time PCR Kit for the Detection of Severe Fever With Thrombocytopenia Syndrome Virus

The recent increase in severe fever with thrombocytopenia syndrome (SFTS) cases has led to the development of the SFTS-QS kit (MiCoBioMed, Seongnam, Korea) for detecting the SFTS virus (SFTSV, now renamed Huaiyangshan banyangvirus). SFTS-QS is a qualitative real-time reverse transcription PCR assay based on lab-on-a-chip technology. We evaluated the performance of the SFTS-QS kit and compared it with that of the PowerChek SFTSV Real-time PCR kit (PowerChek; Kogene Biotech, Seoul, Korea). A total of 117 serum samples were simultaneously assayed using the SFTS-QS and PowerChek kits. Sanger sequencing targeting the S and M segments of SFTSV was performed as the reference method. The total turnaround time of the two kits was compared. The SFTS-QS results agreed with those of PowerChek with a kappa value of 0.92. The diagnostic sensitivity and specificity of the SFTS-QS kit were both 100% (14/14 and 103/103, respectively), whereas those of the PowerChek kit were 100% (14/14) and 98.1% (101/103), respectively. The results of SFTS-QS and PowerChek were comparable; however, the SFTS-QS kit required a shorter total turnaround time. The SFTS-QS kit produced accurate and fast results and thus could serve as a useful tool for detecting SFTSV.

Four Year Surveillance of the Vector Hard Ticks for SFTS, Ganghwa-do, Republic of Korea

The seasonal abundance of hard ticks that transmit severe fever with thrombocytopenia syndrome virus was monitored with a collection trap method every April to November during 2015–2018 and with a flagging method every July and August during 2015–2018 in Ganghwa-do (island) of Incheon Metropolitan City, Republic of Korea. This monitoring was performed in a copse, a short grass field, coniferous forest and broad-leaved forest. A total of 17,457 ticks (8,277 larvae, 4,137 nymphs, 3,389 females, and 1,654 males) of the ixodid ticks comprising 3 species (Haemaphysalis longicornis, H. flava, and Ixodes nipponensis) were collected with collection traps. Of the identified ticks, H. longicornis was the most frequently collected ticks (except larval ticks) (94.26%, 8,653/9,180 ticks (nymphs and adults)), followed by H. flava (5.71%, 524/9,180) and Ix. nipponensis (less than 0.04%, 3/9,180). The ticks collected with collecting traps were pooled and assayed for the presence of SFTS virus with negative results. In addition, for monitoring the prevalence of hard ticks, a total of 7,461 ticks (5,529 larvae, 1,272 nymphs, 469 females, and 191 males) of the ixodid ticks comprising 3 species (H. longicornis, H. flava, and Ix. nipponensis) were collected with flagging method. H. longicornis was the highest collected ticks (except larval ticks) (99.53%, 1,908/1,917 ticks (nymphs and adults)), followed by H. flava (1.15%, 22/1,917).

Epidemiology of severe fever and thrombocytopenia syndrome virus infection and the need for therapeutics for the prevention

Over the past ten years there has been a marked increase in cases of severe fever and thrombocytopenia syndrome in East Asia. This tick-borne hemorrhagic fever presents along with clinical signs including high fever and leukopenia. In addition to humans, the virus has also been detected with shared genetic homology in farm animals including goats, cattle, horses, and pigs. Furthermore, several genotypes of severe fever and thrombocytopenia syndrome virus (SFTSV) are currently co-circulating between humans and animals. In China, where the virus was first detected in rural areas in 2009, the SFTSV mortality rate has been reported to be as 6% and higher than 30%, especially in immuno-compromised patients. Moreover, this virus has been isolated in neighbor countries including Japan and South Korea where the fatality rates in 2015 were more than 30% in both countries. In this review, we comprehensively summarize the virology, genotypes, pathogenesis, and epidemiology of SFTSV infection in humans and animals. Currently, a collaborative global approach against SFTSV infection is being undertaken; however, the need for continuous disease surveillance and production of an effective vaccine is imperative as this virus may lead to an epidemic of irreversible status in both humans and animals.

Comparison and evaluation of laboratory test techniques for severe fever with thrombocytopenia syndrome / 中华实验和临床病毒学杂志

Objective@#Compare the detection result of blood samples of severe fever with thrombocytopenia syndrome (SFTS) patients using different detection techniques, and observe the dynamic characteristics of the virus specific RNA, IgM antibody and IgG antibody, to provide theoretical basis for selection of diagnostic methods of disease.@*Methods@#Acute phase serum of suspected SFTS cases and convalescent serum samples of lab-confirmed cases were collected. Real-time fluorescence quantitative PCR and enzyme-linked immunosorbent assay (ELISA) were used to detect the virus specific RNA, IgM antibody and IgG antibody. The detection results of different methods, the relationship between positive results and the acquisition time, and the dynamic characteristics of viral nucleic acid and antibodies were analyzed.@*Results@#A total of 87 serum samples of the suspected SFTS patients were collected, the positive rate of virus specific RNA, IgM antibody and IgG antibody were 53.41%, 31.03% and 3.41%, respectively. Among 55 confirmed cases of SFTS, the consistent rate of virus specific RNA and IgM antibody detection methods was 36.36%, and the difference between the two methods was significant (χ2=6.82, P=0.009), kappa=-0.257. The sampling intervals of RNA positive samples were all within 12 days, of which the positive detection rate was highest after 7-9 days, and the difference was statistically significant (χ2=10.35, P=0.016). In 34 SFTS convalescent serum samples, all the nucleic acid tests were negative, the positive rate of IgM antibody was 41.18%, which was not significantly different from the acute phase serum samples (P=1.00). The positive rate of IgG antibody was 94.12%, which was significantly higher than that of acute IgG antibody (0%). The dynamic characteristics of IgM and IgG antibody showed that IgM antibody could be detected on the second day after onset, the latest detection time was 74 days after onset, and the highest absorbance value and antibody detection rate occurred in 30-60 days. The earliest detection time of IgG antibody was 12 days after onset, and the last detection time was 100 days.The detection rate of IgG antibody and absorbance value increased rapidly after 30 days, and maintained in a high level. The detection rate of IgG antibody was 100% in 30-60 days.@*Conclusions@#Blood samples taken from SFTS suspected patients within two weeks of onset may be prioritized for detection of viral nucleic acids using Real-time fluorescence PCR or for detection of IgM antibodies by ELISA. Although IgM antibody can be detected 2 days after the onset, the peak appeared much later, so the negative result can’t rule out the diagnosis. IgG antibody has a high seroconversion rate in convalescent samples, and can be used as an auxiliary tool for disease diagnosis.

Construction of the recombinant plasmids encoding nonstructural proteins of SFTS virus and immunogenicity study / 中华实验和临床病毒学杂志

Objective@#To study the construction and humoral immunogenicity of the recombinant plasmids encoding nonstructural proteins(NSs) of severe fever with thrombocytopenia syndrome(SFTS) virus(SFTSV).@*Methods@#Recombinant plasmids encoding NSs gene and optimized NSs gene of SFTSV were constructed by polymerase chain reaction(PCR) and cloned into eukaryotic expression vector pJW4303, which were named pJW4303-NSs and pJW4303-NSs-opt, respectively. Then, the plasmid pJW4303-NSs was tagged with Flag, named pJW4303-NSs-Flag. Meanwhile, Nhe I restrict site and tissue plasminogen activator(tPA) signal sequence were inserted to construct bi-optimized recombinant plasmid named pJW4303-tPA-NSs-opt. All plasmids were identified by sequencing. The transient expression of NSs was confirmed by Western blotting in human embryonic kidney 293T cells. The NSs-specific IgG antibodies in BALB/c mice which were immunized by intramuscular injection with electroporation were examined by enzyme-linked immunosorbent assay(ELISA).@*Results@#The recombinant plasmids pJW4303-NSs, pJW4303-NSs-opt, pJW4303-tPA-NSs-opt and pJW4303-NSs-Flag were successfully constructed. The expression of NSs was confirmed in lysates and supernatants of 293T cells. The NSs-specific IgG responses of all three recombinant plasmids were detected by ELISA in BALB/c mice. It was found that optimized recombinant plasmid pJW4303-NSs-opt elicited higher levels of the NSs-specific IgG than that of pJW4303-NSs in week 6, 8 which induced stronger immune response.@*Conclusions@#The recombinant plasmids encoding SFTSV NSs possess the satisfied immunogenicity. In addition, the plasmid pJW4303-NSs-opt could induce the strongest humoral immune response.

The results of the epidemiological surveillance of severe fever with thrombocytopenia syndrome in Ningbo City / 预防医学

Objective To learn the epidemiological status of severe fever with thrombocytopenia syndrome (SFTS). Methods The surveillance sites were established in two county level hospitals and two township health centers in Ningbo city.Using the diagnostic criteria of the national ministry of health and questionnaire of epidemiological case survey,cases with SFTS were investigated.The SFTS virus was isolated with Vero cells from the positive serum and the gene sequence was analyzed.Results Eleven cases were reported during year 2012—2013,including one person died.A total of 156 ticks were detected negative.The positive rate of serum collected from healthy people in local area was 6.14% and increased with age(P <0.01).There were a close relationship between five isolates of Ningbo and isolates of Japan and Zhoushan,and the gene sequence similarity was more than 96%.Conclusion SFTS cases were reported in Ningbo City, and the surveillance system should be strengthened especially for the disease detection.

No Detection of Severe Fever with Thrombocytopenia Syndrome Virus from Ixodid Ticks Collected in Seoul

Larvae, nymphs, and adult stages of 3 species of ixodid ticks were collected by tick drag methods in Seoul during June-October 2013, and their infection status with severe fever with thrombocytopenia syndrome (SFTS) virus was examined using RT-PCR. During the period, 732 Haemaphysalis longicornis, 62 Haemaphysalis flava, and 2 Ixodes nipponensis specimens were collected. Among the specimens of H. longicornis, the number of female adults, male adults, nymphs, and larvae were 53, 11, 240, and 446, respectively. Ticks were grouped into 63 pools according to the collection site, species, and developmental stage, and assayed for SFTS virus. None of the pools of ticks were found to be positive for SFTS virus gene.