Original article
Nucleocapsid protein-specific IgM antibody responses in the disease progression of severe fever with thrombocytopenia syndrome

https://doi.org/10.1016/j.ttbdis.2019.02.003Get rights and content

Abstract

Objectives

Severe fever with thrombocytopenia syndrome (SFTS) is an emerging infectious disease that is caused by the SFTS virus (SFTSV) and has a high fatality rate. SFTSV-specific antibody profiles among patients with different clinical outcomes are yet to be described. The nucleocapsid protein (NP) is the most immunogenic viral antigen of the SFTSV. This study, therefore, sought to determine NP-specific antibody responses among SFTS patients with different disease progressions.

Methods

In the present study, 43 patients with confirmed SFTS were enrolled in our cohort, and 9 of them deceased. The clinical presentations and key laboratory parameters associated with SFTS fatality were also recorded. Serum samples from each patient were collected every 2 days during their hospitalization. NP-specific IgM and IgG responses as well as Gn or Gc-specific IgM responses were examined by enzyme-linked immunosorbent assay (ELISA), whereas, the dynamic viral loads of SFTSV RNA were quantified via real-time reverse transcription polymerase chain reaction (RT-PCR).

Results

First, 77% of patients generated positive NP-specific IgM antibody responses within two weeks since illness onset, defined as ‘N-specific IgM-positive patients’, while the rest of the patients were termed as ‘N-specific IgM-delayed patients’. Only 17% of the patients generated NP-specific IgG responses. The absence of NP-specific humoral responses was strongly associated with a high risk of fatality and severity of SFTS. IgM-positive patients had significantly lower levels of viral loads, less disturbed coagulopathy, and hepatic and cardiac damage compared to IgM-delayed patients. Moreover, compared to severe or fatal SFTS patients, mild SFTS patients had significantly higher magnitudes of NP-specific IgM responses, but not NP-specific IgG, Gn-specific IgM, or Gc-specific IgM responses. The abundance of NP-specific IgM responses negatively correlated with viral loads, coagulation disturbances, and hepatic injuries among SFTS patients.

Conclusions

Our data highlight distinct humoral profiles of NP-specific IgM responses among SFTS patients with different disease progressions and clinical outcomes.

Introduction

As an emerging hemorrhagic fever disease identified in eastern Asia, severe fever with thrombocytopenia syndrome (SFTS) is caused by a novel phlebovirus in the Bunyaviridae family (Kim et al., 2015; Takahashi et al., 2014; Yu et al., 2011), known as SFTS virus (SFTSV). Another new phlebovirus, Heartland virus, was isolated from two patients suffering from severe febrile illness in Missouri, USA (McMullan et al., 2012), which shares high identity with SFTSV. These two newly emerged phleboviruses have, subsequently, exerted a global public threat.

SFTSV is transmitted through a tick bite (Luo et al., 2015) and human-to-human contact via blood or body fluid (Bao et al., 2011; Chen et al., 2017a). SFTS has an average of 12% case fatality rate, ranging from 6% to 30% (Yu et al., 2011). The clinical symptoms of SFTS infection are variable, ranging from asymptomatic or acute self-limited febrile condition to life-threatening illness. The typical clinical presentation is characterized by the sudden onset of fever, fatigue, gastrointestinal symptoms, leukopenia, and thrombocytopenia. Severe SFTS cases quickly develop multiple organ dysfunction (MOD) and disseminated intravascular coagulation (DIC), which might lead to fatality within 2 weeks of the disease onset (Zhang et al., 2013, 2012). Previous studies have identified a panel of risk factors associated with the fatality and severity of SFTS cases, such as old age, coagulation disturbance, and remarkable liver damage (Chen et al., 2017b; Cui et al., 2014; Ding et al., 2014; Jia et al., 2017; Zhang et al., 2012); however, the underlying immunological mechanism remains to be obscure. Although ribavirin has been proven effective in reducing the viral load of SFTSV, its usage could not decrease the case fatality ratio of SFTS patients (Liu et al., 2013; Lu et al., 2015). Currently, there is no effective treatment regimen for SFTS in clinical practice other than supportive care.

Antibodies typically play a key role in controlling viral infections, which are immune correlates contributing to the protective efficacy of many successful vaccines (Crowe, 2017). However, the role of SFTS-specific adaptive immune responses during an SFTS disease progression has to be elucidated. SFTSV-specific IgM antibody responses reportedly could be detected within an average period of 9 days, while SFTSV-specific IgG responses could only be detected within an average period of 6 weeks (Lu et al., 2015), and the neutralizing antibodies from SFTS patients could be maintained for at least 4 years (Huang et al., 2016). Nevertheless, the dynamic profile of antibody responses against specific SFTS viral proteins has not been further dissected and carefully characterized.

Nucleocapsid protein (NP), encoded by the small segment of bunyavirus, is the most conserved Phlebovirus genus. NP is abundantly present in viral particles and infected cells. The primary function of NP is to encapsidate the viral genome forming ribonucleoprotein complexes (RNPs) (Pekosz et al., 1999; Walter et al., 2011). Beyond its critical role in viral RNA protection, NP is actively involved in RNA transcription and replication, as well as in the formation of an inclusion body in the cytoplasm of virus-infected cells (Eifan and Elliott, 2009; Pekosz et al., 1999; Pinschewer et al., 2003; Walter et al., 2011). It has been shown that NP is highly immunogenic and that NP-specific antibodies are also readily detected early after infection in convalescent individuals, providing a robust basis for diagnostic detection of SFTS disease (Magurano and Nicoletti, 1999; Martin-Folgar et al., 2010). Interestingly, NP-specific antibodies without neutralizing activities have been demonstrated to protect animals from challenges with Rift Valley fever virus- (RVFV) and Hantavirus partially (Boshra et al., 2011b; Nakamura et al., 1985; Yoshimatsu et al., 1993). Whether viral infection-induced NP-specific humoral responses have any association with the clinical outcome of phlebovirus, including SFTSV infection in vivo, is rarely investigated in a real-world setting.

In the current report, we performed a prospective study on SFTS patients, including 34 survivors and 9 deceased patients. The kinetics of NP-specific IgM and IgG antibody responses during the acute infection phase was measured. Further, the clinical manifestations and the dynamic changes of clinical laboratory tests were determined to further define the clinical features among NP-specific IgM-positive versus IgM-delayed patients. The correlations between the magnitudes of NP-specific IgM responses and clinical laboratory parameters were analyzed. With such information, we carefully dissected the antibody profile of NP-specific humoral responses among fatal, severe and mild SFTS patients.

Section snippets

Human subjects

This retrospective study was performed in Nanjing Drum Tower Hospital, Nanjing, Jiangsu Province, China, from June 2016 to Oct 2018. Clinical manifestations and laboratory parameters were collected from 43 SFTS patients (9 deaths and 34 survivors). A SFTS patient was clinical defined if he or she agreed with the following criteria: (1) acute fever with body temperature of > 38 °C; (2) decline of white blood cell (WBC) and platelet count (PLT); (3) the contact history with tick bite or with the

The dynamic NP-specific IgM and IgG responses among SFTS patients

From Jan 2016 to Oct 2018, a total of 43 laboratory-confirmed SFTSV infected patients were recruited in this study. From this cohort, 34 patients (79%) were survived, and 9 patients (21%) died ( Table 1). The NP-specific IgM responses among SFTS patients were determined every other day beginning from the moment they were hospitalized. We found that NP-specific IgM responses were elicited among 27 (63%) out of 43 patients before day 10. Positive NP-specific IgM responses were further found in

Discussion

Here, for the first time, we comprehensively analyzed the humoral response profile for the most immunogenic viral proteins, NP, in SFTSV, among SFTS patients with different clinical outcomes. Additionally, we compared the clinical outcomes and the disease progressions between NP-specific IgM-positive versus IgM-delayed patients. The clinical features among NP-specific IgM-positive and IgM-delayed patients were further characterized, revealed by the clinical presentations and the dynamic

Conclusions

Our study has demonstrated that NP-specific IgM-delayed SFTS patients tend to have severe or even fatal clinical outcomes, validated by remarkably high levels of viral loads and extensively disturbed laboratory findings observed in NP-specific IgM-delayed patients. Furthermore, our data revealed that compared to mild SFTS patients, fatal or severe SFTS patients had significantly lower levels of NP-specific IgM but not NP-specific IgG responses, Gn-specific IgM, nor Gc-specific IgG. Our data

Acknowledgements

This work was supported by the National Natural Science Foundation of China [81600201 and 81672025], Nanjing Medical Science and Technique Development Foundation [QRX17141], Jiangsu Province’s Outstanding Medical Academic Leader Program [LJ201154], Jiangsu Province’s Clinical Medicine and Technology Special Program [BL2012034].

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    These authors contributed equally to this work.

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