Viral Hemorrhagic Fevers in Korea

Abstract

Background Most of the viruses associated with viral hemorrhagic fevers (VHF) are zoonotic, dependent on their hosts for overall survival and further replication. Rodents and arthropods are the main reservoirs for viruses causing VHFs. Hantaviruses and severe fever with thrombocytopenia syndrome virus (SFTSV) are endemic in the Republic of Korea (ROK), yet the knowledge remains scanty in many fields on these infections. Till date, no study is conducted for SFTSV prevalence in rodents at ROK. Moreover, tick borne encephalitis virus (TBEV) has been identified in ticks and rodents, although no human case of TBE is confirmed in ROK. Presented study aims to investigate positive rate of emerging human pathogenic viruses in wild rodents collected in Korea, which serves as the reservoir and the carrier for the potential pathogenic viruses. In addition, this study compared the sensitivity of different reverse transcription polymerase chain reactions (RT-PCR) conducted for detection of SFTSV presence. Methods Wild rodents were captured during 2017 and 2018 at sylvatic habitats in Boseong-gun, Jeollanam-do and Jeju Island, respectively. All rodents were euthanized under animal use protocol and guidelines. Organs like spleen, kidney and lungs were subjected to total RNA extraction. PCRs were performed for the detection of viruses including hantavirus, SFTSV and TBEV using a specific set of primers for each viral genome amplification. Serological analysis was performed using mice sera. Results In total 21 wild mice were captured in Boseong-gun in 2017 and 57 wild mice were captured at Jeju Island in 2018. All of wild mice captured at both locations were Apodemus agrarius. Hantavirus was detected in 24% (5 out of 21) of A.agrarius captured at Jeollanam-do (Boseong-gun) and 17.54% (10/57) for Jeju samples by nested RT-PCR (RT-N-PCR) targeting the L segment of hantavirus. SFTS RT-N-PCR targeting M-segment of the virus detected SFTSV in 5% (1/21) Boseong-gun samples and 5.3% (3/57) samples collected at Jeju Island. TBEV RT-N PCR performed for all the samples collected at Boseong-gun and Jeju were negative. Forty seven sera were available for serological analysis collected from Jeju, among which 5 (11%) of the A.agrarius sera were positive for Immunoglobulin G (IgG) antibodies against hantavirus and 2 (4.2%) for IgG antibodies against SFTSV. Conclusions Our findings includes the first detection of SFTSV in A.agrarius in ROK. This is the first report for occurrence of co-infection of SFTSV and hantavirus in A.agrarius and support the idea that such viruses can co-infect humans as well, provided the fact that they share the same routes of transmissions. Moreover, the study confirms the circulation of hantavirus in ROK through A.agrarius and thus, virus shedding from A.agrarius can increase the risk of human contracting HFRS. Although no TBEV was identified in this study, yet it poses a serious threat to human health in future. First study conducted for SFTS presence in rodents in ROK and positive result found in this study highlights the importance of continued surveillance to find these viral infections possible routes to humans and their prevention. Background Most of the viruses associated with viral hemorrhagic fevers (VHF) are zoonotic, dependent on their hosts for overall survival and further replication. Rodents and arthropods are the main reservoirs for viruses causing VHFs. Hantaviruses and severe fever with thrombocytopenia syndrome virus (SFTSV) are endemic in the Republic of Korea (ROK), yet the knowledge remains scanty in many fields on these infections. Till date, no study is conducted for SFTSV prevalence in rodents at ROK. Moreover, tick borne encephalitis virus (TBEV) has been identified in ticks and rodents, although no human case of TBE is confirmed in ROK. Presented study aims to investigate positive rate of emerging human pathogenic viruses in wild rodents collected in Korea, which serves as the reservoir and the carrier for the potential pathogenic viruses. In addition, this study compared the sensitivity of different reverse transcription polymerase chain reactions (RT-PCR) conducted for detection of SFTSV presence. Methods Wild rodents were captured during 2017 and 2018 at sylvatic habitats in Boseong-gun, Jeollanam-do and Jeju Island, respectively. All rodents were euthanized under animal use protocol and guidelines. Organs like spleen, kidney and lungs were subjected to total RNA extraction. PCRs were performed for the detection of viruses including hantavirus, SFTSV and TBEV using a specific set of primers for each viral genome amplification. Serological analysis was performed using mice sera. Results In total 21 wild mice were captured in Boseong-gun in 2017 and 57 wild mice were captured at Jeju Island in 2018. All of wild mice captured at both locations were Apodemus agrarius. Hantavirus was detected in 24% (5 out of 21) of A.agrarius captured at Jeollanam-do (Boseong-gun) and 17.54% (10/57) for Jeju samples by nested RT-PCR (RT-N-PCR) targeting the L segment of hantavirus. SFTS RT-N-PCR targeting M-segment of the virus detected SFTSV in 5% (1/21) Boseong-gun samples and 5.3% (3/57) samples collected at Jeju Island. TBEV RT-N PCR performed for all the samples collected at Boseong-gun and Jeju were negative. Forty seven sera were available for serological analysis collected from Jeju, among which 5 (11%) of the A.agrarius sera were positive for Immunoglobulin G (IgG) antibodies against hantavirus and 2 (4.2%) for IgG antibodies against SFTSV. Conclusions Our findings includes the first detection of SFTSV in A.agrarius in ROK. This is the first report for occurrence of co-infection of SFTSV and hantavirus in A.agrarius and support the idea that such viruses can co-infect humans as well, provided the fact that they share the same routes of transmissions. Moreover, the study confirms the circulation of hantavirus in ROK through A.agrarius and thus, virus shedding from A.agrarius can increase the risk of human contracting HFRS. Although no TBEV was identified in this study, yet it poses a serious threat to human health in future. First study conducted for SFTS presence in rodents in ROK and positive result found in this study highlights the importance of continued surveillance to find these viral infections possible routes to humans and their prevention.|연구 배경 바이러스성 출혈열(VHF)과 관련된 대부분의 바이러스는 동물에 존재하고, 생존 및 복제를 위해 숙주에 의존하며, 특히 설치류와 절지동물은 VHF 바이러스의 주된 저장소다. 바이러스성 출혈열 의 원인이 되는 바이러스는 한타바이러스(Hantavirus), 중증열성혈소판감소증후군바이러스(SFTSV), 진드기매개뇌염 바이러스(TBEV) 등이 있으며, 한타바이러스와 중증열성혈소판감소증후군바이러스 는 한국에서 지역 풍토적으로 발견된다. 그러나 이 바이러스 감염과 관련된 연구는 아직까지 부족한 상태이며, 지금까지 국내 설치류에서 SFTSV 유병률을 확인하는 연구는 수행되지 않았다. 또한 TBEV는 진드기와 설치류에서 확인되는데, 국내에서 진드기매개뇌염의 인체 감염 사례는 보고되지 않았다. 따라서 본 연구는 국내에서 잠재적인 병원성 바이러스에 대한 저장소 및 운반자 역할을 하는 야생 설치류에서 병원성 바이러스의 양성률을 조사하고자 수행되었으며, SFTSV를 검출하기 위해 수행된 다양한 역전사 중합 효소 연쇄반응(Reverse Transcription Polymerase Chain Reactions, RT-PCR)의 민감도를 비교하였다.

연구 방법 2017년과 2018년 전라남도 보성군과 제주도에서 야생 설치류를 수집하였고, 모든 설치류는 동물실험 지침에 따라 안락사 시켰다. 비장, 신장 및 폐 조직에서 total RNA를 추출하였고, 한타바이러스, SFTSV, TBEV의 각 바이러스에 특이적인 유전자를 검출하기 위해 nested RT-PCR (RT-N-PCR)을 수행하였다. 또한 포획된 설치류의 혈청을 이용하여 한타바이러스와 SFTSV에 대한 항체가 분석을 수행하였다.

연구 결과 2017년 보성군에서 21마리, 2018년 제주도에서 57마리의 야생 들쥐가 포획되었고, 두 지역에서 포획된 야생 들쥐는 모두 등줄쥐(Apodemus agrarius)로 확인되었다. 한타바이러스는 L segment를 타깃으로 nested RT-PCR (RT-N-PCR)을 수행하였으며, 보성군 샘플에서 24% (5/21), 제주 샘플에서 17.54% (10/57)로 확인되었다. SFTSV의 경우 M segment를 타깃으로 RT-N-PCR을 수행하였을 때 민감도가 가장 높았으며, 보성군 샘플에서 5% (1/21), 제주 샘플에서 5.3% (3/57)로 확인되었다. TBEV의 RT-N-PCR 결과는 두 지역 샘플에서 모두 음성이었다. 제주에서 포획된 47마리의 야생 들쥐 혈청을 이용한 항체가 조사 결과, 한타바이러스에 대한 IgG 양성이 11% (5/47)로 확인되었고, SFTSV에 대한 IgG 양성은 4.2% (2/47)로 확인되었다.

연구 결론 본 연구를 통해 국내의 등줄쥐에서 SFTSV의 감염을 처음 확인하였고, 또한 한 마리의 등줄쥐에서 SFTSV와 한타바이러스가 동시 감염(co-infection) 되었음을 최초로 확인하였다. 이 결과는 바이러스가 숙주로 감염되는 경로를 공유하므로, 인체에도 마찬가지로 바이러스의 동시 감염이 일어날 수 있음을 시사한다. 또한 본 연구는 국내에서 등줄쥐의 SFTSV와 한타바이러스 감염을 확인하였으며, 이는 바이러스의 인체 감염으로 질병 발생 위험이 증가할 수 있음을 시사한다. TBEV는 본 연구결과 확인되지 않았지만, 추후에는 인체 건강에 심각한 위협이 될 수 있으므로 꾸준한 감시 및 모니터링이 필요하다. 본 연구는 국내의 설치류에서 SFTS 존재를 확인한 첫 결과이며, 이러한 설치류의 바이러스 감염이 인체 감염 전파 경로에 대한 이해와 예방법을 찾아내기 위해 설치류 바이러스의 지속적인 감시가 중요하다.