• View in gallery

    Phylogenetic tree constructed based on complete S segments of SFTSV. The tree was constructed using the maximum likelihood method with MEGA 6.13 The complete S sequences from stored patient serum are shown in bold (JP13-Korea-2017). The S sequence data for the viruses identified from China, South Korea, and Japan were obtained from the National Center for Biotechnology Information/Basic Local Alignment Search Tool. SFTSV = severe fever with thrombocytopenia syndrome virus.

  • View in gallery

    Phylogenetic tree constructed based on the Orientia tsutsugamushi 56-kDa gene sequences. The tree was constructed using the maximum likelihood method with MEGA 6.13 The O. tsutsugamushi 56-kDa gene sequences from stored patient serum are shown in bold as JP13-Korea-2017 (Boryong) and JP13-Korea-2017 (Taguchi). The O. tsutsugamushi 56-kDa gene sequence data identified from China, South Korea, Japan, and India were obtained from NCBI/BLAST.

  • 1.

    International Committee on Taxonomy of Viruses (ICTV), 2017. Taxonomy. Available at: https://talk.ictvonline.org/taxonomy/. Accessed June 22, 2018.

  • 2.

    Yu XJ 2011. Fever with thrombocytopenia associated with a novel bunyavirus in China. N Engl J Med 364: 15231532.

  • 3.

    McMullan LK 2012. A new phlebovirus associated with severe febrile illness in Missouri. N Engl J Med 367: 834841.

  • 4.

    Mourya DT 2014. Malsoor virus, a novel bat phlebovirus, is closely related to severe fever with thrombocytopenia syndrome virus and heartland virus. J Virol 88: 36053609.

    • Search Google Scholar
    • Export Citation
  • 5.

    Wang J 2014. Novel phlebovirus with zoonotic potential isolated from ticks, Australia. Emerg Infect Dis 20: 10401043.

  • 6.

    Yun Y 2015. Phylogenetic analysis of severe fever with thrombocytopenia syndrome virus in South Korea and migratory bird routes between China, South Korea, and Japan. Am J Trop Med Hyg 93: 468474.

    • Search Google Scholar
    • Export Citation
  • 7.

    Yoo JR, Heo ST, Park D, Kim H, Fukuma A, Fukushi S, Shimojima M, Lee KH, 2016. Family cluster analysis of severe fever with thrombocytopenia syndrome virus infection in Korea. Am J Trop Med Hyg 95: 13511357.

    • Search Google Scholar
    • Export Citation
  • 8.

    Bao CJ 2011. A family cluster of infections by a newly recognized bunyavirus in eastern China, 2007: further evidence of person-to-person transmission. Clin Infect Dis 53: 12081214.

    • Search Google Scholar
    • Export Citation
  • 9.

    Denic S, Janbeith J, Nair S, Conca W, Tariq WU, Al-Salam S, 2011. Acute thrombocytopenia, leucopenia, and multiorgan dysfunction: the first case of SFTS Bunyavirus outside China? Case Rep Infect Dis 2011: 204056.

    • Search Google Scholar
    • Export Citation
  • 10.

    Wi YM, Woo HI, Park D, Lee KH, Kang CI, Chung DR, Peck KR, Song JH, 2016. Severe fever with thrombocytopenia syndrome in patients suspected of having scrub typhus. Emerg Infect Dis 22: 19921995.

    • Search Google Scholar
    • Export Citation
  • 11.

    Weitzel T, Dittrich S, López J, Phuklia W, Martinez-Valdebenito C, Velásquez K, Blacksell SD, Paris DH, Abarca K, 2016. Endemic scrub typhus in South America. N Engl J Med 375: 954961.

    • Search Google Scholar
    • Export Citation
  • 12.

    Lee YM, Kim DM, Lee SH, Jang MS, Neupane GP, 2011. Phylogenetic analysis of the 56 kDa protein genes of Orientia tsutsugamushi in southwest area of Korea. Am J Trop Med Hyg 84: 250254.

    • Search Google Scholar
    • Export Citation
  • 13.

    Tamura K, Stecher G, Peterson D, Filipski A, Kumar S, 2013. MEGA6: molecular evolutionary genetics analysis version 6.0. Mol Biol Evol 30: 27252729.

    • Search Google Scholar
    • Export Citation
  • 14.

    Wang QK, Ge HM, Li ZF, Shan YF, Cui L, Wang YP, 2012. Vector research of severe fever with thrombocytopenia syndrome virus in gamasid mites and chigger mites [in Chinese]. Zhongguo Meijie Shengwuxue Ji Kongzhi Zazhi 23: 452454.

    • Search Google Scholar
    • Export Citation
  • 15.

    Zhang M, Zhao ZT, Wang XJ, Li Z, Ding L, Ding SJ, Yang LP, 2014. Mixed scrub typhus genotype, Shandong, China, 2011. Emerg Infect Dis 20: 484485.

 

 

 

 

 

Mixed Infection with Severe Fever with Thrombocytopenia Syndrome Virus and Two Genotypes of Scrub Typhus in a Patient, South Korea, 2017

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  • 1 Department of Microbiology and Immunology, Jeju National University College of Medicine, Jeju, South Korea

Severe fever with thrombocytopenia syndrome (SFTS) is a tick-borne viral disease with a high mortality rate. Infection can also occur through close contact with an infected patient. Scrub typhus is an acute febrile illness caused by Orientia tsutsugamushi, a bacterium transmitted to humans through chigger mite bites. South Korea is an endemic region of SFTS and scrub typhus. In this study, we confirmed that a patient was coinfected with SFTS virus and two (Boryong and Taguchi) genotypes of O. tsutsugamushi.

INTRODUCTION

Severe fever with thrombocytopenia syndrome (SFTS) is an emerging tick-borne viral hemorrhagic fever disease caused by the genus Phlebovirus in the family Phenuiviridae.1,2 Phleboviruses have a broad geographic range and have been found in sand flies, mosquitoes, and ticks in the Americas, Asia, Africa, and Mediterranean regions for many years; another tick-borne Phlebovirus, the Heartland virus (HRTV), was identified in northwestern Missouri in the United States in 2009.3 Malsoor virus, a novel bat Phlebovirus closely related to SFTS virus (SFTSV) and HRTV, was isolated from the Rousettus leschenaultia species of bats in western India; there was also isolation and characterization of a phlebovirus from ticks in Australia that was similar to SFTSV and HRTV.4,5

Most SFTSV infections occur via the Haemaphysalis longicornis tick, although SFTSV transmission can also occur through close contact with an infected patient.2,68 SFTS was first confirmed in middle China in 2009 and was also reported and confirmed in South Korea and the western regions of Japan in 2013; a SFTS Bunyavirus patient was also reported in the United Arab Emirates, an area not an endemic country for SFTS.2,6,9

The disease is characterized by fever, gastrointestinal signs and symptoms, leukopenia, thrombocytopenia, and a high mortality rate.2,68,10

Scrub typhus is a life-threatening bacterial disease caused by Orientia tsutsugamushi and is transmitted primarily by the bite of the Leptotrombidium spp. mite (Acari: Trombiculiidae) infected with Orientia species and endemic in the “tsutsugamushi triangle” within the Asia-Pacific region.10,11 However, patients were documented in the Middle East and South America in 2016.11 SFTS is endemic to South Korea and is a major public health concern between May and October; scrub typhus, also a major public health problem, is mostly present between October and November.10

The clinical presentations of SFTS and scrub typhus are similar: signs and symptoms typically develop within 1–2 weeks of infection and include fever, headache, malaise, and gastrointestinal symptoms, but SFTS exhibits a higher mortality rate than dose scrub typhus.2,68,10,11

In this study, we report the first confirmed case of mixed infection with SFTSV and two genotypes of O. tsutsugamushi in a patient.

MATERIALS AND METHODS

Patient.

A healthy 74-year-old woman (JP13-Korea-2017) presented with fever, myalgia, headache, and abdominal pain in the periumbilical area of 3 days duration. The patient’s medical history was unremarkable. She resided in an urban area and went to the mountains 4 days prior, in the autumn season. She was unaware of a local insect bite. At presentation, a single eschar was found on the right flank area. Blood tests showed leukopenia and thrombocytopenia (white blood cell [WBC] count 1,300/μL [reference 4,000–10,000]), platelet count 100 × 103/µL (reference 150–450), aspartate aminotransferase 134 IU/L (reference 8–38), alanine aminotransferase 77 IU/L (reference 4–44), C-reactive protein 0.39 mg/dL (reference 0.0–0.3), and ferritin 2,000 ng/mL (reference 13–150). She was administered ceftriaxone and azithromycin. A contrast-enhanced computed tomography scan revealed a wall enhancement in the distal esophagus. The patient became progressively more ill with altered mental status, profound leukopenia (WBC count 700/μL [reference 4,000–10,000]), and thrombocytopenia (platelet count 51 × 103/µL [reference 150–450]). On day 2, SFTSV was diagnosed by reverse transcriptase–polymerase chain reaction (RT-PCR) was achieved. On day 3, the indirect immunofluorescence immunoglobulin G assay for O. tsutsugamushi was positive with a titer of 1/160 at the Jeju Institute of Health and Environment and Laboratory. We performed a nested PCR assay for the 56-kD type specific antigen from her blood specimen because of the differential diagnoses of coinfection of acute O. tsutsugamushi infection.12 On day 4, her mental status, temperature, and laboratory values improved. Her overall illness improved after 3 days of azithromycin, followed by 4 days of doxycycline treatment and supportive treatment. The patient fully recovered without other complication at the time of this draft. This study was approved by the institutional review board (IRB) at the Jeju National University Hospital (IRB file no. 2015-11-001-003).

Molecular diagnosis and phylogenetic analysis of SFTSV.

RNA was extracted from stored patient serum using a QIAamp Viral RNA Mini kit (QIAGEN, Hilden, Germany). Reverse transcriptase–polymerase chain reaction was performed to amplify the complete small (S) segment of the viral RNA from the stored serum and confirmed SFTSV infection.6 Reverse transcriptase–polymerase chain reaction products (1,746 bp) were sequenced using the BigDye Terminator Cycle Sequencing kit (Perkin Elmer Applied Biosystems, Warrington, United Kingdom). Phylogenetic analysis of S segment sequences (1,746 bp) was performed with MEGA6, and phylogenetic trees were constructed using the maximum likelihood method, which confirmed SFTSV infection (Figure 1 and Supplemental Figure 1).13

Figure 1.
Figure 1.

Phylogenetic tree constructed based on complete S segments of SFTSV. The tree was constructed using the maximum likelihood method with MEGA 6.13 The complete S sequences from stored patient serum are shown in bold (JP13-Korea-2017). The S sequence data for the viruses identified from China, South Korea, and Japan were obtained from the National Center for Biotechnology Information/Basic Local Alignment Search Tool. SFTSV = severe fever with thrombocytopenia syndrome virus.

Citation: The American Journal of Tropical Medicine and Hygiene 99, 2; 10.4269/ajtmh.18-0088

Molecular diagnosis and phylogenetic analysis of genotype of O. tsutsugamushi.

DNA was extracted from stored patient serum using a QIAamp DNA Mini kit (QIAGEN). Nested PCR was performed to amplify the 56-kDa antigen of O. tsutsugamushi from the stored serum, confirming O. tsutsugamushi infection.12 Nested-PCR products (483 bp) were cloned using a TA cloning kit (Invitrogen, Carlsbad, CA). Five clones of each product were randomly picked and sequenced using the BigDye Terminator Cycle Sequencing kit (Perkin Elmer Applied Biosystems) because PCR product signals were weak when visualized in a gel and TA cloning and sequencing would lead to differentiation of a mixed infection with two genotypes (Supplemental Figure 1). Phylogenetic analysis of O. tsutsugamushi 56-kDa gene sequences (483 bp) was performed with MEGA6, phylogenetic trees were constructed using the maximum likelihood method, and the two sequences obtained from the patient clustered within the Boryong and Taguchi genotype, respectively, of O. tsutsugamushi (Figure 2 and Supplemental Figure 1).13

Figure 2.
Figure 2.

Phylogenetic tree constructed based on the Orientia tsutsugamushi 56-kDa gene sequences. The tree was constructed using the maximum likelihood method with MEGA 6.13 The O. tsutsugamushi 56-kDa gene sequences from stored patient serum are shown in bold as JP13-Korea-2017 (Boryong) and JP13-Korea-2017 (Taguchi). The O. tsutsugamushi 56-kDa gene sequence data identified from China, South Korea, Japan, and India were obtained from NCBI/BLAST.

Citation: The American Journal of Tropical Medicine and Hygiene 99, 2; 10.4269/ajtmh.18-0088

RESULTS AND DISCUSSION

Severe fever with thrombocytopenia syndrome is a tick-borne viral disease with a high mortality rate that infects humans primarily through tick bites.2,68,10 Transmission from person-to-person has been confirmed in close contact person such as family members and health-care workers, and asymptomatic infection via person-to-person close contact has also been reported.68

Scrub typhus is an acute febrile illness caused by O. tsutsugamushi, a bacterium transmitted to humans through chigger mite (Acari: Trombiculiidae) bites.10,11

The vectors of SFTSV and O. tsutsugamushi are different; H. longicornis is a major vector of SFTSV and Leptotrombidium spp. mites are vectors of O. tsutsugamushi.2,6,10,11 However, a previous report showed that SFTSV was detected by RT-PCR in L. scutellare mite bites in China, suggesting that the mites are a potential vector for SFTSV; Wi et al.10 suggested the possibility of coinfection with SFTSV and O. tsutsugamushi in South Korea.14 However, there has been no report of mixed infection with both SFTSV and different genotypes of O. tsutsugamushi in patients, although Zhang et al.15 showed two genotypes of O. tsutsugamushi coinfected a patient in China. In this study, we first confirmed the case of mixed infection with both SFTSV and two (Boryong and Taguchi) genotypes of O. tsutsugamushi in a patient (Figures 1 and 2, and Supplemental Figure 1); we suggest that mixed infection with both SFTSV and single or different genotypes of O. tsutsugamushi could have occurred in the patient at the same time in endemic regions, such as China, South Korea, and Japan.10

The signs and symptoms of SFTS and scrub typhus are similar, but SFTS exhibits a higher mortality rate than dose scrub typhus.2 Therefore, it is important to consider SFTSV infection in patients having scrub typhus, who may be coinfected with SFTSV.10 Thus, there is a need to better understand the ecological transmission dynamics and geographic distribution of SFTSV and O. tsutsugamushi in endemic countries, such as China, South Korea, and Japan.

Supplementary Material

Acknowledgment:

The authors thank L. Bakkensen for providing comments on this manuscript.

REFERENCES

  • 1.

    International Committee on Taxonomy of Viruses (ICTV), 2017. Taxonomy. Available at: https://talk.ictvonline.org/taxonomy/. Accessed June 22, 2018.

  • 2.

    Yu XJ 2011. Fever with thrombocytopenia associated with a novel bunyavirus in China. N Engl J Med 364: 15231532.

  • 3.

    McMullan LK 2012. A new phlebovirus associated with severe febrile illness in Missouri. N Engl J Med 367: 834841.

  • 4.

    Mourya DT 2014. Malsoor virus, a novel bat phlebovirus, is closely related to severe fever with thrombocytopenia syndrome virus and heartland virus. J Virol 88: 36053609.

    • Search Google Scholar
    • Export Citation
  • 5.

    Wang J 2014. Novel phlebovirus with zoonotic potential isolated from ticks, Australia. Emerg Infect Dis 20: 10401043.

  • 6.

    Yun Y 2015. Phylogenetic analysis of severe fever with thrombocytopenia syndrome virus in South Korea and migratory bird routes between China, South Korea, and Japan. Am J Trop Med Hyg 93: 468474.

    • Search Google Scholar
    • Export Citation
  • 7.

    Yoo JR, Heo ST, Park D, Kim H, Fukuma A, Fukushi S, Shimojima M, Lee KH, 2016. Family cluster analysis of severe fever with thrombocytopenia syndrome virus infection in Korea. Am J Trop Med Hyg 95: 13511357.

    • Search Google Scholar
    • Export Citation
  • 8.

    Bao CJ 2011. A family cluster of infections by a newly recognized bunyavirus in eastern China, 2007: further evidence of person-to-person transmission. Clin Infect Dis 53: 12081214.

    • Search Google Scholar
    • Export Citation
  • 9.

    Denic S, Janbeith J, Nair S, Conca W, Tariq WU, Al-Salam S, 2011. Acute thrombocytopenia, leucopenia, and multiorgan dysfunction: the first case of SFTS Bunyavirus outside China? Case Rep Infect Dis 2011: 204056.

    • Search Google Scholar
    • Export Citation
  • 10.

    Wi YM, Woo HI, Park D, Lee KH, Kang CI, Chung DR, Peck KR, Song JH, 2016. Severe fever with thrombocytopenia syndrome in patients suspected of having scrub typhus. Emerg Infect Dis 22: 19921995.

    • Search Google Scholar
    • Export Citation
  • 11.

    Weitzel T, Dittrich S, López J, Phuklia W, Martinez-Valdebenito C, Velásquez K, Blacksell SD, Paris DH, Abarca K, 2016. Endemic scrub typhus in South America. N Engl J Med 375: 954961.

    • Search Google Scholar
    • Export Citation
  • 12.

    Lee YM, Kim DM, Lee SH, Jang MS, Neupane GP, 2011. Phylogenetic analysis of the 56 kDa protein genes of Orientia tsutsugamushi in southwest area of Korea. Am J Trop Med Hyg 84: 250254.

    • Search Google Scholar
    • Export Citation
  • 13.

    Tamura K, Stecher G, Peterson D, Filipski A, Kumar S, 2013. MEGA6: molecular evolutionary genetics analysis version 6.0. Mol Biol Evol 30: 27252729.

    • Search Google Scholar
    • Export Citation
  • 14.

    Wang QK, Ge HM, Li ZF, Shan YF, Cui L, Wang YP, 2012. Vector research of severe fever with thrombocytopenia syndrome virus in gamasid mites and chigger mites [in Chinese]. Zhongguo Meijie Shengwuxue Ji Kongzhi Zazhi 23: 452454.

    • Search Google Scholar
    • Export Citation
  • 15.

    Zhang M, Zhao ZT, Wang XJ, Li Z, Ding L, Ding SJ, Yang LP, 2014. Mixed scrub typhus genotype, Shandong, China, 2011. Emerg Infect Dis 20: 484485.

Author Notes

Address correspondence to Keun Hwa Lee, Department of Microbiology and Immunology, Jeju National University College of Medicine, 15 Aran 13-gil, Jeju 63241, South Korea. E-mail: yomust7@jejunu.ac.kr

Financial support: This work was supported by a grant obtained from the National Research Foundation of Korea (NRF), the Ministry of Science, ICT, and Future Planning (grant number: NRF-2016M3A9B6021161).

Authors’ addresses: Jeong Rae Yoo, Sang Taek Heo, Ji-Hoon Kang, Dahee Park, Jeong Soon Kim, Jeong Hoon Bae, Jong Jin Woo, Suhyun Kim, and Keun Hwa Lee, Department of Microbiology and Immunology, Jeju National University College of Medicine, Jeju, South Korea, E-mails: mdyoojr@gmail.com, neosangtaek@naver.com, jhkang@jejunu.ac.kr, archons1004@ naver.com, alawy3424@naver.com, ezrabae@naver.com, wooman1995@naver.com, shyunkim19@pupils.nlcsjeju.kr, and yomust7@jejunu.ac.kr.

These authors contributed equally to this work.

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