molecular epidemiology of hantaviruses in the czech republic

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  • Feb 04, 2020

Figure - Molecular Epidemiology of Hantaviruses in the Czech ...
Figure – Molecular Epidemiology of Hantaviruses in the Czech …

During 2008-2018, we collected samples from rodents and patients throughout the Czech Republic and isolate hantavirus marked. We detected Dobrava-Belgrade and orthohantaviruses Puumala in patients and Dobrava-Belgrade, Tula, and Seewis orthohantaviruses in rodents. Increased knowledge about eco-epidemiology of hantaviruses will raise awareness among doctors and better outcomes of patients.

The most prevalent in European hantaviruses Tula virus, Puumala virus (PUUV), and Dobrava-Belgrade virus (DOBV), all orthohantaviruses; PUUV and DOBV causes dengue with renal syndrome (). Four DOBV genotype virulences different in humans are known: the pathogenic Saaremaa; Kurkino, which causes mostly mild disease; and Dobrava and Sochi, which is both very pathogenic ().

Tula virus is the most frequently detected hantavirus in rodents in the Czech Republic, followed by DOBV PUUV in Moravia and South Bohemia. Seroprevalence of hantaviruses in humans in the Czech Republic is 1% -1.4% (). In 2009, one case DOBV infections in patients admitted to hospital were reported in the Czech Republic (), and in 2011, two more occurred in the Czech Republic-Slovakia border (); in 2017, DOBV fatal cases reported (). All 4 of these cases are classified as Dobrava genotype by PCR and sequencing. Overall, 82 hantavirus infections reported in humans in the Czech Republic during 2008-2017 (). In this study, we aimed to determine the location of the reservoir DOBV in the Czech Republic and molecular characteristics of the positive samples

During 2010-2017, we collected 1,551 wild rodents from different localities of the Czech Republic. 618 -necked yellow mice (Apodemus flavicollis), 37 wood mouse (A. sylvaticus), 222 striped field mouse (A. agrarius), 445 mice bank (Myodes glareolus), 40 public mice (Microtus arvalis), and 189 field mice ( Microtus Agrestis). We were stuck all mice were determined by the Animal Protection Law No. 246/1992 of the Czech Republic. In addition, we obtained 61 clinical samples obtained from patients with hantavirus infection during 2008-2018; hantavirus diagnosis is based on serology (specific hantavirus detection of IgG and IgM) and clinical and laboratory findings (fever, renal dysfunction, thrombocytopenia).

We tested a sample of human serum with Anti-Hantavirus Pool 1 “Eurasia” ELISA (IgG and IgM) and confirmed the results of hantavirus earlier by immunoblot EUROLINE Anti-Hanta Profile 1 (IgG and IgM) (both Euroimmun,) , We isolated RNA from rat lung samples and human serum, plasma and whole blood samples using a QIAamp Viral RNA Kit (QIAGEN,) and QIAzol (QIAGEN) or Trizol (Invitrogen,). We screened rodent and human RNA samples for hantavirus RNA using reverse transcription PCR amplified a 390-bp fragment of the large (L) segment (). We tested the segment L-positive samples with additional PCR targeting the area of ​​the medium (M) and (S) small segment (). We analyze hantavirus sequences using BLAST (), aligned with Bioedit (), and the phylogenetic tree constructed using MEGA 7.0 ().

Of 1,551 samples of mice, 43 (2.77%) were PCR positive for hantavirus. These 43 animals of three sample areas: Celadna in the Beskydy Mountain (2010, n = 9), Petrovice u Karvine (2014, n = 33), and Velká Stolová Mountain (2014, n = 1). From 9 hantavirus-positive mice were collected in 2010, we recovered 6 sequence: 2 sequence identical DOBV Dobrava 2 A. flavicollis mice. (GenBank accession no MK605679) and 4 sequences (2 identical) Seewis virus in 3 A. flavicollis mice and 1 A. sylvaticus rat (GenBank accession nos. MK605682-4). The Seewis virus sequences clustered with sequences derived from mice of Beskydy Mountain region (GenBank accession nos JQ425316, JQ425337, JQ425340 ;. data not shown). Of the 33 positive A. agrarius rat trapped in Petrovice u Karvine, we recovered 10 Kurkino genotype sequence with 99.1% -100% similarity (GenBank accession nos. MK605680-1). We detected Tula virus (GenBank accession no. MK605685) in mice 1 field stuck in Velká Stolová Mountain.

Image

. The phylogenetic tree constructed by partial 195-bp fragment of DOBV large segments of the human and mouse, the Czech Republic, 2010-2018. The sequence of this study (bold) compared with sequences available from …

Of 61 patients seropositive, 32 is a positive PCR for DOBV and three PCR positive for PUUV (obtained outside the Czech Republic). We recovered a partial L segment sequence of 28 of 32 patients DOBV-positive (GenBank accession nos. MK605641-65). Because of low quality, we manually shortened this sequence to 195 bp. We obtained a partial (264 bp) sequence of M segment of 6 patients (GenBank accession nos. MK605666-71) and partial (531 bp) S segment sequences of 7 patients (GenBank accession nos. MK605672-8). We construct a phylogenetic tree for comparing the viral sequences from humans and rodents. L segment analysis revealed that the samples are grouped into two groups separated by DOBV genotype (Dobrava or Kurkino), and the viral sequences from the same area (regardless of human or animal origin) clustered into the same clades (). Grouping by DOBV genotype was also observed in the phylogenetic tree constructed by M and S segment (data not shown).

Most Dobrava-positive patients from the mountains Jeseníky (sea Silesia) and Beskydy (southern Silesia), whereas Kurkino cases occur in low-lying between two mountain areas (Figure). DOBV fatal cases only confirmed in the Czech Republic that the patient stay in Kladno District, Central Bohemia region (). The geographical distribution of genotypes DOBV apparently related to the distribution of Apodemus spp. rat (). The higher number of cases of hantavirus in Silesia may be caused by the increased prevalence of DOBV in rodents or could be the result of increased awareness among local physicians DOBV.

Ms. Zelena is an expert in medical microbiology served as head of the National Reference Laboratory for Arboviruses at the Institute of Public Health, Ostrava, Czech Republic, and a doctoral candidate at the University of Defense in Hradec Kralove, Czech Republic. Her main research interests are vectorborne viral diseases and zoonoses.

We would like to thank Hana Blazejova, Lenka Betasova, Kristyna Venclikova, and Juraj Pesko for technical assistance for trapping animals and surgery.

The research was partially funded by the Budget Section Czech Ministry of Education, Youth and Sports for certain higher education research. The authors acknowledge the financial support of the Project FIT (Pharmacology, Immunotherapy, nanotoxicology ;. No CZ.02.1.01 / 0.0 / 0.0 / 15_003 / 0000495, for kindergarten and DR), which is funded by the European Regional Development Fund and the Ministry of the Republic Agriculture (RO0518).

DOI: 10.3201 / eid2511.190449

First Publication Date: 30/09/2019

Please use the form below to send a letter to the author or contact them at the following address:

Hana Zelena, Institute of Public Health in Ostrava, National Reference Laboratory for Arboviruses, Partyzanske nam. 7, 70200 Ostrava, Czech Republic

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