ABSTRACT
@#Objective To develop and verify a plaque method for detection of infectious titer of tick-borne encephalitis virus(TBEV)strain(PHKT strain for short)adapted to primary hamster kidney(PHK)cells.Methods PHK cells were infected with TBEV,a primary mouse brain adaption strain,and passed consecutively for 12 passages.The titer of PHKT was detected by plaque method(Monolayer BHK-21 cells were infected with PHKT of various passages at different dilution ratios,and the plaque number was calculated by neutral red staining)and challenge titration in mouse brain(Mice were challenged with PHKT of various passages at different dilution ratios through brain cavity,0.03 mL for each,observed continuously for 14 days,and calculated for the median lethal dose(LD50)by Reed-Muench method)respectively,and the correlation between the results of two methods was analyzed.The developed plaque method for the detection of TBEV titer was verified for specificity,repeatability and intermediate precision.Results The plaque titer of PHKT virus was up to8.9 lgPFU/mL;The correlation between the results of plaque method and mouse brain challenge titration method was good(r = 0.92);The specificity of plaque method for detecting infectious titer of PHKT virus was good,and the coefficients of variation(CVs)of repeatability and intermediate precision were both less than 5%.Conclusion A plaque method for detecting infectious titer of PHKT virus was developed,which may be used as an alternative method for challenge titration in mouse brain.
ABSTRACT
@#Objective To express and purify the E protein Domain Ⅲ(ED Ⅲ) of tick-borne encephalitis virus(TBEV) in tandem and prepare the corresponding polyclonal antibody.Methods The TBEV RNA was extracted by Trizol method,and then reversely transcribed into cDNA,which was used as template to amplify ED Ⅲ gene fragment by PCR.Two ED Ⅲ gene fragments were ligated into fusion gene by the hydrophobic flexible polypeptide(G_4S)_3 using overlapping PCR,which was then linked to prokaryotic expression vector pET-28a(+) to construct the recombinant expression plasmid pET-28a-2ED Ⅲ.After sequencing,pET-28a-2ED Ⅲ was transformed into E.coli BL21(DE3) competent cells,induced by IPTG and purified by Ni~(2+) affinity chromatography.Female New Zealand white rabbits were immunized with the renatured recombinant protein to prepare polyclonal antibody.The antibody titer was detected by indirect ELISA and the specificity was identified by Western blot.The homology of ED Ⅲ amino acid sequence between TBEV and other flaviviruses was analyzed by DNAMAN software.Results The recombinant plasmid pET-28a-2ED Ⅲ was identified by sequencing,and the amplified sequence contained two genes consistent with the E sequence of TBEV "Senzhang" strain(JQ650523.1) included on GenBank,indicating that the recombinant plasmid was constructed correctly.The recombinant 2ED Ⅲ protein was expressed mainly in the form of inclusion bodies,with a relative molecular mass of about 21 000 and a purity of 97.5%.The titer of rabbit anti-2ED Ⅲ serum polyclonal antibody was 1:10~7,which reacted specifically with TBEV whole virus.DNAMAN software alignment showed that the homology of ED Ⅲ amino acid sequences between TBEV and Japanese encephalitis virus(JEV),yellow fever virus(YFV) and Dengue virus(DENY) was 36.56%,9.28% and 30.77%,respectively.Conclusion The TBEV envelope ED Ⅲ tandem recombinant expression plasmid pET-28a-2ED Ⅲ was successfully constructed.The expressed recombinant 2ED Ⅲ protein had good reactivity and immunogenicity,and the prepared polyclonal antibody had high titer.
ABSTRACT
Background@#Medical geography deals with the application of major concepts and theories derived from human and physical geography to issues of health and disease. Between1970-1980, Russian scientists were first figured landscape, geographical distribution of TBE in Mongolia. Since human cases of TBD were registered from 2005, around 2000 cases of TBD were registered. From 15% of diseases and 78% of fatal cases were tick-borne encephalitis. Therefore, were tried to create current geographical distribution of TBE in Mongolia and detect risk areas. @*Мaterials and Methods@#287 TBE cases data, information of TBE positive tick and human data were analyzed which registered in NCZD between 2005-2017. Arc GIS 9 were used for create map. Mongolian map was divided by 5 landscape range such as forest-taiga, forest-steppe, steppe, steppe-desert, gobi and high mountain. @*Result@#In forest-taiga range, 57% of TBE cases and incidence was 9.51 per 10000 population. 56.4% of I.persulcatus tick, 1.9% of D.nuttalli tick were found and infection rate of tick was Ixodes persulcatus-6.97%, Dermacentor nuttalli-5.2%. Seroprevalence of TBE was 25±12.1 among population. </br> In forest-steppe range, 40% of TBE cases and incidence was 0.56 per 10000 population. 43.6% of I.persulcatus tick, 44.3% of D.nuttalli, 24.4% of D.silvarum tick tick were found and infection rate of tick was Ixodes persulcatus-3.08%, D.silvarum-1.56% and D.nuttalli-1.56%. Seroprevalence of TBE was 14.5±11 among population.</br> In steppe range, 0.7% of TBE cases and incidence was 0.12 per 10000 population. 62.2% of D.silvarum tick, 23.9% of D.nuttalli tick were found and infection rate of tick was D.nuttalli-2.81% and D.silvarum-1.2%. Seroprevalence of TBE was 16.3±6.5 among population.</br> In other range including steppe-desert, gobi and high mountain, 2.8% of TBE cases and incidence was 0.1-0.27 per 10000 population. 62.2% of D.silvarum tick, 47.6% of D.nuttalli tick were found and infection rate of tick was D.nuttalli-0.84%. Seroprevalence of TBE was 2.5-13.1 among population.@*Conclusion@#Natural foci of tick-borne encephalitis have been registered in all landscape ranges of Mongolia and higher risk area of those ranges were forest-taiga and forest-steppe. Dermacentor silvarum, Dermacentor nuttalli tick becoming dominant vector of TBE in steppe range.
ABSTRACT
@#In Mongolia, the incidence and fatality rates of tick-borne encephalitis (TBE) have been increasing. We aimed to identify the epidemiological and molecular characteristics of tick-borne encephalitis virus (TBEV) associated with fatal meningoencephalitis in Mongolia. We conducted a descriptive study of 14 fatal cases of TBE that occurred between 2008 and 2017 in Mongolia. Reverse transcription polymerase chain reaction (RT–PCR) was used to detect viral RNA in brain tissue. RT–PCR products from six patients who died from TBE between 2013 and 2017 were directly sequenced and analysed phylogenetically. Ticks collected from Selenge and Bulgan provinces were also tested for TBEV by RT–PCR. Between 2008 and 2017, there were 14 fatal TBE cases in hospitals in Mongolia. The 14 patients who died reported receiving tick bites in Bulgan or Selenge province; 71.4% of deaths resulted from tick bites in Bulgan province. The TBE case fatality rate was 28.6% for patients in Bulgan province and 2.7% for those in Selenge province. All of the fatalities were men; the median age was 45 ± 12.6 years. Tick bites occurred between April and June in forested areas. In 2013, a 388 base pair fragment of the envelope (E) gene was obtained from a hospitalized patient. The closest relatives of this virus are Far-Eastern TBEV isolates. The case fatality rate differed between two provinces where tick bites occurred. A higher number of TBE cases and the virulent Far-Eastern subtype occurred in patients in Bulgan province. This province should increase vaccination coverage, training, education and investigations.
ABSTRACT
OBJECTIVE@#Tick-borne encephalitis virus (TBEV) is an emerging pathogen in Europe and North Asia that causes tick-borne encephalitis (TBE). A simple, rapid method for detecting TBEV RNA is needed to control this disease.@*METHODS@#A reverse-transcription recombinase-aided amplification (RT-RAA) assay was developed. This assay can be completed in one closed tube at 39 °C within 30 minutes. The sensitivity and specificity of RT-RAA were validated using non-infectious synthetic RNA representing a fragment of the NS5 region of the wild-type (WT) TBEV genome and the Senzhang strain. Additionally, 10 batches of tick samples were used to evaluate the performance of the RT-RAA assay.@*RESULTS@#The analytical limit of detection of the assay was 20 copies per reaction of the TBEV synthetic transcript and 3 plaque-forming units (pfu) per reaction of TBEV titers. With the specific assay, no signal due to other arboviruses was observed. Of the 10 batches of tick samples obtained from the Changbai Mountains of China, three were TBEV-positive, which was consistent with the results of the quantitative real-time PCR assay.@*CONCLUSION@#A rapid, highly sensitive, specific, and easy-to-use method was developed for the detection of the TBEV Far-Eastern subtype.
Subject(s)
Encephalitis Viruses, Tick-Borne , Genetics , Nucleic Acid Amplification Techniques , RNA, ViralABSTRACT
Objective@#To analyze the epidemiological characteristics and distribution characteristics of tick-borne encephalitis in China in 2014, and to provide scientific basis for formulating specific prevention and control measures.@*Methods@#The epidemic data were obtained from the "infectious disease report information management system" , using Excel 2016, GIS and other software to summarize and analyze the cases of tick borne encephalitis (TBE) reported, using the number of cases, incidence, composition ratio and other indicators to analyze and describe the TBE epidemiological characteristics in China in 2014.@*Results@#In 2014, a total of 322 cases of TBE were reported in 9 provinces in China, with an annual incidence of 0.024/100, 000 and 1 death of patient. The provinces with high number of cases were Jilin province, Inner mongolia autonomous region and Heilongjiang province, and the number of cases in the other six provinces is no more than two. TBE was distributed in spring and summer, and it is concentrated in May to July. The age of the affected population was mostly concentrated in 40-49 years old, the male-female ratio was 1.6∶1 (198/124), and the patients were dominantly farmers, household and unemployed workers, and forestry workers, they accounted for 49.40% (159/322), 26.40% (85/322) and 18.60% (60/322) of the national TBE cases respectively. The three hospitals that reported the most TBE cases in 2014 were Inner mongolia forestry general hospital, Jiangyuan People′s hospital of Baishan city, Jilin province and Mudanjiang forestry central hospital of Heilongjiang province. The number of reported cases in these three hospitals accounted for 68.6% of the whole country. The laboratory diagnosis rate of Inner mongolia forestry general hospital was the highest (91.9%).@*Conclusions@#In 2014, the incidence of TBE in China has continued to rise compared with the previous two years. The geographical focus is mainly on the forest areas of Daxing′anling, Xiaoxing′anling and Changbai Mountain.
ABSTRACT
To investigate the genotype and the biological characteristics of Tick-borne encephalitis virus (TBEV) in Charles Hilary endemic foci in the China-Kazakhstan border area in Xinjiang,ticks were collected by flagging during May to June in 2012 and 2014,and were stored in liquid nitrogen.TBEV strains were isolated from tick samples by inoculating BALB/c mice and BHK-21 cells.The FE gene fragments of TBEV-Far and the S gene fragments of TBEV-Sib were detected by RT-PCR from infected mice brain tissue and BHK cells,and then subjected to sequence alignment.Totally 16 TBEV strains were isolated from Ixodes persulcatus and Dermuceuter silvarum,among 13 strains were Far eastern subtype,three strains were Siberian subtype.It was first time that the TBEV-Sib was isolated in China.The Charles Hilary TBE natural foci were in the China-Kazakhstan border area,and both TBEV-Far and TBEV-Sib co-circulated.
ABSTRACT
Tick-borne encephalitis,also called forest encephalitis,is caused by tick-borne encephalitis virus.Central nervous system lesion is the major clinical symptom of tick-borne encephalitis,as an acute infectious disease,the case fatality rate is as high as 10％-20％.Virology experts consider it as a key and difficult point in recent years.This paper summarizes the progress in research of epidemiological characteristics,pathogenesis,clinical manifestations,outcome,diagnosis and treatment of tick-borne encephalitis to provide evidence for the prevention and treatment of tick-borne encephalitis.
ABSTRACT
Tick-borne encephalitis,also called forest encephalitis,is caused by tick-borne encephalitis virus.Central nervous system lesion is the major clinical symptom of tick-borne encephalitis,as an acute infectious disease,the case fatality rate is as high as 10％-20％.Virology experts consider it as a key and difficult point in recent years.This paper summarizes the progress in research of epidemiological characteristics,pathogenesis,clinical manifestations,outcome,diagnosis and treatment of tick-borne encephalitis to provide evidence for the prevention and treatment of tick-borne encephalitis.
ABSTRACT
Tick-borne encephalitis ( TBE) is a well-known central nervous system ( CNS) infection in children. The disease in children is generally milder,although severe illness may occur and even lead to perma-nent impairment of the quality of life due to neuropsychological sequelae. We review the epidemiological and clinical characteristics of tick-borne encephalitis,and summarise biological and virological aspects that are impor-tant for understanding the life-cycle and transmission of the virus. Tick-borne encephalitis virus is a flavivirus that is transmitted by ixodes persulcatus. Tick-borne encephalitis causes acute meningoencephalitis. The serologi-cal diagnosis is usually straightforward. No specific treatment for the disease exists,and immunisation is the main preventive measure.
ABSTRACT
Tickborne encephalitis (TBE) is a human viral infectious disease caused by tickborne encephalitis virus (TBEV). It is transmitted by the bite of an infected tick and also initiated the swelling of brain (encephalitis) and spinal cord. There is a pressing need to develop potent and sufficient amount of drugs and vaccines for control of TBE. We have selected the structural proteins such as anchored core protein C, core protein C, premembrane, matrix and envelope proteins of TBEV for identification of T-cell epitopes using immunoinformatics tools. These epitopes were showed the highest binding affinity with major histocompatibility complex (MHC) class I and II molecules. These finding may be used as an immunodiagnostic agent and also development of peptide based novel vaccines.
ABSTRACT
Objective To identify the infection and the replication of Tick-borne encephalitis virus(TBEV) in human neuroblastoma cells.Methods After being inffected with TBEV,the cell culture supernatant of human neuroblastoma cell line SK-N-SH was collected and assayed at different time points.Byusing real-time RT-PCR and plaque assay to measure the titer of virus in the supernatant,the replication andproliferation of TBEV in human neuroblastoma cell was identified.Meanwhile,the morphological change of SK-N-SH after TBEV infection was also visualized by observation under microscope and immunmquorescenceassay.Results Real-time RT-PCR and plaque assay both demonstrated that TBEV could replicate effectively in SK-N-SH cells,the peak titer could reach 2.92× 107 PFU/ml on 3 days post-inoculation.And significant morphological change occured on infected SK-N-SH cells after 2 days post inoculation.By immunofluorescence assay,the virus particles could be detected and visualized.Conclusion TBEV can replicate andproliferate effcctively and cause significant cell morphological changes in human neuroblastoma cell SK-N-SH,which demonstrated that SK-N-SH could be a suitable cell model for TBEV culture.
ABSTRACT
The number of tick borne diseases is increasing in the world. More than 100000 tick borne encephalitis, tick borne encephalitis cases were registered every year. It occurred in 29 Europien, 4 Asian countries and became public health concern [1]. In our country, virus, tick detection started since 1980 with collaboration Russian scientists. From 1998, collaborative team of Public Health Institute (PHI), National centre for communicable diseases (NCCD), National center for infectious diseases with natural foci (NCIDNF) and Rssian scientists started study of tick prevalence and infection of tick borne encephalitis in Khuvsgul, Khentii, Bulgan, Orkhon, Tuv province. In study of B.Byambaa, M.Dash (1994), 18 species tick were found in Mongolia. Ticks found in 27 soums of 7 provinces. TBE virus infection of tick was 1.2-16.7% in I.persulcatus, 13.7-20% in D.nutalli. Far eastern subtype founded from patient, Siberian subtype founded from tick in Bulgan province of Mongolia. TBEV infection was 1.1-39% among population; highest infection was in Bugan, Khuder soum of Selenge province, Dadаl soum of Khentii provinces. 57.2-59.4% of population was tick bitten and 21-73.7% of them were developed clinical symptoms during surveillance. Symptoms include redness, fever, headache, skin rash, join paint. Most TBE cases were developed fever, headache, vomiting, stiff neck, paralysis. 96% of them typical, 4.4% of them atypical, 60% of them fever, 13.3% meningial, 10% meningoencephalitis among 90 cases in 1998-2004. In review, clinical symptoms of TBE cases that occurred Mongolia similar to cases caused Sibirein subtype.
ABSTRACT
@# A 33-year-old male patient with tick-borne encephalitis (TBE) was reviewed, who presented with severe neurological deficits following TBEV infection, and improved in his motor and quality of life after an individualized rehabilitation.
ABSTRACT
The prevalence of tick-borne encephalitis virus (TBEV) in southern Korea was determined by collecting ticks using tick drags. A total of 4,077 of 6,788 ticks collected were pooled (649 pools) according to collection site, species, and developmental stage and assayed for TBEV. The TBEV protein E and NS5 gene fragments were detected using RT-nested PCR in six pools of nymphs collected from Jeju Island (2,491 ticks). The minimum field detection rates for TBEV were 0.17% and 0.14% for Haemaphysalis longicornis and Haemayphysalis. flava nymphs, respectively. The 252 bp NS5 and 477 bp protein E gene amplicons were sequenced. Phylogenetic analysis showed that the NS5 and protein E genes of the Jeju strain were clustered with Western subtype (98.0% and 99.4% identity, respectively). The Western subtype of TBEV is endemic in Korea, including Jeju Island. The study of vector and zoonotic host susceptibility to TBEV is required to better understand its potential impact on public health.