ABSTRACT
Orthohantaviruses, etiological agents of hemorrhagic fever with renal syndrome (HFRS) and hantavirus cardiopulmonary syndrome, pose a critical public health threat worldwide. Hantaan orthohantavirus (HTNV) outbreaks are particularly endemic in Gyeonggi Province in northern area of the Republic of Korea (ROK). Small mammals were collected from three regions in the Gyeonggi Province during 2017 and 2018. Serological and molecular prevalence of HTNV was 25/201 (12.4%) and 10/25 (40%), respectively. A novel nanopore-based diagnostic assay using a cost-efficient Flongle chip was developed to rapidly and sensitively detect HTNV infection in rodent specimens within 3 h. A rapid phylogeographical surveillance of HTNV at high-resolution phylogeny was established using the amplicon-based Flongle sequencing. In total, seven whole-genome sequences of HTNV were newly obtained from wild rodents collected in Paju-si (Gaekhyeon-ri) and Yeoncheon-gun (Hyeonga-ri and Wangnim-ri), Gyeonggi Province. Phylogenetic analyses revealed well-supported evolutionary divergence and genetic diversity, enhancing the resolution of the phylogeographic map of orthohantaviruses in the ROK. Incongruences in phylogenetic patterns were identified among HTNV tripartite genomes, suggesting differential evolution for each segment. These findings provide crucial insights into on-site diagnostics, genome-based surveillance, and the evolutionary dynamics of orthohantaviruses to mitigate hantaviral outbreaks in HFRS-endemic areas in the ROK.
Subject(s)
Hantaan virus , Hemorrhagic Fever with Renal Syndrome , Orthohantavirus , Animals , Phylogeny , Hantaan virus/genetics , Orthohantavirus/genetics , Rodentia , Mammals , Republic of Korea/epidemiologyABSTRACT
Potential threat of smallpox bioterrorism and concerns related to the adverse effects of currently licensed live-virus vaccines suggest the need to develop novel vaccines with better efficacy against smallpox. Use of DNA vaccines containing specific antigen-encoding plasmids prevents the risks associated with live-virus vaccines, offering a promising alternative to conventional smallpox vaccines. In this study, we investigated the efficiency of toll-like receptor (TLR) ligands in enhancing the immunogenicity of smallpox DNA vaccines. BALB/c mice were immunized with a DNA vaccine encoding the vaccinia virus L1R protein, along with the cytosine-phosphate-guanine (CpG) motif as a vaccine adjuvant, and their immune response was analyzed. Administration of B-type CpG oligodeoxynucleotides (ODNs) as TLR9 ligands 24 h after DNA vaccination enhanced the Th2-biased L1R-specific antibody immunity in mice. Moreover, B-type CpG ODNs improved the protective effects of the DNA vaccine against the lethal Orthopoxvirus challenge. Therefore, use of L1R DNA vaccines with CpG ODNs as adjuvants is a promising approach to achieve effective immunogenicity against smallpox infection.
ABSTRACT
Staphylococcal enterotoxin B (SEB) is a potent bacterial toxin that causes inflammatory stimulation and toxic shock, thus it is necessary to detect SEB in food and environmental samples. Here, we developed a sensitive immunodetection system using monoclonal antibodies (mAbs). Our study is the first to employ a baculovirus expression vector system (BEVS) to produce recombinant wild-type SEB. BEVS facilitated high-quantity and pure SEB production from suspension-cultured insect cells, and the SEB produced was characterized by mass spectrometry analysis. The SEB was stable at 4 °C for at least 2 years, maintaining its purity, and was further utilized for mouse immunization to generate mAbs. An optimal pair of mAbs non-competitive to SEB was selected for sandwich enzyme-linked immunosorbent assay-based immunodetection. The limit of detection of the immunodetection method was 0.38 ng/mL. Moreover, it displayed higher sensitivity in detecting SEB than commercially available immunodetection kits and retained detectability in various matrices and S. aureus culture supernatants. Thus, the results indicate that BEVS is useful for producing pure recombinant SEB with its natural immunogenic property in high yield, and that the developed immunodetection assay is reliable and sensitive for routine identification of SEB in various samples, including foods.
Subject(s)
Bacterial Toxins , Staphylococcus aureus , Mice , Animals , Baculoviridae , Enterotoxins/analysis , Enzyme-Linked Immunosorbent Assay/methods , Antibodies, MonoclonalABSTRACT
Background and aims: Despite global vaccination efforts, the number of confirmed cases of coronavirus disease 2019 (COVID-19) remains high. To overcome the crisis precipitated by the ongoing pandemic, characteristic studies such as virus diagnosis, isolation, and genome analysis of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are necessary. Herein, we report the isolation and molecular characterization of SARS-CoV-2 from the saliva of patients who had tested positive for COVID-19 at Proving Ground in Taean County, Republic of Korea, in 2020. Methods: We analyzed the whole-genome sequence of SARS-CoV-2 isolated from the saliva samples of patients through next-generation sequencing. We also successfully isolated SARS-CoV-2 from the saliva samples of two patients by using cell culture, which was used to study the cytopathic effects and viral replication in Vero E6 cells. Results: Whole-genome sequences of the isolates, SARS-CoV-2 ADD-2 and ADD-4, obtained from saliva were identical, and phylogenetic analysis using Bayesian inference methods showed SARS-CoV-2 GH clade (B.1.497) genome-specific clustering. Typical coronavirus-like particles, with diameters of 70-120 nm, were observed in the SARS-CoV-2 infected Vero E6 cells using transmission electron microscopy. Conclusion: In conclusion, this report provides insights into the molecular diagnosis, isolation, genetic characteristics, and diversity of SARS-CoV-2 isolated from the saliva of patients. Further studies are needed to explore and monitor the evolution and characteristics of SARS-CoV-2 variants.
ABSTRACT
Seoul virus (SEOV), an etiological agent for hemorrhagic fever with renal syndrome, poses a significant public health threat worldwide. This study evaluated the feasibility of a mobile Biomeme platform for facilitating rapid decision making of SEOV infection. A total of 27 Rattus norvegicus were collected from Seoul Metropolitan City and Gangwon Province in Republic of Korea (ROK), during 2016-2020. The serological and molecular prevalence of SEOV was 5/27 (18.5%) and 2/27 (7.4%), respectively. SEOV RNA was detected in multiple tissues of rodents using the Biomeme device, with differences in Ct values ranging from 0.6 to 2.1 cycles compared to a laboratory benchtop system. Using amplicon-based next-generation sequencing, whole-genome sequences of SEOV were acquired from lung tissues of Rn18-1 and Rn19-5 collected in Gangwon Province. Phylogenetic analysis showed a phylogeographical diversity of rat-borne orthohantavirus collected in Gangwon Province. We report a novel isolate of SEOV Rn19-5 from Gangwon Province. Our findings demonstrated that the Biomeme system can be applied for the molecular diagnosis of SEOV comparably to the laboratory-based platform. Whole-genome sequencing of SEOV revealed the phylogeographical diversity of orthohantavirus in the ROK. This study provides important insights into the field-deployable diagnostic assays and genetic diversity of orthohantaviruses for the rapid response to hantaviral outbreaks in the ROK.
ABSTRACT
BACKGROUND: Whole-genome sequencing plays a critical role in the genomic epidemiology intended to improve understanding the spread of emerging viruses. Dabie bandavirus, causing severe fever with thrombocytopenia syndrome (SFTS), is a zoonotic tick-borne virus that poses a significant public health threat. We aimed to evaluate a novel amplicon-based nanopore sequencing tool to obtain whole-genome sequences of Dabie bandavirus, also known as SFTS virus (SFTSV), and investigate the molecular prevalence in wild ticks, Republic of Korea (ROK). PRINCIPAL FINDINGS: A total of 6,593 ticks were collected from Gyeonggi and Gangwon Provinces, ROK in 2019 and 2020. Quantitative polymerase chain reaction revealed the presence of SFSTV RNA in three Haemaphysalis longicornis ticks. Two SFTSV strains were isolated from H. longicornis captured from Pocheon and Cheorwon. Multiplex polymerase chain reaction-based nanopore sequencing provided nearly full-length tripartite genome sequences of SFTSV within one hour running. Phylogenetic and reassortment analyses were performed to infer evolutionary relationships among SFTSVs. Phylogenetic analysis grouped SFTSV Hl19-31-4 and Hl19-31-13 from Pocheon with sub-genotype B-1 in all segments. SFTSV Hl20-8 was found to be a genomic organization compatible with B-1 (for L segment) and B-2 (for M and S segments) sub-genotypes, indicating a natural reassortment between sub-genotypes. CONCLUSION/SIGNIFICANCE: Amplicon-based next-generation sequencing is a robust tool for whole-genome sequencing of SFTSV using the nanopore platform. The molecular prevalence and geographical distribution of SFTSV enhanced the phylogeographic map at high resolution for sophisticated prevention of emerging SFTS in endemic areas. Our findings provide important insights into the rapid whole-genome sequencing and genetic diversity for the genome-based diagnosis of SFTSV in the endemic outbreak.
Subject(s)
Bunyaviridae Infections , Nanopore Sequencing , Phlebovirus , Severe Fever with Thrombocytopenia Syndrome , Ticks , Animals , Bunyaviridae Infections/epidemiology , Genetic Variation , Multiplex Polymerase Chain Reaction , Phlebovirus/genetics , Phylogeny , RNA , Republic of Korea/epidemiologyABSTRACT
The potential use of biological agents has become a major public health concern worldwide. According to the CDC classification, Bacillus anthracis and Clostridium botulinum, the bacterial pathogens that cause anthrax and botulism, respectively, are considered to be the most dangerous potential biological agents. Currently, there is no licensed vaccine that is well suited for mass immunization in the event of an anthrax or botulism epidemic. In the present study, we developed a dual-expression system-based multipathogen DNA vaccine that encodes the PA-D4 gene of B. anthracis and the HCt gene of C. botulinum. When the multipathogen DNA vaccine was administered to mice and guinea pigs, high level antibody responses were elicited against both PA-D4 and HCt. Analysis of the serum IgG subtype implied a combined Th1/Th2 response to both antigens, but one that was Th2 skewed. In addition, immunization with the multipathogen DNA vaccine induced effective neutralizing antibody activity against both PA-D4 and HCt. Finally, the protection efficiency of the multipathogen DNA vaccine was determined by sequential challenge with 10 LD50 of B. anthracis spores and 10 LD50 of botulinum toxin, or vice versa, and the multipathogen DNA vaccine provided higher than 50% protection against lethal challenge with both high-risk biothreat agents. Our studies suggest the strategy used for this anthrax-botulinum multipathogen DNA vaccine as a prospective approach for developing emergency vaccines that can be immediately distributed on a massive scale in response to a biothreat emergency or infectious disease outbreak. Key points ⢠A novel multipathogen DNA vaccine was constructed against anthrax and botulism. ⢠Robust immune responses were induced following vaccination. ⢠Suggests a potential vaccine development strategy against biothreat agents.
Subject(s)
Anthrax Vaccines , Anthrax , Bacillus anthracis , Botulism , Vaccines, DNA , Animals , Anthrax/prevention & control , Antibodies, Bacterial , Antigens, Bacterial/genetics , Bacillus anthracis/genetics , Biological Warfare Agents , Botulism/prevention & control , Guinea Pigs , Immunity , Mice , Vaccines, DNA/geneticsABSTRACT
BACKGROUND: Hantavirus infection occurs through the inhalation of aerosolized excreta, including urine, feces, and saliva of infected rodents. The presence of Hantaan virus (HTNV) RNA or infectious particles in urine specimens of patient with hemorrhagic fever with renal syndrome (HFRS) remains to be investigated. METHODOLOGY/PRINCIPAL FINDINGS: We collected four urine and serum specimens of Republic of Korea Army (ROKA) patients with HFRS. We performed multiplex PCR-based next-generation sequencing (NGS) to obtain the genome sequences of clinical HTNV in urine specimens containing ultra-low amounts of viral genomes. The epidemiological and phylogenetic analyses of HTNV demonstrated geographically homogenous clustering with those in Apodemus agrarius captured in highly endemic areas, indicating that phylogeographic tracing of HTNV genomes reveals the potential infection sites of patients with HFRS. Genetic exchange analyses showed a genetic configuration compatible with HTNV L segment exchange in nature. CONCLUSION/SIGNIFICANCE: Our results suggest that whole or partial genome sequences of HTNV from the urine enabled to track the putative infection sites of patients with HFRS by phylogeographically linking to the zoonotic HTNV from the reservoir host captured at endemic regions. This report raises awareness among physicians for the presence of HTNV in the urine of patients with HFRS.
Subject(s)
Genome, Viral , Hantaan virus/isolation & purification , Hemorrhagic Fever with Renal Syndrome/virology , Urine/virology , Hantaan virus/classification , Hantaan virus/genetics , Hemorrhagic Fever with Renal Syndrome/urine , High-Throughput Nucleotide Sequencing , Humans , Multiplex Polymerase Chain Reaction , Phylogeny , Republic of KoreaABSTRACT
The South Korean government effectively contained the coronavirus disease-2019 (COVID-19) outbreak primarily associated with a religious group. We conducted SARS-CoV-2 whole genome sequencing of 66 cases to investigate connections among the initial South Korean cases and the religious group outbreak. We assessed the accuracy of genomic investigation by comparing the whole genome sequences with comprehensive contact tracing records. Five transmission clusters were estimated among the 15 initial cases. The six close-contact cases and two potential exposure pairs identified by contact tracing showed two or fewer nucleotide base differences. Additionally, we identified two transmission clusters that were phylogenetically distinct from the initial clusters, sharing common G11083T, G26144T, and C14805T markers. The strain closest to the two additional clusters was identified from a pair of identical sequences isolated from individuals who traveled from Wuhan to Italy. Our findings provide insights into the origins of community spread of COVID-19.
Subject(s)
COVID-19/pathology , SARS-CoV-2/genetics , Adolescent , Adult , Aged , Aged, 80 and over , COVID-19/epidemiology , COVID-19/transmission , COVID-19/virology , Child , Child, Preschool , Contact Tracing , Disease Outbreaks , Female , Humans , Infant , Male , Middle Aged , Phylogeny , RNA, Viral/chemistry , RNA, Viral/genetics , RNA, Viral/metabolism , Republic of Korea/epidemiology , SARS-CoV-2/classification , SARS-CoV-2/isolation & purification , Whole Genome Sequencing , Young AdultABSTRACT
BACKGROUND: Hantaan orthohantavirus (Hantaan virus, HTNV), harbored by Apodemus agrarius (the striped field mouse), causes hemorrhagic fever with renal syndrome (HFRS) in humans. Viral genome-based surveillance at new expansion sites to identify HFRS risks plays a critical role in tracking the infection source of orthohantavirus outbreak. In the Republic of Korea (ROK), most studies demonstrated the serological prevalence and genetic diversity of orthohantaviruses collected from HFRS patients or rodents in Gyeonggi Province. Gangwon Province is a HFRS-endemic area with a high incidence of patients and prevalence of infected rodents, ROK. However, the continued epidemiology and surveillance of orthohantavirus remain to be investigated. METHODOLOGY/PRINCIPAL FINDINGS: Whole-genome sequencing of HTNV was accomplished in small mammals collected in Gangwon Province during 2015-2018 by multiplex PCR-based next-generation sequencing. To elucidate the geographic distribution and molecular diversity of viruses, we conducted phylogenetic analyses of HTNV tripartite genomes. We inferred the hybrid zone using cline analysis to estimate the geographic contact between two different HTNV lineages in the ROK. The graph incompatibility based reassortment finder performed reassortment analysis. A total of 12 HTNV genome sequences were completely obtained from A. agrarius newly collected in Gangwon Province. The phylogenetic and cline analyses demonstrated the genetic diversity and hybrid zone of HTNV in the ROK. Genetic exchange analysis suggested the possibility of reassortments in Cheorwon-gun, a highly HFRS-endemic area. CONCLUSIONS/SIGNIFICANCE: The prevalence and distribution of HTNV in HFRS-endemic areas of Gangwon Province enhanced the phylogeographic map for orthohantavirus outbreak monitoring in ROK. This study revealed the hybrid zone reflecting the genetic diversity and evolutionary dynamics of HTNV circulating in Gangwon Province. The results arise awareness of rodent-borne orthohantavirus diseases for physicians in the endemic area of ROK.
Subject(s)
Genome, Viral , Hantaan virus/genetics , Murinae/virology , Animals , Antibodies, Viral , Endemic Diseases , Phylogeny , Republic of KoreaABSTRACT
Severe fever with thrombocytopenia syndrome virus (SFTSV) is an emerging human pathogen, endemic in areas of China, Japan, and the Korea (KOR). It is primarily transmitted through infected ticks and can cause a severe hemorrhagic fever disease with case fatality rates as high as 30%. Despite its high virulence and increasing prevalence, molecular and functional studies in situ are scarce due to the limited availability of high-titer SFTSV exposure stocks. During the course of field virologic surveillance in 2017, we detected SFTSV in ticks and in a symptomatic soldier in a KOR Army training area. SFTSV was isolated from the ticks producing a high-titer viral exposure stock. Through the use of advanced genomic tools, we present here a complete, in-depth characterization of this viral stock, including a comparison with both the virus in its arthropod source and in the human case, and an in vivo study of its pathogenicity. Thanks to this detailed characterization, this SFTSV viral exposure stock constitutes a quality biological tool for the study of this viral agent and for the development of medical countermeasures, fulfilling the requirements of the main regulatory agencies.
Subject(s)
Bunyaviridae Infections/virology , Hemorrhagic Fevers, Viral/virology , Phlebovirus/isolation & purification , Adult , Animals , Bunyaviridae Infections/genetics , Bunyaviridae Infections/metabolism , Female , Genome, Viral , Humans , Male , Mice , Phlebovirus/physiology , Phylogeny , Receptor, Interferon alpha-beta/genetics , Receptor, Interferon alpha-beta/metabolism , Republic of Korea , Ticks/virologyABSTRACT
BACKGROUND: Endemic outbreaks of hantaviruses pose a critical public health threat worldwide. Hantaan orthohantavirus (HTNV) causes hemorrhagic fever with renal syndrome (HFRS) in humans. Using comparative genomic analyses of partial and nearly complete sequences of HTNV from humans and rodents, we were able to localize, with limitations, the putative infection locations for HFRS patients. Partial sequences might not reflect precise phylogenetic positions over the whole-genome sequences; finer granularity of rodent sampling reflects more precisely the circulation of strains. METHODS: Five HFRS specimens were collected. Epidemiological surveys were conducted with the patients during hospitalization. We conducted active surveillance at suspected HFRS outbreak areas. We performed multiplex polymerase chain reaction-based next-generation sequencing to obtain the genomic sequence of HTNV from patients and rodents. The phylogeny of human- and rodent-derived HTNV was generated using the maximum likelihood method. For phylogeographic analyses, the tracing of HTNV genomes from HFRS patients was defined on the bases of epidemiological interviews, phylogenetic patterns of the viruses, and geographic locations of HTNV-positive rodents. RESULTS: The phylogeographic analyses demonstrated genetic clusters of HTNV strains from clinical specimens, with HTNV circulating in rodents at suspected sites of patient infections. CONCLUSIONS: This study demonstrates a major shift in molecular epidemiological surveillance of HTNV. Active targeted surveillance was performed at sites of suspected infections, allowing the high-resolution phylogeographic analysis to reveal the site of emergence of HTNV. We posit that this novel approach will make it possible to identify infectious sources, perform disease risk assessment, and implement preparedness against vector-borne viruses.
Subject(s)
Hantaan virus , Hemorrhagic Fever with Renal Syndrome , Orthohantavirus , Orthohantavirus/genetics , Hemorrhagic Fever with Renal Syndrome/epidemiology , Humans , Phylogeny , Watchful WaitingABSTRACT
Orthohantaviruses, negative-sense single-strand tripartite RNA viruses, are a global public health threat. In humans, orthohantavirus infection causes hemorrhagic fever with renal syndrome or hantavirus cardiopulmonary syndrome. Whole-genome sequencing of the virus helps in identification and characterization of emerging or re-emerging viruses. Next-generation sequencing (NGS) is a potent method to sequence the viral genome, using molecular enrichment methods, from clinical specimens containing low virus titers. Hence, a comparative study on the target enrichment NGS methods is required for whole-genome sequencing of orthohantavirus in clinical samples. In this study, we used the sequence-independent, single-primer amplification, target capture, and amplicon NGS for whole-genome sequencing of Hantaan orthohantavirus (HTNV) from rodent specimens. We analyzed the coverage of the HTNV genome based on the viral RNA copy number, which is quantified by real-time quantitative PCR. Target capture and amplicon NGS demonstrated a high coverage rate of HTNV in Apodemus agrarius lung tissues containing up to 103-104 copies/µL of HTNV RNA. Furthermore, the amplicon NGS showed a 10-fold (102 copies/µL) higher sensitivity than the target capture NGS. This report provides useful insights into target enrichment NGS for whole-genome sequencing of orthohantaviruses without cultivating the viruses.
Subject(s)
Hantaan virus/genetics , High-Throughput Nucleotide Sequencing/methods , Lung/virology , Murinae/virology , Whole Genome Sequencing/methods , Animals , Genome, Viral/genetics , Phylogeny , Real-Time Polymerase Chain Reaction , Republic of KoreaABSTRACT
Human adenovirus (HAdV) is a common pathogen causing respiratory infections with outbreaks reported in the military and community. However, little information is available on the shedding kinetics. We performed a prospective study of immunocompetent adults confirmed with HAdV respiratory infection by multiplex real-time PCR during an outbreak of HAdV-55. Consecutive respiratory specimens of sputum or nasopharyngeal swab were collected from each patient every 2 days. Viral load was measured by real-time quantitative PCR. Of 32 enrolled patients, 27 (84.4%) had pneumonia. Five patients (15.6%) received cidofovir. Viral load was highest in the earliest samples at 8.69 log10 copies/mL. In a linear regression model, viral load declined consistently in a log-linear fashion at the rate of - 0.15 log10 copies/mL per day (95% confidence interval (CI): - 0.18, - 0.12; R2 = 0.32). However, the regression model estimated the viral shedding duration to be 55 days. The rate of decline in viral load did not differ between patients who received cidofovir and who did not. Patients with prominent respiratory symptoms or extensive involvement on chest radiograph had higher volume of viral excretion. Prolonged viral shedding was observed in otherwise healthy adults with HAdV-55 respiratory infection. This finding should be considered in the establishment of infection control and prevention strategies.
Subject(s)
Adenovirus Infections, Human/diagnosis , Adenoviruses, Human/physiology , Respiratory Tract Infections/virology , Virus Shedding , Adenovirus Infections, Human/drug therapy , Adenoviruses, Human/classification , Adolescent , Disease Outbreaks , Humans , Immunocompetence , Linear Models , Male , Nasopharynx/virology , Pneumonia, Viral/diagnosis , Prospective Studies , Real-Time Polymerase Chain Reaction , Republic of Korea/epidemiology , Sputum/virology , Viral Load , Young AdultABSTRACT
Botulinum neurotoxins (BoNTs) produced by the spore-forming, gram-positive, anaerobic bacterium Clostridium botulinum are the most toxic substances known and cause botulism, flaccid paralysis, or death. Owing to their high lethality, BoNTs are classified as category A agents by the Centers for Disease Control (CDC). Currently, there are no vaccines available to protect against BoNTs, so the rapid development of a safe and effective vaccine is important. DNA-based vaccines have recently drawn great attention because they can be developed quickly and can be applied in mass vaccination strategies to prevent disease outbreaks. Here, we report on the immunogenic and protective efficacy of a DNA vaccine, encoding a 50-kDa carboxy-terminal fragment of the BoNT serotype E heavy chain, which is delivered via an intradermal route. This plasmid DNA vaccine induced robust humoral and cellular BoNT/E-specific immune responses and completely protected animals against lethal challenge with BoNT/E. These results not only indicate that DNA vaccines could be further developed as safe and effective candidates for vaccines against BoNTs but also suggest a possible approach for developing vaccines that protect against bio-threat toxins.
Subject(s)
Bacterial Vaccines/immunology , Botulinum Toxins/immunology , Immunization/methods , Injections, Intradermal , Vaccines, DNA/immunology , Animals , Antibodies, Bacterial/blood , Antibodies, Neutralizing/blood , Bacterial Vaccines/administration & dosage , Botulinum Toxins/administration & dosage , Botulism/prevention & control , Female , Immunity, Cellular , Immunity, Humoral , Mice, Inbred BALB C , Serogroup , Vaccines, DNA/administration & dosageABSTRACT
Seoul virus (SEOV) poses a worldwide public health threat. This virus, which is harbored by Rattus norvegicus and R. rattus rats, is the causative agent of hemorrhagic fever with renal syndrome (HFRS) in humans, which has been reported in Asia, Europe, the Americas, and Africa. Defining SEOV genome sequences plays a critical role in development of preventive and therapeutic strategies against the unique worldwide hantavirus. We applied multiplex PCR-based next-generation sequencing to obtain SEOV genome sequences from clinical and reservoir host specimens. Epidemiologic surveillance of R. norvegicus rats in South Korea during 2000-2016 demonstrated that the serologic prevalence of enzootic SEOV infections was not significant on the basis of sex, weight (age), and season. Viral loads of SEOV in rats showed wide dissemination in tissues and dynamic circulation among populations. Phylogenetic analyses showed the global diversity of SEOV and possible genomic configuration of genetic exchanges.
Subject(s)
Genetic Variation , Hemorrhagic Fever with Renal Syndrome/virology , Multiplex Polymerase Chain Reaction , Seoul virus/genetics , Animals , Genome, Viral , Global Health , Hemorrhagic Fever with Renal Syndrome/epidemiology , Humans , Phylogeography , RNA, Viral/genetics , Rats , Republic of Korea/epidemiology , Retrospective Studies , Seasons , Serologic TestsABSTRACT
Botulinum neurotoxins (BoNTs) are the most potent toxins to mammals. A toxoid vaccine was previously used for prevention of botulinum intoxication; however, this vaccine is no longer available. Currently, no approved botulinum vaccines are available from the Food and Drug Administration (FDA). Recently, a recombinant host cell receptor-binding subunit created for use as a potential vaccine completed phase 2 clinical trials. The current study designed a vaccine candidate against BoNT type A (BoNT/A) using a structural design. Our vaccine candidate was the BoNT/A heavy chain C-terminal region (HCR) that contained the point mutation BA15 (R1269A) within the ganglioside-binding site. A Biacore affinity test showed that the affinity of BA15 for ganglioside GT1b was 100 times lower than that of the HCR. A SNAP25 cleavage assay revealed that immunized sera blocked SNAP25 cleavage of the BoNT/A toxin via BA15. In an in vivo experiment, mice and guinea pigs immunized with BA15 produced neutralizing antibodies that protected against 3,000 LD50 of BoNT/A. In conclusion, the results of both in vitro and in vivo assays showed that our BA15 vaccine candidate was similar to the recombinant host cell receptor-binding subunit vaccine. The inability of BA15to bind ganglioside shows that BA15 is a potential safe vaccine candidate.
Subject(s)
Bacterial Vaccines/immunology , Botulinum Toxins, Type A/immunology , Recombinant Proteins/immunology , Amino Acid Sequence , Animals , Binding Sites , Botulinum Toxins, Type A/genetics , Botulism/prevention & control , Cell Line , Gangliosides/chemistry , Gangliosides/metabolism , Guinea Pigs , Immunoglobulin G/blood , Mice , Models, Molecular , Neurons/drug effects , Neurons/metabolism , Point Mutation , Protein Conformation , Protein SubunitsABSTRACT
Four mutations (N23A, Y90A, R110A and F177A) were introduced into S19, a vaccine candidate for staphylococcal enterotoxin B (SEB), resulting in a lower binding affinity towards the T-cell receptor beta chain (TCB) and reducing its superantigen activity. The structure of S19 was solved and was superposed on the native or complex structure of SEB. In the superposition model, mutations that were introduced seemed to reduce the number of hydrogen bonds at the SEB-TCB interface. S19 also displayed an unexpected structural change around the flexible-loop region owing to the Y90A mutation. This local structural change provided evidence that the mutated form of S19 could have a lower affinity for major histocompatibility complex (MHC) class II than wild-type SEB.
Subject(s)
Enterotoxins/chemistry , Enterotoxins/immunology , Mutation , Staphylococcal Vaccines/chemistry , Staphylococcal Vaccines/immunology , Crystallography, X-Ray , Enterotoxins/genetics , Histocompatibility Antigens Class II/immunology , Hydrogen Bonding , Receptors, Antigen, T-Cell, alpha-beta/immunology , Staphylococcal Vaccines/geneticsABSTRACT
Zika virus (ZIKV) (genus Flavivirus, family Flaviviridae) is an emerging pathogen associated with microcephaly and Guillain-Barré syndrome. The rapid spread of ZIKV disease in over 60 countries and the large numbers of travel-associated cases have caused worldwide concern. Thus, intensified surveillance of cases among immigrants and tourists from ZIKV-endemic areas is important for disease control and prevention. In this study, using Next Generation Sequencing, we reported the first whole-genome sequence of ZIKV strain AFMC-U, amplified from the urine of a traveler returning to Korea from the Philippines. Phylogenetic analysis showed geographic-specific clustering. Our results underscore the importance of examining urine in the diagnosis of ZIKV infection.
Subject(s)
Zika Virus Infection/virology , Humans , Philippines , Phylogeny , Republic of Korea , Travel , Whole Genome Sequencing/methods , Zika Virus/geneticsABSTRACT
An outbreak of febrile respiratory illness associated with human adenovirus (HAdV) occurred in the South Korea military during the 2014-15 influenza season and thereafter. Molecular typing and phylogenetic analysis of patient samples identified HAdV type 55 as the causative agent. Emergence of this novel HAdV necessitates continued surveillance in military and civilian populations.