Dengue virus (DENV) is the most widely transmitted arbovirus in the world. Due to the lack of diagnostic technology to quickly identify the virus serotypes in patients, severe dengue hemorrhagic fever cases caused by repeated infections remain high. To realize the rapid differential diagnosis of different serotypes of DENV infection by immunological methods, in this study, four DENV serotype NS1 proteins were expressed and purified in mammalian cells. Monoclonal antibodies (MAbs) against NS1 protein were obtained by hybridoma technology after immunizing BALB/c mice. Enzyme-linked immunosorbent assay, indirect immunofluorescence assay, dot blotting, and Western blotting were used to confirm the reactivity of MAbs to viral native NS1 and recombinant NS1 protein. These MAbs include not only the universal antibodies that recognize all DENV 1-4 serotype NS1, but also serotype-specific antibodies against DENV-1, DENV-2 and DENV-4. Double antibody sandwich ELISA was established based on these antibodies, which can be used to achieve rapid differential diagnosis of serotypes of DENV infection. Preparation of DENV serotype-specific MAbs and establishment of an ELISA technology for identifying DENV serotypes has laid the foundation for the rapid diagnosis of DENV clinical infection.
Animals ; Antibodies, Monoclonal ; Antibodies, Viral/metabolism* ; Dengue/diagnosis* ; Dengue Virus/immunology* ; Enzyme-Linked Immunosorbent Assay ; Humans ; Mice ; Mice, Inbred BALB C ; Sensitivity and Specificity ; Serogroup ; Viral Nonstructural Proteins/immunology*

A rapid diagnostic test (RDT) kit was developed to detect non-structural protein 1 (NS1) of yellow fever virus (YFV) using monoclonal antibody. NS1 protein was purified from the cultured YFV and used to immunize mice. Monoclonal antibody to NS1 was selected and conjugated with colloidal gold to produce the YFV NS1 RDT kit. The YFV RDTs were evaluated for sensitivity and specificity using positive and negative samples of monkeys from Brazil and negative human blood samples from Korea. Among monoclonal antibodies, clones 3A11 and 3B7 proved most sensitive, and used for YFV RDT kit. Diagnostic accuracy of YFV RDT was fairly high; Sensitivity was 0.0% and specificity was 100% against Dengue viruses type 2 and 3, Zika, Chikungunya and Mayaro viruses. This YFV RDT kit could be employed as a test of choice for point-of-care diagnosis and large scale surveys of YFV infection under clinical or field conditions in endemic areas and on the globe.
Animals ; Antibodies, Monoclonal ; Brazil ; Clone Cells ; Dengue Virus ; Diagnosis ; Diagnostic Tests, Routine ; Gold Colloid ; Haplorhini ; Humans ; Korea ; Mice ; Point-of-Care Systems ; Sensitivity and Specificity ; Yellow fever virus ; Yellow Fever

ELISAs and rapid diagnostic tests (RDTs) are widely used for diagnosing dengue virus (DENV) infection. Using 138 single blood samples, we compared the ability to detect non-structural (NS)-1 antigen and anti-DENV IgM/IgG antibodies among (1) DENV Detect NS1 ELISA, DENV Detect IgM capture ELISA and DENV Detect IgG ELISA (InBios International, Inc.); (2) Anti-Dengue virus IgM Human ELISA and Anti-Dengue virus IgG Human ELISA (Abcam); (3) Dengue virus NS1 ELISA, Anti-Dengue virus ELISA (IgM) and Anti-Dengue virus ELISA (IgG) (Euroimmun); (4) Asan Easy Test Dengue NS1 Ag 100 and Asan Easy Test Dengue IgG/IgM (Asan Pharm); (5) SD BIOLINE Dengue Duo (Standard Diagnostics); and (6) Ichroma Dengue NS1 and Ichroma Dengue IgG/IgM (Boditech Med). For NS1 antigen detection, InBios and Euroimmun showed higher sensitivities (100%) than the RDTs (42.9–64.3%). All tests demonstrated variable sensitivities for IgM (38.1–90.5%) and IgG (65.7–100.0%). InBios and Boditech Med demonstrated higher sensitivity (95.6% and 88.2%, respectively) than the other tests for combined NS1 antigen and IgM antibody. Five NS1 antigen tests had good agreement (92.8–98.6%) without showing positivity for chikungunya. However, all IgG tests demonstrated potential false-positivity with variable ranges. Clinical laboratories should note performance variations across tests and potential cross-reactivity.
Antibodies ; Chungcheongnam-do ; Dengue Virus ; Dengue ; Diagnosis ; Diagnostic Tests, Routine ; Enzyme-Linked Immunosorbent Assay ; Humans ; Immunoglobulin G ; Immunoglobulin M

We developed a Rapid Diagnostic Test (RDT) kit for detecting IgG/IgM antibodies against Zika virus (ZIKV) using monoclonal antibodies to the envelope (E) and non-structural protein 1 (NS1) of ZIKV. These proteins were produced using baculovirus expression vector with Sf9 cells. Monoclonal antibodies J2G7 to NS1 and J5E1 to E protein were selected and conjugated with colloidal gold to produce the Zika IgG/IgM RDT kit (Zika RDT). Comparisons with ELISA, plaque reduction neutralization test (PRNT), and PCR were done to investigate the analytical sensitivity of Zika RDT, which resulted in 100% identical results. Sensitivity and specificity of Zika RDT in a field test was determined using positive and negative samples from Brazil and Korea. The diagnostic accuracy of Zika RDT was fairly high; sensitivity and specificity for IgG was 99.0 and 99.3%, respectively, while for IgM it was 96.7 and 98.7%, respectively. Cross reaction with dengue virus was evaluated using anti-Dengue Mixed Titer Performance Panel (PVD201), in which the Zika RDT showed cross-reactions with DENV in 16.7% and 5.6% in IgG and IgM, respectively. Cross reactions were not observed with West Nile, yellow fever, and hepatitis C virus infected sera. Zika RDT kit is very simple to use, rapid to assay, and very sensitive, and highly specific. Therefore, it would serve as a choice of method for point-of-care diagnosis and large scale surveys of ZIKV infection under clinical or field conditions worldwide in endemic areas.
Antibodies ; Antibodies, Monoclonal ; Baculoviridae ; Brazil ; Cross Reactions ; Dengue Virus ; Diagnosis ; Diagnostic Tests, Routine ; Enzyme-Linked Immunosorbent Assay ; Flavivirus ; Gold Colloid ; Hepacivirus ; Immunoglobulin G ; Immunoglobulin M ; Korea ; Methods ; Neutralization Tests ; Point-of-Care Systems ; Polymerase Chain Reaction ; Reagent Kits, Diagnostic ; Sensitivity and Specificity ; Sf9 Cells ; Yellow Fever ; Zika Virus

OBJECTIVE: To investigate the rapid and accurate method for the detection of dengue virus (DENV) by using nicking enzyme assisted strand-displacement amplification (SDA) combined with gold nanoparticles-based lateral flow strip. METHODS: Total RNA of the virus was extracted by using magnetic beads method and transcribed to cDNA for SDA detection system. Nicking enzyme-assisted method was used for detecting DENV, and agarose gel electrophoresis was used for analyzing the sensitivity of SDA amplification products. A gold nanoparticles-based lateral flow strip was developed based on the principle of nucleic acid base complementary pairing to design the test line and control line. The gold particles were prepared by using sodium citrate reduction method for gold nanoparticles-based lateral flow strip construction. RESULTS: The sensitivity of the SDA method was 10 fmol/L, and the sensitivity of gold nanoparticles-based lateral flow strip based on SDA method was also 10 fmol/L. In a linear range from 10 fmol/L to 10 fmol/L, the corresponding linear correlation coefficient () of DENV was 0.98. The specificity of nanoparticles-based lateral flow strip based on SDA for DENV detection was high, which was no crossing with other control groups. CONCLUSIONS A gold nanoparticles-based lateral flow strip based on SDA method for DENV detection has been established, which is convenient, fast, and the result is visible to naked eyes.
Dengue ; diagnosis ; Dengue Virus ; isolation & purification ; Gold ; Humans ; Metal Nanoparticles ; Reproducibility of Results ; Sensitivity and Specificity

Zika virus (ZIKV) was spread to both eastward and westward from Uganda where the virus was identified approximately in 1947 by a group of arbovirus researchers. In 2015, ZIKV reached Americas with major outbreaks in Brazil. Most countries with mosquito transmitted ZIKV infection are located in tropical and subtropical areas, where ZIKV is endemic with other flaviviruses, including JEV, dengue and yellow fever virus. Approximately 40 countries in Central and South Americas and territories in South Pacific Islands and South East Asia show autochthonous ZIKV endemics. American lineage of ZIKV is known significantly to be mutated in susceptibility to host and in pathogenicity from Asian and Asian lineages approximately since 2014. Early and specific identification of ZIKV infection is very important for the effective management of patients. First of all, optimal collection of specimens for the laboratory diagnosis is required for both nucleic acid testing (NAT) and serological tests. Specimens for NAT tests and serological tests should be determined by the available laboratory resources, work-flow in each laboratory and the geographic areas of specimen collected in addition to days after showing symptoms. Testing strategy for specific differentiation among flaviviruses will vary depending on the prevalence of viruses known to be circulating in the area where the patients were exposed. NAT will be employed for the patients presenting with onset of symptoms less than 7 days. Advanced diagnostic technologies should be continuously developed for the increase of specificity and sensitivity of ZIKV diagnosis.
Americas ; Arboviruses ; Asian Continental Ancestry Group ; Biology* ; Brazil ; Clinical Laboratory Techniques* ; Culicidae ; Dengue ; Diagnosis ; Disease Outbreaks ; Epidemiology* ; Far East ; Flavivirus ; Humans ; Pacific Islands ; Prevalence ; Sensitivity and Specificity ; Serologic Tests ; South America ; Uganda ; Virulence ; Yellow fever virus ; Zika Virus*

From 2013 to 2015, the National Institute of Health, Pakistan, received 1,270 blood samples of suspected dengue cases reported from inpatient and outpatient departments of various hospitals in Khyber Pakhtunkhwa (KPK) province. In this study, we determined the circulating dengue virus (DENV) serotypes using real-time reverse transcriptase (RT)-PCR to understand the serotype-based epidemiology of DENV. All four serotypes (DENV-1 [6%], DENV-2 [33%], DENV-3 [47%], and DENV-4 [0.1%]) were found circulating during the study period. Our findings suggest the need for an active surveillance system coupled with the laboratory diagnosis, especially in the chronic endemic areas of the country. Public awareness programs are needed for effective control and prevention of outbreaks in the future.
Adolescent ; Adult ; Dengue/diagnosis/*epidemiology/virology ; Dengue Virus/genetics/*isolation & purification ; Disease Outbreaks ; Female ; Humans ; Male ; Middle Aged ; Pakistan/epidemiology ; RNA, Viral/genetics/metabolism ; Real-Time Polymerase Chain Reaction ; Serogroup ; Young Adult

OBJECTIVEA Dengue outbreak was reported in Dongfen town Jianou county, Fujian province on September 19, 2014. The goal of this project was to explore the role of syndromic surveillance program in the practice of early detection on disease outbreak through the case mentioned above.

METHODSThe authors retrospectively collected data related to Outpatient log and Pharmacy drug use in Dongfen township hospital through the electronic information system of the hospital from August to November, 2014. All the abnormal events were recorded, according to related data on fever and drug use. Description of fever, syndromic characteristics, correlation and Linear regression analyses were conducted, using the surveillance data on fever syndrome and drug use from the pharmacy.

RESULTSA total of 1 102 cases with fever and 2 437 fever-related clinic visits were reported which showing an increased number of 19.6, 10.2 times respectively, when compared to the same period of the previous year in which men accounted for 45.3% (499/1 102) and female accounted for 54.7% (603/1 102). Age groups presented an atypical type " M" type. 5 and 10 year olds groups formed the largest proportion, accounted for 11.5% (127/1 102) of the total number os the patients. The correlation coefficient ranged from 0.85 to 0.97 (P<0.05). Data from the syndromic surveillance program showed an " outbreak" was occured in August 23, 2014.

CONCLUSIONSCompared to routine surveillance program, the syndromic surveillance program could detect the appearence of an outbreak, a month or even more earlier. The role of syndromic surveillance program needs to be further explored.

Data Collection ; Dengue ; diagnosis ; epidemiology ; prevention & control ; Disease Outbreaks ; prevention & control ; Drug Prescriptions ; statistics & numerical data ; Drug Utilization ; statistics & numerical data ; trends ; Early Diagnosis ; Female ; Fever ; etiology ; Health Information Systems ; Humans ; Male ; Pharmacy Service, Hospital ; Population Surveillance ; methods ; Retrospective Studies

Travel-related health problems such as febrile illness have been reported in many travelers going to developing countries. With the emergence of new infectious diseases occurring in many parts of the world and their spread worldwide, early diagnosis of emerging infectious diseases or tropical diseases has become a very important part of controlling these diseases. In doing so, the itinerary of the ill returning traveler is crucial to formulating a differential diagnosis because exposure to pathogens differs depending on the area of travel. With up-to-date information on infectious diseases occurring worldwide, a differential diagnosis can be made by adding information on duration of travel, incubation period, underlying medical illness, history of prophylactic vaccines received, and knowledge of the patient's exposures during travel including insect bites, contaminated food or water, or freshwater swimming. Some travelers may have specific symptoms and signs such as fever, rash, or hemorrhagic manifestations. For example, eosinophilia suggests a possible helminth infection. In this article, the general approach to returnning travelers with suspected tropical disease will be described.
Communicable Diseases ; Communicable Diseases, Emerging ; Dengue ; Developing Countries ; Diagnosis, Differential* ; Early Diagnosis ; Eosinophilia ; Exanthema ; Fever ; Fresh Water ; Helminths ; Insect Bites and Stings ; Malaria ; Swimming ; Travel Medicine ; Vaccines ; Water

Despite recent advances in the development of diagnostics, therapeutics, and vaccines, the ease of international travel and increasing global interdependence have brought about particular challenges for the control of infectious diseases, highlighting concerns for the worldwide spread of emerging and reemerging infectious diseases. Korea is also facing public health challenges for controlling imported cases of infectious diseases; dengue virus, which is the most commonly reported case of imported infectious diseases; the largest outbreak of Middle East respiratory syndrome coronavirus infections outside the Arabian Peninsula in 2015; and the Zika virus infection, which was declared by the WHO as a "Public Health Emergency of International Concern." Although national and global partnerships are critical to controlling imported infectious disease threats, the role of local hospitals, public health sectors, and laboratory capacity remains the cornerstone for initial disease recognition and response. The current status of laboratory diagnosis for imported infectious diseases is reviewed.
Clinical Laboratory Techniques ; Communicable Diseases ; Communicable Diseases, Emerging* ; Coronavirus Infections ; Dengue ; Dengue Virus ; Diagnosis* ; Emergencies ; Hospitals, Public ; Korea* ; Middle East ; Public Health ; Vaccines