An Outbreak of Dengue Virus Serotype 2 Cosmopolitan Genotype in Nepal, 2017

Dengue virus (DENV) is one of the most prevalent neglected tropical diseases, with half of the world’s population at risk of infection. In Nepal, DENV was first reported in 2004, and its prevalence is increasing every year. The present study aimed to obtain and characterize the full-length genome sequence of DENV from the 2017 outbreak. Hospital-based surveillance was conducted in two provinces of Nepal during the outbreak. Acute-phase serum samples were collected from 141 clinically suspected dengue patients after the rainy season. By serological and molecular techniques, 37 (26.9%) and 49 (34.8%), respectively, were confirmed as dengue patients. The cosmopolitan genotype of DENV-2 was isolated from 27 laboratory-confirmed dengue patients. Genomic analysis showed many amino acid substitutions distributed mainly among the E, NS3, and NS5 genes. Phylogenetic analyses of the whole genome sequence revealed two clades (Asian and Indian) among DENV-2 isolates from Nepal. The DENV isolates from hilly and Terai areas were similar to Asian and Indian strains, respectively. Further genomic study on different DENV serotypes is warranted to understand DENV epidemics in Nepal, where there are limited scientific resources and infrastructure.     

Dengue virus infection during post-epidemic period in Delhi, India

Dengue fever (DF) and dengue hemorrhagic fever (DHF) are major public health problems in India. During the period following an epidemic, a study was carried out using virological and serological tests for confirmation of suspected cases of dengue virus infection in fever cases presenting to the All India Institute of Medical Sciences. Serum samples of suspected DF/DHF cases were processed from January to December 1997. In 37 samples from patients with fever of less than 5-day duration, received on ice, virus isolation was attempted in C6/36 clone of Aedes albopictus cell line, followed by indirect fluorescent antibody staining with monoclonal antibodies to dengue viruses 1 to 4. One hundred and forty-three serum samples from patients with more than 5 days fever were tested for dengue specific IgM antibody by either MAC-ELISA or a rapid immunochromatographic assay. Dengue virus type 1 was demonstrated by culture in 8 (21.6%) of 37 serum samples and IgM antibody could be detected in 42 (29.4%) of the 143 serum samples by the serological methods. The peak of dengue virus infection was seen from September to November 1997.     

Investigation of a Chikungunya-like illness in Tirunelveli district, Tamil Nadu, India 2009-2010

Objective To identify the aetiological agent/s of an outbreak of chikungunya‐like illness with high morbidity and several fatalities in Tamil Nadu, India, 2009–2010. Methods Two hundred and seventeen serum samples were collected from the affected areas and screened for chikungunya virus (CHIKV), dengue virus (DENV) and Japanese encephalitis virus (JEV) IgM antibodies using MAC‐ELISA kits. A few selected samples were also tested for Ross River, Sindbis, and Murrey Valley viruses by RT‐PCR and Hantan virus by serology. Twelve acute serum and mosquito samples were processed for virus isolation in C6/36 cells. CHIKV isolate was characterised by RT‐PCR and sequencing. Results Diagnostic levels of IgM antibodies were detected in 107 (49.3%) CHIKV samples and 22 (10.1%) DENV samples. IgM antibodies against JEV were not detected (n = 46). Characterisation of the CHIKV isolate at genetic level demonstrated it as ECSA (E1: 226A). Thirty‐six selected samples were also negative for Ross River, Sindbis, Murrey Valley and Hantan viruses. Conclusion High prevalence of CHIKV IgM antibody positivity, clinical symptoms, virus isolation and the presence of vector mosquitoes clearly suggest CHIKV as the aetiological agent responsible for the outbreak.     

2014年深圳市登革热疫情流行病学特点分析

目的掌握2014年深圳市登革热（denguefever,DF）暴发时期的疫情流行特征,为制定DF防控措施提供科学依据。方法收集深圳市疾病预防控制中心2014年度DF疫情相关数据,结合DF病例个案流行病学调查资料及病例临床信息,用描述性流行病学方法分析DF发病时间、地区、人群分布和血清型特征。结果20t4年深圳市共报告454例DE病例和2起DF暴发疫情,其中346例本地感染病例,108例输入性病例,无2代病例和死亡病例。发病人群以20～50岁青壮年人群为主,占病例总数的76．73％;男性患者所占比例为60．13％,略高于女性。疫情暴发时间主要集中于6～12月,其中9～11月病例报告数占总数的97．14％。全市各区均有疫情发生,以宝安区、福田区和南山区报告病例数最多,占总数的60．00％以上。从100份患者血液标本中分离到65株登革病毒（denguevirus,DENV）,其中登革Ⅰ型59株,登革Ⅱ型4株,登革Ⅲ型2株,登革IV型1株。2014年疫情暴发前、后共采集监测点正常群体血液样本352份,检出DF特异性抗体IgG阳性11份,整体抗体阳性率为3．13％。结论从2014年DF疫情流行特征来看,疫情暴发呈现时间、地域及年龄特征性,且发病人群的流行病学特征与正常人群血清抗体水平均未体现DF具有地方性特征。     

Emergence of dengue virus type-3 in northern India

During the last few decades dengue has reemerged in several parts of Southeast Asia, including India. A major outbreak of dengue infection occurred in northern India during October to December 2003. To determine the etiology, we carried out serological, virological and molecular investigations of this outbreak. A total of 76 dengue suspected patient blood samples were collected from Gwalior, Madhya Pradesh and Delhi, India. Serological investigations carried out using an in-house Dipstick ELISA protocol revealed the presence of anti-dengue antibodies in 53 patients. Twelve of them (22%) had a positive IgM response, indicative of primary infection, and 22 of them (42%) revealed only IgG antibodies, indicative of secondary infection. RT-PCR analysis employing dengue group specific amplimer revealed the presence of dengue specific RNA in four acute phase samples. These four RT-PCR positive samples were further processed for virus isolation in C6/36 cells and suckling mice, yielding four dengue virus isolates. The Nested PCR analysis employing serotype specific amplimer revealed the presence of dengue-3 specific 389 bp amplicon. This study confirmed the reemergence of dengue virus type-3 in a dominant form in India after a gap of nine years. Earlier, dengue virus type-2 was implicated as the etiology of a major dengue epidemic in Delhi in 1996 and Gwalior in 2001. The implication of dengue type-3 as etiology of a DHF epidemic in neighboring Sri Lanka and Bangladesh recently confirms the reemergence of dengue type-3 as the dominant form on the Indian subcontinent.     

Dengue outbreak in Hadramout, Yemen, 2010: an epidemiological perspective

We analyzed surveillance data of a dengue outbreak (2010) reported to the Hadramout Health Office (Yemen) and retrospectively analyzed dengue-related epidemiological and entomological events reported in Hadramout from 2005 to 2009. A total of 630 immunoglobulin M (IgM) -confirmed dengue cases of 982 febrile cases was reported during the period from February to June of 2010; 12 cases died, giving case fatality a rate of 1.9%. Among febrile cases, the highest proportion of dengue cases (37.3%) was reported in the 15- to 24-year-old age group. The overall attack rate was 0.89/1,000. The average number of cases reported by month over the preceding 5-year period compared with the 2010 data is consistent with endemicity of dengue in the region and supports epidemic designation for the dengue activity in 2010. Recognition of endemic dengue transmission and potential for substantial dengue epidemics highlight the need for consistent laboratory-based surveillance that can support prevention and control activities accordingly.     

Molecular characterization and clinical evaluation of dengue outbreak in 2002 in Bangladesh

During the febrile illness epidemic in Bangladesh in 2002, 58 people died out of the 6,132 affected. Two hundred hospitalized patients were analyzed clinically, serologically and virologically to determine the features of this dengue infection. Among the 10- to 70-year-old age group of the 200 clinically suspected dengue patients, 100 (50%) were confirmed as dengue cases by virus isolation and dengue IgM-capture ELISA. Of the 100 dengue-confirmed cases, the mean age was 29.0 (+/-12.4). The possible dengue secondary infection rate determined by Flavivirus IgG-indirect ELISA was 78% in 2002. Eight dengue virus strains were isolated, representing the first dengue virus isolation in the country, and all of the strains were dengue virus type-3 (DEN-3). Sequence data for the envelope gene of the DEN-3 Bangladeshi isolates were used in a phylogenetic comparison with DEN-3 from other countries. A phylogenetic analysis revealed that all 8 strains of DEN-3 were clustered within a well-supported independent sub-cluster of genotype II and were closely related to the Thai isolates from the 1990s. Therefore, it is likely that the currently circulating DEN-3 viruses entered Bangladesh from neighboring countries.     

Identification of an antigenic and genetic variant of dengue-4 virus from the Caribbean

Twenty-one dengue (DEN) viruses isolated from the Caribbean (Dominica and Jamaica) during the 1981-1982 epidemic year were distinct serological and genetic variants of DEN-4 virus. These isolates were clearly identified as DEN-4 viruses using type-specific monoclonal antibodies in indirect immunofluorescence assays. However, they either were not neutralized, or were neutralized poorly using hyperimmune mouse ascitic fluids (HMAF) or rhesus monkey serum directed against the H-241 prototype strain of DEN-4 virus isolated in the Philippines in 1956. HMAF prepared against a representative Caribbean isolate, however, neutralized with similar effectiveness the homologous virus, the H-241 prototype strain, and virus strains isolated from the Pacific and Southeast Asian areas from 1973 to 1984. The Caribbean isolate exhibited no more than 30% and 16% oligonucleotide spot homology with the H-241 and Bangkok viruses, respectively, by RNA fingerprint analysis, while demonstrating 82% and 89% homology with the Gilbert and Niue Island isolates, respectively. The isolation of dengue viruses which are serologically and genetically distinct from the prototype virus emphasizes the need for continued dengue virus surveillance. The recognition of unique dengue isolates should allow the selection of reference strains and vaccine candidate strains which will induce antibodies that are equally effective in neutralizing viruses from all geographic areas.     

Detection of Chikungunya virus in Aedes aegypti during 2011 outbreak in Al Hodayda, Yemen

In October 2010, the Ministry of Public Health and Population reported an outbreak of dengue-like acute febrile illness in Al Hodayda governorate. By January 2011, a total of 1542 cases had been recorded from 19 of the 26 districts in the governorate with 104 purportedly associated deaths. In response this event, in January 2011 entomological investigations aimed at identifying the primary vector and the epidemic associated etiological agent were carried out. Based on the reported cases and the progress of the outbreak in the governorate, mosquito collection was undertaken in two of the most recent outbreak areas; Al Khokha district (130 km south of Al Hodayda) and Al Muneera district (100 km north). Mosquito adults were collected from houses using BG-sentinel? traps, aspiration of resting mosquitoes and knock-down spraying. Indoor and outdoor containers adjacent to the houses were inspected for larvae. Subsequently mosquito pools were analyzed by RT-PCR for detection of the four dengue virus serotypes (DENV-1, DENV-2, DENV-3, DENV-4), and for Chikungunya virus (CHIKV). Aedes aegypti was the dominant mosquito species collected. Four pools represent 40% of the tested pools, all containing adult female Ae. aegypti, were positive for CHIKV. Three CHIKV isolates were obtained from the RNA positive mosquito pools and identified by rRT-PCR. This finding marks the first record of CHIKV isolated from Ae. aegypti in Yemen. The larval container and Breteau indices in the visited localities surveyed were estimated at 53.8 and 100, respectively. The emergence of this unprecedented CHIKV epidemic in Al Hodayda is adding up another arboviral burden to the already existing vector-borne diseases. Considering the governorate as one focal port in the Red Sea region, the spread of the disease to other areas in Yemen and in neighboring countries is anticipated. Public health education and simple measures to detect and prevent mosquito breeding in water storage containers could prevent and reduce the spread of mosquito-borne viruses like CHIKV and DENV in Yemen.     

Epidemic of fever of unknown origin in rural Thailand, caused by influenza A (H1N1) and dengue fever

In the late summer (rainy season) of 1987, a sharp outbreak of fever of unknown origin (FUO) in rural southern Thailand was investigated by a field epidemiology team. In a random survey of households, 40 percent of the children and 20 percent of adults were reported to have had febrile illnesses within the last month. There was at least one death, possibly from Reye's syndrome. Testing 34 pairs of acute and convalescent sera showed significant HI antibody titer rises to influenza A (Taiwan/(H1N1) (9 cases) and dengue virus (12 cases). Testing 79 single sera with the antibody capture ELISA test for dengue, revealed that 23 percent had high titers in the IgM serum fraction suggesting recent infection. There were also six antibody titer rises to coxsackie B viruses, three from well controls. Dengue has previously been observed as a cause of FUO in rural areas in the tropics, but finding a combined epidemic of dengue and influenza was unexpected. With cooperative villagers, adequate personnel and laboratory support, especially the antigen capture ELISA test for dengue infections, it is feasible to successfully investigate disease outbreaks with serologic methods in remote villages.     

Epidemiological and entomological surveillance of the co-circulation of DEN-1, DEN-2 and DEN-4 viruses in French Guiana

We surveyed the disease epidemiology of dengue in French Guiana after the first dengue haemorrhagic fever epidemic from 1991 to 1993 and during an endemic period from 1993 to 1995. DEN-1, DEN-2 and DEN-4 viruses were isolated from patients and DEN-4 was also isolated from Aedes aegypti mosquitoes. Cases of dengue were reported from all over the country, not only from urban areas, but also from rural areas and isolated human settlements, indicating widespread circulation of the viruses. The mosquito vector A. aegypti was found in all inhabited areas of French Guiana and small outdoor containers were the most common breeding grounds. Some ecological features of A. aegypti, such as larvae breeding in Bromeliad plants in the rainforest, a non-exclusive anthropophily and a high vertical transmission rate for dengue viruses, indicate that A. aegypti can behave as a reservoir for dengue viruses in silent areas. Dengue viruses may survive at an endemic level and cause outbreaks when unknown conditions become more favourable. This finding adds to our knowledge of the natural history of dengue viruses in the Americas.     

Imported cases of dengue fever in Russia during 2010-2013

Objective:To confirm dengue infection among Russian tourists returned from Southeast and Mexico in 2010-2013 with clinical signs of infection.Methods:Blood and serum samples from patients were collected.NSI antigen and human IgM/IgG antibodies to dengue virus were identified using commercial tests manufactured by "Standard Diagnostics,INC.",Korea.ELISA test was used for the quantitative analyses of human IgM/IgG antibodies to dengue virus( "Orgenics Ltd.",Israel).Viral RNA was delected using commercial real-lime PCR lesls manufactured by "Genome Diagnostics Pvt.Ltd.",India and "Vector",Russia.Genotypes of revealed dengue viruses were determined employing nucleotide sequencing and phylogenetic analysis of 5'-UTR of the viral genome.Results:A total of 98 collected blood samples were analyzed.Fifty samples were positive for at least one of four markers of dengue infection.IgM to dengue virus were revealed in 38 samples,in 25 samples IgM were combined with IgG.NSI antigen was detected in43 samples.22 serum samples were positive for dengue virus RNA.The majority of samples(12patients) from tourists returned from Thailand were positive for genotype 1 of dengue virus,2nd and 4lh genotype were identified each in 1 patient.Conclusions:Due to laboratory confirmed cases of imported dengue fever in Russia,the differential diagnosis of dengue is strictly recommended for tourists returning from endemic areas.     

Secondary dengue virus type 4 infections in Vietnam

This study was designated to describe clinical and biological features of patients with a suspected diagnosis of dengue fever/dengue hemorrhagic fever during an outbreak in Central Vietnam. One hundred and twenty-five consecutive patients hospitalized at Khanh Hoa and Binh Thuan Provincial hospitals between November 2001 and January 2002 with a diagnosis of suspected dengue infection were included in the present study.Viruses were isolated in C6/36 and VERO E6 cell cultures or detected by RT-PCR. A hemagglutination-inhibition test (HI) was done on each paired sera using dengue antigens type 1-4, Japanese encephalitis (JE) virus antigen, Chickungunya virus antigen and Sindbis virus antigen. Anti-dengue and anti-JE virus IgM were measured by a capture enzyme-linked immunosorbent assay (MAC-ELISA). Anti-dengue and anti-JE virus IgG were measured by an ELISA test. Dengue viruses were isolated in cell culture and/or detected by RT-PCR in 20.8% of blood samples. DEN-4 and DEN-2 serotypes were found in 18.4% and 2.4% of the patients, respectively. A total of 86.4% of individuals had a diagnosis of acute dengue fever by using the HI test and/or dengue virus-specific IgM capture-ELISA and/or virus isolation and/or RT-PCR. The prevalence of primary and secondary acute dengue infection was 4% and 78.4%, respectively. Anti-dengue IgG ELISA test was positive in 88.8% of the patients. In 5 cases (4%), Japanese encephalitis virus infection was positive by serology but the cell culture was negative. No Chickungunya virus or Sindbis virus infection was detected by the HI test. In patients with acute dengue virus infection, the most common presenting symptom was headache, followed by conjunctivitis, petechial rash, muscle and joint pain, nausea and abdominal pain. Four percent of hospitalized patients were classified as dengue hemorrhagic fever. The clinical presentation and blood cell counts were similar between patients hospitalized with acute dengue fever and patients with other febrile illnesses.     

福建省1株登革Ⅱ型病毒的鉴定

目的对2005年福建省分离的1株登革病毒(DV)进行鉴定,并从分子水平追踪其可能的传染源。方法采用酶联免疫吸附试验(ELISA)检测疑似登革热患者血清中DV(IgM、IgG抗体;同时应用C6/36细胞、单克隆抗体间接免疫荧光(McAb-IFA)、逆转录(套式PCR法分别进行病毒的分离和鉴定,并对分离株的部分基因进行核苷酸序列分析。结果患者血清登革病毒特异性IgM抗体阳性、IgG抗体可疑,表明该患者在近期感染过登革病毒。患者血清接种C6/36细胞,观察到登革病毒特有的CPE。受感染的C6/36细胞能与登革病毒Ⅱ型单克隆抗体反应,表明分离的病毒株为登革Ⅱ型病毒。分离株的核酸提取物经RT(PCR扩增,登革病毒通用引物可扩增出511bp的特异性条带,型特异性引物扩增出119bp的特异性条带,进一步证实分离的病毒株为登革Ⅱ型病毒。分离株RT(PCR扩增产物的核苷酸序列与30株不同地域来源的登革Ⅱ型病毒相应序列构建的系统发生树表明,此毒株与东南亚地区的毒株比较接近。此次分离株的序列与1999年登革Ⅱ型病毒福建株的对应序列在亲缘关系上有一定程度距离。结合流行病学调查资料,进一步确定此病例为输入性感染病例。结论福建省首次从输入性登革热患者血清中分离出登革Ⅱ型病毒,该病毒来源于东南亚地区。     

Laboratory-Based Surveillance and Molecular Characterization of Dengue Viruses in Taiwan, 2014

We present the results of a laboratory-based surveillance of dengue in Taiwan in 2014. A total of 240 imported dengue cases were identified. The patients had arrived from 16 countries, and Malaysia, Indonesia, the Philippines, and China were the most frequent importing countries. Phylogenetic analyses showed that genotype I of dengue virus type 1 (DENV-1) and the cosmopolitan genotype of DENV-2 were the predominant DENV strains circulating in southeast Asia. The 2014 dengue epidemic was the largest ever to occur in Taiwan since World War II, and there were 15,492 laboratory-confirmed indigenous dengue cases. Phylogenetic analysis showed that the explosive dengue epidemic in southern Taiwan was caused by a DENV-1 strain of genotype I imported from Indonesia. There were several possible causes of this outbreak, including delayed notification of the outbreak, limited staff and resources for control measures, abnormal weather conditions, and a serious gas pipeline explosion in the dengue hot spot areas in Kaohsiung City. However, the results of this surveillance indicated that both active and passive surveillance systems should be strengthened so appropriate public health measures can be taken promptly to prevent large-scale dengue outbreaks.     

The 2011 outbreak of dengue virus infection in Malwa region of Punjab,India-an evaluation of various diagnostic tests

ObjectiveTo compare the efficacy of rapid immunochromatographic test (ICT) and ELISA for detection of dengue specific parameters (NS1 antigen and IgM antibodies) and the association of these parameters with thrombocytopenia.MethodsA total of 1787 serum samples were obtained from the same number of clinically suspected cases of dengue during an outbreak (2011) of dengue in the Malwa region of Punjab. All the samples were subjected to rapid ICT, dengue early ELISA and dengue IgM-capture ELISA for detection of NS1 antigen and IgM antibodies. Platelet counts of all the cases positive for dengue infection and 150 cases negative for dengue parameters (control group) were recorded.ResutlsRapid ICT for NS1 antigen detection showed same specificity (100%) but lower sensitivity (85.81%) than the dengue early ELISA, while it was 100% and 94.10% respectively for IgM antibody detection. The difference in the platelet counts of cases positive and negative for dengue specific parameters was stastically insignificant (standard error of proportions=0.017, Z value=0.84, P value=0.2).ConclusionsThe confirmation of serological diagnosis of dengue should always be based on ELISA test and ICT and/or thrombocytopenia should not be used as a stand-alone test.     

Hospitalizations for suspected dengue in Puerto Rico, 1991-1995: estimation by capture-recapture methods. The Puerto Rico Association of Epidemiologists.

Capture-recapture estimations compare the results of 2 or more independent surveillance systems for the same event, and by measuring the degree of overlap between them, provide an estimate of the total number of events, and therefore the completeness of ascertainment in each system. The Puerto Rico Department of Health and the Dengue Branch of the Centers for Disease Control and Prevention (CDC) monitor dengue activity in Puerto Rico through 2 distinct surveillance systems: diagnostic specimens from patients with suspected dengue and infection control nurses' reports on patients hospitalized for suspected dengue. The patient listings from these systems were used in a 2-sample, capture-recapture calculation to estimate the total number of persons with suspected dengue hospitalized from 1991 to 1995. The laboratory positivity rate for suspected dengue cases who submitted appropriately timed serum samples in those years ranged from 72.1% to 81.2%. The laboratory-based (diagnostic sample) surveillance system (routinely used to monitor hospitalizations for suspected dengue) detected an average of 1,197 hospitalized cases during non-epidemic years, and 4,329 cases during the epidemic year of 1994. The detection rate of this system averaged 42% of the numbers derived by the capture-recapture method. In non-epidemic years, an estimated average of 2,791 patients (range = 1,553-3,481) was estimated to have been hospitalized with a clinical diagnosis of dengue, compared with 9,479 during 1994. These results demonstrate the under-detection inherent in passive surveillance systems for hospitalized cases of suspected dengue, and illustrate the value of capture-recapture techniques to better estimate the true incidence of hospitalizations for this disease.     

Virologic and serologic surveillance for dengue fever in Jeddah, Saudi Arabia, 1994-1999.

Dengue fever infection was first documented in Jeddah, Saudi Arabia, by virus isolation of dengue type 2 virus in 1994 at the virology laboratory of Dr. Soliman Fakeeh Hospital. Dengue virus surveillance was established after that time. Blood samples were collected from 985 patients (710 male patients and 275 female patients) with suspected cases of dengue from February 1994 to December 1999. Dengue virus isolates were obtained in 207 patients (21%; 162 male patients and 45 female patients). Dengue type 2 was the predominant serotype (138 of 207 isolates, 66.7%), followed by dengue type 1 with (56 of 207 isolates, 27%) and dengue type 3 (13 of 207 isolates, 6.3%). The largest number of isolates (186 of 207 isolates, 90%) was in 1994, a year during which there was a dengue epidemic. In the next 5 years, 1995-1999, only 21 isolates (10%) were isolated. Immunoglobulin M capture enzyme-linked immunosorbent assay was positive in 160 acute samples; 52 of them were from virus culture-positive cases and 108 (11%) from culture-negative cases. The total number of cases diagnosed by both methods was 315 (32%). The prevalence of dengue immunoglobulin G antibodies, as assessed on the basis of immunofluorescent assay, hemagglutination inhibition titers > or = 1/20, or both, in the acute samples was 314 (32%) of 985, indicating past Flavivirus infection. Two patients died, one man with dengue hemorrhagic fever and one woman with dengue shock syndrome. Both fatal dengue cases were due to infection with type 2 virus. All other cases were simple dengue fever. To our knowledge, this is the first report confirming the circulation of 3 dengue serotypes in Jeddah.