The molecular and clinical features of dengue during outbreak in Jambi, Indonesia in 2015.

Dengue is hyperendemic in Indonesia. In 2015, reported cases of dengue fever doubled those of 2014 in the Jambi municipality of Sumatra. We examined viral aetiology and its relationship with disease outcome in Jambi. Dengue-suspected patients' sera were collected and NS1 detection and IgM/IgG serology were performed. Dengue virus (DENV) serotyping was performed using real-time RT-PCR. Envelope genes were sequenced to determine the genotypes of DENV. Clinical, haematologic, and demographic data were recorded. Of 210 dengue-suspected patients, 107 were confirmed. The disease manifested as Dengue Fever (62%), Dengue Haemorrhagic Fever (36%), and Dengue Shock Syndrome (2%). The serotypes of 94 DENV were determined. All DENV serotypes were detected with DENV-1 as the predominant serotype (66%). Genotypically, the DENV-1 viruses belong to Genotype I, DENV-2 was of Cosmopolitan genotype, DENV-3 as Genotype I, and DENV-4 belonged to Genotype II. Comparison with historical data revealed serotype predominance switched from DENV-3 to DENV-1, and the replacement of Genotype IV of DENV-1 with Genotype I. In summary, DENV-1 predominated during the 2015 dengue outbreak in Jambi. The full spectrum of dengue disease occurred and was characterized by a switch in predominant serotypes.

Distribution of Voltage-Gated Sodium Channel (Nav) Alleles among the Aedes aegypti Populations In Central Java Province and Its Association with Resistance to Pyrethroid Insecticides.

The emergence of insecticide resistant Aedes aegypti mosquitoes has hampered dengue control efforts. WHO susceptibility tests, using several pyrethroid compounds, were conducted on Ae. aegypti larvae that were collected and raised to adulthood from Semarang, Surakarta, Kudus and Jepara in Java. The AaNaV gene fragment encompassing kdr polymorphic sites from both susceptible and resistant mosquitoes was amplified, and polymorphisms were associated with the resistant phenotype. The insecticide susceptibility tests demonstrated Ae, aegypti resistance to the pyrethroids, with mortality rates ranging from 1.6%-15.2%. Three non-synonymous polymorphisms (S989P, V1016G and F1534C) and one synonymous polymorphism (codon 982) were detected in the AaNaV gene. Eight AaNaV alleles were observed in specimens from Central Java. Allele 3 (SGF) and allele 7 (PGF) represent the most common alleles found and demonstrated strong associations with resistance to pyrethroids (OR = 2.75, CI: 0.97-7.8 and OR = 7.37, CI: 2.4-22.5, respectively). This is the first report of 8 Ae. aegypti AaNaV alleles, and it indicates the development of resistance in Ae. aegypti in response to pyrethroid insecticide-based selective pressure. These findings strongly suggest the need for an appropriate integrated use of insecticides in the region. The 989P, 1016G and 1534C polymorphisms in the AaNaV gene are potentially valuable molecular markers for pyrethroid insecticide resistance monitoring.

Performance of Simplexa dengue molecular assay compared to conventional and SYBR green RT-PCR for detection of dengue infection in Indonesia.

Diagnostic tests based on detection of dengue virus (DENV) genome are available with varying sensitivities and specificities. The Simplexa Dengue assay (Focus Diagnostics) is a newly developed real-time RT-PCR method designed to detect and serotype DENV simultaneously. To assess the performance of the Simplexa Dengue assay, we performed comparison with conventional RT-PCR and SYBR Green real-time RT-PCR on patients sera isolated from eight cities across Indonesia, a dengue endemic country. A total of 184 sera that were confirmed using NS1 and/or IgM and IgG ELISA were examined. Using conventional and SYBR Green real-time RT-PCR, we detected DENV in 53 (28.8%) and 81 (44.0%) out of 184 sera, respectively. When the Simplexa Dengue assay was employed, the detection rate was increased to 76.6% (141 out of 184 samples). When tested in 40 sera that were confirmed by virus isolation as the gold standard, the conventional RT-PCR yielded 95% sensitivity while the sensitivity of SYBR Green real-time RT-PCR and Simplexa Dengue assay reached 97.5% and 100%, respectively. The specificities of all methods were 100% when tested in 43 non-dengue illness and 20 healthy human samples. Altogether, our data showed the higher detection rate of Simplexa Dengue compared to conventional and SYBR Green real-time RT-PCR in field/surveillance setting. In conclusion, Simplexa Dengue offers rapid and accurate detection and typing of dengue infection and is suitable for both routine diagnostic and surveillance.

Molecular surveillance of dengue in Semarang, Indonesia revealed the circulation of an old genotype of dengue virus serotype-1.

Dengue disease is currently a major health problem in Indonesia and affects all provinces in the country, including Semarang Municipality, Central Java province. While dengue is endemic in this region, only limited data on the disease epidemiology is available. To understand the dynamics of dengue in Semarang, we conducted clinical, virological, and demographical surveillance of dengue in Semarang and its surrounding regions in 2012. Dengue cases were detected in both urban and rural areas located in various geographical features, including the coastal and highland areas. During an eight months' study, a total of 120 febrile patients were recruited, of which 66 were serologically confirmed for dengue infection using IgG/IgM ELISA and/or NS1 tests. The cases occurred both in dry and wet seasons. Majority of patients were under 10 years old. Most patients were diagnosed as dengue hemorrhagic fever, followed by dengue shock syndrome and dengue fever. Serotyping was performed in 31 patients, and we observed the co-circulation of all four dengue virus (DENV) serotypes. When the serotypes were correlated with the severity of the disease, no direct correlation was observed. Phylogenetic analysis of DENV based on Envelope gene sequence revealed the circulation of DENV-2 Cosmopolitan genotype and DENV-3 Genotype I. A striking finding was observed for DENV-1, in which we found the co-circulation of Genotype I with an old Genotype II. The Genotype II was represented by a virus strain that has a very slow mutation rate and is very closely related to the DENV strain from Thailand, isolated in 1964 and never reported in other countries in the last three decades. Moreover, this virus was discovered in a cool highland area with an elevation of 1,001 meters above the sea level. The discovery of this old DENV strain may suggest the silent circulation of old virus strains in Indonesia.

Performance of commercial dengue NS1 ELISA and molecular analysis of NS1 gene of dengue viruses obtained during surveillance in Indonesia.

BACKGROUND: Early diagnosis of dengue infection is crucial for better management of the disease. Diagnostic tests based on the detection of dengue virus (DENV) Non Structural Protein 1 (NS1) antigen are commercially available with different sensitivities and specificities observed in various settings. Dengue is endemic in Indonesia and clinicians are increasingly using the NS1 detection for dengue confirmation. This study described the performance of Panbio Dengue Early NS1 and IgM Capture ELISA assays for dengue detection during our surveillance in eight cities in Indonesia as well as the genetic diversity of DENV NS1 genes and its relationship with the NS1 detection. METHODS: The NS1 and IgM/IgG ELISA assays were used for screening and confirmation of dengue infection during surveillance in 2010-2012. Collected serum samples (n = 440) were subjected to RT-PCR and virus isolation, in which 188 samples were confirmed for dengue infection. The positivity of the ELISA assays were correlated with the RT-PCR results to obtain the sensitivity of the assays. The NS1 genes of 48 Indonesian virus isolates were sequenced and their genetic characteristics were studied. RESULTS: Using molecular data as gold standard, the sensitivity of NS1 ELISA assay for samples from Indonesia was 56.4% while IgM ELISA was 73.7%. When both NS1 and IgM results were combined, the sensitivity increased to 89.4%. The NS1 sensitivity varied when correlated with city/geographical origins and DENV serotype, in which the lowest sensitivity was observed for DENV-4 (19.0%). NS1 sensitivity was higher in primary (67.6%) compared to secondary infection (48.2%). The specificity of NS1 assay for non-dengue samples were 100%. The NS1 gene sequence analysis of 48 isolates revealed the presence of polymorphisms of the NS1 genes which apparently did not influence the NS1 sensitivity. CONCLUSIONS: We observed a relatively low sensitivity of NS1 ELISA for dengue detection on RT-PCR-positive dengue samples. The detection rate increased significantly when NS1 data was combined with IgM. In our study, the low sensitivity of NS1 antigen detection did not relate to NS1 genetic diversity. Rather, the performance of the NS1 antigen test was affected by the infection status of patients and geographical origin of samples.