Multiplex targeted mass spectrometry assay for one-shot flavivirus diagnosis.

Targeted proteomic mass spectrometry is emerging as a salient clinical diagnostic tool to track protein biomarkers. However, its strong analytical properties have not been exploited in the diagnosis and typing of flaviviruses. Here, we report the development of a sensitive and specific single-shot robust assay for flavivirus typing and diagnosis using targeted mass spectrometry technology. Our flavivirus parallel reaction monitoring assay (fvPRM) has the ability to track secreted flaviviral nonstructural protein 1 (NS1) over a broad diagnostic and typing window with high sensitivity, specificity, extendibility, and multiplexing capability. These features, pivotal and pertinent to efficient response toward flavivirus outbreaks, including newly emerging flavivirus strains, circumvent the limitations of current diagnostic assays. fvPRM thus carries high potential in positioning itself as a forerunner in delivering early and accurate diagnosis for disease management.

Intra-epidemic evolutionary dynamics of a Dengue virus type 1 population reveal mutant spectra that correlate with disease transmission.

Dengue virus (DENV) is currently the most prevalent mosquito-borne viral pathogen. DENVs naturally exist as highly heterogeneous populations. Even though the descriptions on DENV diversity are plentiful, only a few studies have narrated the dynamics of intra-epidemic virus diversity at a fine scale. Such accounts are important to decipher the reciprocal relationship between viral evolutionary dynamics and disease transmission that shape dengue epidemiology. In the current study, we present a micro-scale genetic analysis of a monophyletic lineage of DENV-1 genotype III (epidemic lineage) detected from November 2012 to May 2014. The lineage was involved in an unprecedented dengue epidemic in Singapore during 2013-2014. Our findings showed that the epidemic lineage was an ensemble of mutants (variants) originated from an initial mixed viral population. The composition of mutant spectrum was dynamic and positively correlated with case load. The close interaction between viral evolution and transmission intensity indicated that tracking genetic diversity through time is potentially a useful tool to infer DENV transmission dynamics and thereby, to assess the epidemic risk in a disease control perspective. Moreover, such information is salient to understand the viral basis of clinical outcome and immune response variations that is imperative to effective vaccine design.

Feeding host range of Aedes albopictus (Diptera: Culicidae) demonstrates its opportunistic host-seeking behavior in rural Singapore.

Aedes albopictus (Skuse) is a competent vector of arboviruses of public health importance, including dengue virus (DENV) and chikungunya virus viruses. Ae. albopictus is the primary vector of chikungunya virus in Singapore. However, despite being ubiquitous, it plays a secondary role in DENV transmission. The vectorial capacity of Ae. albopictus for DENV in field settings appears to be weak because dengue primarily occurs in Aedes aegypti (L.)-dominated, urban settings of the country. As host-seeking behavior is one of the determinants of vectorial capacity, we screened 6,762 female Ae. albopictus from rural, semiurban, and urban locations in Singapore for avian and nonavian bloodmeals using two polymerase chain reaction-sequencing assays developed in-house. The majority (83.2%, n = 79) of bloodmeals from rural and semiurban areas were from humans. However, Ae. albopictus was also found to feed on shrews, swine, dogs, cats, turtles, and multiple hosts in rural settings. In urban areas, all positive bloodmeals were from humans. There were no avian bloodmeals. Our findings testify that Ae. albopictus is highly anthropophagic even in rural settings, but become opportunistic in extremely low human abundance. This opportunistic feeding behavior warrants further investigations into the vectorial capacity of Ae. albopictus to assess its role in arbovirus transmission in endemic habitats.

Molecular characterization of two hantavirus strains from different rattus species in Singapore.

BACKGROUND: Hantaviruses cause human disease in endemic regions around the world. Outbreaks of hantaviral diseases have been associated with changes in rodent population density and adaptation to human settlements leading to their proliferation in close proximity to human dwellings. In a parallel study initiated to determine the prevalence of pathogens in Singapore's wild rodent population, 1206 rodents were trapped and screened. The findings established a hantavirus seroprevalence of 34%. This paper describes the molecular characterization of hantaviruses from Rattus norvegicus and Rattus tanezumi, the predominant rodents caught in urban Singapore. METHODOLOGY: Pan-hanta RT-PCR performed on samples of Rattus norvegicus and Rattus tanezumi indicated that 27 (2.24%) of the animals were positive. sequence analysis of the S and M segments established that two different hantavirus strains circulate in the rodent population of Singapore. Notably, the hantavirus strains found in Rattus norvegicus clusters with other Asian Seoul virus sequences, while the virus strains found in Rattus tanezumi had the highest sequence similarity to the Serang virus from Rattus tanezumi in Indonesia, followed by Cambodian hantavirus isolates and the Thailand virus isolated from Bandicota indica. CONCLUSIONS: Sequence analysis of the S and M segments of hantavirus strains found in Rattus norvegicus (Seoul virus strain Singapore) and Rattus tanezumi (Serang virus strain Jurong TJK/06) revealed that two genetically different hantavirus strains were found in rodents of Singapore. Evidently, together with Serang, Cambodian and Thailand virus the Jurong virus forms a distinct phylogroup. Interestingly, these highly similar virus strains have been identified in different rodent hosts. Further studies are underway to analyze the public health significance of finding hantavirus strains in Singapore rodents.