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Background: Vocal cord dysfunction/inducible laryngeal obstruction (VCD/ILO) is characterized by breathlessness and often mimics or accompanies severe asthma. The disorder occurs intermittently, and the diagnosis is established by using laryngoscopy. Dynamic computed tomography (CT) imaging of the larynx at low-radiation doses has the potential to provide an alternative method to make the diagnosis of VCD/ILO. Methods: We report two case series: in series A, laryngoscopy (diagnostic standard) and CT imaging of the larynx were each performed within 1 hour of each other (n=31), and in series B, the procedures were performed on separate days 4 to 6 weeks apart (n=72). Diagnosis of VCD/ILO by laryngoscopy used conventional criteria, and diagnosis by CT imaging was based on vocal cord narrowing in excess of a validated normal threshold. In each series, we evaluated the accuracy of CT imaging of the larynx to establish a diagnosis of VCD/ILO compared with laryngoscopy. Results: In series A, the sensitivity of CT imaging of the larynx was 53.8%, and specificity was 88.9%; in series B, the sensitivity of CT imaging of the larynx was 76.2%, and specificity was 93.3%. At a disease prevalence of 30% (which was known to be the case in our clinic), the positive predictive value was 67.5% in series A and 83% in series B. Negative predictive values were 81.8% and 90.1% in series A and B, respectively, and false-positive rates were 11.1% and 6.7%. Conclusions: When the population prevalence was assumed to be 30%, low-dose CT imaging of the larynx detected VCD/ILO with negative predictive values greater than 80% in both series settings and agreed with each other within 9 percentage points. Positive predictive values for laryngeal CT imaging varied substantially between the settings of the two case series. (Supported by Monash Lung and Sleep Institute and Grant APP ID 1198362 and others.) [ABSTRACT FROM AUTHOR]

Rodent population control is an important measure in reducing the risk of rodent-borne disease transmission. In this study, we examined rodent activity in the sanitary waste network around the household waste-collection bin chamber of an urban residential apartment block. We utilised infra-red camera traps to determine the pattern of rodent activity in a rodent-infested bin chamber and its associated sanitary waste network. Multivariable logistic regression was performed to assess the risk factors that were independently associated with rodent activity in the bin chambers. The camera trap surveillance showed that the rodents were active in the bin chamber and sanitary network both in the day and at night. In the cross-sectional study, rodent activity in the bin chambers was independently associated with broken floor traps [Adjusted odds ratio (AOR): 36.7, CI: 21.3–66.3], calendar month [Log-likelihood ratio test (LRT) p = 0.002] and Town Council [LRT p = 0.004] variables. In restricted analysis, rodent activity in bin chambers was independently associated with defects in the wastewater pipe under the chamber [AOR: 12.3, CI: 4.3–51.7]. Our study suggests that urban municipal management councils should prioritize rodent control resources in areas according to the factors that increase the risk of rodent infestation. [ABSTRACT FROM AUTHOR]

BACKGROUND: With its tropical rainforest climate, rapid urbanization, and changing demography and ecology, Singapore experiences endemic dengue; the last large outbreak in 2013 culminated in 22,170 cases. In the absence of a vaccine on the market, vector control is the key approach for prevention. OBJECTIVES: We sought to forecast the evolution of dengue epidemics in Singapore to provide early warning of outbreaks and to facilitate the public health response to moderate an impending outbreak. METHODS: We developed a set of statistical models using least absolute shrinkage and selection operator (LASSO) methods to forecast the weekly incidence of dengue notifications over a 3-month time horizon. This forecasting tool used a variety of data streams and was updated weekly, including recent case data, meteorological data, vector surveillance data, and population-based national statistics. The forecasting methodology was compared with alternative approaches that have been proposed to model dengue case data (seasonal autoregressive integrated moving average and step-down linear regression) by fielding them on the 2013 dengue epidemic, the largest on record in Singapore. RESULTS: Operationally useful forecasts were obtained at a 3-month lag using the LASSO-derived models. Based on the mean average percentage error, the LASSO approach provided more accurate forecasts than the other methods we assessed. We demonstrate its utility in Singapore's dengue control program by providing a forecast of the 2013 outbreak for advance preparation of outbreak response. CONCLUSIONS: Statistical models built using machine learning methods such as LASSO have the potential to markedly improve forecasting techniques for recurrent infectious disease outbreaks such as dengue. http://dx.doi.org/10.1289/ehp.1509981., Introduction Dengue is an acute infectious disease common to tropical and subtropical regions. Dengue viruses are transmitted by Aedes mosquitoes, mainly Aedes aegypti and Aedes albopictus (Rosen et al. 1983). [...]

Abstract 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.

Japanese encephalitis virus (JEV) and West Nile virus (WNV) are arboviruses primarily transmitted by Culex spp. mosquitoes. Birds are the primary hosts for JEV and WNV. Recent WNV outbreaks in Europe and United States and their association with migratory birds highlight the importance of understanding the feeding host preference of potential vectors for outbreak preparedness, especially in nonendemic settings. Singapore is nonendemic to JEV and WNV, but is a stopover site for migratory birds of the East Asian–Australasian Flyway. Therefore, we elucidated the feeding host range of Culex spp. mosquitoes captured in four natural (bird) habitats in Singapore from January 2011 to December 2012. We characterized feeding host DNA in field-caught mosquitoes using a PCR sequencing-based assay targeting the mitochondrial gene regions. Of 22,648 mosquitoes captured, 21,287 belonged to the Culex vishnui subgroup. The host DNA analysis showed that mosquitoes from the Cx. vishnui subgroup are opportunistic biters, feeding on a range of birds and mammals. Cx. vishnui subgroup, Culex sitiens and Culex bitaeniorhynchus, was primarily ornithophagic, although they fed opportunistically on mammals, including humans. Culex gelidus and Culex quinquefasciatus, in contrast, fed mainly on mammals. The presence of ornitho- and anthropophilic mosquito vectors and susceptible avian and mammalian hosts poses a risk spill-over transmission of JEV and WNV among humans, should these viruses be introduced through migratory birds and establish persistent transmission in resident birds and animal hosts in Singapore. [ABSTRACT FROM AUTHOR]

House Index, Container Index, and Breteau Index are the most commonly used indices for dengue vector surveillance. However, these larval indices are a poor proxy for measuring the adult population—which is responsible for disease transmission. Information on the adult distribution and density are important for assessing transmission risk as well as for developing effective control strategies. This study introduces a new entomological index, Gravitrap aegypti index (GAI), which estimates the adult female Aedes aegypti population in the community and presents its association with dengue cases. Gravitraps were deployed across 34 treatment sites in Singapore from September 2013 to September 2016. The GAI, derived from the Gravitrap surveillance data, was analysed to investigate the spatio-temporal patterns of the Ae. aegypti population in Singapore. The index was further categorised into low, moderate, and high-risk groups and its association with dengue cases were examined. A Before-After Control Impact analysis was performed to evaluate the epidemiology impact of Gravitrap system on dengue transmission. The Ae. aegypti population exhibits a seasonal pattern, and spatial heterogeneity in Ae. aegypti abundance was observed among treatment sites. The Ae. aegypti population was also found to be unevenly distributed among floors of an apartment block, with low floors (floors 1–4) having a higher abundance of mosquitoes trapped than mid (floors 5–8) and high (floors ≥9) floors. Areas with high GAI were shown to have higher dengue case count. Gravitrap has also demonstrated to be a good dengue control tool. The contribution of cases by treatment sites to the national numbers was lower after Gravitraps deployment. The GAI, which is of better relevance to dengue transmission risk, could be recommended as an indicator for decision making in vector control efforts, and to monitor the spatio-temporal variability of the adult Aedes population in the country. In addition, findings from this study indicate that Gravitraps can be used as a dengue control tool to reduce dengue transmission. Author summary: In the absence of an effective vaccine: vector surveillance and control remain the key strategy for dengue prevention and control. The collection of the adult female Aedes mosquito is thus important to understand disease transmission dynamics. Information on its distribution and density are also essential for assessing transmission risk as well as for devising an effective control strategy. Here, we described a new approach to dengue vector surveillance based on adult female Aedes trapping using Gravitraps. Using the Gravitrap surveillance data, we derived a new entomological index, Gravitrap aegypti index (GAI), which estimates the adult female Ae. aegypti population in the community and presents its association with dengue cases. When analysed, the index can provide useful information on the spatio-temporal distribution of the Ae. aegypti population in the country and hence, assist planning of vector control. The GAI, which is of better relevance to dengue transmission risk, could be used as an indicator for decision making in vector control efforts, and to monitor the spatio-temporal variability of the adult Aedes population. In addition, findings from this study indicate that Gravitraps can be used as a dengue control tool to reduce dengue transmission. [ABSTRACT FROM AUTHOR]

Background: Singapore experiences endemic dengue, with 2013 being the largest outbreak year known to date, culminating in 22,170 cases. Given the limited resources available, and that vector control is the key approach for prevention in Singapore, it is important that public health professionals know where resources should be invested in. This study aims to stratify the spatial risk of dengue transmission in Singapore for effective deployment of resources. Methodology/principal findings: Random Forest was used to predict the risk rank of dengue transmission in 1km2 grids, with dengue, population, entomological and environmental data. The predicted risk ranks are categorized and mapped to four color-coded risk groups for easy operation application. The risk maps were evaluated with dengue case and cluster data. Risk maps produced by Random Forest have high accuracy. More than 80% of the observed risk ranks fell within the 80% prediction interval. The observed and predicted risk ranks were highly correlated (≥0.86, P <0.01). Furthermore, the predicted risk levels were in excellent agreement with case density, a weighted Kappa coefficient of more than 0.80 (P <0.01). Close to 90% of the dengue clusters occur in high risk areas, and the odds of cluster forming in high risk areas were higher than in low risk areas. Conclusions: This study demonstrates the potential of Random Forest and its strong predictive capability in stratifying the spatial risk of dengue transmission in Singapore. Dengue risk map produced using Random Forest has high accuracy, and is a good surveillance tool to guide vector control operations. [ABSTRACT FROM AUTHOR]

We report a case of a Singaporean who acquired Zika virus (ZIKV) during a visit to Cuba. The infection was confirmed using molecular and serological methods. This report highlights potential drawbacks of using IgG serology for diagnosis of flavivirus infections in endemic regions. The low viremia detected during the early phase of this case resulted in low mosquito infectivity rates, suggesting the possibility of ZIKV transmission prior to clinical onset. The report also emphasizes the challenges of public health interventions for Zika fever and the importance of sustaining a low vector population to reduce the risk of arbovirus transmission in vulnerable regions. [ABSTRACT FROM AUTHOR]

High frequency financial data modelling has become one of the important research areas in the field of financial econometrics. However, the possible structural break in volatile financial time series often trigger inconsistency issue in volatility estimation. In this study, we propose a structural break heavy-tailed heterogeneous autoregressive (HAR) volatility econometric model with the enhancement of jump-robust estimators. The breakpoints in the volatility are captured by dummy variables after the detection by Bai-Perron sequential multi breakpoints procedure. In order to further deal with possible abrupt jump in the volatility, the jump-robust volatility estimators are composed by using the nearest neighbor truncation approach, namely the minimum and median realized volatility. Under the structural break improvements in both the models and volatility estimators, the empirical findings show that the modified HAR model provides the best performing in-sample and out-of-sample forecast evaluations as compared with the standard HAR models. Accurate volatility forecasts have direct influential to the application of risk management and investment portfolio analysis. [ABSTRACT FROM AUTHOR]

BACKGROUND: With its tropical rainforest climate, rapid urbanization, and changing demography and ecology, Singapore experiences endemic dengue; the last large outbreak in 2013 culminated in 22,170 cases. In the absence of a vaccine on the market, vector control is the key approach for prevention. OBJECTIVES: We sought to forecast the evolution of dengue epidemics in Singapore to provide early warning of outbreaks and to facilitate the public health response to moderate an impending outbreak. METHODS: We developed a set of statistical models using least absolute shrinkage and selection operator (LASSO) methods to forecast the weekly incidence of dengue notifications over a 3-month time horizon. This forecasting tool used a variety of data streams and was updated weekly, including recent case data, meteorological data, vector surveillance data, and population-based national statistics. The forecasting methodology was compared with alternative approaches that have been proposed to model dengue case data (seasonal autoregressive integrated moving average and step-down linear regression) by fielding them on the 2013 dengue epidemic, the largest on record in Singapore. RESULTS: Operationally useful forecasts were obtained at a 3-month lag using the LASSO-derived models. Based on the mean average percentage error, the LASSO approach provided more accurate forecasts than the other methods we assessed. We demonstrate its utility in Singapore's dengue control program by providing a forecast of the 2013 outbreak for advance preparation of outbreak response. CONCLUSIONS: Statistical models built using machine learning methods such as LASSO have the potential to markedly improve forecasting techniques for recurrent infectious disease outbreaks such as dengue. [ABSTRACT FROM AUTHOR]

Background: Dengue resurged in Singapore during 2013-14, causing an outbreak with unprecedented number of cases in the country. In the present study, we summarise the epidemiological, virological and entomological findings gathered through the dengue surveillance programme and highlight the drivers of the epidemic. We also describe how the surveillance system facilitated the preparedness to moderate epidemic transmission of dengue in the country.Methods: The case surveillance was based on a mandatory notification system that requires all medical practitioners to report clinically-suspected and laboratory-confirmed cases within 24 hours. The circulating Dengue virus (DENV) populations were monitored through an island wide virus surveillance programme aimed at determining the serotypes and genotypes of circulating virus strains. Entomological surveillance included adult Aedes surveillance as well as premise checks for larval breeding.Results: A switch in the dominant serotype from DENV-2 to DENV-1 in March 2013 signalled a potential spike in cases, and the alert was corroborated by an increase in average Aedes house index. The alert triggered preparedness and early response to moderate the impending outbreak. The two-year outbreak led to 22,170 cases in 2013 and 18,338 in 2014, corresponding to an incidence rate of 410.6 and 335.0 per 100,000 population, respectively. DENV-1 was the dominant serotype in 2013 (61.7 %, n = 5,071) and 2014 (79.2 %, n = 5,226), contributed largely by a newly-introduced DENV-1 genotype III strain. The percentage of houses with Ae. aegypti breeding increased significantly (p < 0.001) from 2012 (annual average of 0.07 %) to 2013 (annual average of 0.14 %), followed by a drop in 2014 (annual average of 0.10 %). Aedes breeding data further showed a wide spread distribution of Ae. aegypti in the country that corresponded with the dengue case distribution pattern in 2013 and 2014. The adult Aedes data from 34 gravitrap sentinel sites revealed that approximately 1/3 of the monitored sites remained at high risk of DENV transmission in 2013.Conclusions: The culmination of the latest epidemic is likely to be due to a number of demographic, social, virological, entomological, immunological, climatic and ecological factors that contribute to DENV transmission. A multi-pronged approach backed by the epidemiological, virological and entomological understanding paved way to moderate the case burden through an integrated vector management approach. [ABSTRACT FROM AUTHOR]

Commensal plants and animals have long been used to track human migrations, with Rattus exulans (the Pacific rat) a common organism for reconstructing Polynesian dispersal in the Pacific. However, with no knowledge of the homeland of R. exulans, the place of origin of this human-commensal relationship is unknown. We conducted a mitochondrial DNA phylogeographic survey of R. exulans diversity across the potential natural range in mainland and Island Southeast Asia in order to establish the origin of this human-commensal dyad. We also conducted allozyme electrophoresis on samples from ISEA to obtain a perspective on patterns of genetic diversity in this critical region. Finally, we compared molecular genetic evidence with knowledge of prehistoric rodent faunas in mainland and ISEA. We find that ISEA populations of R. exulans contain the highest mtDNA lineage diversity including significant haplotype diversity not represented elsewhere in the species range. Within ISEA, the island of Flores in the Lesser Sunda group contains the highest diversity in ISEA (across all loci) and also has a deep fossil record of small mammals that appears to include R. exulans. Therefore, in addition to Flores harboring unusual diversity in the form of Homo floresiensis, dwarfed stegodons and giant rats, this island appears to be the homeland of R. exulans. [ABSTRACT FROM AUTHOR]

Introduction: We investigated the epidemiological features of the 2007 dengue outbreak to determine the factors that could have triggered it two years after the previous large outbreak in 2005. Methods: All laboratory-confirmed cases of dengue reported during the year, as well as entomological and virological data, were analysed. Results: A total of 8826 cases including 24 deaths were reported in 2007, giving an incidence of 192.3 cases per 100 000 residents and a case-fatality rate of 0.27%. The median age of the cases was 37 years (interquartile range 25 to 50), with an age range from two days to 101 years, which was higher than the median age of 31 years (interquartile range 20 to 42), with a range from four days to 98 years, in 2005. The overall Aedes premises index in 2007 was 0.68%, lower than the 1.15% observed in 2005. The predominant dengue serotype in 2007 was dengue virus DENV-2 which re-emerged with a clade replacement in early 2007, and overtook the predominant serotype (DENV-1) of 2005. Seroprevalence studies conducted in the three largest outbreak clusters revealed that 73.2% of residents with recent infection were asymptomatic. Discussion: With the exception of an increase in the median age of the cases, and a change in the predominant dengue serotype, the epidemiological features of the 2007 epidemic were largely similar to those of 2005. Singapore remains vulnerable to major outbreaks of dengue, despite sustained vector control measures to maintain a consistently low Aedes premises index. [ABSTRACT FROM AUTHOR]

Background: The necessity of a venous blood collection in all dengue diagnostic assays and the high cost of tests that are available for testing during the viraemic period hinder early detection of dengue cases and thus could delay cluster management. This study reports the utility of saliva in an assay that detects dengue virus (DENV)–specific immunoglobulin A (Ig A) early in the phase of a dengue infection. Methods and Findings: Using an antigen capture anti-DENV IgA (ACA) ELISA technique, we tested saliva samples collected from dengue-confirmed patients. The sensitivity within 3 days from fever onset was over 36% in primary dengue infections. The performance is markedly better in secondary infections, with 100% sensitivity reported in saliva samples from day 1 after fever onset. Serum and salivary IgA levels showed good correlation (Pearson's r = 0.69, p<0.001). Specificity was found to be 97%. Conclusion: Our findings suggest that this technique would be very useful in dengue endemic regions, where the majority of dengue cases are secondary. The ACA-ELISA is easy to perform, cost effective, and especially useful in laboratories without sophisticated equipment. Our findings established the usefulness and reliability of saliva for early dengue diagnosis. Author Summary: The importance of laboratory diagnosis of dengue cannot be undermined. In recent years, many dengue diagnostic tools have become available for various stages of the disease, but the one limitation is that they require blood as a specimen for testing. In many incidences, phlebotomy in needle-phobic febrile individuals, especially children, can be challenging, and the tendency to forgo a dengue blood test is high. To circumvent this, we decided to work toward a saliva-based assay (antigen-capture anti-DENV IgA ELISA, ACA-ELISA) that has the necessary sensitivity and specificity to detect dengue early. Overall sensitivity of the ACA-ELISA, when tested on saliva collected from dengue-confirmed patients (EDEN study) at three time points, was 70% in the first 3 days after fever onset and 93% between 4 to 8 days after fever onset. In patients with secondary dengue infections, salivary IgA was detected on the first day of fever onset in all the dengue confirmed patients. This demonstrates the utility of saliva in the ACA-ELISA for early dengue diagnostics. This technique is easy to perform, cost effective, and is especially useful in dengue endemic countries. [ABSTRACT FROM AUTHOR]

Background: The sensitivity and specificity of two in-house MAC-ELISA assays were tested and compared with the performance of commercially-available CTK lateral flow rapid test and EUROIMMUN IFA assays for the detection of anti-Chikungunya virus (CHIKV) IgM. Each MAC-ELISA assay used a whole virus-based antigen derived from genetically distinct CHIKV strains involved in two chikungunya disease outbreaks in Singapore (2008); a January outbreak strain with alanine at amino acid residue 226 of the E1 glycoprotein (CHIKV-A226) and a May-to-September outbreak strain that possessed valine at the same residue (CHIKV-226V). We report differences in IgM detection efficacy of different assays between the two outbreaks. The sensitivities of two PCR protocols were also tested. Methods and Findings: For sera from January outbreak, the average detection threshold of CTK lateral flow test, MAC-ELISAs and EUROIMMUN IFA assays was 3.75, 4.38 and 4.88 days post fever onset respectively. In contrast, IgM detection using CTK lateral flow test was delayed to more than 7 days after fever onset in the second outbreak sera. However, MAC-ELISA using CHIKV-226V detected IgM in the second outbreak sera 3.96 days after fever onset, which was approximately one day earlier compared to the same assay using CHIKV-A226 (4.86 days). Specificity was 100% for both commercial assays, and 95.6% for the in-house MAC-ELISAs. For sensitivity determination of the PCR protocols, the probe-based real time RT-PCR method was found to be 10 times more sensitive than one based on SYBR Green. Conclusion: Our findings suggested that the two strains of CHIKV using variants A226 and 226V resulted in variation in sensitivities of the assays evaluated. We postulated that the observed difference in antigen efficacy could be due to the amino acid substitution differences in viral E1 and E2 envelope proteins, especially the E1-A226V substitution. This evaluation demonstrates the importance of appraisal of different diagnostic assays before their application in clinical and operational settings. Author Summary: Chikungunya is a mounting public health concern in many parts of the world. Definitive diagnosis is critical in differentiating the diseases, especially in dengue endemic areas. There are some commercial chikungunya kits and published molecular protocols available, but no comprehensive comparative evaluation of them was performed. Using sera collected in outbreaks caused by two variants of Chikungunya virus (A226 and 226V), we tested 2 commercial IgM tests (CTK lateral flow rapid test and EUROIMMUN IFA) alongside our in-house IgM assays (using both variants of the virus). Sensitivities of 2 published PCR protocols were also evaluated based on RNA standards derived from cell-cultured viruses. The commercial assays had different performances in each outbreak, with CTK's lateral flow test having the best performance in the first outbreak and EUROIMMUN IFA being more sensitive in the second outbreak. Use of the current circulating virus in a test assay improves sensitivity of the MAC-ELISAs. For PCR, a probe-based real time RT-PCR method was found to be 10 times more sensitive than the SYBR Green method. Despite this, the latter protocol is found to be more suitable and cost-effective for our diagnostic laboratory. This evaluation demonstrates the importance of appraisal of commercial kits and published protocols before application of a diagnostic tool in the clinical and operational setting. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Dengue diagnosis is largely dependent on clinical symptoms and routinely confirmed with laboratory detection of dengue virus in patient serum samples collected via phlebotomy. This presents a challenge to patients not amenable to venipuncture. Non-invasive methods of dengue diagnosis have the potential to enhance the current dengue detection algorithm. In this study, samples from dengue infected patients were collected between January 2012 until September 2012 and September 2013 until December 2013 in two different setups. Panel A samples (blood, urine, and saliva) were collected daily when the 39 patients were hospitalised and during their follow-up visits while Panel B samples (saliva) were collected from 23 patients during the acute stage of dengue. Using DENV PCR on Panel A, from day 2 to day 4 post fever onset, serum showed the best overall positivity followed by saliva and urine (100%/82.1%/67.9%). From day 5 until day 10 post fever onset, serum and urine had similar positivity (67.4%/61.2%), followed by saliva (51.3%). Beyond day 10 post fever onset, DENV was undetectable in sera, but urine and saliva showed 56.8% and 28.6% positivity, respectively. DENV in urine was detectable up until 32 days post fever. Panel B results showed overall sensitivity of 32.4%/36% (RNA/NS1) for DENV detection in saliva. Our results suggest that the urine-based detection method is useful especially for late dengue detection, where DENV is undetected in sera but still detectable in urine. This provides a potential tool for the physician to pick up new cases in an area where there is ongoing dengue transmission and subsequently prompt for intensified vector control activities. [ABSTRACT FROM AUTHOR]

Commensal plants and animals have long been used to track human migrations, with Rattus exulans (the Pacific rat) a common organism for reconstructing Polynesian dispersal in the Pacific. However, with no knowledge of the homeland of R. exulans, the place of origin of this human-commensal relationship is unknown. We conducted a mitochondrial DNA phylogeographic survey of R. exulans diversity across the potential natural range in mainland and Island Southeast Asia in order to establish the origin of this human-commensal dyad. We also conducted allozyme electrophoresis on samples from ISEA to obtain a perspective on patterns of genetic diversity in this critical region. Finally, we compared molecular genetic evidence with knowledge of prehistoric rodent faunas in mainland and ISEA. We find that ISEA populations of R. exulans contain the highest mtDNA lineage diversity including significant haplotype diversity not represented elsewhere in the species range. Within ISEA, the island of Flores in the Lesser Sunda group contains the highest diversity in ISEA (across all loci) and also has a deep fossil record of small mammals that appears to include R. exulans. Therefore, in addition to Flores harboring unusual diversity in the form of Homo floresiensis, dwarfed stegodons and giant rats, this island appears to be the homeland of R. exulans. [ABSTRACT FROM AUTHOR]

Purpose: South Asian (SA) have been observed to have higher cardiovascular mortality rates compared to East Asians (EA) and Caucasians. Pericoronary adipose tissue (PCAT) attenuation around the right coronary artery (RCA) from coronary CT angiography (CTA) has been associated with coronary inflammation and cardiac death. We aimed to investigate i) the relationship between plaque characteristics and PCAT attenuation and ii) to assess gender and ethnic differences in PCAT attenuation using a matched cohort of SA, EA and Caucasians.Method: Three-hundred symptomatic patients who underwent CTA were matched for age, gender, BMI and diabetes (100 in each ethnic group). Semi-automated software was used to quantify the total volumes and burden of non-calcified plaque (NCP), low-density non-calcified plaque (LD-NCP) and calcified plaque (CP) in blinded core-lab analysis. PCAT CT attenuation was measured around the RCA (10-50 mm from RCA ostium), the most standardized model for PCAT analysis.Results: The total volumes and burden of NCP, LD-NCP and CP were comparable in the ethnic groups (each p > 0.05). PCAT attenuation was higher in patients with coronary plaque. PCAT attenuation correlated with the total volumes and burden of NCP, LD-NCP and CP (r>0.17; p < 0.003). Within the RCA this correlation persisted only for NCP features (r>0.39;p < 0.001). Males showed higher PCAT attenuation (p < 0.001). PCAT attenuation was similar between Caucasian, EA and SA (p = 0.32).Conclusions: PCAT CT attenuation correlated most with its surrounded NCP features further highlighting its role as surrogate measure of coronary inflammation. As coronary plaque burden and RCA PCAT attenuation did not differ between ethnic groups, causes of increased cardiac mortality in South Asians needs further investigations. [ABSTRACT FROM AUTHOR]

Mosquito-borne flaviviruses are emerging pathogens of an increasing global public health concern because of their rapid increase in geographical range and the impact of climate change. Japanese encephalitis virus (JEV) and West Nile virus (WNV) are of concern because of the risk of reemergence and introduction by migratory birds. In Singapore, human WNV infection has never been reported and human JEV infection is rare. Four sentinel vector surveillance sites were established in Singapore to understand the potential risk posed by these viruses. Surveillance was carried out from August 2011 to December 2012 at Pulau Ubin, from March 2011 to March 2013 at an Avian Sanctuary (AS), from December 2010 from October 2012 at Murai Farmway, and from December 2010 to December 2013 at a nature reserve. The present study revealed active JEV transmission in Singapore through the detection of JEV genotype II in Culex tritaeniorhynchus collected from an Avian Sanctuary. Culex flavivirus (CxFV), similar to the Quang Binh virus isolated from Cx. tritaeniorhynchus in Vietnam and CxFV-LSFlaviV-A20-09 virus isolated in China, was also detected in Culex spp. ( vishnui subgroup) . No WNV was detected. This study demonstrates the important role that surveillance plays in public health and strongly suggests the circulation of JEV among wildlife in Singapore, despite the absence of reported human cases. A One Health approach involving surveillance, the collaboration between public health and wildlife managers, and control of mosquito populations remains the key measures in risk mitigation of JEV transmission in the enzootic cycle between birds and mosquitoes.

Japanese encephalitis virus (JEV) and West Nile virus (WNV) are arboviruses primarily transmitted by Culex spp. mosquitoes. Birds are the primary hosts for JEV and WNV. Recent WNV outbreaks in Europe and United States and their association with migratory birds highlight the importance of understanding the feeding host preference of potential vectors for outbreak preparedness, especially in nonendemic settings. Singapore is nonendemic to JEV and WNV, but is a stopover site for migratory birds of the East Asian-Australasian Flyway. Therefore, we elucidated the feeding host range of Culex spp. mosquitoes captured in four natural (bird) habitats in Singapore from January 2011 to December 2012. We characterized feeding host DNA in field-caught mosquitoes using a PCR sequencing-based assay targeting the mitochondrial gene regions. Of 22,648 mosquitoes captured, 21,287 belonged to the Culex vishnui subgroup. The host DNA analysis showed that mosquitoes from the Cx. vishnui subgroup are opportunistic biters, feeding on a range of birds and mammals. Cx. vishnui subgroup, Culex sitiens and Culex bitaeniorhynchus , was primarily ornithophagic, although they fed opportunistically on mammals, including humans. Culex gelidus and Culex quinquefasciatus , in contrast, fed mainly on mammals. The presence of ornitho- and anthropophilic mosquito vectors and susceptible avian and mammalian hosts poses a risk spill-over transmission of JEV and WNV among humans, should these viruses be introduced through migratory birds and establish persistent transmission in resident birds and animal hosts in Singapore.

Non-typhoidal salmonellosis is a leading cause of foodborne zoonosis. To better understand the epidemiology of human salmonellosis, this study aimed to determine the prevalence, antimicrobial resistance and sequence types of Salmonella in retail food and wild birds (proximity to humans) in Singapore. We analyzed 21,428 cooked and ready-to-eat food and 1,510 residual faecal samples of wild birds collected during 2010-2015. Thirty-two Salmonella isolates from food and wild birds were subjected to disc diffusion and multi-locus sequence typing (MLST). Salmonella was isolated from 0.08% (17/21,428) of food and 0.99% (15/1510) of wild birds. None of the isolates from wild birds ( n = 15) exhibited phenotypic resistance, while the isolates from food (47.1%, 8/17) showed a high prevalence of phenotypic resistance to, at least, one antimicrobial. These findings suggested that the avian Salmonell a isolates had been subjected to less antimicrobial selection pressure than those from food samples. MLST revealed specific sequence types found in both food and wild birds. The study can guide future studies with whole-genome analysis on a larger number of isolates from various sectors for public health measures., Competing Interests: The authors declare no conflict of interest.

Bats are important reservoirs and vectors in the transmission of emerging infectious diseases. Many highly pathogenic viruses such as SARS-CoV and rabies-related lyssaviruses have crossed species barriers to infect humans and other animals. In this study we monitored the major roost sites of bats in Singapore, and performed surveillance for zoonotic pathogens in these bats. Screening of guano samples collected during the survey uncovered a bat coronavirus ( Betacoronavirus ) in Cynopterus brachyotis , commonly known as the lesser dog-faced fruit bat. Using a capture-enrichment sequencing platform, the full-length genome of the bat CoV was sequenced and found to be closely related to the bat coronavirus HKU9 species found in Leschenault's rousette discovered in the Guangdong and Yunnan provinces.

Avian pox is a highly contagious avian disease, yet relatively little is known about the epidemiology and transmission of Avipoxviruses. Using a molecular approach, we report evidence for a potential link between birds and field-caught mosquitoes in the transmission of Fowlpox virus (FWPV) in Singapore. Comparison of fpv167 (P4b), fpv126 (VLTF-1), fpv175-176 (A11R-A12L) and fpv140 (H3L) gene sequences revealed close relatedness between FWPV strains obtained from cutaneous lesions of a chicken and four pools of Culex pseudovishnui, Culex spp. (vishnui group) and Coquellitidea crassipes caught in the vicinity of the study site. Chicken-derived viruses characterized during two separate infections two years later were also identical to those detected in the first event, suggesting repeated transmission of closely related FWPV strains in the locality. Since the study location is home to resident and migratory birds, we postulated that wild birds could be the source of FWPV and that bird-biting mosquitoes could act as bridging mechanical vectors. Therefore, we determined whether the FWPV-positive mosquito pools (n=4) were positive for avian DNA using a polymerase chain reaction-sequencing assay. Our findings confirmed the presence of avian host DNA in all mosquito pools, suggesting a role for Cx. pseudovishnui, Culex spp. (vishnui group) and Cq. crassipes mosquitoes in FWPV transmission. Our study exemplifies the utilization of molecular tools to understand transmission networks of pathogens affecting avian populations, which has important implications for the design of effective control measures to minimize disease burden and economic loss.

Commensal plants and animals have long been used to track human migrations, with Rattus exulans (the Pacific rat) a common organism for reconstructing Polynesian dispersal in the Pacific. However, with no knowledge of the homeland of R. exulans, the place of origin of this human-commensal relationship is unknown. We conducted a mitochondrial DNA phylogeographic survey of R. exulans diversity across the potential natural range in mainland and Island Southeast Asia in order to establish the origin of this human-commensal dyad. We also conducted allozyme electrophoresis on samples from ISEA to obtain a perspective on patterns of genetic diversity in this critical region. Finally, we compared molecular genetic evidence with knowledge of prehistoric rodent faunas in mainland and ISEA. We find that ISEA populations of R. exulans contain the highest mtDNA lineage diversity including significant haplotype diversity not represented elsewhere in the species range. Within ISEA, the island of Flores in the Lesser Sunda group contains the highest diversity in ISEA (across all loci) and also has a deep fossil record of small mammals that appears to include R. exulans. Therefore, in addition to Flores harboring unusual diversity in the form of Homo floresiensis, dwarfed stegodons and giant rats, this island appears to be the homeland of R. exulans.

Although the dengue iceberg phenomenon is well known, there is a paucity of data on inapparent dengue. Results from a seroepidemiological study conducted during a dengue epidemic in 2007 in Singapore showed a seroprevalence of 65.9% and an inapparent dengue rate of 78%. Older adults (> 45 years old) had significantly higher rates of inapparent dengue infections (P < 0.05).

Introduction: To determine the seroepidemiology of dengue virus infection in a representative sample of the adult resident population aged 18 years old to 74 years old in Singapore and to estimate the proportion of asymptomatic dengue infection during the 2004 epidemic., Materials and Methods: The study was based on 4152 stored blood samples collected between September and December 2004 from participants aged 18 years old to 74 years old during the 2004 National Health Survey. Sera were tested for IgG and IgM antibodies using a commercial test kit (PanBio Capture/Indirect ELISA)., Results: Of the study population, 59.0% and 2.6% tested positive for dengue IgG (past infection) and IgM/high-titre IgG (recent infection), respectively. Only 17.2% of young adults aged 18 years old to 24 years old were dengue IgG positive. Multivariate analyses showed that older age, Indian ethnicity and male gender were significantly associated with past infection, whereas only age was significantly associated with recent dengue infection. Based on the dengue cases notified during the period of survey, it was estimated that for every 23 individuals recently infected with dengue, only 1 was reported to the health authority as a clinical case., Conclusion: The Singapore population is highly susceptible to dengue epidemics despite its aggressive Aedes prevention and control programme. The finding of a high proportion of unreported cases due to asymptomatic and subclinical infection poses a challenge for dengue control.

Virus detection methodology provides detection of dengue virus in the early phase of the disease. PCR, targeting cDNA derived from viral RNA, has been used as a laboratory-based molecular tool for the detection of Dengue virus. We report the development and use of three real-time one-step reverse transcriptase PCR (RT-PCR) assays to detect dengue cases and serotype the virus involved. The first RT-PCR assay uses SYBR green I as the reporting dye for the purpose of cost-effective screening for dengue virus. The detection limit of the SYBR green I assay was 10 PFU/ml (0.01 equivalent PFU per assay) for all four dengue virus serotypes. The second RT-PCR assay is a duplex fluorogenic probe-based real-time RT-PCR for serotyping clinical samples for dengue viruses. The detection threshold of the probe-based RT-PCR format was 0.1 PFU for serotypes Dengue-1 and Dengue-2, 1 PFU for serotype Dengue-3, and 0.01 PFU for serotype Dengue-4. The third is a fourplex assay that detects any of the four serotypes in a single closed tube with comparable sensitivity. Validation of the assays with local clinical samples collected from 2004 to 2006 revealed that there was an 88% positive correlation between virus isolation and RT-PCR with regard to dengue virus detection and a 100% correlation with seroconversion in subsequent samples. The serotyping results derived from duplex and fourplex assays agree fully with each other and with that derived from immunofluorescence assays.

Introduction: Dengue is a major public health problem in Singapore. Age-specific dengue morbidity rates are highest in the young adult population, unlike in many other Southeast Asian countries where dengue is mainly a paediatric disease. Hence, the World Health Organization (WHO) guidelines on dengue diagnosis and management which were developed using the paediatric experiences, may not be suitable for the management of adult dengue infections., Materials and Methods: The Early DENgue (EDEN) infection and outcome study is a collaborative longitudinal study to investigate epidemiological, clinical, viral and host-specific features of early dengue-infected adults, in an effort to identify new early markers for prognostication. Patients presenting with early undifferentiated fever were included in the study. We carried out an interim analysis to look for early indicators of severe disease., Results: During the period of this interim study analysis, 455 febrile patients were recruited. Of these, 133 were confirmed as acute dengue cases based on dengue-specific polymerase chain reaction (PCR) results. There were significant clinical and epidemiological differences between dengue and febrile non-dengue cases. Nine per cent of the dengue cases experienced persistent tiredness, drowsiness and loss of appetite beyond 3 weeks of illness. Quantitation of viral loads using the crossover (Ct) value of real-time RT-PCR correlated with the duration of symptoms. More than half of both primary and secondary dengue cases were hospitalised. There was no dengue-related mortality in this study., Conclusion: The duration of illness and prolonged symptom duration in 9% of the subjects indicate that the burden of dengue illness is substantially different from other non-dengue febrile illness in our study cohort. Our study also highlights the paucity of early prognostic markers for dengue fever in adults.