To assess the extent of dengue virus exposure in the population.
To assess the extent of dengue virus exposure in the population.
In this seroepidemiological study, 685 blood samples were collected (from April 2007 to July 2009) from two subject groups: (i) 344 samples from anonymous blood donors of the Hong Kong Red Cross and (ii) 341 samples from healthy volunteers recruited from a university, a community centre and a hospital. Demographic information and travel history were collected for the second subject group. All blood samples were subjected to the PanBio Dengue IgG Indirect ELISA.
Anti-dengue virus IgG was detected in 1.6% of the blood samples. Individuals who visited countries in Southeast Asia in the past year were significantly associated with seropositivity (P = 0.03, OR 5.38, CI 1.13–25.54).
The overall dengue seroprevalence was 1.6%, and visit to Southeast Asia was the only independent predictor for seropositivity. Although the current situation is not alarming, frequent travel, presence of mosquito vector and emergence of local cases suggest that the risk of dengue virus infection within the local community cannot be overlooked, and continuous vigilance is warranted.
Evaluer l’étendue de l'exposition au virus de la dengue dans la population.
Dans cette étude séroépidémiologique 685 échantillons de sang ont été prélevés (d'avril 2007 à juillet 2009) chez deux groupes de sujets: (1) 344 échantillons de donneurs de sang anonymes de la Croix-Rouge de Hong Kong et (2) 341 échantillons provenant de volontaires sains recrutés dans une université, un centre communautaire et un hôpital. Les informations démographiques et les historiques de voyage ont été recueillis pour le second groupe de sujets. Tous les échantillons de sang ont été soumis au test indirect ELISA PanBio Dengue IgG.
Des IgG anti-virus de la dengue ont été détectées dans 1,6% des échantillons de sang. La visite de pays de l'Asie du sud-est dans l'année précédente était significativement associée à la séropositivité (P = 0,03; OR: 5,38; IC: 1,13 à 25,54).
La séroprévalence globale de la dengue était de 1,6% et la visite de l'Asie du Sud était le seul facteur prédictif indépendant de séropositivité. Bien que la situation actuelle ne soit pas alarmante, les déplacements fréquents, la présence de moustiques vecteurs et l’émergence de cas locaux suggèrent que le risque d'infection par le virus de la dengue au sein de la communauté locale ne peut être négligé et une vigilance constante est justifiée.
Evaluar el grado de exposición al virus del dengue en la población de Hong Kong.
En este estudio seroepidemiológico se recolectaron 685 muestras de sangre (entre Abril 2007 y Julio 2009) de dos grupos con diferentes tipos de sujeto: (1) 344 muestras de donantes anónimos de sangre de la Cruz Roja de Hong Kong; (2) 341 muestras de voluntarios sanos reclutados de una universidad, un centro comunitario y un hospital. La información demográfica y la historia de viajes se recogió para el segundo grupo de sujetos. A todas las muestras de sangre se les realizó la prueba de dengue mediante la determinación de IgG por ELISA indirecto (de PanBio).
Se detectó IgG contra el virus del dengue en 1.6% de las muestras de sangre. Los individuos que visitaron países del sudeste asiático en el último año estaban significativamente asociados con seropositividad (P = 0.03, OR 5.38, CI 1.13–25.54).
La seroprevalencia general del dengue fue de 1.6% y el haber visitado el sudeste asiático fue el único vaticinador independiente para la seropositividad. Aunque la situación actual no es alarmante, los viajes frecuentes, la presencia del mosquito vector y la aparición de casos locales sugieren que el riesgo de infección por el virus del dengue dentro de la comunidad local no debería obviarse, y que es necesario garantizar una vigilancia continua.
Dengue fever (DF) is a mosquito-borne disease caused by dengue viruses (DV) and epidemics occurred in Asia and Southeast Asia in the 1900s (Kautner et al. 1997; Thomas et al. 2003; Mackenzie et al. 2004). DV are transmitted to humans by infected female Aedes aegypti or Aedes albopictus mosquitoes (Holtzclaw 2002). Without adequate supportive treatment, the disease may progress to severe dengue haemorrhagic fever and lethal dengue shock syndrome (Kautner et al. 1997; Groen et al. 2000). More importantly, specific treatment for dengue infection is still unavailable, dengue vaccines are undergoing various phases of clinical trials and their efficacy remains to be assessed.
Hong Kong is located in Southeast Asia and is geographically close to countries endemic for dengue. Although Hong Kong is not endemic for dengue, a fourfold increase (from 11 to 49 cases) in the incidence rate was recorded from 2000 to 2003 (Centre for Health Protection 2012). More than 30 cases were reported annually during 2004–2012, and 83 cases were documented in 2010 (Centre for Health Protection 2012). The majority of cases were imported from nearby Asian countries. Hong Kong residents travel frequently (World Tourism Organization 2006) to nearby areas for vacation or work, which is believed to be a risk factor associated with DV infection and seroprevalence. The importance of travel-acquired dengue might be underestimated due to the asymptomatic or subclinical presentation of the infection (Wichmann et al. 2003). The major concern is that DV-infected residents or visitors may carry the virus from endemic areas and be responsible for silent spreading of DV in their home countries (Lopez-Velez et al. 1996; Schwartz et al. 1996; Cobelens et al. 2002; Wichmann et al. 2003).
In 2002, a DF outbreak involving 16 cases occurred in Ma Wan, an island in Hong Kong, and was caused by DV serotype 1. Three more sporadic local cases caused by DV serotypes 1 and 2 were reported in other districts in the same year (Tsang 2002). Thereafter, local cases of DV infection were reported in Hong Kong (Centre for Health Protection 2012; Chuang et al. 2008). Emergence of local DF cases in different sites indicated that silent spreading of DV may exist in Hong Kong. Therefore, we conducted a seroepidemiological study to estimate the rate of previous DV exposure in the population of Hong Kong.
Blood samples were collected from two sources during the period from April 2007 to July 2009. One source was the Hong Kong Red Cross Blood Transfusion Service, which provided samples from anonymous healthy and voluntary blood donors. The age of the donors ranged from 16 to 65 years. Only information on the age and gender of each subject was provided.
The other samples were obtained by the researchers from healthy volunteers recruited from a university, a community centre and a hospital. Subjects from the university and the community centre were recruited through poster promotion. Subjects recruited from the hospital were healthy volunteers participating in a clinical research project. All subjects were adults older than 18 years. A questionnaire survey investigated the risk factors associated with seropositivity. Information collected included demographic data and travel history. Past vaccination history for Japanese encephalitis and yellow fever was also recorded. Subjects of this group provided informed written consent for their samples and data to be analysed.
Ethics approval for collection and use of all human blood samples in this study was obtained from the Faculty Research Committee, Faculty of Health & Social Sciences, the Hong Kong Polytechnic University.
For the researcher-collected samples, donors fasted for 2 h before sampling to prevent lipaemic serum. Both serum and plasma samples were collected from each subject. For the Red Cross samples, only plasmas were collected. Each plasma was then converted into serum analogue by defibrination using a method described by Johnston and Thorpe (1996) with slight modification. Briefly, 1 ml of plasma was pre-warmed to 37 °C, and 10 μl of thrombin solution (100 U of thrombin in 1 ml of 1 m CaCl2) was added to the plasma and stirred vigorously with a sterile wooden stick to induce clot formation. The mixture was incubated at 37 °C for 10 min and then at ambient temperature for 1 h. Supernatant was then separated by centrifugation at 18 000 × g for 15 min.
Sera and serum analogues were tested for IgG antibodies against DV using the PanBio Dengue IgG Indirect ELISA (PanBio) according to the manufacturer's instructions. The specificity of the ELISA was 99.2%; its sensitivity, 96.2% (McBride et al. 1998). The absorbance of each well was measured by a Benchmark Plus microplate spectrophotometer (Bio-Rad) at 450 nm with the reference filter at 620 nm. Measured absorbance was converted into PanBio units according to the formula provided by the manufacturer: PanBio unit = (absorbance of sample/mean of cut-off value) × 10. PanBio unit >11 indicated seropositive for dengue infection (PanBio).
Serum analogues prepared from the Red Cross plasma samples were used in the ELISA. As serum analogue was not the specified sample type, we compared the ELISA results between sera and serum analogues. To do this, the ELISA-positive sera from the researcher-collected samples were run in parallel with the serum analogues converted from the paired plasma samples.
Data were analysed using SPSS software (V17.0, SPSS Inc., Chicago, IL, USA). Individual univariate qualitative variables were analysed for association with dengue seropositivity by chi-squared test. The odds ratio (OR) and 95% confidence intervals (CI) were also calculated for each potential factor to estimate the strength of risk related to seropositivity. Statistically significant (P ≤ 0.05) factors were subjected to multivariate analysis by logistic regression with seropositivity as the dependent variable. Comparison of PanBio units obtained from the paired sera and serum analogues was performed using paired t test.
A total of 685 subjects were recruited from April 2007 to July 2009 for this study, in which the researcher-collected samples accounted for 49.8% (341) and the Red Cross samples accounted for 50.2% (344). According to the 2006 Population By-census of the Census and Statistics Department of Hong Kong, our sample size represented 0.01% of the total population of Hong Kong (6 864 346 people). The gender proportion of the subjects in this study was similar to that of the 2006 Population By-census. On the other hand, we had a higher percentage of subjects aged 15–24 and 25–34 years, and a lower percentage aged 65 years or older (Table 1).
|Characteristics||2006 population by censusa||Subjects recruited in this studyb|
|Male||3 272 956||47.7||341||49.8|
|Female||3 591 390||52.3||344||50.2|
|Total||6 864 346||100||685||100|
|25–34||1 052 126||15.3||165||24.1|
|35–44||1 248 855||18.2||109||15.9|
|45–54||1 193 788||17.4||102||14.9|
|Total||6 864 346||100||684||100|
|Hong Kong||1 268 112||18.5||40||12.0|
|Kowloon||2 019 533||29.4||94||28.8|
|New Territories||3 573 635||52.1||196||59.0|
|Total||6 861 280||100||330||100|
Questionnaires from the 341 subjects of the researcher-collected group were analysed. Of the returned questionnaires, eight missed information on race and birth place and one missed information on residential district and hometown. Compared to the population by districts in Hong Kong in 2006, the residential distribution of the researcher-collected subjects was close to that of the total population (Table 1).
Eleven samples (of 685) tested positive for DV by indirect IgG ELISA; the overall prevalence of DV was 1.6%. Eight of the positive samples were researcher-collected.
When the 8 DV seropositive sera from the researcher-collected samples were compared with the paired serum analogues, the latter group had lower PanBio units (P < 0.05). Five serum analogue samples were 0.5–3.3% lower, while three samples were 12.5–14.6% lower (Table 2). Hence, there might be chance for false negative to occur in serum analogues with borderline PanBio units. However, the serum analogues from Red Cross had clear-cut results; therefore, false negatives were unlikely.
|Sample type||% difference|
The univariate predictors for dengue seropositivity are shown in Table 3. Seropositivity was significantly associated with subjects who were not born and had not grown up in Hong Kong (P < 0.01, OR 18.00, CI 2.18–148.36) and whose hometowns were not in Hong Kong. Among these seropositive individuals, only one was local, five came from China and two from Indonesia (P < 0.01, OR 19.71, CI 2.93–162.52). Individuals who had visited Southeast Asia (P = 0.04, OR 3.05, CI 1.17–7.79) in the past 12 months were more likely to be seropositive.
|Demographic information||ELISA + (%)||ELISA - (%)||P-valuea||OR (95% CI)b|
|Male||4 (36.4)||337 (50.0)||0.37||0.57 (0.17–1.97)|
|Female||7 (63.6)||337 (50.0)|
|>30||10 (90.9)||345 (51.3)||0.09||9.51 (1.21–74.68)|
|≤30||1 (9.1)||329 (48.1)|
|Detailed demographic information and travel historyc|
|HK Island||0 (0.0)||45 (13.6)||0.46||–|
|Kowloon||2 (25.0)||97 (29.2)|
|New Territory||6 (75.0)||190 (57.2)|
|Non-Chinese||0 (0.0)||1 (0.3)||0.88||1.00 (1.00–1.01)|
|Chinese||8 (100)||324 (99.7)|
|Born and grow up in HK|
|No||7 (87.5)||91 (28)||<0.01||18.00 (2.18–148.36)|
|Yes||1 (12.5)||234 (72.0)|
|China and others||7 (87.5)||87 (27.6)||<0.01||19.71 (2.39–162.52)|
|Hong Kong||1 (12.5)||245 (73.8)|
|Travel within past 12 months|
|Yes||7 (87.5)||234 (70.3)||0.29||2.96 (0.37–24.39)|
|No||1 (12.5)||99 (29.7)|
|Eastern Asia 1|
|Yes||7 (87.5)||185 (55.6)||0.07||5.60 (0.68–46.03)|
|No||1 (12.5)||148 (44.4)|
|Eastern Asia 2|
|Yes||0 (0.0)||32 (9.6)||0.36||0.90 (0.87–0.94)|
|No||8 (100)||301 (90.4)|
|Yes||3 (37.5)||41 (12.3)||0.04||3.05 (1.19–7.79)|
|No||5 (62.5)||292 (87.7)|
|Yes||0 (0.0)||7 (2.1)||0.68||0.98 (0.96–1.00)|
|No||8 (100)||326 (97.9)|
|Yes||0 (0.0)||15 (4.5)||0.54||0.96 (0.93–0.98)|
|No||8 (100)||318 (95.5)|
|Yes||0 (0.0)||13 (3.9)||0.57||0.96 (0.94–0.98)|
|No||8 (0.0)||320 (96.1)|
|Yes||0 (0.0)||4 (1.2)||0.76||0.99 (0.98–1.00)|
|No||8 (100)||329 (98.8)|
After multivariate analysis, individuals who had visited countries in Southeast Asia in the past 12 months (P = 0.03, OR 5.38, CI 1.13–25.54) were significantly associated with seropositivity (Table 4).
|Covariate||P-value||OR (95% CI)|
|Born and grow up in Hong Kong|
|China and other places||9.27 (0.01–76183.1)|
|Travel to Southeast Asia|
This is the first published report on seroprevalence of dengue infection in Hong Kong. The seroprevalence of dengue infection in the adult population was 1.6%, much lower than that reported in nearby Asian countries (9.2–59%) (Chen et al. 1997; Wilder-Smith et al. 2004; Yew et al. 2009; Ukey et al. 2010). The low seroprevalence could be due to the absence of Aedes aegypti, which was not found in Hong Kong after the mid-1950s; however, the less anthropophilic Aedes albopictus is abundant in Hong Kong (Cheung & Fok 2009). In Hong Kong's subtropical climate, the mean temperature, total rainfall and relative humidity for the past 30 years (1981–2010) were 23.3 °C, 2398.5 mm and 82%, respectively (Hong Kong Observatory 2012). The climate is optimal for proliferation of Aedes aegypti (Ansari & Razdan 1998). The absence of Aedes aegypti could be due to interspecies competition for larval resources (Juliano 1998) or sterility as a result of interspecies mating with Aedes albopictus (Nasci et al. 1989). Furthermore, the low seroprevalence may be attributed to the effective dengue vector surveillance programme of the Food and Environmental Hygiene Department (FEHD) of Hong Kong, which was started in 1998. Regular mosquito surveys are conducted by FEHD, and breeding activity is monitored using ovitraps in various territories and port areas of Hong Kong (Cheung & Fok 2009).
We found that individuals who were not born and did not grow up in Hong Kong were at significantly increased risk of dengue seroprevalence. Besides, individuals whose hometown was in China or other countries were associated with seropositivity. Three of the foreigners came from the UK and Canada and the others from dengue-endemic areas including Macau, Indonesia and Venezuela. According to the information of the original hometown, these individuals may have acquired dengue infections before coming to live in Hong Kong.
We also found that individuals who had visited Eastern Asia (China, Macau) or Southeast Asia (Indonesia) during the last year were more likely to be seropositive. Both regions are popular destinations for Hong Kong citizens and endemic for dengue. In multivariate analysis, the only independent predictor for dengue seropositivity was visit to Southeast Asia (Indonesia). Hong Kong citizens have a high propensity to travel: according to data from the Department of Health of Hong Kong, there were more than 70 million outbound trips, equivalent to about 10 journeys per person in 2005 (World Tourism Organization 2006). About 94% travelled to Mainland China, Macau and Taiwan and 5% to South and Southeast Asia (World Tourism Organization 2006). Frequent trips to endemic areas also increase the risk of travel-associated dengue fever. In Hong Kong, most of the dengue fever cases were imported from nearby endemic countries; the number of cases has been rising since 2000. However, there was a local outbreak in Ma Wan in 2002, followed by emergence of several local cases in 2003 and 2010 (Leung 2010; Chuang et al. 2008). As Aedes albopictus is abundant in Hong Kong, the risk for spreading of DV infection within the local community cannot be ignored.
In spite of several factors that were significantly associated with seroprevalence, we noted a wide range of the 95% CI for those factors due to the small sample size, which might also affect the accuracy of the statistical analyses. Other limitations included the absence of subjects under 14 years of age and the possibility of selection bias as the Red Cross blood donors were screened for health problems. Although the sample size represented 0.01% of the total population and the demographic characteristics of the subjects were comparable to those of the Hong Kong population, a larger sample size and the inclusion of age groups below 14 years are required for more appropriate conclusions about the association between risk factors and dengue seroprevalence in the general population of Hong Kong.
The dengue seroprevalence in Hong Kong was 1.6%. Visit to Southeast Asia was the only independent predictor for seropositivity. Although the figure is not alarming, frequent travel between Hong Kong and Southeast Asian countries and the presence of Aedes albopictus in Hong Kong suggest that the risk of dengue virus infection cannot be overlooked. Precautionary measures against mosquito bites should be taken, and physicians need to be vigilant in case of febrile patients with recent travel history to endemic regions.
We thank Professor Iris Benzie of the Hong Kong Polytechnic University for help in recruiting subjects from hospital and the staff of Kwai Tsing District Council (Lai King) office for assistance in subject recruitment at the community centre. The study was funded by the Hong Kong Polytechnic University, Internal Competitive Research Grants G-YD97 and G-YH68.