When evaluated by the Institutional Review Board of the CDC, this non-research activity was considered a component of the public health response to the dengue outbreak in St. Croix and thus did not require review. Data analysis was performed on an anonymized dataset.

Routine surveillance in the USVI relies upon healthcare provider-initiated requests for dengue laboratory testing submitted to the DB. Each diagnostic specimen is accompanied by a CDC Dengue Case Investigation Form (DCIF). The DCIF is used to report basic demographic and pertinent clinical information. The majority of the suspected dengue cases are reported from the island's sole tertiary care hospital, the Governor Juan F. Luis Hospital and Medical Center (JFL). Located in the capital, this 130-bed healthcare facility services the entire island.

A suspected dengue case was defined as a febrile illness clinically compatible with DF according to the treating physician. World Health Organization (WHO) criteria [1] were used to classify clinical cases of DHF as follows: (1) fever, or a recent history of fever of 2–7 days duration, accompanied by (2) thrombocytopenia (≤100,000 cells/mm³), (3) hemorrhage, and (4) objective evidence of plasma leakage (e.g. pleural effusion or ascites, an abnormally low serum albumin or protein, or hemoconcentration [defined as hematocrit values ≥20% above mean values for sex and age [9] or ≥20% increase in hematocrit over the stabilized value at discharge]).

All laboratory testing was performed at the DB. For laboratory diagnosis of dengue infections, a real-time, reverse transcriptase PCR assay (RT-PCR, TaqMan Applied Biosystems, Foster City, California, USA ) [10] or viral culture using C6/36 Aedes albopictus mosquito cells or tissues from intrathoracically-inoculated adult Toxorhynchites amboinensis mosquitoes [11], [12] was used to identify virus in serum specimens collected ≤5 days from the onset of fever (acute-phase serum specimen). To determine the outbreak strain, RNA was extracted from tissue culture supernatant (Qiagen). cDNA was generated using Sensiscript RT-PCR (Qiagen) with random hexamers (Applied Biosciences). Fourteen pooled overlapping 2000 nucleotide (nt) amplicons were generated by RT-PCR at the DB and sequenced at the Broad Institute by bidirectional Sanger. [13]

An immunoglobulin M (IgM) antibody-capture enzyme-linked immunosorbent assay (MAC-ELISA) was also used to detect anti-dengue IgM antibodies in acute and convalescent-phase (collected >5 days after the onset of fever) serum specimens. [14] If sufficient anti-dengue IgM antibodies were detected such that the optical density was ≥0.20, [14] this was considered evidence of a recent dengue infection. Serum specimens were also tested for the presence of anti-dengue immunoglobulin G (IgG) antibodies using an IgG ELISA. [15] The presence of anti-dengue IgG antibodies in acute serum specimens was used to distinguish primary (first) from secondary (sequential) dengue infections. An IgG titer ≥163,840 in a single serum specimen was also considered laboratory-positive for recent, secondary dengue infections even without positive virus identification results. [15] There was a limited supply of dengue antigen in 2005, thus a randomly selected subset of specimens were tested by IgG ELISA. All suspected dengue cases with laboratory evidence of an acute or recent infection were considered laboratory-positive. All suspected dengue cases with negative acute-phase results and no convalescent specimen were classified as indeterminate. All convalescent specimens that lacked an IgM response were laboratory-negative.

To investigate risk factors for progression to DHF, we conducted a case-control study among laboratory-positive patients presenting to the Emergency Department (ED) or admitted to JFL Hospital during 2005. A case was defined as a laboratory-positive patient meeting all four WHO criteria for DHF. A control was defined as a laboratory-positive patient who did not meet criteria for DHF but was evaluated for thrombocytopenia, plasma leakage and hemorrhage. Hospitalized patients with negative or indeterminate dengue laboratory results were excluded from this analysis. The medical records of all cases and controls were reviewed. Data abstracted from the medical records included demographic information (date of birth, self-reported race on medical records, etc.) and information on clinical course and outcome. The range of normal values set by the hospital's clinical laboratory was used to define the threshold values for low serum albumin and protein levels.

Data from the case-control study were entered directly into an electronic form designed using Questionnaire Development System v2.3 (Nova Research, Bethesda, MD, USA). All data analysis was then performed using SPSS version 12.0 (SPSS Inc, Chicago IL, USA) and SAS version 9.1 (SAS Institute, Cary NC, USA). The Student's t, Pearson's chi-squared, Fisher's exact, and Mann-Whitney U tests were used to assess unadjusted associations between study factors and DHF. All variables with a p-value less than 0.15 in bivariate analysis were included in the final model. Race [16] was added a priori to the final model. Adjusted odds ratios (ORs) and 95% confidence intervals (CIs) were calculated using exact multivariable logistic regression. All tests for significance were performed at an α-level of 0.05.

Between January 1 and December 31, 2005, 331 suspected DF cases (62.2 cases/10,000 population) were reported from St. Croix, USVI with a peak in July (Figure 1). Fifty-three percent were male, and the median age was 19 years (range: 3 months to 94 years). Cases were reported island-wide, but the most frequently reported place of residence (13%) was Williams Delight in southwest St. Croix. The most commonly reported signs and symptoms in suspected cases were headache (88%), fever (83%), body pain (83%), joint pain (68%), chills (56%), and retro-orbital pain (54%). Fifty-four percent of the suspected cases were hospitalized; 94 (28%) had thrombocytopenia, and 77 (21%) had at least one hemorrhagic manifestation.

Of the 331 suspected dengue cases, 123 (37%) were laboratory-positive, 32 (10%) were laboratory-negative, and 176 (53%) were indeterminate. The overall incidence of laboratory-positive cases was 23.1/10,000 population. Of the 44 laboratory-positive cases tested by the IgG ELISA, 3 (7%) were primary infections and 41 (93%) were secondary dengue infections. Fifteen (5%) of the suspected cases were laboratory-positive and met the WHO criteria for DHF. Of the 278 specimens tested by RT-PCR, 61 were identified as DENV-2. This was the only serotype identified from specimens collected during this outbreak. One of the DENV-2 isolates was sequenced and it corresponded to the southeast Asian lineage descending from the Jamaican 1983 isolate. [13]

One fatality occurred among the 331 suspected cases. The patient, a 14 year-old Afro-Caribbean female, presented to the hospital on day 3 post onset of illness with complaints of fever, generalized body pain and abdominal pain. Upon admission to the JFL Hospital, her platelet count was 33,000/mm³, and within the first 24 hours, she experienced hematuria, pleural effusions, severe abdominal pain, cold/clammy skin, and altered mental status. She suffered circulatory failure then expired soon thereafter. This patient met WHO criteria for DHF and was found to have had an acute, secondary dengue infection.

Eighty-nine laboratory-positive patients were identified from the JFL Hospital including 15 (17%) who met WHO criteria for DHF (cases) and 74 (83%) who did not (controls). Of these 89 patients, 30% had at least one hemorrhagic manifestation. Of the 15 DHF cases, a single patient experienced DHF grade III and the remainder had grade II. [1] There was no significant difference in the median length of hospital stay between cases and controls (4.5 vs. 4.0 days, p = 0.74). Selected clinical characteristics of cases and controls are compared in Table 1. Cases and controls were similar with respect to gender, race, and history of hypertension, diabetes, or asthma. On bivariate analysis, age was the only variable significantly associated with DHF (p = 0.03). Similarly, in the final multivariable model (Table 2), age was the only variable associated with DHF (OR [95% CI]: 1.033 [1.003,1.064]). As age was considered as a continuous variable in the model, its OR is interpreted as follows: for every additional year of life, there would be a 3% increase in the odds of acquiring DHF. As demonstrated in table 3 and figure 2, there was a higher proportion of DHF cases in those above the age of 40.