RESEARCH ARTICLE

Enhanced Surveillance for Fatal Dengue-Like

Acute Febrile Illness in Puerto Rico, 2010-
2012
Kay M. Tomashek1*, Aidsa Rivera1, Brenda Torres-Velasquez1, Elizabeth
A. Hunsperger1, Jorge L. Munoz-Jordan1, Tyler M. Sharp1, Irma Rivera2, Dario Sanabria2,
Dianna M. Blau3, Renee Galloway4, Jose Torres2, Rosa Rodriguez2, Javier Serrano2,
Carlos Chavez2, Francisco Davila2, Janice Perez-Padilla1, Esther M. Ellis1,
Gladys Caballero5, Laura Wright6, Sherif R. Zaki3, Carmen Deseda7, Edda Rodriguez2,
a11111 Harold S. Margolis1
1 Dengue Branch, Division of Vector-Borne Diseases, Centers for Disease Control and Prevention (CDC),
San Juan, Puerto Rico, 2 Puerto Rico Institute of Forensic Sciences, San Juan, Puerto Rico, 3 Infectious
Diseases Pathology Branch, Division of High Consequence Pathogens and Pathology, CDC, Atlanta,
Georgia, United States of America, 4 Bacterial Special Pathogens Branch, Division of High Consequence
Pathogens, CDC, Atlanta, Georgia, United States of America, 5 Demographic Registry of Puerto Rico, San
Juan, Puerto Rico, 6 Geospatial Research, Analysis, and Services Program, Division of Toxicology and
Human Health Sciences, ATSDR, Atlanta, Georgia, United States of America, 7 Puerto Rico Department of
OPEN ACCESS Health, San Juan, Puerto Rico
Citation: Tomashek KM, Rivera A, Torres-
Current address: Office of Clinical Research Resources, Division of Microbiology and Infectious Diseases,
Velasquez B, Hunsperger EA, Munoz-Jordan JL,
National Institute of Allergy and Infectious Diseases, National Institutes of Health, Rockville, Maryland,
Sharp TM, et al. (2016) Enhanced Surveillance for United States of America
Fatal Dengue-Like Acute Febrile Illness in Puerto * [email protected]
Rico, 2010-2012. PLoS Negl Trop Dis 10(10):
e0005025. doi:10.1371/journal.pntd.0005025

Editor: Charles Apperson, North Carolina State

University, UNITED STATES
Abstract
Received: June 27, 2016

Accepted: September 8, 2016

Background
Published: October 11, 2016
Dengue is a leading cause of morbidity throughout the tropics; however, accurate popula-
Copyright: This is an open access article, free of all tion-based estimates of mortality rates are not available.
copyright, and may be freely reproduced,
distributed, transmitted, modified, built upon, or
otherwise used by anyone for any lawful purpose.
The work is made available under the Creative Methods/Principal Findings
Commons CC0 public domain dedication.
We established the Enhanced Fatal Acute Febrile Illness Surveillance System (EFASS) to
Data Availability Statement: The Centers for estimate dengue mortality rates in Puerto Rico. Healthcare professionals submitted serum
Disease Control and Prevention has determined and tissue specimens from patients who died from a dengue-like acute febrile illness, and
that the data cannot be made publicly available for
ethical reasons. Specifically, public availability of
death certificates were reviewed to identify additional cases. Specimens were tested for
data would compromise patient privacy and be markers of dengue virus (DENV) infection by molecular, immunologic, and immunohisto-
detrimental to surviving family members. Contact: chemical methods, and were also tested for West Nile virus, Leptospira spp., and other
Laura Youngblood, Human Subjects Advisor,
pathogens based on histopathologic findings. Medical records were reviewed and clinical
Division of Vector Borne Diseases, NCEZID, CDC.
Email: [email protected] | Office: (404) 639- data abstracted. A total of 311 deaths were identified, of which 58 (19%) were DENV labo-
6394 ratory-positive. Dengue mortality rates were 1.05 per 100,000 population in 2010, 0.16 in
Funding: The authors received CDC Dengue 2011 and 0.36 in 2012. Dengue mortality was highest among adults 1964 years and
Branch programmatic funding for this project. No seniors 65 years (1.17 and 1.66 deaths per 100,000, respectively). Other pathogens

PLOS Neglected Tropical Diseases | DOI:10.1371/journal.pntd.0005025 October 11, 2016 1 / 19

 Enhanced Surveillance for Fatal Dengue

additional sources of funding were received. The identified included 34 Leptospira spp. cases and one case of Burkholderia pseudomallei
funders had no role in the system design, data and Neisseria meningitidis.
collection and analysis, decision to publish or
preparation of the manuscript.
Conclusions/Significance
Competing Interests: The authors have declared
that no competing interests exist. EFASS showed that dengue mortality rates among adults were higher than reported for
influenza, and identified a leptospirosis outbreak and index cases of melioidosis and
meningitis.

Author Summary
Dengue is a major public health problem in the tropics. Despite its global importance,
population-based mortality rates attributable to dengue are largely unknown. Dengue vac-
cines are now in late stage clinical trials and one vaccine has been licensed in several coun-
tries. Evidence-based decisions regarding the future use of dengue vaccines will depend on
robust estimates of disease burden which should include mortality. To estimate mortality
due to dengue in Puerto Rico, where dengue is endemic, we developed an enhanced sur-
veillance system to detect fatalities due to a preceeding dengue-like acute febrile illness
using more sensitive case identification and laboratory methods than the previous passive
method. This surveillance system found the dengue mortality rate was 1.05 per 100,000
Puerto Rico residents in 2010, the highest rate ever detected. Among adults aged 1964
years, mortality from dengue (1.17 deaths per 100,000) was higher than from other infec-
tious diseases, including influenza. The utility of this enhanced surveillance system was
further proven through the identification of an outbreak of leptospirosis as well as detec-
tion of other diseases of public health importance, including melioidosis and meningitis.

Introduction
Dengue is a major public health problem worldwide. While most dengue virus (DENV) infec-
tions are asymptomatic or result in a mild acute febrile illness (AFI), some are life-threatening
due to plasma leakage [1, 2]. With no antivirals to treat dengue or prevent its severe manifesta-
tions [3], early recognition of severe dengue and timely supportive care is used to reduce mor-
tality [46]. Several dengue vaccines are in late stage clinical trials and one was recently
licensed in several countries [7]. Decisions regarding their use will depend on vaccine perfor-
mance and safety, and reduction of disease burden, including deaths.
Globally, an estimated 3.9 billion people are at risk of DENV infection, and 96 million den-
gue cases are estimated to have occurred in 2010 alone [8]. Despite its global presense, robust
estimates of population-based dengue mortality rates are lacking. Most estimates have been
derived from passive surveillance data [914] or hospital-based, retrospective case reviews [15,
16]. During epidemic periods, these methods have produced annual mortality rates that ranged
from 0.300.59 deaths per 100,000 population. However, these approaches have not been vali-
dated as to under recognition due to misdiagnosis or under reporting [14].
Dengue has been endemic in Puerto Rico since the late 1960s [17, 18], and after the first
deaths were reported in 1986, surveillance for deaths due to dengue was established in 1987
[19]. Mortality data have been collected through the island-wide Passive Dengue Surveillance
System (PDSS), a hospital-based Infection Control Nurse Dengue Surveillance System
(ICNDSS) that operated until 2007, and review of death certificates. Evaluations of these

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 Enhanced Surveillance for Fatal Dengue

systems identified misdiagnosis and underreporting of cases, and failure to include dengue on
death certificates of known laboratory-positive dengue cases [14, 20]. Few suspected fatal cases
had tissue specimens or appropriately timed pre-mortem serum specimens for diagnostic test-
ing, which resulted in a high proportion of indeterminate diagnostic results [14, 1921].
In 2009, the Centers for Disease Control and Prevention Dengue Branch (CDC-DB), Puerto
Rico Department of Health (PRDH), Instituto de Ciencias Forenses de Puerto Rico (in English,
Puerto Rico Institute of Forensic Sciences [PRIFS]), Demographic Registry of Puerto Rico, and
CDC Infectious Diseases Pathology Branch (CDC-IDPB) established the Enhanced Fatal Acute
Febrile Illness Surveillance System (EFASS) to define mortality due to dengue-like AFI, and deter-
mine the etiology of these cases. We describe the findings from the first three years of EFASS.

Methods
Ethics Statement
This project underwent CDC institutional review and formal institutional review board review
was not required since the case-patients were deceased. Because cases were reported in the con-
text of public health surveillance, the informed consent of patients families was not sought.
Patient identifiers were removed from the dataset prior to analysis.

Data Sources and Case Detection

EFASS used enhanced surveillance to detect dengue-like AFI deaths, improve reporting, and
standardize collection of specimens at autopsy. While PDSS provided retrospective diagnostic
data on fatal suspected dengue cases reported early in their illness, the primary source of EFASS
cases was reporting by participating epidemiologists, pathologists, and registry statisticians.
Specifically, they were asked to report and submit samples from all fatal cases whose death
occurred during or immediately following a dengue-like AFI defined by the presence of fever
(body temperature 38.0C axillary) or history of fever for 7 days. This included deaths with
pre-defined diagnostic codes on the medical record, autopsy report or death certificate (S1
Appendix); the list of ICD codes was developed in 2009 after a review of the 19942007 fatal
laboratory-positive dengue cases was conducted. Surveillance was enhanced by collaboration
with pathologists and epidemiologists at hospitals most likely to encounter severe dengue cases,
and included training and provision of standardized protocols. Dengue-like AFI fatalities that
occurred at home or within 24 hours of hospital admission and referred to PRIFS were
included. Collaborators were contacted weekly, and death certificates, PDSS, and National Noti-
fiable Diseases Surveillance System (NNDSS) were routinely queried to identify suspected cases.

Specimen and Data Collection

Once a suspected case was identified, serum, whole blood, and tissue specimens were obtained,
and PRIFS pathologists completed a Surgical Pathology and Autopsy Report (SPAR) (S2 Appen-
dix). Cases with history of respiratory failure had a nasopharyngeal swab submitted for testing.
Data was abstracted from medical records of all health care visits during the illness for labo-
ratory-positive dengue cases using a standard instrument that captured demographic charac-
teristics, past medical history, clinical course, and management.

Diagnostic Testing
Serum specimens were tested by a DENV-serotype specific real time, reverse transcriptase-
polymerase chain reaction (rRT-PCR) assay [22], an anti-DENV IgM enzyme-linked immuno-
sorbent assay (MAC ELISA) [23], and an anti-DENV IgG ELISA [24]. Serum specimens were

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 Enhanced Surveillance for Fatal Dengue

also tested by anti-West Nile virus (WNV) MAC-ELISA and, if positive, WNV-specific
rRT-PCR and 90% plaque reduction neutralization tests (PRNT90) were performed [25].
Serum specimens with sufficient volume were sent to CDC Bacterial Special Pathogens Branch
and tested for Leptospira IgM using the ELISA ImmunoDOT kit (GenBio, Inc., San Diego,
CA). Acute specimens were tested for nucleic acid by rRT-PCR and 20 Leptospira reference
antigens representing 17 serogroups by microscopic agglutination test (MAT) [26]. RNA was
extracted from nasopharyngeal specimens and tested for Influenza A and B viral genome by
rRT-PCR [27].
Tissue specimens were tested at the CDC-IDPB for DENV antigen or nucleic acid by immu-
nohistochemistry (IHC) and RT-PCR, respectively [28]. If clinical presentation or histopathol-
ogy were suggestive of another etiology pathogen-specific diagnostic testing was performed
[26, 29].

Definitions
A fatal suspected dengue-like AFI case had a dengue-like AFI that immediately preceded death
in a Puerto Rico resident. A fatal laboratory-positive dengue case was a suspected case with
DENV nucleic acid in serum or tissue; DENV antigen in tissue; IgM seroconversion in paired
specimens; or IgM in a single specimen. A fatal laboratory-negative dengue case was a sus-
pected case with no molecular, immunodiagnostic or IHC markers of DENV infection. A fatal
laboratory-indeterminate dengue case was a suspected case with no DENV nucleic acid or anti-
DENV IgM in the acute serum specimen (collected 5 days post-illness onset [DPO]) and no
available convalescent serum specimen (6 DPO). A fatal dengue co-infection was a fatal sus-
pected dengue-like AFI case with DENV nucleic acid in serum or tissue and another pathogen
detected by PCR or IHC. A primary DENV infection was a fatal laboratory-positive dengue
case without anti-DENV IgG in the acute serum specimen [30] and a secondary DENV infec-
tion had anti-DENV IgG in the acute specimen.
A fatal laboratory-confirmed leptospirosis case was a suspected dengue-like AFI case with
4-fold increase in MAT titers in paired specimens, MAT titer 800 in a single specimen, or
detection of Leptospira spp. nucleic acid in serum by PCR or antigen in tissue by IHC. A proba-
ble fatal leptospirosis case had a MAT titer >200 but <800 in a serum specimen.
Dengue fever (DF), dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS)
were defined according to the 1997 World Health Organization (WHO) guidelines [31]
(Table 1). Dengue, dengue with warning signs, and severe dengue were defined according to
2009 WHO guidelines [2]. Definitions for severe dengue, other clinical features and medical
complications are shown in Table 1.

Data Analysis
Frequencies were calculated for demographic, clinical and laboratory features of fatal labora-
tory-positive dengue cases. Rates of fatal laboratory-positive dengue cases per 100,000 Puerto
Rico population were calculated by age group, sex, and municipality using US Census data
[45]. Incidence rate ratios (IRR) were calculated to compare females to males. Case fatality
rates (CFR) were calculated by dividing the number of fatal laboratory-positive dengue cases
by PDSS laboratory-positive dengue cases. Statistical differences in proportions were tested by
Chi-square or Fisher's exact tests. Differences between municipality-specific fatal laboratory-
positive dengue cases and PDSS laboratory-positive dengue cases were examined by calculation
of Pearson correlation coefficients. A geographically weighted regression model was used to
determine if the number of fatal cases differed from the expected based on PDSS laboratory-
positive dengue cases in the municipality and neighboring municipalities. Data analyses were

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 Enhanced Surveillance for Fatal Dengue

Table 1. Clinical findings used to define fatal laboratory-positive dengue cases.

Clinical Syndrome or Condition Laboratory or Clinical Definitions Used Ref.
Dengue fever, dengue hemorrhagic fever, Case had signs or symptoms as defined in the 1997 WHO Guidelines. Our definitions include: [31]
dengue shock syndrome
Leucopenia White cell count less than 5.0 109 cells/L.
Plasma Leakage Case met at least one of following criteria: [32
1. Hemoconcentration: 20% increase in hematocrit above the age/sex-specific U.S. mean, or 36]
20% hematocrit increase 410 days post-illness onset (DPO) compared to sample taken 3
DPO with no blood transfusions;
2. Pleural effusion or ascites detected by imaging;
3. Serum albumin <2.5th percentile for age/sex
Dengue, dengue with warning signs, severe Case had signs or symptoms as outlined in the 2009 WHO Guidelines. Our more specific [2]
dengue definition of severe dengue criteria included:
Severe plasma leakage Case had plasma leakage (defined above) leading to shock or effusions resulting in acute
respiratory distress, respiratory failure or ARDS.
Severe bleeding Case had intracranial bleed, or bleeding that resulted in hemodynamic instability requiring fluid
replacement and/or blood transfusion.
Severe organ impairment Case had acute liver failure, myocarditis, or neurologic impairment necessitating intubation or
resulting in death.
Jaundice Case had clinically apparent jaundice, or plasma bilirubin greater than 3 mg/dL.
Acute liver failure (ALF) Case with no chronic liver disease had acute hepatitis plus hepatic encephalopathy of any grade, [37]
jaundice, and new onset coagulopathy defined by international normalized ratio 1.5.
Myocarditis Case had dyspnea, chest pain, dizziness, or weakness; echocardiographic evidence of global
dysfunction; left ventricular ejection fraction <30%, and pericardial effusion or elevated serum
troponin T or I.
Coma Case had a Glasgow Coma Score of less than 9 and/or was unconscious and unresponsive to
painful or verbal stimuli for more than 6 hours.
Other Clinical Outcomes
Acute hepatitis Case had serum alanine aminotransferase 10 times the upper limit of normal (ULN) or >400 U/L
and no underlying chronic liver disease (e.g., hepatitis C or B, cirrhosis of other or unknown
etiology).
Acute acalculous cholecystitis (AAC) Case had severe abdominal pain plus two major, or one major and two minor sonographic or CT [38]
scan criteria. Gallbladder wall thickening not used as ACC criteria when ascites or
hypoalbuminemia present.
Acute renal failure Case had at least one of the following criteria: [39]
1. 3-fold increase in serum creatinine;
2. 75% decrease in glomerular filtration rate;
3. Serum creatinine 4.0 mg/dL with acute increase >0.5 mg/dL;
4. Urine output <0.3 mL/kg/hour in 24 hours or anuria for 12 hours.
Medical Complications of Dengue
Prolonged shock Case had hypotension for age for 8 hours.
Metabolic acidosis Case had an arterial pH <7.35 and bicarbonate <24 mmol/L with a serum bicarbonate within 2 [40]
mmol/L, and a normal or low arterial carbon dioxide.
Fluid overload Case had periorbital edema, dyspnea, weight gain, or abdominal compartment syndrome.
Abdominal compartment syndrome Case had intraabdominal pressure >20 mm Hg with attributable organ failure. [41]
Acute hypoxemic respiratory failure Case had an arterial oxygen <60 mm Hg and normal or low arterial carbon dioxide level while on
oxygen.
Acute respiratory distress syndrome (ARDS) Case met criteria outlined in American-European Consensus Conference definition. [42]
Healthcare-associated infections (HAI) Case had infection that became clinically evident >48 hours after hospitalization. [43]
Disseminated intravascular coagulation (DIC) Case had a DIC score of 5 that accounted for platelet count, D-dimer, PT, and fibrinogen. [44]
doi:10.1371/journal.pntd.0005025.t001

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 Enhanced Surveillance for Fatal Dengue

conducted using STATA (Stata Corporation, College Station, TX) and ArcGIS (Environmental
Systems Research Institute, Redlands, CA); maps were created using ArcMap.

Results
Identification of Fatal Cases and Diagnostic Testing
During 20102012, a total of 311 fatalities following a dengue-like AFI were detected by EFASS
and 40,881 suspected dengue cases were reported to PDSS, of which 17,929 (44%) were dengue
laboratory-positive. Of all fatalities detected, 146 (47%) were identified and reported by PRIFS
pathologists, 93 (30%) by death certificate review, 50 (16%) by hospital epidemiologists, 15
(5%) by PDSS, four (1%) by NNDSS, and three (1%) by chart review as part of another study.
Serum and tissue specimens were available for 148 (48%) cases, serum alone for 138 (44%)
cases, tissue alone for 16 (5%) cases, and 9 (3%) cases had no diagnostic specimens. Of the 164
cases (53%) with tissue, one case was not tested because of sample quality. Of evaluable cases,
159 (98%) had liver, 156 (96%) lung, 155 (95%) kidney, 142 (87%) spleen, 98 (60%) lymph
nodes, and 66 (41%) intestine. A nasopharyngeal swab was submitted for 27 (9%) cases.
A pathogen was identified in 120 (40%) of 302 cases with a diagnostic specimen. A pathogen
was more likely to be identified in cases with tissue specimens than in those without (69% ver-
sus 45% respectively, P <0.0001). Overall, 58 (19%) fatal cases were dengue laboratory-posi-
tive, 167 (54%) were dengue laboratory-negative, and 77 (25%) were dengue laboratory-
indeterminate. Other pathogens identified included: Leptospira spp. in 34 (11%) cases (32 con-
firmed, two probable); Staphylococcus spp. in nine (3%) cases; Streptococcus spp. in nine (3%)
cases; influenza A virus in three (1%) cases; and one case each with Neisseria meningitidis, Bur-
kholderia pseudomallei, Proteus spp., Clostridium perfringens, Cryptococcus neoformans, Klebsi-
ella pneumoniae, and an unidentified Gram-positive coccus.

Characteristics of Fatal Laboratory-Positive Dengue Cases

Of the 58 fatal laboratory-positive dengue cases, 53 (91%) were DENV RT-PCR positive in tis-
sue, serum or both; four (7%) were anti-DENV IgM positive in a single serum specimen; and
one (2%) demonstrated anti-DENV IgM seroconversion in paired specimens (Table 2). Autop-
sies were performed on 26 (45%) of the 58 fatal laboratory-positive dengue cases and DENV
was most commonly identified by IHC or RT-PCR in liver (18/26, 69%), lung (15/22, 68%), and
kidney (9/24, 38%) tissue. Of 43 cases with rRT-PCR positive serum specimen, 26 (60%) were
DENV-1, 16 (37%) DENV-4, and one (2%) DENV-2; similar to DENV-type distribution in
PDSS during the same time period [18]. Among the 36 fatal laboratory-positive dengue cases
with an available acute serum specimen, 30 (83%) had a secondary DENV infection and 6
(17%) had primary infection. Five of the DENV RT-PCR positive cases had co-infection with
another pathogen, including Leptospira species (4 cases) [46] and Streptococcus species (1 case).

Epidemiology of Fatal Laboratory-Positive Dengue Cases

Fatal laboratory-positive dengue cases occurred in the months with increased PDSS dengue
reporting (Fig 1). The dengue mortality rate was 1.05 per 100,000 population in 2010, 0.16 in
2011 and 0.36 in 2012; the 3-year average mortality rate was 0.52 dengue deaths per 100,000.
The overall CFR for the three-year period was 0.32% and varied from 0.38% during the 2010
epidemic [18],; to 0.39% in 2011, a non-epidemic year, to 0.22% in 2012, an epidemic year.
The median age of fatal laboratory-positive dengue case-patients was 46 years (range: 5
months89 years). Six (10%) case-patients were <20 years old, 39 (67%) 2064 years old, and
13 (22%) 65 years old. EFASS case-patients were significantly older than laboratory-positive

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 Enhanced Surveillance for Fatal Dengue

Table 2. Diagnostic laboratory results for fatal laboratory-positive dengue cases detected by the Enhanced Fatal Acute Febrile Illness Surveil-
lance System, Puerto Rico 20102012.
Diagnostic Result 20102012 2010 (N = 39) 2011 (N = 6) 2012 (N = 13)
(N = 58)
No. (%) No. (%) No. (%) No. (%)
Tissue RT-PCR and IHC positive, and serum RT-PCR positive with or without IgM positive 13 22.4 11 28.2 0 0.0 2 15.4
Tissue RT-PCR positive with or without IHC positive only 10 17.2 8* 20.5 0 0.0 2 15.4
Tissue IHC positive and serum RT-PCR and IgM positive 1 1.7 0 0.0 0 0.0 1 7.7
Serum RT-PCR positive with or without IgM positive only 29 50.0 18 46.2 6 100 5 38.5
Seroconversion by IgM 1 1.7 1 2.6 0 0.0 0 0.0
Serum IgM positive only 4 6.9 1 2.6 0 0.0 3 23.1

* Three were dual infections; two had DENV and Leptospira spp. bacteria identified and one had DENV and Streptococcus pneumonia identified in tissue.

One dual infection with DENV and Leptospira spp. bacteria identified.

One dual infection with DENV and Leptospira spp. bacteria identified.

doi:10.1371/journal.pntd.0005025.t002

dengue case-patients reported to PDSS (median 46 vs. 18 years, P < 0.001) and a higher pro-
portion of fatal laboratory-positive dengue case-patients were adults (90% vs. 49%, respectively,
P < 0.001).
The majority of fatal laboratory-positive dengue case-patients were female (Table 3), and
were significantly more likely to be female than laboratory-positive dengue cases reported to

Fig 1. Number of laboratory-positive dengue cases reported to the Passive Dengue Surveillance System and fatal
laboratory-positive dengue cases detected by the Enhanced Fatal AFI Surveillance System by month of illness
onset, Puerto Rico 20102012.
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 Enhanced Surveillance for Fatal Dengue

Table 3. Characteristics of all fatal laboratory-positive dengue case-patients detected by the Enhanced Fatal AFI Surveillance System, Puerto
Rico, 20102012.
Lab-positive dengue cases Children and adolescents Adults  20 years old
(n = 58) (n = 6) Dengue only DENV co-infections
(n = 47) (n = 5)
Demographics, no. (%)
Female 34 (59) 5 (83) 29 (62) 0 (0)
Born in Puerto Rico 53 (91) 5 (83) 44 (94) 4 (80)
Medical history, no. (%)
Obese 27 (47) 1 (17) 22 (47) 4 (80)
No chronic disease 10 (17) 3 (50) 7 (15) 0 (0)
One chronic disease 13 (22) 2 (33) 10 (21) 1 (20)
More than one chronic 35 (60) 1 (17) 30 (64) 4 (80)
disease
Diabetes 23 (40) 0 (0) 21 (45) 2 (40)
Asthma 13 (22) 2 (33) 10 (21) 1 (20)
Cardiovascular disease 9 (16) 0 (0) 8 (17) 1 (20)
Psychiatric disease 8 (14) 0 (0) 7 (15) 1 (20)
Thyroid disease 8 (14) 0 (0) 8 (17) 0 (0)
Rheumatologic condition 8 (14) 0 (0) 7 (15) 1 (20)
Neurologic disease 6 (10) 0 (0) 6 (13) 0 (0)
Gastrointestinal disease 5 (9) 1(17) 4 (9) 0 (0)
doi:10.1371/journal.pntd.0005025.t003

PDSS during the same period (59% vs. 45%, P <0.05). In 2010, rates of fatal laboratory-positive
dengue were 1.3 times higher among females than males; 1.19 and 0.90 cases per 100,000 popu-
lation, respectively (female-to-male IRR = 1.3; 95% confidence interval [CI] = 0.702.50,
P = 0.20). Slightly less than half (27, 47%) of fatal laboratory-positive dengue case-patients
were obese, similar to that reported in Puerto Rico [47]. Most fatal laboratory-positive dengue
case-patients had more than one pre-existing medical condition (35, 60%). The most common
included: diabetes mellitus (23, 40%), asthma (13, 22%), cardiovascular disease (9, 16%), thy-
roid disease (8, 14%), psychiatric disease (8, 14%), and rheumatologic conditions (8, 14%).
Fatal laboratory-positive dengue case-patients were residents of 35 of the 78 Puerto Rico
municipalities, and most (53, 91%) were born in Puerto Rico (Table 3). Fatal laboratory-posi-
tive dengue rates were highest in 2010 in Maunabo (0.82 per 10,000 residents), Maricao in
2011 (1.59), and Adjuntas in 2012 (0.51). In all years, there was a positive correlation between
the number of PDSS laboratory-positive dengue cases and the number of fatal laboratory-posi-
tive dengue cases in a municipality (R = 0.56, 0.59, 0.62, and 0.73 in 2010, 2011, 2012, and over-
all, respectively) (Fig 2). The number of fatal laboratory-positive dengue cases detected was no
more than expected based on municipality-specific, laboratory-positive dengue incidence rates.
However, there were fewer than expected fatal laboratory-positive dengue cases detected in 11
of 78 municipalities in individual years. Only Quebradillas and Patillas had fewer than expected
fatalities in all years.

Healthcare Seeking Behavior of Fatal Laboratory-Positive Dengue

Case-Patients
All 58 fatal laboratory-positive dengue case-patients sought care at least once during their ill-
ness; 25 (43%) had two healthcare visits, and seven (12%) had three or more visits. The median
interval between fever onset and arrival at the first visit was 3.5 days (range: 08.5 days). Of the

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 Enhanced Surveillance for Fatal Dengue

Fig 2. Incidence of laboratory-positive dengue, and observed and expected number of fatal laboratory-positive dengue cases by
municipality of residence and year, Puerto Rico, 20102012. Left panels: Incidence per 100,000 population of non-fatal, laboratory-
positive dengue cases reported to the Passive Dengue Surveillance System, and number of fatal laboratory-positive dengue cases identified
by the Enhanced Fatal AFI Surveillance System. Right panels: the standard deviation (SD) of the standard residuals are displayed.
Differences >2 SD denotes significantly fewer than expected fatal laboratory-positive dengue cases, while 2 SD denotes significantly more
than expected fatal laboratory-positive dengue cases.
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 Enhanced Surveillance for Fatal Dengue

Table 4. Clinical features and outcomes for fatal laboratory-positive dengue case-patients detected by the Enhanced Fatal AFI Surveillance Sys-
tem, Puerto Rico, 20102012.
First outpatient healthcare visit* At time of death in the hospital
N = 49 N = 55
Days post onset, median (range) 3.5 (08.5) 4.5 (0.513.0)
No. prior visits, median (range) NA 2 (15)
Clinical diagnosis, no. (%)
Dengue 25 (51.0) 34 (61.8)
Viral syndrome 7 (14.3) 2 (3.6)
Gastroenteritis 6 (12.2) 3 (5.5)
Urinary tract infection 3 (6.1) 1 (1.8)
Leptospirosis 2 (4.1) 4 (7.3)
Multi-organ failure 2 (4.1) 4 (7.3)
Pancytopenia 1 (2.0) 3 (5.5)
Dengue and leptospirosis 1 (2.0) 1 (1.8)
Respiratory tract infection 1 (2.0) 0 ---
Dehydration 1 (2.0) 0 ---
Meningitis with shock 0 --- 2 (3.6)
Myocarditis 0 --- 1 (1.8)
Signs and symptoms, no. (%)
Fever measured at facility 28 (57.1) 32 (58.2)
Headache 21 (42.9) 27 (49.1)
Eye pain 6 (12.2) 10 (18.2)
Muscle pain 31 (63.3) 33 (60.0)
Joint pain 14 (28.6) 23 (41.8)
Bone pain 9 (18.4) 15 (27.3)
Rash 9 (18.4) 19 (34.6)
Any bleeding manifestation 14 (28.6) 47 (85.5)
Vomiting 21 (42.9) 34 (61.8)
Abdominal pain 15 (30.6) 31 (56.4)
Diarrhea 12 (24.5) 26 (47.3)
Cough 7 (14.3) 21 (38.2)
Sore throat 6 (12.2) 6 (10.9)
Clinical laboratory findings
White blood cells (109/L), median (range) 4.6 (0.9419.1) 4.9 (0.9418.6)
Leukopenia, no. (%) 25 (51.0) 29 (52.7)
Platelet count (109/L), median (range) 79 (8367) 55 (7269)
Thrombocytopenia, no. (%) 29 (59.2) 46 (83.6)
Hemoconcentrated, no. (%) 11 (22.5) 8 (14.6)
Aspartate aminotransferases (U/L), median (range) 212 (295,733) 284 (2215,481)
Alanine aminotransferases (U/L), median (range) 187 (2322,046) 176 (2322,046)
Aminotransferases 1000 U/L, no. (%) 9 (18.4) 18 (32.7)
Serum sodium 125 mEq/L, no. (%) 2 (4.1) 8 (14.6)
Clinical syndrome, no. (%)
Dengue fever 35 (71.4) 47 (85.5)
Dengue hemorrhagic fever 9 (18.4) 27 (49.1)
Dengue shock syndrome 5 (10.2) 19 (34.6)
Had warning sign(s) 33 (67.3) 53 (96.4)
Severe dengue 17 (34.7) 45 (81.8)
(Continued)

PLOS Neglected Tropical Diseases | DOI:10.1371/journal.pntd.0005025 October 11, 2016 10 / 19

 Enhanced Surveillance for Fatal Dengue

Table 4. (Continued)

First outpatient healthcare visit* At time of death in the hospital

N = 49 N = 55
Disposition, no. (%)
Discharged to home 18 (37.0) NA ---
Transferred to another healthcare facility 7 (14.0) NA ---
Died in Emergency Department 7 (14.0) 12 (22.0)
Died on inpatient ward 2 (4.0) 10 (18.0)
Died in intensive care unit 15 (31.0) 33 (60.0)

*First healthcare visit may have been at a private clinic or a hospital emergency department. Data does not include that collected after admission to the
hospital or transfer to another hospital.

Case-patients could have had more than one diagnosis listed as a discharge, admission or transfer diagnosis.

doi:10.1371/journal.pntd.0005025.t004

49 (84%) cases with a medical record for review, at the first visit, most (71%) had dengue fever,
while some had severe dengue (35%) or DHF (18%) (Table 4); the majority (67%) had warning
signs for severe dengue, the most common being persistent vomiting (21 of 33, 64%), abdomi-
nal pain (15, 45%), and mucosal bleeding (14, 42%). The leading diagnoses during the first visit
were dengue, viral syndrome, gastroenteritis, and urinary tract infection. At the first visit, 17
(35%) were admitted to the hospital, 7 (14%) were transferred to another hospital, 7 (14%)
died in the ED, and 18 (37%) were discharged home. Of those transferred to another hospital,
six (86%) were admitted and died in the ICU, and one died in the ED.
Among the 18 case-patients discharged home after their first visit, seven (39%) had at least
one recorded warning sign, including compensated shock or hemorrhagic manifestations.
Median age was 36.6 years (range 0.674.9), 14 (78%) were female, and 16 (89%) had at least one
chronic medical condition. Two died at home after being discharged; diagnoses of dengue and
moderate dehydration, and acute gastroenteritis. The remaining 16 returned to a hospital on
average 2 days (range 0.03.5 days) after discharge. Nine (56%) died during their second visit,
four (25%) were transferred to another facility, and three (19%) were again discharged home.
Overall, the median interval between illness onset and death was 7.1 days (range: 1.228.4
days); in six (10%) cases the interval exceeded 14 days. Most (43, 74%) case-patients died as an
inpatient in a hospital; however, 12 (21%) died in the Emergency Department prior to hospital
admission, and three (5%) died at home: one after a 2-day hospitalization, and two after being
seen in an ED (Table 4). Of the 43 case-patients who died after being admitted, 33 (57%) died
in the intensive care unit, and 10 (17%) died in an inpatient ward. The median interval from
hospital admission to death was 1.9 days (range: 0.128.8 days).
Only 25 of 58 (43%) fatal laboratory-positive dengue case-patients had dengue, DHF, DSS, or
dengue-like syndrome listed as primary (17 cases) or contributing (8 cases) cause of death on
their death certificate. The five most common primary causes of death included dengue (29%),
viral syndrome (16%), cardiorespiratory failure (12%), sepsis (9%), and thrombocytopenia (5%).

Clinical Outcomes Among Fatal Laboratory-Positive Dengue Case-

Patients
Most of the 55 laboratory-positive dengue case-patients who died in hospital had signs and
symptoms consistent with dengue (86%) or severe dengue (82%), but only 62% had that clini-
cal diagnosis (Table 4). Of those who met criteria for severe dengue by the time of death, 38 of
45 (84%) had severe plasma leakage, 25 (56%) had severe bleeding, and 22 (49%) had severe

PLOS Neglected Tropical Diseases | DOI:10.1371/journal.pntd.0005025 October 11, 2016 11 / 19

 Enhanced Surveillance for Fatal Dengue

organ impairment. Of note, hemoconcentration was documented in only a few case-patients

even when hematocrits were performed over time. However, 34 (62%) case-patients who died
in hospital had an effusion and most (62%) case-patients with effusions had acute respiratory
failure or ARDS (Table 5).
The majority of case-patients who died in a hospital (86%) were bleeding, and 25 (46%) had
severe bleeding: 19 (35%) gastrointestinal, 14 (26%) pulmonary, six (11%) vaginal, and six
(11%) intracranial (Table 5). Of those with severe bleeding, 14 (56%) received a platelet trans-
fusion while 11 (44%) received a blood transfusion; blood transfusion was more likely to occur
with increased hospital stay (median 5.2 vs. 1.3 days, respectively; P <0.001). Of the 26 case-
patients who received a platelet transfusion, three (12%) had no recorded bleeding. The best
predictor of platelet transfusion was low platelet count [median 13,000 (range: 7,00037,000)
vs. 55,000 (range: 8,000224,000) cells/mm3, recipients versus non-recipients, respectively;
P <0.001].
Other severe clinical outcomes among the 55 laboratory-positive dengue case-patients who
died in hospital included acute hepatitis (76%), acute renal failure (18%), cholecystitis (9%),
acute liver failure (9%), myocarditis (4%), metabolic acidosis (66%), prolonged shock (53%),
and acute respiratory failure (55%) (Table 5). All received intravenous crystalloids, 40%
received a diuretic, and 29% received intravenous colloids. Twenty-five (45%; all adults)
received intravenous corticosteroids. Those given steroids had a lower median platelet count
than those not given steroids (16,000 vs. 47,000 cells/mm3; respectively, P <0.01), and were
three times more likely to have a hospital acquired infection (75% vs. 25%, P <0.05).

Discussion
Enhanced surveillance for dengue deaths showed the majority were not reported to the stan-
dard dengue surveillance system and most did not have dengue coded on the death certifi-
cate. Identification of these unrecognized deaths resulted in a 2 to 3-fold higher dengue
mortality rate than previously reported [14, 17, 20, 21, 48, 49]. EFASS demonstrated the impor-
tance of appropriate diagnostic testing of tissue and serum to make the correct diagnosis in
deaths from a dengue-like acute febrile illness. In addition, EFASS showed its ability to identify
unrecognized deaths from other pathogens of public health importance.
The EFASS estimated age-specific annual dengue mortality rates were comparable to those
from other infectious diseases in the US, including influenza [50, 51]. However, in contrast to
influenza, most dengue deaths occurred among adults 1964 years of age. The estimated aver-
age annual influenza-associated US death rate is 2.4 per 100,000 residents (range: 0.45.1). In
most years, 88% of these deaths are among persons aged 65 years [51, 52]; 17.0 deaths per
100,000 (range: 2.436.7). Influenza death rates among persons <19 years and 1964 years are
0.1 (range: 0.10.3) and 0.4 (range: 0.10.8) per 100,000, respectively. In comparison, EFASS
estimated that dengue mortality in 2010 was 0.42, 1.17, and 1.66 per 100,000 persons aged <19
years, 1964 years and 65 years, respectively.
Most fatal laboratory-positive dengue case-patients appeared to have timely access to
healthcare. However, many (~40%) were sent home after their first ED visit with warning signs
of severe dengue. Although the majority sought care again within 48 hours, two died at home.
Most case-patients who died in a hospital had severe plasma leakage, severe bleeding, or both,
and most received inotropes and half received a platelet transfusion. Although bleeding was
present in the majority who received platelets, half of those with severe bleeding did not receive
red blood cells. A large proportion of case-patients received corticosteroids, which are not con-
sidered of benefit in dengue [2, 53]. As reported by others, we found an increased risk of hospi-
tal-acquired infections in these patients [54, 55].

PLOS Neglected Tropical Diseases | DOI:10.1371/journal.pntd.0005025 October 11, 2016 12 / 19

 Enhanced Surveillance for Fatal Dengue

Table 5. Demographic characteristics, medical history and clinical outcomes for fatal laboratory-positive dengue case-patients detected by the
Enhanced Fatal AFI Surveillance System who died in a hospital, Puerto Rico, 20102012.
Adults 20 years old
All cases (n = 55) Children and adolescents Dengue only Co-infections* (n = 5)
(n = 6) (n = 44)
Female, no. (%) 32 (58.2) 5 (83.3) 27 (61.4) 0 (0.0)
Obese, no. (%) 24 (43.6) 1 (16.7) 19 (43.2) 4 (80.0)
No chronic disease, no. (%) 10 (18.2) 3 (50.0) 7 (15.9) 0 (0.0)
One chronic disease, no. (%) 12 (21.8) 2 (33.3) 9 (20.5) 1 (20.0)
More than one chronic disease, no. (%) 33 (60.0) 1 (16.7) 28 (63.6) 4 (80.0)
Clinical outcomes
Days post-illness onset at admission, median 4.5 (0.513.0) 4.6 (0.58.9) 4.5 (0.513.0) 4.7 (3.67.2)
(range)
Length of hospital stay, median (range) 2.7 (1.129.2) 8.6 (8.68.6) 2.7 (1.129.2) 2.3 (2.32.3)
Admitted to ICU, no. (%) 37 (67.3) 4 (66.7) 29 (65.9) 4 (80.0)
ICU length of stay, median (range) 1.1 (0.125.8) 2.5 (0.44.9) 1.1 (0.125.8) 1.1 (0.75.2)
Clinical laboratory findings
Leukopenia, no. (%) 29 (52.7) 3 (50.0) 24 (54.6) 2 (40.0)
Thrombocytopenia, no. (%) 46 (83.6) 4 (66.7) 37 (84.1) 5 (100.0)
Hematocrit increase by  20%, no. (%) 7 (12.7) 0 (0.0) 7 (15.9) 0 (0.0)
Hemoconcentration by age, no. (%) 1 (1.8) 0 (0.0) 1 (2.3) 0 (0.0)
Days post onset of max HCT, median (range) 4 (013) 4 (09) 4 (113) 4 (37)
Effusion, no. (%)
Any effusion 34 (61.8) 6 (100.0) 26 (59.1) 2 (40.0)
Pleural effusion 24 (43.6) 4 (66.7) 19 (43.2) 1 (20.0)
Ascites 14 (25.5) 4 (66.7) 10 (22.7) 0 (0.0)
Pericardial effusion 9 (16.4) 1 (16.7) 7 (15.9) 1 (20.0)
Effusion with respiratory failure or ARDS 21 (38.2) 4 (66.7) 16 (36.4) 1 (20.0)
Bleeding Manifestation, no. (%)
Any bleeding** 47 (85.5) 6 (100.0) 38 (86.4) 3 (60.0)
Severe bleed 25 (45.5) 4 (66.7) 18 (40.9) 3 (60.0)
Other Clinical Features, no. (%)
Acute hepatitis 42 (76.4) 5 (83.3) 32 (72.7) 5 (100.0)
Acute liver failure 5 (9.1) 2 (33.3) 3 (6.8) 0 (0.0)
Cholecystitis 5 (9.1) 1 (16.7) 3 (6.8) 1 (20.0)
Myocarditis 2 (3.6) 0 (0.0) 2 (4.6) 0 (0.0)
Acute renal failure 10 (18.2) 1(16.7) 7 (15.9) 2 (40.0)
Medical complications, no. (%)
Prolonged shock 29 (52.7) 2 (33.3) 23 (52.3) 4 (80.0)
Metabolic acidosis 36 (65.5) 5 (83.3) 27 (61.4) 4 (80.0)
Fluid overload 17 (30.9) 4 (66.7) 11 (25.0) 2 (40.0)
Abdominal compartment syndrome 2 (3.6) 2 (33.3) 0 (0.0) 0 (0.0)
Acute respiratory failure 30 (54.6) 5 (83.3) 21 (47.7) 4 (80.0)
Acute respiratory distress syndrome 11 (20.0) 4 (66.7) 5 (11.4) 2 (40.0)
Coma 10 (18.2) 2 (33.3) 8 (18.2) 0 (0.0)
Seizure 8 (14.6) 1 (16.7) 7 (15.9) 0 (0.0
Hospital acquired infection 16 (29.1) 2 (33.3) 11 (25.0) 3 (60.0)
Disseminated intravascular coagulation 5 (9.1) 2 (33.3) 3 (6.8) 0 (0.0)
Treatment given, no. (%)
Intravenous colloid 16 (29.1) 3 (50.0) 11 (25.0) 2 (40.0)
(Continued)

PLOS Neglected Tropical Diseases | DOI:10.1371/journal.pntd.0005025 October 11, 2016 13 / 19

 Enhanced Surveillance for Fatal Dengue

Table 5. (Continued)

Adults 20 years old

All cases (n = 55) Children and adolescents Dengue only Co-infections* (n = 5)
(n = 6) (n = 44)
Blood transfusion 13 (23.6) 2 (33.3) 10 (22.7) 1 (20.0)
Platelet transfusion 26 (47.3) 2 (33.3) 22 (50.0) 2 (40.0)
Fresh frozen plasma 12 (21.8) 2 (3.33) 9 (20.5) 1 (20.0)
Inotropes 38 (69.1) 4 (66.7) 30 (68.2) 4 (80.0)
Diuretics 22 (40.0) 3 (50.0) 16 (36.4) 3 (60.0)
Corticosteroid 25 (45.5) 0 (0.0) 22 (50.0) 3 (60.0)

* Co-infections include: Four DENV/Leptospira spp. bacteria, and one DENV/Streptococcus pneumonia.

Number and percent presented unless otherwise stated.

Most (44, 80%) of the 55 case-patients who died in hospital had a chest x-ray and/or an ultrasound done. Several case-patients (30, 55%) had at least one
other imaging study done including an echocardiogram (17 done) and/or a computed tomography (CT) scan (2 abdominal and 23 brain CT scans done).
** Any bleeding was defined by the presence of any of the following: petechiae, purpura, ecchymosis, epistaxis, gingival bleeding, hematuria, menorrhagia,
hemoptysis, hematemesis, melena, or an intracranial bleed.

doi:10.1371/journal.pntd.0005025.t005

Dengue deaths often occur among patients with comorbidities [14, 19]. Nearly half of case-
patients were obese and over half had more than one chronic medical condition; prevalences
similar to those found in the Puerto Rican adult population [47, 56], with the exception of dia-
betes and asthma. The prevalence of diabetes in case-patients was nearly four times that of the
adult population, and asthma was twice as prevalent. Adult diabetics have been over-repre-
sented in other fatal case series [19, 57], and a recent meta-analysis found diabetes was associ-
ated with increased risk of severe dengue [58]. As many endemic areas have reported a
substantial proportion of dengue cases in adults, healthcare providers should be attentive to
dengue patients with these comorbidities [2].
Some patients developed acute liver or renal failure or had atypical presentations [19, 57, 59,
60]. Acute renal failure (ARF) affected ~20% of case-patients though none had pre-existing
renal disease and 80% were non-elderly (median age 49 years). However, dengue patients with
severe dengue, diabetes or secondary infections are known to be at risk for developing acute
kidney injury [61]. Six of the ten ARF cases had at least one risk factor and two were co-
infected with Leptospira spp. One of the four ARF case-patients without risk factors was an
infant with abdominal compartment syndrome and multiple organ dysfunction.
While more sensitive than PDSS, EFASS may not have detected all fatal laboratory-positive
dengue cases. For example, a few rural municipalities had fewer deaths than expected. In the
case of Patillas, this may have been due to higher dengue case-reporting to PDSS because of an
enhanced dengue surveillance project conducted prior to EFASS [62]. Alternatively, individu-
als in rural municipalities who died at home and were not known to have an AFI would not
have been identified. These factors may have led to lower case ascertainment and estimated
dengue mortality. Although we increased the proportion of suspect cases with an etiologic
diagnosis by obtaining tissue and convalescent serum specimens, about one quarter of cases
were dengue laboratory-indeterminate, and were not counted as fatal laboratory-positive den-
gue cases even if dengue was listed on their death certificate. Hence, our final dengue mortality
estimate should be considered conservative.
EFASS demonstrated the feasibility and importance of enhanced surveillance for dengue
deaths, and found a previously unrecognized high dengue mortality in Puerto Rico that was
higher than rates observed in other dengue endemic regions during this time period [914].

PLOS Neglected Tropical Diseases | DOI:10.1371/journal.pntd.0005025 October 11, 2016 14 / 19

 Enhanced Surveillance for Fatal Dengue

Establishment of EFASS-like systems in selected dengue endemic countries would go a long

way towards obtaining robust estimates of the global burden of deaths due to dengue, and iden-
tify areas for improvement in clinical care of patients with severe dengue.

Supporting Information
S1 Appendix. ICD-9 codes used to review death certificates and medical records to identify
individuals that died following a dengue-like acute febrile illness.
(PDF)
S2 Appendix. Surgical Pathology and Autopsy Report (SPAR) Form used by forensic
pathologists to document autopsy findings from individuals that died following a dengue-
like acute febrile illness.
(PDF)
S1 Checklist. STROBE Checklist.
(DOCX)

Acknowledgments
We thank the staff members at the Demographic Registry of Puerto Rico for facilitating the
review of death certificates including Zulma L. Escalera, Ivonne Vasallo, Wanda Llovet, Alejita
Santos, and Nancy Vega. In addition, we would like to acknowledge the medical management
information offices from hospitals in Puerto Rico for working with us to access medical records
from the deceased case-patients. Last, we would like to acknowledge the support of Dr. Mara
Conte, CEO of the PRIFS during the planning phases of our project, Daniel Lpez, Manager at
PRIFS, and all the pathologists at PRIFS and Centro Mdico. Without their interest in and sup-
port of this project, EFASS would never have been possible.

Author Contributions
Conceptualization: KMT AR BTV EAH JLMJ IR DS DMB RG JT RR JS CC FD GC CD ER.
Data curation: KMT AR BTV EAH JLMJ IR DS DMB RG JT RR JS CC FD GC SRZ LW CD
ER.
Formal analysis: KMT AR BTV EAH JLMJ TMS JPP EME LW HSM.
Funding acquisition: KMT HSM.
Investigation: KMT AR BTV EAH JLMJ IR DS DMB RG JT RR JS CC FD CD ER TMS JPP
EME GC LW SRZ.
Methodology: KMT AR BTV EAH JLMJ IR DS DMB RG JT RR JS CC FD LW SRZ GC CD
ER.
Project administration: KMT EAH JLMJ GC SRZ CD ER HSM.
Resources: KMT AR BTV EAH JLMJ TMS IR DS DMB RG JT RR JS CC FD JPP EME GC
SRZ LW CD ER HSM.
Software: KMT AR BTV LW.
Supervision: KMT GC EAH JLMJ SRZ CD ER HSM.

PLOS Neglected Tropical Diseases | DOI:10.1371/journal.pntd.0005025 October 11, 2016 15 / 19

 Enhanced Surveillance for Fatal Dengue

Validation: KMT AR BTV EAH JLMJ TMS IR DS DMB RG JT RR JS CC FD JPP EME GC

LW CD ER HSM.
Visualization: KMT AR BTV TMS LW HSM.
Writing original draft: KMT AR BTV TMS LW HSM.
Writing review & editing: KMT AR BTV EAH JLMJ TMS IR DS DMB RG JT RR JS CC FD
JPP EME GC LW SRZ CD ER HSM.

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