Dengue in the United States of America: A Worsening Scenario?

Dengue is a febrile illness caused by any of the four dengue virus types (DENV-1 to -4, genus Flavivirus, family Flaviviridae) mainly transmitted by the mosquito Aedes aegypti. DENV can be transmitted by blood transfusion. Dengue has been historically present in the continental United States (US), in the state of Hawaii, and in the US insular territories in the Caribbean and the Pacific. During the second half of the 20th century, most of the cases reported in the US were imported cases brought to the country by travelers. Since 2009, cases of autochthonous dengue have been recognized in the state of Florida after 75 years of absence, followed by intensification of transmission in endemic places including the US territories of US Virgin Islands and Puerto Rico, which experienced a large dengue epidemic in 2010. The widespread distribution of dengue mosquito vectors, deficient mosquito control measures and increased frequency of DENV-infected visitors to the US coming from dengue-endemic locations or places experiencing epidemics appear to be jointly responsible for the emergence and reemergence of dengue in the US and its territories.


Introduction
Dengue, the most prevalent arthropod-borne viral disease in the world, is an acute, febrile disease caused by any of the four dengue virus types (DENV-1 to -4, genus Flavivirus, family Flaviviridae) [1]. DENV is naturally transmitted by mosquitoes from the genus Aedes, mainly by the urban species Aedes aegypti, and in some geographical regions by Aedes albopictus. Infection by dengue viruses can be asymptomatic or cause disease of variable degree of severity. Dengue ranges from a mild, influenza-like illness known as dengue fever (DF) to a severe and potentially life-threatening condition called dengue hemorrhagic fever (DHF). DHF can ultimately evolve to hypovolemic shock (dengue shock syndrome (DSS)) and death [2]. DHF and DSS are classified as severe dengue in the newest World Health Organization (WHO) dengue clinical classification [3].
DENV can also be transmitted by transfusion of blood and blood components and by solid organ transplants containing infectious virus [4][5][6]. Dengue is endemic in most countries of tropical America, the Caribbean and Southeast Asia, and causes episodic epidemics in islands of the Pacific and in Africa [7,8]. Many DENV endemic regions are hyperendemic, which is defined by the circulation of all DENV types in a specific geographical area. This may increase the opportunity for occurrence of secondary infections with increased clinical severity via antibody-dependent enhancement of the infection by sub-neutralizing concentrations of anti-dengue antibodies in individuals previously infected with a different dengue type (heterologous infection) [9].
Approximately, 50-100 million cases of dengue were calculated to occur each year around the globe [3]. However, the newest estimates raise these numbers to about 390 million cases per year (range 284-528), from which approximately 96 (range 67-136) will present clinical manifestations of any severity [10]. Despite its low lethality rates, dengue causes severe economic and social disruption and has profound impact on the welfare of regions affected by the disease since it is endemic mostly in developing countries where healthcare systems have limited resources [11][12][13]. After many years of continuous efforts, there are still neither effective vaccines nor specific antiviral treatments available against DENV [14].
The mosquitoes that serve as DENV vectors have been able to reach sub-tropical and temperate regions, including North America and countries in Europe [15][16][17]. In addition, DENV brought in by infected travelers has been able to establish autochthonous infection cycles in some of these countries. DENV is considered the most common cause of 2 BioMed Research International febrile illness in travelers returning to the USA from destinations in the Americas and Asia [18].
The scope of this review is to provide an analysis of the epidemiology of dengue in the United States and its territories, with emphasis on the changes in dengue activity in the last decade and on aspects on the molecular epidemiology of currently circulating DENV.

Dengue in the United States
Dengue is thought to have been present in the USA since the end of the 18th century, when Dr. Benjamin Rush, a physician and signatory of the US Declaration of Independence, described a disease resembling dengue fever in Philadelphia during 1780 [27]. During the first half of the 20th century, a number of dengue outbreaks were reported in the continental USA, especially in the gulf and southeastern states (i.e., Alabama, Florida, Georgia, Louisiana, Mississippi and Texas), in the state of Hawaii, and in the USA Territories in the Caribbean and the Pacific Ocean (Table 1).
The continental USA comprises the 48 contiguous states and the District of Columbia, and the non-contiguous states of Alaska in North America and Hawaii in the Pacific Ocean and a number of unincorporated territories including Puerto Rico and the US Virgin Islands (USVI) in the Caribbean and American Samoa, Guam, and the Northern Mariana Islands in the Pacific Ocean [19].
Autochthonous dengue has been reported sporadically in the Mexico-Texas border, where indigenous cases of dengue reappeared in 1980 after more than 30 years of absence. Since then, most dengue cases reported in the USA have been imported, brought by infected travelers returning from visits to endemic countries or places experiencing dengue epidemics. Dengue has also been present in the state of Hawaii, the US territories in the Pacific (American Samoa, Guam, and Northern Mariana Islands) and the territories of Puerto Rico and USVI. In 2009, after over 70 years of absence, autochthonous dengue reappeared in the state of Florida ( Figure 1).
In all those instances, dengue outbreaks were facilitated by the presence of the mosquito vector, the favorable climatic conditions for its subsistence, and the presence of susceptible individuals. The dengue mosquito vectors Aedes aegypti and Aedes albopictus have been reported to be present in several counties in the USA, especially in the Southern and Southeastern regions of the country [28] (Figure 2).

Dengue in the Mexico-Texas
Border. The first autochthonous case of dengue reported in the USA since 1945 occurred in Brownsville, Texas in 1980 when DENV-1 was isolated from a 5-year-old girl that did not have history of travel outside Brownsville (Table 1). Brownsville is located across the Rio Grande from the city of Matamoros in the state of Tamaulipas, Mexico where many years of dengue activity had been observed [21].
Subsequently, surveillance studies reported 63 additional dengue cases (all caused by DENV-1) that were laboratory confirmed in Texas, 52 of which occurred in counties contiguous to the Mexico-Texas border [30]. Six years later, autochthonous transmission of DENV-1 was again reported in at least 9 individuals in Texas [22]. The Mexico-Texas border region is at risk for dengue endemicity due to the presence of the competent mosquito vectors Aedes aegypti and Aedes albopictus and to the circulation of all DENV types (DENV-1 to -4) in recent years [31]. In sum, dengue has been detected in residents of seven Texas counties: Bee, Cameron (where Brownsville is located), Hidalgo, Maverick, Nueces, Travis, and Webb [23,30,32].
In 2005, a woman who had not traveled to Mexico in the months before the onset of dengue symptoms developed DHF, and this was the first case of autochthonous DHF reported in Texas. During that year, a large dengue epidemic developed in the neighboring Mexican state of Tamaulipas with more than 1,200 dengue cases of which 223 (18%) were classified as DHF. Meanwhile, 25 dengue cases were reported in Brownsville, of which at least three were locally acquired [32]. The disparities in the number of cases observed on different sides of the border have been explained by the higher standard of living in the USA including the use of air-conditioners and the presence of nets in doors and windows, which remain closed most of the time reducing the density of indoor mosquitoes thus minimizing the risk of mosquito bites [33].

Dengue in Florida.
After 75 years without reports of dengue activity in Florida, a case was identified in 2009 in a New York patient who had traveled to Key West (Table 1). This individual had not traveled to any other place either in the USA or abroad before the onset of symptoms, and the case was laboratory-confirmed by the Centers for Disease Control and Prevention (CDC) using RT-PCR. A serosurvey conducted in 2009 revealed that at least 5.4% of the residents of Key West had serological evidence of recent dengue infection [24].
DENV-1 was detected in mosquito pools collected in Key West [49] and in a blood donor from Key West during 2010 [50]. The strains isolated from these two specimens were sequenced, and subsequently, a number of DENV-1 strains obtained from samples from dengue cases from Key West/ Monroe county (8 patients) and one strain each from patients from Broward, Miami-Dade, Orange, and Pinellas counties were sequenced [51]. All DENV-1 strains from Florida from 2009-2010 sequenced to date (obtained from mosquito pools, dengue cases, and a blood donor) belong to the genotype V of the virus and have phylogenetic relationships with Central American strains; however, all but one of the strains sequenced from Key West grouped together in the phylogenetic trees, suggesting that in situ microevolution has occurred and that this Key West sublineage was transmitted Dengue suspected in Philadelphia, PA [19] 1826 USA ports report 1st dengue outbreak [20] 1827-1828 Epidemic in Southern USA [20] 1845 Dengue reported in St. Louis, MO [20] 1850-1851 1st report of dengue epidemic inland (including GA and MS), epidemic in Southern USA, New Orleans, LA, and along South Coast [20] 1870-1872 Epidemic in Southern USA [20] 1873 Dengue reported in LA, AL, and MS; ≈40,000 cases reported in New Orleans [20] 1879-1880 Epidemic in Southern USA [20] 1885-1886 Dengue in gulf ports of TX, dengue in Austin, 16,000 estimated cases of 22,000 inhabitants [20] 1897-1903 Epidemic in Southern USA, TX most heavily affected [20] 1904 Dengue reported in FL and TX [20] 1916 Fatal case of possible DHF reported in TX [20] 1922 Dengue epidemic, 500,000 to 600,000 cases in TX, 30,000 in Galveston, and 7,561 in LA [20] 1923 1,376 dengue infections reported in LA [20] 1924 1 dengue case in LA [20] 1941-1944 Texas and gulf states involved in epidemic [20] 1945 Last continental epidemic of dengue reported in LA [20] 1980 1st indigenous dengue cases in USA since 1945 (Brownsville, TX), DENV-1 isolated [21] 1981 DENV-4 cases reported, 1st isolation of DENV-4 in the USA [20] 1982 1st reports of DENV-2 in the USA [20] 1983 1st reports of DENV-3 in the USA [20] 1986 DENV-1 reported in TX [22] 1987 Autochthonous dengue reported in TX [20] 1990 DENV-1, -2, and -3 isolated in the USA, reports of 102 dengue cases [20] 1991 DENV-1 and DENV-3 isolated in the USA, 25 dengue cases reported [20] 1994 91 cases of dengue, DENV-2 and -3 isolated in the USA [20] 2005 Adapted from [20]. Mosquitoes from both Aedes species obtained from colonies in the state of Florida were experimentally infected with a DENV-1 strain isolated from Key West, and vector competence for this virus was assessed. There were no significant differences in the rates of infection, dissemination, and transmission between these two mosquito species occurring in Florida. The entire genome of the DENV-1 specimen utilized in this vector competence study has been sequenced [16] and shown to cluster within the Key West lineage ( Figure 3).
In addition to the autochthonous cases, an important number of dengue imported cases (392) from travelers to countries in Central and South America, the Caribbean, Africa, the Middle East, and South and Southeast Asia have  (2)  Year(s) Activity reported References

Late 1840s
First large dengue epidemic recorded in HI, associated with Aedes aegypti [34] 1903 Large dengue epidemic in HI, ≈30,000 cases, associated with Aedes aegypti [35] 1943-1944 DENV-1 epidemic in HI, ≈1,500 cases [34,36] 1944 Aedes aegypti reported to be eradicated from GU. Aedes albopictus reported to be present in the island [37] <1950 Dengue cases were reported in AS and GU before 1950, no dengue epidemics reported in GU in recent times [38] 1995 Possible dengue infection in German visitors to HI [39] 1972 Dengue epidemic in AS (DENV-2) [40] 1975 Dengue epidemic in AS (DENV-1) [38,40] 1995-1996 Dengue epidemic in AS (DENV-3) [38] 1997 Reports of dengue cases in AS [41] 1998 Dengue seropositive individuals reported in Saipan (MP) during 1998, DENV-2 implicated in epidemic activity [42,43] 2001 More than 1,600 dengue cases reported in AS (DENV-1), 3 deaths [44] Dengue outbreak in the MP, >1,400 cases reported [45] 2001-2002 Autochthonous transmission of dengue in HI, 122 confirmed cases, DENV-1 isolated, and Aedes albopictus was the implicated vector [46] 2007 63 dengue cases confirmed in AS, 23 cases hospitalized [47] 2008 Dengue activity reported in AS [48] 2009 Outbreak of ≈400 confirmed cases in AS [48] 2010 Dengue cases reported in AS. Serosurvey conducted in 2010 revealed >95% of the tested individuals as seropositive for dengue [8,48] AS  Table 2). The wide geographical area of origin of dengue imported cases in Florida reflects the importance of the Miami International Airport as a gateway to the USA and a possible route of entry for these and other pathogens.

Dengue in Hawaii.
The first large dengue epidemic on record occurred in Hawaii in the mid-19th century. Another large epidemic occurred in 1903 with approximately 30,000 cases reported [34,35]. Aedes aegypti was the vector implicated in those dengue outbreaks. But at a certain point during the early 20th century, Aedes albopictus was introduced in these islands and displaced Aedes aegypti [52].
A DENV-1 epidemic occurred in Hawaii from 1943 to 1944, with almost 1,500 cases reported [36,53]. As in the continental USA no autochthonous cases of dengue were reported after 1945 and imported dengue cases were reported at a low frequency in Hawaii [54]. In 1995, two German travelers developed symptoms compatible with dengue infection after a trip to Hawaii [46] (Table 3).
However, in 2001, following 56 years without reports of autochthonous dengue cases, an epidemic occurred in the island of Maui. A total of 122 laboratory-confirmed dengue cases were reported between 2001 and 2002, 92 of which occurred in Maui, 26 in Oahu, and 4 in Kauai. DENV-1 was the type identified in viral isolates obtained from 15 cases from Hawaii, and the mosquito vector implicated in this epidemic was Aedes albopictus [54].
Molecular epidemiologic studies suggest that at least two distinct DENV-1 strains were introduced in Hawaii during  [28,29]. The delimited area represents the approximate geographical area in which either dengue mosquito vector (Aedes aegypti and/or Aedes albopictus) have been found present in the USA and are therefore considered to be at risk for the establishment of dengue outbreaks. The noncontiguous states of Alaska and Hawaii are not shown at scale. U.S. territories are not shown. the 2001-2002 epidemics, although most of the isolates analyzed belong to the "Pacific subtype" (genotype IV) of DENV-1, and cluster together with Tahitian DENV-1 strains, suggesting that Tahiti was the source of these strains. In contrast, the same study reported a single DENV-1 isolate from Hawaii obtained during 2001 from a traveler to Samoa that clustered outside the Tahitian cluster and that closely associated with a DENV-1 isolate previously obtained from another traveler to Samoa [39] (Figure 3).

Dengue in American Samoa, Guam and Northern Mariana Islands. Dengue was reported in American Samoa and
Guam before 1950 [38] (Table 3). The main vector, Aedes aegypti, had been considered as eradicated from Guam since 1944. However, Aedes albopictus is abundantly present in the island [37]. Guam has not had dengue outbreaks in recent times, but it is at risk of dengue epidemics due to the presence of Aedes albopictus mosquitoes and the arrival of infected travelers from neighboring islands experiencing dengue activity.
In 2010, less than 100 dengue cases were reported for an incidence of 77/100,000 inhabitants [8,48]. Epidemiological surveillance performed in 794 serum samples collected in three islands of American Samoa in 2010 revealed that 759 (95.6%) of the tested individuals were positive for IgG antibodies to dengue [44]. The dengue vectors, Aedes aegypti and Aedes albopictus, are present in American Samoa [73].
There are reports indicating that individuals from the Northern Mariana Islands, without travel history, were seropositive for DENV (IgM and IgG) in 1998 in the island of Saipan [43], where DENV-2 was the DENV type implicated in those infections [42]. Furthermore, these islands suffered a dengue outbreak in 2001 with more than 1,400 cases reported [45]. Since then, no other dengue cases have been reported.

Dengue in US Virgin
Islands. Dengue has been endemic in the USVI since at least 1924, when the first documented dengue epidemic in the Caribbean is thought to have started in the USVI [20]. These islands are located in close proximity to Puerto Rico, and include the four inhabited islands of St. Croix, St. Thomas, St. John and Water Island. Dengue outbreaks occurred in 1978 (DENV-1) and in 1990, this time involving DENV-1, DENV-2 and DENV- 4; 1990 was the year of the first occurrence of DENV-4 in the USVI [20]. The sequence of a DENV-4 strain isolated in 1994 from St. Croix closely associated with strains that were circulating in Puerto Rico around the same time (Figure 4).
Forty dengue cases were reported in the island of St. Thomas in 2004. In 2005, a DENV-2 epidemic was reported in St. Croix, with 331 suspected cases of which 37% were laboratory-confirmed [65]. The sequence of a DENV-2 strain from the 2005 epidemic clustered within the clade IA of the American/Asian genotype, together with a number of strains from the Caribbean and South America [74].
During November 2012, 27 dengue cases were reported in the island of St. Croix, some of which were later laboratory confirmed. A serosurvey found that around 20% of the students and staff from a school in the island were positive for IgM antibodies to dengue; four students were positive for DENV-1 or DENV-4 RNA by PCR [69]. There is no available genetic information about the DENV strains circulating in St. Croix in 2012.

Dengue in Puerto Rico. Dengue is endemic in Puerto
Rico, and cases have been reported every year for more than 50 years (Table 4). The public health definition of a dengue epidemic in Puerto Rico is based on a historical average: the event is called an epidemic when the number of cases detected by the passive surveillance system is higher than the 75th percentile of the distribution of cases for the same epidemiological week in previous years [75]. A large epidemic of DENV-3 involving approximately 27,000 individuals was reported in 1963-1964 [55,56]. Another epidemic caused by DENV-2 occurred in 1969 [57]. Subsequently, sporadic dengue cases caused by DENV-2 were reported during the early 1970s [57]. In 1975, a case of DHF (associated with DENV-2) was reported for the first time in Puerto Rico and in the Western Hemisphere [20]. The outbreak in 1977 was caused by DENV-2, DENV-3 and DENV-1 (DENV types appear in order of frequency of detection), and that was the first documented occurrence of DENV-1 in the island [58].
In 1999, all four DENV types co-circulated in the island resulting in 4,993 reported cases. The 2000 epidemic had cocirculation of DENV-1, DENV-2, and DENV-3 and resulted in 2,433 dengue cases reported [20]. DENV-3 was the predominant type in the inter-epidemic period between 1999 and 2003, to be later displaced by DENV-2 as the predominantly DENV type detected during the inter-epidemic period spanning from 2004 to 2006 [66].
In 2007, a large epidemic (10,508 suspected dengue cases, with 227 fulfilling criteria for DHF) occurred after an interepidemic period of almost ten years; this epidemic was the second outbreak to have co-circulation of all DENV types, after the 1998 epidemic. During the 2007 dengue epidemic, 2,175 individuals tested positive for DENV RNA, of which 62% was infected with DENV-3, 31% with DENV-2, 6% with DENV-1, and 1% with DENV-4. DENV-1, and DENV-4 reappeared after approximately 9 years of absence (Table 4). Overall, an increased incidence of severe disease was reported in the 2007 epidemic when compared with previous outbreaks, resulting either from more efficient reporting of severe cases or from a true increase in the incidence of severe dengue [66].
The largest epidemic of dengue in Puerto Rico's history occurred during 2010, with almost 27,000 suspected dengue cases, of which more than 12,000 individuals (about 47% of tested cases) were laboratory-positive; more than 1,300 cases were classified as severe dengue, and there were 40 dengueassociated deaths ( Table 4). The DENV types implicated in this epidemic were in order of frequency DENV-1 (69%), DENV-4 (23.7%), and DENV-2 (7.3%), with only two DENV-3 cases reported (<0.1%). The 2010 epidemic is considered the longest-lasting dengue outbreak ever registered in Puerto Rico, with cases starting to appear during the first week of 2010, peaking around August and returning to levels below the historical average in December of that year [67].

Epidemiological Data Obtained from Asymptomatic
Infections in Blood Donors. Because DENV can be transmitted by blood transfusion [4,6] and infection can be asymptomatic [1][2][3], there is great concern about DENV activity among blood donors in endemic places in the absence of overt epidemic and during epidemic times. Prevalence studies in blood donors have been performed in various areas around the world where DENV is endemic [76].
A study performed in asymptomatic blood donors from Puerto Rico during the inter-epidemic year 2005 reported that 0.7% (12 out of 16,521) blood donations tested positive for DENV RNA. Of these, three donors were identified by Taq-Man as infected with DENV-2 and one with DENV-3, which were the DENV types circulating in the island during that year (Figure 1). Two of the DENV-2 and the DENV-3 strains were isolated in cell culture or by mosquito inoculation [4]. Another study conducted among blood donors during the epidemic year of 2007 found that 29 out of 15,350 donations tested positive for DENV RNA using an assay that detects all four DENV types but does not discriminate between DENV types. Discriminatory real-time PCR detected 12 positive samples: one was identified as DENV-1, four as DENV-2, and seven as DENV-3 ( Figure 1). All these 12 samples were infectious in C6/36 cell cultures [6].
Study of DENV strains circulating in 2010 performed in blood donors [50] and in symptomatic cases [67] showed that these DENV-1 strains belong to the genotype V (the only one found to circulate in the Americas to date) but were from a lineage different from those that circulated in Puerto Rico during and before 1998 ( Figure 3). Likewise, the sequenced DENV-4 strains from 2010 were found to belong to the genotype II (also the only one circulating in the Americas) and are distinct from those that circulated in Puerto Rico in the 1990s, although with less genetic variation than that observed in the newest DENV-1 strains circulating in the island (Figure 4). The results suggest that overall, a clade replacement for DENV-1 and DENV-4 may have occurred at some point in Puerto Rico during the inter-epidemic period between 1999 and 2006 [50,67].

Conclusions
Dengue has emerged and re-emerged in many locations around the world, including countries in Europe (e.g., France and Croatia) [77,78] and in North America [24] during the last two decades. Dengue has reemerged and caused epidemics in the continental USA for the first time after several decades of absence, and a worrisome panorama is expected if the trend of transmission continues. Several articles expressing these concerns have been published before and after the re-emergence of dengue in Florida in 2009 [79,80]. Such concern is augmented by evolving climatic and ecological conditions that favor vector sustainability and by high travel activity with subsequent importation of cases. In fact, cases of dengue in returning travelers to the USA have been on the rise [81], and cases of dengue in countries in Central America, South America, and the Caribbean, very popular destinations for American tourists, have also been on the rise in the past decades [82].
Although the primary mosquito vector for dengue is Aedes aegypti, a highly domesticated urban mosquito, the virus can be also transmitted by Aedes albopictus, albeit in a less efficient manner. Aedes albopictus is probably responsible for the maintenance of dengue in rural/sylvatic cycles in endemic countries [83], and future research should address the biological and epidemiological implications of the displacement and replacement of Aedes species in the context of dengue epidemics.
In the case of the USA, both dengue vector species are widely distributed in the southern parts of the country (Figure 2), and one or another dengue vector is present in all US insular territories [28,29]. Travel, especially by air, has been considered an important risk factor for the rapid dissemination of pathogens and their vectors in an efficient and rapid manner [84][85][86][87]. New promising vector control strategies based on the release of Wolbachia-infected Aedes aegypti mosquitoes have been tested in regions of Australia with potential for the occurrence of dengue epidemics and if deemed suitable, this approach could be utilized in endemic regions from Asia and the Americas [88].
Many large cities in the US are important hubs for air travel and therefore receive a high number of individuals potentially infected with pathogens that cause asymptomatic disease, including several arboviruses (e.g., dengue viruses, Japanese Encephalitis virus and Chikungunya virus). Thus, there is an increasing risk of introduction of these "exotic" pathogens to urban conglomerates where mosquito vectors are present or have the potential to become established (e.g., Miami, Atlanta, Baltimore/Washington, D.C., and New York City in the East Coast, and Los Angeles and San Francisco in the West Coast).
Despite the current economic and budgetary constraints, strict mosquito control policies and activities, that may include both traditional and biological vector control strategies, must be implemented and maintained in localities that have the potential to become the port of entry for these viruses and become the focus for another dengue epidemic in the USA.