**3. Dengue fever and yellow fever**

There are many similarities between dengue fever and yellow fever:


In the case of dengue fever its annual incidence has increased dramatically around the world in recent decades. It is estimated that over 2500 millions people who live in over 100 tropical and non-tropical countries, are currently at risk from dengue viruses globally. The rise in dengue incidence has been marked by geographic expansion of the virus and the vectors due to globalization, habitat modifications, lack of effective mosquito control programs and climate change. Although the major disease burden occurs in South East Asia, the Americas and the western Pacific, dengue was also a common disease in Europe in the past centuries. Large epidemics of dengue and yellow fever occurred in European ports of Spain, Portugal, France, Italy and even Wales and Ireland as the more northern countries of the continent (Eager, 1902; Monath, 2006). Last dengue epidemic in Europe, estimated at one million cases, occurred in Greece in 1927-28 (Papaevangelou & Halstead, 1977; Rosen, 1986).

Dengue is the most frequent tropical arboviruses imported in Europe and together with schistosomiasis both are considered, after malaria, the most important tropical diseases in

The temporal distribution analysis of imported malaria cases indicates that high-risk months for disease transmission (between July and September) also coincides with the period of the most cases reported in Europe. Therefore most of cases occur during the epoch theoretically favorable for malaria transmission. In regard to the diagnostic delay, i.e. the average time between appearance of symptoms and malaria diagnosis (when therapy began), it shows disparate values according to each country. For example, in Eastern Spain the diagnostic delay of imported malaria was estimated in 13.7 days (Bueno Marí & Jiménez Peydró, 2012), while in other European countries like Sweden, France or Italy values are clearly lower, ranging from 3 to 8.2 days (Romi et al., 2001; Askling et al., 2005; Chalumeau et al., 2006). From an epidemiological point of view it is very important to reduce the diagnostic delay, because this is the period when malaria patients could be a source of infection for *Anopheles* females. Additionally, from an exclusively clinical perspective, delay to diagnosis leads of

Both are viruses of the genus Flavivirus (family Flaviviridae) and are strictly

 In their original habitat, both are zoonotic infections transmitted by forest mosquitoes. Their importance as human pathogens can be related with two forest mosquitoes characterized by high ecological plasticity that have become closely associated with the

Both diseases have a history of transmission in temperate regions, including Europe,

Transovarian transmission in female mosquitoes has been demonstrated for both

The viruses and their urban vectors have a worldwide distribution due to

 Both arboviruses are characterized by short incubation period and can provoke similar clinical symptoms, including hemorrhagic illness in humans, often with fatal consequences. However mortality rate is higher in yellow fever (20%) than in dengue

In the case of dengue fever its annual incidence has increased dramatically around the world in recent decades. It is estimated that over 2500 millions people who live in over 100 tropical and non-tropical countries, are currently at risk from dengue viruses globally. The rise in dengue incidence has been marked by geographic expansion of the virus and the vectors due to globalization, habitat modifications, lack of effective mosquito control programs and climate change. Although the major disease burden occurs in South East Asia, the Americas and the western Pacific, dengue was also a common disease in Europe in the past centuries. Large epidemics of dengue and yellow fever occurred in European ports of Spain, Portugal, France, Italy and even Wales and Ireland as the more northern countries of the continent (Eager, 1902; Monath, 2006). Last dengue epidemic in Europe, estimated at one million cases, occurred in Greece in 1927-28 (Papaevangelou & Halstead,

Dengue is the most frequent tropical arboviruses imported in Europe and together with schistosomiasis both are considered, after malaria, the most important tropical diseases in

course to high parasitemia, which itself leads to severe forms of malaria.

There are many similarities between dengue fever and yellow fever:

primatophilic, infecting only primates, including man.

and share essentially the same selvatic and urban vectors.

**3. Dengue fever and yellow fever** 

peridomestic environment.

transportation of goods and people.

viruses.

(5%).

1977; Rosen, 1986).

quantitative terms in Old continent. Of the hundreds of dengue imported cases reported yearly en Europe (Table 4), the vast majority are represented by tourists (about 84%). A difference to what happens with malaria, immigration (9%) seems to have comparatively little influence on dengue importation. This could be explained, of course by distinct perspectives and approaches of European tourists (e.g. travels to urbanized areas) and immigrants who come to Europe (e.g. Africa, where malaria is much prevalent than dengue, is the main origin from immigrants who arrive to Europe), but also by differences between incubation periods and existing prophylactic measures in both diseases. All dengue cases reported have shown the typical symptomatology of disease, including febrile symptoms in more than 90% of cases (TropNetEurop, 2010). However, it is important to note that the majority of imported dengue infections remain undiagnosed, with a ratio between symptomatic and asymptomatic travelers estimated in 1/3.3 (Cobelens et al., 2002). In general terms, it is estimated that about 80% of all dengue infections are asymptomatic (Farrar, 2008). This high asymptomatic, added to the fact that dengue is not a notifiable disease in much of European countries (Bueno Marí & Jiménez Peydró, 2010c; 2010d), allow us to consider that the knowledge of dengue virus circulation is very limited.


*Note 1*: Imm./Refu. (Immigrants/Refugees), For. Vis . (Foreign Visitors), Eur. E.C. (Europeans living in EC), Eur. Exp. (European Expatriates).

Table 4. Imported dengue in Europe between 2001-2009 (TropNetEurop, 2010).

*Aedes aegypti* is the primary urban vector of dengue and yellow fever basically because it exist a 'domesticated' form of the species that is rarely found more than 100 m from human habitation and feeds almost exclusively on human blood (Reiter, 2010). Both factors allow that *Ae. aegypti* will be considered as an excellent urban vector of viruses. Its distribution was traditionally limited by latitude between 45º N and 35º S according to the existence of January and July 10° C isotherms. Although records out of this latitude range are very rarely, it must be pointed that European northernmost collection of the species occurred in Brest (France) at 48º N (Christopher, 1960). Moreover recent studies have demonstrated that *Ae. aegypti* larvae can withstand temperatures of 2.5° C (Chang et al., 2007). In Eastern Europe it was also seen at its temperature limit at Odessa (Ukraina) at 46º N. (Korovitzkyi and Artemenko, 1933). Despite the species was relatively common in Mediterranean countries, it disappeared from the entire region in the mid-20th century, for reasons that currently are not clear but probably related with thermic tolerance and intensive mosquito

Re-Emergence of Malaria and Dengue in Europe 497

the species was firstly detected in 1990 (Sabatini et al., 1990) and nowadays has colonized more than 2/3 parts of the territory, even having different areas of the country with mosquitoes densities in considerable epidemiological levels. Precisely these locally high densities have allowed the appearance of first cases of human viruses in Europe transmitted by *Ae. albopictus*. Specifically, in the province of Ravenna (Northeastern Italy) occurred an outbreak of Chikungunya virus in 2007. This virus is very similar to dengue and yellow fever (same vectors, bioecology and symptomatology), but much less pathogenic. Just in two and a half months, a total of 205 cases of Chikungunya were reported in two small towns of Ravenna where the infection of *Ae. albopictus* was also confirmed (Rezza et al., 2007). This outbreak of Chikungunya infection, outside a tropical country, was probably begun by a man from India, country that previous year had suffered an epidemic with more than 1 million cases (Ravi, 2006). The Indian man developed a febrile syndrome two days after his arrival in Italy and also had high titres of antibodies against Chikungunya. The phylogenetic analysis showed that the strain that caused Italian outbreak was similar to the strains detected on the Indian subcontinent (Yergolkar et al., 2006), showing in all cases a better adaption to *Ae. albopictus* than other variants. However most worrying scenario took place in 2010 with the re-appearance of first autochthonous cases of dengue in Europe transmitted by *Ae. albopictus*. In this year, two cases of autochthonous dengue fever were diagnosed in Nice (Southeast France) (La Ruche et al., 2010), region where *Ae. albopictus* is established at least since 2004 (Delaunay et al., 2007). Just days after two indigenous cases of Chikungunya in the districts of Alpes-Maritime and Var (also in Southeastern France) were detected through a routinely surveillance of dengue and Chikungunya (ECDC, 2010), which is yearly conducted since 2006 due to the establishment of *Ae. albopictus* in this region. In Greece, other Mediterranean country where *Ae. albopictus* is established at least since 2004 (Klobucar et al., 2006), two cases of indigenous dengue were diagnosed also in 2010 (Schmidt-Chanasit et al., 2010; Gjenero-Margan et al., 2011). The identification of these cases of dengue fever and Chikungunya occurred in 2010, which were in all cases well clustered in space and time, is strongly suggestive that autochthonous transmission of tropical viruses

According to these epidemiological perspectives it seems evident that there is a need to be able to predict the potential distribution and activity of *Ae. albopictus* in Europe to asses about possible re-emergence of dengue and other tropical arboviruses. At respect several Geographic Information Systems (GIS) have been developed in order to predict the number of weeks of activity of *Aedes albopictus* (ECDC, 2009). These GIS models have revealed that throughout much of Europe, more than 23 weeks are predicted to elapse between egg hatching in spring (in response to at least 11.25 hours of daylight and 10.5° C of mean temperature) and adult die-off in autumn (below critical temperature threshold of 9.5° C). Assuming that immature development takes about 2–4 weeks, this constitutes more than 20 weeks of adult activity in Central Europe and Southern United Kingdom, even increasing this activity to more than 40 weeks in southern areas (mainly Greece, Turkey and south of Iberian and Italic Peninsula), depending on availability of surface water for breeding. If these predictions would be fulfilled in Southern Europe, consequently could increase the speed of spread of the species, could also extend the episodes of medical and social alerts derivates from its feeding behavior in urban areas, and even could change the eco-

It must be pointed that *Ae. albopictus* and *Ae. aegypti* are not the only aedine vectors with invasive behavior in Europe. Other exotic mosquitoes, such as *Ochlerotatus japonicus* and

in Europe is ongoing.

epidemiology of viruses that *Ae. albopictus* can transmit.

control campaigns with the employment of DDT. *Ae. aegypti* was common in the Iberian Peninsula mainly introduced from North Africa and was present in this Southern European region up to 1956 (Ribeiro & Ramos, 1999). Since the eradication of the species in Europe, its sporadic presence has been recognized in several countries, namely Britain, France, Italy, Malta, Croatia, Ukraine, Russia and Turkey (Snow & Ramsdale, 1999). However it must be pointed that the species has been reported in Madeira (Portugal) in 2005 (Margarita et al., 2006) and it seems that *Ae. aegypti* is now deeply established in this region because of continuous collections in later years (Almeida et al., 2007). This is the first report of the establishment of the species in Europe since mid-20th century. More recently *Ae. aegypti* has been also captured in The Netherlands (Scholte et al., 2010). In summary, we must pay some attention to surveillance and behavior of *Ae. aegypti* because globalization is provoking the arrival of the species to Europe and global warming could allow the definitive establishment of the species again in Southern areas.

On the other hand the situation is clearly divergent in regard to the secondary vector of dengue and yellow fever, *Aedes albopictus*, usually known as Asian tiger mosquito, due to its quick expansion in Europe in last years. There are several ecological factors that can help us to understand the different importance of *Ae. aegypti* and *Ae. albopictus* as primary and secondary vectors of human viruses respectively. Unlike patterns of oviposition and feeding exhibited by *Ae. aegypti*, Asian tiger mosquito is often abundant in the peridomestic environment, particularly in areas with plentiful vegetation, and feeds freely on humans and other animals. Consequently *Ae. albopictus* can also exist far from human habitation. Additionally *Ae. aegypti* has been globally dispersed from Africa by humans activities since several centuries ago while *Ae. albopictus* was firstly report out of its original Asiatic distribution range in 1979 in Albania (Adhami & Reiter, 1998). Current data indicate that *Ae. albopictus* has been detected much farer north than *Ae. aegypti* and one major difference between both species is that Asian tiger mosquito has the ability to adapt to cold temperatures by becoming dormant during the winter of temperate regions. The ability of *Ae. albopictus* to resist cold temperatures is partially related with its ability to synthesize a high amount of lipids, especially to produce larger amounts of yolk lipid in cold temperatures. At respect, it was demonstrated that larval lipogenesis of *Ae. albopictus* is much more efficient than that of *Ae. aegypti* (Briegel & Timmermann, 2001). Although *Ae. albopictus* occurs in both temperate and tropical areas, only temperate population, but not tropical ones, show a photoperiodic diapauses (Hawley, 1988). During the shortening daylight hours in late summer/early autumn, the reduced photoperiod stimulates the females of *Ae*. *albopictus* to produce eggs that enter facultative diapause (Estrada-Franco & Craig 1995). These eggs can resist hatching stimuli until the following spring and remain in a state of reduced morphogenesis as fully formed first instar larvae, exhibiting increased resistance to environmental extremes. Although the diapause occurs in the egg stage, only adults and pupae are known to be photoperiodically sensitive stages (Wang, 1966; Imai & Maeda, 1976; Mori et al, 1981).

*Ae. albopictus* has been found to be capable to transmit 26 viruses (Moore & Mitchell, 1997; Gratz, 2004; Paupy et al., 2009) and to be experimentally susceptible to several filariasis of veterinary interest (Cancrini et al., 1995; Nayar & Knight, 1999). Globalization has allowed the arrival of this species to Europe, mainly through the transport of eggs and larvae in used tires and gardening products (Reiter & Sprenger, 1987; Madon et al., 2002). The presence of Asian tiger mosquito has been confirmed in 16 European countries, but only in Southern ones the species is deeply established. Particularly interesting is the situation of Italy, where

control campaigns with the employment of DDT. *Ae. aegypti* was common in the Iberian Peninsula mainly introduced from North Africa and was present in this Southern European region up to 1956 (Ribeiro & Ramos, 1999). Since the eradication of the species in Europe, its sporadic presence has been recognized in several countries, namely Britain, France, Italy, Malta, Croatia, Ukraine, Russia and Turkey (Snow & Ramsdale, 1999). However it must be pointed that the species has been reported in Madeira (Portugal) in 2005 (Margarita et al., 2006) and it seems that *Ae. aegypti* is now deeply established in this region because of continuous collections in later years (Almeida et al., 2007). This is the first report of the establishment of the species in Europe since mid-20th century. More recently *Ae. aegypti* has been also captured in The Netherlands (Scholte et al., 2010). In summary, we must pay some attention to surveillance and behavior of *Ae. aegypti* because globalization is provoking the arrival of the species to Europe and global warming could allow the definitive establishment

On the other hand the situation is clearly divergent in regard to the secondary vector of dengue and yellow fever, *Aedes albopictus*, usually known as Asian tiger mosquito, due to its quick expansion in Europe in last years. There are several ecological factors that can help us to understand the different importance of *Ae. aegypti* and *Ae. albopictus* as primary and secondary vectors of human viruses respectively. Unlike patterns of oviposition and feeding exhibited by *Ae. aegypti*, Asian tiger mosquito is often abundant in the peridomestic environment, particularly in areas with plentiful vegetation, and feeds freely on humans and other animals. Consequently *Ae. albopictus* can also exist far from human habitation. Additionally *Ae. aegypti* has been globally dispersed from Africa by humans activities since several centuries ago while *Ae. albopictus* was firstly report out of its original Asiatic distribution range in 1979 in Albania (Adhami & Reiter, 1998). Current data indicate that *Ae. albopictus* has been detected much farer north than *Ae. aegypti* and one major difference between both species is that Asian tiger mosquito has the ability to adapt to cold temperatures by becoming dormant during the winter of temperate regions. The ability of *Ae. albopictus* to resist cold temperatures is partially related with its ability to synthesize a high amount of lipids, especially to produce larger amounts of yolk lipid in cold temperatures. At respect, it was demonstrated that larval lipogenesis of *Ae. albopictus* is much more efficient than that of *Ae. aegypti* (Briegel & Timmermann, 2001). Although *Ae. albopictus* occurs in both temperate and tropical areas, only temperate population, but not tropical ones, show a photoperiodic diapauses (Hawley, 1988). During the shortening daylight hours in late summer/early autumn, the reduced photoperiod stimulates the females of *Ae*. *albopictus* to produce eggs that enter facultative diapause (Estrada-Franco & Craig 1995). These eggs can resist hatching stimuli until the following spring and remain in a state of reduced morphogenesis as fully formed first instar larvae, exhibiting increased resistance to environmental extremes. Although the diapause occurs in the egg stage, only adults and pupae are known to be photoperiodically sensitive stages (Wang, 1966; Imai &

*Ae. albopictus* has been found to be capable to transmit 26 viruses (Moore & Mitchell, 1997; Gratz, 2004; Paupy et al., 2009) and to be experimentally susceptible to several filariasis of veterinary interest (Cancrini et al., 1995; Nayar & Knight, 1999). Globalization has allowed the arrival of this species to Europe, mainly through the transport of eggs and larvae in used tires and gardening products (Reiter & Sprenger, 1987; Madon et al., 2002). The presence of Asian tiger mosquito has been confirmed in 16 European countries, but only in Southern ones the species is deeply established. Particularly interesting is the situation of Italy, where

of the species again in Southern areas.

Maeda, 1976; Mori et al, 1981).

the species was firstly detected in 1990 (Sabatini et al., 1990) and nowadays has colonized more than 2/3 parts of the territory, even having different areas of the country with mosquitoes densities in considerable epidemiological levels. Precisely these locally high densities have allowed the appearance of first cases of human viruses in Europe transmitted by *Ae. albopictus*. Specifically, in the province of Ravenna (Northeastern Italy) occurred an outbreak of Chikungunya virus in 2007. This virus is very similar to dengue and yellow fever (same vectors, bioecology and symptomatology), but much less pathogenic. Just in two and a half months, a total of 205 cases of Chikungunya were reported in two small towns of Ravenna where the infection of *Ae. albopictus* was also confirmed (Rezza et al., 2007). This outbreak of Chikungunya infection, outside a tropical country, was probably begun by a man from India, country that previous year had suffered an epidemic with more than 1 million cases (Ravi, 2006). The Indian man developed a febrile syndrome two days after his arrival in Italy and also had high titres of antibodies against Chikungunya. The phylogenetic analysis showed that the strain that caused Italian outbreak was similar to the strains detected on the Indian subcontinent (Yergolkar et al., 2006), showing in all cases a better adaption to *Ae. albopictus* than other variants. However most worrying scenario took place in 2010 with the re-appearance of first autochthonous cases of dengue in Europe transmitted by *Ae. albopictus*. In this year, two cases of autochthonous dengue fever were diagnosed in Nice (Southeast France) (La Ruche et al., 2010), region where *Ae. albopictus* is established at least since 2004 (Delaunay et al., 2007). Just days after two indigenous cases of Chikungunya in the districts of Alpes-Maritime and Var (also in Southeastern France) were detected through a routinely surveillance of dengue and Chikungunya (ECDC, 2010), which is yearly conducted since 2006 due to the establishment of *Ae. albopictus* in this region. In Greece, other Mediterranean country where *Ae. albopictus* is established at least since 2004 (Klobucar et al., 2006), two cases of indigenous dengue were diagnosed also in 2010 (Schmidt-Chanasit et al., 2010; Gjenero-Margan et al., 2011). The identification of these cases of dengue fever and Chikungunya occurred in 2010, which were in all cases well clustered in space and time, is strongly suggestive that autochthonous transmission of tropical viruses in Europe is ongoing.

According to these epidemiological perspectives it seems evident that there is a need to be able to predict the potential distribution and activity of *Ae. albopictus* in Europe to asses about possible re-emergence of dengue and other tropical arboviruses. At respect several Geographic Information Systems (GIS) have been developed in order to predict the number of weeks of activity of *Aedes albopictus* (ECDC, 2009). These GIS models have revealed that throughout much of Europe, more than 23 weeks are predicted to elapse between egg hatching in spring (in response to at least 11.25 hours of daylight and 10.5° C of mean temperature) and adult die-off in autumn (below critical temperature threshold of 9.5° C). Assuming that immature development takes about 2–4 weeks, this constitutes more than 20 weeks of adult activity in Central Europe and Southern United Kingdom, even increasing this activity to more than 40 weeks in southern areas (mainly Greece, Turkey and south of Iberian and Italic Peninsula), depending on availability of surface water for breeding. If these predictions would be fulfilled in Southern Europe, consequently could increase the speed of spread of the species, could also extend the episodes of medical and social alerts derivates from its feeding behavior in urban areas, and even could change the ecoepidemiology of viruses that *Ae. albopictus* can transmit.

It must be pointed that *Ae. albopictus* and *Ae. aegypti* are not the only aedine vectors with invasive behavior in Europe. Other exotic mosquitoes, such as *Ochlerotatus japonicus* and

Re-Emergence of Malaria and Dengue in Europe 499

Although malaria's receptivity is still high in different parts of Europe, we may conclude that the malariogenic potential of the Old Continent is low. Fortunately socio-economic and sanitary conditions of most European countries also support this assertion. While it is true that infectivity studies should be further promoted, percentages of imported malaria cases remain very low. However we must pay some attention to the increasing trend of malaria importation in last years, as well as also awareness among tourists and VFR's for to take corresponding prophylactic measures during their travels to endemic areas. Anyway, sporadic and local cases of autochthonous transmission mainly transmitted by *An. atroparvus, An. labranchiae, An. sacharovi* and/or *An. plumbeus*, can not been discarded

On the other hand, the answer to the question about if should be expected the re-emergence of dengue and other mosquito-borne tropical viruses in Europe in next years is indubitable: definitively yes. The arrival, establishment and expansion of dengue urbanite vectors due to global changes such as globalization, climate change and the lack of effective mosquito control programs, together with the increasing of imported cases in humans provokes that local and intense transmission of dengue could be a reality in next years in Southern Europe. To cope this possibility is necessary to enhance the entomological surveillance in potential areas of mosquitoes importation, such as airports or seaports, strength the monitoring of tropical viruses imported and awareness among citizens about their role in mosquito control

We wish to acknowledge that current work was partially funded by the Research Project CGL 2009-11364 (BOS), supported by the Ministry of Science and Innovation of Spain

Adege-EID Méditerranée. (2006). *Éléments entomologiques relatifs au risque d'apparition du virus* 

Adhami, J.R. & Reiter, P. (1998). Introduction and establishment of *Aedes (Stegomyia)* 

Almeida, A.P.; Gonçalves, Y.M.; Novo, M.T.; Sousa, C.A.; Melim, M. & Gracio AJ. (2007). Vector

http://www.eurosurveillance.org/ViewArticle.aspx?ArticleId=3311 Alten, B.; Çağlar, S.S.; Şimşek, F.M. & Kaynas, S. (2003). Effect of insecticide-treated bednets

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*Association*, Vol.14, No.3, (September 1998), pp. 340-343, ISSN 1046-3607. Aitken, T.G.H. (1954). The Culicidae of Sardinia and Corsica (Diptera). *Bulletin of* 

*Chikungunya en métropole* . Entente interdépartementale pour la démoustication du

*albopictus* Skuse (Diptera: Culicidae) in Albania. *Journal of American Mosquito Control* 

*Entomological Research*, Vol.45, No.3, (September 1954), pp. 437-494, ISSN 0007-4853.

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for malaria control in Southeast Anatolia - Turkey. *Journal of Vector Ecology*, No.28,

and best prophylactic measures to take during the travels to tropical regions.

(Ministerio de Ciencia e Innovación del Gobierno de España).

littoral (EID) Méditerranée, Montpellier, France.

**4. Conclusions** 

in next years.

**5. Acknowledgments** 

**6. References** 

*Ochlerotatus atropalpus*, have been also reported. *Oc. japonicus* is an Asian species and a competent vector of several arboviruses, including West Nile virus and Japanese encephalitis virus and is considered a significant public health risk (Sardelis & Turell, 2001; Sardelis et al., 2002a; 2002b; 2003). *Oc. japonicus* has been collected only in France, Belgium, Switzerland and Germany (Schafther et al., 2003; 2009; Becker et al., 2011). On the other hand *Oc. atropalpus* is endemic to North America and has been observed in Italy, France and Netherlands (Romi et al., 1997; Adege-EID Méditerranée, 2006; Scholte et al., 2009). Although in the field, *Oc. atropalpus* has not been evidenced as an important vector of infectious diseases, under laboratory conditions, the species has been proven as a competent vector for West Nile virus, Japanese encephalitis virus, Saint-Louis encephalitis virus La Crosse encephalitis virus, among other arboviruses (King, 1960; Turell et al., 2001). Globalization, especially traffic of used tires, has led the arrival of *Oc. japonicus* and *Oc. atropalpus* to Europe. Out of these exotic vectors, we can not forget or ignore the presence of potential indigenous vectors of dengue and yellow fever in Europe. For example, *Aedes vittatus* is an important vector of yellow fever in different parts of Africa (Lewis, 1943; Satti & Haseeb, 1966) and also a potential vector of Chikungunya and four dengue serotypes (Mourya & Banerjee, 1987; Mavale et al., 1992). Although the species is deeply distributed in Mediterranean region (Spain, Portugal, France and Italy), the studies about its biology and phenology have been scanty in Europe. Anyway it seems unlikely that *Ae. vittatus* could start a cycle of virus transmission to humans because of its high degree of ruralism. Moreover *Ochlerotatus geniculatus* is a dendrolimnic species endemic to Europe that can efficiently transmit yellow fever, but this possibility has been evidenced only in laboratory conditions (Roubaud et al., 1937).

#### **3.1 New challenges: The development of dengue vaccines**

Although a vaccine based on live attenuated virus of the strain 17D is available for yelow fever since years, currently we haven´t any vaccine to be used with full warranty against dengue. However, the need for a dengue vaccine is clear. The most effective measures of an integrated mosquito control program (including changes in human habitation and behavior, the use of insecticides, and long-lasting modification of natural and man-made mosquito habitats) are difficult to implement and largely unsuccessful in most poverty-stricken settings, and consequently have not been carried out comprehensively enough to limit dengue's spread. While vector control is an integral part of any dengue prevention strategy, it is not enough on its own.

In recent years it has been obtained a better understanding of the disease and its etiopatogenicity, as well as of the necessary aspects to develop a vaccine that provides an effective and lasting protection against the virus. Dengue vaccine development is a very difficult task due to the possible participation of four related serotypes, since immunity to one serotype does not confer immunity to the remaining three. Complicating the scenario further is immune enhancement, which can result in severe dengue hemorrhagic fever or dengue shock syndrome in anyone who has been infected with one of the serotypes and subsequently becomes infected with another. Most of researchers agree that only effective solution is a tetravalent vaccine that simultaneously protects against all four serotypes. Regarding to this, it must be noted that tetravalent vaccines against dengue are currently in last phases of trials and is expected to be available for human population in the next following years.
