The Global Distribution and Burden of Dengue and Japanese Encephalitis Co-Infection in Acute Encephalitis Syndrome

*Shailendra K. Saxena, Swatantra Kumar, Vimal K. Maurya and Madan L.B. Bhatt*

## **Abstract**

Dengue is widespread throughout the tropics globally in more than hundred countries and coincides with various climatic factors for co-infection with other flaviviral infections of the central nervous system (CNS). Dengue and Japanese encephalitis virus co-infection are highly prevalent, with diagnosis dilemma including significant mortality and morbidity in Southeast Asia. Both dengue and Japanese encephalitis transmissions intensify during the rainy season, during which the vector population increases. CNS involvement during dengue and Japanese encephalitis co-infectionassociated acute encephalitis syndrome (AES) is still poorly understood, and therefore, there is a desperate need to understand the etiology, therapeutics, clinical management, and prevention of these tropically neglected diseases. AES can be differentiated from other etiologies of encephalopathy through considering its essential features: sudden onset of fever, cerebrospinal fluid (CSF) comprising inflammatory cells, magnetic resonance imaging (MRI)-based confirmation, and presence of pathogen or pathogenspecific antibodies. Complementary and alternative medicine is progressively being used globally and can be effective for the overall management of this co-infection.

**Keywords:** dengue, Japanese encephalitis, co-infection, encephalitis, acute encephalitis syndrome (AES), differential diagnosis, treatment, management, prevention, complementary and alternative medicine

### **1. Introduction**

Neglected tropical diseases (NTDs) are the diverse group of communicable diseases which exist in tropical and subtropical settings affecting more than one billion people worldwide [1]. Populations inhabiting places with poor sanitation are in close contact with infected vectors, and domestic animals are principally affected. Arthropod-borne or arboviruses such as dengue, Zika, and Chikungunya have been recently included in the list of NTDs by the World Health Organization [2].

Human infection of flavivirus may cause severe clinical manifestations and can be broadly subdivided into two groups as neurological diseases caused by Japanese encephalitis virus (JEV), West Nile virus, and Zika virus (ZIKV) and hemorrhagic and viscerotropic diseases caused by dengue virus (DENV), ZIKV, and yellow fever virus (YFV) [3]. More than half of the global population is now at the risk of getting flavivirus infections where the majority of areas are endemic for more than one flaviviruses which results in the phenomenon of co-infection [4]. The worldwide incident of dengue has extensively grown in few decades [5]. Majority of the dengue cases are asymptomatic, and therefore, it is hard to anticipate the accurate burden of the disease. The rise in number of cases from 2.2 million in the year 2010 to 3.34 million cases in 2016 suggests the sharp increase in the disease burden. The 2016 year is characterized as the largest outbreak for dengue where 2.38 million cases were reported from the region of the Americas where 1.5 million cases were contributed by Brazil alone. Currently 3.9 billion in 128 countries people are at risk of DENV infection [6]. Unlike dengue, Japanese encephalitis (JE) is confined to Southeast Asia and Western Pacific regions. Approximately 68,000 clinical cases of JE are reported annually with 13,000 to 24,000 deaths. Currently more than 3 billion in 24 countries are at risk of JEV infection [7]. The epidemiology of dengue and JE has been depicted in **Figure 1**.

DENV and JEV belongs to the *Flaviviridae* family, which consists of more than 70 viruses, comprising of single-stranded positive-sense RNA genome protected by envelope protein [8]. Viruses from this family belong to the genus *Flavivirus*, which are transmitted by mosquitoes or ticks and are characterized as arthropod-borne infections. The transmission cycle of *Flavivirus* involves animals including human which are considered to be the dead-end hosts [9]. Hematophagous mosquitoes are the transmission vector for these diseases. *Aedes albopictus* and *Aedes aegypti* mosquitoes are known to transmit the dengue virus, whereas *Culex tritaeniorhynchus* is predominantly involve in the transmission of JEV [10]. These viruses have been shown to be transmitted via transplacental route as well [11].

Pathogen-associated acute encephalitis syndrome (PA-AES) may result from diverse pathogenic infections including DENV and JEV. PA-AES shows a wide range of symptoms including headache, vomiting and severe illness, reduced consciousness, altered sensorium, convulsions, and tremors [12]. Flaviviruses share substantial sequence similarities to stimulate sero-cross-reactivity which results in the antibody-dependent enhancement (ADE) of infection with other flaviviruses [13].

#### **Figure 1.**

*Dengue and Japanese encephalitis epidemiology. The worldwide epidemiology of dengue and Japanese encephalitis has been depicted in this map. More than 100 countries are endemic for dengue where America, Western Pacific regions, and Southeast Asia are mostly affected. The Japanese encephalitis is confined to mostly in Southeast Asia and Western Pacific regions which include approximately 24 countries. The dengue-affected regions are highlighted in red color, JE-affected regions are highlighted in yellow color, and regions where both viruses are circulating have been highlighted in orange color.*

**23**

**Figure 2.**

*The Global Distribution and Burden of Dengue and Japanese Encephalitis Co-Infection in Acute…*

Outbreaks of DENV co-infection are predominantly associated with the JEV endemic area or areas of JE immunization [14]. Although DENV-induced encephalitis is rare, the co-infection may increase the severity of encephalitis. DENV infection causes dengue hemorrhagic fever (DHF), whereas JEV infection may result in neurological complications [15]. Intermittently, DENV has been reported to cause encephalitis, and JEV infection may cause extraneural hemorrhage [16, 17].

However, this may not be the case in co-infection with both the flaviviruses. Simultaneous detection of both the viruses is not unusual; nevertheless we need to

Co-infection with both viruses may occur in the single-cell types or different cells in the infected individuals. Simultaneous infection of target cells or different cell types by DENV and JEV defines the phenomenon of co-infection [18]. Both the viruses share common cell surface receptors expressed on target cell types such as DC-SIGN, mannose receptors, and CLEC5A [19]. On the other hand, virus can be internalized in the presence of non-neutralizing antibodies via Fc*γ* receptor-mediated endocytosis [20]. Clathrin-mediated endocytosis has been shown to be involved in the internalization of flaviviruses [21]. After internalization, the virus from the endocytic vesicles is delivered to the early endosomes [22]. Acidification of the endosomal

*DENV and JEV co-infection. The mechanism of co-infection has not been completely understood.* 

*Co-infection with DENV and JEV may occur in the single-cell types or different cells in the infected individuals. Co-infection results in the generation of antibodies against both viruses. However, detection of nucleic acids and antigens may confirm the phenomenon of co-infection (A). Upon infection, JEV is internalized by receptor-mediated endocytosis. The decrease in pH causes the fusion of viral and endosomal membrane which results in the release of viral RNA into the cytosol. The released viral genome is then translated prior to the commencement of replication. Virus maturation occurs at the Golgi complex and mature virus is released via the egress process. JEV infection in the hyperendemic area may result in false-positive diagnosis due to presence of pre-existing sero-cross-reactive antibody (B). Pre-existing sero-cross-reactive antibodies may bind to the viral particles upon infection which results in antibody-dependent enhancement of infection (C).*

be attentive in the diagnosis of the etiology of PA-AES.

**2. Molecular mechanism of DENV and JEV co-infection**

*DOI: http://dx.doi.org/10.5772/intechopen.89792*

*The Global Distribution and Burden of Dengue and Japanese Encephalitis Co-Infection in Acute… DOI: http://dx.doi.org/10.5772/intechopen.89792*

Outbreaks of DENV co-infection are predominantly associated with the JEV endemic area or areas of JE immunization [14]. Although DENV-induced encephalitis is rare, the co-infection may increase the severity of encephalitis. DENV infection causes dengue hemorrhagic fever (DHF), whereas JEV infection may result in neurological complications [15]. Intermittently, DENV has been reported to cause encephalitis, and JEV infection may cause extraneural hemorrhage [16, 17]. However, this may not be the case in co-infection with both the flaviviruses. Simultaneous detection of both the viruses is not unusual; nevertheless we need to be attentive in the diagnosis of the etiology of PA-AES.

## **2. Molecular mechanism of DENV and JEV co-infection**

Co-infection with both viruses may occur in the single-cell types or different cells in the infected individuals. Simultaneous infection of target cells or different cell types by DENV and JEV defines the phenomenon of co-infection [18]. Both the viruses share common cell surface receptors expressed on target cell types such as DC-SIGN, mannose receptors, and CLEC5A [19]. On the other hand, virus can be internalized in the presence of non-neutralizing antibodies via Fc*γ* receptor-mediated endocytosis [20]. Clathrin-mediated endocytosis has been shown to be involved in the internalization of flaviviruses [21]. After internalization, the virus from the endocytic vesicles is delivered to the early endosomes [22]. Acidification of the endosomal

#### **Figure 2.**

*Current Topics in Neglected Tropical Diseases*

virus (YFV) [3]. More than half of the global population is now at the risk of getting flavivirus infections where the majority of areas are endemic for more than one flaviviruses which results in the phenomenon of co-infection [4]. The worldwide incident of dengue has extensively grown in few decades [5]. Majority of the dengue cases are asymptomatic, and therefore, it is hard to anticipate the accurate burden of the disease. The rise in number of cases from 2.2 million in the year 2010 to 3.34 million cases in 2016 suggests the sharp increase in the disease burden. The 2016 year is characterized as the largest outbreak for dengue where 2.38 million cases were reported from the region of the Americas where 1.5 million cases were contributed by Brazil alone. Currently 3.9 billion in 128 countries people are at risk of DENV infection [6]. Unlike dengue, Japanese encephalitis (JE) is confined to Southeast Asia and Western Pacific regions. Approximately 68,000 clinical cases of JE are reported annually with 13,000 to 24,000 deaths. Currently more than 3 billion in 24 countries are at risk of JEV infection [7]. The epidemiology of dengue and JE has been depicted in **Figure 1**. DENV and JEV belongs to the *Flaviviridae* family, which consists of more than 70 viruses, comprising of single-stranded positive-sense RNA genome protected by envelope protein [8]. Viruses from this family belong to the genus *Flavivirus*, which are transmitted by mosquitoes or ticks and are characterized as arthropod-borne infections. The transmission cycle of *Flavivirus* involves animals including human which are considered to be the dead-end hosts [9]. Hematophagous mosquitoes are the transmission vector for these diseases. *Aedes albopictus* and *Aedes aegypti* mosquitoes are known to transmit the dengue virus, whereas *Culex tritaeniorhynchus* is predominantly involve in the transmission of JEV [10]. These viruses have been

shown to be transmitted via transplacental route as well [11].

Pathogen-associated acute encephalitis syndrome (PA-AES) may result from diverse pathogenic infections including DENV and JEV. PA-AES shows a wide range of symptoms including headache, vomiting and severe illness, reduced consciousness, altered sensorium, convulsions, and tremors [12]. Flaviviruses share substantial sequence similarities to stimulate sero-cross-reactivity which results in the antibody-dependent enhancement (ADE) of infection with other flaviviruses [13].

*Dengue and Japanese encephalitis epidemiology. The worldwide epidemiology of dengue and Japanese encephalitis has been depicted in this map. More than 100 countries are endemic for dengue where America, Western Pacific regions, and Southeast Asia are mostly affected. The Japanese encephalitis is confined to mostly in Southeast Asia and Western Pacific regions which include approximately 24 countries. The dengue-affected regions are highlighted in red color, JE-affected regions are highlighted in yellow color, and regions where both* 

*viruses are circulating have been highlighted in orange color.*

**22**

**Figure 1.**

*DENV and JEV co-infection. The mechanism of co-infection has not been completely understood. Co-infection with DENV and JEV may occur in the single-cell types or different cells in the infected individuals. Co-infection results in the generation of antibodies against both viruses. However, detection of nucleic acids and antigens may confirm the phenomenon of co-infection (A). Upon infection, JEV is internalized by receptor-mediated endocytosis. The decrease in pH causes the fusion of viral and endosomal membrane which results in the release of viral RNA into the cytosol. The released viral genome is then translated prior to the commencement of replication. Virus maturation occurs at the Golgi complex and mature virus is released via the egress process. JEV infection in the hyperendemic area may result in false-positive diagnosis due to presence of pre-existing sero-cross-reactive antibody (B). Pre-existing sero-cross-reactive antibodies may bind to the viral particles upon infection which results in antibody-dependent enhancement of infection (C).*

compartments causes trimerization of the envelope protein that results in the fusion of endosomes to the viral membrane and release of nucleocapsid in the cytosol [23].

The released viral genome is then translated prior to the commencement of replication [24]. The backbone of flaviviral genome is invariable which are of ~10 kb that codes for three structural capsid (C), premembrane (prM), and envelope (E) and seven nonstructural proteins NS1, NS2a, NS2b, NS3, NS4a, NS4b, and NS5. The viral genome encodes for 3400 amino acid long polyprotein which is arranged in the lumen endoplasmic reticulum where the replication takes place by the RNAdependent RNA polymerase [25]. Replicated copies of the genome interact with viral proteins to form nucleocapsid. Immature virions enter into the secretary pathway where furin-mediated cleavage of prM results in the maturation of virus [26]. Mature virus is then released from the infected cells via the egress process [27]. The conventional mechanism of pathogenesis and viral replication may not be followed in case of co-infection and may involve distinct process. In order to effectively control and treat the co-infection of DENV and JEV, we need to first understand the immunopathogenesis of dual infection in various cell and animal models. The probable phenomenon of DENV and JEV co-infection has been demonstrated in **Figure 2**.

## **3. Diagnostic schemes for dengue and Japanese encephalitis co-infection**

JEV is the most documented causative agent of acute encephalitis syndrome (AES) [28]. DENV has also been considered among the top three etiological agents causing


#### **Figure 3.**

*Laboratory diagnostic algorithms during DENV and JEV co-infection. Suspected cases of DENV and JEV co-infection can be decided by the above laboratory diagnostic algorithms of co-infection. Detection of IgG antibodies against both viruses may be considered as the previous exposure of the viruses, whereas detection of IgM or NS1 antigen suggests the recent or current infectious conditions. However, this should be further validated based on the nucleic acid-based tests such as RT-PCR and isolation of virus which may define the co-infectious condition in addition to the clinical correlation. "Y" indicates positive, "N" indicates negative, and "?" indicates inconclusive test report.*

**25**

of co-infection.

**co-infection**

*The Global Distribution and Burden of Dengue and Japanese Encephalitis Co-Infection in Acute…*

AES [29]. Broad spectrum of clinical features in the endemic areas of both viruses may dilute the encephalitis-like clinical presentation. Therefore, laboratory based diagnosis and discrimination of pathogenesis is crucial to understand. Detection of IgG antibodies against both viruses may be considered as the previous exposure of the viruses, whereas detection of IgM suggests the recent or current infectious conditions. However, this should be further validated based on the nucleic acid-based tests where detection of DENV and JEV nucleic acids defines the co-infectious condition. Nucleic acid-based detection by RT-PCR for other probable arboviral infections such as Zika and Chikungunya can also be included which may overlap with the clinical manifestations of dengue or JE [30]. Although the clinical manifestations presented in the individual cases of DENV and JEV infections are distinct, the coinfection might be a complex situation to diagnose. Co-detection of serum antibodies against both viruses may not be the evidence of co-infection. Surveillance programs conducted in the hyperendemic areas of flaviviruses have reported at least 9% of co-infections cases [31]. Moreover, the MRI findings of DENV-infected patients may show the characteristic features of encephalitis as in case of JEV where the basal ganglia, thalamus, and midbrain are predominantly affected [32]. Considering the sero-cross-reactivity among flaviviruses, the dependence of serum-based diagnosis may give false-positive results. However, simultaneous detection of nucleic acid or viral-specific antigens in blood or CSF samples may define the incident of co-infection. The laboratory diagnostic algorithms during

DENV and JEV co-infection have been depicted in **Figure 3**.

**4. Treatments regimes for dengue and Japanese encephalitis co-infection**

Viral infections have always been a global threat to mankind due to scarcity of effective antiviral drugs. Flaviviral infections are not the exception since there is no specific treatment available for both DENV and JEV [33]. The overall treatment relies on the symptomatic relief of the patients. Due to the complex and unclear pathogenesis of dual infection, the potential candidate drugs may not effective. Therefore, we need to look for the complementary and alternative medicine (CAM) in alliance with

In case of DENV infection, various forms of CAM have been used such as *Carica papaya* leaf extracts is the most accepted one. Platelet-activating factor receptor (PTAFR) gene has been shown to be upregulated upon consumption of *Carica papaya* leaf extracts or its juices [34]. In case of sever dengue infection, maintenance of body fluids volume of the patients is critical [35]. However, this may or may not be effective in case of co-infections. Recently, andrographolide has been shown to exhibit anti-DENV activity [36]. *Eupatorium perfoliatum* which is a homeopathic medicine has been shown to exhibit anti-DENV activity [37]. Luteolin has been shown to be effective during JEV infection which also exhibits direct virucidal activity [38]. Similarly, belladonna has been shown to be effective in chick embryos infected with JEV [39]. Several of the CAM-based therapies have been shown to be effective in case of JEV infection, but these have to be validated in case

conventional medicines as the choice of treatment during co-infection.

**5. Preventive strategies for dengue and Japanese encephalitis** 

To prevent the worldwide burden of DENV infection, the WHO has recently approved a tetravalent vaccine, Dengvaxia (CYD-TDV) in 20 countries. This has

*DOI: http://dx.doi.org/10.5772/intechopen.89792*

*The Global Distribution and Burden of Dengue and Japanese Encephalitis Co-Infection in Acute… DOI: http://dx.doi.org/10.5772/intechopen.89792*

AES [29]. Broad spectrum of clinical features in the endemic areas of both viruses may dilute the encephalitis-like clinical presentation. Therefore, laboratory based diagnosis and discrimination of pathogenesis is crucial to understand. Detection of IgG antibodies against both viruses may be considered as the previous exposure of the viruses, whereas detection of IgM suggests the recent or current infectious conditions. However, this should be further validated based on the nucleic acid-based tests where detection of DENV and JEV nucleic acids defines the co-infectious condition.

Nucleic acid-based detection by RT-PCR for other probable arboviral infections such as Zika and Chikungunya can also be included which may overlap with the clinical manifestations of dengue or JE [30]. Although the clinical manifestations presented in the individual cases of DENV and JEV infections are distinct, the coinfection might be a complex situation to diagnose. Co-detection of serum antibodies against both viruses may not be the evidence of co-infection. Surveillance programs conducted in the hyperendemic areas of flaviviruses have reported at least 9% of co-infections cases [31]. Moreover, the MRI findings of DENV-infected patients may show the characteristic features of encephalitis as in case of JEV where the basal ganglia, thalamus, and midbrain are predominantly affected [32]. Considering the sero-cross-reactivity among flaviviruses, the dependence of serum-based diagnosis may give false-positive results. However, simultaneous detection of nucleic acid or viral-specific antigens in blood or CSF samples may define the incident of co-infection. The laboratory diagnostic algorithms during DENV and JEV co-infection have been depicted in **Figure 3**.

#### **4. Treatments regimes for dengue and Japanese encephalitis co-infection**

Viral infections have always been a global threat to mankind due to scarcity of effective antiviral drugs. Flaviviral infections are not the exception since there is no specific treatment available for both DENV and JEV [33]. The overall treatment relies on the symptomatic relief of the patients. Due to the complex and unclear pathogenesis of dual infection, the potential candidate drugs may not effective. Therefore, we need to look for the complementary and alternative medicine (CAM) in alliance with conventional medicines as the choice of treatment during co-infection.

In case of DENV infection, various forms of CAM have been used such as *Carica papaya* leaf extracts is the most accepted one. Platelet-activating factor receptor (PTAFR) gene has been shown to be upregulated upon consumption of *Carica papaya* leaf extracts or its juices [34]. In case of sever dengue infection, maintenance of body fluids volume of the patients is critical [35]. However, this may or may not be effective in case of co-infections. Recently, andrographolide has been shown to exhibit anti-DENV activity [36]. *Eupatorium perfoliatum* which is a homeopathic medicine has been shown to exhibit anti-DENV activity [37]. Luteolin has been shown to be effective during JEV infection which also exhibits direct virucidal activity [38]. Similarly, belladonna has been shown to be effective in chick embryos infected with JEV [39]. Several of the CAM-based therapies have been shown to be effective in case of JEV infection, but these have to be validated in case of co-infection.

### **5. Preventive strategies for dengue and Japanese encephalitis co-infection**

To prevent the worldwide burden of DENV infection, the WHO has recently approved a tetravalent vaccine, Dengvaxia (CYD-TDV) in 20 countries. This has

*Current Topics in Neglected Tropical Diseases*

compartments causes trimerization of the envelope protein that results in the fusion of endosomes to the viral membrane and release of nucleocapsid in the cytosol [23]. The released viral genome is then translated prior to the commencement of replication [24]. The backbone of flaviviral genome is invariable which are of ~10 kb that codes for three structural capsid (C), premembrane (prM), and envelope (E) and seven nonstructural proteins NS1, NS2a, NS2b, NS3, NS4a, NS4b, and NS5. The viral genome encodes for 3400 amino acid long polyprotein which is arranged in the lumen endoplasmic reticulum where the replication takes place by the RNAdependent RNA polymerase [25]. Replicated copies of the genome interact with viral proteins to form nucleocapsid. Immature virions enter into the secretary pathway where furin-mediated cleavage of prM results in the maturation of virus [26]. Mature virus is then released from the infected cells via the egress process [27]. The conventional mechanism of pathogenesis and viral replication may not be followed in case of co-infection and may involve distinct process. In order to effectively control and treat the co-infection of DENV and JEV, we need to first understand the immunopathogenesis of dual infection in various cell and animal models. The probable phenomenon of DENV and JEV co-infection has been demonstrated in **Figure 2**.

**3. Diagnostic schemes for dengue and Japanese encephalitis co-infection**

*Laboratory diagnostic algorithms during DENV and JEV co-infection. Suspected cases of DENV and JEV co-infection can be decided by the above laboratory diagnostic algorithms of co-infection. Detection of IgG antibodies against both viruses may be considered as the previous exposure of the viruses, whereas detection of IgM or NS1 antigen suggests the recent or current infectious conditions. However, this should be further validated based on the nucleic acid-based tests such as RT-PCR and isolation of virus which may define the co-infectious condition in addition to the clinical correlation. "Y" indicates positive, "N" indicates negative, and* 

JEV is the most documented causative agent of acute encephalitis syndrome (AES) [28]. DENV has also been considered among the top three etiological agents causing

**24**

**Figure 3.**

*"?" indicates inconclusive test report.*

been designed by using the yellow fever vaccine backbone expressing the prM and envelope protein of DENV 1–4 serotypes [40]. In case of JEV, a live attenuated vaccine based on SA 14–14-2 has been used in China, India, Sri Lanka, Republic of Korea, and Thailand. Due to higher sero-cross-reactivity, the one vaccine may induce the other infection due to the antibody-dependent enhancement [41]. Therefore, to design an effective vaccine for co-infection, there is a need to understand the mechanism and probability of sero-cross-reactivity among the viruses. Apart from the vaccination, the personal preventive measures are always the paramount to prevent any vector-borne infections [42]. To prevent the mosquito biting, several protective measures include mosquito repellents, mosquito nets, and use of full sleeves cloths [43].

## **6. Conclusions**

Genomic and proteomic sequence similarity among the flaviviruses causes the sero-cross-reactivity that leads to the phenomenon of antibody-dependent enhancement of infections. Incidence of DENV and JEV co-infection in the hyperendemic areas may be frequently reported. The diagnosis of co-infection should not rely on the presence of serum antibodies. However, simultaneous detection of nucleic acids or antigens may define the condition of co-infection. Clinical features may overlap in the patients infected with both viruses, and therefore we need to precisely distinguish the patients' clinical reports. Development of effective antivirals targeting both the viruses is the most imperative therapeutic strategy.

## **7. Future perspectives**

Due to the similarity in the structural domains of the viral proteins, molecules may be designed to inhibit the action of viral proteins or enzymes. Similarly, vaccines may be designed to target the population living in the hyperendemic areas of flaviviruses. Peptide-based vaccines may be designed by using various immunoinformatics approaches by considering the consensus peptide sequences among the viruses. Apart from the vaccination, personal preventive measures are always recommended the best practice to reduce the chances of infections.

## **Acknowledgements**

The authors are grateful to the Vice Chancellor, King George's Medical University (KGMU), Lucknow, India, for the encouragement and support for this work. SK Saxena is also supported by CCRH, Government of India, and US NIH grants: R37DA025576 and R01MH085259. The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed.

**27**

**Author details**

*The Global Distribution and Burden of Dengue and Japanese Encephalitis Co-Infection in Acute…*

Shailendra K. Saxena\*, Swatantra Kumar, Vimal K. Maurya and Madan L.B. Bhatt Center for Advanced Research (CFAR)-Stem Cell/Cell Culture Unit, Faculty of

© 2019 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium,

Medicine, King George's Medical University, Lucknow, India

\*Address all correspondence to: myedrsaxena@gmail.com

provided the original work is properly cited.

*DOI: http://dx.doi.org/10.5772/intechopen.89792*

*The Global Distribution and Burden of Dengue and Japanese Encephalitis Co-Infection in Acute… DOI: http://dx.doi.org/10.5772/intechopen.89792*

## **Author details**

*Current Topics in Neglected Tropical Diseases*

use of full sleeves cloths [43].

**7. Future perspectives**

**Acknowledgements**

manuscript apart from those disclosed.

**6. Conclusions**

been designed by using the yellow fever vaccine backbone expressing the prM and envelope protein of DENV 1–4 serotypes [40]. In case of JEV, a live attenuated vaccine based on SA 14–14-2 has been used in China, India, Sri Lanka, Republic of Korea, and Thailand. Due to higher sero-cross-reactivity, the one vaccine may induce the other infection due to the antibody-dependent enhancement [41]. Therefore, to design an effective vaccine for co-infection, there is a need to understand the mechanism and probability of sero-cross-reactivity among the viruses. Apart from the vaccination, the personal preventive measures are always the paramount to prevent any vector-borne infections [42]. To prevent the mosquito biting, several protective measures include mosquito repellents, mosquito nets, and

Genomic and proteomic sequence similarity among the flaviviruses causes the sero-cross-reactivity that leads to the phenomenon of antibody-dependent enhancement of infections. Incidence of DENV and JEV co-infection in the hyperendemic areas may be frequently reported. The diagnosis of co-infection should not rely on the presence of serum antibodies. However, simultaneous detection of nucleic acids or antigens may define the condition of co-infection. Clinical features may overlap in the patients infected with both viruses, and therefore we need to precisely distinguish the patients' clinical reports. Development of effective antivi-

rals targeting both the viruses is the most imperative therapeutic strategy.

Due to the similarity in the structural domains of the viral proteins, molecules

may be designed to inhibit the action of viral proteins or enzymes. Similarly, vaccines may be designed to target the population living in the hyperendemic areas of flaviviruses. Peptide-based vaccines may be designed by using various immunoinformatics approaches by considering the consensus peptide sequences among the viruses. Apart from the vaccination, personal preventive measures are always

The authors are grateful to the Vice Chancellor, King George's Medical University (KGMU), Lucknow, India, for the encouragement and support for this work. SK Saxena is also supported by CCRH, Government of India, and US NIH grants: R37DA025576 and R01MH085259. The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the

recommended the best practice to reduce the chances of infections.

**26**

Shailendra K. Saxena\*, Swatantra Kumar, Vimal K. Maurya and Madan L.B. Bhatt Center for Advanced Research (CFAR)-Stem Cell/Cell Culture Unit, Faculty of Medicine, King George's Medical University, Lucknow, India

\*Address all correspondence to: myedrsaxena@gmail.com

© 2019 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/ by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

## **References**

[1] Molyneux DH, Savioli L, Engels D. Neglected tropical diseases: Progress towards addressing the chronic pandemic. Lancet. 2017;**389**(10066):312-325

[2] Mitra AK, Mawson AR. Neglected tropical diseases: Epidemiology and global burden. Tropical Medicine and Infectious Disease. 2017;**2**(3):pii. E36

[3] Holbrook MR. Historical perspectives on Flavivirus research. Viruses. 2017;**9**(5):pii. E97

[4] Vogels CBF, Rückert C, Cavany SM, Perkins TA, Ebel GD, Grubaugh ND. Arbovirus coinfection and co-transmission: A neglected public health concern? PLoS Biology. 2019;**17**(1):e3000130

[5] Guo C, Zhou Z, Wen Z, Liu Y, Zeng C, Xiao D, et al. Global epidemiology of dengue outbreaks in 1990-2015: A systematic review and meta-analysis. Frontiers in Cellular and Infection Microbiology. 2017;**7**:317

[6] World Health Organization. Dengue and severe dengue. https://www.who. int/news-room/fact-sheets/detail/ dengue-and-severe-dengue [Accessed: June 01, 2019]

[7] World Health Organization. Japanese encephalitis. https://www.who.int/ news-room/fact-sheets/detail/japaneseencephalitis [Accessed: June 01, 2019]

[8] Daep CA, Muñoz-Jordán JL, Eugenin EA. Flaviviruses, an expanding threat in public health: Focus on dengue, west Nile, and Japanese encephalitis virus. Journal of Neurovirology. 2014;**20**(6):539-560

[9] Neufeldt CJ, Cortese M, Acosta EG, Bartenschlager R. Rewiring cellular networks by members of the

Flaviviridae family. Nature Reviews. Microbiology. 2018;**16**(3):125-142

[10] Huang YJ, Higgs S, Horne KM, Vanlandingham DL. Flavivirusmosquito interactions. Viruses. 2014;**6**(11):4703-4730

[11] Raj Y, Kumar S, Haikerwal A, Goel MM, Bhatt ML, Saxena SK. Current advances in Zika virus transmission: Urgency for effective therapeutics and prevention. American Journal of Infectious Diseases. 2017;**13**(2):13-20

[12] Saxena SK, Kumar S, Maurya VK. Pathogen-associated acute encephalitis syndrome: Therapeutics and management. Future Microbiology. 2019;**14**:259-262

[13] Saron WAA, Rathore APS, Ting L, Ooi EE, Low J, Abraham SN, et al. Flavivirus serocomplex cross-reactive immunity is protective by activating heterologous memory CD4 T cells. Science Advances. 2018;**4**(7):eaar4297

[14] Saito Y, Moi ML, Takeshita N, Lim CK, Shiba H, Hosono K, et al. Japanese encephalitis vaccine-facilitated dengue virus infection-enhancement antibody in adults. BMC Infectious Diseases. 2016;**16**(1):578

[15] Garg RK, Malhotra HS, Gupta A, Kumar N, Jain A. Concurrent dengue virus and Japanese encephalitis virus infection of the brain: Is it co-infection or co-detection? Infection. 2012;**40**(5):589-593

[16] Aggarwal A, Kumar P, Faridi MM. Neurological manifestation as presenting feature of dengue infection. Journal of Pediatric Neurosciences. 2015;**10**(1):76-77

[17] Tiroumourougane SV, Raghava P, Srinivasan S. Japanese viral encephalitis.

**29**

2018;**9**:595

*The Global Distribution and Burden of Dengue and Japanese Encephalitis Co-Infection in Acute…*

[26] Plevka P, Battisti AJ, Junjhon J, Winkler DC, Holdaway HA, Keelapang P, et al. Maturation of flaviviruses starts from one or more icosahedrally independent nucleation

[27] Plevka P, Battisti AJ, Sheng J, Rossmann MG. Mechanism for maturation-related reorganization of flavivirus glycoproteins. Journal of Structural Biology. 2014;**185**(1):27-31

[28] Ghosh S, Basu A. Acute encephalitis syndrome in India: The changing scenario. Annals of Neurosciences.

centres. EMBO Reports. 2011;**12**(6):602-606

2016;**23**(3):131-133

[29] Vasanthapuram R, Shahul Hameed SK, Desai A, Mani RS, Reddy V, Velayudhan A, et al. Dengue virus is an under-recognised causative agent of acute encephalitis syndrome (AES): Results from a four year AES surveillance study of Japanese encephalitis in selected states of India. International Journal Infectious Diseases. 2019;**84S**(2019):S19-S24

[30] Paixão ES, Teixeira MG,

[31] Singh KP, Mishra G, Jain P, Pandey N, Nagar R, Gupta S, et al. Co-positivity of anti-dengue virus and anti-Japanese encephalitis virus IgM in endemic area: co-infection or cross reactivity? Asian Pacific Journal of Tropical Medicine. 2014;**7**(2):124-129

Rodrigues LC. Zika, chikungunya and dengue: The causes and threats of new and re-emerging arboviral diseases. BMJ Global Health. 2018;**3**(Suppl 1):e000530

[32] Verma R. MRI features of Japanese encephalitis. BML Case Reports. 2012;

[33] Wang S, Liu Y, Guo J, Wang P, Zhang L, Xiao G, Wang W. Screening of FDA-approved drugs for inhibitors of Japanese encephalitis virus infection.

2012. pii: bcr0320126088

*DOI: http://dx.doi.org/10.5772/intechopen.89792*

Postgraduate Medical Journal.

[18] Salas-Benito JS, De Nova-Ocampo M. Viral interference and persistence in mosquito borne flaviviruses. Journal of Immunology

Research. 2015;**2015**:873404

[19] Laureti M, Narayanan D, Rodriguez-Andres J, Fazakerley JK, Kedzierski L. Flavivirus receptors: Diversity, identity, and cell entry. Frontiers in Immunology. 2018;**9**:2180

against flaviviruses. Nature

2015;**10**(10):e0140824

[20] Slon Campos JL, Mongkolsapaya J, Screaton GR. The immune response

Immunology. 2018;**19**(11):1189-1198

[21] Piccini LE, Castilla V, Damonte EB. Dengue-3 virus entry into vero cells: Role of clathrin-mediated endocytosis in the outcome of infection. PLoS One.

[22] Hackett BA, Cherry S. Flavivirus internalization is regulated by a size-dependent endocytic pathway. Proceedings of the National Academy of Sciences of the United States of America. 2018;**115**(16):4246-4251

[23] Bressanelli S, Stiasny K, Allison SL, Stura EA, Duquerroy S, Lescar J, et al. Structure of a flavivirus envelope glycoprotein in its low-pH-induced membrane fusion conformation. The EMBO Journal. 2004;**23**(4):728-738

[24] Nour AM, Li Y, Wolenski J, Modis Y. Viral membrane fusion and nucleocapsid delivery into the cytoplasm are distinct events in some flaviviruses. PLoS Pathogens.

2013;**9**(9):e1003585

[25] Mazeaud C, Freppel W,

Chatel-Chaix L. The multiples fates of the flavivirus RNA genome during pathogenesis. Frontiers in Genetics.

2002;**78**(918):205-215

*The Global Distribution and Burden of Dengue and Japanese Encephalitis Co-Infection in Acute… DOI: http://dx.doi.org/10.5772/intechopen.89792*

Postgraduate Medical Journal. 2002;**78**(918):205-215

[18] Salas-Benito JS, De Nova-Ocampo M. Viral interference and persistence in mosquito borne flaviviruses. Journal of Immunology Research. 2015;**2015**:873404

[19] Laureti M, Narayanan D, Rodriguez-Andres J, Fazakerley JK, Kedzierski L. Flavivirus receptors: Diversity, identity, and cell entry. Frontiers in Immunology. 2018;**9**:2180

[20] Slon Campos JL, Mongkolsapaya J, Screaton GR. The immune response against flaviviruses. Nature Immunology. 2018;**19**(11):1189-1198

[21] Piccini LE, Castilla V, Damonte EB. Dengue-3 virus entry into vero cells: Role of clathrin-mediated endocytosis in the outcome of infection. PLoS One. 2015;**10**(10):e0140824

[22] Hackett BA, Cherry S. Flavivirus internalization is regulated by a size-dependent endocytic pathway. Proceedings of the National Academy of Sciences of the United States of America. 2018;**115**(16):4246-4251

[23] Bressanelli S, Stiasny K, Allison SL, Stura EA, Duquerroy S, Lescar J, et al. Structure of a flavivirus envelope glycoprotein in its low-pH-induced membrane fusion conformation. The EMBO Journal. 2004;**23**(4):728-738

[24] Nour AM, Li Y, Wolenski J, Modis Y. Viral membrane fusion and nucleocapsid delivery into the cytoplasm are distinct events in some flaviviruses. PLoS Pathogens. 2013;**9**(9):e1003585

[25] Mazeaud C, Freppel W, Chatel-Chaix L. The multiples fates of the flavivirus RNA genome during pathogenesis. Frontiers in Genetics. 2018;**9**:595

[26] Plevka P, Battisti AJ, Junjhon J, Winkler DC, Holdaway HA, Keelapang P, et al. Maturation of flaviviruses starts from one or more icosahedrally independent nucleation centres. EMBO Reports. 2011;**12**(6):602-606

[27] Plevka P, Battisti AJ, Sheng J, Rossmann MG. Mechanism for maturation-related reorganization of flavivirus glycoproteins. Journal of Structural Biology. 2014;**185**(1):27-31

[28] Ghosh S, Basu A. Acute encephalitis syndrome in India: The changing scenario. Annals of Neurosciences. 2016;**23**(3):131-133

[29] Vasanthapuram R, Shahul Hameed SK, Desai A, Mani RS, Reddy V, Velayudhan A, et al. Dengue virus is an under-recognised causative agent of acute encephalitis syndrome (AES): Results from a four year AES surveillance study of Japanese encephalitis in selected states of India. International Journal Infectious Diseases. 2019;**84S**(2019):S19-S24

[30] Paixão ES, Teixeira MG, Rodrigues LC. Zika, chikungunya and dengue: The causes and threats of new and re-emerging arboviral diseases. BMJ Global Health. 2018;**3**(Suppl 1):e000530

[31] Singh KP, Mishra G, Jain P, Pandey N, Nagar R, Gupta S, et al. Co-positivity of anti-dengue virus and anti-Japanese encephalitis virus IgM in endemic area: co-infection or cross reactivity? Asian Pacific Journal of Tropical Medicine. 2014;**7**(2):124-129

[32] Verma R. MRI features of Japanese encephalitis. BML Case Reports. 2012; 2012. pii: bcr0320126088

[33] Wang S, Liu Y, Guo J, Wang P, Zhang L, Xiao G, Wang W. Screening of FDA-approved drugs for inhibitors of Japanese encephalitis virus infection.

**28**

*Current Topics in Neglected Tropical Diseases*

Engels D. Neglected tropical diseases:

Flaviviridae family. Nature Reviews. Microbiology. 2018;**16**(3):125-142

[10] Huang YJ, Higgs S, Horne KM, Vanlandingham DL. Flavivirusmosquito interactions. Viruses.

[11] Raj Y, Kumar S, Haikerwal A, Goel MM, Bhatt ML, Saxena SK. Current advances in Zika virus transmission: Urgency for effective therapeutics and prevention. American Journal of Infectious Diseases. 2017;**13**(2):13-20

[12] Saxena SK, Kumar S, Maurya VK. Pathogen-associated acute encephalitis

[13] Saron WAA, Rathore APS, Ting L, Ooi EE, Low J, Abraham SN, et al. Flavivirus serocomplex cross-reactive immunity is protective by activating heterologous memory CD4 T cells. Science Advances. 2018;**4**(7):eaar4297

[14] Saito Y, Moi ML, Takeshita N, Lim CK, Shiba H, Hosono K, et al. Japanese encephalitis vaccine-facilitated dengue virus infection-enhancement antibody in adults. BMC Infectious

[15] Garg RK, Malhotra HS, Gupta A, Kumar N, Jain A. Concurrent dengue virus and Japanese encephalitis virus infection of the brain: Is it

co-infection or co-detection? Infection.

[16] Aggarwal A, Kumar P, Faridi MM.

presenting feature of dengue infection. Journal of Pediatric Neurosciences.

[17] Tiroumourougane SV, Raghava P, Srinivasan S. Japanese viral encephalitis.

Neurological manifestation as

Diseases. 2016;**16**(1):578

2012;**40**(5):589-593

2015;**10**(1):76-77

syndrome: Therapeutics and management. Future Microbiology.

2019;**14**:259-262

2014;**6**(11):4703-4730

[2] Mitra AK, Mawson AR. Neglected tropical diseases: Epidemiology and global burden. Tropical Medicine and Infectious Disease. 2017;**2**(3):pii. E36

perspectives on Flavivirus research.

Cavany SM, Perkins TA, Ebel GD, Grubaugh ND. Arbovirus coinfection and co-transmission: A neglected public health concern? PLoS Biology.

Liu Y, Zeng C, Xiao D, et al. Global epidemiology of dengue outbreaks in 1990-2015: A systematic review and meta-analysis. Frontiers in Cellular and Infection Microbiology. 2017;**7**:317

[6] World Health Organization. Dengue and severe dengue. https://www.who. int/news-room/fact-sheets/detail/ dengue-and-severe-dengue [Accessed:

[7] World Health Organization. Japanese encephalitis. https://www.who.int/ news-room/fact-sheets/detail/japaneseencephalitis [Accessed: June 01, 2019]

Eugenin EA. Flaviviruses, an expanding threat in public health: Focus on dengue, west Nile, and Japanese encephalitis virus. Journal of Neurovirology.

[9] Neufeldt CJ, Cortese M, Acosta EG, Bartenschlager R. Rewiring cellular networks by members of the

[8] Daep CA, Muñoz-Jordán JL,

2014;**20**(6):539-560

[1] Molyneux DH, Savioli L,

**References**

Progress towards addressing the chronic pandemic. Lancet. 2017;**389**(10066):312-325

[3] Holbrook MR. Historical

Viruses. 2017;**9**(5):pii. E97

[4] Vogels CBF, Rückert C,

[5] Guo C, Zhou Z, Wen Z,

2019;**17**(1):e3000130

June 01, 2019]

Journal of Virology. 2017;**91**(21):pii. e01055-17

[34] Sarala N, Paknikar S. Papaya extract to treat dengue: A novel therapeutic option? Annals of Medical and Health Sciences Research. 2014;**4**(3):320-324

[35] Hung NT. Fluid management for dengue in children. Paediatrics and International Child Health. 2012;**32**(Suppl 1):39-42

[36] Panraksa P, Ramphan S, Khongwichit S, Smith DR. Activity of andrographolide against dengue virus. Antiviral Research. 2017;**139**:69-78

[37] Saxena SK, Haikerwal A, Gadugu S, Bhatt ML. Complementary and alternative medicine in alliance with conventional medicine for dengue therapeutics and prevention. Future Virology. 2017;**12**(8):399-402

[38] Fan W, Qian S, Qian P, Li X. Antiviral activity of luteolin against Japanese encephalitis virus. Virus Research. 2016;**220**:112-116

[39] Chakraborty U, Katoch S, Sinha M, Nayak D, Khurana A, Manchanda RK, et al. Changes in viral load in different organs of Japanese encephalitis virusinfected chick embryo under the influence of belladonna 200C. Indian Journal of Research in Homoeopathy. 2018;**12**(2):75

[40] Clapham H, Wills B. Implementing a dengue vaccination programme-who, where and how? Transactions of the Royal Society of Tropical Medicine and Hygiene. 2018;**112**(8):367-368

[41] Ginsburg AS, Meghani A, Halstead SB, Yaich M. Use of the live attenuated Japanese encephalitis vaccine SA 14-14-2 in children: A review of safety and tolerability studies. Human Vaccines & Immunotherapeutics. 2017;**13**(10):2222-2231

[42] Demers J, Bewick S, Calabrese J, Fagan WF. Dynamic modelling of personal protection control strategies for vector-borne disease limits the role of diversity amplification. Journal of the Royal Society Interface. 2018;**15**(145):pii. 20180166

[43] Tangena JA, Thammavong P, Chonephetsarath S, Logan JG, Brey PT, Lindsay SW. Field evaluation of personal protection methods against outdoor-biting mosquitoes in Lao PDR. Parasites & Vectors. 2018;**11**(1):661

**31**

**Chapter 3**

**Abstract**

Diagnosis and Molecular

*Marta E. Álvarez-Argüelles, Susana Rojo Alba,* 

*Mercedes Rodríguez Pérez, Jose Antonio Boga Riveiro* 

Virus Infections

*and Santiago Melón García*

**Keywords:** chikungunya, genotype, chronic

**1. Introduction**

Characterization of Chikungunya

In recent years, large-scale outbreaks of chikungunya arbovirus (CHIKV), which is transmitted by the *Aedes* mosquito, have enabled the rapid propagation of the virus across the world. After acute infection phase with commonly fever, joint pain, headache, or rash, chronic rheumatism (arthralgia or myalgia, anorexia, and concentration disorders) up to 40% of cases is observed. The chronic form is defined by symptoms persisting for more than 3 months, and up to years, after initial diagnosis. Chronic discomfort has been linked to one of the four genotypes described. These genotypes represent different geographic lineages (classification based on partial sequence of viral E1 glycoprotein): West African, East-Central-South-African (ECSA), ECSA-diverged or Indian Ocean Lineage (IOL), and Asian. The first marker detected in CHIK infection is the viral RNA, usually by reverse transcription-polymerase chain reaction (RT-PCR). This marker can be identified in samples within 8 days of symptom onset. The infection can also be diagnosed with serological testing to detect CHIKV-specific immunoglobu-

lin IgG and/or IgM. Sequencing studies can determine the infecting genotype.

because of the incapacitating arthralgia caused by the disease.

**1.1 Epidemiology, structure, and classification of chikungunya**

Chikungunya virus (CHIKV) is an emerging mosquito-borne alphavirus that causes severe acute febrile infection, often debilitating polyarthralgias and arthritis [1], and its symptoms are similar. The name chikungunya is derived from an African language *makonde* which means "that which bends up" or "stooped walk"

Chikungunya virus, the first reported case of which was in Tanzania in 1952 [2], is endemic in certain parts of West Africa, human serosurveys having identified antibodies to chikungunya virus in 35–50% of the population in some areas [3, 4].

Since 2004 CHIKV has spread into novel locations, with outbreaks having occurred in other parts of Africa as well as Asia, Europe, and the islands of the Indian

## **Chapter 3**

*Current Topics in Neglected Tropical Diseases*

Journal of Virology. 2017;**91**(21):pii.

[35] Hung NT. Fluid management for dengue in children. Paediatrics and International Child Health.

2012;**32**(Suppl 1):39-42

[36] Panraksa P, Ramphan S,

[37] Saxena SK, Haikerwal A,

Virology. 2017;**12**(8):399-402

[38] Fan W, Qian S, Qian P, Li X. Antiviral activity of luteolin against Japanese encephalitis virus. Virus Research. 2016;**220**:112-116

[39] Chakraborty U, Katoch S, Sinha M, Nayak D, Khurana A, Manchanda RK, et al. Changes in viral load in different organs of Japanese encephalitis virusinfected chick embryo under the influence of belladonna 200C. Indian Journal of Research in Homoeopathy.

[40] Clapham H, Wills B. Implementing a dengue vaccination programme-who, where and how? Transactions of the Royal Society of Tropical Medicine and

Hygiene. 2018;**112**(8):367-368

[41] Ginsburg AS, Meghani A, Halstead SB, Yaich M. Use of the live attenuated Japanese encephalitis vaccine SA 14-14-2 in children: A review of safety and tolerability studies. Human Vaccines & Immunotherapeutics.

2017;**13**(10):2222-2231

Khongwichit S, Smith DR. Activity of andrographolide against dengue virus. Antiviral Research. 2017;**139**:69-78

Gadugu S, Bhatt ML. Complementary and alternative medicine in alliance with conventional medicine for dengue therapeutics and prevention. Future

[34] Sarala N, Paknikar S. Papaya extract to treat dengue: A novel therapeutic option? Annals of Medical and Health Sciences Research. 2014;**4**(3):320-324

[42] Demers J, Bewick S,

20180166

2018;**11**(1):661

Calabrese J, Fagan WF. Dynamic modelling of personal protection control strategies for vector-borne disease limits the role of diversity amplification. Journal of the Royal Society Interface. 2018;**15**(145):pii.

[43] Tangena JA, Thammavong P, Chonephetsarath S, Logan JG,

Brey PT, Lindsay SW. Field evaluation of personal protection methods against outdoor-biting mosquitoes in Lao PDR. Parasites & Vectors.

e01055-17

**30**

2018;**12**(2):75
