High rates of co-infection of Dengue and Chikungunya virus in Odisha and Maharashtra, India during 2013

Highlights

• Detection of 38.23% of DENV and 27.94% of CHIKV during 2013 outbreak in Odisha.

• Detection of 44.8% co-infection of CHIKV and DENV in collected clinical samples.

• All CHIKV strains were ECSA type and DENV serotype 2 was predominant in the samples.

• Two consistent mutations in E1 gene in almost all co-infected strains of CHIKV.

• Five consistent mutations in serotype 1 in C-prM gene of DENV.

Abstract

Dengue viral (DENV) infection is endemic in different parts of India and because of similar primary signs and symptoms, Chikungunya virus (CHIKV) is mostly undiagnosed. Hence, we investigated 204 suspected Dengue cases in a hospital based cross-sectional study in Odisha, India in 2013. It was observed that 50 samples were positive for DENV only, 28 were positive for CHIKV only and interestingly, 28 patients were co-infected with both DENV and CHIKV. Additionally, a total of 18 confirmed Dengue samples from Maharashtra, India were screened for CHIKV and out of those, 15 were co-infected. All CHIKV strains were of East Central South African (ECSA) type and serotype 2 (genotype IV) was predominant in the DENV samples. Additionally, Dengue serotype 1 and 3 were also detected during this time. Further, sequence analysis of E1 gene of CHIKV strains revealed that two substitution mutations (M269V and D284E) were observed in almost 50% strains and they were from co-infected patients. Similarly, sequence analysis of C-prM gene showed the presence of five substitution mutations, (G70S, L72F, N90S, S93N and I150L) in all serotype 1 and two consistent mutations (A101V and V112A) in serotype 2 Dengue samples. Together, it appears that a significantly high number of dengue patients (43, 44.8%) were co-infected with DENV and CHIKV during this study. This emphasizes the need of a routine diagnosis of CHIKV along with DENV for febrile patients. This will be useful in early and proper recognition of infecting pathogen to study the correlation of clinical symptoms with single or co-infection which will ultimately help to implement proper patient care in future.

Introduction

The Dengue virus (DENV) belongs to genus Flavivirus, under the family Flaviviridae and there are four serotypes which cause wide spectrum of illness from asymptomatic to severe fatal Dengue Hemorrhagic Fever or Dengue Shock Syndrome (DHF/DSS) (Halstead, 2007). All four serotypes of DENV can cause all above mentioned clinical manifestations. Infection with one serotype of DENV delivers lifelong immunity to subsequent infection by the same serotype, but, it does not confer strong immunity against infection with other serotypes of DENV. Hence, an individual with severe problem (DHF/DSS) is thought to be associated with secondary or tertiary infection (Green and Rothman, 2006). DENV was reported almost simultaneously in Asia, Africa, North America in 1780s and after that major clinical cases were reported in 1953–54 in Philippines followed by global spread of DENV outbreaks (Gubler, 1998). In every year, 50–100 million cases of Dengue fever and 250,000 to 550,000 DHF cases are found in more than 100 countries and the incidences have increased around 30-fold in last 50 years (Hales et al., 2002, World Health Organization, 2009).

On the other hand, Chikungunya virus (CHIKV) is an Alphavirus, that belongs to Togaviridae family (Strauss and Strauss, 1986) and the disease is characterized by abrupt onset of high fever, arthralgia, myalgia, headache, rash and the typical clinical sign is poly-arthralgia which is very painful and may persist for several months (Jupp PG, 1988). Sometimes, prolonged rheumatoid symptoms (Sissoko et al., 2009) or neurological complications (Chandak et al., 2009, Robin et al., 2008) can also be associated with this infection. CHIKV was first isolated in Tanzania, Africa in 1952 (Sarkar et al., 1964). Since then CHIKV has been identified in nearly 40 countries in Asia, Africa, Europe and America and recently several million cases have been observed in the Indian Ocean Island and many other regions of the world (World Health Organization, 2014). It was also observed in India after a gap of 32 years. During the outbreak, around 1.3 million people were affected in almost 13 states of India including Odisha (Dash et al., 2007).

These two viruses are transmitted by the same vectors, Aedes aegypti and Aedes albopictus mosquitoes and in both the cases viral infection has similar primary signs and symptoms in the patients (Gubler, 1998). Therefore, it is difficult to recognize individual infection or co-infection among the patients. The cumulative arboviral disease burden has sharply increased in recent time (Dash et al., 2013) and generally the outbreak of Chikungunya is neglected in the Dengue hyper-endemic regions. CHIKV and DENV have emerged as important arboviral infections and also their co-circulation in the same region or co-infection with more than one virus in the same patient has made the problem more complex globally (Gandhi et al., 2015). As a result, co-circulation of both the viruses has been reported in India, Sri Lanka, Malaysia, Indonesia, Gabon, Cameroon, Madagascar and Thailand (Chang et al., 2010). In addition, concurrent infection of DENV and CHIKV in patients have been reported in India, Yemen, Singapore, Sri Lanka, Malaysia etc. (Chang et al., 2010, Hapuarachchi et al., 2008, Nayar et al., 2007, Rezza et al., 2014, Singh et al., 2012).

Accordingly, in the current study, an investigation was carried out to understand the status of DENV and CHIKV infection in a hospital based cross sectional study where patients from Odisha (Eastern part), India with suspected viral fevers were screened for DENV and CHIKV during a dengue outbreak in 2013. Few confirmed DENV cases from Maharashtra (Western part), India were also included in this study to look for DENV–CHIKV co-infection. Further, few selected viral strains were sequenced and analyzed for understanding the phylogenetic relationship and identifying the mutations in the strains.