This observational cross-sectional study was conducted in the Department of Microbiology, Dhiraj General Hospital, from June 2015 to March 2020 at SBKS Medical Institute and Research Centre, Piparia, Dist- Vadodara, Gujarat, India. Approval was obtained from the Institutional Ethics Committee (IEC: no: SVIEC/ON/2015/15011) and informed consent was obtained from the patients.

Inclusion criteria: A total of 604 clinically suspected patients having fever, rigors, vomiting, nausea, abdominal pain, jaundice, headache, joint and muscle ache and other various symptoms like haemorrhagic manifestation, kidney failure, skin rash, etc were included.

Exclusion criteria: Patients who were suspected to have hospital acquired infections (in whom fever occurred after 48 hours of hospital admission) were excluded.

For detection of species, 4 mL of blood sample was collected in Ethylenediaminetetraacetic Acid (EDTA) bulb through venipuncture from suspected samples. All samples were tested with thick and thin peripheral blood smears and stained with Giemsa stain to detect Plasmodium species and simultaneously the rapid antigen detection test done by Malascan Pan/pf (Viola Diagnostic system) for co-confirmation of Malaria. Dengue infection was detected by using the Dengucheck combo rapid test system from serum/plasma to detect of NS-1 antigen and antibodies (IgM/IgG). The data including clinical diagnosis, haematological parameters were taken from the medical record department, and compared with the findings. The serological method were carried as per the manufacturer’s instructions [15].

Data for clinical diagnosis and haematological parameters were collected from hospital records; biochemical and pathological tests performed were registered. All the data were entered in excel sheet and analysis was done by software EPI info.

A total of 604 clinically suspected samples were analysed, The most affected patients were from the age group of 31-60 years (275, 45.53%) and less affected age group was 1-17 years (50, 8.28%). Male patients were more common 61.59% (372), than female 38.41% (232) [Table/Fig-1]. In 604 samples, 80 samples were found positive for dengue and 58 samples were positive for malaria. The concurrent infection of both dengue and malaria were 21 (3.47%) found and 445 samples were negative [Table/Fig-2].

In 58 malaria positive, 17 (29.31%) Plasmodium vivax, 25 (43.10%) Plasmodium falciparum, and 16 (27.59%) mixed infection with Plasmodium vivax and Plasmodium falciparum were detected. In 80 dengue positive cases, 38.75% (31) NS1, IgM 25% (20), NS1 and IgM 25% (20), IgG 5% (4), IgM and IgG 6.25% (5) were detected [Table/Fig-3].

Of the 21 with co-infection, 7 (33.33%) were severe dengue cases, 11 (52.38%) showed warning signs of dengue and 3 (14.28%) showed dengue without warning signs with malaria as co-infection [Table/Fig-4].

Out of these 21, Plasmodium falciparum was positive in 33.33% and 52.38% of Plasmodium vivax were positive and mixed infection of both Plasmodium vivax and Plasmodium falciparum were detected in 14.29% cases [Table/Fig-4].

In malaria and dengue co-infection, 52.38% (11) had hepatomegaly and jaundice and 23.80% (5) had haemorrhagic manifestation, haemoglobin was <12 g/dL in all, kidney failure was found in 4.76% (1), thrombocytopenia (platelet count <150,000/cmm) in 95.23% (20) and condition also more common in Plasmodium vivax infections. No death was detected in dengue and malaria co-infection cases [Table/Fig-5,6].

Dengue and malaria are common vector-borne diseases but are preventable. Both diseases have clinically similar features and symptoms however, they are separate and given different treatment. Consecutive presence of malaria and dengue in one individual can easily be missed as the detection of any one of them in an acute febrile patient can cover the diagnosis of other [16]. In this study, the occurrence of concurrent infections of 3.47% were found. Other studies have comparatively different prevalence of concurrent infection, they have found rate as of 6% in India, French Guiana had 1% and Pakistan had 27% [17-20]. In a study by Abrahamsen SK et al., from Karnataka, 100 patients of acute febrile illness were diagnosed with dengue (25%); malaria (8.0%), and enteric fever (14%) [21].

A study by Singh R et al., in a retrospective review of 160 patients reported 23% malaria and 2% dengue fever while up to 54% died because of unexplained fever due to acute febrile illness in Mumbai, India and 18% patients were noted to have mixed infection [22]. More than one aetiological agent with concurrent infections leads to illness with overlapping symptoms and hence, patients diagnosis and treatment could be challenging [23]. There was typical conception of malaria incidence in rural spot and in urban region dengue was found in various reports which were coming from different countries because of overlapping of mosquito biotypes [24]. In this study, only hospitalised patients were included and the co-infection incidence does not represent prevalence in the community or local population. The concurrent infection in a locality was determined but the vector load could not determined [25,26].

The diagnosis of dengue infection was positive by the IgM test, but the acute and past infection cannot be differentiated because IgM can persist for one week after the onset of the symptom, as reported in case reports and cross-reaction with other arboviruses [27,28]. A patient at one time can be diagnosed both with malaria and dengue after five days of fever, and can be positive when tested with dengue rapid kit and malaria rapid kit. Patients have also been infected with both concurrent DENV and Plasmodium parasite infections in the present study. Some other possibility for both infections was that patients were previously infected with the serum IgM DENV and then infection with the malaria parasite.

The patients who have dengue infection; have low immunity for the period of convalescence which makes it predisposed to other infections. If dengue infection shows no signs of improvement in conservative treatment beyond one week of fever, clinicians should consider other co-infection possibilities, prominently malaria. The present study diagnosed Plasmodium vivax in 52.38% which was different from other studies [25,26]. However, it attributed Plasmodium species prevailing in an exacting ecological region. Both malaria and dengue co-infection have clinical symptoms which are more possible to single dengue than only malaria infection. So, clinical examination of both concomitant dengue and malaria is complicated. The present study found complications in patients infected with P.vivax that were common in malaria and dengue co-infection with haemorrhagic manifestations of five cases, enlargement of liver and jaundice was in 11 cases. In Plasmodium vivax infections, the main investigations found haemoglobin <12 g/dL, and 95.23% had decreased platelet count. In P.falciparum infection, altered liver and renal function tests were observed in co-infections. Deranged liver function was also found in one study [26].

The condition is remarkable to significance that haemorrhagic manifestations are frequent in dengue cases and rare in falciparum malaria. The low levels of platelets can be caused by both malaria and dengue and hence, it can be complicating to choose which one is responsible for the haemorrhage. However, malaria with bleeding is considered as severe malaria and treated accordingly [27].

This study was hospital based and such work in community may give better idea of malaria /dengue dual infections so its results were supposed to be interpreted with warning. If the very low prevalence of dual infection was such questionable, prospective studies through similar investigation methods and patient groups must be tried to verify the superior cruelty of co-infection. The benign outcome has also been observed in the other two studies [25,26]. A positive outcome was attributed to the early medical treatment of co-infection cases [29,30].

There were three distinguished results showed in the study, both malaria and dengue co-infection which were mosquito vectors co-exist and they are common in a geographical region was the first result. Second, the clinical symptoms of dengue fever are predominant more than malaria in concurrent infection. Third, P. falciparum is concerned with severe dengue and the warning signs were more in patients infected with both dengue and malaria infections. WHO classified the dengue cases based on a study of 80 dengue positive patients, dengue without warning symptoms, dengue with warning symptoms and severe dengue were respectively 42, 28 and 10 cases were found. In this study, 11 showed dengue with symptoms, severe dengue cases were 7 and less cases of dengue without warning signs was 3 cases [30,31].

In the present study, concurrent infections of malaria and dengue must be suspected in febrile patients diagnosed with both malaria and dengue, whether one or the other is positive. The study indicated that both dengue and malaria prevalence rates of co-infection are correlated with symptoms and haematological parameters, as well as the severity of such co-infections.

This was a hospital based study and co-infection of malaria and dengue does not reflect the community prevalence.

All the patients having fever must be diagnosed for both malaria and dengue. Moreover, when both co-infection occurs in one patient it can cause other complications as well. When patients is febrile or returning from endemic region the clinicians treat should treat thoroughly and order examinations for both malaria and dengue diagnoses. The vector control is an important preventive measure in the populations and also proper hygiene, public health education are the preventive measures that are necessary to control these both diseases.