Seroprevalence of dengue virus among children presenting with febrile illness in some public health facilities in Cameroon

Introduction A routine diagnosis of Dengue virus (DENV) infection is not usually conducted in hospitals. Because symptoms overlap, many potential febrile illnesses due to DENV may be confused for malaria, typhoid or paratyphoid (enteric) fever. The absence of data on DENV exposure rates among children attending health facilities could undermine management of this disease. This study aimed to investigate the seroprevalence of dengue virus infection in children presenting febrile illness in some public health facilities in Cameroon. Methods A cross-sectional study was performed in children ≤ 15 years attending seven urban and three semi-urban public hospitals of Cameroon. From each volunteer, 2ml of whole blood was collected and tested for dengue virus IgM, malaria (Pf/Pan antigens) and enteric fever (Typhoid IgM) using rapid diagnostic tests (RDT); in order to allow the healthcare workers to quickly put the positive cases under appropriate treatment. Positive cases of dengue virus infection were confirmed by indirect ELISA. Data analysis were performed using the statistical package for social sciences software, version 22.1. Results A total of 961 children were enrolled in the study and 492 (51.2%) were infected with at least one of the three pathogens. Overall, DENV IgM seroprevalence among febrile children was 14.4% (138/961). About 390 (40.6%) and 22 (2.3%) had malaria (Pf/Pan Ag) and enteric fever (Typhoid IgM) respectively. Co-infection with dengue virus was found in 51 (5.3%) participants. The dengue virus IgM seroprevalence was higher in Bankim (19.3%), Ntui (18.3%) and Douala (18.2%). Conclusion Dengue virus infection seroprevalence appears to be low in children presenting with febrile illness in the studied health centres in Cameroon but call for more attention and research to further characterise the circulating strains of the dengue virus.


Introduction
Dengue is a tropical disease transmitted by mosquitoes of the genus Aedes [1,2]. Infection by dengue virus causes flu-like illness and can occasionally develop to severe complications and death.
Dengue has grown dramatically over the years and it is estimated that about half of the world population is now at risk [3]. Dengue is a leading cause of serious illness and death among children in Asia and Latin America [3]. Yet the real number of dengue cases is underreported and many cases are misclassified [3]. In Africa, at least 15 countries declared locally acquired dengue cases since 1960.
Moreover, dengue has frequently been detected in travellers returning from over 30 African countries. In Central African region, dengue is highly prevalent and cases of dengue infection have been reported in recent dengue-like symptoms outbreaks in Cameroon and Gabon [4][5][6]. A study conducted in three major towns in Cameroon by Demanou et al. [6], reported dengue virus IgM seroprevalence of 0.3, 0.1 and 0.0% among healthy children in Douala, Garoua and Yaounde respectively but no known study has described DENV IgM seroprevalence among febrile children in Cameroon. WHO gives priority to malaria when considering the aetiology of fever in tropical countries [7].
It is therefore recommended to think of malaria diagnosis in the first place when feverish syndromes are observed in patients with empirical treatment of malaria administered in sufficient dose and duration. Like malaria or enteric fever, dengue infection symptoms include fever headache, rash, vomit and joint pain [8,9]. Fever accounts for 70% of purposes of consultation for children visiting health care facilities in Cameroon [10]. More than 80% of primary infection of dengue virus in children are usually asymptomatic and are generally characterized by a fever greater than 38°C in addition to other symptoms identical to those of malaria or enteric fever [11,12]. A particular feature for dengue fever is that its re-infections can be associated with haemorrhage or shock syndromes in patients [2,6,[12][13][14][15][16][17][18][19]. Children under the age of 15 are particularly at risk of developing a severe form of dengue fever (DF) as well as any patient who had already been infected by another serotype of the virus [20].
Most arboviruses especially DENV are rarely taken into consideration by local clinicians because the disease is not considered as endemic and the diagnosis is always focused on other endemic diseases such as enteric fever and malaria. However, early diagnosis of DF by a rapid diagnostic test for the detection of immunoglobulin M (IgM) and non-structural protein 1 (NS1) is important for preventing potential complications in children [21,22] and also to limit the over-consumption of anti-malarial drugs and antibiotics by patients who do not need them. This could also help in slowing down the development and emergence of resistance to antibiotics and antimalarial drugs. years regardless of gender, ethnicity or tribe. However, we excluded children admitted at the hospital for known diseases or with fever more than seven days.

Methods
Description of study sites: the study was conducted in ten areas Patient information and informed consent: the risks, if any and benefits of the study participation were explained to the patient during the administration of informed consent. Written consent was obtained from the parents or guardians or legal representative. In case of patients between 10-14 years, assent was also obtained.
Blood sampling: two millilitres (2ml) of venous blood was collected by venipuncture from children and dispensed in EDTA tubes and centrifuged at 4000 rpm for 10 min for rapid diagnostic test of enteric fever, malaria, dengue markers (IgM) and IgM ELISA assay. The plasma fraction was collected, aliquoted in small, single used volumes and stored for at -20° for subsequent analyses.

Results
Characteristics of study populations: a total of 961 consenting children attending hospital for febrile illness and for whom the clinician suspected malaria and/or typhoid fever were included in the study. The minimum number of children recruited per site was 60 in Bafia and the highest was 150 in both Kaele and Bankim (Table 1).

Discussion
Dengue virus is an important emerging disease of the tropical and sub-tropical country today. The differential diagnosis associated with dengue fever includes a wide variety of parasitic and bacterial infections that produce similar syndrome. This study investigated the seroprevalence of dengue among children presenting with undifferentiated fever who are living in ten areas of Cameroon.
Laboratory diagnosis of acute infection (dengue, enteric fever or malaria) is important to provide appropriate treatment and to reduce development and emergence of resistance to antibiotics and antimalarial drugs. In malaria endemic areas like in Cameroon, most febrile illnesses are treated as malaria or enteric fever cases due to lack of trained staff and laboratory to properly rule-out malaria or enteric fever as a cause of fever and as well, identify alternative fevercausing pathogens [14,26,27]. prevalence was reported [30,31]. We observed few positive cases to dengue IgM (7%, 16% and 14% respectively) among the various age group of children febrile illness (less than 1 year, 1-5 years and 6-15 years old respectively). According to WHO recommendations [32], the diagnosis of dengue virus can be confirmed when at least IgM is positive during seroconversion in paired sera. As previously reported in some studies [4,6,8,33] dengue is endemic in Cameroon but this study is the first carried out among children with febrile illness.
Dengue virus IgM biomarkers were detected in all age groups. This suggests that children were recently bitten by Aedes mosquitoes during the day. However, several authors suggested transmission of antibodies or Dengue virus from mother to child through breastfeeding [34][35][36].
Transmission of DENV in several semi-urban and urban regions ( Figure 1) is an indication that it is widespread in urban and semiurban populations. Human-DENV-Mosquito cycle has been reportedly found in nearly all urban and semi-urban environments throughout the tropics and subtropics [37]. Dengue in tropical and subtropical regions has increased due to uncontrolled urbanization, lack of effective and sustainable vector control programs [38].

Conclusion
We have reported seroprevalence of dengue virus in fever illness children, from some health facilities in Cameroon for the first time.
Our study has revealed that dengue prevalence is low in children presenting with febrile illness in Cameroon. However, getting a realistic prevalence would demand a more representative sample size