Co-infections with Plasmodium falciparum, Schistosoma mansoni and intestinal helminths among schoolchildren in endemic areas of northwestern Tanzania

Background Malaria, schistosomiasis and intestinal helminth infections are causes of high morbidity in most tropical parts of the world. Even though these infections often co-exist, most studies focus on individual diseases. In the present study, we investigated the prevalence of Plasmodium falciparum-malaria, intestinal schistosomiasis, soil-transmitted helminth infections, and the respective co-infections, among schoolchildren in northwest Tanzania. Methods A cross sectional study was conducted among schoolchildren living in villages located close to the shores of Lake Victoria. The Kato Katz technique was employed to screen faecal samples for S. mansoni and soil-transmitted helminth eggs. Giemsa stained thick and thin blood smears were analysed for the presence of malaria parasites. Results Of the 400 children included in the study, 218 (54.5%) were infected with a single parasite species, 116 (29%) with two or more species, and 66 (16.5%) had no infection. The prevalences of P. falciparum and S. mansoni were 13.5% (95% CI, 10.2-16.8), and 64.3% (95% CI, 59.6-68.9) respectively. Prevalence of hookworm infection was 38% (95% CI, 33.2-42.8). A. lumbricoides and T. trichiura were not detected. Of the children 26.5% (95% CI, 21.9-30.6) that harbored two parasite species, combination of S. mansoni and hookworm co-infections was the most common (69%). Prevalence of S. mansoni - P. falciparum co-infections was 22.6% (95%CI, 15.3-31.3) and that of hookworm - P. falciparum co-infections 5.7% (95%CI, 2.6-12.8). Prevalence of co-infection of P. falciparum, S. mansoni and hookworm was 2.8% (95%CI, 1.15-4.4). Conclusion Multiple parasitic infections are common among schoolchildren in rural northwest Tanzania. These findings can be used for the design and implementation of sound intervention strategies to mitigate morbidity and co-morbidity.


Background
Parasitic infections present a major cause of disease and morbidity in Africa [1]. Among these diseases, Plasmodium falciparum inflicts the largest burden [2,3]. It is estimated that 40% of the world's population is at risk of malaria, and about 90% of the infected populations with malaria live in sub-Saharan Africa [4]. In Tanzania, over 95% of the 38.7 million people are at risk for malaria; the disease contributes to 39% -48% of all outpatients [5].
Schistosomiasis is endemic in 76 countries worldwide and besides malaria is the second important parasitic disease for public health [6,7]. Of the 662 million people infected worldwide, 85% are from Africa [8]. In Tanzania, schistosomiasis is highly endemic and its prevalence varies from one region to another, with a prevalence of up to 80% in highly endemic areas [9].
The World Health Organization estimates that more than one billion people are chronically infected with soil-transmitted helminths [10]. Hookworms are estimated to affect 1300 million people [11].
Due to the fact that P. falciparum malaria and helminth infections are endemic throughout Tanzania, the affected populations often endure infections with a number of different species [12], and individuals are commonly coinfected with combinations of helminths and malaria parasites [13]. Such infections may have considerable health consequences, leading to more severe clinical symptoms and pathology than for infection with single parasite species. Interaction of malaria and helminth infections increase the severity of anaemia and organomegaly observed in schoolchildren and thus may potentially create a great challenge for disease control in the tropics. There is also evidence that co-infection with multiple parasites may alter the immune responses [14].
Overlap of schistosomes, soil-transmitted helminth and P. falciparum malaria depends on the conditions that favour multiple parasitic species survival and transmission [15]. These conditions include poverty, environmental contamination, water bodies and lack of effective preventive measures [15]. Polyparasitism of P. falciparum malaria, schistosomiasis and soil-transmitted helminth have been reported from various epidemiological settings in Africa [12,16,17], yet most parasitic diseases are still studied individually and data on prevalence, morbidity and mortality associated with multiple parasitic infections in Tanzania are limited.
This study was therefore conducted to determine the prevalence of P. falciparum malaria, S. mansoni and soiltransmitted helminth, and co-infections, among schoolchildren living in villages surrounding the Lake Victoria shore. Understanding the extent of polyparasitism in high-risk groups will serve as a guide in developing sound interventions strategies to reduce the burden of these diseases and co-morbidity.

Study area and population
Nyamatongo ward is located in Sengerema district in the Mwanza region, northwest Tanzania. The ward is located on the eastern side of the district and boarders Lake Victoria to the east. It has a population of about 21,000. The study area is located at 1140 m altitude above sea level, and the inland area is covered by seasonal rivers and stream flowing down to the lake. The area is characterized by a tropical climate, with an average annual temperature of 26.5°C, and experiences long rainy seasons between January and May. Annual average rainfall is 1065 mm. The majority of inhabitants are involved in rural subsistence farming, fishing and livestock keeping.
The study was carried out between March -May 2009 among 400 schoolchildren from four primary schools, located 500 m-1500 m from the shore of Lake Victoria in Sengerema district, northwest Tanzania. The inclusion criteria for the study were: (1) children aged 8-16 years; (2) parents or guardians gave written informed consent; (3) in addition to the written consent from caretakers, children also were supposed to agree and provide informed assent; (4) children lived in the study area.

Study design
A cross-sectional study was undertaken to determine the prevalence of P. falciparum malaria, S. mansoni and soiltransmitted helminth and any co-infections of P. falciparum, and intestinal helminths among schoolchildren. A two-step random sampling method was used to select children participating in the study. First, four schools of Nyamatongo ward were selected randomly. Second, using the class attendance registers, each child was given a number. Selection of the study participants was achieved by using tables of random numbers. Demographic data including age, gender and school/village where participants studied/lived were recorded by a structured questionnaire.

Parasitological examination of S. mansoni and soiltransmitted helminths
The day before parasitological screening commenced, schoolchildren were issued with plastic containers and instructed by a research team (HDM and NJSL) on how to collect a portion of their morning stool samples the next day. Stool containers were then collected at the school and labeled with identification numbers. These samples were brought to the field laboratory on the same day, and duplicate Kato-Katz cellophane thick smears was prepared from each specimen [18,19]. As a quality control measure, 10% of randomly selected smears were reexamined by a third experienced parasitologist who was blinded of the previous results. The mean number of eggs from each Kato Katz thick smear were multiplied by 24 in order to express infection intensities as the number of eggs per gram of faeces [18].

Parasitological examination of malaria parasites
A finger prick blood sample was collected after cleaning the finger surface using a sterile cotton wool soaked in methylated spirit. Two thick and one thin smears were prepared and stained with 10% Giemsa (Sigma, Aldrich, Nairobi) in phosphate buffer [18]. Species specific and parasite densities were estimated under a light microscope at high magnification by counting the number of parasites per 200 white blood cells (WBC). If < 10 parasites were found the reading was continued up to 500 WBC. The presence of either ring forms or gametocytes was conclusive diagnosis of P. falciparum. The counts of P. falciparum were converted to the number of parasite per μl of blood, assuming as standard a WBC count of 8000/μl [20].

Treatment
Children found positive for hookworm and S. mansoni infections were treated with mebendazole 500 mg and praziquantel 40 mg/kg [21]. Sweet potatoes and porridge were given before tablets in order to reduce the nauseating effects of praziquantel. Children positive for malaria parasites and body temperature above 37°C were treated with combination therapy of lumefantrine and artemether in collaboration with nearby health centers within the village.

Ethical considerations
The study was approved by the Institutional Review Board (Research and Publication Committee, certificate no. BREC/001/03/2009) of the Weill-Bugando University College Health Sciences and Bugando Medical Centre, Mwanza, Tanzania.
Prior to conducting the study, meetings were held with parents or guardians, teachers and community leaders to explain the aims and procedures to be used to collect data. Informed written consents were obtained from children's parents or guardians and in addition assent was subsequently obtained from children.
The prevalence of single and multiple species of parasites were assessed and classified by gender and three age groups (≤ 10, 11 -13 and 14-16 years). Chi-squared (X 2 ) test was used to assess whether single and double or triple parasitic infections were associated with sex and age. Logistic regression analysis was applied to investigate whether gender and age were significantly associated with S. mansoni, P. falciparum and hookworms. A model of with S. mansoni infected children defined as cases incorporated age and sex. The adjusted odds ratios, including 95% confidence interval and P-values were calculated. Similar procedures were repeated for hookworm and P. falciparum infections.

Results
A total of 400 schoolchildren aged 8-16 years were included. The age and sex distribution of study participants is shown in Table 1.

Discussion
The findings from the current study confirm that Nyamatongo ward in northwest Tanzania is highly endemic for intestinal schistosomiasis and hookworm infections. S. mansoni was the most prevalent parasitic disease identified with almost 2/3 of the schoolchildren infected. Prevalence of malaria was relatively low. Parasitic coinfections were common. Ascariasis and trichuriasis were absent.
Earlier studies conducted on other localities within the lake basin showed that both schistosomiasis and hookworm infections are common in the area [9,23]. However, prevalences in earlier studies were much lower (10.9%-S. mansoni) than in the present study probably due to focal exposure to S. mansoni by the population living in endemic areas as reported previously [24,25]. Infection intensities for S. mansoni in children were light to moderate, and only 7.5% of the children had heavy infections. These findings support previous observations that only few individuals in an endemic community excrete large numbers of eggs and that S. mansoni infections starts at early ages [26]. In the current study, children < 13 years recorded highest infection rates with S. mansoni. Similar observations have been made for other areas [27]. On the other hand, the prevalence of S. mansoni was observed to decrease with increase in age of the children, which has also been reported in previous reports from Cote d'Ivoire [27]. The decrease in prevalence of S. mansoni appears to coincide with the onset of puberty [25]. The prevalence of hookworm infections observed in the current study area (38%) was similar to Magu district (37%) within northwest Tanzania [9] but slightly lower than reported from Western Kenya (42.5%) [25] and Brazil [28].
Our data confirm that among soil-transmitted helminths, hookworm infection is the major public health problem within the Lake basin [9,25]. A positive association between older age group (14-16 years) and hookworm infection was observed. A similar observation was made in locality in rural Côte d'Ivoire [27]. Hookworms were more widespread as compared to other soil-transmitted helminth infections, namely A. lumbricoides and T. trichiura, which occur in various ecological settings of sub-Saharan African [29]. Interestingly, earlier studies in Magu district, northwest Tanzania, reported a prevalence of ascariasis of < 1% [9] and a report by Handzel et al. [25] in Kenya's Nyanza province reported a prevalence of 22.9% and 17.9% for A. lumbricoides and T. trichuris. Variability in endemicity or prevalence of these infections, low sensitivity of the diagnostic method, the use of single stool sample, environmental contamination and inability of the helminth eggs to withstand high temperature could partly explain the observed difference. The prevalence of P. falciparum malaria in Nyamatongo ward was relatively low (13.5%) in spite of the locality being within a malaria holoendemic area [30]. Our findings confirm other studies conducted in Tanzania [31,32], but are different from data among primary schoolchildren in holoendemic areas of Cameroon [33].
This difference could be attributed to low malaria prevalence in the study area or high mosquito net coverage which was not assessed in the present study. A particular concern is the high occurrence of parasitic co-infections, with concomitant infections of S. mansoni and hookworms being the most common. Co-infections of helminth and P. falciparum infections also have clinical importance [34]. Previous studies have clearly documented the relationship between intestinal helminth infections, polyparasitism and cognitive functions, growth and malnutrition among school children [35,36]. Hookworm anemia may be exacerbated by infection with other parasites [29]. Children with multiple parasitic infections especially those with heavy infections intensity tend to experience more severe cognitive outcomes and other health problems such as malnutrition than children with only one helminth infection [36,37]. Studies suggest that co-infections of P. falciparum with hookworms or schistosomes tend to exacerbate hepato-splenic, anaemia and malnutrition morbidities among school children [38]. Co-infections of S. mansoni and hookworm could partly be attributed to the co-endemicity of the two species in the study area and poor sanitations [28,39,40]. This observation is similar to previously reports from Brazil [28] Côte d'Ivoire [27] and China [40]. The polyparasitism of S. mansoni and P. falciparum observed in the present study have also been observed previously from Senegal [41] and Zimbabwe [42] where prevalence of P. falciparum among schoolchildren were high in those infected with S. mansoni. One reason for this observation could be the availability of breeding sites for the intermediate host (fresh water snail) and malaria vectors (Anopheles mosquitoes) in the study area. Furthermore, heavy intensities of S. mansoni infection did not correlate with having heavy intensities of P. falciparum. The findings of our study were similar to a study from Uganda [38].
Hookworm and P. falciparum co-infections were more common in our study than reports from Cameroon (0.2%) [33] but lower than in Zimbabwe (12.6%) and Western Kenya (41.5%) [25,43]. Difference in geographical variations in exposure and endemicity could be one of the reasons for these differences. However, in this study, P. falciparum infection showed no significant association with hookworms.
Similar to our results, triple infections of P. falciparum, hookworm and S. mansoni have been reported in studies from Côte d'Ivore, Cameroon and Zimbabwe [39,33,42]. These studies conducted throughout Africa and China indicate clearly that most parasitic infections do not occur singly but as co-infections [28,29,39,40]. Although both malaria and helminths have distinct means of transmission patterns, a variety of environmental and host factors may influence their epidemiological and geographical patterns of infections and diseases [34]. In conclusion, this study revealed that S. mansoni, hookworm and P. falciparum are prevalent among schoolchildren in the study area. The findings of the present study, supports the need for initiatives to implement a new framework for an integrated approach in disease management. We also recommend longitudinal studies to identify the associations between parasites and associated morbidity.