Malaria, gastrointestinal parasite infection and nutritional status among febrile children In Accra, Ghana

Introduction Malaria and intestinal parasite infection are common in developing countries. These Parasites causes anaemia and malnutrition mostly in children. For this reason, it is important to study these infections and their effects in order to monitor interventions to control them. This study aims to determine prevalence of malaria and intestinal parasite infections and their association with nutritional status among febrile children in Accra, Ghana. Methods The study was conducted among febrile children aged 6 months to 5 years attending three health facilities in Accra from May to October, 2022. A total of 315 children were selected for the study. Anthropometric measurement was done for each participant. Blood and stool samples were collected for investigation. Thick and thin blood smears stained with 10% Giemsa were prepared and examined for Plasmodium parasite using microscopy. Stool samples were processed using direct wet mount and formalin-ether concentration method and examined for intestinal parasites using microscopy. Haemoglobin concentration was measured using automatic haematology analyzer. Results A total of 24% (76/315) were positive for malaria. Plasmodium falciparum accounted for 77.6% (59/76) of parasitaemia, whereas Plasmodium malariae was 22.4% (17/76). Prevalence of intestinal parasite infection was 10.7% (34/315). Giardia lamblia accounted for 17/315 (5.3%) of the entire children, followed by Ascaris lumbricoides 8/315 (2.5%), Hookworm 6/315 (1.9%) and Trichuris trichiura 3/315 (0.9%). A total of 15/315 (5%) of the participants had co-infection of malaria and intestinal parasite infection. Prevalence of anaemia, malnutrition, stunting, wasting and underweight were (72%), (30.7%), (16.2%), (24.4%) and (57.1%) respectively. Malaria was significantly associated with anaemia (p = 0.000) and underweight (p = 0.013). Ascaris lumbricoides was significantly associated with wasting (p = 0.010). Giardia lamblia was significantly association with malnutrition (p = 0.000) and Stunting (p = 0.000), whereas Hookworm was found to be significantly associated with anaemia (p = 0.021). Conclusion Prevalence of IPI in this study was less than previously reported, most likely due to regular deworming of most of the children. However, Malaria and intestinal parasitic infection were significantly associated with anaemia and malnutrition including wasting, stunting, and underweight.


Introduction
Malaria and intestinal parasite infection are common in developing countries. These Parasites causes anaemia and malnutrition mostly in children. For this reason, it is important to study these infections and their effects in order to monitor interventions to control them. This study aims to determine prevalence of malaria and intestinal parasite infections and their association with nutritional status among febrile children in Accra, Ghana.

Methods
The study was conducted among febrile children aged 6 months to 5 years attending three health facilities in Accra from May to October, 2022. A total of 315 children were selected for the study.
Anthropometric measurement was done for each participant. Blood and stool samples were collected for investigation. Thick and thin blood smears stained with 10% Giemsa were prepared and examined for Plasmodium parasite using microscopy. Stool samples were processed using direct wet mount and formalin-ether concentration method and examined for intestinal parasites using microscopy. Haemoglobin concentration was measured using automatic haematology analyzer.

Conclusion
Prevalence of IPI in this study was less than previously reported, most likely due to regular deworming of most of the children. However, Malaria and intestinal parasitic infection were signi cantly associated with anaemia and malnutrition including wasting, stunting, and underweight.

Background
Malaria and intestinal parasite infection (IPI) are of great public health concern, causing death and morbidity in South-Saharan Africa [1,2]. Recent reports indicates that, about 627,000 and 200,000 cases of mortality due to malaria and intestinal parasite infection respectively have been recorded worldwide, with majority from West Africa [3,4]. In Ghana, these parasites are common and affect a large number of people each year [5][6][7][8]. Due to the high prevalence of malaria and IPI, these conditions have been listed among the top ve outpatient diseases in the country [7]. Furthermore, the most prevalent and severe form of malaria is caused by the parasite Plasmodium falciparum. Plasmodium falciparum accounts for more than 90% of malaria mortality in Ghana [9,10]. High morbidity has been associated with children under the age of ve due to their weak immune system [11][12][13].
Undernutrition in children is reported to be an important public health issue in Ghana [14]. Nutritional status is closely linked to immune function and infectious disease [15]. A study by Black et al., indicated that children who are malnourished can develop a number of diseases [16]. Another study conducted in the northern part of Ghana reported that malnutrition in children is a fundamental factor contributing to malaria-associated morbidity in Ghana [17]. Since malnutrition reduces immunity to infection, malnutrition makes people vulnerable to malaria [18]. Malaria reduces haemoglobin levels through enhanced destruction of parasitized red blood cells (RBCs), which results in anaemia [19,20]. Plasmodium falciparum infection causes loss of appetite, which might lead to underweight [21].
Intestinal parasite infections of public health importance in Ghana are amoebiasis, giardiasis, acariasis, hookworm, and trichuriasis [22,23,7]. These parasites can potentially cause nutritional de ciencies [24]. Ascaris lumbricoides has the potential to bind to the absorptive surfaces of the intestinal walls and affect intake of nutrients meant for the body of its host, leading to malnutrition [24,25]. Hookworm infection and trichuriasis can cause chronic blood loss, resulting in iron de ciency anaemia, whereas giardiasis and amoebiasis may cause diarrhea and malabsorption leading to stunted growth [26]. To prevent this, the Child Health Program (CHP) in Ghana recommends regular deworming of children, and this has been integrated into services delivered at child welfare clinics. However, this practice is actioned by individual parents rather than through mass treatment [27]. Thus, the effect of the intervention needs to be monitored, particularly as there have been concerns about the effectiveness of the medication used in deworming children in other parts of the world [28,29].
In Ghana, studies have established the endemicity of malaria and intestinal parasitic infections [6, 30,31]. However, research on the impact of these parasitic infection on nutritional status among children is limited [23]. There is a need to investigate the association of malnutrition and parasitic infections occurring in children. This will help provide information to aid policy on the treatment of these infections.
Therefore, this study sought to determine the prevalence of malaria and intestinal parasite infection and their association with nutritional status among febrile children in Accra.

Study design
This study was a cross-sectional study carried out at the outpatient department (OPD) and inpatient department (emergency and admission wards) of three hospitals in Accra from May 2022 to October 2022. A semi-structured questionnaire was used to collect data. Anthropometry was also measured.

Study area
This study was conducted at three hospital facilities in Accra: Princess Marie Louise Children Hospital, Shukura Hospital, and Mamprobi Hospital. Princess Marie Louise Children's hospital (PML) (5.5488 0 N, 0.2116 0 W) is located within Ashiedu Keteke sub-metropolitan district with a population of 443,768 people [32]. The hospital offers both secondary and primary care for paediatric patients from birth to 18 years old. It has 74 bed capacity and the largest nutritional rehabilitation centre in the country [14]. Shukura community hospital (5.5488 0 N, 0.2501 0 W) is one of the best medical facilities located in the Ablekuma Central Municipality in Accra. Shukura Community hospital provides medical services for people in the municipality. It is a 61-bed capacity hospital and patients are mostly self-referred. Mamprobi hospital (5.5367 0 N, 0.2421 0 W) is located in the Ablekuma South Sub Metropolitan District in Accra. The population of Ablekuma South District is 15,051 [32]. It has 54 bed capacity and over 250 patients visit per day. It offers clinical and healthcare services, including disease treatment, reproductive health, child health and nutrition to people living in the municipality.

Study population
Febrile children (body temperature > 37.5˚c) with diarrhea, from the age of 6 months to 5 years old who reported to the hospital were eligible for this study. Febrile children who had convulsion based on evidence from the medical history were excluded from the study.

Sampling method
Participants were selected from each study sites using a convenient sampling method. Selection of this method was based on study participants availability at the selected hospitals.

Data collection
In-person interviews were used to collect primary data using a semi-structured questionnaire with questions about participant's sociodemographic characteristics such as gender and age, and medical history such as clinical signs and symptoms. Weight was measured with a calibrated Seca 750 Robust mechanical oor scale to the nearest 0.1Kg for children above 2 years at all study sites. Weight was measured with a calibrated Seca 374 scale for children below 2 years at all study sites. Height measurement was taken with Seca 213 stadiometer to the nearest 0.1cm at PML and Mamprobi Hospital, and Seca 216 stadiometer to the nearest 0.1cm at Shukura Hospital for children above 2-year-old. Length was taken to the nearest 0.1cm with Seca 416 baby infantometer at PML Hospital, and Seca 210 at Mamprobi and Shukura Hospital for children below 2 years. Mid-upper arm circumference was measured with MUAC tape to the nearest 0.1cm and recorded using World Health Organization (WHO) standard [33].
The anthropometric indices: height-for-age (HA), weight-for-age (WA), weight-for-height (WH) were expressed as Z-scores using the WHO child growth standard [34]. Measurement was done by the principal investigator and assisted by trained nurses using standard techniques.

Sample collection and examinations
Blood and stool samples were collected from the participants. About 3ml of a venous blood sample was collected into test tubes containing EDTA anticoagulant using aseptic techniques for haemoglobin (Hb) measurements and malaria testing. Parents/guardians were instructed to put a teaspoon of the participant's stool into the clean, leak-proof wide-neck stool containers provided. Stool samples were processed within 6 hours of collection and examined microscopically within 30 minutes of preparation.
Both direct wet mount and formol-ether concentration technique was used to con rm the presence of intestinal parasites in stool [35].
Thin and thick blood smears were prepared, stained with 10% Giemsa, and observed under a light microscope using 40x and 100x objective lenses for malaria parasites. Malaria parasite density was calculated by comparing number of parasites against 200 leucocytes in thick lms [36]. Heamoglobin concentration was measured using automatic haematology analyzer (Mindray, Nanshan, Shenzhen, China).

Data processing and analysis
Data from interviews and parasitic investigations were entered into Microsoft Excel 2016, and exported into Statistical Package for Social Sciences (SPSS, version 26) for statistical analyses. Anaemia was de ned as Hb < 11 g/dl; severe anaemia, Hb < 7 g/dl; moderate anaemia, Hb 7 to < 10 g/dl; and mild anaemia, Hb 10-11 g/dl [37]. Children were classi ed as stunted, underweight or wasted when their HA, WA or WH Z-scores were <-2 below the reference mean, respectively (WHO, 2006). A child was identi ed as being malnourished with a Mid upper arm circumference (MUAC) measurement < 13.5cm. Moderate malnutrition was identi ed as MUAC < 12.5-11.5cm, whiles severe malnutrition was classi ed as MUAC < 11.5cm [33]. Descriptive statistics (frequencies and percentages) were used to determine the prevalence of parasitic infections. Association between parasitic infections and nutritional status was determined using chi square test and univariate logistic regression analysis and their crude odd ratio (95% con dence interval) was reported. Multiple logistic regression models were run for statistically signi cant variables to determine the signi cant risk factors. Statistical signi cance threshold was set at p ≤ 0.05.
Nevertheless, a signi cant association was observed between Giardia lamblia and malnutrition (χ2 = 4.302, df = 2, p = 0.000) ( Table 3). Prevalence of stunting, wasting and underweight among febrile children in relation to gender and age are summarized in Table 4. A signi cant association was found between malaria caused by Plasmodium falciparum and underweight (χ2 = 2.082, df = 1, p = 0.013) ( Table 4). Intestinal parasite infection caused by Giardia lamblia and Ascaris lumbricoides were signi cantly associated with stunting (χ 2 = 3.412, df = 3, p = 0.000) and wasting (χ2 = 4.052, df = 3, p = 0.010) respectively as shown in table Table 4.   an increased susceptibility to parasitic infection [39]. At the same time, children infected with parasitic infection are more likely to have nutritional de ciency [40]. This study investigated the prevalence of malaria, intestinal parasite infection and their association with nutritional status among febrile children in Accra.
Findings from this study indicated a high malaria prevalence (24%). However, this nding was contrary to a study by Kanwugu and colleagues in Tamale Metropolis, Ghana. They reported a lower malaria prevalence (2.6%) [41]. These differences could be due to the fact that the present study was hospital based and only febrile children were enrolled to participate in the study compared to theirs which was a community-based study and healthy children were included. Malaria cases were lower in children under one year, which could be due to passive transfer of maternal antibodies through the placenta [42]. A study by Journal et al., [43], also found that malaria occurred more frequently in older children aged 3years old compared to younger children of one year old.
In this study, a lower prevalence of intestinal parasite infection was observed (10.7%). These ndings differ from a study by Martinez and colleagues in overcrowded urban slums in Accra, Ghana, who reported a higher prevalence of IPI (47%) [44]. Our prevalence of 10.7% is lower than 17.3% reported by Mirisho et al. [23] at PML hospital in Accra. Another study by Gelaw and colleagues in Northwest Ethiopia, reported a higher prevalence of intestinal parasites (34.2%) [45]. The low IPI prevalence reported in our study, may have been due to the nationwide policy of regular deworming of children by the Ghana Health Service which could account for these disparities [45].
In this study, Giardia lamblia was the most prevalent intestinal parasite species. This result is consistent with a research study carried out by Anim-Baidoo et al [11]. They reported that the most dominant parasite was Giardia lamblia species (5%) of all children in their study at Princess Marie Louise Children's Hospital in Accra, Ghana. They emphasized that, assemblage B could be the predominant genotype causing giardiasis in children. Intestinal parasite infection was common among febrile children at Shukura hospital in this study. This could be due to the poor sanitation and hygiene within the community [47]. Lack of proper and clean sanitation facilities promotes open defecation practices such in the rivers and bushes which can directly contaminate water and soil.
Findings from this study revealed that more than 70% of the children were aneamic. This result is higher than 53.8% reported in Hohoe Municipality in Ghana [48] and 49.2% in Enugu, Nigeria [49]. This study provides evidence that there was an association between anaemia and age which aligns with other ndings [50,51], but in contrast to a studies by Id & Jibril [52]. In the current study, species-speci c analysis revealed a signi cant association between different parasite species and anaemia. Plasmodium falciparum and hookworm were signi cantly associated with anaemia. This nding corroborates with a study by Osazuwa et al. [53] in Nigeria. They reported an association between infection with multiple parasites and anaemia, as was shown by the high prevalence of anaemia with co-infection.
The prevalence of malnutrition in this study was 30.7%, which raises concerns for public health. Prevalence of stunting (16.2%), wasting (24.4%), and underweight (74%) among febrile children in this study emphasizes the extent of growth retardation in the studied area. Giardia lamblia was signi cantly associated with malnutrition in this study. This nding agrees with what was earlier reported by Kanokwanvimol et al. [54].
Findings of the current study revealed a signi cant association between malaria and underweight. This evidence is stronger than an earlier study conducted in Ethiopia by Hassan and Ali, [55]. Their study reported that severely underweight children had an increased risk of a clinical malaria attack, but this nding was not statistically signi cant. Intestinal parasite infection could have a serious impact on growth and development in children [56]. Djoban and colleagues in Indonesia indicated that, ascariasis causes wasting in children through lactose intolerance, malabsorption of vitamins and other micronutrients, and appetite loss that could impair growth gain. Nevertheless, ascariasis was not associated with wasting in their study [57]. In the current study, 20.5% of children with Ascaris lumbricoides infection had wasting, and a signi cant association was found between Ascaris lumbricoides and wasting. Giardiasis is a strong predictor of stunting [58]. In this study, a signi cant association was found between prevalence of Giardia lamblia and stunting. This nding was also consistent with ndings from a study by Caron and colleagues in Cambodia, which reported that giardiasis was found to be a protective factor for acute diarrhea, yet, associated with stunting [59]. Our ndings suggest that, there is a strong link between nutritional status and parasitic infection [38].

Limitation
In a few instances, it was not possible to process stool samples within the stipulated time due to distance of study sites from the laboratory, thus it is possible that some motile parasites were lost in the process, and this was a limitation to our study. The proportion of children with low-intensity infections may have been mistakenly categorized as uninfected since investigations for intestinal parasites only used one stool sample from each child which could have led to an underestimation of the prevalence of intestinal parasite infections.

Conclusion
This study revealed a lower prevalence of intestinal parasites than previously reported and that, malaria and intestinal parasitic infection are signi cantly associated with anaemia, wasting, stunting, and underweight. These results emphasize the necessity for initiatives of intervention aimed at further lowering the burden of malaria and intestinal parasite infection to reduce their impact on nutritional status of children in the research area. Thus, malaria control initiatives by Ghana Health Service should go hand in hand with effort to improve sanitation especially in areas with poor environments.

Declarations Acknowledgement
We gratefully acknowledge the support of the management and staff of Princess Marie Loise Children Hospital, Mamprobi Hospital and Shukura Hospital for allowing us access to their facility to undertake this study. We are also grateful to the study participants for being part of the study.

Author Contributions
BAD, EMAT and YAA were responsible for the study design, supervised the data collection and contributed to the writing of the manuscript. BAD performed data collection with support from EMAT and YAA. BAD, KA-O and EA-L performed laboratory analysis. Data management and analysis was done by BAD and CMO-A. BAD wrote the manuscript with contribution from EMAT, YAA. All authors read the nal version of the manuscript.
Con ict of interest.
The authors declare that they have no competing interests.

Ethical consideration
The study protocol was approved by

Funding
This study was funded by a training grant from the National Institute of Health (D43 TW 011513). The funders had no role or in uence on the design of this study, the collection, analysis, and interpretation of the data collected, as well as in writing this manuscript.

Availability of data and materials
The datasets generated and/or analyzed during the current study are available from the corresponding author on reasonable request.