Effect of enteric parasitic infection on serum trace elements and nutritional status in upper Egyptian children.

INTRODUCTION: Enteric parasitic infections still the cause of major health problems among Egyptian children as they have great morbid effect on their physical and cognitive development. Malnutrition makes children more prone to micronutrient deficiency and subsequently more vulnerable to parasitic infection. The present study aimed to identify the effect of intestinal parasitism on micronutrient serum level and children nutritional status.
MATERIALS AND METHODS: A case control study was carried out on children from 1 to 6 years old who were attending the Assiut University Children Hospital outpatient clinic, after parasitological stool examination they were divided into Group 1 (G1, n: 60) positive with enteric parasite and Group 2 (G2, n: 60) age and sex matched and free of parasites. Anthropometric measurements were expressed as weight for age (WFA), height for age (HFA), and weight for height (WFH) parameters. Serum zinc (Zn) and copper (Cu) were determined by atomic absorption spectrophotometer.
RESULTS: Intestinal parasitic infection rate was 55.7%; more commonly detected parasites were Giardia lamblia 28%, Cryptosporidium sp. 20%, and polyparasitism 18%. All children (G1 and G2) were underweight (WFA) while 63% of G1 were malnourished, either in the form of wasting (WFH) or stunting (HFA) or both aspects. Stunting and wasting were more dominant among children infected with G. lamblia and Cryptosporidium sp. and most of them were below 2 years old.
CONCLUSIONS: Coincident decrease in serum Zn level and an increase of serum Cu was more prominent among G. lamblia and Cryptosporidium sp. patients. G. lamblia and Cryptosporidium sp. were found to be more associated with nonstandard children nutritional status beside to an altered micronutrient level.

INTRODUCTION

Parasites inhabit about 2 billion people worldwide; 300 million of them experience associated severe morbidity. These infections represented >40% of the disease burden caused by all tropical diseases, excluding malaria.[1] A total of 400 million school-age children are infected; they are often physically and intellectually compromised by anemia leading to attention deficits, learning disabilities, school absenteeism and higher dropout rates.[2] Parasitic infection is a major public health problem in children worldwide, especially in developing societies where parasites are endemic as in many parts of the world.[3] Parasitic infection produces nutritional deficiency, especially among chronically infected children.[45] According to UNICEF 2000 report about 56% of the studied children in Egypt suffered from intestinal parasites. Enteric parasitic infections, with or without evident diarrhea, have intense effects on intestinal digestion and absorption. Hence, parasitic infections have indirect morbidity, ranging from malnutrition, anemia, afterward growth retardation, irritability, cognitive impairment to increase susceptibility to other infections and acute complications.[67] Trace elements as zinc (Zn), copper (Cu) and iron have a significant task in the metabolic function and tissue maintenance. Alterations in their serum concentrations are commonly found in patients with gastrointestinal parasitic infections.[89] They serve in catalytic, structural and regulatory activities in which they interact with macromolecules as enzymes, pro-hormones and biological membrane.[10] They are essential for reproduction, growth and development.[11] They are also co-factor in the development, maintenance and expression of the natural body defense system. Their deficiency leads to suppressed immunity predisposing to infections which sequentially exacerbate the nutritional status, leading to no-win situation.[12] In addition, their decline stimulates the oxidative stress which is implicated in the pathogenesis of several diseases, including cardiovascular diseases, cancer and in some parasitic infections.[1314] Growth in childhood is determined by environmental factors such as nutrition (which is influenced by infection) and illness.[15] Nutritional anthropometric measures remain the most practical and useful mean for the assessment of the nutritional status of the population particularly children. It is the single universally applicable, inexpensive and noninvasive method and reflects both health and nutrition and predicts performance.[16]

Therefore, the present study was directed to estimate the effect of intestinal parasitic infection on nutritional status and micronutrient serum level (Zn and Cu) among 1-6 years old Egyptian children.

PATIENTS AND METHODS

Three hundred children attending the outpatient clinic of Assiut University Pediatric Hospital, from November 2012 to April 2013 were examined primarily for intestinal parasitic infection.

Collection and processing of stool samples

The 300 stool samples were collected in sterile clean stool cups labeled with the patient's serial number, name, age, sex and date of collection. Parasitological examination was immediately processed within half an hour at the parasitology laboratory, Department of Medical Parasitology, Faculty of Medicine, Assiut University, Egypt.

Direct wet smear was performed using iodine and lacto-phenol cotton blue. Afterward, formalin-ethyl acetate sedimentation was done to the stool sample and examined by direct wet smear (as previous) and Modified Ziehl-Neelsen stain.[17]

The names of the infected children and the detected parasites were sent back to the pediatrician for proper treatment.

Certain inclusion and exclusion criteria were applied to enroll children in the study. Inclusion criteria included age 1-6 years old with no history of administration of any vitamin/mineral supplements for the previous 3 months. Exclusion criteria included children suffering from chronic disease.

According to their parasitological stool results and criteria (inclusion and exclusion); case-control study was conducted on 120 children (1-6 years) out of 300.

60 infected children (G1) were selected out of 167 parasitically infected ones and fulfilling the study criteria. 60 other children (G2) were selected out of 133 non infected children to meet age and sex of the infected one beside the other criteria. Children joined the study after an informed consent was taken from their parents or guardians; also ethical approval was granted by the Research and Ethic Committee of Assiut Faculty of Medicine, Assiut University.

Both groups were submitted to nutritional status assessment and serum trace elements estimation (Zn and Cu).

Anthropometric measurements

Body weight was recorded with a portable electronic scale to the nearest 100 g. Subjects were barefoot and wore minimum clothing. Height or length was measured (without shoes) using an anthropometric stadiometer (Seca) to the nearest 0.1 cm. A single person performed all measurements which were recorded as the mean of three consecutive readings. Measures were expressed as weight for age (WFA), height for age (HFA) and weight for height (WFH) according to Egyptian growth charts[18] and WHO[19] parameters. Underweight, Stunting, and wasting were defined by values below the 3rd centile line for WFA, HFA, and WFH respectively.

Biochemical estimation

Three milliliter of cubital venous blood samples were collected in sterile tubes and centrifuged at 500 g for 15 min. Sera were separated and stored at −20°C until analysis was done at the Analytical Chemistry Unit, Faculty of Science, Assiut University, Egypt. Zn and Cu concentrations of serum samples were determined by Contr AA 700 High Resolution Continuum Source Atomic Absorption Spectrometer.[20]

Statistical analysis

Statistical analysis was performed with  SPSS software package (Statistical Package for Social Sciences, version 16 for Windows). Data were expressed as mean ± standard deviation for comparison of two groups of continuous variables. Independent samples t-test and Chi-square test were used. A probability value of (P < 0.05) indicated a statistically significant difference.

RESULTS

Parasitological examination of 300 stool samples diagnosed infection in 167 patients (55.7%). The detected gastrointestinal parasite species were Giardia lamblia (cysts and trophozoites), Cryptosporidium species (oocysts), Blastocystis hominis (cysts), Entamoeba coli (cysts), Hymenolepis nana (eggs) and Isospora species (cysts) in order of frequencies [Table 1]. Monoparasitism was detected in about 82% whereas 18% showed polyparasitism. Polyparasitism was noticed commonly between G. lamblia, Cryptosporidium sp. and other parasites [Table 1].

Table 1

Prevalence of parasitic infection among the total examined (300) and infected studied children (Group 1)

Analysis of the children's anthropometric measurements revealed that about 63% of G1 were malnourished against 41% of G2 [Table 2].

Table 2

Nutritional status of infected (Group 1) and noninfected (Group 2) children

Irrespective of being infected or not, the 63 malnourished children were underweight (WFA). Wasting (WFH) was more frequent than stunting (HFA) in the studied groups [Table 3]. Stunting was more frequent in children below 2 years old while wasting was more common in children above 2 years old (statistically insignificant difference) (P < 0.05) [Table 4].

Table 3

Types of malnutrition affecting infected (Group 1) and noninfected (Group 2) children

Table 4

Relation between malnutrition and age in children

Children with mixed infection were 33% wasted nevertheless wasting and stunting were also linked to children infected with G. lamblia and Cryptosporidium [Table 5].

Table 5

Relation between species of parasite and type of malnutrition

There was a significant decrease in Zn serum level and increase in Cu serum level in G1 in contrast to G2 [Table 6]. The alteration of serum trace element (decreased Zn and increased Cu) was statistically different in male children than in female ones (P < 0.05) [Table 7]. There was no significant difference between type of malnutrition and sex of the examined children [Table 8].

Table 6

Serum level of copper and zinc in infected (Group 1) and noninfected (Group 2) children

Table 7

Serum level of copper and zinc in infected (Group 1) and noninfected (Group 2) children in relation to sex of children

Table 8

Relation between malnutrition and gender in infected and noninfected children

G. lamblia infected children had significant marked decrease in Zn level than other parasites in the study, which were associated with a nonsignificant increase in Cu serum level also (P < 0.05) [Table 9].

Table 9

Serum level of copper and zinc in relation to different parasites

Correlation of trace elements level to the most prominent type of malnutrition showed that wasted children had a significant decrease in Zn serum level and a significant increase in Cu level than nonwasted children. Stunted children also had significant decrease in Zn level and insignificant increase in Cu level than nonwasted children (P < 0.05) [Table 10].

Table 10

Serum level of copper and zinc in relation to the detected type of malnutrition

DISCUSSION

Children in developing countries especially in rural areas have high rates of parasite infestations due to poor sanitation, contact with contaminated water supply, low level of education and malnutrition.[21] The overall percentage of parasitic infections in the present study was 55.7%. Different percentages of parasites prevalence were reported among upper Egyptian children from 60% by Shalaby et al. in 1986[22] to reach 88% in 1995 in El-Gammal et al. study.[23] However in 2007 El-Masry et al.[24] stated only 38% among Rural Egyptian school children.

Malnutrition and enteric parasitic infections disturb childhood development and morbidity in many developing countries. Undernutrition may upturn susceptibility to parasitic infections which in sequence impair the nutritional status of the patients. In the present study; 63% of infected children and 41% of noninfected children had malnutrition. These results were in agreement with previous studies done by Hadju et al.,[25] Shubair et al.[26] and Quihui-Cota et al.[27] who demonstrated the correlation between nutritional status and both prevalence and intensity of intestinal parasitism in cross-sectional surveys. Parasitic infection may cause damage to full individual development, affecting even cognitive function and school performance of children.[2829] Enteric parasitic infections, with or without evident diarrhea, have intense effects on intestinal absorption and digestion. Hence parasitic infections have indirect morbidity, ranging from malnutrition, anemia, afterward growth retardation, irritability, cognitive impairment to increase susceptibility to other infections and acute complications.[67]

All the present examined malnourished children (infected and noninfected) were underweight while wasting was significantly more common than stunting. In accord with Okolo and John,[30] who reported that children with intestinal parasites were observed to be generally malnourished, with WFA and HFA indices ranging between 5th and 25th centile marks. In the present study; stunting and wasting were more to occur in children infected with G. lamblia, Cryptosporidium sp. or with both than other parasite infection. Farthing et al.[31] and Fraser et al.[32] had also reached that both parasites are often associated with growth and nutritional status deficit, especially during infancy. In the same way Carvalho-Costa et al.[33] reported that although WFA and WFH were clearly influenced by G. lamblia infection, but HFA was not. The anthropometric parameter defining chronic malnutrition was presented a borderline interaction with giardiasis on the multivariate model and a significant association with this parasite on the bivariate analysis. Data suggested that giardiasis can acutely influence weight, probably through malabsorption, but chronic infection can potentially contribute to height deficits, being possibly one of the multiple determinants of the high prevalence of stunting observed in these communities.

As regard, the relation between age and type of malnutrition in our patients; wasting was more prevalent in children below 2 years while stunting was more common in children from 2 to 6 years but the difference was statistically insignificant in both groups. This is because in short-term malnutrition, weight declines before length, so values of WFA and weight for recumbent length centiles are low compared to the length for age centile. In long term malnutrition, stunting is eventually so in addition to low WFA centile the HFA centile starts to deviate.[18]

Concerning the relation of gender to the type of malnutrition; there was no significant difference between type of malnutrition and sex of the examined children. This could be explained by that both sexes are sharing poor dietary intake and infectious diseases, which are affected by family access to water sanitation and adequate health services.

There was a significant decrease in serum level of Zn in infected children than noninfected children while there was an insignificant increase in serum level of Cu in infected children than noninfected group. These alterations in serum level of Zn and Cu were more significant with G. lamblia infection than other parasitic infections. Our results were in agreement with Schmidt et al.,[34] Bourdon and Blache,[35] and Stief.[36] This could be explained by the antagonistic effect of Zn and Zn deficiency in the patient with parasitic infection and increase Cu absorption by the gastrointestinal tract. Zn antagonizes Cu absorption by inducing the synthesis of thionein, which have a higher affinity to Cu than Zn.[37] A significant increase in serum level of Cu demonstrates that inflammation initiate free radical generation. Our results were in line with Ertan et al.[11] and Kilic et al.[38] who found that Cu level was significantly increased in patients infected with G. lamblia. Although, the mechanism is unknown, this increase in Cu level in patients with G. lamblia could possibly be explained by increase of Cu containing enzyme systems.

As regard, the significant decrease in serum Zn level in the infected group were in agreement with Ertan et al.;[11] Serarslan et al.[39] and Al-wahab et al.[40] This could be due to dietary factors (intake of food low in antioxidant) so body will use endogenous cytoplasmic superoxide dismutase (SOD) to scavenge the over production of reactive oxygen species, as a result, of parasitic infection which leads to decrease serum Zn level. Furthermore, it could be due to the antioxidant effect of Zn which plays a structural role in the maintenance of Cu-Zn-SOD structural integrity.[37] Also, our results were in agreement with Jendryczko et al.;[41] Karakas et al.;[8] Quihui et al.[42] and Abou-Shady et al.[43] who detected a significant decrease in serum Zn level in G. lamblia infected patient in comparison with other parasitically infected children. Nash and Mowatt,[44] reported that G. lamblia variant-specific surface proteins bind Zn and other heavy metals in the intestine leading to its deficiency. While Ertan et al.[11] and Roxström-Lindquist et al.[45] suggested that this decrease might be due to interference in the digestion and absorption process due to impairment and mucosal injury which by itself, could restrict the Zn absorption but also affect the activity of digestive enzymes such as lipases, proteases, and disaccharidases, beside the release of cytopathic substances that damage the intestinal epithelium.[46] It is hypothesized that the increased intestinal absorption of Zn associated with anti-Giardia treatment may be explained by the restoration of the intestinal mucosa that had been impaired by giardiasis.[47]

Significant decrease in Zn serum level associated with increased Cu level was detected in both stunted and wasted children. This could be explained by the significant increased parasitic infection in these malnourished children compared to nonwasted and nonstunted children. Although, our findings submitted the altered element metabolism due to the changes in the absorptive villous architecture that is often severely disrupted, inflamed, or destroyed by parasitic infection (mainly G. lamblia and Cryptosporidium sp.) in the malnourished or marginally nourished children, who're having the compounded problem of having inadequate or rate-limiting stores of key nutrients to repair this mucosal damage.[48]

Targeting this circle and break it up in order to diminish the serious and lifelong health complications should be considered. Therefore, this study throws the light on an immense problem in our community, which jeopardizes the health of the new generation which can be easily treated by routine diagnosis for intestinal parasite in primary health care unit. Treatment of intestinal parasitic infection would have a positive impact on the micronutrient status and on the children general health at this important preschool age and by scale up Zn supplementation programs alongside enhancement of living, social and economic conditions.