Anemia among HIV Infected Children on Highly Active Anti Retroviral Therapy in Wolaita Zone, South Ethiopia: a facility based cross sectional study

Background Anemia is a global public health problem and most of the HIV positive people become anemic at some point in the course of the disease. We lack adequate evidence on the magnitude of anemia among children on highly active anti retroviral therapy in Ethiopia and particularly in South Ethiopia. Thus, this study aimed at determining the proportion and associated factors of anemia among children on highly active anti retroviral therapy in Wolaita zone, South Ethiopia. Method a facility based cross-sectional study has been conducted from November to December 2018 on 256 children 6 months to 14 years of age who were on Anti retroviral therapy. Data were collected through an interview with caregivers and review of medical records. CD4+ cell count was analyzed using FACS caliber and Hemoglobin level was measured with Hem cue 301 analyzer. Stool sample was examined for presence of intestinal parasites by direct wet mount technique. Data analyzed with Stata version 14.0 were conveyed in mean and standard deviation of the mean, median and inter-quartile range. Multivariate analysis was carried out to identify independent predictors of the outcome variable. Adjusted Odds Ratio with 95% confidence interval was reported. HIV: ROC:


Background
Anemia is a condition in which the number of red blood cells is insufficient to meet the body's physiologic needs and consequently diminish the oxygen-carrying capacity. In addition to iron deficiency, other nutritional deficiencies, acute and chronic inflammations, parasitic infections, and inherited or acquired disorders that affect hemoglobin synthesis, red blood cell production or survival can cause anemia [1].
Children with Human Immunodeficiency Virus (HIV) are exposed to macro and micro nutrient deficiencies; anemia being the commonest [2,3]. Anemia is a critical health concern affecting growth and energy levels adversely, and it highly damages the immune mechanisms [4,5].
Anemia affects both developing and developed countries and thus it is a global public health problem [6]. Globally, roughly 1.3 to 2.2 billion people suffer from anemia and in developing countries 50% of women and children are anemic [7]. An estimated 63%-95% of people living with HIV become anemic at some point in the course of the disease [8]. In India the prevalence of anemia among HIV infected children was reported to be 47.1% [9].
A prevalence level of 69.1% was reported among HIV infected children in Democratic Republic of Congo [10]. In Ethiopia the prevalence of anemia among HIV infected children ranges between 16.2% and 39.1% [11,12,13]. A combination of environmental and host factors predispose HIV infected children to anemia.
We have very limited evidence on the prevalence of anemia among children on Highly Active Anti Retroviral Therapy (HAART) in Ethiopia and there is a wide variation in the reported prevalence. Thus the main aim of this study was to estimate the proportion of anemia among HIV infected children on HAART and to identify factors associated with the outcome in Anti Retroviral Therapy (ART) clinics of Wolaita zone, South Ethiopia.

Study Setting
A facility based cross-sectional study was conducted from November to December 2018 at ART Clinics of Wolaita zone. There are 16 health facilities, 12 health centers and 4 hospitals, which provide ART service. Wolaita zone has a latitude and longitude of 6 0 54'N 37 0 45'E with an average elevation of 1850 meters above sea level.
Population and sampling Children, 6 months to 14 years of age, on HAART paired with their mothers/caretakers who lived in the study area for at least half a year were included in this study. Since there are 262 children on HAART attending ART clinics of Wolaita zone, sample size was not calculated and all eligible children were included. Children who were on anemia treatment were excluded from the study.

Data Collection
A structured and interviewer administered questionnaire was used to collect data. Medical records were reviewed and presence of opportunistic infections, baseline CD4+ count and World Health Organization (WHO) clinical stage were recorded to complement the primary data. Twenty four hours recall was used to assess dietary diversity. Nurses and laboratory technicians were recruited for interview and laboratory analysis respectively. BSc nurses were recruited for supervision. Two days training was given for data collectors on the art of interviewing. Trained laboratory technicians were employed and additional training was given. The questionnaire was pretested on 5% of the sample size at Shone Primary Hospital which is not part of the actual data collection. Close supervision was under taken during data collection in a way avoiding bias. Standardized operating procedures were strictly followed during blood sample collection, storage and analytical process.

Laboratory analysis
Three ml of venous blood sample was collected using EDTA anti-coagulated test tube for CD4+ cell count and hemoglobin analyses. CD4+ cell count was analyzed using FACS caliber and Hemoglobin level was measured with Hem cue 301 analyzer, Angelholm, Sweden. Stool sample was taken and examined by direct wet mount technique to detect present of intestinal parasites microscopically.

Statistical analysis
Data were entered into Epi data version 3.1 and exported to Stata version 14.0 for analyses. The data were conveyed in mean and standard deviation of the mean, median and inter-quartile range. Normality was checked for all continuous variables. Hosmer-Lemeshow good-of-fit test and Receiver Operating Characteristics (ROC) test were carried out to have confidence in the regression model. Collinearity was diagnosed, and there was no multicollinearity among the exposure variables included in the model. We used logistic regression analysis to identify exposure variables with association to the outcome variable. Exposures and covariates with p<0.25 during the bivariate analysis were included in multivariate analysis to increase the power of the model. Multivariate analysis was carried out to control for potential confounders and identify independent predictors of the outcome variable. Adjusted Odds Ratio (AOR) with 95% confidence interval (CI) was reported. The level of significance was set at p<0.05.

Socio-Demographic Characteristics
Overall, 256 children On HAART participated in the study with a response rate of 97.7%.

Clinical and Immunological Characteristics
One hundred seventy one (66.8%) were on HAART for more than 60 months. A lithe higher  (Table 3).

Proportion and factors Associated with Anemia
The mean ± (standard deviation) hemoglobin level of the study participants was 12.2 ± (1.4). The proportion of anemia was 38.8%. Children taking co-trimoxazole antibiotic prophylaxis had 55% lower chance of being anemic than those who were not taking Cotrimoxazole (AOR =.45; 95% CI:.21,.95). Children whose caregivers were given nutritional counseling had 91% lesser chance of having anemia (AOR = 09; 95% CI:.01,.98). The odd of being anemic was 3 times higher for children who had intestinal parasitic infection than their counterparts (AOR = 3.10; 95% CI: 1.39, 6.88).

Discussion
In this study, the prevalence of anemia among children on HAART was found to be 38.8%.
The prevalence of anemia in this study is a moderate public health problem according to the WHO classification of anemia [1]. This finding is comparable with a similar study conducted in different parts of Ethiopia (35%) [14] and 39.1% [15]. However, it is lower compared to a study conducted in southwest Ethiopia (53.1 %) [16]. Lower prevalence was reported in different parts of Ethiopia (16.2 %) [11], (10%) [17] and (22.2%) [12]. This divergence might be due to variation in the culture, dietary practice and socio-economic status across different segments of the population within the country.
Children who were taking Co-trimoxazole antibiotic prophylaxis had a lesser odds of being anemic compared to those who did not. This is in agreement with studies from Ethiopia and Gambia [11,18]. One of the most important mechanisms of anemia in HIV infection is failure of erythropoiesis. Co-trimoxazole may probably reduce the levels of cytokines that impair erythropoiesis, by reducing immune activation as well as by preventing infections [19].
Children whose caregivers did not receive nutritional counseling were more likely to be anemic compared to their counter parts. This finding is in agreement with a study conducted in Hawassa referral hospital where children whose caregivers did not receive nutritional counseling had higher odds of being anemic [20]. Better information on the key dietary recommendations might have improved the caregiver's practice of child feeding.
In consistent with studies from Ethiopia [12] and Tanzania [21], children with intestinal parasitic infections had much higher odds to have anemia compared with those children who were not infected with intestinal parasitic infections. This might possibly be due to the attachment of the worm in the intestine to the intestinal mucosa causing intestinal necrosis and blood loss. Chronic infections may also lead to anemia which results from excessive iron loss [4].
Unlike many other studies, this study did not found an association between lower CD4+ cell count and anemia. This finding disagrees with other similar study from Addis Ababa (Central Ethiopia) [12]. This difference might be due to the less number of children with lower CD4+ count involved in this study.

Study limitations
Regarding dietary habit, certain level of recall bias was expected; data collectors aware of the cultural issues collected the data to reduce recall bias. Data on the duration of HAART, baseline CD4+ count, WHO stage, drug regimen, taking Co-trimoxazole and presence of opportunistic infections were taken from the patients charts. Since the health facilities record patient's information consistently and accurately, we believe that the data is unbiased. CD4+ count was not done for 44 children due to lack of reagent. Since the reagent comes as a donation to the country, we couldn't access it at a market.

Conclusions
The magnitude of anemia among children on HAART in the study area is a moderate public health problem. Anemia was significantly associated with caregivers' not receiving clearance letter was written to Wolaita zone health department which is the highest government body in the area for health and health research conducts. Permission was granted from Wolaita zone health department and its lower health administrative structures to conduct the study. Finally, informed written consent was taken from caregivers since we can't obtain consent from children under 18 years of age. The data was kept anonymous.

Availability of data and materials
The dataset analyzed for this study is available from the corresponding author on reasonable request.

Competing interests
The authors declare that they have no competing interests.