Causes for diarrhoea and infection with intestinal parasites among HIV positive patients at a referral hospital in Central Ethiopia

Background: Intestinal parasitic infections are a major public health challenge in many tropical countries. Opportunistic intestinal coccidia such as Cryptosporidia, Cytoisospora or Cyclospora species are common pathogens which are regularly missed using widely practiced wet mount stool microscopy techniques. Therefore, treatment choices are limited and mostly rely on empirical use of cotrimoxazole. The aim of this study was to determine the prevalence of intestinal parasitosis among HIV-infected individuals with and without diarrhoea at the Asella Teaching and Referral Hospital in Ethiopia. Methods: This institution-based cross sectional study was conducted among 163 ambulatory HIV-infected patients with and without diarrhoea. Stool samples were processed for both wet mount and Kinyoun stain. EDTA blood was collected for analysis of CD4 cell count using BD FACSCount™ Flow Cytometer. Sociodemographic and behavioural data was collected using a standardized questionnaire. Chi-squared test was used for statistical analysis. Results: The majority of study participants (62.0%, n=101) were female and the mean age was 38.2 (SD +10.7) years. 52.1% (n=85) of the participants suffered from diarrhoea. The overall prevalence of intestinal parasitic infection in the study population was 18.4% (n=30). Protozoa (Cryptosporidium spp., E. histolytica, G. lamblia and Pentatrichomonas hominis) and helminths (Taenia spp., A. lumbricoides, S. stercoralis, T. trichuria and H. nana) were detected in 12.9% (n=21) and 5.5% (n=9) of patients, respectively. The likelihood for having a parasitic infection was more than eight times higher in participants having diarrhoea. No oocysts of coccidian parasites were detected in the routinely performed wet mount stool microscopy, as expected. Conclusions: There was a high prevalence of opportunistic intestinal parasitic infection in the studied population. Considering the clinical relevance of opportunistic infections

3 particularly in individuals with low CD4 cell count and diarrhoea, the implementation of both stool concentration and modified acid fast staining techniques should be considered to enhance the quality of health care service for HIV-infected patients in resource-limited settings as Ethiopia.

Background
Intestinal parasitic infections are a major public health challenge in many tropical countries. Frequently, the pathogen causing parasitic enteritis is not identified in resource-limited health care systems with restricted diagnostic capacities. Opportunistic intestinal coccidia such as Cryptosporidia, Cystoisospora or Cyclospora species are common causes of those infections which are regularly missed using widely available wet mount stool microscopy techniques. In general, patients with an impaired immune response, primarily due to a Human Immunodeficiency Virus (HIV) infection in this study setting, are at greater risk for developing severe and chronic infections (1). Opportunistic parasite-related intestinal infections are the most common reasons for the development of diarrhoea in HIV-infected individuals; causing about half of the cases of diarrhoeal disease within this group of patients (2). Once symptomatic, they often lead to significant impairments in quality of life or even death (3,4). In addition, cryptosporidiosis and cystoisosporidiosis are considered AIDS-defining illnesses.
In Ethiopia, the prevalence of opportunistic parasitic infections among HIV-infected patients is likely to be under-estimated, as only wet mount microscopy of stool samples is routinely performed in health care facilities. Therefore, choices for treatment of wet mount microscopy negative diarrhoeal disease in HIV-infected patients are limited and mostly rely on empirical use of cotrimoxazole.
Regional differences in the frequency of causing pathogens are to be expected.

Laboratory test
Stool and blood samples were collected from all study participants. Stool examination for parasitic infections was done by wet mount light microscopy at a maximum magnification of 400-fold and modified acid-fast (Kinyoun) staining after processing the stool samples with Telemann concentration technique. Wet mount microscopic examination was also performed from native stool samples to detect trophozoite and larval stages of parasites.
CD4 cell count was determined from EDTA blood samples using BD FACSCount™ Flow Cytometer (Becton Dickinson, Franklin Lakes, NJ, USA).

Data collection and statistical analysis
Sociodemographic and behavioural data was collected using a standardized questionnaire to identify possible predictors of and risk factors for parasitic infection. Collected data was analysed using IBM SPSS Statistics for Windows, version 21.0 (IBM corp., Armonk, NY, USA). Prevalence of parasitic infection among different age groups and sex was analysed by simple frequency distribution. Chi-squared test and multivariate regression analysis were used to identify significant predictors and risk factors. A p-value of <0.05 was considered statistically significant.

Ethical considerations
Ethical clearance to conduct this study was obtained from the appropriate institutional ethical review board at Arsi University, College of Health Science and collected data was used only for the purpose of this study. All study participants signed written informed 5 consent before data and specimen collection commenced. Positive results were communicated with treating physicians for further treatment.

D e m o g r a p h i c c h a r a c t e r i s t i c s a n d c l i n i c a l s t a t u s a m o n g t h e s t u d y p a r t i c i p a n t s
A total of 163 HIV-infected patients were included into the study. The majority of study participants (62.0%, n=101) were female and the mean age was 38.2 years (SD +10.7).
68.7% (n=112) were living in urban areas. 90.2% (n=147) of the participants were treated with combined antiretroviral therapy (cART). The mean CD4 cell count was 482 cells/µl (SD +286.1) with minimum and maximum of CD4+ cell count of 21 cells/µl and 1,742 cells/µl, respectively. 52.1% (n=85) of the participants suffered from diarrhoea. Of those, 11.0% (n=18) reported ongoing diarrhoea for more than two weeks. 11.0% (n=11) of participants reported that they did not have a latrine facility and 17.8% (n=29) to defecate in open field. 25.2% (n=41) had repeated contact with animal excreta and 32.5% (n=53) had the habit of regularly eating uncooked food (for more details, see Table 1).
P r e v a l e n c e a n d r of isolated pathogens, n=12), followed by Giardia lamblia (4.3% in study population, 23.3% of isolated pathogens, n=7) (see Figure 1).  Table 3).  (6), but lower than the prevalence described in previous studies from Ethiopia (i.e. from Desie (7), Butajira (8) and Bahir Dar (9)). The highest prevalence of intestinal parasitosis among HIV-positive individuals (80.3%) was described from an investigation in Bahir Dar in northwest Ethiopia (9). These distinctive regional differences could be triggered by different climatic conditions. The study site is situated in the town of Asella, roughly 2,400 m above sea level in the Eastern Ethiopian Highlands. Despite low numbers, this study showed evidence for four different protozoan parasites and five helminths circulating in the community.
In study participants with diarrhoea, the likelihood of parasitic infection was more than 7 eight times higher in comparison to participants without diarrhoea. Similar results were reported from different studies [9][10][11]. Eating dishes prepared from uncooked beef or

Consent for publication
Not applicable

Availability of data and material
All data generated or analysed during this study are included in this published.     Figure 1 Distribution of parasites detected among study participants