Risk factors for intestinal parasitosis among antiretroviral-treated HIV/AIDS patients in Ethiopia

Abstract

Summary

A cross-sectional survey was conducted to determine the risk factors associated with intestinal parasitosis in HIV/AIDS patients receiving antiretroviral therapy (ART). Socio-demographic information was collected and faecal samples were analysed from 384 randomly selected patients on ART. Data on CD4+ T-cell counts and World Health Organization clinical staging were obtained from the medical records at the hospital. The overall prevalence of intestinal parasitosis was 56% (95% confidence interval [CI]: 51% to 61%). No opportunistic intestinal parasites or Schistosoma haematobium eggs were detected. Unavailability of latrine and lack of hand washing with soap were associated with Entamoeba histolytica/dispar (adjusted odds ratio [AOR], 2.75; 95% CI: 1.77 to 4.27 and AOR, 2.67; 95% CI: 1.60 to 4.44, respectively) and Giardia lamblia (AOR, 2.08; 95% CI: 1.08 to 3.99 and AOR, 2.46; 95% CI: 1.06 to 5.75, respectively) infections. Intestinal parasitosis was significantly associated with low CD4 cell count (p = 0.002). In contrast, intestinal parasitic infections were not associated (p > 0.05) with the World Health Organization disease staging. In summary, poor personal hygiene and sanitation practice contributed to the high prevalence of intestinal parasitosis. Routine diagnosis for intestinal parasitic infections should be performed in patients attending ART clinics in this setting.

Keywords

Intestinal parasitosis risk factors HIV/AIDS antiretroviral therapy Africa

Introduction

Intestinal parasitosis¹ and human immunodeficiency virus (HIV) infection² are highly endemic in resource-poor settings. Approximately one-third of the almost three billion people living in marginalized regions of the world are infected with one or more helminth parasites.¹ Over 60% of the people living with HIV globally are estimated to live in sub-Saharan Africa.³ Like in many other developing countries, Ethiopia is highly affected by the pandemic (with nearly 800,000 people living with the virus).⁴ Similarly, intestinal parasitic infections (IPIs) are also found widely distributed among the population,^5–7 due to lack of safe drinking water and poor hygienic and sanitary conditions.

The expansion of the HIV/AIDS pandemic has brought a significant change in the fauna of intestinal parasites. More importantly, the epidemiology as well as outcome of diseases caused by opportunistic parasites was significantly modified.^8,9 Various intestinal parasites, previously considered to be sporadic or zoonotic, have become causative agents of life-threatening diarrhoea.^10,11

Infection with intestinal parasites causes chronic immune activation and contributes to the rapid progression of disease caused by HIV infection.^12,13 Significant correlations between number of excreted eggs and plasma viral load has been documented, with subsequent reduction of viral load following eradication of IPIs.¹⁴

In Ethiopia, previous studies have shown high prevalence of IPIs among HIV/AIDS patients.^15–17 However, information regarding the magnitude and risk factors associated with IPIs among patients receiving antiretroviral therapy (ART) is still very limited. The purpose of this study, therefore, was to determine the prevalence and associated risk for IPIs among HIV/AIDS patients receiving ART at Adigrat hospital in northern Ethiopia.

Methods

Study design and study population

This cross-sectional study was carried out among patients receiving ART at Adigrat hospital from northern Ethiopia. The study area was selected by purposive sampling for reasons which include relatively large number of HIV/AIDS patients receiving ART during the study period and for its relative accessibility. The study population consisted of 384 (aged: 17−64 years) randomly selected patients visiting the ART clinic for treatment. Patients that were not treated with anti-parasitic drugs nor were involved in any de-worming program within the two weeks before sample collection were included in the study. The selected subjects were invited to participate in the study after obtaining written informed consent.

Data collection

Socio-demographic information

Structured questionnaires were administered by trained medical professionals working in the clinic in a local language to generate information on personal bio-data and other socio-demographic and socio-economic information. Daily close supervision (spot checks, re-interviewing and thorough scrutiny of filled-in questionnaires) was made by the field supervisors deployed with the data collectors.

Parasitological examination

From each subject, about 10 grams (thumb size) fresh stool and 10 mL of midday urine specimens were collected on the spot in clean and labeled containers and were analyzed by well-trained laboratory technologists. Ten percent sub-samples of stool and urine specimens were re-examined for quality control purposes. A subject was classified as infected if an infection was detected by any of the methods used.

Direct microscopy (Wet mount)

A direct wet mount of stool specimen in normal saline (0.85% NaCl solution) was freshly prepared and was analyzed under the light microscope (at ×100 and ×400 magnifications) within less than 1 hour after preparation for the detection of ova, larvae, trophozoites and cysts of intestinal parasites.

Concentration methods

Formalin-ethyl acetate method

A portion of each fresh stool sample was processed using formalin ethyl acetate concentration technique.¹⁸ Briefly, about 4 grams of stool specimen was mixed with 10 mL of 10% formalin and sieved with double-layered gauze into conical centrifuge test tubes. Ten percent formalin was added on the top of the tube and centrifuged for 10 minutes at 500 × g. Supernatant fluid was discarded and sediment was re-suspended with formalin. Two millilitres of ethyl acetate was added and centrifuged for 10 minutes, at 500 × g, after vigorous hand shaking. The supernatant was discarded and the sediment was observed for the presence of ova and/or parasites under the light microscope, at a magnification of ×100 and ×400.

Kato-Katz method

Duplicate Kato-Katz thick smear slides were prepared for each stool specimen by one experienced laboratory technologist to avoid inter-observer bias. The Kato-Katz smears were prepared and analyzed as follows. A small amount of faecal specimen was placed on a clean plastic sheet and a piece of a mesh was pressed on top and the upper surface of the mesh was scraped using a flat-sided spatula to collect the sieved faeces. A template was placed on a slide and the sieved faeces were added with the spatula to completely fill the hole in the template. The template was removed carefully and the faecal material was covered with a pre-soaked cellophane strip. The slide was then inverted and the faecal sample was pressed firmly against other slide to spread the specimen evenly. Prepared slides were placed on the bench with cellophane upwards to enable the evaporation of water while glycerol clears the faeces. Prepared slides were microscopically analyzed within 30 minutes for hookworm eggs identification and after 72 hours for the identification of ova of other helminths.¹⁹ The total number of eggs detected on each slide was counted and the number of eggs per gram of faeces (epg) was calculated to determine egg burden using the conversion factor.

Modified Ziehl-Neelsen staining method

Detection of opportunistic protozoan parasites like Cryptosporidium and Isospora oocysts was done using the modified Ziehl-Neelsen staining technique.²⁰ Briefly, concentrated smear was prepared from fresh stool on slide and fixed with methanol after air drying. Smear was floated with cold carbol fuchsin for 15 minutes and rinsed with water. Preparation was decolorized with 1% acid alcohol (5 mL of 37% hydrochloric acid and 495 mL of 70% ethanol) for 15 seconds. After rinsing again in tap water, the slide was counter-stained with methylene blue for 30 seconds. Smear examined microscopically for oocysts using a low-power magnification and oil immersion objectives after air drying.

Urine sedimentation test

A total of 10 mL urine specimens collected between 10 : 00 h and 14 : 00 h in clean containers were transferred to a conical tube and centrifuged at 500 × g. Supernatant was discarded and sediment was examined microscopically (using the ×10 objective) for the detection of Schistosoma haematobium ova.²⁰

Immunological profile

Data on CD4 + T-cell counts obtained during the study period and World Health Organization (WHO) clinical staging of the study participants were acquired from the medical records at the hospital.

Statistical analysis

Data were entered in an Excel spread sheet, cleaned and anonymised. Statistical analysis was done using Statistical Package for Social Sciences (SPSS) version 16 (Chicago, USA). Descriptive statistics were used to analyze prevalence of the outcome variables. Chi square and logistic regression tests were used to investigate associations between potential variables or risk factors by odds ratio (OR) and 95% confidence interval (CI). Independent sample t test was used for comparison of means. Significance was set at p < 0.05.

Ethical considerations

Ethical clearance was obtained from the institutional Ethical Review Boards of the College of Health Sciences, Mekelle University, Ethiopia (ERC0032/2011). Written permission to conduct the study was sought from the Regional Health Bureau and the hospital. Written consent was obtained from study participants and participants diagnosed positive for IPIs were treated with standard regimens.²¹

Results

From the 384 cases selected for the study, 372 (97%) were able to provide stool and urine specimens. Analysis was done on the 372 subjects who were able to provide specimens. About 40% (n = 150) of the study participants were men. The median age of the participants was 34 years (range 17–64 years). Table 1 describes the prevalence of parasitic infections (56%, 95% CI: 51% to 61%). Entamoeba histolytica/dispar (overall prevalence of 40%) proved to be the most commonly detected parasite, followed by Giardia lamblia and Taenia species. Except for Taenia species infection (p = 0.015), no significant difference (p > 0.05) was observed on the prevalence of intestinal parasitosis between the men and women participants (Table 1).

Table 1.

Prevalence of intestinal parasitosis among HIV/AIDS patients on antiretroviral therapy, Ethiopia (n = 372).

Dependent variables	Men	Women	Total	p Value
	No (%)	No (%)	No (%)
	(n = 146)	(n = 226)	(n = 372)
Intestinal parasitic infections
Protozoa
Entamoeba histolytica/dispar	61 (42)	86 (38)	147 (40)	0.473
Giardia lamblia	18 (12)	29 (13)	47 (13)	0.887
Nematodes
Hookworm	3 (2)	5 (2)	8 (2)	0.919
Ascaris lumbricoides	2 (1)	1 (0.4)	3 (0.8)	0.329
Enterobius vermicularis	2 (1)	1 (0.4)	3 (0.8)	0.329
Strongyloides stercoralis	0 (0)	1 (0.4)	1 (0.2)	0.421
Cestodes
Hymenolepis nana	6 (4)	5 (2)	11 (3)	0.292
Taenia species	18 (12)	12 (5)	30 (8)	0.015^a
Total parasitosis	89 (61)	119 (53)	208 (56)	0.115
Multi parasite infections	22 (15)	23 (10)	45 (12)	0.158

Prevalence of multiple parasite infection was about 12% (45/372) and E. histolytica/dispar and G. lamblia were the most common parasite combinations detected in multiple parasitic infections. No opportunistic intestinal parasites or S. haematobium eggs were detected. Egg concentrations of helminthic infections were of light intensity (less than 1999 and 4999 epg for hookworms and Ascaris lumbricoides, respectively).²² The arithmetic mean egg counts for the identified soil-transmitted helminths were 75 epg of stool for hookworm (range: 24 to 96 epg) and 40 epg for A. lumbricoides (range: 24 to 48 epg). Mean CD4 count was 302 cells/mm³. Intestinal parasitosis was significantly associated with low CD4 cell count (p = 0.002). In contrast, there was no association (p > 0.05) between intestinal parasitosis and WHO disease staging (Data not shown in the table).

Univariate and multivariate logistic regression (adjusted for age, gender, education level and family size) analysis results for IPIs are shown in Table 2. Our findings revealed a strong positive association between poor personal hygiene habits and intestinal parasitosis (Table 2). Overall, we found a relevant pattern of associations between intestinal parasitosis and poor hygiene and sanitation practices. Unavailability of latrine and lack of hand washing with soap practice at critical times were significantly associated with E. histolytica/dispar (adjusted odds ratio [AOR], 2.75: 95% CI: 1.77 to 4.27 and AOR, 2.67; 95% CI: 1.60 to 4.44, respectively) and G. lamblia (AOR, 2.08, 95% CI: 1.08 to 3.99 and AOR, 2.46; 95% CI: 1.06 to 5.75, respectively) infections. Women participants were less likely (AOR, 0.34; 95% CI: 0.15 to 0.74) to be infected with human Taenia parasites (Table 2).

Table 2.

Associations of Intestinal parasitic infections among HIV/AIDS patients on antiretroviral therapy, Ethiopia (n = 372).

		E. histolytica/dispar		G. lamblia		Taenia sp.		H. nana
		COR (CI)		COR (CI)		COR (CI)		COR (CI)
	n	AOR (CI)^a	%	AOR (CI)^a	%	AOR (CI)^a	%	AOR (CI)^a	%
Age
<34	193	1	34	1	10	1	9	1	4
≥34	179	1.53 (1.01, 2.33)^b	43	1.44 (0.78, 2.67)	14	0.72 (0.34, 1.55)	7	0.63 (0.18, 2.18)	2
		1.38 (0.88, 2.16)		1.38 (0.88, 2.16)		0.66 (0.29, 1.50)		0.47 (0.12, 1.83)
Gender
Men	145	1	41	1	12	1	12	1	4
Women	227	0.86 (0.56, 1.31)	38	1.05 (0.56, 1.96)	13	0.40 (0.19, 0.86)^b	5	0.53 (0.16, 1.76)	2
		0.97 (0.62, 1.52)		1.13 (0.59, 2.18)		0.34 (0.15, 0.74)^b		0.53 (0.15, 1.88)
Family size
<3	156	1	46	1	13	1	13	1	3
≥3	216	1.63 (1.07, 2.48)^b	33	1.81 (0.98, 3.35)	12	0.56 (0.25, 1.25)	4	1.65 (0.49, 5.51)	3
		1.50 (0.96, 2.33)		1.50 (0.96, 2.33)		0.55 (0.23, 1.29)		2.13 (0.59, 7.69)
Presence of latrine
Yes	182	1	30	1	10	1	7	1	2
No	190	2.84 (1.85,4.38)^b	46	2.21(1.16, 4.19)^b	14	1.39 (0.65, 2.94)	7	1.84 (0.53, 6.39)	4
		2.75 (1.77, 4.27)^b		2.08 (1.08, 3.99)^b		1.44 (0.67, 3.10)		2.01 (0.43, 7.12)
Hand washing with soap
Yes	108	1	27	1	6	1	9	1	4
No	264	2.77 (1.68, 4.58)^b	44	2.65 (1.15, 6.12)^b	15	0.83 (0.37, 1.83)	7	1.92 (0.41, 9.04)	3
		2.67 (1.60, 4.44)^b		2.46 (1.06, 5.75)^b		0.85 (0.38, 1.92)		1.01 (0.42, 9.70)
Family with diarrhoea
No	309	1	38	1	12	1	8	1	3
Yes	63	1.18 (0.68, 2.05)	41	1.01 (0.45, 2.27)	12	1.25 (0.49, 3.20)	10	0.48 (0.06, 3.84)	2
		1.12 (0.64, 1.92)		1.10 (0.48, 2.51)		1.40 (0.53, 3.65)		0.52 (0.06, 4.20)
Pipeline water source
Yes	253	1	37	1	12	1	7	1	3
No	119	1.04 (0.67, 1.62)	40	1.19 (0.63, 2.25)	14	1.64 (0.77, 3.49)	10	1.18 (0.34, 4.10)	3
		1.01 (0.65, 1.59)		1.12 (0.58, 2.14)		1.77 (0.82, 3.84)		1.23 (0.36, 4.51)

E. histolytica/dispar: Entamoeba histolytica and/or Entamoeba dispar; G. lamblia; Giardia lamblia; COR: Crude odds ratio as computed by logistic regression model; H. nana: Hymenolepis nana; AOR: adjusted odds ratio as computed by logistic regression model; Taenia sp.: Taenia species.

Adjusted for age, gender, education level and family size.

Statistically significant at 0.05.

Discussion

Our findings revealed widespread prevalence of intestinal parasitosis among antiretroviral-treated patients. Only extracellular parasites were identified. The prevalence of E. histolytica/dispar and G. lamblia was significantly higher. IPIs and individual hygiene and sanitation practices were very clearly related. Absence of latrine and hand washing with soap practice contributed to the higher proportion of IPIs, which suggests the need to address personal hygiene and sanitation practices among HIV/AIDS patients.

The overall prevalence of IPIs (56%) in our data was higher than prevalence reports from different studies conducted in the central (35.4%),²³ north-eastern (17.6%)²⁴ and south-western (39.6%)²⁵ regions of the country, Kenya (50.9%),²⁶ Ghana (35%),²⁷ Nigeria (5.3%)²⁸ and Democratic Republic of Congo (15.4%).²⁹ Variations in geographical locations, socio-economic conditions and cultural practices of the population under consideration might explain the differences in findings among the studies. The methods employed for stool examination and the time of the study may also have contributed to the differences. High prevalence of IPIs among the study participants may call for better follow-up through laboratory tests and more comprehensive actions by the patients themselves in adopting prevention measures against intestinal parasites.

In agreement with other studies, opportunistic coccidian parasitic infections were not detected in the present study.^23,28,30 ART have been documented to improve immune status,^31,32 and thereby preventing the occurrence of opportunistic parasitic infections.^30–32 This may explain the absence of opportunistic parasites in this study. The significantly less prevalence of Taenia parasite infection among female patients might be related to less consumption of raw or undercooked meat among this gender group.

Non-opportunistic parasites, including the intestinal parasites E. histolytica/dispar and G. lamblia, are frequently encountered in resource-limited countries.³³ Diarrhoea is a common clinical symptom associated with infection by these parasites.³⁴ Infection with these organisms, therefore, might contribute to the morbidity among HIV/AIDS patients and early detection and treatment of these parasites are important to improve the quality of life of patients on ART. Associations between poor hygiene and sanitation practices and intestinal parasitosis are well documented.^35,36 Faecal-oral route is the most common method of transmission through contaminated hands and fingers.³³

A major strength of this study was the random selection of the study participants. Generalization may be made to the study population as an attempt was made to identify randomized HIV/AIDS patients in the ART clinic. The limitation of the study was the cross-sectional nature making any inference on causal relationship among variables impossible. The cross-sectional nature of the study could only generate a hypothesis about the possible role of certain independent variables on the infection status of these patients but not their causal relationships.

In conclusion, our results emphasize the need of increased personal hygiene and sanitation practice, such as increased access to clean water and soap for hand washing and access to latrines to restrict contamination, and hygiene-related education would improve health of HIV-infected patients on ART. Stool examination should be routinely performed in the follow-up of patients attending ART clinics in order to optimize treatment and institution of other preventive measures.

Footnotes

Acknowledgments

The authors thank the Norwegian Agency for Development Co-operation (NORAD) phase II project for supporting the study. The authors thank all participants for their collaboration. The authors are grateful to the hospital administration and the ART clinic. The authors acknowledge the cooperation of the Tigray Regional Health Bureau. The authors express their sincere gratitude to the laboratory staff involved in the field work.

Conflict of interest

The authors declare no conflict of interest.

Funding

This work was supported by the Norwegian Agency for Development Co-operation (NORAD) phase II project.

References

Hotez

Brindley

Bethony

. Helminth infections: the great neglected tropical diseases. J Clin Invest 2008; 118: 1311–1321.

Gilks

Crowley

Ekpini

. The WHO public-health approach to antiretroviral treatment against HIV in resource-limited settings. Lancet 2006; 368: 505–10.

World Health Organization. HIV/AIDS fact sheet. No. 360, 2012, www.who.int/mediacenter/factsheets/fs360/en/index.html (2012, accessed 01 March 2013).

Federal Democratic Republic of Ethiopia. Country Progress Report on HIV/AIDS Response, http://www.unaids.org/en/dataanalysis/knowyourresponse/countryprogressreports/2012countries/GAP Report 2012.pdf (2012, accessed 08 March, 2013).

Mengistu

Gebre-Selassie

Kassa

. Prevalence of intestinal parasitic infections among urban dwellers in southwest Ethiopia. Ethiop J Health Dev 2007; 21: 12–17.

Wegayehu

Tsalla

Seifu

. Prevalence of intestinal parasitic infection among highland and lowland dwellers in Gamo area, south Ethiopia. BMC Public Health 2013; 13: 151–151.

Dejenie

Petros

. Irrigation Practices and Intestinal Helminth Infections in Southern and Central Zones of Tigray. Ethiop J Health Dev 2009; 23: 48–56.

Gupta

Narang

Nunavath

. Chronic diarrhea in HIV patients: prevalence of coccidian parasites. Indian J Med Microbiol 2008; 26: 172–175.

Kelly

Todd

Sianongo

. Susceptibility to intestinal infection and diarrhoea in Zambian adults in relation to HIV status and CD4 count. BMC Gastroenterol 2009; 9: 7–7.

10.

Lindo

Dubon

Ager

. Intestinal parasitic infections in human immunodeficiency virus (HIV)-positive and HIV-negative individuals in San Pedro Sula, Honduras. Am J Trop Med Hyg 1998; 58: 431–435.

11.

Arenas-Pinto

Certad

Ferrara

. Association between parasitic intestinal infections and acute or chronic diarrhoea in HIV-infected patients in Caracas, Venezuela. Int J STD AIDS 2003; 14: 487–492.

12.

Bentwich

Kalinkovich

Weisman

. Immune activation is a dominant factor in the pathogenesis of Africa AIDS. Immunol Today 1995; 16: 187–19.

13.

Weissman

Barker

Fauci

. The efficiency of acute infection of CD4+ T cells is markedly enhanced in the setting of antigen specific immune activation. J Exp Med 1996; 183: 687–692.

14.

Wolday

Mayaan

Mariam

. Treatment of intestinal worms is associated with decreased HIV plasma viral load. J Acquir Immun Defic Syndr 2002; 31: 56–62.

15.

Mariam

Abebe

Mulu

. Opportunistic and other intestinal parasitic infections in AIDS patients, HIV seropositive healthy carriers and HIV seronegative individuals in southwest Ethiopia. East Afr J Public Health 2008; 5: 169–173.

16.

Assefa

Erko

Medhin

. Intestinal parasitic infections in relation to HIV/AIDS status, diarrhea and CD4 T-cell count. BMC Infect Dis 2009; 9: 155–155.

17.

Alemu

Shiferaw

Getnet

. Opportunistic and other intestinal parasites among HIV/AIDS patients attending Gambi higher clinic in Bahir Dar city, North West Ethiopia. Asian Pac J Trop Med 2001, pp. 661–665.

18.

Zeibig

. Clinical parasitology. A principal approach, The Curtis Center. Independence Square west Philadelphia: Saunders, 1997, pp. 269–269.

19.

World Health Organization. Action against Worms. 2008, Issue 11, http://www.who.int/neglected_diseases/preventive_chemotherapy/pctnewsletter11.pdf (accessed 05 December, 2013).

20.

Cheesbrough

. District laboratory practice in tropical countries, 2nd ed. Part 1. Cambridge: Cambridge University Press, 2005.

21.

Food, Medicine and Health Care Administration and Control Authority of Ethiopia (FMHACA). Standard Treatment Guideline for General Hospitals. Drug Administration and Control Authority of Ethiopia Contents; 2010.

22.

Montresor A, Crompton DWT, Hall A, et al. Guidelines for the evaluation of soil-transmitted helminthiasis and schistosomiasis at community level. World Health Organization, Geneva; 1998. Document WHO/CTD/SIP/98.1.

23.

Adamu

Petros

. Intestinal protozoan infections among HIV positive persons with and without Antiretroviral Treatment (ART) in selected ART centers in Adama, Afar and Dire-Dawa, Ethiopi. Ethiop J Health Dev 2009; 23: 133–140.

24.

Missaye

Dagnew

Alemu

. Prevalence of intestinal parasites and associated risk factors among HIV/AIDS patients with pre-ART and on-ART attending Dessie hospital ART clinic, Northeast Ethiopia. AIDS Res Ther 2013; 10: 7–7.

25.

Zynudin

Hemalatha

Kannan

. Prevalence of opportunistic intestinal parasitic infections among HIV infected patients who are taking antiretroviral treatment at Jimma Health Center, Jimma, Ethiopia. Eur Rev Med Pharmacol Sci 2013; 17: 513–516.

26.

Kipyegen

Shivairo

Odhiambo

. Prevalence of intestinal parasites among HIV patients in Baringo, Kenya. PAMJ 2012; 13: 37–37.

27.

Boaitey

Nkrumah

Idriss

. Gastrointestinal and urinary tract pathogenic infections among HIV seropositive patients at the Komfo Anokye Teaching Hospital in Ghana. BMC Res Notes 2012; 5: 454–454.

28.

Akinbo

Omoregie

. Intestinal parasitic infections in human immunodeficiency virus-infected persons on highly active antiretroviral therapy in Benin City, Nigeria. GMBHS 2011; 3: 119–122.

29.

Wumba

Longo-Mbenza

Mandina

. Intestinal parasites infections in hospitalized AIDS patients in Kinshasa, democratic Republic of Congo. Parasite 2010; 17: 321–328.

30.

Nissapatorn

. Lessons learned about opportunistic infections in Southeast Asia. Southeast Asian J Trop Med Public Health 2008; 39: 625–641.

31.

Idigbe

Adewole

Eisen

. Management of HIV-1 infection with a combination of nevirapine, stavudine, and lamivudine: a preliminary report on the Nigerian antiretroviral program. J Acquir Immune Defic Syndr 2005; 40: 65–69.

32.

Willemot

Klein

. Prevention of HIV-associated opportunistic infections and diseases in the age of highly active antiretroviral therapy. Expert Rev Anti Infect Ther 2004; 2: 521–532.

33.

Haque

. Human intestinal parasites. J Health Popul Nutr 2007; 25: 387–391.

34.

Paniker

CKJ

. Text book of medical parasitology, 6th ed. New Delhi, India: Jaypee Brothers Medical Publishers, 2007.

35.

Crompton

DWT

Salvoli

. Intestinal parasitic infections and urbanization. Bull World Health Organ 1993; 71: 1–7.

36.

Alum

Rubino Jr Ijaz

. The global war against intestinal parasite-should we use a holistic approach? IJID 2010; 14: e732–e738.