Study of intestinal parasites among the immunosuppressed patients attending a tertiary-care center in Northeast India.
Intestinal parasitic infestations cause a variety of clinical conditions, ranging from asymptomatic infestations to life-threatening situations. Majority of the symptoms are related to gastrointestinal (GI) tract. Parasitic infections that cause self-limited diarrhea in immunocompetent patients may cause profuse diarrhea in immunocompromised individuals, generally accompanied by loss of weight, anorexia, malabsorption syndrome, and, in some cases, fever and abdominal pain.  Intestinal parasites are among the most common human infections distributed worldwide with prevalence rates as high as 40.59% in developing countries.[R] In India, occurrence of intestinal parasites proclaimed from various workers reveal wide variations from 11.3% to 90%.  By definition, a compromised host is the one in whom normal defense mechanisms are impaired (eg. AIDS), absent (eg. congenital deficiencies), or bypassed. These patients are becoming more common in medical facilities and represent a growing problem in terms of diagnosis and subsequent management. Individuals with defective immune system exhibit high susceptibility to infections with nonvirulent and minimally pathogenic organisms. 
Immunodeficiency diseases can be classified as primary and secondary. The primary immunodeficiencies may be either congenital or acquired and are currently classified according to the mode of inheritance and whether the defect involves T cells, B cells, or both. AIDS caused by human immunodeficiency virus is a secondary immunodeficiency disease. Other examples of secondary immunodeficiency diseases are protein-losing enteropathy, lymphoreticular malignancy, and patients on immunosuppressive drugs.  Impaired cell-mediated immunity results in progressive decline of immunological response making them susceptible to variety of common and intestinal infections leading to increased morbidity and mortality. 
In the early 1980s, it appeared that the importance of parasitic infections was declining in immunocompromised cancer patients. However, in recent years, the frequency of these diseases has risen owing to the increased use of corticosteroid in cytotoxic regimens. 
A lot of these pathogens, especially the intracellular protozoa that chiefly influence the small intestine, produce their most devastating effects in patients with immune deficiency. Parasitic infections generally are asymptomatic in otherwise healthy individuals; however, their manifestations in immune compromised individuals are more severe and devastating. In any parasitic infection in immunosuppressed host, certain organisms tend to produce greater pathological sequelae in these patients. 
Some of the common parasites found in immunosuppressed patients are Giardia lamblia, Entamoeba histolytica, Strongyloides stercoralis, Cryptosporidium parvum, Cyclospora cayeatanensis, Isospora belli, and Microsporidia spp. 
There is paucity of reports regarding prevalence of parasites in this part of Northeast India. In view of this, the study was conducted to understand the prevalence of intestinal parasitic infestations among the immunosuppressed patients, to demonstrate the intestinal parasite profile in immunosuppressed patients, and to determine the association of intestinal parasite and clinical presentation.
Materials and Methods
This study was carried out in the Department of Microbiology, for a period of 1 year. Stool samples received from different departments were included in the study as per inclusion criteria.
The stool samples received from the patients with the following immunosuppressive (IS) status like HIV infection, malignancy, patients on IS therapy including persons who underwent transplantation, diabetes mellitus (DM) patients with retinopathy/ neuropathy/nephropathy, severe anemia (<6g/dL), protein energy malnutrition (grade III PEM), tuberculosis (TB), and chronic diseases (e.g. chronic kidney disease, chronic obstructive pulmonary disease, etc.) were included in the study.
A clinical data in reference to the duration and frequency of diarrhea, weight loss, loss of appetite, associated abdominal symptoms, vomiting, and fever were obtained from the individual patient's record.
Specimen Collection and Transportation
Patients were instructed to avoid contamination of the stool specimen with urine or water.
The specimen was sent to the Microbiology Department for further processing on the same day. The specimen was labeled properly and processed immediately.
Examination of Stool Sample 
The specimen was examined by naked eye for color, consistency, presence of blood, mucus, adult worms, or segments of worms and recorded accordingly.
Microscopic examination of the stool specimen was performed by the following techniques:
Direct wet smear: Saline preparation and iodine preparation. Concentration techniques: [R] Formol-ether sedimentation and Sheather's sugar floatation technique were employed for all specimens for the concentration of the parasitic ova and cysts.
Permanent staining techniques:  Specialized stain such as Kinyoun's acid-fast stain was performed for coccidian parasites.
Significance was evaluated by Fisher's exact test and/or [chi square]-test and "p" value less than 0.05 was considered as significant.
A total of 149 immunosuppressed patients were enrolled in the study, of which 49% of the patients were male and 51% were female subjects, and the highest number of patients [46 (30.8%)] belonged to the age group of 1-10 years. Parasitic infestations were present in 79 (53.02%) patients with male and female ratio of almost 1:1 [Figure 1] and highest prevalence belonging to age group 21-30 years [Table 1]. The most common parasite isolated was E. histolytica/E. dispar [31 (33.7%)], followed by Ascaris lumbricoides [22 (23.9%)], G. lamblia [19 (20.65%)], Trichuris trichiura [10 (10.87%)], hookworm [6 (6.52%)], S. stercoralis [3 (3.26%)], and Taenia spp. [1 (1.08%)].
The highest prevalence of parasitic infestation was found in patients with cancers [80% (12 of 15)], followed by severe anemia [68.42% (13 of 19)], PEM [59.20% (29 of 46)], tuberculosis (41.67%), HIV positive (40%), other chronic diseases (38.50%), and diabetes mellitus (17.64%). The most common infestation in HIV-positive patients, cancer patients, diabetes mellitus patients, severe anemia patients, and in patients with IS drugs was found to be E. histolytica/ Entamoeba dispar. A. lumbricoides was the most common infestation in patients with PEM [Table 2].
Mixed infestations were seen in nine patients, where four (44.44%) showed infestation with Ascaris + Entamoeba, one (11.11%) showed infestation with hookworm + Trichuris, one showed infestation with Entamoeba + Ascaris + Trichuris (11.11%), two (22.22%) showed infestation with Entamoeba + hookworm + Trichuris, and one (11.11%) showed infestation with Ascaris + hookworm + Trichuris + Strongyloides.
On comparison of routine microscopy method with others, stool concentration method showed higher prevalence and isolation by the formal-ether sedimentation method (52.3%) [Figure 2]. This association when compared with routine microscopy was statistically significant (p = 0.011).
Overall, 106 (71.1%) patients presented GI symptoms. The association of GI symptoms was statistically significant (p = 0.0183). The prevalence of parasitic infestation was highest in patient complaining of diarrhea (65.5%). The study also showed that 43 (28.9%) patients who did not have any GI symptoms [16 (11%)] had intestinal parasites, indicating that they still harbor these opportunistic and nonopportunistic parasites and act as carriers in the community via contaminated water and open defecation practices [Table 3].
The study showed association of intestinal parasitic infection in 79 (53.02%) among the 149 patients with different immunosuppressed status. The result of this study is comparable with the reports of Idris et al.  and Rao et al.,  where both reported a prevalence of 57% in their studies. However, when compared with our study, many reported a varied range of prevalence ranging from 23% to 97.4%. [11,01,11] The difference among prevalence of parasites in the abovementioned studies may be owing to multifactorial reasons in different geographical locations.
In this study, the prevalence of intestinal parasites in immunosuppressed patients was found to be the highest (63.15%) in the age group 21-30 years, which was similar to the study by Al-Megrin  An increase in the infectivity rate in this group in our study may be owing to the cumulative effect of the factors such as exposure to outdoor life, poor socioeconomic and poor sanitary conditions, which demands broad-based community study to ascertain such association.
In our study, we have observed parasitic infections in male patients (50.63%) compared with female patients (49.37%), which do not indicate any gender preponderance among immunosuppressed patients. This observation is in concordance with that of Al-Megrin.  Overall, the most common parasite isolated was E. histolytica/E. dispar [31(33.7%)]. The most common infestation in HIV-positive patients, cancer patients, DM patients, severe anemia patients, and in patients with IS drugs was found to be E. histolytica/E. dispar. In our study, only microscopy was employed to identify the parasites. Molecular methods for differentiation of E. histolytica from other nonpathogenic parasites such as E. dispar were not employed. The higher prevalence of E. histolytica can be explained owing to the favorable climate condition for survival of protozoan cyst outside the human host, absence of intermediate host, and lack of latency period to maturation when passed in feces.
The intestinal parasitic infections among cancer patients (80%) was found to be higher in our study when compared with similar studies [1,8,10,13,14] E. histolytica/E. dispar was the most common parasite in cancer patients in our study, similar to studies by Guarner et al.  and Rudrapatna et al. 
In our study, patients with severe anemia showed a parasitic prevalence of 68.42%, which was comparable to many studies. [9,15,16] E. histolytica/E. dispar was the most common parasite in patients with with severe anemia, which is a unique feature of our study, as Dori et al.  and Rao et al.  found hookworm and Tsuyuoka et al. found A. lumbricoides to be the most common parasites isolated in their respective studies.
The prevalence of parasites among patients on IS therapy in this study was found to be 60%, which was similar to the studies by Abaza et al. (31.7%)  and Idris et al. (50%) . In this study, E. histolytica/E. dispar and A. lumbricoides were the most common parasites isolated, which was different from the study conducted by Idris et al., who found Blastocystis hominis as the most common parasite isolated. 
Among PEM patients, 59.2% showed prevalence of parasites, which was similar to findings by Bhandari et al.  However, our study varied in the prevalence when compared with many studies. [8,18] In our study, A. lumbricoides was the most common parasite isolated among PEM patients. Bechir et al.  observed that A. lumbricoides and E. histolytica/ E. dispar were the most common parasites isolated. However, Bhandari et al.  observed that E. histolytica, G. lamblia, and A. lumbricoides were most commonly isolated in patients with grade 3 and grade 4 PEM.  Thus, it was seen that the results of this study is comparable with most of the other studies. The high prevalence of Ascaris may be explained by the fact that they cause absorption and retention of protein and nitrogen and by themselves ingesting, absorbing, and utilizing the host food. Heavy ascarial infestation can probably induce PEM in persons whose diet is otherwise inadequate.
The prevalence of parasites among HIV-positive patients in this study was found to be 40%, which was in accordance with many similar studies. [20-22] However, the prevalence rate among HIV patients shows a wide variation from 11.4% to 62.7% in various studies. [23-29] The wide variation of prevalence may be attributed to difference in geographical distribution of parasites, sanitary practices, and different selection criteria of cases. In this study, E. histolytica/E. dispar was the most common parasite isolated in HIV-positive patients, which was similar to studies by Assefa et al.,  and Asma et al.  C. parvum was the most common parasite isolated in many studies. [28,32] G. lamblia was the most common parasite isolated in a study by Cimerman et al.  and I. belli was the common parasite in the study by Gupta et al.  The difference in predominant parasite isolated in different studies compared with our study was because these parasites were more encountered when CD4 T-cell count falls below 200 cells/[micro]L. However, in our study, we included all HIV-positive patients irrespective of their CD4 T-cell count.
Parasites in Diabetic patients in this study was found to be 17.64%. This was similar to study by Akhlagi et al.  However, there is a variation in prevalence ranging from 8% to 47%  The most common infestation among diabetic patients was found to be E. histolytica/E. dispar which differed from many studies where G. lamblia,  A. lumbricoides  to be the most common parasite isolated.
The prevalence of intestinal parasite in patients with GI symptoms was found to be 53.77%. This was comparable to most of the studies. [1,12,13,24,25] Our study also revealed that 22 individuals showed no GI disorders such as diarrhea and pain in the abdomen, but they still harbor these opportunistic and nonopportunistic parasites and act as carriers in the community via contaminated water and open defecation practices.
In our study, we also found that 37% of stool samples were positive for parasites by direct microscopy, whereas with formol-ether sedimentation method, 52.3% of stool samples were positive for parasite. This finding is similar to that reported by Saxena et al.  and Vinayak et al.,  who reported that, by formol-ether sedimentation, 31% of stool samples were positive for helminthic ova and pathogenic protozoan cysts compared with 21% by direct saline and iodine preparation. This can be explained by the fact that the use of formalin fixes and preserves the fecal specimen, and ether decreases the specific gravity of small fecal particles, causing them to float in the suspension. The coarse nonabsorbent elements including eggs and cysts are left at the bottom, and ether also dissolves fat. The addition of these two chemicals and centrifugation improved the isolation rate.
This study thus reveals that intestinal parasites are prevalent among immunosuppressed patients in Meghalaya and rapid detection of such infestations is important for empirical therapy.
Our study was conducted on a limited number of sample size within a period of 1 year. However, the study supported the fact that immunosuppressed individuals are prone to infections with intestinal parasites.
Demonstration of intestinal infections in the immunosuppressed patients employing concentration method such as Sheather's sugar floatation technique and formol-ether concentration method are found to be essentially effective in our study. Such approaches are easier for reliable laboratory diagnosis for appropriate management of immunosuppressed patients with intestinal parasitic infection.
In our study, we found that prevalence of parasites was highest among cancer patients in comparison to other similar studies. However, this may not be significant because of the limited sample size.
Among HIV-positive patients, E. histolytica/E. dispar was the most common parasite isolated in contrast to many studies, where coccidian parasites were more commonly isolated in this group of patients. This difference may be because, in our study, HIV-positive patients were included irrespective of their CD4 T-cell counts.
This study thus highlights the importance of testing for intestinal parasites in immunosuppressed patients and emphasizes the necessity of awareness among clinicians regarding the occurrence of these parasites in this population and health education of the population for food hygiene. The frequency and the danger of those opportunistic infections require their efficient diagnosis and appropriate management.
[1.] Botero JH, Castano A, Montoya MN, Ocampo NE, Hurtado MI, Lopera MM. A preliminary study of the prevalence of intestinal parasites in immunocompromised patients with and without gastrointestinal manifestations. Rev Inst Med Trop Sao Paulo 2003; 45(4):197-200.
[2.] Sethi S, Sehgal R, Malla N, Dubey ML, Mahajan RC. Changing trends of parasitic infections in Chandigarh (Northern India): hospital based study. Indian J Med Microbiol 2000; 18(3):106-9.
[3.] Garcia LS. Diagnostic Medical Parsitology, 4th edn. Washington DC: ASM Press, 2001. pp. 503-535, p 723, p 741.
[4.] Kindt TJ, Gosby RA, Osborne BA. Kuby Immunology, 6th edn. New York: WH Freeman and Company, 2007. pp. 493-524.
[5.] Ayyagari A, Sharma AK, Prasad KN, Dhole TN, Kishore J, Chaudhury G. Spectrum of opportunistic infections in HIV infected cases in a tertiary care hospital. Indian J Med Microbiol 1999; 17:78-80.
[6.] Pizzo PA, editor. Infectious complications in the immunocompromised host. I. Hematol Oncol Clin North Am 1993; 7:4.
[7.] Collee JG, Fraser AG, Marnion BP, Simmons A (Eds.). Mackie & McCartney Practical Medical Microbiology, 14th edn. New Delhi: Elsevier, 1999. p. 743.
[8.] Idris NS, Dwipoerwantoro PG, Kurniawan A, Said M. Intestinal parasitic infection of immunocompromised children with diarrhoea: clinical profile and therapeutic response.J Infect Dev Ctries 2010; 4(5):309-17.
[9.] Rao VG, Aggrawal MC, Yadav R, Das SK, Sahare LK, Bondley MK, Minocha RK. Intestinal parasitic infections, anaemia and undernutrition among tribal adolescents of Madhya Pradesh. Indian J Community Med 2003; 28:26-9.
[10.] Abaza SM, Makhlouf LM, el-Shewy KA, el-Moamly AA. Intestinal opportunistic parasites among different groups of immunocompromised hosts. J Egypt Soc Parasitol 1995; 25(3):713-27.
[11.] Kang G, Mathew MS, Rajan DP, Daniel JD, Mathan MM, Mathan VI, et al. Prevalence of intestinal parasites in rural Southern Indians. Trop Med Int Health 1998; 3(1):70-5.
[12.] Al-Megrin WA. Intestinal parasites infection among immunocompromised patients in Riyadh, Saudi Arabia. Pak J Biol Sci 2010; 13(8):390-4.
[13.] Guarner J, Matilde-Nava T, Villasenor-Flores R, SanchezMejorada G. Frequency of intestinal parasites in adult cancer patients in Mexico. Arch Med Res 1997; 28(2):219-22.
[14.] Rudrapatna JS, Kumar V, Sridhar H. Intestinal parasitic infections in patients with malignancy. J Diarrhoeal Dis Res 1997; 15(2):71-4.
[15.] Tsuyuoka R, Bailey JW, Nery Guimaraes AM, Gurgel RQ, Cuevas LE. Anemia and intestinal parasitic infections in primary school students in Aracaju, Sergipe, Brazil. Cad Saude Publica. 1999; 15(2):413-21.
[16.] Dori GU, Tullu KD, Ali I, Hirko A, Mekuria G. Prevalence of hookworm infection and its association with anemia among patients visiting Fenan Medical Center, East Wollega Zone, Ethiopia. Ethiop Med J 2011; 49(3):265-71.
[17.] Bhandari B, GP Gupta, Mandowara SL. Prevalence of intestinal parasites in Udaipur. Indian J Pediatr 1985; 52(416):299-302.
[18.] Dhingra DC, Anand NK, Gupta S. Interaction of nutrition and infection in school children. Indian Pediatr 1977; 14(3):189-93.
[19.] Bechir M, Schelling E, Hamit MA, Tanner M, Zinsstag J. Parasitic infections, anemia and malnutrition among rural settled and mobile pastoralist mothers and their children in Chad. Ecohealth 2012; 9(2):122-31.
[20.] Cimerman S, Cimerman B, Lewi DS. Prevalence of intestinal parasitic infections in patients with acquired immunodeficiency syndrome in Brazil. Int J Infect Dis 1999; 3(4):203-6.
[21.] Gupta S, Narang S, Nunavath V, Singh S. Chronic diarrhoea in HIV patients: prevalence of coccidian parasites. Indian J Med Microbiol 2008; 26(2):172-5.
[22.] Awole M, Gebre-Selassie S, Kassa T, Kibru G. Prevalence of intestinal parasites in HIV-infected adult patients in Southwesten Ethiopia. Ethiop J Health Dev 2003; 17(1):71-8.
[23.] Zali MR, Mehr AJ, Rezaian M, Meamar AR, Vaziri S, Mohraz M. Prevalence of intestinal parasitic pathogens among HIV-positive individuals in Iran. Jpn J Infect Dis 2004; 57(6):268-70.
[24.] Mukhopadhya A, Ramakrishna BS, Kang G, Pulimood AB, Mathan MM, Zachariah A, et al. Enteric pathogens in southern Indian HIV-infected patients with and without diarrhoea. Indian J Med Res 1999; 109:85-9.
[25.] Mohandas, Sehgal R, Sud A, Malla N. Prevalence of intestinal parasitic pathogens in HIV-seropositive individuals in Northern India. Jpn J Infect Dis 2002; 55(3):83-8.
[26.] Kumar SS, Ananthan S, Lakshmi P. Intestinal parasitic infections in HIV infected patients with diarrhoea in Chennai. Indian J Med Microbiol 2002; 20(2):88-91.
[27.] Gupta M, Sinha M, Raizada N. Opportunistic intestinal protozoan parasitic infection in HIV positive patient in Jamnagar, Gujarat. SAARC J Tuber Lung Dis HIV/AIDS 2008; 5(1):21-4.
[28.] Dwivedi KK, Prasad G, Saini S, Mahajan S, Lal S, Baveja UK. Enteric opportunistic parasites among HIV infected individuals: associated risk factors and immune status. Jpn J Infect Dis 2007; 60:76-81.
[29.] Kulkarani SV, Kairon R, Sane SS, Padmawar PS, Kale VA, Thakar MR, et al. Opportunistic parasitic infections in HIV/AIDS patients presenting with diarrhoea by the level of immunosuppression. Indian J Med Res 2009; 130:63-6.
[30.] Assefa S, Erko B, Medhin G, Assefa Z, Shimelis T Intestinal parasitic infections in relation to HIV/AIDS status, diarrhea and CD4 T-cell count. BMC Infect Dis 2009; 9:155.
[31.] Asma I, Johari S, Sim BL, Lim YA. How common is intestinal parasitism in HIV-infected patients in Malaysia? Trop Biomed 2011; 28(2):400-10.
[32.] Arora DR, Arora B. AIDS-associated parasitic diarrhoea. Indian J Med Microbiol 2009; 27(3):185-90.
[33.] Akhlaghi L, Gharavi MJ, Faghihi AH, Jabbari M. Survey on the prevalence rates of intestinal parasites in diabetic patients in Karaj and Savodjbolagh cities. Razi J Med Sci 2005; 12(45):23-9.
[34.] Nazligul Y, Sabuncu T, Ozbilge H. Is there a predisposition to intestinal parasitosis in diabetic patients? Diabetic Care 2001; 24(8):1503-4.
[35.] Saxena SN, Varghese A, Gosh Ray B. A comparison of salt floatation and formal ether techniques in demonstrating helminthic ova in faeces. Indian J Med Res 1969; 57:1213.
[36.] Vinayak VK, Tandon BN, Prakash O. A comparative evaluation of formol-ether, zinc sulphate and magnesium sulphate concentration techniques for diagnosis of helminthic ova and protozoal cysts Indian J Med Research 1967; 55(2):134-8.
Ishani Bora, Vikramjeet Dutta, Wihiwot Valarie Lyngdoh, Annie Bakorlin Khyriem, Elantamilian Durairaj, Anil Chandra Phukan
Department of Microbiology, North Eastern Indira Gandhi Regional Institute of Health and Medical Sciences, Shillong, Meghalaya, India.
Correspondence to: Ishani Bora, E-mail: email@example.com
Received January 8, 2016. Accepted January 22, 2016
Table 1: Age-wise prevalence of parasitic infestation in the study population Age group Positive Negative (years) Number Percentage Number Percentage 0-10 29 61.70 18 38.3 11-20 9 52.94 8 47.06 21-30 12 63.15 7 36.85 31-40 13 48.15 14 51.85 41-50 6 50 6 50 >50 10 37 17 63 Total 79 53.02 70 46.98 Age group Total (years) Number Percentage 0-10 47 31.54 11-20 17 11.40 21-30 19 12.75 31-40 27 18.12 41-50 12 8.05 >50 27 18.12 Total 149 100 Table 2: Type of parasitic infestations in different IS states Diagnosis E. histolytica/ G. lamblia Taenia E. dispar HIV infection 4 1 0 Cancer 6 3 0 IS therapy 2 1 0 DM 2 0 0 Severe anemia 5 2 1 PEM 9 11 0 TB 2 0 0 Chronic diseases 1 1 0 Total (number) 31 19 1 Diagnosis A. Hookworm T S. lumbricoides trichiura stercoralis HIV infection 2 0 0 0 Cancer 1 0 2 0 IS therapy 2 0 0 1 DM 0 0 1 0 Severe anemia 2 2 1 0 PEM 12 2 4 1 TB 2 1 1 1 Chronic diseases 1 1 1 0 Total (number) 22 6 10 3 Table 3: Comparison of presence of parasites and GI symptoms Parasite Parasite Total present absent GI symptom present 63 43 106 GI symptom absent 16 27 43 Total 79 70 149 Figure 1: Sex-wise prevalence of intestinal parasites. Females Males Positive 39 40 Negative 37 33 Note: Table made from bar graph. Figure 2: Comparison of direct, floatation and formol-ether concentration techniques for detection of different parasites. Positive Negative Direct 37.00% 63.00% Floatation 43.00% 57.00% Concentration 52.30% 47.70% Note: Table made from bar graph.
|Printer friendly Cite/link Email Feedback|
|Title Annotation:||Research Article|
|Author:||Bora, Ishani; Dutta, Vikramjeet; Lyngdoh, Wihiwot Valarie; Khyriem, Annie Bakorlin; Durairaj, Elanta|
|Publication:||International Journal of Medical Science and Public Health|
|Date:||May 1, 2016|
|Previous Article:||Radiological evaluation of various types of primary bronchogenic carcinoma: a study of 65 cases.|
|Next Article:||Knowledge, attitudes, and practices relating to dengue fever among high school students in Makkah, Saudi Arabia.|