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 ~  Abstract
 ~  Materials and me...
 ~  Results
 ~  Discussion
 ~  Acknowledgement
 ~  References

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ORIGINAL ARTICLE
Year : 2003  |  Volume : 21  |  Issue : 2  |  Page : 87-90
 

A study of diarrhoea among children in eastern Nepal with special reference to rotavirus


Departments of Microbiology, BP Koirala Institute of Health Sciences, Dharan, Nepal

Correspondence Address:
BP Koirala Institute of Health Sciences, Dharan, Nepal

 ~ Abstract 

PURPOSE: To examine the incidence of Rotavirus infection in children below five years of age. METHODS: Faecal samples from 160 children under five years of age with acute gastroenteritis were collected over a period of one year from July 1999 to June 2000. These were studied for the presence of Rotavirus antigen by enzyme immuno assay (EIA). RESULTS: Rota antigen could be detected in 62 (38.7%) samples. Co-infection with other parasites or bacterial pathogens in presence of Rota antigen was also demonstrated. Forty one (66.4%) children were admitted for hospital care. Forty two samples positive by EIA were further tested by latex agglutination (LA) to consider introducing this test routinely in clinical laboratory. Although a rapid and convenient test, LA failed to demonstrate antigen in 15(35.6%) of the samples. CONCLUSIONS: Rotavirus infection of children in Nepal is reported for the first time. EIA was found to be more sensitive than LA for the detection of Rotavirus antigen in faecal samples.

How to cite this article:
Shariff M, Deb M, Singh R. A study of diarrhoea among children in eastern Nepal with special reference to rotavirus. Indian J Med Microbiol 2003;21:87-90


How to cite this URL:
Shariff M, Deb M, Singh R. A study of diarrhoea among children in eastern Nepal with special reference to rotavirus. Indian J Med Microbiol [serial online] 2003 [cited 2019 Jun 24];21:87-90. Available from: http://www.ijmm.org/text.asp?2003/21/2/87/7981


Rotavirus is the most important cause of early childhood nonbacterial gastroenteritis in both developed and developing countries. The infection is also observed in older children and adults. In developed countries 50% of pediatric hospitalization are due to acute diarrhoea, while in developing countries it is responsible for an estimated one million deaths annually.[1],[2],[3] Drug therapy has little, if any, place in the management of viral diarrhoea and its prevention is obviously a long-term goal.
A few studies on occurrence of gastro-intestinal infections in Nepal are available. According to the Nepal Human Development Report published in 1998, 16 to 25% of childhood deaths occur due to diarrhoea. A study of travellers and foreign residents reported an incidence of Rotavirus in 8% of cases.[4] Group C Rotavirus infection was demonstrated from convalescent sera in patients with diarrhoea by others.[5]
Rotavirus is shed in large quantities in diarrhoeal stool. Therefore, nosocomial infection in paediatric wards and neonatal nurseries are known to occur and management is difficult. Consequently, an early and reliable identification of Rotavirus is an essential prerequisite for the control of potential outbreaks.
The aim of this study was to examine the incidence of Rotavirus infection in children less than five years of age. This study also provides a comparison of the incidence of Rotavirus infection with that of non-Rotavirus infection.

 ~ Materials and methods Top

Subjects
One hundred and sixty children under five years of age suffering from acute gastroenteritis who attended the out patient department and those who were admitted at the BP Koirala Institute of Health Sciences (BPKIHS), a tertiary care hospital situated in Eastern Nepal, were included in the study. Faeces of children showing the presence of blood and mucus (dysentery) were excluded from the study.
Methods
A single faecal sample was collected from each child with gastrointestinal complaints on reporting to hospital. Rectal swabs were discouraged. The relevant clinical data were entered in a proforma. The standard definition of malnutrition and dehydration were strictly followed.[6]
The faecal samples were subjected to naked eye examination for consistency, colour and atypical components (mucus, blood and parasites). The samples were examined microscopically for the presence of leucocytes, RBCs, mucus, fat and parasitic ova and cysts in saline and iodine preparations. Samples were also screened for the presence of Cryptosporidium and other newer protozoan parasites by modified Zeihl-Neelsen (Kinyoun's) acid-fast stain.[7]
The faecal specimens were subjected to culture in order to look for common enteropathogens. MacConkey, deoxycholate citrate (DCA) and thiosulphate citrate bile sucrose (TCBS), selenite F broth and alkaline peptone water were used for the isolation of bacterial pathogens including Vibrio cholerae. The bacterial isolates were identified by standard biochemical tests and by slide agglutination with polyvalent and monovalent sera.[8] The growth of  E.coli  , as pure/predominant flora was considered significant. However, characterization of E.coli could not be attempted due to the lack of facilities.
The presence of Rotavirus antigen was prospectively investigated by Enzyme immuno assay (Ridascreen® Rotavirus, R-Biopharm, Germany). Enzyme immunoassay (EIA) utilized monoclonal antibodies in a solid phase sandwich type ELISA. Plastic microtiter wells were coated with a monoclonal antibody directed against VP6, (group specific antigen of human Rotavirus). Faecal suspensions were added to the wells and incubated simultaneously with an anti Rotavirus monoclonal antibody conjugated to horseradish peroxidase, resulting in the Rotavirus antigen being sandwiched between the solid phase and enzyme-linked antibodies. The wells were washed to remove unbound enzyme labeled antibodies. The substrate (urea peroxide) and chromogen (TMB) were added to the wells. The enzyme bound in the wells converts the colourless substrate/ chromogen to a blue colour. Addition of stop solution converted the colour from blue to yellow. Wells were read spectrophotometrically using a 450 nm filter and the results were calculated according to the manufacturer's criteria. Samples with equivocal results were subjected to repeat tests.
Forty-two samples positive by EIA were also tested by Latex agglutination (Rotalex from Orion Diagnostica, Finland) according to the instructions of the manufacturer.

 ~ Results Top

One hundred and sixty children with acute diarrhoea were included in the study, of which 108 (67.5%) were male and 52 (32.5%) were female.
[Table - 1] shows the age wise distribution of children with diarrhoea and Rotavirus positivity. Rota antigen was detected in 62(38.7%) samples. In 9 (14.5%) samples antigen was detected in combination with parasite and bacterial agents.
The majority (70.9%) of infections were observed in patients between 6 months and 2 years of age. The clinical and laboratory parameters in the Rotavirus positive (Group I) and negative (Group II) children are shown in [Table - 2].
Vomiting (56%) and dehydration (76%) were predominantly seen in Group I, due to which more children in this group needed hospitalization (66%) as compared to their Rotavirus negative counterparts. Fever was seen in both groups. Presence of pus cells and RBCs were detected more often in Group II. Bacterial pathogens were isolated in 11% in Group I. These included six strains of E. coli and one strain of Shigella flexneri. In Group II bacterial pathogens were isolated in 17 (19%) samples which included 10 strains of E.coli, six strains of Shigella flexneri and one strain of Shigella boydii. One each of Cryptosporidium and Entamoeba histolytica cysts was seen in Group I. E.histolytica in three samples and Trichomonas in one sample were detected in Group II.
[Figure - 1] and [Figure:2] show the month wise distribution of the diarrhoeal cases and Rotavirus positivity. Infections occurred throughout the year though the intensity was not uniform and appeared to peak in late winter (Jan-Feb), and to a lesser degree, during the summer/ monsoon months (Apr-Aug).
Forty-two samples, which showed presence of Rota antigen by EIA, were further tested by the latex agglutination test. Of these, 15(35.6%) failed to produce positive results by LA, thus demonstrating its lower sensitivity.
Most children received antibiotics prior to reporting to the hospital. Ampicillin, cloxacillin, co-trimoxazole, chloramphenicol, cephalosporins, gentamicin, and metronidazole were used as monotherapy and also in combination. Some of these antibiotics were also used during their hospital stay, having been prescribed before test results were made available. During the hospital stay, no mortality was seen in children. However, the children were not followed up after they were discharged from the hospital.

 ~ Discussion Top

Rotavirus is the most important cause of severe, life threatening gastroenteritis in children, accounting for 20-50% of hospitalization for gastroenteritis in children worldwide.[3],[5],[7],[9] Rapid acquisition of antibodies is known to take place in children between 6 and 24 months. Information about the incidence and impact of Rotavirus infection in Nepal is scarce.[4],[5] A rapid diagnosis of Rotavirus by detection of Rota antigen provides the physician with useful information. Development of sensitive enzyme immunoassay (EIA), available in kit form has made diagnosis easy and widely available.[10] Latex slide agglutination (LA) technique has further simplified the diagnosis, making it a bedside procedure. Rapid diagnosis may prevent inappropriate treatment and help prevent spread of infection particularly in institutions.
The present study, which was attempted in a limited number of samples from cases of gastroenteritis in the under-five age group, showed an incidence of 38.7%. Concomitant infection with bacterial agents and intestinal protozoan was also observed. Associated signs and symptoms of vomiting, fever and dehydration observed among the Rota antigen positive cases were broadly comparable to findings reported in other studies.[2],[5],[9] The infections were seen throughout the year. An increase in the number of cases was observed during late winter and summer -monsoon months.
Sixty six percent children needed hospitalization owing to vomiting and dehydration as compared to 50% of Rotavirus negative cases. Drug therapy has little place in management of viral diarrhoea. However, it was observed that many of the children received antibiotics prior to attending hospital and also during hospital stay. Therefore, there is a strong possibility that certain bacterial infections were missed during the study period.
In conclusion, it was observed that Rotavirus is associated with an important proportion of diarrhoea cases in small children in Eastern Nepal. A number of these cases were severe enough to warrant admission to the hospital. Rotavirus infection is common in children. The control of spread is by improved hygienic measures in both home and in hospital environment and possibly immunization in some situations.
In summary, two methods of Rotavirus antigen detection were used in the present study. The enzyme immuno assay appeared to be more sensitive than the LA. Detection of Rotavirus has given an insight into the problem in this part of Nepal where no data of this problem are available. However, due to the high cost factor (approximately 130 Nepali Rupees per test) these tests may not be affordable for routine diagnosis and perhaps its use will remain limited in pediatric practice.

 ~ Acknowledgement Top

This study was partially funded by a BPKIHS research grant. 

 ~ References Top

1.Steinhoff MC. Rotavirus: The first five years. J Ped 1980;96:611-622.  Back to cited text no. 1    
2.Bantavala JE. The role of viruses in acute diarrhoeal disease. Clinics in gastroenterology 1979;3:569-598.  Back to cited text no. 2    
3.Cilia G, Perez-Trallero E, Pineiro LD, Iturzaeta A, Vicente D. Rotavirus gastroenteritis in Gipuzkoa, Spain. Emerging Infectious diseases 1999;6:834-835.  Back to cited text no. 3    
4.Taylor DN, Houston R, Shlim DRU, Bhaibulaya M, Ungar BL, Echeverria P. Etiology of diarrhea among travellers and foreign residents in Nepal. JAMA 1988;260(9):1245-1248.  Back to cited text no. 4    
5.Penaranda ME, Cubitt ND, Sinarachatanant P, Taylor DN, Likanonsakul S, Saif L et al. Group C Rotavirus infection in patients with diarrhea in Thailand, Nepal and England. J Infect Dis 1989;160(3):392-397.  Back to cited text no. 5    
6.OP Ghai, Nutrition pp37-52 & Fluid and electrolyte disturbances pp171-176 In- Essential Pediatrics, 2nd edition, 1990.  Back to cited text no. 6    
7.Henriksen SA, Pohlenz JFL. Staining of Cryptosporidia by a modified Ziehl-Neelsen technique. Acta Vet Scand 1981;22:594-96.  Back to cited text no. 7    
8.Forbes BA, Sahm DF, Weissfeld AS, Eds. Bailey and Scott's Diagnostic Microbiology. (The Mosby CV, Co. St. Louis) 1998.  Back to cited text no. 8    
9.Rao RS, Vijaylakshmi N, Amarnath SK, Ramesh I. Viral gastroenteritis: A review. Indian J Med Microbiol 1994;12(1):1-8.  Back to cited text no. 9    
10.Cukor G, Perron DM, Hudson R, Blacklow NR. Detection of Rotavirus in human stools by using monoclonal antibody. J Clin Microbiol 1984;19:888-892.  Back to cited text no. 10    
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