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Journal of Applied Biology & Biotechnology Vol. 1 (02), pp. 013-017, July- Aug, 2013 Available online at http://www.jabonline.in DOI: 10.7324/JABB.2013.1203 Gastroenteritis with Eschericha coli in pediatric hospital in N’Djamena-Chad
Bessimbaye N1,5, Tidjani A2*, Moussa AM1,2, Brahim BO2, Mbanga D 1,2, Ndoutamia G3, Sangare L4, Barro N5, Traore
AS5
1General Hospital of National Reference of N'Djamena (HGRN) 2Faculty of Human Health (FSSH), University of N'Djamena 3University of Doba, Chad 4Unit Training and Research in Health Sciences (UFR / SDS)University of Ouagadougou, University Hospital of Yalgado Ouedraogo of Ouagadougou (CHU-YO) 5Centre for Research in Biological Food and Nutritional Sciences (CRSBAN)University of Ouagadougou. ARTICLE INFO
ABSTRACT
Gastroenteritis with Escherichia coli diarrhea is severe and recurrent fatal diseases in children under 5 years especially in Africa. A prospective study of acute diarrhea was conducted at the General Hospital of National Reference of N'Djamena (HGRN). The aim of this study was to investigate the epidemiological, clinical and evolutionary characteristics of acute diarrhea with enteropathogenic Escherichia coli. This study involved 167 children aged 0-2 years having seen in outpatients or hospitalized for acute diarrhea in children. Stool samples were collected and analyzed according to standard procedures of clinical microbiology. Identification of
Key words:
pathotypes of E. coli was performed by Nonavalents serum agglutination, Trivalent I, Trivalent II, Trivalent III and mixture IV. Antimicrobial susceptibility was performed by disk diffusion method in solid medium according MH Kirby Bauer. Examination of 167 stool showed 26 cases (15.56%) bloody, 141 cases (84.83%), mucous, 122 cases (73.05%) and liquid 45 cases (26.94%) soft. The enteropathogenic Escherichia coli (EPEC) were detected in 58 cases (34.73%). On the other hand parasites (16.76%) were identified to direct microscopic observation that Trichomonas intestinalis 13 cases (7.78%), Giardia intestinalis 9 cases (5.38%), Entamoeba histolytica 6 cases (3.59%) and cases of intestinal candidiasis (15.56%). The sensibility of origins in antibiotics was varied, 89.65 % of the ECEP, were resistant in Sulpha drugs, 56% were it towards beta-lactamines. An average rate of 26% of resistance was observed by the families of fluoroquinolones and quinolones. This study allowed us not only to include the characteristics of diarrheas at the children of less than two years, but also, the necessity of an effective coverage as well as the implementation of a program of prevention against the diarrheas of the children in a hospital environment. 1. INTRODUCTION
380,000 deaths per year. They produce heat-labile and heat-stable Diarrhoeal diseases are very common in the world, enteroadherents (EAAGG, DAEC) are responsible for chronic especially in developing countries where they are endemic state diarrhea in children in underdeveloped countries and produce an [1]. Currently diarrheal diseases arrive in third place among the enterotoxin and hemolysin [7]. The entero-invasive Escherichia most deadly infectious diseases in the world with 2.5 million coli (EIEC) cause bloody diarrhea [8; 9]. Currently, deaths any confused age. In terms of years of life lost adjusted Escherichia coli are considered emerging pathogens, the leading for disability, diarrhea come in second with 73 million behind cause of bacterial diarrhea in children and are a major problem of public health [10]. Instead of Eschericha coli diarrhea in Escherichia coli (STEC) are responsible for foodborne infections developing countries, particularly in Chad is not well known that result in diarrhea but also more serious syndromes in man as because very few studies have been devoted. Chad has few hemolytic uremic syndrome that can cause death [3;4]. reliable national statistics on the most frequently encountered Enterotoxigenic Escherichia coli (ETEC) are an important cause enteropathogens in diarrheal diseases. However, data from the of diarrhea in children in developing countries and an estimated registers of stool cultures of some hospitals and typhoidal perforation recorded in the service of visceral surgery, these different hospitals attest diarrhoeal disease due to e-mail : abdelti@yahoo.fr /abdelti@gmail.com enteric pathogens (bacteria, viruses, parasites and fungal agents) 2013 Bessimbaye N et al. This is an open access article distributed under the terms of the Creative Commons Attribution License -NonCommercial-ShareAlike Unported License (http://creativecommons.org/licenses/by-nc-sa/3.0/). Bessimbaye et al. / Journal of Applied Biology & Biotechnology 1 (02); 2013: 013-017 posed a real public health problem in Chad. The objective of this strain (rough). In the absence of agglutination, we proceed in the work is to contribute to better understanding the epidemiological same way with Nonavalents serums, Ttrivalent I, Ttrivalent II, characteristics of diarrhoea due to Escherichia coli in pediatric Trivalent III and mixture IV (Bio-Rad®). The antibiotic sensitivity hospital and the phenotype of the antibiotic susceptibility of the tests were performed on solid Mueller-Hinton (Kirby-Bauer) by epidemiological characteristics of Escherichia coli diarrhea in diffusion method using discs impregnated with known antibiotics pediatric hospitals and the phenotype of antibiotic sensitivity. as recommended by the Committee on Antimicrobial Technique doses of the French Society of Microbiology (CASFM). 58 strains 2. MATERIALS AND METHODS
of E. coli were tested with six antibiotics belonging to the family of beta-lactam antibiotics (amoxicillin (25μg): Amoxicillin + 2.1 Period of the study site
clavulanic acid ((20/10 μg), Ceftriaxone (30 μg), quinolones This is a prospective study carried out between 01 (nalidixic acid (30 μg), fluoroquinolones (ciprofloxacin (5 µg) and November 2009 to 30 October 2010, which took place in the sulfonamides (trimethoprim-sulfamethoxazole (SXT) (1.25/23.75 Pediatric Unit and Bacteriology Laboratory of General National Hospital of Reference in N'Djamena (HGRN). 2.6 Data processing
2.2 Population
The data was entered and analyzed using Excel software. The study involved 167 children aged 0-2 years, 97 girls Statistical analysis was performed using the chi-square test (χ ²) for (58.08%) and 70 males (41.91%) with acute diarrhea in comparison of two variables. Differences were considered hospitalized or come visit the Pediatric Department of HGRN. Cases of acute diarrhea were determined according to the WHO definition, namely the emission of at least three loose or liquid 3. RESULTS
stools per day lasting for less than fourteen days. 3.1 Epidemiological characteristics
2.3 Investigations
During the study period, out of 167 cases of children of Investigations were made to parents or guardians of 0-2 years of age with acute diarrhea reported in Pediatrics, 107 children after obtaining their informed consent. They were: the (64.07%) were hospitalized and 60 (35.92%) were followed on an origin, sex, age, date of disease, ongoing treatment, number of outpatient basis. The evaluation of the temporal distribution of stools per day, education level of the mother and the suspected diarrhea is shown in Fig. 1. It appears a respective predominance origin of the contamination (the source drinking water, food and of diarrhea cases in the months of January, March and June of whole way of life of parents) signs and symptoms (diarrhea, 2010 with 18 (11%), 24 (14.4%) and 26 (16%) cases, respectively. abdominal pain, vomiting, fever, etc.) and conditions associated The number of Escherichia coli isolated from the same month enteric was 7 (12.1%), 9 (15.5%) and 10 (17.2%) respectively. The average age of children was 12 months and ranged from 1 day to 24 months (Table 1). Cases of diarrhea were mostly observed in 2.4 Sampling and analysis of stool
Feces were collected in sterile jars. Each sample was examined for macroscopic aspects (Purulent, mucus, presence of Table 1: Distribution of cases of diarrhea according to age and sex.
blood) and consistency (liquid, soft, molded). It has been Age classes
Total (%)
Female (%)
researched parasites in association with diarrhea. The stools were also processed and analyzed using standard methods of clinical microbiology. The agar EMB (Eosin Methylene Blue) and MH (Muller-Hinton) (Bio-Rad ®) were used for the isolation of Escherichia coli. The Petri dishes containing the medium seeded were incubated at 37 °C in an oven for 24 hours. The reflective 3.2 Clinical Features
metallic green colonies were searched and re-isolated on MH for Fifty six (56) children (33.53%) were taken to the the subsequent identification of enteropathogenic Escherichia coli. consultation within 1 to 3 days after event emission liquid stools The isolates were stored at -86 °C in brain heart broth with 15% while 71 (42.51%) were found within 4 to 7 days and 40 (29.95%) glycerol for future molecular characterization. were seen in consultation after the first week. Total duration of diarrhea could be specified in 114 children: it was 5.8 days on 2.5 Serotyping and antimicrobial susceptibility
average with the extremes of 1 to 14 days. Table 2 reported the On pure cultures of E. coli, it was proceeded to results of the manifestations of diarrhea in the group of children serotypagee as recommended by WHO. For this a colony of observed. The number of stools per day ranged from 3 to 18; bacteria was deposited on a drop of saline water (0.85% Nacl) on a Altogether 86 children (51.49%) expressed 3-5 bowel movements slide and crushed. Agglutination indicates the presence of a rough per day, 65 children (38.92%) were 6-10 stools and 16 (9, 58%) Bessimbaye et al. / Journal of Applied Biology & Biotechnology 1 (02); 2013: 013-017 more than 10 stools per 24 days. The stools were bloody in 26 there are 47 (81.03%) of trivalent 1 (O111 + O55 + O26) and 11 cases (15.56%), mucous in 141 cases (84.83%), liquid in 122 cases (18.96) enteropathogenic Escherichia coli (EPEC). Twenty-eight (73.05%) and soft in 45 cases (26.94%). Table 3 shows the cases (16.76%) of diarrhea parasites were identified to direct distribution of signs and conditions associated with diarrhea. Fever microscopic observation of which 13 cases (7.78%) Trichomonas was the most common symptom associated with 55.68% of cases, intestinalis, 9 cases (5.38%) of Giardia intestinalis and 6 cases followed by vomiting in 72 cases (43.11%) and anemia in 15 cases (9%). Acute respiratory infections were the most common Candida albicans were isolated in 26 cases (15.56%). ailments associated with diarrhea (24.55%) followed by We highlighted six (3.6%) when combining Entamoeba gastrointestinal candidiasis (15.56%) and malnutrition (10.17%). histolytica / Giardia intestinalis and 4 (2.4%) when combining Escherichia coli / Candida albicans, an overall prevalence of Table 2: Distribution of cases of diarrhea by age according to the appearance
Age classes
Consistence
3.4 Evolution
Thirteen cases (12.14%) deaths were recorded among 107 hospitalized children. The evolution towards healing was observed in 81 children (75.70%). Cases of escape were noted in 13 children (12.14%) escaped. The distribution function and evolution of nutritional status is reported in Table 4. The analysis shows that 61.53% of deaths (8 of 13) occurred in malnourished Table 3 : Signs and diseases associated with diarrhea.
children. The observed difference is statistically significant. The Signs and symptoms
Percentage (%)
risk of death was significantly greater in malnourished children than those with a normal nutritional status. Table 4: Distribution of cases according to the evolution of the nutritional
Affections Case Percentage (%)
Evolution
Nutritional status
Malnutrition
3.3 Etiologic agents
3.5 Susceptibility testing
E. coli was isolated in 34.73% of cases of diarrhea. Table 5 shows the evaluation of the effectiveness of 65.27% of cases were due to parasitic infections, acute respiratory, antibiotics with isolated bacterial agents. The study involved six candidiasis and infection Ear Nose and Throat with proportions of antibiotics tested on 58 enteropathogenic Escherichia coli isolated. 16.76%, 15.56%, 24.55 % and 8.37% respectively. Of 167 The proportions of resistance (R), intermediate (I) and sensitive cultures performed, 58 enteropathogenic Escherichia coli were (S) were obtained (Table 5). 56% of strains were resistant to beta- isolated (34.73%), 46 cases (42.99%) among 107 children lactam antibiotics tested. The quinolones and fluoroquinolones hospitalized and 12 cases (20%) among children monitored were very active on 64% and 85% respectively of the strains. externally. Of the 58 serotypes isolated from Escherichia coli, Figure 1: Monthly distribution of cases of diarrhea and enteropathogenic Escherichia coli.
Bessimbaye et al. / Journal of Applied Biology & Biotechnology 1 (02); 2013: 013-017 Table 5 : Effectiveness of some antibiotics of the family of beta-lactam antibiotics, sulfonamides, quinolones and fluoroquinolones
Nb tested
R = Resistance, I = Intermediate, S =Sensitive: Trimethoprim-sulfamethoxazole (SXT) : R < 1 0, 10 ≤ I≤ 18, S ≥ 19; Nalidixic acid (Nal) : R< 15, 15 ≤ I≤ 19, S ≥ 20 ; Ciprofloxacin (Cip) : R< 15, 15 ≤ I≤ 20, S ≥ 21 ; Ampicillin (AM) :R<11, 11≤ I≤ 16, S≥ 17 ; Ceftriaxone (CRO) : R< 15, 15 ≤ I≤ 20, S≥ 21 ; Amoxicillin (AMX) : R< 14, 14 ≤ I≤ 20, S≥ 21(NCCLS, 1998, CASFM, 2010). 4. DISCUSSION
EIEC (0.4%). The identification of parasites such as Trichomonas Diarrhoeal diseases are still a leading cause of intestinalis and Giardia intestinalis in liquid stools in our study hospitalization of children in developing countries. In this study was also observed. By cons, in mucoid-bloody stools, were children 0-6 months were the most affected by diarrhea 37.72% of frequently reported the presence of Entamoeba histolytica. This cases. Maaroufi et al. [11] in Tunisia reported higher frequencies would explain why, from the appearance and consistency of stools, of 55.7%. The child before one year of age are particularly one might suspect the presence of such parasites or bacterial vulnerable to diarrhea for two main reasons. It was during this infection such. The frequency of 16.76% of parasites found in the period that gradually develops the child's own immunity as it faeces in our study is relatively higher than that reported by Diouf occurs a decline in maternal antibodies. The infant is more et al. [21] in Senegal, which was 10.7%. Gendrel [22] in Gabon vulnerable to infections during this period [12]. This age found a 15% prevalence of carriage in disadvantaged children. corresponds to the period when food diversification of the child Entamoeba histolytica were found at a low rate of 3.59% of cases. begins. When it is poorly managed, malnutrition and diarrhea may The 6% of parasite/parasite or bacteria/candidiasis association occur [13].The distribution of diarrhea by sex and age shows a could be explained by the level of adverse social parents to Chad predominance of diarrhea cases in the age group of 0-6 for females or lack of food hygiene at the household level. Acute respiratory (20.35%) than males (14.97%) but difference is not statistically infections with 24.55% were disease most associated with significant (x2 = 0.00781, df = 1; p ≥ 0.90).This could be explained diarrhea. It is followed by malnutrition in 11.37% of cases. by the fact that Chad, women's vital statistics are 51% against 49% Kangah et al. [23] in Côte d'Ivoire reported a rate increase of 22% for male according to INSEED (National Institute of Statistics and association of malnutrition with diarrhea. These two conditions Economic and Demographic Studies) in 2010. Barro et al. [14] feed off each other, creating a vicious circle of malnutrition reported the infection rate among women (52.7%) which is slightly increases the risk of diarrhea and diarrhea worsens malnutrition higher than men (47%) during an etiological study of [24; 25]. The average time taken to send the children in gastroenteritis at the National Hospital Center Universitaire consultation noted in this study, 33.53% of children were Yalgado Ouedraogo (CHU-YO) of Ouagadougou, Burkina Faso. consulted dice the first 3 days of diarrhea and two in five children The rate of 15.56% of bloody diarrhea in our study is slightly after a week reflects the neglect of diarrhea and ignorance of its higher than the 10.2% reported by Diagne et al. [15] in Senegal. consequences for mothers. This could be explained by the The mucous stools were observed in 84.83% of cases and loose conditions of precarious socio-economic life of the population in stools in 73.05% of cases. The different aspects and consistency of Chad. Access to health care due to the level of socio-economic life stools were also reported by Dao et al. [16] at Bamako in Mali : has also been reported by Poda et al. [26] in Burkina Faso. Most aspects mucus in 14 cases (18.4%), the consistency of watery recorded deaths in our study occurred in malnourished children (x2 stools in 40 cases (52.6%), and polymorphic consistency in 8 cases = 21.27, df = 1; p ≥ 0.01 significant difference). The association is (10.5%). Enteropathogenic Escherichia coli were much more diarrhea-malnutrition in our areas of major concern. These deaths isolated in the stools liquid consistency. Ingested bacteria adhere could be explained by the fact that the department of pediatrics of to enterocytes (intestinal cells) by a factor attachment and secrete HGRN does not have a Therapy Centre Oral Rehydration (CTRO) enterotoxin (ST a thermostable toxin and heat-labile toxin LT) for the management of children with diarrhea and a Center for which are responsible for watery diarrhea [17]. The results of stool Recovery and Nutritional Education (CREN) dealing with cultures were positive in 37.12% of cases. This rate is three times malnourished children. Most isolates were resistant to beta-lactam that observed Luki et al. [18] in Zaire, it was 12%. The high rate in antibiotics (Ampicillin, amoxicillin, ceftriaxone) at an average our study could be explained by an unsatisfactory immunization ratio of about 56%. Such resistance when observed, is linked to the status, parental neglect due to present infants in health centers. production of beta-lactamase Amp C types and penicillinases by The frequency of enteropathogenic Escherichia coli isolated is the strain in question [27; 28]. Our results can be explained by this relatively high (34.73%)) in our case study. Sanou et al. [19] phenomenon. This increase in resistance is due to the improper reported a rate of 32% at the University Hospital Yalgado and inappropriate prescribing of antibiotics by health workers. Ouedraogo of Ouagadougou in Burkina Faso. Bonkoungou et al. This is what was observed during our investigation. By cons, our [20] showed that the major pathogroupes encountered is the ECEA strains manifested good sensitivity to quinolones (64%) and (12%), followed by EPEC (8%), ETEC (4%), ECST (0.4%) and fluoroquinolones (85%). Strains have developed a resistance rate Bessimbaye et al. / Journal of Applied Biology & Biotechnology 1 (02); 2013: 013-017 of 89.65% to trimethoprim-Sulfamoxazole. On the other hand, it 12. Hoerr RA. Book Review: Nutrition and Immunology: Principles and appears, still, according to our surveys, that about 8% of mothers Practice. Am.J.Clin.Nutr. 2001; 73: 1114-1115. practiced self-medication by giving their child drugs such as 13. Scrimshaw NS. Historical concepts of interactions, synergism and amoxicillin, trimethoprim-sulfamethoxazole and metronidazole antagonism between nutrition and infection. J of Nutr. 2003; 33: syrup before to the hospital. Moreover, Kam et al. [29] reported 14. 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food, use of GMOs, food risk analysis. From 8 to 11 November 2005; CIDEFA 113-118.Ouagadougou, Burkina Faso. The results of this study show that enteropathogenic 15. Diagne I, Camara B, Diouf S, Ba M, Ndiaye O, Cisse MF, Sow HD, Sall MG, Sarr ., Kuakuvi n, Fall M. Infantile diarrhea in hospital Escherichia coli, representing 34.73% of all diarrheal diseases practice: the experience of Pediatrics of the University Hospital of indicating the public health problem that cause enteric pathogens Dakar, epidemiological and biological Clinical Aspects. Med Afr Several other parasites and enteric pathogens were 16. Dao S, Oumar AA, Doumbia, Goita D, Boushab M, Maïga II, Bougoudogo F. Emiotic, etiologic and prognostic aspect of diarrhea identified in the stool. However, it has not been sought enterovirus in AIDS hospital in Bamako.J Mali Med. 2007; 22 : 1-4. in the stool. In this study, the antibiotics tested groups . L avigne JP. Course of Microbiology, Faculty of Medicine showed a very high resistance in beta-lactam and low in Montpelier-Nîmes: Diarrhea-Diagnosis, techniques and interpretation of bacteriological examinations. 2005; 1-3. Quinolones and Fluoroquinolones. Actions favoured are needed to 18. Luki N’Ganda Y, Muyembe L, Krubwa Foemi O, Busmans E. protect the health of infants, highly vulnerable to diarrheal Bacterial and viral etiology of childhood acute gastroenteritis in Kinshasa (Zaire). Ann soc belge Med Trop. 1986; 66 : 3331-3337. 19. Sanou I, Kam KL, Tougouma A, Sangare L, Nikiema JHP. Acute 6. REFERENCES
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Enterohaemorrhagic Escherichia coli: emerging issues on virulence Bessimbaye N, Tidjani A, Moussa AM, Brahim BO, Mbanga D, and modes of transmission. Vet. Res. 2005; 36 : 289-311. Ndoutamia G, Sangare L, Barro N, Traore AS. Gastroenteritis 11. Maaroufi S, Ben Dridi MF, Ben Chaabane T, Zaimi ., Boujnah A, with Eschericha coli in pediatric hospitals in N’Djamena-Chad. J Zribi A, Ben Osman. Epidemiology of acute infantile diarrhea. App Biol Biotech, 2013; 1 (02): 013-017.

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