1. INTRODUCTION:

The diarrhea is defined as an elevation in the occurrence of bowel movements, an elevation in the looseness of stool or both. It is abnormal loss of the feces more than three times within 24 hours with the creation of liquid or semi-liquid feces resulting in fluid loss [1].

Diarrhea is caused by life-threatening or non-life-related causes [2].

The majority cases of diarrhea are caused by microbes. Diarrhea is think to be result from direct interaction between bacteria and bowel epithelial cells. The most common symptoms of diarrhea are poor absorption of vitamins, carbohydrates, fats, calcium, proteins in the alveolar and fasting regions and injure to the lining of the intestines. The microbes responsible for this infection include: bacteria, viruses and parasitic organisms [3].

E. coli is a main element of the intestinal flora of humans and other mammals. E.coli that cause diarrhea are extremely common worldwide E. coli accounts780-900 million diarrheal cases of global annually. The E.coli strains are classified by the characteristics of their virulence properties, and each group causes disease by a different mechanism, at least six of which have been characterized [4].

Among six diverse classes of pathogenic E. coli, ETEC, EPEC and EIEC are responsible for the most common of diarrheal cases in the developing world [5]. E.coli pathogenic forms can cause a diversity of diarrheal infections in hosts according to the presence of specific colonization factors,virulence factors and genes that associated with pathogenicity which are commonly not found in other E.coli strains .The diseases caused by a particular strain of E. coli depend upon expression and distribution of an array of virulence factors, including toxins, invasins, adhesins, and capability to withstand host defenses [6].

2- MATERIALS AND METHODS:

2-1. Collection of stool sample:

One hundred fifty stool samples were collected from patients suffering from significant diarrhea during the period of October 2017 to the end of December 2017 from three main hospitals in Hilla city in Iraq (Hilla Teaching Hospitals, Babylon Hospital for maternity and children, and Teaching Merjan Hospital). The stool specimens were collected in sterile containers.

Patient data were collected with their medical records reviewed for each patient which include: name, age, gender, taking antibiotic, and date of collection.

2-2 Cultivation and identification:

A calibrated loop (0.01 ml) of all stool suspension were cultured on MacConkey and blood agar plates. They were incubated overnight at 37˚C in bacteriological incubators under aerobic conditions.

Depending on morphological features of the colonies and microscopically examination with Gram^'s stain, a pure cultures on MacConkey agar plates and eosin methylene blue (EMB) agar plates were made from each single group of colonies. All isolates were identified by standard biochemical reactions that mentioned in MacFaddin (2000).

2-3 Serotyping:

E.coli positive cultures were set further serological test by using slide agglutination test with E.coli antisera (HKM company, China ) of E.coli separately. A slid agglutination test was performed according to company by transfer a small amount of bacterial mass from colony of a pure culture onto a glass slide containing drop of normal saline. The bacterial mass under test was emulsified with a loop in the drop of saline to give a smooth, fairly dense bacterial suspension. To this drop added a drop of antiserum and then mixed. The slide was rotate for one minute. The agglutination was detected with the naked eye through holding the slide against a dark background using indirect light source.

3- RESULTS AND DISCUSSION:

Because of the importance of E. coli to the intestine in the occurrence of watery diarrhea in infants so it was necessary to identify the strains belonging to them and using specific antisera, which depends on the identification of somatic antigens (O antigen) and capsular antigen (K antigen) using the serological method by agglutination test.

In present study the high majority of E.coli isolates that isolated from patients with diarrheal stool are belong to Enterotoxigenic E.coli (ETEC) that represent about 16/46(34.8%) pathotypes followed by Enteropathogenic E.coli (EPEC) that represent about 12/46(26.1%) followed by Enteroinvasive E.coli (EIEC) that represent about 10/46:(21.7% ) then followed by Enterohemorrhagic E.coli (EHEC) that represented by (O157) about 8/46(17.4%).The serotypes were mentioned in table (3-1).

By using serological test alone for identification of ETEC isolates belonged to 8 different serogroups, of which O6:K15 was the most prevalent. ETEC were the most common pathogen among patients with diarrhea. In fact, the ETEC pathotype is recognized as one of the main important diarrheagenic E.coli in developing countries. High rates of ETEC have been previously reported in North Africa especially in Egypt where a strong association with diarrhea has been shown (8)

The diarrhea caused by enterotoxigenic E.coli (ETEC) result in significant morbidity, and are economically important diseases of humans (9).

Also EPEC isolates belonged to 11 different serogroups, of which O128:K67 was the most prevalent. This result agreed with another study in the south of Iran (10) that reported EPEC rate of 30.7% for this path types in patients with diarrhea, in it the O128 were the most prevalent. While the rate of EPEC in Tehran hospital was 26.7% (11).

Enteropathogenic E.coli (EPEC) is the leading cause of diarrhea in infants. EPEC infection is primarily a disease of infants younger than 2 years of age in developing world (12).

The EIEC isolates that belong to 9 different serogroups, of which O29: K? was the most prevalent.

Table (3-1): serogroups and serotypes of E.coli in 46 isolates.

Serogrops	Number	Serotypes
ETEC	16	O6:K15, O7:K1, O7:K1, O9:K9, O6:K15, O6:K15, O6:K15, O15:K?, O6:K15, O25:K19, O6:K15, O78:K80, O7:K1, O9:K9, O25:K19, O78:K80,
EPEC	12	O111:K58, O86:K61, O128:K67, O128:K67, O111:K58, O128:K67, O128:K67, O111:K58,O128:K67, O86:K61, O26:K60, O55:K59
EIEC	10	O136:K78, O29:K?, O143:K?, O28:K73, O29:K?, O112:K66, O29:K, O28:K73, O112:K66, O29:K?,
EHEC	8	O157

In this study, also subdivided the results according to the age and sex factors. We detect the ETEC in age factor are 8 adults and 8 children while according to sex factor were 8 male and 8 female. Also the EPEC in age factor are 2 adults and 10 children while according to sex factor were 7 male and 3 female. And the EIEC in age factor are 10 adults and 2 children while according to sex factor were 7 male and 5 female. While the EHEC in age factor are 3 adults and 5 children while according to sex factor were 7 male and 1 female.

CONCLUSION:

We are concluded that the more serogroup was ETEC while the more serotypes was O6: K15.

ABBREVIATION:

ETEC: Enterotoxigenic E.coli.

EPEC: Enteropathogenic E.coli.

EIEC: Enteroinvasive E.coli.

EHEC: Enterohemorrhagic E.coli.

O antigen: Somatic antigen.

K antigen: Capsular antigen.

H antigen: Flagellar antigen.

EMB: Eosin methylene blue.

E.coli: Escherichia coli

ACKNOWLEDGMENT:

I would like to thank my supervisor Prof. Dr. Jawad K. Al Khafaji, and Asst. Prof. Dr. Hadi F. Al Yasari.

EHICAL APPROVAL:

Verbal consent was obtained from each patient before collection of samples.

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Received on 08.09.2018 Modified on 11.12.2018

Research J. Pharm. and Tech. 2019; 12(7): 3347-3349.

DOI: 10.5958/0974-360X.2019.00564.X