ORIGINAL

Niger J Paed 2012; 39 (3): 124 - 127

Ogunlaja A

Orelaru OF

Prevalence of aeromonas species and

escherichia coli in stool samples of

primary school children in Ofada,

Ogun state, Nigeria.

DOI:http://dx.doi.org/10.4314/njp.v39i3.7

Accepted: 17th February 2012

Abstract Background: Diarrhoea is stool samples yielded other patho-

one of the main causes of mortality gens such as Salmonella species,

and morbidity in childhood. Bacte- Shigella species, and Proteus vul-

rial diarrhoea is a common disorder. garis. Aeromonas spp was not iso-

Aeromonas species and Escherichia lated. Pathogenic E. coli was more

coli (E. coli) are some of the aetio- prevalent among pupils aged 11 to

logical agents associated with diar- 15 years. The susceptibility of E.

Ogunlaja A (

) Orelaru OF

Biological Sciences Department,

College of Natural Sciences,

Redeemer’s University,

Redemption camp, Mowe,

Ogun State.

rhoea in children.

coli to commonly used antibiotics

E-mail: aigbomere@yahoo.com

Tel :+2347031822517

Objective: To determine the preva- (such as tetracycline and ampicillin)

lence of Aeromonas species and E. was < 30% compared to > 70% for

coli in the stool samples of primary Augmentin, ofloxacin and nalidixic

school children as well as the anti- acid. E. coli showed 100% suscepti-

biotic sensitivity pattern of the iso- bility to only gentamicin.

lates.

Conclusion: E. coli was the pre-

Methods: Stool samples were ob- dominant bacterial agent isolated

tained from pupils (diarrhoeic and from stool samples of school chil-

non- diarrhoeic) aged four to eight- dren in Ofada, Ogun State. Gen-

een years in two primary schools in tamicin may be recommended as an

Ofada. The stool specimens were effective antibiotic agent against E.

processed by standard microbi- coli diarrhoeal diseases in the popu-

ological methods and antibiotic lation studied.

susceptibility was determined using

disc diffusion method.

Keywords: Aeromonas, antibiotics,

Results: Out of 104 stool samples, children, diarrhoea, Escherichia

E. coli was isolated from seven coli.

(

6.73%) stool samples while three

Introduction

tion suggests that poor hygiene and the ingestion of

contaminated food may be more important in the causa-

tion of diarrhoeal diseases. Some of the other important

risk factors for diarrhoeal diseases include poverty, poor

access to health care services, poor nutrition and over-

Diarrhoeal diseases constitute an important cause of

childhood morbidity and mortality worldwide, most

especially in the developing countries where sanitation

1

4

is suboptimal. The major factors predisposing to diar-

crowding as obtains in day care centers. In most devel-

rhoeal diseases include limited access to good quality

water, poor food hygiene and poor sanitation. The inci-

dence of diarrhoeal diseases might have reduced partly

because of improved public health measures, improved

hygiene, improved case management and increased use

of Oral Rehydration Therapy (ORT) as₂ well as better

understanding of the role of nutrition Contaminated

water was hitherto, suspected to be a leading predispos-

ing factor to diarrhoeal diseases. However, efforts at

improving the quality of water and sanitation observed

in other regions outside Africa, have no₃t yielded signifi-

cant reduction in diarrhoeal morbidity. This observa-

oping countries, the use of improved hygiene, improved

sanitation and improved quality of water, may not be

sufficient in the prevention of enteric illnesses. The

widespread indiscriminate use of antibiotics also allows

the pathogens to develop resistance to commonly used

antibiotics such as tetracycline and ampicillin among

others.

Aeromonas and E. coli are some of the aetiological

5

agents associated with diarrhoea in children. Other

agents include Shigella, Salmonella, Campylobacter and

Yersinia. Aeromonas species, formerly believed to be an

1

25

opportunistic organism capable of infecting only immu-

nodeficient individuals like children, have recently been

associated with both gastrointestinal disease and extra-

observed for growth and distinctive cultural characteris-

tics. The resultant colon₁i₁es were identified using stan-

dard biochemical tests. Specifically, Gram-negative

isolates were identified using different biochemical tests

such as oxidase test, indole test, urease test, citrate utili-

zation test, motility test, gas production and sugar fer-

mentation test. All the te₁s₂t_,s₁₃were conducted using stan-

dard laboratory methods.

6

intestinal ₇complications like the haemolytic-uraemic

syndrome.

Aeromonas diarrhoea is a mild, self-limiting infection.

Isolation rates for Aeromonas range from less than 1%

to more than 60% in diarrhoeic populations in various

8

geographic locations. A study had also shown that a

Antimicrobial susceptibility testing

small percentage of diarrhoeal disease₉s in Nigeria can be

attributed to Aeromonas hydrophilia.

Sensitivity of isolates to antimicrobial agents was deter-

mined on Muel₁le₂ r-Hinton agar plates using the disc dif-

fusion method. Interpretation of results was done by

measuring the zone sizes on the agar plates. Zone di-

ameter above or equal to 12mm defined susceptibility to

the respective antibiotic while zone diameter below

The aim of the study was to determine the prevalence of

Aeromonas species and E. coli in the stools of primary

school children and to determine the antimicrobial sus-

ceptibility profiles of these pathogens in two population

groups in Obafemi-Owode local government area, Ogun

State, Nigeria.

1

4

12mm defined resistance to the respective antibiotic.

All the isolates were tested for sensitivity to the follow-

ing antibiotics: ampicillin (10µg), tetracycline (10µg),

gentamicin (10µg), cotrimoxazole (25µg), streptomycin

(

10µg), nalidixic acid (30µg), nitrofurantoin (200µg)

and colistin (25µg), all of Abtek Biologicals Ltd, UK.

Subjects and methods

Study population and sampling

The study population was drawn from randomly se-

lected pupils in two primary schools (St. David Angli-

can Nursery and Basic School and Imedu Nla Nursery

and Primary School) at Mowe, Obafemi-Owode Local

Government Area, Ogun State, Nigeria. Consent for

inclusion in the study was obtained from the pupils and

their parents through the respective head teachers.

Results

Aeromonas spp. was not isolated from the 104 stool

samples analyzed. The isolates included E. coli from

seven (6.73%) stool samples as well as Salmonella spe-

cies, (Table 1). There was no considerable difference in

the total number of isolates from diarrhoeic (watery or

mucoid) and non-diarrhoeic (formed or semi-formed)

stool samples. Watery stools yielded the highest number

of E. coli isolates (Table 2). In addition, E. coli was pre-

dominantly found in both sexes between 11 and 15 years

of age (Figure 1).

Determination of the Sample Size:

Using formula: n =

Pq

E / Z)

2

(

1

0

Where: p = prevalence obtained from a previous study

15%

q = 100 - p = 100 – 15 = 85

=

Table 1: Enteropathogens identified in stool samples

E = Error margin of 5% ; CI = 1.96 ; Z = Standard nor-

mal distribution at 95%

Pathogen

School

A

School B

(n=59)

To-

tal

%

n=104

n = desired sample size

(n=45)

Therefore: n = 195

Freq

4

%

Freq

%

Sampling technique: 250 sterile universal sampling bot-

tles were randomly distributed to consenting pupils irre-

spective of their health status. Relatively equal number

of males and females were selected in each class and

school. Overall, a total of 104 pupils (53 females and 51

males) returned stool samples for the study.

Media: The culture media used in this study included

MacConkey agar, Kliggler Ion agar, Simon Citrate agar,

Motility agar, Nutrient agar, Mueller Hinton agar and

Peptone water.

E.coli

8.8

3

5.1

7

6.7

Shigella

1

-

2.2

-

1

0.9

Salmonella

1

1.6

-

Proteus

vulgaris

Aeromonas

-

1

-

1

0.9

-

Bacterial isolation and identification:

Total

10

9.6

Each stool sample was enriched in peptone water and

sub-cultured on MacConkey agar. This was then incu-

bated at 37 C for 18-24 hours. The culture plates were

School A -St. David Anglican Nursery and Basic school,

School B - Imedu Nla Nursery and Primary School.

0

1

26

Table 2: Appearance of stool Samples and Frequency of

Fig 1: Fig 1: Bar chart showing the relationship be-

tween the frequency of isolation of Escherichia coli and

the age distribution of the pupils.

Isolation of E.coli

School A School B

No. of

Isolates

No. of

Isolates

Total No

Isolates

4

3

2

1

0

Appearance of Stool

Watery Stool

1

-

2

-

3

-

Watery and Mucoid Stool

Formed

2

1

4

-

2

7

Semi-formed

1

3

Total No. of Samples

School A - St. David Anglican Nursery and Basic School,

School B - Imedu Nla Nursery and Primary School.

Antimicrobial Susceptibility Patterns

Table 3 shows the antimicrobial susceptibility pattern of

the four bacterial species tested: E. coli, Salmonella spe-

cies, Shigella species and Proteus vulgaris, against eight

antibiotics. The isolates showed the least susceptibility

to tetracycline and ampicillin. The overall susceptibility

of the isolates to these antibiotics was less than or equal

to 30%. All the isolates showed 100% susceptibility to

gentamicin. Generally, the isolates showed overall sus-

ceptibility of 90% to ofloxacin and nalidixic acid. Aug-

mentin and nitrofurantoin were also effective against

most bacterial pathogens except Salmonella species.

≤⁵

6-10

AGE (yr)

11-15

>15

Cotrimoxazole was only effective on Salmonella species

and Proteus vulgaris and the overall susceptibility of the

isolates to cotrimoxazole was 40%. Resistance to more

than two antibiotics was observed in relation to four

57.1%) E. coli species, one (100%) Shigella species

and one (100%) Salmonella species.

(

Table 3: Percentage antimicrobial susceptibility patterns of enteric bacteria from sampled population.

Organism

OFL

AUG

COT

NIT

NAL

GEN

AMX

TET

Escherichia coli (n=7)

6 (85.7)

5 (71.4)

3 (42.9)

4 (57.1)

6 (85.7)

7 (100)

2 (28.6)

1 (14.3)

Salmonella (n=1)

Shigella (n=1)

1 (100)

0

1 (100)

0

1 (100)

0

1 (100)

0

Proteus vulgaris

1 (100)

0

1 (100)

0

(

n=1)

Overall Susceptibilty

n=10)

9 (90)

7 (70)

4 (40)

6 (60)

9 (90)

10

3 (30)

1 (10)

(

(100)

TET = Tetracycline (10µg), GEN = Gentamicin (10µg), COT = Cotrimoxazole (25µg), NAL = Nalidixic acid (30µg), NIT = Nitro-

furantoin (200µg), OFL = Ofloxacin (5µG), AUG = Augmentin (30 µg), AMX=Ampicillin (10µg).

n = number of isolate

similar to the reports from an earlier study₉ which was

similarly conducted during the dry season. Aeromonas

infection is water-borne, thus, its frequency had been

reported to be higher during the wet season (May

through October).

Discussion

This study showed that E. coli infection was most fre-

quent among children aged eleven to fifteen compared

to other age groups. This observation agreed with re-

ports from Nsukka suggesting that school-age children

were likely to consume contaminated water and food. In

contrast, another report had shown that the highest

prevalence o₁f₆ E. coli infections occurred in the younger

age groups.

Aeromonas was not isolated in the present study despite

the use of standard microbiological procedures. Indeed,

Aeromonas has been suggested to be rarely associated

6

with diarrhoeal diseases in Nigeria. However, this claim

contrasted with other Ni₁g₅erian reports from Nassarawa

9

State and in Edo State. The non-occurrence of Aero-

monas species in the present report may be attributed to

the fact that the study was conducted during the dry sea-

son (December through February). This observation was

1

27

The susceptibility pattern of E. coli isolates showed that,

among the commonly used antibiotics, gentamicin was

the only one that the isolates were still largely sensitive

to, unlike other reports that recorded resistance to gen-

tamicin. In addition, ofloxacin, Augmentin and nalidixic

acid could serve as alternatives to gentamicin in the

treatm₁₆e_,₁n₇t of bacterial diarrhoeal diseases in the study

The development of new antibiotics may offer a

short term solution to the problem of antibiotic resis-

tance but more cost-effective measures like health edu-

cation and other preventive measures should be

encouraged.

Contribution of authors

area.

The high resistance of E. coli species to tetra-

Most research materials were provided by the

Redeemer’s University while the remaining were

sourced by the researchers.

Conflict of interest : None

Funding: None

cycline and ampicillin in the study area calls for a large-

scale detailed research works on antibiotic susceptibility

pattern of stool pathogens in the locality. In addition,

emphasis must be placed on preventive measures such

as improved hygiene rather than increasing reliance on

antibiotic therapy as it obtains in some countries.

Acknowledgement

Conclusion

We thank the pupils, parents and members of staff of

both St. David Nursery and Basic school and Imedu nla

Nursery and Primary school for their consent and co-

operation during this project. We also thank the Re-

deemer’s University especially department of Biological

Sciences for the use of the laboratory.

This study shows that E. coli, rather than Aeromonas,

was the predominant bacterial agent associated with

diarrhoeal diseases among school pupils in Obafemi-

Owode Local Government Area, Ogun State.

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