General Health and Medical Sciences, Vol(1), No (2), December, 2014. pp.

27-29

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General Health and Medical Sciences

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In Vitro Susceptibility of Coliform and Pseudomonas Species to Some

Antibiotics by E-test Strips
Bachir Raho Ghalem*
Department of Biology, University of Mascara, Algeria.

Benattouche Z.
Department of Biology, University of Mascara, Algeria.

Bevilacqua A.
Department of Food Science, Faculty of Agricultural Science, Foggia University, Italy.

Corbo M.R.
Department of Food Science, Faculty of Agricultural Science, Foggia University, Italy.

Sinigaglia M.
Department of Food Science, Faculty of Agricultural Science, Foggia University, Italy.

Pignatiello S.
Department of Food Science, Faculty of Agricultural Science, Foggia University, Italy.
*Corresponding author: bachir_raho@yahoo.fr

Keywords

Abstract

Coliform and Pseudomonas species antibiotics

E-test strips
Antimicrobial susceptibility

The aim of this study was to characterize the in vitro antimicrobial susceptibility profile of Coliform and
Pseudomonas species strains to 10 antibiotics (Vancomycine, Ciprofloxine, Tetracyclin,
Chloramphenicol , Erythromycin, Streptomycine, Gentamycine , Ampicilline, trimethoprimsulfamethoxazole and Clarithromycin) that could potentially be used in clinical therapy.
MIC was determined for antibiotics in Mueller-Hinton (MH) agar plates, by using E-test strips
(Liofilchem, Roseto degli Abruzzi, Italy). MIC was read after 24 h of incubation at 37C.
The both strains were more susceptible to Ciprofloxine (MIC values: Coliform, 0.002 g/mL; and
Pseudomonas, 0.047 g/mL). No strain of Pseudomonas was found sensitive to Chloramphenicol,
Erythromycin, Ampicilline, trimethoprim-sulfamethoxazole and Clarithromycin, contrary to coliform,
which was found resistant to those antibiotics with MIC values ranged between 0.38 and 128 g/mL .
Gentamycine (MIC values: Coliform, 0.125g/mL; and Pseudomonas, 0.50 g/mL), Tetracyclin (MIC
values: Coliform, 1.00 g/mL; and Pseudomonas, 0.75 g/mL) and Streptomycine (MIC values:
Coliform, 0.75 g/mL; and Pseudomonas, 2.5 g/mL) were also active against Coliform and
Pseudomonas strains, while resistance to Vancomycine (no inhibition) was observed in the two strains.

1.

Introduction

Antimicrobial drugs have played an indispensable role in decreasing illness and death associated with infectious diseases in animals and humans.
However, selective pressure exerted by antimicrobial drug use also has been the major driving force behind the emergence and spread of drugresistance traits among pathogenic and commensal bacteria. In addition, resistance has developed after advent of every major class of
antimicrobial drugs, varying in time from as short as 1 year (penicillin) to >10 years (vancomycin) [1].
Pseudomonas and coliforms species are important cause of nosocomial infections and are responsible for many hospital-acquired infections
[2,3]. The coliform bacteria include Shigella, which causes severe gastroenteritis or bacterial dysentery; certain strains of Escherichia coli
(referred to as E. coli), which can cause urinary tract infections and diarrhea; and Salmonella, which causes typhoid fever and gastroenteritis
(food poisoning) [4].
As opportunistic pathogens, Pseudomonas spp. often invades the host tissue and cause infection and bacteremia in immunocompromised hosts
(e.g., HIV/AIDS, cystic fibrosis, bronchiectasis, and severe chronic obstructive pulmonary disease, burns, malignancy, or diabetes mellitus)
[5,6]. Infections also include endocarditis, osteomyelitis, urinary tract infections, gastrointestinal infections, meningitis, and, commonly,
septicaemia [3,7,8].
Resistant Pseudomonas and coliform species is a public health problem that affects many countries of the world. Infections caused by these
pathogens are often severe and life threatening and are difficult to treat because the high frequency of an emergence of antibiotic resistance
during therapy.
The aim of this study was to characterize the in vitro antimicrobial susceptibility profile of coliform and Pseudomonas species to 10 antibiotics
(Ampicillin, Chloramphenicol, Clarithromycin, Erythromycin, Tetracycline, Vancomycin, Streptomycin, Gentamycin, trimethoprimsulfamethoxazole and Ciprofloxin) that could potentially be used in clinical therapy.

2.

Material and Methods

2.1 Antimicrobial agents.

The E test strips were purchased from Liofilchem (Roseto degli Abruzzi, Italy). Antimicrobial agents tested against Coliform and Pseudomonas
species included Ampicillin, Chloramphenicol, Clarithromycin, Erythromycin, Tetracycline, Vancomycin, Streptomycin, Gentamycin,

Bachir Raho Ghalem *, Benattouche Zouaoui , Bevilacqua A., Corbo M. R., Sinigaglia M., Pignatiello S.

28

General Health and Medical Sciences Vol(1), No (2), December, 2014.

trimethoprim-sulfamethoxazole and Ciprofloxin. The concentration of the first six antimicrobial agents ranged from 0.016 to 256.0 g/ml, that of
Streptomycin and Gentamycin was from 0.064 to 1024 g /ml, but the strips of trimethoprim-sulfamethoxazole and Ciprofloxin contained
concentration gradients of 0.00232 g / ml of the respective drug.
2.2 Bacterial strains
The microorganisms used for this experiment were coliform spp and Pseudomonas spp belonging to the Culture Collection of the Laboratory of
Applied Microbiology (University of Foggia, Italy).
2.3 Antibacterial susceptibility testing
This assay was performed through a modified version of the E-test, using the antibiotic strips produced by Liofilchem (Roseto degli Abruzzi,
Italy) [9]. Coliform and Pseudomonas species were streaked onto the surface of Mueller-Hinton agar through a sterile swab; thereafter, the strip
containing the antibiotic was placed onto the surface of the plates.
The plates were incubated at 37C for 24 h; a clear halo was the sign of the susceptibility of the microbial target to the tested antibiotic. The MIC
of each antibiotic was read at the point of significant inhibition in fungal growth.

3.

Results and discussion

As demonstrated in Table 1, in vitro susceptibility tests showed that the Pseudomonas spp exhibited high-level resistance to Ampicillin,
Chloramphenicol, Clarithromycin, Erythromycin, Vancomycin and Trimethoprim-sulfamethoxazole. Most notably, it exhibited more
susceptibility to Ciprofloxin (0.047 g/ml), Gentamycin (0.50 g/ml), Tetracycline (0.75 g/ml), and Streptomycin (2.5 g/ml).
Coliform spp was also resistant to Vancomycin, and Erythromycin, but susceptible to others antibiotics with different concentrations. The
MIC values ranged from 0.002 to 3.00 g/ml for the remaining antibiotics. The more active is Ciprofloxin with 0.002 g/ml, but the less is
Trimethoprim-sulfamethoxazole with 3.00 g/ml.
Table 1. Susceptibilities of Coliform and Pseudomonas species to various antimicrobial agents
Antimicrobial agent

MIC, g/ml
Coliform spp

Pseudomonas spp

Ampicillin
Chloramphenicol
Clarithromycin
Erythromycin

0.38
2.5
1.5
128

NI
NI
NI
NI

Tetracycline
Vancomycin
Streptomycin
Gentamycin
TSM
Ciprofloxacin

1.00
NI
0.75
0.125
3.00
0.002

0.75
NI
2.5
0.50
NI
0.047

NI: no inhibition; TSM: Trimethoprim-sulfamethoxazole

The E test is technically simpler, can be set up more rapidly than traditional agar or broth dilution test methods, and does not require special
equipment. Strips are available for a wide array of antimicrobial agents and can be used to test a variety of fastidious and nonfastidious
bacteria[10].
Pseudomonas spp. were observed to be resistant to the majority of the antibiotic tested. Our results confirm the results of previous reports, which
showed a resistance of some Pseudomonas species to these antibiotics [11, 12, 13, 14]. Also a high incidence of antibiotic resistance in
Pseudomonas spp. has been reported by Ibiebele and Sokari (1989) for drinking-well waters, and by Jones (1986) and Jones et al. (1986a) for
lake waters.
High resistance rate may be either due to the potential use of these antimicrobials in open market and private pharmacies without
prescription[18]. High frequencies of antibiotic resistance have been reported for bacteria isolated from areas where antibiotics have been used
extensively by humans, and in terrestrial and aquatic environments subject to intensive animal production. As antibiotic resistance can be spread
from resistant to formerly sensitive bacteria via the transfer of R-plasmids, maintenance of antibiotic resistance in non-pathogenic native bacteria
could provide a reservoir for antibiotic-resistance genes[19].
Coliform spp was more susceptible than Pseudomonas spp to all antimicrobial agents tested in this experiment. All MIC of antibiotics for
Coliform spp were lower than those for Pseudomonas spp, which is in agreement with the study of Sokari et al. (1988) that found a lower
incidence of resistance among E. coli than among Pseudomonas spp. in his working on diverse Nigerian waters. Our results confirm the results
of previous reports, which showed the susceptibility of coliform from different origins to some antibiotics[21, 22, 23, 24, 25, 26].
Here, also we found that Coliform spp was resistant to Vancomycin, and Erythromycin, as has been shown by Kuljinder and Kahlon (2014) who
have reported that the coliforms isolated from food in Rajasthan market, in India were most resistant to all selected antibiotics, including
Vancomycin and Erythromycin.

4.

Conclusion

The results of this study indicate that Ciprofloxin is the most active drug, in vitro, against the two bacteria, followed by Gentamycin and
Tetracycline, as shown by E-test method. Vancomycin has no activity against Coliform and Pseudomonas species. The E-test may be an
effective method for testing susceptibility of organisms to antibiotics.

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In Vitro Susceptibility of Coliform and Pseudomonas Species to Some Antibiotics by E-test Strips
General Health and Medical Sciences Vol(1), No (2), December, 2014.

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