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Antimicrobial Susceptibility Patterns of Pseudomonas aeruginosa from Diabetes Patients with Foot Ulcers

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International Journal of Microbiology
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Pseudomonas aeruginosa is an invasive organism that frequently causes severe tissue damage in diabetic foot ulcers. A major problem in P. aeruginosa infection may be that this pathogen exhibits a high degree of resistance to a broad spectrum of antibiotics. The study aimed to isolate and determine the antimicrobial susceptibility patterns of the P. aeruginosa population from diabetes patients with foot ulcers attending tertiary care hospitals in and around Coimbatore and their antimicrobial susceptibility pattern. The study was carried out at the Department of Microbiology, Dr. N.G.P. Arts and Science College, Coimbatore, for a period of one year (June 2006 to April 2007). The present study comprised 270 pus specimens collected from diabetic patients with foot ulcers. All pus samples were subjected to gram staining; bacterial culture and subsequently the antibiotic sensitivity to 15 different antibiotics for the confirmed P. aeruginosa were performed as per the standard procedures. Eighteen strains (14.28%) of P. aeruginosa from 270 diabetic foot ulcers were detected. Almost all the strains exhibited a varying degree of resistance to the antibiotics tested. Multidrug resistance for about 8 to 11 antibiotics was observed among the 55.5% of the isolates. Disk diffusion results show 100% resistance to ampicillin, cefoperazone, erythromycin, norfloxacin, and only cefotaxime, ciprofloxacin exhibited greater activity against Pseudomonas aeruginosa.
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Hindawi Publishing Corporation
International Journal of Microbiology
Volume 2011, Article ID 605195, 4pages
doi:10.1155/2011/605195
Research Article
Antimicrobial Susceptibility Patterns of
Pseudomonas aeruginosa
from Diabetes Patients with Foot Ulcers
Tamil Selvi Sivanmaliappan1and Murugan Sevanan2
1Department of Microbiology, Dr. N.G.P. Arts and Science College, Coimbatore, Tamil Nadu 641048, India
2Department of Biotechnology, School of Biotechnology and Health Sciences, Karunya University, Karunya Nagar,
Coimbatore, Tamil Nadu 641 114, India
Correspondence should be addressed to Murugan Sevanan, micromurugans@gmail.com
Received 13 July 2011; Accepted 23 August 2011
Academic Editor: Toni L. Poole
Copyright © 2011 T. S. Sivanmaliappan and M. Sevanan. This is an open access article distributed under the Creative Commons
Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is
properly cited.
Pseudomonas aeruginosa is an invasive organism that frequently causes severe tissue damage in diabetic foot ulcers. A major
problem in P. a er u g i n o s a infection may be that this pathogen exhibits a high degree of resistance to a broad spectrum of antibiotics.
The study aimed to isolate and determine the antimicrobial susceptibility patterns of the P. a e r u g in o s a population from diabetes
patients with foot ulcers attending tertiary care hospitals in and around Coimbatore and their antimicrobial susceptibility pattern.
The study was carried out at the Department of Microbiology, Dr. N.G.P. Arts and Science College, Coimbatore, for a period
of one year (June 2006 to April 2007). The present study comprised 270 pus specimens collected from diabetic patients with
foot ulcers. All pus samples were subjected to gram staining; bacterial culture and subsequently the antibiotic sensitivity to 15
dierent antibiotics for the confirmed P. a e r u g i n o sa were performed as per the standard procedures. Eighteen strains (14.28%)
of P. a e r u g i no s a from 270 diabetic foot ulcers were detected. Almost all the strains exhibited a varying degree of resistance to the
antibiotics tested. Multidrug resistance for about 8 to 11 antibiotics was observed among the 55.5% of the isolates. Disk diusion
results show 100% resistance to ampicillin, cefoperazone, erythromycin, norfloxacin, and only cefotaxime, ciprofloxacin exhibited
greater activity against Pseudomonas aeruginosa.
1. Introduction
Diabetes is a chronic disorder that aects a large segment of
population and is a major public health problem. Diabetes
and foot problems are almost synonymous [1]. A recent
WHO report indicates that India has the largest diabetic
population (19 million in 1995) that is expected to rise to 57
million by 2025 [2]. A commonly accepted definition of foot
infection is the presence of systemic signs of infection (e.g.,
fever and leucocytosis) or purulent secretions or two or more
local symptoms or signs (redness, warmth, indurations, pain,
or tenderness) [3]. Viswanathan et al. [4] reported that 25%
of diabetic individuals are anticipated to develop severe foot
problems at some point in their lifetime that often end
with amputation. Diabetic foot infections are more severe
and more dicult to treat than infections in nondiabetics.
Polymicrobial etiology has been implicated in the infected
diabetic foot. Gram-negative infections are three-times more
frequent in the diabetic than in non-diabetic individuals [5].
Most infections with Pseudomonas species occur in
compromised hosts. The pathogenicity of these organisms
is based on its ability to produce a variety of toxins and
proteases and also on its ability to resist phagocytosis [6].
Pseudomonas aeruginosa is commonly resistant to antibiotics,
and because of this it is a dangerous and dreaded pathogen.
The only antibiotic agents to which strains are regularly
sensitive are cephalosporins, carbenicillin, colistin, gentam-
icin, polymyxin, quinolones, and streptomycin; however
degrees of cross-resistance between these agents have been
reported [7]. P. a e r ug i n o s a is one of the most important
microorganisms that cause clinical problems resulting from
high-resistance to antimicrobial agents. Though it is rarely
2International Journal of Microbiology
found in the normal flora of humans, it is frequently isolated
from patients with burns, cystic fibrosis, and neutropenia
[8]. P. a e r u g in o s a may cause severe tissue damage in diabetics
and should never be ignored as insignificant in diabetic
foot ulcers. Moreover, it should never be considered a
contaminants or normal flora, and it should clearly be
considered a pathogen, because it may result in sepsis and
amputation [9].
One of the challenges in managing P. a e r u g in o s a infec-
tions is an inherent resistance mechanism, referred to as
intrinsic resistance. Its multiplicity of resistance mecha-
nisms may render this microbe less amenable to control
by antibiotic cycling [10]. P.aeruginosa is noted for its
metabolic versatility and its exceptional ability to colonize
a wide variety of environments and also for its intrinsic
resistance to a wide variety of antimicrobial agents. The
bacillus almost never causes infections in healthy individuals
and often infects the immunocompromised. Because of its
virulence and the limited choices of eective antimicrobial
agents, treatments of infections by P. a e r u g i n os a are often
dicult [11]. Though a lot of work has been carried out
elsewhere pertaining to P. a e r ug i n o s a , the prevalence and
the antimicrobial susceptibility patterns of P. a e r u g i n os a
from diabetes patients with foot ulcers have rarely been
documented in this part of South India. Therefore, the
present study has been carried out to study the prevalence
of P. ae r u g i n os a and their antimicrobial susceptibility by the
Kirby Bauer-Disk Diusion method.
2. Materials and Methods
2.1. Sample. The study was based on 270 pus specimens
received for the screening of P. ae r u g i n o sa from diabetes
patients with foot ulcers attending tertiary care hospitals
in and around Coimbatore. Specimens included in the
study were from “Soft tissue infection” which includes foot
wound and “limb threatening infections” specimens were
included in the study. Specimens were obtained using aseptic
techniques to avoid contamination and were promptly
transported to the laboratory in a sterile swab in ice-cold
conditions.
2.2. Isolation and Identification of Pseudomonas aeruginosa.
The isolation and identification of test organisms was carried
out by the procedures suggested by Valentina and Lalitha
[12]. Identification analysis like Gram staining, motility,
catalase, oxidase, pigment production, growth on cetrimide
agar, ability to grow at 42C, gelatin hydrolysis, arginine
dihydrolase, acid from Hugh-Leifson’s glucose, and nitrate
reduction tests were carried out.
2.3. Antibiotic Sensitivity Testing (Kirby Bauer-Disk Dif-
fusion Method). Antibiogram was performed using com-
mercially available antibiotic discs (Hi-Media, Mumbai)
with a standard P. a e r u g in o s a ATCC 27853 as a positive
control. Kirby-Bauer, recommended by the CLSI [13], was
used for antimicrobial susceptibility testing. The identi-
fied 18 P. a e r u g i n os a strains were tested against ampi-
cillin (10 µg), amikacin (30 µg), ceftazidime (30 µg), cefo-
taxime (30 µg), ciprofloxacin (5 µg), cefoperozone (75 µg),
co-trimoxazole (25 µg), erythromycin (10 µg), gentamicin
(10 µg), imipenem (10 µg), norfloxacin (10 µg), piperacillin
(100 µg), tobramycin (30 µg), ticarcillin (75 µg), and tetracy-
cline (30 µg).
3. Results and Discussion
3.1. Patients. Of the total 270 diabetic patients suering from
foot infections, 180 were male and 90 were female. The
male-to-female ratio was 2 : 1 and the age of the patients
ranged between 36 to 75 years. Very few of these patients
have undergone amputation. Studies conducted in Chennai
have shown that males were more susceptible than females
in the ratio of 8 : 3 [14]. Previous studies have shown that
males were more susceptible than females in the ratio of 2 : 1,
which is in accordance with the current study. Predominance
of male over female patients as shown in the study can be
explained by the fact that in our country males are exposed
more to the outside environment because of their mobility as
compared to females.
3.2. Isolation Rate. Of the 270 pus specimens of diabetic
patients with foot infections, 180 (66.6%) specimens were
culture positive and the other 90 (33.3%) were negative.
Among the strains, aerobic gram-negative Pseudomonas
species were 126 (70.0%) and other aerobic organisms
comprised 54 (30.0%). From the 126 Pseudomonas species,
18 (14.30%) were found to be P. ae r u g i n o sa . Dhanasekaran
et al.[5] reported the prevalence of Pseudomonas species to
be 18.79% from a diabetic centre in Chennai. In a similar
study conducted in a private hospital in Chennai, 29.8%
strains among diabetic foot ulcer patients were P. a e r u g i no s a
[15]. This finding shows the high prevalence of Pseudomonas
species and P. ae r u g i n o sa among diabetes patients with foot
ulcers.
3.3. Antibiogram Pattern of Pseudomonas aeruginosa. The
Mueller Hinton agar-based antibiogram-resistogram pattern
study of P. a e r ug i n o s a isolated from foot ulcers of diabetes
patients is shown in Figure 1. Almost all of the eighteen
P. a e ru g i n o s a strains screened showed 100% resistance to
ampicillin, erythromycin, and norfloxacin, similarly 83.3%
resistance to piperacillin, ticarcillin, and tetracycline, 66.6%
resistance to ceftazidime, imipenem, gentamicin, amikacin,
tobramycin, and cotrimoxazole and 50.0% resistance to cef-
operazone. However, 15 (83.3%) P. a e ru g i n o s a strains were
susceptible to cefotaxime. Multidrug resistance for about 8
to 11 antibiotics was observed among 55.5% of the strains
(Tab l e 1 ). No single antibiotic showed 100% sensitivity to all
P. a e r u g i n o sa strains. Resistance was least with cefotaxime
(16.6%), followed by an intermediate resistance of 66.7%
observed for ciprofloxacin.
India has the largest number of diabetic individuals
and appreciably poor economic conditions; the study on
International Journal of Microbiology 3
120
100
80
60
40
20
0
Resistance (%)
Antimicrobial agents
Resistant Sensitivity
Intermediate
AK
A
CO
Cef
CTX
Cf
Nx
Te t
Ti
Tb
P
G
I
E
CAZ
Figure 1: Antimicrobial susceptibility patterns of Pseudomonas
aeruginosa among diabetes patients with foot ulcers.
Tab l e 1: Multidrug resistance patterns of P. a e r u g in o s a of diabetic
foot ulcers.
No. of drugs resistant No. of isolates
(n=18) Resistance (%)
8 18 100
9 15 83.3
10 13 72.2
11 10 55.5
12 7 38.8
13 5 27.7
14 2 11.1
15 0 0
this intrinsic resistant organism in diabetic foot infections
assumes significance. The present study has shown the
incidence of P. a e r u g i n os a to be 14.3% in diabetic foot
ulcers, which is significant when compared to previous
studies. In accordance with earlier observations [16], the
current study has demonstrated that P. ae r u g i n o s a strains
isolated from foot ulcers are more resistant to antimicrobial
agents. This may be due to the fact that the strains
isolated from clinical specimens have been subjected to
the selective actions of both disinfectants and antibiotics
[16]. As expected, the strains were resistant to imipenem,
piperacillin, erythromycin, ticarcillin, tetracycline, gentam-
icin, co-trimoxazole, and amikacin indicating the emergence
of multidrug-resistant strains. Antibiogram also revealed
that cefotaxime and ciprofloxacin retained high levels of
antipseudomonal activity and cefoperazone, gentamicin,
ceftazidime, amikacin, imipenem, and tobramycin had the
least activity. Ciprofloxacin and cefotaxime were found to be
better choices for diabetes patients with foot ulcers in this
part of the region when compared to gentamicin, imipenem,
piperacillin, and other third-generation cephalosporins. The
present study thus revealed the importance of P. ae r u g i n os a
from diabetes patients with foot ulcers, which is necessary for
proper management of diabetes patients.
Acknowledgments
The second author is grateful to the Chancellor (Dr. Paul
Dhinakaran), Vice Chancellor (Dr. Paul P. Appasamy), and
Registrar (Dr. Anne Mary Fernandez) of Karunya University,
Coimbatore, India, for their constant encouragement and
support to carry out this research publication.
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