Int J Biol Med Res.

2021 ;12(4):7386-7392
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Original article
A STUDY ON RESISTANCE PATTERN OF BACTERIA ISOLATED FROM DIABETIC AND
NON- DIABETIC ULCERS
a
Joana Mary Magdaline, bAkhila Aravind
a
Department of Microbiology, Government Medical College-Ernakulam, Kalamassery, Cochin Kerala, India- 683503
b
City hospital lane, Kottayam, Kerala India 686001

ARTICLE INFO ABSTRACT

Keywords: Background & Objectives: Epidemiological data from various studies shows that the prevalence
Chronic ulcer
of chronic ulcer is high in Indian population which has a significant impact on health care.
bacterial flora
Multidrug resistance
Bacteriological profile & their antibiotic resistance pattern shows a region wise variation in
different types of ulcers. The present study was conducted to compare the prevalence of
diabetic and non- diabetic ulcers in patients admitted in surgical wards as well as to determine
the resistance pattern of bacteria isolated from the ulcers. Methods: This descriptive study,
which was conducted in the Departments of Microbiology & Surgery, included 103 patients
admitted with ulcers in surgical wards. Samples from the ulcers were collected using sterile
swabs. The isolates were identified using standard microbiological methods &
sensitivity/resistance pattern determined by Kirby- Bauer disc diffusion method. Results &
Discussion: In the present study 142 bacterial isolates were obtained from 103 patients. Most
prevalent type of ulcers among the study population was diabetic ulcers (54%). On comparison
of flora of the three major types of ulcers (diabetic, venous &decubitus) in our study, it was
observed that all of them had a predominant monomicrobial infection with equal proportions
of Gram positive & Gram-negative isolates. This was in discordance with other studies who
reported a Gram-negative predominance. An overall Gram-negative predominance was
observed but the most frequent isolate was Staphylococcus aureus. There is a higher prevalence
(79.5%) of multi drug resistant (MDR) isolates in our study when compared to other studies. All
Gram-positive isolates were resistant to Penicillin & Erythromycin. Gram negative bacteria
showed 100% resistance to Ampicillin. Conclusion: An increased prevalence of MDR bacteria in
ulcers was observed in this study. Hence to promote healing, judicious use of antibiotics should
be followed in patients with chronic ulcers.

c
Copyright 2010 BioMedSciDirect Publications IJBMR - ISSN: 0976:6685. All rights reserved.

Introduction
Ulcers are defined as wounds with a “full thickness depth” and
a “slow healing tendency” which mainly occurs by endogenous
mechanisms associated with predisposing conditions like
Diabetes, venous insufficiency. In ulcers, there is complete loss of
the epidermis, portions of the dermis and even subcutaneous fat.1
Chronic ulcers are defined as wounds that have failed to
proceed through the orderly process that produces satisfactory
anatomic and functional integrity or that have proceeded through
the repair process without producing an adequate anatomic and
functional result2-6. Major types of chronic ulcers include7.
Diabetic foot ulcers, Venous ulcers & Pressure sores / Decubitus
ulcers.
Healing of an ulcer is a dynamic process & is mediated by
interactive reactions of parenchymal cells, soluble mediators,
blood elements, and extracellular matrix. This normal course is
affected by various systemic as well as local factors which delays
the healing and determines the chronicity of an ulcer.
The impact of multidrug resistant (MDR) infections in ulcers
can be understood from the fact that the rates of amputations in
patients who had MDR infection was 35.6% compared to 11.2% in
non-MDR infection.8 Studies had reported that the incidence of
MDR bacterial infections are more in chronic wounds compared to
acute ones.9

* Corresponding Author : Dr Akhila Aravind

Email id: akhila4u@gmail.com
c Copyright 2011. CurrentSciDirect Publications. IJBMR - All rights reserved.
 Joana Mary Magdaline and Akhila Aravind / Int J Biol Med Res.12(4):7386-7392
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OBJECTIVES technique is relatively inexpensive, non-invasive & a simple

I. To compare the prevalence of diabetic and non- diabetic ulcers in
patients admitted in surgical wards.
II. To determine the resistance pattern of bacteria isolated from the
ulcers.
RELEVANCE OF STUDY
Epidemiological data available from the studies shows that the
prevalence of chronic ulcer in Indian population is much higher than
that seen in the Western population. In our institution also chronic
ulcers is a significant burden. 44% (377/849) of the total samples
from the surgery department is from ulcers. Bacterial infection is the
most common cause of delayed healing. The impact of multidrug
resistant (MDR) infections in ulcers can be understood from the fact
that the rates of amputations in patients who had MDR infection was
35.6% compared to 11.2% in non-MDR infection.8 Studies had
reported that the incidence of MDR bacterial infections are more in
chronic wounds compared to acute ones.9 Available studies showed
that bacterial flora and resistance pattern of isolates varied widely in
different types of ulcers. Infection of chronic ulcers with multidrug
resistant bacteria will make them recalcitrant to healing. There is a
paucity of studies regarding the bacterial flora and their resistance
pattern in non-diabetic ulcers. These observations warrant the need
for region wise studies, which may help in formulating the policy for
empirical therapy.
MATERIALS AND METHODS
procedure. Since various studies have shown comparable
results from properly collected swab samples and tissue
specimens hence, we adopted the same. 1,10.
Surrounding skin of the ulcer was disinfected with alcohol
swabs. Ulcer area was thoroughly cleaned with sterile saline.
Two sterile swabs were used for each patient.2,11 Samples were
collected by making firm rotatory movements, covering entire
area of the ulcer, holding both the swabs together. The swabs
were then transported to the laboratory in sterile test tubes and
processed within 2 hours.3-6 Swabs with the specimens were
inoculated on to culture media. Identification of the isolates up
to species level was done as per standard microbiological
methods.
Antibiotic sensitivity testing was done for the above isolates
using Kirby – Bauer Disc Diffusion technique according to
standard microbiological methods.1The antibiotic disc strength
and panel of antibiotics tested for Gram negative & Gram-
positive bacteria were according to Clinical and Laboratory
Standards Institute (CLSI) guidelines. 12-15 The list of
antibiotics used, their strength and zone size for interpretation
for Gram negative and Gram-positive isolates are given in the
tables given below.

❖ Study Design: Descriptive Study (Prevalence study

❖ Study Setting: Department of Microbiology & Department of

Surgery.

❖ Study Period: 1 Year from January 2015/all samples collected

during this period.

❖ Study population: All patients admitted with ulcers in the

surgical wards were included in the study.

● Inclusion Criteria: A patient with ulcer was included only once

during the period of study

● Exclusion criteria: Patients with malignant ulcers and ulcers

associated with burns and post-operative wound infections were not
included in the study.

❖ Sample size & Sampling methodology: Using the formula

4pq/d2, the sample size was calculated to be 99, the sampling
methodology was Universal sampling

❖ Ethical Clearance: Obtained from the Institutional Ethical

Committee before commencement of the study.

❖ Procedure In Detail:

The ideal sample for culture from ulcers is tissue biopsy

specimens.10 In this study we had opted for sample collection using
sterile swabs considering the requirement for technical skill and
increased cost for collection of tissue biopsy specimens. Swab
 Joana Mary Magdaline and Akhila Aravind / Int J Biol Med Res.12(4):7386-7392
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History and clinical details obtained from each patient was

numerically coded and entered into Microsoft excel spread
sheet. The bacteriological profile of different ulcers & resistance
pattern of the isolates were also coded & entered into excel
spread sheet and analysed using Statistical package for Social
Sciences (SPSS) software16.0.
RESULTS
A total of 103 samples were collected from patients with
different types of ulcers. Males were predominant in our study
population compared to females. There were 66 males & 37
females in our study.

Figure 1: Pie diagram showing the prevalence of types of

ulcers in the study population
Of the 103 cases 56 patients had diabetic ulcers. Among the
non-diabetic ulcers which were 47 in number, venous ulcers and
decubitus ulcers predominated
66 patients had monomicrobial infection; among mono-
microbial infection 30 were due to Gram positive bacteria and
36 were due to Gram negative bacteria. Polymicrobial infection
was observed in 36 patients; 32 cases yielded two isolates each
and 4 cases had three isolates.

Table 3: Frequency Table of all the Isolated Organisms

An isolate was considered as Multidrug resistant (MDR) if it

has shown non-susceptibility to at least one agent in three or
more antimicrobial categories. Extensive drug resistance (XDR)
was defined as non-susceptibility to at least one agent in all but
two or fewer antimicrobial categories (i.e., bacterial isolates
remain susceptible to only one or two categories) and Pan drug
resistance (PDR) was defined as non-susceptibility to all agents
in all antimicrobial categories.16
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Table 4: Table showing the frequency of various isolates

obtained from major types of ulcers in the study

In the present study 142 isolates were obtained from 103

patients. Pseudomonas aeruginosa were the predominant isolates
from all types of ulcers. Gram positive isolates other than
Staphylococcus aureus were Enterococcus spp, Coagulase negative
Staphylococci, Corynebacterium spp and Streptococcus spp. Apart
from Pseudomonas aeruginosa the gram-negative bacteria isolated
were Escherichia coli, Proteus spp, Klebsiella pneumoniae,
Acinetobacter spp, Serratia spp, Morganella Morgagni,
Flavobacterium spp Enterobacter spp and Coliform.

Table 5: Resistance pattern of Gram-positive bacteria

isolated in the study

Figure 2: Resistance pattern of predominant isolates from

Enterobacteriaceae group
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· Almost all the Gram-negative isolates were resistant to
Ampicillin in which it is tested.
· Most of the Gram-negative isolates except for Acinetobacter spp
were sensitive to Imipenam.
· Majority of the Acinetobacter spp isolates were resistant to most
of the antibiotics tested (40% were PDR strains, 40% were XDR & the
remaining were MDR strains.
· Majority of the Enterobacteriaceae members showed almost
100% resistance to Augmentin except Proteus spp (44%).
· Meropenam resistance was shown by majority of isolates
belonging to Klebsiella spp, Acinetobacter spp.
· Serratia spp were sensitive to Carbapenams only
· Only one isolate of Flavobacterium spp was obtained which was
sensitive to most of the antibiotics tested.
Table 7: Overall resistance pattern of the tested antibiotics in
Gram positive bacteria & Gram-negative isolates
7391
· Gram positive bacteria showed maximum resistance against
Penicillin & least resistance shown against Linezolid & Vancomycin.
Gram negative bacteria (except Acinetobacter spp &
Pseudomonas aeruginosa were Ampicillin is not tested) showed
maximum resistance to Ampicillin.
· Least resistance among Gram negative isolates was seen against
Imipenem
DISCUSSION
Ulcers especially chronic ulcers pose a significant health care
burden worldwide. It also causes major social as well as economic
problem to the affected individual. The non- healing of such ulcers
could be due to multiple factors among which bacterial infections are
the forerunners especially infection with MDR bacteria.
The study population in the present study included patients from
the age of 25 to those up to the age of 87. Maximum number of
patients were from the age group 51- 60. This data was in accordance
with the observation from other studies.17-23
We had a male predominance among our study population. This
was in accordance with the studies on ulcers from India. 18-23 but
studies from outside India especially from the West17,25 had
reported that women were twice likely to get affected with ulcers
than men
We had identified diabetic foot ulcers as the most prevalent type
among the study population followed by venous ulcers & decubitus
ulcers. This was in discordance with the studies conducted in the
West where venous ulcers tend to be more prevalent than the other
major types of ulcers 3,25-26.
The higher prevalence of diabetic ulcers in our study could be
because of the increased prevalence of poorly controlled diabetes in
our population.
Drug resistance in bacteria is an important cause responsible for
non- healing of ulcers even with adequate & appropriate antibiotic
therapy. In view of this problem, we have assessed the resistance
pattern of the predominant isolates from our study.
In our study an increased prevalence (79.5%) of multidrug
resistance was seen among the isolates. Published data from most of
the other studies had shown less than 60% prevalence of MDR
isolates in the study population. Gram negative bacilli (GNB) were
the predominant (56%) among MDR isolates in our study, which
holds true for other studies also 17,19,20,27,28,.
In the present study among the Gram negative MDR bacteria,
Pseudomonas aeruginosa was most frequent followed by Klebsiella
& Escherichia coli. This was in accordance with majority of other
studies, but some studies had identified Enterobacteriaceae group &
Acinetobacter spp as the predominant MDR bacteria among the
Gram negatives. Pseudomonas isolates showed maximum resistance
to Gentamicin &least resistance to Carbapenams in our study which
is in comparison with that reported by Umadevi et al at
Puducherry23. In the study by Swarna et al Pseudomonas showed
maximum resistance to amikacin & fluoroquinolones28.
Among the Gram-positive isolates in our study most frequent
MDR bacteria was Staphylococcus aureus. Out of which 54% were
MRSA.Prevalence of MRSA in the present study was less compared to
other studies17,27. Staphylococcus aureus showed maximum
resistance against Penicillin (94%), Ciprofloxacin (84%) &
Erythromycin (60%). All the Staphylococcus aureus isolates were
susceptible to Linezolid & Vancomycin.
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7392
Among the isolates of Enterococci in our study, high level gentamicin
resistance was observed in 50% of isolates which was more than that
detected in study by Umadevi et al at Puducherry23.
The higher prevalence of multidrug resistance among the
isolates in our study could be due to the fact that ours is a tertiary
care hospital were most of the patients have got treatment already
from elsewhere, which might have been inadequate. This would have
led to a selective survival advantage of the resistant infecting
bacteria & this could explain the increased prevalence of MDR
isolates in our study, as inadvertent use of antibiotics is one of the
major cause implicated in the development of drug resistance.
CONCLUSION
An increased prevalence of diabetic ulcers was observed in the
study which contrasted with the Western studies. This could be
because of the increased number of patients with poorly controlled
diabetes in the population.
The present study & available studies showed that bacterial flora
& resistance pattern of isolates varied widely in different types of
ulcers.
In our situation the resistance pattern (all Gram negatives were
resistant to Ampicillin & all Gram positives were resistant to
Penicillin & mostly to Erythromycin) exhibited by the predominant
isolates indicates that Ampicillin should not be used for treating
Gram negative infections & Penicillin & Erythromycin not to be used
for Gram positive infections in ulcers.
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c Copyright 2019 BioMedSciDirect Publications IJBMR - ISSN: 0976:6685.
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