How to cite this article: Hameed-Ud-Din, Muhammad Saaiq, Muhammad

Ibrahim Khan. Microbial profile and antibiotic susceptibility of the bacterial

contaminants of infected wounds: Experience at a Plastic and Reconstructive
surgical unit. Ann Pak Inst Med Sci 2006; 2(2): 93-8.

TITLE:

MICROBIAL PROFILE AND ANTIBIOTIC SUSCEPTIBILITY OF THE

BACTERIAL CONTAMINANTS OF INFECTED WOUNDS: EXPERIENCE AT
A PLASTIC AND RECONSTRUCTIVE SURGICAL UNIT.

AUTHORS:

HAMEED-UD-DIN,
MUHAMMAD SAAIQ,
MUHAMMAD IBRAHIM KHAN.

DEPARTMENT OF PLASTIC AND RECONSTRUCTIVE SURGERY,

PAKISTAN INSTITUTE OF MEDICAL SCIENCES (PIMS),
ISLAMABAD.

ABSTRACT

Objective : To study the microbial profile of the infected wounds and to detect the

antibiotic susceptibility pattern of the microbes in our setup.

Study design : Single centre non-interventional descriptive study.

Place and duration of the study: Department of plastic and Reconstructive Surgery,

Pakistan Institute of Medical Sciences (PIMS), Islamabad from Jan 2004 to Dec. 2005.

Subjects and Methods: A total of 193 adult patients of either gender having clinical

features of infected wound were included in the study. Patients with healthy wounds,

those who had been taking antibiotic therapy within the preceding 72 hours and those not

consenting to participate in the study were excluded. Specimens for culture and

1
sensitivity were collected by employing standard collection techniques and analyzed at a

single microbiological laboratory. All the samples were incubated for 24 hours at 37 C

for obtaining aerobic and anaerobic growths. Microbes were identified by their colonial

morphology and characteristic biochemical tests.

Results : There was male predominance with a male : female of 2.11 : 1 . The mean

age was 33 + 16.21 years with a range of 15-68 years. Bacterial growths were obtained

in 127 (65.8 % ) specimens while the remaining 66 (34.2 %) specimens yielded no

growth. 119 (93.7 % ) isolates were monobacterial whereas 8 ( 6.29 %) were

polybacterial . Pseudomonas aeruginosa 49 ( 36.02 % ) was the most frequent

isolate followed by Escherichia coli 31 (22.79 %), Staphylococcus aureaus 17 ( 12.5 %),

Klebsiella pneumoniae 12 (8.82%) , Methicillin resistant Staphylococcus aureaus

(MRSA) 10 ( 7.35 %), Acinetobacter 8 (5.88%), Proteus and Enterobacter

each 4 (2.94%) and Serratia 1 (0.73%). Sensitivity tests showed variable response to

different antibiotics.

Conclusion: In this era of evidence based medicine and evidence based surgery the

importance of objectivity can’t be overemphasized. For evidence based

antibiotic prescribing practices it is mandatory to have periodic audits of

the infected wounds to know about the invading microbes and their

antibiotic susceptibility. This will also help not only to prevent the emergence of

antibiotic resistance owing to the irrational and indiscriminate use of antibiotics

but also reduce the financial sufferings of our patients.

Key words: Wound infection, Culture and sensitivity, Antibiotic resistance.

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INTRODUCTION :

Surgeon has no greater enemy than wound infection. The history of wound infection is

probably as old as that of surgery itself. The ancient Egyptians were the first civilization

to have a definite protocol for wound management with the application of various
1,2
potions and grease to assist wound healing. For centuries the process of wound

healing remained a mystery as highlighted by the famous saying by French military

surgeon Ambroise Pare’ “I dressed the wound , God healed it. 3 German microbiologist

Koch laid down the first scientific definition of infection through his famous Koch’s

postulates. 4 The discovery of Penicillins by Alexander Fleming (1881-1955) heralded a

new era in wound management that led to the proliferation of many other antibiotics

especially Cephalosporins in 1950s.4,5

Despite of significant advances in antimicrobial and surgical therapy wound

infection continues to pose a major challenge in the context of surgery and trauma.

The wound may become infected either by the patient’s own endogenous flora

present on the skin, mucosa or in hollow viscera or by exogenous microbes.

The chance of developing an established wound infection depends on a variety of

factors most notably the count and virulence of the microbes and the host defense

mechanisms. Generally infection is associated with a bacterial count of over 10,000 per

gram tissue or per square cm in case of burns. 6

In Plastic and Reconstructive surgery , wound infection in an elective procedure

rather happens to be a catastrophe. As the bacterial spectrum of wounds and their

antibiotic susceptibility vary from unit to unit and center to center, the present study

was conducted to establish our own database regarding the microbial profile of

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infected wounds as well as their antibiotic sensitivity profile . Such studies would

ensure more appropriate and rational use of antibiotics thereby not only reducing the

financial sufferings of the ailing humanity but also help to prevent the emerging

antibiotic resistance which poses a major threat to the existing antimicrobial

therapies.

MATERIALS AND METHODS :

This non-interventional descriptive study was undertaken at the department of Plastic

and Reconstructive Surgery , Pakistan Institute of Medical Sciences (PIMS), Islamabad

from Jan 2004 to Dec. 2005. It prospectively included 193 adult patients of either

sex by convenience sampling technique. All patients with local or systemic clinical

features of wound infection were included in the study. Patients with healthy wounds,

those who had been taking antibiotic therapy within the preceding 72 hours and

those not consenting to participate in the study were excluded from the study.

Specimens for culture and sensitivity were collected by employing standard

7
collection techniques and analyzed at a single microbiological laboratory for

culture and sensitivity . After inoculation on appropriate culture media , the specimens

were incubated for 24 hours at 37 C for obtaining aerobic and anaerobic growths.

The microbes were identified by their colonial morphology and characteristic

biochemical tests. Antibiotic susceptibility was tested by employing disc diffusion

method according to NCCLS (National Committee for Clinical Laboratory Standards )

guidelines using standard antibiotic discs. 8

Stastical analysis: The data were analyzed through SPSS for Windows
version 10. The nominal

4
variables were reported as frequency and percentages. The numerical data

was reported as mean ± standard deviation.

RESULTS :

Their was male predominance with a male : female of 2.11 : 1. The mean age was

33 ± 16.21 years with a range of 15-68 years. Fig. I shows the various types of

wounds from which the bacterial growths were obtained. Bacterial growths were

obtained in 127 (65.8 % ) specimens while the remaining 66(34.2 %) specimens

yielded no growth. 119 (93.7 % ) isolates were monobacterial whereas 8 ( 6.29 %) were

polybacterial. Pseudomonas aeruginosa 49 ( 36.02 % ) was the most frequent isolate

followed by Escherichia coli 31 (22.79 %), Staphylococcus aureaus 17 ( 12.5 %),

Klebsiella pneumoniae 12 (8.82%) , Methicillin resistant Staphylococcus aureaus

(MRSA) 10 ( 7.35 %), Acinetobacter 8 (5.88%), Proteus and Enterobacter 4 (2.94%)

each and Serratia 1 (0.73%).

Sensitivity tests showed variable response to different antibiotics. Table I shows the

antibiotic susceptibility profile of the various isolates of Aerobic gram negative

bacilli (AGNB). The antibiotic sensitivity of the Methicillin sensitive staphylococcus

aureaus (MSSA) and Methicillin resistant staphylococcus aureaus is

depicted in table II.

DISCUSSION :

Aerobic gram negative bacilli have always been problematic nosocomial pathogens

encountered by surgeons as well as physicians, often acquiring antibiotic resistance that

limit our therapeutic choices in their management.

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In our study Pseudomonas aeruginosa (36.02%) was the most frequently cultured

microbe followed by Escherichia coli 22.79% and Staphylococcus aureus 19.85% .

9
Khan JS et al reported Staphylococcus aureus (45%) , E.Coli (14%) and

Pseudomonas aeruginosa (6 %) as the three most frequent isolates from skin and

soft tissue infections. In an other study on the microbial profile of patients in

10
intensive care unit Mahmood A reported Staphylococcus aureus (39.53 % ),

Pseudomonas aeruginosa (15.11 %) and Escherichia Coli (10.4 % ) as the commonest

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bacterial isolates. Batool T et al recorded Pseudomonas aeruginosa, Staphylococcus

aureus and Klebsiella pneumoniae as the most common microbial isolates

from burn wounds. Unlike the other studies , Pseudomonas aeruginosa and

Escherichia Coli were the two most frequent microbial isolates in our study.

This reflects our local microbial flora and negates the common perception that

gram positive cocci are the dominant pathogens causing wound infection. With

this evidence based new scenario , if our initial empiric antibiotic cover is still merely

directed against gram positive organisms it would simply be irrational and

counterproductive leading to wasteful use of our limited resources.

In our study MRSA isolates were 37 % of the Staphylococcal contaminants . Other

12 13
local studies have reported variable incidence ranging from 5 % to 38.5%

Internationally the reported incidence of MRSA among Staphylococcal isolates

14,15
range from less than 10 % to over 65 % . The emergence of MRSA was

first reported in Europe in the early 1960s and even today it continues to be a

major threat to antimicrobial therapy as Methicillin resistance means resistance to

all beta lactam antibiotics .16 ,17

It has rather proven to be one of the most

6
widespread and durable nosocomial pathogens of the late 20th century. 18

MRSA is spread by direct contact and healthcare workers are the most frequent

culprits responsible for their transmission from one patient to another by their transiently

colonized hands. This cross infection can be effectively prevented by adoption

of standard aseptic measures by all hospital workers .

Emergence of antibiotic resistance is a major concern as it limits our therapeutic choices

for infection control. Pseudomonas aeruginosa is one of the difficult to treat microbes.

Ceftazidime has historically been perceived as an effective antibiotic against it ,

however in our study only 24.48 % of its isolates were sensitive to it. Rather

there was 100 % sensitivity to Cefoperazone + Sulbactum , 95.91 % to Imipenem

and 53 % to Cefoperazone. The antibiotic susceptibility of various other AGNB

showed remarkable variability as shown in table I .

The MRSA isolates in our study were generally multi drug resistant . Except for

Vancomycin to which 100 % sensitivity of MRSA was found, there was variable

antibiotic susceptibility pattern. Sensitivity to Fuscidin and Teicoplanin was 70 % each

while there was only 20 % sensitivity to both imipenem and Erythromycin. Table II

shows the in vitro activity of the various tested antimicrobial agents against MRSA in our
13
study. Bukhari MH et al reported 96 % , 94 % and 86 % susceptibility of MRSA to
19 20
Vancomycin, Teicoplanin and Fuscidic acid respectively. Qureshi AH and Latif S

also reported 100 % sensitivity of MRSA to Vancomycin which is in conformity with

our study. Our study proves that Vancomycin is the only agent that can be confidently

employed on empirical basis to combat life threatening infection caused by multi drug

resistant strains of MRSA, however the worldwide reports of intermediate resistance to

7
Vancomycin warrants regular periodic monitoring of the prevalence and antibiotic

sensitivity of this challenging microbe . 21-23

CONCLUSION :

In this era of evidence based medicine and evidence based surgery the

importance of objectivity can’t be overemphasized. For evidence based

antibiotic prescribing practices it is mandatory to have periodic audits of

the infected wounds to know about the invading microbes and their

antibiotic susceptibility. This will also help to prevent the emergence of

antibiotic resistance owing to the irrational and indiscriminate use of antibiotics

but also reduce the financial sufferings of our patients.

REFERENCES :

1) Breasted D: The Edwin Smith Surgical Papyrus. University of Chicago: University of

Chicago press;1930.

2) Bryan PW: The Papyrus Ebers. London/Washington DC. Government Printing

Office;1883.

3) Cohen IK.A brief history of wound healing.1st ed. Yardley Pa: Oxford Clinical

Communication Inc;1998.

4) Leaper DJ. Wound infection. In : Russel RCG, Williams NS, Bulstrode CJK, editors.

Bailey and Love’s Short practice of surgery.24th ed. London : Arnold ;2004:118-32.

5) Cuschieri A, Steele RJC. Infected patients. In : Cuschieri A, Steele RJC, Moosa

AR,editors. Essential surgical practice Vol. I. 4th ed. London : Butterworth-Heinemann ;

2000: 125-70.

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6) Krizek TJ, Robson MC : Evolution of quantitative bacteriology in wound

management. Am J Surg 1975 ; 130: 579-84.

7) Cheesebrough M .District laboratory practice in tropical countries:

Microbiological test ; 63-124.

8) National Committee for Clinical Laboratory Standards. Methods for dilution

antimicrobial susceptibility tests for bacteria that grow aerobically;

Approved standards . 5th ed. 2000 ; 20.

9) Khan JS, Khan JA, Bhoopal FG, Iqbal M. Surgical audit of skin and soft tissue

infections. J Rawal Med Coll : 2005 : 9:26-29.

10) Mahmood A. Blood stream infections in a medical intensive care unit: spectrum

and antibiotic susceptibility pattern. J Pak Med Assoc. : 2001: 51:213 .

11) Batool T , Akhtart J ,Ahmed S, Mirza F, Jalil Sh , Soomro A. Antibiotic sensitivity

pattern in burn wound cultures. J Surg Pak .2005 : 10(1) :8-11.

12) Ashiq B, Tareen AK. Methicillin resistant staphylococcus aureaus in a teaching

hospital at Karachi, Laboratory studies .J Pak Med Assoc. 1989 ;39:6-9.

13) Bukhari MH, Iqbal A, Khatoon N, Iqbal N, Naeeem S, Qureshi GR, Naveed IA.

A laboratory study of susceptibility of Methicillin resistant staphylococcus aureaus

(MRSA). Pak J Med Sci 2004:20(3):229-33.

14) Boyce JM . Diagnosis and treatment of serious antimicrobial resistant

staphylococcus aureus infection . Infectious disease Clinical Updates, 1998 ; 4 : 4.

15) Struelens MJ, Ronveaus O, Jans B, Mertens R. Methicillin resistant staphylococcus

aureaus epidemiology and control in Belgian hospitals, 1991 to 1995. Infect Control

Hosp Epidemiol 1996,17 ( 8): 503-8.

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16) Hartstein AI, Mulligan MI. In hospital epidemiology and infection control ed.

Mayhall CG Lippincott Williams and Wilkins Philadelphia 1999 : 364-74.

17) Hryniewicz W. Epidemiology of resistance to methicillin and other antibiotics in

isolates of staphylococcus aureaus (MRSA): Infection 1999;27: 13-6.

18) Nimmo GR, Schooneveldt J, O’Kane G, McCall B, Vickery A. Community

acquisition of gentamicin sensitive Methicillin resistant staphylococcus aureaus in

Southeast Queensland , Australia. J Clin Microbiology 2000,38:11:3926-31.

19) Qureshi AH, Rafi s, Qureshi SM, Ali AM. The current susceptibility pattern of

Methycillin resistant staphylococcus aureaus to conventional antistaphylococcal

antimicrobials at Rawalpindi. Pak J Med Sci 2004 ;20(4): 31-64.

20) Latif S, Anwar MS, Chaudhry Na. The susceptibility pattern of nosocomial

Methycillin resistant staphylococcus aureaus isolates to Vancomycin and other anti

staphylococcal antibiotics. Biomedical 2000;16:32-5.

21) Srinivasan A, Dick JD,perl TM. Vancomycin resistance in staphylococci. Clinical

Mircobiology Reviews, 2002;15 (3) :430-38.

22) Geisel R, Schmitz FJ, Thomas L, Berns G, Zetasche O, Vlrich B, Fluit AC,

Labischinsky H,Witte W. Emergence of heterogenous intermediate Vancomycin

resistance in staphylococcus aureaus isolates in the Dusseldorf area. J Antimicrob

Chemother 1999,43:846-48.

23) Tallet SM, Bischoff T, Climo M, Ostrowsky B, Wenzel RP, Edmond MB.

Vancomycin susceptibility of Oxacillin resistance Staphylococcus aureaus isolates

causing nosocomial infections. J Clin Microbiol 2002 ; 40 (6) : 2249-50.

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 TYPES OF WOUNDS

Burns Miscellane
19% ous
2%

Postoperati Truamatic
ve wounds wounds
5% 74%

Fig. I The various types of wounds from which bacterial cultures were obtained.

TABLE I: THE SENSITIVITY PROFILE OF THE CULTURED AEROBIC GRAM NEGATIVE

BACILLI (AGNB). (n=109)

ANTIBIOTICS Pseudomona E.Coli Klebsiella Acinetobact Enterobact Proteus Serratia

TESTED s Aeruginosa Pneumoni er er Marcesce
(n=49) (n=31) ae (n=12) (n=8) (n=4) (n=4) ns (n=1)

CEPHALOSPORINS
Cefoclor Sensitive 4(8.1%) 3(9.67%) - 1(12.7%) 1(25%) 1(25%) -
Resistant 45(91.8%) 28(90.32%) 12(100%) 7(87.5%)_ 3(75%) 3(75%) 1(100%)
Cefotaxi Sensitive 8(16.32%) 8(25.80%) 5(41.66%) 1(12.7%) 2(50%) - -
me Resistant 41(83.67%) 23(74.19%) 7(58.33%) 7(87.5%)_ 2(50%) 4(100%) 1(100%)

Ceftriaxo Sensitive 5(10.20%) 8(25.80%) 6(50%) 1(12.7%) 2(50%) - -

ne Resistant 44(89.79%) 23(74.19%) 6(50%) 7(87.5%) 2(50%) 4(100%) 1(100%)

Cefopera Sensitive 26(53.06%) 9(29.03%) 2(16.66%) - 3(75%) 1(25%) -

zone Resistant 23(46.93%) 22(70.96%) 10(83.33%) 8(100%) 1(25%) 3(75%) 1(100%)

Cefopera Sensitive 49(100%) 28(90.32%) 12(100%) 6(75%) 3(75%) 4(100%) -

zone+Sul Resistant - 3(9.67%) - 2(25%) 1(25%) - 1(100%)
bactum
Ceftazidi Sensitive 12(24.48%) 15(48.38%) 11(91.66%) - 3(75%) 3(75%) -
me Resistant 37(75.51%) 16(51.61%) 1(8.33%) 8(100%) 1(25%) 1(25%) 1(100%)

Cefpodox Sensitive - - - - 1(25%) 1(25%) -

ime Resistant 49(100%) 31(100%) 12(100%) 8(100%) 3(75%) 3(75%) 1(100%)

QUINOLONES
Ofloxacin Sensitive 19(38.77%) 2(6.45%) 2(16.66%) 2(25%) 4(100%) - -
Resistant 30(61.22%) 29(93.54%) 10(83.33%) 6(75%) - 4(100%) 1(100%)
Ciproflox Sensitive 22(44.89%) 4(12.90%) 2(16.66%) 5(62.5%) 4(100%) - -
acin Resistant 27(55.10%) 27(87.04%) 10(83.33%) 3(37.5%) - 4(100%) 1(100%)

Enoxacin Sensitive 17(34.69%) 2(6.45%) 6(50%) 6(75%) 2(50%) - -

Resistant 32(65.30%) 29(93.54%) 6(50%) 2(25%) 2(50%) 4(100%) 1(100%)
Sparfloxa Sensitive 18(36.73%) 15(48.38%) 6(50%) 6(75%) 4(100%) 3(75%) -
cin Resistant 31(63.26%) 16(51.61%) 6(50%) 2(25%) - 1(25%) 1(100%)

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PENICILLINS & ALIKE AGENTS
Carbenici Sensitive - 26(83.87%) 10(83.33%) 7(87.5%) 4(100%) 1(25%)
llin Resistant 49(100%) 5(16.12%) 2(16.66%) 1(12.7%) - 3(75%) -
1(100%)
Co- Sensitive - 15(48.38%) 6(50%) 2(25%) 2(50%) - -
amoxilav Resistant 49(100%) 16(51.61%) 6(50%) 6(75%) 2(50%) 4(100%) 1(100%)
Ampicil+ Sensitive - 9(29.03%) 1(8.33%) 7(87.5%) 1(25%) - -
Sulbactm Resistant 49(100%) 22(70.96%) 11(91.66%) 1(12.7%) 3(75%) 4(100%) 1(100%)

Piperacill Sensitive 35(71.42%) 26(83.82%) 8(66.66%) 4(50%) 4(100%) 3(75%) 1(100%)

in+Tazob Resistant 14(28.57%) 5(16.12%) 4(33.33%) 4(50%) - 1(25%) -
actum

Imipene Sensitive 47(95.91%) 26(83.82%) 10(83.88%) 7(87.5%) 4(100%) 1(25%) 1(100%)

m Resistant 2(4.08%) 5(16.12%) 2(16.66%) 1(12.7%) - 3(75%) -

Aztreona Sensitive 12(24.48%) 2(6.45%) 3(25%) - 2(50%) 2(50%) -

m Resistant 37(75.51%) 29(93.54%) 9(75%) 8(100%) 2(50%) 2(50%) 1(100%)

AMINOGLYCOSIDES
Gentamic Sensitive 24(48.97%) 21(67.74%) 10(83.88%) 6(75%) - - -
in Resistant 25(51.02%) 10(32.25%) 2(16.66%) 2(25%) 4(100%) 4(100%) 1(100%)

Tobramy Sensitive 26(53.06%) - 6(50%) - 1(25%) 1(25%) -

cin Resistant 23(46.93%) 31(100%) 6(50%) 8(100%) 3(75%) 3(75%) 1(100%)

Amikicin Sensitive 30(61.22%) 26(83.87%) 12(100%) 6(75%) 1(25%) 2(50%) -

Resistant 19(38.77%) 5(16.12%) - 2(25%) 3(75%) 2(50%) 1(100%)

TABLE II: Antibiotic Susceptibility profile of the isolated Gram positive organisms .
(n=27)

ANTIBIOTICS TESTED MSSA MRSA

( n=17) (n=10)
Sensitive Resistant Sensitive Resistant
PENICILLINS
Oxacillins 16(94.11%) 1 (5.89%) - 10(100%)
Amoxycillin 15(88.23%) 2(11.76%) - 10(100%)
Co-amoxiclav 17(100%) - - 10(100%)
Penicillin G 2(11.78%) 15(88.23%) - 10(100%)
Ampicillin + Salbactum 8(47.05%) 9(52.94%) - 10(100%)
Piperacillin + Tazobactum 17(100%) -
Impipenem 17(100%) - 2(20%) 8(80%)
CEPHALOSPORINS
Cefazolin 9(52.94%) 8(47.05%) - 10(100%)
Cefaclor 8(47.05%) 9(52.94%) - 10(100%)
Cefotaxime 11(64.70%) 6(35.29%) - 10(100%)
Ceftriaxone 11(64.70%) 6(35.29%) - 10(100%)
Cefoperazone 10(58.82%) 7(41.17%) - 10(100%)
Cefoperazone + Salbactum 17(100%) - - 10(100%)
Ceftazidime 16(94.11%) 1(5.89%) - 10(100%)

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QUINOLONES
Ofloxacin 16(94.11%) 1(5.89%) - 10(100%)
Ciprofloxacin 14(82.35%) 3(17.64%) - 10(100%)
Sparfloxacin 15(88.23%) 2(11.76%) - 10(100%)
Enoxacin 16(94.11%) 1(5.89%) - 10(100%)
MISCELLANEOUS
Erythrocin 6(35.29%) 11(64.70%) 2(20%) 8(80%)
Vancomycin 17 (100%) - 10(100%) -
Fusidic acid 17(100%) - 7(70%) 3(30%)
Teicoplanin 17(100%) - 7(70% 3(30%)

13