Indian Journal of Pathology and Microbiology

: 2008  |  Volume : 51  |  Issue : 2  |  Page : 204--208

Spectrum of microbial flora in diabetic foot ulcers

Ekta Bansal1, Ashish Garg1, Sanjeev Bhatia1, AK Attri1, Jagdish Chander2,  
1 Department of Surgery, Government Medical College and Hospital, Chandigarh, India
2 Department of Microbiology, Government Medical College and Hospital, Chandigarh, India

Correspondence Address:
Ashish Garg
House No. 153, Sector - 21, Chandigarh - 160 022


A prospective study was carried out on patients with diabetic foot lesions to determine their clinical characteristics, the spectrum of aerobic microbial flora and to assess their comparative in vitro susceptibility to the commonly used antibiotics. A total of 157 organisms (143 bacteria and 14 fungi) were isolated and an average of 1.52 isolates per case was reported. Polymicrobial infection was found in 35% of the patients. In this study, Pseudomonas aeruginosa among the gram-negative (22%) and Staphylococcus aureus among the gram-positive (19%) were the predominantly isolated organisms, while Candida was the most predominantly isolated fungus. Antimicrobial sensitivity pattern of the isolates is discussed in detail. There was a linear increase in the prevalence of organisms with increase in Wagner«SQ»s grade. Neuropathy (76%) and peripheral vascular disease (57.28%) was a common feature among the patients. Poor glycemic control was found in 67% of the patients. Awareness about lower limb complications of diabetes was very low (23%) among the patients

How to cite this article:
Bansal E, Garg A, Bhatia S, Attri A K, Chander J. Spectrum of microbial flora in diabetic foot ulcers.Indian J Pathol Microbiol 2008;51:204-208

How to cite this URL:
Bansal E, Garg A, Bhatia S, Attri A K, Chander J. Spectrum of microbial flora in diabetic foot ulcers. Indian J Pathol Microbiol [serial online] 2008 [cited 2020 Feb 24 ];51:204-208
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Infected foot ulcer is a common cause of morbidity in diabetic patients, ultimately leading to dreaded complications like gangrene and amputations. Lifetime risk to a person with diabetes for developing a foot ulcer could be as high as 25%. [1] Infection is most often a consequence of foot ulceration, which typically follows trauma to a neuropathic foot. [2] The alarming fact is that India has more people with diabetes than any other country [3],[4] and the incidence of foot problems and amputations remains very high, accounting for up to 20% of diabetes-related hospital admissions. [5],[6] This can be easily attributed to several practices prevalent in India, such as barefoot walking, inadequate facilities for diabetes care, low socioeconomic status and illiteracy. [7]

The incidence of type 2 diabetes is rising to epidemic proportions in India and the whole world. [3],[4] Because of its relatively low case fatality rate, prevalence of associated chronic complications is expected to increase. The burden of diabetic foot is set to rise further in the future since its contributory factors such as peripheral neuropathy and peripheral vascular disease (PVD) are present in >10% of the cases at the time of diagnosis. [8]

The decision regarding proper management of diabetic foot infection is a difficult one and still a matter of debate. While optimal therapy is yet to be established, most authors agree that the management of these infections require isolation and identification of the microbial flora, appropriate antibiotic therapy according to the sensitivity patterns, precise selection and identification of the chronic complications and proper surgical intervention for these complications. [9] Most of the diabetic foot infections are polymicrobial in nature and mixed organisms are frequently encountered. [10],[11],[12],[13],[14],[15] However, the spectrum of microorganisms depends mainly on microbial flora of the lower limb, metabolic factors, foot hygiene and the use of antibiotics. [13]

Emergence of resistance among organisms against the commonly used antibiotics has been clearly outlined in various studies as being largely due to their indiscriminate use. [10],[11],[12],[13],[14],[15] There is a direct relationship between the total amount of a certain antibiotic used in a particular hospital during a certain period and the number of resistant strains that emerge. [10]

In view of the above facts, a prospective study was carried out to determine the relative frequency of aerobic microbial isolates cultured from community-acquired diabetic foot infections and to assess their comparative in vitro susceptibility to the commonly used antibiotics.

 Materials And Methods

A prospective study was carried out on 103 diabetic patients with foot ulcer during the period of January 2006 to January 2007 at the Center for Research and Treatment of Peripheral Vascular Diseases and Diabetic Foot, Department of Surgery, Government Medical College and Hospital, Chandigarh. Principles of "convenience sampling" were applied for collecting the detailed data in the present study.

Foot ulcers in diabetic patients were categorized into six grades (grade 0 - grade 5) based on Meggit Wagner Classification System. [16] The criterion for inclusion was a diabetic patient with foot ulcer of grade 1 or more. However, all patients with grade 0 and/or limb amputations were excluded from the study group.

A semistructured questionnaire was developed to record the medical history, examination details and investigation reports. A pilot study was carried out on 10 subjects. Shortcomings of the questionnaire and the other methods to be applied in the study were detected and corrections were made. Medical history was taken for all the subjects. Details regarding type of diabetes, [17] its duration, treatment, compliance by the patient, awareness about complications, personal habits like smoking and alcohol consumption were recorded. Meticulous clinical examination was done. Neuropathy was assessed by the ability to sense touch with a 10-gram monofilament and tuning fork, [17],[18],[19] ischemia by pulsations of dorsalis pedis and posterior tibial arteries, [17] while osteomyelitis (to assess bone involvement) was diagnosed on x-rays. [17] Cases with ulcer on the other foot also, were considered as separate cases.

Specimens (pus, wound exudates, or tissue biopsy) for microbiological studies were obtained from the ulcer region. Pus and exudates were collected from the margins and the base of the ulcer in 97 and 6 patients respectively using a sterile swab stick, which was then transported in a clean and sterile test tube. Tissue biopsy was taken with a sterile blade/knife in wedge shape including base and margin of ulcer along with wound swabs from the same site in 15 patients and was then transported in sterile solution of normal saline and sterile test tube respectively. Thus 118 specimens were collected from 103 patients. These specimens were immediately transported to the microbiology laboratory for further processing. Culture, isolation, antibiotic sensitivity and identification of the microorganisms were done according to the standard microbiological procedures. [20],[21] Due to lack of resources, anaerobic culture was not done; therefore, results were analyzed for aerobic flora only.

Antimicrobial susceptibility testing of aerobic isolates was performed by the Kirby Bauer disc diffusion method as recommended by the Clinical and Laboratory Standards Institute (CLSI). [22]

The data was collected and analyzed using descriptive statistics. Statistical significance was evaluated using Student's Chi-square Test.


In the 103 diabetic foot patients studied, 81 (78.64%) were men and 22 (21.36%) were women, male-to-female ratio being 3.68. The age ranged from 25 to 84 years (average = 57.04 years; s.d. = 11.63 years). Among those having diabetic foot ulcers, a majority of patients (56.31%) were in the age group 51 to 70 years.

Among the total patient population, 100 (97.08%) had type 2 diabetes mellitus, whereas only 3 (2.91%) patients had type 1 diabetes mellitus. While 50 (48.54%) of the patients were diagnosed to have diabetes mellitus for >10 years, 27 (26%) and 26 (25%) had diabetes for 5-10 years and n = 97), only 61 (62.89%) were fully compliant to their treatment whereas 36 (37.11%) were noncompliant to it. All the patients were evaluated for awareness about the complications related to diabetes. It was seen that 60% of the patients were completely unaware of the diabetes-related complications, while 33% were aware of some complications. Only 7% were aware of almost all the complications. Of the total patients, 77% were unaware of the lower limb complications, while only 23% were aware about lower limb complications of diabetes [Table 1]. Poor glycemic control, i.e., random blood sugar (RBS) >200 mg/dL, was found in 69 (67%) patients and HbA 1c >7 was found in 59 (64%) patients (HbA 1c data available for 92 patients only) [Table 1].

Out of 103 patients, 56 (54%) were found to be anemic, 78 (76%) had neuropathy, 59 (57%) showed signs of ischemia, 31 (30%) had osteomyelitis. The involvement of the other foot in the form of callus, neuropathy, ischemia, ulcer, amputation was found in 77 (75%) patients. Maximum number of organisms were isolated from grade IV foot wounds ( n = 65). Patients were graded according to Meggit Wagner Classification [16] and their types of flora are given in [Table 2].

Microbiological observations

In the 103 patients studied, 157 organisms (143 aerobic bacteria and 14 fungi) were isolated in 118 specimens, which represent an average of 1.52 organisms per case. In 101 (85.6%) specimens, only bacteria were isolated, whereas mixed infection of both bacteria and fungi was found in 10 (8.4%) specimen, while 7 (5.9%) specimens were found to be sterile. Of the total 61 patients who had received antibiotic prior to the sample collection, 6 had sterile flora, 39 had monomicrobial flora, whereas 16 had polymicrobial flora.

Among the bacterial isolates, gram-negative comprised of 76% and gram-positive accounted for 24%. Pseudomonas aeruginosa was the most common isolate, accounting for 21.67%; followed by Staphylococcus aureus , Escherichia coli and Klebsiella pneumoniae, comprising 18.88%, 18.18% and 16.78% respectively. When compared, HbA 1c , duration of diabetes, prevalence of PVD and neuropathy were similar in both the gram-positive and gram-negative groups. The prevalence of various bacterial isolates is shown in [Table 3].

Fungal isolates accounted for 9% of the total isolates. Among the Candida sp., predominant were C. tropicalis (29%), C. albicans (14%) and C. guililermondii (7%); followed by Aspergillus flavus (21%), A. niger (14%) and Fusarium sp. (14%) [Table 3].

The results of the test for susceptibility to the commonly used antibiotics are shown in [Table 4]. Antibiotic sensitivity pattern of S. aureus showed that Oxacillin resistance, i.e., Methicillin-resistant S. aureus (MRSA), was 55.50%. Almost all the strains were sensitive to Ceftriaxone and Imepenem. Amikacin and Ciprofloxacin also showed good sensitivity. E. fecalis showed absolute resistance to Cotrimoxazole and Amikacin, while resistance to Erythromycin and Amoxycillin was 75% and 42% respectively. Majority of the strains were sensitive to Cefoperazone + Sulbactum (85%), Amoxycillin + Clavulanic Acid (71%), Ciprofloxacin (62%) and Gentamicin (66%). Almost all the isolates of P. aeruginosa were sensitive to Cefoperazone + Sulbactum, Ceftazidime and Imepenem, while Piperacillin showed good activity. Very high resistance was shown to Cotrimoxazole (100%), Amoxycillin + Clavulanic Acid (97%). In E. coli , majority of strains were resistant to Penicillins, while sensitivity was shown to Imepenem (100%), Cefoperazone + Sulbactum (96%), Amikacin (90%). In case of Proteus Sp., majority of the strains were sensitive to majority of the antibiotics. Amikacin, Cefoperazone + Sulbactum, Ceftazidime showed absolute sensitivity [Table 4].

Out of the two most commonly used combinations (Amoxycillin + Clavulanic Acid, Cefoperazone + Sulbactum), Cefoperazone + Sulbactum exhibited broad activity with only 8 out of 143 isolates harboring resistance (antibiotic sensitivity = 94.40%), while 92 out of 143 isolates showed resistance to Amoxycillin + Clavulanic Acid (antibiotic sensitivity = 35.64%).


In the present study, a total of 157 organisms were isolated from 103 patients and an average of 1.52 organisms per case was found. This is slightly higher than the findings by Vishwanathan et al., [12] where cultures yielded an average of 1.21. Polymicrobial nature of diabetic foot infections has been observed in various studies in the subcontinent and abroad. [10],[11],[12],[13],[14],[15] Gram-negative organisms were the predominant type found, which is also in concordance with the findings of Shankar et al. [5] In the present study, P. aeruginosa , gram-negative (22%); and S. aureus , gram-positive (19%), were predominantly isolated, while the prevalence of other organisms like K. pneumoniae , E. coli , Proteus sp. was 17%, 18% and 11% respectively. Almost similar results were obtained by Chincholikar et al. [13] [ P. aeruginosa (19%), K. pneumoniae (18%), E. coli (15%) and Proteus sp . (9.3%)], though they reported highest positivity of S. aureus (31%) in their study. Ramani et al. [10] also made similar observations and found Proteus (20.73%), Klebsiella (12.35%), Pseudomonas (11.73%) as the most common isolates. Prabhakar et al. [23] also showed similar results.

As regards to the antibiotic sensitivity pattern of gram-positive cocci, more than 75% strains were sensitive to Cephalosporins. Cefoperazone + Sulbactum showed around 85% sensitivity, while Ciprofloxacin and Gentamicin were about 65% sensitive. Similar results were found by Chincholikar et al., [13] as their sensitivity patterns were cephalosporins, 74%; Ciprofloxacin, 77.27%; penicillin, 50%. Methicillin resistance was seen in 55.56% of the S. aureus, which is also in concordance with findings by Gadepalli et al., [24] in which MRSA was seen in 56% of the cases of S. aureus; whereas in the study by Tentolouris et al., [25] MRSA was present in 40% of the cases. P. aeruginosa , the most predominant isolate, showed sensitivity to Amikacin, Ceftazidime, Cefoperazone + Sulbactum, Piperacillin and Imepenem.

Fungi seldom behave as a pathogen in normal host but occur most often in the immunocompromised host with or without an underlying pathology. In our study, Candida sp. was most predominantly isolated, with C. tropicalis (29%) being the predominant species. Similarly, Chincholikar et al. [13] reported high prevalence of Candida [ C. tropicalis (68%), C. albicans (16.6%) of the total Candidal isolates]. Chakarbarti et al. [22] too have reported C. tropicalis as the predominant isolate (42.1%).


Diabetic foot infections are polymicrobial in nature. As the Wagner's grade increased, the prevalence of isolates also increased. While Pseudomonas aeruginosa was the most frequent isolate among the gram-negative pathogens, Staphylococcus aureus was the most common among gram-positive organisms. Candida was the predominant isolate among fungal pathogens. Ciprofloxacin, Imepenem, or Cefoperazone + Sulbactum would be appropriate for empiric treatment.


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