Microbial Profile and Utility of Soft Tissue,Pus,and Bone Cultures in Diagnosing Diabetic Foot Infections

Abstract

Objective:

This study assessed the utility of pus, soft tissue, and bone specimens in diagnosing diabetic foot infections and the spectrum of the microbial flora and in vitro susceptibility to antibiotics.

Subjects and Methods:

This prospective study was carried out in 60 consecutive patients with diabetes having clinically infected foot ulcers. Detailed history, physical examination, and investigation were carried out to diagnose the presence of osteomyelitis and the microbial etiology of foot ulcers. Foot ulcers were classified as per Wagner's classification. Soft tissue, pus, and bone samples were obtained and cultured for aerobic and anaerobic bacteria, and antimicrobial susceptibility testing was carried out as per the standard protocol.

Results:

Causative bacteria were isolated in 55 of 60 patients, and 157 isolates were cultured from 117 specimens with an average of 1.34 isolates per cases; however, the number of isolates per specimen did not differ among the various types of samples (P=0.78). Pus and soft tissue had predominantly polymicrobial flora, whereas bone infections were monomicrobial. The isolates from soft tissue specimens were different from those from bone and pus in 57% and 54% of cases, respectively. The common bacterial isolates from 117 specimens included Escherichia coli (21%) and Proteus species (15.9%). Nearly 70% of Staphylococcus aureus isolates were methicillin sensitive. All S. aureus and Enterococcus isolates were sensitive to vancomycin. Susceptibility of Gram-negative organisms to ciprofloxacin was 50%.

Conclusions:

Diabetic foot infections are mostly polymicrobial with Gram-negative predominance. Multiple sampling from superficial and deep tissues, including bone, when involved, yields more relevant information diagnostically and therapeutically.

Introduction

D iabetic foot ulcer is a common cause of morbidity and mortality, and the lifetime risk of a individual with diabetes for development of foot ulcer is approximately 25%.¹ Infection is a common sequela of foot ulceration, which usually follows repeated trauma in a neuropathic and/or ischemic limb.

India is the diabetes capital of the world, being the home of nearly 62 million people with diabetes, and this number is expected to grow further.² Because of the relatively low case fatality rate of foot ulcers, there is a growing concern of its increasing burden. Diabetic foot-related sepsis is responsible for one-fifth of diabetes-related hospital admissions.³ In our previous study, foot sepsis was related to 8% of hospital deaths in patients with diabetes.⁴

Most of the diabetic foot infections are polymicrobial, and if cultures are not taken properly, mixed bacterial flora of no or doubtful significance are frequently encountered. The spectrum of organisms depends mainly on duration of the wound, prior use of antibiotics, metabolic derangements, and practice of foot hygiene.⁵ Hence proper methodology of obtaining appropriate culture specimens is of utmost importance. Optimal management of diabetic foot ulcers includes appropriate surgical debridement, off-loading of the involved foot, restoration of vascularity, appropriate antibiotics, and treatment of acute metabolic derangements. Although there is growing concern about increasing multidrug-resistant organisms leading to an increase in morbidity and mortality,⁶ the data on this issue are sparse from India. Hence this prospective study was carried out to find out the bacterial flora of various tissue specimens in diabetic foot infections and their susceptibility to antibiotics.

Subjects and Methods

Patient selection

This prospective study was carried out in 60 consecutive diabetes patients with clinically infected foot ulcers admitted under the Endocrine and General Surgery Services of the Postgraduate Institute of Medical Education and Research, Chandigarh, India, during the period of January 2009 to May 2010. Written informed consent was obtained from each participant, and the Institute's Ethics Committee approved the study protocol.

Assessment of study subjects

Age, sex, type and duration of diabetes, glycemic control, presence of any microvascular or macrovascular complications, and treatment modalities were noted for each case. Foot care practice was ascertained by using a questionnaire (Supplementary Table S1; Supplementary Data are available online at www.liebertonline.com/dia<http://www.liebertpub.com/dia>). Neuropathy was defined by vibration perception threshold of >25 mV and/or absence of response in the monofilament test. Nephropathy was defined as either estimated glomerular filtration rate (calculated using the Modification of Diet in Renal Disease formula) of <60 mL/min and/or the presence of overt proteinuria. Peripheral vascular disease was assessed clinically by history of claudication or rest pain or signs of ischemia and confirmed by either an ankle brachial index of <0.8 and/or imaging modalities. Foot ulcers were graded using the classification of Wagner.⁷ Site, size, and duration of ulcer, presence of clinical signs of infections, treatment, and outcomes were noted in each case. Severities of infections were graded as described previously as non–limb-threatening, limb-threatening, and life-threatening.⁵ For diagnosis of osteomyelitis, plain X-ray of the foot was done in all cases, and the findings of bone destruction, periosteal reaction, bone sclerosis, periosteal new bone formation, and sequestra formation were taken as diagnostic of osteomyelitis. With negative X-ray foot and a high clinical suspicion of osteomyelitis, a three-phase bone scan with ⁹⁹Tc bisphosphonate was done. An intense uptake in all three phases was taken as evidence for osteomyelitis. Probe testing using a sterile blunt probe was also used to examine its efficiency in diagnosing osteomyelitis.

Method of obtaining specimens for culture

Soft tissue specimens (n=60) were obtained from all patients at the time of admission, after the surface of the wound was washed vigorously by saline followed by debridement of superficial exudates. Specimens were obtained by scraping the ulcer base or the deep portion of the wound edge with a sterile surgical blade. Pus cultures (n=22) were taken by needle aspiration from ulcers with evident pus formation. Bone specimens for culture were obtained from 35 ulcers with suspected osteomyelitis. Bone biopsies were performed surgically using a sterile bone nibbler or a bone curette under full aseptic conditions. Bone specimens were also obtained during surgical debridement procedures and amputations. If the bone was liquefied and surgical methods demanded large incision, ultrasound-guided bone aspiration was done by an intervention radiologist. Needle puncture was performed after the normal skin around the ulcer was first cleaned with detergent, before antiseptic was applied. On the first occasion, the operator located the area of bone or joint infection from previously taken foot X-rays to determine the direction and depth of the needle entry. Then the operator inserted an 18-gauge needle mounted on a syringe across the normal skin surrounding the foot ulcer under ultrasound guidance. The aspirate was diluted into 1 mL of sterile saline solution. Air was eliminated, and the syringe was closed with a sterile cap. The specimens were promptly sent to the laboratory and processed for aerobic and anaerobic bacteria. Standard conventional methods for isolation and identification of aerobic and anaerobic bacteria were followed. Anaerobic culture was done with the Anoxomat™ automated system (Mart Microbiology B.V., Drachten, The Netherlands).

The number(s) and/or species of organisms isolated from two different specimens in the same patient, if similar, was considered identical. Otherwise, they were considered non-identical.

Susceptibility testing

Antimicrobial susceptibility was tested according to standard protocol.⁸ As per protocol, all the isolated organisms were tested against a set of first-line antibiotics. They were tested for second-line antibiotics only when these organisms were resistant to the first-line antibiotics.

Statistical methods

Statistical analysis was performed using SPSS version 10 software (SPSS Inc., Chicago, IL). Data were expressed as percentages for categorical variables and as mean±SD values for continuous variables. Logistic regression was used to calculate predictors of osteomyelitis. A value of P≤0.05 was taken as statistically significant.

Results

Of the 60 patients with diabetic foot ulcers, 53 (88.3%) were male. Fifty-six patients (93.3%) had type 2 diabetes mellitus, and four patients (6.7%) had type 1 diabetes. Mean age of the patients was 52.6±11 years (range, 22–82 years), and mean duration of diabetes was 10.8±7.0 years, with nearly 60% having had diabetes of more than 10 years in duration. The mean fasting plasma glucose and glycosylated hemoglobin (HbA1c) were 148.0±56.2 mg/dL and 9.5±2.2%, respectively. More than 90% of patients had poor glycemic control (fasting plasma glucose ≥130 mg/dL and HbA1c >7%). In the study population, 35 (58.3%) were on oral hypoglycemic agents, 13 (21.7%) were receiving insulin, seven (11.7%) were treatment naive, and five (8.3%) were on alternative medications.

Foot care practices were followed by only 25% of the study population. Peripheral neuropathy and peripheral vascular disease were present in 90% and 31.6% of the subjects, respectively. The etiology of ulcer was unknown in 31.9%, footwear-related in 26.6%, ulceration of callus in 21.6%, trivial trauma in 18.3%, and due to thermal injury in 1.6%. Neuropathic ulcers were more common than ischemic ulcers.

The most common site of ulcers was toes (36.7%), followed by forefoot (26.6%), heel (20%), and midfoot (16.7%). Seventeen patients (28.3%) had Wagner's Grade 2 ulcers, 41 patients (68.4%) had Grade 3 or 4 ulcers, and two (3.3%) had Grade 5 ulcers. Limb-threatening foot ulcer was present in 39 (65%), and life-threatening ulcers occurred in 10 (16.7%) patients. Twelve (20%) patients required amputation as a part of treatment.

Plain X-ray foot was done in all (n=60), of which 30 showed evidence of osteomyelitis. Three-phase bone scan was done in 10 patients who had strong suspicion of osteomyelitis, and in five of them it was suggestive of osteomyelitis. Thus in total 35 (58.3%) patients had underlying osteomyelitis on imaging. Surgical bone biopsy was done in 27 patients, and image (ultrasound guidance/computed tomography)-guided bone aspiration was performed in eight patients (n=35). The bone culture was positive in 28/35 (80%) of bone specimens sent for cultures. Taking bone culture as the gold standard, the probe test showed 86% sensitivity, 62% specificity, 68% positive predictive value, and 83% negative predictive value. Using logistic regression analysis, duration of ulcer was the strongest predictor of osteomyelitis (odds ratio, 1.2; P=0.007).

Diagnostic utility of soft tissue, pus, and bone cultures

Soft tissue culture was done in all the patients with ulcers (n=60); pus culture was done in 22, and bone specimens were cultured in 35 patients with osteomyelitis. Therefore, in total 117 specimens were processed for culture. Of the 60 patients, organisms could be isolated in 55, whereas in five the wound cultures were sterile. In total, 157 isolates were grown, indicating an average yield of 1.34 isolates per specimen. The number of isolates from soft tissue, with a yield of 1.4 bacterial species per specimen, was higher than those from pus (1.3 isolates per specimen) and bone (1.26 isolates per specimen); however, the difference was statistically nonsignificant (P=0.78). The majority (78.3%) of isolates were aerobic or microaerophilic, whereas 21.7% were anaerobes (aerobe to anaerobe ratio, 3:1). Gram-negative organisms were isolated in 66.2%, and the ratio of Gram-negative to Gram-positive organisms was 2:1 (Table 1).

Table 1.

Microbiological Results of Pus, Soft Tissue, and Bone Cultures

Specimen (n=117)	No. of isolates	No. of isolates per specimen	Gram-positive isolates (aerobe+anaerobe)	Gram-negative isolates (aerobe+anaerobe)	Gram-negative/Gram-positive ratio	Aerobes	Anaerobes	Aerobic/anaerobic ratio
Pus (n=22)	29	1.31	12 (41.4%)	17 (58.6%)	1.4	22 (75.9%)	7 (24.1%)	3.1
Tissue (n=60)	84	1.4	30 (35.7%)	54 (64.3%)	1.8	64 (76.2%)	20 (23.8%)	3.2
Bone (n=35)	44	1.26	11 (25%)	33 (75%)	3	37 (84.1%)	7 (15.9%)	5.3
Total	157		53 (33.8%)	104 (66.2%)	1.9	123 (78.3%)	34 (21.7%)	3.6

The commonest bacterial isolate was Escherichia coli in 33 (21%), followed by Proteus species in 25 (15.9%) specimens, methicillin-sensitive Staphylococcus aureus in 15 (9.5%), Klebsiella pneumoniae in 10 (6.4%), methicillin-resistant S. aureus in six (3.8%), and Pseudomonas aeruginosa in eight (5.1%). Among the anaerobic organisms, Peptostreptococcus species, isolated from 22 (14.1%) specimens, was the most common, followed by Bacteroides species in 10 (16.6%) and Prevotella species in two (3.3%). Of the 60 patients, 33 (55%) had polymicrobial infection, and 22 (37%) had monomicrobial infection, whereas five (8%) had a sterile culture (Table 2). As the Wagner's grade increased the prevalence of polymicrobial infection also increased (Supplementary Fig. S1). Pus and tissue cultures had predominantly polymicrobial infections, whereas bone culture had mainly monomicrobial infection.

Table 2.

Profile of Bacterial Isolates from Pus, Soft Tissue, and Bone Cultures

	n (%)
Bacterial category	Pus (19/22)	Soft tissue (54/60)	Bone (28/35)
Number of isolates	29	84	44
Aerobic and facultative isolates	22 (75.9%)	64 (76.2%)	37 (84.1%)
Gram-negative	14 (48.3%)	47 (55.9%)	31 (70.4%)
E. coli	7 (24.1%)	16 (19%)	10 (22.7%)
Proteus species	3 (10.3%)	14 (16.7%)	8 (18.2%)
K. pneumoniae	2 (6.9%)	5 (5.9%)	3 (6.8%)
P. aeruginosa	0	5 (5.9%)	3 (6.8%)
Acinetobacter species	0	3 (3.6%)	3 (6.8%)
Citrobacter species	0	0	1 (2.3%)
Enterobacter species	0	1 (1.1%)	2 (4.6%)
Morganella	1 (3.4%)	3 (3.6%)	1 (2.5%)
Providencia	1 (3.4%)	0	0
Gram-positive	8 (27.6%)	17 (20.2%)	6 (13.6%)
MSSA	4 (13.8%)	8 (9.5%)	3 (6.8%)
MRSA	1 (3.4%)	4 (4.8%)	1 (2.3%)
β-Hemolytic Streptococcus	0	1 (1.2%)	0
Enterococcus species	3 (10.3%)	4 (4.8%)	2 (4.6%)
Anaerobic isolates	7 (24.1%)	20 (23.8%)	7 (15.9%)
Gram-positive	4 (13.8%)	13 (15.4%)	5 (11.4%)
Peptostreptococcus	4 (13.8%)	13 (15.4%)	5 (11.4%)
Clostridium species	0	0	0
Gram-negative	3 (10.3%)	7 (8.3%)	2 (4.6%)
Bacteroides species	3 (10.3%)	6 (7.1%)	1 (2.3%)
Prevotella species	0	1 (1.2%)	1 (2.3%)

MRSA, methicillin-resistant S. aureus; MSSA, methicillin-sensitive S. aureus.

When different specimens taken for culture were compared for microbiological profile in the same patient, the isolates from pus and soft tissue cultures were identical in 46% (9/22), and those from soft tissue and bone were identical in 43% (15/22). The organisms isolated from pus and bone culture were totally non-identical (100%) (Table 3).

Table 3.

Comparisons of Culture Results of Different Specimens in the Same Patient

	Pus–soft tissue	Pus–bone	Soft tissue–bone
Identical	10 (46%)	0 (0%)	15 (43%)
Non-identical	12 (54%)	12 (100%)	20 (57%)
Total	22	12	35

Antibiotic susceptibility and resistance pattern of different microbial flora

Of S. aureus isolates, 28.6% exhibited oxacillin (methicillin) resistance. All Gram-positive isolates were sensitive to vancomycin. The Enterococcus isolates showed good susceptibility to amoxicillin and erythromycin. In the Gram-negative aerobic series, the majority (88–100%) showed sensitivity to the cefoperazone and sulbactam combination, amikacin, and ceftazidime. However, the isolates resistant to first-line antibiotics showed good susceptibility to second-line antibiotics, including the piperacillin and tazobactam combination, meropenem, imipenem, polymyxin, and cefepime.

The susceptibility of Gram-negative organisms to ciprofloxacin was poor (40–60%). The prevalence of Pseudomonas infection was low (5.1%), with high susceptibility to first-line antibiotics. The anaerobic organisms were highly (90–100%) susceptible to metronidazole and most of the first-line antibiotics (Tables 4 –6).

Table 4.

Antimicrobial Susceptibility Pattern of Aerobic Gram-Positive Organisms

Antibiotic/organism	S. aureus (n=21)	β-Hemolytic Streptococcus (n=1)	Enterococcus spp. (n=10)
Oxacillin	15 (71.4%)	—	—
MRSA	6	—	—
Vancomycin	6 (100%)	—	10 (100%)
Erythromycin	13 (61.9%)	1 (100%)	7 (70%)
Amoxicillin	—	1 (100%)	8 (80%)
Ampicillin	—	1 (100%)	—
Netilmycin	19 (90.5%)	—	—
Gentamicin	17 (81%)	—	8 (80%)
Amikacin	19 (90.5%)	—	—
Ciprofloxacin	9 (42.9%)	—	—

MRSA, methicillin-resistant S. aureus.

Table 5.

Antimicrobial Susceptibility Pattern of Aerobic Gram-Negative Organisms

Antibiotic	E. coli (n=33)	Proteus spp. (n =25)	K. pneumoniae (n=10)	P. aeruginosa (n=8)	Acinetobacter spp. (n=6)
Gentamicin	24 (72.7%)	19 (76%)	5 (50%)	7 (87.5%)	3 (50%)
Amikacin	28 (84.8%)	23 (92%)	7 (70%)	7 (87.5%)	4 (66.7%)
Ciprofloxacin	14 (42.4%)	15 (60%)	6 (60%)	5 (62.5%)	3 (50%)
Cefotaxime	26 (78.8%)	19 (76%)	5 (50%)	—	2 (33.3%)
Ceftazidime	25 (75.8%)	20 (80%)	6 (60%)	5 (62.5%)	3 (50%)
Cefoperazone	—	—	—	6 (75%)	4 (66.7%)
Cefoperazone+sulbactum	29 (88%)	25 (100%)	7 (70%)	8 (100%)	4 (66.7%)
β-Lactamase resistance (n)	4		3		2
Piperacillin+tazobactum	4/4 (100%)	—	3/3 (100%)	—	2/2 (100%)
Imipenem	4/4 (100%)	—	3/3 (100%)	—	1/2 (50%)
Meropenem	4/4 (100%)	—	1/3 (33.3%)	—	0/2 (0%)
Polymixin	4/4 (100%)	—	3/3 (100%)	—	2/2 (100%)
Colistin	4/4 (100%)	—	3/3 (100%)	—	2/2 (100%)
Aztreonam	4/4 (100%)	—	3/3 (100%)	—	2/2 (100%)
Cefipime	3/4 (75%)	—	3/3 (100%)	—	1/2 (50%)

Table 6.

Antimicrobial Susceptibility Pattern of Anaerobic Organisms

	Peptostreptoccus spp. (n=22)	Bacteroides spp. (n=10)	Prevotella spp. (n =2)
Erythromycin	11 (50%)	3 (30%)
Ampicillin	16 (72.7%)	—	2 (100%)
Amoxicillin	—	—	2 (100%)
Amoxicillin+clavulanic acid	18 (81.8%)	4 (40%)	2 (100%)
Metronidazole	20 (90.9%)	10 (100%)	2 (100%)
Cefotaxime	20 (90.9%)	7 (70%)	2 (100%)
Chloroamphenicol	—	10 (100%)	—

Management and outcome

All patients received standard care, including antibiotics, surgical debridement, and strict glycemic control. In the case of polymicrobial isolates, antibiotic therapy was selected with the aim of covering the maximum number of organisms with priority for isolates from bone. At 3 months of follow-up, in 28 (46.7%) the ulcers were completely healed, whereas 23 (38.3%) were partially healed, and worsening of foot ulcers occurred in four (6.7%) requiring further surgery in the form of deep tissue debridement in one and toe amputation in three.

Discussion

In our study Gram-negative organisms were the predominant organisms, with E. coli being the commonest. Bacterial isolates per specimen from soft tissue culture and bone culture were similar; however, they were non-identical. Soft tissue culture yielded predominantly polymicrobial organisms, whereas bone cultures yielded monomicrobial organisms. The duration of ulcer was the strongest predictor of osteomyelitis.

The yield of organisms in our study was comparable to other studies by Bansal et al.⁹ and Viswanathan et al.¹⁰ The polymicrobial nature of diabetic foot infections has been observed in various studies. However, in contrast to Western studies, where Gram-positive bacteria were predominant,^11
–13 most of the studies from the Indian subcontinent, including the current one, have shown Gram-negative preponderance.^9,10,14 This is probably due to the long duration of ulcer and/or prior antibiotics therapy, which is known to alter the flora. The common bacterial isolates are variable in different studies from our country and worldwide. In the studies by Bansal et al.⁹ and Shankar et al.,¹⁴ P. aeruginosa was the most common isolate from swab culture, followed by S. aureus, coagulase-negative staphylococci, and members of the Enterobacteriaceae family, whereas in our study E. coli followed by Proteus species and methicillin-sensitive S. aureus were the common isolates. This could be because of the predominance of local flora in individual patients. Studies regarding anaerobic cultures in the diabetic foot are limited. It is surprising that anaerobes constituted one of the major isolates (22%), comprising Peptostreptococcus, Bacteroides species, and Prevotella species. The prevalence of anaerobic bacteria in cultures of specimens from foot infections in diabetes patients is dependent upon the method of collection, the care with which the sample is transported, and the sophistication of testing. Literature search revealed that the prevalence of anaerobic pathogens in the diabetic foot varies widely from 5% to 95%.^15,16

Prolonged or broad-spectrum, topical or systemic antibiotic therapy may predispose patients to infections with antibiotic-resistant organisms, but our study was unable to provide significant data pertaining to previous antibiotic treatment of the study cases.

Of the 60 patients with ulcers, 35 were diagnosed as having osteomyelitis on imaging, thus showing a prevalence of osteomyelitis in 58.3% in the study patients. However, 28 (46.6%) patients had culture-proven osteomyelitis. There is a wide variation in the reported prevalence of osteomyelitis in diabetic foot patients, ranging from 20% to 66%.^17,18 The reason for this variation may be the use of different techniques and operational definitions to diagnose osteomyelitis, duration of ulcer, and prior antibiotics used. In our study we have confirmed osteomyelitis by bone culture. The study done by Newman et al.¹⁹ found a 68% prevalence of osteomyelitis, as determined by bone biopsy and culture in 41 diabetic foot ulcers.

Soft tissue specimen cultures are not appropriate in the determination of causative organisms in patients of diabetic foot with osteomyelitis. Bone specimen for culture is considered the gold standard for isolating microorganisms in patients with diabetic foot infections with suspicion of osteomyelitis. However, it is difficult to obtain a bone specimen in all cases. In brief, multiple specimens are to be taken for better microbial yield. However, it is very surprising that microorganisms obtained from different specimens in the same patient were not identical. This has been observed by two previous studies that have looked into the microbial profile in bone biopsy and ulcer swab/soft tissue cultures and showed marked differences in microbial concordance (22–49%) between two specimens.^20,21 The reason for non-identical microbial profile between various specimens in a same patient may be explained as follows. Healthy skin has its own share of resident flora. In patients with underlying chronic debilitating illnesses, compromised local and general protective factors contribute to additional colonization with transient flora comprising true and conditional pathogens. In conditions presenting with superficial inflammation and ulceration, the serous and purulent products of inflammation and necrotic tissue result in saprophytic multiplication of resident commensal bacteria and transient colonizers in addition to those causing true invasive infection. The commensals and colonizers, better adapted to the ecological niche, have a potential to outgrow the non-adapted and fastidious invaders. This has possibly led to discordant flora isolated from superficial and deeper tissues. The deeper the source of specimen, the more representative and reliable it is of true infection than confounding colonizers.

The majority of S. aureus isolates were methicillin sensitive, and Gram-negative organisms were sensitive to third-generation cephalosporins and aminoglycosides. Recent reports suggest emergence of extended-spectrum β-lactamase-mediated resistance in patients with diabetic foot infection. However, we did not encounter this in our study. The duration of ulcers and prior use of antimicrobial therapy determine the frequency of antimicrobial resistance, which might be variable in different studies.

Although fungal infections are not uncommon in patients with a diabetic foot, it should be seriously considered in those patients who do not respond to optimal dose and duration of appropriate antibacterial therapy. Studies from India have shown the prevalence of fungal infections in patients with a diabetic foot varying from 9% to 27%.^5,22 Therefore, specimens for fungal culture would have strengthened the findings of our study.

The decision to administer antibiotic therapy should depend on these results, and hence the culture results of specimens taken concurrently from multiple sites are more informative for therapeutic decision-making. Studies regarding bone infections in diabetic foot osteomyelitis from this subcontinent are lacking.

Our study is the first in highlighting the spectrum of microbial flora in diabetic foot osteomyelitis from this part of the world and will encourage further research in this area; however, the major limitations of the study are small sample size, lack of previous details of antimicrobial therapy, and non-performance of fungal culture.

In conclusion, diabetic foot infections are mostly polymicrobial with Gram-negative predominance. The grade and duration of ulceration predicted the number of organisms and the occurrence of osteomyelitis, respectively. However, microbial profiles yielded by bone and soft tissue culture were highly non-identical. Therefore, in a given individual various specimens should be sampled for microbial profile.

Footnotes

Author Disclosure Statement

No competing financial interests exist.

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