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 ~  Abstract
 ~ Introduction
 ~ Methodology
 ~ Results
 ~ Discussion
 ~ Conclusion
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  Table of Contents  
ORIGINAL ARTICLE
Year : 2019  |  Volume : 37  |  Issue : 1  |  Page : 19-23
 

Early outcome of culture-negative infection in open fractures of the lower limb: A prospective study


1 Department of Orthopaedics, Christian Medical College, Vellore, Tamil Nadu, India
2 Department of Clinical Microbiology, Christian Medical College, Vellore, Tamil Nadu, India

Date of Web Publication16-Aug-2019

Correspondence Address:
Dr. Abel Livingston
Department of Orthopaedics, Christian Medical College, Vellore - 632 004, Tamil Nadu
India
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/ijmm.IJMM_19_143

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 ~ Abstract 


Background: Culture-negative infections in open long bone fractures are frequently encountered in clinical practice. We aimed to identify the rate and outcome of culture-negative infections in open long bone fractures of lower limb. Methodology: A prospective cohort study was conducted from November 2015 to May 2017 on Gustilo and Anderson Grade III open long bone fractures of the lower limb. Demographic data, injury details, time from injury to receiving antibiotics and index surgical procedure were noted. Length of hospital stay, number of additional surgeries and occurrence of complications were also noted. Patients with infected open fractures were grouped as culture positive or culture negative depending on the isolation of infecting microorganisms in deep intraoperative specimen. The clinical outcome of these two groups was statistically analysed. Results: A total of 231 patients with 275 open fractures involving the femur, tibia or fibula were studied. There was clinical signs of infection in 84 patients (36.4%) with 99 fractures (36%). Forty-three patients (51.2%) had positive cultures and remaining 41 patients had negative cultures (48.8%). The rate of culture-negative infection in open type III long bone fractures in our study was 17.7%. There was no statistical difference in the clinical outcome between culture-negative and culture-positive infections. Conclusion: Failure to identify an infective microorganism in the presence of clinical signs of infection is routinely seen in open fractures and needs to be treated aggressively.


Keywords: Culture-negative infection, open fracture, trauma


How to cite this article:
Hariharan T D, Joseph CM, Samuel S, Elangovan D, Livingston A, Ramasamy B, Nithyananth M, Jepegnanam T. Early outcome of culture-negative infection in open fractures of the lower limb: A prospective study. Indian J Med Microbiol 2019;37:19-23

How to cite this URL:
Hariharan T D, Joseph CM, Samuel S, Elangovan D, Livingston A, Ramasamy B, Nithyananth M, Jepegnanam T. Early outcome of culture-negative infection in open fractures of the lower limb: A prospective study. Indian J Med Microbiol [serial online] 2019 [cited 2019 Oct 21];37:19-23. Available from: http://www.ijmm.org/text.asp?2019/37/1/19/264487





 ~ Introduction Top


Open long bone fracture is commonly encountered in tertiary level orthopaedic trauma care.[1] Infection is a common complication of open fractures. Chronic osteomyelitis, non-union, loss of function or even limb loss are some serious complications of deep surgical site infections. Further, it contributes to increase in healthcare costs and decrease in the quality of life.[2] Primary goal in management of open fractures, in addition to achieving union, is prevention of infection by early debridement, irrigation of wound and administration of broad-spectrum antibiotics with stabilisation of fracture.[3] However, deep surgical site infection is common, and clinicians make treatment decisions based on signs and symptoms, laboratory and imaging workup and bacteriological culture results. The standard of care for treating infections which complicate open fractures involves accurately identifying the infecting microorganism and providing adequate antibiotic therapy based on culture and sensitivity. This is supplemented with additional surgical procedures as necessary. However, negative culture results are frequently encountered even when there are clinical signs of infection. Negative bacteriological culture results are challenging to the clinician, as they may be false-negative and empirical treatment of the same may result in inadequate resolution of infection and development of resistant microorganisms.[4] The reported incidence of culture-negative infections in orthopaedic trauma is 9%, but its incidence in open fractures and its outcome has not been described in the literature.[4]

We aimed to identify the rate of culture-negative infections in open long bone fractures of the lower limb and study its outcome. We hypothesised that culture-negative patients with clinical signs of infection have a better outcome than culture-positive patients.


 ~ Methodology Top


A prospective cohort study was conducted from November 2015 to May 2017 on Gustilo and Anderson Open Type III fractures of the lower limb. The study protocol was reviewed and approved by our Institutions Research and Ethics Committees (IRB Min No: 9693 dated 20th October, 2015).

Inclusion criteria

All adult patients with open type III fractures of the femur, tibia or fibula were included in the study. Patients in whom index surgical procedure was done elsewhere and patients with unsalvageable limbs requiring urgent amputation were excluded from the study.

Treatment protocol

A standard emergency room protocol was followed for open fractures for all included study patients. This included wound wash (5 L normal saline) after appropriate analgesia, temporary stabilisation with splints and prophylactic antibiotics followed by radiographic assessment. The standard prophylactic antibiotic regimen followed was crystalline penicillin for clostridial infection, cloxacillin for Gram-positive and gentamicin or ciprofloxacin for Gram-negative organisms as per the East Practice Management Guidelines 2011.[5] All patients were also given tetanus prophylaxis when previous immunisation status was uncertain. Primary surgical care was provided as early as possible and the open injury was thoroughly debrided as per standard principles. After debridement, the method of fracture stabilisation and soft-tissue cover was decided by the operating surgeon based on the injury. Further surgical procedures were carried out as indicated.

The patients were regularly monitored for signs of surgical site infection; both during hospital stay and at outpatient follow-up, based on the CDC criteria.[6] Clinical signs of infection necessitated operative intervention during which deep intraoperative tissue/pus swab was sent to isolate the infecting micro-organism. Patients with clinical signs of infection, but negative deep intra-operative cultures were termed as culture-negative open fractures.

Culture technique

Tissue samples/pus swab from the infected site were sent immediately to the microbiology laboratory and processed as per the standard protocol. In case of pus swab, a minimum of two swabs were required in a sterile glass tube. One was used to obtain smear by Gram staining and second was used for inoculation on both aerobic and anaerobic media. For aerobic culture, bacterial inoculation was done on blood agar and MacConkey agar and the plates were incubated at 37°C overnight and anaerobic cultures were done with blood agar, thioglycollate and Robertson cooked meat broth incubated under anaerobic condition.[7] Tissue received in a sterile test tube was homogenised in a mortar and pestle with sterile normal saline. Both microscopy and culture plating were done as for pus swab. Cultures were followed up and growth of the organism was identified using preliminary biochemical screening media. Antibiotic susceptibility profile of the isolate was done by Kirby–Bauer disc diffusion method/E-test and susceptibility results were reported based on the Clinical and Laboratory Standards Institute interpretation.[8] Antibiotic susceptibility profile guided antimicrobial therapy in culture-positive patients, whereas culture-negative patients were treated for clinical signs of infection with broad-spectrum antibiotic therapy.

Data collection

Data were obtained prospectively from all study patients and entered in a pro forma which included demographic data, medical comorbidities, American Society of Anesthesiologists grade and mechanism of injury. Injury details collected included site of injury, Gustilo and Anderson grade, Injury Severity Score, time from injury to receiving prophylactic antibiotics and time from injury to index surgical procedure. Clinical outcomes measured included length of hospital stay, number of additional surgeries and occurrence of complications, namely, non-union and failure of limb salvage necessitating amputation after the index procedure. All patients were followed up for 9 months.

Statistical analysis

Quantitative variables were presented as mean and categorical variables by absolute (n) and relative (%) frequencies. The independent two-sample t-test was used to compare the outcome between culture-negative and culture-positive infected open fractures. P < 0.05 was considered statistically significant.


 ~ Results Top


Demographic data

A total of 231 patients with 275 open fractures involving the femur, tibia and/or fibula were included in our study. Included study patients were predominantly male (201 patients, 87%) and the mean age of our study group was 40 years (range: 17–77) [Table 1]. Road traffic accidents were found to be the most common cause of injury (219 patients, 95%). Forty-three patients (18.6%) had open type III fracture involving more than one long bone of the lower limb and tibia was most commonly involved bone (64%). Majority of them had Gustilo and Anderson Grade IIIB injury (65%). The average time from injury to receiving prophylactic antibiotics was 11.6 h (range: 1.4–122.5). The average time to index surgical procedure from injury was 21.9 h (range: 4.2–164). The average hospital stay was 15.7 days (range: 3–60) and 126 patients (54.5%) required additional surgical procedures following the index operation for various indications.
Table 1: Demographic data, injury details and clinical outcome of included study patients

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Infections in open fractures

There were clinical signs of infection as per the CDC criteria in 84 patients (36.4%) with 99 fractures.[6] Clinical signs of infection either during primary hospital admission or during follow-up care necessitated surgical intervention and deep intra-operative cultures in these 84 patients. Forty-three patients (51.2%) had positive cultures and remaining 41 patients had negative cultures (48.8%). The rate of culture-negative infection in open long bone fractures in our study was 17.7%. The demographic status of culture-positive and culture-negative patients with open type III fractures are enumerated in [Table 1].

Among culture-positive isolates, forty-nine isolates were monomicrobial and 13 isolates were polymicrobial infection. Most of the infections were caused by Gram-negative bacilli (55.8%) followed by Gram-positive cocci in 34.8% patients. The distribution of microbes identified in culture with their antibiotic resistance pattern is given in [Table 2].
Table 2: Isolated organisms in cases of deep and superficial infection and their susceptibility profile

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Comparative analysis of culture-positive versus culture-negative infections

The comparative analysis of culture-positive and culture-negative infected open fracture at 9 months is given in [Table 1]. The mean duration of hospital stay was marginally higher in the culture-positive group in comparison to the culture-negative group (24.8 vs. 20.44 days, P = 0.17), but this did not reach statistical significance. The mean number of additional procedures was also higher in the culture-positive group in comparison to the culture-negative group (1.93 vs. 1.44, P = 0.09), but again, this did not reach statistical significance. Further, there was no statistical significance (P = 1.0) between the two groups for complications such as non-unions and amputations. Our hypothesis that culture-negative patients with clinical signs of infection have a better outcome than culture-positive patients could not be proven.


 ~ Discussion Top


This is the first prospective cohort study in literature which compares the outcome between culture-negative and culture-positive infection in open fractures. Culture-negative infections in arthroplasty have been well described, but literature on culture-negative infections in orthopaedic trauma is limited to a single retrospective study not exclusively specific for open fractures.[9] The strength of our study is that we prospectively recruited patients and regularly followed them up with a fixed protocol for 9 months. Patients with clinical signs of infection often do not have positive intraoperative deep cultures. Therefore, we cannot rely on intraoperative cultures for diagnosis of deep surgical site infection.[10] Causative microorganisms were not found in 17.7% patients in our study. Failure to identify the infecting microorganism can lead to inappropriate treatment with a subsequent increase in treatment failure.[11]

The exact cause for culture-negative infections is not definitely clear. They could be due to sampling error, insufficient amount of isolate in the culture, comparatively milder form of infection where the pathogen load is less compared to culture-positive samples, poor transport conditions and inappropriate diagnostic tools for fastidious organisms that are difficult to identify using routine methods. Another common cause for culture-negative infection is the prior use of antibiotics which can compromise the sensitivity of routinely used diagnostic laboratory tests. Withholding antibiotic administration before culture is important to identify the causative organism. To increase the detection rate of low-virulence microorganisms, multiple samples (minimum 3) should be taken and importantly an adequate growth time of at least 7 days should be allowed in broth culture.[11]

Pus swab routinely used for collecting culture samples can only collect a small fraction of a millilitre of specimen (<100 μl), which greatly reduces the amount of bacteria that can be recovered for culture.[12] Another reason for sampling error could be use of a single swab for multiple types of cultures (aerobic, anaerobic and fungal) requiring inoculation of many different types of media which reduces yield of culture. Palmer et al. identified bacterial DNA in 88% of non-unions, while only 24% had positive intraoperative cultures.[13] Their study suggests that routine microbiology culture is not good enough for identifying all infections.

Kim et al.[14] compared the results of culture-negative and culture-positive infection in total knee arthroplasty and was unable to demonstrate any significant difference in clinical outcome between these two groups. In contrast, Choi et al.[15] compared culture-negative and culture-positive periprosthetic joint infection and found that culture-negative periprosthetic joint infections had a better clinical outcome. Culture-negative infections may predict good prognosis for the patients as the pathogen load is very less and should, therefore, be easily treatable. In contrast, culture-positive infections have an organism load of 103–1010/μl, thereby necessitating more intensive treatment.[16] On the other hand, inability to identify the infecting microorganism and lack of its antibiotic susceptibility may lead to treatment failure.[17] We found no significant difference in the outcome of culture-positive versus culture-negative patients in our group of patients with infected open type III fractures of the lower limb. As infection in open type III fractures is a serious cause of morbidity, it is important to treat culture-negative infections aggressively when clinical signs of infection are present, especially in this era of multidrug-resistant infections.[18]

The limitation of our study is that not all patients had more than one intraoperative deep tissue cultures to reduce sampling error. Further, molecular methods were not used for identification of pathogen which would have increased the yield of culture.


 ~ Conclusion Top


Failure to identify the infective microorganism in patients with clinical signs of infection is a frequently encountered situation which has not been adequately addressed in orthopaedic trauma literature. We found the rate ofculture-negative infections to be 17.7% in open type III long bone fractures of the lower limb. This study did not find any statistical difference in the clinical outcome between culture-negative and culture-positive infections in open fractures of the lower limb. We suggest that all patients with clinical signs of infection should be treated aggressively, even in the absence of bacterial growth in deep intraoperative specimens.

Acknowledgement

The study was approved and funded by the Institutional Review Board of Christian Medical College, Vellore (83-i/11/13) (IRB Minute No. 9693).

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.



 
 ~ References Top

1.
Tampe U, Widmer LW, Weiss RJ, Jansson KŠ. Mortality, risk factors and causes of death in Swedish patients with open tibial fractures – A nationwide study of 3, 777 patients. Scand J Trauma Resusc Emerg Med 2018;26:62.  Back to cited text no. 1
    
2.
Diwan A, Eberlin KR, Smith RM. The principles and practice of open fracture care, 2018. Chin J Traumatol 2018;21:187-92.  Back to cited text no. 2
    
3.
Court-Brown CM, Bugler KE, Clement ND, Duckworth AD, McQueen MM. The epidemiology of open fractures in adults. A 15-year review. Injury 2012;43:891-7.  Back to cited text no. 3
    
4.
Gitajn IL, Heng M, Weaver MJ, Ehrlichman LK, Harris MB. Culture-negative infection after operative fixation of fractures. J Orthop Trauma 2016;30:538-44.  Back to cited text no. 4
    
5.
Hoff WS, Bonadies JA, Cachecho R, Dorlac WC. East practice management guidelines work group: Update to practice management guidelines for prophylactic antibiotic use in open fractures. J Trauma 2011;70:751-4.  Back to cited text no. 5
    
6.
Garner JS, Jarvis WR, Emori TG, Horan TC, Hughes JM. CDC definitions for nosocomial infections, 1988. Am J Infect Control 1988;16:128-40.  Back to cited text no. 6
    
7.
Isenberg HD. Clinical Microbiology Procedures Handbook, Vol. 1, 2, 3. Washington, DC: American Society for Microbiology; 2010.  Back to cited text no. 7
    
8.
Clinical and Laboratory Standards Institute. Performance Standards for Antimicrobial Susceptibility Testing. CLSI Supplement M100. 29th ed. Wayne PA: Clinical and Laboratory Standards Institute; 2019.  Back to cited text no. 8
    
9.
Reisener M, Perka C. Do culture-negative periprosthetic joint infections have a worse outcome than culture-positive periprosthetic joint infections? A systematic review and meta-analysis. Biomed Res Int 2018;2018:6278012.  Back to cited text no. 9
    
10.
Kallstrom G. Are quantitative bacterial wound cultures useful? J Clin Microbiol 2014;52:2753-6.  Back to cited text no. 10
    
11.
Yoon HK, Cho SH, Lee DY, Kang BH, Lee SH, Moon DG. A review of the literature on culture-negative periprosthetic joint infection: Epidemiology, diagnosis and treatment. Knee Surg Relat Res 2017;29:155-64.  Back to cited text no. 11
    
12.
Phua J, Ngerng W, See K, Tay C, Kiong T, Lim H, et al. Characteristics and outcomes of culture-negative versus culture-positive severe sepsis. Crit Care 2013;17:R202.  Back to cited text no. 12
    
13.
Palmer MP, Melton-Kreft R, Nistico L, Hiller NL, Kim LH, Altman GT, et al. Polymerase chain reaction-electrospray-time-of-flight mass spectrometry versus culture for bacterial detection in septic arthritis and osteoarthritis. Genet Test Mol Biomarkers 2016;20:721-31.  Back to cited text no. 13
    
14.
Kim YH, Kulkarni SS, Park JW, Kim JS, Oh HK, Rastogi D. Comparison of infection control rates and clinical outcomes in culture-positive and culture-negative infected total-knee arthroplasty. J Orthop 2015;12:S37-43.  Back to cited text no. 14
    
15.
Choi HR, Kwon YM, Freiberg AA, Nelson SB, Malchau H. Periprosthetic joint infection with negative culture results: Clinical characteristics and treatment outcome. J Arthroplasty 2013;28:899-903.  Back to cited text no. 15
    
16.
Dark PM, Dean P, Warhurst G. Bench-to-bedside review: The promise of rapid infection diagnosis during sepsis using polymerase chain reaction-based pathogen detection. Crit Care 2009;13:217.  Back to cited text no. 16
    
17.
Veeraraghavan B, Pragasam AK, Bakthavatchalam YD, Anandan S, Ramasubramanian V, Swaminathan S. Newer β-lactam/β-lactamase inhibitor for multidrug-resistant gram-negative infections: Challenges, implications and surveillance strategy for India. Indian J Med Microbiol 2018;36:334-43.  Back to cited text no. 17
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18.
Pragasam AK, Veeraraghavan B, Nalini E, Anandan S, Kaye KS. An update on antimicrobial resistance and the role of newer antimicrobial agents for Pseudomonas aeruginosa. Indian J Med Microbiol 2018;36:303-16.  Back to cited text no. 18
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