Improving the sensitivity of microbiologic cultures aids in diagnosing, treating periprosthetic joint infection

Increased incubation duration decreases rate of false-negative microbiology.

by Benjamin M. Zmistowski; and Javad Parvizi, MD, FRCS

The complexity and devastation of periprosthetic joint infection following total knee and total hip arthroplasty is well known. When presented with suspected infection around a prosthetic joint, quick and accurate diagnosis is essential for effective treatment.

Many diagnostic criteria for periprosthetic joint infection (PJI) have been offered. These criteria utilize varying combinations of positive intraoperative and aspirate cultures, elevated serology — erythrocyte sedimentation rate (ESR) and C-reactive protein (CRP), elevated joint aspirate fluid cell counts — white blood cell (WBC) and polymorphonuclear neutrophil percentage (PMN%), histological analysis, gram stain, and presence of purulence and sinus tract.

Gold standard diagnostic

Javad Parvizi

Benjamin M. Zmistowski

While many diagnostic criteria combinations have proven accuracy, the gold standard in PJI diagnosis remains pathogen isolation from culture of periprosthetic tissue. An inability to provide a positive culture in spite of other positive signs of infection muddles the diagnosis. It is our experience that treatment of unknown pathogens is less successful in eradicating PJI. This experience may be due to a lack of guidance in selection of treatment and appropriate antimicrobials. False-negative results from culture occur in 7% to 40% of confirmed PJI.

Multiple factors may contribute to an inability to isolate an organism in a confirmed PJI. Those implicated include pre-emptive antimicrobial use, biofilm formation, atypical pathogens for which standard testing media are inadequate, and a lack of sufficient incubation length.

It has been the directive of many reports that antimicrobial therapy be withheld up to 4 weeks prior to planned culture, and intraoperative antimicrobial prophylaxis withheld until after collection of tissue samples. However, Ghanem and colleagues in 2007 reported that in cases with preoperative organism isolation from joint aspiration, the use of preoperative antibiotics does not preclude isolation of the same organism from intraoperative periprosthetic tissue.

Biofilm

Sessile bacteria may cause false-negative results as the biofilm formation around the prosthetic leaves no organisms to culture in typically collected periprosthetic tissue. Sonication of the resected prostheses has proved useful in providing samples of sessile bacteria that can be isolated using standard microbiological techniques, but this is not widely practiced. When defining infection regardless of culture results, Trampuz and colleagues reported in 2007 improved sensitivity and specificity of cultures of sonicate-fluid vs. standard tissue samples. Even though improved, their reported sensitivity of microbiologic analysis of sonicate-fluid remains low, 78.5%.

An inability to isolate an infecting pathogen may also be related to the culture medium. Fungi and mycobacteria are known periprosthetic joint pathogens and may be the source of lower culture sensitivity when these organisms remain untested. Many have proposed that a subset of periprosthetic tissue samples be stowed for later testing with atypical media if standard culture techniques do not isolate a pathogen in suspected PJI.

While false-negatives may be associated with atypical pathogens, eagerness to prematurely cease standard culture techniques lacking signs of growth may result in a false-negative outcome. While many elements limit the accuracy of microbiological culture, none may be as influential and easily remedied as inadequate incubation duration. Nonetheless, culture duration is rarely mentioned in research surrounding the accuracy of diagnostic criteria, specifically culture results, in the orthopedic literature. This lack of reporting is possibly from an orthopedist’s disregard of this necessary information or the lack of a relationship between the treating orthopedist and microbiologist. When mentioned, incubation periods typically range from 3 to 5 days for both aerobic and anaerobic cultures. Prolonged incubation periods, up to 2 weeks, have been investigated with meaningful results, according to studies by Schäfer, Neut and Ince and their colleagues.

Surface cultures

In a study of a new culturing technique, Neut’s team compared standard work-up of periprosthetic tissue or swab with an incubation period of 5 or 7 days. Following an incubation period of 5 days bacterial growth had a sensitivity of 41%, whereas after 7 days the sensitivity had risen to 64%. They went on to show even greater sensitivity (86%) with 7 days culturing of the prosthetic surface.

Culturing of the surface consisted of analyzing scrapings of all components. This technique resulted in a significant increase in the incidence of polymicrobial growth, including nonpathogens. These results demonstrate that increased incubation period and direct culturing of the prosthesis both independently improve sensitivity.

Schäfer and colleagues extensively studied an increased incubation period and its effects on sensitivity and organism profile. Organism isolation increased in an exponential manner from the first day to day 14. They defined infection as isolation of phenotypically identical organisms from two or more cultures or organism isolation in a single culture and positive histological analysis.

When using histological analysis as the gold standard, phenotypically identical isolates in two or greater cultures provided a sensitivity and specificity after a single week of 62.5% and 95%, respectively. After 2 weeks of incubation, sensitivity and specificity of the same criteria were 76.9% and 93.3%, respectively. A long-standing concern of increased incubation duration is an expected dramatic increase in growth of contaminants. Contrarily, the specificity of the test only mildly decreased. Furthermore, in regard to their definition of infection, 52% of contaminant strains grew in the first week. This is consistent with the lack of contaminants observed by Neut and colleagues when simply increasing the culture duration.

Time

The profile of organisms isolated considerably shifted as time increased. The majority of the following organisms were isolated within the first week: Staphylococcus aureus, coagulase-negative staphylococci, Enterococcus species, Streptococcus species, and Enterobacteriaceae. The remaining species — coryneform bacteria, Propionibacterium, Peptostreptococcus, and others — were isolated in the second week of incubation. These results indicate that an increase in incubation period past the first week may allow for identification of more nonstandard, fastidious microorganisms.

Length of incubation strongly affects success of pathogen isolation from an infected joint. Of course, cost and labor limitations play a role in an ability to extend incubation period and these limitations likely vary widely between institutions. Most importantly, a strong relationship with microbiologists in both research and practice will assist in maximizing the detection of PJI pathogens. In cases with a strong suspicion of PJI in which precautions against false-negative cultures have been followed and nonstandard media cultures initiated, it is recommended that an extended culture be performed.

References:

• Ghanem E, Parvizi J, Clohisy J, et al. Perioperative antibiotics should not be withheld in proven cases of periprosthetic infection. Clin Orthop Relat Res. 2007;461:44-47.
• Ince A, Rupp J, Frommelt L, et al. Is “aseptic” loosening of the prosthetic cup after total hip replacement due to nonculturable bacterial pathogens in patients with low-grade infection? Clin Infect Dis. 2004;39(11):1599-1603.
• Neut D, van Horn JR, van Kooten TG, van der Mei HC, Busscher HJ. Detection of biomaterial-associated infections in orthopaedic joint implants. Clin Orthop Relat Res. 2003;(413):261-268.
• Schäfer P, Fink B, Sandow D, et al. Prolonged bacterial culture to identify late periprosthetic joint infection: a promising strategy. Clin Infect Dis. 2008;47(11):1403-1409.
• Trampuz A, Piper KE, Jacobson MJ, et al. Sonication of removed hip and knee prostheses for diagnosis of infection. N Engl J Med. 2007;357(7):654-663.

• Javad Parvizi, MD, FRCSC, can be reached at Rothman Institute of Orthopaedics at Jefferson, 925 Chestnut St., 2nd Floor, Philadelphia PA 19107; 267-399-3617; e-mail: parvj@aol.com.