Orthopedics

Feature Article 

Leukocyte Esterase Strip Test: A Rapid and Reliable Method for the Diagnosis of Infections in Arthroplasty

Xiang Li, MD; Rui Li, MD; Ming Ni, MD; Wei Chai, MD; Libo Hao, MD; Yonggang Zhou, MD; Jiying Chen, MD

Abstract

The reliability of the leukocyte esterase (LE) strip test needs further investigation. A total of 204 joints suspected of having periprosthetic joint infection were enrolled from July 2014 to June 2016. One drop of synovial fluid extracted by joint aspiration was applied to LE strips. The results were recorded after 2 to 3 minutes according to the 5 color grades on the color chart (−neg, 25, 27, 250, and 500). A centrifuge was used before the LE strip test when the synovial fluid was mixed with blood. Based on the Musculoskeletal Infection Society criteria, 88 joints were diagnosed as being infected and 116 as not infected. On positive threshold comparison (500 vs 250), there was no statistically significant difference in sensitivity (92.0% vs 97.7%, P=.0736), but specificity was significantly increased in the 500 group (93.1% vs 80.2%, P=.0003). When only 500 was considered positive, the sensitivity, specificity, positive predictive value, and negative predictive value were 92.0% (95% confidence interval [CI], 83.8%–96.5%), 93.1% (95% CI, 86.4%–96.8%), 91.0% (95% CI, 82.6%–95.8%), and 93.9% (95% CI, 87.4%–97.3%), respectively. Compared with white blood cell count, the LE strip test had similar sensitivity (93.2% vs 92.0%, P=1) and specificity (92.2% vs 93.1%, P=1). In cases using the centrifugation method, sensitivity and specificity also remained favorable. In this study, using 500 rather than 250 as the positive threshold increased accuracy. Compared with white blood cell count and polymorphonuclear neutrophil percentage, the LE strip test has excellent sensitivity and specificity and is reliable. Synovial fluid centrifugation is an effective means of overcoming interference from erythrocytes. [Orthopedics. 2018; 41(2):e189–e193.]

Abstract

The reliability of the leukocyte esterase (LE) strip test needs further investigation. A total of 204 joints suspected of having periprosthetic joint infection were enrolled from July 2014 to June 2016. One drop of synovial fluid extracted by joint aspiration was applied to LE strips. The results were recorded after 2 to 3 minutes according to the 5 color grades on the color chart (−neg, 25, 27, 250, and 500). A centrifuge was used before the LE strip test when the synovial fluid was mixed with blood. Based on the Musculoskeletal Infection Society criteria, 88 joints were diagnosed as being infected and 116 as not infected. On positive threshold comparison (500 vs 250), there was no statistically significant difference in sensitivity (92.0% vs 97.7%, P=.0736), but specificity was significantly increased in the 500 group (93.1% vs 80.2%, P=.0003). When only 500 was considered positive, the sensitivity, specificity, positive predictive value, and negative predictive value were 92.0% (95% confidence interval [CI], 83.8%–96.5%), 93.1% (95% CI, 86.4%–96.8%), 91.0% (95% CI, 82.6%–95.8%), and 93.9% (95% CI, 87.4%–97.3%), respectively. Compared with white blood cell count, the LE strip test had similar sensitivity (93.2% vs 92.0%, P=1) and specificity (92.2% vs 93.1%, P=1). In cases using the centrifugation method, sensitivity and specificity also remained favorable. In this study, using 500 rather than 250 as the positive threshold increased accuracy. Compared with white blood cell count and polymorphonuclear neutrophil percentage, the LE strip test has excellent sensitivity and specificity and is reliable. Synovial fluid centrifugation is an effective means of overcoming interference from erythrocytes. [Orthopedics. 2018; 41(2):e189–e193.]

Periprosthetic joint infection (PJI) is a devastating complication of total joint arthroplasty. An estimated 1% to 3% of patients who undergo total joint arthroplasty subsequently develop a PJI.1–3 Because the number of primary and revision total joint arthroplasties is expected to increase,4 it is reasonable to believe that the number of PJIs will rise dramatically. Because many patients present with nonspecific manifestations, the diagnosis of PJI remains challenging for orthopedic surgeons. There is no single test with sufficient specificity and sensitivity to confirm or rule out PJI in atypical cases. Clinically, surgeons use several criteria involving multiple parameters and tests to diagnose PJI, including symptoms, serum examinations, histologic analysis, and synovial fluid test.5–7 However, the criteria used to diagnose PJI are complicated. The development of inexpensive, uncomplicated, and more accurate diagnostic tools for PJI remains a challenge.

Leukocyte esterase (LE) is an enzyme present within granulocytes and secreted by activated neutrophils that will be recruited to the infected site. It is normally used on a colorimetric reagent strip for the diagnosis of urinary system infections.8–10 Parvizi et al11 first illustrated that the LE strip could be highly accurate in identifying the presence of PJI in patients who previously underwent total knee arthroplasty. Subsequently, Wetters et al12 claimed that LE strips had a promising diagnostic capability for PJI in both the knee and the hip joints. Some organizations have regarded the LE strip test as a minor criterion equivalent to white blood cell (WBC) count in the diagnosis of PJI.7,13 However, the use of LE strips for diagnosing PJI remains controversial for several reasons. First, the positive threshold for the LE strip test was still in dispute in the latest PJI definition of the Musculoskeletal Infection Society (MSIS).13 Second, sufficient evidence supported by well-designed studies comparing diagnostic power between the LE strip test and synovial WBC count or polymorphonuclear neutrophil percentage (PMN%) is lacking. Finally, according to the literature and the current authors' experience, approximately one-third of LE strips cannot be read because the synovial fluid is mixed with blood.11,12 A technical note propounded by Aggarwal et al14 recommended that centrifugation be used in bloody joint aspiration cases because the accuracy of LE testing is not affected by centrifugation. However, because the samples in their study were thoroughly hand mixed with the patients' own blood, the results might not correlate directly with daily clinic work. Further validation is needed regarding the sensitivity and specificity of the LE strip test after centrifugation. The purposes of this study were (1) to discuss the optimistic threshold for the LE strip test and (2) to comprehensively investigate the power of LE strips, compared with synovial WBC count or PMN%, in the diagnosis of PJI. Moreover, this study also evaluated the reliability of using centrifugation for bloody joint aspirations.

Materials and Methods

The research ethics board of the authors' institution approved this study. From July 2014 to June 2016, the authors performed synovial aspirations on 231 consecutive patients (239 joints) suspected of having PJI. To be included in this study, patients had to have previous total joint replacement; a higher probability of infection (including abnormal erythrocyte sedimentation rate and/or C-reactive protein; a history of fevers, chills, pain, and loss of range of motion; or erythema at the incision site and swelling and warmth of the affected joint); and antibiotics paused for 2 weeks or more. Patients were excluded if total joint replacement had occurred within the past 3 months or they had been lost to follow-up or had insufficient information for MSIS infection criteria.6 Thirty-three patients (35 joints) were excluded, and 198 patients (204 joints) remained in the final cohort.

Because the modified MSIS criteria from 201413 listed the LE strip test as a minor criterion (elevated synovial fluid WBC count or “++ or +” change on LE test strip), the authors used 2011 criteria instead. If the accuracy of the LE strip test was evaluated using modified MSIS criteria that included the LE strip test, bias would exist in the results. Moreover, more evidence is needed to support that the LE strip test is comparable to synovial WBC count in clinical practice.

Aspirations were performed in the outpatient operating room. Strict sterile manipulation was emphasized to avoid contamination. One drop of synovial fluid sample was applied to the LE strip test pad (AUTION Sticks 10 PA; ARKRAY, Kyoto, Japan) immediately after aspiration. After 2 to 3 minutes, the pad will have a color that indicates the concentration of esterase. There were 5 color grades on the color chart (−neg, 25, 27, 250, and 500) (Figure 1). The grades of color change on the LE strip test pads were recorded according to the chart by 3 independent observers who were blinded to diagnostic information about the patients. If inconsistent records existed among the observers, the majority would be considered as the final results by another independent analyst (M.N.).

The red rectangle marks the interpretation of the results of the leukocyte esterase strip test. The color chart had 5 color grades (−neg, 25, 27, 250, and 500). The results of the leukocyte esterase strip test were recorded based on matching the color change with the grade on the color chart.

Figure 1:

The red rectangle marks the interpretation of the results of the leukocyte esterase strip test. The color chart had 5 color grades (−neg, 25, 27, 250, and 500). The results of the leukocyte esterase strip test were recorded based on matching the color change with the grade on the color chart.

A mini centrifuge (D1008E; SCILO-GEX, Rocky Hill, Connecticut) was used to wipe out the disturbance of the erythrocytes when heavily blood-stained synovial fluid made the color change on the LE strip pad unreadable.

The remaining synovial fluid obtained was sent to the clinical laboratory for WBC count, PMN%, and pathogen cultures.

Statistical Analysis

Musculoskeletal Infection Society criteria were used to identify PJI. Sensitivity, specificity, positive predictive value, and negative predictive value were calculated for the results of the LE strip test. The accuracy of different methods was analyzed with the 3-dimensional paired chi-square test. P<.05 was considered statistically significant.

Results

Of the 198 patients (204 joints) examined, 87 patients (88 joints, 54 knees, and 34 hips) were classified as infected. There were 39 males and 48 females in this cohort with a mean age and a mean body mass index of 62.94±14.25 years and 25.16±4.10 kg/m2, respectively. The other 111 patients (116 joints, 71 knees, and 45 hips) were classified as not infected. There were 42 males and 69 females in this cohort with a mean age and a mean body mass index of 62.36±9.30 years and 25.76±4.13 kg/m2, respectively.

If the color change on the LE strip test pad matched the final grade (dark purple, 500), the results were considered positive. The sensitivity, specificity, positive predictive value, and negative predictive value were 92.0% (95% confidence interval [CI], 83.8%–96.5%), 93.1% (95% CI, 86.4%–96.8%), 91.0% (95% CI, 82.6%–95.8%), and 93.9% (95% CI, 87.4%–97.3%), respectively (Table 1). If the last 2 grades of the color chart (500 and 250) were both considered as positive standards, the sensitivity, specificity, positive predictive value, and negative predictive value were 97.7% (95% CI, 91.3%–99.6%), 80.2% (95% CI, 71.5%–86.8%), 78.9% (95% CI, 69.8%–85.9%), and 97.9% (95% CI, 91.9%–99.6%), respectively.

Sensitivity, Specificity, Positive Predictive Value, and Negative Predictive Value of the Leukocyte Esterase Strip and Traditional Synovial Fluid Tests

Table 1:

Sensitivity, Specificity, Positive Predictive Value, and Negative Predictive Value of the Leukocyte Esterase Strip and Traditional Synovial Fluid Tests

Analysis showed that the specificity was significantly increased, but not statistically significantly decreased, when 500 was considered the positive standard (Table 2). Thus, the authors believe that 500 as a positive result seems more appropriate for this analysis.

Results of the 3-Dimensional Paired Chi-square Test for the Comparison of Each Pair of Methods

Table 2:

Results of the 3-Dimensional Paired Chi-square Test for the Comparison of Each Pair of Methods

The sensitivity, specificity, positive predictive value, and negative predictive value of traditional synovial fluid tests including WBC count and PMN% were also evaluated (Table 1). Their accuracy was compared with that of the LE strip test (Table 2).

In this cohort study, 69 samples mixed with blood were centrifuged to avoid interference from erythrocytes (Figure 2). The sensitivity, specificity, positive predictive value, and negative predictive value of these samples were also analyzed and compared with those of other samples (Table 1). No statistically significant differences were observed between the centrifugation and non-centrifugation groups.

Blood-stained synovial fluid aspirated from a joint, which could not be used for the leukocyte esterase (LE) strip test directly (A). Erythrocytes were separated by centrifugation. Supernatant was left for the LE strip test (B). After 2 to 3 minutes, the LE strip test pad (arrow) was dark purple (500), indicating a positive result (C).

Figure 2:

Blood-stained synovial fluid aspirated from a joint, which could not be used for the leukocyte esterase (LE) strip test directly (A). Erythrocytes were separated by centrifugation. Supernatant was left for the LE strip test (B). After 2 to 3 minutes, the LE strip test pad (arrow) was dark purple (500), indicating a positive result (C).

Discussion

Periprosthetic joint infection remains a devastating complication associated with greater mortality, morbidity, and economic costs.3 Currently, the diagnosis of PJI is based on a combination of clinical symptoms, serological tests, pathogen culture results, and histological examinations; however, it is to some extent imperfect. Novel tools for the timely and accurate diagnosis of PJI need to be introduced and evaluated.

The LE strip test is a newer method for diagnosing PJI. Parvizi et al11 first adopted the use of LE strips in the diagnosis of PJI and determined their sensitivity and specificity. They evaluated 2 standards indicating positive LE strip test results. When “++” (the darkest grade of the color chart) was considered positive, the LE strip test was 80.6% sensitive and 100% specific. When “++” and “+” were both considered positive, the LE strip test was 93.5% sensitive and 86.7% specific. Thereafter, some authors considered “++” a positive result,15 whereas others believed that “++” and “+” should both be considered positive.16,17 Indeed, the positive threshold for the LE strip test was still in dispute in the latest PJI definition of the MSIS. Therefore, 500 (dark purple) and 250 (light purple) were both evaluated in the current study. According to the results, considering 500 alone or 500 and 250 together as positive results led to no statistically significant difference in sensitivity (P=.0736). Specificity was significantly increased in the 500 group (P=.0003). Thus, the authors believed that considering 500 a positive result, equivalent to “++” in other studies, was more appropriate.

In this study, 8 false-positive cases were encountered when 500 was considered a positive result. Among them, 3 patients had been diagnosed with rheumatoid arthritis, 1 patient had been diagnosed with spondyloarthropathies, and 1 patient had been diagnosed with ankylosing spondylitis by a physician from the rheumatology department. The 3 remaining cases had no further diagnosis. This may indicate that autoimmune disease had some effect on the results of the LE strip test and will lead to a false-positive result. Because the authors did not systematically screen all patients with autoimmune diseases, more research is needed to confirm their hypothesis.

On the basis of this study, the LE strip test had a high consistency with synovial WBC count (P=1). This supported the second minor criterion (elevated synovial fluid WBC count or “++” change on the LE strip test) listed in the modified MSIS criteria.13

Eliminating the interference generated by blood mixed in the synovial fluid remains a major issue with the LE strip test. The red color on the LE strip test pad caused by erythrocytes could make the results unreadable. In a study conducted by Parvizi et al,11 47 (26.6%) of 177 samples were unreadable because of blood mixing. Wetters et al12 reported that 29.2% of samples was unreadable for the same reason. To resolve this issue, researchers14 have used centrifugation to eliminate erythrocytes before the LE strip test. However, the influence of centrifugation on the accuracy of the LE strip test continues to be discussed. In this study, 69 samples mixed with blood were centrifuged to avoid interference from erythrocytes. Most results became clearly readable after centrifugation; however, 2 samples remained light red and 1 sample remained yellow, which slightly interfered with the authors' judgment of the results. Because the colors of these 3 cases were far from being dark purple, they were finally classified as having negative results on the LE strip test; none of them was classified as having PJI with the MSIS criterion. The authors hypothesized that the color might have been caused by hemoglobin or hemosiderin, which were released by destroyed erythrocytes and thus could not be separated out by centrifugation. All 3 cases had a synovial hyperemia for a long time and repeated aspiration of bloody synovial fluid, so the erythrocytes mixed in the synovial fluid might have been destroyed. Regarding all of the samples that underwent centrifugation, the sensitivity and specificity remained reliable.

As a diagnostic tool for PJI, the LE strip test is advantageous because it only requires 1 drop of synovial fluid from aspiration. A small amount of synovial fluid is usually inadequate for pathogen culture and other laboratory tests. Other advantages of the LE strip test include its speed and low cost.

This study had some limitations that must be considered. First, the 2011 version of the MSIS criteria, which is somewhat outdated, was adopted as the gold standard. However, using the modified version including the LE strip test would have led to bias in this study. Second, the LE strip test yields qualitative results based on naked-eye observations. Although the authors adopted the principle that the minority was subordinate to the majority if disagreement existed between observers, subjectivity remained. Regarding the almost distinguishable border-stain judgment, quantitative results from special equipment could be more reliable, but the speed and low cost of the LE strip test would be sacrificed. Third, the LE strip test may not be suitable for patients with comorbid systemic inflammatory disease. The effectiveness of the LE strip test has yet to be confirmed in these patients. Finally, concerns remain regarding the blood-mixed synovial fluid after centrifugation. In this study, although sensitivity and specificity were favorable in the centrifugation group, the evaluation was difficult. A self-control study evaluating the same synovial sample before and after centrifugation is needed.

Conclusion

On the basis of this study, using 500 rather than 250 as the positive threshold is recommended due to increased accuracy. Compared with WBC count and PMN%, the LE strip test has excellent sensitivity and specificity and is reliable. Synovial fluid centrifugation is an effective means of overcoming interference from erythrocytes. The authors consider the LE strip test a quick, low-cost method that offers satisfactory accuracy for diagnosing PJI.

References

  1. Phillips JE, Crane TP, Noy M, Elliott TS, Grimer RJ. The incidence of deep prosthetic infections in a specialist orthopaedic hospital: a 15-year prospective survey. J Bone Joint Surg Br. 2006; 88(7):943–948. doi:10.1302/0301-620X.88B7.17150 [CrossRef]
  2. Jamsen E, Varonen M, Huhtala H, et al. Incidence of prosthetic joint infections after primary knee arthroplasty. J Arthroplasty. 2010; 25(1):87–92. doi:10.1016/j.arth.2008.10.013 [CrossRef]
  3. Kurtz SM, Lau E, Watson H, Schmier JK, Parvizi J. Economic burden of periprosthetic joint infection in the United States. J Arthroplasty. 2012; 27(8)(suppl):61–65. doi:10.1016/j.arth.2012.02.022 [CrossRef]
  4. Patel A, Pavlou G, Mújica-Mota RE, Toms AD. The epidemiology of revision total knee and hip arthroplasty in England and Wales: a comparative analysis with projections for the United States. A study using the National Joint Registry dataset. Bone Joint J. 2015; 97-B(8):1076–1081. doi:10.1302/0301-620X.97B8.35170 [CrossRef]
  5. Della Valle C, Parvizi J, Bauer TW, et al. American Academy of Orthopaedic Surgeons. Diagnosis of periprosthetic joint infections of the hip and knee. J Am Acad Orthop Surg. 2010; 18(12):760–770. doi:10.5435/00124635-201012000-00006 [CrossRef]
  6. Parvizi J, Zmistowski B, Berbari EF, et al. New definition for periprosthetic joint infection: from the Workgroup of the Musculoskeletal Infection Society. Clin Orthop Relat Res. 2011; 469(11):2992–2994. doi:10.1007/s11999-011-2102-9 [CrossRef]
  7. Parvizi J, Gehrke T, Chen AF. Proceedings of the International Consensus on Periprosthetic Joint Infection. Bone Joint J. 2013; 95-B(11):1450–1452. doi:10.1302/0301-620X.95B11.33135 [CrossRef]
  8. Perry JL, Matthews JS, Weesner DE. Evaluation of leukocyte esterase activity as a rapid screening technique for bacteriuria. J Clin Microbiol. 1982; 15(5):852–854.
  9. Smalley DL, Dittmann AN. Use of leukocyte esterase-nitrate activity as predictive assays of significant bacteriuria. J Clin Microbiol. 1983; 18(5):1256–1257.
  10. Devillé WL, Yzermans JC, van Duijn NP, Bezemer PD, van der Windt DA, Bouter LM. The urine dipstick test useful to rule out infections: a meta-analysis of the accuracy. BMC Urol. 2004; 4:4. doi:10.1186/1471-2490-4-4 [CrossRef]
  11. Parvizi J, Jacovides C, Antoci V, Ghanem E. Diagnosis of periprosthetic joint infection: the utility of a simple yet unappreciated enzyme. J Bone Joint Surg Am. 2011; 93(24):2242–2248. doi:10.2106/JBJS.J.01413 [CrossRef]
  12. Wetters NG, Berend KR, Lombardi AV, Morris MJ, Tucker TL, Della Valle CJ. Leukocyte esterase reagent strips for the rapid diagnosis of periprosthetic joint infection. J Arthroplasty. 2012; 27(8)(suppl):8–11. doi:10.1016/j.arth.2012.03.037 [CrossRef]
  13. Parvizi J, Gehrke TInternational Consensus Group on Periprosthetic Joint Infection. Definition of periprosthetic joint infection. J Arthroplasty. 2014; 29(7):1331. doi:10.1016/j.arth.2014.03.009 [CrossRef]
  14. Aggarwal VK, Tischler E, Ghanem E, Parvizi J. Leukocyte esterase from synovial fluid aspirate: a technical note. J Arthroplasty. 2013; 28(1):193–195. doi:10.1016/j.arth.2012.06.023 [CrossRef]
  15. Colvin OC, Kransdorf MJ, Roberts CC, et al. Leukocyte esterase analysis in the diagnosis of joint infection: can we make a diagnosis using a simple urine dipstick?Skeletal Radiol. 2015; 44(5):673–677. doi:10.1007/s00256-015-2097-5 [CrossRef]
  16. Shafafy R, McClatchie W, Chettiar K, et al. Use of leucocyte esterase reagent strips in the diagnosis or exclusion of prosthetic joint infection. Bone Joint J. 2015; 97-B(9):1232–1236. doi:10.1302/0301-620X.97B9.34910 [CrossRef]
  17. Guenther D, Kokenge T, Jacobs O, et al. Excluding infections in arthroplasty using leucocyte esterase test. Int Orthop. 2014; 38(11):2385–2390. doi:10.1007/s00264-014-2449-0 [CrossRef]

Sensitivity, Specificity, Positive Predictive Value, and Negative Predictive Value of the Leukocyte Esterase Strip and Traditional Synovial Fluid Tests

ItemSensitivityaSpecificityaPPVaNPVa
LE strip test (500)b92.0% (83.8%–96.5%)93.1% (86.4%–96.8%)91.0% (82.6%–95.8%)93.9% (87.4%–97.3%)
LE strip test (250)c97.7% (91.3%–99.6%)80.2% (71.5%–86.8%)78.9% (69.8%–85.9%)97.9% (91.9%–99.6%)
Synovial WBC countb93.2% (85.2%–97.2%)92.2% (85.4%–96.2%)90.1% (81.6%–95.1%)94.7% (88.3%–97.8%)
PMN%b94.3% (86.6%–97.9%)87.9% (80.3%–93.0%)85.6% (76.6%–91.6%)95.3% (88.9%–98.3%)
Centrifugation groupb90.3% (73.1%–97.5%)94.7% (80.9%–99.1%)93.3% (76.5%–98.8%)92.3% (78.0%–98.0%)
Non-centrifugation groupb92.9% (81.9%–97.7%)92.3% (83.4%–96.8%)89.7% (78.2%–95.7%)94.7% (86.4%–98.3%)

Results of the 3-Dimensional Paired Chi-square Test for the Comparison of Each Pair of Methods

ItemSensitivitySpecificity


Chi-square TestPChi-square TestP
LE (500)a vs LE (250)b3.20.073613.07.0003c
LE (500)a vs synovial WBC counta0.0010.001
LE (500)a vs PMN%a0.17.68011.79.1809
Authors

The authors are from the Department of Orthopedics, Chinese PLA General Hospital, Beijing, China.

Drs Xiang Li and Rui Li contributed equally to this work and should be considered as equal first authors.

The authors have no relevant financial relationships to disclose.

Correspondence should be addressed to: Jiying Chen, MD, Department of Orthopedics, Chinese PLA General Hospital, 28 Fuxing Rd, Beijing, China 100853 ( chenjiying_301@163.com).

Received: June 02, 2017
Accepted: November 13, 2017
Posted Online: January 09, 2018

10.3928/01477447-20180102-03

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