Practical Guidance for Clinical Microbiology Laboratories: Viruses Causing Acute Respiratory Tract Infections.

Respiratory viral infections are associated with a wide range of acute syndromes and infectious disease processes in children and adults worldwide. Many viruses are implicated in these infections, and these viruses are spread largely via respiratory means between humans but also occasionally from animals to humans. This article is an American Society for Microbiology (ASM)-sponsored Practical Guidance for Clinical Microbiology (PGCM) document identifying best practices for diagnosis and characterization of viruses that cause acute respiratory infections and replaces the most recent prior version of the ASM-sponsored Cumitech 21 document, Laboratory Diagnosis of Viral Respiratory Disease, published in 1986. The scope of the original document was quite broad, with an emphasis on clinical diagnosis of a wide variety of infectious agents and laboratory focus on antigen detection and viral culture. The new PGCM document is designed to be used by laboratorians in a wide variety of diagnostic and public health microbiology/virology laboratory settings worldwide. The article provides guidance to a rapidly changing field of diagnostics and outlines the epidemiology and clinical impact of acute respiratory viral infections, including preferred methods of specimen collection and current methods for diagnosis and characterization of viral pathogens causing acute respiratory tract infections. Compared to the case in 1986, molecular techniques are now the preferred diagnostic approaches for the detection of acute respiratory viruses, and they allow for automation, high-throughput workflows, and near-patient testing. These changes require quality assurance programs to prevent laboratory contamination as well as strong preanalytical screening approaches to utilize laboratory resources appropriately. Appropriate guidance from laboratorians to stakeholders will allow for appropriate specimen collection, as well as correct test ordering that will quickly identify highly transmissible emerging pathogens.

Performance of Treponemal Tests for the Diagnosis of Syphilis.

BACKGROUND: Treponemal immunoassays are increasingly used for syphilis screening with the reverse sequence algorithm. There are few data describing performance of treponemal immunoassays compared to traditional treponemal tests in patients with and without syphilis. METHODS: We calculated sensitivity and specificity of 7 treponemal assays: (1) ADVIA Centaur (chemiluminescence immunoassay [CIA]); (2) Bioplex 2200 (microbead immunoassay); (3) fluorescent treponemal antibody absorption test (FTA-ABS); (4) INNO-LIA (line immunoassay); (5) LIAISON CIA; (6) Treponema pallidum particle agglutination assay (TPPA); and (7) Trep-Sure (enzyme immunoassay [EIA]), using a reference standard combining clinical diagnosis and serology results. Sera were collected between May 2012-January 2013. Cases were characterized as: (1) current clinical diagnosis of syphilis: primary, secondary, early latent, late latent; (2) prior treated syphilis only; (3) no evidence of current syphilis, no prior history of syphilis, and at least 4 of 7 treponemal tests negative. RESULTS: Among 959 participants, 262 had current syphilis, 294 had prior syphilis, and 403 did not have syphilis. FTA-ABS was less sensitive for primary syphilis (78.2%) than the immunoassays or TPPA (94.5%-96.4%) (all P ≤ .01). All immunoassays were 100% sensitive for secondary syphilis, 95.2%-100% sensitive for early latent disease, and 86.8%-98.5% sensitive in late latent disease. TPPA had 100% specificity. CONCLUSIONS: Treponemal immunoassays demonstrated excellent sensitivity for secondary, early latent, and seropositive primary syphilis. Sensitivity of FTA-ABS in primary syphilis was poor. Given its high specificity and superior sensitivity, TPPA is preferred to adjudicate discordant results with the reverse sequence algorithm over the FTA-ABS.

Correlation of Treponemal Immunoassay Signal Strength Values with Reactivity of Confirmatory Treponemal Testing.

Automated treponemal immunoassays are used for syphilis screening with the reverse-sequence algorithm; discordant results (e.g., enzyme immunoassay [EIA] reactive and reactive plasma reagin [RPR] nonreactive) are resolved with a second treponemal test. We conducted a study to determine automated immunoassay signal strength values consistently correlating with reactive confirmatory treponemal testing. We conducted a cross-sectional analysis of four automated immunoassays (BioPlex 2200 microbead immunoassay [MBIA], Liaison chemiluminescence immunoassay [CIA], Advia-Centaur CIA, and Trep-Sure EIA) and three manual assays (Treponema pallidum particle agglutination [TP-PA], fluorescent treponemal antibody absorption [FTA-ABS] test, and Inno-LIA line immunoassay). We compared signal strength values of automated immunoassays and positive and negative agreement. Among 1,995 specimens, 908 (45.5%) were true positives (≥4/7 tests reactive) and 1,087 (54.5%) were true negatives (≥4/7 tests nonreactive). Positive agreement ranged from 86.1% (83.7 to 88.2%) for FTA-ABS to 99.7% (99.0 to 99.9%) for Advia-Centaur CIA; negative agreement ranged from 86.3% (84.1 to 88.2%) for Trep-Sure EIA to 100% for TP-PA (99.6 to 100%). Increasing signal strength values correlated with increasing reactivity of confirmatory testing (P < 0.0001 for all automated immunoassays by Cochran-Armitage test for trend). All automated immunoassays had signal strength cutoffs corresponding to ≥4/7 reactive treponemal tests. BioPlex MBIA and Liaison CIA had signal strength cutoffs correlating with ≥99% and 100% TP-PA reactivity, respectively. The Advia-Centaur CIA and Trep-Sure EIA had signal strength cutoffs correlating with at least 95% TP-PA reactivity. All automated immunoassays had signal strength cutoffs correlating with at least 95% FTA-ABS reactivity. Assuming that a 95% level of confirmation is adequate, these signal strength values can be used in lieu of confirmatory testing with TP-PA and FTA-ABS.

The role of the clinical laboratory in the future of health care: lean microbiology.

This commentary will introduce lean concepts into the clinical microbiology laboratory. The practice of lean in the clinical microbiology laboratory can remove waste, increase efficiency, and reduce costs. Lean, Six Sigma, and other such management initiatives are useful tools and can provide dividends but must be accompanied by organizational leadership commitment to sustaining the lean culture in the laboratory setting and providing resources and time to work through the process.

Laboratory diagnosis of Clostridium difficile infections: there is light at the end of the colon.

Single molecular or multistep assays (glutamate dehydrogenase, toxin A/B, ± molecular) are recommended for the diagnosis of CDI in patients with clinically significant diarrhea. Rapid and accurate tests can improve resource allocations and improve patient care. Enzyme immunoassay (EIA) for toxins A/B is too insensitive for use as a stand-alone assay. This guideline will examine the use of molecular tests and multitest algorithms for the diagnosis of Clostridium difficile infection (CDI). These new tests, alone or in a multistep algorithm consisting of >1 assay, are more expensive than the older EIA assays; however, rapid and accurate testing can save money overall by initiating appropriate treatment and infection control protocols sooner and by possibly reducing length of hospital stay. We recommend testing only unformed stool in patients with clinically significant diarrhea by a molecular method or by a 2- to 3-step algorithm.

Clostridium difficile ribotype diversity at six health care institutions in the United States.

Capillary-based PCR ribotyping was used to quantify the presence/absence and relative abundance of 98 Clostridium difficile ribotypes from clinical cases of disease at health care institutions in six states of the United States. Regionally important ribotypes were identified, and institutions in close proximity did not necessarily share more ribotype diversity than institutions that were farther apart.

Impact of a Rapid Respiratory Pathogen Panel on Antibiotic and Chest Radiography Usage and Hospital Length of Stay in the Pediatric Inpatient Setting.

IntroductionThe objective of this study was to compare the antibiotic initiation rate and duration, hospital length of stay, emergency department (ED) admission rate and chest radiography usage in a pediatric inpatient unit before and after the decentralization of rapid respiratory pathogen panel (RRPP) processing. MethodsThis retrospective cohort study examined antibiotic initiation rates and durations, hospital lengths of stay, ED admission rates, and chest radiography usage from 2 respiratory virus seasons. For the 2014 cohort, RRPPs were processed at a centralized laboratory, and result times averaged 26 hours, whereas for the 2015 cohort, RRPPs were processed on-site with result times averaging 2 hours. Demographic data were collected and demonstrated similar populations. Chi-square testing was used to detect the change of antibiotic initiation rates, ED admission rates, and chest radiography usage after on-site RRPP processing was introduced. Antibiotic duration and hospital length of stay were determined by Wilcoxon rank sum. ResultsThe study population included 94 patients from the 2014 respiratory virus season and 108 patients from the 2015 respiratory virus season. There were no statistically significant differences in gender, ethnicity, or age between the 2 cohorts. Antimicrobial initiation rates during hospital stay decreased from 46% to 27% (p = 0.005). The rate of admission from the ED decreased from 75% to 30% (p < 0.001). There were no statistically significant differences in antibiotic duration, hospital length of stay, or chest radiography usage. ConclusionRapid respiratory pathogen testing is a useful tool that can decrease unnecessary antibiotic initiation and hospital admissions in the pediatric population.

Evaluation of the Cepheid Xpert MTB/RIF assay for direct detection of Mycobacterium tuberculosis complex in respiratory specimens.

A total of 217 specimens submitted for routine smear and culture from three different sites within the western United States were used to evaluate the GeneXpert MTB/RIF assay (for research use only) (Cepheid, Sunnyvale, CA). Overall agreement compared to culture was 89% (98% for smear positives and 72% for smear negatives) for detection of Mycobacterium tuberculosis.

Comparison of premier CAMPY enzyme immunoassay (EIA), ProSpecT Campylobacter EIA, and ImmunoCard STAT! CAMPY tests with culture for laboratory diagnosis of Campylobacter enteric infections.

Campylobacter enteritis is a food-borne or waterborne illness caused almost exclusively by Campylobacter jejuni and, to a lesser extent, by Campylobacter coli. These organisms produce indistinguishable clinical diseases and together represent the second most common cause of bacterial diarrhea in the United States and the leading cause of enteric infection throughout the world. The conventional approach to the laboratory diagnosis of Campylobacter enteritis is based on the recovery of the organism from a stool specimen, which requires the use of a specialized medium incubated at 42°C for several days in an artificially created microaerophilic environment. Recently, several commercially available enzyme immunoassays (EIAs) have been developed for the direct detection of C. jejuni and C. coli in stool specimens. This study compared conventional culture with three EIA methods, the Premier CAMPY EIA (Meridian Bioscience, Cincinnati, OH), the ProSpecT Campylobacter EIA (Remel, Lenexa, KS), and the ImmunoCard STAT! CAMPY test (Meridian Bioscience, Cincinnati, OH), for the detection of C. jejuni and C. coli in 485 patient stool samples. Discordant results were arbitrated by using an in-house, real-time PCR assay that was developed and validated by a public health reference laboratory. Following analyses of the discrepant specimens by PCR, the sensitivity and specificity of both the Premier CAMPY and ProSpecT Campylobacter EIAs were 99.3% and 98%, respectively, while the ImmunoCard STAT! CAMPY test had a sensitivity of 98.5% and a specificity of 98.2%. By use of the PCR test as the reference standard, culture detected 127 of 135 Campylobacter-positive stool specimens, yielding a sensitivity of 94.1%. These results showed that the three EIAs evaluated in this study provide a rapid and reliable alternative for the laboratory diagnosis of enteric infections with C. jejuni and C. coli and that conventional culture may no longer be recognized as the "gold standard" for diagnosis.

Clostridium difficile testing in the clinical laboratory by use of multiple testing algorithms.

The incidence of Clostridium difficile infection (CDI) has risen almost 3-fold in the United States over the past decade, emphasizing the need for rapid and accurate tests for CDI. The Cepheid Xpert C. difficile assay is an integrated, closed, nucleic acid amplification system that automates sample preparation and real-time PCR detection of the toxin B gene (tcdB). A total of 432 stool specimens from symptomatic patients were tested by a glutamate dehydrogenase (GDH) assay, a toxin A and B enzyme immunoassay (EIA), the Xpert C. difficile assay, and a cell culture cytotoxicity neutralization assay (CCCN). The results of these methods, used individually and in combination, were compared to those of toxigenic culture. Results for the Xpert C. difficile assay alone showed a sensitivity, specificity, positive predictive value, and negative predictive value (NPV) of 94.4, 96.3, 84.0, and 98.8%, while the EIA alone gave corresponding values of 58.3, 94.7, 68.9, and 91.9%, respectively. An algorithm using the GDH assay and the EIA (plus the CCCN if the EIA was negative) showed corresponding values of 83.1, 96.7, 83.1, and 96.1%. The Xpert C. difficile assay was statistically superior to the EIA (P, <0.001 by Fisher's exact test) and to the GDH-EIA-CCCN algorithm (P, 0.0363). Combining the GDH and Xpert C. difficile assays lowered both the sensitivity and the NPV of the Xpert assay. The GDH-EIA-CCCN procedure required, on average, 2 days to complete testing on GDH-positive results, while testing by the Xpert C. difficile assay was completed, on average, in less than 1 h. Xpert C. difficile testing yielded the highest sensitivity and NPV, in the least amount of time, of the individual- and multiple-test algorithms evaluated in this study.

Multicenter evaluation of a new shortened peptide nucleic acid fluorescence in situ hybridization procedure for species identification of select Gram-negative bacilli from blood cultures.

A shortened protocol for two peptide nucleic acid fluorescence in situ hybridization (PNA FISH) assays for the detection of Gram-negative bacilli from positive blood cultures was evaluated in a multicenter trial. There was 100% concordance between the two protocols for each assay (368 of 368 and 370 of 370 results) and 99.7% (367 of 368 and 369 of 370 results) agreement with routine laboratory techniques.

Impact of strain type on detection of toxigenic Clostridium difficile: comparison of molecular diagnostic and enzyme immunoassay approaches.

A multicenter clinical trial assessed the performance of the Cepheid Xpert C. difficile assay on stool specimens collected from patients suspected of having Clostridium difficile infection (CDI). A total of 2,296 unformed stool specimens, collected from seven study sites, were tested by Xpert C. difficile enrichment culture followed by cell culture cytotoxicity testing of the isolates (i.e., toxigenic culture with enrichment) and the study sites' standard C. difficile test methods. The methods included enzyme immunoassay (EIA), direct cytotoxin testing, and two- and three-step algorithms using glutamate dehydrogenase (GDH) screening followed by either EIA or EIA and an in-house PCR assay. All C. difficile strains were typed by PCR-ribotyping. Compared to results for toxigenic culture with enrichment, the sensitivity, specificity, and positive and negative predictive values of the Xpert assay were 93.5, 94.0, 73.0, and 98.8%, respectively. The overall sensitivity of the EIAs compared to that of enrichment culture was 60.0%, and the sensitivity of combined GDH algorithms was 72.9%; both were significantly lower than that of Xpert C. difficile (P < 0.001 and P = 0.03, respectively). The sensitivity of the EIA was significantly lower than that of the Xpert C. difficile assay for detection of ribotypes 002, 027, and 106 (P < 0.0001, P < 0.0001, and P = 0.004, respectively, Fisher's exact test), and the sensitivity of GDH algorithms for ribotypes other than 027 was lower than that for Xpert C. difficile (P < 0.001). The Xpert C. difficile assay is a simple, rapid, and accurate method for detection of toxigenic C. difficile in unformed stool specimens and is minimally affected by strain type compared to EIA and GDH-based methods.

Trimethoprim in vitro antibacterial activity is not increased by adding sulfamethoxazole for pediatric Escherichia coli urinary tract infection.

PURPOSE: The combination of trimethoprim/sulfamethoxazole is often used to treat uncomplicated urinary tract infections in children. The rationale for combining trimethoprim and sulfamethoxazole is that they may act synergistically to increase antibacterial activity. However, approximately 3% of patients show allergic reactions to sulfamethoxazole, of which some are serious (liver failure and Stevens-Johnson syndrome). We determined whether adding sulfamethoxazole is necessary to increase in vitro antibacterial activity for pediatric urinary tract infection compared to that of trimethoprim alone. MATERIALS AND METHODS: We prospectively identified 1,298 children with urinary tract infection (greater than 100,000 cfu/ml Escherichia coli) from a total of 4 American regions. In vitro susceptibility of bacterial isolates to sulfamethoxazole, trimethoprim and trimethoprim/sulfamethoxazole was determined using disk diffusion. Ampicillin susceptibility was tested at 2 sites. At 1 site all uropathogens from consecutive urinary isolates were evaluated. RESULTS: E. coli susceptibility to trimethoprim was 70%, comparable to the 70% of trimethoprim/sulfamethoxazole (p = 0.9) and higher than the 56.9% of sulfamethoxazole (p <0.05). This susceptibility pattern was without regional differences. At 2 sites susceptibility to trimethoprim was significantly higher than to ampicillin. At 1 site the susceptibility of other uropathogens to trimethoprim and trimethoprim/sulfamethoxazole was similar to that of E. coli. CONCLUSIONS: In children with urinary tract infection in vitro susceptibility to trimethoprim was comparable to that to trimethoprim/sulfamethoxazole and significantly higher than to sulfamethoxazole. This finding was similar at all sites. Adding sulfamethoxazole appears unnecessary and may represent a risk to patients. Trimethoprim can be used as an alternative to trimethoprim/sulfamethoxazole based on in vitro antibacterial susceptibility. Routine trimethoprim/sulfamethoxazole use for urinary tract infection should be carefully reevaluated.

Rapid group B streptococci screening using a real-time polymerase chain reaction assay.

OBJECTIVE: To estimate the clinical performance characteristics of a real-time polymerase chain reaction (PCR) assay using vaginal/rectal swabs from antepartum (35-37 weeks of gestation) and intrapartum women. METHODS: The assay evaluated is a qualitative, automated, real-time PCR test for the detection of group B streptococci, with results available in approximately 75 minutes. Enrollment in this multicenter clinical study occurred between October 2005 and January 2006. Vaginal/rectal swabs were analyzed by nursing personnel (intrapartum tests) or by laboratory technologists (all others). Polymerase chain reaction assay results were compared with culture using standard methods, including selective broth medium, and to a predicate nucleic acid amplification test. RESULTS: Of 1,028 enrolled women, 234 were deemed ineligible, and 10 had unresolved test results. Of the 784 remaining women, valid PCR assay results were obtained on the first test attempt for 93.0%. Performance characteristics relative to culture were sensitivity 91.1%, specificity 96.0%, positive predictive value 87.8%, negative predictive value 97.1%, and accuracy 94.8%. These results exceeded those obtained using the predicate nucleic acid amplification test. CONCLUSION: Performance characteristics of the PCR assay exceed the threshold recommended by the Centers for Disease Control and Prevention when compared with culture. The test is sufficiently robust to be performed for intrapartum patients in a point-of-care setting by medical professionals. LEVEL OF EVIDENCE: II.

Performance of the cobas HPV Test for the Triage of Atypical Squamous Cells of Undetermined Significance Cytology in Cervical Specimens Collected in SurePath.

OBJECTIVES: Determine performance of the cobas human papillomavirus (HPV) test for triage of atypical squamous cells of undetermined significance (ASC-US) in SurePath. METHODS: Women presenting for routine screening had cervical specimens collected in SurePath and specimen transport medium (STM); those with ASC-US cytology underwent colposcopy. Performance of cobas HPV in SurePath specimens that had undergone a preanalytic procedure to reverse possible cross-linking of HPV DNA was compared with Hybrid Capture 2 (hc2) specimens in STM. RESULTS: Among 856 women, HPV prevalence was 45.8%; HPV 16 and HPV 18 prevalences were lower than expected in the 21- to 29-year-old group in this highly vaccinated population. cobas HPV performance in SurePath was comparable to hc2 in STM. Sensitivity and specificity for detection of cervical intraepithelial neoplasia grade 3 or worse were 87.5% (95% confidence interval [CI], 71.9%-95.2%) and 55.5% (95% CI, 52.1%-58.9%) for cobas and 85.3% (95% CI, 69.9%-93.6%) and 54.7% (95% CI, 51.4%-57.9%) for hc2. Sensitivity was negatively affected by random biopsies performed at colposcopy; comparable sensitivities were achieved in the nonvaccinated and vaccinated populations with disease determined by directed biopsy only. CONCLUSIONS: Performance of cobas HPV for ASC-US triage in pretreated SurePath specimens meets criteria for validation. Preliminary data indicate reliable performance of HPV testing in a highly vaccinated population.

Multicenter Evaluation of the Bruker MALDI Biotyper CA System for the Identification of Clinically Important Bacteria and Yeasts.

OBJECTIVES: A report on the multicenter evaluation of the Bruker MALDI Biotyper CA System (MBT-CA; Bruker Daltonics, Billerica, MA) for the identification of clinically important bacteria and yeasts. METHODS: In total, 4,399 isolates of medically important bacteria and yeasts were assessed in the MBT-CA. These included 2,262 aerobic gram-positive (AGP) bacteria, 792 aerobic gram-negative (AGN) bacteria 530 anaerobic (AnA) bacteria, and 815 yeasts (YSTs). Three processing methods were assesed. RESULTS: Overall, 98.4% (4,329/4,399) of all bacterial and yeast isolates were correctly identified to the genus and species/species complex level, and 95.7% of isolates were identified with a high degree of confidence. The percentage correctly identified and the percentage identified correctly with a high level of confidence, respectively, were as follows: AGP bacteria (98.6%/96.5%), AGN bacteria (98.5%/96.8%), AnA bacteria (98.5%/97.4%), and YSTs (97.8%/87.6%). The extended direct transfer method was only minimally superior to the direct transfer method for bacteria (89.9% vs 86.8%, respectively) but significantly superior for yeast isolates (74.0% vs 48.9%, respectively). CONCLUSIONS: The Bruker MALDI Biotyper CA System accurately identifies most clinically important bacteria and yeasts and has optional processing methods to improve isolate characterization.

Multicenter Evaluation of the Bruker MALDI Biotyper CA System for the Identification of Clinical Aerobic Gram-Negative Bacterial Isolates.

The prompt and accurate identification of bacterial pathogens is fundamental to patient health and outcome. Recent advances in matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) have revolutionized bacterial identification in the clinical laboratory, but uniform incorporation of this technology in the U.S. market has been delayed by a lack of FDA-cleared systems. In this study, we conducted a multicenter evaluation of the MALDI Biotyper CA (MBT-CA) System (Bruker Daltonics Inc, Billerica, MA) for the identification of aerobic gram-negative bacteria as part of a 510(k) submission to the FDA. A total of 2,263 aerobic gram negative bacterial isolates were tested representing 23 genera and 61 species. Isolates were collected from various clinical sources and results obtained from the MBT-CA System were compared to DNA sequencing and/or biochemical testing. Isolates that failed to report as a "high confidence species ID" [log(score) ≥2.00] were re-tested using an extraction method. The MBT-CA System identified 96.8% and 3.1% of isolates with either a "high confidence" or a "low confidence" [log(score) value between 1.70 and <2.00] species ID, respectively. Two isolates did not produce acceptable confidence scores after extraction. The MBT-CA System correctly identified 99.8% (2,258/2,263) to genus and 98.2% (2,222/2,263) to species level. These data demonstrate that the MBT-CA System provides accurate results for the identification of aerobic gram-negative bacteria.

Performance of a cytomegalovirus IgG enzyme immunoassay kit modified to measure avidity.

The measurement of cytomegalovirus (CMV) IgG avidity accurately discriminates recent and past CMV infections. We sought to determine if the Wampole Laboratories CMV IgG enzyme immunoassay (EIA) could be modified to measure avidity. The evaluation panel consisted of 156 serum samples we used in 2002 to validate a laboratory-developed EIA, in which 78 serum samples exhibited low avidity, 7 exhibited intermediate avidity, and 71 exhibited high avidity. The qualitative agreement between the two avidity assays was 94% (147/156); all 9 sera with discordant results exhibited intermediate avidity in one of the assays. The avidity index (AI) values in the two assays showed excellent correlation (r = 0.96, P < 0.0001). The definition of high avidity was verified for the Wampole assay by demonstrating high avidity in 91/93 (98%) recently collected CMV IgG-positive/IgM-negative serum samples. The performance of the Wampole avidity assay in a reference laboratory setting was assessed using 470 consecutive serum samples submitted for CMV IgG avidity testing. Surprisingly, 101 serum samples were negative when screened for CMV IgG using the Wampole kit per the package insert; 98 of these 101 serum samples were tested using a CMV IgG chemiluminescent immunoassay, and only 5 were positive. Of the 369 CMV IgG-positive samples, 6% exhibited low IgG avidity, 6% exhibited intermediate avidity, and 88% exhibited high avidity; CMV IgM detection rates were inversely related to AI levels. These findings show that (i) the Wampole CMV IgG EIA can be modified to measure CMV IgG avidity, (ii) many samples are apparently submitted for avidity testing without knowledge of their CMV IgG status, and (iii) most CMV IgG-positive sera submitted for avidity testing exhibit high avidity.

Comparison of the premier toxin A and B assay and the TOX A/B II assay for diagnosis of Clostridium difficile infection.

Clostridium difficile causes nosocomial diarrhea and is responsible for complications such as pseudomembranous colitis, megacolon, and perforation. Using 442 stool specimens, we compared the sensitivities and specificities of the Premier toxin A and B (Meridian Bioscience, Inc.) and C. difficile TOX A/B II (TechLab, Inc., Blacksburg, VA) immunoassays in the Virology Department of the Kaiser Permanente Regional Reference Laboratories. The Premier toxin A and B assay demonstrated a higher sensitivity (97.44%) and a higher positive predictive value (79.17%) than the C. difficile TOX A/B II assay (87.18% and 75.56%, respectively), while assay specificities and negative predictive values were similar. We also performed experiments using serially diluted, purified toxin A and B antigens to understand the basis for assay differences. The two assays' toxin A antibodies detected toxin A at comparable levels. Preliminary results indicated that the toxin B antibody in the Premier toxin A and B assay could detect toxin B at a concentration of 125 pg/100 microl, while the toxin B antibody in the C. difficile TOX A/B II assay could not detect toxin B below a concentration of 250 pg/100 microl. Therefore, the Premier toxin A and B assay provides greater sensitivity than the C. difficile TOX A/B II assay, perhaps due to a superior detection ability of its toxin B antibody.