High-throughput sequencing of cerebrospinal fluid for diagnosis of chronic Propionibacterium acnes meningitis in an allogeneic stem cell transplant recipient.

BACKGROUND: A 40-year-old man with chronic myelogenous leukemia presented multiple times over a period of 3 years with episodes of confusion, wide-based gait and falls because of recurrent hydrocephalus despite repeated therapeutic lumbar punctures. These problems occurred in the context of persistent cerebrospinal fluid (CSF) pleocytosis and leptomeningeal enhancement. Extensive diagnostic workups and therapeutic trials had failed to identify a clinically plausible cause or produce any significant improvement in the CSF and neuroimaging abnormalities. METHODS: We used high-throughput metagenomic shotgun sequencing to identify microbes in 2 CSF samples collected from the patient during his illness. These results were compared to sequence data from 1 CSF sample collected during treatment and 5 control CSF samples from other patients. RESULTS: We found sequences representing 53% and 67% of the Propionibacterium acnes genome in 2 CSF samples collected from the patient during his illness. Directed antimicrobial therapy was administered for 6 weeks with resolution of CSF and neuroimaging abnormalities. Sequencing of a sample obtained during treatment demonstrated that the P. acnes levels were decreased to background levels. After insertion of a ventriculo-peritoneal shunt, the patient returned to baseline status. CONCLUSIONS: High-throughput metagenomic shotgun sequencing revealed P. acnes as the cause of chronic meningitis that had eluded conventional attempts at diagnosis. Treatment directed at this organism resulted in cure of the infection and clinical improvement.

BK-virus and the impact of pre-emptive immunosuppression reduction: 5-year results.

A 1-year, single-center, randomized trial demonstrated that the calcineurin inhibitor or adjuvant immunosuppression, independently, does not affect BK-viruria or viremia and that monitoring and pre-emptive withdrawal of immunosuppression was associated with resolution of BK-viremia and absence of clinical BK-nephropathy without acute rejection or graft loss. A retrospective 5-year review of this trial was conducted. In cases of BK viremia, the antimetabolite was withdrawn and for sustained viremia, the calcineurin inhibitor was minimized. Five-year follow-up was available on 97% of patients. Overall 5-year patient survival was 91% and graft survival was 84%. There were no differences in patient-survival by immunosuppressive regimen or presence of BK-viremia. Immunosuppression and viremia did not influence graft survival. Acute rejection occurred in 12% by 5-years after transplant, was less common with tacrolimus versus cyclosporine (9% vs. 18%; p = 0.082), and was lowest with the tacrolimus-azathioprine regimen (5%, p = 0.127). Tacrolimus was associated with better renal function at 5-years (eGFR 63 FK vs. 52 CsA mL/min, p = 0.001). Minimization of immunosuppression upon detection of BK-viremia was associated with excellent graft survival at 5-years, low rejection rates and excellent renal function. It is a safe, short and long-term strategy that resulted in freedom from clinically evident BK-virus nephropathy.

Clinical outcomes of solid organ transplant recipients with ehrlichiosis.

Because of our experience with severe Ehrlichia infections in lung transplant recipients, we reviewed all cases of ehrlichiosis in solid organ transplant recipients at Barnes-Jewish Hospital in St. Louis, Missouri. Between 1996 and 2007, 25 cases of ehrlichiosis were identified. We retrospectively collected demographic, clinical, laboratory, and outcomes data, and we compared the 5 cases in lung transplant recipients with 20 cases in other solid organ transplant recipients (heart, 2; kidney, 13; liver, 5). The presenting symptoms in the majority of both groups consisted of fever and headache. Clinical outcomes were worse in the lung transplant group and included a greater need for intensive care unit treatment (80% vs. 20%, P=0.02), longer length of hospital stay (21 vs. 5 days, P=0.02), and propensity to develop acute lung injury or acute respiratory distress syndrome (60% vs. 10%, P=0.04). No mortalities occurred in either group of patients. In an endemic area, ehrlichiosis is not unusual in solid organ transplant recipients, and lung transplant recipients tend to have a more severe illness.

Antibiotic-induced lysis of enterococci.

Enterococci are resistant to penicillin killing in vivo and in vitro. Because some bacteria resistant to penicillin killing have reduced autolytic activity, we examined the lysis of clinical enterococcal isolates suspended in buffer (spontaneous lysis), and compared it with their susceptibility to antibiotic-induced lysis and killing. We found significant correlations between spontaneous and antibiotic-induced lysis, using five antibiotics that inhibit cell wall synthesis (penicillin, cephalothin, bacitracin, cycloserine, and vancomycin). Among isolates, strains more rapidly lysed by one antibiotic were more rapidly lysed by the other antibiotics, and more susceptible to spontaneous lysis. In studies involving a single strain grown in different media, spontaneous lysis also correlated closely with antibiotic-induced lysis. These results are consistent with a common mechanism for spontaneous and antibiotic-induced lysis, such as the autolytic enzyme system. Human serum was one of the least permissive media tested for enterococcal growth and antibiotic-induced lysis and killing. We suggest that the inhibitory effect of human serum on growth and the activation of the enterococcal autolytic enzyme system may be a critical factor in the resistance of enterococcal endocarditis to treatment with penicillin alone.

RNA fingerprinting of respiratory syncytial virus using ribonuclease protection. Application to molecular epidemiology.

We have used the technique of ribonuclease protection to define genomic variation among circulating isolates of subgroup A respiratory syncytial (RS) virus. RNAs extracted from HEp-2 cells infected with strains to be analyzed were hybridized with a 32P-labeled RNA probe corresponding to the RS virus G glycoprotein (A2 strain). Areas of nonhomology were detected by cleavage with ribonuclease A. Using this technique, multiple distinct RNA cleavage patterns could be distinguished among viral isolates recovered from infants residing in the same metropolitan area and infected during the same epidemic season. Epidemiologically related isolates (from coinfected twins, from infants infected during a nosocomial outbreak at an extended care facility, and from institutionalized adults infected during an outbreak) yielded identical patterns. In two separate outbreaks, differences in cleavage patterns among certain isolates corresponded to epidemiologically significant differences among the individuals from whom the isolates were recovered. We conclude that substantial genomic heterogeneity exists among circulating isolates of subgroup A RS virus. Ribonuclease protection can be used as a molecular fingerprinting tool for expanded studies of the molecular epidemiology of this virus.

Once daily ganciclovir as initial pre-emptive therapy delayed until threshold CMV load > or =10000 copies/ml: a safe and effective strategy for allogeneic stem cell transplant patients.

Quantitative polymerase chain reaction (QPCR) for cytomegalovirus (CMV) is emerging as the preferred screening method for detection of CMV viremia in patients following allogeneic bone marrow and peripheral blood stem cell transplant. However, there are currently no universally accepted QPCR treatment thresholds at which to start pre-emptive therapy. We report here results of a pre-emptive therapy strategy using ganciclovir (GCV) 5 mg/kg initiated once daily (ODG) delayed till a threshold CMV load of > or =10 000 copies/ml whole blood in clinically stable patients. Sixty-nine at risk patients underwent allogeneic stem cell transplant. 48/69 (70%) patients had an initial episode of CMV viremia. 5/48 (10%) cleared viremia without requiring treatment. 28/43 (65%) patients requiring treatment initiated treatment with ODG. 17/28 (61%) patients successfully cleared CMV viremia on ODG, 10/28 (36%) patients required dose escalation to twice daily GCV for increasing viral loads. There were two cases of CMV disease (colitis) and no deaths due to CMV disease in patients initiating treatment with ODG. We conclude delaying pre-emptive therapy with ODG until whole blood QPCR> or =10 000 copies/ml is a safe and effective strategy for CMV viremia after allogeneic stem cell transplant in clinically stable patients.

Epstein-Barr virus encephalomyelitis diagnosed by polymerase chain reaction: detection of the genome in the CSF.

A case of encephalomyelitis with polymerase chain reaction detection of Epstein-Barr virus (EBV) in the CSF, and concurrent serologic changes consistent with acute systemic EBV infection is presented and discussed. We document involvement of the brain, spinal cord, and nerve roots, summarize some unusual imaging findings, and note the evolution of CSF oligoclonal bands.

Use of polymerase chain reaction to demonstrate cytomegalovirus DNA in CSF of patients with human immunodeficiency virus infection.

We used the polymerase chain reaction (PCR) to demonstrate cytomegalovirus (CMV) DNA in the CSF of a patient with the CMV radiculomyelopathy syndrome. To investigate the significance of this finding, we also performed PCR for CMV on CSF samples from 30 patients with human immunodeficiency virus (HIV) infection and neurologic disease, four patients with solid organ transplants including three with active CMV infection, and 10 patients with no clinical suspicion of HIV or CMV infection. There was CMV DNA only in patients with HIV, and it was present more often in patients with evidence of spinal cord dysfunction. Our results suggest that PCR may be useful in the rapid diagnosis of CMV infection of the CNS in patients with HIV and that the radiculomyelopathy syndrome may represent only part of a spectrum of CMV-induced spinal cord dysfunction in these patients.

Use of the polymerase chain reaction in the diagnosis of herpes simplex encephalitis: a decision analysis model.

PURPOSE: To evaluate the utility of an assay based on a polymerase chain reaction (PCR) of cerebrospinal fluid in the management of patients with suspected herpes simplex encephalitis. METHODS: A decision model was constructed and used to compare a PCR-based approach with empiric therapy. Inputs required by the model included the sensitivity (96%) and specificity (99%) of PCR (derived from review of the literature), the prevalence of herpes simplex encephalitis (5%, based on the actual prevalence at Barnes Hospital among patients treated empirically with acyclovir), the outcomes for patients with and without herpes simplex encephalitis (derived from clinical studies of the Collaborative Antiviral Study Group and the actual experience at Barnes Hospital), and the average duration of empiric acyclovir therapy for patients with possible herpes simplex encephalitis (5.3 days based on actual experience at Barnes Hospital). RESULTS: Using these input values, the decision model predicted better outcomes with empiric therapy. However, low rates of inappropriate discontinuation of empiric therapy in patients with herpes simplex encephalitis or improved diagnosis and outcome resulting from a negative PCR assay result in patients without herpes simplex encephalitis led to better outcomes with the PCR-based approach. The PCR-based approach was associated with 9.2 fewer doses of acyclovir per patient. CONCLUSION: Based on the decision model using conservative assumptions, a PCR-based approach can yield better outcomes and reduced acyclovir use compared with empiric therapy.

Use of conventional and IgM-specific radioimmunoassays for anti-hepatitis A antibody in an outbreak of hepatitis A.

During a common source outbreak of hepatitis A, we studied the characteristics and utility of commercially available radioimmunoassays for total and IgM-specific antibody to hepatitis A virus. IgM hepatitis A antibody was detectable in all serum specimens obtained up to 119 days following onset from the seven persons with hepatitis A, and as long as 347 days in one person. Acute infection could also be documented by a four-fold or greater increase in titers of hepatitis A antibody, although as long as nine weeks was required between the times acute and convalescent specimens were obtained. The radioimmunoassay for IgM-specific hepatitis A antibody had greater specificity (99 percent versus 84 percent) and a higher positive prediction value (88 percent versus 23 percent) for the diagnosis of acute hepatitis A than did the radioimmunoassy for hepatitis A antibody. Uses of the radioimmunoassay for IgM-specific hepatitis A antibody include rapid diagnosis of acute hepatitis A, differentiation between recent and past hepatitis A infection, and screening for recent hepatitis A infection in epidemiologic investigation.

Failure of high-dose oral acyclovir with or without immune globulin to prevent primary cytomegalovirus disease in recipients of solid organ transplants.

PURPOSE: To assess the efficacy of acyclovir and intravenous immune globulin (IVIG) for cytomegalovirus (CMV) prophylaxis in high-risk recipients of solid organ transplants. PATIENTS AND METHODS: We randomized 21 CMV-seronegative organ transplant recipients with seropositive donors (D+R-) to receive oral acyclovir, 800 mg four times daily, or, in addition to acyclovir, IVIG, 300 mg/kg, every 2 weeks for six doses. Patients were followed closely for the development of CMV infection and disease. RESULTS: All but one prophylactically treated patient (95%) developed CMV infection. Fifteen of 21 patients (71%) who received prophylaxis fulfilled criteria for CMV disease. Disease onset was delayed in those who received IVIG, but this did not reach statistical significance. Ganciclovir was used for treatment in 15 of the 21 patients (71%). CONCLUSIONS: Acyclovir, with or without IVIG, did not prevent primary CMV infection or disease in D+R- solid organ transplant recipients at our institution. Moreover, most patients were treated with ganciclovir despite the use of prophylaxis. Given the ready availability of ganciclovir to treat CMV disease, we recommend a reappraisal of the role of CMV prophylaxis by these means in the solid organ transplant population.

Epstein-Barr virus DNA in peripheral blood leukocytes of patients with posttransplant lymphoproliferative disease.

We tested the hypotheses that Epstein-Barr virus (EBV) DNA levels in peripheral blood leukocytes (PBL) of transplant recipients with posttransplant lymphoproliferative disease (PTLD) (1) exceed those of patients without PTLD, (2) rise with or before clinical detection of the disease, and (3) fall with effective therapy. Using the polymerase chain reaction (PCR) and an endpoint dilution technique, we compared EBV DNA levels in sequential specimens from 5 patients with PTLD, 16 solid organ transplant recipients without PTLD, and 5 young adults with primary infectious mononucleosis (IM), and in single specimens from 21 healthy seropositive subjects. EBV DNA levels in the first two groups rose with induction of immunosuppression despite prophylactic acyclovir. Markedly elevated levels of EBV DNA were seen in 4 of 5 patients with PTLD at or before clinical diagnosis. The peak levels in these patients exceeded those of transplant recipients without PTLD (P = 0.02) and healthy adults with IM (P = 0.02). EBV DNA levels fell dramatically with effective therapy. Four of 21 healthy seropositive subjects demonstrated low levels of EBV DNA, similar to levels seen late in the course of patients with IM. We conclude that a semiquantitative PCR assay for EBV DNA in PBL can assist in the detection of PTLD and in monitoring the effect of therapy.

The impact of ganciclovir-resistant cytomegalovirus infection after lung transplantation.

BACKGROUND: Cytomegalovirus (CMV) resistance to ganciclovir has become increasingly common in acquired immunodeficiency syndrome patients but has only rarely been reported in recipients of solid organ transplants. METHODS: A retrospective study of ganciclovir susceptibility testing of CMV isolates recovered from lung transplant recipients was performed. Patients with CMV isolates having partial (1 or =3 microg/ml) to ganciclovir determined by plaque reduction assay were included in a case-control study to identify risk factors for ganciclovir resistance. RESULTS: Between 2/91 and 5/98, 18 patients (5.2% of patients transplanted) were found to have CMV infections with some degree of ganciclovir resistance (4 partially, 14 fully resistant). More positive viral blood cultures (3.2+/-2.5 vs. 1.6+/-1.4 CMV positive cultures, P=0.02) and more episodes of CMV pneumonitis (0.24+/-0.23 vs. 0.10+/-0.17 episodes/bronchoscopy, P=0.02) occurring before the detection of resistance were seen among resistant patients than controls. Ganciclovir-resistant patients received more antithymocyte globulin during induction (70+/-44 vs. 45+/-39 mg/kg, P=0.03) and received ganciclovir for a greater number of days (79+/-52 vs. 64+/-53 days, P=0.005) before the detection of resistance than controls. Ganciclovir-resistant patients had a shorter survival and an earlier onset of bronchiolitis obliterans syndrome compared with patients in the transplant database at Washington University. CONCLUSIONS: Ganciclovir-resistant CMV infection is a serious complication of solid organ transplantation associated with more episodes of viremia, more frequent disease, earlier onset of bronchiolitis obliterans and shorter survival. The use of antithymocyte globulin and prolonged exposure to ganciclovir are risk factors for the development of ganciclovir resistance.

Relationship of cytomegalovirus viral load in blood to pneumonitis in lung transplant recipients.

We developed a multiplex, quantitative, real-time, polymerase chain reaction assay for cytomegalovirus (CMV) and used it to measure the CMV viral load in weekly blood specimens from 43 lung transplant recipients. The median viral load in blood samples immediately preceding bronchoscopy was 1150 copies/microg human DNA for 12 subjects with pneumonitis compared to 91 copies for 31 subjects without (P=0.02, Mann-Whitney U test). Each log10 increase in CMV viral load resulted in an increase of 1.92 in the odds ratio for CMV pneumonitis (95% confidence interval 1.03-3.56). CMV viral load was elevated (>100 copies/microg human DNA) for a median of 21 days before bronchoscopy in those subjects with pneumonitis versus 0 days in those without (P=0.004). We conclude that the risk of CMV pneumonitis after lung transplantation is related to the level of CMV DNA in blood. Quantitative PCR should be evaluated prospectively for the preemptive management of CMV in lung transplant recipients.

Prophylactic oral ganciclovir compared with deferred therapy for control of cytomegalovirus in renal transplant recipients.

BACKGROUND: Treatment with prophylactic oral acyclovir, intravenous ganciclovir, or immunoglobulins to prevent cytomegalovirus (CMV) infection and disease in renal transplantation is associated with variable efficacy and significant expense. We studied control of CMV in renal transplant recipients using either prophylactic oral ganciclovir or deferred therapy with intensive monitoring with polymerase chain reaction (PCR) analysis. METHODS: Forty-two recipients were followed for 6 months after transplantation. Ganciclovir (1000 mg p.o. t.i.d.; n=19) or acyclovir (200 mg p.o. b.i.d.; n=23) was begun at transplantation and continued for 12 weeks. PCR for CMV was performed on buffy-coat specimens every week for 15 weeks and at months 5 and 6. RESULTS: No patients in the ganciclovir group, compared with 14 of 23 patients (61%) in the deferred-therapy group (P<0.0001), developed CMV disease during the first 12 weeks. In the ganciclovir group, 4 of 19 patients (21%) subsequently experienced 5 episodes, whereas 14 patients in the deferred-therapy group experienced 18 episodes (P=0.013 for subjects and P=0.026 for episodes). The time to disease was also delayed in the ganciclovir group compared with the deferred-therapy group (133+/-17 days vs. 51+/-7 days; P<0.0001). Oral ganciclovir also prevented CMV viremia during prophylaxis (2/19 patients [11%] vs. 23/23 patients [100%]). Time to CMV viremia was delayed in the ganciclovir group; however, 13/19 patients (68%) ultimately showed PCR evidence for CMV viremia (P=0.005). CONCLUSIONS: An initial 12-week course of oral ganciclovir prevents CMV disease and infection in renal transplant recipients during prophylaxis, and the benefits persist after discontinuation.

Enteroviral meningitis in infancy: potential role for polymerase chain reaction in patient management.

STUDY OBJECTIVE: To evaluate the performance characteristics and potential clinical utility of a polymerase chain reaction (PCR) assay for enteroviral RNA in comparison to viral culture in infants under 3 months of age with meningitis. SPECIMENS AND TESTING: Specimens were obtained from a collection of cerebrospinal fluid specimens from infants under 3 months of age (excluding those in the neonatal intensive care unit) undergoing lumbar puncture at St. Louis Children's Hospital during a 12-month period. Those tested by PCR included all 27 with pleocytosis, 8 others from infants without pleocytosis but from whom an enterovirus was cultured, and 10 from infants who did not have pleocytosis and had a negative viral culture of cerebrospinal fluid. Viral cultures were performed at the discretion of physicians caring for individual patients. RESULTS: PCR was positive for enteroviral RNA on cerebrospinal fluid (CSF) specimens from 11 of 12 patients with definite or probable enteroviral meningitis, as well as on 6 of 13 with possible enteroviral meningitis, and was negative on all 10 with absence of pleocytosis and negative enteroviral cultures. CSF viral cultures were negative in 6 of the patients in whom PCR was positive. Viral cultures had minimal impact on patient management. In contrast, under study assumptions, PCR could have saved an average of 1.2 days of hospitalization per patient in the 27 patients with CSF pleocytosis. CONCLUSIONS: Enterovirus PCR performed on CSF is a sensitive and specific method for the diagnosis of enteroviral meningitis. This method has the potential for improving the accuracy of diagnosis in young infants and for saving costs by allowing earlier diagnosis and discharge from the hospital when clinically appropriate.

Prevalence of hepatitis B antibodies in personnel at a children's hospital.

Antibody to hepatitis B core antigen and antibody to hepatitis B surface antigen were measured in serum samples from 825 personnel at St Louis Children's Hospital (702 were possibly at high risk of occupational hepatitis B virus (HBV) exposure and 123 were office personnel); 5.6% had positive findings for both antibodies, 5.6% had findings for antibody to hepatitis B surface antigens alone, and 1.3% had positive findings for antibody to hepatitis B core antigens alone. The group with positive findings for antibody to hepatitis B surface antigens alone did not have traditional risk factors for HBV infection, suggesting that this serologic finding may not be a reliable indicator of past HBV infection. After accounting for the effects of age, sex, and ethnicity, it was found that no occupational group had a significantly increased prevalence of HBV antibodies compared with prevalence in other personnel. In comparison with volunteer blood donors, only physicians older than 40 years of age had an increased HBV antibody prevalence. It is concluded that St Louis Children's Hospital has not been a high-risk environment for HBV exposure in recent years. However, caution is advised in generalizing these conclusions because other children's hospitals may serve a patient population at higher risk of HBV infection. Decisions regarding HBV immunization policy should take into consideration the fact that personnel at different hospitals may face markedly different risks of HBV exposure.

Genetic variability in envelope-associated protein genes of closely related group A strains of respiratory syncytial virus.

The genetic and antigenic diversity present in respiratory syncytial virus (RSV) strains may in part be explained by genetic drift similar to that which occurs with influenza virus B. To study drift in RSV strains, we sequenced the five membrane-associated genes, M, SH, G, F, and M2, from three sets of RSV isolates: one set of seven closely related isolates obtained over 5 years in St. Louis, MO, and two sets of four closely related RSV isolates from other communities. We found nucleotide-variable and conserved regions in all five genes, and the greatest diversity in the SH and G genes. We did not find clear evidence of genetic drift in the seven isolates from St. Louis for any of the five genes. Although the relationships between strains were usually maintained independent of the genes studied, for several isolates there was a dramatic shift in genetic relationships for one of the five genes. Our inability to demonstrate genetic drift and the dramatic shift in genetic relationships between some strains for some genes suggest that we need to better define the mechanisms and rate of change in this virus to accurately define phylogenetic relationships between strains.

Resistance to penicillin and non-beta-lactam antibiotics of Streptococcus pneumoniae at a children's hospital.

During a 12-month period we tested all isolates of Streptococcus pneumoniae recovered from patients at St. Louis Children's Hospital for resistance to penicillin and other antibiotics. Twenty-seven (20%) of 136 had relative penicillin resistance (minimum inhibitory concentration 0.1 to 1.0 microgram/ml) and 8 (6%) were fully resistant (minimum inhibitory concentration > or = 2.0 micrograms/ml). Sixteen percent from blood and cerebrospinal fluid were resistant, compared with 30% from other body sites. The resistant isolates were of diverse serotypes and included 38% intermediate and 6% resistant to cefotaxime, 40% resistant to trimethoprim-sulfamethoxazole and 20% resistant to erythromycin. Patients with resistant isolates were more likely to have taken antibiotics of the aminopenicillin class and to be of the white race. We conclude that penicillin-resistant pneumococci, including some with resistance to third generation cephalosporins and some with multidrug resistance to non-beta-lactam antibiotics, are widespread in the St. Louis area. The presence of these stains requires reconsideration of current approaches to the antibiotic therapy of a variety of infectious diseases in which pneumococci play a prominent role.

Polymerase chain reaction for Streptococcus pyogenes used to evaluate an optical immunoassay for the detection of group A streptococci in children with pharyngitis.

BACKGROUND: In evaluations of sensitive rapid tests for group A streptococci such as the optical immunoassay (OIA), some samples are positive by the antigen test but negative by culture. A method is needed for resolving these discrepant results. OBJECTIVE: To develop a PCR-based assay to detect group A streptococci and to use it to establish a reference standard for evaluating an OIA for group A streptococcal antigen. METHODS: A PCR assay that detects a segment of the MF gene of Streptococcus pyogenes was developed for the detection of group A streptococci in throat swabs. Paired swabs were obtained from 200 children with symptomatic pharyngitis and used to perform OIA, agar culture, broth-enhanced culture and PCR. As a reference standard any patient with group A streptococci detected by either culture or PCR was considered to be truly positive. RESULTS: In comparison to agar and broth-enhanced culture procedures, OIA had sensitivities of 82 and 80% and specificities of 87 and 89%, respectively. Eight (44%) of 18 samples that were positive by OIA but negative by culture were positive for group A streptococci by PCR. Compared with the reference standard, sensitivities were OIA 76%, agar culture 79%, broth-enhanced culture 86% and PCR 96%. The specificity of OIA was 92%. CONCLUSIONS: PCR can be used to establish a reference standard for evaluating rapid tests for group A streptococci. With this reference standard OIA was nearly as sensitive as but less specific than agar culture for detection of group A streptococci. Maximum detection requires use of both tests.