Multicenter Evaluation of the Simplexa VZV Direct Assay for Detection of Varicella-Zoster Virus in Cerebrospinal Fluid and Lesion-Swab Specimens.

Varicella-zoster virus (VZV) is the etiologic agent of varicella (chickenpox) and herpes zoster (shingles) infections commonly involving skin, mucous membranes, and less frequently the central nervous system. Traditional methods for the laboratory diagnosis of these infections are time-consuming, labor-intensive, and often insensitive. As such, these tests are being replaced by more sensitive and rapid molecular methods. This study evaluated the performance of two different molecular assays, the Simplexa VZV Direct and Simplexa VZV Swab Direct, to detect VZV DNA in cerebrospinal fluid (CSF) and lesion-swab specimens, respectively. The Simplexa VZV Direct and Simplexa VZV Swab Direct assays were compared against individual composite reference methods that varied depending on the sample cohort examined. A total of 883 CSF and 452 cutaneous and mucocutaneous prospective, retrospective, and contrived specimens were evaluated in this multicenter study. The results of this study showed that the Simplexa assays demonstrated near perfect agreement (k = 0.98) compared to the composite reference methods for the detection of VZV in CSF and lesion swab specimens. A further comparison between the standard of care molecular assays employed at the site of specimen collection and the Simplexa assays demonstrated excellent agreement (k = 1.0). The Simplexa assays offer rapid and reliable alternatives for the detection of VZV in certain clinical specimens without the need for nucleic acid extraction.

A Novel Multiplexed Enzyme-Linked Immunosorbent Assay for the Detection of IgG Seroreactivity to Cytomegalovirus (CMV) UL144.

Infection with human cytomegalovirus (CMV) is common and may have grave consequences in transplant recipients and congenitally infected children. Diagnosis of CMV infection is based on detection of specific antibodies and molecular assays. The incorporation of CMV serological assays into diagnostic algorithms requires careful evaluation and interpretation. Very few serological assays measure CMV infection by a specific strain. We developed an enzyme-linked immunosorbent assay (ELISA) using CMV-encoded UL144 as the antigen. UL144 encodes three major genotypes, A, B, and C, and recombinants. The ELISA was developed with the three UL144 proteins and optimized as a multiplex assay. Sera from 55 positive and 59 negative CMV IgG, determined by the clinical microbiology laboratory, were used for evaluation and optimization. A cutoff optical density (OD) that distinguishes UL144 antibody-positive from antibody-negative sera was established. UL144 A, B, C, and combinations of these antigens were detected in sera. An assay threshold of 0.1 was established and, from a total of 303 sera, the overall sensitivity, specificity, and positive and negative predictive values of the multiplex ELISA were 86.72% (95% confidence interval [CI] 79.59% to 92.07%), 96.57% (92.69% to 98.73%), 94.40% (88.45% to 97.38%), and 91.60% (87.50% to 94.44%), respectively. The inter- and intraassay median coefficients of variation were 0.06 (interquartile range [IQR] 0.56, 0.2) and 0.171 (IQR 0.038, 0.302), respectively. No cross-reactivity was observed with HSV-positive CMV-negative sera. This ELISA gives simple and reproducible results for detection of anti-CMV UL144 IgG. It may assist in differentiating natural infection from CMV vaccines that lack UL144, and may provide an important tool for epidemiological studies of CMV strains.

Multicenter Clinical Evaluation of the Revogene Strep A Molecular Assay for Detection of Streptococcus pyogenes from Throat Swab Specimens.

Group A streptococcus (GAS) species cause bacterial pharyngitis in both adults and children. Early and accurate diagnosis of GAS is important for appropriate antibiotic therapy to prevent GAS sequalae. The Revogene Strep A molecular assay (Meridian Bioscience Canada Inc, Quebec City, QC, Canada) is an automated real-time PCR assay for GAS detection from throat swab specimens within approximately 70 min. This multicenter prospective study evaluated the performance of the Revogene Strep A molecular assay compared to that of bacterial culture. Dual throat swab specimens in either liquid Amies or Stuart medium were collected from eligible subjects (pediatric population and adults) enrolled across 7 sites (USA and Canada). Revogene Strep A and reference testing was performed within 7 days and 48 h of sample collection, respectively. Of the 604 evaluable specimens, GAS was detected in 154 (25.5%) samples by the reference method and in 175 (29%) samples by the Revogene Strep A assay. Revogene Strep A assay sensitivity and specificity were reported to be 98.1% (95% confidence interval [CI], 94.4 to 99.3) and 94.7% (95% CI, 92.2 to 96.4), respectively. The positive predictive value was 86.3% (95% CI, 80.4 to 90.6), negative predictive value was 99.3% (95% CI, 98.0 to 99.8) with a 1.0% invalid rate. Discrepant analysis with alternative PCR/bidirectional sequencing was performed for 24 false-positive (FP) and 3 false-negative (FN) specimens. Concordant results were reported for 17 (FP only) of 27 discordant specimens. The Revogene Strep A assay had high sensitivity and specificity for GAS detection and provides a faster alternative for GAS diagnosis.

Mycobacterium mucogenicum bacteremia in immune-compromised patients, 2008-2013.

Mycobacterium mucogenicum (MM) is a rapidly growing nontuberculous mycobacterium that may rarely cause bacteremia in immune-compromised hosts. All MM cases from 2008 to 2013 were analyzed across 4 risk groups: stem cell transplantation (SCT), hematologic malignancy, solid tumors, and others. Descriptive analysis was performed, as well as comparative analysis of neutropenic patients (absolute neutrophil count ≤1000/µL) with nonneutropenic patients. Of 39 MM cases, 27 patients had undergone SCT. Neutropenia was present in 12 patients. There was a significant difference in the presence of fever at the time of MM bacteremia between neutropenic and nonneutropenic groups (92% versus 42%; P=0.005). Central venous catheter (CVC) was present in 33 cases. All patients were treated with >1 antibiotic. Most frequently used combination antibiotic regimen involved clarithromycin and amikacin. Median duration of antibiotic treatment was 42days. Bacteremia resolved in all cases with CVC removal and combination antibiotic treatment.

Morbidity and mortality attributable to Rothia bacteremia in neutropenic and nonneutropenic patients.

Rothia spp. are increasingly being recognized as emerging opportunistic pathogens associated with serious infections in immune-compromised hosts. Risk factors include neutropenia, hematologic malignancies, prosthetic devices, and intravascular catheters. We describe 29 patients at our institute from 2006 to 2014 with positive blood cultures for Rothia spp. Neutropenia was observed in 21/29 (72%) patients at the time of bacteremia, and 16/29 (61%) had leukemia. Neutropenic patients were less likely than nonneutropenic patients to have polymicrobial infection (24% versus 63%; P= 0.083) and were also more likely to have multiple positive blood cultures (76% versus 0%; P= 0.0003), indicating true infection. Sources of bacteremia included intravascular catheters, mucositis, and presumed gut translocation. A significant association was seen with steroid use (81% versus 13%; P= 0.0014) and fluoroquinolone use (86% versus 13%; P≤ 0.0001) preceding bacteremia in neutropenic patients. There was no difference between the 2 groups for admission to intensive care unit or mortality. One death was reported possibly due to Rothia infection.

ESwab as an optional collection device for use with the Affirm VPIII microbial test system.

The ESwab collection device was compared to the collection swab provided as part of the Affirm VPIII microbial identification test kit for testing vaginal specimens with the Affirm test system. There was excellent agreement between the two sampling devices for Candida spp., Gardnerella vaginalis, and Trichomonas vaginalis.