Multicenter Performance Evaluation of the Simplexa Bordetella Direct Kit in Nasopharyngeal Swab Specimens.

Detection of Bordetella pertussis and Bordetella parapertussis using molecular methods is sensitive and specific with a short turnaround time compared to other diagnostic methods. In this multicenter study, we compared the performance of the Simplexa Bordetella Direct kit to those of other molecular assays in detecting and differentiating B. pertussis and B. parapertussis in nasopharyngeal swab specimens. The limits of detection (LODs) were 150 CFU/ml or 3 fg/µl of DNA for B. pertussis and 1,500 CFU/ml or 10 fg/µl of DNA for B. parapertussis A total of 1,103 fresh and residual frozen specimens from eight clinical sites were tested. Combining the data from individual clinical sites using different comparative assays, the overall positive percent agreement (PPA) and negative percent agreement (NPA) for B. pertussis were 98.7% and 97.3%, respectively. The overall PPA and NPA for B. parapertussis were 96.7% and 100%, respectively. For prospective fresh specimens, the overall PPA and NPA for both targets were 97.7% and 99.3%, respectively. For retrospective frozen specimens, the overall PPA and NPA for both targets were 92.6% and 93.2%, respectively. The percentage of invalid results was 1.0%. A cross-reactivity study using 74 non-Bordetella bacterial species and five yeast species revealed that the Simplexa Bordetella Direct kit was 100% specific. The hands-on time and assay run time of the Simplexa Bordetella Direct kit are favorable compared to those of other commercial and laboratory-developed tests. In summary, the Simplexa Bordetella Direct kit has a performance comparable to those of other molecular assays for the detection of B. pertussis and B. parapertussis.

Neonatal Pasteurella multocida subsp. septica Meningitis Traced to Household Cats: Molecular Linkage Analysis Using Repetitive-Sequence-Based PCR.

Pasteurella multocida is a rare cause of neonatal bacterial meningitis. We describe such a case and verify two household cats as the source of infection using repetitive-element PCR (rep-PCR) molecular fingering.

Evaluation of the Lyra Direct Strep Assay To Detect Group A Streptococcus and Group C and G Beta-Hemolytic Streptococcus from Pharyngeal Specimens.

The Lyra Direct strep assay was compared to culture for its ability to detect Streptococcus group A and ß-hemolytic groups C/G using rapid antigen-negative pharyngeal specimens (n = 161). The Lyra assay correctly detected all ß-hemolytic streptococci (group A, n = 19; group C/G, n = 5). In batch mode, the Lyra assay reduced intralaboratory turnaround time by 60% (18.1 h versus 45.0 h) but increased hands-on time by 96% (3 min 16 s versus 1 min 40 s per specimen).

Characterization of a novel melt curve by use of the Roche LightCycler HSV 1/2 analyte-specific reagent real-time PCR assay: frequencies of this novel (low) melt curve and commonly encountered (intermediate) melt curves.

We characterize a novel probe binding-site polymorphism detectable solely by melt curve analysis using the Roche LightCycler HSV 1/2 analyte-specific reagent real-time PCR assay. The frequencies of this novel (47°C) and previously described intermediate (60 to 62°C) melt curves were 0.016% and 4.9%, respectively.

Performance of the Affirm VP-III using residual vaginal discharge collected from the speculum to characterize vaginitis in symptomatic women.

OBJECTIVE: To evaluate the ability of collecting the Affirm VP-III test sample using the residual vaginal discharge found on the speculum. METHODS AND METHODS: One hundred nine symptomatic women (≥18 y) participated in this study. During pelvic examination, vaginal fluid was collected onto 3 swabs for office-based diagnostic tests and Affirm (referred to as Affirm-R). A fourth swab was used to collect residual vaginal discharge from the speculum, followed by Affirm testing (referred to as Affirm-RVD). Sensitivity, specificity, and Cohen κ agreement for office-based diagnostic tests and Affirm-RVD were determined against Affirm-R. RESULTS: Complete results were available for 99 samples. Cohen κ agreement between Affirm-RVD and Affirm-R was 0.66 (p<.0001) for Gardnerella vaginalis, 0.81 (p<.0001) for Candida species, and 1.0 (p<.0001) for Trichomonas vaginalis. Affirm-RVD sensitivity, specificity, and positive and negative predictive values were 73.8%, 91.2%, 86.1%, and 82.5% for G. vaginalis; 84.2%, 96.3%, 84.2%, and 96.3% for Candida species; and 100%, 100%, 100%, and 100% for T. vaginalis, respectively. Cohen κ agreement between office-based diagnostic tests and Affirm-R was 0.16 (p=.141) for G. vaginalis, 0.46 (p<.0001) for Candida species, and 0.55 (p<.0001) for T. vaginalis. CONCLUSIONS: The Affirm VP-III sample collected from the residual vaginal discharge found on the speculum after performing office-based diagnostic tests can produce comparable results to traditionally collected sample.

SARS-CoV-2 pandemic non-pharmacologic interventions temporally associated with reduced pediatric infections due to Mycoplasma pneumoniae and co-infecting respiratory viruses in Arkansas.

Non-pharmacologic interventions (NPIs), such as universal masking, implemented during the SARS-CoV-2 pandemic have reduced respiratory infections among children. This study evaluated the impact of NPIs on Mycoplasma pneumoniae infections in children, analyzing data from two hospitals in Arkansas and examining age-related differences and co-infections with other respiratory viruses. The study was approved by the Institutional Review Board and included patients (≤18 years) with upper respiratory tract symptoms. Data generated from the FilmArray Respiratory Panel were divided into pre-NPI, NPI, and post-NPI periods for analysis. Overall test positivity rate and positivity rate interval changes were evaluated. Statistical differences were determined by Chi-square (χ2 independence) analysis. A total of 100,077 tests were performed, with a statistical increase in testing volume during the NPI and post-NPI periods. The number of positive M. pneumoniae tests decreased by 77% (77 to 18) during the NPI period, then increased by 50% (18 to 27) during the post-NPI period. Preschool and elementary school age groups had the highest number of positive tests during the study at 59 (48%) and 40 (33%), respectively. Reduced M. pneumoniae infections were consistent across age groups. Co-infections with other respiratory viruses, particularly human rhinovirus/enterovirus, were observed at much lower levels. Pediatric M. pneumoniae infections in Arkansas were temporally associated with implementation and discontinuation of NPIs. Specific viral co-infections still occurred, albeit at lower levels during the SARS-CoV-2 pandemic. Because of the slower growth of this bacterium, we expect M. pneumoniae infections to return to pre-pandemic levels within approximately 2 years. IMPORTANCE: Non-pharmacologic interventions (NPIs) effectively curtailed the spread of SARS-CoV-2 and, fortuitously, many other aerosol-transmitted respiratory pathogens. This study included the largest data set of symptomatic, pediatric patients from within the United States spanning a period from November 2017 through December 2023, and encompassed individuals residing in both rural and urban settings. We observed a strong correlation between the implementation and cessation of NPIs with the rate of respiratory infections due to Mycoplasma pneumoniae and viral co-infections. These infections are returning to baseline levels approximately 2 years following NPI cessation. This observation was not unexpected since the replication time for viruses is exponentially faster than that of bacteria. The resurgence of M. pneumoniae and likely other atypical bacterial pathogens is currently in process. Healthcare providers should strongly consider these pathogens in individuals presenting with respiratory tract illnesses.

NR3C2 microdeletions-an underrecognized cause of pseudohypoaldosteronism type 1A: a case report and literature review.

OBJECTIVES: Pseudohypoaldosteronism type 1A (PHA1A) is caused by haploinsufficiency of the mineralocorticoid receptor (MR). Heterozygous small insertions/deletions, transitions, and/or transversions within NR3C2 comprise the majority (85%-90%) of pathogenic copy number variants. Structural chromosomal abnormalities, contiguous gene deletion syndromes, and microdeletions are infrequent. We describe a neonate with PHA1A due to a novel NR3C2 microdeletion involving exons 1-2. METHODS: Literature review identified 39 individuals with PHA1A due to NR3C2 microdeletions. Transmission modality, variant description(s), testing method(s), exon(s) deleted, and affected functional domain(s) were characterized. RESULTS: In total, 40 individuals with NR3C2 microdeletions were described: 19 involved contiguous exons encoding a single MR domain; 21 involved contiguous exons encoding multiple MR domains. Transmission modality frequency was familial (65%), de novo (20%), or unknown (15%). Sequencing (Sanger or short-read next-generation) failed to detect microdeletions in 100% of tested individuals (n = 38). All were detected using deletion/duplication testing modalities. In 2 individuals, only microarray-based testing was performed; microdeletions were detected in both cases. CONCLUSION: Initial testing for PHA1A should rely on sequencing to detect the most common genetic alterations. Deletion/duplication analysis should be performed when initial testing is nondiagnostic. Most NR3C2 microdeletions are parentally transmitted, thus highlighting the importance of familial genetic testing and counseling.

SARS-CoV-2 Pandemic Non-Pharmacologic Interventions Temporally Associated with Reduced Pediatric Infections Due to Mycoplasma pneumoniae and Co-Infecting Respiratory Viruses in Arkansas.

Introduction: Non-pharmacologic interventions (NPIs), such as universal masking, implemented during the SARS-CoV-2 pandemic have reduced respiratory infections among children. This study focuses on evaluating the impact of NPIs on Mycoplasma pneumoniae infections in children, analyzing data from two hospitals in Arkansas, and examining age-related differences and coinfections with other viruses. Methods: The study was approved by the Institutional Review Board and included patients aged ≤18 years with upper respiratory tract symptoms. Data from the FilmArray® Respiratory Panel (FARP) were collected and divided into pre-NPI and NPI periods for analysis. Total test positivity rate and interval change in the positivity rate were evaluated. Statistical differences were determined by Chi-square (χ2-independence) analysis. Results: A total of 68,949 tests were performed with a statistical increase in testing during the NPI period. The overall test positivity rate for M. pneumoniae decreased by 74% (0.86% to 0.03%) during the NPI period, and the preschool age group had the highest number of positive tests in the pre- and NPI periods (Pre-NPI: n=40, NPI: n=12 positive tests, p=<0.001). The reduction in M. pneumoniae infections was consistent across age groups. Coinfections with other respiratory viruses, particularly human rhinovirus/enterovirus, were observed at much lower levels. Conclusions: NPIs effectively reduced M. pneumoniae in pediatric patients in Arkansas, and coinfections with specific viruses still occurred, albeit at lower levels during the SARS-CoV-2 pandemic. As NPIs are relaxed and the pandemic ends, we expect M. pneumoniae infections to return to pre-pandemic levels within the next 1-2 years.

Severe Acute Respiratory Syndrome Coronavirus-2 Lambda Variant Collected from a Child from Arkansas and Sequenced.

An eleven-year-old tested positive for SARS-CoV-2 Lambda variant. Sequencing was performed on the Oxford Nanopore and the Illumina NextSeq 500. Both platforms identified all 7 of the synonymous mutations in the sample, while all 28 nonsynonymous mutations were identified from Oxford Nanopore and 20 nonsynonymous mutations were identified from Illumina.

Loop-mediated isothermal amplification compared to real-time PCR and enzyme immunoassay for toxigenic Clostridium difficile detection.

Clostridium difficile infection is the primary cause of health care-associated diarrhea. While most laboratories have been using rapid antigen tests for detecting C. difficile toxins, they have poor sensitivity; newer molecular methods offer rapid results with high test sensitivity and specificity. This study was designed to compare the performances of two molecular assays (Meridian illumigene and BD GeneOhm) and two antigen assays (Wampole Quik Chek Complete and TechLab Tox A/B II) to detect toxigenic C. difficile. Fecal specimens from hospitalized patients (n = 139) suspected of having C. difficile infection were tested by the four assays. Nine specimens were positive and 109 were negative by all four methods. After discrepant analysis by toxigenic culture (n = 21), the total numbers of stool specimens classified as positive and negative for toxigenic C. difficile were 21 (15%) and 118 (85%), respectively. The sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) were as follows: GeneOhm (95.2%, 100%, 100%, and 99.2%), illumigene (95.2%, 96.6%, 83.3%, and 99.2%), Tox A/B II (52.4%, 97.5%, 78.6%, and 92.4%), and Quik Chek Complete (47.6%, 100%, 100%, and 91.9%). The illumigene assay performed comparably to the GeneOhm assay with a slight decrease in test specificity; the sensitivities of both far exceeded those of the antigen assays. The clinical characteristics of the concordant and discrepant study patients were similar, including stool consistency and frequency. In the era of rapid molecular-based tests for toxigenic C. difficile, toxin enzyme immunoassays (EIAs) should no longer be considered the standard of care.

Seroprevalence of SARS-CoV-2 antibodies in front-line pediatric health care workers.

Objective: The goal of this study was to determine the prevalence of SARS-CoV-2 infections in pediatric front-line health care workers (HCWs) using SARS-CoV-2 serum antibodies as an indicator of infection. Methods: In this cross-sectional study, we collected blood samples and survey responses from HCWs in a 38-bed pediatric emergency department. Serum antibodies to SARS-CoV-2 (IgM and/or IgG) were measured using a 2-step enzyme-linked immunosorbent assay (ELISA) to detect antibodies against the Spike protein receptor binding domain (RBD), the ectodomain of Spike (S), and the nucleoprotein (N). Results: We collected survey responses and serum samples from 54 pediatric front-line HCWs from October 2020 through April 2021. Among the 29 unvaccinated HCWs, 4 (13.7%) had antibodies to SARS-CoV-2. For the 25 vaccinated HCWs, 10 (40%) were seropositive; 3 were <10 days from the first vaccine dose and 7 were ≥10 days after the first dose. Two of the 10 seropositive vaccines had a prior positive reverse transcription polymerase chain reaction test. Individuals ≥10 days from receiving the first vaccine dose were 37.5 (95% CI: 3.5-399.3) times more likely to have SARS-CoV-2 antibodies than unvaccinated individuals or those <10 days from first vaccine dose. Conclusions: Evidence of widespread SARS-CoV-2 infections was not found in unvaccinated front-line HCWs from a pediatric ED as of April 2021. Future work will be required to determine the reasons underlying the lower SARS-CoV-2 antibody prevalence compared to adult HCWs.

Pediatric SARS-CoV-2 Seroprevalence in Arkansas Over the First Year of the COVID-19 Pandemic.

BACKGROUND: Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) seroprevalence studies largely focus on adults, but little is known about spread in children. We determined SARS-CoV-2 seroprevalence in children and adolescents from Arkansas over the first year of the coronavirus disease of 2019 (COVID-19) pandemic. METHODS: We tested remnant serum samples from children ages 1-18 years who visited Arkansas hospitals or clinics for non-COVID-19-related reasons from April 2020 through April 2021 for SARS-CoV-2 antibodies. We used univariable and multivariable regression models to determine the association between seropositivity and participant characteristics. RESULTS: Among 2357 participants, seroprevalence rose from 7.9% in April/May 2020 (95% CI, 4.9-10.9) to 25.0% in April 2021 (95% CI, 21.5-28.5). Hispanic and black children had a higher association with antibody positivity than non-Hispanic and white children, respectively, in multiple sampling periods. CONCLUSIONS: By spring 2021, most children in Arkansas were not infected with SARS-CoV-2. With the emergence of SARS-CoV-2 variants, recognition of long-term effects of COVID-19, and the lack of an authorized pediatric SARS-CoV-2 vaccine at the time, these results highlight the importance of including children in SARS-CoV-2 public health, clinical care, and research strategies.

Rapid stool-based diagnosis of Clostridium difficile infection by real-time PCR in a children's hospital.

Clostridium difficile is a major cause of nosocomial antibiotic-associated infectious diarrhea and pseudomembranous colitis. Detection of C. difficile by anaerobic bacterial culture and/or cytotoxicity assays has been largely replaced by rapid enzyme immunoassays (EIA). However, due to the lack of sensitivity of stool EIA, we developed a multiplex real-time PCR assay targeting the C. difficile toxin genes tcdA and tcdB. Stool samples from hospitalized pediatric patients suspected of having C. difficile-associated disease were prospectively cultured on cycloserine-cefoxitin-fructose agar following alcohol shock. Six testing modalities were evaluated, including stool EIA, culture EIA, and real-time PCR (tcdA and tcdB) of cultured isolates and stool samples. Real-time PCR detection was performed with tcdA and tcdB gene-specific primers and hydrolysis probes using the LightCycler platforms (Roche Diagnostics, Indianapolis, IN). A total of 157 samples from 96 pediatric patients were analyzed. The sensitivities of stool real-time PCR and stool EIA were 95% and 35%, respectively, with a specificity of 100% for both methods. The lower limit of detection of the stool real-time PCR was 30 CFU/ml of stool sample per reaction for tcdA and tcdB. This study highlights the poor performance of stool toxin EIAs in pediatric settings. Direct detection of C. difficile toxin genes in stool samples by real-time PCR showed sensitivity superior to that of stool and culture EIAs and performance comparable to that of real-time PCR assay of cultured isolates. Real-time PCR of DNA from stool samples is a rapid and cost-effective diagnostic modality for children that should facilitate appropriate patient management and halt the practice of serial testing by EIA.

Native vs Prosthetic Valve Histoplasma capsulatum Infective Endocarditis: A Case Report and Systemic Literature Review Comparing Patient Presentation, Treatment Modalities, Clinical Outcomes, and Diagnostic Laboratory Testing.

Histoplasma capsulatum is a rare cause of fungal endocarditis that affects both native and prosthetic valves. It is associated with a high mortality rate if not diagnosed early and treated with a combination of antifungal therapy and surgical intervention. We present a case of a 47-year-old man with histoplasmosis infective endocarditis. He was successfully treated with antifungal therapy and surgical replacement of the infected bioprosthetic aortic valve. Our systemic literature review includes 52 articles encompassing 60 individual cases of H. capsulatum infective endocarditis from 1940 to 2020. Patient presentations, diagnostic laboratory testing accuracy, treatment modalities, and patient outcomes comparing and contrasting native and prosthetic valve infection are described.

Implementing Pharmacogenomics Testing: Single Center Experience at Arkansas Children's Hospital.

Pharmacogenomics (PGx) is a growing field within precision medicine. Testing can help predict adverse events and sub-therapeutic response risks of certain medications. To date, the US FDA lists over 280 drugs which provide biomarker-based dosing guidance for adults and children. At Arkansas Children's Hospital (ACH), a clinical PGx laboratory-based test was developed and implemented to provide guidance on 66 pediatric medications for genotype-guided dosing. This PGx test consists of 174 single nucleotide polymorphisms (SNPs) targeting 23 clinically actionable PGx genes or gene variants. Individual genotypes are processed to provide per-gene discrete results in star-allele and phenotype format. These results are then integrated into EPIC- EHR. Genomic indicators built into EPIC-EHR provide the source for clinical decision support (CDS) for clinicians, providing genotype-guided dosing.

Detection of Respiratory Pathogens Does Not Predict Risks After Outpatient Adenotonsillectomy.

OBJECTIVES/HYPOTHESIS: To determine whether the presence of detectable upper respiratory infections (URIs) at the time of adenoidectomy/adenotonsillectomy is associated with increased morbidity, complications, and unexpected admissions. STUDY DESIGN: Prospective double-blinded cohort. METHODS: In this prospective cohort study, nasopharyngeal swabs were obtained intraoperatively from 164 pediatric patients undergoing outpatient adenoidectomy/tonsillectomy with or without pressure equalization tubes (PETs) and were analyzed with PCR for the presence of 22 known URIs, including SARS-CoV-2. Surgeons and families were blinded to the results. At the conclusion of the study, rates of detectable infection were determined and intraoperative and postoperative events (unexpected admissions, length of PACU stay, rates of laryngospasm/bronchospasm, oxygen desaturation, bradycardia, and postoperative presentation to an emergency department) were compared between infected and uninfected patients. RESULTS: Of the 164 patients (50% male, 50% female, ages 8 mo-18 y), 136 patients (82.9%) tested positive for one or more URI at the time of surgery. Forty one patients (25.0%) tested positive for three or more URIs concurrently, and 11 (6.7%) tested positive for five or more URIs concurrently. There were no significant differences in admission rates, length of PACU stay, rates of laryngospasm/bronchospasm, oxygen desaturation, bradycardia, or postoperative presentation to an emergency department between positive and negative patients. No patients tested positive for SARS-CoV-2. CONCLUSIONS: A recent positive URI test does not confer any additional intraoperative or postoperative risk in the setting of outpatient adenoidectomy/tonsillectomy in healthy patients. There is no utility in preoperative URI testing, and delaying surgery due to a recent positive URI test is not warranted in this population. LEVEL OF EVIDENCE: 3 Laryngoscope, 131:E2074-E2079, 2021.