Utility of the Signal-to-Cutoff Ratio and Antigen Index from Fourth- and Fifth-Generation HIV-1/HIV-2 Antigen/Antibody Assays for the Diagnosis of Acute HIV Infection: Applicability for Real-Time Use for Immediate Initiation of Antiretroviral Therapy.

Identification of individuals with acute HIV infection (AHI) and rapid initiation of antiretroviral therapy (ART) are priorities for HIV elimination efforts. Fourth- and fifth-generation HIV-1/HIV-2 antigen (Ag)/antibody (Ab) combination assays can quickly identify patients with AHI, but false-positive results can occur. Confirmatory nucleic acid amplification testing (NAAT) may not be rapidly available. We reviewed the data for 127 patients with positive fourth-generation ARCHITECT and fifth-generation Bio-Plex immunoassay results who had negative or indeterminate confirmatory Ab testing results, which yielded 38 patients with confirmed AHI and 89 patients with false-positive results. The receiver operating characteristic (ROC) curves showed excellent discriminatory power, with an area under the curve (AUC) for the signal-to-cutoff (S/CO) ratio of 0.970 (95% confidence interval [CI], 0.935 to 1.00) and an AUC for the Ag index (AI) of 0.968 (95% CI, 0.904 to 1.00). A threshold of 3.78 for the S/CO ratio would maximize the sensitivity (96.3%) and specificity (93.4%). The threshold for AI was 2.83 (sensitivity of 100% and specificity of 96.4%). The S/CO ratio was significantly correlated with the viral load (Spearman correlation coefficient, 0.486 [P = 0.014]), but the AI was not. The viral loads were all high, with a median of >2.8 million copies/mL. Two false-positive results with AI and S/CO ratio values markedly higher than the medians were observed, indicating that biological false-positive results can occur. Review of the S/CO ratio or AI may be used to improve the accuracy of AHI diagnosis prior to confirmatory NAAT results being available.

Clinical Performance of the Point-of-Care cobas Liat for Detection of SARS-CoV-2 in 20 Minutes: a Multicenter Study.

Highly accurate testing for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) at the point of care (POC) is an unmet diagnostic need in emergency care and time-sensitive outpatient care settings. Reverse transcription-PCR (RT-PCR) technology is the gold standard for SARS-CoV-2 diagnostics. We performed a multisite U.S. study comparing the clinical performance of the first U.S. Food and Drug Administration (FDA)-authorized POC RT-PCR for detection of SARS-CoV-2 in 20 min, the cobas Liat SARS-CoV-2 and influenza A/B nucleic acid test, to the most widely used RT-PCR laboratory test, the cobas 68/8800 SARS-CoV-2 test. Clinical nasopharyngeal swab specimens from 444 patients with 357 evaluable specimens at five U.S. clinical laboratories were enrolled from 21 September 2020 to 23 October 2020. The overall agreement between the Liat and 68/8800 systems for SARS-CoV-2 diagnostics was 98.6% (352/357). Using Liat, positive percent agreement for SARS-CoV-2 was 100% (162/162) and the negative percent agreement was 97.4% (190/195). The Liat is an RT-PCR POC test that provides highly accurate SARS-CoV-2 results in 20 min with performance equivalent to that of high-throughput laboratory molecular testing. Rapid RT-PCR testing at the POC can enable more timely infection control and individual care decisions for coronavirus disease 2019.

Misdiagnosis of Bordetella bronchiseptica Respiratory Infection as Bordetella pertussis by Multiplex Molecular Assay.

Multiplex polymerase chain reaction (PCR) tests are useful for the rapid detection of pathogens, though diagnostic challenges may arise. We report 2 immunocompromised patients with Bordetella bronchiseptica respiratory infection misdiagnosed as Bordetella pertussis using PCR, including discussion of transmission, diagnostic testing, clinical implications, and infection control considerations.

Use of the Accelerate Pheno System for Identification and Antimicrobial Susceptibility Testing of Pathogens in Positive Blood Cultures and Impact on Time to Results and Workflow.

The Accelerate Pheno system uses automated fluorescence in situ hybridization technology with morphokinetic cellular analysis to provide rapid species identification (ID) and antimicrobial susceptibility testing (AST) results for the most commonly identified organisms in bloodstream infections. The objective was to evaluate the accuracy and workflow of bacterial and yeast ID and bacterial AST using the Accelerate Pheno system in the clinical microbiology laboratory. The consecutive fresh blood cultures received in the laboratory were analyzed by the Accelerate Pheno system within 0 to 8 h of growth detection. ID/AST performance, the average times to results, and workflow were compared to those of the routine standard of care. Of the 232 blood cultures evaluated (223 monomicrobial and 9 polymicrobial) comprising 241 organisms, the overall sensitivity and specificity for the identification of organisms were 95.6% and 99.5%, respectively. For antimicrobial susceptibility, the overall essential agreement was 95.1% and categorical agreement was 95.5% compared to routine methods. There was one very major error and 3 major errors. The time to identification and the time to susceptibility using the Accelerate Pheno system were decreased by 23.47 and 41.86 h, respectively, compared to those for the standard of care. The reduction in hands on time was 25.5 min per culture. The Accelerate Pheno system provides rapid and accurate ID/AST results for most of the organisms found routinely in blood cultures. It is easy to use, reduces hands on time for ID/AST of common blood pathogens, and enables clinically actionable results to be released much earlier than with the current standard of care.

BioFire FilmArray Respiratory Panel for Detection of Enterovirus D68.

During the enterovirus D68 (EV-D68) outbreak of 2014, the BioFire FilmArray (FA) respiratory panel was used to detect rhinovirus/enterovirus in respiratory specimens; suspected EV-D68-positive specimens were sent to CDC for confirmation. Positive rhinovirus/enterovirus FA targets revealed patterns loosely associated with EV-D68 that may be useful for confirmation triaging.

Comparison of Cepheid Xpert Flu/RSV XC and BioFire FilmArray for Detection of Influenza A, Influenza B, and Respiratory Syncytial Virus.

The Xpert Flu/RSV XC was compared to the FilmArray respiratory panel for detection of influenza (Flu) A, Flu B, and respiratory syncytial virus (RSV), using 128 nasopharyngeal swabs. Positive agreements were 100% for Flu A and RSV and 92.3% for Flu B. The Xpert may be useful in clinical situations when extensive testing is not required and may serve an important role in laboratories already performing broader respiratory panel testing.

Influenza among afebrile and vaccinated healthcare workers.

BACKGROUND: To prevent transmission of influenza from healthcare workers (HCWs) to patients, many hospitals exclude febrile HCWs from working, but allow afebrile HCWs with respiratory symptoms to have contact with patients. During the 2013-2014 influenza season at our hospital, an influenza-positive HCW with respiratory symptoms but no fever was linked to a case of possible healthcare-associated influenza in a patient. Therefore, we implemented a temporary policy of mandatory influenza testing for HCWs with respiratory symptoms. METHODS: From 3 January through 28 February 2014, we tested HCWs with respiratory symptoms for influenza and other respiratory pathogens by polymerase chain reaction of flocked nasopharyngeal swabs. HCWs also reported symptoms and influenza vaccination status, and underwent temperature measurement. We calculated the proportion of influenza-positive HCWs with fever and prior influenza vaccination. RESULTS: Of 449 HCWs, 243 (54%) had a positive test for any respiratory pathogen; 34 (7.6%) HCWs tested positive for influenza. An additional 7 HCWs were diagnosed with influenza by outside physicians. Twenty-one (51.2%) employees with influenza had fever. Among influenza-infected HCWs, 20 had previously received influenza vaccination, 18 had declined the vaccine, and 3 had unknown vaccination status. There was no significant difference in febrile disease among influenza-infected employees who had received the influenza vaccine and those who had not received the vaccine (45% vs 61%; P = .32). CONCLUSIONS: Nearly half of HCWs with influenza were afebrile prior to their diagnosis. HCWs with respiratory symptoms but no fever may pose a risk of influenza transmission to patients and coworkers.

Expect the unexpected: endocarditis caused by Legionella feeleii.

We report a fatal case of Legionella feeleii endocarditis in a post-lung transplant patient. The diagnosis was delayed, as routine microbiological testing of nonrespiratory specimens does not account for extrapulmonary Legionella, and urine antigen testing only reliably detects Legionella pneumophila serogroup 1. This case also illustrates the utility of molecular sequencing for blood culture-negative endocarditis.

Reflex Human Immunodeficiency Virus (HIV) Type 1 RNA Testing Enables Timely Differentiation of False-Positive Results From Acute HIV Infection.

Accurate, timely human immunodeficiency virus (HIV) diagnosis is critical. Routine HIV screening program data were examined before and after reflex HIV type 1 RNA testing. Reflex testing facilitated confirmation of reactive HIV screening assays (as true or false positives) (odds ratio, 23.7 [95% confidence interval, 6.7-83.4]; P < .0001), improving detection of acute HIV and reducing unconfirmed discordant results.

Novel paradigm enables accurate monthly gestational screening to prevent congenital toxoplasmosis and more.

BACKGROUND: Congenital toxoplasmosis is a treatable, preventable disease, but untreated causes death, prematurity, loss of sight, cognition and motor function, and substantial costs worldwide. OBJECTIVES: We asked whether high performance of an Immunochromatographic-test (ICT) could enable accurate, rapid diagnosis/treatment, establishing new, improved care-paradigms at point-of-care and clinical laboratory. METHODS: Data were obtained in 12 studies/analyses addressing: 1-feasibility/efficacy; 2-false-positives; 3-acceptability; 4-pink/black-line/all studies; 5-time/cost; 6-Quick-Information/Limit-of-detection; 7, 8-acute;-chronic; 9-epidemiology; 10-ADBio; 11,12-Commentary/Cases/Chronology. FINDINGS: ICT was compared with gold-standard or predicate-tests. Overall, ICT performance for 1093 blood/4967 sera was 99.2%/97.5% sensitive and 99.0%/99.7% specific. However, in clinical trial, FDA-cleared-predicate tests initially caused practical, costly problems due to false-positive-IgM results. For 58 persons, 3/43 seronegative and 2/15 chronically infected persons had false positive IgM predicate tests. This caused substantial anxiety, concerns, and required costly, delayed confirmation in reference centers. Absence of false positive ICT results contributes to solutions: Lyon and Paris France and USA Reference laboratories frequently receive sera with erroneously positive local laboratory IgM results impeding patient care. Therefore, thirty-two such sera referred to Lyon's Reference laboratory were ICT-tested. We collated these with other earlier/ongoing results: 132 of 137 USA or French persons had false-positive local laboratory IgM results identified correctly as negative by ICT. Five false positive ICT results in Tunisia and Marseille, France, emphasize need to confirm positive ICT results with Sabin-Feldman-Dye-test or western blot. Separate studies demonstrated high performance in detecting acute infections, meeting FDA, CLIA, WHO REASSURED, CEMark criteria and patient and physician satisfaction with monthly-gestational-ICT-screening. CONCLUSIONS/SIGNIFICANCE: This novel paradigm using ICT identifies likely false positives or raises suspicion that a result is truly positive, rapidly needing prompt follow up and treatment. Thus, ICT enables well-accepted gestational screening programs that facilitate rapid treatment saving lives, sight, cognition and motor function. This reduces anxiety, delays, work, and cost at point-of-care and clinical laboratories. TRIAL REGISTRATION: NCT04474132, https://clinicaltrials.gov/study/NCT04474132 ClinicalTrials.gov.

Community-associated methicillin-resistant Staphylococcus aureus colonization burden in HIV-infected patients.

BACKGROUND: The epidemic of community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) has had a disproportionate impact on patients with human immunodeficiency virus (HIV). METHODS: We evaluated CA-MRSA colonization burden (number of colonized sites per total number sampled) among HIV-infected and HIV-negative inpatients within 72 hours of hospitalization. From March 2011 through April 2012, we obtained cultures from nasal and extranasal sites (throat, axilla, inguinal, perirectal, and chronic wound if present) and collected risk factor data. RESULTS: Of 745 patients (374 HIV-infected, 371 HIV-negative), 15.7% were colonized with CA-MRSA at any site: 20% of HIV and 11% of HIV-negative patients (relative prevalence=1.8, P=.002). HIV-infected patients had a higher prevalence of nasal, extranasal, and exclusive extranasal colonization as well as higher colonization burden. Perirectal and inguinal areas were the extranasal sites most frequently colonized, and 38.5% of colonized patients had exclusive extranasal colonization. Seventy-three percent of isolates were identified as USA300. Among HIV-infected patients, male sex, younger age, and recent incarceration were positively associated whereas Hispanic ethnicity was negatively associated with higher colonization burden. Among HIV-negative patients, temporary housing (homeless, shelter, or substance abuse center) was the only factor associated with higher colonization burden. Predictors of USA300 included HIV, younger age, illicit drug use, and male sex; all but 1 colonized individual with current or recent incarceration carried USA300. CONCLUSIONS: HIV-infected patients were more likely to have a higher CA-MRSA colonization burden and carry USA300. In certain populations, enhanced community and outpatient-based infection control strategies may be needed to prevent CA-MRSA cross-transmission and infection.

Rapid, Sensitive, Label-Free Electrical Detection of SARS-CoV-2 in Nasal Swab Samples.

Rapid diagnosis of coronavirus disease 2019 (COVID-19) is key for the long-term control of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) amid renewed threats of mutated SARS-CoV-2 around the world. Here, we report on an electrical label-free detection of SARS-CoV-2 in nasopharyngeal swab samples directly collected from outpatients or in saliva-relevant conditions by using a remote floating-gate field-effect transistor (RFGFET) with a 2-dimensional reduced graphene oxide (rGO) sensing membrane. RFGFET sensors demonstrate rapid detection (<5 min), a 90.6% accuracy from 8 nasal swab samples measured by 4 different devices for each sample, and a coefficient of variation (CV) < 6%. Also, RFGFET sensors display a limit of detection (LOD) of pseudo-SARS-CoV-2 that is 10 000-fold lower than enzyme-linked immunosorbent assays, with a comparable LOD to that of reverse transcription-polymerase chain reaction (RT-PCR) for patient samples. To achieve this, comprehensive systematic studies were performed regarding interactions between SARS-CoV-2 and spike proteins, neutralizing antibodies, and angiotensin-converting enzyme 2, as either a biomarker (detection target) or a sensing probe (receptor) functionalized on the rGO sensing membrane. Taken together, this work may have an immense effect on positioning FET bioelectronics for rapid SARS-CoV-2 diagnostics.

Bifidobacteria metabolize lactulose to optimize gut metabolites and prevent systemic infection in patients with liver disease.

Progression of chronic liver disease is precipitated by hepatocyte loss, inflammation and fibrosis. This process results in the loss of critical hepatic functions, increasing morbidity and the risk of infection. Medical interventions that treat complications of hepatic failure, including antibiotic administration for systemic infections and lactulose treatment for hepatic encephalopathy, can impact gut microbiome composition and metabolite production. Here, using shotgun metagenomic sequencing and targeted metabolomic analyses on 847 faecal samples from 262 patients with acute or chronic liver disease, we demonstrate that patients hospitalized for liver disease have reduced microbiome diversity and a paucity of bioactive metabolites, including short-chain fatty acids and bile acid derivatives, that impact immune defences and epithelial barrier integrity. We find that patients treated with the orally administered but non-absorbable disaccharide lactulose have increased densities of intestinal bifidobacteria and reduced incidence of systemic infections and mortality. Bifidobacteria metabolize lactulose, produce high concentrations of acetate and acidify the gut lumen in humans and mice, which, in combination, can reduce the growth of antibiotic-resistant bacteria such as vancomycin-resistant Enterococcus faecium in vitro. Our studies suggest that lactulose and bifidobacteria serve as a synbiotic to reduce rates of infection in patients with severe liver disease.

Community-associated methicillin-resistant Staphylococcus aureus colonization in high-risk groups of HIV-infected patients.

BACKGROUND: We examined the epidemiology of community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) nasal colonization among 3 groups of human immunodeficiency virus (HIV)-infected and 1 group of HIV-negative outpatients. METHODS: We determined prevalence and risk factors associated with MRSA colonization among women, recently incarcerated, and Hispanic HIV-infected patients and HIV-negative patients; isolates were typed by pulsed-field gel electrophoresis. Relative prevalence was calculated using Poisson regression, and logistic regression was used for multivariate analysis. RESULTS: Of 601 patients, 9.3% were colonized with MRSA; 11% of HIV-infected and 4.2% of HIV-negative patients were colonized (relative prevalence, 2.6; 95% confidence interval [CI], 1.12-6.07; P = .03). Among HIV-infected patients, recently incarcerated patients had the highest colonization prevalence (15.6%) followed by women (12%); Hispanic patients had the lowest (2.8%). Eighty percent of confirmed MRSA isolates were identified as USA300. On multivariate analysis, history of incarceration or residence in alternative housing (odds ratio [OR], 2.3; 95% CI, 1.1-4.7; P = .03) was associated with MRSA colonization; Hispanic ethnicity was negatively associated (OR, 0.3; 95% CI, .11-.98; P = .045). There was a trend (OR, 1.6; 95% CI, .9-3.0; P = .097) toward geographic location of residence being associated with colonization. After controlling for incarceration, residence, and geography, HIV status was no longer significantly associated with colonization. CONCLUSIONS: The CA-MRSA and HIV epidemics have intersected. Examination of networks of individuals released from incarceration, both HIV positive and negative, is needed to assess the role of social networks in spread of CA-MRSA and inform prevention strategies.

Remote Floating-Gate Field-Effect Transistor with 2-Dimensional Reduced Graphene Oxide Sensing Layer for Reliable Detection of SARS-CoV-2 Spike Proteins.

Despite intensive research of nanomaterials-based field-effect transistors (FETs) as a rapid diagnostic tool, it remains to be seen for FET sensors to be used for clinical applications due to a lack of stability, reliability, reproducibility, and scalability for mass production. Herein, we propose a remote floating-gate (RFG) FET configuration to eliminate device-to-device variations of two-dimensional reduced graphene oxide (rGO) sensing surfaces and most of the instability at the solution interface. Also, critical mechanistic factors behind the electrochemical instability of rGO such as severe drift and hysteresis were identified through extensive studies on rGO-solution interfaces varied by rGO thickness, coverage, and reduction temperature. rGO surfaces in our RFGFET structure displayed a Nernstian response of 54 mV/pH (from pH 2 to 11) with a 90% yield (9 samples out of total 10), coefficient of variation (CV) < 3%, and a low drift rate of 2%, all of which were calculated from the absolute measurement values. As proof-of-concept, we demonstrated highly reliable, reproducible, and label-free detection of spike proteins of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in a saliva-relevant media with concentrations ranging from 500 fg/mL to 5 µg/mL, with an R2 value of 0.984 and CV < 3%, and a guaranteed limit of detection at a few pg/mL. Taken together, this new platform may have an immense effect on positioning FET bioelectronics in a clinical setting for detecting SARS-CoV-2.

Routine, rapid HIV testing of medicine service admissions in the emergency department.

OBJECTIVE: We identify undiagnosed HIV among adult emergency department (ED) patients awaiting medicine admission through rapid testing, expedite their redirection to the inpatient HIV service, and improve linkage to ambulatory HIV care. METHODS: Two ED health educators offered rapid testing to patients aged 18 to 64 years from the high-acuity ED area from which most medicine admissions originate. Heath educators obtained consent, obtained fingerstick blood, and performed point-of-care testing. Patients with reactive results received counseling, confirmatory testing, and appointments at the affiliated HIV clinic. RESULTS: Between March 1, 2008, and February 28, 2009, 4,755 patients received testing. Thirty patients (0.6%) had received a new diagnosis of HIV; 26 were admitted and redirected to the HIV service. Characteristics of HIV positive patients were mean age 38 years, 87% men, 64% black, and 33% Hispanic; 76% had CD4 counts less than 200 cells/mm(3); 67% had HIV-related diagnoses; and 93% reported for ambulatory HIV care in a median of 10 days. During 2 preceding years, these patients had a mean of 3 previous health system visits without testing. During a 6-month quality assurance interval of the 5,340 ED medicine admissions, 31% of patients were eligible for testing, of whom 88% received testing (1% positive) and 12% declined; 29% of the 5,340 were not approached for testing; and 40% were deemed ineligible. Common reasons for ineligibility included older age, recent previous test, and known HIV-positive status. CONCLUSION: Patients who receive a diagnosis of HIV in our ED before admission are extremely ill, most having AIDS. Targeted HIV screening of ED patients awaiting hospital admission facilitated timely diagnosis and reliable linkage to inpatient and outpatient HIV care.

Cording in Disseminated Mycobacterium chelonae Infection in an Immunocompromised Patient.

Cording is a phenomenon in which acid fast bacilli grow in parallel and was previously used as a means of presumptive microscopic identification of Mycobacterium tuberculosis (TB). However, this process has been shown in multiple other nontuberculous mycobacterial (NTM) species. Here we present the case of an immunocompromised adult who presented with wrist pain, weight loss, and cough. A positron emission tomography scan showed uptake in the right ulna, multiple soft tissue sites, and the left lung. Biopsies and cultures were obtained from multiple sites, and the patient was ultimately diagnosed with disseminated Mycobacterium chelonae infection. The organism showed cording in culture. As seen in this patient, cording may occur in multiple NTM species and is not reliable as the sole indicator of the presence of TB.

Implementation of a Sample Pooling Strategy for the Direct Detection of SARS-CoV-2 by Real-Time Polymerase Chain Reaction During the COVID-19 Pandemic.

OBJECTIVES: To report our institutional experience in devising and implementing a pooling protocol and process for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) reverse transcription polymerase chain reaction (RT-PCR) testing over a 3-month period in the fall of 2020. METHODS: The widespread testing implemented in the United States for detecting SARS-CoV-2 infection in response to the coronavirus disease 2019 pandemic has led to a significant shortage of testing supplies and therefore has become a major impediment to the public health response. To date, several institutions have implemented sample pooling, but publications documenting these experiences are sparse. Nasal and nasopharyngeal samples collected from low-positivity (<5%) areas were tested in pools of five on the Roche cobas 6800 analyzer system. Routine SARS-CoV-2 RT-PCR turnaround times between sample collection to result reporting were monitored and compared before and after sample pooling implementation. RESULTS: A total of 4,131 sample pools were tested over a 3-month period (during which 39,770 RT-PCR results were reported from the Roche system), allowing our laboratory to save 13,824 tests, equivalent to a conservation rate of 35%. A 48-hour or less turnaround time was generally maintained throughout the pooling period. CONCLUSIONS: Sample pooling offers a viable means to mitigate shortfalls of PCR testing supplies in the ongoing pandemic without significantly compromising overall turnaround times.

Universal screening for Clostridioides difficile at an urban academic medical center.

We implemented universal inpatient Clostridioides difficile screening at an 800-bed hospital. Over 3 years, 2,010 of 47,048 screening tests (4.2%) were positive, with significantly higher rates of C. difficile colonization on transplant units than medical-surgical units: 5.4% (152 of 2,801) versus 4.3% (880 of 20,564), respectively (P = .005). Compliance with screening ranged from 79% to 96%.