Evaluation of an emergency department opt-out provider-driven HIV and syphilis screening and linkage-to-care program.

OBJECTIVE: While the effectiveness of emergency departments (ED) in screening for HIV and syphilis is understood, less is known about dual screening programs. We aim to evaluate the impact of an opt-out provider-initiated HIV and syphilis program on screening, diagnosis, and linkage to care outcomes. METHODS: We performed a retrospective review of patients screened pre (2014-2017) and post (2017-2021) program implementation. Primary outcomes include HIV and syphilis screening, incidence of positive tests, and proportion of patients linked to care. Secondary outcomes included pre-exposure prophylaxis (PrEP) referral and successful linkage rates for HIV-negative syphilis-positive patients. RESULTS: Pre-implementation, 882 HIV tests were performed, of which 22 (2.49%) were new cases and 18 (81.82%) were linked to care; 754 syphilis tests were performed, of which 33 (4.38%) were active infections and 30 (90.91%) were treated. No eligible patients received PrEP referral. Post-implementation, 12,999 HIV tests were performed, of which 73 (0.56%) were new cases and 55 (75.34%) were linked to care; 10,885 syphilis tests were performed, of which 216 (1.98%) were active infections and 188 (87.04%) were treated. 25 (9.09%) eligible patients were referred for PrEP, and four (16.0%) attended their appointment. CONCLUSIONS: Post-implementation, there was a 1373.81% and 1343.63% increase in screening, and a 231.82% and 554.55% increase in positive cases of HIV and syphilis, respectively. Dual screening programs can be successfully implemented within the existing ED framework to increase screening and early detection for HIV and syphilis.

Evaluation of Quantitative Real-Time PCR as a Hepatitis C Virus Supplementary Test After RIBA Discontinuation.

Laboratory testing plays a major role in hepatitis C virus (HCV) diagnosis and patient follow-up. The high false positive rates of HCV screening tests require confirmation through a supplementary test. According to the 2003 CDC guidelines, recombinant immunoblot assay (RIBA) is indispensible to confirm positive screening results and differentiate biologic false positivity from true HCV exposure. However, RIBA has been permanently discontinued since 2011. In the 2013 update of its guidelines, CDC called for further studies on HCV laboratory testing without RIBA. In this study, we analyzed the applicability of quantitative real-time PCR (qPCR) as a supplementary HCV diagnostic test. By comparing our HCV testing performances before and after RIBA discontinuation, we found that omitting RIBA has no significant effect on the accurate and efficient identification of HCV infection, provided that HCV antibody signal-to-cutoff ratio is considered. Furthermore, we proposed a new HCV testing algorithm that incorporates semiquantitative assessment of HCV antibody positivity and HCV viral load measurement by qPCR. By following the algorithm, we were able to address confirmation of positive HCV screening results and to provide useful information generally required by clinicians, including the needs of further laboratory testing or clinical follow-up, as well as HCV viral titers.

Translational control during endoplasmic reticulum stress beyond phosphorylation of the translation initiation factor eIF2α.

The accumulation of unfolded/misfolded proteins in the endoplasmic reticulum (ER) causes stress to which an unfolded protein response is activated to render cell survival or apoptosis (chronic stress). Transcriptional and translational reprogramming is tightly regulated during the unfolded protein response to ensure specific gene expression. The master regulator of this response is the PERK/eIF2α/ATF4 signaling where eIF2α is phosphorylated (eIF2α-P) by the kinase PERK. This signal leads to global translational shutdown, but it also enables translation of the transcription factor ATF4 mRNA. We showed recently that ATF4 induces an anabolic program through the up-regulation of selected amino acid transporters and aminoacyl-tRNA synthetases. Paradoxically, this anabolic program led cells to apoptosis during chronic ER stress in a manner that involved recovery from stress-induced protein synthesis inhibition. By using eIF2α-P-deficient cells as an experimental system, we identified a communicating network of signaling pathways that contribute to the inhibition of protein synthesis during chronic ER stress. This eIF2α-P-independent network includes (i) inhibition of mammalian target of rapamycin kinase protein complex 1 (mTORC1)-targeted protein phosphorylation, (ii) inhibited translation of a selective group of 5'-terminal oligopyrimidine mRNAs (encoding proteins involved in the translation machinery and translationally controlled by mTORC1 signaling), and (iii) inhibited translation of non-5'-terminal oligopyrimidine ribosomal protein mRNAs and ribosomal RNA biogenesis. We propose that the PERK/eIF2α-P/ATF4 signaling acts as a brake in the decline of protein synthesis during chronic ER stress by positively regulating signaling downstream of the mTORC1 activity. These studies advance our knowledge on the complexity of the communicating signaling pathways in controlling protein synthesis rates during chronic stress.

Sensitive deep-sequencing-based HIV-1 genotyping assay to simultaneously determine susceptibility to protease, reverse transcriptase, integrase, and maturation inhibitors, as well as HIV-1 coreceptor tropism.

With 29 individual antiretroviral drugs available from six classes that are approved for the treatment of HIV-1 infection, a combination of different phenotypic and genotypic tests is currently needed to monitor HIV-infected individuals. In this study, we developed a novel HIV-1 genotypic assay based on deep sequencing (DeepGen HIV) to simultaneously assess HIV-1 susceptibilities to all drugs targeting the three viral enzymes and to predict HIV-1 coreceptor tropism. Patient-derived gag-p2/NCp7/p1/p6/pol-PR/RT/IN- and env-C2V3 PCR products were sequenced using the Ion Torrent Personal Genome Machine. Reads spanning the 3' end of the Gag, protease (PR), reverse transcriptase (RT), integrase (IN), and V3 regions were extracted, truncated, translated, and assembled for genotype and HIV-1 coreceptor tropism determination. DeepGen HIV consistently detected both minority drug-resistant viruses and non-R5 HIV-1 variants from clinical specimens with viral loads of ≥1,000 copies/ml and from B and non-B subtypes. Additional mutations associated with resistance to PR, RT, and IN inhibitors, previously undetected by standard (Sanger) population sequencing, were reliably identified at frequencies as low as 1%. DeepGen HIV results correlated with phenotypic (original Trofile, 92%; enhanced-sensitivity Trofile assay [ESTA], 80%; TROCAI, 81%; and VeriTrop, 80%) and genotypic (population sequencing/Geno2Pheno with a 10% false-positive rate [FPR], 84%) HIV-1 tropism test results. DeepGen HIV (83%) and Trofile (85%) showed similar concordances with the clinical response following an 8-day course of maraviroc monotherapy (MCT). In summary, this novel all-inclusive HIV-1 genotypic and coreceptor tropism assay, based on deep sequencing of the PR, RT, IN, and V3 regions, permits simultaneous multiplex detection of low-level drug-resistant and/or non-R5 viruses in up to 96 clinical samples. This comprehensive test, the first of its class, will be instrumental in the development of new antiretroviral drugs and, more importantly, will aid in the treatment and management of HIV-infected individuals.

SARS-CoV-2 Seroprevalence Among First Responders in Northeastern Ohio, 2020.

OBJECTIVES: First responders, including firefighters, emergency medical technicians (EMTs), paramedics, and law enforcement officers, are working on the front lines to fight the COVID-19 pandemic and facing an increased risk of infection. This study assessed the seroprevalence of SARS-CoV-2 infection among first responders in northeastern Ohio. METHODS: A survey and immunoglobulin G antibody test against SARS-CoV-2 nucleocapsid protein were offered to University Hospitals Health System-affiliated first-responder departments during May to September 2020. The survey contained questions about demographic characteristics and history of SARS-CoV-2 infection. A total of 3080 first responders with diverse job assignments from more than 400 fire and police departments participated in the study. RESULTS: Of 3080 participants, 73 (2.4%) were seropositive and 26 (0.8%) had previously positive real-time polymerase chain reaction results. Asymptomatic infection accounted for 46.6% (34 of 73) of seropositivity. By occupation, rates of seropositivity were highest among administration/support staff (3.8%), followed by paramedics (3.0%), EMTs (2.6%), firefighters (2.2%), and law enforcement officers (0.8%). Work-associated exposure rates to COVID-19 patients were: paramedics (48.2%), firefighters (37.1%), EMTs (32.3%), law enforcement officers (7.7%), and administration/support staff (4.4%). Self-reported community exposure was positively correlated with self-reported work-associated exposure rate (correlation coefficient = 0.99). Neither self-reported community nor work-associated exposure was correlated with SARS-CoV-2 seroprevalence. We found no significant difference in seroprevalence among sex/gender or age groups; however, Black participants had a higher positivity rate than participants of other racial groups despite reporting lower exposure. CONCLUSIONS: Despite the high work-associated exposure rate to SARS-CoV-2 infection, first responders with various roles demonstrated seroprevalence no higher than their administrative/supportive colleagues, which suggests infection control measures are effective in preventing work-related infection.

Spectrum of neurological and survival outcomes in pyruvate dehydrogenase complex (PDC) deficiency: lack of correlation with genotype.

Pyruvate dehydrogenase complex (PDC) deficiency is a relatively common mitochondrial disorder that primarily presents with neurological manifestations and lactic acidemia. We analyzed the clinical outcomes and neurological features of 59 consented symptomatic subjects (27 M, 32 F), who were confirmed to have PDC deficiency with defined mutations in one of the genes of PDC (PDHA1, n = 53; PDHB, n = 4; DLAT, n = 2), including 47 different mutations, of which 22 were novel, and for whom clinical records and/or structured interviews were obtained. 39% of these subjects (23/59) have died. Of these, 91% (21/23) died before age 4 years, 61% (14/23) before 1 year, and 43% (10/23) before 3 months. 56% of males died compared with 25% of females. Causes of death included severe lactic acidosis, respiratory failure, and infection. In subjects surviving past 6 months, a broad range of intellectual outcomes was observed. Of 42 subjects whose intellectual abilities were professionally evaluated, 19% had normal or borderline intellectual ability (CQ/IQ ≥ 70), 10% had mild intellectual disability (ID) (CQ/IQ 55-69), 17% had moderate ID (CQ/IQ 40-54), 24% had severe ID (CQ/IQ 25-39) and 33% had profound ID (CQ/IQ<25). Assessment by parents was comparable. Of 10 subjects who reached age 12 years, 9 had had professional IQ assessments, and only 4 had IQs ≥ 70 (only 2 of these 4 had assessments after age 12 years). The average outcome for females was severe-to-profound ID, whereas that of males was mild-to-moderate ID. Of subjects for whom specific neurological data were available, the majority had hypotonia (89%), and hypertonia or mixed hyper-/hypotonia (49%) were common. Seizures (57%), microcephaly (49%), and structural brain abnormalities including ventriculomegaly (67%) and agenesis, dysgenesis, or hypoplasia of the corpus callosum (55%) were common. Leigh syndrome was found in only 35%. Structural brain abnormalities were more common in females, and Leigh syndrome was more common in males. In a subgroup of 16 ambulatory subjects >3.5 years in whom balance was evaluated, ataxia was found in 13. Peripheral neuropathy was documented in 2 cases but not objectively evaluated in most subjects. Outcomes of this population with genetically confirmed PDC deficiency are heterogeneous and not distinctive. Correlations between specific genotypes and outcomes were not established. Although more females survive, related to the prevalence of X-linked PDHA1 mutations, symptomatic surviving females are generally more severely impaired cognitively and have a different pattern of neurological impairment compared to males. Neonatal or infant onset of symptoms was associated with poor outcomes. Males with PDHA1 mutations and low fibroblast PDC activity were less likely to survive beyond infancy. Recurrence rate in siblings of subjects with PDHA1 mutation was less than 5%. Paradoxically, in this retrospective review, potential factors considered possibly relevant to development, such as in vitro PDC activity, specific mutations, use of ketogenic diets, supplements, or medications, were generally not confirmed to be significantly correlated with objective outcomes of survival or neuro-cognitive function. Therefore, the basis of variability of these outcomes remains largely undetermined.

SARS-CoV-2 antibody profile of naturally infected and vaccinated individuals detected using qualitative, semi-quantitative and multiplex immunoassays.

This study measured antibodies against different antigen targets in healthcare workers (HCW) who have been fully vaccinated with mRNA vaccines, recovered from natural infection, or patients during active infection. All vaccinated individuals were positive for anti-RBD, anti-S1, and anti-S2 antibodies. The nonvaccinated recovered cohort showed 90% seropositivity by Atellica total antibody, 73% by Atellica IgG, 84% by Bioplex anti-RBD, 77% by Bioplex anti-S1, 37% by Bioplex anti-S2, and 79% by Bioplex antinucleocapsid respectively. The active infection cohort exhibited a similar pattern as the recovered cohort. About 88% and 78% of the recovered and active infection cohort produced both anti-spike and anti-N antibodies with Anti-S1/anti-N ratios ranging from 0.07 to 16.26. In summary, fully vaccinated individuals demonstrated an average of 50-fold higher antibody levels than naturally infected unvaccinated individuals with immune reactivity strongly towards RBD/S1 and a weak response to S2. The results support vaccination regardless of previous COVID-infection status.

High-Throughput Adaptable SARS-CoV-2 Screening for Rapid Identification of Dominant and Emerging Regional Variants.

OBJECTIVES: Emerging severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variant strains can be associated with increased transmissibility, more severe disease, and reduced effectiveness of treatments. To improve the availability of regional variant surveillance, we describe a variant genotyping system that is rapid, accurate, adaptable, and able to detect new low-level variants built with existing hospital infrastructure. METHODS: We used a tiered high-throughput SARS-CoV-2 screening program to characterize variants in a supraregional health system over 76 days. Combining targeted reverse transcription-polymerase chain reaction (RT-PCR) and selective sequencing, we screened SARS-CoV-2 reactive samples from all hospitals within our health care system for genotyping dominant and emerging variants. RESULTS: The median turnaround for genotyping was 2 days using the high-throughput RT-PCR-based screen, allowing us to rapidly characterize the emerging Delta variant. In our population, the Delta variant is associated with a lower cycle threshold value, lower age at infection, and increased vaccine-breakthrough cases. Detection of low-level and potentially emerging variants highlights the utility of a tiered approach. CONCLUSIONS: These findings underscore the need for fast, low-cost, high-throughput monitoring of regional viral sequences as the pandemic unfolds and the emergence of SARS-CoV-2 variants increases. Combining RT-PCR-based screening with selective sequencing allows for rapid genotyping of variants and dynamic system improvement.

Seroprevalence of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) among healthcare providers prior to the vaccine era in an integrated midwestern healthcare system.

We performed severe acute respiratory coronavirus virus 2 (SARS-CoV-2) antinucleocapsid IgG testing on 5,557 healthcare providers and found a seroprevalence of 3.9%. African Americans were more likely to test positive than Whites, and HCWs with household exposure and those working on COVID-19 cohorting units were more likely to test positive than their peers.

A Model for Syphilis Screening in the Emergency Department.

The incidence of syphilis infections is on the rise, particularly among African American men and men who have sex with men, and it is reaching epidemic levels in these communities throughout the United States. Although syphilis is relatively inexpensive to treat and cure and is a predictor for HIV incidence among men and transgender women who have sex with men, rates of co-screening for syphilis are low in the emergency department setting, with a dearth of literature on this topic since the 1990s and early 2000s. In this case study, we describe an operational model for routine syphilis screening implemented in June 2017 at the University Hospitals Cleveland Medical Center in Cleveland, Ohio. We describe the advantages of screening using a reverse testing algorithm rather than the traditional method and the necessity of partnering with the Cleveland Department of Public Health for both diagnostic and follow-up logistics.

Procedure-specific preoperative red blood cell preparation and utilization management in pediatric surgical patients.

BACKGROUND: Data-driven practices in preoperative red blood cell (RBC) preparation for pediatric surgical procedures are not well established. Adaptation of established adult preparation guidance methods to pediatric populations may improve perioperative RBC utilization. STUDY DESIGN AND METHODS: A retrospective audit of preoperative RBC preparation volumes (Vp) and intraoperative RBC transfusion volumes (Vt) for pediatric surgical procedures was undertaken at a large children's hospital from January to June 2006. RBC preparation-to-transfusion volume (mL/kg) ratios (P:T) were calculated for all surgeries, subspecialties, and select procedures. P:T equals Vp divided by Vt. Resulting P:Ts were compared to a target P:T of 2:1. A model for maximum procedure-specific Vp (Vp-max) defined Vp-max as the RBC transfusion volume able to meet the needs of 80% of patients undergoing an individual surgical procedure. Vp-max values were applied to the study data set to predict the impact on P:Ts and Vp. RESULTS: RBCs were prepared for 332 surgical procedures and transfused during 113 procedures. P:T was 3.5:1 for total surgical procedures (subspecialty range, 2.7:1-46:0), exceeding the 2:1 target. Vp-max modeling for spinal fusion, craniotomy for neoplasia, craniotomy for seizure, and craniosynostectomy yielded P:T ratios of 1.5:1, 1.5:1, 1.7:1, and 1.0:1, respectively, predicting a 30% decrease in Vp for these four surgical procedures. CONCLUSIONS: P:Ts for pediatric surgical procedures at this institution indicate potentially excessive preoperative RBC preparations. Determination of data-driven procedure-specific Vp may increase the efficiency of preoperative RBC preparation practices.

Von Hippel-Lindau (VHL) disease: an update on the clinico-pathologic and genetic aspects.

von Hippel-Lindau (VHL) disease is an inherited multisystem familial cancer syndrome caused by mutations of the VHL gene on chromosome 3p25. A wide variety of neoplastic processes are known to be associated with VHL disease. The consequences of the VHL mutations and the pathway for tumor development continue to be elucidated. This paper will detail the variety of tumors associated with VHL disease and discuss the genetic mechanisms that lead to the predisposition for neoplasia.

Interference of Therapeutic Monoclonal Antibodies With Routine Serum Protein Electrophoresis and Immunofixation in Patients With Myeloma: Frequency and Duration of Detection of Daratumumab and Elotuzumab.

OBJECTIVES: We investigated the frequency and pattern of detection of therapeutic monoclonal antibodies (t-mAbs) daratumumab and elotuzumab by routine serum protein electrophoresis (SPE) and immunofixation (IF) in treated patients with myeloma. METHODS: Detection of t-mAb was assessed in 22 patients by retrospective review of SPE/IF ordered prior to, during, and after 26 individual courses of therapy. RESULTS: t-mAb was distinguishable from M-protein in 16 of 26 courses, with daratumumab detected in nine of nine and elotuzumab in six of seven patients. t-mAb was detected on first follow-up SPE/IF in 12 patients, with earliest detection 7 days after therapy initiation and latest detection 70 days after therapy. t-mAb persisted throughout induction therapy in most patients, with loss of detection during maintenance daratumumab. CONCLUSIONS: When distinguishable from M-protein, t-mAbs are detectable in 93% of treated patients as soon as 7 days after the initial dose and are consistently observed throughout induction therapy, warranting increased monitoring and careful interpretation of SPE/IF.

Validation of hemolysis index thresholds optimizes detection of clinically significant hemolysis.

OBJECTIVES: Automated hemolysis index (HI) measurement has standardized the identification and gradation of sample hemolysis. METHODS: This study evaluates whether clinically significant changes in the concentration of intracellular analytes occur at manufacturer-recommended automated HI thresholds (HI ≥3, >25 mg/dL hemoglobin). RESULTS: Adult outpatient results for serum potassium (K+), magnesium (Mg), lactate dehydrogenase (LDH), and aspartate aminotransferase (AST) were analyzed. Mean ± SD analyte concentration and distribution within the reference interval (RI) were calculated for each HI group (1-7). Potassium results with an HI of 4 or more demonstrated clinically significant differences (≥0.5 mmol/L) in mean K+ concentration and RI classification compared with non-hemolyzed samples (HI = 1). LDH and AST showed clinically significant differences (+20%) for an HI of 3 or more. For Mg, only the group with an HI of 7 demonstrated a clinically significant difference (>25%); however, the number was low. CONCLUSIONS: Mean measured potassium concentrations are not clinically significantly affected by hemolysis at the manufacturer-recommended HI threshold, while AST and LDH are. Aligning reporting of sample hemolysis with clinically significant changes provides clinically meaningful alerts regarding this common pre-analytic error.

Next-Generation Sequencing to Help Monitor Patients Infected with HIV: Ready for Clinical Use?

Given the extreme variability of the human immunodeficiency virus (HIV) and its ability to replicate as complex viral populations, HIV variants with reduced susceptibility to antiretroviral drugs or with specific coreceptor tropism (CCR5 and/or CXCR4) may be present as minority members of the viral quasispecies. The sensitivity of current HIV genotypic or phenotypic assays is limited, and thus, these tests usually fail to detect low-abundance viral variants. Next-generation (deep) sequencing (NGS) produces an enormous amount of information that allows the detection of minority HIV variants at levels unimaginable using standard Sanger sequencing. NGS technologies continue to evolve, opening new and more affordable opportunities to implement this methodology in clinical laboratories, and HIV is not an exception. The ample use of a battery of more effective antiretroviral drugs, together with careful patient monitoring based on HIV resistance testing, has resulted in HIV-infected patients whose disease is usually well-controlled. The vast majority of adherent patients without detectable resistance become virologically suppressed; however, a subset of these patients with undetectable resistance by standard methods may fail antiretroviral therapy, perhaps due to the presence of minority HIV-resistant variants. Novel NGS-based HIV assays with increased sensitivity for identifying low-level drug resistance and/or coreceptor tropism may play an important role in the success of antiretroviral treatments.