Advances in molecular infectious diseases testing in the time of COVID-19.

The Coronavirus Disease of 2019 (COVID-19) pandemic has been a challenging event for laboratory medicine and diagnostics manufacturers. We have had to confront numerous unique and previously unthinkable issues on a daily basis in order to continue offering diagnostic testing for not only Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2), but other testing that was significantly impacted by supply chain and staffing disruptions related to COVID-19. Out of this tremendously stressful and, at times, chaotic environment, decades of innovations and advances in testing methodologies and instrumentation became essential to handle the overwhelming volume of samples with clinically appropriate turn-around-time. Additionally, a number of novel testing approaches and technological innovations emerged to address laboratory and public health needs for widespread testing. In this review we consider both technological advances in infectious diseases testing and other innovations in sample collection, processing, automation, workflow, and testing that have embodied the laboratory response to the COVID-19 pandemic.

Implementation of Pre-Admission Caregiver Testing for COVID-19.

BACKGROUND: Caregivers are often at the bedside of hospitalized children posing an additional risk for coronavirus disease 2019 (COVID-19) transmission. We describe the implementation of caregiver COVID-19 testing before inpatient pediatric admissions and the effect on patient cohorting and bed capacity. METHODS: We implemented an ordering pathway to facilitate COVID-19 testing of caregivers of patients admitted to the inpatient units from the pediatric emergency department, elective procedural admissions, or direct admissions at a tertiary children's hospital in the Northeastern United States in August 2021. Testing was expedited by the clinical laboratory, and caregiver results were used to inform cohorting, infection prevention, and bed management decisions. RESULTS: From August 2021 to January 2022, 2558 caregiver tests were ordered through this pathway, and 83 (3.2%) were positive. Of the positive tests, 72 (86.7%) occurred after December 18, 2021, coinciding with the local Omicron variant wave. Among positives, 67 caregiver or child pairs were identified, and 36 positive caregivers had a COVID-19 negative child leading to use of isolation precautions. Reintroduction of patient cohorting increased overall bed capacity from 74% to 100% of available beds. CONCLUSIONS: The overall incidence of COVID-19 among caregivers before admission correlated well with rates of COVID-19 positivity among asymptomatic adults in the community during the study period. Implementation of caregiver testing increased bed capacity by reintroducing cohorting of patients and identified patients needing isolation that would have been missed by patient testing alone. More research is necessary to determine the extent that routine caregiver testing mitigates the risk of nosocomial severe acute respiratory syndrome coronavirus 2 transmission.

Age-dependent impairment in antibody responses elicited by a homologous CoronaVac booster dose.

The emergence of the SARS-CoV-2 Omicron sublineages resulted in increased transmission rates and reduced protection from vaccines. To counteract these effects, multiple booster strategies were used in different countries, although data comparing their efficiency in improving protective immunity remain sparse, especially among vulnerable populations, including older adults. The inactivated CoronaVac vaccine was among the most widely distributed vaccine worldwide and was essential in the early control of SARS-CoV-2-related hospitalizations and deaths. However, it is not well understood whether homologous versus heterologous booster doses in those fully vaccinated with CoronaVac induce distinct humoral responses or whether these responses vary across age groups. We analyzed plasma antibody responses from CoronaVac-vaccinated younger or older individuals who received a homologous CoronaVac or heterologous BNT162b2 or ChAdOx1 booster vaccine. All three evaluated boosters resulted in increased virus-specific IgG titers 28 days after the booster dose. However, we found that both IgG titers against SARS-CoV-2 Spike or RBD and neutralization titers against Omicron sublineages were substantially reduced in participants who received homologous CoronaVac compared with the heterologous BNT162b2 or ChAdOx1 booster. This effect was specifically prominent in recipients >50 years of age. In this group, the CoronaVac booster induced low virus-specific IgG titers and failed to elevate neutralization titers against any Omicron sublineage. Our results point to the notable inefficiency of CoronaVac immunization and boosting in mounting protective antiviral humoral immunity, particularly among older adults, during the Omicron wave. These observations also point to benefits of heterologous regimens in high-risk populations fully vaccinated with CoronaVac.

Choice Architecture to Assist Clinicians with Appropriate COVID-19 Test Ordering.

BACKGROUND: Despite improving supplies, SARS-CoV-2 nucleic acid amplification tests remain limited during surges and more so given concerns around COVID-19/influenza co-occurrence. Matching clinical guidelines to available supplies ensures resources remain available to meet clinical needs. We report a change in clinician practice after an electronic health record (EHR) order redesign to impact emergency department (ED) testing patterns. METHODS: We included all ED visits between December 1, 2021 and January 18, 2022 across a hospital system to assess the impact of EHR order changes on provider behavior 3 weeks before and after the change. The EHR order redesign included embedded symptom-based order guidance. Primary outcomes were the proportion of COVID-19 + flu/respiratory syncytial virus (RSV) testing performed on symptomatic, admitted, and discharged patients, and the proportion of COVID-19 + flu testing on symptomatic, discharged patients. RESULTS: A total of 52 215 ED visits were included. For symptomatic, discharged patients, COVID-19 + flu/RSV testing decreased from 11.4 to 5.8 tests per 100 symptomatic visits, and the rate of COVID-19 + flu testing increased from 7.4 to 19.1 before and after the intervention, respectively. The rate of COVID-19 + flu/RSV testing increased from 5.7 to 13.1 tests per 100 symptomatic visits for symptomatic patients admitted to the hospital. All changes were significant (P < 0.0001). CONCLUSIONS: A simple EHR order redesign was associated with increased adherence to institutional guidelines for SARS-CoV-2 and influenza testing amidst supply chain limitations necessitating optimal allocation of scarce testing resources. With continually shifting resource availability, clinician education is not sufficient. Rather, system-based interventions embedded within exiting workflows can better align resources and serve testing needs of the community.

Partial ORF1ab Gene Target Failure with Omicron BA.2.12.1.

Mutations in the genome of SARS-CoV-2 can affect the performance of molecular diagnostic assays. In some cases, such as S-gene target failure, the impact can serve as a unique indicator of a particular SARS-CoV-2 variant and provide a method for rapid detection. Here, we describe partial ORF1ab gene target failure (pOGTF) on the cobas SARS-CoV-2 assays, defined by a ≥2-thermocycle delay in detection of the ORF1ab gene compared to that of the E-gene. We demonstrate that pOGTF is 98.6% sensitive and 99.9% specific for SARS-CoV-2 lineage BA.2.12.1, an emerging variant in the United States with spike L452Q and S704L mutations that may affect transmission, infectivity, and/or immune evasion. Increasing rates of pOGTF closely mirrored rates of BA.2.12.1 sequences uploaded to public databases, and, importantly, increasing local rates of pOGTF also mirrored increasing overall test positivity. Use of pOGTF as a proxy for BA.2.12.1 provides faster tracking of the variant than whole-genome sequencing and can benefit laboratories without sequencing capabilities.

Combining genomic and epidemiological data to compare the transmissibility of SARS-CoV-2 variants Alpha and Iota.

SARS-CoV-2 variants shaped the second year of the COVID-19 pandemic and the discourse around effective control measures. Evaluating the threat posed by a new variant is essential for adapting response efforts when community transmission is detected. In this study, we compare the dynamics of two variants, Alpha and Iota, by integrating genomic surveillance data to estimate the effective reproduction number (Rt) of the variants. We use Connecticut, United States, in which Alpha and Iota co-circulated in 2021. We find that the Rt of these variants were up to 50% larger than that of other variants. We then use phylogeography to show that while both variants were introduced into Connecticut at comparable frequencies, clades that resulted from introductions of Alpha were larger than those resulting from Iota introductions. By monitoring the dynamics of individual variants throughout our study period, we demonstrate the importance of routine surveillance in the response to COVID-19.

Rapid emergence of SARS-CoV-2 Omicron variant is associated with an infection advantage over Delta in vaccinated persons.

Background: The SARS-CoV-2 Omicron variant became a global concern due to its rapid spread and displacement of the dominant Delta variant. We hypothesized that part of Omicron's rapid rise was based on its increased ability to cause infections in persons that are vaccinated compared to Delta. Methods: We analyzed nasal swab PCR tests for samples collected between December 12 and 16, 2021, in Connecticut when the proportion of Delta and Omicron variants was relatively equal. We used the spike gene target failure (SGTF) to classify probable Delta and Omicron infections. We fitted an exponential curve to the estimated infections to determine the doubling times for each variant. We compared the test positivity rates for each variant by vaccination status, number of doses, and vaccine manufacturer. Generalized linear models were used to assess factors associated with odds of infection with each variant among persons testing positive for SARS-CoV-2. Findings: For infections with high virus copies (Ct < 30) among vaccinated persons, we found higher odds that they were infected with Omicron compared to Delta, and that the odds increased with increased number of vaccine doses. Compared to unvaccinated persons, we found significant reduction in Delta positivity rates after two (43.4%-49.1%) and three vaccine doses (81.1%), while we only found a significant reduction in Omicron positivity rates after three doses (62.3%). Conclusion: The rapid rise in Omicron infections was likely driven by Omicron's escape from vaccine-induced immunity. Funding: This work was supported by the Centers for Disease Control and Prevention (CDC).

A multicenter evaluation of computable phenotyping approaches for SARS-CoV-2 infection and COVID-19 hospitalizations.

Diagnosis codes are used to study SARS-CoV2 infections and COVID-19 hospitalizations in administrative and electronic health record (EHR) data. Using EHR data (April 2020-March 2021) at the Yale-New Haven Health System and the three hospital systems of the Mayo Clinic, computable phenotype definitions based on ICD-10 diagnosis of COVID-19 (U07.1) were evaluated against positive SARS-CoV-2 PCR or antigen tests. We included 69,423 patients at Yale and 75,748 at Mayo Clinic with either a diagnosis code or a positive SARS-CoV-2 test. The precision and recall of a COVID-19 diagnosis for a positive test were 68.8% and 83.3%, respectively, at Yale, with higher precision (95%) and lower recall (63.5%) at Mayo Clinic, varying between 59.2% in Rochester to 97.3% in Arizona. For hospitalizations with a principal COVID-19 diagnosis, 94.8% at Yale and 80.5% at Mayo Clinic had an associated positive laboratory test, with secondary diagnosis of COVID-19 identifying additional patients. These patients had a twofold higher inhospital mortality than based on principal diagnosis. Standardization of coding practices is needed before the use of diagnosis codes in clinical research and epidemiological surveillance of COVID-19.

Comparative transmissibility of SARS-CoV-2 variants Delta and Alpha in New England, USA.

The SARS-CoV-2 Delta variant rose to dominance in mid-2021, likely propelled by an estimated 40%-80% increased transmissibility over Alpha. To investigate if this ostensible difference in transmissibility is uniform across populations, we partner with public health programs from all six states in New England in the United States. We compare logistic growth rates during each variant's respective emergence period, finding that Delta emerged 1.37-2.63 times faster than Alpha (range across states). We compute variant-specific effective reproductive numbers, estimating that Delta is 63%-167% more transmissible than Alpha (range across states). Finally, we estimate that Delta infections generate on average 6.2 (95% CI 3.1-10.9) times more viral RNA copies per milliliter than Alpha infections during their respective emergence. Overall, our evidence suggests that Delta's enhanced transmissibility can be attributed to its innate ability to increase infectiousness, but its epidemiological dynamics may vary depending on underlying population attributes and sequencing data availability.

Real-world assessment of severe acute respiratory coronavirus virus 2 (SARS-CoV-2) nasopharyngeal swab testing in a region with a high burden of coronavirus disease 2019 (COVID-19).

Concerns persist regarding possible false-negative results that may compromise COVID-19 containment. Although obtaining a true false-negative rate is infeasible, using real-life observations, the data suggest a possible false-negative rate of ˜2.3%. Use of a sensitive, amplified RNA platform should reassure healthcare systems.

Universal SARS-CoV-2 Testing of Emergency Department Admissions Increases Emergency Department Length of Stay.

STUDY OBJECTIVE: Our institution experienced a change in SARS-CoV-2 testing policy as well as substantial changes in local COVID-19 prevalence, allowing for a unique examination of the relationship between SARS-CoV-2 testing and emergency department (ED) length of stay. METHODS: This was an observational interrupted time series of all patients admitted to an academic health system between March 15, 2020, and September 30, 2020. Given testing limitations from March 15 to April 24, all patients receiving SARS-CoV-2 tests were symptomatic. On April 24, testing was expanded to all ED admissions. The primary and secondary outcomes were ED length of stay and number needed to test to obtain a positive, respectively. RESULTS: A total of 70,856 patients were cared for in the EDs during the 7-month period. The testing change increased admission length of stay by 1.89 hours (95% confidence interval 1.39 to 2.38). The number needed to test was 2.5 patients and was highest yield on April 1, 2020, when the state positivity rate was 39.7%; however, the number needed to test exceeded 170 patients by Sept 1, 2020, at which point the state positivity rate was 0.5%. CONCLUSION: Although universal SARS-CoV-2 testing of ED admissions may meaningfully support mitigation and containment efforts, the clinical cost of testing all admissions amid low community positivity is notable. In our system, universal ED SARS-CoV-2 testing was associated with a 24% increase in admission length of stay alongside the detection of only 1 positive case every other day. Given the known harms and risks of ED boarding and crowding, solutions must be developed to support regular operational flow while balancing infection prevention needs.

Considerations from the College of American Pathologists for Implementation of an Assay for SARS-CoV-2 Testing after a Change in Regulatory Status.

The U.S. Food & Drug Administration (FDA) regulates the marketing of manufacturers' in vitro diagnostic tests (IVDs), including assays for the detection of SARS-CoV-2. The U.S. government's Clinical Laboratory Improvement Amendments (CLIA) of 1988 regulates the studies that a clinical diagnostic laboratory needs to perform for an IVD before placing it into use. Until recently, the FDA has authorized the marketing of SARS-CoV-2 IVDs exclusively through the Emergency Use Authorization (EUA) pathway. The regulatory landscape continues to evolve, and IVDs will eventually be required to pass through conventional non-EUA FDA review pathways once the emergency declaration is terminated, in order to continue to be marketed as an IVD in the United States. When FDA regulatory status of an IVD changes or is anticipated to change, the laboratory should review manufacturer information and previously performed internal verification studies to determine what, if any, additional studies are needed before implementing the non-EUA version of the IVD in accordance with CLIA regulations. Herein, the College of American Pathologists' Microbiology Committee provides guidance for how to approach regulatory considerations when an IVD is converted from EUA to non-EUA status.

Mass severe acute respiratory coronavirus 2 (SARS-CoV-2) testing of asymptomatic healthcare personnel.

Mass asymptomatic SARS-CoV-2 nucleic acid amplified testing of healthcare personnel (HCP) was performed at a large tertiary health system. A low period-prevalence of positive HCP was observed. Of those who tested positive, half had mild symptoms in retrospect. HCP with even mild symptoms should be isolated and tested.

Severe Acute Respiratory Syndrome Coronavirus 2 Testing in Children in a Large Regional US Health System During the Coronavirus Disease 2019 Pandemic.

BACKGROUND: The objective was to evaluate patterns of pediatric coronavirus disease 2019 testing in a large health system throughout the pandemic, before and after school reopening. METHODS: This was a cross-sectional time-series study of clinical virology results from children tested for severe acute respiratory syndrome coronavirus 2 in Southern Connecticut and areas of New York and Rhode Island. Data collected include demographics, hospital admission, changes in percent positive tests over time, detection intervals in persistently positive children and cycle threshold values. The setting was the Yale New Haven Health System has 6 hospitals at 4 Connecticut locations, 1 hospital in Rhode Island and ambulatory locations in Connecticut, Rhode Island and New York. Participants included twenty-three-thousand one-hundred thirty-seven children ≤ 18 years of age, tested for coronavirus disease 2019 at an ambulatory testing site, the emergency department or on an inpatient unit within the Yale New Haven Health System. RESULTS: Among all tests, 3.2% were positive. Older children consistently made up the larger portion of positive pediatric cases, regardless of community prevalence. Increased pediatric cases later in the pandemic when prevalence in adults was relatively low correlates with a higher number of tests performed in children and not with an increased positivity rate. No significant changes in trends of positivity were detected after the reopening of schools. Symptomatic and asymptomatic children had similar cycle threshold values regardless of age, and a subset of children demonstrated persistent viral detection, some for as long as 6 weeks. CONCLUSION: An increase in pediatric cases documented in the late summer was predominately due to increased access to testing for children. The percent positivity in children did not change in the first 3 weeks after school opened. A subset of children has detectable severe acute respiratory syndrome coronavirus 2 RNA in the upper respiratory tract for weeks after the initial infection.

Early introductions and transmission of SARS-CoV-2 variant B.1.1.7 in the United States.

The emergence and spread of SARS-CoV-2 lineage B.1.1.7, first detected in the United Kingdom, has become a global public health concern because of its increased transmissibility. Over 2,500 COVID-19 cases associated with this variant have been detected in the United States (US) since December 2020, but the extent of establishment is relatively unknown. Using travel, genomic, and diagnostic data, we highlight that the primary ports of entry for B.1.1.7 in the US were in New York, California, and Florida. Furthermore, we found evidence for many independent B.1.1.7 establishments starting in early December 2020, followed by interstate spread by the end of the month. Finally, we project that B.1.1.7 will be the dominant lineage in many states by mid- to late March. Thus, genomic surveillance for B.1.1.7 and other variants urgently needs to be enhanced to better inform the public health response.

Case Report: Disseminated Strongyloidiasis in a Patient with COVID-19.

The SARS-CoV-2 virus has emerged and rapidly evolved into a current global pandemic. Although bacterial and fungal coinfections have been associated with COVID-19, little is known about parasitic infection. We report a case of a COVID-19 patient who developed disseminated strongyloidiasis following treatment with high-dose corticosteroids and tocilizumab. Screening for Strongyloides infection should be pursued in individuals with COVID-19 who originate from endemic regions before initiating immunosuppressive therapy.

Clinical implications of SARS-CoV-2 cycle threshold values in solid organ transplant recipients.

Detection of SARS-CoV-2 viral RNA by RT-PCR assays is the primary diagnostic test for COVID-19. Cycle threshold (CT ) values generated by some of these assays provide inversely proportional proxy measurements of viral load. The clinical implications of CT values are incompletely characterized, particularly in solid organ transplant (SOT) recipients. We conducted a retrospective chart review of 25 adult SOT recipients admitted to the Yale New Haven Health System between March 1 and May 15, 2020, analyzing 50 test results to investigate the clinical implications of SARS-CoV-2 CT values in this population. Initial CT values from upper respiratory tract samples were significantly higher in patients on tacrolimus, but were not associated with admission severity nor highest clinical acuity. Viral RNA was detected up to 38 days from symptom onset with a gradual increase in CT values over time. In five patients with serial testing, CT values <35.0 were detected >21 days after symptom onset in 4/5 and ≥27 days in 2/5, demonstrating prolonged RNA detection. These data describe SARS-CoV-2 viral dynamics in SOT patients and suggest that CT values may not be useful to predict COVID-19 severity in SOT patients. SARS-CoV-2 CT values may be more useful in informing infection prevention measures.