Context-dependent accuracy of the cobas plasma separation card for HCV RNA viral load measurement.

Collection and preservation of plasma are challenging in remote or under-resourced settings. The cobas® Plasma Separation Card (PSC) is an alternative specimen type for blood-borne pathogen nucleic acid quantitation. We assessed PSC as a specimen type for HCV RNA quantitation in Pakistan. Plasma from venous blood and PSC from finger prick blood were prepared at two sites: Site 1 (in Lahore, n = 199) consisted of laboratory-based outpatient clinics. Specimens were prepared in the same facility and stored frozen. Site 2 was a catchment area within a resource-limited, semi-urban locality of Islamabad with limited access to healthcare services (n = 151). Community public health outreach staff collected blood and prepared the PSC in the participants' homes. Specimens were transported to the central hepatitis laboratory in Lahore to be stored frozen until tested. HCV RNA testing was performed using the cobas HCV RNA test in a central laboratory. Concordance with respect to RNA detectability was high at Site 1 (97.4%), but lower at Site 2 (82.4%). At Site 1, HCV viral load in plasma and PSC were well correlated across the linear range with a 0.21 log10 IU/mL mean bias toward higher concentrations in PSC. At Site 2, HCV viral load in plasma and PSC were poorly correlated. There was a 0.11 log10 IU/mL mean bias toward higher concentrations in PSC. PSC performance can be excellent in underserved settings where refrigerated transport of traditional specimens is difficult. In very challenging field settings, extra support must be provided to ensure correct specimen collection and handling.

Assessment of the cobas® HBV RNA investigational assay in the setting of nucleoside analog therapy cessation.

HBV RNA is used as a marker of cccDNA transcription and is applicable in the setting of nucleos(t)ide analog (NA) treatment, which suppresses HBV DNA. Traditional assays for quantification of HBV RNA rely on labor-intensive 3'RACE assays targeting the polyA tail. In this study, the high-throughput Roche cobas®HBV RNA investigational assay was assessed on the Roche cobas® 6800 automated platform. Of 969 samples collected for a NA treatment cessation trial, and tested on the cobas assay, 249 were analyzed for sensitivity, reproducibility, sample type applicability, and results were compared to a RACE-based assay. Results of 97 paired serum and plasma samples demonstrated an excellent correlation of 0.98. However, 14.5% of plasma samples yielded detectable (below the limit of quantification) results, when the paired serum was undetectable, and plasma was shown to yield a statistically significant (p < 0.001) greater mean 0.119 log10 copies/ml. Quantification of 152 samples showed good correlation (0.91) between the cobas and RACE assays. The cobas assay demonstrated superior lower limit of quantification, 10 copies/ml, which resulted in detection of 13.2% more samples than the RACE assay. Reproducibility and linear range of the automated assay were also confirmed. The Roche cobas assay for HBV RNA is sensitive and highly recommended.

National-Level Disparities in Internet Access Among Low-Income and Black and Hispanic Youth: Current Population Survey.

BACKGROUND: Internet access is increasingly critical for adolescents with regard to obtaining health information and resources, participating in web-based health promotion, and communicating with health practitioners. However, past work demonstrates that access is not uniform among youth in the United States, with lower access found among groups with higher health-related needs. Population-level data yield important insights about access and internet use in the United States. OBJECTIVE: The aim of this study is to examine internet access and mode of access by social class and race and ethnicity among youth (aged 14-17 years) in the United States. METHODS: Using the Current Population Survey, we examined internet access, cell phone or smartphone access, and modes of connecting to the internet for adolescents in 2015 (unweighted N=6950; expanded weights N=17,103,547) and 2017 (unweighted N=6761; expanded weights N=17,379,728). RESULTS: Internet access increased from 2015 to 2017, but socioeconomic status (SES) and racial and ethnic disparities remained. In 2017, the greatest disparities were found for youth in low-income households (no home access=23%) and for Black youth (no home access=18%) and Hispanic youth (no home access=14%). Low-income Black and Hispanic youth were the most likely to lack home internet access (no home access, low SES Black youth=29%; low SES Hispanic youth=21%). The mode of access (eg, from home and smartphone) and smartphone-only analyses also revealed disparities. CONCLUSIONS: Without internet access, web-based dissemination of information, health promotion, and health care will not reach a significant segment of youth. Currently, SES and racial and ethnic disparities in access prolong health inequalities. Moreover, the economic impact of COVID-19 on Black, Hispanic, and low-income communities may lead to losses in internet access for youth that will further exacerbate disparities.

Dual composite reference standards (dCRS) in molecular diagnostic research: A new approach to reduce bias in the presence of Imperfect reference.

A main challenge in molecular diagnostic research is to accurately evaluate the performance of a new nucleic acid amplification test when the reference standard is imperfect. Several approaches, such as discrepant analysis, composite reference standard (CRS) method, or latent class analysis (LCA), are commonly applied for this purpose by combining multiple imperfect (reference) test results. In discrepant analysis or LCA, test results from the new assay are often involved in the construction of a new pseudo-reference standard, which results in the potential risk of overestimating the parameters of interest. On the contrary, the CRS methods only combine the results of reference tests, which is more preferable in practice. In this article, we study the properties of two extreme CRS methods, i.e., combining multiple reference test results by the "any positive" rule or by the "all-positive" rule, and propose a new approach "dual composite reference standards (dCRS)" based on these two extreme methods to reduce the biases of the estimates. Simulations are performed for various scenarios and the proposed approach is applied to two real datasets. The results demonstrate that our approach outperforms other commonly used approaches and therefore is recommended for future applications.

A study to assess the impact of the cobas point-of-care RT-PCR assay (SARS-CoV-2 and Influenza A/B) on patient clinical management in the emergency department of the University of California at Davis Medical Center.

BACKGROUND: Rapid detection of SARS-CoV-2 is crucial for reduction of transmission and clinical decision-making. Several rapid (<30 min) molecular point-of-care (POC) tests based on nucleic acid amplification exist for diagnosis of SARS-CoV-2 & Influenza A/B infections. METHODS: This unblinded, pre-post study enrolled consecutive patients with symptoms/signs consistent with SARS-CoV-2 infection presenting to the University of California, Davis emergency department (ED). Outcomes following implementation of the cobas® SARS-CoV-2 & Influenza A/B test for use on the cobas Liat System (intervention: December 2020-May 2021) were compared with previous standard-of-care using centralized laboratory reverse transcriptase polymerase chain reaction (RT-PCR) methods (control: April 2020-October 2020). RESULTS: Electronic health records of 8879 symptomatic patient visits were analyzed, comprising 4339 and 4540 visits and 538 and 638 positive SARS-CoV-2 PCR test results in the control and intervention periods, respectively. Compared with the control period, turnaround time (TAT) was shorter in the intervention period (median 0.98 vs 12.30 h; p < 0.0001). ED length of stay (LOS) was generally longer in the intervention period compared with the control period, but for those SARS-CoV-2-negative who were admitted, ED LOS was shorter (median 12.53 vs 17.93 h; p < 0.0001). The rate of antibiotic prescribing was lower in the intervention than in the control period (42.86% vs 49.16%; p < 0.0001) and antiviral prescribing was higher (7.64% vs 5.49%; p < 0.0001). CONCLUSION: This real-world study confirms faster TAT with a POC RT-PCR method in an emergency care setting and highlights the importance of rapid SARS-CoV-2 detection to aid patient management and inform treatment decisions.

Baseline serum HBV RNA is associated with the risk of hepatitis flare after stopping nucleoside analog therapy in HBeAg-negative participants.

BACKGROUND AND AIMS: HBV RNA in peripheral blood reflects HBV cccDNA transcriptional activity and may predict clinical outcomes. The prospective Melbourne HBV-STOP trial studied nucleot(s)ide analog discontinuation in HBeAg-negative non-cirrhotic participants with long-term virological suppression. Ninety-six weeks after stopping treatment, the proportion of participants with virological relapse (HBV DNA > 2000 IU/mL), biochemical relapse (ALT > 2 × ULN and HBV DNA > 2000 IU/mL), or hepatitis flare (ALT > 5 × ULN and HBV DNA > 2000 IU/mL) was 89%, 58%, and 38%, respectively. We evaluated the ability of serum HBV RNA levels to predict these outcomes. APPROACH RESULTS: HBV RNA levels were measured using the Roche cobas 6800/8800 HBV RNA Investigational Assay. Sixty-five participants had baseline and longitudinal off-treatment specimens available for RNA testing. HBV RNA was detectable at baseline in 25% of participants and was associated with a higher risk of biochemical relapse (81% vs. 51%, p value 0.04) and hepatitis flare (63% vs. 31%, p value 0.04). Participants who had undetectable serum HBV RNA as well as HBsAg ≤ 100 IU/mL at baseline were less likely to experience virological relapse (4 of 9, 44%) than participants with detectable HBV RNA and HBsAg level > 100 IU/mL (15/15, 100%; p value 0.0009). Off-treatment levels of HBV RNA were correlated with HBV DNA and were associated with the risk of hepatitis flare. CONCLUSIONS: Serum HBV RNA may be a useful biomarker for guiding clinical decision-making before stopping nucleot(s)ide analog therapy. Baseline HBV RNA and HBsAg levels are associated with the risk of clinical relapse, hepatitis flare, and disease remission off-treatment.

HBV-RNA Co-amplification May Influence HBV DNA Viral Load Determination.

Despite effective hepatitis B virus (HBV)-DNA suppression, HBV RNA can circulate in patients receiving nucleoside/nucleotide analogues (NAs). Current assays quantify HBV DNA by either real-time polymerase chain reaction (PCR), which uses DNA polymerase, or transcription-mediated amplification, which uses reverse-transcriptase (RT) and RNA polymerase. We assessed the effect of RT capability on HBV-DNA quantification in samples from three cohorts, including patients with quantified HBV RNA. We compared the HBV-DNA levels by real-time PCR (cobas HBV, Roche 6800/8800; Xpert HBV, Cepheid), transcription-mediated amplification (Aptima HBV, Hologic), and real-time PCR with added RT capability (cobas HBV+RT). In the first cohort (n = 45) followed over 192 weeks of NA therapy, on-treatment HBV-DNA levels were higher with cobas HBV+RT than cobas HBV (mean difference: 0.14 log10 IU/mL). In a second cohort (n = 50) followed over 96 weeks of NA therapy, HBV-DNA viral load was significantly higher with the cobas HBV+RT and Aptima HBV compared with the cobas HBV test at all time points after initiation of NA therapy (mean difference: 0.65-1.16 log10 IU/mL). A clinically significant difference was not detected between the assays at baseline. In a third cohort (n = 53), after a median of 2.2 years of NA therapy, we detected HBV RNA (median 5.6 log10 copies/mL) in 23 patients (43.4%). Median HBV-DNA levels by Aptima HBV were 2.4 versus less than 1 log10 IU/mL in samples with HBV RNA and without HBV RNA, respectively (P = 0.0006). In treated patients with HBV RNA, Aptima HBV measured higher HBV-DNA levels than Xpert HBV and cobas HBV. Conclusion: Tests including an RT step may overestimate HBV DNA, particularly in samples with low viral loads as a result of NA therapy. This overestimation is likely due to amplification of HBV RNA and may have an impact on clinical decisions.

Dyslipidemia and adherence to the Mediterranean diet in Croatian HIV-infected patients during the first year of highly active antiretroviral therapy.

We investigated the association of adherence to the Mediterranean diet and other risk factors for dyslipidemia in HIV-infected Croatian patients during the first year of highly active antiretroviral therapy (HAART). Adherence to the Mediterranean diet was determined by a 150-item questionnaire; a 0 to 9-point diet scale was created that stratified respondents as having low adherence (<4 points) and moderate to high adherence (> or = 4 points). We interviewed 117 participants between May 2004 and June 2005 and abstracted their serum lipid measurements taken during the first year of HAART The values of total cholesterol, HDL-cholesterol, LDL-cholesterol and triglycerides increased most prominently in the first 3 to 6 months after initiation of HAART (average increase at 3 months: 25% for total cholesterol, 22% for LDL-cholesterol, 18% for HDL-cholesterol and 43% for triglycerides). A Mediterranean diet and physical activity had no effect on serum lipids. The mean total cholesterol was higher in participants receiving a combination of a non-nucleoside reverse transcriptase inhibitor and a protease inhibitor compared to participants receiving a combination of nucleoside analogs with a non-nucleoside analog or a combination of nucleoside analogs and a protease inhibitor Among individual drug treatments, indinavir/ritonavir had the most unfavorable lipid profile. We conclude that adherence to a Mediterranean diet does not influence serum lipid profiles during the first year of HAART.

Multicenter evaluation of the cobas® HIV-1 quantitative nucleic acid test for use on the cobas® 4800 system for the quantification of HIV-1 plasma viral load.

BACKGROUND AND OBJECTIVES: Measurement of HIV-1 viral load (VL) is necessary to monitor treatment efficacy in patients receiving antiretroviral therapy. We evaluated the performance of the cobas® HIV-1 quantitative nucleic acid test for use on the cobas® 4800 system ("cobas 4800 HIV-1"). METHODS: Limit of detection, linearity, accuracy, precision, and specificity of cobas 4800 HIV-1, COBAS® AmpliPrep/COBAS® Taqman® HIV-1 version 2.0 (CAP/CTM HIV-1 v2) and Abbott RealTime HIV-1 were determined in one or two out of three sites. RESULTS: The limit of detection of the cobas 4800 HIV-1 for 400 µL and 200 µL input volumes was 14.2 copies/mL (95% CI: 12.5-16.6 copies/mL) and 43.9 copies/mL (37.7-52.7 copies/mL), respectively. Cobas 4800 HIV-1 demonstrated 100% specificity, and results were linear for all analyzed group M HIV-1 subtypes. Precision was high (SD < 0.19 log10) across all measured ranges, reagent lots and input volumes. Correlation between cobas 4800 HIV-1 and CAP/CTM HIV-1 v2 or RealTime HIV-1 was high (R2 ≥ 0.95). Agreement between cobas 4800 HIV-1 and CAP/CTM HIV-1 v2 was 96.5% (95.0%-97.7%) at a threshold of 50 copies/mL, and 97.2% (95.8%-98.3%) at 200 copies/mL. Agreement between cobas 4800 HIV-1 and RealTime HIV-1 was 96.6% (93.4%-98.5%) at 50 copies/mL, and 97.0% (94.0%-98.8%) at 200 copies/mL. The mean difference between cobas 4800 HIV-1 and CAP/CTM HIV-1 v2 or RealTime HIV-1 was -0.10 log10 or 0.01 log10, respectively. CONCLUSIONS: The cobas 4800 HIV-1 test is highly sensitive, accurate and correlated well with other assays, including agreement around clinically relevant thresholds, indicating minimal overall VL quantification differences between tested platforms.

Utility of the new cobas HCV test for viral load monitoring during direct-acting antiviral therapy.

BACKGROUND: The COBAS AmpliPrep/COBAS TaqMan assay HCV (CAP/CTM) is widely used in clinical routine for HCV testing. Recently, the new cobas HCV test was established for high throughput testing with minimal operator intervention. As different assays may yield different quantitative/qualitative results that possibly impact treatment decisions, the aim of this study was to externally evaluate the cobas HCV test performance in comparison to CAP/CTM in a clinically relevant setting. METHODS: Serum samples were obtained from 270 patients who received direct acting antiviral therapy with different treatment regimens at two study sites (Hannover and Frankfurt) in 2016. Overall, 1545 samples (baseline, on-treatment and follow-up) were tested in parallel by both assays. RESULTS: The mean difference between cobas HCV and CAP/CTM for the quantification of HCV RNA was 0.008 log10 IU/ml HCV RNA (95% limits of agreement: -0.02-0.036) showing excellent agreement of both assays. With respect to clinical cut offs (HCV RNA detectable vs. target not detected and HCV RNA above the lower limit of quantification (LLOQ) vs.

Microalbuminuria in HIV infection.

OBJECTIVE: Microalbuminuria is associated with increased risk of cardiovascular disease and mortality. The objective of the study was to evaluate if HIV infection was an independent risk factor for microalbuminuria. DESIGN: Cross sectional. METHODS: The relationship between HIV infection and microalbuminuria was assessed using subjects enrolled in the study of Fat Redistribution and Metabolic Change in HIV Infection, which consists of HIV-positive and control men and women. Participants with proteinuria (dipstick >or= 1+) were excluded. RESULTS: Microalbuminuria (urinary albumin/creatinine ratio, ACR > 30 mg/g) was present in 11% of HIV infected, and 2% of control participants (P < 0.001); a fivefold odds after multivariate adjustment (odds ratio, 5.11; 95% confidence interval, 1.97-13.31; P=0.0008). Several cardiovascular risk factors were associated with higher ACR in HIV participants: insulin resistance (HOMA > 4; 32%, P < 0.0001), systolic blood pressure (21%, P = 0.01 for 120-140 versus < 120 mmHg, and 43%, P = 0.06 for > 140 versus < 120 mmHg), and family history of hypertension (17%, P = 0.03). Higher CD4 cell count was associated with lower albumin/creatinine ratio (-24%, P = 0.009 for 200-400 versus < 200 cells/ml and -26%, P = 0.005 for > 400 versus < 200 cells/ml). CONCLUSION: HIV infection had a strong and independent association with microalbuminuria, the severity of which was predicted by markers of insulin resistance, hypertension, and advanced HIV infection. These associations warrant further investigation, as the increased prevalence of microalbuminuria in HIV infection may be a harbinger of future risk of cardiovascular and kidney diseases.

Recalling sexual behavior: a methodological analysis of memory recall bias via interview using the diary as the gold standard.

This study examined the effect of time lag on the validity of retrospective self-reports of sexual behavior. Seventy-five heterosexual students (44 women, 31 men) made daily recordings of sexual behavior, condom use, and alcohol or substance use for 1 month. Ability of respondents to recall sexual behavior recorded during this period was assessed at 1, 2, and 3 months after diary completion using recall interviews (25 interviewed at each interval). For vaginal intercourse, total recall error was significantly greater at 3 months than at 1 month post-diary. For all other variables assessed, the 2- and 3-month time intervals did not produce significant increase in total recall error. Higher frequency of vaginal intercourse, orgasm, and alcohol use prior to sexual activity were associated with total recall error for some but not all behaviors and outcomes. The results provide a partial validation of the diary-interview recall model as a method for studying recall error.

A sensitive branched DNA HIV-1 signal amplification viral load assay with single day turnaround.

Branched DNA (bDNA) is a signal amplification technology used in clinical and research laboratories to quantitatively detect nucleic acids. An overnight incubation is a significant drawback of highly sensitive bDNA assays. The VERSANT® HIV-1 RNA 3.0 Assay (bDNA) ("Versant Assay") currently used in clinical laboratories was modified to allow shorter target incubation, enabling the viral load assay to be run in a single day. To dramatically reduce the target incubation from 16-18 h to 2.5 h, composition of only the "Lysis Diluent" solution was modified. Nucleic acid probes in the assay were unchanged. Performance of the modified assay (assay in development; not commercially available) was evaluated and compared to the Versant Assay. Dilution series replicates (>950 results) were used to demonstrate that analytical sensitivity, linearity, accuracy, and precision for the shorter modified assay are comparable to the Versant Assay. HIV RNA-positive clinical specimens (n = 135) showed no significant difference in quantification between the modified assay and the Versant Assay. Equivalent relative quantification of samples of eight genotypes was demonstrated for the two assays. Elevated levels of several potentially interfering endogenous substances had no effect on quantification or specificity of the modified assay. The modified assay with drastically improved turnaround time demonstrates the viability of signal-amplifying technology, such as bDNA, as an alternative to the PCR-based assays dominating viral load monitoring in clinical laboratories. Highly sensitive bDNA assays with a single day turnaround may be ideal for laboratories with especially stringent cost, contamination, or reliability requirements.

Battering victimization among a probability-based sample of men who have sex with men.

OBJECTIVES: This study measured the prevalence of battering victimization (i.e., experience of psychological/symbolic, physical, and sexual battering) among men who have sex with men (MSM) and identified characteristics of these men. METHODS: A probability-based sample of 2881 MSM living in 4 cities completed telephone interviews between 1996 and 1998. RESULTS: Prevalence estimates were 34% for psychological/symbolic battering, 22% for physical battering, and 5% for sexual battering. The strongest demographic correlate independently associated with all forms of battering was age 40 or younger, whereas education and HIV serostatus were associated with physical and psychological/symbolic violence. CONCLUSIONS: Rates of battering victimization among urban MSM are substantially higher than among heterosexual men and possibly heterosexual women. Public health efforts directed toward addressing intimate partner battering among these men are needed.