Dementia assessment and management in primary care settings: a survey of current provider practices in the United States.

BACKGROUND: Primary care providers (PCPs) are typically the first to screen and evaluate patients for neurocognitive disorders (NCDs), including mild cognitive impairment and dementia. However, data on PCP attitudes and evaluation and management practices are sparse. Our objective was to quantify perspectives and behaviors of PCPs and neurologists with respect to NCD evaluation and management. METHODS: A cross-sectional survey with 150 PCPs and 50 neurologists in the United States who evaluated more than 10 patients over age 55 per month. The 51-item survey assessed clinical practice characteristics, and confidence, perceived barriers, and typical practices when diagnosing and managing patients with NCDs. RESULTS: PCPs and neurologists reported similar confidence and approaches to general medical care and laboratory testing. Though over half of PCPs performed cognitive screening or referred patients for cognitive testing in over 50% of their patients, only 20% reported high confidence in interpreting results of cognitive tests. PCPs were more likely to order CT scans than MRIs, and only 14% of PCPs reported high confidence interpreting brain imaging findings, compared to 70% of specialists. Only 21% of PCPs were highly confident that they correctly recognized when a patient had an NCD, and only 13% were highly confident in making a specific NCD diagnosis (compared to 72 and 44% for neurologists, both p < 0.001). A quarter of all providers identified lack of familiarity with diagnostic criteria for NCD syndromes as a barrier to clinical practice. CONCLUSIONS: This study demonstrates how PCPs approach diagnosis and management of patients with NCDs, and identified areas for improvement in regards to cognitive testing and neuroimaging. This study also identified all providers' lack of familiarity with published diagnostic criteria for NCD syndromes. These findings may inform the development of new policies and interventions to help providers improve the efficacy of their decision processes and deliver better quality care to patients with NCDs.

Laboratory validation of a clinical metagenomic sequencing assay for pathogen detection in cerebrospinal fluid.

Metagenomic next-generation sequencing (mNGS) for pan-pathogen detection has been successfully tested in proof-of-concept case studies in patients with acute illness of unknown etiology but to date has been largely confined to research settings. Here, we developed and validated a clinical mNGS assay for diagnosis of infectious causes of meningitis and encephalitis from cerebrospinal fluid (CSF) in a licensed microbiology laboratory. A customized bioinformatics pipeline, SURPI+, was developed to rapidly analyze mNGS data, generate an automated summary of detected pathogens, and provide a graphical user interface for evaluating and interpreting results. We established quality metrics, threshold values, and limits of detection of 0.2-313 genomic copies or colony forming units per milliliter for each representative organism type. Gross hemolysis and excess host nucleic acid reduced assay sensitivity; however, spiked phages used as internal controls were reliable indicators of sensitivity loss. Diagnostic test accuracy was evaluated by blinded mNGS testing of 95 patient samples, revealing 73% sensitivity and 99% specificity compared to original clinical test results, and 81% positive percent agreement and 99% negative percent agreement after discrepancy analysis. Subsequent mNGS challenge testing of 20 positive CSF samples prospectively collected from a cohort of pediatric patients hospitalized with meningitis, encephalitis, and/or myelitis showed 92% sensitivity and 96% specificity relative to conventional microbiological testing of CSF in identifying the causative pathogen. These results demonstrate the analytic performance of a laboratory-validated mNGS assay for pan-pathogen detection, to be used clinically for diagnosis of neurological infections from CSF.

Hepatitis C Virus Infection Among Reproductive-Aged Women and Children in the United States, 2006 to 2014.

BACKGROUND: In the United States, hepatitis C virus (HCV) infection has increased among young persons who inject drugs, but the extent of this epidemic among reproductive-aged women and their children is unknown. OBJECTIVE: To estimate numbers and describe characteristics of reproductive-aged women with HCV infection and of their offspring. DESIGN: Analysis of the National Notifiable Diseases Surveillance System (NNDSS) from 2006 to 2014 and the Quest Diagnostics Health Trends national database from 2011 to 2014. SETTING: United States. PARTICIPANTS: 171 801 women (aged 15 to 44 years) and 1859 children (aged 2 to 13 years) with HCV infection reported to the NNDSS; 2.1 million reproductive-aged women and 56 684 children who had HCV testing by Quest Diagnostics. MEASUREMENTS: NNDSS HCV case reports and Quest laboratory data regarding unique reproductive-aged women and children who were tested for HCV infection. RESULTS: The number of reproductive-aged women with acute and past or present HCV infection in the NNDSS doubled, from 15 550 in 2006 to 31 039 in 2014. Of 581 255 pregnant women tested by Quest from 2011 to 2014, 4232 (0.73% [95% CI, 0.71% to 0.75%]) had HCV infection. Of children tested by Quest, 0.76% (CI, 0.69% to 0.83%) had HCV infection, but the percentage was 3.2-fold higher among children aged 2 to 3 years (1.62% [CI, 1.34% to 1.96%]) than those aged 12 to 13 years (0.50% [CI, 0.41% to 0.62%]). Applying the Quest HCV infection rate to annual live births from 2011 to 2014 resulted in an estimated average of 29 000 women (CI, 27 400 to 30 900 women) with HCV infection, who gave birth to 1700 infants (CI, 1200 to 2200 infants) with the infection each year. LIMITATIONS: Only a fraction of HCV infections is detected and reported to the NNDSS. Quest data are potentially biased, because women who are asymptomatic, do not access health care, or have unreported risks may be less likely to be tested for HCV infection. CONCLUSION: These data suggest a recent increase in HCV infection among reproductive-aged women and may inform deliberations regarding a role for routine HCV screening during pregnancy. PRIMARY FUNDING SOURCE: Centers for Disease Control and Prevention.

The Burden of Hepatitis C Infection-Related Liver Fibrosis in the United States.

BACKGROUND: Knowledge of the estimated proportion of hepatitis C virus (HCV)-infected persons with advanced fibrosis or cirrhosis is critical to estimating healthcare needs. METHODS: We analyzed HCV-related testing conducted by Quest Diagnostics from January 2010 through December 2013. Tests included hepatitis C antibody, HCV RNA, HCV genotype (nucleic acid tests [NAT]), liver function tests, and platelet counts; patient age was also determined. Aspartate aminotransferase (AST)-to-platelet ratio (APRI) was calculated as = 100*(aspartate aminotransferase [AST]/upper limit of AST)/platelet. Fibrosis-4 (FIB-4) was calculated as (age × AST)/(platelet ×√ alanine aminotransferase [ALT]). Persons were "currently infected" if they had ≥1 positive HCV NAT; "in care" if a positive RNA test was followed <6 months by ≥1 additional NAT(s), or ALT, AST, and platelets <90 days, or any test ordered by an infectious diseases or gastroenterology specialist; and "evaluated for treatment" if they had a genotype test. RESULTS: Approximately 10 million HCV test results were analyzed, representing 5.6 million unique patients. Of the 2.6 million patients with data to estimate liver disease, 5% were currently infected. Among those currently infected, APRI and FIB-4 scores indicated that 23% overall-and 27% among the cohort born during 1945-1965-had advanced fibrosis or cirrhosis at first diagnosis. A total of 54% of infected were in care and 51% of infected with advanced fibrosis or cirrhosis were evaluated for treatment. CONCLUSIONS: Testing from a large US commercial laboratory indicates that about 1 in 4 HCV-infected persons have levels of liver disease put them at highest risk for complications and could benefit from immediate antiviral therapy.

Quantitative measurement of HER2 expression in breast cancers: comparison with 'real-world' routine HER2 testing in a multicenter Collaborative Biomarker Study and correlation with overall survival.

INTRODUCTION: Accurate assessment of HER2 status is critical in determining appropriate therapy for breast cancer patients but the best HER2 testing methodology has yet to be defined. In this study, we compared quantitative HER2 expression by the HERmark™ Breast Cancer Assay (HERmark) with routine HER2 testing by immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH), and correlated HER2 results with overall survival (OS) of breast cancer patients in a multicenter Collaborative Biomarker Study (CBS). METHODS: Two hundred and thirty-two formalin-fixed, paraffin-embedded breast cancer tissues and local laboratory HER2 testing results were provided by 11 CBS sites. HERmark assay and central laboratory HER2 IHC retesting were retrospectively performed in a blinded fashion. HER2 results by all testing methods were obtained in 192 cases. RESULTS: HERmark yielded a continuum of total HER2 expression (H2T) ranging from 0.3 to 403 RF/mm2 (approximately 3 logs). The distribution of H2T levels correlated significantly (P<0.0001) with all routine HER2 testing results. The concordance of positive and negative values (equivocal cases excluded) between HERmark and routine HER2 testing was 84% for local IHC, 96% for central IHC, 85% for local FISH, and 84% for local HER2 status. OS analysis revealed a significant correlation of shorter OS with HER2 positivity by local IHC (HR=2.6, P=0.016), central IHC (HR=3.2, P=0.015), and HERmark (HR=5.1, P<0.0001) in this cohort of patients most of whom received no HER2-targeted therapy. The OS curve of discordant low (HER2 positive but H2T low, 10% of all cases) was aligned with concordant negative (HER2 negative and H2T low, HR=1.9, P=0.444), but showed a significantly longer OS than concordant positive (HER2 positive and H2T high, HR=0.31, P=0.024). Conversely, the OS curve of discordant high (HER2 negative but H2T high, 9% of all cases) was aligned with concordant positive (HR=0.41, P=0.105), but showed a significantly shorter OS than concordant negative (HR=41, P<0.0001). CONCLUSIONS: Quantitative HER2 measurement by HERmark is highly sensitive, accurately quantifies HER2 protein expression and correlates well with routine HER2 testing. When HERmark and local HER2 results were discordant, HERmark more accurately predicted overall survival.

Determination of HIV-1 coreceptor tropism using proviral DNA in women before and after viral suppression.

BACKGROUND: An HIV-1 tropism test is recommended prior to CCR5 antagonist administration to exclude patients harboring non-R5 virus from treatment with this class of antiretrovirals. HIV-1 tropism determination based on proviral DNA (pvDNA) may be useful in individuals with plasma viral RNA suppression. We developed a genotypic tropism assay for pvDNA and assessed its performance in a retrospective analysis of samples collected longitudinally. RESULTS: We randomly selected paired plasma/PBMC samples from the Women's Interagency HIV Study with plasma viral load ≥5,000 cp/mL at time 1 (T1), undetectable viral load maintained for ≥1 year and CD4+ >200 cells/µL at time 2 (T2). pvDNA was isolated from cryopreserved PBMCs. Sequences were analyzed in triplicate from 49/50 women, with tropism assigned using the geno2pheno (g2p) algorithm. The median time between T1 and T2 was 4.1 years. CXCR4-using virus was detected in 24% of the RNA samples and 33% of the pvDNA samples at T1, compared to 37% of the pvDNA samples at T2. Concordance between plasma RNA and pvDNA tropism was 88% at T1 and 80% at T2. The g2p scores for RNA (T1) vs DNA (T1, T2) were strongly correlated (Spearman rho: 0.85 (T1); 0.78 (T2)). In women with evidence of tropism switch at T2 (either R5 to non-R5 or non-R5 to R5), there was a correlation between change in tropism and time. Mean pvDNA viral load decreased by 0.4 log10 copies/106 cells between T1 and T2 (p < 0.0001), but this decrease was not significantly associated with tropism status. CONCLUSIONS: We demonstrated that pvDNA tropism in women with HIV-1 suppression is concordant with prior RNA tropism results, even after a median time of >4 years. pvDNA tropism testing may be useful to determine eligibility of patients with viral suppression to switch to a CCR5-antagonist based regimen as well as for research purposes.

A genotypic HIV-1 proviral DNA coreceptor tropism assay: characterization in viremic subjects.

BACKGROUND: HIV-1 coreceptor tropism testing is used to evaluate eligibility for CCR5 antagonist therapy. However, HIV-1 RNA-based tests are not suitable for virologically suppressed patients, therefore the use of proviral DNA tropism testing has been investigated. We describe a novel proviral DNA-based genotypic tropism assay and compare its performance to that of a sensitive HIV-1 RNA-based genotypic test. METHODS: Tropism was determined using HIV-1 plasma RNA and proviral DNA from 42 paired samples from patients with plasma viral loads ≥1000 HIV-1 RNA copies/mL. Proviral DNA sample types included whole blood, separated peripheral blood mononuclear cells resuspended in phosphate-buffered saline and peripheral blood mononuclear cells resuspended in spun plasma. The HIV-1 envelope V3 region was PCR-amplified, sequenced in triplicate, and analyzed for tropism with the geno2pheno algorithm using a 10% false-positive rate (FPR). RESULTS: Amplicons were obtained from proviral DNA and plasma RNA in 41/42 samples. Tropism predictions were highly concordant (93%-98%) between proviral DNA and plasma RNA, regardless of the proviral DNA isolation method. Non-R5 proviral DNA results were obtained for 100% of patients with detectable non-R5 plasma HIV-1 RNA results. Geno2pheno FPRs for proviral DNA and plasma RNA were highly correlated (Spearman rho = 0.86). CONCLUSIONS: Our findings demonstrate that proviral DNA tropism determinations from whole blood or peripheral blood mononuclear cells were highly concordant with plasma HIV-1 RNA tropism determinations. This assay may be useful for screening virologically suppressed patients for CCR5-antagonist eligibility and for research purposes.

Comparison of genotypic and phenotypic HIV type 1 tropism assay: results from the screening samples of Cenicriviroc Study 202, a randomized phase II trial in treatment-naive subjects.

Cenicriviroc is a once-daily oral CCR5/CCR2 antagonist in development for treatment of HIV infection. CVC Study 202 (652-2-202; NCT01338883) excluded treatment-naive subjects demonstrated to harbor non-R5 (CXCR4-tropic or dual-mixed) tropic HIV-1 by either genotypic or phenotypic tropism testing. Here we compare the results of genotypic and phenotypic tropism testing in Study 202. A total of 304 subjects screened had paired genotypic and phenotypic results. Genotypic tropism testing (GTT) incorporated triplicate population sequencing using the geno2pheno algorithm and the PSSM algorithm, followed by ultradeep sequencing (UDS) for samples with R5 results. All samples were further evaluated with a phenotypic test, the enhanced-sensitivity Trofile assay (ESTA). Concordance between GTT and ESTA was 80% and increased to 84% when only geno2pheno was used for triplicate population sequencing. GTT (geno2pheno) classified 18% of the samples as non-R5 compared to 16% by ESTA. Only one-third of samples with non-R5 results by either test were classified as non-R5 by both tests. Median CD4((+)) cell counts were lower in patients with concordant non-R5 results by UDS and ESTA than in subjects with an R5 result by either assay (p=0.0004). UDS detected non-R5 virus in an additional 27/304 subjects (median 15% non-R5, interquartile range: 3.7-62%) with R5 results by ESTA. In conclusion, the geno2pheno algorithm improves concordance of GTT with a clinically validated phenotypic tropism assay as does the use of UDS. These findings provide support for recent guidelines indicating that genotypic tropism testing may be considered as an alternative to phenotypic testing.

A genotypic test for HIV-1 tropism combining Sanger sequencing with ultradeep sequencing predicts virologic response in treatment-experienced patients.

A tropism test is required prior to initiation of CCR5 antagonist therapy in HIV-1 infected individuals, as these agents are not effective in patients harboring CXCR4 (X4) coreceptor-using viral variants. We developed a clinical laboratory-based genotypic tropism test for detection of CCR5-using (R5) or X4 variants that utilizes triplicate population sequencing (TPS) followed by ultradeep sequencing (UDS) for samples classified as R5. Tropism was inferred using the bioinformatic algorithms geno2pheno([coreceptor]) and PSSM(x4r5). Virologic response as a function of tropism readout was retrospectively assessed using blinded samples from treatment-experienced subjects who received maraviroc (N = 327) in the MOTIVATE and A4001029 clinical trials. MOTIVATE patients were classified as R5 and A4001029 patients were classified as non-R5 by the original Trofile test. Virologic response was compared between the R5 and non-R5 groups determined by TPS, UDS alone, the reflex strategy and the Trofile Enhanced Sensitivity (TF-ES) test. UDS had greater sensitivity than TPS to detect minority non-R5 variants. The median log(10) viral load change at week 8 was -2.4 for R5 subjects, regardless of the method used for classification; for subjects with non-R5 virus, median changes were -1.2 for TF-ES or the Reflex Test and -1.0 for UDS. The differences between R5 and non-R5 groups were highly significant in all 3 cases (p<0.0001). At week 8, the positive predictive value was 66% for TF-ES and 65% for both the Reflex test and UDS. Negative predictive values were 59% for TF-ES, 58% for the Reflex Test and 61% for UDS. In conclusion, genotypic tropism testing using UDS alone or a reflex strategy separated maraviroc responders and non-responders as well as a sensitive phenotypic test, and both assays showed improved performance compared to TPS alone. Genotypic tropism tests may provide an alternative to phenotypic testing with similar discriminating ability.

Testing and reporting practices for improved management of hepatitis C.

Recent advances in diagnosis and treatment are providing physicians with new options for managing patients with chronic HCV infection. The potential of these new technologies, however, can only be fully realized in the US if surveillance of new cases is improved; this can be achieved by establishing and implementing comprehensive test reporting requirements and by ensuring that physicians have a good understanding of appropriate test ordering and interpretation. Harmonized reporting standards, combined with physician education, will lead to improved identification of infected individuals and increased timeliness of medical interventions.

A public health model for the molecular surveillance of HIV transmission in San Diego, California.

BACKGROUND: Current public health efforts often use molecular technologies to identify and contain communicable disease networks, but not for HIV. Here, we investigate how molecular epidemiology can be used to identify highly related HIV networks within a population and how voluntary contact tracing of sexual partners can be used to selectively target these networks. METHODS: We evaluated the use of HIV-1 pol sequences obtained from participants of a community-recruited cohort (n = 268) and a primary infection research cohort (n = 369) to define highly related transmission clusters and the use of contact tracing to link other individuals (n = 36) within these clusters. The presence of transmitted drug resistance was interpreted from the pol sequences (Calibrated Population Resistance v3.0). RESULTS: Phylogenetic clustering was conservatively defined when the genetic distance between any two pol sequences was less than 1%, which identified 34 distinct transmission clusters within the combined community-recruited and primary infection research cohorts containing 160 individuals. Although sequences from the epidemiologically linked partners represented approximately 5% of the total sequences, they clustered with 60% of the sequences that clustered from the combined cohorts (odds ratio 21.7; P < or = 0.01). Major resistance to at least one class of antiretroviral medication was found in 19% of clustering sequences. CONCLUSION: Phylogenetic methods can be used to identify individuals who are within highly related transmission groups, and contact tracing of epidemiologically linked partners of recently infected individuals can be used to link into previously defined transmission groups. These methods could be used to implement selectively targeted prevention interventions.

Association of HIV-1 replication capacity with treatment outcomes in patients with virologic treatment failure.

BACKGROUND: The extent to which HIV-1 replication capacity (RC) influences the response to therapy remains to be established. METHODS: Phenotypic susceptibility and RC of baseline isolates (n = 139) from patients enrolled in the ARGENTA trial were measured and correlated to treatment outcomes over 36 months. RESULTS: RC in baseline isolates correlated with the number of phenotypically active drugs (R = 0.34, P < 0.001). Crude viral RC did not predict treatment outcomes. When viral RC was adjusted by baseline CD4 cell counts, HIV-1 RNA levels, and phenotypic susceptibility to the rescue regimen, it showed significant association with the immunologic outcome (per log10 RC higher, mean difference in 36 months' time-averaged change from baseline CD4 count = -68 cells/microL; P = 0.020). In the subgroup of patients with 3 or more phenotypically active drugs in the salvage regimen (n = 35, median RC = 65%), subjects carrying isolates with RC < or =65% as compared to those with RC >65% had better time-averaged HIV-1 RNA responses (mean: -1.04 vs. -0.32 log10 copies/mL; P = 0.012) and CD4 cell responses (mean: 132 vs. -7 cells/microL; P = 0.006). Among patients with HIV-1 RNA levels persistently >500 copies/mL (n = 61), RC, on a log10 basis, was inversely associated with time-averaged 36-month CD4 cell responses (beta = -0.26; P = 0.046). CONCLUSION: After normalizing for viral susceptibility to the employed regimen or in patient subsets with suboptimal virologic response, higher viral RC may predict worse subsequent treatment outcomes.

Clinical utility of HIV standard genotyping among antiretroviral-naive individuals with unknown duration of infection.

In clinical settings, we have found a high rate of human immunodeficiency virus (HIV) drug resistance among antiretroviral-naive patients for whom the duration of infection was unknown. These high rates were most likely the result of both transmitted resistance and informal antiretroviral use, and they suggest that routine resistance testing among antiretroviral-naive patients would be a cost-effective clinical practice.

Active methamphetamine use is associated with transmitted drug resistance to non-nucleoside reverse transcriptase inhibitors in individuals with HIV infection of unknown duration.

BACKGROUND: Frequent methamphetamine use among recently HIV infected individuals is associated with transmitted drug resistance (TDR) to non-nucleoside reverse transcriptase inhibitors (NNRTI); however, the reversion time of TDR to drug susceptible HIV may exceed 3 years. We assessed whether recreational substance use is associated with detectable TDR among individuals newly diagnosed with HIV infection of unknown duration. DESIGN: Cross-sectional analysis. METHODS: Subjects were enrolled at the University California, San Diego Early Intervention Program. Demographic, clinical and substance use data were collected using structured interviews. Genotypic resistance testing was performed using GeneSeq, Monogram Biosciences. We analyzed the association between substance use and TDR using bivariate analyses and the corresponding transmission networks using phylogenetic models. RESULTS: Between April 2004 and July 2006, 115 individuals with genotype data were enrolled. The prevalence of alcohol, marijuana and methamphetamine use were 98%, 71% and 64% respectively. Only active methamphetamine use in the 30 days prior to HIV diagnosis was independently associated with TDR to NNRTI (OR: 6.6; p=0.002). CONCLUSION: Despite not knowing the duration of their HIV infection, individuals reporting active methamphetamine use in the 30 days prior to HIV diagnosis are at an increased risk of having HIV strains that are resistant to NNRTI.