National integration of mental health screening and treatment into specialized care for cystic fibrosis: What predicts success?

OBJECTIVES: The CF Foundation sponsored competitive awards for Mental Health Coordinators (MHCs) from 2016 to 2018 to implement the international guidelines for mental health screening and treatment in US CF centers. Longitudinal surveys evaluated success in implementing these guidelines using the Consolidated Framework for Implementation Research (CFIR). METHODS: MHCs completed annual surveys assessing implementation from preparation/basic implementation (e.g., using recommended screeners) to full implementation/sustainability (e.g., providing evidence-based treatments). Points were assigned to questions through consensus, with higher scores assigned to more complex tasks. Linear regression and mixed effects models were used to: (1) examine differences in centers and MHC characteristics, (2) identify predictors of success, (3) model the longitudinal trajectory of implementation scores. RESULTS: A total of 122 MHCs (88.4% responded): Cohort 1, N = 80; Cohort 2, N = 30; and Cohort 3, N = 12. No differences in center characteristics were found. Significant improvements in implementation were observed across centers over time. Years of experience on a CF team was the only significant predictor of success; those with 1-5 years or longer reported the highest implementation scores. Change over time was predicted by >5 years of experience. CONCLUSIONS: Implementation of the mental health guidelines was highly successful over time. Funding for MHCs with dedicated time was critical. Longitudinal modeling indicated that CF centers with diverse characteristics could implement them, supported by evidence from the CF Patient Registry showing nearly universal uptake of mental health screening in the United States. Years of experience predicted better implementation, suggesting that education and training of MHCs and retention of experienced providers are critical to success.

Inhibition of LILRB2 by a Novel Blocking Antibody Designed to Reprogram Immunosuppressive Macrophages to Drive T-Cell Activation in Tumors.

Tumor-associated macrophages (TAM) play an important role in maintaining the immunosuppressive state of the tumor microenvironment (TME). High levels of CD163+ TAMs specifically are associated with poor prognosis in many solid tumor types. Targeting TAMs may represent a key approach in development of the next generation of cancer immune therapeutics. Members of the leukocyte immunoglobulin-like receptor B (LILRB) family, including LILRB2 (ILT4), are known to transmit inhibitory signals in macrophages and other myeloid cells. Leveraging bulk and single cell RNA-sequencing datasets, as well as extensive immunophenotyping of human tumors, we found that LILRB2 is highly expressed on CD163+ CD11b+ cells in the TME and that LILRB2 expression correlates with CD163 expression across many tumor types. To target LILRB2, we have developed JTX-8064, a highly potent and selective antagonistic mAb. JTX-8064 blocks LILRB2 binding to its cognate ligands, including classical and nonclassical MHC molecules. In vitro, JTX-8064 drives the polarization of human macrophages and dendritic cells toward an immunostimulatory phenotype. As a result, human macrophages treated with a LILRB2 blocker are reprogrammed to increase the activation of autologous T cells in co-culture systems. Furthermore, JTX-8064 significantly potentiates the activity of anti-PD-1 in allogeneic mixed lymphocyte reaction. In a human tumor explant culture, pharmacodynamic activity of JTX-8064 was observed in monotherapy and in combination with anti-PD-1. Collectively, our work provides strong translational and preclinical rationale to target LILRB2 in cancer.

Inactivation of LATS1/2 drives luminal-basal plasticity to initiate basal-like mammary carcinomas.

Basal-like breast cancers, an aggressive breast cancer subtype that has poor treatment options, are thought to arise from luminal mammary epithelial cells that undergo basal plasticity through poorly understood mechanisms. Using genetic mouse models and ex vivo primary organoid cultures, we show that conditional co-deletion of the LATS1 and LATS2 kinases, key effectors of Hippo pathway signaling, in mature mammary luminal epithelial cells promotes the development of Krt14 and Sox9-expressing basal-like carcinomas that metastasize over time. Genetic co-deletion experiments revealed that phenotypes resulting from the loss of LATS1/2 activity are dependent on the transcriptional regulators YAP/TAZ. Gene expression analyses of LATS1/2-deleted mammary epithelial cells notably revealed a transcriptional program that associates with human basal-like breast cancers. Our study demonstrates in vivo roles for the LATS1/2 kinases in mammary epithelial homeostasis and luminal-basal fate control and implicates signaling networks induced upon the loss of LATS1/2 activity in the development of basal-like breast cancer.

Differential expression of CCR8 in tumors versus normal tissue allows specific depletion of tumor-infiltrating T regulatory cells by GS-1811, a novel Fc-optimized anti-CCR8 antibody.

The presence of T regulatory (Treg) cells in the tumor microenvironment is associated with poor prognosis and resistance to therapies aimed at reactivating anti-tumor immune responses. Therefore, depletion of tumor-infiltrating Tregs is a potential approach to overcome resistance to immunotherapy. However, identifying Treg-specific targets to drive such selective depletion is challenging. CCR8 has recently emerged as one of these potential targets. Here, we describe GS-1811, a novel therapeutic monoclonal antibody that specifically binds to human CCR8 and is designed to selectively deplete tumor-infiltrating Tregs. We validate previous findings showing restricted expression of CCR8 on tumor Tregs, and precisely quantify CCR8 receptor densities on tumor and normal tissue T cell subsets, demonstrating a window for selective depletion of Tregs in the tumor. Importantly, we show that GS-1811 depleting activity is limited to cells expressing CCR8 at levels comparable to tumor-infiltrating Tregs. Targeting CCR8 in mouse tumor models results in robust anti-tumor efficacy, which is dependent on Treg depleting activity, and synergizes with PD-1 inhibition to promote anti-tumor responses in PD-1 resistant models. Our data support clinical development of GS-1811 to target CCR8 in cancer and drive tumor Treg depletion in order to promote anti-tumor immunity.

Genetic testing to gain diagnostic clarity in neurofibromatosis type 2 and schwannomatosis.

Neurofibromatosis type 2 (NF2) and schwannomatosis (SWN) have distinct genetic etiologies but overlapping phenotypes. Genetic testing may be required for accurate diagnosis, which is critical for determining prognosis, screening recommendations, and treatment options. Our study aimed to compare the efficacy of germline-only versus paired (germline and tumor) genetic testing for clarifying the diagnosis in patients with features of NF2 and SWN. We performed a retrospective chart review of patients referred for NF2/SWN genetic testing at Massachusetts General Hospital from 2015 to 2020. Logistic regression analysis was performed to assess factors associated with diagnostic clarity. Overall, paired testing had 8.5 times greater odds of providing diagnostic clarity than germline-only testing (p < 0.01). Among patients who underwent paired testing, those who had analysis of two or more tumors had the greatest likelihood of gaining diagnostic clarity, with odds 13 times greater than patients who underwent germline-only testing (p < 0.01). Paired testing with analysis of one tumor significantly increased the odds of diagnostic clarity over germline-only testing by a factor of 6.5 (p < 0.01). These results have implications for genetic testing strategies and counseling patients about genetic testing utility. They also support the routine use of testing in individuals with suspected NF2 or SWN and improved insurance coverage for paired testing within this population.

Impact of COVID-19 on mental health: Effects on screening, care delivery, and people with cystic fibrosis.

BACKGROUND: Depression and anxiety are two to four times more prevalent in people with CF (pwCF) than the general population. COVID-19 may exacerbate mental health challenges, increasing demand for psychological services, while decreasing their availability. We assessed the impact of the pandemic on depression and anxiety in pwCF, including how COVID-19 affected the frequency of mental health screening and the types of services provided. METHODS: A 38-item internet survey, completed in June 2020, assessed how COVID-19 affected: 1) the mental health clinician's role and screening processes; 2) barriers to screening and resource needs; 3) impact of COVID-19 on depression and anxiety, and 4) positive outcomes and confidence in sustaining mental health screening and treatment, including telehealth services, after the pandemic. RESULTS: Responses were obtained from 131 of the 289 US CF programs. Overall, 60% of programs (n=79) continued mental health screening and treatment, although less frequently; 50% provided individual tele-mental health interventions, and 9% provided telehealth group therapy. Clinically elevated depression symptoms (PHQ-9≥10; moderate to severe), were found in 12% of 785 pwCF, with 3.1% endorsing suicidal ideation. Similarly, elevated anxiety (moderate to severe; GAD-7≥10) was found in 13% of pwCF (n=779). CONCLUSIONS: The COVID-19 pandemic created an opportunity to implement innovative solutions to disruptions in mental health screening and treatment in CF programs. We found that pwCF had increased access to psychological interventions during the pandemic via telehealth, supporting the continued integration of tele-mental health screening and treatment into CF care.

Status and advances in technologies for phosphorus species detection and characterization in natural environment- A comprehensive review.

Poor recovery of phosphorus (P) across natural environment (water, soil, sediment, and biological sources) is causing rapid depletion of phosphate rocks and continuous accumulation of P in natural waters, resulting in deteriorated water quality and aquatic lives. Accurate detection and characterization of various P species using suitable analytical methods provide a comprehensive understanding of the biogeochemical cycle of P and thus help its proper management in the environment. This paper aims to provide a comprehensive review of the analytical methods used for P speciation in natural environment by dividing them into five broad categories (i.e., chemical, biological, molecular, staining microscopy, and sensors) and highlighting the suitability (i.e., targeted species, sample matrix), detection limit, advantages-limitations, and reference studies of all methods under each category. This can be useful in designing studies involving P detection and characterization across environmental matrices by providing insights about a wide range of analytical methods based on the end user application needs of individual studies.

Promoting emotional wellness in children with CF, part II: Mental health assessment and intervention.

This is the second of two companion papers that examine the emotional wellness of children with cystic fibrosis (CF) during the early years of life, defined here as the period between birth and age 12. Both papers promote optimal mental health and well-being, with an emphasis on early identification and intervention. The first paper explores child and family resilience. Here, we discuss strategies for pediatric CF teams to provide routine, systematic mental health assessment, anticipatory guidance, brief intervention, and triage to evidence-based treatment when needed, while addressing barriers to accessing care. Many mental health conditions emerge before the age of 12, with the potential for lifelong effects on individuals, their families, and society. Living with a chronic illness such as CF can further increase the risk of mental health concerns and, in a bidirectional manner, their consequences for the quality of life, sustaining daily care, and health outcomes. There has been a significant focus in recent years on the mental health and wellness of adolescents and adults with CF, but less attention to specifics of depression and anxiety in younger children, or to other common pediatric comorbidities including trauma, developmental disorders such as attention-deficit/hyperactivity disorder or autism spectrum disorder, and oppositional behavior. Given the availability of psychometrically sound screening instruments and effective interventions, routinely addressing the mental health of children with CF and their families is feasible to integrate within multidisciplinary CF care, allowing for a personalized approach respecting individual needs, values, and goals.

Promoting emotional wellness in children with cystic fibrosis, Part I: Child and family resilience.

Attention should be given to individual and family well-being from a child's first interaction with the medical team and continuing throughout development, especially for families who experience chronic illnesses, such as cystic fibrosis (CF). While much attention has been given to the mental health of people with CF 12 years and older, this paper explores various areas for CF teams to assess and provide additional resources during the first 12 years of a child's life to promote child and family wellness. In this paper, we discuss parental mental health, social determinants of health, adherence/self-care, nutrition, attention to family lifestyle factors, engagement with school and peers, and modulator therapy for this age group of people with CF. This is the first of two companion papers which examines emotional wellness of children during the early years. The second paper examines mental health assessment and intervention for children under 12. Both encourage teams to strive to promote optimal child and family emotional health and wellness, emphasizing holistic health promotion and prevention, early identification, and intervention.

Quantitative Method for Comparative Assessment of Particle Removal Efficiency of Fabric Masks as Alternatives to Standard Surgical Masks for PPE.

In response to the COVID-19 pandemic, cloth masks are being used to control the spread of virus, but the efficacy of these loose-fitting masks is not well known. Here, tools and methods typically used to assess tight-fitting respirators were modified to quantify the efficacy of community-produced and commercially produced fabric masks as personal protective equipment. Two particle counters concurrently sample ambient air and air inside the masks; mask performance is evaluated by mean particle removal efficiency and statistical variability when worn as designed and with a nylon overlayer, to independently assess fit and material. Worn as designed, both commercial surgical masks and cloth masks had widely varying effectiveness (53%-75% and 28%-91% particle removal efficiency, respectively). Most surgical-style masks improved with the nylon overlayer, indicating poor fit. This rapid testing method uses widely available hardware, requires only a few calculations from collected data, and provides both a holistic and aspect-wise evaluation of mask performance.

Introduction to Cystic Fibrosis for Mental Health Care Coordinators and Providers: Collaborating to Promote Wellness.

To maximize health, individuals with cystic fibrosis (CF) follow a complex, burdensome daily care regimen. Managing CF is associated with a range of significant biopsychosocial challenges and places individuals with CF, and their caregivers, at greater risk for developing anxiety and depression. To promote wellness and address the potential barriers that affect management of this complex chronic illness, many individuals would benefit from treatment from a behavioral health provider. Social workers within multidisciplinary CF care teams are well positioned to respond to this need, and an expanding number of social workers serving as behavioral health providers in the community will be sought as a resource to provide treatment to this population. This article serves as a primer for social workers to maximize knowledge of the psychosocial and potential behavioral health needs of individuals with CF across the life span. To best support individuals with CF, authors describe the disease-specific manifestations and outline the numerous potential clinical targets for social work to promote wellness. The article concludes by highlighting the importance of communication with the medical team and considerations for effective collaborative care.

Development and Analytical Validation of a DNA Dual-Strand Approach for the US Food and Drug Administration-Approved Next-Generation Sequencing-Based Praxis Extended RAS Panel for Metastatic Colorectal Cancer Samples.

A next-generation sequencing method was developed that can distinguish single-stranded modifications from low-frequency somatic mutations present on both strands of DNA in formalin-fixed paraffin-embedded colorectal cancer samples. We applied this method for analytical validation of the Praxis Extended RAS Panel, a US Food and Drug Administration-approved companion diagnostic for panitumumab, on the Illumina MiSeqDx platform. With the use of the TruSeq amplicon workflow, both strands of DNA from the starting material were interrogated independently. Mutations were reported only if found on both strands; artifacts usually present on only one strand would not be reported. A total of 56 mutations were targeted within the KRAS and NRAS genes. A minimum read depth of 1800× per amplicon is required per sample but averaged >30,000× at maximum multiplexing levels. Analytical validation studies were performed to determine the simultaneous detection of mutations on both strands, reproducibility, assay detection level, precision of the assay across various factors, and the impact of interfering substances. In conclusion, this assay can clearly distinguish single-stranded artifacts from low-frequency mutations. Furthermore, the assay is accurate, precise, and reproducible, can achieve consistent detection of a mutation at 5% mutation frequency, exhibits minimal impact from tested interfering substances, and can simultaneously detect 56 mutations in a single run using 10 samples plus controls.

Recontact practices of cancer genetic counselors and an exploration of professional, legal, and ethical duty.

The duty to recontact continues to be revisited in the field of clinical genetics and is currently relevant for cancer genetic counseling given the transition from single-gene to multi-gene panel testing. We recruited cancer genetic counselors through the National Society of Genetic Counselors list-serv to complete an online survey assessing current practices and perspectives regarding recontacting patients about diagnostic genetic tests. Forty-one percent of respondents reported that they have recontacted patients to offer updated (new) diagnostic genetic testing (40/97). A majority (61%, 17/28), of genetic counselors who reported recontact specifically for panel testing indicated that the availability of management recommendations for genes not previously tested routinely was an important factor in the decision to recontact. All respondents who recontacted patients reported "improved patient care" as a perceived benefit. Respondents indicated that recontact is mostly a patient responsibility (49%), followed by a shared responsibility between the provider and patient (43%). Few respondents (2%) reported a uniform ethical duty to recontact patients regarding new and updated testing, while the majority (89%) felt that there was some degree of ethical duty. A greater percentage of those who reported past recontact practices reported intention to recontact in the future (p = 0.001). There is little consensus among the genetic counselor respondents about how to approach the recontacting of patients to offer updated genetic testing.

Statistical generation of training sets for measuring NO3(-), NH4(+) and major ions in natural waters using an ion selective electrode array.

Knowledge of ionic concentrations in natural waters is essential to understand watershed processes. Inorganic nitrogen, in the form of nitrate and ammonium ions, is a key nutrient as well as a participant in redox, acid-base, and photochemical processes of natural waters, leading to spatiotemporal patterns of ion concentrations at scales as small as meters or hours. Current options for measurement in situ are costly, relying primarily on instruments adapted from laboratory methods (e.g., colorimetric, UV absorption); free-standing and inexpensive ISE sensors for NO3(-) and NH4(+) could be attractive alternatives if interferences from other constituents were overcome. Multi-sensor arrays, coupled with appropriate non-linear signal processing, offer promise in this capacity but have not yet successfully achieved signal separation for NO3(-) and NH4(+)in situ at naturally occurring levels in unprocessed water samples. A novel signal processor, underpinned by an appropriate sensor array, is proposed that overcomes previous limitations by explicitly integrating basic chemical constraints (e.g., charge balance). This work further presents a rationalized process for the development of such in situ instrumentation for NO3(-) and NH4(+), including a statistical-modeling strategy for instrument design, training/calibration, and validation. Statistical analysis reveals that historical concentrations of major ionic constituents in natural waters across New England strongly covary and are multi-modal. This informs the design of a statistically appropriate training set, suggesting that the strong covariance of constituents across environmental samples can be exploited through appropriate signal processing mechanisms to further improve estimates of minor constituents. Two artificial neural network architectures, one expanded to incorporate knowledge of basic chemical constraints, were tested to process outputs of a multi-sensor array, trained using datasets of varying degrees of statistical representativeness to natural water samples. The accuracy of ANN results improves monotonically with the statistical representativeness of the training set (error decreases by ∼5×), while the expanded neural network architecture contributes a further factor of 2-3.5 decrease in error when trained with the most representative sample set. Results using the most statistically accurate set of training samples (which retain environmentally relevant ion concentrations but avoid the potential interference of humic acids) demonstrated accurate, unbiased quantification of nitrate and ammonium at natural environmental levels (±20% down to <10 µM), as well as the major ions Na(+), K(+), Ca(2+), Mg(2+), Cl(-), and SO4(2-), in unprocessed samples. These results show promise for the development of new in situ instrumentation for the support of scientific field work.

The role of viral introductions in sustaining community-based HIV epidemics in rural Uganda: evidence from spatial clustering, phylogenetics, and egocentric transmission models.

BACKGROUND: It is often assumed that local sexual networks play a dominant role in HIV spread in sub-Saharan Africa. The aim of this study was to determine the extent to which continued HIV transmission in rural communities--home to two-thirds of the African population--is driven by intra-community sexual networks versus viral introductions from outside of communities. METHODS AND FINDINGS: We analyzed the spatial dynamics of HIV transmission in rural Rakai District, Uganda, using data from a cohort of 14,594 individuals within 46 communities. We applied spatial clustering statistics, viral phylogenetics, and probabilistic transmission models to quantify the relative contribution of viral introductions into communities versus community- and household-based transmission to HIV incidence. Individuals living in households with HIV-incident (n = 189) or HIV-prevalent (n = 1,597) persons were 3.2 (95% CI: 2.7-3.7) times more likely to be HIV infected themselves compared to the population in general, but spatial clustering outside of households was relatively weak and was confined to distances <500 m. Phylogenetic analyses of gag and env genes suggest that chains of transmission frequently cross community boundaries. A total of 95 phylogenetic clusters were identified, of which 44% (42/95) were two individuals sharing a household. Among the remaining clusters, 72% (38/53) crossed community boundaries. Using the locations of self-reported sexual partners, we estimate that 39% (95% CI: 34%-42%) of new viral transmissions occur within stable household partnerships, and that among those infected by extra-household sexual partners, 62% (95% CI: 55%-70%) are infected by sexual partners from outside their community. These results rely on the representativeness of the sample and the quality of self-reported partnership data and may not reflect HIV transmission patterns outside of Rakai. CONCLUSIONS: Our findings suggest that HIV introductions into communities are common and account for a significant proportion of new HIV infections acquired outside of households in rural Uganda, though the extent to which this is true elsewhere in Africa remains unknown. Our results also suggest that HIV prevention efforts should be implemented at spatial scales broader than the community and should target key populations likely responsible for introductions into communities.

Safety and efficacy of low-dose paclitaxel utilizing the cobra-P drug-eluting stent system with a novel biodegradable coating in de novo coronary lesions: the PLUS-ONE first-in-man study.

BACKGROUND: The Cobra-P drug-eluting stent (DES) system consists of cobalt chromium alloy with bio-absorbable siloxane sol-gel matrix coating that elutes low dose paclitaxel within 6 months. The aim of this first-in-man trial was to evaluate the safety and performance of 2 doses of the Cobra-P DES. METHODS: A total of 60 lesions (54 patients) were sequentially assigned to 2 different paclitaxel doses: group A (3.7 µg/18mm, n=30) or group B (8 µg/18mm, n=30). The primary endpoint was MACE at 4 months defined as cardiac death, myocardial infarction, and target lesion revascularization. RESULTS: Patient and lesion characteristics were matched between the 2 groups except for male sex. MACE at 4 months was 3.3% and 0% respectively (P=1.000) and at 1-year follow-up remained unchanged. In-stent late loss at 4 months was similar in both groups (0.36 ± 0.30mm and 0.34 ± 0.20mm P=.773). CONCLUSIONS: In this FIM study, implantation of the Cobra-P low dose paclitaxel-eluting stent with a bioabsorbable sol-gel coating was proven to be feasible and safe. Moderate neointimal proliferation was observed as well as an acceptable MACE rate up to 1 year.

Extended artificial neural networks: incorporation of a priori chemical knowledge enables use of ion selective electrodes for in-situ measurement of ions at environmentally relevant levels.

A novel artificial neural network (ANN) architecture is proposed which explicitly incorporates a priori system knowledge, i.e., relationships between output signals, while preserving the unconstrained non-linear function estimator characteristics of the traditional ANN. A method is provided for architecture layout, disabling training on a subset of neurons, and encoding system knowledge into the neuron structure. The novel architecture is applied to raw readings from a chemical sensor multi-probe (electric tongue), comprised of off-the-shelf ion selective electrodes (ISEs), to estimate individual ion concentrations in solutions at environmentally relevant concentrations and containing environmentally representative ion mixtures. Conductivity measurements and the concept of charge balance are incorporated into the ANN structure, resulting in (1) removal of estimation bias typically seen with use of ISEs in mixtures of unknown composition and (2) improvement of signal estimation by an order of magnitude or more for both major and minor constituents relative to use of ISEs as stand-alone sensors and error reduction by 30-50% relative to use of standard ANN models. This method is suggested as an alternative to parameterization of traditional models (e.g., Nikolsky-Eisenman), for which parameters are strongly dependent on both analyte concentration and temperature, and to standard ANN models which have no mechanism for incorporation of system knowledge. Network architecture and weighting are presented for the base case where the dot product can be used to relate ion concentrations to both conductivity and charge balance as well as for an extension to log-normalized data where the model can no longer be represented in this manner. While parameterization in this case study is analyte-dependent, the architecture is generalizable, allowing application of this method to other environmental problems for which mathematical constraints can be explicitly stated.

Factors associated with incorrect identification of recent HIV infection using the BED capture immunoassay.

The BED capture enzyme immunoassay (BED-CEIA) was developed for estimating HIV incidence from cross-sectional data. This assay misclassifies some individuals with nonrecent HIV infection as recently infected, leading to overestimation of HIV incidence. We analyzed factors associated with misclassification by the BED-CEIA. We analyzed samples from 383 men who were diagnosed with HIV infection less than 1 year after a negative HIV test (Multicenter AIDS Cohort Study). Samples were collected 2-8 years after HIV seroconversion, which was defined as the midpoint between the last negative and first positive HIV test. Samples were analyzed using the BED-CEIA with a cutoff of OD-n ≤ 0.8 for recent infection. Logistic regression was used to identify factors associated with misclassification. Ninety-one (15.1%) of 603 samples were misclassified. In multivariate models, misclassification was independently associated with highly active antiretroviral treatment (HAART) for >2 years, HIV RNA <400 copies/ml, and CD4 cell count <50 or <200 cells/mm(3); adjusted odds ratios (OR) and 95% confidence intervals (CI) were 4.72 (1.35-16.5), 3.96 (1.53-10.3), 6.85 (2.71-17.4), and 11.5 (3.64-36.0), respectively. Among 220 men with paired samples, misclassification 2-4 years after seroconversion was significantly associated with misclassification 6-8 years after seroconversion [adjusted OR: 25.8 (95% CI: 8.17-81.5), p<0.001] after adjusting for race, CD4 cell count, HIV viral load, and HAART use. Low HIV viral load, low CD4 cell count, and >2 years of HAART were significantly associated with misclassification using the BED-CEIA. Some men were persistently misclassified as recently infected up to 8 years after HIV seroconversion.

Short communication: Colony-forming hematopoietic progenitor cells are not preferentially infected by HIV type 1 subtypes A and D in vivo.

HIV subtype C has previously been shown to infect hematopoietic progenitor cells (HPCs) at a significantly higher rate than subtype B. To better understand the subtype-specific nature of HPC infection, we examined the prevalence of HPC infection in vivo by HIV-1 subtypes A and D. HIV-1 infection of HPC was examined in 40 individuals, 19 infected with subtype A and 21 with subtype D, using a single colony assay format. DNA from 1177 extracted colonies was tested for integrated viral DNA of the p24 gene. Four colonies were found to be stably infected, three of 462 colonies (0.65%) from HIV-1A-infected individuals (1/19 individuals) and one of 715 colonies (0.14%) from HIV-1D-infected individuals (1/22 individuals). These rates of colony infection were comparable to the rates observed in PBMCs from the same subjects. Additionally, no correlation was observed between cell colony density and circulating viral load or proviral load. Our findings suggest that HIV-1 subtypes A and D do not preferentially infect colony-forming HPCs over mature HIV target cells in vivo.

Towards an automated, standardized protocol for determination of equilibrium potential of ion-selective electrodes.

An automated real-time method for determination of ISE steady state value and response time is developed, following most recent IUPAC recommendations. Specifically, detection of the 'steady state' is related to (1) the time derivative of the emf as it reaches a limiting value (ΔE/Δt(limit), e.g., 0.1-1.0 mV min(-1)) and (2) the duration of time for which the absolute value of the time derivative remains less than this limiting value (stability window, denoted win(st)). A suite of representative ISEs, including glass, solid state, and polymer-based electrodes, is examined to determine sensitivity of results to parameterization choice. Measurements taken over a wide range of concentration values and in un-processed samples (i.e., without use of ionic strength adjustment) provide insight into behavior of ISEs in applications where analyte concentrations span a wide range and/or sample pre-processing may not be an option, e.g., use of sensors for in situ environmental sampling. Results show that declared steady state emf is strongly sensitive to variations in ΔE/Δt(limit) but relatively unaffected by changes in the stability window when win(st) ≥30 s. Linearity of calibration curves produced, quantified by root mean squared error (RMSE) against a linear fit, improves as ΔE/Δt(limit) decreases, however the percentage of measurements which reach a declared steady state within the prescribed sample window (∼6.5 min) falls with corresponding decreases in the ΔE/Δt(limit) parameter. Response time, defined as the time required to reach declared steady emf, is also a strong function of parameterization. Dependence of response times on sample composition and/or ISE membrane composition and type are also discussed; results for ISEs in samples comprised exclusively of interfering ions are included. In general, limiting emf derivatives of {0.25-0.4 mV min(-1)} and stability windows of {30-40s} achieve both good analytical accuracy and compliance with potentially short sampling window requirements. Methodology based on use of these parameters can improve sampling speed and accuracy as well as promote inter-comparison of data and ISE characterizations among research teams.